Chinese Medicine Huoxiangzhengqi Water Quality Evaluation And Pharmacokinetic Study

Objective: The Formula usually used in clinic is the characteristic and soul of TCM(Traditional Chinese Medicine). Quality control of the TCM Formula is focused by researchers, and it is difficult for modernization and internationalization of TCM. In this study Huoxiangzhengqi Liquid(HXZQL) was selected as an example to analyze the effective components and establish multi-components determination method. These will provide scientific evidence for fingerprint and quality control of drugs.Methods: The Formula of HXZQL is composed of Atractylodes lancea(Thunb.)DC., Citrus reticulata Blanco, Magnolia of'ficinalis Rehd . et Wils, Angelica dahurica.(Fisch. Ex Hoffm.)Benth. et Hook. f, Poria cocos(Schw.)Wolf, Areca catechu L. Pinellia ternate(Thunb.)Breit, Extractum Glycyrrhizae, Oleum Pogostemonis, oil of Folium Perillae. HXZQL is used extensively in clinic because of its remarkable effect. In the study, LC-MS/MS(Liquid Chromatography-Mass Spectrum/Mass Spectrum) and GC(Gas Chromatography) were used for constituents identification of HXZQL. The constituents included Patchouli alcohol, Magnolol, Honokiol, Imperatorin, Isoimperatorin, Hesperidin, Narrigin, Glycyrrhizic acid, Liquiritin, Nobiletin and Phellopterin were identified. GC with the internal standard method was used to determine the content of Patchouli alcohol of Pogostemonis which plays the part of kingship in the HXZQL Formula. HPLC(High Performance Liquid Chromatography) was used to determine the content of Liquiritin and Hesperidin at one time. The contents of Magnolol, Honokiol, Imperatorin, Isoimperatorin and Glycyrrhizic acid were determined at the same time by HPLC. LC-MS/MS was used to identify Magnolol in the plasma samples after oral administration with the extraction of HXZQL. A method with the internal standard for the determination of the Magnolol concentrations in the dog plasma samples was performanced with HPLC. The primary Pharmacokinetic parameters and the regularity of Magnolol distribution were estimated with the Pharmacokinetic calculating software. The relationship between analgesic effect and time was observed by the experiments of writhing induced by acetic acid in mice.Results: (1) Patchouli alcohol of HXZQL was identified with its contrast by GC; Magnolol, Honokiol, Imperatorin, Isoimperatorin, Hesperidin, Narrigin, Glycyrrhizic acid, Liquiritin, Nobiletin and Phellopterin were identified by analyzing the data of Mass Spectrum, thereinto, Magnolol, Honokiol, Imperatorin, Isoimperatorin, Hesperidin, Narrigin, Glycyrrhizic acid and Liquiritin were also identified with their contrasts.(2) GC was used to determine the content of Patchouli alcohol of HXZQL, and n-octadecand was used as internal standard. A good linearity was obtained in the range of 0.0411~0.4112mg·mL-1, r=0.9992. The average recovery was 97.67% with a RSD of 1.48%(n=6).(3) HPLC was used to determine the content of Liquiritin and Hesperidin at one time. A good linearity of Liquiritin was obtained in the range of 13.35~267μg·mL-1, r=0.9999. The average recovery was 101.3% with a RSD of 2.91%(n=6). A good linearity of Hesperidin was obtained in the range of 18.00~360.0μg·mL-1, r=0.9999. The average recovery was 100.8% with a RSD of 1.47%(n=6).(4) HPLC was used to determine the content of Magnolol, Honokiol, Imperatorin, Isoimperatorin, Glycyrrhizic acid at the same time. The good linearity was obtained in the ranges of 93.50~467.5μg, 41.30~206.5μg, 18.15~90.75μg, 235.0~1175μg, 311.0~1555μg, respectively. The average recoveries were 98.31%, 99.56%, 98.57%, 98.05%, 99.57%, respectively.(5) Magnolol and Emodin were identified by LC-MS/MS. HPLC was used to determine the concentrations of Magnolol in the dog plasma samples and Emodin was used as internal standard. A good linear relationship was obtained between 0.063~10.08μg·mL-1. The limit of detection was 18ng·mL-1. The average absolute recovery was more than 85%. The RSD values of within-day and between-day precision and stability were both below 10%. The metabolism of Magnolol in dogs was confirmed to two-compartment model with weight 1/C2.The main parameters were as follows: Cmax0.4870mg·mL-1, Tmax2.0000h, AUC0~61.59mg·h·L-1, t1/2α0.433h, t1/2β1.256h, CL/F3.937L·h-1·kg-1, k100.741h-1, k120.219h-1, k211.192h-1, MRT0~∞8.7260h, MAT11.8401h, respectively.Conclusion: The method with internal standard for determination of the magnolol concentration in dog plasma was established for the first time. Through the determination of magnolol which is a active chemical from extraction of HXZQL, the pharmacokinetic study of the formula was accomplished, and the functionary regularity and feature of it were also illustrated.The qualitative and quantitative methods of the main constituents were found. .It provided multi-components determination method which can provide quality control for 50% of material medecas in HXZQL. The GC method for the determination of patchouli alcohol implemented the quality control of the king part in the HXZQL formula. All of these were the foundations of establishing scientific and comprehensive methods to appraise the quality of HXZQL in further study.