Studies On Pharmacodynamic Material Basis Of Reduning Injection

TCM prescription "Re-Du-Ning Injection" consisted of three common used Chinese material medica, including Lonicera japonica Thunb (Jinyinhua), Gardenia jasminoides Ellis (Zhizi), Artemisia annual (Qinghao). It has been proved to be widely used for the treatment of wind-heat cold, cough, fever, upper respiratory tract infection and acute bronchitis in clinical practice.In this thesis, the influenza virus induced stress mouse model was adopted, using disease time, survival quality and survival time as evalulation index. Moreover, among the fractions of "Re-Du-Ning Injection" over Dainon HP-20column, RDN-3(95%ethanol elute) was proved to be the anti-virus active fraction of "Re-Du-Ning Injection" in vivo.The futher investigation on the chemical constituents of RDN-3led to the isolation of63compounds through various chromatographic techniques. The structures of56compounds were established on the basis of their physic-chemical properties and spectroscopic analysis, including twelve iridoids, twelve lignans, seven flavones, eleven phenolic components, three coumarins, four caffeoylquinic acids and other two compounds. Among them, six compounds were new compounds, namely japonicaside A (compound6), jasmigeniposide B (compound7),7,8,11-trihydroxy-l-hydroperoxy-4-guaien-3-one (compound13),7,8,11-trihydroxy-1-hydroperoxy-4-guaien-3-one (compound14),2-carboxy-4-methyl-a-methylene-3-(3-oxobutyl)-cyclohexaneacetic acid (compound15),1,3,4,5-tetrahydro-7-hydroxy-5-(4-hydroxy-3,5-dimethoxyphenyl)-8-methoxy-1,4-Methano-2-benzoxepin-10-methanol (compound27)。Furthermore, this paper has also established "Re-Du-Ning Injection" HPLC characteristic chromatogram, and we also separated and identified the main peaks. The structures of27compounds were established on the basis of their physic-chemical properties and spectroscopic analysis. Among them, three compounds were new iridoid glycosides, namely2’-O-trans-caffeoylgardoside (compound B23), jasmigeniposide B (compound B20),6"-O-trans-feruloylgenipin gentiobioside (compound B18).The anti-virus (RSV/EV-71/H1N1/H3N2) activities of thirty-six isolated compounds were evaluated in vitro. The results showed that4,5-di-O-caffeoylquinic acid,4,5-di-O-caffeoylquinic methyl ester,3,5-di-O-caffeoylquinic acid, luteolin, quercetin and genipin could inhibit H1N1influenza virus. Among them, we can focus on4,5-di-O-caffeoylquinic methyl ester and luteolin with EC50lower than50μM, and selective index (SI) value greater than10, they are value for further research. Additionally, quercetin, luteolin,3,4-di-O-caffeoylquinic methyl ester,3,5-di-O-caffeoylquinic acid,3,5-di-O-caffeoylquinic methyl ester,4,5-di-O-caffeoylquinic methyl ester,3,5-di-O-caffeoylquinic acid showed anti-influenza virus activity (H3N2). Among them, we can also focus on luteolin with EC50lower than50μM, and selective index (SI) value greater than10, and cell protection can be observed clearly under a microscope. Isorhamnetin and luteolin showed anti-virus activity (EV-71), but its anti-virus activity may be caused by cytotoxicity because of a lower SI value. In addition,5-O-caffeoylquinic acid, methyl5-O-caffeoylquinate,4-O-caffeoylquinic acid, methyl4-O-caffeoylquinate,3,5-di-O-caffeoylquinic acid,3,5-di-O-caffeoylquinic methyl ester,4,5-di-O-caffeoylquinic acid,4,5-di-O-caffeoylquinic methyl ester,3,4-di-O-caffeoylquinic acid,3,4-di-O-caffeoylquinic methyl ester can efficiently inhibit RSV. It is highly value for further research.The innovation of this research topic is to characterize the new constituent produced in traditional Chinese medicine prescription. The strategy and methodology of this study can be used to other chemical research of compound prescription of traditional Chinese herb drugs.This work partly revealed the anti-virus material basis of "Re-Du-Ning Injection",and provided evidence for developing the scientific and rational methods for quality control. This investigation also provided a possible research method for the study of bioactive and effective material basis of traditional Chinese medicine prescription.