Isolation,Purification And Structure Elucidation Of Secondary Metabolites From Marine Bacterium G0018B

In recent decades,with the massive development of terrestrial natural products,the rate of discovering novel natural prouducts form terrestrial resources has been continuously decreasing.However,in the ocean there are more complex and interesting natural products,of which some marine natural products or their derivatives have been in clinic or clinic trials.With the rapid advancement in bioinformatics,genetics and biochemistry,biosynthetic pathways and enzymology of a number of marine natural products have been extensively studied.The development and application of DNA sequencing technology and analytical techniques have driven the method,isolating and purifying novel marine natural products by genomics-guided fractionation,to become one of the main platforms for drug discovery and development.In this study,we identified a new marine strain G0018B from two aspects of morphology and molecular biology,and found that:1)the colonies was yellow and shaped like transparent and smooth minidisk;high-power microscopic observations showed that the individual strain was rod-shaped;2)the 16S rDNA sequence analysis indicated that G0018B strain had the highest homology with Idiomarina xiamenensis strain 10-D-4,and the phylogenetic tree also proved the kinship.Genome mining of strain G0018B indicated that this strain possessed a functional gene cluster encoding a thiopeptide and several other unknown gene clusters,which indicated that the strain may produce a new thiopeptide compound.The strain was massively cultivated by fermentation cylinder.Guided by antibacterial bioassay,the crude extract was isolated and purified through various chromatographic methods,including high performance liquid chromatography(HPLC),ODS(C18)reversed phase column chromatography and Sephadex LH-20 molecular sieve.Chemical structures were identified by high resolution MS and one-dimensional as well as two-dimensional NMR spectra.Ultimately,seven compounds were obtained:Compound 1:Butyl isobutyl phthalate;Compound 2:Dibutyl phthalate;Compound 4:Cyclo-(4-hydroxy-prolyl-phenylalanine),with an uncertainty of steric structure;Compound 5:Cyclo-(4-hydroxy-prolyl-phenylalanine),but determined by NMR it is a spatial isomer different from compound 4,and its spatial structure is to be determined;Compound 6:2-Methyl-N-(2'-phenylethyl)-butyramide;The amount of Compound 3 extracted was too small to identify the structure;It was initially judged that the structure of Compound 7 might contain multiple rings,and it was yellow powder.In addition,these 7 compounds were tested for antibacterial activity and were found that compound 2 had activities against Staphylococcus aureus and Bacillus subtilis under the positive control of ampicillin,with inhibition rate of 47.0%and 50.0%,respectively,at a concentration of 25?g/mL.From this study we obtained a preliminary comprehension of the profiles of secondary metabolites of strain G0018B which provided a guide for subsequent studies.Furthermore,we will continue to isolate and purify genomically predicted thiopeptide antibiotics to cope with the roughly international problem of growing bacterial resistance.