Evaluation Of Wenshenkechuan Tablets On Cytochrome P450 Activity By Cocktail Probe Drugs And Its Metabonomics Study Of Compatibility

Objective: Wenshenkechuan tablets(WSKCT),a new Chinese prescription of effective parts, is composed of extracts of Cortex Magnoliae Officinalis and Fructus Cnidii by CO2 supercritical technology,and the extracts of Radix Glycyrrhizae by water and ethanol. The principal active constituent is made up of Magnolol, Honokiol, Osthole, imperatorin and glycyrrhizic acid. The prescription has the effective of warming the kidney to improve inspiration, relieving cough and asthma. Treatment is primarily aimed at cough, asthma and another diseases of kidney failuring receiving Qi and Lung Qi upward reversal.At present, this prescription has approved by State food and drug administration to do the clinical trials. Fructus Cnidii is recorded in a little toxicity, but its toxic mechanism is not clear; Radix Glycyrrhizae has a wide range of pharmacological activity,such as detoxify, reconciling the medicinal function. So more research of Fructus Cnidii and Radix Glycyrrhizae is necessary for elucidating the compatibility mechanism of WSKCT, it can provide a dependable basis for development and clinical application of this drug.Methods: Each probe drug and its metabolites was separated and analyzed by Cocktail probe and LC-MS technology to discuss the influence for the different subtypes of CYP450 enzyme by liquorice extract and its monomers; We had 8 experimental groups by splitting the prescription of WSKCT: Cortex Magnoliae Officinalis group, Fructus Cnidii group, Radix Glycyrrhizae group, Cortex Magnoliae Officinalis + Radix Glycyrrhizae group, Fructus Cnidii + Radix Glycyrrhizae group, Cortex Magnoliae Officinalis + Fructus Cnidii group, the whole prescription group, and the carbon tetrachloride group as a toxic compare for SD rats dosing 13 days, determining the liver function indexes AST, AIT, ALP, TBI, Urea, and renal function indexes Urea,Cre, observing the liver pathological changes; And the metabolites fingerprint analysis was to compare the rat urine between Fructus Cnidii group and control group through UPLC/Q-TOF MS analysis technique combining with metabolomics "whole concept" idea.Results: Cocktail probe experiment found that Glycyrrhizic acid could effectively restrain the activity of CYP 2A6, and induct the activity of CYP 2E1; Glycyrrhizin could induct the activity of CYP 2A6, and restrain the activity of CYP 2D6, CYP 2E1. Serum indexes showed: giving the extracts of Cortex Magnoliae Officinalis and Fructus Cnidii respectively, the AST and ALP increased significantly; After compatibility with Radix Glycyrrhizae, the AST and ALP was non-significance different with control group; There had the biggest toxicity when giving the extracts of Cortex Magnoliae Officinalis and Fructus Cnidii together, the AST, ALP,TBI,Urea all increased significantly; After compatibility with Radix Glycyrrhizae, Serum indexes were normal again, it explained Radix Glycyrrhizae had the effect of detoxification and reconciling the medicinal function; Liver biopsy results and serum index results was basic agreement. Metabolomics research showed the rat urine metabolites fingerprint classification is good between Fructus Cnidii group and control group, the research also find six possible biomarkers.Conclusion: Glycyrrhizic acid and Glycyrrhizin really had an impact on CYP450 enzyme activity, as glycyrrhizic acid was the major effective component in WSKCT, this research provided theoretical basis for the rational use of WSKCT. According to the toxicology research shown,the toxicity of Cortex Magnoliae Officinalis and Fructus Cnidii could reduce significantly after compatibility of Radix Glycyrrhizae, it proved that the formula of Wenshenkechuan tablets was reasonable; The potential biomarkers found by Metabonomics studies provided the possibility for toxicity mechanism of Fructus Cnidii.