Taxonomic and Pharmaceutical Analysis of Medicinal Plants of Genera Clematis and Houttuynia
|Tutor||Yi Yu Cheng|
|Course||Pharmaceutical and Taxonomical Analysis|
|Keywords||Pharmaceutical analysis Molecular systematics Two-dimensional electrophoresis Ethnobotany Proteomics Quality control Morphological classification system Chemotaxonomy Clematis Houttuynia Traditional Chinese medicines Taxonomy HPLC-MS GC-MS|
Traditionally, plant taxonomy uses morphological characteristics, secondary metabolites and molecular biomarkers to delimit boundaries within or between various taxa and demonstrate their taxon relationships. Although remarkable achievements have been obtained but many problems in evolutionary taxonomy hitherto need further investigations. Recently, there is an increasing emphasis on application of "-OMICS" sciences in the post-genome era. The proteomic approach has great potential for providing global, holistic profiling and characterization of protein expressions in cells, tissues or organs. The progress in the research of plant proteome will not only afford new insights to decipher the unraveling of phenomenon of plant’s response to biotic and abiotic stimuli but also it will be marvelous leap towards improvement of agronomic and medicinal plant species. Moreover, proteome technique will made it feasible to differentiate intra-taxon or inter-taxa phylogenetic relationships in different plants and it will be initialization of corner stone step towards the development of modern biosystematics.In this research project we studied preliminary application of the proteome technique for taxonomic studies to differentiate intra-genus or inter-genera species. Mainly, those taxa having medicinal importance were selected for the studies and a comparative analysis was performed by using chemical and morphological features of same taxa in order to evaluate the significance of proteome approach as taxonomic and phylogenetic tool. Secondly, medicinal plants of genera Clematis and Houttuynia being used in different traditional Chinese medicines (TCMs) were analyzed by different analytical approaches and protocols were evaluated and optimized for their quality control (QC).In perspectives of taxonomic analysis; firstly, morphological studies including description of collected species and taxonomic analysis of the taxa based on 27 phylogenetically informative phenotypic characteristics was conducted by using Matlab/Multivariate Statistical Package (MVSP 3.1) and results were presented as phenetic tree. The ambiguities of this classical classification system in comparision with those of chemotypic or/and proteomic approaches were discussed.Secondly, chemodiversity of saponins in 12 species of genus Clematis were analyzed by LC-UV/MS approach, differential distribution data of saponins were computed as matrix and analyzed by Matlab/MVSP, and results were presented as phenograms. The taxonomic significance of saponins distribution patterns in comparision with phenotypic and protein based classification systems was presented and discussed.Thirdly, in the proteomic studies for taxonomic and phylogenetic analysis, a protocol for leaf protein extraction, solubilization and analysis by two-dimensional electrophoresis (2-DE) was optimized by evaluating different methods of choice. For taxonomic studies, leaf proteome of 12 different taxa of Clematis and Ranunculus muricatus (as one out group) were analyzed by 2-DE approach. The gels with repetitive polypeptide spots were selected for each accession and presence or absence of spots was formulated as matrix. The obtained matrix was analyzed by Matlab/MVSP and results were presented as phenograms. The proteome approach was used to characterize an important TCM, Clematis chinensis by identifying biomarkers. Identified proteins were mostly concerned with energy metabolism, evolution and defensive mechanism of plants. The proteome technique was also applied to measure genetic distances between different specifies of genera Clematis, Ranunculus and Paeonia. The genetic distances among the taxa of different genera were computed by using differential distribution of polypeptide chains (expressed genomes). The data were computed as matrix and processed by MVSP and results were presented as phenograms. This proteomic approach has proved to be an eminent technique to differentiate different taxa of three genera of family Ranunculaceae.In perspectives of pharmaceutical analysis; firstly, an ethnobotanical expedition was conducted to Tian Mu Shan Biosphere Reserve (TMSBR) Zhejiang not only to collect medicinal taxa of the genera but also to document precious indigenous ethnomedicinal knowledge about the flora of the region. The significance of ethnomedicinal knowledge in drug discovery and drug development is debated. Some of the obtained taxa (TCMs) were chemically analyzed by different analytical techniques.Secondly, Clematis huchouensis Tamura (CHT) which is an important TCM was studied for its chemical constituents (flavonoids). A protocol based on high performance liquid chromatography (HPLC) fingerprinting was optimized for its QC. The optimized protocol can also serve as modal for other TCMs.Thirdly, chemical composition of essential oil (EO) of Houttuynia cordata Thunb. (HCT), a valuable TCM was carried out by GC/MS technique. EO was extracted by hydrodistillation (HD) method and obtained EO was analyzed by On-Column GC/MS and their quantity contents were determined. The established GC/MS methodology can serve as good QC protocol for TCMs of this species and modal for other botanic drugs. The significance of different taxonomic and pharmaceutical approaches in QC of ethnomedicines/botanical drugs with exploring their individualistic role was the corroborative goal of this work.