Chuanxiong Active Ingredient And Xiaoxuming Soup In Rats In Vivo Metabolism Study

Traditional Chinese Medicine (TCM) and its prescriptions are Chinese national treasures; they are also the important fountainhead of developing independent innovative new drugs in our country. Studies on pharmacokinetic of TCM prescriptions can help reveal the pharmacodynamic material basis and mechanism and have a great significance to the modernization and internationalization of TCM. In this paper, rhizome of Ligusticum chuanxiong Hort. (Chuanxiong) and the TCM prescription Xiao Xu Ming-Dectotion (XXMD) were selected to study the metabolic characteristic of the active monomer in rats after oral administration of the extract or the active monomer and explore the differences of pharmacokinetic of some active constituents in rats after oral administration of the TCM prescription or its active fraction, these provide useful reference for pharmacokinetic studies of TCM.Chuanxiong is a well-known traditional Chinese medicinal herb, which is firstly recorded in《Shennong's Herbal》. It is mainly used for the treatment of headache, rheumatic arthralgia, menstrual disorders, swelling and pain due to traumatic injury, pricking pain in the chest and coronary heart diseases. In this paper, we compared the differences of metabolic patern and metabolites after oral administration of essential oil or ligustilide to rats. The structures of metabolites of ligustilide were identified. The inhibition of essential oil on different subgroup of CYP450 was studied.A simple HPLC method for analysis metabolites of ligustilide in rats after oral administration of ligustilide or essential oil was developed in the present work. The metabolites were separated on a Phenomenex C₁₈ (4.6×250mm, 5μm) column with a gradient elution system which composed of acetonitrile and aqueous acetic acid. The mobile phase flow rate was 0.8 mL/min. The HPLC was equipped with a programmable diode array detector (DAD), the detected wavelength was set at 280 nm, the temperature of column oven was maintained at 25℃.By HPLC-DAD, the metabolites in urine, feces and bile of rats dosed with essential oil or ligustilide were analyzed. The results indicated that ligustilide was eliminated mainly as metabolites localized in urine, a fewer metabolites detected in feces and the content of the metabolites was very low, the parent ligustilide was not detected neither in urine nor in feces. In the bile, only a little of metabolites were detected, but the content of parent ligustilide was very high. It was notable that the obvious differences of amount and relative content of metabolites were observed in rats after oral administration of essential oil or ligustilide, which indicated that other chemical constituents in essential oil could influence the metabolism of the main active constituent ligustilide.Seven metabolites in rat urine were detected and six of them were separated and purified. According to the data of NMR, MS and MS/MS, four metabolites were identified as senkyunolide I, 3, 5-dihydroxy-3-butylphthalide, 6, 7-dihydro-6-hydroxy -7-sulf methyl ligustilide and senkyunolide C. The results provide useful references for the further study of pharmacokinetic and clinical application of ligustilide.In the paper, the inhibition of essential oil on CYP4501A2, 2C9, 2C19, 2D6, 2E1, 3A4 was studied, the calculated IC50 of essential oil were all much more than 10μM, which indicated that essential oil had no inhibition on CYP4501A2, 2C9, 2C19, 2D6, 2E1, 3A4. The results afforded the scientific data for safety of clinical drug combination of essential oil.In order to study the pharmacokinetic of XXMD, a rapid and specific liquid chromatography coupled with electrospray ionization mass spectrometry (LC-ESI-MS) method had been developed and validated for the identification and quantification of 5-O-methylvisamminol and cimifugin in rat plasma after oral administration of XXMD prescription or the active fraction (AF) of XXMD. Biological samples were prepared by liquid-liquid extraction with acetoacetate. Phenacetin was used as internal standard (IS). The chromatographic separation was accomplished on an Inertsil ODS-3 column (2.1 mm×100 mm, 3μm). The mobile phase consisted of acetonitrile and 0.1% aqueous formic acid (30 : 70, V/V) was delivered at a flow rate of 0.2 mL/min, the column oven was maintained at 30℃. Detection was performed on a linear ion trap mass spectrometer by full scan mode via electrospray ionization (ESI) source, and the positive ionization mode was used as the detection mode, the ion was set at m/z 290.5-291.5 for 5-O-methylvisamminol, m/z 306.5-307.5 for cimifugin and m/z 179.5-180.5 for phenacetin. Linearity of 5-O-methylvisamminol and cimifugin were established for the range of concentrations 11.5-230.0 ng/mL (r > 0.99) and 10.4-936.0 ng/mL(r > 0.99), respectively. The intra-day and inter-day precision were all less than 10.7%. The mean plasma concentration-time profile of 5-O-methylvisamminol indicated that the absorption of 5-O-methylvisamminol after oral administration of the AF of XXMD was lower than that of XXMD prescription (T_max(min) : 480 versus 360; C_max (ng/mL): 338.03 versus 318.46 ). But the mean plasma concentration-time profile of cimifugin showed a converse result, the absorption of cimifugin after oral administration of the AF of XXMD was faster than that of XXMD prescription (T_max (min): 180 versus 420; C_max (ng/mL): 2018.33 versus 1625.73 ). The results demonstrated that the pharmacokinetic of the AF was influenced by the inactive fraction of XXMD prescription. These provided scientific data for the further study of XXMD and useful references for the modern pharmacokinetic study of TCM.