Biological Activity Of Secondary Metabolites Of Endophytic Actinomycetes From Dendrobium Officinale

Endophytes are abundant microbial resources and actinomycetes are important producing bacteria of active natural products.Therefore,plant endophytic actinomycetes have great potential as microbial resources.Dendrobium officinalis is a precious traditional Chinese medicine with important physiological activity.Horizontal gene transfer or gene recombination may occur between the endophytic actinomycetes and the host,so that the endophytic actinomycetes can produce the same or similar physiological active substances with the host.Therefore,this study took endophytic actinomycetes of Dendrobium candidum as the research object,and studied the biological activities of secondary metabolites.The main research contents and results are as follows:In this study,13 strains of endophytic actinomycetes isolated from Dendrobium officinale which were isolated and preserved by Hebei Province's Microbial Diversity Research and Application Laboratory as research objects.Firstly,phylogenetic analysis of 13 strains was carried out.The results showed that among the 13 tested strains,there were 6strains of Streptomyces,4 strains of Pseudonocardia,1 strain of Mycobacterium,1 strain of Mycolicibacter and 1 strain of Nocardiopsis,indicating that the species diversity of endophytic actinomycetes in Dendrobium candidum was relatively rich.The inhibitory activity of the tested strain of the secondary metabolites were measured by alpha-glucosidase,the MCF-7 cell line of breast cancer and the pathogenic indicator bacteria.The results show that six of the strains showed the alpha-glycosidase enzymes were more than 50% of the inhibitory activity,two of the strains of alpha-glycosidase enzyme inhibition rate above 70% and seven of the strains showed on breast cancer MCF-7 cell inhibition rate more than 40%,with twelve of the tested strains inhibited at least one pathogenic indicator,among them,strains 2055473,205518-1,205524 and 205527 showed broad spectrum antibacterial activity,In general,the inhibitory effect of the tested strains on pathogenic bacteria was significantly higher than that of pathogenic fungi.According to the results of the three activity assays,ten of the endophytic actinomycetes have two or morekinds of activity at the same time.Among them,the active bacteria of Streptomyces is better than the tested strains of other genera,and its active substance is worth further study.Based on the preliminary analysis of the activity results and the secondary metabolites of the active strain,a streptomyces strain 205527 with good activity and abundant secondary metabolites was selected as the starting strain to study the secondary metabolites.The ultraviolet characteristic absorption of compound peaks was detected by HPLC,and it was determined that F fermentation medium was used for batch fermentation.The active strain was fermented by fermentation in a fermenter,and a total of 145 L fermentation product was obtained.The fermentation broth was extracted by ethyl acetate and concentrated by spin evaporation,and 30.63 g crude extract was obtained.The extract was separated and purified by thin layer chromatography,gel column chromatography,Biotage rapid preparation chromatography,high-performance liquid chromatography,and preparative liquid chromatography.Pure compounds structure identifited by 1H-NMR,13C-NMR,and LC-MS,finally obtained four active compounds which respectively were 4-N-hydroxy-l-N-methy-3-E--benzylidene-6-E-isobutylidene-2,5-piperazinediketone,1H-pyrrole-2-formamide,(3Z,6E)-1-N-methyl-3-benzylidene-6-(2S-methyl-3-hydroxypropyl)-piperazine-2,5-diketone and salicylamide.