Screening Of Antimicrobial Marine Actinomycetes,Isolation And Structure Elucidation Of The Antimicrobial Compounds

Objective:To screen out the marine actinomycetes with the antimicrobial activity from marine actinomycetes strain library.To isolate the antimicrobial compounds,and furtherly identify the structures.Accordingly discover the new resource of antimicrobial activity compounds from marine actinomycetes.Methods: First,screen out the marine actinomycetes with the antimicrobial activity: according to the classification information,select some classified marine actinomycetes strains from marine actinomycetes strain library.The marine actinomycetes strains were fully recovered and cultured with 2216 E liquid medium for 7 days.Collect the culture solution,the Filter method was performed to screen the antibacterial activity fermentation liquor to Bacillus subtilis,Staphylococcus aureus,Escherichia coli and Candida albicans.Second,the culture conditions of the antibacterial marine actinomycete were optimized:mainly to optimize the culture medium formula and culture time,determine the culture conditions with which the antibacterial marine actinomycete has the highest antibacterial activity.Third,isolate the antimicrobial activity compounds:according to the optimized culture conditions,cultivate the antibacterial marine actinomycete 150 L on a small scale,centrifuge and collect the fermented liquid,concentrate with 40? water bath rotating pan,extract with organic solvents.Antibacterial activity compound was then isolated from the fermentation liquor based on activity tracking and combined chromatographic separation techniques.Fourth,identify the structure of the isolated compounds:the structure of the isolated compounds were identified using NMR spectral analysis and high resolution mass spectrometry.Fifth,test the antimicrobial spectrum of the antimicrobial compounds:the Filter method was performed to analyze antibacterial activity of the antimicrobial compounds.Results: First,a marine actinomycetes strain was discovered with the antimicrobial activity,the serial number is 649-20,it was identified as a strain of Streptomyces tendae.This actinomycetes strain has significantly antibacterial activities to Bacillus subtilis and Staphylococcus aureus,but not showed significantly antibacterial activities to Escherichia coli and Candida albicans.Second,the strongest antibacterial activity of the fermentation liquor was observed when the strain was cultured with 2216 E medium for 96 h,under 28 ? and shaking under 180 rpm.Third,two antimicrobial compounds Cd-A and Cd-B were purified from the actinomycetes fermentation liquor using combined chromatographic methods.Fourth,the structures of two antimicrobial compounds were respectively identified as 2-Hydroxy-3-propyl-hexahydro-pyrrolo [1,2-a] pyrazine-1,4-dione and 3-Propylhexahydro-pyrrolo [1,2-a] pyrazine-1,4-dione.Both of them were discovered structures.Biology metabotelism results speculated that the compounds sourced from cyclin dipeptides.Fifth,the results of the antimicrobial spectrum indicated that two antimicrobial compounds Cd-A and Cd-B have significant antimicrobial activities for gram positive bacteria,in addition to this,compound Cd-B has significant antimicrobial activity for gram negative aquatic pathogenic bacteria.Conclusions:A marine actinomycetes which has significant antimicrobial activity was screened.Two cyclin dipeptides were purified from the actinomycetes fermentation liquor.They are different from the traditional antibiotic compounds,both of them have development potential for new antimicrobial agents.