Study On Serum Pharmacochemistry Of Gegen Qinlian Decoction

Gegen Qinlian Decoction (GQD) was one of the most widely used traditional Chinese medicines , composed of Radix Puerariae , Radix Scutellariae,Rhizoma Coptis and Glycyrrhiza. It has been used to treat for antipyretic, anti-diarrhea, anti-inflammatory and relieving muscles. There were many compounds in GQD including flavones,alkaloids and triterpenes. In conventional studies, a few mark compounds had been used to control the quality or investigate the pharmacokinetic of GQD. However,these studies were not able to reflect the decoction's overall efficacy,because the multiple constituents may work synergistically,which was not combination of each herb simply. Based on the hypothesis that only the constituents migrating to blood after oral administration of TCM have the probability to be effective constituents,pharmacochemistry strategy was applied to study the effective compounds in rat plasma after oral administration of GQD.1. A chromatographic method was established on rat plasma after oral administration of GQD. The chromatographic separation was carried out on Globalsil C18 ( 250 mm×4.6 mm ID, 5μm ) column with a binary mobile phase system consisting of 20 mmol·L-1 ammonium acetate and acetonitrile with gradient elution at 270 nm. Precipitation with methanol was chosen as a sample preparation method,because it not only removed a lot of endogenous impurities but also extracted most of the target compounds. 0.1 mol·L-1 ammonium acetate ( pH 4.0 adjusted by acetic acid ) was added during the sample preparation so as to improve the extraction recovery rate.2. Constituents migrating to blood after oral administration of GQD were ascribed. Learning from fingerprint strategy,rat plasma samples after oral administration of GQD and individual herb were compared. According to the retention behavior,31 compounds in rat plasma were ascribed,including 15 of them came from herb,13 of them were metabolites,and 3 of them were the new substances in plasma.3. Constituents migrating to blood after oral administration of GQD were identified by HPLC-DAD-MS. Due to the advantages of sensitivity and specificity , LC-MS (ESI/APCI) was applied to determine the compounds in plasma after oral administration of GQD. According to the retention time, UVmax and m/z in positive or negative ion mode,18 compounds were identified or tentatively elucidated comparing with standard compounds or literature. There were 14 compounds came from herb,including puerarin, 3'-methoxy puerarin, daidzein 8-C-apiosy (1→6) glucoside , daidzein , baicalin , oroxylin A-7-O-β-D-glucuronide , apigenin-7-O-β-D-glucuronide, wogonoside,jatrorrhizine, coptisine, palmatin, berberine, liquiritin, glycyrrhizin. Furthermore, Four metabolites were tentatively designated to be daidzein4'-O-sulfate,daidzein 7-O-β-D-glucuronide, 6-O-Methyl-baicalin-7-O-sulfate,baicalin -6-O-β- glucopyranuronoside.4. Peak area vs. time curves were studied after oral administration of GQD in rat. Based on the previous chromatographia conditions , repeatability and stability of the method were investigated,and they were suitable to apply to study the pharmacokinetic of GQD. The peak areas of 12 components in rat plasma were measured at the same time at 270nm. Drug peak area vs. time curves of compounds including components from GQD and metabolites were studied after oral administration of GQD in 48h. The results showed that components in Pueraria were absorbed well with rapid metabolism, while the components in Scutellaria were easy to present double-peak trends due to hepatoenteral circulation. Then , the chromatography data was imported to SIMCA-P to perform Principal Component Analysis (PCA), which was helpful to reveal time-effect relation of GQD.In conclusion, the modern liquid chromatography coupled with DAD detection and MS identification was successfully used in the study on the pharmacochemistry of Gegen Qinlian Decoction. With the fingerprint strategy,18 compounds were identified and 12 peak area vs. time curves were studied, which provided a new method for analyzing the effective compounds in TCM.