| Zicao, a commonly used Chinese Materia Medica (CMM) for treatment of macular eruption, measles, sore-throat, carbuncles, burns, etc., is mainly derived from the roots of Arnebia euchroma (Royle) Johnston., A. guttata Bunge and Lithospermum erythrorhizon Sieb. et Zucc. However, according to our previous literature research and field investigations, six species of genus Onosma have been found in use as the customary source species of Zicao in some regions of southwest China; these are Onosma paniculatum Bur. et Franch., O. exsertum Hemsl., O. confertum W. W. Smith, O. hookerii Clarke var. longiflorum Duthie, O. hookerii Clarke and O. waltonii Duthic. To date, there are limited studies related to these species, in particular for the Tibetan species of O. waltonii Duthic, use of which had not even been recorded in any publications on herbs. This thesis, then, focuses on authentication of the different source species of Zicao and related herbs of Boraginaceae, through the following two-pronged approach:;Firstly, to identify the different Boraginaceous herbs and refine the methods of identification.;As a first step for the authentication of Zicao and related Boraginaceous herbs, these were identified by their botanical species to ensure use of the authentic species in various samples in the study. Then identification was conducted among the different species on the basis of morphological characteristics of crude drugs and microscopic characteristics of both transverse sections and powders of roots. The species proved to be individually identifiable by their distinguishing characteristics.;Fourier Transform Infrared Spectrascopy (FTIR), as a powerful analytical tool for characterizing and identifying some components of an unknown mixture in a nondestructive way, was tried for the identification of the different Zicao species in the study. With the differences in infrared absorption spectrum, the different species of crude drugs were distinguished from each other. Moreover, when 1D FTIR spectra were combined with 2D correlation IR synchronous spectrascopy, the samples taken from different growth years of the same species could be readily distinguished from each other, as could samples from wild and cultivated specimens.;An improved protocol, based on the typical CTAB method, was developed for the extraction of high quality genomic DNA from these herbs that contain multiple of polysaccharides, polyphenolic compounds, etc., The extracted genomic DNA was then amplified and sequenced on several selected regions of the chloroplast DNA and nuclear ribosomal DNA; the selected regions were the intergenic spacer between trnTUGU-trnLUAA , the trnLUAA intron, the intergenic spacer between trnLUAA-trnFGAA , the internal transcribed spacer region (ITS) and the internal transcribed spacer region 1 (ITS1). Thirtythree sequences were obtained from these species, an investigative resource never before made available for these herbs. Based on the sequence alignments of the four genomic regions, the different herbal species could be distinguished from each other. Furthermore, preliminary studies on the phylogenetic relationships of these species were analyzed in connection with the sequence data of the different regions. With the exception of the trnL intron, the sequence data on the other three regions generally support the homological properties of the Boraginaceous species.;Secondly, to evaluate the pharmaceutical quality of Zicao and related Boraginaceous herbs.;A high performance liquid chromatographic method using diode-array detection (HPLC-DAD) was developed for distinguishing eight selected naphthoquinones in Zicao. Two positional isomers (namely, 2-methyl-n-butyrylshikonin and isovalerylshikonin) have been successfully separated and quantified for the first time in this study. The contents of the eight naphthoquinones were then determined and compared in Zicao samples from the nine species for the evaluation of their pharmaceutical quality.;Furthermore in this study, by combining information from both the traditional application and modern pharmacological studies of the herbs, a multi-component HPLCDAD fingerprint method was developed which can sufficiently reflect chemical information from both nonpolar components of naphthoquinones and polar components like phenolic acids in a sample by using a single column in a single gradient chromatographic system. A total of eight naphthoquinones and two phenolic acids were selected as reference compounds here for the authentication of Zicao and related herbs of Boraginaceae. The pharmaceutical qualities of each different herbal sample could be evaluated by detecting the existence and absorption intensities of related reference marker compounds while comparing the chromatographic fingerprint to a standard Zicao sample.;Overall, to ensure the safe and effective use of Zicao in clinic, authentication is fundamental for standardization and reliability of this herb. This will also conduce to the rational application and development of these Zicao and related herbal resources. |
