Study On The Secondary Metabolites Of A Soil Actinomycete

The sources of natural products include animals,plants,and microorganisms.Among them,microorganisms have become an important source for the development of clinical antibacterial,anti-tumor,immunosuppressive drugs and other drugs due to their rapid growth and wide distribution.Today Most of the known antibiotics were originally isolated from actinomycetes,especially from the genus Streptomyces,so the study of Streptomyces is of great significance.In this project,30 strains of actinomycetes were isolated from soil samples,and 10 strains of actinomycetes with the fastest growth rate were selected.Using the bacterial block confrontation method,nine phytopathogenic fungi were used as indicator strains,and 10 actinomycetes were screened.Four actinomycetes were active against phytopathogenic fungi.Among them,GQ.3 was effective against Fusarium proliferatum and Fusariumsolani.Fusarium proliferatum,and Fusarium Link have good inhibitory effects.The 16 S r DNA sequence analysis showed that it belongs to the genus Streptomyces.Strain GQ.3 was inoculated on potato glucose medium,S medium,sporulation medium and Gao's No.1medium.It was found that strain GQ.3 grew fastest on Gao's No.1 medium and had the most metabolites.And the metabolites are well formed after being developed by thin-layer chromatography.Gao's No.1 medium was selected as the optimal fermentation medium,and the secondary metabolites were extracted with ethyl acetate.After vacuum distillation,20 g crude extract was obtained and separated and purified.At the same time,the whole gene sequencing was performed on strain GQ.3.Antis Mash software was used to analyze the genome sequencing results,and it was found that strain GQ.3 had 52 biosynthetic gene clusters of secondary metabolites.By analyzing these gene clusters,it could be seen that strain GQ.3 had the potential to produce secondary metabolites such as polyketones,terpenoids,lipopeptides and carbohydrates.The secondary metabolites were isolated and purified by column chromatography and spectral analysis,and their structures were identified.Eleven compounds were isolated and identified,including one long-chain fatty acid,one terpene,one indole,and eight polyketones.Among them,compounds 5 and 6 are similar in structure to the known macrolide antibiotic Concanamycin A,and both have the same 18-membered ring in the structure.Compound 6 is a new Concanamycins compound.The rest of the compounds are all known structures.The antibacterial activities of 11 compounds against Staphylococcus aureus,Escherichia coli,Pseudomonas aeruginosa and Bacillus subtilis were tested by filter paper method and micro-dilution method.Only compounds 5 and 6 have weaker activity against Bacillus subtilis,with a zone of inhibition of 8 mm.The cytotoxicity of all compounds to leukemia cells(HL60),breast cancer cells(A549),liver cancer cells(SMMC7721)and breast cancer cells(MCF7)was detected by MTT assay.None of the compounds had cytotoxicity.