Study On Biosynthesis Of Swainsonine In Fungal Endophyte,Undifilum Oxytropis, From Locoweed

Locoweeds are toxic plants of the genera Astragalus and Oxytropis containing swainsonine. Swainsonine is the principal agent responsible for locoism in animals. Recently, the fungal endophytes are isolated frequently from locoweeds, Undifilum spp, are believed that responsible for the production of swanisonine in locoweeds. If the swainsonine production in Undifilum spp is inhibited, the locoweed’s toxicity would be reduced or eliminate. Currently, the mechanism was few known about the biosynthesis of swainsonine in Locoweed’s endophytic fungi. The research about influence of different factors on swainsonine production will be benefit for building the important basis for further to elucidate the swainsonine biosynthetic pathways and molecular regulation mechanism.The principal purpose of this paper was to isolate and identify the swainsonine-producing fungal endophytes, U. oxytropis, from locoweeds in Ningxia. The influence of different factors on swainsonine production was estimated in U.oxytropis. The protoplasts of U. oxytropis was developed and regenerated. This study will provide a important science basis for further to elucidate the swainsonine biosynthetic pathways and build a non-toxic fungus.1. The results indicated that five isolates are harvested from Oxytropis ochrocephala Bunge in Ningxia. Only one strain was determined to synthesize swainsonine. The strain could produce abundant conidia or conidiospores after induced culture. The morphology and genetic evolution relationship of the strain was closely related to the Undifilum oxytropis. Therefore, the strain was identified as U. oxytropis in this study.2. In this research, the influence of pH, PEG, L-pipecolic acid, L-lysine ando-ketoglutaric acid on the growth and swainsonine production of U. oxytropis was estimated. The results showed that is no significant difference about growth of different pH treatment of U. oxytropis groups. However, low pH (pH4.5) will inhibit the synthesis of swainsonine. PEG promoted the growth of U.oxytropis, but inhibited the synthesis of swainsonine. L-pipecolic acid could increase significantly the mycelial dry mass. When the initial concentration of L-pipecolic acid was10-3mol/L and10-2mol/L, the production of swainsonine in U. oxytropis was inhibited. However, when the initial concentration of L-pipecolic acid was10-4mol/L, the production of swainsonine in U. oxytropis was increased significantly. When the initial concentration of L-lysine was10-3mol/L, the production of swainsonine in U.oxytropis was increased significantly. However, when the initial concentration of L-pipecolic acid was10-1mol/L,10-2mol/L and10-4mol/L, the production of swainsonine in U. oxytropis was inhibited significantly. When the initial concentration ofa-ketoglutaric acid was10-1mol/L,10-2mol/L and10-3mol/L, the growth of production of swainsonine in U. oxytropis was inhibited significantly. The above results indicated that L-pipecolic acid, L-lysine and a-ketoglutaric acid could be important substrates in the biosynthesis of swainsonine in U. oxytropis.3.The highest production rate of protoplast was obtained when NX-FEL001was cultured in the solid medium for15d at22℃, and then the mycelia were digested by a compound enzyme consisting of1mol/L sorbitol,1%cellulase,1%snailase and0.1%lywallzyme for3h at35℃,80r/min, W/V 1:10. As for protoplast regeneration, a digestion time of about3h was preferable. The highest regeneration rate8.5%was achieved when the protoplasts were regenerated in the YCM medium for15d.The regeneration of hyphae still had the ability to biosynthesis swainsonine.