Isolation And Identification Of Secondary Metabolites From Marine Derived Actinomycete HN-3-14

The experimental strain is a Mycale-phvllophla sponge isolated from Yangpu Bay,Hainan Province,China,at a depth of 20 meters(1096'E,.1941'N).The 16 S rDNA sequence had the highest homology with Microbacterium esteraromaticum DSM 8609(T),up to 99.2%.The strain was identified as Microbacterium sp.,named HN-3-14.The strain was deposited in the symbiotic microbial Bank of marine organisms of Xiamen Medical College.The culture conditions of HN-3-14 were compared and analyzed from the environmental factors such as basic medium type,salt concentration,pH value and culture temperature.The results showed that the strain had good environmental adaptability.(Here,the culture conditions for normal growth of the medium are provided.In order to better activate the gene expression of secondary metabolites of the strain,according to the principle of metabolic engineering of actinomycetes,the strain had good environmental adaptability.Glucosamine,mannitol,peanut oil and other metabolic pathway feedback inhibitors were added to the culture medium respectively.The abundances of secondary metabolites in different culture conditions were analyzed.It was found that the strain could obtain better secondary metabolites abundances in oligotrphic medium with peanut oil(0.1%),salt concentration: 3%,pH: 7.5 and temperature: 28 ?.The strains were cultured for 28 days with 180 rpm shaking.After removing mycelium by centrifugation,the fermentation broth was incubated with macroporous resin for adsorption,60% ethanol elution to collect metabolites The extract of secondary metabolites was 162.1 g by rotary evaporation.Secondary metabolites were isolated and purified by TLC TLC,Sephadex LH-20 gel column chromatography,normal silica gel column chromatography and HPLC high performance liquid chromatography.10 compounds were isolated and purified.According to the analysis data of nuclear magnetic resonance spectroscopy(N MR)and ESI-high resolution mass spectrometry(ESI-HRMS),they were resolved i nto pyrimidine-2,4(1H,3H)-dione(1),4-hydroxybenzaldehyde(2),(S)-N-(S)-(2-hydro xy-5-oxo-2,5-dihydrofuran-3-yl)acetamide(3),N-5-(2-hydroxytyl)furan-3-yl)acetamide(4),2-hydroxybenzoic acid(5),indoline-3-cardehyde(6),(Z)-5-hydroxy-2-(2-hydro xyethylidene)-2H-pyrrol-4-yl acetate(7),(E)-5-hydroxy-2-(2-hydroxyethylidene)-2H-p yrrol-4-yl acetate(8),N-(5-hydroxy-2-oxo-2H-pyrrol-4-yl)acetamid(9),N-(5-(1,2-di hydroxyethyl)-2-oxo-2,5-dihydrofuran-3-yl)acetamide(10)? Compound 7 and compo und 8 are isomers.Compounds 3,4,7,8 and 9 have not been reported by SCI F INDER.The research in this paper shows that symbiotic actinomycetes of marine organisms are an important source of natural products of new chemical structures,which will provide more new chemical entities for new drug research and development.