Tissue culture of safflower and analysis of secondary metabolites in suspension cells / 中国中药杂志

Safflower(Carthamus tinctorius), a valuable traditional Chinese medicinal plant, has attracted much attention in recent years. This study established a stable tissue culture system of safflower and analyzed the chromatogram of its secondary metabolites, providing high-quality experimental materials for further research on natural products in safflower. The calluses were established from the safflower seeds germinated in a sterile environment, and then they were differentiated into the aseptic seedlings, or cultured to obtain suspension cells in liquid medium. The ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry(UPLC-Q-TOF-MS), Progenesis QI, and principal component analysis(PCA) were used to detect and analyze the secondary metabolites in the suspension cells before and after induction with different elicitors(methyl jasmonate, silver nitrate, salicylic acid and yeast extract). A total of 23 secondary metabolites including flavonoids, phenylpropanoids, alkaloids, fatty acids and aromatic glycosides were detected in safflower suspension cells. In response to the four elicitors, 11 compounds showed increased or decreased relative content. The results indicate that different elicitors have various effects on the accumulation of secondary metabolites in safflower suspension cells, and yeast extract shows more obvious positive induction. Therefore, different elicitors may play a role in the expression of related genes in the biosynthetic pathway of specific secondary metabolites. The results facilitate the discovery of targeted elicitors and the large-scale production of valuable secondary metabolites in the future.

Discussion on research idea of quality marker of Salvia miltiorrhiza based on biosynthetic pathway of tanshinone compounds / 中国中药杂志

Based on the theory of Q-marker, the hairy root of Salvia miltiorrhiza and S. miltiorrhiza in many provinces were studied. The relative expressions of SmCPS, SmKSL and CYP76AH1 genes in hairy roots were detected by real-time fluorescence quantitative PCR and the contents of tanshinoneⅡ_A, cryptotanshinone, tanshinoneⅠ, 1,2-dihydrotanshinone, ferruginol and miltiradiene were detected by UPLC and GC-MS, respectively. Statistical analysis shows as fllows: in the hairy root of S. miltiorrhiza, the content of miltiradiene and ferruginol is positively correlated with the content of tanshinone compounds in the downstream, and the relative expression of important genes in the biosynthetic pathway of tanshinone can reflect the content of tanshinone compounds to a certain extent; in many provinces of S. miltiorrhiza, the content of ferruginol and tanshinone compounds can also be found that there is a positive correlation between the contents. Based on the biosynthetic pathway of tanshinone compounds, which is a special index component in S. miltiorrhiza, this study focused on the important relationship between the upstream gene, the middle intermediate compound and the downstream tanshinone compound content of the biosynthetic pathway, and explored the possible research ideas of improving the quality marker system of S. miltiorrhiza, and then provided the possible research ideas for understanding and studying the quality marker of traditional Chinese medicine from the biosynthetic pathway.

Effect on biosynthesis of terpenoid active components through RNA interference with MCT gene in Tripterygium wilfordii / 药学学报

MCT is an important key enzyme in the terpenoid biosynthesis in MEP pathway. In this study, Gateway technology was used to construct RNAi vector of TwMCT, and a vector fragment with a size of 484 bp was obtained. The TwMCT RNAi vector was transferred into the suspension cells of Tripterygium wilfordii by gene gun. Accumulation of terpenoids was assayed by UPLC, and the result showed that the content of triptolide and celastrol in cells decreased by 23.4% and 42.8%, respectively, compared with the control group pK7GWIWG2D. Moreover, the gene expression of TwMCT and major genes in terpenoid biosynthesis pathway was detected by qRT-PCR, which demonstrated that the expression of TwMCT reduced by 29.2% relative to that of the control group pK7GWIWG2D, and the relative expression of TwDXR, TwGGPS, TwHMGR and TwHMGS diminished by 36.3%, 31.3%, 62.2%, and 29.1%, respectively, but the expression of TwDXS was up-regulated by 114.2%, and there was no significant change in TwFPS. Thus, it was verified in vivo that interference with TwMCT expression significantly inhibited the accumulation of triptolide and celastrol in Tripterygium wilfordii, laying a foundation for further exploring the regulation mechanism of MCT gene on the terpenoid biosynthesis in Tripterygium wilfordii.