| Oreorchis patens(Lindl.) Lindl. is the perennial herbaceous plants of Oreorchis genus, Orchidaceae family, its pseudobulb is a kind of traditional Chinese medicine, with the properties of cold, bitter and sweet flavor and has a little toxin. In the Dictionary of Chinese Medicine recorded it can be used as the original plant of Shancigu drug, but it is not included in Chinese Pharmacopoeia. The three kinds of the original plants of the Shancigu reproduced very difficultyly like other species in Orchidaceae family. In recent years, the demand of Shancigu drugs is gradually increasing, people thus have begun digging O. patens as Shancigu acquisitions, leading the plants have become a scarce species. In order to ensure the sustainable development and utilization of the plant, and exploring the possibility as the original plant of Shancigu drug, it is carried out to research on the key technology of the development and utilization of O. patens resources from the following aspects:(1) To establish a HPLC Library of the chemical composition of O. patens and compared it with three original plants of Shancigu records in Chines Pharmacopoeia;(2) To study the basic pharmacological effects of O. patens, also compared it with the three original plants of Shancigu;(3) In order to determine the genetic characteristics of the species, the six samples of O. patens in Jilin province was studed with RAPD and ISSR molecular markers, and also also compared with the three original plants of Shancigu;(4) To solve the problem of reproduction difficulity, the asexual propagation techniques of O. patens was carried out. Through the study of the key technologys, we could provide a theoretical basis for the development and cultivation of the plant.The results are as follows:(1) There are 20 common peaks between O. patens(6 samples from different places) and original plants of Shancigu(6 samples of 3 genus) in the HPLC maps, unique peaks were more than 70, the total peak only exists difference in the amount of that O. patens and Shancigu herbs also contain the same chemical composition. According to the principal component analysis and cluster analysis, the chemical components of O. patens is very similar to Shancigu, which shows that O.patens has the potential development and utilization value as Shancigu, and deserved further attention.(2) The MIC of O. patens and three original plants of Shancigu to common pathogens of human were determined with the liquid micro-culture. The results showed that the ethanol extracts of these medicinal herbs have antimicrobial activity against Staphylococcus aureus(SA) and Pseudomonas aeruginosa(PA), the MIC is less than 50μg/ mL, but not to Escherichia coli(E.coli).(3) The genome DNA was extracted from different parts of O. Patens with CTAB method, and measured the OD260/OD280 of all samples was 1.9, the concentration of DNA in 89.99-367.80g/mL. After RAPD-PCR amplification, the electrophoretic pattern of the amplified products was good, the bands were clear, which indicated that the DNA could be used in the following molecular marker experiments.(4) Through the detection of RAPD and ISSR labeling methord, we can find that the genetic diversity of O. patens, Cremastra appendiculata(D.Don) Makino, Pleione bulbocodioides(Franch.) Rolfe and P. yunnanensis Rolfe is very rich and the differentce is very obvious. The results of cluster analysis showed that the 4 kinds of herbs could be classified in 3 categories. Different sorts of samples show low identity, long distance and far relationship about hereditary, the same sort of samples are opposite. |
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