Abstract
BackgroundThe dried root of Polygala tenuifolia, named Radix Polygalae, is a well-known traditional Chinese medicine. Triterpenoid saponins are some of the most important components of Radix Polygalae extracts and are widely studied because of their valuable pharmacological properties. However, the relationship between gene expression and triterpenoid saponin biosynthesis in P. tenuifolia is unclear.Methodology/FindingsIn this study, ultra-performance liquid chromatography (UPLC) coupled with quadrupole time-of-flight mass spectrometry (Q-TOF MS)-based metabolomic analysis was performed to identify and quantify the different chemical constituents of the roots, stems, leaves, and seeds of P. tenuifolia. A total of 22 marker compounds (VIP>1) were explored, and significant differences in all 7 triterpenoid saponins among the different tissues were found. We also observed an efficient reference gene GAPDH for different tissues in this plant and determined the expression level of some genes in the triterpenoid saponin biosynthetic pathway. Results showed that MVA pathway has more important functions in the triterpenoid saponin biosynthesis of P. tenuifolia. The expression levels of squalene synthase (SQS), squalene monooxygenase (SQE), and beta-amyrin synthase (β-AS) were highly correlated with the peak area intensity of triterpenoid saponins compared with data from UPLC/Q-TOF MS-based metabolomic analysis.Conclusions/SignificanceThis finding suggested that a combination of UPLC/Q-TOF MS-based metabolomics and gene expression analysis can effectively elucidate the mechanism of triterpenoid saponin biosynthesis and can provide useful information on gene discovery. These findings can serve as a reference for using the overexpression of genes encoding for SQS, SQE, and/or β-AS to increase the triterpenoid saponin production of P. tenuifolia.
Highlights
Radix Polygalae, which is the root of Polygala tenuifolia Willd. or P. sibirica L
The observation of [M-H-Glc-H2O-CO2-Fuc-Rha-Xyl-Api-HMG-CH2O]2 at m/z 455.3153 (Y1) and [M-H-Glc-H2O-CO2-Fuc-Rha-XylApi-HMG]2 (Y2) at m/z 425.3045 in the MS/MS spectrum can be considered as the diagnostic ions for this type of triterpene saponins, and the fragment ions at m/z 1235.5663, corresponding to the losses of 144 Da proved the occurrence of 3-hydroxy-3methyl-5-pentanoic acid ester (X1)
Stable genes identified in geNorm were confirmed by NormFinder, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was chosen as the reference gene in the different tissues of P. tenuifolia
Summary
Radix Polygalae, which is the root of Polygala tenuifolia Willd. or P. sibirica L. Radix Polygalae, which is the root of Polygala tenuifolia Willd. Triterpenoid saponins are documented as an important group of active components. The structures of the triterpenoid saponins in P. tenuifolia are rather complex [7]. The triterpenoid saponins have a triterpene presenegenin aglycone as basic structure, and the slight differences among these saponins are a result of various substitutes including saccharides and/or acyl groups linked to C28. Considering the complicated isolation and complete structural characterization of triterpenoid saponins, more methods need to be established to analyze the chemical constituents of triterpenoid saponins in P. tenuifolia. The dried root of Polygala tenuifolia, named Radix Polygalae, is a well-known traditional Chinese medicine. Triterpenoid saponins are some of the most important components of Radix Polygalae extracts and are widely studied because of their valuable pharmacological properties. The relationship between gene expression and triterpenoid saponin biosynthesis in P. tenuifolia is unclear
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