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It is generally believed that high-quality Bupleurum scorzonerifolium roots possess specific morphological characteristics, being red, robust, and long with strong odor. However, the scientific connotation of these characteristics has not been elucidated. According to the theory of "quality evaluation through morphological identification", we studied the correlations between appearance traits(the RGB value of root surface, root length, root diameter, dry weight, and ratio of phloem to xylem) and content of main chemical components(volatile oils, total saponins, total flavonoids, total polysaccharides, and seven saikosaponins) of B. scorzonerifolium roots. Epson Scanner and ImageJ were used to scan the root samples and measure the appearance traits. Ultraviolet spectrophotometry and HPLC were employed to determine the content of chemical components. The correlation, regression, and cluster analyses were performed to study the correlations between the appearance traits and the content of chemical components. The results showed that the content of volatile oils and saikosaponins were significantly correlated with RGB value, root length, and root diameter, indicating that within a certain range, the roots being redder, longer, and thicker had higher content of volatile oils and saikosaponins. According to the appearance traits and chemical component content, the 14 samples from different producing areas were classified into four grades, and the differences in morphological traits and chemical component content were consistent among different grades. The findings in this study demonstrate that appearance traits(RGB value, root length, and root diameter) can be used to evaluate the quality of B. scorzonerifolium roots. Meanwhile, this study lays a foundation for establishing an objective quality evaluation method for B. scorzonerifolium roots.
Subject(s)
Bupleurum/chemistry , Saponins/analysis , Oleanolic Acid/analysis , Oils, Volatile/analysis , Plant Roots/chemistryABSTRACT
Objective To investigate the relationship between the content of flavonoids and the key enzyme genes expression in different tissues of Bupleurum chinense and B. scorzonerifolium. Methods The roots, stems, leaves, and fruits of B. chinense and B. scorzonerifolium were used as test materials, determination of flavonoids (rutin, quercetin, kaempferol, and isorhamnetin) in different tissues by HPLC, determination of total flavonoids by UV spectrophotometry, the tissues expression of key enzyme genes (IFS, F3H, and DFR) in flavonoids synthesis was determined by real-time quantitative PCR, correlation analysis was performed with SPSS. Results The content of flavonoids in the aerial parts (stems, leaves, and fruits) of B. chinense and B. scorzonerifolium was significantly higher than that in roots, the content of flavonoids was mainly rutin, and the content of rutin in the leaves of B. chinense leaves was up to 106.961 mg/g; The distribution of total flavonoids in B. chinense and B. scorzonerifolium was obviously different, the content was from high to low: leaves ≥ fruit > stem > root; The expression of B. chinense IFS, F3H, and DFR gene in the aerial parts was much higher than that in roots, IFS gene was significantly positive correlated with rutin (P < 0.05), F3H gene was significantly positive correlated with DFR gene (P < 0.05), but the expression of IFS, F3H, and DFR gene in each tissues of B. scorzonerifolium was at lower level. Conclusion The content of flavonoids in different parts of B. chinense and B. scorzonerifolium was consistent with the expression of flavonoids synthesis key enzyme genes, the differential expression of key enzyme genes regulates the synthesis and accumulation of flavonoids in different tissues.
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Objective: To select suitable references genes of Bupleurum scorzonerifolium for tissue expression analyses, and study the tissue expression characteristics of the key enzyme genes of saikosaponins biosynthesis. Methods: Five candidate reference genes including Actin, α-tubulin, β-tubulin, Cyclophilin, and EF-1α were chosen. The stability of these candidate reference genes was investigated by using four softwares (Delta CT, BestKeeper, NormFinder, and GeNorm). The stability of these candidate reference genes was tested and verified by real-time quantitative PCR. Used the stable reference gene, the tissue expression characteristics of the saikosaponins biosynthesis key enzyme genes (HMGR, IPPI, FPS, SS, and β-AS) was analyzed by qRT-PCR. Results: The average expression stability of the five candidate reference genes from high to low was β-tubulin > Cyclophilin > Actin > EF-1α > α-tubulin. Β-tubulin was the most suitable reference gene for tissue expression analysis in B. scorzonerifolium. HMGR expression level was roots > stems and fruits > leaves, IPPI expression level was roots > stems > fruits and leaves, FPS expression level was leaves > roots > stems and fruits, SS expression level was leaves > fruits > roots > stems, β-AS expression level was leaves > roots > fruits > stems. HMGR was significant positive correlated with IPPI, and FPS was significant positive correlated with β-AS (P < 0.05). Conclusion: β-tubulin gene was confirmed as the most suitable reference gene in different tissues of B. scorzonerifolium. It provided a methodological basis for the tissue expression analysis on the functional genes of B. scorzonerifolium. The expression pattern of five key enzyme genes of saikosaponins biosynthesis in different tissues had obvious differentiation, which might be involved in regulating the flow of saikosaponins synthesis and accumulation in various tissues of B. scorzonerifolium.
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Bupleuri Radix, originally appeared in Chinese materia medica (CMM) dating from the Han Dynasty, is a commonly used herb in clinical practice. According to ancient Chinese medical literatures, the morphological description of Bupleuri Radix was vague based on the species-rich and similar patterns, thus it exhibited significantly problems in species identification of Bupleuri Radix which applied in ancient Chinese medicine clinical practice. This paper summarizes important descriptions of morphology and efficacy of Bupleuri Radix from CMM in past dynasties. Research results from modern scholars about identification Bupleuri Radix are summarized to find out which species of Bupleuri Radix in clinic by traditional Chinese physicians. A kind of Bupleurum plants, having consistent efficacy, were used by traditional Chinese physicians before the Tang Dynasty, then Bupleurum scorzonerifolium., B. yinchowense, and Stellariae dichotoma var. lanceolata were used as Bupleuri Radix simultaneously. Stellariae Radix, its efficacy is different from the efficacy of Bupleuri Radix, was distinguished from Bupleuri Radix in the Ming and Qing Dynasties. With the deepening of research of taxonomy, “Bei Chai Hu” (corresponded to B. chinese DC.) and “Nan Chai Hu” (corresponded to B. scorzonerifolium Willd.) first appeared in the Ming Dynasty and the Republic of China era respectively.
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With Bupleurum smithii var. parvifolium and B. scorzonerifolium as test objects, in order to provide a theoretical basis for the introduction and domestication of B. smithii var. parvifolium, the growth and development dynamics of seedlings, biomass accumulation, the content of malonaldehyde(MDA), the activity of antioxidase such as SOD, POD, CAT and APX between them were comparatively analyzed by direct sowing culture in the open field. The results indicated that the morphological index and the biomass accumulation of B. smithii var. parvifolium such as root diameter, root length, plant height and leaf number were inferior to B. scorzonerifolium, the antioxidase SOD and POD activity of B. smithii var. parvifolium was significantly inferior to B. scorzonerifolium (<0.05), the antioxidase CAT and APX activity of B. smithii var. parvifolium was inferior to B. scorzonerifolium but the difference wasn't significant, while MDA content was superior to B. scorzonerifolium(<0.05). Thus, compared with cultivated B. scorzoneri folium, the plant growth velocity of wild B. smithii var. parvifolium was relatively slower and its resistance was relatively weaker after introduction and domestication.
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OBJECTIVE: To explore a set of identification methods of discriminating Bupleurum scorzonerifolium from its adulterant Bupleuru bicaule. METHODS: Methods of morphological comparison of plant samples with specimens, macroscopic identification of the roots, TLC identification of the root extracts, and DNA molecular identification by ITS2 sequence were applied. RESULTS: There were distinctive differences in the morphological, macroscopic, and TLC characteristics between B. chinense and B. scorzomerifolium. The DNA of ITS2 sequence was able to be identified by measuring their mean intraspecific genetic distance (K-2-P distance) which was lower than their mean interspecific genetic distance. In the cluster dendrogram, both of them showed monophyletic property and could be distinguished from other adulterants. CONCLUSION: On the basis of comparison of Bupleurum scorzonerifolium and its adulterant Bupleurum bicaule, the main identification features were summarized by a combination of original plant morphology, macroscopic identification of the roots, TLC identification, and DNA molecular identification methods. DNA identification technology is an important supplement of the traditional identification methods, which may have great potential and application prospect in origin identification of medicinal plants.
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Objective: Bupleuri Radix is a kind of medicinal material derived from many sources, and Bupleurum chinense and B. scorzonerifolium are described in Chinese Pharmacopoeia. In this study, the chemical constituents in Chaige Jieji Decoction (CJD) composed of B. scorzonerifolium or B. chinense were compared to elucidate the influence of different Bupleuri Radix on other components in CJD. Methods: Firstly, an HPLC method for the simultaneous determination of seven constituents (puerarin, albiflorin, paeoniflorin, liquiritin, baicalin, wogonoside, and baicalein) in CJD was developed. Then CJD composed of B. scorzonerifolium or B. chinense was compared by the established method. Results: Good linearity for the seven compounds was obtained, and the average recoveries ranged from 94.1% to 106.6%. The RSD values of all the precision and repeatability of the compounds were less than 2%, and the test sample was stable within 12 h. No significant differences were observed between the two different CJD. Conclusion: The developed HPLC method is simple and repeatable, which is helpful for the quality control of CJD. The effects of different Bupleuri Radix compatibility on other ingredients in CJD and pharmacological activity of CJD deserve further study.
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Objective: To evaluate the antipyretic effect of Bupleurum scorzonerifolium based on GC-MS metabolomics and explore its mechanism. Methods: Rats were randomly divided into control, model, high-, mid-, low-dose B. scorzonerifolium extract (40, 20, and 10 g/kg), and Aspirin (100 mg/kg) positive control groups. The fever rats induced by yeast were administered twice. The body temperature changes of rats were observed at the 5th, 6th, 7th, 8th, 9th, and 10th hours after modeling; Meanwhile, plasma samples were collected at the 10th hour and subjected to GC-MS analysis to find potential biomarkers of fever. Results: Aspirin showed antipyretic effect well. The temperature showed no significant difference compared with the control group 2 h after administration (P > 0.05). Different doses of extract from B. scorzonerifolium showed the antipyretic effect at varying degrees, which presented the obvious dose-dependent antipyretic effect. The high-dose group had a faster onset, and there was a significant difference (P 0.05). Compared with the model group, low-dose group showed no significant differences at each time point. The body temperature showed a large fluctuation, indicating the drugs could interfere with the increase of body temperature but did not reach a significant antipyretic effect. Seventeen potential biomarkers were identified from the loading plot based on the metabolomics and the different doses of B. scorzonerifolium extract could regulate the biomarkers. Conclusion: B. scorzonerifolium has the significant antipyretic effect in a dose-dependent manner and the antipyretic role may be related to the synergism of enzyme inhibition, neurotransmitter, glucose and lipid metabolism, amino acid metabolism, energy metabolism with the multi-targets.
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Composition of the polysaccharide from Bupleurum scorzonerifolium Willd. was studiedby high resolution capillary gas chromatography after hydrolysis and followed by acetylation and esterifica-tion of the hydrolysate. The results showed that it is composed of arabinose, ribose, D-xylose, D-man-nose, D-glucose and D-galactose, with a molar ratio of 0. 106 : 2. 652 : 4. 070: 0. 519 : 0. 930 : 2. 518.
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Object Four regenerated somatic cell hybrids between carrot and Bupleurum scorzonerifolium Willd. proved by chromosome analysis were analyzed at the molecular level. Methods RAPD analysis and 5S rDNA spacer sequence were adopted. Results In the electrophoresis pattern of the PCR products the four somatic cell hybrids had the characteristic bands of two parents and new bands. Conclusion Their hybrid nature was clarified at molecular level. This provides a firm foundation to further analyze the main active components, saikosaponin of somatic cell hybrids, and screen out efficacious hybrid cell lines.
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Objective: To study the effects of different varieties of Radix Bupleuri on the absorbance of Caihu Injection.Methods:The absorbances of redistilled liquids of different varieties of Radix Bupleuri were determind.Results: There was a significant difference in absorbance between B. scorzonerifolium Willd and B. chinense DC.Conclusion:The qualities of Bupleurum preparation can't reach the requirement of legal drug standard due to the variation in crude medicinal materials of Radix Bupleuri.So it is necessary that the crude medicinal materials of Radix Bupleuri in Bupleurum preparation of which absorbance needs to determine should be definitely limited.