1.Genotype, environment and their interactions of functional components in Rheum palmatum.
Lei MENG ; Hui-Juan HU ; Tong SHANG ; Wen-Cang TIAN ; Ming-Ming ZHANG ; Yong-Jie LI ; Li LI ; Sheng-Li WEI
China Journal of Chinese Materia Medica 2018;43(12):2495-2502
Eight kinds of provenance of Rheum palmatum collected from 4 provinces Sichuan, Ningxia, Gansu, Shannxi as test materials, which were transplanted under 3 different environments by using complete randomized block design with three replicates. The contents of the chemical components was determined by HPLC. This study aimed at analyzing the effect of genotype, environment and their interactions on the 4 kinds of functional components (phenolic acids, bianthrone, free anthraquinones and combined anthraquinones) in 14 kinds of active components of R. palmatum, in order to provide a theoretical basis for the selection of cultivated R. palmatum in high quality producing area and excellent provenance. The functional components of R. palmatum were influenced by genotype and environment. The content of phenolic acids was mainly influenced by environment, and the other three kinds of functional components were affected by environment and their interactions. The proportion of environment was larger. The cultivation quality of R. palmatum should give priority to environment, then choose a provenance. Sichuan may be beneficial in accumulation of free anthraquinones in R. palmatum, Gansu may facilitate the binding of combined anthraquinone, phenolic acids and bianthrone content. Preliminary inference based on the content and proportion of efficacy components, P2 could be potential special medicinal germplasm that have function of heat-clearing and detoxifying drugs. P6 could be potential special medicinal germplasm that activate blood circulation to dissipate blood stasis. P7 and P1 could all be potential specialmedicinal germplasms that exist diarrhea attack characters. The results of this study have important guiding significance for the production of rhubarb precision medicinal materials.
2.Study on geographical variation of medicinal ingredients contents in Scutellaria baicalensis from different provenances.
Yong-Jie LI ; Rong-Xiu LIU ; Wen-Cang TIAN ; Xue-Sen WANG ; Zhi-Yan HOU ; Lei MENG ; Guo-Hu SONG ; Xue-Wen ZHANG ; Sheng-Li WEI
China Journal of Chinese Materia Medica 2018;43(13):2664-2669
In order to investigate the genetic difference on medicinal components of Scutellaria baicalensis from different provenances on the genetic difference, the S. baicalensis provenance tests were arranged by randomized block design.Excavating the crude drugs that have been growing for three years, with the same drying process, the content of baicalin, baicalein, wogonoside, wogonin and laminarin A in S. baicalensis were detected by HPLC, and then the data were analyzed. The results indicated that the content of baicalin in different provenances of S. baicalensis was significantly different (<0.05), while the variation of baicalein reached extremely significant level (<0.01). Cluster analysis showed that if the distance was divided by 5.0, the provenances in Chengde, Hebei province were divided into two independent populations, while the other two populations had large geographic spans. The results show that the significant geographical variations exist in the content of medicinal components in S. baicalensis. The study laid a theoretical foundation of provenance selection of S. baicalensis.
3.Combinatorial mutation on the -glycosidase specific to 7--xylosyltaxanes and increasing the mutated enzyme production by engineering the recombinant yeast.
Jing-Jing CHEN ; Xiao LIANG ; Fen WANG ; Yan-Hua WEN ; Tian-Jiao CHEN ; Wan-Cang LIU ; Ting GONG ; Jin-Ling YANG ; Ping ZHU
Acta Pharmaceutica Sinica B 2019;9(3):626-638
Taxol is a "blockbuster" antitumor drug produced by species with extremely low amount, while its analogue 7--xylosyl-10-deacetyltaxol is generally much higher in the plants. Both the fungal enzymes LXYL-P1-1 and LXYL-P1-2 can convert 7--xylosyl-10-deacetyltaxol into 10-deacetyltaxol for Taxol semi-synthesis. Of them, LXYL-P1-2 is twice more active than LXYL-P1-1, but there are only 11 significantly different amino acids in terms of the polarity and acidic-basic properties between them. In this study, single and multiple site-directed mutations at the 11 sites from LXYL-P1-1 to LXYL-P1-2 were performed to define the amino acids with upward bias in activities and to acquire variants with improved catalytic properties. Among all the 17 mutants, E12 (A72T/V91S) was the most active and even displayed 2.8- and 3-fold higher than LXYL-P1-2 on -xylosidase and -glucosidase activities. The possible mechanism for such improvement was proposed by homology modeling and molecular docking between E12 and 7--xylosyl-10-deacetyltaxol. The recombinant yeast GS115-P1E12-7 was constructed by introducing variant , the molecular chaperone gene and the bacterial hemoglobin gene . This engineered yeast rendered 4 times higher biomass enzyme activity than GS115-3.5K-P1-2 that had been used for demo-scale fermentation. Thus, GS115-P1E12-7 becomes a promising candidate to replace GS115-3.5K-P1-2 for industrial purpose.