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Objective: In order to identify the function of the mevalonate kinase (MK) which is a key enzyme of the mevalonate pathway (MVA) in Swertia mussotii, and to improve the study of MVA in S. mussotii. Methods: According to the SmMK gene sequence of transcriptome of S. mussotii, the specific primers were designed, the cDNA complete sequences was obtained by RT-PCR and the sequence was analyzed using bioinformatics. Prokaryotic expression vector MBP-SmMK was constructed and transformed into Escherichia coli Rosetta (DE3) for expression. Results: The results showed that SmMK cDNA complete sequences had a length of 1 164 bp encoding 387 amino acid residues. The SmMK protein shared high identity with other MK proteins of plants. And the protein signal peptide, transmembrane region, location, secondary, and tertiary structures were analyzed and forecasted. The SDS-PAGE results showed that the expressed proteins were consistent with the anticipated size, which was 40 970. Conclusion: This work will provide a foundation for research the SmMK protein functional and study MCA in S. mussotii. At the same time, it will supply the basis to improve the production of the isoprenoids.
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Objective: To clone the geranyl pyrophosphate synthase gene from Swertia mussotii (SmGPPS), analyze the bioinformation of SmGPPS, and perform the gene expression. Methods: According to the SmGPPS gene sequence of transcriptome of S. mussotii, the specific primers were designed, the cDNA complete sequences was obtained by RT-PCR and the sequence was analyzed using bioinformatics. Prokaryotic expression vector pET-28a-SmGPPS was constructed and transformed into Escherichia coli BL-21 (DE3) for expression under 37℃ and induced by 1 mmol/L IPTG. The relative expression of gene SmGPPS in the leaf, stem, and flower of S. mussotii was also studied. Results: The results showed that SmGPPS cDNA complete sequences had a length of 1 119 bp encoding 372 amino acid residues. And the protein secondary and tertiary structures were analyzed and forecasted. The SmGPPS protein shared high identity with other GPPS proteins of plants. The SDS-PAGE results showed that the expressed proteins were consistent with the anticipated size. Relative RT-PCR analysis indicated that SmGPPS showed the highest transcript abundance in the leaf. Conclusion: This work will provide a foundation for further functional research of SmGPPS protein and increasing the product of iridoid compound by genetic engineering in S. mussotii.
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The resource of rare and endangered medicinal plant Swertia mussotii Franch. in Tibet, Qinghai and Sichuan province were surveyed by ways of documents, interview, quadrat and market investigation. The results indi-cated that Swertia mussotii Franch. mainly distributed in Zuogong and Mangkang of Tibet, Yushu of Qinghai province, Shiqu, Daofu, Kangding, Maerkang, Jinchuan and Xiaojin of Sichuan province. According to the height above sea level, the distribution altitude was from 2 300 m (Kangding of Sichuan province) to 3 900 m (Mangkang of Tibet). There are distributions of Swertia mussotii Franch. within the scope of 2 600 m. The illumination, water, soil, temperature and altitude had significant influence on the distribution, growth and reserve of Swertia mussotii Franch. from different angles. In recent years, there was huge increase of market requirement in Swertia mussotii Franch. which were used in Tibetan medicine Zangyinc he n. Excess collection was the primary cause of rapid decreasing in resource of Swertia mussotii Franch.. It was suggested to strengthen the management of rational collection, as well as to accelerate the development of cultivation and production.
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OBJECTIVE: To study the quality control method of Swertia mussotii Franch. METHODS: A high performance liquid chromatographic method was developed to establish the fingerprint of Swertia mussotii Franch. Nineteen samples from various batches were analyzed, furthermore, PCA were used to differentiate and evaluate the whole fingerprints. Some characteristic peaks were i-dentified preliminarily with HPLC-MS method. RESULTS: Eleven peaks were identified by MS. Swertiamarin and swertianolin were the key components for quality control. CONCLUSION: The method provides an academic reference for controlling the quality of Swertia mussotii Franch.
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Objective: To study the chemical constituents of Swertia mussotii. Methods: Various chromatographic methods were employed to isolate the compounds and their structures were established by spectroscopic analysis. Results: Nineteen compounds were isolated from 75% ethanol extract of S. mussotii and identified as 1-hydroxyl-3, 4, 7, 8-tetramethoxyxanthone (1), 1, 7- dihydroxyl-3-methoxyxanthone (2), 1, 3, 7-trihydroxyxanthone (3), 1, 3, 7, 8-tetrahydroxyxanthone (4), 1, 3, 8-trihydroxyl-7- methoxyxanthone (5), 1, 3-dihyfroxyl-7, 8-dimethoxyxanthone (6), 1, 5, 8-trihydroxyl-3, 4-dimethoxyxanthone (7), 1-hydroxyl-3, 4, 5, 8-tetramethoxyxanthone (8), 1-hydroxyl-3, 5, 8-trimethoxyxanthone (9), (S)-(+)-gentiolactone (10a), (R)-(-)-gentiolactone (10b), 1, 8-dihydroxyl-3, 7-dimethoxyxanthone (11), 1-hydroxyl-3, 7, 8-trimethoxyxanthone (12), 1, 7-dihydroxyl-3, 8-dimethoxyxanthone (13), 1, 7, 8-trihydroxyl-3-methoxyxanthone (14), 1, 3, 5, 8-tetrahydroxyxanthone (15), 1, 7-dihydroxyl-3, 4, 8-trimethoxyxanthone (16), mangiferin (17), and oleanolic acid (18), respectively. Conclusion: Compounds 1-9 are firstly isolated from S. mussotii. Compounds 10a and 10b are firstly isolated from the plants in Swertia L.
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Objective To observe the therapeutic effect of Swertia mussotii Franch is extracted by alcohol on immunological liver injury in mice,and further evaluate its immun-regulating effect in decreasing aminotransferase and protecting heptocytes. Methods BCG vaccine and lipopolysaccharide were intravenously injected to establish liver injury model.The effects of Swertia mussotii Franch on liver injury were contrasted. Results The high-,medium-and low-dose Swertia mussotii Franch groups which had been administered for 10 consecutive days,had significantly inhibited serum ALT and AST acticities of immunological liver injury;Swertia mussotii Franch could effectively prevent immunological liver injury by BCG+LPS. Conclusion Swertia mussotii Franch can protect mice from immunological liver injury and protect the function of liver.
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Objective To study the separation and purification technology of iridoid glycosides and xanthones from Swertia mussotii by macroporous resin.Methods The dynamic adsorption and desorption characteristics of macroporous adsorption resins HPD-300,HPD-400,HPD-600,AB-8,DM-301,and D-101-Ⅰ for swertiamarin and swertianolin were investigated.On the base of the investigation,the better macroporous adsorption resin was chosen to finish the experiments.Results The adsorption and elution characteristics of HPD-300 used to separate and purify iridoid glycosides and xanthones were very good.In the course of adsorption,the amount of used adsorption is 0.9 g dried medicinal herb/mL resin,with about 8 pH value of adsorption solution,and the volumetric rate is 2 BV/h; In the course of elution,the resin column chromatography was eluted gradiently with 7 BV 20% and 5 BV 70% EtOH after removing impurities with water.The elution rates were both more than 90%,and the contents of iridoid glycosides and xanthones were both up to 50% in the two dried fractions of 20% and 70% ethanolic elutions.Conclusion HPD-300 is an ideal resin with the best enrichment for separating and purifying both iridoid glucosioles and xanthanoes simultanously.