Polyhedron Vol[ 06\ No[ 05\ pp[ 1628Ð1638\ 0887Þ 0887 Elsevier Science Ltd\ Pergamon All rights reserved[ Printed in Great Britain9166Ð4276:87 ,08[99¦9[99PII ] S9166Ð4276"87#99951ÐX

Kinetics and mechanism of the oxidation ofD!galactono!0\3!lactone by CrVI and CrV$

S[ Signorella\a M[ Santoro\a C[ Palopoli\a C[ Brondino\b J[ M[ Salas!Peregrin\c

M[ Quirozc and L[ F[ Salaa�

a Departamento de Qu(�mica\ Facultad de Ciencias Bioqu(�micas\ y Farmace�uticas\ UNR\ Suipacha 420\1999 Rosario\ Argentina\

b Instituto de Desarrolo Tecnolo�gico para la Industria Qu(�mica\ Gu�emes 2349\ 2999 Santa Fe\ Argentina\

c Departamento de Qu(�mica Inorga�nica\ Facultad de Ciencias\ Universidad de Granada\Fuentenueva s:n\ 07960 Granada\ Espan½a

"Received 10 January 0887 ^ accepted in revised form 15 January 0887#

Abstract*The oxidation of D!galactono!0\3!lactone by CrVI yields D!lyxonic acid\ carbon dioxide and Cr2¦ as_nal products when an excess of sugar acid over CrVI is used[ The redox reaction occurs through CrVI :CrIII and CrVI : CrV : CrIII paths[ The complete rate law for the CrVI oxidation reaction is expressed by−dðCrVIŁ:dt � "k9¦kH ðH¦Ł# ðgalŁðCrVIŁ\ where k9 � "2022#×09−3 M−0 s−0 and kH � "8824#×09−3 M−1

s−0\ at 39>C[ CrV is formed in a rapid step by reaction of the CO=−1 radical with CrVI[ CrV reacts with the

substrate faster than does CrVI[ The CrV oxidation follows the rate law ] −dðCrVŁ:dt � "k?9¦k?H ðH¦Ł# ðgalŁ\where k?9 � "0421#×09−2 M−0 s−0 and k?H � "2323#×09−2 M−1 s−0\ at 39>C[ The EPR spectra show thatseveral intermediate ðCr"O#"gala#1Ł− linkage isomers are formed in rapid pre!equilibria before the redox steps[Þ 0887 Elsevier Science Ltd[ All rights reserved

Keywords] galactonolactone ^ chromium ^ oxidation ^ kinetics ^ mechanism ^ CrV ^ CrVI[

———————————————————————————————————————————————

Hexavalent chromium\ in many di}erent compounds\is a well established carcinogen and mutagen ð0Ð4Ł[The observation of CrV and CrIV intermediates in theselective oxidation of organic substrates by CrVI andtheir implication in the mechanism of Cr!induced can!cers ð5Ð7Ł has generated a considerable amount ofinterest in their chemistry and biochemistry ð8Ð01Ł[The biological reduction of CrVI to lower states hasbeen observed with a wide variety of naturally occur!ring cellular reductants ð02Ð04Ł[ Ligands that possesstwo oxygen atoms able to form _ve!membered ringsabout the metal ion\ such as 0\1!diols and a!hyd!roxyacids\ are e}ective as non enzymatic reductantsand complexation agents towards hypervalent chro!mium and can stabilise the labile oxidation states ofchromium ð01\ 05Ð08Ł[ For this reason sugars\ or their

� Author to whom correspondence should be addressed[e!mail ] inquibirÝsatlink[com

$ The present work was presented at the X Meeting onPhysical!Chemistry\ Tucuma�n\ Argentina\ April 10\ 0886[

1628

derivatives\ may play an important role in the chem!istry of CrVI[ Aldonic acids "or their lactones# havebeen found as metabolic intermediates in animals andplants\ as products of the action of micro!organismson carbohydrates and as constituents of acidic poly!saccharides ð19Ł[ CrVI oxidation of a!hydroxyacids hasbeen extensively studied ð10Ð18Ł[ These oxidationsmay yield products coming from either C0C ð10Ð11\14Ð16\18Ł or C0H ð12Ð13\17\18Ł cleavage\ depend!ing on the particular molecular characteristics of theorganic substrate\ through mechanisms involvingeither two!electron or three!electron initial oxidationof substrate by CrVI[ Polyhydroxyacids o}eradditional binding sites to form the CrVI "CrV# esterprecursor of slow electron!transfer steps and providehelpful information about stereoelectronic e}ectsa}ecting the reaction course ð17Ł[ In previous workwe have found that the number and arrangement ofhydroxyl groups of aldoses a}ect the CrVI and CrV

oxidation rates and mechanism ð29Ð23Ł[ At present\we are interested in correlating the stereochemistry ofaldonic acids with their ability for reducing and co!

S[ Signorella et al[1639

ordinating CrVI and CrV\ as well as determining thepreferred donor sites of these polyhydroxyacids[ Inthis paper we report our results on the chromic oxi!dation of D!galactono!0\3!lactone "Scheme 0# and wepresent structural evidences for the intermediate CrV

complexes formed in acid and neutral aqueous solu!tions[

EXPERIMENTAL

Materials

D!Galactono!0\3!lactone "Sigma grade#\ D!gluconicacid "Sigma grade#\ potassium dichromate "BDH#\sodium perchlorate "Fluka grade#\ acrylonitrile "Ald!rich grade# and perchloric acid "P[ A[ Merck# wereused without further puri_cation[ Water was puri_edby deionisation\ followed by double distillation froma potassium permanganate solution[ NaðC!r"O#"ehba#1Ł = H1O was synthesised from 1!ethyl!1!hydroxybutanoic acid "Aldrich grade# and sodiumdichromate "Merck# in acetone "Merck\ A[ R[ grade#according to the literature method ð24Ł[

The stability of the organic substrate under con!ditions used in the kinetic studies was tested[ The acid"gala# ] lactone "galL# proportion of the reductant inthe starting solution was calculated by optical rotationmeasurements at given hydrogen ion concentrationsð25Ł and the composition of the substrate solution atdi}erent perchloric acid concentrations is shown inTable 0[ The equilibrated solutions are stable over thetime required for the kinetic measurements[ In the textthat follow gal refers to the total reductant con!centration "ðgalŁ � ðgalLŁ¦ðgalaŁ#[

For experiments performed in the 1Ð7 pH range\the pH of the solutions was adjusted by addition of9[4 M NaOH and 9[4 M HClO3[

Spectrophotometric measurements

Most kinetics measurements were made at 249 nm\by monitoring the absorbance changes on a GuilfordResponse II spectrophotometer with fully ther!mostated cell compartments[ Mixtures of sodium per!

Scheme 0[

Table 0[ Proportion of D!galactonolactone "galL# and D!galactonic acid "gala# in the starting solutiona

ðHClO3Ł:M 091 CTb:M Rc galL ")# gala ")#

9[14 5[59 −9[659 67[8 10[09[27 4[99 −9[428 61[1 16[79[49 4[34 −9[473 60[5 17[39[52 4[03 −9[467 65[3 12[59[64 4[95 −9[462 66[0 11[89[89 5[18 −9[559 71[9 07[90[99 4[47 −9[571 74[2 03[6

a Values used in calculations ] molar rotations ] −02[73"galL# ^ −1[73 "gala#\ ¦9[794 "gal−# ^ K � ðH¦Łðgal−Ł"ðgalaŁ¦ðgalLŁ# � 1[14×09−4[

b CT � ðgalaŁ¦ðgalLŁ¦ðgal−Ł\ where gal− refers to gal!actonate and represents ³9[91) of CT[

c Optical rotation[

chlorate and perchloric acid were used to maintain aconstant ionic strength "I# of 0[9 M[ Reactant solu!tions were previously thermostated and transferredinto a cell of 0 cm path length immediately after mix!ing and disappearance of CrVI was followed until atleast 79) conversion[ Experiments were performedat 39>C unless otherwise mentioned[ In most exper!iments the concentration of CrVI was kept constant at2[5×09−3 M while the gal concentration was variedfrom 9[907 to 9[011 M[ The _rst!order dependence ofthe rate upon ðCrVIŁ was veri_ed[ The pseudo!_rst!order rate constants were calculated at various ðCrVIŁ9\but at constant temperature\ ðgalŁ9\ ðH¦Ł and I[ Asexpected on the basis of a −d"lnðCrVIŁ#dt � kobs ratelaw where kobs � f"ðgalŁðH¦#\ kobs was found to beessentially constant with increasing ðCrVIŁ9[

CrV was monitored by following the absorbancegrowth and decay at 649 nm of mixtures of CrVI

"2[5×09−2 M# and excess gal "9[07Ð9[46 M# at _xedperchloric acid concentrations[ The formation of CrIII

was monitored at 469 nm "ðCrVIŁ9 � 0[7×09−1 M andratios of gal to CrVI from 09 ] 0 to 19 ] 0\ at ðHClO3Ł inthe 9[14Ð9[64 M range#[ At the end of the reactionthe two dÐd bands ascribed to CrIII were observed at

The oxidation of D!galactono!0\3!lactone by CrVI and CrV 1630

lmax � 309 "o � 22# and 455 nm "o � 16[4 dm2 mol−0

cm−0# "ðHClO3Ł � 9[14 M#[ Both\ visible and UVspectral maxima and intensities\ are in close agreement"vide infra# with those observed for the acid solutionsof the KðCr"OH#"gala#"lyxa#Ł "lyxa � D!lyxonic acid#ternary complex\ independently synthesised ð26Łaccording to a literature method ð27Ł[ These dÐd bandschanged with time toward bands of lower intensity atlmax � 397 "o � 08[6# and 462 nm "o � 04 dm2 mol−0

cm−0#\ characteristic of the ðCr"H1O#5Ł2¦ ion[

Products analysis

Under the conditions used in the kinetic measure!ments "ratios of gal to CrVI from 49 ] 0 to 239 ] 0#\qualitative identi_cation of D!lyxonic acid as the reac!tion product was carried out by paper chro!matography[ D!lyxonic acid was identi_ed against anauthentic sample using n!butanol:acetic acid:water"3 ] 0 ] 4# as eluent[ Paper chromatograms were visu!alised by two kinds of development reagents ] a three!stage dip of silver nitrate\ sodium hydroxide andsodium thiosulphate ð28Ł and p!anisidine reagent ð39Ł[D!lyxonic acid identi_cation as the only reaction prod!uct was also made by 02C NMR[ A mixture of gal"6[53×09−2 mmol# and potassium dichromate"0[55×09−2 mmol# in 0 M HClO3 "09 cm2# wasallowed to react at 39>C[ After the reaction had pro!ceeded to completion the mixture was stirred with aDowex 49W!X7 cation exchange resin until all theCrIII was removed from the solution and a con!centrated sample was analysed by 02C NMR[ Lyxonicacid and unreacted galL and gala were identi_ed bycomparing against authentic samples[

Under all these experimental conditions "large tomoderate excess of organic substrate# a quantitativeyield of carbon dioxide was observed[

The presence of free radicals was tested for in thereaction of gal with CrVI[ A solution of potassiumdichromate "3×09−3 mmol# in 1 cm2 of 9[4 M HClO3

was added to a mixture of 3×09−1 mmol gal andacrylonitrile "9[14 cm2# in 9[4 M HClO3 "9[14 cm2# at39>C[ After 0 h a white precipitate appeared[ Controlexperiments "without potassium dichromate orreductant present# did not show the formation of aprecipitate[ Besides\ the addition of an excess of acry!lonitrile on the reaction mixture results in the completesuppression of the carbon dioxide evolution[

EPR spectroscopy

The EPR spectra were obtained on a Bruker ESP299 spectrometer[ The microwave frequency was gen!erated with a Bruker 93 ER "8Ð09 GHz# and measuredwith a Racal!Dana frequency meter[ The magnetic_eld was measured with a Bruker NMR!probegaussmeter[ A least!squares EPR spectrum!_ttingprogram\ e12new\ was used to estimate the spectral

parameters and relative concentrations of the CrV

complexes\ using Lorentzian line shapes ð30\31Ł[

RESULTS AND DISCUSSION

Rate studies

The absorbance vs time curves at 249 nm exhibit amonotonic decrease of absorbance which cannot bedescribed by a single exponential decay[ At this wave!length\ kinetic traces show an initial deviation from_rst!order decay over short time periods[ These kineticpro_les could be adequately described by the set ofconsecutive _rst!order reactions of Scheme 1 ]

It is known that CrV species absorb strongly at 249nm and may superimpose CrVI absorbance yieldingthe wrong interpretation of spectrophotometricabsorbance decay values ð18Ł[ Thus\ considering theCrV absorption superimposition\ the absorbance at249 nm\ at any time during the reaction\ is given by

Abs249 � oVI ðCrVIŁ¦oVðCrVŁ "0#

Combining the eqn "0# with rate expressions ð32Łderived from scheme 1\ yields ]

Abs249 � Abs9e−k5t¦k5oVðCrVIŁ9"e−k4t−e−1k5t#:

"1k5−k4# "1#

In this equation\ oV refers to the molar absorptivity ofCrV at 249 nm and was assumed to be the same as forthe complex ðCrV"O#"ehba#1Ł− ð24Ł[ Parameters k5 andk4 refer to the rate of disappearance of CrVI and CrV\respectively\ and were evaluated from a non!lineariterative computer _t of eqn "1#[ It must be noted thatin eqn "1#\ k5 appears in the numerator of the pre!exponential term and 1k5 appears in the denominatorand in the exponential terms because\ according tothe proposed reaction scheme\ only half of the CrVI

reaches CrIII through a CrV intermediate[Figure 0"a# shows a typical curve for one run at

249 nm and the curve _t according to eqn "1#[ Thecalculated kinetic parameters\ k5 and k4\ for variousconcentrations of gal at _xed concentrations ofHClO3\ are summarised in Table 1[ In multiplemeasurements\ the reproducibility of the two rate con!stants is better than 09)[

Values of k4 and k5 may also be obtained fromabsorbance data at 649 nm[ At this wave length\ onlyCrV absorbs and the experimentally observed inter!mediate CrV growth and decay curve could be _ttedby eqn "2#

Abs649 � k5 ðCrVIŁ9oV"e−k4t−e−1k5t#:"1k5−k4# "2#

where oV refers here to CrV molar absorptivity at 649nm[ A good _t of data is obtained with oV � 27 M−0

cm−0 ð15Ł\ as shown in Fig[ 0"b#[ Since at least tenfoldhigher initial CrVI and gal concentrations were necess!ary to follow absorbance changes at 649 nm\ k4:09and 1k5:09 values calculated by eqn "2# are coincident

S[ Signorella et al[1631

Scheme 1[ S � organic substrate ^ I> � intermediate radical

Fig[ 0[ Absorbance vs time curves for the chromic oxidation of gal\ I � 0M\ T � 39>C at "a# 249 nm\ ðCrVIŁ � 2[5×09−3 M\ðgalŁ � 2[5×09−1 M\ ðH¦Ł � 9[27 M ^ "b# 649 nm\ ðCrVIŁ � 2[5×09−1 M\ ðgalŁ � 2[5×09−0 M\ ðH¦Ł � 9[27 M ^ "c# 469 nm\

ðCrVIŁ � 0[7×09−1 M\ ðgalŁ � 0[7×09−0 M\ ðH¦Ł � 9[64 M[

Table 1[ Observed pseudo!_rst!order rate constants "k5 and k4# for di}erent concentrations of HClO3 and organic substrate[T � 39>C ^ ðCrVIŁ9 � 2[5×09−3 M ^ I � 0 M

1k5 "s−0# and k4 "s−0# for ðHClO3Ł:M ofðgalŁ:M 9[14 9[27 9[49 9[64 0[99

0931k5 093k4 0931k5 093k4 0931k5 093k4 0931k5 093k4 0931k5 093k4

9[907 9[89 3[99 0[28 4[29 0[50 5[0 0[87 6[19 1[30 7[809[925 0[77 5[18 1[67 09[6 2[01 01[5 3[09 03[3 3[69 06[49[949 1[49 09[4 2[71 04[1 * 06[9 4[07 19[5 5[37 13[79[946 * * 3[28 06[5 3[50 07[8 * * * *9[961 2[69 04[5 4[99 19[6 5[11 13[0 7[90 18[5 8[31 22[89[975 * * * * * * 8[29 22[6 * *9[982 * * * * * * * * 01[1 34[79[097 4[28 10[9 6[60 21[0 7[60 25[7 09[8 31[0 * *9[011 5[50 15[0 7[67 25[1 8[88 30[9 01[5 35[5 04[5 47[1

The oxidation of D!galactono!0\3!lactone by CrVI and CrV 1632

with those obtained from eqn "1#\ when the samegal:CrVI ratio and ðHClO3Ł were employed[ For galand CrVI concentrations used in the kinetic measure!ments\ either at 249 or 649 nm\ the calculatedmaximum CrV concentrations "ðCrVŁmax# represent upto 7) of the total chromium concentration[

The CrIII formation was followed at 469 nm[ At thiswave length\ changes in CrIII concentration can bedescribed by the pair of consecutive reactions shownin the Scheme 2 and the experimental absorbance canbe _tted by eqn "3#

Abs469 � Abs"ðCrIIIŁ#¦Abs"Cr2¦#

� ðCrVIŁ9k0"e−k0t−e−k2t#"oIII0 −oIII

1 #:

"k2−k0#¦Abs�"0−e−k0t# "3#

where k0 and k2 refer to the rates of CrVI and ðCrIIIŁdisappearance\ and oIII

0 \ oIII1 are the molar absorptivities

of the CrIII complex formed at the end of the redoxreaction and ðCr"H1O#5Ł2¦\ respectively[ A typical _tof experimental data at 469 nm by using eqn "3# isshown in Fig[ 0"c#[ Here\ we have considered that therate of CrIII complex formation equals the rate ofCrVI decay\ without taking into account CrV[ This isconsistent with the observation of an isosbestic pointat 414 nm under conditions employed to follow CrIII

at 469 nm "Fig[ 1# and the low ðCrVŁmax "up to 7)#observed at 649 nm "Fig[ 0#\ for the same oxidant andreductant concentration range[ Values of k0 calculatedby eqn "3# are coincident with 1k5 values obtained byusing either eqn "1# or "2#[ On the other hand\ k2

depends strongly on ðH¦Ł and a more detailed study isrequired for the complete description of the hydrolysismechanism[

CrVI oxidation of `al

In the proton concentration range studied\ plots of1k5 vs ðgalŁ gave good straight lines from which valuesof k were determined "Table 2#[ A plot of k vs ðH¦Łgave a straight line with a positive intercept\ showingthat the second!order rate constant may be expressedas consisting of an acidity independent and an aciditydependent term ]

k � k9¦kHðH¦Ł "4#

where k9 � "2022#×09−3 M−0 s−0 andkH � "8824#×09−3 M−1 s−0

The complete rate law is then given by

Scheme 2[ ðCrIIIŁ � CrIII!gala!lyxa ternary complex

−d ðCrVIŁ:dt � 1k5 ðCrVIŁT

� "k9¦kHðH¦Ł#ðgalŁðCrVIŁT "5#

where ðCrVIŁT represents the total CrVI concentration[The two terms in the rate law "eqn "5## results from

the fact that there are at least two transition states\di}ering in composition but similar in energy\ throughwhich the reduction of CrVI can proceed[ Thus\ theoxidation of gal by CrVI should occur through twoparallel slow steps leading to the redox products[Besides\ in the range of substrate concentrationsemployed\ only substrate _rst!order dependence wasobserved and D!lyxonic acid and carbon dioxide werethe only detected reaction products\ clearly showingthat the C0C scission is preferred to the C0H cleav!age at the a!carbon[ A mechanism taking account ofthese facts is proposed in Scheme 3[

In this mechanism we have assumed that the opencarboxylic acid is the reactive form of the organicsubstrate[ This assumption is based on previous _n!dings showing the enhanced ability of hydroxyacidsto bind CrVI and CrV ð02\14Ð17Ł[ In any case\ a possiblemechanism involving the lactone may not bediscarded[ However\ if the lactone was the reactiveform of the organic reductant\ co!ordination of CrVI

should be favoured at a pair of adjacent hydroxylgroups of the molecule ð33Ł[ The intermediate complexformed in this way might be present in the equilibriummixture but should not be a direct precursor of theslow redox steps[ Opening of the lactone ring and re!arrangement should be required for the slow electrontransfer steps to take place within a complex with thecarboxylate bound to CrVI[ On the other hand\ in theðH¦Ł range studied\ CrVI exists mainly as Cr1O�

6

ð34\35Ł[ Thus\ the _rst step of the proposed mechanismmay be interpreted as the formation of a monochelatewith gala acting as a bidentate ligand bound to CrVI

at Cl0O and C10O to yield the anionic species A1−[This intermediate\ or its monoprotonated form"AH−#\ may decompose to the products[ The rate lawcorresponding to this mechanism may be expressed asin eqn "6# ]

−dðCrVIŁ:dt � "k0K0¦k1K0K1 ðH¦Ł#ðgalaŁðCrVIŁ

"6#

Equation "6# may be expressed as a function of thetotal organic substrate concentration as in eqn "7# ]

−dðCrVIŁ:dt � "k0K0¦k1K0K1 ðH¦Ł#ðgalŁKh ðCrVIŁ:

"0¦Kh# "7#

We propose that redox paths\ involving either A1− orAH− intermediate complexes\ are three electron steps\just as observed for the chromic oxidation of mandelicð14Ł\ 1!hydroxy!2!methylbutanoic ð10Ł and D!ribonicð11Ł acids\ and CrIII is formed with the oxidation prod!ucts[ The alternative path corresponding to thesequence CrVI:CrIV:CrIII was discarded on the basis ofthe complete CO1 suppression after addition of excess

S[ Signorella et al[1633

Fig[ 1[ Time evolution of the UVÐvis spectrum of a mixture of gal "9[64 M# and CrVI "0[7×09−1 M#[ ðHClO3Ł � 9[4 M ^T � 14>C\ over a period of ca 34 min[

Table 2[ Values of k and k? for di}erent ðHClO3Ł

ðHClO3:M 9[14 9[27 9[4 9[64 0[9

092 k:M−0 s−0 4[19 6[13 7[11 09[4 01[8092 k?:M−0 s−0 19[5 18[6 22[7 28[1 37[1

acrylonitrile on the reaction mixture and the fact thatthe 1!ketoacid was not observed over the whole set ofconditions used here[ Besides\ CrII formation was notproposed since CrO1¦

1 \ which is taken as evidence thatCrII is an intermediate product of the reaction\ wasnot observed as we did for the oxidation of homo!serine by CrVI ð13Ł[ CO=−

1 radicals formed in the slowsteps may react with CrVI in a subsequent fast step toyield CrV and carbon dioxide[ Polymerisation afteraddition of acrylonitrile support the formation ofthese radicals[ CrV can further oxidise gala as will nowbe discussed[

CrV oxidation of `al

At the ðHClO3Ł used in the kinetics measurements\the reaction of CrVI with excess gal generates an EPR

spectrum dominated by a single detectable signal at`iso � 0[8670 "Fig[ 2 Inset# with the four weak 42Cr"8[44) abundance\ I � 2:1# hyper_ne peaks at 07 Gspacing\ typical of _ve!co!ordinate oxochromate"V#complexes ð36\37Ł[

The rate of formation and disappearance of theintermediate CrV calculated from the peak!to!peakheights "and:or areas# of the EPR signals as a functionof time "Fig[ 2# are consistent with values obtainedfrom the spectrophotometric measurements[ Thedelayed tmax observed by this technique\ account forthe lower temperature "14>C\ instead of 39>C# used inthe EPR measurements[

In the 9[14Ð0 M proton concentration range\ plotsof k4 vs ðgalŁ gave good straight lines from whichvalues of k? were determined "Table 2#[ A plot of k?vs ðH¦Ł gave a straight line with a positive intercept\showing that the second!order rate constant may beexpressed as consisting of an acidity independent andan acidity dependent term[ The rate constant for thedisappearance of CrV then may be expressed as ]

k4 � "k?9¦k?HðH¦Ł#ðgalŁ "8#

where k?9 � "0421#×09−2M−0s−0

and k?H � "2323#×09−2M−1s−0 "09#

The oxidation of D!galactono!0\3!lactone by CrVI and CrV 1634

Scheme 3[ A1− � ð"galaH−1#"O#1"OH1#CrOCrO2Ł1−

Fig[ 2[ Peak!to!peak heights of CrV EPR signals vs time[ ðgalŁ � 9[033 M ^ ðCrVIŁ � 2[5×09−2 M ^ ðH¦Ł � 9[4 M ^ I � 0 M ^T � 14>C[ Tmax � 497 s[ Inset ] EPR signal recorded at t � 04 min[

This means that the CrV intermediate decays to theproduct directly or through an acid!catalysed step[Since the latter is of the same order as the non!cat!alysed step\ at low acid concentration "i[e[ neutral pH#the oxidation occurs mainly through the non!acid!

catalysed path[ In this way\ the intermediate CrV isconsumed faster than observed for the reaction of CrVI

with sugars for which the non!acid catalysed path isextremely slow and the CrV intermediates remain insolution after several weeks ð29Ð21Ł[

S[ Signorella et al[1635

A mechanism taking into account the structuralevidence for the CrV intermediate complex and thekinetics results is given in Scheme 4[

In this mechanism\ only one of the two gala mol!ecules involved in the CrV complex formation is oxi!dised to D!lyxonic acid through the two!electronintramolecular redox path\ in accordance with theobserved _rst!order rate on ðgalaŁ[ Possibly\ severallinkage isomers might be formed by co!ordinationof gala with CrVI via any pair of properly disposedhydroxyl group "see discussion below#[ However\since D!lyxonic acid is the only reaction productobserved in the solution\ the CrV complex\ precursorof the redox steps\ must be one with at least onegala molecule bound to CrV at the carboxylate group[Besides\ the `iso and Aiso values observed for the CrV

intermediate are close to those of ðCr"O#"ehba#1Ł−[Since the `iso and\ specially\ Aiso values are very sen!sitive to the CrV co!ordination environment\ ourassumption seems to be a reasonable one[

It must be noted that\ in Scheme 4\ the two!electronsubstrate C0C cleavage a}ords formic acid[However\ under our experimental conditions\ we havenever detected formic acid[ This fact may be explainedif\ under conditions used here\ formic acid is oxidisedfaster than gal[ E}ectively\ CrVI oxidation of formicacid to carbon dioxide is faster than gal oxidation ð38Ł\a}ording the observed quantitative yield of carbondioxide[

The proton superhyper_ne "shf# coupling has beenshown to be useful in determining the binding of 0\1!diolate moieties of sugars to the CrV centre ð29\33Ł[We found that performing the reaction at pH × 1\the single broad "linewidth ½ 1 G# signal observed athigher ðH¦Ł sharpens and the shf pattern resolves\thus providing new insights about the gala co!ordi!nation sites[

The reaction of CrVI with an excess of 4Ð19 timesof gal\ in the 1Ð5 pH range\ at 14>C\ results in theformation of a CrV EPR spectrum which is a com!

Scheme 4[ R � CH1OH"CHOH#2

posite of at least two EPR signals at `0 � 0[8670 and`1 � 0[8677[ The relative proportion of the signalsdepends on the pH\ but is independent of the gal:CrVI

ratio employed and the reaction time "the signals weredeconvoluted by _tting the spectra by Lorentzianderivatives#[ Figure 3"a# shows the spectrum of a reac!tion mixture at pH � 4 with the simulated spectrum\for which a `0:`1 ratio � 9[30 was obtained[ The best_t of the spectrum yields the spectral parameters sum!marised in Table 3[ The `iso values obtained for thetwo signals are those expected for _ve!co!ordinateoxochromate"V# complexes ^ while the di}erent shfpatterns indicate that they are linkage isomers ð49Ł[ Inthis way\ the shf splitting of the signal at `0 arises fromthe coupling of two equivalent protons and may beattributed to a complex with two ligand moleculesbound to CrV at Cl0O:C10O[ The shf splitting ofthe signal at `1 may be assumed to arise from twodi}erent binding modes of the two ligand moleculesco!ordinated to CrV\ via a 0\1!diolate and the a!hyd!roxocarboxylate donor sites\ respectively[ In the oxi!dation of D!gluconic acid "glca# with CrVI\ in this pHrange\ only one species is detected[ Interestingly\ the`iso value "0[867# and the shf pattern "Fig[ 4# revealthe same chelation mode of glca as for gala in thecomplex giving rise to the signal at `1[ For the samegal:CrVI ratios\ in the 5Ð7 pH range\ the spectra aredominated by a signal at 0[8684 with a shf splittingvalue of 9[81 G[ The `iso and shf pattern clearly indi!cate that CrO2¦ co!ordinates two gala molecules viaa 0\1!diolate moiety and that the shf structure derivesfrom the coupling with the four equivalent protons[In this pH range\ the minor signal "`iso � 0[867# canbe attributed to the isomer with the ligand actingthrough a 0\1!diolate and the a!hydroxocarboxylatedonor sites[ Figure 3"b# shows the spectrum of a reac!tion mixture at pH � 6[3 with the simulated spectrum\for which the major CrV species represents the 81)of the CrV in solution[

A last point to consider is that\ at each pH\ the

The oxidation of D!galactono!0\3!lactone by CrVI and CrV 1636

Fig[ 3[ X!band EPR signal of a mixture of gal and CrVI "4 ] 0 mole ratio# at "a# pH � 4[3\ 1 h after mixing\ "b# pH � 6[3\ 0day after mixing\ and the corresponding simulated spectra "linewidth � 9[7 and 9[5 G\ respectively# ^ T � 14>C\ modulation

amplitude � 9[52 G\ centre _eld � 2444 G ^ frequency � 8[73 GHz[

Table 3[ EPR spectral parameters

`0iso "m# a0iso:G `1iso "m# a1iso:G

ðH¦Ł × 9[0 M 0[8670 *pH � 1Ð5 0[8670 "t# 9[64 0[8677 "dt# 9[64\ 9[81pH � 5Ð7 0[8684 "quint# 9[81 0[867 "dt# 9[6\ 9[8

m � multiplicity ^ t � triplet ^ dt � double!triplet ^ quint � quintet

S[ Signorella et al[1637

Fig[ 4[ X!band EPR signal of a mixture of D!gluconic acid and CrVI "09 ] 0 mole ratio# at pH � 3\ 0 day after mixing and thecorresponding simulated spectrum "linewidth � 9[44# ^ T � 14>C ^ modulation amplitude � 9[404 G ^ centre _eld � 2429 G ^

frequency � 8[66 GHz[

relative proportion of the intermediate CrV complexesremains constant over the reaction time\ meaning thatthese CrV intermediates are in rapid equilibria com!pared to the time scale of their subsequent reductionto CrIII[

Acknowled`ements*We thank the National Research Coun!cil of Argentina "CONICET#\ The Third World Academyof Sciences "TWAS#\ the National University of Rosario\Antorchas Foundation and The International Foundationfor Sciences "IFS# for _nancial support[ We thank Prof[Brumby and Prof[ Lay for providing us with the e12newProgram[

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