Gao Yan, Zhang Tao, Kang Xue-Ping, Han Mei, Yang Li-Min
College of Chinese materials,Jilin Agricultural University Changchun 130118,China.
Yanbian Korean Autonomous Prefecture Academy of Agricultural Sciences Yanbian 133001,China.
Zhongguo Zhong Yao Za Zhi. 2019 Jul;44(13):2768-2776. doi: 10.19540/j.cnki.cjcmm.20190323.103.
The study is aimed to explore the effect of soil moisture content on ginsenoside biosynthesis and explain its mechanism from the perspectives of antioxidant enzyme system and gene expression of key enzymes in the pathway of ginsenoside synthesis. In the study,two years old Panax ginseng was used as the experimental material and three moisture gradient,40% of saturated water content( W1),60%( W2),80%( W3) were set up. The content of 11 monomeric saponins were determined by HPLC. With GAPDH as a reference gene,six key enzymes( HMGR,SS,β-AS,CYP716 A47,CYP716 A52 v2,CYP716 A53 v2) in ginseng saponin synthesis pathway expression were analyzed by fluorescent quantitative PCR and the activities of superoxide dismutase( SOD),peroxidase( POD),catalase( CAT) activity and MDA content were also determined. With the increase of soil water,the content of ginseng saponin and biomass showed an increasing trend. PPD( Rb1,Rc,Rb2,Rd,Rh2,Rb3,Rg3),PPT( Rg1,Re,Rf) ginsenoside,Ro and total ginsenoside reached the maximum value on August 30,were 9.92,5.48,0.63 mg·g-1,respectively. During the whole regulation period,the antioxidant activity of W3 was greater than that of W1,and the MDA content was less than that of W1. At W3,expression levels of β-AS,CYP716 A47 and CYP716 A53 v2 showed an increasing trend,while HMGR and SS genes showed relatively stable expression levels under various water conditions. According to the correlation analysis,HMGR and SS genes in the W3 treatment group were significantly positively correlated with PPD,PPT ginsenoside and Ro,CYP716 A52 v2 gene was significantly positively correlated with Ro,and CYP716 A47 gene was significantly positively correlated with PPD ginsenoside. There was a significant positive correlation between β-AS gene and PPD ginsenoside in W1 and W2 treatment. Therefore,W3 is the optimum moisture content,ginseng total saponins and monomer saponin content is the highest,the gene closely correlation with content of saponins and more conducive to the accumulation of ginsenosides.
本研究旨在探讨土壤水分含量对人参皂苷生物合成的影响，并从抗氧化酶系统和人参皂苷合成途径中关键酶的基因表达角度解释其机制。本研究以两年生人参为实验材料，设置了三个水分梯度，即饱和含水量的40%（W1）、60%（W2）、80%（W3）。采用高效液相色谱法测定11种单体皂苷的含量。以甘油醛-3-磷酸脱氢酶（GAPDH）为内参基因，通过荧光定量PCR分析人参皂苷合成途径中6种关键酶（3-羟基-3-甲基戊二酰辅酶A还原酶（HMGR）、鲨烯合酶（SS）、β-香树素合酶（β-AS）、细胞色素P450 716A47（CYP716A47）、细胞色素P450 716A52 v2（CYP716A52 v2）、细胞色素P450 716A53 v2（CYP716A53 v2））的表达，并测定超氧化物歧化酶（SOD）、过氧化物酶（POD）、过氧化氢酶（CAT）活性及丙二醛（MDA）含量。随着土壤水分增加，人参皂苷含量和生物量呈上升趋势。原人参二醇（PPD，包括Rb1、Rc、Rb2、Rd、Rh2、Rb3、Rg3）、原人参三醇（PPT，包括Rg1、Re、Rf）型人参皂苷、Ro及总人参皂苷于8月30日达到最大值，分别为9.92、5.48、0.63 mg·g-1。在整个调控期内，W3的抗氧化活性大于W1，MDA含量小于W1。在W3条件下，β-AS、CYP716A47和CYP716A53 v2的表达水平呈上升趋势，而HMGR和SS基因在不同水分条件下表达水平相对稳定。相关性分析表明，W3处理组中HMGR和SS基因与PPD、PPT型人参皂苷及Ro显著正相关，CYP716A52 v2基因与Ro显著正相关，CYP716A47基因与PPD型人参皂苷显著正相关。W1和W2处理中β-AS基因与PPD型人参皂苷显著正相关。因此，W3为最佳含水量，人参总皂苷和单体皂苷含量最高，与皂苷含量密切相关的基因较多，更有利于人参皂苷的积累。
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