\(.---l.93

I

S tereo selective Functionalizationof cr-Amino Acids

A Thesis

Submitted Towards the

Degree of

Doctor of Philosophy

by

Craig Anthony HuttonB.Sc. (Hons.)

Department of Chemistry

University of Adelaide

April 1993

Awarde d lqq 3

Contents

Acknowledgements

Statement

Publications

Abstract

Introduction

Results and Discussion

Chapter 1 Regiocontrolled Side Chain Bromination of

N-Phthaloyl Protected o-Amino Acid Derivatives

Chapter 2 Stereocontrolled Synthesis of Homochiral

Hydroxy- a-Amino Acid Derivatives

Chapter 3 Towards a Synthesis of Chloramphenicol

Chapter 4 Scope and Limitations of the Elaboration of

B romo- a-Amino Acid Derivatives

Chapter 5 Synthesis of Each Stereoisomer ofp-Deuteriophenylalanine

Chapter 6 Stereochemical Course of the reaction of

(S)- and (R)-Phenylalanine with

(S) -Phenylalanine Ammonia Lyase

Conclusion

Experimental

References

i

ü

iiiiv

105

IT2

115

166

1

27

45

72

78

94

Appendices 104lo+

I

ACKNOWLEDGEMENTS

Many thanks go to Dr. Chris Easton for his guidance over the past few

years. I consider myself fortunate to have had a supervisor who provided

continual optimism, inspiration, enthusiasm and support, as well as sincere

friendship.

I would like to thank the staff and students of my department for providing

an enjoyable working environment. Particular thanks are extended to the

members of my research group and the members of lab 3, Kathy, Jeff and

Steve, for their inspiring discussions. Thanks also to Caroline Ward and

Darren Schliebs for proof reading parts of this thesis.

Special thanks go to Pete Roselt, "Barney" Miller and Eng Wui Tan for

their insights and friendship"

I would also tike to thank my family for their encouragement throughout

the years of my education. Last but not least, thanks go to Kathy for keeping

me sane, especially while I was writing this thesis.

l1

STATEMENT

This work contains no material which has been accepted for the award of

any other degree or diploma in any university or other tertiary institution, and,

to the best of my knowledge and belief, contains no material previously

published or written by another person, except where due reference has been

made in the text.

I give consent to this copy of my thesis, when deposited in the University

Library, being available for loan or photocopying.

Craig Hutton (B.Sc. Hons.) April 14 1993

u1

PUBLICATIONS

Some of the work described in this thesis has been reported in the

following publications:

"Synthesis of Homochiral Hydroxy-a-Amino Acid Derivatives", Easton, C.

J., Hutton, C. 4., Tan, E. W., and Tiekink, E. R. T., Tetrahedron Lett., 1990,

31, 7059.

"Regioselective Functionalization of N-Phthaloyl-Substituted Amino Acid

and Peptide Derivatives", Easton, C. J., Hutton, C. 4., Rositano, G., and Tan,

E. W., J. Org. Chem.,1991,56,5614"

"Synthesis and Molecular Structure of Stable Derivatives of (E)- and

(Z)-Dehydrophenylalanine", Easton, C. J., Hutton, C. 4., Roselt, P. D., and

Tiekink, E. R. T., Aust. J. Chem.,I99I,44,681.

"Crystal Structure of N-Phthaloyl-B-Phenylserine Methyl Ester", Easton,

C. J., Hutton, C.4., and Tiekink, E. R. T.,Z.Krist.,1993,203,310.

lv

ABSTRACT

The cl-position of N-phthaloyl-cr-amino acid derivatives has been shown to

be deactivated towards hydrogen atom abstraction, such that bromination

occurs regiospecifically via the most stable side chain radical.

N-Phthaloyl derivatives of B-bromovaline, y-bromoleucine and

y-bromohomophenylalanine have been converted to the conesponding hydroxy

amino acid derivatives. B-Bromo-phenylalanine and tyrosine derivatives

convert stereoselectively to (25)-threo-þ-hydroxy-phenylalanine and tyrosine

derivatives. It has been found that the stereoslectivity is much greater when the

carboxyl group is protected as an amide rather than as an ester, which is

attributable to the amide substituent providing greater intramolecular

stabilization of intermediate carbocations. Deprotection of B-hydroxy-

phenylalanine derivatives yietds the free amino acid ( 2 S 3 R ) -

B-hydroxyphenylalanine without loss of chiral integrity. A B-hydroxy-

p-nitrophenylalanine derivative which is a potential precursor to

chloramphenicol has been synthesized.

lH N.m.r. spectroscopic correlations have been ascertained which allow the

determination of the relative stereochemistry of B-bromo- and B-hydroxy-

phenylalanine and tyrosine derivatives.

B-Bromophenylalanine derivatives have been converted to 3,4-disubstituted

p-lactams and B-lactones with control of both relative and absolute

stereochemistry. Elaboration of 1-bromo-leucine and homophenylalanine

derivatives has enabled the introduction of functionality at the B- and

ô-positions of leucine derivatives and at the B-position of homophenylalanine

derivatives. B-Bromophenylalanine derivatives have also been converted to

deuteiurn labetleC phenylalanine deri.,,atives. These labelled compounds have

been deprotected to provide each of the stereoisomets of

B-deuteri-qphenylalanine with ca.987o d.e. and 997o 2[rincorporation. The

v

B-deuteriophenylalanines have been used to determine the stereochemical

course of the reactions of (S)- and (R)-phenylalanine with the enzyme

(S)-phenylalanine ammonia lyase. The enzyme catalyzes the stereospecific

removal of the pro-S hydrogen from (S)-phenylalanine, whereas the pro-R

hydrogen is preferentially abstracted from the unnatural substrate,

(R)-phenylalanine.

Introduction 1

INTRODUCTION

cx,-Amino acids (1) occur in nature as the monomeric building blocks of

peptides and proteins, and in many other species.l Although only twenty amino

acids occur commonly in proteins there are currently more than five hundred

known naturally occurring amino acids.2'3 Biosynthesis of many of the non-

proteinogenic amino acids is presumed to proceed viø functionalization of the

side chains of proteinogenic amino acids.4

H¡N

RH(1)

In addition to the fundamental biochemical and physiological significance

of amino acids, both natural and unnatural amino acids have found importance

in the pharmaceutical and agricultural industries,5,6 aS flavours,7 taste

enhancers and sweeteners.S In addition to their industrial uses, major advances

in the understanding of enzyme mechanisms,g protein conformalis¡5lo-l2 ¿n¿

regulatory interactions of peptidesl3 have placed the study of amino acid

chemistry at the forefront of chemical research.l4 As a result of the wide

spectrum of applications of amino acids, their economic impact is quite

significant, with a multi-bitlion dollar indusfry existing for their production

either by extraction from natural sources or by chemical synthesis.

Given their widespread natural occurrence and physiological importance,

intense interest has been focussed on the synthesis and subsequent studies of

cx,-amino acids. Many uncommon amino acids are unavailable from natural

sources in sufficient quantities for thorough structural and biochemical testing,

and it is often only through synthesis of these physiologically important

compoun$s that it becomes possible to confirm their structure and function.

+

Introduction 2

Biochemical tabelling studies,15,16 development of enzyme inhibitorslT-20 and

conformational restriction studiesl0-12 arc but a few of the areas where the

synthesis of rare and unnatural amino acids has provided insight into the

functioning of biological systems.

One approach to the synthesis of chiral unconunon cr-amino acids involves

the elaboration of proteinogenic a-amino acids through manipulation of pre-

existing side-chain functionality. This technique utilizes the important fact that

the proteinogenic cr-amino acids ¿ìre a remarkably cheap and readily available

source of enantiomerically pure starting materials. Examples of this approach

include modification of the hydroxyl moiety of serine,2t-24 and modification of

aspartic and glutamic acids vía eithq Barton decarboxylation procedures25,26

or anionic intermediates.2T Vederas and co-ws¡þsrc22-24 have detailed the

preparation of p-substituted amino acids vía ring opening reactions of

p-lactones derived from serine (Scheme 1). (S)-N-tert-Butoxycarbonylserine

(2a) and (S)-N-benzyloxycarbonylserine (2b) were converted to the

corresponding B-lactones (3a) and (3b) with dimethyl azidodicarboxylate

RNH lht,.

DMADNrnI' RNH

Nu

Nu

co2H

Nu----+PPh3

(2) (3) (4)

(a)R=Boc= Me3CO-CO-

(b) R = Cbz= PhCH2O-CO-

Nu = nucleophile +

(s)

cF3co2H

o HtNrr,". coi+

Scheme 1

o

(ó)

Nu

Introduction 3

(DMAD) and triphenylphosphine. Ring opening with various nucleophiles gave

the corresponding N-protected B-substituted alanine derivatives (4).

Alternatively, the B-lactone (3a) was deprotected by treatment with

ftifluoroacetic acid, with subsequent ring opening leading directly to the free

amino acids (6). This method was found to be useful for the synthesis of mono-

B-substituted amino acid derivatives, but not of p,p-disubstituted amino acid

derivatives.2S

Complementary to the modification of amino acid derivatives via the

manipulation of pre-existing functionality is the direct functionalization of the

side chains of proteinogenic amino acid derivatives. This concept not only

utilizes the inherent chirality of ø-amino acids but also the fact that many, if

not most of the naturally occurring uncommon ø-amino acids are derived from

the proteinogenic o-amino acids viq functionalization of the side chains.

Ohfune and Simamoto29 have developed a novel synthesis of (S)-threo-

B-hydroxy-O-methyltyrosine (10) by direct benzylic oxidation of the

K2S20s

CuSOa

MeO MeO0)

Ac

o

Ac

(8)

i) Ba(OH)2ü) Boc2O

+H3Nza,.. coi

.,rrrOH i)Ptoz/02

BocNHlht,.OH

.,rrtlOH

ü) cF3co2H

(e)MeO(10)

Scheme 2

MeO

Introduction 4

N-tert-butoxycarbonyltyrosinol derivative (7) (Scheme 2). Oxidation of the

tyrosinol derivative (7) with potassium persulfate-copper sulfate gave the cyclic

carbamate (8) with high threo-selectivity. Subsequent hydrolysis and

reprotection of the amino group gave the diol (9), with oxidation and removal

of the tert-butoxycarbonyl (Boc) protecting group giving (S)-threo-p.hydroxy-

O-methyltyrosine (10) in reasonable overall yield. However, this procedure is

rather limited by its lack of generality, with attempts to synthesize the

corresponding (S)-threo-isomer of p-hydroxyphenylalanine (25) yía an

analogous pathway being unsuccess ful.29

Direct functionalization of the side chains of amino acids via fuee radical

halogenation has also been reported, and has gained renewed interest in recent

years. This approach is exemplified by the y-chlorination and subsequent

elaboration of lysine (Scheme 3), as described by Kollonitsch and

co-workers.30-32 Ultraviolet irradiation of a concentrated hydrochloric acid

solution of lysine (11), with the concurrent introduction of chlorine gas, led to

the formation of y-chlorolysine (12). Subsequent elaboration of the chloride

(12) gave the hydroxylysine derivative (13).

+H¡N

cl2 / HCI

hv iÐ H*

co2H+

H¡N+

H¡N

-oHÐ

+ +NH3 NH3

(13)

Scheme 3

The regioselectivity of hydrogen atom abstraction reactions is determined

by several factors, including radical stability, steric effects and polar effects.33

(1 1) (12)

Introduction 5

Radical stability is important when there is extensive carbon-hydrogen bond

breaking, and consequently significant radical character, in the transition state.

Steric factors can influence regioselectivity by hindering the approach of the

hydrogen atom abstractor or by consfraining the conformation of the product

radical. Polar effects refer to activating/deactivating effects brought on by

partial charge development in the transition state of the hydrogen atom

abstraction.33'34 Polar effects were used to explain the regioselectivity

observed in the chlorination of lysine (11) described above" Hydrogen atom

abstraction by the electrophilic chlorine atom creates a partial positive charge

in the transition state on the carbon from which the hydrogen is being

abstracted. The cr- and e-aminium groups and the carboxyl group aÍe

inductively electron withdrawing, and therefore deactivate adjacent positions to

attack by the elecfrophilic chlorine radical" Consequently, reaction occurs at

the position farthest from both the a- and e-positions, that is, at the y-position.

Polar effects have been used to explain the regioselectivity observed in several

radical processes in biological systems" The biosynthesis of

(S)-Ê-hydroxyvaline (37) involves enzymic oxidation at the B-position of a

valine residue in a peptide, and there is strong evidence that the oxidation

involves radical intermediates,3S with the observed regioselectivity arising due

to polar effecß.36

The widespread application of radical reactions to the functionalization of

o-amino acids is limited by the virtual insolubility of amino acids in the

organic solvents generally used in such processes. This problem can be

overcome by suitable protection of the amino acids, however such protection

has been shown to markedly effect the regioselectivity of radical reactions.

Whereas hydrogen atom abstraction from lysine (11) was shown in an acidic

environment to occur at the y-position, due to deactivation by the q,- and

e-substituents, hydrogen atom transfer reactions of N-acyl-cr-amino acid

derivative! generally favour formation of the conesponding a-centred radicals

Introduction 6

(Figure 1).37'38 Shbitization of radicals of this type results from overlap of the

semi-occupied p-orbital of the radical with the n-orbitals of the electron

donating (dative) amido substituent and the electron withdrawing (capto)

carboxy substituent. This type of radical is extremely stable and is referred to

as a captodative radical,39 although the terms merostabilízeda} and push-pull4l

radicals have also been used. Radical functionalization of N-acyl-protected

o-amino acid derivatives is therefore limited due to reaction occurring at the

o-centres rather than on the side-chains. Formation of the cr,-centred radicals

also desfroys the chirality at that position, leading to formation of enantiomeric

mixtures of products.

Ro

R'

Figure 1. cx,-Centred radical formed by hydrogen atom abstraction from anN-acylamino acid.

In contrast, N"lV-diacyl-protected amino acid derivatives undergo hydrogen

atom abstraction at the o-position much less readily. Whereas treatment of

N-benzoylglycylglycine methyl ester (14) with N-bromosuccinimide (NBS)

results in hydrogen atom abstraction and subsequent bromine incorporation at

the N-terminal glycine residtre, to give the bromide (15), reaction of

N-phthaloylglycylglycine methyl ester (16) with NBS results in bromination of

the C-terminal glycine residue to give the bromide (17) (Scheme 4).42

Similarly, reaction of the N-acyl-N'-methylpiperazinedione (18) with one

equivaient of NBS resulted in specific incorporation of bromine at the

cr-position of the N-methylglycine residue to give the bromide (19) (Scheme

5).+r i'

o

NI

H

PhCONH

(14)

PhthN

(16)

PhthN phthalimido

NAc

NBSPhCONH

NBS

Introduction 1

Br

(1s)

Br

CO2Me

Â"o*r^"orr"PhrtrN

Scheme 4

CONH

(r7)

NAcNBS

MeN MeN

(18) Br

(1e)

Scheme 5

It was found that with N-phthaloyl-protected cr,-amino acid derivatives

hydrogen atom abstraction at the cx,-centres is deactivated to such an extent that

reaction occurs on the side chains if suitably stabilized radicals can be formed.

Accordingly, bromination of N-phthaloylvaline methyl ester (20) and

N-phthaloylphenylalanine methyl ester (22) resulted in regiospecific formation

of the p-bromides (21) and (23), respectively (Scheme 6).42

The bromination of N-phthaloylamino acid derivatives was envisaged to be

an effective method for the direct introduction of functionality onto the side

chains of various proteinogenic amino acids. The generality of side-chain

bromination reactions of N-phthaloyl-protected cr-amino acid derivatives and

Introduction 8

the basis of the deactivating effect of the N-phthaloyl-protecting group is

therefore the subject of the investigation described in Chapter 1 of the Results

and Discussion of this thesis.

PhthN PhthN Me

NBS

Br

(20) (2r)

PhthN PhthN

NBSBr

(22) (23)

Scheme 6

The brominated amino acid derivatives described in Chapter 1 of the

Results and Discussion were envisaged to be suitable precursors to a wide

variety of substituted q-amino acids. One such important class of substituted

amino acids is that of the hydroxyamino acids, especially the p-hydroxy-

a-amino acids, which are ubiquitous in nature and play essential physiological

roles. Examples of naturally occurring B-hydroxy-q,-amino acids include the

common amino acids serine, threonine and the 4-hydroxyproline (24).

B-Hydroxyphenylalanine (25) and p-hydroxytyrosine (26) have been implicated

as precursors in the biosynthesis of the hypertensive agents epinephrine (27b)

and norepinephrine (27a) (adrenalin and noradrenalin),44 and of the antibiotic

Introduction 9

chloramphenicol (28¡,+s and as components of peptidases46 and esterases.4

Chloramphenicol (28), isolated from Streptomyces venezuelae in 1947, was one

of the first broad spechum antibiotics to be used medicinally.aT It is also the

only naturally occurring antibiotic which is economically produced on an

industrial scale by synthesis rather than by fermentation.48

+H¡N

+ coi

HO

(24)

RNHCI

.,rrttOHcr

OH

(21)

ozN

R

R=HR=OH

(2s)(26)

H

OH

(a) R=H(b) R=Me (28)

p-Hydroxy-o-amino acids have also found many synthetic applications, for

example in the synthesis of B-lactam antibioliçs.49'5O Millers0 has converted

various hydroxamate derivatives of B-hydroxyamino acids (29) to the

corresponding p-lactams (30), and then elaborated these B-lactams (30) in the

synthesis of a large number of different p-lactam antibiotics, including

penicillins, cephalosporins and monobactams (Scheme 7).

Introduction IO

RCONH -oR3 RCO o

plOH Bl 3

R

(2e)

penicillins monobactams

cephalosporins

Scheme 7

More recently, B-hydroxyamino acids have received attention due to thei¡

common occurence in a family of biologically active compounds known as the

"cyclic peptides". One of the most important groups in this family is the

vancomycin group of antibiotics.5l Vancomycin (31) was isolated in 1956

from Streptomyces orientalis52 and is currently employed in widespread

clinical üse,53 especially in the treatment of severe staphylococcal infections

such as septicaemia. Vancomycin (31) has also been shown to be an effective

antibiotic in many cases of p-lactam resistance.S3 A tricyclic heptapeptide,

vancomycin (31) contains two B-hydroxytyrosine (26) residues (shown bold),

one with the (253R)-stereochemistry, and the other with the (2R3R)-

stereochemistry.

Although vancomycin-resistant bacterial strains are rare, they aÍe

becoming more common,53 and attention has been focussed on finding even

more potent antibiotics. Currently other cyclic peptide antibiotics are most

promising, with lysobactin (32) being found to be four times as potent as

vancomycin (31) and effective over a much wider range of bacterial

infections.54 Lysobactin (32) was isolated in 1988 from a Lysobacter strain,

and contains five p-hydroxyamino acid residues, three of which are residues

of uncommon hydroxyamino acids; namely B-hydroxyphenylalanine (25),

(30)

CIsugar\o CI

Introduction 11

Ho

+NH2Me

o

oo H

N NHNH

CO; o HzN

o

OH

(31)

p-hydroxyleucine (33) and B-hydroxyasparagine QÐ (shown bold). It isinteresting to note that the three uncommon B-hydroxyamino acids present in

lysobac tin (32) all po s ses s the ( 2 S ) + hr e o - stet eochemi s try.

HHzN

oo

o

o

NH

o

OHHN

HN

NH

HN

HN

oHN

NH

NH

HzN

oo

NH

H

(32)

o

HNL NH2

Introduction 12

+ +H3Nr,,...

"?oH "?oH

NHz

(34)

Another clinically important cyclic peptide is cyclosporin A, an

immunosuppressant undecapeptide which plays a very critical role in many

chemotherapeutic drug treatments and bone marrow transplants.5 5

Cyclosporin A contains an unusual B-hydroxy-cr-amino acid residue, MeBmt

(35), which has been shown to be essential to the action of the drug.s6 Various

other cyclic peptide drugs also contain hydroxyamino acid residues.

BouvardinsT contains a B-hydroxytyrosine (26) residue with the (2535)-

stereochemistry, echinocandins C and D58 each contain (253R)-

B-hydroxyhomotyrosine (36) and (254R)-4-hydroxyproline (24) residues, and

phomopsin 459'60 contains a (253R)-B-hydroxyisoleucine (37) residue and a

B-hydroxyphenylalanine (25) residue with the (2S3S)-stereochemistry. The

majority of the hydroxyamino acids found in these cyclic peptides, with the

exceptions of MeBmt (35) and p-hydroxyhomotyrosine (36), are hydroxylated

derivatives of proteinogenic amino acids, and are presumably formed by

functionalization of the corresponding amino acid residues following their

incorporation into the peptides.a

Hydroxyamino acids are also present in toxic peptides,6l-65 ¿¡¡1i1u¡¡1e¡

peptides,tr and are themselves of interest as enzyme inhibitors.6T For example,

B-hydroxyphenylalanine (25) has been shown to inhibit Neisseria gonorrhoeae

bacterial strains68 and the lactose operon in Escherichia coli.69 Although rale,

lS)-y-hydroxyleucine (38) is a naturally occurring amino acid, being found in

the mushroom toxin phalloine,To and (S)-Þ-hydroxyvaline (39) has been found

H

(33)

Introduction 13

in the hydrolysate of a number of peptides, including the antibiotic

Berninamycin A.7l

MeNH ¡¡3N/2r,.. coi

"raoH1¡rN2ø,.

(37)

(3s)

*1tNlrr,,.

OH

(38) (3e)

A multitude of syntheses of rare and unnatural ø-amino acids has been

developed .2,14,72-74 The most useful syntheses are those which address the

stereochemistry of the amino acids, at both the cr-centre and at chiral centres on

the side chains. Preparation of chiral cr,-amino acids has been achieved using

methods such as the resolution of racemates, asymmetric synthesis, and

synthesis from optically active starting materials from the so-called "chiral

pool". Optically active (R)-Þ-hydroxyvaline Ø3) was first prepared by

reaction of benzylamine with p,B-dimethylglycidate (40) to give the alcohol

(41), followed by removal of the benzyl group and reprotection of the amine to

give the alcohol (42). Resolution of the racemic product with

(-)-a-methylbenzylamine followed by removal of the protecting group gave

(R)-p-hydroxyvaline (a3) (Scheme 8).7s Most current syntheses of optically

+

+

(36)

+ +H

Introduction 14

active hydroxyamino acid derivatives, however, utilize asymmetric synthesis or

chiral starting materials to avoid tedious resolution steps.

PhcH2NH o;benzylamine

(40)(41)

i) H} Pdlcii) NaOH, PhCOCI

+ coi PhCONHÐ (-)-PhCH(Me)NH2

iÐ H* lIl2o

(43) (42)

Scheme 8

Many asymmetric syntheses of B-hydroxyamino acids via condensation of

glycine equivalents with aldehydes have been reported.T6-88 One of the most

general and versatile methods for the synthesis of functionalized cr-amino acids

has been developed by Schollkopf and collaborators.S0-8s The general protocol

involves the coupling of two amino acids to give a piperazinedione, with

subsequent bis-lactim ether formation by treatment with trimethyloxonium

tetrafluoroborate. For example, the bis-lactim ether (45) was produced viø

condensation of (S)-O,O' -dimethyl-a-methyldopa (44) and ethyl glycinate,

followed by treatment with trimethyloxonium tetrafluoroborate. Subsequent

ffeatment with n-butyl lithium followed by succesive additions of acetone and

methyl iodide gave the substituted piperazinedione (46) which was converted in

several steps to (R)-B-hydroxyverline (43) with an enantiomeric cxcess (e.e.) of

907o (Scheme 9).82

Introduction 15

+H3N coi

\ llll¡'i) H2NCH2CO2EI

ü) Me3OBFa

(45)OMe

(M)Ð BuLi

ü) acetone

üi) MeI

\ llltr,'N OMe

(43)hydrolysis

(46)

Scheme 9

Other practical glycine equivalents for the asymmetric synthesis of

B-hydroxy-cr-amino acids have been developed by Seebach and co-workers86'87

and by Evans and Weber.88 Evans and Weber's procedure involves the

formation of a chiral oxazolidinone thiocyanate (47) from (S)-phenylalaninol.

Condensation of the tin enolate of the thiocyanate (47) with an aldehyde gives

the thionourethane (48), with cleavage of the chiral auxiliary giving the

oxazolidinone (49). Hydrolysis of the oxazolidinone (49) then gives the

corresponding B-hydroxy-o-amino acid (50) (Scheme 10). The unusual amino

acid present in cyclosporin A, MeBmt (35), was produced in 947o e.e., and

(253R)-þ-hydroxyleucine (33) was produced ín 987o e.e. vía this route.88

AIN

Ar

N

Introduction 16

ooN

oo

N

R

i) Sn(OTf)2

-__-->

NCS ü) RCHO

KOH / H2O

i) Mg(OMe)2/Ilv4rIOHiÐ H2o2

R

N

(47)

(s0)

(48)

o

H+

RN

(4e)

Scheme 10

However, these procedures are obviously limited to the synthesis of

B-substituted a-amino acid derivatives. Also, although the above examples give

products of high enantiomeric excess there is currently a strong demand,

particularly in the pharmaceutical industry, for compounds which are

enantiomerically pure" This arises from the fact that in most cases only one

enantiomer of an administered drug is responsible for the desired therapeutic

effects, while it is often found that the other enantiomer exhibits detrimental

side effects. Probably the most striking example is that of the drug

thalidomide, in which it was found that the required activity resided in the

(R)-enantiomer (51), whereas the (S)-enantiomer was responsible for the

disasterous teratogenic effects.39'90 Similarly, treatment of Wilson's disease

and bitiary crrhosis with (R/-penicillamine (52) is extremeiy effective,9t Oor 'n

contrast the (S)-antipode of penicillamine causes optic atrophy, which can lead

to blindnoss.92

o +

Introduction 17

SH

(50)

Ho

N

o

(s2)(51)

Various enantiospecific syntheses of functionalized amino acids have

therefore been developed by modification of chirally pure starting materials

from the so-called "chi¡al pool". Most of the compounds comprising the chiral

pool are isolated from biological sources; examples include the proteinogenic

c¡-amino acids, carbohydrates, terpenes and tartaric acid. Tanner and co-

workers93 have reported the synthesis of B-substituted aspaftic acid derivatives

from (+)- and (-)-tartaric acid. Rao and co-workers94 have utilized the

inherent chirality in D-glucose (53) to develop a synthesis of MeBmt (35).

Although the procedure is very long, requiring over 17 steps from D-glucose

(53), it represents a general method for the stereospecific synthesis of

p-hydroxy-c¿-amino acids (50) (Scheme 1l).

R

+++

-+

OH

(s3)

OH

(54)

Scheme 11

The amenability of halogenated amino acid derivatives to further

elaboration has been well documented. Examples include the elaboration of

iodoalanine derivatives to give 4-hydroxy-ø-amino acids2l and the synthesis of

Introduction 18

yhydroxylysine (13) as described u5ons.2e-31 The side-chain halogenation of

proteinogenic amino acid derivatives and subsequent manipulation of the

introduced functionality is therefore a powerful approach to the synthesis of

rare and unnatural amino acids. The scope of elaboration of the side-chain

brominated amino acid derivatives, prepared as described in Chapter 1, is the

subject of the studies described in Chapters 2, 3, 4 and 5 of the Results and

Discussion of this thesis. The importance of hydroxyamino acids both as

physiologically active compounds and in the stereocontrolled syntheses of other

biologically active molecules has been detailed above. The conversion of the

bromoamino acid derivatives described in Chapter I to the corresponding

hydroxyamino acid derivatives was therefore investigated. Mild, non-basic

conditions were considered to be required for these reactions to reduce the

possibility of racemization at the o-cenffe, and to avoid undesirable reactions of

the phthalimido group due to its base-sensitivity. The above requirements

preclude the use of sfrong nucleophiles such as hydroxide to effect substitution

of the bromide. However, silver salts have been shown to induce substitution of

alkyl halides under very mild conditions.9s-97 Silver ion induced reactions of

alkyl halides have been shown to occur with graded S¡1-S¡.,r2 character, with the

varying degree of character dependent on the solvent, silver salt and bromide

structure in each reaction.95'97 The conversion of bromoamino acid derivatives

to the corresponding hydroxyamino acid derivatives, with control of both

relative and absolute stereochemistry, was therefore investigated using silver

nitrate induced hydrolysis reactions, with particular attention focussed on

maximizing the stereoselectivity of the reactions. Methods for the deprotection

of the hydroxyamino acid derivatives to give the corresponding free amino

acids, via procedures which avoid racemization, were also studied. The results

of the above studies are discussed in Chapter 2 of this thesis.

Various syntheses of the natural product chloramphenicol (28) have been

reported.4S Many are based on the condensation of glycine equivalents with

Introduction 19

p-nitrobenzaldehyde, with subsequent resolution steps required. The synthetic

methods developed in Chapters 1 and 2 of the Results and Discussion of this

thesis indicated a potential new method for the short, stereoconfrolled synthesis

of chloramphenicol (28), starting from (R)-phenylalanine (55) or itsderivatives. The results of this investigation are described in Chapter 3 of the

Results and Discussion of this thesis.

coi

(ss)

The brominated amino acid derivatives described in Chapter 1 of this thesis

are also suitable for elaboration to a wide variety of functionalized amino acid

derivatives. The scope and limitations of such further elaborations of the

bromoamino acids were therefore investigated. Stereocontrolled cyclization

reactions of the bromoamino acid derivatives to give lactone and lactam

products suitable for elaboration to known antibiotics were studied, as well as

various methods for the synthesis of unstable Ê,T- and y,õ-dehydroamino acids,

which have been found to be potent enzyme inhibitors9S'99 and precursors to

other functionalized amino acid derivatives.2 The results of these investigations

are described in Chapter 4 of the Results and Discussion of this thesis.

The free radical side chain bromination of N-phthaloyl-a-amino acid

derivatives is limited to incorporation of the bromine atom only at the position

on the side chain which gives the most stable radical. Transfer of functionality

from one position on the side chain to other, non-activated positions was

therefore investigated. The aim of this work was to further demonstrate the

applications of side-chain bromination and subsequent elaboration of

+

Introduction 20

N-phthaloyl-cr,-amino acid derivatives to the stereocontrolled synthesis of

functionalized amino acids. The results of this work are also described in

Chapter 4 of this thesis.

The bromoamino acid derivatives described in Chapter 1 of the Results and

Discussion are also suitable for conversion to unnatural amino acid analogues,

such as isotopically-labelled amino acids. The utility of labelled amino acids in

the study of enzyme reactions includes the use of 13C-labelled substrates to

determine the stereochemistry of enzyme-catalyzed decarboxylation reactions,16

and the use of deuterium-labelled tyrosine in the determination of the

stereochemical course of tyramine oxidation by the enzyme semicarbazide-

sensitive amine o*i¿¿ss.l00 The mechanism of the enzyme phenylalanine

ammonia lyase (PAL) has also been determined with the aid of labelling

experimen1s.15,101-105 Numerous studies of PAL have been conducted,l06-110

particularly by Havir and Hanson.lOT-l10 PAL is a plant enzyme which

catalyses the elimination of ammonia and a proton from (S)-phenylalanine (56)

to give trqns-cinnamic acid (57) (Scheme 12)" It has been postulated that this

enzyme represents a switch in plants for re-routing (S)-phenylalanine (56) from

protein synthesis to secondary plant metabolites such as phenylpropanoids,

alkaloids and flavanoid compounds. 1@

+HrNrr,,,. coi

PAL NH3+

(56)

Scheme 12

(st)

Introduction 2l

The enzyme consists of four identical subunits, which each possess an

N-terminal serine residue. The enzyme tetramer contains two active sites,

indicating that two subunits combine to form each of the active sites. Labelling

studies have indicated that the N-terminal serine residues (58) are post-

ribosomally modified, with two undergoing a,B-elimination to give

dehydroalanine residues (60) and the other two most probably being oxidised to

give serine-aldehyde residues (59).101-l0s A dehydroalanine residue (60) then

reacts with a serine-aldehyde residue (59) to give a conjugated electrophilic

prosthetic group (61) in each of the active sites (Scheme 13). Upon binding of

(S)-phenylalanine (56) in an active site, the amino group of (S)-phenylalanine

(56) reacts with and becomes covalently bound to the electrophilic cenre (61).

Formation of the enamine (62) then establishes an elecfton sink for facile C-N

bond cleavage of the substrate, such that removal of a B-proton and subsequent

elimination occurs to give trans-cinnamic acid (57) and the amino-enzyme

complex (63). The enzyme then releases ammonia, and in doing so reforms the

activated electrophilic centre (61) in the active site (Scheme 14).

Enz

-H2 ¡f2Nh,,. -EnzEnz

(se) -Hzo

OH -Hzo

(s8) HzN -Enz

(61)

(60)

-Enz

Enz= enzyme

Scheme 13

(56)

(s7)

Introduction 22

Enzo

Enz

o

Enz

Enz

Enz

(61)

Enz

Ph

Ph

Ph

/a-HLB,

o(

N

HN=

NH¡

B lHEnz

Enz

(

HN

=

Enz

Enz

( HN

HzN

o

Enz

HN

HN

o)

Enz

(

OHEnz

( HN

Hcoz

H

(62)BI

Scheme 14

Enz

(63)

HN

Introduction 23

Battersby and co-workersl5 have reported a study of the stereochemical

course of the elimination catalyzed by PAL using the deuterium-labelled

phenylalanines (65a) and (65b). The synthesis of the labelled phenylalanine

derivatives (65a) and (65b) was performed by hydrogenation of the deuteriated

dehydrophenylalanine derivative (64), followed by removal of the protecting

group, to give a racemic mixture of the p-deuteriated phenylalanines (65a) and

(65c). The N-chloroacetyl derivatives of the racemic mixture of (65a) and

(65c) were resolved by treament with hog kidney-acylase-I to give (253R)-

3-deuteriophenylalanine (65a), and the N-chloroacetyl derivative of (2R35)-

3-deuteriophenylalanine (65c), which was subsequently hydrolysed to give

(2R35)-3-deuteriophenylalanine (65c). Racemization at the cr-centre of the

deuteriated phenylalanine (65c) by treatment with acetic anhydride lacetic acid

gave a mixture of the deuteriated phenylalanines (65b) and (65c) (Scheme

15¡.ts The deuteriated phenylalanines (65a), (65b) and (65c) produced viø this

procedure were isolated with enantiomeric excesses of ca. 867o and deuterium

incorporation of 9OVo"

DD

D N-benzoyl-glycine

Ph N

Ph

NaOH

cot

NHCOPh

(64)

i) H2lPd

ii) hydrolysis

DD

=

D

= Ac2O / cozPh

NH¡-1-NH.+)

(65a)

NH"^¡ '^ -J1

Ph

(6sb)

AcOH

Scheme 15

(65c)

+

Introduction 24

Kirby and Michaellll reported a very similar procedure at about the same

time, and Ife and Haslaml12 also reported a synthesis of (2S3R)- and (2535)'

3-deuteriophenylatanine (65a) and (65b), from (S)-c-deuteriobenzyl alcohol

(66) (Scheme 16). These and related synthesesl13,l14 are limited by their lack

of enantioselectivity, and require enzyme-catalyzed resolution steps to obtain

homochiral products.

NHAcD

ErO2C

i) HBrÆI2OiÐ crcH2cocl

-+

Phüi) Acylase I

AcNH

(66)

D

I

PCI 3

NHAc i) HBrÆI2OiÐ clcH2coclEtO2C

Ph C1Ph

üi) Acylase IAcNH

Ts = SO2C6Ha-p-Me

Scheme 16

Battersby and co-workersl5 used the deuterium labelled phenylalanines

(65) to determine that PAL catalyses the elimination of the pro-S hydrogen of

(S)-phenylalanine (56) (Figure 2), together with ammonia, to give trans-

cinnamic acid (57). Reaction of (253R)-3-deuteriophenylalanine (65a, 907o 2II)

with the enzyme gave deuteriated trans-cinnamic acid (67) with 88+47o

deuterium incorporation, indicating a highty stereoselective elimination of the

3-pro-S hydrogen from (S)-phenylalanine (56) (Scheme 17). Treatment of the

(2SiS)-isomer (65b, 907o 2H) with the enzyme gave cinnamic acid (57) with

8+27o do¡terium incorporation, supporting the above conclusion. Hanson and

D

?

Ph

(65a)

(65b)

D

=

D

Introduction 25

¡¡uu¡l10 showed that (R)-phenylalanine (55) is also metabolized by PAL, but at

a, rate one five thousandth that of lS)-phenylalanine (56). However,

(R)-phenylalanine (55) does act as a competitive inhibitor of the enzyme,

indicating that it binds effectively to the active site. Hanson and Havirl l0

suggested that when (R)-phenylalanine (55) is bound to the active site the

groups on the enzyme are poorly situated to play an effective chemical role,

however no stereochemical studies of the reaction of PAL with

(R)-phenylalanine (55) have been reported.

+g3Nzø,,.

PAL

D D

(65a) (6t¡

+

co2H

g3N/ø,,.

(6sb)

PAL

Scheme 17

+

HR

(56)

H

(s7)

¡¡3NZr,.. cot

Fig ure. 2. D e s ig natio n of theo3,í#;i lr.;"f il

of (S) - phen y I al ani ne ( 5 6 )

Introduction 26

The above syntheses of the deuteriated phenylalanines (65) are limited

because they are not stereospecific and/or require enzymic resolution steps to

obtain enantiomerically pure products. The conversion of

B-bromophenylalanine derivatives described in Chapter 1 of this thesis to all

four stereoisomers of B-deuteriated phenylalanine (65) was therefore studied.

The results of this investigation are described in Chapter 5 of the Results and

Discussion of this thesis.

The use of the deuteriated phenylalanines (65) in a study of the reactions of

both lS)- and (R)-phenylalanine, (56) and (55), with the enzyme PAL is

described in Chapter 6 of the Results and Discussion. Particular attention was

focussed on determining the stereochemical course of the reaction of the

B-deuteriated (R)-phenylalanines (65c) and (65d) with PAL, with the interest

being to determine how the enzyme metabolizes a compound with the opposite

stereochemistry to that of its normal subsffate.

Chapter I

RESULTS AND DISCUSSION: CHAPTER 1

Regiocontrolled Side Chain Bromination of N-Phthaloyl-Protected

s-Amino Acid Derivatives

The starting materials required for the study of side chain bromination of

N-phthaloylamino acid derivatives were synthesized as described below. The

N-phthaloyl-protected (S)-valine, (S)-leucine, (S)-phenylalanine and

(RS)-homophenylalanine derivatives (68)-(70) and (76) were synthesized

according to the procedure of Sheehan, Chapman and Roth,ll5 r¡¡þiçþ involved

heating equimolar mixtures of finely groung phthalic anhydride and the

corresponding amino acids at 145-150o for 30 minutes (Schemes 18 and 19).

Temperature control was important as it has been reported that racemization

can occur at temperatures above 180o.115 Although this procedure is suitable

for the preparation of many N-phthaloyl-a-amino acids, it is not applicable for

protection of amino acids with hydroxyl groups on the side chains.l16

Phthaloylation of (S)-tyrosine was therefore performed by addition of

(S)-tyrosine to a. boiling solution of phthalic anhydride in

N,N-dimethylformamide (DMF), with boiling continued for 10 minutes, giving

(S)-N-phthaloyltyrosine (71) tn 787o yield.

+¡I3Nzzr,.. phthatic

anhydride

CO; tnrhN¿¿,,,.MeOH

HCI

(68) R = CHMez

(69) R = CHzCHMez

(70) R: CHzPh

(71) R = CHzCoHq-p-OH

(72) R = CHMez

(73) R = CHzCHMez

(74) R = CHzPh

(75) R = CHzCoH¡-P-OH

..........-..-.-'-RRR

27

Scheme l8

+H¡N coi PhthN

Chapter I 28

PhthN CO2Me

phthalicanhydride

MeOH

HCI

(77)

Scheme 19

Optical rotations of the N-phthaloylamino acids (68)-(71) were consistent

with literature values,115,117 indicating that racemization at the q-centres had

not occurred. The N-phthaloylamino acids (68)-(71) and (76) were then

esterified by treatment with methanolic hydrogen chloride solution, which was

prepared by the slow addition of thionyl chloride to cooled, dry methanol

under anhydrous conditions (Schemes 18 and 19), to give the esters (72)-(75)

and (77) in good yields.

(S)-N-Phthaloylphenylalanine (70) was also protected as the corresponding

tert-butylamide (78). Initially synthesis of the amide (78) was attempted by

treatment of (S)-N-phthaloylphenytalanine (70) with thionyl chloride and

pyridine in refluxing carbon tetrachloride (CCl¿) to give the corresponding

acid chloride, followed by addition of tert-butylamine. However, this

procedure was shown to cause racemization at the o-centre. Analysis of the

product by high performance liquid chromatography (HPLC) using a column

with (S)-phenylglycine as the chiral stationary phase showed the presence of

two compounds, with retention times of 2l and 22 minutes. Subsequently the

amide (78) was synthesized under milder conditions in order to prevent

racemization. Accordingly, a solution of (S)-N-phthaloylphenylalanine (70) in

dichloromethane at 0o was treated with triethylamine then ethylchloroformate,

(76)

Chapter I 29

followed by the addition of tert-butylamine, giving the product (78) in 937o

yield (Scheme 20). Analysis of this material by HPLC using conditions

identical to those previously described showed the presence of only the

(S)-enantiomer (78), which eluted with a retention time of 2l minutes. The

(S)-enantiomer (78) was therefore shown to have a shorter retention time on

the column than the corresponding (R)-enantiomer, which is consistent with

previous results using columns with (S)-phenylglycine as the chiral stationary

phase.lta

nhthN¿r,,,. NHCMe3

Ð NEt,ü) EIOCOCI

iii) t-BuNH2

(70) (78)

Scheme 20

Attempted amidation of the tyrosine derivative (71), under conditions

identical to those described for the preparation of the phenylalaninamide (78),

proved difficult due to its limited solubility in the reaction solvent,

dichloromethane. The phenolic group was therefore acetylated by treatment of

the tyrosine derivative (71) in refluxing acetic anhydride to give (S)-O-acetyl-

N-phthaloyltyrosine (79) in 797o yield" This compound had much increased

solubility in dichloromethane, and the amidation reaction therefore proceeded

smoothly via treatment of the tyrosine derivative (79) with triethylamine,

ethylchloroformate and tert-butylamine, to give the fully protected tyrosine

derivative (80) in 937o yield (Scheme 2l).

PhthN co2Hhh,.

(71)

Ac2O

Chapter I 30

PhthN co2H

(7e)

Ð NEr3ü) ETOCOCIiii) r-BuNH2

PhthN CONHCMe3

(80)

Scheme 21

Reactions of the (S)-valine and (S)-phenylalanine ester derivatives (72) and

(74) with NBS in CCI¿ to give the corresponding B-bromoamino acid

derivatives (81) and (82) (Scheme 22) were conducted as reported by Easton,

Tan and Hay.az The B-bromovaline derivative (81) was isolated as a white

solid in 837o yieLd. The B-bromophenylalanine derivative (82) was obtained as

a 1:1 ratio of the diastereomers (82a) and (82b), as determined by lH n.m.r.

spectroscopy, which showed peaks at õ 3.55 and 3.82 colresponding to their

methyl ester resonances.

The reaction to give the bromide (81) presumably proceeds viø abstraction

of the hydrogen atom from the p-position of the valine derivative (12) by a

bromine atom to give the tertiary radical (83) and hydrogen bromide.

Hydrogen bromide then reacts with NBS to give succinimide and molecular

bromine. Reaction of the radical (83) with bromine then gives the

B-bromoVåline derivative (81) and a bromine atom, which continues the radical

Chapter I 3I

.¡uin.119 Similarly, reaction of the phenylalanine derivative (74) would

proceed vía the benzylic radical (84) to give the p-bromide (82) as a mixture of

diastereomers. The lack of asymmetric induction in the formation of the

bromide (82) can be attributed to the exfremely low activation energy of such

halogen hansfer reactions.34

PhthN ththN¿¿r,,. CO2Me

NBS

Br

Q2) (8 1)

,hthN¿¿r,.. CO2Me tnrhN¿r,,,. CO2Me

NBS pl2

(74) (82)

(a) Rl=Br, R2=H(b) Rl=H, R2=Br

Scheme 22

PhthNnnr¡Nzrr,,. CO2Me

(83)(84)

The bromide diastereomers (82a) and (82b) were separated by tiactional

crystallization from hexane/dichloromethane, giving diastereomer (82a) in 427o

i"

Chapter I 32

yield (m.p. 142-143") and diastereomer (82b) in 407o yield (m.p. l2I-122").

The lH n.m.r. spectrum of diastereomer (82a) showed doublets at õ 5.52 and

6.02 due to the cr- and p-protons, respectively, as well as a singlet at ô 3.55 due

to the ester moiety. Diastereomer (82b) showed doublets at ô 5.59 and 5.91

due to the o- and p-protons, respectively, and a singlet at ô 3.82 due to the ester

moiety. The assignment of peaks due to the cr- and B-protons was based on

comparison of the lH n.m.r. spectral data of the bromides (82a) and (82b) with

that of the non-brominated compound (74). Substitution of a methylene proton

with bromine is reported to cause a downfield shift of the adjacent proton of

2.3 ppm.rz0 Hence, as the signals corresponding to the p-protons of the

phenylalanine derivative (74) occur at ca. õ 3.6, the peaks corresponding to the

P-protons of the bromides (82a) and (82b) would be expected to occur at ca.

õ 5.9. This is in close agreement with the observed values of õ 5.91 and 6.02.

The signals due to the a-protons of the bromides (82a) and (82b) therefore

occur at ô 5.52 and 5.59, respectively, being shifted downfield by ca.0.3-0.4

ppm from that of the non-brominated compound (74). These assignments were

confirmed using heterocorrelation n.m.r. spectroscopy" The 13C n.m.r.

spectrum of the bromide (82b) was obtained and the peaks were assigned to

their corresponding carbon atoms.120 The signal corresponding to the

a-carbon occurs at õ 57.0, and the signal corresponding to the B-carbon occurs

at õ 47.6. In the heterocorrelation n.m.r. spectrum, a strong coupling is

observed between the cr,-carbon and the doublet at ô 5.59, indicating that the

resonance observed at ô 5.59 in the lH n.m.r. spectrum corresponds to the

a-proton. Similarly, there is a strong coupling between the p-carbon and the

doublet at ô 5.91, indicating that the resonance observed at õ 5.91 in the

lH n.m.r. spectrum corresponds to the B-proton.

It was of interest to determine whether bromination also occurs with thc

amide-protected phenylalanine derivative (78). Treatment of the

phenylal4ninamide (78) with NBS under similar conditions as for bromination

Chapter I 33

of the ester (74), but using a3:1 mixture of CCI+ and dichloromethane to

achieve complete dissolution of the amide (78), did indeed give the

p-bromophenylalanine derivative (85) (Scheme 23), as a 1:1 ratio of the

diastereomers (85a) and (85b). To verify that no racemization at the cr-centre

had occurred during the bromination reaction, the diastereomeric mixture of

the bromides (85a) and (85b) was reduced to the phenylalaninamide derivative

(78), by treatment with tributyltin hydride (Scheme 23), and the product was

analysed by TIPLC as described above. Only the (S)-enantiomer (78) was

eluted, with a retention time of 21 minutes, indicating that both bromination

and reduction occurred withoutracemization at the g-centre.

ththN¿¿r,,. CONHCMe3 PhthN

NBS

p1Bu3SnH 2

(78) (8s)

(a) Rl=Br, R2=H(b) Rl=H, R2=Br

Scheme 23

The bromoamide diastereomers (85a) and (85b) were separated by

fractional recrystallization from hexane/iso-propanol, giving diastereomer

(85a) ín 4IVo yield (m.p. 188-191') and diastereomer (85b) in 43Vo yield (m.p.

208-209")" Both of the bromides (85a) and (85b) were identified using mass

spectrometry, each giving molecular ions at mlz 428 and 430 in equal

abundance, and using lH n.m.r. spectroscopy. Diastereomer (85a) showed

doublet resonances at õ 5.22 and 6.17, due to the c¿- and B-protons,

respectively. Diastereomer (85b) showed doublet resonances at õ 5.32 and

6.05, also due to the cr- and p-protons, respectively.i'

Chapter I 34

The relative stereochemistry of the B-bromophenylalanine ester and amide

derivatives (82a), (82b), (85a) and (85b) was determined by converting them to

the corresponding dehydrophenylalanine derivatives (86)-(89). Potassium

fluoride was chosen as the appropriate base for these ¡e¿ç¡iq¡s.121 Treatment

of the bromoamide (85a) with potassium fluoride and 18-crown-6 in

acetonitrile produced the (Z)-dehydrophenylalanine derivative (86) in 84Vo

yield, with none of the (E)-dehydrophenylalanine derivative (87) present.

Treatment of the other bromide diastereomer (85b) under the same conditions

gave the (E)-dehydrophenylalanine derivative (87) in 57Vo yield, and a small

amount of the (Z)-isomer (86) (97o) (Scheme 24). The stereochemistry of each

of the dehydrophenylalanine derivatives (86) and (87) was determined by X-ray

crystallographic analysis (Appendices 3 and 4). lH N.m.r. spectroscopy

showed the signal corresponding to the vinylic proton of the

(E)-dehydrophenylalanine derivative (87) at ô 7.05, whereas that of the

(Z)-dehydrophenylalanine derivative (86) occurred downfield at õ 7.59.

Consistent with an earlier repoÍ,rz2 this observation was the first in the present

tnrhN¿¿r,,. CONHCMe3 PhthN

.,?8, K+F-

18-crown-6

(85a) (86)

tn,hN¿rr,,. CONHCMe3 PhthN CONHCMe3

K+F-

18-crown-6

(8sb)

Br

Scheme 24

(87)

+ (86)

Chapter I 35

work of a trend in which the signals corresponding to the vinylic protons of

various (Z)-dehydrophenylalanine derivatives consistently occurred downfield

of the signals attributed to the vinylic protons of the corresponding

(E)-isomers.

The elimination of hydrogen bromide from the bromides (85a) and (85b)

is stereoselective and is therefore most probably proceeding via an anti-

elimination mechanism as the major pathway. On this basis the bromide (85a)

which gave only the (Z)-dehydrophenylalanine derivative (86) on elimination

of hydrogen bromide must have the (253R)-configuration, and the bromide

(85b) which gave the (E)-dehydrophenylalanine derivative (87) as the major

product must have the ( 2 S 3 S )- stereochemistry.

Treatment of the bromoester (82a) with potassium fluoride and

18-crown-6 in acetonitrile under similar conditions to those described above

produced the (Z)-dehydrophenylalanine derivative (88) in 88Vo yield, with none

of the (E)-dehydrophenylalanine derivative (89) present. The stereochemistry

of the (Z)-isomer (88) was confirmed by X-ray crystallographic analysis

(Appendix 5). Treatment of the bromide diastereomer (82b) under the same

conditions gave a 1:2 mixture of the (E)-dehydrophenylalanine derivative (89)

and the (Z)-dehydrophenylalanine derivative (88), in 88Vo overall yield

(Scheme 25). The isomers (88) and (89) were inseparable by chromatography,

but were partially separated by fractional crystallization from hexane/ethyl

acetate to give a pure sample of the (Z)-dehydrophenylalanine derivative (88).

The (E)-isomer (89) was always contaminated with residual amounts of the

(Z)-isomer (88), and consequently a crystal structure of the (E)-isomer (89)

could not be obtained. The stereochemistry of the (E)-isomer (89) was

confirmed by comparison of its lH n.m.r. spectrum with that of the

corresponding (Z)-isomer (88). The signal due to the vinylic proton of the

(E,)-isomer (89) occurred at õ 7.23, whereas the vinylic proton signal of the

Chapter I 36

(Z)-isomer (88) occurred downfield at ô 8.12, consistent with the trend

described above.

PhthN CO2Me

K+F-

18-crown-6

(82a) (88)

tnthN¿rr,,. PhthN

K+F- + (88)Br 18-crown-6

(82b) (8e)

Scheme 25

To determine whether the (Z)-dehydrophenylalanine derivatives (86) and

(88), formed during reactions of the corresponding erythro-bromides (85b)

and (82b) with potassium fluoride, were produced viø isomerization of the

corresponding (E)-isomers (87) and (89), the (E)-isomers (87) and (89) were

re-subjected to the original reaction conditions. No conversion of the

(E)-dehydrophenylalanine derivatives (87) and (89) to the corresponding

(Z)-isomers (86) and (88) was detected in the reaction mixtures by 1¡¡ n.m.r.

spectroscopy. Formation of the (Z)-isomers (86) and (88) is therefore

attributed to removal of the acidic cr-hydrogens from the bromides (85b) and

(82b), respectively, by fluoride ion, to give the conesponding stabilized

c-anions, followed by rotation about the respective Co-CÞ bonds, with

subsequent elimination of bromide ion. Formation of the (Z)-isomers (86) and

Chapter I 37

(88) is not unexpected, as the (E)-isomers of dehydrophenylalanine derivatives

are generally much less stable than the corresponding (Z)-isomers.l23,l24 A

number of isomerizations of (E)-dehydrophenylalanine derivatives to the

corresponding (Z)-dehydrophenylalanine derivatives have been reported; for

example, the (E)-dehydrophenylalanine derivative (90) isomeized to the the

correspondtng (Z)-isomer on heating in chloroform and, more rapidly, on

standing in neat trifluoroacaliç ¿gld.12s

CbzNH

(e0)

Evidently, elimination of hydrogen bromide from the erythro-bromides

(85b) and (82b) does not proceed exclusively via an E2 mechanism, but also vlø

an El.6 mechanism to some extent. The decreased selectivity in the elimination

of hydrogen bromide from the ester (82b) compared to the amide (85b) can be

attributed to the greater acidity of the a-hydrogen of the ester (82b), such that

the E1ç5 mechanism occurs to a greater extent. However, the major product

still arises from anti-elimination, and therefore the relative stereochemisfry of

the bromoesters (82a) and (82b) and the bromoamides (85a) and (85b) can still

be inferred. Thus, the bromide (82a) which gave the

(Z)-dehydrophenylalanine derivative (88) on elimination of hydrogen bromide

must have the (253R)-configuration, and the bromide (82b) which gave the

(E)-dehydrophenylalanine derivative (89) must have the (253 S )-

stereochemistry.

Reactions of the (S)-leucine and (RS)-homophenylalanine derivatives (73)

and (77) ¡vere also studied, with the aim of investigating the generality of the

Chapter I 38

side-chain bromination procedure. Treatment of the leucine derivative (73)

with one equivalent of NBS in refluxing CCla, with irradiation by a 250W

mercury lamp for 2 h, gave the y-bromoleucine derivative (91) (Scheme 26) in

8I7o yield after chromatography. Identification of the y-bromoleucine

derivative (91) was achieved using lH n.m.r. spectroscopy, which showed

singlet resonances at ô 1.76 and 1.84 corresponding to the diastereotopic

y-methyl groups, and by mass $pectrometry which gave molecular ions at mlz

353 and 355 ofequal abundance.

PhthN PhthN

NBSBr

(73) (e1)

PhthN PhthN

NBSBr

(77) (e2)

Scheme 26

Treatment of the homophenylalanine derivative (77) under identical

conditions gave the corresponding 7-bromohomophenylalanine derivative (92)

(Scheme 26) as an approximately l:1.5 ratio of diastereomers. The bromide

(92) was identified using mass spectrometry, which showed molecula¡ ions at

mlz 401 and 403 in equal abundance, and using lH n.m.r. spectroscopy. Two

resonances were observed at near identical chemical shifts at õ 4.91, due to the

Chapter I 39

y-protons of the diastereomers of the bromide (92), with each resonance

existing as a triplet due to the presence of two hydrogens on the adjacent

p-carbon. A riplet at ô 5.10 and a doublet of doublets at ô 5.21 were also

observed, attributable to the cr,-protons of the diastereomers of the bromide

(92). Assignment of the peaks due to the a- and y-protons was made by

comparison of the lH n.m.r. spectral data of the bromide (92) with that of the

non-brominated compound (77), as for the bromophenylalanine derivative (82)

described above. As the signal due to the y-protons of the homophenylalanine

derivative (77) is observed at ca. õ 2.6, the signal due to the y-protons of the

diastereomers of the bromide (92) would be expected to occur at cq. 6 4.9,120

indicating that the peaks observed at ô 4.91 are due to the y-protons of the

diastereomers of the bromide (92). The signals corresponding to the q,-protons

of the diastereomers of the bromide (92) therefore occur at õ 5.10 and 5.21,

being shifted downfield by ca" 0.2-0.3 ppm from that of the non-brominated

compound (77).

Attempted separation of the diastereomers of the bromohomophenylalanine

derivative (92) by fractional recrystallization and through chromatography on

silica was unsuccessful, with mixtures always being obtained.

Having developed procedures for the regioselective bromination of the

above N-phthaloylamino acid derivatives (72)-(74), (77) and (78), similar

bromination of the tyrosine derivatives (75) and (80) was investigated. The

challenge in radical bromination of tyrosine derivatives is to avoid oxidation of

the phenolic group, which is a facile process.126 Initially, attempted

bromination of fS)-N-phthaloyltyrosine methyl ester (75) was unsuccessful,

with no identifiable products being formed. Presumably the phenolic group

was oxidized, giving a complex mixture of products. In order to overcome this

oxidation the nhenol was nrotected as the corresnondins acetate. Treatment of'--r --__ f _ --_ f __ - ----t - - o' "'

(S)-N-phthaloyltyrosine methyl ester (75) with acetic anhydride, triethylamine

and a catalytic amount of N/V-dimethylaminopyridine (DMAP) gave the acetate

Chapter I 40

(93) in 87Vo yield. Subsequent treatment of the protected tyrosine derivative

(93) with NBS under standard conditions for 2 h gave a near quantitative yield

of the B-bromotyrosine derivative (94) (Scheme 21) as a 1:1 ratio of

diastereomers.

tnrhN¿r¿,..

NBSp1

p2

(e4)

(a) Rl =Br, R2=H(b) Rl=H, R2=Br

Scheme 27

The bromide (94) was identified using mass spectrometry, which showed

molecular ions at mlz 445 and 447 in equal abundance. The bromide (94)

slowly decomposed on standing in air, or in solution, and was therefore

characterized using high resolution mass spectrometry, which gave an ion at

mlz 445.018 corresponding to the molecular ion with a calculated mass of

445.016, rather than by elemental analysis. The bromide (94) was also

characterized as the corresponding B-hydroxytyrosine derivative (I22a) as

described in Chapter 2 of the Results and Discussion of this thesis. Due to their

instability the diastereomers of the bromide (94) were not able to be separated

by fractional recystallization, but enrichment of diastereomer (94a) was

achieved by chromatography on silica, which enabled the assignment of the 1H

n.m.r. spectra of each of the diastereomers (94a) and (94b). Diastereomer

(94a) showed doublet resonances at õ 5.46 and 6.03, due to the s- ancl

B-protons, respectively. Diastereomer (94b) showed doublet resonances at

ô 5.56 and" 5.93, also due to the cr- and B-protons, respectively.

(e3)

Chnpter I 4I

Treatment of the tyrosinamide derivative (80) with one equivalent of NBS

under similar conditions to those described above for bromination of the

tyrosine derivative (93) gave a near quantitative yield of the

p-bromotyrosinamide derivative (95) (Scheme 28), again as a 1:1 ratio of

diastereomers. The bromide (95) was identified using mass spectrometry,

which showed molecular ions at mlz 486 and 488 in equal abundance. In a

similar manner to that for the bromoester (94), the bromoamide (95) was

characterized using high resolution mass spectrometry, giving an ion at mlz

486.080 corresponding to the molecular ion with a calculated mass of 486.079,

and also by conversion to the B-hydroxytyrosinamide derivative (124) as

described in Chapter 2 of the Results and Discussion of this thesis. Enrichment

of diastereomer (95a) was acheived by chromatography on silica, which

enabled the assignment of the lH n.m.r. spectra of each of the diastereomers

(95a) and (95b). Diastereomer (95a) showed doublet resonances at ô 5.17 and

6.22, due to the a- and B-protons, respectively" Diastereomer (95b) showed

doublet resonances at ô 5.30 and 6.08, also due to the cr- and B-protons,

respectively.

PhthNhtr,. CONHCMe3 tnthNr¿r,,. CMe3

NBSp1

p2

AcO(80) (e5)

(a) Rl=Br, R2=H(b) Rl=H, R2=Br

Scheme 28

Although the diastereomers of the tyrosyl bromides (94) and (95) were not

separated and their relative configurations determined independently,I

Chapter I 42

comparison of their lH n.m.r. spectral data with that of the diastereomers of

the phenylalanyl bromides (82) and (85) enabled the ascription of the lH n.m.r.

spectra of the threo-bromides (94a) and (95a) and of the corresponding

erythro-bromides (94b) and (95b) (Appendix l). It can be seen from the data

in Appendix I that the lH n.m.r. spectra of the bromotyrosine ester

diastereomers (94a) and (94b) correlate very closely with those of the

bromophenylalanine ester diastereomers (82a) and (82b). The tyrosyl bromide

(94a) which exhibits its methyl ester resonance at ô 3.56 is therefore assumed to

be the threo-isomer, and similarly the bromide (94b) with resonance at ô 3.81

is assumed to be the erythro-isomer. Similar comparison of the lH n.m.r.

spectra of the diastereomers of the bromoamide derivatives (95) and (85)

enabled stereochemical assignment of the spectra of the tyrosyl bromide

diastereomers (95a) and (95b)"

Indeed a general trend was seen in the lH n.m.r. spectra for atl the

B-bromo-phenylalanine and tyrosine derivatives (82), (85), (94) and (95), and

those described in latter chapters of this thesis. The signal corresponding to the

carboxylate protecting group, be it ester or amide, occurs at lower chemical

shift for the threo-diastereomer than for the corresponding erythro-

diastereomer. Also, the threo-bromides exhibit the B-proton signal at higher

chemical shift, the cr-proton signal at lower chemical shift, and a larger

coupling constant between the c¡- and p-protons, than the corresponding values

for the erythro-isomers. These effects may be explained by studying, as an

example, the most favoured conformations of the threo- and erythro-bromides

(82a) and (82b). The most favoured of the conformations of the bromides

(82a) and (82b) is presumably staggened with the a- and B-protons anti to each

other (Figure 3). This minimizes the number of gauche interactions between

the bulky groups. It can be seen that when the threo-isomer (82a) adopts this

conformation, the phenyl group is situated close to the ester group, and the

shielding effect of the phenyl group may explain the lower chemical shift of the

Chapter I 43

ester group of the threo-isomer (82a) compared to that of the erythro-tsomer

(82b). Also, in the case of the erythro-bromide (82b), æ,æ-stacking may occur

between the aromatic rings of the phthalimido and phenyl groups. This would

cause rotation of the Co-CÞ bond such that the dihedral angle between the a-

and B-protons was less than 180o, thereby explaining the lower coupling

constant observed between the a- and B-protons for the erythro-diastereomer

(S2b) than for the threo-diastereomer (82a) (Figure 3).120

Ph Br

PhthN PhthN

HH

Ph

H

threo-isomer

(82a)

erythro-ísomer

(82b)

Figure 3. Newman projection of the preferred conformation of the

B-bromophenylalanine derivatives (82a) and (82b).

In summary, it has been shown that the protection of various cr-amino

acids as their N-phthaloyl derivatives enables the regioselective radical

bromination of the side chain of these protected cr-amino acids, rather than

functionalization at the o-centre (which occurs with amido- or carbamate-

protected cr,-amino acids). It is apparent, therefore, that the presence of the

phthaloyl protecting group strongly impedes the formation of the

corresponding cx,-centred radical. The destabilization of the cr-centred radical is

attributed to several steric and electronic factors. To obtain maximum

captodative stabilization of the radical (97), the molecule must lie in a planar

orientation such thai maximum orbital overlap between ihe imide, single

electron, and ester orbitals occurs. In this orientation it is clearly seen that

severe steric interactions exist between the phthalimido and ester carbonyl

Chapter I 44

H oI

N NOMe Me

I I Ro R

(e6)

groups, compared to the negligible steric interactions that exist in the benzoyl-

protected amino acid radical (96). The phthalimido group would therefore

twist out of the plane of the sp2-hybridized a-carbon, and would not participate

in stabilization of the radical. Also, the nitrogen electron pair of the imide (97)

is substantially more delocalized than that of the amide (96). The nitrogen

electron pair of the imide (97) is therefore less available for dative radical

stabilization. Thus, abstraction of the cr-hydrogen does not normally occur in

N-phthaloyl-protected ø-amino acids, instead abstraction occurs from the most

reactive position on the side chain if the resultant radical is appropriately

stabilized" Therefore the side chain bromination of various amino acid

derivatives can be achieved, and with retention of the inherent chirality at the

cr-position.

oX

(e7)

Chapter 2 45

RESULTS AND DISCUSSION: CHAPTER 2

Stereocontrolled Synthesis of Homochiral Hydroxy-cr-Amino Acid

Derivatives

The brominated amino acid derivatives (81), (82), (85), (91), (92), (94)

and (95) described in Chapter 1 of the Results and Discussion of this thesis were

thought to be suitable precursors for the synthesis of hydroxy amino acids, the

utility of which has been described in the Introduction. Hence, the synthesis of

the p-hydroxyvaline derivative (100) from the p-bromovaline derivative (81)

was initially attempted. As this transformation involves a nucleophilic

substitution ata tertiary centre, it was presumed that a method which proceeds

via an S¡1 mechanism was required. Silver ion is a strong halophile and has

been shown to promote the substitution of tertiary alkyl halides,95-97 hence

silver nitrate was chosen as a suitable reagent to encourage substitution of the

bromide (81). Many studies of the mechanism of silver ion induced

substitutions of alkyl halides have shown that these reactions occur with graded

S¡1-S¡2 character,9s-97 but that substitution of tertiary alkyl halides can

normally be assumed to proceed via carbocation-type intermediates.95

Kornblum and Hardies9T have reported that substitution of

(S)-a-phenylethylchloride (98) occurs with varying amounts of inversion and

retention of configuration, depending on the solvent and silver salt used in the

reaction. Rearrangement of benzylic halides upon treatment with silver nitrate

provides evidence for carbocation intermediates in these reactions,95 whereas

reaction of (S)-cr-phenylethylchloride (98) with silver nitrite to give

(R)-o-phenylnitroethane (99) with complete inversion of configuration in

acetonitrile (Sc.he.me. 29) indicate.s the importance, of the. rearwarcl displacement

process, or S¡2 nature, of these reactions.95 The low reactivity of neopentyl

halides with silver nitrate compared to the reaction of n-butyl halides also

Chapter 2 46

provides evidence for the S¡2 nature of silver nitrate induced substitutions of

alkyl ¡u1i¿s5.127

Ag+ Nor-

(ee)

.rrrtlCl

(e8)

Scheme 29

Treatment of the bromide (81) with 1.5 equivalents of silver nitrate, in a

1:1 mixture of acetone and water, gave the p-hydroxyvaline derivative (100) in

437o yiel{ as well as the p,y-dehydrovaline derivative (102) (347o) and a small

amount (87o) of the cr,p-dehydrovaline derivative (101) (Scheme 30). The

p-hydroxyvaline derivative (100) was identified using lH n.m.r. spectroscopy,

which showed singlet resonances at ô 1.31 and 1.53, corresponding to the two

B-methyl groups, a singlet at ô 4.91 attributable to the G-proton, and a broad

singlet at õ 4.4I due to the hydroxyl proton.

PhthN CO2Me

+

,^r^*r+CO2Me

Br

AgNO3

acetone / water

tnrhN¿¿¿,.. CO2Me

OH

(100)

(101)

(102)

OnrhN¿¿r,..

+(8 1)

Scheme 30

Chapter 2 41

Identification of the cr,p-dehydrovaline derivative (101) was also achieved

using lH n.m.r. spectroscopy, which showed the absence of any o-proton

resonances, with singlets at õ 1.88 and 2.43 conesponding to the vinylic methyl

groups. The p,1-dehydrovaline derivative (102) was identified by comparison

with the 1i¡"¡¿¡u¡s.128

Using lH n.m.r. spectroscopy with a chiral shift reagent, it was shown that

formation of the alcohol (100) occurred without racemization at the cr-position.

An authentic racemic sample of p-hydroxy-N-phthaloylvaline methyl ester was

prepared via an identical procedure as for the chiral compound (100), but

starting with (RS)-valine" The lH n.m.r. spectrum of a solution of the racemic

material and the chiral shift reagent Eu(hfc)3,rzg in deuteriochloroform,

resolved the enantiomers, showing four singlet resonances corresponding to the

two p-methyl groups of each of the two enantiomers" Two methyl ester

resonances were observed, as well as two hydroxyl proton peaks and two

o-proton peaks. The lH n.m.r. spectrum of a solution of Eu(hfc)3 and the

hydroxyvaline derivative (100), synthesized from (S)-valine, showed

resonances corresponding to only one enantiomer, indicating that the

hydroxyvaline derivative (100) was homochiral at the cr,-position" The

hydroxyl and a-proton peaks of the (S)-enantiomer (100) were observed

upfield of the conesponding peaks of the (R)-enantiomer, and the peaks

corresponding to the p-methyl groups of the (S)-isomer (100) resonated with a

smaller chemical shift difference than the corresponding peaks for the

(R)-isomer.

Presumably silver ion abstracts bromide from the valine derivative (81), to

give the carbocation (103), which is then either quenched by water to give the

alcohol (100), or an cr,- or y-proton is abstracted, leading to formation of the

cr,p-dehydrovaline derivative (101) and p,y-dehydrovaline derivative (102),

respectively (Scheme 31). The greater yield of the p,1-dehydrovaline

derivative (I02) compared to the o,p-dehydrovaline derivative (101) can be

Chapter 2 48

atfibuted to the presence of six y-protons compared to only one o-proton in the

carbocation (103), such that formation of the B,y-dehydrovaline derivative

(102) is favoured statistically even though the cr-proton is more acidic"

(100)

(81)

+Ag* AgBr (101) + (102)

(103)

Scheme 31

Initial treatment of the y-bromoleucine derivative (91) with silver nitrate

in acetonelwater on a small scale gave the corresponding y-hydroxyleucine

derivative (104) in 787o yield, a small amount of the y-hydroxyleucine lactone

(105) (87o), and a trace amount of a mixture of the p,y-dehydroleucine

derivative (107) and the y,ô-dehydroleucine derivative (106) (Scheme 32). The

lH n.m.r. spectrum of the hydroxyleucine derivative (104) showed singlet

resonances at õ I.24 and 1.31, corresponding to the two T-methyl groupS, â

singlet at õ 3.73 due to the methyl ester moiety, and a broad singlet at õ 1.70,

due to the hydroxyl proton. An ABX pattern consisting of doublet of doublet

resonances at ô 2.38, 2.50 and 5.15 is consistent with the diastereotopic

p-protons and the cr,-proton" The mass specffum of the alcohol (104) showed

no molecular ion, but showed fragment ions at mlz 276 and 233, due to the loss

of methyl radical and acetone, respectively, which is consistent with the

proposed structure.

The lactone (105) was identified from its mass spectrum, which showed a

protonated molecular ion at mlz 260, and using lH n.m.r. spectroscopy, which

showed singlets at ô I .52 and I.64, attributable to the two methyl groups, and

HIHzo

H+

Chnpter 2 49

nhthN¿¿¿.,. CO2Me PhthNrr,,.

(106)

+

Ph

BrAgN03

acetone lwater

(104) (10s)

PhthN CO2Me PhthN CO2Me

+(e1)

+

(107)

Scheme 32

an ABX pattern consisting of doublets of doublets at E 2.46,2.60 and 5.25,

consistent with the diastereotopic p-protons and the G-proton. The melting

point of the lactone (105) was identical to that reported in the literature.l3O

The p,y-dehydroleucine derivative (107) and the 1,õ-dehydroleucine derivative

(106) were inseparable by chromatography on silica or on silver nitrate

impregnated silica, a method described for the separation of isomeric

alkenes.13r They were identified using mass spectrometry, which showed a

peak at mlz 273 corresponding to the molecular ions of each of the

dehydroleucine derivatives (106) and (107), and using lH n.m.r" spectroscopy,

with the p,y-dehydroleucine derivative (107) exhibiting a doublet resonance at

õ 5.66, conesponding to the cr-proton, coupled to only one p-proton. The

signal due to the vinylic p-proton was observed at õ 5.82 as a doublet of

multiplets, indicating that the B-proton was coupled to the ü-proton and also vi¿

long-range coupling to the vinylic methyl groups. The y,ô-dehydroleucine

cferivative (106) exhibited singlets at ô 4.65 and 4.69, coresponding to the

terminal vinylic protons, with an ABX system comprising of doublets of

doublets at õ 5.10,3.10 and 2.86 corresponding to the cr-proton andÌ

Chapter 2 50

diastereotopic p-protons. Singlets were observed at õ 1.J6, L.77 and 1J8

which were attributed to the y-methyl group of the y,õ-dehydroleucine

derivative (106) and the y-methyl groups of the p,y-dehydroleucine derivative

(107).

By analogy with reaction of the bromovaline derivative (81),

hydroxylation of the bromoleucine derivative (91) is presumed to proceed viø

the tertiary carbocation (108), with subsequent attack by water leading to

formation of the hydroxyleucine derivative (104), or abstraction of either a õ-

or p-proton leading to formation of the dehydroleucine derivatives (106) or

(107), respectively. Formation of the lactone (105) presumably occurs via

cyclizatíon of the alcohol (104) with loss of methanol. Indeed, the alcohol

(104) slowly converted to the lactone (105) on standing in deuteriochloroform

for extended periods, and further reactions of the alcohol (104) described in

Chapter 4 of the Results and Discussion of this thesis also gave small amounts of

the lactone (105) as a byproduct, providing further evidence for the conversion

of the alcohol (104) to the lactone (105)" The lactone (105) was shown to be

homochiral by comparison with the literature,l30 ¿nd it was concluded that the

alcohol (104) must therefore also be homochiral.

The conversion of the bromide (91) to the alcohol (104) was shown to be

susceptible to a concentration effect. 'When the reaction was performed with

the bromide (91) concentration greater than 0.1M, the lactone (105) was

produced in greater than 607o yield, with only a trace of the alcohol (104).

However, it was found that if the reaction was performed under high dilution

conditions (<0.02M of the bromide (91)), the hydroxyleucine derivative (104)

was consistently isolated in greater thanTO%o yield.

It can be seen that in the reaction of the leucyl bromide (91) with silver

nitrate, the yield of the hydroxyleucine derivative (104) is much greater than

that of the elimination products (106) and (107). In contrast, reaction of the

valyl bromide (81) with silver nitrate occurs to give the substitution product

Chapter 2 5l

(100) and the elimination products (101) and (102) with approximately equal

propensity. This result can be attributed to the leucine y-carbocation (108)

being less hindered than the valine p-carbocation (103), due to the fact that the

y-position is further from the bulky phthalimido and ester moieties. That is, in

the synthesis of the hydroxyvaline derivative (100), steric hindrance at the

p-position of the valine carbocation (103) disfavours substitution, such that

elimination of a y-proton to give the p,y-dehydrovaline derivative (IOZ)

competes effectively with substitution. In contrast, as there is less steric

hindrance at the 1-position of the leucine carbocation (108), substitution to give

the hydroxyleucine derivative (104) is the major reaction pathway.

CO2Me

(108)

+

The method described above therefore represents an expeditious route for

the synthesis of homochiral p-hydroxyvaline and yhydroxyleucine derivatives,

with the chirality at the cr,-centre predetermined by that of the corresponding

chiral, commercially available amino acids.

Synthesis of the y-hydroxyhomophenylalanine derivative (109) was also

performed. Treatment of a 1.5:1 mixture of the diastereomers of the

bromohomophenylalanine derivative (92) with silver nitrate in acetone lwater

gave the y-hydroxyhomophenylalanine derivative (109) (Scheme 33) in 8O7o

yield after chromatography, as a 1.5:1 ratio of diastereomers. A mixture of the

lactones (110) and (111) was also obtained, which was separated by fractional

crystallization to give the cis-lactone (110) in37o yield and the trans-lactone

(111) in27o yield. The y-hydroxyhomophenylalanine derivative (109) was

'ir

Chapter 2 52

identified from its mass spectrum, which showed a molecular ion at mlz 339,

and using lH n.m.r. spectroscopy. The major diastereomer (109a) exhibited a

singlet at õ 3.70 corresponding to the methyl ester moiety, and a broad doublet

of doublets at ô 4.92 conesponding to the y-proton, coupled to the two

p-protons and broadened by coupling to the hydroxyl group. A doublet of

doublets at ô 5.08 was attributable to the G-proton, coupled to the adjacent

p-protons" The minor diastereomer (109b) exhibited a singlet at õ 3.71, a

broad doublet of doublets at ô 4.57, and a doublet of doublets at ô 5.25,

attributable to the methyl ester moiety, the 1-proton and the o-proton,

respectively.

PhthN

+

PhthN PhthN CO2Me

Br AgNo3

o

(110)

acetone/watero

PhthN

+(e2) (10e)

(111)

Scheme 33

The lactones (110) and (111) were identified from their mass spectra, with

the trans-lactone (111) giving a peak at mlz 307, corresponding to the

Chnpter 2 53

molecular ion, and the cis-lactone (110) giving a peak af mlz 308,

corresponding to the protonated molecular ion. The relative stereochemistry of

the lactones (110) and (111) was inferred on the basis of their lH n.m.r.

spectra. Altman et a1.132 have reported that in the lH n.m.r. spectra of related

2,4-disubstituted butyrolactones the chemical shift difference between the

diastereotopic geminal protons is greater for the cis-isomer than for the

corresponding trans-isomer. The lH n.m.r. spectrum of the lactone (111)

showed an ABMX splitting system, with resonances at õ 2.85 (J - 10.6, 12.0

and 12.5 Hz) and õ 2.93 (J - 6.2, 9.2 and 12.5 Hz) conesponding to the

geminal protons. The lactone (110) also showed an ABND( splitting system,

with resonances atõ2.65 (J =3.3,9.8 and 13.l IJz) and ô 3.12 (J = 8.7,9.4 and

13.1 Hz) again corresponding to the geminal protons. On this basis the lactones

(111) and (110) were assigned the trans- and cis-stereochemistry, respectively.

As the diastereomers of the alcohol (109), produced from a 1.5:1 mixture

of the diastereomers of the bromide (92),werc also formed in a 1.5:1 ratio, it

seems likely that substitution of the bromide (92) proceeds viø an S¡2

mechanism. However, as separation of the diastereomers of the bromide (92)

was unsuccessful, the mechanism of this reaction could not be determined

unambiguously. It may be that substitution of the bromide (92) occurs with

substantiat S¡1 character, and fortuitously provides the diastereomers of the

alcohol (109) in the same ratio as that of the sta¡ting bromide (92).

Presumably formation of the lactones (110) and (111) occurs vía

cyclization of the diastereomers of the alcohol (109), with elimination of

methanol, as for production of the leucine y-lactone (105).

It was perceived that the synthesis of p-hydroxyphenylalanine derivatives

could be performed via an analogous procedure to that used in the formation of

the hydroxyvaline, hydroxyleucine and hydroxyhomophenylalanine derivatives

(100), (104) and (109). However, as substitution at the p-carbon of

phenylal4nine introduces a chiral centre adjacent to the G-centre, some

Chapter 2 54

diastereoselectivity was anticipated if the substitution reactions were to proceed

via an S¡1 mechanism. As silver nitrate induced substitutions of alkyl halides

have been reported to proceed with graded SNl-SN2 character, substitution

reactions of the phenylalanine bromide (82) were initially performed on the

separated diastereomers (82a) and (82b) in order to minimize the

stereochemical complexity of the reactions.

Treatment of the threo-bromide (82a) with silver nitrate in acetone lwater

gave a 2:1 mixture of the threo- and erythro-þ-hydroxyphenylalanine

derivatives (112a) and (112b) (Scheme 34). The diastereomeric ratio was

determined by integration of the methyl ester signals of the alcohols (112a) and

(112b) in the 1H n.m.r. spectrum of the crude reaction mixture.

Recrystallization of the crude product gave the threo-þ-hydroxyphenylalanine

derivative (lI2a) in 617o yield, the stereochemistry of which was determined

unambiguously using X-ray crystallographic analysis (Appendix 6).

Purification of the erythro-isomer (112b) in the mother liquor from small

amounts of the threo-isomer (112a) and other impurities was achieved by

HPLC using a Crg reverse-phase column. Repeated elutions of the mother

liquor and collection of the appropriate components provided a small amount of

the pure erythro-isomer (112b) for charactenzation purposes.

PhthN¿¿r,,. CO2Me PhthN CO2Me PhrhNhr,,.

AgN03

Bracetone/water

H

+

(It2a) (1 12b)(82a)

Scheme 34

Chapter 2 55

The lH n.m.r. spectrum of the threo-þ-hydroxyphenylalanine derivative

(llza) showed a doublet resonance at õ 5.51, corresponding to the o-proton, a

doublet of doublets at õ 5.71, due to the B-proton, and a doublet at ô 5.13, due

to the hydroxyl proton. The cr- and p-protons were coupled by 4.6 Hz, and the

hydroxyl and p-protons were coupled by IO.4 Hz" Exchange with D2O

removed the peak corresponding to the hydroxyl proton, and simplified the

p-proton resonance to a doublet, being coupled only to the cr-proton. FAB

mass spectrometry of the alcohol (Il2a) showed a protonated molecular ion at

mlz 326. The lH n.m.r. spectrum of the erythro-isomer (112b) showed a

doublet at õ 4.95 corresponding to the cr-proton, a doublet of doublets at õ 5.46

corresponding to the p-proton, and a doublet at õ 4.27 due to the hydroxyl

proton. The a- and p-protons were coupled by 8.4 Hz, and the hydroxyl and

B-protons were coupled by 2.3 Hz.

The lH n.m.r. spectra of the threo- and erythro-isomers (ll2a) and (112b)

follow a general trend seen for all the p-hydroxyphenylalanine and

p-hydroxytyrosine derivatives described in this thesis, as seen in Appendix 2.

The chemical shifts of the o-, Þ-, and hydroxyl protons of the threo-isomets

are all significantly higher than those of the corresponding erythro-isomers.

Also, the threo-isomers each exhibit a smaller coupling constant between their

c¿- and p-protons, and a larger coupling constant between their p- and hydroxyl

protons, than is observed for the corresponding erythro-isomers.

PhthN o PhthN oHp

Ph

threo-ísomer

(rr2a)

Ph oOMe

HÞ

erythro-iwmer

(1 12b)

H

Figure 4. Hydrogen bonded conformations of the alcohols (112a) and (112b).

Chapter 2 56

The p-hydroxyphenylalanine derivatives (112a) and (112b) may adopt

stable hydrogen-bonded chair conformations as shown in Figure 4. The

presence of these conformations may explain the observed trends in the lH

n.m.r. spectra described above. The dihedral angle between the cr- and

p-protons of the threo-isomer (ll2a) in this conformation would be ca.60". In

contrast, the dihedral angle between the cr- and p-protons of the erythro-isomer

(112b) would be ca. 180", hence the coupling constant between the cr- and

p-protons of the threo-isomer (ll2a) would be smaller than that observed for

the corresponding erythro-isomer (112b). Similarly, the dihedral angle

between the p- and hydroxyl protons in the threo-isomer (ll2a) would be ca.

180o, whereas in the erythro-isomer (112b) it would be ca" 60", thereby

explaining the greater coupling constant between the p- and hydroxyl protons

of the threo-isomer (Il2a) than that of the erythro-isomer (112b)"

The major product of the hydroxylation of the threo-bromide (82a) is

produced with retention of chirality. This is consistent with abstraction of

bromide by silver ion to give the carbocation (113)" The carbocation (113)

may adopt a preferred conformation as shown in Figure 5. In this

conformation, steric interactions betwen the phenyl group and the phthalimido

Hzo

HPhthN

H Ph

OMe

(1 13)

Figure 5. The carbocation (113) in its preferred conformation.

and ester groups are minimized. Molecular modelling studies using MM2

calculations in the modelling program PCMODEL (Serena Software,

Chapter 2 57

Bloomington, Indiana, 1990) provide supporting evidence for this preferred

conformation of the carbocation (113). The ester moiety then blocks one face

of the carbocation (113), such that water attacks preferentially from the

opposite face, giving rise to the threo-isomer (ll2a) as the major product.

Treatment of the erythro-bromide (82b) under the same conditions as

described for the hydroxylation of the threo-bromide (82a) gave only the

threo-hydroxyphenylalanine derivative (ll2a), ín 93Vo yield (Scheme 35).

None of the erythro-isomer (112b) was observed.

"nrhN¿r¿,..Me PhthNfht,. CO2Me

AgNO3

Br acetone lwater

(82b) (rr2a)

Scheme 35

If hydroxylation of each of the bromides (82a) and (82b) proceeded vía the

carbocation (113) then identical ratios of the alcohols (llza) and (112b) would

be observed. Substitution of the bromides (82a) and (82b) must therefore

proceed via different mechanisms. The preferred conformation of the threo-

bromide (82a) is shown in Scheme 36" From this conformation, abstraction of

bromide by silver ion occurs easily, as the phenyl and p-hydrogen substituents

are already in the required orientation to give the most stable conformation of

the carbocation (113). There also exists the potential for stabilization of the

developing carbocation (113) by the ester carbonyl group, as the ester moiety is

situated in the plane of the developing unoccupied orbital. Also, SN2

substitution of the bromide (82a) is disfavoured as the ester moiety blocks

attack of water from behind the carbon-bromine bond. Hence, formation of the

"?ott

Chapter 2 58

carbocation (113) occurs with subsequent attack preferentially from the less

hindered face giving rise to the threo- and erythro-hydroxyphenylalanine

derivatives (112a) and (112b) in a2:I ratio.

PhthN PhthN PhthN

# H Ph-_+

CO2Me CO2Me CO2Me

threo-isomer

(82a)

OH

PhthN PhthN HPh

PhthN HH +.#

Br H Ph

CO2Me CO2Me

Br

^+

4goIIIIt

BrIIIt

HPhH

HH

PhH

(

2b

3)

(11

11

I+I2a)(1 )

Ph

H

H

erythro-isomer

(82b)

(IrZa)

ô*

The preferred conformation of the erythro-bromide (82b) is also shown in

Scheme 36. In this conformation, abstraction of bromide by silver ion to give

the carbocation (113) occurs less readily, as rotation of the Co-CÊ bond is

required to give the most stable conformation of the carbocation (113), and this

would ca-use steric crowding between the ester moiety and the dissociating

Cp2Me

Scheme 36

II

BrIItII

Ag

Chapter 2 59

bromine atom. In contrast, attack of water from behind the breaking carbon-

bromine bond is not hindered. It is therefore presumed that reaction of the

erythro-bromide (82b) occurs via an S¡2-type mechanism, where the silver ion

initially coordinates to the bromine, and assists in breaking the carbon-bromine

bond, as water attacks from the other side to give the t hr e o -

p-hydroxyphenylalanine derivative (112a) stereospecifically. These results are

consistent with many observations concerning substitutions of alkyl halides in

the presence of silver s¿l¡5.95-97 It is therefore not surprising that in the

reaction of the threo-bromide (82a) with silver nitrate, where S¡2 attack is

sterically disfavoured, substitution appears to proceed via the carbocation

intermediate (113), whereas in the reaction of the erythro-bromide (82b),

where S¡2 attack by water can occur, the product is formed with complete

inversion of configuration.

As hydroxylation of each of the bromides (82a) and (82b) gives rise to the

threo-hydroxyphenylalanine derivative (ll2a) as the major product, it was

decided to carry out this and further hydroxylation reactions using I :1

diastereomeric mixtures of the appropriate bromides, thereby avoiding their

separation. Treatment of a 1:1 mixture of the diastereomers of the

phenylalanyl bromide (82) under the standard conditions described above gave

the threo-p-hydroxyphenylalanine derivative (lI2a) and erythro-

B-hydroxyphenylalanine derivative (1 12b) in the expected 5:1 ratio.

In order to check the hypothesis for the observed diastereoselectivity in the

hydroxylation of the bromides (82a) and (82b) a more bulky ester group was

used, anticipating that it would increase the stereoselectivity of the reaction.

Hence the synthesis of the phenylalanine tert-butyl ester derivative (114) was

performed. Treatment of a solution of (S)-N-phthaloylphenylalanine (70) in

dichloromethane wíth tert-butanol, dicyclohexylcarbodiimide (DCC) and a

catalytic amount of 4-(N,N-dimethylamino)-pyridine (DURP¡t33 gave the

desired ester (114) in 757o yield. Bromination of the tert-butyl ester (114) was

Chapter 2 60

performed under identical conditions as described for that of the methyl ester

(74),and gave a 1:1 mixture of the bromide diastereomers (115a) and (115b)

(Scheme 37).

PhthN nhthN¿¿r,,. cMqr-BuOH NBSDCCDMAP

....'....+p1

(114)

Scheme 37

Separation of the bromides (115a) and (115b) was achieved by fractional

recrystallization from hexane/dichloromethane, giving diastereomer (115a) in

4l7o yield and diastereomer (115b) in 39Vo yield. Determination of the

configuration of the bromides (115a) and (115b) was accomplished by

comparison of their lH n.m.r. spectral data with that of the bromoesters (82a)

and (82b) and the bromoamides (85a) and (85b), as described in Chapter 1

(Appendix 1)" The lH n.m.r. specftum of the bromide (115a) showed a singlet

at ô 1.16, corresponding to the tert-butyl group, and doublets at ô 5.47 and

6.06, conesponding to the o- and p-protons, respectively, with a coupling

constant between those protons of 11 .4 Hz. The lH n.m.r. specÍum of the

bromide (115b) showed a singlet at õ 1.48, corresponding to the tert-bufyl

group, and doublets at õ 5.49 and 5.85, corresponding to the cr- and B-protons,

respectively, with a coupling constant between those protons of 10.4 Hz. The

bromide (115a) whose tert-butyl group signal occurred at ô 1.16 was therefore

assumed to be the threo-isomer, and the bromide (115b) whose tert-butyl group

signal occured at õ 1.48 was assumed to be the erythro-isomer, with the lH

n.m.r. speptral data following the trend described in Chapter 1.

(70) (1 1s)

(a) Rl =Br, R2=H(b) Rl=H, R2=Br

Chapter 2 61

PhthN CMe3 PhthN nnrhN¿r,,,.

AgN03

-+pl acetone/water

+

(1 1s)

(a) Rl=Br, R2=H(b) Rl =H, R2=Br

(1 16a) (1 16b)

Scheme 38

Hydroxylation of a 1:1 mixture of the bromide diastereomers (115a) and

(115b) was performed under standard conditions as previously described to

give amixture of the alcohols (116a) and (116b) (Scheme 38). Integration of

thetert-butyl ester signals of the alcohols (116a) and (116b) in the lH n.m.r.

spectrum of the crude reaction mixture indicated that the alcohols (116a) and

(116b) were produced in an 8:1 ratio. The threo-isomer (116a) was isolated in

SlVo yietd after recrystallization and was identified using lH n.m.r.

spectroscopy. A singlet was observed at ô 1.52 due to the tert-butyl group"

Doublet resonances at ô 5.25 and 5.44 were attributed to the hydroxyl and

q-protons, respectively. A doublet of doublets at ô 5.67 was attributed to the

p-proton, coupled to the ct,-proton by 4.6 Hz" and to the hydroxyl proton by

10.6 Hz. The chemical shifts and coupling constants of the c-, p- and hydroxyl

protons correlated closely to those of the threo-þ-hydroxyphenylalanine methyl

ester derivative (ll2a) (Appendix 2). Signals corresponding to Íhe erythro-

isomer (116b) were observed in the lH n.m.r. spectrum of the crude product.

Doublet resonances were observed at õ 4.95 and 5.49, corresponding to the cr-

and p-protons, respectively. The hydroxyl proton was observed as a broad

singlet at õ 4.60, and the tert-butyl ester signal was observed at õ 1.28.

It can be seen that there is an approximately two-fold increase in the

diastereoselectivity observed in the substitution of the tert-butyl ester bromide

Chapter 2 62

(115) compared to that observed in the substitution of the methyl ester bromide

(82). Presumably, by analogy with the reaction of the erythro-bromide methyl

ester (82b), the reaction of the erythro-bromide (115b) occurs with complete

inversion of stereochemistry. The reaction of the threo-bromide (115a) must

therefore occur to give an approximately 4:l ratío of the threo- and erythro-

alcohols (116a) and (116b), such that an 8:1 ratio of the alcohols (116a) and

(116b) is produced from a 1:1 mixture of the bromides (115a) and (115b)" The

increased selectivity observed for reaction of the terl-butyl ester derivative

(115) can be attributed to the larger tert-butyl group more effectively blocking

one face of the intermediate carbocation (117), than the methyl group in the

methyl ester,,carbocation (113), and provides further evidence for the

hypothesis described above explaining the preferential formation of the threo-

p-hydroxy-phenylalanine deriv atív e (l l2a).

H

(1 17)

In order to further examine the above hypothesis, hydroxylation of the

bromophenylalaninamide (85) was also performed (Scheme 39), under the

standard conditions. The threo-p-hydroxyphenylalaninamide (1 18a) was

isolated in excellent yield (937o) after recystallization, and was identified using

FAB mass spectrometry, which showed a protonated molecular ion at mlz 367,

and rH n.m.r. spectroscopy (see Appendix 2) as described below. No peaks

atftibutable to the erythro-isomer (118b) could be identified in the 300MHz

lH n.m.r. spectrum of the crude reaction mixture, indicating that the

hydroxylation reaction had proceeded to give the threo-isomer (118a) in

Scheme 39

> 95Vo diastereomeric excess. To calculate the diastereomeric excess more

accurately, the lH n.m.r. spectrum of the erythro-þ-hydroxyphenylalanine

derivative (118b) was required" The synthesis of the erythro-isomer (118b)

was proposed yi¿ racemizatton at the p-centre of the threo-isomer (118a). This

was achieved by oxidation of the alcohol (118a) to the ketone (119), with

subsequent reduction of the ketone (119) to give a mixture of the threo- and

erythro-atcohols (118a) and (118b) (Scheme 40).

ththN¿¿,,..

CrO" / H+(118a)

-'+

PhrhN CONHCMe3

plB2

(85)

(a) Rl=Br, R2=H(b) Rl=H, R2:Br

AgN03

acetone/water

Chapter 2 63

CONHCMe3

(1 18a)

+ (118a)

(1 18b)

CMe3

NaBHa

(1 1e)

Scheme 40

Treatment of the alcohol (118a) with Jones reagentl34 gave the ketone

(119) which was identified using mass spectroscopy, which showed a peak at

mlz 364 corresponding to the molecular ion, and using lH n.m.r" Spectroscopy.

A singlet was observed at õ 6.04, which was attributed to the cr-proton, with no

other siglals observed between õ 2-l " The crude ketone was reduced by

Chapter 2 64

treatment with a mixture of sodium borohydride in ethanol, with the reaction

being quenched by the addition of dilute hydrochloric acid after 10 minutes.

Care was taken to ensure that the reaction time was kept to a minimum, or the

yield of the alcohols (118a) and (118b) was greatly reduced, presumably due to

reduction of the phthalimido group.l3s The product was isolated in 847o yield

as an approximately 1.2:l ratio of the threo- and erythro-diastereomers (118a)

and (118b). It is possible that reduction of the ketone (119) with sodium

borohydride may be accompanied by racemization at the o-centre, due to the

acidic nature of the cr -hydrogen of the ketone (1 19), but as the

oxidation/reduction of the threo-a\cohol (118a) was only performed to obtain

the lH n.m.r" spectrum of the erythro-alcohol (118b) the chirality at the

cl,-centre is of no consequence. The lH n.m.r. spectrum of the threo-isomer

(118a) showed a singlet at ô 1.30, attributable to the tert-butyl group, doublets

at ô 4.41 and 5.11 due to the hydroxyl and a-protons, respectively, and a

doublet of doublets at õ 5.63 due to the p-proton. The lH n.m.r. spectrum of

the erythro-isomer (118b) exhibited a singlet at ô 1.15, due to the tert-butyl

group, characteristic doublets at õ 4.25 and 4.60, due to the hydroxyl and

G-protons, respectively, and a doublet of doublets at ô 5.39, due to the

B-proton. The stereochemistry of the alcohols (118a) and (118b) was

determined by correlation of their 1H n.m.r. spectra with other

p-hydroxyamino acid derivatives, as shown in Appendix 2, which indicated that

the product formed upon hydroxylation of the bromoamide (85) was the threo-

isomer (118a).

The lH n.m.r. spectrum of the crude product from reaction of the

bromoamide (85) with silver nitrate in acetone lwater is shown in Figure 6. No

peak corresponding to the tert-butyl group of the erythro-\somer (118b) can be

seen. Indeed, one of the two 13C satellites of the peak due to rhe tert-butyl

group of the threo-isomer (118a) can be seen at õ 1.08 and, as the natural

abundance of the 13C isotope is ca. l.l 7o, the area of this peak constitutes about

Chapter 2 65

0.5Vo of the area of the signal due to the tert-butyl group of the threo-isomer

(118a). If it is assumed that a peak being of one tenth the area of the l3C

satellite can be visualized, then it can be determined that the amount of the

erythro-isomer (118b) present in the crude mixture is less than 0.057o of that

of the threo-isomer (118a). Therefore the threo-alcohol (118a) is produced in

greater Íhan 99.9Vo diastereomeric excess (d.e.).

tert-butyl signal of (118a)

\

13C satellite

chemical shift oftert-búyl signal of (118b)

13C satellite

1.5 1.)PPM

Figure 6. lH n.m.r. of the crude product from substitution of the bromide (85)"

Within the limits of detection the hydroxylation of the bromoamide (85)

occurs diastereospecificalty, giving only the threo-þ-hydroxyphenylalanine

derivative (118a). Much greater selectivity is seen in the hydroxylation of the

bromoamide (85) than the bromoester (82), with the increase in selectivity

being at least 100-fold. This pronounced increase in selectivity may be due to

stabilization of the intermediate carbocation (120) by the amide carbonyl

Chapter 2 66

group. The carbonyl oxygen of an amido group is about 106 times more basic

than the carbonyl oxygen of an esls¡.136 Stabilization of the amide carbocation

(I2O) as shown in Figure 7 may therefore lock the carbocation (120) into a

specific conformation, such that water can only attack from one side to give the

thr eo -þ-hydroxyphenylalanine derivative ( I 1 8a) s tereospecifically.

PhthN

H

H

\ ô*NHCMe3

(120)

Figure 7. The carbocation (120) in the preferred conformation, withstabilization by the amido substituent.

It can be seen from the above examples that bromination of N-phthaloyl-

protected phenylalanine derivatives at the p-position and subsequent treatment

of the bromides with silver nitrate in acetone lwater provides a fast and efficient

method for the production of threo-þ-hydroxyphenylalanine derivatives with

control of stereochemistry at both the a- and p-centres.

To determine the generality of the method described above for the

synthesis of the p-hydroxyphenylalanine derivatives (ll2), (116) and (118),

hydroxylation reactions were also carried out with the p-bromotyrosine

derivatives (9a) and (95). Treatment of a 1:1 mixture of the diastereomers of

the bromotyrosine ester (94) under the standard hydroxylation conditions

proceeded to give the p-hydroxytyrosine derivative (121) (Scheme 4l) in 867o

yield, as a 6:1 ratio of diastereomers, consistent with the 5:1 ratio observed in

the hydroxylation of the phenylalanine derivative (82). The diastereomeric

ratio was determined using lH n.m.r. spectroscopy, as for the phenylalanine

derivatives (112a) and (1I2b), with the major alcohol (lzla) exhibiting peaks at

I

PhrhN

Blp2

(e4)

(a) Rl=Br, R2=H(b) Rl =H, R2=Br

AgN03

acetone lwater

Chapter 2 67

plB2

(r2r)

(a) Rl=OH, R2=H(b) Rl=H, R2=OH

Ph

MeOH /p-TsOH

Ph CO2Me PhthN CO2Me

+

(122a) (r22b)

Scheme 41

E 2.L9 and 3.78 due to the acetyl and methoxycarbonyl groups, repectively, and

the minor alcohol (121b) giving peaks at õ 2.17 and 3.14 due to the acetyl and

methoxycarbonyl groups, respectively. The FAB mass spectrum of a mixture

of the hydroxytyrosine derivatives (121a) and (121b) showed an ion at mlz 384,

corresponding to the protonated molecular ions. However, the hydroxytyrosine

derivatives (121a) and (121b) were unstable and deacetylated slowly on

standing in air or in solution. Hencg, the crude product was deacetylated by

treatment with a catalytic amount of p-toluenesulfonic acid in methanol

(Scheme 41) to give a 6:1 ratio of the hydroxytyrosine derivatives (122a) and

(122b)" The threo-isomer (L22a) was isolated in 727o yield after

recrystallization, and was identified using FAB mass spectrometry, which

showed a protonated molecular ion at mlz 342, and using lH n.m.r.

spectroscopy. Characteristic doublet resonances were observed at ô 5.02 and

HO

Chapter 2 68

5.44, due to the p-hydroxyl and a-protons, respectively, and a doublet of

doublets was observed at õ 5.64, due to the p-proton. The o- and p-protons

were coupled by 4.8 Hz, and the hydroxyl and p-protons were coupled by 10.3

tlz. Purification of the erythro-isomer (122b) was performed by HPLC using a

C1g reverse-phase column, as for the phenylalanine derivative (112b). The

lH n.m.r. spectrum of the erythro-isomer (122b) showed characteristic doublet

resonances at E 4.27 and 4.97, due to the p-hydroxyl and G-protons,

respectively, and a doublet of doublets at ô 5.49, due to the p-proton. The a-

and p-protons were coupled by 8.6 Hz, and the hydroxyl and p-protons were

coupled by 2.1Hz. The lH n.m.r. spectra of the p-hydroxytyrosine derivatives

(122a) and (122b) correlate very closely with those of the

p-hydroxyphenylalanine derivatives (1 l2a) and (112b), and were used to assign

the stereochemistry of the p-hydroxytyrosine derivatives (122a) and (I22b)

(Appendix 2).

p1 AgNO3

acetone/water

PhCMe3

",?oH

(es)

(a) Rl=Br, R2=H(b) Rl =H, R2=Br

(123)

MeOH /p-TsOH

CONHCMe3

",?oH

Scheme 42

(r24)

Chapter 2 69

Hydroxylation of the bromotyrosinamide (95) was also performed under

the standard conditions, giving the alcohol (123) (Scheme 42). Deacetylation of

the alcohol (123) gave the threo-þ-hydroxytyrosinamide (124) in 937o yield

after recrystallization. The hydroxytyrosine derivative (124) was identified

using FAB mass spectrometry, which showed a protonated molecular ion at mlz

383, and from its lH n.m.r. spectrum, which showed characteristic doublets at

õ4.12 and 5.11 due to the hydroxyl and cr-protons, respectively, and a doublet

of doublets at õ 5.62 due to the p-proton. The coupling constant between the p-

and hydroxyl protons was 7 .5 Hz, and the coupling constant between the cr- and

p-protons was 6.9 Hz. A singlet at õ 1 .27 was attributed to the tert-butyl

group. No peaks near ô 1"15 were observed in the 300MHz lH n.m.r.

spectrum of the crude product, which is where the peak due to the tert-butyl

group of the erythro-ísomer would be expected by comparison with the spectral

data of the phenylalanine derivative (118b). This indicates that the

hydroxylation reaction had proceeded in greater than 957o d.e., and it is

probable that the hydroxylation reaction occurs with greater than 99.97o

stereospecificity, as in the case of the formation of the hydroxyphenylalanine

derivative (118a).

The above procedure is therefore also applicable to the synthesis of threo-

B-hydroxytyrosine derivatives with known chirality at both the a- and

p-positions. As for the syntheses of the phenylalanine derivatives (112a),

(116a) and (118a), the diastereoselectivity observed in the reactions to give the

tyrosine derivatives (121a) and (L23) can be attributed to preferential attack

from one face of each of the corresponding intermediate carbocations. The

greater selectivity seen in the reaction to give the tyrosinamide (123) compared

to that observed in the synthesis of the tyrosine methyl ester (121a) is consistent

with the stereoselectivity observed in the reactions to give the corresponding

phenylalanine derivatives (ll2a) and (118a), and provides supporting evidence

for the effect of the amido substituent to increase the diastereoselectivity"

Chapter 2 lO

The above procedures for the synthesis of hydroxyamino acid derivatives

are of limited utility unless these derivatives can be converted to the

corresponding free hydroxy-a-amino acids without racemization at the chiral

centres. Hydrolysis of the hydroxyphenylalanine ester derivative (112a) and

the hydroxyphenylalanine amide derivative (1 18a) to (253R)-p-hydroxy-

phenylalanine (25a) was therefore investigated. Initially a mild two-step

procedure for the hydrolysis of the ester (lI2a) was examined. Removal of

phthaloyl protecting groups by treatment with hydrazine is an established

procedu¡s.137 Treatment of the hydroxy ester (llza) with a 2M solution of

hydrazine hydrate in ethanol to remove the phthaloyl group, followed by mild

acid hydrolysis of the ester in 1N HCI solution, gave the hydrochloride salt of

the amino acid (25a). Treatment of a solution of the hydrochloride salt in

ethanol with excess aniline resulted in crystallization of the free amino acid

(25a) in 8O7o yield (Scheme 43). A more vigorous but one-step procedure was

also performed, by treatment of the ester (ll2a) in a refluxing 2:1 mixture of

6N hydrochloric acid and glacial acetic acid. This gave the hydrochloride salt

of the amino acid (25a), which was treated with aniline in ethanol to give the

free amino acid (25a) in 937o yield (Scheme 43)" Identification of the free

amino acid (25a) was achieved by comparison of its lH n.m.r. spectrum with

that of a commercially available racemic sample, and by mass spectrometry,

which showed a protonated molecular ion at mlz 182. An optical rotation value

of -49.7" for the free amino acid (25a) obtained via the one-step hydrolysis was

determined in 6N HCI solution at 16o, which is consistent with literature

values,l38 indicating that the hydrolysis had occurred without racemization.

Hydrolysis of the amide (118a) was performed under conditions identical

to those described for the one-step hydroysis of the ester (112a), to give the

free amino acid (25a) in 78Vo yield (Scheme 44). The optical rotation value of

the free amino acid (25a) obtained from hydrolysis of the amide (118a) again

indicated that no racemízation had occurred.

Chapter 2 7l

PhthNr. either Ð NH2NH2iÐ lN HCriiÐ PhNHÆroH

CO;

or Ð 6N HCI /AcOHii) PtrNH2ÆtOH

(rr2a) (25a)

Scheme 43

Ð 6N HCI / AcOH(25a)

ü) PhNHr/EtOH

(1 18a)

Scheme 44

Hence, the side chain bromination of N-phthaloyl amino acid derivatives

followed by treatment of the bromides with silver nitrate and water provides an

efficient route to the synthesis of hydroxyamino acid derivatives, with control

of both relative and absolute stereochemistry, with subsequent deprotection

steps providing the corresponding homochiral hydroxy-substituted free amino

acids.

+

"%oll

Chapter 3 12

RESULTS AND DISCUSSION: CHAPTER 3

Towards a Synthesis of Chloramphenicol

The development of the methods for the synthesis of p-hydroxyamino acid

derivatives described in Chapters 1 and 2 of the Results and Discussion of this

thesis prompted an investigation of the synthesis of the alcohol (129) as a

potential precursor to the antibiotic chloramphenicol (28). It was envisaged

that the alcohol (I29) could be prepared from (R)-p-nitrophenylalanine (125) as

outlined in Scheme 45, using a route analogous to that described for the

synthesis of the hydroxyphenylalanine derivative (ll2a). This work was

conducted concurrently with that described in Chapter 2 of the Results and

Discussion of this thesis, and consequently the potential benefits of utilizing

amide rather than ester derivatives of phenylalanine in the synthesis of

p-hydroxyphenylalanine derivatives were not exploited. Although outside the

scope of the present work, the alcohol (129) can be expected to be a suitable

precursor for the synthesis of chloramphenicol (28). Indeed, Chenevert and

ThiboutotaT have elaborated a racemic sample of the amino alcohol (130) to

chloramphenicol (28).

Protection of the commercially available amino acid (125) was

accomplished via the same method as described for the protection of

(S)-phenylalanine. Treatment of a mixture of phthalic anhydride and

(R)-p-nitrophenylalanine (IZ5) at 140' for 30 minutes gave the N-phthaloyl-

protected amino acid (L26) in 817o yieId. The reaction time and temperature

were both kept to a minimum, as prolonged heating caused discolouration of

the mixture and reduced the yield of the product (126). Esterification was

perforrned by treatment of the protected amino acid (126) with a methanolic

solution of hydrogen chloride, giving the ester (121) in 860/o yield (Scheme 45).

Chapter 3 13

+H

PhrhN

(a) Rl =O) Rl=

PhthN

PhrhN

ozN

ozN

ozN

ozN

(r2s) (126)

p2

PhrhN

CI

HN

(t27)

(28)

(128)

Br, R2=HH, R2=Br

(r29)

--

ozN

ozN

Scheme 45

R

CI

ozN

HzN

(130)

CO2Me

Chapter 3 14

Bromination of the protected amino acid (127) was achieved by treatment

with a refluxing solution of NBS in CCI¿ for 4 hours, while the mixture was

irradiated with a 250W mercury lamp, and gave a 1:1 ratio of the bromide

diastereomers (128a) and (128b) in quantitative yield. The reaction time

required to effect bromination of the p-nitrophenylalanine derivative (I27) was

twice that required for bromination of the corresponding phenylalanine

derivative (74). This is presumably due to the strong electron withdrawing

effect of the p-nitro substituent, such that hydrogen atom abstraction from the

benzylic position by the electrophilic bromine radical, with consequent

development of a partial positive charge at that position, is disfavoured.

The bromides (128a) and (128b) were separated by fractional

recrystallization, giving diastereomer (128a) in 44Vo yield, and diastereomer

(128b) in 407o yield. Identification of the bromides (128a) and (128b) was

achieved using mass spectrometry, which showed molecular ions at rnlz 432 and

434 of equal abundance in each case. lH N.m.r. spectroscopy showed

characteristic doublets for diastereomer (128a) at õ 5.51 and 6.O2, due to the cr-

and p-protons, respectively, with a coupling constant between these protons of

II.2 llz" A singlet was observed at ô 3.59, due to the methyl ester moiety.

Diastereomer (128b) showed doublets at ô 5.59 and 5.9J, due to the cr- and

p-protons, respectively, with a coupling constant between these protons of 10.3

Hz. The signal due to the methyl ester group was observed at ô 3.83. By

analogy with the corresponding values obtained for bromides described in

Chapter 1 of the Results and Discussion of this thesis (Appendix 1) this data

indicates that the bromide (128a) has the threo-stereochemistry, and the

bromide (128b) has the erythro-stereochemistry. The stereochemical

assignments were confirmed by X-ray crystallographic analysis of the erythro-

bromide. (128b) (Appe.ndix 7).

Initially hydroxylation of a mixture of the bromides (128a) and (128b) was

attempte{ using the same conditions as for the preparation of the

Chaprer 3 75

p-hydroxyphenylalanine derivatives (llZa) and (112b). However, after stirring

a 1:1 mixture of the bromides (128a) and (128b) and silver nitratè in

acetone/water solvent for 16 hours at room temperature, no reaction had

occurred, and the bromides (128a) and (128b) were recovered. The lack of

reactivity of the bromides (128a) and (128b) under these conditions is

attributable to the electron withdrawing effect of the p-nitro group such that

formation of the benzylic carbocation (131) is disfavoured.

PhthN Me

+ H

ozN(131)

It was thought that more vigorous conditions were required to achieve

substitution of the bromides (128a) and (128b). Accordingly, the threo-

bromide (128a) was treated with silver nitrate in acetone/water at 50" for 2

days. However, under these conditions the (Z)-dehydrophenylalanine

derivative (132) was isolated in high yield, with none of the alcohol (I29)

formed (Scheme 46). Treatment of the erythro-bromide (128b) under

conditions as described above for the reaction of the threo-bromide (128a) gave

the threo-p-hydroxy-p-nitrophenylalanine derivative (L29) in 657o yield, with

I57o of the (Z)-dehydrophenylalanine derivative (132) and lOTo of the

(E)-dehydrophenylalanine derivative (1 33) (Scheme 47).

PhthN Me PhthN

AgNO3

acetone/water

500, 2d

Me

ozN

(128a)

Br

Scheme 46

ozN

(r32)

Chapter 3 76

PhthN PhthN CO2Me+ (r29)

AgNO3,a

Br acetone/water

500,2d+ (132)

ozN

(128b) (133)

Scheme 47

The alcohol (129) was purified by column chromatography on silica and

was identified using lH n.m.r. spectroscopy. Characteristic doublets were

observed at ô 5.34 and 5.53, due to the hydroxyl and G-protons, respectively.

A doublet of doublets was observed at ô 5.79, corresponding to the p-proton,

which was coupled by 10.0 Hz to the hydroxyl proton and by 4.4 Hz to the

o-proton. The lH n.m.r. spectrum of the alcohol (129) is very similar to that

of the threo-þ-hyd¡oxyphenylalanine derivative (llZa), and indicates that the

B-hydroxy-p-nitrophenylalanine derivative (I29) has the threo-stereochemistry

(Appendix 2). The dehydrophenylalanine derivatives (132) and (133) were

identified using mass spectrometry, with each giving a molecular ion at mlz

352, and from their lH n.m.r. spectra. The (Z)-isomer (132) showed a singlet

at ô 8.13 corresponding to the vinylic proton. The singlet signal due to the

vinylic proton of the (E)-isomer (133) was observed at õ 7.28, upfield of that

of the (Z)-isomer (132), consistent with the trend described in Chapter I of the

Results and Discussion of this thesis.

Presumably S¡1 substitution of the bromides (128a) and (128b) does not

occur as formation of the intermediate carbocation (131) is disfavoured due to

the electron withdrawing effect of the p-nitro group. The conformations of the

bromides (128a) and (128b) in which S¡2 substitution would be expected to

occur most readily are shown in Figure 8, such that attack from behind the

carbon-bromine bond is not hindered. These conformations, however, are also

the conformations required for E2 anti-elimination of hydrogen bromide from

Noz

Chapter 3 77

the p-nitrophenylalanine derivatives (128a) and (128b). Thus, in the reaction

of the threo-bromide (128a), elimination is the predominant pathway and the

(Z)-dehydro-p-nitrophenylalanine derivative (I32) is formed. In the reaction

of the erythro-bromide (128b), E2 elimination of hydrogen bromide is less

favourable as this produces the less stable (E)-dehydro-p-nitrophenylalanine

derivative (133).r23,r24 Substitution to give the alcohol (129) therefore

competes effectively with elimination of hydrogen bromide. The substitution

reaction of the erythro-bromide (128b) to give the alcohol (129) is analogous to

the formation of the threo-p-hydroxyphenylalanine derivative (112a) from the

ery thro -bromide (82b).

p-O2N-Ph H H Ph-p-N02

PhthN PhthN CO2Me

HH

BrBr

threo-isomer

(128a)

erythro-ísomet

(128b)

Figure 8. Conformations of the bromides (128a) and (128b) most susceptible toboth S¡¡2 andE2 pathwâys

Formation of the alcohol (129) was not always reproducible, the yield

ranging between I0 - 65Vo, with the dehydrophenylalanine derivatives (132)

and (133) making up the remainder of the products. Various conditions were

employed in an attempt to maximize the yield of the alcohol (129), by adjusting

the temperature, solvent ratio and amount of silver nitrate added, but optimum

conditions were not found. However, further work in this area would be

expected to provide a system which gives the alcohol (129) in a reasonable and

consistent yield.

Chapter 4 18

RESULTS AND DISCUSSION: CHAPTER 4

Scope and Limitations of the Elaboration of Bromo-cr-Amino Acid

Derivatives

Having developed procedures for the synthesis of p-hydroxyamino acids

with control of relative and absolute stereochemistry, the scope and limitations

of further elaboration of side chain brominated N-phthaloyl-cr,-amino acid

derivatives were investigated. It was conceived that the bromoamides (85a) and

(85b) could be converted to 3,4-disubstituted p-lactams. Formation of

B-lactams via cyclization of p-bromopropionamides has been reported,l39 as

has cyclization of other p-substituted amides, such as those derived from

p-hydroxy-cr-amino u.1¿5.140 Cyclízation via N-C4 bond formation was

expected to provide a method for the stereocontrolled synthesis of

3,4-disubstituted p-lactams, with the stereochemistry of the products

predetermined by that of the precursor cr,p-disubstituted amides.

Treatment of the bromide (85a) with a solution of potassium amide in

ammonia, prepared by the addition of potassium and a catalytic amount of

ferric nitrate to liquid ammonia,l4l gave the trans-p-lactam (134) in good

yield, with a small amount of the (Z)-dehydrophenylalanine derivative (86)

(Scheme 48). The mass spectrum of the p-lactam (134) showed a molecular ion

at mlz 348. The lH n.m.r. spectrum showed doublets at ô 4.96 and 5.01, due to

the C4 and C3 protons, respectively. The coupling constant between these

protons was 2.3 Hz, consistent with the 3,4-disubstituted p-lactam (134)

pos ses sing the t rans - stereochemistry. t+z

Presumably the bromide (85a) reacts initially by loss of the amide proton.

The product anion can then undergo an intramolecular S¡¡2 cyclization to form

the trans-þ-lactam (134) yia substitution at the benzylic position. Alternatively,

intermole ular proton transfer affords the corresponding cr-anion which reacts

Chapter 4 79

by elimination of bromide to form the (Z)-dehydrophenylalanine derivative

(86). The intermolecular nature of the proton transfer to give the a-anion was

determined by performing the above reaction with varying concentrations of

the bromide (85a). It was found that as the concentration of the bromide (85a)

decreased, the ratio of the p-lactam (134) to the (Z)-dehydrophenylalanine

derivative (86) increased. The rate of the proton transfer reaction is therefore

dependent on the concentration of the bromide (85a), indicating a bimolecular

reaction, whereas cyclization to give the p-lactam (134) is unimolecular with

the rate being independent of the concentration of the bromide (85a).

CMe3PhrhN

K+ NH2-(86)+

Br

(85a) (134)

Scheme 48

Treatment of the bromide (85b) under conditions identical to those used in

the preparation of the p-lactam (I34) from the bromide (85a) did not give the

cis-p-lactam (136), but instead gave the B-aminophenylalanine derivative (135)

in 93Vo yield (Scheme 49). The amine (135) was identified from its FAB mass

spectrum, which showed a protonated molecular ion at mlz 366, and from its

lH n.m.r. spectrum, which showed doublet resonances at õ 4.63 and 5.77, due

to the cr- and p-protons, respectively. The amine (135) is presumed to have the

threo-stereochemistry, assuming the reaction proceeds vi¿ an SN2 mechanism

based on analogy with the reactions of the erythro-bromides (82b) and (85b) to

give the correspondíng ihreo-p-hydroxyphenylalanine derivatives (i 12a) and

(118a), as described in Chapter 2 of the Results and Discussion of this thesis.

Chapter 4 80

ththN¿¿,... CMe3 CONHCMe3

either K+NH2-

or NH4OHrfrf'?l.tn

2

(8sb) (13s)

Scheme 49

The difference in the above reactions of the bromides (85a) and (85b) may

be explained by examining the preferred conformations of the bromides (85a)

and (85b) (Figure 9a)" It can be seen that the preferred conformation of the

threo-bromide (85a) has the amido group in the required orientation for

displacement of bromide, and hence ring closure to form the trans-p-lactam

(134) occurs readily. The erythro-bromide (85b), however, must undergo

rotation of the Co-CÞ bond from its preferred conformation for the molecule

to be aligned for cyclization. This results in severe steric repulsion between the

PhthN CONHCMe3 PhthN CONHCMe3

threo-isomer erythro-isomer

(85a) (8sb)

Figure 9a. Preferred conformations of the bromides (85a) and (85b).

H

PhrhN NHCMe3

Figure 9b. Conformation of the bromide (85b) required to form the

cis-p-lactam (136).

HHBrPhPhBr

H

Ph

H

Br

Chapter 4 81

(136)

phthalimido, phenyl and tert-búylamido groups (Figure 9b), such that

formation of the cis-p-lactam (136) is disfavoured. Substitution of the erythro-

bromide (S5b) at the benzylic position to give the threo-p-aminophenylalanine

derivative (135) is therefore the most favourable process.

The amine (135) was also produced by treatment of a solution of the

erythro-bromide (85b) in tetrahydrofuran (TFß) with concentrated aqueous

ammonia. Treatment of the threo-bromide (85a) under identical conditions

gave only the (Z)-dehydrophenylalanine derivative (86). The difference

between these reactions is presumably a result of the relative stabilities of the

(Z)- and (E)-dehydrophenylalanine derivatives (86) and (87). anti-Elimination

of hydrogen bromide from the threo-bromide (85a) gives the

(Z)-dehydrophenylalanine derivative (86), whereas anti-elimination from the

erythro-bromide (85b) is less favourable as the product would be the less stable

(E)-isomer (87). Substitution of Íhe erythro-bromide (85b) at the benzylic

position to give the threo-p-aminophenylalanine derivative (135) therefore

occurs preferentially.

Having produced the trans-p-lactam (134) from the bromoamide (85a) it

was of interest to ascertain whether synthesis of the corresponding p-lactone

(139a) could be achieved. p-Lactones have recently received much interest due

to their antibiotic propertiesr43-r4s and their potential as precursors to a wide

variety of substituted amino acids.22-24 Various syntheses of p-lactones from

p-bromoacids have been reported, with most involving treatment of the

bromoacids with mild bases or silver salts.146 It was envisaged that the

synthesis.of the phenylalanine B-lactones (139a) and (139b) could be achieved

Chnpter 4 82

viø intramolecular substitution of the p-bromo acids (l3la) and (137b). This

procedure is analogous to the reactions of the bromides (82a) and (82b) to give

the alcohols (ll2a) and (1lzb), except that it involves intramolecular

substitution by the carboxylic acid group rather than intermolecular substitution

by water. Bromination of (S)-N-phthaloylphenylalanine (70) without

protection of the carboxylic acid group was therefore investigated.

Accordingly, (S)-N-phthaloylphenylalanine (70) was treated with NBS

under the standard conditions as described for the preparation of the bromides

(82a) and (82b) from the ester (74), and gave the p-bromophenylalanine

derivative (131) in987o yield, as a 1:1 ratio of diastereomers (Scheme 50).

Attempts to separate the diastereomers (137a) and (137b) by fractional

crystallization from methanol/water resulted in decomposition of the bromide

(137a) to give p-phthalimidostyrene (138) (Scheme 51), which crystallized as

large yellow prisms, with the other bromide (137b) remaining in solution. The

lH n.m.r. spectrum of the bromide (137b) showed doublets at õ 5.64 and 5.86

attributable to the cr- and p-protons, respectively, while the corresponding

resonances of the bromide (137a) were observed at ô 5.55 and 5.97"

Correlation of the lH n.m.r. spectral data of the bromoacids (131a) and (137b)

with that of the corresponding bromoesters (82a) and (82b) (Appendix 1)

indicated that the bromoacid (137b) had the erythro-stereochemistry and that

the bromide (137a) had the threo-stereochemistry.

PhthN PhrhN lth,,

NBSB1

p2

(r3t)(a) Rl=Br, R2=H(b) Rl =H, R2=Br

(70)

Scheme 50

Chnpter 4 83

Presumably only the threo-bromide (I37a) undergoes a decarboxylative

anti-elimination, to give the trans-alkene (138) (Scheme 51), because the

prefened conformation of fhe threo-bromoacid (137a) has the carboxylic acid

group anti to the bromine (Figure 10), in the required alignment for

elimination to occur. Decarboxylative anti-elimination from the erythro-

bromide (137b) is less favourable as rotation from the preferred conformer is

required (Figure 10). The facile conversion of the bromide (137a) to the

alkene (138) is presumably facilitated by the formation of an extended

conjugated rc-system in the alkene (138). Due to the lability of the bromide

(137 a) further reactions were carried out with mixtures of the diastereomers

(r37a) and (137b).

Ph

PhthN PhthN co2H

HHBrPhBr

HH

threo-isomer

(137a)

erythro-isomer

(137b)

Figure 10. Preferred conformations of the bromoacids (137a) and (137b).

co2H PhthN

.rrtt

Br

CO2 + HBr

(r37a) (138)

Scheme 51

Treatment of a mixture of the bromides (131a) and (137b) with silver

carbonate. in acetone gave a mixture of products which was analyzed using

Chapter 4 84

lH n.m.r. spectroscopy and shown to consist of the cis-p-lactone (139b), the

trans-þ-Iactone (139a) and a small amount of the trans-alkene (138). The cis-

p-lactone (139b) exhibited doublets at ô 6.33 and 6.70, due to the o- and

p-protons, with a coupling constant of 9.2 H:z. The trans-p-lactone (139a)

exhibited doublet signals at õ 5.67 and 5.92, due to the a- and p-protons, with a

coupling constant between these protons of 4.5 H:z. Determination of the

stereochemisry of the lactones (139a) and (139b) was based on the values of

the coupling constants between the a- and p-pro¡sns.147,148 Periodic analysis of

the mixture using lH n.m.r. spectroscopy showed that the trans-p-lactone

(139a) was slowly decomposing with a concomitant increase in the amount of

the alkene (138).

PhthN

Ag2CO3

Br

(r37a)(139a)

PhthN

Ag2C03

Br

(137b) (13eb)

Scheme 52

Presumably the threo-bromide (I3la) cyclizes to give the trans-lactone

(139a) and the erythro-bromide (137b) cyclizes to give the cis-lactone (139b)

(Scheme 52). The trans-lactone (139a) then undergoes a facile retro-[2+2]-

cycloaddition to give the alkene (138) and carbon dioxide (Scheme 53). In

Ph

o

o

Chapter 4 85

contrast, steric repulsion between the phthalimido and phenyl groups in the cis-

lactone (139b) presumably causes the four-membered ring to twist out of

planarity, such that there is not sufficient orbital overlap to enable

cycloreversion of that compound.

Attempted separation of the mixture of the products (138), (139a) and

(139b) by chromatography on silica resulted in total decomposition of the

rrans-þ-lactone (139a), while the cis-p-lactone (139b) was isolateÀin16%o yield

and the alkene (138) was isolated in 67Vo yield. The low yield of the cis-

p-lactone (139b) suggests that this compound is slowly decomposing on silica,

but the absence of the cis-alkene (141), expected from a concerted process,

indicates that the decomposition is probably occurring via a stepwise acid-

catalyzed process, as shown in Scheme 53. Cleavage of the O-C4 bond gives

the benzylic cation (140) which then undergoes decarboxylation to give the

trans-alkene (138).

Ph,hNr,,..

- coz(138)o

-+

(139a)

oH+

#PhrhNrr,,.. - coz

-H+(138).#

o+ H

(140)(13eb)

Scheme 53

Chapter 4 86

PhthN

(141)

The potential for utilizing the y-functionality in the amino acid derivatives

(91) and (92) to introduce functionality at the p-position of homophenylalanine

derivatives and the p- and y-positions of leucine derivatives was also studied.

Radical bromination of alkyl bromides has been shown to occur

regioselectively, giving the corresponding vicinal dibromides in high yields.las

Incorporation of the second bromine atom occurs at the most substituted vicinal

position, in accordance with the factors governing radical stability. The

selctivity of these reactions has been attributed to a neighbouring group effect,

where formation of the radical on the vicinal carbon is stabilized by the

adjacent bromine substituent.

Bromination of the y-bromohomophenylalanine derivative (92) was

initially attempted, as vicinal bromination can only occur at the p-position. The

bromide (92) was initially treated with two equivalents of NBS and a catalytic

amount of benzoyl peroxide in refluxing CCla, with irradiation by a 250W

mercury lamp for 4 hours. However, no products were observed and the

starting material (92) was recovered. The reaction was then repeated at reflux

in chlorobenzene, such that the temperature of the reaction was increased.

After 4 hours, lH n.m.r. spectroscopy indicated that only the starting material

(92) was present. Increasing the reaction time to as long as 3 days gave no

reaction, with the starting material (92) being isolated in good yield. The lack

of reactivity of the bromide (92) is attributable to severe hindrance at the

B-position due to the presence of the bulky o-substituents.

The lack of reactivity of the homophenylalanine bromide (92) at the

p-position suggested that this procedure may be applicable to the selective

Chapter 4 87

functionalization of the leucine derivative (91) at a primary ô-position.

Accordingly, the bromide (92) was treated under the same conditions as

previously described for reaction of the bromohomophenylalanine derivative

(92) (Scheme 54). Analysis of the reaction mixture using mass spectrometry

indicated the presence of the dibromide (I42), with peaks at mlz 431, 433 and

435 in a l:2:l ratio, corresponding to the molecular ion, as well as peaks at mlz

372,374 and376 in a 1;2:I ratio, corresponding to the loss of CO2Me from the

molecular ion. The dibromide (142) was purified through repeated

chromatography on silica, and its structure was determined using lH n.m.r.

spectroscopy. Resonances characteristic of the methyl ester groups of the

diastereomers of the dibromide (142) were observed at E 3.15 and 3.76.

Overlapping resonances were observed between ô 5.16-5.30, corresponding to

the a-protons of the diastereomers of the dibromide (142)" Resonances

between õ2.43-3.02 are consistent with the p-protons, with singlet resonances

at ô 1.93 and 1.86 attributable to the õ-methyl groups. A multiplet between

õ 3.68-3.85 is attributable to the ô-methylene protons Integration of the

singlets attributable to the õ-methyl groups indicated that the diastereomers of

the bromide (142) were produced in a ca. l:1 ratio.

PhthN

NBSBr Br

(e1) (r42)

Scheme 54

The selective bromination of the bromoleucine derivative (91) at a primary

ô-position in preference to reaction at the secondary B-position is consistent

PhthN¿rr,,.

Br

Chapter 4 88

with the explanation of the lack of reactivity of the bromohomophenylalanine

derivative (92) described above.

Alternative methods for the introduction of functionality at the Ê- and

ô-positions of the bromoamino acid derivatives (91) and (92) were also studied.

Conversion of the bromides (91) and (92) to the corresponding dehydroamino

acid derivatives (106), (107) and (146) was envisaged as one such method. The

p,ydehydroamino acid derivatives (107) and (146) are vinylglycine analogues,

themselves receiving attention as synthetic targets due to their propensity for

enzyme inhibition.9,98, I 50- 1 52

Elimination reactions were initially performed on the bromo-

homophenylalanine derivative (92), as less side reactions were expected with

this compound than with the bromoleucine derivative (91), where the

possibility of both p,y- and y,ô-elimination exists. Accordingly, the

bromohomophenylalanine derivative (92) was treated with one equivalent of

1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) in THF" The hindered base DBU

has been reported to promote regioselective dehydrohalogenation of various

alkyl halides, such that the most substituted alkene is the major product.l53

However, none of the p,y-dehydrohomophenylalanine derivative (146) was

produced, instead the methanophenylalanine derivative (143) was isolated, in

72Vo yield (Scheme 55)" The cyclic product (143) was identified using

PhthNPhthN

DBU......................................-

Br

(e2)

Scheme 55

(143)

Chapter 4 89

lH n.m.r. spectroscopy, which showed an ABX splitting pattern consisting of

doublets of doublets at ô 2.2J, 2.43 and 3.38, corresponding to the

diastereotopic methylene protons and the benzylic proton, respectively. Only

one diastereomer of the cyclic product (143) was isolated, and X-ray

crystallographic analysis of this material (Appendix 8) indicated that the phenyl

and phthalimido groups were cis to each other" Presumably the base, DBU,

abstracts the a-proton from the bromide (92)" Cyclization of the resultant

cr-anion to give the cyclopropyl amino acid derivative (143) then occurs

diastereoselectively, although the basis for this selectivity is not obvious.

The reaction of the bromide (92) with DBU was also performed in the

presence of silver nitrate, anticipating that formation of the benzylic

carbocation through abstraction of bromide by silver ion may increase the

acidity of the p-protons" However, the cyclopropyl amino acid derivative (143)

was again isolated in good yield, with a tÍace of the 1-hydroxy-

homophenylalanine derivative (109) also being produced, presumably due to a

small amount of water present in the reaction mixture.

As elimination of hydrogen bromide from the bromophenylalanine

derivative (92) proceeded via a,y-elimination to give the cyclic product (143)

rather than via p,1-elimination, a different route to the formation of the

dehydroamino acid derivatives (106), (107) and (146) was formulated.

Elimination of acetates and xanthates under pyrolytic conditions has been

reported to proceed cleanly and in good yield.ls¿ Consequently the acetates

(lM) and (145) were synthesized from the corresponding hydroxyamino acid

derivatives (10a) and (109), the syntheses of which are described in Chapter 2

of the Results and Discussion of this thesis. Treatment of a solution of the

hydroxyleucine derivative (104) in dichloromethane with acetyl chloride,

pyridine and a catalytic amount of DMAP gave the acetate (lM) in 7O7o yield,

and the leucine lactone (105) in 167o yield (Scheme 56). The acetate (144) was

identified using FAB mass spectrometry, which showed a protonated molecular

Chapter 4 90

ion at mlz334, and from its lH n.m.r. spectrum, which was similar to that of

the alcohot (104) but showed an additional singlet resonance corresponding to

the acetyl group" The lactone (105) is presumably formed by cyclization of the

alcohol (104)"

PhthNhtq.

AcCI + (10s)pyr /DMAP

Scheme 56

Pyrolysis of the acetate (L44) was performed under reduced pressure (0.08

mm), with the acetate distilled (ca.250") through a pyrolysis tube packed with

quartz fragments and heated to ca.450". Purification of the crude product by

chromatography on silica gave a mixture of the y,õ-dehydroleucine derivative

(106) and the p,y-dehydroleucine derivative (107) in a 1.6:1ratio, in847o yield

(Scheme 57).

tnrhN¿¿,,..

45tr

(104)

(t44)

Me

(I44)

+---+

(106) (107)

Scheme 57

Although the more substituted p,y-dehydroleucine derivative (107) is

thermodynamically more stable, formation of the y,ö-dehydroleucine derivative

(106) is favoured statistically as there are six õ-protons compared to two

Chapter 4 9l

p-protons in the acetate (144). These factors are consistent with the

dehydroamino acid derivatives (106) and (107) being produced in the observed

1.6:1 ratio.

Synthesis of the homophenylalanine acetate (145) from the alcohol (109)

(Scheme 58) was carried out under conditions identical to those described for

acetylation of the hydroxyleucine derivative (104). The acetate (145) was

produced in74%o yield, with small amounts of the cis-p-lactone (1lO) (1Vo) and

the trans-p-lactone (111) (zEo) also isolated. The diastereomers of the aceta;te

(145) were sepÍuated by fractional crystallization and identified using lH n.m.r.

spectroscopy, with one diastereomer exhibiting a singlet resonance at õ 1.94

due to its acetyl group, and the other diastereomer giving rise to a singlet at

ô 2.09 for that substituent.

PhthN PhthN

AcCl Ac + (110)

+ (111)

pyrlDMAP

(10e) (r4s)

Scheme 58

Pyrolysis of the acetoxyhomophenylalanine derivative (145) was initially

performed as described for that of the acetoxyleucine derivative (144).

However, the reaction did not proceed to completion, with approximately 507o

of the acetate (145) recovered. Consequently, pyrolysis of the acetate (145)

was performed at the higher temperature of 520' (Scheme 59). lH N.m.r"

specfioscopy of the crude reaction mixture indicated the presence of the trans-

p,y-dehydrohomophenylalanine derivative (146). A doublet signal was

Chnpter 4 92

observed at ô 5.58, attributable to the o-proton. Another doublet was observed

at õ 6.68, attributable to the vinylic y-proton, with a coupling constant of 15.9

Hz characteristic of a trans-substituted alkene. A doublet of doublets was

observed at õ 6.74 corresponding to the vinylic p-proton, coupled to the

G-proton by 7.1 Hz and to the yproton by 15.9 }Iz. Further interpretation of

the lH n.m.r. spectrum of the crude reaction mixture indicated the formation of

the cr,p-dehydrohomophenylalanine derivative (147). A doublet was observed

atõ3.52, attributable to the T-protons of the alkene (147) coupled to the vinylic

p-proton. The signal due to the vinylic P-proton presumably occurs between õ

7.19-7.43, beneath the complex multiplet observed due to the phenyl groups of

the two alkenes (146) and (147). The a,p-dehydrohomophenylalanine

derivative (147) was presumed to have the (Z)-configuration, based on the

general relative stability of (E)- and (Z)-dehydroamino acid derivatives, as

described in Chapter 1 of the Results and Discussion of this thesis. Integration

of the peaks due to the methyl ester groups of the alkenes (146) and (147)

indicated that they were produced in a ca. l:1 ratio. Attempted separation of

these compounds by chromatography on silica and silver nitrate-impregnated

silica proved unsuccessful.

PhthN PhthN PhthN CO2Me

s?tr +

(146) (147)(14s)

Scheme 59

Chapter 4 93

Presumably pyrolysis of the acetate (145) proceeds initially to give the

trans-þ,y-dehydrohomophenylalanine derivative (146)" This alkene (146) then

undergoes partial thermal isomerization to give the more stable

cr,p-dehydrohomophenylalanine derivativ e Oa7 ) "

In conclusion, it can be seen that regiospecific bromination of the side

chains of N-phthaloyl-o-amino acid derivatives and subsequent elaboration of

this functionality via various methods has the potential to provide

stereocontrolled syntheses of a myriad of functionalized cr-amino acid

derivatives. For example, the erythro-bromoamide (85b) has been converted to

the trans-3,4-disubstituted p-lactam (I34) with control of relative and absolute

configuration. Synthesis of the corresponding cÍs-B-lactam (136) from the

threo-bromoamide (85a) was not possible, but the bromide (85a) was converted

to the corresponding threo-p-aminophenylalanine derivative (135). The

bromoacids (137a) and (137b) have been converted to the corresponding

disubstituted p-lactones (139a) and (139b), although isolation of the trans-

p-lactone (139a) was not possible due to its lability. Bromination of the

bromoleucine derivative (91) occurred regiospecifically to give the

y,ô-dibromoleucine derivative (I42). Elaboration of the bromoleucine and

bromohomophenylalanine derivatives (91) and (92) provided the dehydroamino

acid derivatives (106), (107) and (146), suitable for the synthesis of naturally

occurring side-chain functionalized leucine and homophenylalanine derivatives.

Chapter 5 94

RESULTS AND DISCUSSION: CHAPTER 5

Synthesis of Each Stereoisomer of B-Deuteriophenylalanine

It was anticipated that the side chain brominated amino acid derivatives

described in Chapter 1 of the Results and Discussion of this thesis could be

converted to deuterium labelled amino acids, which have found important use

in the study of enzyme catalyzed reactions. To examine this potential, the

synthesis of deuteriated phenylalanine derivatives was studied, with interest

focussed on the development of stereoselective routes to all four stereoisomers

of B-deuteriophenylalanine (65a)-(65d).

Initial attempts to synthesize deuteriated phenylalanine derivatives involved

reduction of the bromides (85a) and (85b) with tri-n-butyltin deuteride" As

reduction of each of the bromides (85a) and (85b) would proceed via the sÍìme

intermediate benzylic radical (150), separation of the diastereomers (85a) and

(85b) was not required. It was anticipated that the intermediate radical (150)

would adopt a preferred conformation (Figure 11) similar to that of the

carbocation (120), with subsequent delivery of the deuterium atom from the

stannane preferentially from one face of the sp2-hybridized radical leading to

the stereoselective formation of the deuterium labelled phenylalanine

derivatives (148a) and (148b). Although radical reactions were once thought to

be impractical for use in asymmetric synthesis, much work in this area has been

reported in recent years, with a variety of radical reactions with very high

stereoselectivity being developed. 155- 1s7

Consequently, a mixture of the bromides (85a) and (85b) was freated with

one equivalent of tri-n-butyltin deuteride in refluxing benzene for 4 hours.

The reaction proceeded in high yield and with 957o 2Et-incorporation, but with

only low stereoselectivity, producing the deuterides (148a) and (148b) in a2:l

ratio. The reaction was then carried out at 25", in an attempt to increase the

Chapter 5 95

selectivity of the reduction, however this required the addition of a catalytic

amount of AIBN and a reaction time of 3 days to achieve a high yield of the

deuterides (1a8a) and (148b). The stereoselectivity of the reaction increased

marginally, giving the deuterides (148a) and (148b) in a 3:1 ratio (Scheme 60).

Reduction of a 1:1 mixture of the bromoesters (82a) and (82b) under

NHCMe3 PhthN CONHCMe3

B1Bu3SnD pl

2 p2

(8s)

(a) Rl=Br" R2=H(b) Rl =H, R2=Br

(148)

(a) R1 =D, R2=H(b) Rl=H, R2=D

Scheme 60

nhthN¿rr,..

pl Bu3SnD pl2 R2

(82)

(u) Rl =Br, R2=H(b) Rl=H, R2=Br

(l4e)

(a) Rl=D, R2=H(b) Rt=H, R2=D

Scheme 61

identical conditions as described for the reduction of the amides (85a) and (85b)

at25" gave the corresponding deuterides Qa9$ and (149b) in 6l%o yield, in a

2:I ratío (Scheme 61).

The lH n.m.r. spectrum of a mixture of the deuterides (148a) and (148b)

showed doublet resonances at õ 3.54 and 3.46, coresponding to the respective

Chapter 5 96

B-protons, and integration of these signals was used to determine the

diastereomeric ratio of the deuterides (148a) and (148b). The lH n.m.r.

spectrum of the material obtained from reduction of the bromoesters (82a) and

(82b) showed doublets at ô 3.59 and 3.53 for the respective p-protons, with

integration of these peaks used to determine the diastereomeric ratio of the

deuterides (I49a) and (1a9b).

The greater selectivity observed in the reduction of the amide (85) than the

ester (82) is consistent with the greater selectivity observed in the

hydroxylation reactions of the bromoamide (85) than the corresponding

bromoester (82), as described in Chapter 2 of the Results and Discussion of this

thesis. It is presumed therefore, that the amide substituent more effectively

blocks one face of the intermediate radical (150) (Figure l l) than does the ester

moiety in the radical (84), such that the stannane delivers the deuterium atom

with greater selectivity in the reduction of the bromoamide (85) than the

bromoester (82). On this basis, it was assumed that the reduction of the

bromides (82) and (85) occurs selectively to give the threo-deuterides Qa9a)

and (148a) as the major products, analogous to the preferential formation of the

threo-alcohols (112a) and (118a) from the bromides (82a) and (85a)" These

assignments were confirmed by experiments described below"

PhrhN

H

CONHCMe3

(1s0)

Figure 11. The radicat (150) in the preferred conformation for deuteriumatom abstraction from tributyltin deuteride.

It was thought that a less reactive deuterium atom donor would result in

higher stereoselectivity in the reductions of the bromides (82) and (85),

H

Ph

Chaprer 5 91

Germanium hydrides have been reported to be more selective reducing agents

than the cofresponding tin hydrides, as the germanium-hydrogen bond is

stronger than the tin-hydrogen bond, thereby increasing the activation energy

required for reaction to occur.158 Reductions of benzylic bromides with

ftiphenylgermanium hydride have been reported to proceed effectively,l59 ¿¡d

consequently reductions of the bromoamide (85) and bromoester (82) were

attempted. Triphenylgermanium deuteride was synthesized by reduction of

triphenylgermanium chloride with lithium aluminium deuteride (997o ztt¡.

However, upon treatment of the bromoamide (85) and bromoester (82) with

triphenylgermanium deuteride, no reduction was observed, even when high

temperatures and activation with ultraviolet light were employed, and the

starting materials (85) and (82) were recovered. The absence of reactivity

under these conditions may be attributed to considerable steric hindrance

inherent in these systems.

As the radical reductions of the bromides (85) and (82) with tri-n-butyltin

deuteride did not proceed stereospecifically, other methods of reduction were

investigated. Sodium borohydride (NaBIIa) effectively reduces benzylic

bromides,l60 þor¡¡syer NaBII¿ has also been used to deprotect phthalimi¿e5.135

The bromide (82b) was therefore treated with NaBH4 in order to determine

whether reduction of the benzylic bromide or the phthalimide group would

occur preferentially. The alcohols (151) and (152) were isolated, indicating

that preferential reduction of the phthatimido group had occurred (Scheme 62).

Evidence for the alcohols (151) and (152) was obtained from their lH n.m.r.

spectra. The secondary alcohol (151) showed doublets at õ 4.28 and 5.75,

colresponding to the cr- and p-protons, respectively. Singlets at õ 4.70 and

5.77 were attributable to the hydroxyl proton and the isoindole ring proton.

The primary alcohol (152) showed doublets at ô 4.70 and 5.92, corresponding

to the cr- and p-protons, respectively. The diastereotopic benzylic protons

exhibited,-4n AB sptitting pattern, with doublets observed at ô 4.69 and 4.86.

NaBHa(82b)

OH

Br

Chnpter 5 98

CO2Me

Br

N

+

(151) (rs2)

Scheme 62

Sodium cyanoborohydride (NaCNBH3) is a less reactive reducing agent

than NaBH4, and its zinc chloride complex is reported to reduce benzylic

halides without affecting amides e¡ i¡¡ide5.161 Accordingly, the bromide (82b)

was treated with the sodium cyanoborohydndelzinc chloride complex, however

no reaction occurred and the starting material (82b) was recovered. The

absence of reactivity may again be attributed to the inherent steric hindrance in

this system.

Efforts were then directed to the catalytic reduction of the bromides (82a)

and (82b) over a palladium catalyst. Initialty, hydrogenolysis of a mixture of

(82a) and (82b) was performed using a 5Vo patladium on carbon catalyst, with

ethyt acetate aS the solvent, under an atmosphere of hydrogen, to examine

whether reduction of the bromides (82a) and (82b) was effected under these

conditions. This reaction proceeded to give the reduced product (74) in good

yield (Scheme 63).

PhthN

pl Hz, Pdlc.'.............................._*

R

CO2Me

2

(82)

(a) Rl=Br, R2=H(b) Rl=H, R2=Br

Scheme 63

(7 4)

Chapter 5 99

Having established benzylic reduction, deuteriolysis of a mixture of the

bromides (82a) and (82b) was then investigated, conducting the reaction under

an atmosphere of deuterium. The reduced product was isolated with only 58Vo

deuterium incorporation, indicating that some scrambling had occurred

between the deuterium gas and ethyl acetate. It was anticipated that a

deuteriated solvent system was required to prevent hydrogen-deuterium

scrambling. Accordingly, treatment of a mixture of the bromides (82a) and

(82b) in a mixture of TFIF and D2O at room temperature, using palladium on

carbon catalyst under an atmosphere of deuterium, proceeded to give a mixture

of the B-deuteriated phenylalanine derivatives (149a) and (149b) with >957o

2H1-incorporation.

Atthough little has been reported on the stereochemical course of the

catalytic reduction of benzylic halides, Berti and co-wo¡¡e¡5162,163 have shown

that hydrogenolysis of 2-phenyl-2-halocyclohexanols occurs with retention of

configuration. The deuteriolysis reactions were therefore repeated on the

separated bromide diastereomers (82a) and (82b) in an attempt to produce the

corresponding deuteriated phenylalanine derivatives (149a) and (149b)

stereoselectively. Reduction of the bromide (82a) under the conditions

described above for reduction of a mixture of the bromides (82a) and (82b)

was indeed stereoselective, producing the deuterides (149a) and (149b) in a

13:1 ratio. Reduction of the bromide (82b) was also stereoselective, giving a

1:13 ratio of the deuterides (149a) and (149b) (Scheme 64). The

stereoselectivity of these reactions was determined as described above for that

of the deuterides (1a9a) and (149b) obtained by reduction of the bromides

(82a) and (82b) with ni-n-butyttin deuteride. The deuteriated phenylalanine

derivatives (149a) and (149b) were produced with ca.99%o 2[t-incorporation,

as determined using rnass spectrometry.

The temperature of the deuteriolysis reactions was then decreased in order

to increas-g the stereoselectivity. Deuteriolysis of the bromide (82b) at 5o

PhthN

Dz, Pd/C

D2, Pdlc

6N HCI...q

AcOHD

Chapter 5 100

+*¡rNrr,.. CO;

,rrtD

(65a)

+t¡rNrr,,.

(65b)

PhthN/r,,.

...q

(82a)

(82b)

Br

Br

(t49a)

PhthNrr,.. PhthN/r,,. coi6N HCI

AcOHD D

(14eb)

Scheme 64

proceeded in24 hours to give the deuterides (149b) and (I49a) in a27:1 ratio.

Below 0",2Hr-methanol was employed as the solvent, as D2O crystallized from

the TIIF/D2O mixture. When the reaction of the bromide (82b) was pedormed

at -20" in 2Hr-methanol, deuteriolysis proceeded with almost complete

stereospecificity (Table 1). The (2S3S)- and (253R)-deuterides (149b) and

Qa9a) were produced in a 100:1 ratio, indicating a diastereomeric excess of

987o, as determined by lH n.m.r. spectroscopy" Analogous reactions of the

threo-bromide (82a) produced the (2S3RJ- and (253Í)-deuterides (149a) and

(149b) in the corresponding ratios as for reduction of the erythro-bromide

(82b). However, precise determination of the diastereomeric excess of the

(253R)-deuteride (149a), produced from the reduction of the threo-bromide

(82a) at -20", was not possible using lH n.m.r. spectroscopy, as resonances

corresponding to a 13C satellite of the methyl ester group of (I49a) and the

methyl ester group of the bromide (82a) present in trace amounts overlapped

with the signal corresponding to the p-proton of the minor (2539)-deuteride

(149b). However, it could be determined that the deuteride (149a) was,nt ,

"101

'!t.ìì.

Chnpter 5

't :.,. .

produced in >95Vo diastereomeric excess. Further experiments described below

detailing the deprotection of the deuteride (149a), and experiments described in

Chapter 6 of the Results and Discussion of this thesis, enabled a more accurate

determination of the stereochemical purity of the deuteride (149a). When the

deuteriolysis was attempted at temperatures below -30o, no reaction was

observed after a period of 7 days.

1 3 1

27 I

1 00 1

no reaction

<r2h

24h

3d

7d

25"

50

-20"

-300

THF/DzO

TFIF/DzO

MeOD

MeOD

diastereo-selectivitytimetemperaturesolvent

Table 1. Diastereoselectivity of the deuteriolysis ofthe bromides (82a) and (82b).

The deuteriophenylalanine derivatives (149a) and (149b) are of limited

utility unless they can be converted to the corresponding free amino acids (65a)

and (65b), without loss of stereochemical integrity. Accordingly, deprotection

of the deuterides Qa9a) and (149b) was investigated. The deuterides (I49a)

and (149b) were hydrolyzed in a 2:1 mixture of 6N HCI and acetic acid, with

subsequent treatment with aniline in ethanol giving the corresponding free

amino acids (65a) and (65b) (Scheme 64). The lH n.m.r. specfrum of the free

amino acid (65a) showed doublet signals at õ 3.92 and 3.21, conesponding to

the a- and p-protons respectively. The coupling constant between these protons

was 4.9 Hz, which is consistent with the coupling constant of 4.5 Hz reported by

Ife and Haslaml12 for the (253R)-deuteride (65a). The lH n.m.r. spectrum of

the free amino acid (65b) showed doublet signals at 83.92 and 3.03,

Chapter 5 lO2

corresponding to the cr- and p-protons respectively, with a coupling constant

between these protons of 7.9 Hz, consistent with the value of 8.5 Hz reported by

Ife and Haslaml12 for the (2S3S)-deuteride (65b). Assignments of the

stereochemistry of the free amino acids (65a) and (65b) were based on the

coupling constants between the a- and p-pro1e¡s.164 Presumably the most

stable conformers of the deuterides (65a) and (65b) have a staggered

orientation, with the phenyl and carboxyl groups lying anti to each other, such

that steric interactions are reduced (Figure 12). It can be seen that in this

conformation (253R)-deuteriophenylalanine (65a) would have a smaller

coupling constant between the cr,- and B-protons than would (2535)-

deuteriophenylalanine (65 b).

D

+H¡N

DH

(65a) (65b)

Figure 12. Preferred conformations of the deuterides (65a) and (65b).

The conversion of the bromides (82a) and (82b) to (253R)- and (2535)-

deuteriophenylalanine (65a) and (65b), respectively, confirms that the

deuteriolysis reactions of the bromides (82a) and (82b) proceed preferentially

with retention of configuration. The diastereomeric excess of each of the free

amino acids (65a) and (65b) was determined to be >98Vo using lH n.m.r.

spectroscopy, indicating that no racemization had occurred during deprotection.

Mass specffometric analysis of the amino acids (65a) and (65b) indicated that

each contained ca. 997o2H1at the B-position.

The (2R3S)- and (2R3R)-þ-bromophenylalanine derivatives (82c) and

(82d) were synthesized via identtcal procedures to those used for the (253R)-

HH

Ph

+H¡N

HPh

Chapter 5 103

and (2S3S)-B-bromides (82a) and (82b), but using (R)-phenylalanine (55) as the

starting material (Scheme 65). Thus, all four stereoisomers of B-bromo-N-

phthaloylphenylalanine methyl ester (82a)-(82d) have been synthesized with

known relative and absolute configuration at both cr- and B-centres.

+PhttrN

i) phthalicanhydride

B2ü) MeOH /

HCI

(s5)

(82c)

D2, Pd/C

Scheme 65

(149c)

..rat

(14ed)

NBS...*

6N HCI

AcOH

6N HCI

AcOH

(82)

(c) Rl = Br, R2

(d) Rl = H, R2

D

(65c)

+H

(65d)

=fl=Br

PhthN+

H¡N

Br

CO2Me

D2,Pdlc

coi

"4DBr D

(82d)

Scheme 66

Deuterioiysis of the bromides (82c) and (82d) was carried ouL aL -20" in

2Hr-methanol, as described for the reactions of the bromides (82a) and (82b),

and gave,.the corresponding deuteriated phenylalanine derivatives (149c) and

Chapter 5 LM

(149d). Subsequent deprotection of these compounds was achieved as described

for the deuterides (I49a) and (149b), to give the deuterium labelled free amino

acids (65c) and (65d) (Scheme 66). The deuterides (65c) and (65d) were

produced in 98Vo d.e., and with cø. 99Vo 2H1-incorporation, as for the

production of the deuterides (65a) and (65b).

Hence a procedure has been developed for the synthesis of all four

stereoisomers of p-deuteriophenylalanine (65a)-(65d), homochiral at the

q,-cenffe, with 987o diastereomeric excess and 99Vo 2H1-incorporation.

Chapter 6 105

RESULTS AND DISCUSSION: CHAPTER 6

Stereochemical Course of the Reactions of (S)- and

(R)-Phenylalanine with (S)-Phenylalanine Ammonia Lyase

The highly stereoselective synthesis of all four stereoisomers of

p-deuteriophenylalanine (65a)-(65d), as described in Chapter 5 of the Results

and Discussion of this thesis, enabled an investigation of the stereochemical

course of the reaction of both (S)- and (R)-phenylalanine, (56) and (55), with

the enzyme (S)-phenylalanine ammonia lyase (PAL).

In accord with Battersby's studieslS Eeatment of (253R)-3-deuterio-

phenylalanine (65a) with PAL in sodium borate buffer at pH 8.7 gave

deuteriated trans-cinnamic acid (67) (Scheme 67). The deuteriated cinnamic

acid (67) was identified using lH n.m.r. spectroscopy, with a broad singlet

observed at ô 6.46 corresponding to the cr-vinylic proton, consistent with

literature values.l5 A fiace of non-deuteriated trans-cinnamic acid (57) was

observed, with integration of the relevant peaks in the lH n.m.r. spectrum

indicating that the product contained 987o deterium at the p-position. The

presence of ZVo non-deuteriated cinnamic acid (57) is consistent with the

production of the deuteriophenylalanine derivative (65a) in 987o diastereomeric

excess and with 997o 2E1-incorporation, as described in Chapter 5 of the

Results and Discussion of this thesis. This result infers that specific removal of

the 3-pro-S hydrogen from (S)-phenylalanine (56) occurs during reaction with

the enzyme, with l7o of the non-deuteriated cinnamic acid (57) arising from

unlabelled (S)-phenylalanine (56), and the remainder arising from the lVo

(253 S)-deuteriophenylalanine (65b) in the sample.

Similar treatment of (2S3S)-3-deuteriophenylalanine (65b) with PAL gave

trans-cinnamic acid (57) (Scheme 67), providing further evidence for the above

conclusion. The product (57) was identified by comparison of its lH n.m.r.

Chnpter 6 106

spectrum with that of an authentic sample, giving doublets atõ 6.41 and 7.81,

corresponding to the cr- and B-vinylic protons, respectively, with a coupling

constant of 16.0 FIz being characteristic of a trans-sttbstituted u1¡e¡s.120 No

signals attributable to deuteriated trans-cinnamic acid (67) were observed.

Approximately I7o deuteriated cinnamic acid (67) is expected from this

reaction, due to l%o of the contaminant (253R)-deuteriophenylalanine (65a).

However, the signal in the lH n.m.r" spectrum due to the ø-proton of

deuteriated trans-cinnamic acid (67) occurs as a broad singlet atõ 6.46, and is

presumably not visible under the doublet signal due to the a-proton of cinnamic

acid (57).

t¡rNzrr,,.

PAL

+

D

(65a) (67)

+H¡N¿¿,,.

PAL

H

(6sb) (s7)

Scheme 67

These results are in agreement with the conclusion of Battersby and

co-workers15 that the elimination reaction catalyzed by PAL proceeds viø

abstraction of the 3-pro-S hydrogen of lS)-phenylalanine (56) with anti-

elimination of the amino group leading to formation of trans-cinnamic acid

Chapter 6 lO7

(57). Battersby and co-workersl5 used deuteriated phenylalanines which

contained 907o z}lr at the p-position, and which were of less than 93c/o

stereochemical purity, such that the deuteriated trans-cinnamic acid (67)

obtained from reaction of the (253R)-deuteride (65a) contained only 88+4Vo

2IJt" Similarly, the trans-cinnamic acid (57) obtained from reaction of the

(2S3S)-deuteride (65b) with PAL contained 8+2Vo z}It. The results described

above confirm that the minor impurities in Battersby's studies are in fact due to

impurities in the substrates, and are not due to a lack of enzyme

stereospecificity.

Atthough (R)-phenylalanine (55) is a subsffate of PAL,I10 nq mechanistic

or stereochemical studies have been reported with this compound.

(R)-Phenylalanine (55) binds effectively to the active site, but is metabolized at

only one five thousandth the rate of (S)-phenylalanine (50).tto We therefore

wished to confirm the product from the reaction of (R)-phenylalanine (55) with

PAL, and then investigate the stereochemical course of the reaction, with the

interest being to examine how the enzyme manages to metabolize a substrate

with the opposite chiratity to its normal substrate. Initially, (R,)-phenylalanine

(55) was treated with PAL and trans-cinnamic acid (57) was isolated in

reasonable yield. Treatment of (2R35)-deuteriophenylalanine (65c) with PAL

gave deuteriated cinnamic acid (67) with 927o 2Hr-incorporation, indicating

that the enzyme preferentially abstracts the 3-pro-R hydrogen from

(R)-phenylalanine (55) (Scheme 68). Treatment of (2R3R,)-deuterio-

phenylalanine (65d) with the enzyme confirmed this result, giving non-

deuteriated cinnamic acid (57) as the major product (Scheme 68).

These results can be explained by examining the conformation of both

(S)-phenylalanine (56) and (R)-phenylalanine (55) in the active site.

trans-Cinnamic acid (57) and benzoic acid have been shown to inhibit the

enzyme,lOT'IO8 indicating that the active site contains a hydrophobic pocket for

binding oi{ Fe aromatic ring, and positively charged residues, possibly arginine,

Chapter 6 108

to accommodate binding of the carboxyl group. As described in the

Introduction, the active site also contains a prosthetic group for covalent

attachment of the amino group. For anti-elimination to occur, a B-hydrogen

must be antiperiplanar to the nitrogen atom. The phenyl and carboxyl groups

must also be qnti to each other, such that they aÍe trans to each other in the

product cinnamic acid (57). Therefore, when (S)-phenylalanine (56) binds in

the active site, it must adopt a conformation in which the amino, carboxyl and

phenyl groups and the 3-pro-S hydrogen all lie in a plane (Figure 13). A basic

residue such as histidine is presumably situated in an appropriate position to

abstract the 3-pro-S hydrogen with anti-elimination leading to formation of

tans -cinnamic acid (57).

PAL

D 927ozH1

(65c) (67)

co2H

PAL

H 277o 2H1

(6sd) (s7)

Scheme 68

H

+H

+coiH¡N

+

(5s)

coi

pro-R

Chapter 6 109

H H

Ph- B-Enz

..r¡lll NH

oEnz-HN

HR o- HR

Figure 13" (S)-Phenylalanine (56) in the active site of the enzyme PAL.

llntr. Enz-HN

Ph Hn_/- B-EnzH H

Figure 14. (R)-Phenylalanine (55) in the active site of the enzyme PAL.

(R)-Phenylalanine (55) can also be situated in the active site with the

phenyl, amino and carboxyl groups bound in the same way as that for

(S)-phenylalanine (56), even though the chirality at the c-centre is reversed

(Figure 14). In this conformation, the phenyl and carboxyl groups are in the

required anti onentation, and the amino group and pro-R hydrogen are also

antiperiplanar. The pro-R hydrogen of (R)-phenylalanine (55) occupies a

similar position in the active site to that which the pro-S hydrogen of

(S)-phenylalanine (56) occupies. Thus, abstraction of the pro-R hydrogen from

(R)-phenylalanine (55) occurs preferentially, wîth anti-elimination again giving

trans -cinnamic acid (57).

However, it can be seen that the reaction of the enzyme with

(R)-phenylalanine (55) is not completely stereospecific, as is the case with

(S)-phenylalanine (56). Reaction of the (2R35)-deuteride (65c) occurs wíth ca.

87o remov"SL of the B-deuterium atom to give the cinnamic acid (67) with only

Ba*

NH

Chapter 6 110

927o 2[rincorporation. Similarly, reaction of the enzyme with the (2R3R)-

deuteride (65d) occurs with 277o removal of the B-hydrogen atom to give the

cinnamic acid (57) with2TVo 2H1-incorporation. The difference between these

ratios is attributable to two deuterium isotope effects, and indicates that

although the pro-R hydrogen is preferentially abstracted from

(R)-phenylalanine (55), presumably ca. l57o of the reaction occurs vic

abstraction of the pro-S hydrogen. Reaction of the (2R35)-deuteride (65c) with

PAL occurs with only 8Vo (rather than l57o) removal of the p-deuterium due

to a deuterium isotope effect of ca. 2, and similarly reaction of the (2R3R)-

deuteride (65d) occurs with 27Eo (rather than l57o) removal of the B-hydrogen

atom, again due to a deuterium isotope effect of ca.2. As no cis-cinnamic acid

is produced from this reaction, concerted removal of the pro-S hydrogen and

elimination of ammonia must proceed via a syn-elimination. Thus,

(R)-phenylalanine (55) must be distorted in the active site and situated in such a

way that the basic residue is sufficiently close to the pro-S hydrogen, or another

basic residue in the active site may be close enough to the pro-S hydrogen to

abstract it, with elimination then leading to the production of trans-cinnamic

acid (57) (Figure 16). It is interesting to note that if a second basic residue was

to exist in proximity to the pro-S hydrogen of (R)-phenylalanine (55) in the

active site (Figure 16), it would not interfere with binding of the natural

subsffate, (S)-phenylalanine (5O Gf Figure 13).

B

ll tt t t,.

H

Ho-

Figurp"16. Abstraction of the pro-S hydrogen of (R)-phenylalanine (55).

chapter 6 111

In conclusion, it has been demonstrated that the elimination of a p-proton

and ammonia from (S)-phenylalanine (56) catalyzedby the enzyme PAL occurs

with stereospecific removal of the pro-S hydrogen. In contrast, the elimination

of a p-proton and ammonia from (R)-phenylalanine (55) catalyzed by the

enzyme PAL occurs with stereoselective removal of the pro-R hydrogen. The

pro-R hydrogen of (R,)-phenylalanine (55) occupies the same position in the

active site as does the pro-S hydrogen of (S)-phenylalanine, and is therefore

preferentially abstracted. However, the unnatural substrate lR)-phenylalanine

(55) presumably is distorted such that the elimination catalyzed by PAL is not

completely stereospecific.

Conclusion II2

CONCLUSION

The work described in this thesis has shown that N-phthaloyl-protected

cr-amino acid derivatives are deactivated towards hydrogen abstraction at the a-

position. This effect has been attributed to steric and electronic factors.

Bromination of various N-phthaloyl amino acid derivatives has been shown to

occur regiospecifically via the most stable side chain radicals"

The versatility of brominated amino acid derivatives has been exploited in

the synthesis of a variety of homochiral hydroxy-G-amino acids and their

derivatives" Stereoselective syntheses of (S)-threo-þ-hydroxy-phenylalanine

and tyrosine derivatives have been developed" Incidentally, the (S)-threo-

isomers of p-hydroxy-a-amino acids are the predominant stereoisomers which

occur in nature. Deprotection procedures have been developed to provide the

free amino acid (253R)-þ-hydroxyphenylalanine, a component of various

naturally occurring peptide antibiotics. The methods are suitable for the

preparation of unnatural and rare amino acid derivatives. A p-hydroxy-

p-nitrophenylalanine methyl ester derivative, suitable for elaboration to the

antibiotic chloramphenicol, has also been prepared. In future work,

exploitation of the potential benefits of using p-bromoamino acid derivatives

with the carboxyl group protected as an amide rather than an ester, discovered

during the course of the present study, should provide an even more efficient

route to the synthesis of chloramphenicol.

Procedures have been formulated for the stereocontrolled synthesis of

trans-3, -disubstituted p-lactams and threo-þ-aminophenylalanine derivatives

with control of relative and absolute stereochemistry. A suitable method for

the introduction of functionality at the ô-position of leucine derivatives, the

least activated of the positions on the side chain, has been developed. Methods

have also been developed for the introduction of functionality at the p-position

of leucine and homophenylalanine derivatives.

Conclusion I 13

A stereoselective route which provides each of the stereoisomers of

p-deuteriophenylalanine with very high diastereoselectivity and deuterium

incorporation has been developed. The stereoisomers of p-deuterio-

phenylalanine have been used to determine the stereochemical course of the

reaction of both lS)- and (R)-phenylalanine with the enzyme (S)-phenylalanine

ammonia lyase (PAL)" It has been determined that PAL catalyzes the

stereospecific removal of the pro-S hydrogen from the natural substrate

(S)-phenylalanine, whereas reaction with (R,)-phenylalanine occurs with

stereoselective removal of the pro-R hydrogen" The contrasting reactions of

the natural substrate and its antipode with PAL were attributed to both

compounds occupying the active site of the enzyme with the amino, carboxyl

and phenyl groups bound at specific sites. The p ro-R hydrogen of

(R)-phenylalanine then occupies the same spacial position in the enzyme active

site as does the pro-S hydrogen of (S)-phenylalanine when it is bound.

The above syntheses are all enantiospecific, except where indicated in the

text, with the chirality determined by that of the starting amino acids"

Although homochirality of each individual compound has not been determined

independently, it is implied by inference with representative examples. The

chiral integrity of a bromophenylalanine derivative has been established

through analysis of a reduced derivative using HPLC with a chiral stationary

phase. An hydroxyvaline derivative has been shown to be homochiral using

lH n.m.r. spectroscopy with a chiral shift reagent, which also implies the

homochirality of the precursor bromide. Comparison of a leucine lactone with

an authentic sample confirms the chiral integrity of the precursor hydroxy- and

bromo-leucine derivatives. Optical rotation of a p-hydroxyphenylalanine free

amino acid and comparison with an authentic sample has established the

homochirality of the corresponding hydroxy- and bromo-phenylalanine

derivatives. X-Ray crystallographic analysis of a hydroxyphenylalanine

derivative and a bromo-p-nitrophenylalanine derivative has also established the

Conclusion ll4

chirality of these compounds and of their precursors. Reactions of

deuteriophenylalanine stereoisomers with PAL confirm the homochirality of

these compounds and also of the corresponding precursors.

In summary, ûew procedures have been developed for the synthesis of

functionalized cr-amino acid derivatives with control of both relative and

absolute stereochemistry, and the use of such compounds in the study of

biochemical systems has been illustrated.

Experimental 115

EXPERIMENTAL

General

Melting points were determined on a Kofler hot-stage under a Reichert

microscope and are unconected.

Elemental analyses were carried out by Canadian Microanalytical Service

Ltd., New Westminster, Canada, and by Chemical and Micro Analytical

Services Pty. Ltd., Victoria, Australia.

Analytical thin layer chromatography was performed using Merck

Kieselgel 60 Fzs+ silica on aluminium backing plates. Preparative

chromatography was performed using positive pressure flash

chromatography165 or dry column flash chromatographyl66 using Merck

Kieselgel 60 (230-400 mesh ASTM).

High performance liquid chromatography (HPLC) was carried out using a

V/aters 60004 solvent pump, a Waters U6K injector and a Waters model 441

absorbance detector operating at 254 nm, in conjunction with an ICI DP-700

data station. Analyses were performed using a Waters Radial Pak normal phase

10 pm silica column (100 x 8 mm), eluting with a gradient of hexanelethyl

acetate, or where specified, a Waters Nova Pak C13-reverse phase 10 pm silica

column (100 x 8 mm) eluting with a gradient of methanol/water, or a Regis

Pirkle Covalent (S)-Phenylglycine column (250 x 4.6 mm), eluting with a

gradient of hexane/iso-propanol.

Infrared spectra were recorded on an Hitachi 270-30 spectrophotometer or

a Jasco A-I02 spectrophotometer using the 1603 cm-l band of polystyrene as a

reference. Spectra were recorded as solutions in dichloromethane unless

otherwise specified.

Nuclear magnetic resonance spectra were recorded on either a Bruker

CXP-300 " or Bruker ACP-300 spectrometer. Proton nuclear magnetic

Experimental 116

resonance (lH n.m.r.) specna were recorded at 300 MHz, and carbon nuclear

magnetic resonance (13C n.m.r.) spectra were recorded at75.5 MHz. Spectra

were either recorded in deuteriochloroform using tetramethylsilane (TMS) as

an internal standard or in deuterium oxide (DzO) using sodium 12,2,3,3-2Ha)3-

(trimethylsilyl)propionate (TSP) as an internal standard. Chemical shifts are

quoted as õ in parts per million downfield from the internal standard.

Multiplicities are abbreviated to; s, singlet; d, doublet; t, triplet; q, quartet; m,

multiplet; br, broad.

Electron impact mass spectra were recorded with an AEI MS-30 double

focussing mass spectrometer operating at 70 eV or on a Vacuum Generators

ZAB 2ÉIF mass specfrometer. Fast atom bombardment (FAB) mass spectra

were recorded on a Vacuum Generators ZAB 2FIF mass spectrometer.

All solvents were distilled before use. Anhydrous diethyl ether and

tetrahydrofuran (THF) were obtained by distillation from sodium

benzophenone ketyl. Drying and purification of other solvents and reagents

was performed using standard laboratory procedures.l6T'168 Organic solutions

were dried by the addition of anhydrous magnesium sulfate, unless otherwise

specified.

Experimental II7

(S)-N-Phthaloylvaline (68)

A mixture of (S)-valine (10.0 g, 85 mmol) and finely ground phthalic

anhydride (12.6 g, 85 mmol) was heated in an oil bath at 145-150o for 30 min.

The mixture was allowed to cool, then the solid was dissolved in hot methanol

(40 rnl). The solution was filtered and then water (60 ml) was added. The title

compound (68) crystallized on cooling and was isolated as fine colourless

crystals (18.8 g,89Vo), m.p. 116-117" (tit.t6q 118-119'), [a]¡22 -6go (c, 1 in

acetone) (lit.ttz [ct]o23 -69.4" (c, 1 in ethanol)). lH n.m.r. (300 MHz, CDCI3) ô

0.93, d, J 6.7 Hz,3}J, Me; 1 .l'7, d, J 6.7 Hz,3H., tr..fe'; 2.71, m, H3; 4.65, d, J

8.3 Hz, H2:7.14-1.90, m, 4H, Phth; 10.68, br s, CO2H.

(S)-N-Phthaloylleucine (69)

A mixture of (S)-leucine (10.0 g, 26.2 mmol) and finely ground phthalic

anhydride (11.3 9,26.3 mmol) was heated in an oil bath at 145-150" as

described for the synthesis of (S)-N-phthaloylvaline (68) from (S)-valine. The

títle compound (69) was isolated as a white solid (19.07 g,967o), m.p. l2I-I22"(lit.tzo 120-121"), [a]o23 -25" (c, 1 in acetone) (li1.t6e ¡s¿lrzz -24" (c, I inethanol)). lH n.m.r. (300 MHz, CDCI3) õ 0.94, d, J 6.6 Hz,3H, Me; 0.97, d, J

6.6 Hz,3H, Me'; 1.53, m, H4: I.96, m, H3; 2.39, m, H3'; 5.02, dd, J 4.2H2,

lI .5 Hz, II2; 7 .16, m, 2H, Phth; 7 .89, m, 2H, Phth; 9.64, br s, CO2H.

(S)-N-Phthaloylphenylalanine (70)

A mixture of (S)-phenylalanine (10.0 g, 60.5 mmol) and finely ground

phthalic anhydride (9.0 g, 60.8 mmol) was heated in an oil bath at 145-150" as

described for the synthesis of (S)-N-phthaloylvaline (68) from (S)-valine. The

title compound (70) was isolated as fine colourless crystals (17.08 g,967o),

Experimental 118

m.p. 187' (lit.trt 187'), [cr]o2r -212" (c, 1 in acetone) (lit.tts [a]o -212o (c, 1.9

in ethanol)). tH n.m.r. (300 MHz, CDCI3) õ 3.59, d, J 8.5 }{z,2H,CHz;5.23,t,

J 8.5 Hz, II2;7.16, s,5H, Ph;7.62-7.77,m,4H, Phth; 10.68, br s, CO2H.

(S)-N-Phthaloyltyrosine (71)

(S)-Tyrosine (10.0 g, 55.2 mmol) and phthalic anhydride (8.2 g, 55.3

mmol) were dissolved in boiling DMF (50 nìl) and the solution was boiled for a

further l0 min. The mixture was then filtered, and the fìlrate was poured into

water (500 mt) with vigorous stirring. The precipitate was collected by

filtration and was recrystallized from methanol/water to give the title

compound (71) as a white solid (13.32 g,787o), m.p. 163-165'(lit.t6e 173-

I74"), [cr]o23 -234" (c, 1 in acetone) (lit.16e [a]o22 -232' (c, 1 in ethanol)). lH

n.m.r. (300 MHz, CDCI¡) ô 3.58, d, J 8.4 Hz,CHz;5.21,t,J 8.4H2,ÍI2;6.91,

d, J 8.6 Hz,2H, ArH; 7.I8, d, J 8.6 Hz,2H, ArH; 7.67-7.8I, m,4H, Phth; 9.05,

br s, COzH.

(S)-N-Phthaloylvaline Methyl Ester (72)

(S,)-N-Phthaloylvaline (68) (6.0 g, 24.3 mmol) was treated with dry

methanol (100 rrìl) which had been pre-treated with thionyl chloride (1 ml, 14

mmol). The solution was stirred under anhydrous conditions for 16 h, then the

solvent was removed under reduced pressure. The residue was taken up in

dichloromethane, and the organic solution was washed with I07o NazCO¡ and

water, then was dried. Removal of the solvent under reduced pressure and

recystallizatíon of the residue from hexane/ethyl acetate gave the title

compound (72) as a viscous oil (5.52 g,87Vo), [a]p23 -68" (c, 1 in acetone). lH

n.m.r. (300 MHz, CDCI3) õ 0.92, d, J 6.8 IJz,3}J, Me; 1.16, d, J 6.8 Hz,3H,

Experimental 119

Me'; 2.77 , m, H3; 3.72, s, 3H, OMe; 4.58, d, J 8.3 Hz, H2; J .78, m, 2H, Phth;

7.88, m, 2H, Phth.

(S)-N-Phthaloylleucine Methyl Ester (73)

(s)-N-Phthaloylleucine (69) (19.07 g, 73.O mmol) was dissolved in dry

methanol (150 rnl) which had been pre-treated with thionyl chloride (2.8 9,24

mmol). The solution was stirred under anhydrous conditions for 16 h, then the

solvent was removed under reduced pressure. The residue was taken up in

dichloromethane (150 ml), and the solution was washed with l07o NazCO¡ (100

ml) and water (100 mt). The organic phase was dried, then was concenfated

under reduced pressure" Distillation (170o, 0.05 mm, block) gave the title

compound (73) as a clear viscous oit (18.90 g, 94 7o), [o]p23 -23" (c, 1.0 in

acetone). 1H n.m.r. (300 MHz, CDCI3) ô 0.93, d, J 6.8 Ilz,3H, Me; 0.96, d, J

6.6 H2, 3H, Me'; I.49, m, H4; 1.97, ddd, J 4.4 }l;2, I0.3 }lz, I4.3 Hz, }l3; 2.34,

ddd, J 4.1 Hz, II.6 Hz, 14.3 Hz,H3';3.73, s, 3H, OMe; 4.96, dd, J 4.4 Hz, lI.6

Hz, ÍI2; I .7 5, m, 2}l, Phth; 7 .87, m, 2H, Phth.

(S)-N-Phthaloylphenylalanine Methyl Ester (74)

(S)-N-Phthatoylphenylalanine (70) (5.0 g,16.9 mmol) was dissolved in dry

methanol (100 rnl) which had been pre-treated with thionyl chloride (1 ml, 14

mmol), as described for the preparation of (S)-N-phthaloylvaline methyl ester

(72) from (S)-N-phthaloylvaline (68). The title compound (74) was isolated as

colourless crystals (4.48 g,867o), m.p. 123-125' (lit.l72 Il3-114"), [a]o2: -

212" (c, 1.0 in acetone). lH n.m.r. (300 MHz, CDCI3) õ 3.58, dd, J 10.9 Hz,

14.3 Hz,lf3;3.63, dd, J 5.6 Hz,14.3 Hz,H3';3.80, s, 3H, OMe; 5.20, dd, J 5.6

Experímental l2O

Hz, 10.9 }lz, H2;7.I8, s, 5H, Ph;1.70, m,2H, Phth; 1.79, m,2fJ, Phth.

lH n.m.r. spectral data consistent with literature uu1u"r.173

(S)-N-Phthaloyltyrosine Methyl Ester (75)

lS)-N-Phthaloyltyrosine (71) (8.0 g,25.7 mmol) was dissolved in dry

methanol (100 rnl) which had been pre-treated with thionyl chloride (1 ml), and

the solution was stirred under anhydrous conditions for 16 h. The solvent was

removed under reduced pressure, then the residue was taken up in

dichloromethane. The solution was washed with IOTo Na2CO3 and water, then

was dried. Removal of the solvent under reduced pressure and recrystallization

of the residue from hexane/dichloromethane gave a ffanslucent gel, which upon

drying in air and over PzOs gave the title compound (75) as a white powder

(6.73 g,8l7o), m.p. 101-104' (Iit.130 98-99"). lH n.m.r. (300 .I..f.}Jz, CDCI¡) õ

3.44, dd, J 11.0 Hz, l4.4Hz, H3; 3.50, dd, J 5.7 Hz,I4.4 Hz, H3'; 3.77, s, 3H,

OMe; 5.10, dd, J 5.7 Hz,II.0Hz,I12;6.62, d, J 8.6 }lz,2H, ArH; 6.98, d, J 8.6

Hz, zIJ, ArH; 7 .66-7 .78, m, 4H, Phth"

N-Phthaloylhomophenylalanine (7 6)

A mixtu¡e of homophenylalanine (4.7 g,26.2 mmol) and finely ground

phthalic anhydride (3.9 9,26.3 mmol) was heated in an oil bath at 145-150" as

described for the synthesis of (S,)-N-phthaloylvaline (68) from (S)-valine. The

title compound (76) was isolated as a white solid (7.29 g,907o),m.p. 144-145"

(lit.tz+ 142.5-144"). lH n.m.r. (300 MHz, CDCI3) õ 2.56-2.77 , m, 4H, CIlzCftz;

4.93, m, II2; 7 .A9-7 .29, m, 5II, Ph; 7 .7 5, m, 2II, Phth; 7 .86, m, 2H, Phth; 9.43,

br s, CO2H.

Experimental I2l

N-Phthaloylhomophenylalanine Methyl Ester (77)

N-Phthaloylhomophenylalanine (76) (7.0 9,22.7 mmol) was dissolved in

dry methanol (100 ml) which had been pre-ffeated with thionyl chloride (1 ml,

14 mmol). The solution was stirred under anhydrous conditions for 16 h, then

the solvent was removed under reduced pressure. The residue was taken up in

dichloromethane, and the solution was washed with l07o NazCO3, then with

water, and was dried. Removal of the solvent under reduced pressure and

purification by chromatography on silica, followed by distillation (17 5" ,

0.03mm, block), gave the title compound (77) as a clear oil (6.73 g, 927o),

(Found: C, 70.5; H,5.3; N,4.4. CrqHrzNO4 requires C,70.6: H, 5.3; N,4.37o).

lH n.m.r. (300 MHz, CDCI3) õ2.52-2.71, ffi, 4}l,CHzCHz;3J72, s,3H, OMe;

4.88, m, lH, ÍÐ;7.08-7.22,m,5H, Ph; 7.74,m,2H, Phth; J.84,m,2H, Phth"

v max 1794, 17 44, 1720, I 388, 1098, 708 cm-l. Mass spectrum; ml z 324 (M+H+,

27o),323 (M*', I),307 (-CH4, 2),262 (7), 219 (100), 187 (85), 132 (24), 109

(30), 104 (21),9t (21),76 (t2).

(R S) -N- f e r t -B uty l-¡rr ø- p h t h a I o y I p h e n y I a I an i n a m i d e

Thionyl chloride (12.1g, 0.10 mol) and pyridine (0.1 ml) were added to a

suspension of (S)-N-phthaloylphenylalanine (70) (10.0 9,34 mmol) in CCI¿

(100 ml). The mixture was refluxed for 3 h, allowed to cool, then the solvent

and excess thionyt chtoride were removed under reduced pressure. The residue

was suspended in CCl¿ (100 ml) and tert-butylamine (6.2 9,85 mmol) was

added slowly. The mixture was refluxed for I h, then the solvent and excess

tert-butylamine were removed under reduced pressure. The residue was taken

up in ethyl acetate, then was washed with I07o NazCO¡ and water. The solution

was dried, then the solvent was removed under reduced pressure. The residue

Experimental I22

was recrystallized from hexane/ethyl acetate to give the title compound as white

crystals (10.3 g,87Vo), m.p.194-195". Spectral data identical to that described

below for the (S)-enantiomer (78).

Elution of this compound through a Regis Pirkle HPLC column showed the

presence of two compounds, with retention times of 2I and 22 min, eluting

with a gradient of hexane and iso-propanol (5-407o iso-propanol). The

material obtained viø this method was therefore assumed to be racemic.

(S) -N -terf-Butyl-Nø-phthaloylphenylalaninamide (78)

To a solution of (S)-N-phthaloylphenylalanine (70) (8.0 9,27.1 mmol) and

triethylamine (2.74 9,2J.1 mmol) in dichloromethane (100 ml) was added

ethyl chloroformate (2.94 9,27.1 mmol). The mixture was stirred for 10 min,

then was cooled to 0". tert-Búylamine (1.98 9,27.1 mmol) was added, and the

mixture was stirred at 0o for 15 min. The mixture was allowed to warm to

room temperature, then was stirred for a further 30 min. The mixture was

filtered, the filtrate was washed with water, then was dried. The solvent was

removed under reduced pressure and the residue was recrystallized from

hexane/ethyl acetate to give the títle compound (78) as a white solid (8.8 g,

937o), m.p. 186-188" (Found: C, 72.0; H, 6.3; N, 8.I. CztHzzNzO¡ requires C,

72.0: H,6.3; N, 8.07o)" lH n.m.r. (300 MHz, CDCI3) ô 1.30, s,9H, CMe¡; 3.48,

dd, J 10.0 IJz, l4.l Ha H3; 3.56, dd, J 6.7 Hz, l4.l Hz,H3'; 4.99, dd, J 6.7 Hz,

10.0 Hz, H2:5.62, br s, NH; 7.18, s,5H, Ph;7.68-1.80, m,4H, Phth. Vmax

3300, 1775, 1720,1660, 1510, 1380, 720 cm-r" Mass spectrum; mlz 350 (M*',

lTVo),279 (l),251 (50), 250 (54),232 (26),203 (10), 188 (7)' 173 (19),167

(29), 160 (20), r49 (100), 104 (41), 9r (16),77 (33).

Elution of this compound through a Regis Pirkle HPLC column showed the

presence of only the (S)-enantiomer (78). This enantiomer (78) had a retention

Experimental I23

time of 21 min, eluting with a gradient of hexane and iso-propanol (5-407o iso-

propanol).

(S) -O -

^cetyl-N-phthaloyltyrosine (79)

A solution of (S)-N-phthaloyltyrosine (71) (3.0 g, 9.6 mmol) in acetic

anhydride (20 ml) was refluxed for 4 h. The solution was allowed to cool, then

was poured into water (200 ml) and stirred vigorously for 10 min. The

mixture was cooled on ice and the solid was isolated by filtration, washed with

water, and dried over P2O5. Recrystallization of the solid gave the title

compound (79) as a white powder (2.68 9,797o), m.p. 186-187" (Found M+',

mlz 353.091. CleHlsNOo requires mlz 353.090¡. t1¡ n.m.r. (300 MHz, CDCI¡)

õ2.23, s, 3H, OAc; 3.58, d, J 8.3 }lz,2H, CHz;5.22, t, J 8.3 H:z,IÐ;6.92, d, J

8.4H2,2H, ArH;7.18, d, J 8.4 IIz,2H, ArH; 7.67-7.80, m,4H, Phth; 9.09, br

s, COzH. v*a* 3480, 3100-2600 br, 17J5, 1760, 1720,1510, 1390,1195, 1020,

9I5,720 cm-l. Mass specfium; mlz 353 (M*', l7o),311 (-CHzCO, 5), 288

(r4),273 (4),227 (5),2r3 (5), 187 (6), r73 (13), 164 (100), r47 (PhrhNH,

24), r30 (10), 121 (11), 107 (64), r04 (13), 91 (t0),71 (17),76 (17), 43 (30).

(S) -O -A c etyl- N - terf-b utyl -¡¡r ø-p hthaloyltyrosinamide (8 0)

To a solution of (S)-O-acetyl-N-phthaloyltyrosine (79) (2.0 9,5ó7 rnrnol)

and triethylamine (574 mg,5.67 mmol) in dichloromethane (20 ml) was added

ethyl chloroformate (615 mg, 5.67 mmol). The mixture was stirred for 10

min, then was cooled to 0". tert-Butylamine (415 mg, 5.67 mmol) was added,

and the mixture was stirred at 0" for 15 min. The mixture was allowed to warm

to room temperature, then was stirred for a further 30 min. The mixture was

filtered, the filtrate was washed with water, then was dried. The solvent was

Experimental 124

removed under reduced pressure and the residue was recrystallized from

hexane/ethyl acetate to give the title compound (80) as a white solid (2.15 g,

937o), m.p.212-213' (Found: C, 67.2; H, 5.8; N, 6.7. C23H2aN2O5 requires C,

67.6; H,5.9; N,6.97o). lH n.m.r. (300 MHz, CDCI3) ô 1.31, s,9H, CMe¡; 2.24,

s, 3H, OAc; 3.50, dd, J 10.0 Hz,l4.2Hz,H3;3.56, dd, J 6.8 IJ'z,I4.2Hz,H3';

4.98, dd, J 6.8 Hz, 10.0 Hz, H2:5.87, br s, NH; 6.92, d, J 8.5 Hz,2}I, ArH;

7.I9, d, J 8.5 }Iz,2H, ArH; 7.69-7.8I, m,4H, Phth" vnlu* 3430,1762, 1718,

1688, 1510, 1392, 1198, 960,ll9 cm-l. Mass spectrum; mlz 408 (M+',98o),

308 (4), 266 (59),248 (24), 2t9 (56), r92 (29), r13 (20), 163 (23), 162 (22),

r47 (31), 130 (25), t20 (27), t07 (68), 104 (5r),76 (53),43 (100).

(S)-3-Bromo-N-phthaloylvaline Methyl Ester (81)

To a solution of (S)-N-phthaloylvaline methyl ester (72) (1.0 g, 3.83

mmol) in CCI¿ (30 rnl) was added NBS (0.68 g, 3.83 mmol), and the mixture

was refluxed for 2 h while irradiated with a 250W mercury latnp. The mixture

was allowed to cool, then was filtered and the filtrate was washed with water

(30 mt). The organic phase was dried and then concenfrated under reduced

pressure. Recrystallization of the residue from hexane/dichloromethane gave

the títle compound (81) as a white solid (0.87 g,83Vo), m.p. 124-126". rH

n.m.r. (300 MHz, CDCI3) ô 1.99, s, 3H, Mle;2.15, s, 3H, Me'; 3.7I, s, 3H,

OMe; 5.I7, s, H2; 7.81, m,2H, Phth; 7.92,m,2H, Phth. lH n.m.r. spectral

data consistent with literature values.42

Experimental I25

(253R)-3-Bromo-N-phthaloylphenylalanine Methyl Ester (82a) and

(2535)-3-Bromo-N-phthaloylphenylalanine Methyl Ester (82b)

To a solution of (S)-N-phthaloylphenylalanine methyl ester (74) (10.0 g,

32.3 mmol) in CCla (150 ml) was added NBS (5.76 g,32.4 mmol), and the

mixture was refluxed for 2 h while irradiated with a 250W mercury lamp" The

mixture was allowed to cool, then was filtered and the filtrate was washed with

water (100 ml). The organic phase was dried, then was concentrated under

reduced pressure to give the title compounds (82a) and (82b) (12.55 g, lffi%o)

as a 1:l mixture. Fractional recrystallization from hexane/dichloromethane

(ca.2:1,200 ml) gave the threo-diastereomer (82a) (5.27 g,427o), m.p. 142-

143'(Found: C, 55.7;H,3.7; N, 3.6. ClsHlaBrNOa requires C, 55.8; H,3.7;

N, 3.67o). lH n.m.r. (300 MIIZ, CDCI3) ô 3.55, s, 3H, OMe; 5.52, d, J 1l.2Hz,

lI2;6.02, d, J 1l.2Hz,}{3;7.34-7.61, m, 5H, Ph; 7.79-7.97, m, 4H, Phth. Vmax

1778, 1755, 1709,708 cm-I. Mass spectrum; mlz 389 and 387 (M*', 67o),330

(-COzMe, 8), 328 (-CO2Me, 8), 308 (-Br, 65), 307 (-HBr, 18), 276 (89),249

(92),248 (100), 242 (32),24A Q2\218 (59), 190 (46), t62 (39), r3O (22),

105 (46), 103 (28), 77 (Il). Further recrystallization gave the erythro-

diastereomer (82b) (4.98 9,407o), m.p. l2l-122" (Found: C, 56.2; H, 3.7; N,

3.7. CrsHlaBrNOa requires C, 55.8; H,3.7: N, 3.67o). lH n.m.r. (300 MHz,

CDCI¡) ô 3.82, s, 3H, OMe; 5.59, d, J 10.5 Hz, H2,5.91, d, J 10.5 Hz, H3;

7 .lI-7 .37 , m, 5H, Ph; 7 .63-7 .72, m,4H, Phth. 13C n.m.r. (75.5 N.4Hz, CDCI¡)

õ 47.6, 53.1, 51.0, 123.6, 128.1, 128.6, 128.9, 130.9, 134.3, 137.r, 166.3,

167.2. v^u* 1774, 1758, 1718, 727 cm-r. Mass spectrum; mlz 389 and 387

(M*', ZVo),330 (-COzMe, 5), 328 (-CO2Me, 5), 308 (-8r,32),276 (59),249

(79),248 (85),242 (24),240 (24),2t8 (56), 190 (62), 169 (tt), 167 (11), l6l

(26), 130 (33), 104 (82), r02 (64),76 (100).

Experimental 126

(2 S 3 R) -3- B romo -N - te rt-buty¡ -¡¿ø- phthaloylp henylalaninamide (S5a)

and(2 S 3 S ) -3- B romo - N - te rt-buty¡ -¡y a-p hthaloylpheny lalaninami de (S5b )

To a solution of ( s ) -N -tert-butyl-Nø-phthaloylphenylalaninamide (78) (3.5

g, 10.0 mmol) in CCl¿/dichloromethane (3:1, 60 ml) was added NBS (1 .87 g,

10.5 mmol), and the mixture was refluxed for 2 h while irradiated with a

250W mercury lamp. The mixture was allowed to cool, then was filtered and

the filtrate was washed with water (60 rnl). The organic phase was dried, then

was concenfrated under reduced pressure to give the title compounds (85a) and

(85b) (4.17 g, 977o) as a 1:l mixture" Fractional recrystallization from

hexane/iso-propanol gave the erythro-diastereomer (85b) (1.84 g,43Vo), m.p.

208-209" (Found: C, 58,6; H, 5.0; N, 6.8. CztHztBrNzO¡ requires C, 58.8; H,

4.9; N, 6.57o). lH n.m.r. (300 MHz, CDC13) ô 1.40, s, 9H, CMe¡; 5.32, d, J

Il.4 IJz, I{2; 6.05, d, J 1I.4 Hz, H3; 6.41, br s, NH; 7.11-7.38, m, 5H, Ph;

7.62-7 .72, m, 4H, Phth" v'nu* 3350, 1700, 1530, 1450, 1365, 710 cm-l. Mass

spectrum; mlz 430 and 428 (M*', 57o), 415 (-Me, 0.2), 413 (-Me, 0.2),375

(0.6), 313 (0.6),330 (2),328 (2),249 (rW), 232 (6),220 (3),204 (5), 165 (2),

130 (3), 104 (8), I02 (7), 16 (6). Further recrystallization gave the threo-

diastereomer (85a) (I.75 g,4l%o), m.p. 188-191" (Found: C,58.8; H,4.9; N,

6.6. CztH21BrN2O3 requires C, 58.8; H, 4.9; N, 6.57o). lH n.m.r. (300 MHz,

CDCI¡) õ 1.02, s, 9H, CMq; 5.22, d, J 11.8 Hz, H2;5.76, br s, NH; 6.17, d,I11.8 Hz, H3;1.34-7.62, m,5H, Ph; l.7l-7.96, m, 4H, Phth. vn'u^ 3375 , 1J75,

I'705, 1530, 1380, 120 cm-r. Mass spectrum; mlz 430 and 428 (M*', IVo),4I5

(-Me, 0.l), 413 (-Me, 0.1), 375 (0.2), 373 (0.2), 330 (1), 328 (1), 249 (IO0),

232 (r0),220 (3),204 (4),165 (2),130 (3), 104 (r2),102 (10), 76 (13).

Experimental 127

Treatment of QS3R)- and (2535)-3-Bromo- N-tert-butyl-¡¡rø-phthaloylphenylalaninamide (85a) and (85b) with tri-n-butyltinhydride

A 1:1 mixture of the bromides (85a) and (85b) (1:1, 100 m9,0.23 mmol)

was dissolved in benzene (5 ml) under an atmosphere of nitrogen.

Tri-z-butyltin hydride (67 mg,0.23 mmol) was added, and the solution was

refluxed for 4 h. The solvent was removed under reduced pressure, then

hexane (10 ml) was added to dissolve the tri-n-butyltin bromide. The

suspension was stirred for 5 min, then the reduced compound (78) was isolated

by filtration as a white solid (71 mg,887o). Spectral characteristics of this

material were identical to those described above.

Elution of this compound through a Regis Pirkle HPLC column showed the

presence of only the (S)-enantiomer (78). This enantiomer (78) had a retention

time of 2I min, eluting with a gradient of hexane and iso-propanol (5-407o iso-

propanol).

(Z) -N - te rf-B utyl-Na-phthaloyl-ø,p-d ehydrop henylalaninamid e (86)

A solution of 18-crown-6 in acetonitrile (1.15M,25m1) was treated with

anhydrous potassium fluoride (0.11 9,2.3 mmol), and the mixture was stirred

vigorously at reflux for 30 min. The threo-bromide (85a) (0.50 g, 1.2 mmol)

was then added and the mixture was stirred at reflux for a further 30 min, after

which the mixture was allowed to cool and was then concentrated under

reduced pressure. The residue was taken up in ethyl acetate, and the suspension

was washed with water, dried, then concentrated under reduced pressure.

Recrystallization of the residue from hexane/ethyl acetate afforded the title

compound (86) as coiouriess crystals (0.36 9,847o), m.p. 2i5' (Found: C,72.6;

H, 5.8; N, 8.0. C21H2oN2O3 requires C,72.4; H, 5.8; N, 8.07o). lH n.m.r. (300

}d}Jz, CDCI3) ô 1.40, s, 9H, CMe3; 5.9J, br s, NH; 7.25, m,5H, Ph; 7.59, s,

Experimental I28

vinylic-H; 7.75, m,2H, Phth; 7.87, m,2H, Phth. v*u* 3350, 1780, 1720, 1660,

1630, 1525 cm-l. Mass spectrum; mlz 348 (M*', I07o),334 (6),321 (4),294

(67),277 (36),264(1),248 (100), 165 (8), 162 (8), 145 (5), 104 (88),76(32).

(B) -N -terf-Butyl-Nø'phthaloyl-cr,B-dehydrophenylalaninamide (87)

Treatment of the erythro-bromide (85b) with the 18-crown-6 complex of

potassium fluoride in acetonitrile, as described for the synthesis of (Z)-N-tert-

butyl-Nø-phthaloyl-a,B-dehydrophenylalaninamide (86) from the thre o-

bromide (85a), afforded a 5:l mixture of (E)- and (Z)-N-tert-búyl-Nd-

phthaloyl-o,p-dehydrophenylalaninamide (87) and (86). The isomers (86) and

(87) were separated by fractional recrystallization from a dilute solution of

hexane/ethyl acetate to give the (Z)-isomer (86) (9Vo), identical to the sample

described above, and the (E)-isomer (87) (517o), m.p.185-186o (Found: C,

12.5; H, 5.8; N, 8.0. C21H26N2O3 requires C, 72.4; H, 5.8; N, 8.07o). lH

n.m.r. (300 MHz, CDCI3) õ 1.22, s, 9H, CMe¡; 5.50, br s, NH; 7.05, s, vinylic-

H;7.40, m,5H, Ph; 7.85, m,4H, Phth. v^u*3420, lJ15, 1660, 1510 cm-l.

Mass spectrum; mlz348 (M*').

(Z)-N-Phthaloyl-ø,B-dehydrophenylalanine Methyl Ester (88)

Treatment of the threo-bromide (82a) with the 18-crown-6 complex of

potassium fluoride in acetonitrile, as described for the synthesis of (Z)-N-tert-

butyl-Nø-phthaloyl-o,B-dehydrophenylalaninamide (86) from the threo-

bromide (85a), afforded the title compound (88) as colourless crystals (887o),

m.p. 136-137o (Found: C, 70.6;H,4.2; N, 4.4. CrsHr¡NO4 requires C,70.4iH,

4.3; N, 4.67o). lH n.m.r. (300 MHz, CDCIg) õ 3.82, s,3H, OMe; 7.26-7.42,m,

5H, Ph; 7.18, m,2H, Phth; 7.90, m,2P,., Phth; 8.12, s, vinylic-H. vru* 1780,

Experimental 129

1120, 1640,1600 cm-l. Mass spectrum; mlz 307 (M*', 1007o),279 (-CO),248

(-COzMe) ,247 .

(E')-N-Phthaloyl-a,p-dehydrophenylalanine Methyl Ester (89)

Treatment of the erythro-bromide (82b) with the 18-crown-6 complex of

potassium fluoride in acetonitrile, as described for the synthesis of (Z)-N-tert-

butyl-Nø-phthaloyl-cr,B-dehydrophenylalaninamide (86) from the thre o-

bromide (85a), afforded a 2:l mixture of (Z)- and (E)-N-phthaloyl-o,0-

dehydrophenylalanine methyl ester (88) and (89). Recrystallization of the

mixture from hexane/ethyl acetate gave the (Z)-isomer (88) (34Eo). The

(E)-isomer (89) could not be obtained pure, but 5:1 mixture of the (E)- and

(Z)-isomers (89) and (88) was isolated as a clear oil. The (E)-isomer (89) had

1H n.m.r. (300 MHz, CDCI3) õ3.74, s,3H, OMe; 7.23, s, vinylic-H;7.29-7.49,

m, 5H, Ph; 7.80, m,2}J, Phth; 7.93, m,2}{, Phth" vru^ 1780, 1724, 1645,

1600, 1560 cm-l. Mass spectrum; mlz 307 (M*', LjOVo), 279 (-CO),248

(-COzMe) ,247 .

(S)'4-Bromo-N-phthaloylleucine Methyl Ester (91)

To a solution of (S)-N-phthaloylleucine methyl ester (73) (10.3 g, 37.5

mmol) in CCI¿ (150 mt) was added NBS (6.67 9,37.5 mmol), and the mixture

was refluxed for 2 h white inadiated with a 250W mercury lamp" The mixture

was allowed to cool, then was filtered and the filtrate was washed with water

(100 ml). The organic phase was dried, then was concentrated under reduced

pressure. The residue was purified by chromatography on silica to give the

bromide (91) as a viscous oil which crystallized on storage at -20o overnight.

Experimental 130

Recrystallization from ethanol/water gave the title compound (91) as colourless

crystals (10.8 E, 8l7o), m.p. 63-64" (Found: C, 50.9; H, 4.6; N, 4.0.

C15H16BrNOa requires C, 50.8; H,4.6;N,4.07o).lH n.m.r. (300 MHz, CDCI3)

õ 1.76, s,3H, Me; 1.84, s,3H, Me'; 2.85, m,2}l,CH2;3.74, s,3H, OMe; 5.25,

dd, J 4.4 FI2,8.0 Hz, }l2;7.75-7.91, m, 4H, Phth. vmax 1748, 1719, 1269,

1230, Il34,7Il cm-r. Mass spectrum; mlz 355 and 353 (M*', 0.3Vo) 354 (-H,

0.6),352 (-H, 0.6), 296 (-COzMe, 1), 294 (-COzMe, 1), 274 (-8r,2),215 (64),

214 (60),200 (74), t74 (35), 160 (100), l3O (26), 104 (38), 76 (49).

4-Bromo-N-phthaloylhomophenylalanine Methyl Ester (92)

To a solution of N-phthaloylhomophenylalanine methyl ester (77) (7.5 g,

23.2 mmol) in CCI¿ (150 ml) was added NBS (4.15 g, 23.3 mmol), and the

mixture was refluxed for 2 h while inadiated with a 250W mercury lamp" The

mixture was allowed to cool, then was filtered and the filrate was washed with

water (100 ml). The organic phase was dried and then concenffated under

reduced pressure. The residue was purified by chromatography on silica to

give an approximately 1 .2:l ratio of the diastereomers of the title compound

(92) as a viscous oil which crystallized on standing (7.88 g, 847o), m.p. 112-

118" (Found: C,56.8; H,4.0; N,3.5. C19H16BrNOa requires C, 56.1; H,4.0;

N,3.57o). lH n.m.r. (300 MHz, CDCI3) ô (one diastereomer) 3.12-3.27,m,2H,

CHz;3.75, s,3H, OMe; 4.91,t,J 7.6H2,}J4;5.21, dd, J 5.8 Hz, 9.4H2,H.2;

7.05-7.35, m, 5H, Ph; 7.73, m, 2H, Phth; '7.79, m, 2H, Phth; (other

diastereomer) ô 2.99-3.31, m, 2H, CHz; 3.73, s, 3H, OMe; 4.91, t, J 7 .6 Hz, H4;

5.10, t, J 7.5 H:z, H:2; 7.05-7.38, m, 5H, Ph; 7.76, m, 2H, Phth; 7.88, m,2H,

Phth. v,nax1774, 1744, 1716, 1264, 1216,122,704 cm-l. Mass spectrum: mlz

403 and 401 (M+', 0.57o), 402 (-H, 1), 400 (-H, l), 369 (2), 367 (2),342 (5),

Experimental l3I

340 (5), 322 (-Br, l00),262 (76),219 (68), 187 (31), 175 (15), 148 (24), 132

(33), 130 (42), tO5 (37), tO4 (57),91 (29).

(S)-O-Acetyl-N-phthaloyltyrosine Methyl Ester (93)

To a solution of (S)-N-phthaloyltyrosine methyl ester (75) (5.0 g, 15.4

mmol) in dichloromethane (20 ml) was added triethylamine (2.33 g, 23.0

mmol), acetic anhydride (2.35 g, 23.0 mmol) and DMAP (100 mg, 0.82

mmol). The solution was stirred at ambient temperature for t h, then water

(20 ml) was added, and the mixture was stirred vigorously for a further 10

min. The organic phase was separated, washed with water, dried, and then the

solvent was removed under reduced pressure. The solid residue was

recrystallized from hexane/ether to give the title compound (93) as colourless

needles (4.91 g,877o) m.p. 86-87o (Found: C, 65.3; H,4.5: N, 3.8. C29H17NO6

requires C, 65.4; H, 4.7; N, 3.87o). lH n.m.r. (300 MHz, CDCI3) õ 2.22, s, 3H,

OAc; 3.55, dd, J 10.5 Hz, 14.4 Hz,H3;3.59, dd, J 6.0 Hz, 14.4 Hz,H'3';3.78, s,

3H, OMe; 5.16, dd, J 6.0 Hz, 10.5 Hz,IÐ;6.92, d, J 8.6 Hz, 2H, ArH; 7.18, d, J

8.6 Hz, 2H, ArH; 7.68-7.80, m, 4H, Phth. v^u* 1775,1760, 1720, 1610, 1510,

1390, 1190, 915,7I5 cm-l. Mass specffum; mlz367 (M*',2Vo),325 (9),308

(-OAc, 3),266 (8), 248 (3),220 (11), 178 (100), 147 (16), 122 (71), I2l (M),

r07 (39),105 (34).

(253R)-O-Acetyl-3-bromo-N-phthaloyltyrosine Methyl Ester Qaa)and(2535)-O-Acetyl-3-bromo-N-phthaloyltyrosine Methyl Ester (94b)

To a solution of (S)-O-acetyl-N-phthaloyltyrosine methyl ester (93) (1.0 g,

2.7 mmol) in CCI+ (50 ml) was added NBS (520 mg, 2.9 mmol), and the

Experimental I32

mixture was refluxed for 2 h while irradiated with a 250W mercury lamp. The

mixture was allowed to cool, then was filtered and the filtrate was washed with

water (50 mt). The organic phase was dried, then was concentrated under

reduced pressure to give thetitle compounds (9aa) and (94b) as a 1:1 mixture

(1.18 g, 977o) (Found: y7+',mlz 445.018. CzoH168rNO6 requhes mlz

445.016). lH n.m.r. (300 MHz, CDCI3) threo-isomer (94a) õ 2.30, s, 3H, OAc;

3.56, s, 3H, OMe; 5.46, d, J II.2 Hz, fl2;6.03, d, J lI.2 Hz, H3:- J .I4, d, J 8.6

Hz, 2H, ArH; 7.6I, d, J 8.6 H:z, 2H, ArH; 7.78-7.96, m, 4H, Phth; erythro-

isomer (94b) õ2.I9, s,3H, OAc; 3.81, s,3H, OMe; 5.56, d, J 10.4 Hz,II2;

5.93, d, J 10.4 Hz,}I3;6.93, d,I 8.6H2,2H, ArH; 7.37, d,J 8.6H2,2H, ArH;

7.64-7.73, m, 4H, Phth. Mass spectrum; mlz 447 and 445 (M*', 0.37o),405

(0.3), 403 (0.3), 388 (-CO2Me, 0.6), 386 (-CO2Me, 0.6),366 (-Br, 8), 334

(r7),324 (5), 306 (5),292 (16),264 (100), 218 (r2), r87 (7), t63 (9), 147

(10), 138 (r4),tzl (35), 104 (22),86 (38),76 (19).

( 2 S 3 R) - O- A ce ty l - 3 - b r o mo - N - t e r t -buty¡ - ¡y ø- p ht hal o y I ty r o s in am i d e

(95a) and

(2 S 3 S ) - O- A ce ty l - 3 - b r o mo -N - t e rt -butyl -¡y ø- p h tha I o y lty r o s in a m id e

(esb)

To a solution of ( S )-O -acetyl-Nø-phthaloyltyrosinamide (80) (500m g, 0.82

mmol) in CCl¿/dichloromethane (4:I,20 ml) was added NBS (150 mg, 0.84

mmol), and the mixture was refluxed for 2 h while inadiated with a 250W

mercury lamp. The mixture was allowed to cool, then was filtered and the

filtrate was washed with water (20 ml). The organic phase was dried, then was

concentrated under reduced pressure to give the title compounds (95a) and

lq5h\ âs e l'l mixtrrre (57R ms 97q"\ lFound: M+'. mlz 486.080.\-_-, _ \-- _ ---ò,, -,,-/ \- __-__-'.

C1;.}J23BrNzOs requires m12486.079). t1¡ n.m.r. (300 MHz, CDCI3) threo-

isomer (9.5a) ô 1.05, s,9H, CMe3; 2.31, s,3H, OAc; 5.17, d, J ll.8Hz,H2;

Experimental 133

5.87, br s, NH; 6.22, d, J 11.8 flz,}{3; J .I5, d, J 8.6 }Jz,2}I, ArH; J.62, d, J 8.6

Hz,2IJ, ArH;7.77-7.94, m,4H, Phth; erythro-isomer (95b) ô 1.38, s,9H,

CMe3; 2.20, s, 3 H, OAc; 5.30, d, J 11.5 Hz,fI3i 6.08, d, J I1.5 Hz,I12;6.41,

br s, NH; 6.95, d, J 8.6 Hz,2H, ArH; 7.39, d, J 8.6 Hz,2H, ArH; 7.62-7.73,m,

4H, Phth. Vmax 1780, 1726, 1675, 1608, 1394, 1160, 720 cm-r. Mass

spectrum; mlz 488 and 486 (M*', 27o), 446 (l), 444 (1), 403 (14), 388 (1), 386

(1), 361 (10), 308 (3t),265 (100), 248 (5), r2t (11), 118 (10), 104 (r4),76

(e).

Treatment of (S)-3-Bromo-N-phthaloylvaline Methyl Ester (81)

with silver nitrate in acetone/water.

To a solution of the bromide (8I) (1.2 g,3.52 mmol) in acetone (50 ml)

was added a solution of silver nitrate (0.90 g, 5.3 mmol) in water (50 rnl). The

resultant mixture was stirred in the dark for 24 h, then the yellow precipiøte

was removed by filtration. The filtrate was concentrated under reduced

pressure, and the residue was extracted with dichloromethane (2 x 40ml). The

combined extracts were dried, and the solvent was removed under reduced

pressure. The products were separated by chromatography on silica and

recrystallized from hexane/dichloromethane to give the p-hydroxyvaline

derivative (100) as colourless crystals (0.42 g, 437o), m.p" 79-80" (Found: C,

60.6; H, 5.5; N, 5.1. ClaHrsNOs requires C, 60.6; H, 5.5; N, 5.17o). lH n.m.r.

(300 MHz, CDCI3) õ 1.31, s,3H, Me; 1.53, s,3H, N.4e'i3.77, s,3H, OMe; 4.41,

br s, OH; 4.9I, s, H2; 7.78-7.93, m,4H, Phth.v*u* 3544,1767,1725, 1275,

lI7 cm-r. Mass spectrum; mlz 262 (-Me, l}Vo),246 (5), 230 (28),219 (100),

188 (74), 187 (98), 160 (14)" The o,B-dehydrovaline derivative (101) was

isolated as colourless crystals (0.08 g,87o), m.p. 8l-82" (Found: C,64.7i H,

5.1; N, 5.4. Cr+Hr:NO¿ requires C, 64.8; H, 5.1; N, 5.4Vo). lH n.m.r. (300

MHz, CDCI¡) ô 1.88, s, 3H, M.e:2.43, s, 3H, Me'; 3.68, s, 3H, OMe; 7.75,m,

Experimental 134

4H, Phth. vn.,u* 1728 br, l22J,l2O cm-t. Mass spectrum; mlz 259 (M+',IVo),

227 (-MeOH, 55), 132 (87), 104 (100), 76 (43)" The p,y-dehydrovaline

derivative (I02) was isolated as a clear oil (0.30 g, 34Vo) (Found: y4+', mlz

259.086. C1aH13NO4 requires mlz 259.085). lH n.m.r (300 MHz, CDCI3) ô

t.92, s,3H, y-Me; 3.79, s,3H, OMe; 5.11, s, lH; 5.14, s, lH; 5.38, s, lH; 7.73-

7.91, m, 4H, Phth. v*u* (neat) 1780, 1748, 1728, 1470, I44O, 1386, 1293,

1113, 915,717 cm-l. Mass spectrum; mlz259 (M*', 87o),227 (-MeOH,20),

200 (-COzMe, 100).

Treatment of (S)-4-Bromo-N-phthaloylleucine Methyl Ester (91)

with silver nitrate in acetone/water.

To a solution of the bromide (91) (18.9 9,53.4 mmol) in acetone (1.5 L)

was added a solution of silver nitrate (13.5 g,79.9 mmol) in water (1.0 L)"

The resultant mixture was stirred in the dark for 24 h, then the yellow

precipitate was removed by filtration" The filtrate was concentrated under

reduced pressure, and the residue was extracted with dichloromethane (2 x

400rnl). The combined extracts were dried, and the solvent was removed under

reduced pressure to give a pale yellow oil. Fractional crystallization from

ether gave the lactone (105) as a white solid (1.16 g,\Vo), m.p. 132-133'

(Ht.l¡o 131'). lH n.m.r. (300 MHz, CDCI3) ô 1.52, s, 3H, Me; 1.64, s,3H, Me';

2.46, dd, J 9.7 }{z, 12.3 }Iz, }I4; 2.60, dd, J lI.5 Hz, 12.3 Hz, H4'; 5.25, dd, J

9.7 IJz, II.5 Hz,H3;7.75-7.88, m, 4H, Phth. Mass spectrum; mlz 260 (M+H+,

l7o),215 (-COz, 82),200 (100), 160 (72), 130 (22), 104 (34), 76 (51).

Concentration of the filtrate and purification by chromatography on silica gave

the y-hydroxyleucine derivative (104) as a clear, viscous oil which was

crystallized from hexane/clichloromethane to afford eolourless crystals (12.1 g,

787o), m.p.7l-72", ¡u1orc -25.9" (c, 0.5 in EIOH) (Found C, 61.8; H, 5.9; N,

4.8. CrsHlzNOs requires C, 61.8; H, 5.9; N, 4.8¡. t1¡ n.m.r. (300 MHz, CDCI¡)

Experimental 135

õ I.24, s, 3H, Me; 1.31, s, 3H, Me'; l.70,br s, OH; 2.38, dd, J 8.8 Hz,I5.l Hz,

H3;2.50, dd, J 4.0 Hz, 15JH:z,H3';3.J3, s,3H, OMe; 5.15, dd, J 4.0 Hz, 8.8

Hz, IJ2; 7.74, m,2H, Phth; 7.86, m,2H, Phth. v*u* 3514, 1770, 1743, 1707,

1272, 1230, 1161, 720 cm-r. Mass spectrum; mlz 276 (M+'-Me, 77o),233

(-Me2CO, l7),215 (29),2W (41), 174 (100). The y,õ- and B,y-dehydroleucine

derivatives (106) and (107) were isolated as an inseparable mixture (291 mg,

27o¡. r11n.m.r (300 MHz, CDCI3) 7,õ-dehydroleucine derivative (106) õ 1.78,

s,3H, Me; 2.86, brdd, J 4.0}J.2,I4.3H2, H3; 3.10, dd,J l2.3Hz,14.3Hz,I{3'i

3.76, s, 3H, OMe; 4.65, br s, vinylic-H; 4.69, br s, vinylic-H; 5.10, dd, J 4.0

Hz,l2.3 }{z,I{2;7.71-7.89, m, 4H, Phth: p,1-dehydroleucine derivative (107) õ

1.76, s, 3H, Me; I.77, s,3H, Me'; 3.76, s,3H, OMe; 5.66, d, J 9.2Hz,IÐ;5.82"

br d, J 9.2H2,H3;7.71-7.89, m,4H, Phth. vmax 1777,1746, I7I8, 1392,

1188, 1104 cm-l" Mass spectrum; mlz273 (M*',27o),241 (6),2I4 (-CO2Me,

100), 196 (16), 190 (8), 160 (10), 130 (19), 126 (13), r04 (24),16 (28).

Treatment of (S)-4-Bromo-N-phthaloylhomophenylalanine MethylEster (92) with silver nitrate in acetone/water.

To a solution of the bromide (92) (9.0 g, 22.4 mmol) in acetone (600 mt)

was added a solution of silver nitrate (5.67 9,33.6 mmol) in water (a00 ml).

The resultant mixture was stirred in the dark for 24 h, then the yellow

precipitate was removed by filtration. The filtrate was concentrated under

reduced pressure, and the residue was extracted with dichloromethane (2 x

250m1). The combined exfracts were dried, and the solvent was removed under

reduced pressure to give a pale yellow oil. Separation of the products by

chromatography on silica gave the y-hydroxyhomophenylalanine derivative

(109) as a 1.5:1 ratio of diastereomers, as a clear, viscous oil (6.06 g,8O7o)

(Found ¡11+', mlz 339.1I2. CeHlzNOs requires mlz 339.lII). lH n.m.r. (300

MHz, CDCI¡) ô (major isomer) 2.54-2.76,m,2H, CHz; 3.70, s,3H, OMe; 4.92,

Experimental 136

br dd, J 5.2 }lz, 6.5 Hz,}{4;5.08, dd, J 5.3 f{2,8.8 IIz, H2:7.II-7.32, m, 5H,

Ph; 7.67-7.79, m,4H, Phth; (minor isomer) õ2.54-2.16, D, 2H, CH2;3.71, s,

3H, OMe;4.57, br dd, J 4.2 Hz, 8.8 }lz,}I4;5.25, dd, J 4.7 Hz, l0.4Hz,H2;

7.ll-7.32, m,5H, Ph;7.7I-7.85, m,4H, Phth. vn'u* 3600,3480, 17'74, 1752,

1719, 1614, 1426, 1280, 1184, 1118, 918,719 cm-l. Mass spectrum: mlz 339

(M*', 37o),338 (4),322 (-OH, 65), 308 (-OMe, l5),262 (16),219 (100), 187

(36), 174 (54), 160 (20), 148 (r3)" r32 (19), 1 16 (26), 105 (36), 77 (r7).

Further recystallization gave the cis-lactone (110) (198 mg, 3Vo), m.p. 140-

143" (Found M+H+, mlz 308.092. C13H1¿NO+ requires mlz 308.092¡. r¡¡ n.m.r.

(300 MHz, CDCI3) õ2.65, ddd, J 3.3H2,9.8H2, 13.I}{z,H4;3.l2,ddd,J 8.7

Hz,9.4Hz,13.L}Iz, H4'; 5.1'l,dd,J 9.4Hu9.8Hz,, H5;5.93, dd, J 3.3 Hz,8.J

Hz, IJ3; 7.33-7.54, m, 5H, Ph; 7.74-7.90, m, 4H, Phth. v'nu* 3005, 1790, 1780,

1730, 1720, 1395 cm-l. Mass spectrum; mlz 308 (M+H+), 245,239, 148, 116

(I007o). Further recrstallization gave the trans-lactone (1ll) (129 mg,2Vo),

m.p. 143-144'(Found C, 70.2;H,4.2; N, 4.6. CrsHr:NO4 requires C,70.4;H,

4.3; N,4.6¡. tg n.m.r. (300 MHz, CDCI3) ô 2.85, ddd, J l0.6Hz,l2.OIJz,l2.5

Hz,IJ4;2.93, ddd,J 6.2H2,9.2H2, I2.5Hz,}l4';5.31, dd, J 9.2H2, I2.0Hz,

H5; 5.50, dd, 6.2 Hz, 10.6 Hz, H3; 1.33-7.56, m, 5H, Ph; 7.77-7.93, m, 4H,

Phth. vr* 3000,1795,1780, 1730,1390, 1095 cm-I. Mass spectrum; mlz307

(M*.), 245, I48,116 $mEo).

(253R) -3-Hydroxy-N-phthaloylphenylalanine Methyl Ester (ll2a)and(2535)-3-Hydroxy-N-phthaloylphenylalanine Methyl Ester (112b)

To a solution of a 1:1 mixture of the bromides (82a) and (82b) (I.09,2.6

mmol) in acetone (40 nìl) was added a solution of silver nitrate (660 mg, 3.9

mmol) in water (40 ml), and the resultant mixture was stirred in the dark for

24 h. Tþe precipitate was removed by filtration and the filtrate was

Experimental 137

concentrated under reduced pressure. The residue was extracted with

dichloromethane, the organic phase was dried, and then the solvent was

removed under reduced pressure to give a crude mixture of the (253R)- and

(2535)-3-hydroxyphenylalanine derivatives (112a) and (lI2b) in a 5:1 ratio

(769 mg, 92Eo)" Recrystallization from hexane/dichloromethane gave the

(253R)-þ-hydroxyphenylalanine derivative (l l2a) as colourless crystals (633

mg,75Vo), m.p. 185-186", [o]o16 -67.0" (c, 0.6 in EtOH) (Found: C, 66.7i H,

4.7; N, 4.4. ClsHrsNOs requires C, 66.5; H, 4.7; N, 4.3Vo). lH n.m.r. (300

MHz, CDCI¡) ô 3.86, s,3H, OMe; 5.13, d, J 10.4 Hz, OH; 5.51, d, J 4.6H2,ÍÐ;

5.71, dd, J 4.6 IJ.z, 10.4 }lz, Il3: I .14-l .34, m, 5H, Ph; 7 .69-1.80, m, 4H, Phth.

Vmax 3580, 3420 br, 1780, 1745" 1715, 1390, 1215, 1020, 120 cm-r. Mass

spectrum (FAB); mlz 326 (M+H+, 77Vo),308 (-HzO, 100), 248 (54),219 (19),

t60 (27), 149 (23), r3t (21),105 (48), 104 (33), 9t (42)"

Elution of the mother liquor through a reverse-phase HPLC column gave

the (253R)- and (2535)-hydroxyphenylalanine derivatives (112a) and (112b), at

retention times of 10.5 and 11 min, respectively. The (2S3S)-isomer (112b)

was collected from repeated injections (ca" 5 mg) m.p" 110-111'. lH n.m.r"

(300 MHz, CDCI3) E 3.72, s, 3H, OMe; 4.2J , d, I 2.3 Hz, OH; 4.95, d, I 8.4 Hz,

II2; 5.46, dd, J 2.3 Hz, 8.4 }{z, H3; 7 .ll-l .21 , m, 5H, Ph; 7 .60-7 .70, m, 4H,

Phth.

(S)-N-Phthaloylphenylalanine tert-Butyl Ester (114)

To a solution of (S)-N-phthaloylphenylalanine (70) (l .0 g,3.2 mmol) in

dichloromethane (25 ml) was added DCC (730 mg, 3.5 mmol), /¿rl-butanol

(260 mg,3.5 mmol) and DMAP (39 mg, 0.32 mmol). The solution was stirred

at room temperature for 16 h, then the dicyclohexylurea was removed by

filtration. filfrate was washed with 57o acetic acid solution (2 x 25 ml),

Experimental 138

and water (2 x 25 ml). The organic phase was dried, then the solvent was

removed under reduced pressure to give the crude tert-butyl ester (114) as a

viscous oil. The crude product was purified by chromatography on silica,

yielding the ester (114) as a clear oil which solidified on standing.

Recrystallization from hexane/ether gave the title compound (114) as large

colourless crystals (0.85 g, 75Vo), m.p. 95-96" (Found ¡4+', mlz 351.148.

CzrHzrNO4 requires mlz 351.147)" lH n.m.r. (300 MHz, CDCI3) ô 1.45, s, 9H,

CMe¡; 3.51, dd, J 10.7 }lz, l4.3}Jz,H3i 3.55, dd, J 6.0 I{z,l4.3Hz,H3;5.07,

dd, J 6.0 }lz, 10.7 Hz,IÐ;7.I0-7.2I, m, 5H, Ph;7.66-7.79, m,4H, Phth. Vmax

1780, I745, 1720,1390, 710 cm-1. Mass specffum; mlz 352 (M+H+, 27o),35I

(I), 296 (29), 295 (53), 250 (-COzCMq,73),232 (28), 148 (100), I3O (12),

104 (14), 91 (8), 77 (9),57 (25).

(253R)-3-Bromo-N-phthaloylphenylalanine tert-Butyl Ester (115a)

and(2535)-3-Bromo-N-phthaloylphenylalanine tert-Butyl Ester (115b)

To a solution of (S)-N-phthaloylphenylalanine tert-butyl ester (114) (7W

mg, 2.0 mmol) in CCI¿ (50 ml) was added NBS (355 mg,2.0 mmol), and the

mixture was refluxed for 2 h while irradiated with a 250W mercury lamp. The

mixture was allowed to cool, then was filtered and the filtrate was washed with

water (50 ml). The organic phase was dried, then was concentrated under

reduced pressure to give the title compounds (115a) and (115b) (858 mg,

l00%o) as a 1:1 mixture, as a pale yellow oil which partially crystallized on

standing overnight. Fractional recystallization from hexane/dichloromethane

gave the threo-díastereomer (115a) (348 mg, 4l7o), m.p. 152-153"" lH n.m.r.

(300 MHz, CDCI3) ô l 16, s, 9H, CMe:; 5.4J, d, J 11 .4 H2,H2;6.06, d. J Il.4

Hz,H3;7.34-7.59, m, 5H, Ph;7.77-7.96, m,4H, Phth. v*u^ 1780,1J38, 1120,

1420, 1390, 1050, 840, 110 cm-l. Mass spectrum; mlz 431 and 429 1N'{+',

Experimental 139

.02Vo),375 (9),373 (9),330 (-COzCMe3, lI),328 (-COzCMe3, 11),294 (4),

216 (2),249 (93),248 (94),232 (r5),220 (lO),204 (44), 165 (7), 130 (8), 104

(38), 102 (32),76 (36), 57 (100). Further recrystallization gave the erythro-

diastereomer (115b) (337 mg,397o), D.P. 118-119". lH n.m.r. (300 MHz,

CDCI¡) õ 1.48, s, 9H, CMe3; 5.49, d, J 10.4 Hz,}f2;5.85, d, J 10.4 }l.z,fI3i

7.10-7.36, m, 5H, Ph;7.62-7.72, m,4H, Phth. vru^ 1780, l'748, 1726, 1390,

715 cm-l. Mass spectrum; mlz43l and 429 (M+',.017o),315 (7),373 (1),330

(-COzCMe3, 10), 328 (-COzCMe3, l0),294 (6),276 (3),249 (75),248 (100)'

232 (tt),220 (8), 204 (28), 165 (5), 130 (6), 104 (27), r02 (25),76 (26), 51

(66).

(253R) -3-Hydroxy-N-phthatoylphenylalanine tert-Butyl Ester

(116a) and (253S) -3 - Hydroxy -N-phthaloylphenylalanine tert-Butyl

Ester (116b)

To a solution of a 1:1 mixture of the bromides (115a) and (115b) (430 mg,

1.0 mmol) in acetone(12 ml) was added a solution of silver nitrate (255 mg,

1.5 mmol) in water (8 mt), and the resultant mixture was stined in the dark

for 24 h" The precipitate was removed by filtration and the filtrate was

concentrated under reduced pressure. The residue was extracted with

dichloromethane, the organic phase was dried, and then the solvent was

removed under reduced pressure to give a crude mixture of the (253R)- and

(2535)-hydroxyphenylalanine derivatives (116a) and (116b) in an 8:1 ratio.

Recrystallization from hexane/dichloromethane gave the threo-B-

hydroxyphenylalanine derivative (116a) as colourless crystals (297 mg,8I7o),

m.p. 125-127" (Found: C,68.7; H,5.9; N, 3.8. CzrHztNOs requires C, 68.7; H,

5.8; N, 3.87o). lH n.m.r. (300 MHz, CDC13) ô 1.52, s, 9H, CMe¡; 5.25, d, J

lO.6 ldz, OIgr; 5.44, d, J 4.6 Hrz, H2;5.67, dd, J 4.6 Hz, 10.6 Hz, H3; I .15-7 .36,

Experimental l4O

m, 5H, Ph;7.67-7.79, m,4H, Phth. v'nu* 3440, 1785, 1738, 1712, 1604, 1552,

1392, 1194, 1108, 844, 120 cm-1. Mass spectrum (FAB); mlz 368 (M+H+,

87o),312 (44),294 (-MqCOH, 55),216 (23),266 (-COzCMe3, 10), 250 (100),

232 (tt),205 (13), 160 (19), 105 (2t), 9r (20),87 (25), 51 (93). The lH

n.m.r. spectrum of the crude product showed peaks attributable to the erythro-

isomer (116b); õ 1.28, s, 9H, CMe¡; 4.60, br s, OH; 4.95, d, J 8.0 Hz,IÐ;5.49,

d, J 8.0 IJz,H3;7.I5-7.36, m,5H, Ph;7.67-7.79,m,4H, Phth.

(2 S 3 R) -3- Hy droxy - N -te rt-butyl-N a-phthaloylp heny lalaninamid e

(118a)

To a solution of a 1:l mixture of the bromides (85a) and (85b) (500 mg,

1.17 mmol) in acetone (20 ml) was added a solution of silver nitrate (300 mg,

1.76 mmol) in water (20 ml). The mixture was stirred in the dark for 24 h,

then was filtered through celite and the filnate was concentrated under reduced

pressure. To the residue was added dichloromethane (50 ml) and water (30

ml), the organic phase was then dried, and the solvent was removed under

reduced pressure. The crude product was then recystallized from

hexane/dichloromethane to give the title compound (118a) as colourless crystals

(398 mg,937o), m.p. 195-197" (Found: C,68.8; H,6.0; N,7.7. CztHzzNzO¿

requires C, 68.8; H, 6.1 ; N,7 .77o). lH n.m.r. (300 lvftlz, CDCI3) õ 1.30, s, 9H,

CMer; 4.41,d,J 8.2 Hz, OH; 5.11, d, J 6.2H2,H2;5.63,dd,16.2H2,8.2H2,

H3; 5.95, br s, NH; 7.20-7.38, m, 5H, Ph;7.68-1.81, m, 4H, Phth. Vmax 3352,

3275, 1778, llÙ4, 1644,717 cm-r. Mass specrum (FAB); mlz 361 (M+H+,

68Vo),307 (3), 289 (2),266 (4),260 (-PhCHO, 18),250 (100)' 232 (1), 187

(10), 160 (17), 154 (24), 136 (18), 107 (8), 105 (6),11 (6). In the lH n.m.r.

spectrum of the crude product, the l3C sateiiites of the tert-butyl signal of the

threo-alcohot (118a) were observed at õ 1.08 and 1.51 (J 126.7 Hz), but no

Experimental l4l

peak atüibutable to the tert-butyl group of the erythro-alcohol (118b) (ô 1.15)

was observed.

N - te rt-Butyl- 3 - oxo -¡¡r ø-phthaloylp henylalaninamide ( 1 19)

Jones reagen¡l34 was added dropwise to a solution of the threo-alcohol

(118a) (500 mg, 1.37 mmol) in acetone (20 ml) with vigorous stirring until the

red/brown colour persisited (c¿. 0.5 ml). The reaction was stirred for a

further 15 min, then the solvent was removed under reduced pressure, and the

residue was partitioned between dichloromethane and water. The organic layer

was washed with lOTo Na2CO3, then with water, and was dried. The solvent

was removed under reduced pressure and the residue was eluted through a

short silica column with ethyl acetate. The appropriate fractions were

combined and the solvent was removed under reduced pressure.

Recrystallization of the residue from hexane/ethyl acetate gave the title

compound (1 19) as a white solid (332 mg, 677o)" lH n.m.r. (300 MHz, CDCI:)

õ 1.34, s, 9H, CMe3; 6.04, s, H2; 7.21-1.56, m,5H, Ph;7.73-7.90, m,4H,

Phth. Mass spectrum; mlz 364 (M*', 27o),308 (l),291 (2),265 40),221 (6),

147 (PhthNH,72), 105 (PhCO, 43), 104 (87), 103 (58), 76 (100)"

(2 S 3 S) -3 -Hy droxy - N -te rt-butyl -Nø- p hthaloy lp h eny lalanin amid e

(1 18b) and (2S3R) -3-Hydroxy -N-f ert-butyl-¡¡rø-phthaloylphenylalaninamide (118a)

To a solution of the ketone (119) (100 mg,0.27 mmol) in ethanol (5 ml)

was added NaBtIa (10 mg, 0.26 mmol). The mixture was stirred at room

temperature for 10 min, then was quenched by the addition of dilute HCI (ca. I

rrìl). The solvent was removed under reduced pressure, and the residue was

Experimental 142

partitioned between dichloromethane and water. The organic layer was washed

with I07o Na2CO3, then with water, then was dried and the solvent was

removed under reduced pressure" Purification of the mixture by

chromatography on silica gave a, mixture of the threo- and erythro-alcohols

(118a) and (118b) in a 1.2:l ratio (84 mg, 847o). lH n.m.r. (300 MIIZ, CDCI3)

erythro-alcohol (118b) ô 1.15, s,9H, CMe¡; 4.25,d,J 4.9 Hz, OH; 4.60,d,J

8.3 Hz,lÐ; 5.39, dd, J 4.9 H2,8.3 Hz,H3:.1.25-7.54, m, 5H, Ph; 7.68-7.84, m,

4H, Phth. lH n.m.r. specfral data of the threo-isomer (118a) identical to that

described above.

(253R)-O-Acetyl-3-hydroxy-N-phthaloyltyrosine Methyl Ester(lzla) and (2S3S) -O - A cetyl -3 - hy droxy -N- p hthaloyltyrosine MethylEster (121b)

To a solution of a 1:1 mixture of the bromides (9aa) and (94b) (1.18 g,

2.65 mmol) in acetone (40 ml) was added a solution of silver nitrate (680 mg,

4.0 mmol) in water (40 ml), and the resultant mixture was stirred in the dark

for 24 h" The mixture was then filtered, and the filtrate was concentrated

under reduced pressure. The residue was extracted with dichloromethane, the

organic phase was washed with water, dried, and then the solvent was removed

under reduced pressure to give a crude mixture of the hydroxytyresine

derivatives (121a) and (12lb) in a 6:l ratio (870 mg, 867o) (Found: M+'-HzO,

mlz 365.090. CzoHrsNOo requires m12365.090). lH n.m.r. (300 MHz, CDCI3)

threo-ísomer (121a) õ 2.19, s, 3H, OAc; 3.J8, s, 3H, OMe; 4.98, br s, OH;

5.48, d, J 5.0 Hz,IÐ;5.67, d, J 5.0 Hz, H3; 7.00, d, J 8.6 Hz,2H, ArH; 7.38, d,

J 8.6 IJz,2H, ArH; 7.64-7.17, m,4H, Phth; erythro-isomer (12lb) ô 2.17, s,

3H, OAc; 3.74,s, 3H, OMe; 4.98, br s, OH; 5.02,d, J 8.1 IIz,ÍI2; 5.54, d, J

8.1 Hz, H3;6.94, d, J 8.7 Hz,2H, ArH; J.34, d, J 8.7 Hz,2H, ArH; 7.64-1.77,

m,4H, Phth. v.¿^ 3580,3410, 1775, l'752, ll12, 1552, 1392, 1184, 1118,

Experimental I43

852,710 cm-l. Mass spectrum (FAB); mlz 384 (M+H+, l37o),366 (-HzO,79),

334 (100), 324 (-OAc, 7), 306 (35), 292 (38), 264 (58), 219 (34), 187 (23),

160 (r5), t54 (37), 136 (33), r07 (t7), 105 (13), t04 (12),89 (17), 77 (t9).

The crude mixture of the alcohols (l2la) and (121b) was found to decompose

slowly on standing and was therefore de-acetylated without purification.

(253R)-3-Hydroxy-N-phthaloyltyrosine Methyl Ester (L22a) and

(2535)-3-Hydroxy-N-phthaloyltyrosine Methyl Ester (122b)

A crude mixture of the acetylated hydroxytyrosine derivatives (121a) and

(121b) (870 mg, 2.27 mmol) was dissolved in MeO[VH2O (20:1,20 ml) and

p-toluenesulfonic acid monohydrate (100 mg 0.5 mmol) was added. The

solution was stirred at room temperature for 6 h, then the solvent was removed

under reduced pressure. The residue was partitioned between ethyl acetate and

water, the organic phase was washed twice with l07o Na2CO3, dried, and then

the solvent was removed under reduced pressure to give the hydroxytyrosine

derivatives (I22a) and (122b) in a 6:1 ratio (741 mg 967o)" Recrystallization of

the crude product from hexane/ethyl acetate gave the (253R)-p-hydroxy-

tyrosine derivative (I22a) as a white solid (556 mg, 727o), m.p. 200-201",

[cr]o16 -70.7" (c, 0.4 in EtOH) (Found: C,63.3;H,4.4; N, 4.0. CrsHrsNOo

requires C,63.3;H,4.4; N, 4.17o). L11n.m.r. (300 MHz, CDCI3) ô 3.85, s, 3H,

OMe; 4.76, s, Ar-OH; 5.02, d, J 10.3 Hz,3-OH;5.44, d, J 4.8 Hz,}l2;5.64,dd,

J 4.8 Hz, lO.3 Hz,Il3; 6.11, d, J 8.6 }Jz,2}l, ArH; 7.I9, d, J 8.6 Hz,2H, ArH;

7.70-7.81, m, 4H, Phth. v*a* 3582,3405, 1775, ll50, 1714,1516, 1392, lll2,

850, 715 cm-l. Mass spectrum (FAB); mlz 342 (M+H+, 27o),324 (-H20, 10),

307 (r2),292 (ll),289 (1),264 (5),232 (r4),231 (17), 219 (9), 154 (100),

r37 (63), 136 (77),107 (30), 105 (12), 89 (29),77 (3t).

Experimental I44

Elution of the mother liquor through a reverse-phase HPLC column gave

the (253R)- and (2535)-hydroxytyrosine derivatives (122a) and (122b), at

retention times of 10.5 and 10 min, respectively. The (2,S3S)-isomer (122b)

was collected from repeated injections (ca. lmg). lH n.m.r. (300 MHz, CDCI3)

õ 3.78, s, 3H, OMe; 4.27, d, I 2.1 Hz, 3-OH; 4.84, br s, Ar-OH; 4.97, d, J 8.6

Hz, H2; 5.49, dd, J 2.1 Hz, 8.6 }{z, H3; 6.65, d, J 8.5 Hz, 2H, ArH; 7 .2O, d, I8.5 Hz, 2H, ArH; 7.67-7.77, m, 4H, Phth.

(2 S 3 R) -O-Acetyl-3 -hydroxy -N -te rt-butyl-Nø-phthaloyltyrosinamid e

(r23)

To a solution of a 1:1 mixture of the bromides (95a) and (95b) (200 mg,

0.41 mmol) in acetone (10 ml) was added a solution of silver nitrate (105 mg,

0.62 mmol) in water (10m1). The resultant mixture was stirred in the dark for

24 h, then the filtered and the filtrate was concenhated under reduced pressure.

The residue extracted with dichloromethane, the organic phase was dried, then

the solvent was removed under reduced pressure. Recrystallization from

hexane/dichloromethane afforded the title compound (I23) as colourless

crystals (153 mg, 887o), m.p. 202-204" (Found: M+'-HzO, mlz 406.154"

CztHzzNzO5 requires m\2,406.153). lH n.m.r. (300 MHz, CDCI3) õ 1.31, s,

9H, CMe3;2.26, s, 3H, OAc; 4.43, d, J 8.1 Hz, OH; 5.08, d, J 6.3 Hz,}l2:5.63,

dd, J 6.3 Hz, 8. I }Iz, H3; 5.95, br s, NH; 7 .03, d, J 8.6 Hz, 2}{, ArH; J .40, d, J

8.6H2,2H, ArH; 7.69-1.82, m,4H, Phth. v.u* 3600,3440,3390, 1775,l7l0,

1685, 1515, 1390, 1220, 860, 720 cm-I. Mass spectrum (FAB); mlz 425

(M+H+, 437o),308 (67), 266 (100),248 (4), 187 (16), 160 (26), 154 (23), 136

(37), t07 (18), 105 (10), 89 (r7),77 (22).

Experimental 145

(2 S 3 R) -3- Hy d rox y - N - te rt-b utyl -N ø- p hthaloy ltyrosinami d e (124)

The hydroxytyrosine derivative (I23) (90 mg, 0.21 mmol) was dissolved

in MeOHÆI2O (10:1, 10 ml). p-Toluenesulfonic acid monohydrate (10 mg, 0.05

mmol) was added, and the solution was stirred at 50o for 16 h. The solvent was

removed under reduced pressure, and the residue was partitioned between ethyl

acetate and water. The organic phase was dried, then the solvent was removed

under reduced pressure. Recrystallization of the crude product from

hexane/chloroform gave the title compound (124) as a white solid (73 mg,

937o), m.p. 214-215' (Found: C, 65.7: H,5.8; N, 7 .2. CztHzzNzOs requires C,

66.0; H, 5.8; N, 7.3Vo).1H n.m.r. (300 MHz, CDCI3) õ 1.21, s, 9H, CMe3; 4.72,

d, J 7 .5 Hz, 3-OH; 5.1 1, d, J 6.9 Hz, H2; 5.62, dd, J 6.9 Hz, '7 .5 Hz, H3; 6.4I,

br s, NH; 6.79, d, J 8.5 Hz,2IJ, ArH; 7.24,d, J 8.5 Hz,2H, ArH;7.J1-7.82,m,

4H, Phth; 7.97, s, Ar-OH. vma* 3590,3546, 3410, 1772, 1712, 1685, 1366,

855, 717 cm-l. Mass spectrum (FAB); mlz 383 (M+H+, I87o),307 (3),289

(-CoH+OH, 4),267 (48), 266 (100), 260 (16), 187 (15), 160 (15), 154 (26), 136

(17), t07 (8), 105 (8), 91 (10), 77 (9).

(25 3 R) -3-Hydroxyphenylalanine (25a)

Method 1

(2 S 3 R ) -3 -Hydroxy-N-phthaloylphenylalanine methyl ester ( I l2a) (250 mg,

0.77 mmol) was dissolved in a2:I mixture of 6N HCI and acetic acid (10 ml).

The solution was refluxed for 5 h, then was cooled and evaporated to dryness

under reduced pressure. The residue was taken up in water (10 ml) and the

suspension was filtered to remove phthalic acid. The filnate was evaporated to

dryness under reduced pressure, then the residue was dissolved in ethanol (7

ml) and a solution of aniline (0.7 ml) in dichloromethane (10 ml) was added.

The solutien was stored at 0o overnight, and the title compound (25a) was

Experimental 146

isolated as a white powder (129 mg,93Vo), m.p. 192-195", [o]o16 -49.7+O.5"

(c, 0.4 in 6N HCI) (lit.t¡s [a]p2o -50.2+2" (c,2 ín 6N HCI)). lH n.m.r. (300

MHz, DzO) ô 3.95, d, J 4.4 Hz, H2; 5.29, d, J 4.4 }lz, }{3; J.47, s, 5H, Ph.

Spectral data consistent with literature uu1uss.175

Method 2

To a solution of (253R)-3-hydroxy-N-phthaloylphenylalanine methyl ester

(ll2a) (250 mg, 0.77 mmol) in ethanol (5 ml) was added a solution of

hydrazine hydrate in ethanol (2M,0.77 mI). The mixture was refluxed for 2 h,

then was cooled and lÙVo aqueous HCI (1 rnl) was added" The mixture was

refluxed for a further t h, then was cooled and filtered. The white solid was

washed with ethanol, then the filtrate and washings were combined and

evaporated to dryness under reduced pressure. The residue was dissolved in

ethanol (7 ml) and a solution of aniline (0.7 ml) in dichloromethane (10 ml)

was added. The solution was stored at 0o overnight, and the title compound

(25a) was isolated as a white powder (111 mg, 807o). Spectral data identical to

that described above.

Method 3

The hydroxyphenylalaninamide derivative (118a) (500 mg, 1.37 mmol)

was hydrolyzedin a2:1 mixture of 6N HCI and acetic acid, as described for the

hydrolysis of the alcohol (ll2a), and gave the title compound (25a) as a fine

white powder (194 mg, 787o). Spectral data identical to that described above.

Experímental I47

(R)-N-Phthaloyl-p-nitrophenylalanin e (126)

A mixture of (R)-p-nitrophenylalanine monohydrate (2.5 g, 11.0 mmol)

and finely ground phthalic anhydride (1.63 g, 11.0 mmol) was heated in an oil

bath at 140o for 30 min. The resultant yellow gum was dissolved in hot

methanol (30 ml), filtered, and then water (20 ml) was added. The product was

isolated by filtration and dried over P2O5 to give the title compound (126) as a

pale yellow crystalline solid (3.01 g,8I7o), m.p. 206-207" (Iit.t76 204.7o). tH

n.m.r. (300 MHz, CDCI3) ô 3.69, dd, J 10.9 Hz, 14.4 Hz, H3; 3.75, dd, J 5.7

H4 I4.4 Hz, H3'; 5.26, dd, I 5.7 IJz, 10.9 Hz,IÐ; 7 "48, d, J 8.7 Hz,2H, ArH;

7.80, s,4H, Phth; 8.07, d, J 8.7 IJz,2H, ArH"

(R)-N-Phthaloyl-p-nitrophenylalanine Methyl Ester (L27)

(R)-N-Phthaloyl-p-nitrophenylalanine (126) (2.5 g, 7.35 mmol) was

dissolved in dry methanol (50 ml) which had been pre-treated with thionyl

chloride (400 mg, 3.36 mmol). The solution was stirred under anhydrous

conditions for 16 h, then the solvent was removed under reduced pressure. The

residue was taken up in dichloromethane, and was washed with l07o Na2CO3

and water, then was dried" Removal of the solvent under reduced pressure and

recystallization of the product from hexane/dichloromethane gave the title

compound (127) as a white solid (2.24 g,867o), h.p. l2l-122" (Found M+',

mlz 354.087. CraHr¿NzOo requires mlz 354.085¡. tg n.m.r. (300 MHz, CDCI¡)

ô 3.71, dd, J lO.9 Hz, I4.3 }lz, H.3; 3.77, dd, J 5.5 Ilz, 14.3 Hz, H3'; 3.81, s,

3H, OMe; 5.31, dd, J 5.5Il.2, 10.9H2,}I2;7.45, d, J 8.6 Hz,2}l, ArH; 7.12-

7.82, m,4H, Phth; 8.06, d, J 8.6 Hz,2H, ArH. v^u* 1775,I750, 1715, 1600,

1520, 1390,1345, t240, 860,120 cm-1. Mass spectrum; mlz 354 (M*', I27o),

295 (-COzMe,37),278(14),218 (36),207 (100), 190 (37), ll6 (25), 130 (33),

104 (n),16 (2r).

Experimental 148

(2R35)-3-Bromo-N-phthaloyl-p-nitrophenylalanine Methyl Ester(128a) and

(2R3 R) -3-Bromo-N-phthaloyl-p-nitrophenylalanine Methyl Ester(128b)

To a solution of (R)-N-phthaloyl-p-nitrophenylalanine methyl ester (I27)

(2.20 9,6.24 mmol) in CCI¿ (40 mt) was added NBS (1.20 9,6.74 mmol), and

the mixture was refluxed for 4 h while irradiated with a 250W mercury lamp.

The mixture was allowed to cool, then was filtered and the filtrate was washed

with water (40 ml). The organic phase was dried, then was concentrated under

reduced pressure to give the title compounds (128a) and (128b) (2.69 g, 1007o)

as a 1:1 mixture. Fractional recrystallization from hexane/dichloromethane

gave the threo-diastereomer (128a) (1.11 g, 447o), m.p. 198-201" (Found: C,

49.8; H, 3.0; N, 6.5. ClgHl3BrNzOo requires C, 49.9; H, 3.0; N, 6.57o). lH

n.m.r. (300 MHz, CDCI¡) ô 3.59, s, 3H, OMe; 5.51, d, J 1l.2Hz,ÍÐ; 6.02, d, J

II.2Hz,}I3;1.78, d, J 8.8 Hz,2H, ArH; 7.82-7.99, m,4H, Phth; 8.27, d, J 8.8

Hz, 2H, ArH. v^a* I77 5, 1750, 1720, 1600, 1525, 1340, 1215, 1 100, 820, 7 15

cm-l. Mass spectrum; mlz 434 and 432 (M*', 27o),375 (-CO2Me, 6), 373

(-CO2Me, 6), 353 (-Br, 4), 352 (9),321 (6),294 (29),293 (11),287 (IO),285

(r0),247 (7),2t9 (16), 218 (100), 190 (30), 130 (18), 104 (40),76 (37).

Further recrystallization gave the erythro-diastereomer (128b) (1.07 g, 407o),

m.p. 195-197' (Found: C, 49.8; H, 3.0; N, 6.6. ClsHl3BrNzOe requires C,

49.9; H, 3.0; N, 6.57o). 1H n.m.r. (300 MHz, CDCI3) ô 3.83, s, 3H, OMe; 5.59,

d, J 10.3 I{z,H2; 5.9J, d, J 10.3 }lz,}l3;7.56, d, J 8.7 H¿2H, ArH; 7 .68-1.76,

m, 4H, Phth; 8.07, d, J 8.7 }Jz,2IJ, ArH. v^u* 1775, L755, 1720, 1605, 1525,

1390, 1350, 855,720 cm-l. Mass spectrum; mlz 434 and 432 (M*', l7o),375

(-COzMe, 3), 373 (-CO2Me, 3), 353 (-Br, 6), 352 (3), 321 (7), 294 (20),293

(r2),287 (3), 285 (3),247 (5),219 (15), 218 (100), t90 (29),130 (16), 104

(28),76 (26).

Experimental 149

Treatmen t of (2 R3S) -3 - B rom o -N- p hthaloy l-p-nitrop henylalanine

Methyl Ester (128a) with silver nitrate in acetone/water.

To a solution of the threo-bromide (128a) (50 mg, 0.115 mmol) in acetone

(3 mt) was added a solution of silver nitrate (25 mg 0.15 mmol) in water (2

rnl). The resultant mixture was stirred at 50o in the dark for 16 h. The mixture

was then filtered and the fitrate was concentrated under reduced pressure. The

residue extracted with dichloromethane, the organic phase was dried, then the

solvent was removed under reduced pressure. Recrystallization of the residue

from hexane/dichloromethane gave the (Z)-p-nitro-dehydrophenylalanine

derivative (I32) as large prisms (34 mg, 84Vo), m.p. 133-134' (Found; I\z[+',

mlz 352.068. CrsHrzNzOo requires mlz 352.070¡. t1¡ n.m.r. (300 MHz, CDCI3)

ô3.87, s,3H, OMe; 7.55, d, J 8.8 }{z,2H, ArH; 7.83,m,2}l, Phth; 7-92,m,2}{,

Phth; 8.13, s, vinylic-H; 8.16, d, J 8.8 Hz, ArH. vru* 1780,1720,1600, 1530,

1345 cm-l. Mass spectrum; mlz352 (M*', 90Vo),324 (63),293 (-COzMe, 41),

292 (46),247 (24),218 (15), 190 (18), 166 (2r),104 (100),76 (73).

Treatment of (2R3R)-3-Bromo-N-phthaloyl-p-nitrophenylalanineMethyl Ester (12Sb) with silver nitrate in acetone/water.

To a solution of the erythro-bromide (128b) (50 mg, 0.115 mmol) in

acetone (3 ml) was added a solution of silver nitrate (25 mg,0.15 mmol) in

water (2 mt). The resultant mixture was stirred at 50" in the dark for 16 h.

The mixture was then filtered and the filtrate was concentrated under reduced

pressure. The residue extracted with dichloromethane, the organic phase was

dried, then the solvent was removed under reduced pressure. Separation of the

products by chromatography on silica gave the B-hydroxy-p-nirophenylalanine

derivative (129) which was recrystallized from hexane/dichloromethane and

obtained,:t .otou.less needles (27 mg, 637o), m.p. 183-185". lH n.m'r- (300

Experimental 150

MHz, CDCI¡) ô 3.89, s, 3H, OMe; 5.34, d, J 10.0 Hz, OH; 5.53, d, J 4.4 Hz,

IÐ; 5.79, dd, J 4.4 Hz, 10.0 Hz, }I3;7 .54, d, J 8.9 Hz,2H, ArH; 7 .73-7.82, m,

4H, Phth; 8.13, d, J 8.9 Hz,2H ArH. v,nu* 3604,3421, 1779, I752, 1714,

1614, 1526, 1392, 1352, 1182 cm-I. Mass spectrum (FAB); mlz 371 (M+H+,

97o), 353 (-HzO, 3),321 (3),301 (11), 289 (9),219 (3), 154 (100), 137 (66),

136 (79), 107 (28),89 (33),77 (31). A mixture of the (Z)- and

(E)-dehydrophenylalanine derivatives (132) and (133) was obtained as a viscous

oil (3:2, 257o)" lH n.m.r. (300 MHz, CDCI3) (E)-isomer (133) õ 3.72, s, 3H,

OMe; 7.28, s, vinylic-H; 7.60, d, J 8.7 }Jz, 2}J, ArH; 7 .80-1.98, m, 4H, Phth;

8.26, d,I 8.7 }{z,2H, ArH. Spectral data for the (Z)-isomer (132) identical to

that described above.

(3 S4S)-l-tert-Butyl-4-phenyl-3-phthalimidoazetidin-2-one (134) and(Z) -N - te rf- B utyl -N ø-p hthaloyl - cr, B -d ehy drop henylalaninamid e (8 6)

Potassium amide solution was prepared by adding potassium (19 mg, 0.48

mmol) and ferric nitrate (1 mg) to liquid ammonia (200m1). The solution was

cooled to -78", then a solution of the bromide (85a) (200 mg, 0.47 mmol) in

dichloromethane (10m1) was added dropwise. The mixture was stirred at -78"

for 3 h, then the ammonia was allowed to evaporate overnight. The residue

was partitioned between dichloromethane and water, the organic layer was

separated, then washed with water. The solution was dried, and the solvent was

removed under reduced pressure. The mixture was chromatographed on silica

to give the (Z)-dehydrophenylalanine derivative (86) (22 mg,277o), identical to

the sample described above, and the trans-þ-Iactam (134) (51 mg, 637o), m.p.

243-245" (Found C,1L8; H, 5.8; N, 8.1. CzrHzoNzOs requires C,72.4;, H, 5.8;

N, 8"07o). lH n.m.r. (300 MHz, CDCI3) ô 1.36, s, 9H, CMe3; 4.96, d. J 2.3 Hz,

H4; 5.01, d, J 2.3 lFrz,H3;7.39, s,5H, Ph; 7.72-7.87, m, 4H, Phth. v^u^ 2920,

1745, 1120, 1465, 1390, 710 cm-l. Mass spectrum; mlz 348 (M*', 27o),333

Experimental 151

(-Me, 1), 305 (2),291 (4),276 (3),249 (100), 232 (5),220 (9),204 (14), 165

(5), t32 (5), 104 (28), tjz (34),76 (27).

(2 S 3 R) -3- Am ino -N - t e rt -butyl -¡¿ø-phthaloylphenylalaninamid e ( 135)

Method 1

Potassium amide solution was prepared by adding potassium (11 mg, 0.28

mmol) and ferric nitrate (1 mg) to liquid ammonia (30 ml). The solution was

cooled to -78o, then a solution of the bromide (85b) (100 mg, 0.23 mmol) in

dichloromethane (5ml) was added dropwise. The mixture was stirred for 2 h,

then the ammonia was allowed to evaporate overnight. The residue was

partitioned between dichloromethane and water, then the organic layer was

washed with water, and dried. The solvent was removed under reduced

pressure, and the residue was recrystallized from hexane/dichloromethane to

give the title compound (135) as a white solid (78 mg, 937o), n.P. 150-152"

(Found C,68.9; H,6.5; N, 11.4. CzrHz¡N¡O3 requires C,69.0; H,6.3; N,

lI.57o). lH n.m.r. (300 MHz, CDCI3) õI.44, s, 9H, CMe3; 4.63, d, J 6.8 Hz,

IÐ;5.7i, br s, lH, NH; 5.77, d, J 6.8 Hz,IF.13;6.03, br s, lH, NH'; 6.73, br s,

lH, CONH;7.26-7.41, h,5H, Ph; 7.46-8.02,m,4}l, Phth. v,nu* 3360,3310,

3170,2900, 1660, 1640, 1520, 1450, 750 cm-l. Mass spectrum (FAB); mlz

366 (M+H+,1007o),249 (29),130 (53), 120 (30), 105 (15), 103 (17), 57 (49).

Method 2

The bromide (85b) (100 mg, 0.23 mmol) was dissolved in TÉIF (10 ml)

and then concentrated aqueous ammonia solution (0.5m1) was added. The

solution was stirred for 18 h, then the THF was removed under reduced

pressure. The residue was partitioned between dichloromethane and water, and

the aqueous phase was re-extracted with dichloromethane. The combined

Experimental I52

extracts were dried, and the solvent was removed under reduced pressure

Purification of the product by chromatography on silica gave the title

compound (135) (55 mg, 66Vo), identical to the sample produced above.

Treatment of (2 S 3R) -3 - B ro mo - N - te rú-b utyl-N a-p hthal oyl -

phenylalaninamide (85a) with aqueous ammonia in THF.

Treatment of a solution of the bromide (85a) in TIIF with aqueous

ammonia, as described above for the treatment of the bromide (85a), gave the

(Z)-dehydrophenylalaninamide derivative (86) (887o), with spectral data

identical to that described above"

(253 R) -3-Bromo-N-phthaloylphenylalanine (L37 a) and

(253 R) -3-Bromo-N-phthaloylphenylalanine (137b)

lS)-N-Phthaloylphenylalanine (70) (5.0 g, I7 mmol) was dissolved in a

mixture of CCI¿ and dichloromethane (10:1, 150rnl). NBS (3.15 g, 18 mmol)

was added, and the mixture was refluxed for 2 h while irradiated with a 250W

mercury lamp. The mixture was cooled, filtered, and the solid was washed

with CCI¿. The filtrate was then washed with water, dried, then the solvent was

removed under reduced pressure to give a pale yellow gum. Dissolution of the

gum in CCla and removal of the solvent under reduced pressure at room

temperature gave the title compounds (137a) and (137b) as a 1:1 mixture, as a

pale yellow solid (6.2 g,987o) (Found: C, 52.8; H,3.2; N, 3.5. C17H12BrNOa

requires C,54.6:H,3.2;N,3.7Vo). lH n.m.r. (300 MHz, CDCI3) threo-isomer

(131a) ô5.55, d, J 1l.2Hz,fI2;5.97, d, J 1I.2Hz,H3;6.64, br s, CO2H;7.10-

7.55, m,5H, Ph;7.76-7.95, m,4H, Phth; erythro-isomer (137b) ô 5.64, d, J

I0.4IJz, H2; 5.86, d, J 10.4 Hz,Il3:7.I0-7.31, m,5H, Ph;7.61-7.80, m,4H,t"

Experimental 153

Phth; 8.03, br s, CO2H. vn'u* 3450 br, 1798, 1718, 1608, 1168, 1104, 924,868

cm-l. Mass spectrum; mlz 375 and 373 (M*'), 249 (l'ffiVo),232,220,204.

Treatment of a mixture of (253R)-3-Bromo-N-phthaloyl-phenylalanine (137a) and (253R)-3-Bromo-N-phthaloyl-phenylalanine (137b) with silver carbonate.

To a solution of a 1:1 mixture of the bromides (I37a) and (137b) (6.2 g,

16.6mmol) in acetone (100 ml) was added silver carbonate (900 mg, 8.5

mmol)" The mixture was stirred for 3 h, then was filtered through celite and

the solvent was removed under reduced pressure. Analysis of the crude

mixture using lH n.m.r. specFoscopy indicated the presence of the cis-B-lactone

(139b), the trans-B-lactone (139a) (õ 5.67, d, I 4.5 Hz, H3:. 5.92, d, J 4.5}{2,

H4), and trans-2-phthalimidostyrene (138) in a 2:l;l ratio. Attempted

separations of the compounds by fractional recrystallization or chromatography

on silica resulted in the decomposition of the trans-B-lactone (139a) to give

2-phthalimidostyrene (138). Consequently, separation of the products by

chromatography on silica gave trans-2-phthatimidostyrene (138) (2.769, 67Vo),

m.p. 185-187". lH n.m.r. (300 MHz, CDCI3) õ 7.36, d, I 15.2 Hz, lH;7.36-

1.38, m, 3H, ArH; 7.46-7.49, m, 2H, ArH; 7.65, d, J I5.2 }lz, l}I; 7.74-1.93"

m, 4H, Phth. Further elution gave the cis-B-lactone (139b) (170 mg, 167o),

m.p. 123-125o.lH n.m.r. (300 MHz, CDC13) õ 6.33, d,J 9.2Hz,H4i 6.70, d,J

9.2 Hz, H3; 7.25, s, 5H, Ph;1.72-7.87, m, 4H, arom. Vmax 1840, 1180, 1760

cm-l. Mass spectrum; mlz 24911vf+'-CO2, 1007o),232 (9),220 (20),2O4 (28),

165 (11), 133 (7), 130 (4), I n Q),104 (39), 102 (51),76 (76)"

Experimental 154

(25)-4,5-Dibromo-N-phthaloylleucine Methyl Ester (I42)

The y-bromoleucine derivative (91) (100 mg, 0.28 mmol) was dissolved in

chlorobenzene (2 ml). NBS (100 mg, 0.56 mmol) and a catalytic amount of

benzoyl peroxide (5 mg) were added, and the mixture was refluxed while

irradiated with a 250W mercury lamp for 4 h. Inadiation was stopped and the

mixture was refluxed for a further 12 h. The mixture was cooled and the

solvent was removed under reduced pressure. Purification of the residue by

chromatography on silica gave the title compound (I42) as a pale yellow oil (64

mg,537o). lH n.m.r. (300 MHz, CDCI¡) ô 1.86, s, 1.5H, Y-Me; 1.93, s, 1.5H,

y-Me'; 2.46, dd, J 9.7 Hz, J I2.3 Hz, 0.5H, H.3;2.59, dd, J Il.6 }{z, I2.3 Hz,

0.5H, H3';2.87-3.02, m, lH, CHCIIz; 3.68-3.85, m,2H, CHzBr; 3.75, s, 1.5H,

OMe; 3.76, s, 1.5H, OMe; 5.16-5.30, m, lH, H2;7.27-7.58, m,5H, Ph;7.74-

7.94, m,4H, Phth. Mass spectrum; mlz 435,433 and 431 (M+', 1,2 and lVo),

376 (-COzMe, 28), 374 (-COzMe, 56), 372 (-COzMe, 28), 354 (-Br, 16),352 (-

Br, 16), 294 (38),292 (38),272 (Il),270 (ll),257 (13),242 (12),215 (45),

2r4 (75),200 (52),186 (40), 160 (47),130 (41), 105 (33), 104 (100), 76 (50).

(Z)-N-Phthaloyl-o,B-methanophenylalanine Methyl Ester (143)

To a solution of the y-bromohomophenylalanine derivative (92) (100 mg,

0.25 mmol) in TIIF (2 ml) was added 1,8-diazabicyclo[5.4.0]undec-7-ene

(DBU) (38 mg, 0.25 mmol). The mixture was heated at reflux for t h, then

was cooled and dituted with dichloromethane (20 ml). The solution was washed

with water, dried, then the solvent was removed under reduced pressure. The

residue was chromatographed on silica, eluting with hexanell,2-dichloroethane,

tc give a viscous oil which was crystallized from hexane./dichloromethane to

give the title compound (143) as colourless crystals (58 mg,727o), m.p. 131-

132' (Fouqd: C, 10.9;H,4.9; N, 4.4. CrsHrsNO4 requires C,71.0; H, 4.7; N,

Experimental 155

4.47o). lH n.m.r. (300 MHz, CDCI3) õ2.27, dd, J 6.6 Il2,9.9 Hz,Il,4;2.43, dd,

J 6.6 H2,8.5 Hz,H4';3.38, dd,J 8.5 Hz,9.9}{z,H'3;3.12, s,3H, OMe; 7.07-

7.17, m, 5H, Ph;1.60-7.85, m, 4H, Phth. vmax 1182, L724, 1440, 1404, 1272,

732 cm-t. Mass specfrum; mlz32l (M*').

Acetylation of (S)-a-Hydroxy-N-phthaloylleucine Methyl Ester

(104).

To a solution of y-hydroxyleucine derivative (104) (11.5 g,39.5 mmol) in

dichloromethane (40 ml) was added pyridine (3.13 g,39.5 mmol) and DMAP

(0.48 g,3.9 mmol). Acetyt chloride (20 rnl) was then added, and the mixture

was stirred under nitrogen for 16 h. The solvent was removed under reduced

pressure, and the residue was taken up in ether. The solution was then washed

successivety with dilute HCl, water and 10% NazCO¡" The organic phase was

dried, and the solvent was removed under reduced pressure to give a clear oil.

The products were separated by dry column flash chromatography on silica, to

give the lactone (105) (1.62 g, 167o), identical to the sample described above,

and (S)-4-acetoxy-N-phthatoylleucine methyl ester (144). Further purification

of the acetate (144) by distittation using a Kugelrohr apparatus (200", 0.04mm)

yielded a clear oil which partially crystallized on storage at -20o overnight.

Repeated trituration with hexane/ether and vacuum filtration gave the acetate

(I44) as a white solid (9.24 g,707o), m.p.85-87" (Found: C,61.3; H,5.9; N,

4.2. CnHrqNOe requires C, 61.3; H, 5.7; N, 4.2Vo). lH n.m.r. (300 MHz,

CDCI¡) ô 1.48, s, 3H; 1.57, s, 3H; 1.59, s, 3H;2.52, dd, J 2.8 þ¡z,15.4Hz,f{3

2.82, dd, J 10.1 Hz, 15.4Hz,H3':3.'74, s,3H, OMe; 5.11, dd, J 2.8}{2, IO-l

Hz,rr2i l.l5-7.92, m, 4H, Phth" v^u* l7'14,1738,1718, 1390, 1145, 1020,110

cm-l. Mass spectrum (FAB); mlz 334 (M+H+, 37o),274 (-OAc, 90), 260 (10)'

214 (59),185 (47), r27 (34),93 (100).

Experimental 156

Pyrolysis of (S)-4-Acetoxy-N-phthaloylleucine Methyl Ester (144).

(S)-4-Acetoxy-N-phthaloylleucine methyl ester (144) (5.0g, 15.0 mmol)

was distilled (ca.250",0.5 mm) through a quartz pyrolysis tube packed with

quartz fragments, which had been preheated to 450o. The distillate was

obtained as a viscous yellow-green oil, and was purified by dry cloumn flash

chromatography. The y,ô- and B,y-dehydroleucine derivatives (106) and (107)

were obtained as a mixture in a 1.6:1 ratio, as a pale yellow oil which

crystallized on standing overnight (3.42 g,84Vo). Spectral characteristics of the

y,ô- and B,y-dehydroleucine derivatives (106) and (107) were identical to those

described above.

Acetylation of 4-Hydroxy-N-phthaloylhomophenylalanine MethylEster (145).

To a solution of the y-hydroxyhomophenylalanine derivative (109) (5.0 g,

14.7 mmol) in dichloromethane (20 ml) was added pyridine (1.16 g, I4.7

mmol) and DMAP (0.18 g, I.47 mmol). Freshly distilled acetyl chloride (10

ml) was then added, and the mixture was stirred under nitrogen for 6 h. The

solvent was removed under reduced pressure, and the residue was taken up in

ether, then washed successively with dilute HCl, water and L07o NazCO¡. The

organic phase was dried, then the solvent was removed under reduced pressure

to give a clear oil which crystallized on standing overnight. Fractional

recrystallization from hexane/ethyl acetate gave the cis-B-lactone (110) (304

mg, 77o), and the trans-þ-Iactone (77 mg, 27o), identical to the samples

described above. Further recrystallization gave one diastereomer of (S)-4-

acetoxy-N-phthatoylhomophenylalanine methyl ester (145) (2.21 9,397o), m.p.

137-140".IH n.m.r. (300 MHz, CDCI3) ô 1.94, s, 3H, OAc; 2.83,m,2H,CH2;

3.72, s,3H, OMe; 4.90, dd, J 6.2 H2,8.1 Hz, H4; 5.85, t, J 6.6 Hz, H2;7.18-

Experimental 157

7.45, m, 5H, Ph;7.73-7.90, m,4H, Phth. vru* 1780,1745,1715,1395,1025

cm-1. Mass spectrum; mlz38I (M*', O.lVo),338 (-Ac, ll),322 (-CO2Me, 1),

308 (3), 262 (r3),219 (100), t87 (86),114 (43), r32 (28), 130 (27),115 (81),

107 (44), 105 (52), 77 (33),76 (33), 43 (92). Further recrystallizatíon gave

the other diastereomer of the acetate (145) (1.51 g,277o), m.p. I25-I27"

(Found: C,66.1; H, 5.1; N, 3.6. CzrHrqNO6 requires C,66.L; H, 5.0; N,3.77o)"

rH n.m.r. (300 MHz, CDCI3) õ 2.09, s, 3H, OAc; 2.80, ddd, J 4.9 IIz,,9.7 }Iz,

l4|7 IJz,H.3;2.86, ddd, J 4.9 Hz, ll.0 Hz, 14.7 Hz,}{3';3.74, s, 3H, OMe;

5.I4, dd, J 4.9 Hz, 1 1.0 Hz, H4: 5.62, dd, J 4.9 Hz, 9.7 HaIÐ; 7 .18-'7 .30, m,

5H, Ph; 7.74-7.87, m, 4H, Phth. v*u* 1778, 1746, 1722, 1600, 1555, 1390,

1066, 772 cm-r. Mass spectrum; mlz 381(M+', 0.57o),338 (-Ac, l4), 308 (3),

262 (r5),2t9 (100), 187 (84), t74 (28),115 (36), 105 (67), 77 (16),16 (16),

43 (s0).

Pyrolysis of 4-Acetoxy-N-phthaloylhomophenylalanine Methyl Ester(14s).

A 1.5:1 mixture of the diastereomers of 4-acetoxy-N-phthaloyl-

homophenylalanine methyl ester (145) (1.0 g,2.62 mmol) was distilled (250",

0.3 mm) and passed through a pyrolysis tube (520') as for pyrolysis of the

acetoxyleucine derivative (I44). Purification of the crude residue by flash

chromatography on silica gave a 1:1 mixture of the B,1-dehydro-

homophenylalanine derivative ( 146) and the a,B-dehydrohomophenylalanine

derivative (141) (560 mg, 667o), as a clear oil. lH n.m.r. (300 MHz, CDCI3)

p,y-dehydrohomophenylalanine derivative (146) õ 3.80, s, 3H, OMe; 5.58, d, J

7.1Hz, IJr2; 6.68, d, J 15.9 }lz,H4; 6.74, dd, J 7.1 Hz, 15.9 Hz,H3;7.19-7.43,

m, 5H, Ph;7.73-1.95, m, 4I{, Phth; a,B-dchydrohomophenylalanine derivative

(I41) õ 3.52, d,J 7.4IJr2,2H, CHzPh; 3.77, s,3H, OMe; 7.19-7.43, m,6H, H3

and Ph; 7:73-7.95, m,4H, Phth. vnlu^ 1780,1745, l'715, 1605, 1560, 1390,

Experímental 158

850,720 cm-I. Mass spectrum; mlz32l (M*', 57o),3O5 (5),289 (-MeOH,78),

262 (-CO2Me, 54),245 (20),230 (19), 147 (12), 132 (29), 115 (100), 104

(71), 9l (rt),76 (44).

Treatment of ( 2 S 35) -3 - B ro mo -N - te rf-bu tyl -N ø-p hthaloyl -

phenylalaninamide (85b) with NaBHa

To a solution of the bromide (85b) (100 mg, 0.23 mmol) in 8O7o aqueous

ethanol (10 rnt) was added NaBtIa (40 mg, 1.06 mmol), and the mixture was

stirred at room temperature for 12 h. The reaction was quenched by the

addition of dilute HCl, then the mixture was concentrated under reduced

pressure. To the concentrate was added water (40 ml), then the mixture was

extracted with dichloromethane (2x40 mt). The combined organic phases were

dried and the solvent was removed under reduced pressure" Separation of the

products by chromatography on silica gave the secondary alcohol (151) (36 mg,

367o)" lH n.m.r. (300 MHz, CDCI3) õ 1.40, s, 9H, CMe3; 4.28,d,I7.II{z,lI2;

4.70, s, OH; 5.75, d, J 7.1 IIz, }I3:- 5.74, s, ring-H; 6.51, br s, NH; 7.22-7.37,

m, 5H, Ph; 7.63-7.71, m, 3H, ArH; 7.86-7.88, m, lH, ArH. Further elution

gave the primary alcohol (152) (21 mg, 2l7o). lH n.m.r. (300 MHz, CDCI3) ô

1.41, s, 9H, CMe3; 4.69, d, J 12.5 IIz, CHOH; 4.70, d, J 7 .2 Hz, H2; 4.86, d, J

12.5 }Jz, CII'OH; 5.28, d, J 7 .2 IIz, H3; 6.34, br s, NH; 7 .23-7 .54, m, 8H, ArH;

8.04-8.07, m, lH, fuH.

Experimental 159

Treatment of a mixture of (253R)- and (2535)-3-Bromo'N-tert-butyl-Nø-phthaloytphenylalaninamide (85a) and (85b) withtri-z-butyltin deuteride.

To a solution of a 1:1 mixture of the bromides (85a) and (85b) (200 mg,

0.47 mmol) in benzene (10 ml) was added tri-n-butyltin deuteride (275 mg,

0.93 mmol) and AIBN (40 mg, 0.24 mmol). The solution was stined at room

temperature for 3 d, then the solvent was removed under reduced pressure.

The residue was taken up in hexane, and the suspension was stirred for 5 min.

The white sotid was isolated by filtration and was washed repeatedly with

hexane to remove traces of stannanes. The solid was recrystallized from

hexane/ethyl acetate to give the deuteriophenylalaninamidd derivative (148) as a

3:1 ratio of diastereomers (102 mg, 637o), m.p. 186-187". lH n.m.r. (300

MIIz, CDCI¡) ô 1.30, s,9H, CMe3; 3.46,d,110.2H2,0.25H,H3;3.54,d,J 6.4

Hz, 0.75H, H3'; 4.99, d, J 10.2 Hz, 0.25 H, H2: 4.99, d, J 6.4 Hz, O.l5H, H2;

l.l3-7.21, m, 5H, Ph;7.67-7.80, m, 4H, ArH.

Treatment of a mixture of (2S3R)- and (2535)-3-Bromo'N-phthaloylphenylalanine Methyl Ester (82a) and (82b) withtri-z-butyltin deuteride.

Treatment of a 1:1 mixture of the bromides (82a) and (82b) with

tri-r-butyltin deuteride and AIBN, as described above for treatment of a

mixture of the bromoamides (85a) and (85b), gave the deuteriophenylalanine

derivative (149) as a2:1 ratio of diastereomers (98 mg, 617o), m.p. I22-I24".

lH n.m.r. (300 MHz, CDCI3) õ 3.53, d, J 11.7 }J2,0.35H, }l3;3-59, d, J 4-8H2,

0.65H, F-l3';3.77, s,3H, OMe; 5.16, d, J 11.7 H2,0.35H, }{2:5.16, d, J 4-8H2,

0.65H, H2;7.11-7.19, m,5H, Ph;7.65-7.78, m,4H, ArH.

Experimental 160

(253R)-3-Deuterio-N-phthaloylphenylalanine Methyl Ester Ga9a)

A mixture of the threo-bromide (82a) (1.0 g, mmol) and 5Vo Pd/C (100

mg) in 2Hr-methanol (99.57o 2H,20 ml) was stirred at -20" under an

atmosphere of deuterium gas for 3 d in a refrigerated bath. The palladium

catalyst was removed by filftation, and the solvent was removed under reduced

pressure. The residue was taken up in dichloromethane and washed with l07o

NazCO¡, then dried, and the solvent was removed under reduced pressure.

Recrystallization of the residue from hexane/ethyl acetate gave the title

compound (149a) (987o d.e.) as colourless prisms (728 mg,9I7o), m.p. 124-

126"" lH n.m.r. (300 MHz, CDCI3) õ 3.59, d, J 4.8 Hz,}J3;3.77, s,3H, OMe;

5.16, d, J 4.8 llz,}l2;7.I1-7.19, m,5H, Ph; 7.65-7.78, m,4H, Phth. Mass

spectrum; mlz 310 (M+', 997o 2}J).

(2535)-3-deuterio-N-phthaloylphenylalanine Methyl Ester (149b)

Treatment of a solution of the erythro-bromide (82b) (1.0 g, mmol) in

2Hr-methanol (99.5Vo D,20 ml) wíth 57o PdlC under an atmosphere of

deuterium gas, as described for the preparation of the deuteriophenylalanine

derivative (I49a) from fhe threo-bromide (82a), gave the title compound

(149b) as colourless prisms (717 mg,90Vo), m.p. 125-121". tH n.m'r. (300

MHz, CDCI3) ô 3.53, d, J 1I.J }Jz,H3;3.17, s, 3H, OMe; 5.16, d, J 11.7 Hz,

II2; 7.Il-7.19, m, 5H, Ph; 1.65-7.78, m, 4H, Phth. Mass spectrum; mlz 310

(M*.,997o2Hù.

(R) -N-Phthaloylp heny lalanine

Treatment of (R)-phenylalanine (55) with phthalic anhydride, as described

above for the preparation of (S)-N-phthaloylphenylalanine (70) from

Experimental 161

(S)-phenylalanine (56), gave the title compound as colourless crystals (87Vo),

m.p. 185-186' (lit.lls 180-180.5). lH n.m.r. spectrum identical to that of the

(S)-enantiomer (70).

(R)-N-Phthaloylphenylalanine Methyl Ester

Treatment of (R)-N-phthaloylphenylalanine with a methanolic solution of

hydrogen chloride, as described for the preparation of ( S ) - N -

phthaloylphenylalanine methyl ester (74) from (^S)-N-phthaloylphenylalanine

(70), gave the títle compound as large colourless prisms (937o), m.p. 123-126".

lH n.m.r. spectrum identical to that of the (S)-isomer (74).

(2R35)-3-Bromo-N-phthaloylphenylalanine Methyl Ester (82c) and(2R3R)-3-Bromo-N-phthaloylphenylalanine Methyl Ester (82d)

Treatment of (R)-N-phthaloylphenylalanine methyl ester with NBS, as

descibed for the preparation of the bromides (82a) and (82b) from

(S)-N-phthaloylphenylalanine methyl ester (74), gave the title compounds (82c)

and (82d) as colourless crystals" The threo-bromide (82c) was isolated in MVo

yield, m.p. 139-142", and had spectral data identical to that of the (253R)-

isomer (82a). The erythro-bromide (82d) was isolated in 43%oyield, m.p. 120-

I21", and had spectral data identical to that of the (2535,)-isomer (82b).

(2R35)-3-Deuterio-N-phthaloylphenylalanine Methyl Ester (1a9c)

Treatment of a solution of the threo-bromide (82c) (500 mg, mmol) in

2Hr-methanol (99.5Vo 2H, l0 mt) with 57o Pd/C under an atmosphere of

Experimental 162

deuterium gas, as described for the preparation of the deuteriophenylalanine

derivative (I49a) from the threo-bromide (82a), gave the title compound

(149c) as colouless crystals (372 mg, 93Vo), m.p. 125-126" " lH n.m.r.

spectrum identical to that of the (253R)-enantiomer (149a)" Mass spectrum;

mlz310 (M+',99Vo2H).

(2R3R)-3-Deuterio-N'phthaloylphenylalanine Methyl Ester (149d)

Treatment of a solution of the erythro-bromide (82d) (500 mg, mmol) in

2Hr-methanol (99.5Vo 2}I, 10 ml) with 5Vo Pd/C under an atmosphere of

deuterium gas, as described for the preparation of the deuteriophenylalanine

derivative Qaga) from the threo-bromide (82a), gave the title compound

(149d) as colouless crystals (358 mg, 907o), m.p. 124-126". lH n.m.r.

spectrum identicat to that of the (2SiS)-enantiomer (149b). Mass spectrum;

mlz310 (M+',997o2Hù.

(253 R) -3-Deuteriophenylalanine (65a)

A solution of the threo-deuteriophenylalanine derivative (I49a) (500 mg,

1.6 mmol) in 6N HCl/acetic acid (2:I,30 ml) was refluxed for 6 h. The

mixture was allowed to cool, then was evaporated to dryness under reduced

pressure. The residue was taken up in water (30 ml) and the phthalic acid was

removed by filtration. The filtrate was evaporated to dryness, then the residue

was dissolved in dry ethanot (15 ml). Aniline (1.5 ml) was added and the

solution was left to crystallize. The precipitate was isolated by filtration and

was washed with acetone to give the title compound (65a) as a white powder

(223 mg,837o),m.p.272-276".IH n.m.r. (300 MHz, DzO) õ 3.21 ,d,14.9H2,

H3; 3.92, d, J 4.9 Hz, IJZ: 7.24-7.38, m, 5H, Ph. Mass spectrum; mlz 167

Experimental 163

(M+H+, 99Vo 2}t1, l77o), I2l (42),93 (11), 92 (PhCHO+, 38), 74 (100),28

(r2).

(2535)-3-Deuteriophenylalanine (65b)

Treatment of the erythro-deuteriophenylalanine derivative (149b) (500

mg, 1.6 mmol) with 6N HCl/acetic acid, as described for the preparation of

(253R)-deteriophenylalanine (65a) from the deuteride (149a), gave the title

compound (65b) as a white powder (228 mg, 857o), m.p. 270-275". lH n.m.r.

(300 MHz, DzO) õ 3.03, d, J 7 .9 Hz, }J3; 3.92, d, J 7 .9 Hz, H2; 7 .24-7 .38, m,

5H, Ph. Mass spectrum; mlz 167 (M+H+, 997o 2H1, 4O7o), 121 85), 93 (23),92

(PhcHo+, 100), 74 (95),28 (20).

(2R35)-3-Deuteriophenylalanine (65c)

Treatment of the threo-deuteriophenylalanine derivative Oa9c) (100 mg,

0.32 mmol) with 6N HCl/acetic acid, as described for the preparation of

(253R)-deteriophenylalanine (65a) from the deuteride (149a), gave the title

compound (65c) as a white powder (44 mg, 827o), m.p. 268-212". Spectral

data identical to that of the (253R)-enantiomer (65a).

(2R3R)-3-Deuteriophenylalanine (65d)

Treatment of the erythro-deuteriophenylalanine derivative (149d) (250

mg, 0.81 mmol) with 6N HCl/acetic acid, as described for the preparation of

(253R)-deteriophenylalanine (65a) from the deuteride (149a), gave the title

Experimentnl I&

compound (65d) as a white powder (109 mg, SlVo), m.p. 270-273". Spectral

data identical to that of the (2S3S)-enantiomer (65b).

Treatment of (253R)-3-Deuteriophenylalanine (65a) with PAL

Sodium borate buffer (0.04M, pH 8.7) was prepared by dissolving sodium

tetraborate decahydrate (26.7 g, 0.07 mol) in water (1.75 L). The pH was

reduced to 8.7 by the addition of conc. IICI (ca. 7 ml). To a solution of

(253R)-3-deuteriophenylalanine (65a) (33 mg, 0.20 mmol) in sodium borate

buffer (25 ml) was added PAL (Sigma, from Rhodotorula glutinis, grade 1,

solution in 607o glycerol, 3 mM Tris-HCl, pH 7.5; 0.2 ml, 0.5 units). The

solution was stirred at 30o in a constant temperature water bath for 20 h. The

solution was acidified to pH 1 by the addition of conc. HCl, then was extracted

with dichloromethane (2x25 ml). The organic phase was dried and the solvent

was removed under reduced pressure Recrystallization of the residue gave

trans-3-deuteriocinnamic acid (67) (15.9 mg,547o) m.p. 135-137". lH n.m.r.

(300 MI{z, CDCI3) ô 6.46, br s, 0.98H,H.2; 6.47, d, J 16.0}J2,0.02H, H2;7.41-

7.44, m, 3H, ArH; 7.55-7.58, m,2Il, ArH; 7.81, d, J 16.0 }{2,0.02H, H3. lH

n.m.r. spectroscopic analysis indicated that the product (67) contained 98Vo

2H1-incorporation.

Treatment of (2535)-3-Deuteriophenylalanine (65b) with PAL

Treatment of (2S3S)-3-deuteriophenylalanine (65b) (33 mg, 0.20 mmol) in

sodium borate buffer (25 ml) with PAL (0.2 ml, 0.5 units), as described above

for the treatment of (253R)-3-deuteriophenylalanine (65a) with PAL, gave

trans-cinnamic acid (57) (17.8 mg, 607o) m.p. 134-136' (lit.t7e I32o). lH

Experímental 165

n.m.r. (300 MHz, CDCI3) õ 6.47, d, J 16.0 }{z,II2; 7.41-1.44, m, 3H, ArH;

7.55-7.58, m,2}J., ArH; 7.81, d, J 16.0 Hz,H3.

Treatment of QR3S)-3-Deuteriophenylalanine (65c) with PAL

Treatment of (2R35)-3-deuteriophenylalanine (65c) (33 mg, 0.20 mmol) in

sodium borate buffer (25 ml) with PAL (0.2 ml, 0.5 units), as described above

for the treatment of (253R)-3-deuteriophenylalanine (65a) with PAL, but

stirring the solution for 8 days, gave trans-3-deuteriocinnamic acid (67) (20.6

mg,707o) m.p. 134-I37". lH n.m.r. (300 MHz, CDCI3) ô 6.46, br s, 0.92H,

IÐ; 6.47 , d, J 16.0 Hz, 0.08H , I12; 7 .41-7 .44, m, 3H, ArH; 7 .55-7 .58, m, 2}J,

ArH; 7.81, d, J 16.0 Hz, 0.08H, H3. lH n.m.r. spectroscopic analysis indicated

that the product (67) contained 927o 2H1-incorporation.

Treatment of (2R3R)-3-deuteriophenylalanine (65d) with PAL

Treatment of (2R3R)-3-deuteriophenylalanine (65d) (33 mg, 0.20 mmot)

in sodium borate buffer (25 ml) with PAL (0.2 ml, 0.5 units), as described

above for the treatment of (2R35)-3-deuteriop[enylalanine (65c) with PAL,

gave trons-cinnamic acid (57) (17.4 mg, 597o) m.p. 135-136'. lH n.m.r. (300

lvffIz, CDCI¡) õ 6.46, br s, 0.27H,IÐ; 6.47, d, J 16.0 Hz,0.73H,If2;7.4I-7 .M,

m,3H, ArH; 7.55-7.58,m,2H, ArH; 7.81, d, J 16.0 }Iz,0.l3H. H3. lH n.m.r.

spectroscopic analysis indicated that the product (57) contained 217o 2I{r

incorporation.

1

2

3

4

References 166

REFERENCES

Greenstein, J. P., and Winitz,M., Chemistry of the Amino Acids, John

Wiley and Sons Inc., New York, 196I, vols. 1-3.

(a) Wagner, I., and Musso, H., Angew. Chem., Int. Ed. EngI., 1983,22,

816; (b) Ohfune, Y., Acc. Chem. Res., 1992,25, 360, and references

cited therein.

Williams, R. M., Sinclair, P. J., Zha\ D., and Chen, D., J. Am. Chem

9oc.,1988, II0, 1547.

(a) Hammond, S. J., Williamson, M. P., Williams, D. H., Boeck, L. D.,

and Marconi, G. G., .f. Chem. Soc., Chem. Commun., 1982,344; (b)

Hammond, S.J., Williams, D.H., and Nielsen, R. V., J. Chem. Soc.,

Chem. Commun", 1983, 116; (c) Lin, T.-S., and Kolattukudy, P. E.,

Eur. J. Biochem., 1980, 106,341.

5. Meerakami, Y., Hisaeda, Y., and Ohno, T., Chem. Lett., 1987,1351.

6. Suzuki, F., and Baker, H. 4., J" BioI. Chem., 1966,244,878.

Kleeman, 4., Leuchtenberger, W., Hoppo, B., and Tanner, H., "Amino

Acids" in Ullmann's Encyclopedia of Industríal Chemistry, VCH

Verlagsgesellschaft, Weinheim, 1985, vol. A-2, pp. 57-97 .

Mazur, R. H., Schlatter, J. M., and Goldkamp, A. H., J. Am. Chem. Soc.,

1969,9L,2684.

7

8

9

Reþrences 167

(a) Rando, R. R., Acc. Chem. Res., 1915,8, 28; (b) Abeles, R. H., Pure

Appl. Chem.,1980, 53, 149.

10" Poisel, H., and Schmidt, U., Angew. Chem., Int. Ed" EngI., 1976, 15,

294.

11. Shimohigashi, Y., Stammer, C. H., and Costa, T., "Synthetic Enkephalin

Analogues: Designing New Drugs of Pharmacological Interest" in

Synthetic Peptides in Biotechnology, A.R. Liss, Inc., 1988, pp.203-223.

12. Mapelli, C., Kimura, H., and Stammer, C. H., Int. J. Peptíde Protein

Res., 1986, 28,347.

13. Hruby, V. J., Life 9ci.,1982,31, 189.

14. Williams, R. M., Synthesis of Optícally Active a-Amino Acíds,

Pergammon Press, Oxford, UK, 1989"

15 Wightman, R. H., Staunton, J., Battersby, A. R., and Hanson, K. R.,.f.

Chem. Soc., Perkin Trans. I , 1972,2355.

16. Thomas, N. R., and Gani, D., Tetrahedron, 199I, 47 , 497 .

I7. Kollonitsch, J., Perkins, L. M., Patchett, A. 4., Doldouras, G. 4.,

Marburg, S., Duggatr, D.E., Maycock, A. L., and Aster, S. D., Nature,

1978,274,906.

18. Versteeg, D. H. G., Palkouts, M., Van der Gugten, J., Wijnen, J. L. M.,

Smeets, G. W. M., and DeJong, W., Prog. Brain Res., 1971, 47, lll.

References 168

19. Walsh, C., and Wang, 8., Biochemistry, 1978,17, 1313.

20. Kollonitsch, J., and Barash, L., J. Am. Chem. Soc., I976,98, 5591.

2I. Jackson, R. F. W., Wood, 4., and Wythes, M. J., Synlett, 1990,735.

22. Arnold, L. D., Kalantar, T. H., and Vederas, J. C., J" Am. Chem. Soc.,

1985, L07,7105.

23 Arnold, L. D., Drover, J. C. G., and Vederas, J. C., J. Am. Chem. Soc.,

1987, 109" 4649"

24 Arnold, L. D., Muy, R. G., and Vederas, J. C., J. Am. Chem. Soc., 1988,

1^1.0, 2231.

25 (a) Barton, D. H. R., Herve, Y., Potier, P., and Thierry,J.Tetrahedron,

1988, 43,4297; (b) Barton, D. H. R., Herve, Y., Potier, P., and

Thierry, J., Tetrahedron, 1988, 44, 5419.

26 (a) Ohfunê, Y., and Nishio, H., Tetrahedron Lett., 1984,25, 4I3; (b)

Fushiya, S., Nakatsuyama, S., Sato, Y.and Nozoe, 5., Heterocycles,1981,

15, 819.

27 (a) Baldwin, J. E., Moloney, M. G., and North, M., Tetrahedron, 1989,

45,6309; (b) Sardina, F. J., Paz, M. M., Fernandez-Megia, E., de Boer,

R. F., and Alvarez, M. P., Tetrahedron Lett., 1992, 33, 4637 .

Pu, Y., Martin, F. M., and Vederas, J. C.,J. Org. Chem., 1991,56,

1280.

28

References 169

29. Shimamoto, K., and Ohfune, Y., Tetrahedron Lett., 1988, 29, 5171.

30. Kollonitsch, J., Rosegay, 4., and Doldouras, G. 4., J. Am. Chem. Soc.,

1964,86,1857.

31. Kollonitsch, J., Scott, A. N., and Doldouras, G. A., J. Am. Chem. Soc.,

1966,88,3624.

32. Fujita, Y., Kollonitsch, J., and Witkopp, 8., J. Am. Chem. Soc., 1965,

97, 2030.

33. Russell, G. 4., in Free Radícals, Kochi, J. K. (Ed.), Wiley, New York,

1973, vol. l, pp.275-331.

34 Poutsma, M. L., in Free Radícals, Kochi, J.K. (Ed.), Wiley, New York,

1973, vol.2, pp. 159-229.

35 (a) Ito, Y., Ohashi, Y., Kawabe, S., Abe, H., and Okuda, T., J. Antibiot.,

1972, 25,360; (b) Konishi, M., Ohkuma, H., Sakai, F., Tsuno, T.,

Koshiyamâ, H., Naito, T., and Kawaguchi, H., J. Am. Chem. Soc., 1981,

I03, I24l; (c) Arnold, E., and Clardy, 1., J. Am. Chem. Soc., 1981,

r03, t243.

36. Bowman, N. J., Hay, M. P., Love, S. G., and Easton, C.J.,J. Chem

Soc., Perkin Trans. l, 1988,259.

37 . Neta, P., ancl Fessenclen, R. W'., .l . Phys. Chem., 1971,75,738.

References I7O

38. Tanaguchi, H., Hatano, H., Hasegawa, H., and Maruyama, '1., J. Phys

Chem., 1970,74,3063.

39 (a) Viehe, H. G., Merenyi, R., Stella, L., and Janousek, 2., Angew

Chem., Int. Ed. Engl., 1979,18, 917; (b) Viehe, H. G., Janousek, 2.,

and Merenyi, R., Acc. Chem. Res.,1985, 18, 148.

q (a) Baldock, R. W., Hudson, P., Katritsky, A. R., and Soti, F.,

Heterocycles,1973,L,67; (b) Baldock, R. W., Hudson, P., Katritsky, A.

R., and Soti, F., J. Chem. Soc., Perkin Trans. I, I9'74, 1422.

4I. Balaban, A. T., Rev. Roum. Chím., I9ll,16,725.

42. Easton, C. J., Tan, E. W., and Hay, M. P., J" Chem" Soc., Chem"

Commun.,1989, 385.

Badran, T. !V., Ph.D. Thesis, University of Adelaide, 1991, pp. 58-63.

Rosenmund, K. W., and Dornsaft, H., Chem. 9er.,1919,52, 1734.

(a) Hahn, F. E., in Antibiolics, Gottlieb, D., and Shaw, P. D. (Eds.),

Springer-Verlag, Heidelberg, 1967, vol. 1, p. 308; (b) Pestka, S., in

Antibiotics, Corcoran, J. W., and Hahn, F. E. (Eds.), Springer-Verlag,

Heidelberg,I975, vol.3, p.310: (c) Pongs, O., in Antibiotics, Hahn, F.

E. (Ed.), Springer-Verlag, Heidelberg, 1979, vol.5, part 1, p.26.

46. Krant, J., Ann. Rev. Ùiochem.,1977,46,331.

43

44

45

47 Che¡evert, R., and Thiboutot,5., Synth¿sis, 1989, 444.

48

51.

References I7l

(a) Ehrhart, G., Siedel, W., and Nahm, H., Chem. Ber., 1957,90, 2088;

(b) Johnson, F., in The Total Synthesis of Natural Products, ApSimon, J.

(Ed.), Wiley, NewYork, 1973, vol. l, p. 457; (c) Suzuki, M., 'f .

Antibiot. (Japan), Ser. 8,1961, 14,323.

49 Sahu, D. P., Mashava, P., Manhas, M. S., and Bose, A. K., J. Org

Chem.,1983, 48,1142.

50. Miller, M. J., Acc. Chem. Res., 1986, 19,49"

Williams, D. H., Acc. Chem. Res., 1984, 17,364, and references cited

therein.

52 McCormick, M. H., Stark, W. M., Pittenger, G. F., Pittenger, R, C., and

McGuire G. M., Antibiot. Annu.,1955-56, 606.

53. (a) Cafferkey, M. T., Hone, R., and Keane, C. T., .,f. Antimícrob.

Chemother., 1982,9,69; (b) Watanakunakorn, C., Rev. Inf. Dis., 1981,

3, 5210; (c) Horowitz, H. W., Handwerger, S., van Korn, K. G., and

Wormser, G. P., Lancet, 1981, vol.Il(857I), 1329.

54 (a) O'sullivan, J., McCullough, J. 8., Tymiak, A. 4., Kirsch, D. R.,

Trejo, W. H., and Principe, P. A.,.1. Antibiot., 1988, 41, 1740; (b)

Bonner, D. P., O'Sullivan, J., Tanaka, S. K., Clark, J. M., and Whitney,

R. R., J. Antibiol., 1988, 4L, 1745; (c) Tymiak, A. 4., McCormick, T.

J., and Unger, S. E., J. Org. Chem.,1989, 54, 1149.

55. Cyclosporir¿ A, White, D. J. G. (Ed.), Biomedical, Amsterdam, 1982

57

References 172

56. Wenger, R. M., Angew. Chem., Int. Ed. Engl., 1985, 24,77.

Jolad, S. D., Hoffmann, J.J., Torrance, S. J., Wiedhopf, R. M., Cole, J

R., Arora, S. K., Bates, R. 8., Gargiulo, R. L., and Kriek, G. R., J. Am

Chem. Soc.,1977, 99, 8040.

Traber, R., Keller-Juslen, C., Loosli, H.-R., Kuhn, M., and von

'Wartburg, A., Helv. Chim. Acta, 1979, 62, 1252.

58

59. Culvenor, C. C. J., Cockruñ, P. 4., Edgar, J. 4., Frahn, J. L., Gorst-

Allman, C. P., Jones, A. J., Marasas, W. F. O., Murray, K. E., Smith, L.

W., Steyn, P. S., Vleggaar, R., and'Wessels, P. L., J. Chem. Soc., Chem.

Commun.,1983, 1259"

60. Mackay, M. F., Van Donkelaar, 4., and Culvenor, C. C. J., J" Chem

Soc., Chem. Commun., 1986,1219.

Wiedland, T ., N at urwi s s e ns c haft e n, 197 2, 59, 225

V/iedland, T., and Hofer, A., Liebigs Ann. Chem., 1958, 619,35.

Pfeander, P., and Wiedland, T., Liebigs Ann. Chem., 1966,700, 126.

Wiedland,T., Liebigs Ann. Chem.,1981, 1445.

Buku, 4., Faulstich, H., Wiedland, T., and Dabrowski, J., Proc. Natl

Acad" Sci. [/SA, 1980, 71,2370.

61.

62"

63.

64.

65.

67

68

69

References 173

66. Morita, H., Kondo, K., Hitotsuyanagi, Y., Takeyâ, K., Itokawa, H.,

Tomioka, N., Itai, 4., and Iitaka, Y., Tetrahedron, 1991, 47,2757.

DiBello, I. C., Dorling, P., Fellows, L., and Winchester, 8., FEBS Lett.,

1984, L76,61.

Hendry, A. T., and Dillon, J. R., Can. J. Microbiol.,1984,30, 1319.

Janecek, J., Jiresova, M., Technikova, 2., and Spizek, J., Folia

M icrobiol., 197 l, 16, 367 .

70. Wiedland, T., Mannes, K., and Schopf, 4., Liebigs Ann. Chem., 1958,

617, r52.

7I. Liesch, J. M., Milington, D.S., Pandey, R. C., and Reinhart, K. L.,J"

Am. Chem. Soc., 1976,98,8237.

72 "cr-Amino Acid Synthesis", Tefiahedron Symposia in Print, O'Donnell,

M. J., (Ed.), Tetrqhedron, 1988,44,5253.

Williams, R. M., and Hendrix, J. 4., Chem. Rev., 1992,92,889.

Coppola, G. M., and Schuster, H. F., Asymmetric Synthesís:

Constructíon of Chiral Molecules Using Amíno Acids, John Wiley and

Sons, New York, 1987.

13

74

75. Oh-Hashi, J., and Harada, K., Bull. Chem. Soc. Jpn, t966,39,2281.

l6

77

References 174

Guanti, G., Banfi, L., Narisano, E., and Scolastico, C., Tetrahedron,

1988,44,3671.

Belokon, Y. N., Bulychev, A. G., Vitt, S. V., Struchkov, Y. T.,

Batsanov, A. S., Timofeeva, T. V., Tsyryapkitr, V. 4., Ryzhov, M. G.,

Lysova, L.4., Bakhmutov, V. I., and Belikov, V. M., J. Am. Chem.

Soc., 1985, 107, 4252.

Casella, L., Jommi, G., Montanari, S., and Sisti, M., Tetrahedron Lett.,

1988,29,2067.

78

79 Ito, Y., Sawamura, S., and Hayashi, T., J. Am. Chem. \oc.,1986, 108,

6405.

80 Schollkopf, U., Groth, U., and Deng, C., Angew" Chem., Int. Ed. Engl.,

1981,20,798.

81. Schollkopf, U., Hartwig, W., Pospischil, K.-H., and Kehne, H., Synthesis,

1981,966.

Schollkopf, U., Nozulak, J., and Groth, U., Synthesis, 1982, 868.

Schotlkopf, U., and Groth,U., Angew. Chem., Int. Ed. Engl., 198I,20,

977.

Schollkopf, IJ., and Groth,U., Synthesis, 1983 , 673.

Schollkopf, U., Tiller, T., and Bardenhagen, J., Tetrahedron, 1988,44,

s29,3.

82.

83.

84.

85.

References I75

86. Seebach, D., Imwinkelried, R., and Weber, T., Modern Synthetic

Methods, Springer-Verlag, Heidelberg, 1986, vol. 4, 182.

87. Seebach, D., Juaristi, 8., Miller, D. D., Schickli, C., and'Weber, T.,

Helv. Chim. Acta, 1987 ,70,237 .

(a) Evans, D. 4., and Weber, A. E., J. Am. Chem. Soc., 1986, L08,

6757; (b) Evans, D.4., and Weber, A. E., J. Am. Chem. Soc., 1987,

109, 7151.

88

89 Mellin, G. W., and Katzenstein, M., New Engl. J. Med., 1962,267,

1 184.

90. von Blaschke, G., Kraft, H. P., Finkentscher, K., and Kohler, F.,

Arzneim.-F orsch.l Drug Res., 1979, 29, 1640.

9I. Remington's Pharmqceutical Sciences, l6th ed., Osol, ,A.,. (Ed.), Mack

Publishing, Easton, PA, 1980, 1170.

92 Csaky, T.2., Cutting's Handbook of Pharmacology, 6th ed., Appelton-

Century-Crofts, New York, 1979, 16l "

93 Tanner, D., Birgersson, C., and Dhaliwal, H. K., Tetrahedron Lett.,

1990,31, 1903.

Rao, A. V. R., Yadav, J. S., Chandresekhar, S., and Rao, C. S

Tetrahedron Lett., 1989, 30,6769"

94

References 176

95 Kornblum, N., Smiley, R. A., Blackwood, R. K., and Iffland, D. C., J.

Am. Chem" Soc., 1955, 77, 6269"

96. Pocker, Y., and.Kevill, D. N., J. Am. Chem. Soc., 1965,87, 4760.

97. Korblum, N., and Hardies, D. E., J. Am. Chem" Soc., 1966,88, 1707.

98.'Walsh, C.,Tetahedron, 1982,38, 871.

99. Walsh, C., Ann. Rey. Bíochem.,1984,53, 493.

100. Scaman, C. H., and Palcic, M. M., Biochemístry,1992,31,6829.

101. Hanson, K. R., and Havir, E. 4., Arch. Biochem. Biophys., 1970,I4L,

1.

102. Havir, E. A., Reid, P. D., and Marsh, H. V. Jr., Plant Physiol., I971,48,

130.

103. Havir, E. 4., and Hanson, K. R., Biochemistry, I975,14, 1620.

104. Hodgins, D. S., J. Bíol. Chem., 197I,266,2977.

105. Parkhurst, J. R., and Hodgins, D. 5., Arch. Biochem. Bíophys., 1972,

I52, 597.

106. Walsh, C., Enzymatic Reactíon Mechanisms, W. H. Freeman & Co., San

Francisco , 1979, pp. 563-567.

References ll7

lO1. Hanson, K. R., and Havir, E. 4., "The Enzymic Elimination of

Ammonia" inThe Enzymes,3rd ed., Boyer, P. D. (Ed.), Academic Press,

New York,1972, vol. 7, pp. 75-166.

108. Hanson, K. R., and Havi¡, 8.,A.., "Phenylalanine Ammonia Lyase" inThe

Biochemistry of Plar?ts, Stumpf, P. K., and Conn, E. E. (Eds.), Academic

Press, New York, 1981, vol. 7,pp.577-625.

109. Havir, E. 4., and Hanson, K. R., Biochemistry,1968, 7, 1896.

110. Havir, E. 4., and Hanson, K. R., Biochemístry, 1968,7 , 1904"

111. Kirby, G. W., and Michael, J., J. Chem. Soc., PerkinTrans. 1,1973,

115.

lI2. Ife, R., and Haslan, E., J. Chem. Soc. (C), 197I,2818.

113. Bartl, K., Cavalar, C., Krebs, T., Ripp, E., Retey, J., Hull, 'W. E.,

Gunther, H., and Simon, H., Eur. J" Bíochem., 1977,72,247 .

ll4. Nagai, U., and Kobayashi, J., Tetrahedron Lett., 1976,33,2873

115. Sheehan, J" C., Chapman, D. W., and Roth, R. W., J. Am. Chem. Soc.,

1952,74,3822.

116. Sheehan, J. C., Goodman, M., and Hess, G. P., J. Am. Chem.\oc.,1956,

78, 1367.

Reþrences 178

Il7. Fling, M., Minard, F. N., and Fox, S. W., .f. Am. Chem. Soc., 1947, 69,

2466.

118. (a) Pirkle, W.H., and Welch, C.J., J. Org. Chem., 1984,49, 138, and

references cited therein; (b) Pirkle, W. H., Finn, J. M., Schreiner, J. L.,

and Hamper, J. C., f . Am. Chem. Soc., 1981, L03,3964; (c) Pirkle, W.

H., and House, D. W., J. Org. Chem.,1979,44, 1957.

119. Adam, J., Gosselain, P. 4., and Goldfinger,P.,Nature,1953,l7L,704.

I20. Williams, D. H., and Fleming, I., Spectroscopic Methods in Organic

Chemistry, 3rd ed., McGraw Hill Book Company, Maidenhead, U.K.,

1980.

I2I. Liotta, C. L., and Hanis, H. P., J. Am. Chem. Soc., 1974,96,2250.

122" Srinivasan, 4., Richards K. D., and Olsen, R. K., Tetrahedron Lett.,

1976,33,891.

123" Poisel, H., and Schmidt, U., Chem. Ber., 1915,108,2547 "

I24. Srinivasan, 4., Stephenson, R. W., and Olsen, R. K., J. Org. Chem.,

1977,42,2256.

125. Nitz, T. J., Holt, E. M., Rubin,8., and Stammer, C.H., J. Org. Chem.,

1981,46,2667.

References I79

126. (a) Waters, Vy'. A, Mechanism of Oxidatíon of Organic Compounds,

Methuen and Co. Ltd., London, 1964; (b) Walling, C., Free Radicals in

Solution, John Wiley and Sons, New York,1957.

I27. Hammond, G. S., Hawthorne, M. F., 'Waters, J, H., and Graybill, B. M.,

J. Am. Chem. 5oc., 1960, 82, 704.

128. Griesbeck, A. G., and Mauder, H., Angew. Chem., Int. Ed. EngI., 1992,

31,73.

129. Goering, H. L., Eikenberry, J. N., Koermer, G. S., and Lattimer, C. J.,

J. Am. Chem. Soc., 19J4,96, 1493.

130. Clarke, S., Hider, R. C., and John, D. I., J. Chem. Soc., PerkinTrans. l,1973,230.

131. Jain, T. C., and McCloskey, J.9., Tetrahedron,I975,3l,22ll

132. Altman, J., Gilboa, H., and Ben-Ishai, D., Tetrahedron, 1971,33,3173

133. Hassner, A., and Alexanian, Y ., Tetrahedron Lett., 1978, M75.

I34. Bowden, K., Heilbron, I. M., Jones, E. R., and Weedon, B. C., J. Chem.

Soc., 1946,39.

135. Osby, J. O., Martin, M. G., and Ganem,B.,Tetrahedron Lett.,1984,25,

2093

References 180

136. March, J., Advanced Organic Chemistry, 3,d ed.,Iohn Wiley and Sons,

New York, 1985, pp.220-222.

137. Sheehan, J. C., and Frank, V.S., J. Am. Chem.5oc.,1949, 71, 1856.

138. Vogler, K., Helv. Chim. Acta,1950, 33,2I1I.

I39. Takahata, H., Ohnishi, Y., Takehara, H., Tsuritani, K., and Yamazaki,

T., Chem. Pharm. Bull. Jpn., 1981, 29, 1063

140. Miller, M. J., and Mattingly, P. G., Tetrahedron, 1983, 39,2563.

l4l. Easton, C. J., J. Chem. Soc., PerkinTrans.l, 1985, 153.

I42. Barrow, K. D., and Spotswood, T. M., Tetrahedron Lett., 1965,3325"

143. Wells, J. S., Trejo, W. H., Principe, P. 4., and Sykes, R. 8., J. Antibiot.,

1984,37,802.

144. Lowe, C., Pu, Y., and Vederas, J. C., J. Org. Chem.,1992,57, 10.

I45. Herbert, R. 8., and Knaggs, A. R., J" Chem. Soc., PerkinTrans. 1,1992,

103 and 109.

146. Zaugg, H. E., Org. React., 1954,8, 305.

147. Danheiser, R. L,., and Nowick, J. S., J Org. Che.m.., 1991, 56, 1176.

References 181

148. Cardani, S., DeTomâ, C., Gennari, C., and Scolastico, C., Tetrahedron,

1992,48, 5551.

I49. Thaler, W., ./. Am. Chem. Soc., 1963,85,2607

150. Rando, R. R., Nature (London), 1974,250, 586

151. Castelhano,4.L., Plivra, D.H., Taylor, G. J., Hsieh, K. C., and Krantz,

A., J. Am. Chem. Soc., 1984, 106,2734.

I52. Scannell, J. P., Preuss, J. L., Demney, T. C., Sello, L. H., Williams, T.,

and Stempel, 4., J. Antibiot., 1972,25, I22.

153. V/olkoff, P., J. Org. Chem., 1982,47, 1944.

154. DePuy, C. H., and King, R. W., Chem. Rev., 1960,431.

155. Kopping, B., Chatgilialoglu, C.,Znhnder, M., and Giese,8", J. Org.

Chem., 1992,57,3994; (b) Giese, 8., Damm, W., Wetterich, F., and

Zeitz, H.-G., Tetrahedron Lett., 1992,33, 1863; (c) Giese, 8., Damm,

W., Roth, M., and 7nhnder,M., Synlett, 1992,441.

156. Mok, P. L. H., and Roberts, 8., J. Chem. Soc., Chem. Commun., 1991,

151.

I57. Guindon, Y., Yoakifl, C., Lemieux, R., Boisvert, L., Delorme, D., and

Lavallee, J.-F., Tetrahedron Lett., 1990, 31,2845.

Reþrences I82

158. Lusztyk, J., Maillard, B., Lindsay, D. 4., and Ingold, K. U., J. Am

Chem. \oc.,1983, 105, 3578.

159. Beckwith, A. L. J., and Pigou, P. E., Aust. J. Chem.,1986,39, 1151.

160. Bell, H. M., and Brown, H. C., J. Am. Chem. Soc., 1966,88, 1473.

161.

162.

Kim, S., Kim, Y. J., and Ahn, K. H., Tetrahedron Lett., 1983, 24,3369"

Berti, G., Bottari, F., Macchia, 8., and Macchia, F., Tetrahedron, 1966,

22, lgg.

163. Berti, G., Macchia, 8., Macchia, F., and Monti, L., J. Org. Chem., 1968,

33,4M5.

164. Cavanaugh, J. R., J. Am. Chem. Soc., 1968,90, 4533.

165. Still, \V. C., Kahn, M., and Miffa, A., J. Org. Chem.,1978,43,2923

166. Harwood, L. M., Aldrichimica Acta., 1985, 18,25

161. Perrin, D. D., Armarego, W. L. F., and Perrin, D. R., Purification of

Laboratory Chemicals,2nd ed., Pergammon, Oxford, 1980.

168. Vogel, A. I., A Textbook of Practical Organic Chemistry including

Quantitative Organic Analysis, 3,d ed., Longman, London, 1977.

t69. Hoogwater, D. 4., Reinhoundt, D. N., Lie, T. S., Gunneweg, J. J., and

Beyerman, H. C., Recl. Trav. Chim. Pays-Bas., 19J3,92,819.

References 183

170. Wunsch, E., Drees, F., and Jentsch, J., Chem. Ber., 1965, 98, 803

I7l. Hoffmann, E., and Schiff-Shenhav, H., J.Org. Chem., 1962,27, 4686.

172. Banks, G. R., Cohen, D., Pattenden, G. E., and Thomas, J. A. G

J. Chem Soc. (C), 1967, 126"

173. (a) Schwyzer, R., and Ludescher, U., Biochemistry, 1968,7,2514; (b)

Greenlee, W. J., and Thorsett, E. D., J. Org. Chem., 1981, 46, 5351"

174. Horton, W. J., and Thompson, G., J. Am. Chem.Ïoc.,1954,76, 1909.

I75. Marchand, J., Pais, M., and Jarreau, F.-X., Bull. Soc. Chím. Fr., 1971,

3742.

176. Schwyzer, R., and Caviezel, M., Helv. Chím. Acta.,I97l,54, 1395.

177. Ville, 4., Comte,P.,Zwingelstein, G., and Favre-Bonvin, J., Bull. Soc

Chim. Fr.,1958, 1352.

Appendices 184

Appendix 1

lH n.m.r. spectral data of the p-bromophenylalanine derivatives (82), (85),

(115), (128) and (137), and the p-bromotyrosine derivatives (9a) and (95).

tnrhN¿¿,,,. R an,hNrrr,..

.,.tf

Bf Br

R' R'

(82a)

(9aa)

(1 15a)

(128a)

(85a)

(95a)

(r37a)

(82b)

(e4b)

(1 15b)

(128b)

(85b)

(e5b)

(137b)

R=OMe, R'=HR = OMe, R' = OAc

R = OCMe¡, R' = H

R = OMe, R' = NOz

R = NHCMo3, R' = H

R = NHCMe3, R' = OAc

R= OH, R'= H

R=OMe, R'=HR = OMe, R' = OAc

R = OCMe¡, R' = H

R = OMe, R' = NOz

R = NHCMe3, R' = H

R = NHCMe3, R' = OAc

R=OH, R'=H

8.031.381.401.483.833.813.82R

erythro-

ßomers to.411.5LL.4to.410.310.410.5Jnß

5.866.086.055.8.55.975.935.91p

5.645.30s.325.495.595.565.59ct

(r37b)(e5b)(85b)(1 15b)(128b)(94b)(82b)

6.641.05r.o21.163.593.563.55R

threo-

rsomers rt.211.811.8tt.4tr.2rt.2tt.2Jaß

5.976.226.t76.066.O26.036.02p

5.555.175.225.475.515.465.52c[

037a\(95a)(85a)(1 15a)(t28a)(o4a\(82a)

Appendices 185

Appendix 2

lH n.m.r. spectral data of thep-hydroxyphenylalanine derivatives (1 LZ),QI6), (1 I 8) and (129)

and the p-hydroxytyrosine derivatives (121), (I22), (I23) and (I24).

R R

R' R'

(rr2a)(t22a)(r2La)(tze)(1 16a)(1 18a)(r24)(r23)

OMe,OMe,OMe,OMe,OCMe3,NHCMe3,

_H=OH= OAc= NOz_H_H=OH= OAc

(rrzb(r22b(tzrb

R'= HR'= OHR' = OAc

Me,Me,Me,

) R=O) R=O) R=O

RRRRR

R_R_R_R_R_ft=R_R-

(116b) R=OCMe¡, R'=H(118b) R=NHCMe3,R'=H

NHCMe3, RNHCMe3, R

1 15t.283.743.783.72R

erythroisomers

4.254.604.984.274.27OH

4.92.t2.3Ja.on

s.395.495.545.495.46p

8.38.08.28.68.4Ja,ß

4.614.955.024.974.95c[

(1 18b)(116b)(121b)(r22b)(112b)

1 3 1t.271.30t.523.893.783.853.86R

tlveo-isomers

4.434.724.41.5.255.344.985.O25.13OH

8.17.58.210.610.010.310.4Json

5.63s.625.635.675.795.675.645.7 |p

6.36.96.24.64.45.04.84.6Ja.ß

5.085.1 15.115.445.535.485.445.51c[

( r23)(124\(1 I 8a)(1 16a)(129\(I2la\(122a\(ll2a\

Appendices 186

Appendix 3

Crystal sEucture of

(Z )-N - t e rt- butyl-Nø-phthaloyl-o, B-dehydrophenylalaninamide ( 86)

o(21) c(32)

N(2) c(31)

o(28)

c(2) c(3)

N(3)c(1)

c(4)c(4a)

o(1)

C(4c) c(4b)

Appendices 187

Bond distances (,Å,) for

(Z ) -N - t e rt-butyl-Nø-phthaloyl-a, p-dehydrophenylalaninamide ( 86)

c (1)c (28)c (21)c (1)H (3N)c (3)H (3)c (48)H(41)H (43)H (4s)H (47)H(4e)c (23)c (241c (2s)c(26tc (271c (28)c (36)a(32',,H (33)H (34)H (3s)

o (1)o (28)N (2)N (3)N (3)c(2,c (3)c (4)c (4À)c (4À)c (48)c(4c)c (4c)c(22',,c(23)c (241c(2s)c(261c (21\c (31)c(32)c (33)c(34)c (3s)

1 .236 (3 )

1 - 211 (3)1.4ls (3)1.337 (3)o.824 (26'.,1 .336 (3 )

o -962 (2411.s01(4)0-9?00-9700-9?00-9700.9701 -386 (3)1.385 (4)1.383 (4)1.390(4)1.383 (3)1 .4 86 (3)r.390 (3)o -921 (2610:933 (32)0.960 (31)I .019 (29)

c (21)c (21c (28)c(4)c(21c (31)c(44)c(4c)H(421H(44)H(45)H (48)c(221c (211H (23)11(241H (2s)H(261c(321c (33)c(34)c(3s)c(36)H (36)

1.20s (3)1- 431 (3)1. 404 (3)1.480 (3)r. s02 (3)1.{62(3)1.s21(4)r.s11(4)0. 9700. 9?00.9?00.9701.48s (3)r.38s (3)0. 930 (30)0.930 (29)0. 935 (36)1.011 (27)1.401 (3)r-3?5(4)r.364 (4)1.390(4)1.379(4)o.941 (211

(2oN

N

N

cccccccccccccccccccc

2

2

3

1

3

4

4)4À)

( 4B)(48)( 4c)(2rl(221(231(241(2st(261(31)(32 )

(33)(34 )

(3s )

(36)

Bond angles (deg.) for

(Z )-N - te rt-butyl-Nø-phthaloyl-o, B-dehydrophenylalaninamide ( 86)

c (2rlc (281H (3NlN (3)c(2)c(3)c (31)H(3)c (¡¡B)c (4clc (4clH 142lH (43)H ({3)H ({slH (r6lH({6)H(48)H ({9}H({e}c(221c (23)c(2'tlH (231c (2slH (24)H (2s)c l21lH(26)c (28)N (21c (2-r)c (36)c (33)H (32)H (33)c (3st

, H (34)H (3s)

; c (3s)H (36)

- N(2)- N(2)- N(31- c(1)- c(1)- c(2)- c(3)- c(31- c({)- c(4)- c(4)- c({À)- c(4À)- c({À)- c(48)- c(48)- c(48)- c(4c)- c(4cl- c(4cl- c(21)- c(221- c(221- c(231- c(241- cl24l- c(2s)- c(261- c(26)- c(2?)- c (2B)- c (28)- c (311- c(32¡- c(321- c(33)- c(34)- c(34)- c (3s)- c(36)- c(36)

- c(2t- c(21'- c(tl- o(1)- N(3)- N(2)- c(2)- c(3r.1- N(3)- N(3)- c(48)- c(4)- c(4)- H (42)- c(4)- c({)- H ({s)- c(4)- c(4)- H (48)- o(2r)- c(211- c(23)- cl22l- c(23)- c (2s)- c(24l.- c(2s)- c(2?)- c(221- o (28)- N(2)- c(3)- c(3rl- c (331

- c (32)- c(33)- c (15)- c(34)- c(31)- c (35)

109109109109r09129L29l2lr2lt22L22L20rl7r22108L24t05119t2Ltl9tl7tt912312012tlt 9

L25 - 6 (21r1r.6(2)tt?.6(18)L24 -2 (21tr5.?(2)r22 -5 (21t29 -8 (21r1{-2(14)r10.3(2)109-5 (2)lll.9(3)r09.3 (2)109.6 (2)109. s (-)r09.3(2)

l2(2(2

(2t(-)(2t(2t(-)

2lr8)2Ll2trsl

)))

8)

c (28)c({)H (3N)c(2)c(t)c (3)H (3)c (4À)c (48)c (4cl¡{(41)H (42)H (431H(44)H (4s)H(46)H (4?)H ({8)H (49)N(2tc (221c (2't,c(241H (23)H (24)c(26)H (2slH (26)c (26)c (28)c (2?)c (32)c (36)H (32)c (34)H (33)H (3{)c (36)H (3slH (r6)

- N(2)- N(31- N(3)- c(11- c(2)- c(21- c(3)- c(4)- c(4)- c(4)- c (4Àl- c(4Al- c(4À)- c(48'- c(4Bl- c (4Bl- c(4c)- c(4c)- c(4cl- c(21t- c (211

- c(221- c (231

- c (23t- c(24'- c (2s)- c (2s)- c (26)- c(211- c(211- c (28)- c(31)- c(3r)- c(32)- c (33)- c (33)- c(34)- c (3s,- c (3s)- c(36)

- c(21- c(r)- c({}- o(11- N(21- c(1t- c(21- N(3)- c(4À)- c (4À)- c(4t- H (4r)- H({r}- c(41- H ({4}- H (¡r4)- c(41- H (47'- H(¡tl)- o(2r,- N(2)- c(2r)- c(221- c(24',,- c (23)- c (24'- c (26)- c (25)- cl22l- c(26)- o (28)- c(3t- c{32)- c(31)

c(12)- c(r4)- c (33)

c (]{)- c (36)

c(31)

t22t2611st201l-tr20ll6106108).0 9

1.0 9

109I09109I09t09109109109125105108tl6L2l115l2Ilr8t20121130r29\231ì?119t20l2ltr7Ì20tl9t19

3 t2te (2tI (18)1(2tI (2)3 t2t0(r4)1(2'ts (3)I (3)5 l2ts (-)s (-)6 l2',ts (-)s (-)5 (2)s (-)s (-)o (2't4l2lI (2)6 l2t6(r8tB(T8)o(2t2(2rl8(r5)5 l2tI (2)4 12tt (2)6 t2t5(r5)6 (3)8(r9)2(ì8)0(3)4 (r6)r (ì7)

3 t2t6 (2)9l2l2 (210 (3)2 (1s)3 (19)? (3)1(18)5(rs)I (2tB(11)

Appendices 188

Appendix 4

Crystal structure of

( E )-N -t ert-B utyl-Nø-phthaloyl-cr, p-dehydrophenylalaninamide (87)

C(4c)

c(4b)

C(aa) c(4)N(3)

o(1)c(31) c(32)

c(1)

c(2) c(3)

o(28)

N(2)

o(21)

Appendices 189

Bond distances (,Ä.) for

( E ) -N -t e rt-B utyl-Nø-phthaloyl-cr, p-dehydrophenylalaninamide (87)

c (1)c (28)c (21)c (1)H (3N)c (3)H (3)c (48)H (41)H (43)H (4s)H (47)H (4e)c (23)c (241c (2s)c (26)c (211c (28)c (36)H (32)H (33)H (34)H (3s!

o (1)o(28)N (2)N (3)N (3)c(21c (3)c(4)c(4À)c (4À)c(48)c(4c)c (4c)c(221c(23)c(241c (2s)c(261c(2'tlc(31)c (32)c(33)c(34)c(3s)

1.216 (4)1.197 (4)1 .407 (s)r.340 (s)0.8s4 (4e)1.331(s)0 - BB6 (33)1 . s20 (6)1-046 (71)1.040 (64)1-10s(63)I .008 (43)o - e26 (s9)1.376(s)1.399 (6)I .312 (6',t

1.3?s (s)1-392 (A',)

1.486 (s)1.381(s)1 .007 (s0)o.es3(47)0. e83 (s2)0.92s (4s)

c(2L',1c (21c (28)c(4)c(21c (31)c (4À)c (4c)H(421H(44)H(46)H (48)c (221c(211H (23)H(241H (25)H (261c (32)c (33)c (34)c (3s)

o (21)N(2)N (2)N (3)c (1)c (3)c(4)c (4)c (4A)c (48)c (48)c (4c)c (21)c(22',tc (231c (241c (2s)c(26)c (31)c(32)c (33)c(34)c (3s)c (36)

1-205(4)r - 428 (411- 406 (4)1- 4 67 (s)1 - s0s (s)1 - 4?5 (5)1 - s21 (6)1 - s1s (s)0.80s(68)0. 909 (s3 )

1.002 (so)0- 966 (4s)r.49s(s)1-3?1(s)0.941(36)1.000 (43)1.045 (3s)0 - 933 (36)1.388 (s)1-380(6)r - 360 (7)1.3se (8)1.394 (6)0. 887 (39 )

c (36)H (36)

ond angles (deg.) fo

( E )-N + e n-B utyl Nø-phthaloyl-a, p-dehydrophenylalaninamide ( 87)

c (211c (28)H (3N)N (31c(2)c (31

c (31)H (3)c (4Blc (¡lclc(4c)H (421H (431H(431H (4s)H ({6}H(46)H (48)H (4slH(49)c (221c (23)c(211H (23)c (251H(24)H (2s)c t21lH (261c (28)N (2)

c (36)c (33)H (32)H (33)c (35)H (34)

l" H (3s)c (3s)H (36)

- N(2)- N(2)- N(3)- c(1)- c (t)- c(21- c(3)- c(3)- c(4'- c({)- c(41- c (4À)- c(4À)- c(4Àt- c(48)- c({B}- c ({B}- c({cl- c(4cl- c (¡rc)- c(211- c(221- c(221- c(231- c(2{}- c (2{}- c (2sl- c(26)- c (26)- cl21l- c (281

- c (28)- c(3rl- c (32)- c (32)- c (33)- c (34 )

- c(34)- c(351- c(36)- c (36)

- c(2t- c (2rl- c(r)- o(11- N(3)- N(2)- c(2)- c (31)- N(3)- N(3)- c (48)- c(4)- c(4)- H (42)- c(4)- c(4)- H (4s)- c(4)- c(4)- H ({8}- o (21)- c (21)- c(23)- c(221- c (23)- c (2s)- c(24)- c (2s)- c(211- c(22t- o (28)- N(2)- c(3)- c (3r)- c (33)- c (32)- c (33)- c (3s)- c(34)- c(3rt- c(3s¡

r2s-9(3)ll2. r (3)rr6.3(30)124.9 (3)lts-3 (31

rr9.5(3)121.3(4)rr4.2(r?)106.2 (4tr09-8(31109-0(4)111.1(39)104 - 4 (35)Llt-3(52)lo5-3(32)1O8.6 (26)r01.4(38)107.0(23)1tr-6(38)106.4 (421129-3 (3)129.9(3)122-0(3)r19.6(21)r22 .3 (41121.r(24)120- 0 (20)lr'7.¡t(41lI?.o(22)109 - 6 (3)r24.3(31104.9 (l)122.5(4)Ì20-4 (s)r19.1(28)rr9 8(241120-6(s)125.8(28t130-4(321tI9_7(5)114.8(25)

c (28)c(4tH (3N)c (21c(11c(3)H (3)c (4À.)c(4alc(4clH(4r)H (421H ({3)H({4)H (4s)H(461H (47)H ({8}H (491N (2)c l22lc(2llc (241H (23)R (2{)c (26)H (2slH (26)c (26)c (2Blc (271c (32)c (36'H (32)c(34)H (33)H(34)c (36)H (3slH (361

- N(2)- N(lt- N(3t- c(1)

c(21- c(2)- c(3)- c(4t- c(4)- c(4)- c(4À)- c (4À)- c(4Àl- c (48)- c(48)- c(,[B)- c(4cl- c (4c)- c(4c)- c(2Ì)- c(2r)- c(221- c(231- c(23)- cl24'l- c (2s)- c(2s)- cl26l- cl21l- cl2ll- c (28)

c(31)c(3t)

- c(32)- c(33)- c (31)- c(14)- c(3s)- c (351

- c(36)

- ct2lc(I)

- c(4)o(I)N(2)

- c(r)- c(2)- N(3t- c(4À)- c(4À)

c(4)- H (41)- H(4r)- c(4)- H(44)- H(44)- c(4)- n(47)- H(41)- o(2t)- N(2)- c(21)- cl22't- c(24)- c(231- c(24)- c(26)- c(25)- cl22l- c(25)- o(28)

c(l)- c(12)- c (31)- c(32)

c(34tc(ll)

- c(34)c{16)c(lU

l2l-7(3tì26.r(4trls.B(3r)rl9 8(3)D.1.5(3)126.8(3)rr8.5(r-r)r09.-r(3)rrr.3(s)1r0-r(511r2.6(36)rr0.9(58t106.3(49)r04.0(32)1t4.2({7)t22-3(42t1r0.8(21)IO0.6 (34 )

119.0({01125.4(3)ro5-3 (3t108- r (3)1r6.1(41t24.3(2r)rl6-?(25)r20-8(4)118-9(2O)t25 .5 l22l121-3(3)I29-I(4)r30.9(3)1r8.6(3)118 9(4)120.5(28)120.0(5)tzo 2 l24lÌ1r.6(28t120 r (s))o9 2 (33)t25 . 6 125',t

Appendices 190

Appendix 5

Crystal sfructures of

(Z ) -N -phthaloyl- a, p-dehydrophenylalanine methyl ester (8 8)

I'aOla

028a Ol'aC75a 7a ClaC)-8a

N2aC22'

C24aC23a ah O2la C3a

C31a

C32a C36a

C33a C35a

C34a

bolb

a5b cz6bo28b orb

c28b 1b

Q4b N2bczb

cz3b cz2bc21b 021b

c3b

c31b

c32b c36b

c34b

c35b

Appendices 191

Bond distances (,Ä.) for

(Z)-N -phthaloyl-o, B-dehydrophenylalanine methyl ester (88)

atom

o (1a)

o (1' a)

o(1'a)

o(1'b)

o(1,b)

o (Ib)

O (21a)

o (21b)

o (2Bb)

o (2Ba)

N (2a)

N (2a)

N (2a)

N (2b)

N (2b)

N (2b)

c (1a)

C(1'a)

c(1,a)

c(1,a)

c(rb)

c (r, b)

c (t' b)

c ( r'b)

c (2a)

c (2b)

disLance

1.1?6 (6)

1.315 (5)

1.449 (?)

1.320 (5)

1.441(?)

1.191 (5)

1.187 (s)

1.188 (s)

1.188 (s)

1.201 (st

1.412 (5)

1.407 (6)

1.3?9 (6)

1.41? (5)

1.404 (s)

1.465(6)

1.07(4)

1.07 (6)

0.Bs(s)

r.47216',,

0.90(s)

1.05 (s)

0.92(6)

1.311(6)

1.318 (6)

di stance

1.443 (6)

0.99(4)

1.443 (6)

0.98 (3)

r.472('t',,

r - 412 (6\

1.371 (6)

1.3?4 (6)

r.3't2 (6',,

1.3s6 (6)

1.380 (B)

0.9s (s)

1.3?0(?)

0-91(4)

1.360 (7)

1.02 (41

1.349(8)

0.98 (4)

1.376(?)

r.13 (5)

1.375 (7)

0.95(4)

1.363 (6)

0.9s(4)

r.3?0 (6)

0.99 (4)

distance

1.466 (6)

1.46s(6)

r.379 (?)

1.377 (6)

1.3?s(6)

1.383 (6)

1.3?3 (B)

0.94 (s)

1.36s (7)

0.92 (3)

1.3s6 (7)

0.93(4)

1.352 (B)

1.06 (5)

1.341(?)

r-.01 (4)

1.3s6 (7)

0.94 (4)

1.361(?)

1.02 (s)

1.3?5 (?)

0.99(4)

r.0B(4)

0.9? (3)

atom

C (1a)

c (1a)

C (1'a)

c (1b)

c(1,b)

c(rb)

c (21a)

c (21b)

c(2Bb)

C (28a)

c l2a')

c (21a)

C (2Ba)

c (2b)

c (2lb)

c (28b)

c (2al

H (1' a)

H (1', b)

H(1'c)

c (2b)

H (1'd)

H (1'e)

H(1'f)

c(3a)

c (3b)

atom

c (3b)

c (3b)

c (3a)

C (3a)

c (21a)

c (2lb)

c(22b')

c(22b1

C (22a)

C (22a!

c (23a)

c (23a)

c (23b)

c (23b)

c (24b)

c (24b1

c (24a)

c (24a!

c (25a)

c (25a)

c (25b)

c (25b)

c (26b)

c (26b)

C (26a)

C (26a)

atom

c (3Ib)

H (3b)

C (3la)

H (3a)

C (22aI

c(22bt

c (23b)

c(2'tb')

C (23a)

C (2'l a)

c (24a!

H (23a)

c (24b',

H (23b)

c (25b)

H (24b)

C (25a )

H (24a',

C (26a)

H (25a)

c (26c'l-

H (25b)

c (z'tbl

H (26b)

C (27 al

H (26a)

atom

C (2'l al

c (27b)

C (31a)

C(31a)

c ( 3lb)

c ( 31b)

C (32a)

c (32a)

c (32b)

c (32b)

c (33b)

c (33b)

C (33a)

c (33a)

C (34a)

C (34a)

c ( 34b)

c ( 34b)

c (3sb)

c (3sb)

c (35a)

C (35a)

C (36a)

c(36b)

atom

c (2Ba)

c (2Bb)

c (32a)

c (36a)

c (32b)

c (36b)

c (33a)

H (32a)

c (33b)

H (32b)

c (34b)

H (33b)

C (34a)

H (33a)

c (35a)

H (34a)

c (35b)

H ( 34b)

c (36b)

H (35b)

C (36a)

H (35a)

H (36a)

H (36b)

1 5943

Appendices 192

Bond angles (deg.) for

(Z ) - N -phthalo yl-cr, p-dehydro phenylalanine methy I es ter ( 8 8 )

c(r.t otl'¿r ctl'ål

ctlbt o(l'br c(r'bt

c{2åt Nl2å) ct2låt

ct2àl Nl2¿r c{20àl

C(21.1 N(2.¡ ct28ål

c{2b} Nt2bt ct2tbl

ct2b) N(2b) c(28Þl

c(2rb) N(2br c(28b1

o(r.l c(l.l o(¡'.1

o{l.l C(ròl C{2.1

O(l'ål C{1.¡ Cl2¡l

olr'àl c(r'.t fl(r'àl

O(l'.1 C(1'¡r [(l'Þl

Oll'.1 Cl¡'.1 H(l'c)

trlr'àl C{l'.t BII'Þl

r(l'¡, c(l'¡t HtI'c'

f,(l,bt c(1,ât H(1,c)

o(r,bt c(rbt o{tbl

o(r'bt c(rb¡ c(2b)

o(lb) c(lbl c(2b)

o(1.b, c(r,bt H{1,d)

o(r'bt c(r'bt B(r'e,

o(r'b) c(r'b) x(r'rl

É(l'dl CII'bl H(¡'et

Í(l'dt c(l'bt fl(l'ft

H(Ì'êl C(1'bl H(l'f)

Ct2a.l C(21¡) H(21.1

c{22br c(2lbt c(2{bl

c{22bt c(2lbt r{21Þl

c(?rþt c(2lbr fl(23b)

ct2lbt c(2abt cl25bl

ct2lbt c(2rbt fl(2{bl

c t25bì c (2abt H (2{Þl

c(21.! C(2aÀl C(25å)

ct2làt c(2ràt ü{2.Àl

c(25à' C(2aâl H(2a¡l

c(2.!l ct25¡) c(26à)

Ct2a.l C(25.1 H(25¡)

c{26åì c(25àl ¡(25¡l

c(24bt c(25b, cl26bl

c{2{Þt ct25bt fl(25b)

ct25bt c(25b1 f(25b)

c(25b) c(26bt c{2?b)

c(25b, c(26bt fl(26b1

c(2?bl c(26bt H(26b1

C(25.1 ct26àl Ct2?å)

c(25¿) c(26ål 8(26à)

C(21ôì c(26Àl ff{26à)

C{22Àl C(21¿l Cl26ål

¡ræ ¡tú .tm

N(2al C{2.1 C(l¿l

N(2¡l c(2¡) c(l.l

cll¿l c(2.) c(lál

N{2bt c(2bt c(lbt

N(2b) C(2b) Ctfbl

c (rb, c (2bt c (3bl

ct2b) c(lb) c(3rÞ)

c(2bt c(3bt x(3bt

c(3lb) c{3b} f(fbl

C(2¡, c(là, c(3r.)

C(2.1 C(3.1 ff(3àt

c(fl¡) c(3.1 a(3.1

o121., C(21¡l N(2¡)

o(2r¡l c(21¡t c(22r)

N(2¡) C(21¡l Cl22àl

o(21b, c(21b) N(2b)

o(2lb) c(2rb) c(22b1

N(2bt c(2rb, ct22b)

c(2rbt c(22b) c(23bt

c(2lbt c(22b) c(27b)

c(23b) c(22bt c(2?Þl

c(21.t c(22à) c(23à)

c(21à) c(22àl c(21.1

c(23ål cl22¡) c(27öl

C(22.t C(23¡) C(2iât

c(22.1 cl23¡) f,(23¡l

c(22bt c(2lbt c(2lbì

c(26bt c(2rbt c(2rbl

o(28¡, c(28àl N(2âì

o(28âl c(28¡l c(21àl

N(2àl c(28äl C(21àl

o(28bt ct28bl N(2bl

o(28b) c(28Þ' C(2?br

N(2bl c(28bt c{2?bl

C(3.1 c(31.) c(32!)

c(3!l c(31â) cl36àl

c(32.1 c{3ràl cl36àl

c(3bt c(3rbt c(32b'

c(3b, ct3rbl c(36b1

c(32bt c(lrbt c(36b1

c{31.1 c(32àl C(33.1

c(31.1 c(32å) H(32.1

c(3!.1 c{32!} f,(32.1

c(3rbt c(32b1 c(33b)

c(3lbt c(32bt fl(32b)

c(33b, c(l2b) fl{32b,

c(l2b) c(33b1 c(3tbl

c(32bt c(3lbl ¡(33b1

c(lrb) c(33b) l(llb)

c(32.1 c(33a1 c{34.)

c(32ò) c(33å) ff133¿l

c (34à) c (33å) I (314,

c22 C21 C2S

tt5.a(5)

ll5_5({l

r2a.2{5t

r23.6 (5t

rrr 3(a)

120-9(.t

t22 -6 tll

1rÌ.0{ll

r2r.l t6t

r2l 3 (6)

r12-5(51

r0? (31

r03 {. I

r0{ {a)

Il2 (a)

r09 {5t

rl9 (6,

lzr.0 (5)

r11.9(5t

l2{.1(51

r03(r,

r0l (lt

rr? (5,

rrr (5t

12t (6)

97 (5t

129 (l)

t¡?.å{5t

r2l (ì

l¡8 tll

r20.9(61

r2r (2)

ll9 (2)

r22.6 (6)

rr9 (lt

r¡8 (31

r20.5 (6)

12? (31

rr2(ll

r21.7 (51

tr9 (3)

1r9 (31

lrl-a (5)

r22 (3)

r20 (3t

1r1.0(6t

r22 (3t

r21 (3t

r22 5 t5'

¡08.0(5)

r29.5(5t

¡2r -2 (5)

r20-9 (6t

It2.5 f5)

r2J.6t5t

r23.9t5t

tt2. I (al

r2{ - l (51

t23.2 tsl

r32.5(5)

11t (2)

rr4 (2)

13r.9(5)

rrr (2,

lla (2)

12a-?(51

130.r(6)

¡0a .5 (5,

12a - ? (5)

r29.9 (s)

r0s.a (a¡

r30.0(st

r08.9(5'

r20.9 (51

129.? (6'

109.5t5t

120. I (5'

116-6(61

lla (!t

t08 t (r)

r30.6 t5r

¡2{. t (51

L29 2t6l

r06.5(st

¡23- 3 (5r

r30. ? {s,

r06-o(4t

125.I (6'

rr7.2(5t

117.5 (51

L25.2 tsl

!r6.9(s)

rr6- E (5ì

l2t.lf5t

lr9 (3t

tt9 (3)

l2t.r(5t

r20 (2,

rr9 (2t

t20.4 (6)

l1l (3t

r22 13)

l¡9 a (6t

116 t3)

r2l(3)

C(ll.l

c(ls.t

c (l3bt

c (33b1

c (35b1

c{3abl

c (lrbl

c (36b)

c(3{¡l

c ( 31.1

c (36.t

c (3r.)

c ( 31¡,

c (35rt

c (3rbl

c(3lbl

c (35b)

c ( la.l

cflaàl

c(labl

c(3abt

ct3{b)

c ( l5bt

c(35b1

c (l5bt

c(35àt

cll5ål

c (35.)

c(36àl

c (l6rl

c (16à I

c{16b,

c (36b)

c (36b1

{lla¿l

H(la¿l

c(l5bl

ff ( t{bt

n ( labt

c(l6bl

H (lsbt

I (l5bt

c(16Àl

H(15àt

f ( 15.1

c(15¿l

fl(36àl

H (36¿l

cll5bl

H (36bt

H (36b)

tl9(2t

120 t2,

tl9 6(6r

ltltlt

l2l l3r

r20_2 {6t

t20 f l¡

rl9(lr

r20 2 t6¡

r2l o)

rt6(3t

t20-r(61

t20 12l

tl9(2t

l2r - 5 t6t

tl7 (21

l2t {2t

ct26.t c(2?àt c(28a,

ct¿2bt c{21Þr c(25b1

c(33ål c{34a1 C(15ål

Appendices 193

Appendix 6

Crystal structure of

( 2 S 3 R ) -3 -hydroxy-N-phthaloylphenylalanine meth yl ester (l l2a)

q32)

c(31)

o(21)c(21) q3)

c(?21 o(3)

q2)o(1)

c42n q28)c(l)o(28)

c(1)o(1)

Appendices 194

Bond distances 1Å¡ for

( 2 S 3 R ) -3 -hydroxy-N-ph thaloy lphenylal anine methyl ester (l l2a)

c (1)c (1')c (3)c (29')c(2tlc(2tH(1'B)H(21H (3)c(221c (271c (24tc (2s)c(26tc (27')c (32)H (32)H (33)H(34)H (3s)H (36)

ooooN

cccccccccccccccc

1)1')3)28t2',)

1)1')2t3)2tl22)

1.192(8)1.443 (8)1.419 (9)1.203 (9)1.411 (8)1.537 (10)0.970 (-)1.014(64)0.931 (s2)1.494 (10)1.378 (10)1.38s (10)1.380 (12)1.36?(11)1.388 (9)1.375 (10)0.970 (-)0 - 970 (-)0.970 (-)0.970 (-)0.9?0 (-)

c (1)H (3)c(zL',c (2')c (28)H (1'A)H(1'C)c (3)c (31)c (23',)H(23)H(24',)H(2slH(26',)c(28tc (36)c (33)c (34)c (3s)c (36)

(1')(3)(2L)(2')(2t(1')(1')(2',)

(3)(22',)qz3t(24t

oooNNccccccccccccccc

1.320 (8)0.949(6)1.207(8)1.448 (9)1.39s (9)0.970 (-)0.9?0 (-)1. s46 (9)1 - 500 (10)1.378 (10)0.9?0 (-)0.970 (-)0.970 (-)0.9?0 (-)1.496 (10)1.401 (10)1.406 (11)1.386 (14)1. 354 (14 )

1.363(11)

23t24',)2st26t31)32t33)34)3s)36)

2sl26273132333435

)

)

)

)

)

)

)

Bond angles (deg.) for

( 2 S 3 R ) -3 -hydroxy-N-phthaloylphenylalanine methyl es ter (l l2a)

c(1'lc(2r)c (28)c(2)H (r 'À)H(I'B)H (1,C)c (1)H (2tc (3)c(2)H (3)c (311N (2)c (221c l2', tH (23)c (24'c (251H (25)c (26)c(2?)c 126lc (28)c (211c(32)c(36)c (33)H (33)c(34'c (35)H (35)c (36)H (36)

o(1,)N(2)N(2)c (1)c (1,tc (1,)c(1,)c (2,c(21c(21c (3)c (3)c(3)c (2r)c(21)c(221c (23)c(23)c(24)c (25)c(2s)c (26tc 127',c(271c (28)c(31)c(31)c(32)c(33)c(33)c(34)c (35¡c (3s)c (36)

-c-c-c-o-o-H-H-N-c-c-o-c-c-o-N-c-c-H-c-c-H

-c-c-o-c

1

2I

1r6L2LllrL25109109109110118113lr2106111r23105r08T2L121L2L118119117t2I130LZ9120119120L20120119118118r.2 0

6 (6)2 t6t3 (6)r (7)4 (4)s (-)s (-)5 (5)9 (36)0 (6)? (6)4 (31)4 (6)9(7)9 (6)1(6)6 (s)I (s)3 (8)9 (s)1(s)0 (8)o (?)5(?)3 (7)0 (7)4 (?)2 (8)4 (6)4(-tt4 (10)6(7)7 (6)4 (5)

c (3)c(28)o(1')c(2)H (1'¡ B)H(r,c)H(1,C)H az','c(3)c (3)H (3)c(31)c(31)c (221c (23)c(211c l24tH(24tc (25)c(25)H (26tc(211c(28tN (2)c(211c (36tH(32)c (331c(341H(34)c (35)c (36)c (3s)H(36)

o(3)N (2)c(1)c(1)c (1,)c(1,1c(1,1c(2tc (2)c (2'tc (3)c (3)c (3)c (21)c(22tc(22tc(23)c (241c (24tc (25tc (26tc (261ct21tc (28)c (28tc (31tc(32)c(32tc (33)c(34)c(34)c (3s)c (36)c(36)

- H(3)- c(2)- o(1)- o (1')- o(1')- o (1,)- H(1'Bì- N(2)- N (2)- H(2)- o(3)- o(3)- H(3)- o(21)- c (2r)- c(23)- c(22t- c(23)- H (24 )

- cl24,- c (25)- Hl26t- cl22l- o(28)- N(2)- c(3)- c(3r)- H(32t- c(32)- cf ?1ì- H (34)- c(34)- c(3r)- c (35)

92.4 (5)r2?.0(6)r25. 5 (7 )

r09.3(6)r09.s(s)109-5(4)109.5(-)99.6 (36t

rr,4-5(6199.s(33)

r06.9 (32)114.4(6)104.2 (31)r30.2 (?)129.8(7tr22 .L ('t Irt6.6(8)1r9.4 (5'119.3(5)t22 .0 ('t Ir2r.4 (51

121.6 (5)r08.5(6)L24 .-t l-t I106.0 (61r20.4 (?)119-8(5)r20.0(6)rr9.3(9)r20.3 (7)r20.3(?)L22 -'t (L0lr19.0(9)t20.6(6)

r,)(1'À)(t'À)(2t(1)(1)(3)(2t(21(2Ll(21(21)122',(23 )(23t(24t(251(2s)(22t(26t(28 )

(31- c(32)- c (31)- c(32)- H rllì- c(33)- c(34)- H(3s)- c (31)

Appendices 195

Appendix 7

Crystal structure of

(2 S 3 S ) -3 -bromo-N-ph thaloyl-p - ninophenylalanine methy I es ter ( I 28 b)

o1B13

cl'C1

o2r01' c46

c4L

czl N2t

c22 e8 c4z c44c43o28 o44'a3

c27 N44o.44

e6

es

C3

c2

Appendices 196

Bond distances 1Å¡ for

( 2 s 3 s ) -3 -bro mo-N-phthaloyt-p -nirophenylalanine merhyl es ter ( I 28 b)

Bond angles (deg.) for

(2 S 3 S ) -3 -bro mo-N-phthatoyt-p -nitrophenylalanine merhyl es ter ( 1 28b)

at om

Br(31

o(r)

o(1')

o(1't

o(2r)

o(28t

o(4¡tl

o(4{, )

N (21)

N (21)

N (21)

N(¡l/tl

c(1)

c (21

c (3!

a Com

c (3)

c (rl

c (r)

c(1')

c (21)

c (281

N (44)

N(441

c (2)

c (2r,)

c (28)

c(4¿¡)

cl2',

c (3)

c({1)

distance

r _ 943 (6t

1,184(8)

1.298(8)

1.45(1)

1.189(81

1.169 (8t

1.203 (9t

1. r.99 (9)

1.436(?)

r.3?3(8)

r.385 (9)

1. ¡t4 (]. )

1.515 (9)

1. s24 (9)

1. ¡¡87 (91

a Lom

c (21t

c (221

c (22t

c (23)

c (24',

c (25)

c (26)

c l21l

c(41)

c (41)

c (421

c (43)

c(44)

c (45)

a Èom

c l22t

c (23)

c(211

c 124't

c (25)

c t26t

c tz't I

c(28)

c (421

c (46)

c(43)

c(44)

c (4s)

c (46)

distance

r-458(91

r..3?5(8)

r.37(l)

r.38(1)

r-35(r)

r.35(1)

1.35 (r )

1.464 (9)

r.38(rl

1.363 (9)

r.36(r)

r.36(11

r-33(rt

1.37(r)

c(U

c (2t

c (2t

c(2rt

o(aal

o({al

o(at't

o(U

o(u

ou'tN(2rt

N (2r'

c(lt

Br(31

B!(ll

c(2t

o(2u

ot21,

N(2tt

c(2lt

c(2r,

olt't

N(2tt

N(2tt

x(21)

N(aal

N(til

N(ail

c(r)

c(1t

c(1)

c (21

c 12l

c(2t

ct3t

c (lt

cot

c(2U

c(2ll

c(2rl

cl22l

c(22)

c(r'I

ct2rl

c(20t

c(2El

otta'I

c(aal

c(aat

o(¡')

c(2t

c(2t

ctU

c(lt

c(lt

c(2t

c(.u

c{(rl

f(2rl

ct22l

c a22l

c(23'

ct21l

¡t6.?(61

r2l-r(51

r21.0(61

rr2-t{51

r2z I {9t

tl9-a {9,

ltl-2{9,

126-a (71

¡2a.2(?l

109-a t?)

r09.7 (61

It2-? (6)

rr2-5(6)

r0?. 6 (a )

¡0r-9(5t

rra,I (6t

l2a-2t?l

rl0-1 (8t

r05- 6 (6t

rlr,a(8t

r06.1(6t

c(2lt

ct22l

ct23t

c(2at

c t25l

ct22l

ct22l

c (26)

o(28t

o(2St

N(2¡)

c(3t

c(!t

c(a2r

c(.rt

c (42)

x({ll

x({Í

c(a3t

c(aat

c(alt

c(22t

c(2lt

c(2al

ct25t

c (261

c{2?l

c(271

c(2ll

c(28t

c(281

c (28,

c(at)

c(a¡l

c(a¡t

c({21

c({lt

c(att

c(aal

c(aal

c(.5t

c(a6l

c{¿?t

c(2al

c(251

c(261

cl21l

c(261

c{2El

c(20t

N(2lt

c(2lt

c(2rl

c({2,

c(a5t

c(a6,

c(r3t

c(ial

ctall

c{{51

c(a5l

c(a6t

cta5l

120 I tìt

rr6 Stst

I2l a (€t

r22 0 (91

llì.0t9t

122.7 t1l

106.6 (71

r20-6 f 8t

125.I trt

tl0.0t?r

104.1{rl

122-t (?¡

t¡9.a (lt

t1,.2 f ?t

122. I {rt

tr?.0ttt

ll?-lt9l

rr9_rl8r

r2l_ 0 (9t

119.2 (ll

r20 2 (l)

Appendices l9l

Appendix I

Crystal structure of

(Z ) - N -phthaloyl-cr, p-meth anophenylalanine meth yl es ter ( 1 43 )

c(13)c(14)

c(t5)

c(16)

N(10)

c(3)

c(l)

C(12) o(u)

c(1

c(l8)CQI)

c(17)

o(30)

o(18) c(2)

o(31) c(22)c(30)

c(31)

Appendices 198

Bond distances (Å) for

( Z ) - N -phth aloyl-o, p- methanophenylalanine methyl ester ( I 43 )

c (11)c (30)c (30)c (11)c(21c (30)c (3)H (31)c (12)c (17)c (14)c (1s)c (16)c (1?)c (23)tl(221H (23)H (24l-H (2s)H (26)H (312)

o(11)o (30)o (31)N (10)c (1)c (1)c (21c (3)c (11)c (r2',c (13)c(14)c (ls)c (16)c (22'c(221c(23)c (241c (2s)c (26)c (31)

1.200(B)1.347 (10)1.186(9)1.39s(9)1.s40 (11)1.476 (10)r - 411 (rz',,r .o12 ('l9l1.491 (9)1.393 (10)1.394 (12)1.373 (13)L.362 (Lzl1.3?9 (10)1.39s(-)1 .203 (70)0.965 (72)0.879 (7 9)1.133 (72)1.111 (73)1.067 (82)

c (18)c (31)c (1)c(18)c (3)H(21c(2LlH (32)c(13)H (13)H(14)H (1s)H (16)c (18)c (21)c (241c (2s)c (26',,

c (21)H (311)H (313)

o (18)o (30)N (10)N (10)c (1)c (2')c (2'tc(3)c (12)c (13)c (14)c (1s)c (16)c (17)c(221c (23)c (241c (2s)c (26'c (31)c (31)

1.194 (B)1.460(14)1.431 (9)1.410 (9)1.506(11)0.936(79)1.483 (10)0. 942 (86)1. 382 (11 )

0.9s9(?6)0-865(7?)o. 89? (82 )

1.002 (75)1.503 (11 )

1.39s (-)1.3es (-)1.395 (-)1.39s (-)1.3es (-)0.?41 (91)0. 863 (82 )

Bond angles (deg.) for

( Z ) -N -phthaloy l-cr, p-meth anophenylalanine methyl es ter ( I 43 )

c (3r)c (r6.tc (2tc(3tc (3olH(2)c (3)c (21tc (21H (311H (321

N (r0lc (r2tc (r?)H (r3)c (1llc (rslH (rstc(16tc (r7)c(r5lc (r8)c (r7lc (2rtA t22lH (23'c (25'H (2{)H (2s)c (21)H (26)c(26)o(31)c(t)H (312)H (313)rì (rr3)

o (30,N (r0tc(11c(llc(r)c(21c(2)c(21c (3tc (31c (3¡c(rltc(rr)c(r2lc(13)c(131c(1{)c(151c (rslc (r6lc(r7lc(r7tc(r8)cl22lc(221c (231c(2{)c(2{}c (251c(261c (25)c(2r)c (30tc (30)c(31'c(31'c(3r)

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