纳米银胁迫对甘西鼠尾草毛状根生长及丹参酮积累的影响

Effects of Silver Nanoparticles Stress on the Growth and Tanshinone Accumulation in Salvia przewalskii Hairy Root Cultures

目的:采用穿心莲水提液制备纳米银颗粒(AgNPs)处理甘西鼠尾草(Salvia przewalskii)毛状根,探讨AgNPs胁迫对毛状根生长、丹参酮含量和丹参酮生物合成途径关键酶基因表达水平的影响,探寻促进丹参酮生物合成与代谢的新方法。方法:利用不同质量浓度的AgNPs(0 mg/L、50 mg/L、75 mg/L、100 mg/L)处理甘西鼠尾草毛状根,测定毛状根生物量;利用HPLC法测定3种丹参酮类(隐丹参酮、丹参酮Ⅰ、丹参酮ⅡA)活性成分含量;采用qRT-PCR检测不同处理时间丹参酮生物合成途径关键酶基因HMGR、DXR、CPS相对表达水平。结果:甘西鼠尾草毛状根鲜重随AgNPs处理浓度的升高逐渐降低,各处理组干重相比对照组无显著变化;丹参酮类成分含量随AgNPs浓度增加持续升高,100 mg/L AgNPs处理能使隐丹参酮、丹参酮Ⅰ、丹参酮ⅡA大量积累;与对照组相比,100 mg/L AgNPs显著上调丹参酮合成途径HMGR、DXR、CPS基因的表达,在处理24 h时分别达到最高水平。结论:穿心莲水提液制备纳米银颗粒对甘西鼠尾草毛状根生长具有一定的抑制作用,高浓度AgNPs胁迫显著促进毛状根中丹参酮的生物合成与代谢,通过调控丹参酮合成相关酶基因的表达水平,导致丹参酮类活性成分的大量积累。

Objective:In this study,silver nanoparticles(AgNPs)prepared by Andrographis paniculata water extracts was applied to treat the hairy roots of Salvia przewalskii,the growth,accumulation of tanshinones compounds,and the expression levels of tanshinone biosynthetic-related genes in the hairy root under AgNPs stress were investigated. Finally,explore new ways to promote the biosynthesis and metabolism of tanshinone. Methods:A series mass concentration of AgNPs(0 mg/L,50 mg/L,75 mg/L,and100 mg/L)were used to treat S. przewalskii hairy roots,the fresh and dry weight were determined,respectively. Using high performance liquid chromatography method to determine the contents of cryptotanshinone,tanshinone Ⅰ,and tanshinone ⅡA;real-time quantitative PCR was applied to detect the relative expression levels of tanshinone biosynthetic-related enzyme genes. Results:The fresh weight of S. przewa relative lskii hairy roots decreased gradually by the AgNPs concentration increasing,while the dry weight of each treatment did not change significantly compared with the control group. The content of tanshinone components increased gradually with the elevated AgNPs concentration,cryptotanshinone,tanshinone Ⅰ and tanshinone ⅡA accumulated much higher treated by 100 mg/L of AgNPs. Moreover,the expression levels of 3-hydroxy-3-methyl glutaryl-CoA reductase,1-deoxy-D-xylulose-5-phosphate reductoisomerase,and copalyl diphosphate synthase genes reached the highest levels when treated with 100 mg/L of AgNPs for 24 h,respectively. Conclusion:A. paniculata extracts prepared silver nanoparticles showed a significant inhibitory effect on the growth of the hairy roots,high concentration of AgNPs could stimulate the biosynthetic-related gene expression and promote the accumulation of tanshinone components in S.przewalskii hairy roots significantly.