Study On The Quality Control And Serum Pharmacochemistry Of Schisandra Chinensis

Study on the material foundation of the efficacy of traditional Chinese medicine (TCM) is the key and difficult point of the modernization of TCM. But there is no generally accepted model of it until now. The traditional approach is to separate the components for the activity test or track separation guided by the activity. However, whether the components of the method to determine are the efficacy of representative components is a question. Although using the traditional method did illustrate the material basis for the efficacy of some TCM. First, TCM is a multi-component and multi-target complex system. Its efficacy is often the effects of integrating a variety of components acting on different targets, and chemical separation often makes the integration effect disappear. Secondly, TCM for oral administration determined the components that are absorbed into the body can generate the effects. Furthermore, both the original components and the metabolites can work in the body. Therefore, some scholars have proposed a study strategy based on "Medicated serum". The theory guideline is the medicated serum can be study on the active components of TCM because in the body TCM produce the effects depends on the concentration of active components in the target organ or tissue. And blood plays an important role in transporting these active components of TCM to the target organ and tissue. Although the active ingredients in the body are not all of the absorbed components, the kinds and number of the components are greatly decreased after the treatment of the gastrointestinal tract. So the study on medicated serum can not only simplify the complex system of TCM but also directly determine the active components.The research strategy of the active components of TCM based on "medicated serum" includes the research of "serum pharmacology" and "serum pharmacochemistry". The "serum pharmacology" refers to the pharmacodynamic evaluation on the cell model treated on medicated serum in vitro. The "serum pharmacochemistry" refers to the qualitative and quantitative analysis on the plasma-derived ingredients (including original compounds in vitro and metabolites in vivo) by modern analytical methods. The combination of the two methods can basically clarify the active components of Chinese herbal medicine.Schisandra chinensis is the dried ripe fruit of Schisandra chinensis (Turcz.) Baill. of Magnoliaceae, has a variety of chemical constituents and pharmacological activities. Dibenzocyclooctadienes (also known as schisandra lignans) is the main chemical composition of Schisandra chinensis and the main active ingredient. Nearly50ingredients of Schisandra lignans have been isolated from Schisandra chinensis currently. Schisandra lignans have the exactly hepatoprotective effect, which is related to its structure and the process of its metabolism in vivo. But the structure-activity relationship and the active components of Schisandra lignans in vivo are still confused. Thus, it has the vital significance to clarify the hepatoprotective effects material basis and the structure-activity relationships of Schisandra lignans by study on the process in vivo using the pharmacochemistry method.In view of this, this study based on the quality control and extraction process of Schisandra chinensis, used the serum pharmacochemistry method, fingerprint technology, LC-MS and other detection methods to analyze the qualitative and quantitative of the prototype of Schisandra lignans and its metabolites on rats by gastric gavage Schisandra chinensis.The dissertation has two parts:the review and the experimental research.Part1, the reviewIt cited83articles illustrated the progress in the chemical constituents of Schisandra chinensis and Schisandra lignans pharmacochemistry research, and then briefly introduced Research ideas and program design.Part2, experiments1、study on the qualitative and quantitative analysis of S.chinensis and the extraction technical study of schisandra lignans(1) Study on the quality evaluation methods of S.chinensis based on the combination of fingerprint and single marker (QAMS).The common mode of fingerprint was established after determination fingerprints of12batches samples of S.chinensis by HPLC. There were24common peaks in these samples. Compared with the control sample, six of them were identified as schizandrol A, schizandrol B, schisantherin A, deoxyschizandrin, schizandrin B and schizandrin C. The similarity of the twelve batches fingerprints were greater than or equal to0.991. The relative correction factors (RCF) of schizandrol B, schisantherin A, deoxyschizandrin, schizandrin B and schizandrin C were established using the Internal control of Schizandrol A, and the contents of the components in these12batches of samples were detected by QAMS. Simultaneously the contents of the six lignanoids were determined by external standard method. The accuracy of QAMS was evaluated by comparison of the calculated and measured values. As the result, the established RCF had a good reproducibility. It was no significant differences between the quantitative results of external standard method and QAMS.(2) Comparison of the content of6kinds of lignans from S.chinensis and its seeds and pulp.The contents of six lignanoids of schizandrol A, schizandrol B, schisantherin A, deoxyschizandrin, schizandrin B and schizandrin C in fruit and seeds and pulp of S.chinensis have been dectedted by HPLC. By analyzed and compared, the results showed that the contents of chizandrol A, schizandrol B, schisantherin A, deoxyschizandrin, schizandrin B and schizandrin C in the seed accounted for91.23%,87.49%,95.10%,90.76%,93.43%and90.54%of the total content of medicinal herbs. The total average of these six contents in the seed was accounted for84.63%of the total content of medicinal herbs. It showed schisandra lignans mainly existed in the seeds.(3) Study on the extraction technical of schisandra lignans from S.chinensis seedsTotal of schisandra Lignans and six major lignanoids extracting weighted as index, frequency of reflux, reflux time, alcohol concentration and alcohol consumption as the factors, three levels in each factor, the extraction technique of schisandra lignans in S.chinensis were studied by L9(34) orthogonal design. The analysis of variance showed that the frequency of reflux and reflux time had a significant impact on extraction rate of lignanoids, but there was no significant on alcohol concentration and alcohol consumption. The optimal extraction project was six times the amount of95%ethanol, reflux three times and60min every time.(4) Analysis of the fingerprint of the extracts from S.chinensis seeds and determination the content of schisandra LignansThe common mode of fingerprint was established after determination fingerprints of11batches samples of S.chinensis by HPLC. There were24common peaks in these samples. Compared with the control sample, six of them were identified as schizandrol A, schizandrol B, schisantherin A, deoxyschizandrin, schizandrin B and schizandrin C. The similarity of the11batches fingerprints were greater than0.994. Extracts content of these6kinds of S.chinensis were identified by determination of both the QAMS and the external standard method. Then the results of the two methods of measurements were compared, and there were no significant differences.2、Study on the serum pharmacochemistry of the extract of S.chinensis seeds(1) Study on fingerprint of the medicated serum of the extract from S.chinensis seedsThe extract of S.chinensis seeds were orally given to male SD rats once (the doses of schizandrol A, schizandrol B, deoxyschizandrin, schizandrin B were29.44%、8.45、7.76and14.2mg/100g).6h later, the plasma were collected and detected fingerprints of10batches plasma samples by HPLC which had used above. Then the common mode of fingerprint was established. Compared with the fingerprints of blank plasma, there were16common peaks. Then compared with the fingerprints of the fingerprints extract of S.chinensis seeds and control samples, nine of these peas were the metabolites in body and seven peaks were original compounds. Four original compounds were chizandrol A, schizandrol B, deoxyschizandrin, schizandrin B.(2) Lignanoids in the extract of S.chinensis seeds and in plasma were detected by UPLC-ESI-MSThe extract of S.chinensis seeds and the medicate serum were detected by UPLC-MS/MS. The results showed the extract included29schisandra lignans. Compared with the control sample, five of them were schisandrol A, schisandrol B, schisantherin A, deoxyschizandrin,and schisandrin B. The structures of the other24were speculated by referring to the relevant literature.In the rat serum samples,28drug-induced constituents were found. The17of them were original components,11of them were metabolites. And4metabolites were schisandrin, schisandrol B, deoxyschizandrin,and schisandrin B. Schisantherin A has not been detected. Reference to the relevant literature, the other13original components could be isomer schisandrin, gomisin D, tigloylgomisin H, angeloylgomisin H, Gomisin F, schisantherin B, Schisanhenol, tigloylgomisin P, Gomisin M1, epigomisin O, Gomisin N, benzoylisogomisin O and schizandrin C. And the other11metabolites were7,8-dihydroxy-3-demethyl schizandrol A、7,8-dihydroxy-2-demethyl schizandrol A、Gomisin H isomer、Gomisin P、7,8-dihydroxy schizandrol A、Gomisin H (Gomisin isomer)、Deangeloylgomisin B、schisandrin metabolites(schisandrol B or deoxyschizandrin,)、schisandrin B metabolites, and the other needed to be identified.(3) Preliminary study on the pharmacokinetics of4lignanoids extracts of S.chinensis seeds The extracts of S.chinensis seeds, schizandrol A, schizandrol B, deoxyschizandrin and schizandrin B were orally given to male SD rats once. After15min、30min、1h、2h、4h、6h、8h、10h、12h and24h, the plasma of rats were collected.Compared with data of the drug concentrations-time of all samples,It was found that the pharmacokinetics of chizandrol A and schizandrol B may be influenced by other lignans and metabolized slowly. There was no obviously difference in the pharmacokinetics of deoxyschizandrin from the extracts, schizandrin B from the extracts and the monomers of them. The other components, except deoxyschizandrin, could not completely metabolized and excreted in24h. The present study provides the basis for discussing the pharmacokinetics of the primary lignans compounds in the future.