[Objective]To systematically isolate and purify the polysaccharide from the mycelium of Streptomyces rochei D74(SRP),elucidate its fine structure,and evaluate the effect of the purified polysaccharide fraction on the ...[Objective]To systematically isolate and purify the polysaccharide from the mycelium of Streptomyces rochei D74(SRP),elucidate its fine structure,and evaluate the effect of the purified polysaccharide fraction on the growth of Salvia miltiorrhiza hairy roots and the biosynthesis of tanshinones,along with the underlying mechanism.[Methods]The crude polysaccharide was extracted using hot water,which was followed by ethanol precipitation and deproteinization via the Sevag method.Further purification was performed using DEAE-52 anionexchange chromatography and Sephadex G-100 gel filtration chromatography.The physicochemical properties and structural features of the main active fraction,SRP-W-2,were systematically characterized by Fourier transform infrared spectroscopy(FTIR),high performance liquid chromatography-mass spectrometry(HPLC-MS),and nuclear magnetic resonance(NMR).The effects of SRP-W-2 on hairy root growth and the biosynthesis of tanshinones were assessed by measuring biomass,tanshinone content,and the expression levels of key biosynthetic genes.[Results]SRP-W-2 was obtained with a yield of 2.41%.It was primarily composed of glucose and galactose at a molar ratio of 12.53:1.Structural analysis revealed that the backbone of SRP-W-2 consisted of→4)-α-D-Glcp-(1→and→4)-α-D-Galp-(1→residues,with branching points at→4,6)-α-D-Glcp-(1→and→4,6)-α-D-Galp-(1→.The side chain was identified asα-D-Glcp-(1→4)-α-DGlcp-(1→.Bioactivity assays demonstrated that SRP-W-2 significantly enhanced both the biomass of S.miltiorrhiza hairy roots and the accumulation of tanshinones.After 15 d of treatment with 50 mg/L SRP-W-2,the dry weight of the hairy roots increased by 37.52%.Meanwhile,the content of cryptotanshinone(CT),dihydrotanshinone I(DT-I),tanshinone I(T-I),and tanshinone IIA(TIIA)was increased by 19.0-fold,6.4-fold,2.8-fold,and 4.8-fold,respectively.Gene expression analysis further indicated that SRP-W-2 up-regulated key genes involved in the tanshinone biosynthetic pathway,including HMGR,DXS,DXR,and GGPPS.[Conclusion]The polysaccharide fraction SRP-W-2 from S.rochei D74 simultaneously promoted the growth of S.miltiorrhiza hairy roots and the biosynthesis of tanshinones,demonstrating its potential as an effective elicitor.This study provided a new strategy for the utilization and development of S.miltiorrhiza resources.展开更多
文摘[Objective]To systematically isolate and purify the polysaccharide from the mycelium of Streptomyces rochei D74(SRP),elucidate its fine structure,and evaluate the effect of the purified polysaccharide fraction on the growth of Salvia miltiorrhiza hairy roots and the biosynthesis of tanshinones,along with the underlying mechanism.[Methods]The crude polysaccharide was extracted using hot water,which was followed by ethanol precipitation and deproteinization via the Sevag method.Further purification was performed using DEAE-52 anionexchange chromatography and Sephadex G-100 gel filtration chromatography.The physicochemical properties and structural features of the main active fraction,SRP-W-2,were systematically characterized by Fourier transform infrared spectroscopy(FTIR),high performance liquid chromatography-mass spectrometry(HPLC-MS),and nuclear magnetic resonance(NMR).The effects of SRP-W-2 on hairy root growth and the biosynthesis of tanshinones were assessed by measuring biomass,tanshinone content,and the expression levels of key biosynthetic genes.[Results]SRP-W-2 was obtained with a yield of 2.41%.It was primarily composed of glucose and galactose at a molar ratio of 12.53:1.Structural analysis revealed that the backbone of SRP-W-2 consisted of→4)-α-D-Glcp-(1→and→4)-α-D-Galp-(1→residues,with branching points at→4,6)-α-D-Glcp-(1→and→4,6)-α-D-Galp-(1→.The side chain was identified asα-D-Glcp-(1→4)-α-DGlcp-(1→.Bioactivity assays demonstrated that SRP-W-2 significantly enhanced both the biomass of S.miltiorrhiza hairy roots and the accumulation of tanshinones.After 15 d of treatment with 50 mg/L SRP-W-2,the dry weight of the hairy roots increased by 37.52%.Meanwhile,the content of cryptotanshinone(CT),dihydrotanshinone I(DT-I),tanshinone I(T-I),and tanshinone IIA(TIIA)was increased by 19.0-fold,6.4-fold,2.8-fold,and 4.8-fold,respectively.Gene expression analysis further indicated that SRP-W-2 up-regulated key genes involved in the tanshinone biosynthetic pathway,including HMGR,DXS,DXR,and GGPPS.[Conclusion]The polysaccharide fraction SRP-W-2 from S.rochei D74 simultaneously promoted the growth of S.miltiorrhiza hairy roots and the biosynthesis of tanshinones,demonstrating its potential as an effective elicitor.This study provided a new strategy for the utilization and development of S.miltiorrhiza resources.
