Studies On The Metabolite And Antifungal Activity Of Xenorhabdus Budapestensis SN19 From Entomopathogenic Nematodes

The Symbiotic Bacteria of entomopathogenic nematode is a vital microorganism. It can produce various secondary metabolites with biological activities. The Symbiotic Bacteria of entomopathogenic nematode has application value in the fields of medical science and agricultural biological control and so on. Xenorhabdus budapestensis SN19 bacterial strain was used as experimental material which is saved in the microbial secondary metabolite laboratory, College of plant protection, Shenyang Agricultural University. Then, studing the inhibiting effects of Xenorhabdus budapestensis SN19 bacterial strain on plant pathogenic fungus. And the structures of the compounds which possess biological activities were isolated and identified.SN19 bacterial strain is belonged to Xenorhabdus budapestensis through 16S rRNA gene sequences. The fermentation crude extract of SN19 bacterial strain was obtained by utilizing some research methods of macroporous resin, methanol immersion and dichloromethane extraction. Then, Botrytis cinerea, Phytophthora capsici and Pyricularia oryzae as target bacterium to analysis the biological activity of SN19 bacterial strain through mycelium growth inhibition method. Effective components of antifungal activity can be separated and isolated from fermentation crude extract of SN19 bacterial strain through solid phase extraction, column chromatography separation, gel column chromatography and HPLC. And the chemical construction of the active components can be analyzed and confirmed by ’H-NMR,13C-NMR, MS and Marfey’s method.Quantitative liquid fermentation culture (24 L) is done to Xenorhabdus budapestensis SN19 bacterial strain by M medium. The fermentation products can be extracted and adsorbed by the macroporous resin. Fermentation can be obtained by solvent extraction. A type of developing solvent is consisted of two kinds of petroleum ether:acetone (=7:3), chloroform: methanol (=85:15), dichloromethane:methanol (=85:15), dichloromethane:methanol (=92:8). With the different blend solvents above-mentioned, a thin layer chromatography (TLC) is act on the fermentation to filter the optimal scale as the final solvent. According to the result of TLC, methanol, dichloromethane system was used for gradient elution. Basing on the result of TLC, the fermentation will be in polarity extracted into 4 groups (from A to D). Using of solid phase extraction, column chromatography separation, gel column chromatography and HPLC to separate the fermentation. In the end, three compounds were separated and purified. With the spectra method such as 1H-NMR,13C-NMR, MS, Marfey’s method and then referred to the relative literature, the construction of the compounds were identified. The result shows the three compounds are compound 1 (xenematides C)、compound 2 (xenematides B) and compound 3. Compound 3 was an open chain structure and it is a new compound by searching SCI Finder. Compound 3 was named xenematides E.Botrytis cinerea, Phytophthora capsici and Pyricularia oryzae are target bacterium. Through mycelium growth inhibition method, the result demonstrats that compound 1 (xenematides C) has the significantly antifungal activity to Botrytis cinerea (EC50=22.71 mg/L) while it is weakly active or even inactive to Phytophthora capsici and Pyricularia oryzae.According to the above results, shows that fermentation crude extract of SN19 bacterial strain has significantly antifungal activity to Botrytis cinerea. Three lipopeptide monomeric compounds were isolated and identified from fermentation crude extract of SN19 bacterial strain. The result shows that compound 1 (xenematides C) and compound 2 (xenematides B) are known compounds. However, the research on inhibiting activity of compound 1 (xenematides C) to Botrytis cinerea is the first time. Compound 3 is an open chain structure and it is a new compound, which was named xenematides E.