Xerocomus langbianensis. Trịnh Tam Kiệt Trung tâm Công nghệ Sinh học, Đại học Quốc gia Hà Nội Udo Graefer, Peter Saluz, B. Schelegel Viện nghiên cứu Hoá các hợp chất tự nhiên Jena, CHLBĐức. Chrysospermins A, B, C, D được chiết xuất như là những peptaibols ở quả thể của nấm Xerocomus langbianensis và cấu trúc của chúng đã được xác định bởi Mass spectrometrie. Những chrysospermin này đã được phát hiện trước đây trong khi nghiên cứu sinh khối của nấm sợi Sepedonium chrysospermum (telemorph Apiocrea chrysosperma). Những kết quả này có thể liên quan tới việc Xerocomus langbianensis đã bị tấn công bởi các đại diện của nấm sinh đính bào tử. I. Abstract Chrysospermins A- D were isolated recurrently as 19- membered peptaibols in a fruiting body of Xerocomus langbianensis and structure was confirmed by ESI mass spectrometry. The chrysospermins A – D 3 ) were detected formerly in cultures of Sepedonium chrysospermum (telemorph Apiocrea chrysosperma).The result suggested that the old Xerocomus langbianenis was infected by a representative of the conidial fungi. II. Introduction Peptaibols such as aibelin, alamethicins, atiamoebins, emerimicins, paracelsin, saturnisopin, suzukacillin, trichorzianins, trikoningins, zervamicins (3) are of considarable biological interest products because they faciliate the transport of ions across membranes via mechanism involving pore formation. The fungus Sepedonium chrysospermum (telemorph Apiocrea chrysosperma) was found to produce novel antifulgal peptides, chrysospermins A (1), B (2), C (3), D (4) as member of the peptaibol class of linear lipophilic peptide antibiotics (3). Therefore chrysospermins promote pigment formation by surface culture of the mold fungus Phoma destructiva. The fungus Sepedonium ampullosporum known as the producer of the similar peptaibols ampullosporins A- E 1,2 ) . The another filamentous fungi such as Trichoderma herxianum, Acremonium sp. and various other Hyphomycetes 4,5 ) are able to produce petaibols. Here we report the occurrence of chrysospermins A-D (1-4; Fig.1) in the fruit body of Xerocomus langbianensis. III. Material and methods: Specimens of Xerocomus langbianensis were collected in the pine forest near Dalat (South Central Vietnam) 7) . Taxonomical studies were carried according Roef Singer (1987). A old fruiting body of this mushroom was lyophilized to yield 120g dry material and researched the bioactive compound according Udo Graefer (1995). IV. Results and discussion 1. Xerocomus langbianensis Cap 8 – 30 cm wide; convex, becoming nearly flat; smooth to somewhat pitted, usually cracking in dry weather as well as when old; brown or cinnamon-buff. Flesh white; cap skin parenchyma. Tubes: sunken around stalk, yellow. Stalk 8 – 20 cm long, 3 – 5 cm thick and cylandric or bulbous in the middle. Flesh white. Spores: 11 – 14 x 3 – 4 m cylindric eliptical, smooth with several oil drops inside. Good taste. Growing season: raining time. on the ground under conifers in Dalat, Lamdong province. (Pic. 1, 2) It was extracted five-times for each 24 hours by 500 ml CHCl 3 /MeOH (1:1, v/v), and the combined extracts were evaporated in vacuo. The residue (5.4 g) was subjected to column chromatography on silicagel 60 (0.063 – 0.1 mm, column 4 x 80 cm ). The components of the mixture were eluted in order of their polarity by CHCl 3 , CHCl 3 /MeOH (9:1) and CHCl 3 /MeOH (8:2). 20 ml fractions were collected and evaporated. Samples of the residues were spotted on TLC sheets (silicagel 60, Merck) and developed by CHCl 3 /MeOH, 9:1). Subsequently the chromatograms were stained by 1% vanillin in conc. H 2 SO 4 . First ergosterol and other terpenoid materials were eluted. Later fractions occurrence containing linolic acid, cerebroside B, honogerine lipids. Finally a fraction occurred with R f 0.1 o- n TLC (see above) staining reddish with the above spray reagent (yield: 8 mg). The sample this obtained was analyzed by electrospray triple quadrupole mass spectrometry (Quattro instrument, VG Biotech, Altrincham, England). Pseudomolecular ions with m/z 1898, [M+H] + (1), m/z 1912 [M+H] + (2,3) and m/z 1926 [M+H] + (4) were readily disclosed as characteristics of the chrysospermins A-D (1-4). (see c.f. Fig. 2: FAB-MS of 2). Figure 1: Amino acid sequences of chrysospermins A (1), B (2), C (3), D (4) and boletusin (5): 1 AcPhe Aib Ser Aib Aib Leu Gln Gly Aib Aiib Ala Ala Aib Pro Aib Aib Aib Gln Trpol 2 AcPhe Aib Ser Aib Aib Leu Gln Gly Aib Aib Ala Ala Aib Jpro Iva Aib Aib Gln Trpol 3 AcPhe Aib Ser Aib Iva Leu Gln Gly Aib Aib Ala Ala Aib Pro Aib Aib Aib Aln Trpol 4 Acphe Aib Ser Aib Iva Leu Gln Gly Aib Aib Ala Ala Aib Pro Iva Aib Aib Gln Trpol 5 AcPhe Aib Ala Iva Leu Gln Gly Aib Aib Ala ala Aib Pro Aib Aib Aib Gln Trpol The resuls with Xerocomus langbianensis confirm the occurrence of chrysospermins 3) in fruit bodies of basidiomycetes. Sang- Jun Lee et al 6) reported also in 1999 about isolation of chrysospermins A-D (1-4) and boletusis (5) from the fruit body of a mushroom Boletus sp However, the chrysospermins have been reported as products from submerged fermentations of Sepedonium chrysospermum (telemorph Apiocrea chrysospermum 6) ). Hence, it appears as unlikely that components 1-4 are products of fruit bodies of basidomycetes such as Xerocomus langbianensis. It can be suggested that occurrence of peptaibols 1-4 in the fruit body of Xerocomus langbianensis was due to the infection of the fruit body by a fungicolous connidials fungus as real producer of chrysospermins. Moreover it can be suggested that channel- forming chrysospermum 8) of a fungicolous strains 9) play a role in the infection process of the fruit body. Acknowledgements We gratefully acknowledge support of this work by DLR (Bonn, Germany) and FCI (Frankfurt/Main, Germany). References 1) Ritzau, M. et.al.:J. Antibiotics 50, 722-728 (1997) 2) Kronen, M. et.al.:J. Antibiotics 54, 175-178 (2001) 3) Dornberger, K., Ihn, W., Ritzau, M., Grọfe, U., Schlegel, B., Fleck, W.F., Metzger, JW.: J. Antibiot. 48, 977-989 (1995) 4) Laatsch, H.: Antibase, Database of microbial compounds. Chemical Concepts, Weinheim, 2000 5) Huang, Q. et al.: Chem. Pharm. Bull. 43, 223-229 (1995) 6) Lee, S.J., Lea, W.H., ,Yun, B.S., Yoo, I.D.: J. Peptide Science 5, 374-378 (1999) 7) Kiet, T.T., Dửrfelt, H.: Intern. Biol. Congr. Hanoi, Vietnam, Abstracts 2000, pp. 253-255 8) Grigoriev, P.A., Schlegel, R., Dornberger, K., Grọfe, U.: Biochem. Biophys. Acta 1237, 1-5 (1995) 9) Hawksworth, D.L.: In: Biology of Conidial Fungi, Vol.1, ed. By Cole, G.T. and Kendrick, B., Academic Press, N.Y. (1981), 171-235