Study On The Secondary Metabolites And Bioactivity Of Xenorhabdus Bovienii SN52 From Entomopathogenic Nematodes

Insect pathogens symbiotic bacteria is a class of drug-related bacteria which can produce various metabolites.These bacteria include Xenorhabdus and Photorhabdus,and the secondary metabolites of Xenorhabdus are found more than 40 kinds.These compounds have a wide range of biological activities,such as anti-bacterial,anti-fungal,anti-virus,anti-cancer,against insects.China's scientific research workers have also carried out excellent results by study these strains in many years.The results showed that the organic phase of the fermentation broth and the fermentation broth have a good prospects for agricultural diseases and insect pests.However,the basic research and application of these bacteria was not systematic,as well as lack of the studies on their secondary metabolites.Based on the preliminary study of our research group,this study isolated and purified the secondary metabolism of the pathogen bacteria SN52 by column chromatography(silica gel column chromatography,gel column chromatography)and reverse-phase semipreparative HPLC.The chemical structures were identified and their biological activities were tested.The results are as follows:1.The preliminary evaluation of the crude extract of the organic phase of the symbiotic bacteria SN52 was carried out.It was found that the crude extract had a strong inhibitory activity on Bacillus cereus and Candida albicans.The crude extracts of the organic phase of the fermentation broth were isolated and purified by chromatographic column chromatography and HPLC to obtain six yellow solid powder monomer compounds and two white solid powder monomer compounds.The structures were identified as four known dithiolopyrrolone derivatives,Xenorhabdin I(compound 3),Xenorhabdin II(compound 4),Xenorhabdin IV(compound 5),and Xenorhabdin V(compound 6),two new ones,6-(E-5-methylhex-2-enamide)-5-oxo-1,2-dithiolo[4,3-b]pyrrole(compound 1),and 6-(E-5-methylhex-2-enamide)-4-methyl-5-oxo-1,2-dithiolo[4,3-b]pyrrole(compound 2)and 4-0-(4-acetamide-?-L-rhamnoside)-benzaldehyde(compound 7),4-0-(4-acetamide-?-L-rhamnoside)-(Z)-N-styrylformamide(compound 8).The antimicrobial activities of dithiolopyrrolone compounds 1-6 was tested by 96-well microtiter plates and found that compounds 3-6 exhibited significant antibacterial activity against Bacillus subtilis,Bacillus cereus and Staphylococcus aureus with IC50 value of 0.8-4.9?g/mL.While compounds 1 and 2 showed moderate inhibitory activity against the three pathogens of Bacillus subtilis,Bacillus cereus and Staphylococcus aureus.In addition,compounds 3-6 were found to have a certain inhibitory activity against Enterococcus pernyi and Candida albicans,and their structural-activity relationship was analyzed.The structure-activity relationships indicated that the antibacterial activities of dithiolopyrrolone derivatives are significant influenced by the N-7 position mutable acyl side chain.In addition,the weak inhibitory activity of compounds 1-4 against Botrytis cinerea may be related to their solubility.3.The rhabduscin analogues compounds isolated from the insect pathogens symbiotic bacteria have a nanomolar inhibitory activity(IC50 value)against the phenoloxidase,that associated with the insect immune system.Compounds 7 and 8 are analogs of rhabduscin,the only difference between them were the disappearance of the isonitrile functional group.The inhibitory activity of hemolymph phenoloxidase of the insect was tested in vitro.The assay results supported the view that the isonitrile group played a key role in the inhibition of phenoloxidase.The above results indicate that the dithiolopyrrolone N-7 acyl group has a strong effect on its antibacterial activity,which provide a template for further development of semisynthetic derivatives with improved activity against the pathogenic bacteria.The discovery of four new compounds enriches the diversity of secondary metabolites of these bacteria.