Marine natural products of fungal origin

REVIEW www.rsc.org/npr | Natural Product Reports

Marine natural products of fungal origin

Muhammad Saleem,a Muhammad Shaiq Ali,*b Shafqat Hussain,c Abdul Jabbar,c Muhammad Ashrafd andYong Sup Leee

Received (in Cambridge, UK) 16th November 2006First published as an Advance Article on the web 22nd March 2007DOI: 10.1039/b607254m

Covering: 2000 to 2005

In the search for novel and bioactive molecules for drug discovery, marine-derived natural resources arebecoming an important research area. Over 15 marine-derived secondary metabolites are currently inhuman clinical trials. Terrestrial fungi have produced many therapeutically significant molecules.However, the potential of marine fungi has only been investigated to a limited extent. This review articlecontains 103 marine-derived fungal metabolites and 77 references.

1 Introduction2 Polyketides and their isoprene hybrids3 Terpenoids4 Nitrogenous compounds (alkaloids and peptides)4.1 Alkaloids4.2 Peptides5 Cerebroside analogues6 Miscellaneous metabolites7 Conclusions8 Acknowledgements9 References

1 Introduction

Almost three quarters of the earth’s surface is occupied by seas andoceans. Their resources are varied and vast and partly compriseof fish, shellfish, other animals, vegetation, algae, bacteria andfungi. These natural resources have only recently been exploredfor natural products; many of them are biologically active andpotentially useful. Since marine organisms live in a significantlydifferent environment from those of terrestrial organisms, it isreasonable to suppose that their secondary metabolites will differconsiderably.1–5

In recent years, research on the chemistry of marine organismshas experienced a tremendous increase due to the need forcompounds possessing bioactivity with possible pharmaceuticalapplications or other economically useful properties such asfine chemicals, drugs, cosmetics and functional personal-careproducts.6 To this end, a variety of species have been assayed fortheir activity and a number of bioactive molecules, often with toxic

aPharmaceutical Research Center, PCSIR Labs. Complex, , 75280, Karachi,PakistanbH.E.J. Research Institute of Chemistry, International Center for Chemicaland Biological Sciences (ICCBS), University of Karachi, 75270, Karachi,PakistancDepartment of Chemistry, Faculty of Science, The Islamia University ofBahawalpur, PakistandDepartment of Pharmacy, Faculty of Pharmacy and Alternative Medicine,The Islamia University of Bahawalpur, PakistaneDepartment of Pharmaceutical Science, College of Pharmacy, Kyung HeeUniversity, 1 Hoegi-DongDongdaemoon-ku, 130-701, Seoul, Korea

properties and unique structural features, have been isolated.1,7

The rapid growth in the chemistry of marine organisms over thelast five decades has led to the discovery of a surprisingly largenumber of novel structures. Over 15 compounds of marine originare currently in human clinical trials.

The first systematic study of fungal metabolites was initiatedsoon after World War I by Harold Raistrick, who in the courseof the following four decades made a seminal contribution tothe recognition of fungi as a major source of natural products.8

Terrestrial fungi have provided many natural products withsignificant therapeutic value.9–13 Keeping this in mind, the potentialof marine fungi has only been investigated to a limited extent.Mycochemistry of marine fungi blossomed in the 1990s andnotably since 1998 and about 272 compounds have been de-scribed so far.14 These includes polyketides, terpenes, steroids andpeptides.15 Terrestrial sources including fungi have been intensivelyinvestigated for their secondary metabolites. Marine fungi livingin diverse environments containing high salt levels typical of themarine situation in association with invertebrates are very sensitiveto culture media. Therefore, bioactive metabolites may be obtainedform marine fungi and thus the hidden wealth of this source shouldbe explored.

This review article includes 103 marine fungal metabolites withtheir structures published during 2000–2005. These metabolitesare classified according to their classes into four sections. Alto-gether 77 references are cited in this review article.

2 Polyketides and their isoprene hybrids

Polyketides comprise of a large family of structurally diverse andmedicinally important natural products.16 The biosynthesis of thisgroup of secondary metabolites is catalysed by a superfamily ofenzymes called polyketide synthases (PKSs I, II and III). Thesetherapeutically important agents are biologically synthesized bysuccessive Claisen condensations of extender units derived from(methyl) malonyl-CoA with an acyl-CoA starter unit in a mannerreminiscent of fatty acid synthesis. These compounds possessbroad ranges of biological activities. Examples of such polyke-tides include erythromycin (antibacterial), nystatin (antifungal),avermectin (antiparasitic), rapamycin (immunosuppressant) and

1142 | Nat. Prod. Rep., 2007, 24, 1142–1152 This journal is © The Royal Society of Chemistry 2007

Dr Saleem did his B.Sc. and M.Sc. at Islamia University Bahawalpur, Pakistan in the years 1994 and 1996, respectively. He finished hisPh.D. in 2001 at HEJ Research Institute of Chemistry, International Center for Chemical Sciences, University of Karachi, Pakistan. Duringhis Ph.D. he worked on secondary metabolites of higher plants and marine algae under the supervision of Dr M. Shaiq Ali. Afterwards,Dr Saleem joined Pakistan Council of Scientific and Industrial Research Laboratories Karachi and worked on applied pharmaceutical andcosmeceutical aspects of natural products. Dr Saleem was awarded a KOSEF postdoctoral fellowship in 2003 and he worked at KoreaInstitute of Science and Technology (KIST), Seoul, South Korea with Professor Yong Sup Lee. In 2004 he was appointed as visiting scientistat the same institute and worked on the development of new drug candidates including anti-cancers, neuroprotective and anti-viral agents fromnatural products. In 2005, Dr Saleem was awarded an Alexander Von Humboldt Foundation fellowship and he joined Institute of Organicand Biomolecular Chemistry, University of Goettingen, Germany in 2006 where he is presently involved in searching for new drug candidatesfrom microbial sources under the guidance of Prof. H. Laatsch. His research interests are the development of new drug candidates includingnovel antibiotics, anti-cancers, neuroprotective agents, or anti-virals from natural products.

Muhammad Shaiq Ali did his masters in organic chemistry at the University of Karachi (Pakistan). He obtained his M.Phil. degree inmicrobiological transformation of organic molecules from the University of Sussex (UK) under the supervision of Prof. Dr J. R. Hansonand doctoral degree in marine chemistry (1992) under the supervision of Prof. Dr Viqar Uddin Ahmad of the H. E. J. Research Instituteof Chemistry, International Center for Chemical and Biological Sciences (ICCBS) at the University of Karachi. In 1994, he was appointedas an Assistant Professor at ICCBS, and presently he is serving as Associate Professor in the same institution. His research interestsare marine/terrestrial natural product chemistry and the microbiological transformation of organic molecules. He has over 150 researchpublications and a book on “Abbreviations in Organic Chemistry” to his credit. He is the winner of a young scientist TWAS Prize under 40(1997). He was STA fellow (Japan, 1995), UNESCO fellow (Japan, 1994, Tokyo Inst. of Tech.) and DAAD fellow (Germany, 1993).

Mr Shafqat did his B.Sc. and M.Sc. at Islamia University Bahawalpur (IUB), Pakistan in the years 1996 and 1998, respectively and then hejoined Sadiq Public School Bahawalpur (Pakistan) as Lecturer in Chemistry. He obtained his M.Phil. degree in 2005 from Islamia UniversityBahawalpur (Pakistan) while working on isolation and characterization of natural products from terrestrial plants. He is currently workingon isolation and structural elucidation of secondary metabolites of medicinal plants, under the supervision of Dr Abdul Jabbar (IUB) and cosupervision of Dr Abdul Malik of HEJ Research Institute of Chemistry, International Center for Chemical Sciences, University of Karachi,Pakistan for his doctoral degree.

Dr Abdul Jabbar did his B.Sc. (Hons) and M.Sc. at Punjab University Lahore, Pakistan in the years 1974 and 1976, respectively. In 1987,he finished his Ph.D. studies at the University of Aberdeen, Aberdeen, Scotland. Under the supervision of J. R. Lewis at the University ofAberdeen, his work was mainly focused on natural coumarins, alkaloids and flavonoids. In 1978, Dr A. Jabbar joined Islamia UniversityBahawalpur, Pakistan as a lecturer, where he is working now as a Professor in the Chemistry Department. His field of interest is phytochemicalinvestigation of medicinal plants of Cholistan Desert (Pakistan) and syntheses of acridones of medicinal importance.

Dr Ashraf did his M.Sc. (Chemistry) at The Islamia University of Bahawalpur (I.U.B.), Bahawalpur, Pakistan in 1984. He earned hisPh.D. in biochemistry in 1990 under the supervision of Dr C. M. Bray from Manchester University, Manchester (UK) and then carried out3 years of SERC funded post-doctoral research on the flux control analysis of primary nitrogen assimilation in maize mesophyll chloroplastsunder the leadership of Prof. Dr Mike Emes from the same university. Dr Ashraf joined the Chemistry Department as Assistant Professorback in Pakistan in I.U.B. in 1994 and joined the Pharmacy Department in 2000 as Associate Professor of Pharmaceutical Chemistry. Heis engaged in the phytochemistry and ethnopharmacology of desert plants, determination of biological activities of synthetic and naturalcompounds, in vitro and in vivo antioxidant and biological activities of homoeopathic medicines and herbal formulations and developmentof new enzyme based biological assays.

Yong Sup Lee received his B.En. degree in chemical technology from Seoul National University in 1983. He got his Ph.D. degree in organicchemistry from Korea Advanced Institute of Science and Technology (KAIST) under the supervision of Dr Hokoon Park and ProfessorSang Chul Shim. He worked on medicinal chemistry and natural products chemistry at Korea Institute of Science and Technology (KIST)as a principal research scientist from 1985 to 2004. In 2004, he was appointed as an Associate Professor at the College of Pharmacy, KyungHee University, Korea. His research interests are development of new drug candidates including anti-cancers, neuroprotective agents, oranti-virals from natural products or by organic syntheses.

Muhammad Saleem Muhammad Shaiq Ali Shafqat Hussain Abdul Jabbar Muhammad Ashraf Yong Sup Lee

This journal is © The Royal Society of Chemistry 2007 Nat. Prod. Rep., 2007, 24, 1142–1152 | 1143

daunorubicin (antitumor).17 Like the Gram-positive bacteria ofthe genus Streptomyces, marine fungi are also a potential producerof polyketides.

The fungus Epicoccum sp. isolated from the marine brown algaFucus vesiculosus yielded isobenzofuranone (epicoccone, 1).18 Thestructure of 1 was determined by interpretation of its spectroscopicdata and found to be potently active, showing 95% 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging effects at 25 lg mL−1.The same compound also inhibits the peroxidation of linolenicacid in the TBARS assay (62% inhibition at 37 lg mL−1). Amarine fungal isolate, identified as Acremonium sp., yielded twonovel hydroquinone derivatives: 7-isopropenylbicyclo[4.2.0]octa-1,3,5-triene-2,5-diol (2) and 7-isopropenylbicyclo[4.2.0]octa-1,3,5-triene-2,5-diol-5-b-D-glucopyranoside (3).19 Metabolite 2 and itsglucoside (3) possess a most unusual ring system. These com-pounds were characterized by interpretation of their detailed spec-troscopic data. Each isolate was tested for its antioxidant (TBARDassay) property and found to be significantly potent (85.5% at25.0 lg mL−1). Liu et al. purified two benzoquinones: 2-(2′,3′-dihydrosorbyl)-3,6-dimethyl-5-hydroxy-1,4-benzoquinone (4) and3-acetonyl-2,6-dimethyl-5-hydroxy-1,4-benzoquinone (5) from amarine fungus Penicillium terrestre. Their structures were estab-lished on the basis of spectroscopic methods and their cytotoxiceffects on P388 and A-549 cell lines were preliminarily examinedby the MTT method.20

Namikoshi et al. obtained three chlorine containing antibioticsfrom a marine fungus Aspergillus ostianus (strain TUF 01F313)isolated from a marine sponge of Pohnpei.21 The structuresof these antibiotics were determined based on their spectraldata as: 8-chloro-9-hydroxy-8,9-deoxyasperlactone (6), 9-chloro-8-hydroxy-8,9-deoxyasperlactone (7) and 9-chloro-8-hydroxy-8,9-deoxyaspyrone (8). Compound 8 inhibited the growth of Ruegeriaatlantica at 5 lg disk−1 (inhibition zone: 12.7 mm), while 7 and 8were active at 25 lg disk−1 (10.1 and 10.5 mm, respectively). Kojicacid dimethyl ether (9) has been isolated from the organic extractof the broth of the marine fungus Alternaria sp. collected from thesurface of the marine green alga Ulva pertusa.22 The structure wasassigned on the basis of comprehensive spectroscopic analyses.Osterhage et al. isolated the fungus Phoma tropica from the innertissues of the marine brown alga Fucus spiralis. This strain yielded5-hydroxyramulosin (10).23 The structure elucidation was mainlybased on 1D and 2D NMR and single crystal X-ray diffractionanalysis. Son et al. carried out bioassay-guided fractionation ofan organic extract of the broth from the marine fungal culture

(Aspergillus parasiticus) which led to the isolation and subsequentstructural elucidation of parasitenone (11), which showed moder-ate activity in the free radical scavenging assay.24 Dihydroterrein(12) was identified from the marine-derived fungus Emericellavariecolor. The molecular structure was established by means ofspectroscopic techniques and some chemical transformations.25

Two 10-membered macrolides: modiolides A (13) and B(14) were isolated from the cultured broth of a fungus Para-phaeosphaeria sp. (N-119), which was separated from a marinehorse mussel and the structures were elucidated by spectroscopicdata.26 Two polyketides: penicillones A (15) and B (16), withtricyclo[5.3.1.0]undecane skeletons, were isolated from Penicilliumterrestre.27 Their structures and relative stereochemistry wereestablished on the basis of spectroscopic methods. The absoluteconfiguration of penicillone B (16) was established by the modifiedMosher method, while 15 was deduced from the similar CDabsorptions of 15 and 16. Compound 15 showed weak cytotoxicityagainst P338 and A-549 cell lines, while 16 was inactive againstP388.

Two halogenated alkenoates: methyl-2,4-dibromo-5-oxo-2-decenoate (17) and methyl-2,4-dibromo-5-oxo-3-decenoate (18)were isolated from the broth of an unidentified marine fungus,which was separated from the surface of the marine red algaGracillaria verrucosa collected at Jinha, Ulsan.28 Two cytotoxictwelve-membered macrolides: sporiolides A (19) and B (20), havebeen reported from the cultured broth of a fungus Cladosporiumsp., which was separated from an Okinawan marine brownalga Actinotrichia fragilis, and the structures were elucidated byspectroscopic means.29 Sporiolides A (19) and B (20) exhibitedcytotoxicity against murine lymphoma L1210 cells. Sporiolide A


(19) showed antifungal activity against Cryptococcus neoformansand Neurospora crassa. Aspermytin A (21) was isolated from acultured marine fungus, Aspergillus sp., which was separated fromthe mussel Mytilus edulis. Its structure was elucidated on thebasis of their spectral data and the absolute configuration wasdetermined by the analysis of its CD spectrum.30 Aspermytin Ainduced neurite outgrowth in rat pheochromocytoma (PC-12) cellsat concentration of 50 lM. Phomoxin (22) and phomoxide (23)were isolated from the fermentation broth of a marine fungusof the genus Phoma (strain CNC-651). These compounds arehighly oxygenated polyketides of a rather new structural class.31

Compound 22 contains an unusual cyclic carbonate functionalitythat is rare among natural products. Structures of these metabo-lites were assigned by spectroscopic methods including 2D NMRspectroscopic analysis.

Shimalactone A (24) was obtained as a novel polyketide havingbicyclo[4.2.0]octadiene and oxabicyclo[2.2.1]heptane units, froma cultured marine fungus of Emericella variecolor GF10. Thestereo-structure of 24 was deduced from 2D NMR and X-raycrystallographic analyses of its derivatives.32 Shimalactone A (24)induced neuritogenesis at 10 lg mL−1 against neuroblastomaNeuro 2A cells. A modest cytotoxic polyketide glycoside, cladionolA (25), was isolated from the cultured broth of a fungus

Gliocladium sp., which was separated from sea grass Syringodiumisoetifolium and the structure was elucidated by physical data.33

The relative stereochemistry of C-2–C-3 was assigned by mainlyJ-based configuration analysis, while those of two sugar units wereelucidated to be b-mannopyranoside and arabitol on the basis ofNOESY spectral data and/or 1H–1H couplings. Furthermore, theabsolute configuration of the mannose moiety was determined asD-form on the basis of chiral HPLC analysis of a benzoyl derivativeof the acid hydrolyzate of 25.

From a strain of the fungus Emericella variecolor derivedfrom the marine sponge Haliclona valliculata, two metabolites:evariquinone (26) and isoemericellin (27), were isolated afterHPLC-UV, -MS, and -NMR studies of the extract and theirstructures were elucidated by mass spectrometry and NMRexperiments.34 Evariquinone (26) showed antiproliferative activitytowards KB and NCI-H460 cells at a concentration of 3.16 lgmL−1. Abdel-Lateff et al. cultivated a marine fungal isolate, War-domyces anomalus, and found two xanthone derivatives: 2,3,6,8-tetrahydroxy-1-methylxanthone (28) and 2,3,4,6,8-pentahydroxy-1-methylxanthone (29).35 The structures of both the compoundswere determined on the basis of extensive spectroscopic measure-ments (1D and 2D NMR, MS, UV and IR). Compound 28 showedsignificant antioxidant activity (94.7% at 25.0 lg mL−1).

Malmstrom et al. isolated varitriol (30), varioxirane (31) andvarixanthone (32) from a marine fungus Emericella variecolor.Their structures were established by means of spectroscopictechniques and some chemical transformations.25 In the NCI’s60-cell panel, varitriol (30) displayed increased potency towardselected renal, CNS and breast cancer cell lines. Varixanthone(32) showed antimicrobial activity against Gram-positive andGram-negative bacteria, higher than terrein, shamixanthone andtajixanthone hydrate.


Four anthraquinone derivatives: averufin (33), nidurufin (34),versicolorin C (35) and physcion (36) were derived from the man-grove fungus #2526 from the South China Sea.36 Their structureswere elucidated by comprehensive spectroscopic methods.

Paecilospirone (37) was isolated and characterized from amarine fungus collected in tropical and sub-tropical coral reefenvironments.37 A spiroacetal compound named paecilospirone(38) was isolated from the marine fungus Paecilomyces speciescollected at Yap Island.38 Paecilospirone (38) was found to be aninhibitor of microtubule assembly using conidia of Pyriculariaoryzae. The structure was assigned by spectroscopic data. Twometabolites possessing a phenalenone skeleton: sculezonones A(39) and B (40), were isolated from cultured broth of the fungusPenicillium sp., which were separated from the Okinawan marinebivalve Mytilus coruscus and the structures were elucidated byspectral analysis.39 Sculezonones A (39) and B (40) inhibit bovineDNA polymerases a and c, and moderately affect the activity ofDNA polymerase e.40

Six chromone derivatives: aspergione A (41), B (42), C (43),D (44), E (45) and F (46) were reported from fungal isolates ofAspergillus versicolor (Vuill) obtained from the marine spongeXestospongia exigua.41 Structures of all the metabolites wereestablished on the basis of extensive spectroscopic techniques.

Lin et al. described the isolation and characterization of fournovel secondary metabolites: aspergillone (47), aspergillodiol (48),aspergillol (49) and 2-acetyl-aspergillol (50) from the culturedfiltrates of fungus Aspergillus versicolor obtained from marinesponge Xestospongia exigua.42 The structures of compounds 47–49 were established on the basis of 2D NMR spectroscopy inconjugation with MS, UV spectral analysis. The basic structure ofthese compounds possesses a hydroindenoisopyran nucleus.

3 Terpenoids

Terpenoids are a class of compounds characteristic of somefamilies of terrestrial plants and microorganisms. Interestingly,metabolites of this class have also been isolated as bioactiveagents from marine sources. Chinworrungsee et al. reported aophiobolane sesterterpene named halorosellinic acid (51), fromcultured broth of a marine fungus Halorosellinia oceanica BCC5149,43 which showed antibacterial activity with an IC50 valueof 13 lg mL−1. It exhibited low antimycobacterial activity withan MIC value of 200 lg mL−1. Another sesterterpene epoxy-diol, aspergilloxide (52), has been obtained from the extract ofa cultured marine fungus (strain CNM-713) identified as anundescribed member of the genus Aspergillus.44 The structureof 52 was determined by interpretation of NMR data as wellas by chemical methods. The absolute stereochemistry of as-pergilloxide was assigned by application of the modified Mosher


method. The carbon skeleton of 52 represents a new additionto the architectural diversity of the sesterterpenoid (C25) classof secondary metabolites obtained from marine fungi. Wei et al.reported cytotoxic sesterterpenes 6-epi-ophiobolin G (53) and 6-epi-ophiobolin N (54) from the extracts of the fungus Emericellavariecolor GF10, which were separated from marine sediment.45

Their structures were deduced by analysis of 2D NMR spectraand the stereochemistry was determined by extensive examinationof NOESY spectra. Compounds 53 and 54 showed cytotoxicityagainst a neuroblastoma cell line. The cytotoxic polyoxygenatedfarnesylcyclohexenones: 7-deacetoxyyanuthone A (55) and its2,3-hydro derivative (56) were isolated from a marine isolateof the genus Penicillium.46 The structures of 55 and 56 wereassigned by spectroscopic methods including 2D NMR and CDexperiments. Compound 55 showed moderate in vitro cytotoxicityin a panel of 5 human tumor cell lines and 55 also exhibitedmild in vitro antibacterial activity against methicillin-resistant andmultidrug-resistant Staphylococcus aureus (MIC, 50 lg mL−1).Farnesylhydroquinone (57) has been isolated from the myceliumof a marine fungus of the genus Penicillium.47 The structure ofcompound 57 was elucidated by physical methods. Compound 57exhibited potent radical scavenging activity (IC50 12.5 lM) againstDPPH.

An eremophilane sesquiterpene 07H239-A (58) from a marineXylariaceous fungus was isolated and characterized with thehelp of NMR and mass spectrometry.48 This compound showedcytotoxicity towards a variety of cancer cell lines, with someselectivity for a CCRFCEM leukemia line (IC50 = 0.9 lg mL−1).Phomactin H (59), a diterpene with an oxepane moiety, wasreported from cultures of an unidentified marine fungus.49 The

structure and relative stereochemistry were determined by X-ray diffraction analysis. A new macrocyclic trichothecene, 12,13-deoxyroridin-E (60) was obtained as a cytotoxic component fromthe marine fungus Myrothecium roridum, isolated in Palau. Thiscompound is the second example of a macrocyclic trichothecenepossessing a double bond between C-12–C-13 and was about 80-fold less cytotoxic than roridin E, the epoxide variant.50

4 Nitrogenous compounds (alkaloids and peptides)

4.1 Alkaloids

Structurally diverse alkaloids have been isolated from variousstrains of marine fungi. Many of them possess potent biolog-ical activities and can be good candidates for development ofpharmaceutical products. A novel chiral dipyrrolobenzoquinonederivative, terreusinone (61), was isolated as a potent UV-A protec-tant from the marine algicolous fungus Aspergillus terreus strain(culture # MFA 460).14 The structure and absolute stereochemistrywere established by spectral interpretation, Horeau’s method andquantum chemistry calculations. This compound exhibited a UV-A absorbing activity with an ED50 value of 70 lg mL−1.

Another new pentacyclic oxindole alkaloid, speradine A (62),was isolated from the cultured broth of a fungus Aspergillustamarii, which was separated from driftwood at a seashorein Okinawa.51 The structure and relative stereochemistry weredetermined by spectroscopic data and a single crystal X-raydiffraction analysis. Li et al. reported indolyl alkaloid, Nb-acetyltryptamine (63), from the organic extract of the broth of anunidentified fungus collected from the surface of the marine redalga Gracilaria verrucosa.52 The structure of Nb-acetyltryptamine(63) was assigned on the basis of comprehensive spectroscopicanalyses.


Sasaki et al. reported a novel tetracyclic alkaloid, perinadine A(64), isolated from the cultured broth of the fungus Penicilliumcitrinum, which was separated from the gastrointestine of amarine fish and the structure was elucidated on the basis ofspectroscopic data including 2D NMR spectra.53 Perinadine A(64) may be derived from citrinin (65), a well-known mycotoxin,and a scalusamide A-type pyrrolidine alkaloid.

Three pyrrolidine alkaloids: scalusamides A–C (66–68) werefound from the cultured broth of the fungus Penicillium citrinum,which was separated from the gastrointestine of a marine fishand the structures were elucidated by spectroscopic data.54 Theabsolute stereochemistry at C-2 in the pyrrolidine unit wasdetermined by HPLC analysis of a Marfey derivative of thehydrolysate of 66, while those of 67 and 68 were assigned bycomparison of spectral data of 64 and reductive products of 66and 67. On the other hand, each metabolite (66–68) was found asa mixture of C-7 epimers. Scalusamide A (66) exhibited antifungalactivity against Cryptococcus neoformans (MIC 16.7 lg mL−1) andantibacterial activity against Micrococcus luteus (MIC 33.3 lgmL−1).

A novel pentacyclic alkaloid, citrinadin A (69), was isolatedfrom the cultured broth of the fungus Penicillium citrinum, whichwas separated from a marine red alga, and the structure waselucidated spectroscopically.55 The relative stereochemistry of thepentacyclic skeleton was assigned on the basis of NOESY data plus1H–1H coupling constants and the presence of an N,N-dimethyl-L-valine residue in 69 was detected by HPLC analysis of thehydrolysate using a chiral column. A diketopiperazine alkaloid,golmaenone (70), has been isolated from the cultured broth of amarine fungus Aspergillus species.56 The structure and absolutestereochemistry of 70 were assigned by spectroscopic methodsand the advanced Marfey method. This compound exhibitedsignificant radical scavenging activity against DPPH with an IC50

value of 20 lM, which was similar to the positive control (ascorbicacid IC50, 20 lM). Compound 70 also showed UV-A (320–390 nm)protecting activity with an ED50 value of 90 lM, which is moreactive than oxybenzone (ED50, 350 lM) currently in use as asunscreen.

Trichodermamides A (71) and B (72) were isolated fromcultures of Trichoderma virens.57 The trichodermamides possessa rare cyclic O-alkyl-oxime functionality incorporated into a 6-membered ring. The structure of 72 was established by X-raydiffraction analysis, while the structure assignment of 71 anddetermination of the absolute stereochemistry were accomplishedby spectral and chemical methods. Compound 72 exhibitedsignificant in vitro cytotoxicity against HCT-116 human coloncarcinoma with an IC50 of 0.32 lg mL−1. Lin et al. isolatedpenicillazine (73) as a new compound with both quinolone and4H-5,6-dihydro-1,2-oxazine ring systems that was isolated froma culture of the marine fungus Penicillium sp. (strain #386). Itsstructure was elucidated using spectroscopic methods, primarily2D NMR techniques, and confirmed by X-ray diffraction analysis.Variations in the 1H NMR spectrum of penicillazine (73) wereobserved at a 65 ◦C temperature.58

Lin et al. isolated the fungus Aspergillus versicolor fromthe marine sponge Xestospongia exigua collected in Indonesia.Following cultivation in a seawater-based medium they isolated anew angular tricyclic chromone derivative (74) from the myceliaand culture filtrate.59 The structure of this natural product wasestablished on the basis of extensive one- and two-dimensionalNMR spectroscopic studies as well as on mass spectral analysis.


Two new antimicrobial pyrrolidinone derivatives: zopfiellamidesA (75) and B (76) were isolated from fermentations of thefacultative marine fungus Zopfiella latipes CBS 611.97. Theirstructures were elucidated by spectroscopic techniques.60 Theantimicrobial activity of zopfiellamides A (75) and B (76) towardsvarious bacteria and yeasts was found to be moderate towardsthe Gram-positive: Arthrobacter citreus, Bacillus brevis, B. subtilis,B. licheniformis, Corynebacterium insidiosum, Micrococcus luteus,Mycobacterium phlei and Streptomyces sp., and towards the Gram-negative: Acinetobacter calcoaceticus with minimal inhibitoryconcentrations between 2 and 10 lg mL−1 of zopfiellamide A (75).Zopfiellamide B (76) was found to be approximately 5 times lessactive than 75. The yeasts Nematospora coryli and Saccharomycescerevisiae were affected by both the metabolites with minimalinhibitory concentrations starting with 2.0 lg mL−1 for bothcompounds. No cytotoxic effects toward several mammalian celllines were observed for either compound, at concentrations up to100 lg mL−1.

Four cytotoxic disulfides: rostratins A–D (77–80) were obtainedfrom the whole broth of the marine-derived fungus Exserohilum ro-stratum (Drechsler), a fungal strain found associated with a marinecyanobacterial mat.61 Their structures were established throughchemical degradation and a variety of two-dimensional NMRtechniques. The absolute configurations of 77–80 were determinedby the modified Mosher method. In the cases of mercaptols 77and 80, regioselective acylation was achieved by modulating thereaction temperature monitored by 1H NMR. These compoundsshowed in vitro cytotoxicity against human colon carcinoma(HCT-116).

4.2 Peptides

Hundreds of peptide antibiotics have been described in the pasthalf-century.62–64 These fall into two classes: non-ribosomally

synthesized peptides, such as the gramicidins, polymyxins, bac-itracins, glycopeptides, etc., and ribosomally synthesized (natural)peptides. The former are often drastically modified and are largelyproduced by bacteria, whereas the latter are produced by all speciesof life (including bacteria and fungi) as a major component ofthe natural host defence molecules of these species63–64 and havepotential biological activities.

Not much work has been done on peptides from marine fungi.Komatsu et al. have reported dictyonamides A (81) and B (82)from a fungus (KO63) separated from the red alga Ceratodictyonspongiosum.65 Both the compounds are linear dodecapeptidesof similar sequence containing N-methylamino acids and ananthranilic acid at the C terminus. Compound 81 has beenreported to inhibit cyclin-dependent kinase 4 with the IC50 value16.5 lg mL−1.

Rowley et al. have described a series of peptides designatedhalovirs A–E (83–87), obtained during the saline fermentation ofa marine fungus Scytalidium sp. These lipophilic linear peptidesare potent in vitro inhibitors of the herpes simplex viruses 1 and2.66 The complete structures of these compounds were establishedby a combination of spectral and chemical techniques.

The fungus Aspergillus carneus, isolated from an estuarinesediment collected from Tasmania and Australia, yielded a seriesof new depsipeptides, aspergillicins A–E (88–92).67 Characteri-zation and complete stereostructures of aspergillicins A–E wereelucidated on the basis of detailed spectroscopic analyses, togetherwith ESIMS analysis of the free amino acids generated by acidhydrolysis and HPLC analysis of Marfey derivatives preparedfrom the acid hydrolyzate. The peptide amino acid sequence for allaspergillicins was unambiguously assigned by MSn ion-trap ESImass spectrometry.

Two anti-dinoflagellates: clonostachysins A (93) and B (94)were obtained from a marine sponge derived fungus Clonostachysrogersoniana strain HJK9.68 Their structures were determinedby spectroscopic studies as highly N-methylated cyclic peptidescontaining nine amino acids. The absolute stereochemistry was de-termined by the advanced Marfey method. Both clonostachysins


A and B exhibited a selectively inhibitory effect on a dinoflagellateProrocentrum micans at 30 lM but showed no effect on othermicroalgae and bacteria even at 100 lM.

5 Cerebroside analogues

Flavicerebroside A (95) and B (96) have been obtained from themycelium of the marine fungus Aspergillus flavipes isolated fromthe sea anemone Anthopleura xanthogrammica.69 Their structureswere determined by means of extensive spectroscopic analyses(IR, UV, 2D NMR, MS, CD) and chemical degradation. Thesecompounds showed cytotoxic activity against the KB cell line.

6 Miscellaneous metabolites

The unusual habitats of the marine fungi developed the abilityto produce structurally and biologically diverse metabolites andmany of the novel compounds isolated from this group oforganisms belong to diverse structural classes. Marine fungi area source of compounds having significant chemical diversity. A2-hexylidene-3-methylsuccinic acid 4-methyl ester (97) has beenreported from cultured broth of a marine fungus Haloroselliniaoceanica BCC 5149 and the structure was determined by

NMR/mass spectral data.44 This metabolite may derive fromsuccinic acid. Namikoshi et al. isolated phomopsidin (98) frommarine fungi collected in tropical and sub-tropical coral reefenvironments.37 The authors discussed the characterization ofcompound 98 and utility of marine fungi as a source of bioactivesecondary metabolites. The biosynthetic study using labeled-carbon precursors revealed the origin of all carbon atoms in 98,which was built by nine acetates and three methyl groups fromL-methionine. A new metabolite possessing a 1,3-dioxane ringnamed coruscol A (99) was isolated from the mycelium of a Peni-cillium sp. separated from the Okinawan marine bivalve Mytiluscoruscus. The structure of 99 was elucidated by spectroscopicdata.70 This is the first isolation from a natural source, although ithas been reported as a synthetic compound.71 Compound 99 maybe biosynthetically derived from erythritol and n-butyraldehyde.Spiciferol A (100) and a monocyclic spiciferone derivative (101)have been purified from a fungus Drechslera hawaiiensis obtainedfrom the marine sponge Callyspongia aerizusa collected from theSea of Bali, Indonesia. The structures of these novel compoundswere established on the basis of NMR spectroscopic (1H, 13C,COSY) and mass spectrometric (EIMS) data.72 Compounds 100and 101 have basic skeleton like spiciferone A. The biosyntheticpathway to spiciferone A was already investigated by Nakajimaet al.,73 through incorporation experiments with [1-13C], [2-13C],[1,2-13C] and [1-13C,2H3] acetate and [S-13CH3]-L-methionine. Theincorporation patterns suggested that they are produced from acommon precursor, which is derived from a hexaketide and twoC1 units, after undergoing modifications including the uniqueC–C bond cleavage by retro-aldol condensation. Two 3-amino-5-ethenylcyclopentenones: myrothenones A (102) and B (103)were isolated from a marine algicolous fungus of the genusMyrothecium. The structure and absolute stereochemistry ofthese compounds were established by spectral interpretation andX-ray analysis.74 Compound 102 exhibited a tyrosinase inhibitoryactivity with IC50, 6.6 lM, which is more active than kojic acid


(IC50 7.7 lM) currently being used as a functional personal-carecompound. Myrothenone A (102) is a formamide derivative ofmyrothenone B (103). These vinylcyclopentenone analogues havebeen reported as an artifact75 and a conjugated adduct76 butnatural products of this class having amino group are very rareexamples.

7 Conclusions

Among the 103 metabolites observed, approximately 50% werepolyketide derived. Metabolites of this class might constitutethe largest part of the secondary metabolites of terrestrial andmarine microorganisms. Terrestrial sources have been intensivelyinvestigated for their metabolites and even marine bacteria offereda considerable number of this class of compounds. Marine fungiliving in diverse environments in association with invertebrates arevery sensitive to culture media and planned research can producemore potent polyketides from this source. Specifically, fused-ringpolyketides are potential candidates responsible for antioxidant,antibiotic and anti-cancer properties can also be obtained by theseorganisms.

The number of reported terpenoids is only ten and most ofthese are sesterterpenoids. This class of compounds also possessantibiotic and cytotoxic properties and can be good candidates forchemotherapy. Unfortunately, very few metabolites of this groupwere reported from fungi.

During the last five years a total of 20 alkaloids and 14peptides have been reported from marine fungi. With the recentadvances in the areas of peptide synthesis, screening, stability andmodifications, peptides are now recognized as lead molecules fortherapeutics.77

In this review, seven compounds of unusual structures arereported. Among them, myrothenone A (102) showed enzymetyrosinase inhibiting properties, and was found to be more potentthan kojic acid. Some metabolites exhibit cytotoxic activity as well.

In some cases, it is extremely difficult to provide sufficientamounts of active substances from fungi due to their limitedlevels of biosynthesis. The limited quantity might be due to therare occurrence of the such organisms themselves, or geographic,seasonal or sexual variations may affect the amount and nature ofproduced secondary metabolites. On the other hand, the structuralcomplexity in most of the interesting bioactive substances pre-cluded the development of commercially viable syntheses. Casesof direct isolation of target compounds from bacterial or fungalsymbionts of invertebrates are rare. Although fermentation ofmicroorganisms is a possible way, it does not apply to metabolitesproduced strictly by macroorganisms. Research on marine fungiand use of biotransformation and biotechnological methods mayhelp to obtain potent candidates for the treatment of diseases.

8 Acknowledgements

We thank Prof. Dr J. R. Hanson, Sussex University (UK), forvaluable comments and suggestions in manuscript preparation.

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