Study On The Quality Control Of Liuwei Dihuang Decoction And Pharmacokinetics In Rats

Liuwei Dihuang Decoction is the classic formula, which has the function of nourishing yin and supplementing the kidney. It is composed of Radix Rehmanniae Praeparata, Fructus Corni, Rhizoma Dioscoreae, Poria, Cortex Moutan, Rhuzoma Alismatis. It was carefully designed and the drugs were combined based upon accurate efficacy. To study on the quality control and pharmacokinetics of Liuwei Dihuang Decoction: A sensitive, specific HPLC method for the fingerprint of Liuwei Dihuang Decoction and five effective components simultaneous determination was developed, which provides a better qualitative and quantitative analysis method for the inside quality control of Liuwei Dihuang Decoction; After i.g Liuwei Dihuang Decoction several times, a methodology for determining the concentration of morroniside and loganin in rat plasma was developed. After i.g Liuwei Dihuang Decoction combination groups, the pharmacokinetics parameter was determined which explore the dynamic state changes regulation inside the body, and to provide scientific basis for clinical medication.The main content reads as follows:1. The quality control of Liuwei Dihuang DecoctionA sensitive, specific and rapid HPLC method for determination of liuwei duhuang decoction was developed, which provides a better qualitative and quantitative analysis method for the inside quality control of Liuwei Dihuang Decoction. 19 common chromatographic peaks were found. The concentration of gallic acid, morroniside, loganin, paeoniflorin, paeonol were determined simultaneously. The common chromatographic peaks were mainly from Radix Rehmanniae Praeparata, Fructus Corni and Cortex Moutan. The 2^nd, 8^th, 12^th, 14^th chromatography peaks were both from Fructus Corni and Cortex Moutan.After comparing the concentration of five components in all combination groups, six-herb were found which can increase the concentration of four- components in Liuwei Dihuang Decoction. The concentration of gallic acid was lower in single Cortex Moutan herb decoction, but rises when mixed with other herbs.Polysaccharide is the effective component of Liuwei Dihuang Decoction. An anthrone-sulfuric acid method for the determination of polysaccharide in Liuwei Dihuang Decoction.2. Serum pharmacochemistry research on the constituents absorbed in blood after i.g Liuwei Dihuang DecoctionMorroniside and loganin were found absorbed into blood after i.g Liuwei Dihuang Decoction. A sensitive, specific HPLC method for instant determination of morroniside and loganin in rat serum was developed. The linerity range of morroniside was 0.254～102μg/ml(r=0.9998). The RSD% of within-day and between-day precision were lower then 15%;The linerity range of loganin was 0.579～232μg/ml (r=0.9998), the RSD% of within-day and between-day precision were lower then 15%. This method specially provides an accurate and sensitive way in detecting the in vivo blood concentration of morroniside and loganin in serum at the same time.After i.g Liuwei Dihuang Decoction compatibility groups , it found that Liuwei Dihuang Decoction can promote the absorption of morroniside and loganin in rat blood.3. the pharmacokinetics research on the rat after i.g Liuwei Dihuang DecoctionA sensitive, specific and rapid HPLC method for determination of morroniside and loganin in rat plasma at the same time was established. Morroniside and loganin were extracted by methanol to eliminate of foreign matter and dissolved with methanol after blowing by N₂; then they were detected by RP-HPLC. Morroniside and loganin were fully separated from the other components in plasma. The linerity range of morroniside was 0.01021～10.21μg/ml(r=1), the average recovery was higher than 70%, the RSD%of within-day and between-day precision were lower than 5%; the liner range of loganin was 0.006325～6.325μg/ml (r=1), the average recovery was higher than 80%, the RSD%of within-day and between-day precision were lower than 5%. The method specially provides an accurate and sensitive way in detecting the in vivo blood concentration of morroniside and loganin in the plasma at the same time.The method was successfully applied to determinate the concentration of morroniside and loganin in rat plasma after administration intragastric of Liuwei Dihuang Decoction(a), three-tonifying herb group(b), one-tonifying one-purging herb group(c) to rat(n=16). The plasma drug concentration-time curve of morroniside and loganin in group a and group b conform to a 1-compartment open model with a first order absorption; the plasma drug concentration-time curve of morroniside and loganin in one-tonifying one-purging herb group conforms to a 2-compartment open model with a first order absorption. Study on the pharmacokinetics parameter to know the process inside the body, to provide science basis for clinical.Two metabolites were found in rat plasma and urine, purified part was gained by HPLC . After HPLC-DAD and LC-MS analytical identification, it was primarily conferred that the possible molecular ion peak[M+H]⁺ and [M+Na]⁺ of metabolite Ml were 341 and 364.The possible molecular ion peak[M+H]⁺ of M2 was 272. The accurate chemical structure was to be completed by further methods as NMR.