Research Of The Secondary Metabolites From Two Antarctic Marine Fungi

The natural environment of polar region is harsh,with characteristics of severe cold,strong winds and strong ultraviolet radiation,leading to the absence of plants and animals,but microbiological resources such as bacteria,fungi,actinomycetes are rich and varied.In order to adapt to the polar natural ecological environment,polar microbes have formed unique molecular biological characteristics in the gene composition and expression,enzyme and related metabolic regulation,which can produce novel,unique secondary metabolites with various biological activity,such as antitumor,antibacterial activity,etc.These metabolites could be important sources of new chemical drugs.In the preliminary study in our group,fungi Penicillium sp.S-1-18 was screened by PTP1 B inhibitory activity as an indicator and TLC vanillin colorimetric analysis method.A medium containing 0.3 % yeast extract,0.5 % tryptone,0.3 % maltose,1 % glucose and 5 % sucrose was used,the initial pH was 7.0,then cultured at 18 °C and 180 r/min for 14 days.After the fermentation,the mycelia were filtered off and the solution was concentrated and extracted with butyl acetate.Finally,the total extract was about 60 g.On the basis of this,the identification of metabolites and the study of biological activity were carried out.The total extract was extracted with petroleum ether and methylene chloride.The extraction and purification of the dichloromethane phase extract were carried out systematically.By means of using Vacuum Liquid Chromatography(VLC),Sephadex LH 20 gel column chromatography and reverse phase ODS column Analysis,normal phase silica gel column chromatography and high performance liquid chromatography,nine compounds were identified,butanolide A(1),guignarderemophilane F(2),xylarenone A(3),penicyclone A(4),4(3H)-quinazolinone(5),cyclo-(L-Pro-L-Phe)(6),cyclo-(L-Phe-4R-hydroxyl-L-Pro)(7),callyspongidipeptide A(8),and N-(2-hydroxypropanoyl)-2-aminobenzoic(9).Compounds 1 and 2 were new compounds,and their absolute configuration was determined by modified Mosher's method and ECD/CD alignment method respectively.The biological activities of the compounds obtained in this strain were evaluated.The results showed that compound 1 had moderate PTP1 B inhibitory activity with IC50 value of 27.4 ?M.Besides,compounds 4,5,8 had weak inhibitory activity against Acinetobacter baumannii with IC50 values of 64 ?M,respectively.At the same time,an Antarctic marine-derived strain Penicillium sp.S-2-10 was screened by TLC vanillin colorimetric analysis Method.The fermentation conditions were optimized by TLC chromatography and product extract.The strain was cultured with GPY medium containing 1.0% NaCl at 24 °C and 180 r·min-1for 12 days.After 90 L scale fermentation,the mycelia and the broth were filtered through the gauze.The mycelia were extracted with methylene chloride and methanol.The broth was extracted with ethyl acetate and the combined extracts were concentrated to obtain about 16 g extract.The total extract was systematically separated and purified by Sephadex LH 20 gel column chromatography,normal phase silica gel column chromatography,reversed phase ODS column chromatography,high performance liquid chromatography and so on.The 1H-NMR,13 CNMR,mass spectrometry(MS)and other spectral analysis methods were used to determine the structure of seven compounds by comparing with the relevant literature.The compounds were identified as benzoic acid,2-(2,6-dihydroxybenzoyl)-3-hydroxy-5-(hydroxymethyl)-methyl,methyl ester(10),chrysophanol(11),volemolide(12),ergosta-6,8(14),22-trien-3-ol(13),2-aminophenoxazin-3-one(14),2-benzothiazolol(15),benzoic acid(16).Compounds 12 and 13 were firstly isolated from the fungus Penicillium sp.The biological activity of these compounds was evaluated and it was found that compound 12 had weak inhibitory activity of tumor cells.IC50 for inhibiting SMMC-7721 and SGC-7901 were 42.70 ?M and 49.98 ?M,respectively.Compounds 11,12,13,15 had weak inhibitory activity against Acinetobacter baumannii with IC50 values of 64 ?M,respectively.