Licorice root has been frequently used as antitode in traditional Chinese medicine. As the main active component of Licorice root, glycyrrhetic acid (GA) is mainly metabolized in liver. This study was designed to investigate the in vitro metabolism of GA by human liver microsomes (HLM) and human recombinant cytochrome P450 (CYP) isoforms. The results indicated that GA was metabolized mainly by CYP3A4. The K(m), V(max) and CL(int) of GA in HLM were 18.6 micromol x L(-1), 4.4 nmol x mg(-1) (protein) x min(-1) and 0.237 mL x mg(-1) (protein) x min(-1), respectively. At concentration up to 50 micromol x L(-1), GA inhibited CYP2C19, CYP2C9 and CYP3A4 enzyme activities with the inhibitory potencies up to 50%.
Aryl Hydrocarbon Hydroxylases ; antagonists & inhibitors ; metabolism ; Cytochrome P-450 CYP2C19 ; Cytochrome P-450 CYP2C9 ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 CYP3A Inhibitors ; Enzyme Inhibitors ; pharmacology ; Glycyrrhetinic Acid ; isolation & purification ; pharmacokinetics ; pharmacology ; Glycyrrhiza ; chemistry ; Humans ; Isoenzymes ; metabolism ; Microsomes, Liver ; enzymology ; metabolism ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry

<p>AIMTo identify the drug-metabolizing enzymes involved in the hydroxylation of the new anti-inflammatory and anodyne imrecoxib.p><p>METHODSImrecoxib was incubated with heterologous expression human cytochrome P450 (rCYPs) in vitro, and metabolites and remained parent drug were detected with liquid chromatography-multistage mass spectrometry. The contribution of 4 CYPs in the hydroxylation metabolism of imrecoxib was evaluated by total normalized rate (TNR) method.p><p>RESULTSImrecoxib is metabolized by CYP2C9, CYP2D6 and CYP3A4, with the rate of 62.5%, 21.1% and 16.4%, respectively.p><p>CONCLUSIONCYP2C9 is the major enzyme involved in imrecoxib hydroxylation metabolism.p>
Aryl Hydrocarbon Hydroxylases ; metabolism ; Cyclooxygenase 2 Inhibitors ; metabolism ; Cytochrome P-450 CYP2C9 ; Cytochrome P-450 CYP2D6 ; metabolism ; Cytochrome P-450 CYP3A ; Cytochrome P-450 Enzyme System ; metabolism ; Hydroxylation ; Pyrroles ; metabolism ; Spectrometry, Mass, Electrospray Ionization ; Sulfides ; metabolism

The metabolic characteristics of ligustrazin (TMPz) in liver microsomes were investigated in the present study. The reaction phenotyping of TMPz metabolism was also identified by in vitro assessment using recombinant human cytochrome P450 enzymes (CYP) and UDP glucuronosyltransferases (UGT). TMPz was incubated at 37 degrees C with human (HLM) and rat liver microsomes (RLM) in the presence of different co-factors. The metabolic stability and enzyme kinetics of TMPz were studied by determining its remaining concentrations with a LC-MS/MS method. TMPz was only metabolically eliminated in the microsomes with NADPH or NADPH+UDPGA. In the HLM and RLM with NADPH+UDPGA, t1/2, K(m) and V(max) of TMPz were 94.24 +/- 4.53 and 105.07 +/- 9.44 min, 22.74 +/- 1.89 and 33.09 +/- 2.74 micromol x L(-1), 253.50 +/- 10.06 and 190.40 +/- 8.35 nmol x min(-1) x mg(-1) (protein), respectively. TMPz showed a slightly higher metabolic rate in HLM than that in RLM. Its primary oxidative metabolites, 2-hydroxymethyl-3, 5, 6-trimethylpyrazine (HTMP), could undergo glucuronide conjugation. The CYP reaction phenotyping of TMPz metabolism was identified using a panel of recombinant CYP isoforms (rCYP) and specific CYP inhibitors in HLM. CYP1A2, 2C9 and 3A4 were found to be the major CYP isoforms involved in TMPz metabolism. Their individual contributions were assessed b) using the method of the total normalized rate to be 19.32%, 27.79% and 52.90%, respectively. It was observed that these CYP isoforms mediated the formation of HTMP in rCYP incubation. The UGT reaction phenotyping of HTMP glucuronidation was also investigated preliminarily by using a panel of 6 UGT isoforms (rUGT). UGT1A1, 1A4 and 1A6 were the predominant isoforms mediated the HTMP glucuronidation. The results above indicate that the metabolism of TMPz involves multiple enzymes mediated phase I and phase II reactions.
Animals ; Cytochrome P-450 CYP1A2 ; metabolism ; Cytochrome P-450 CYP2C9 ; metabolism ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 Enzyme Inhibitors ; Cytochrome P-450 Enzyme System ; metabolism ; Drug Interactions ; Glucuronosyltransferase ; metabolism ; Humans ; Ligusticum ; chemistry ; Microsomes, Liver ; enzymology ; NADP ; metabolism ; pharmacology ; Pyrazines ; metabolism ; pharmacokinetics ; Rats ; Uridine Diphosphate Glucuronic Acid ; metabolism ; pharmacology

Flavonoids are present in fruits, vegetables and beverages derived from plants, and in many dietary supplements or herbal remedies. A number of naturally occurring flavonoids have been shown to modulate the CYP450 system, including the induction or inhibition of these enzymes. This review focuses on the flavonoid effects on cytochrome P450 (CYP) enzyme CYP1, 2E1, 3A4 and 19. Flavonoids alter CYPs by various mechanisms, including the stimulation of gene expression via specific receptors and/or CYP protein, or mRNA stabilization and so on. But in vivo and in vitro, the effects of flavonoids are not always coincident as a result of concentrations of flavonoids, genetic and environmental factors. As well, flavonoids may interact with drugs through the induction or inhibition of their metabolism. Much attention should be paid to the metabolism interaction of the flavonoids when coadministered with other drugs.
Animals ; Aromatase ; genetics ; metabolism ; Cytochrome P-450 CYP1A1 ; antagonists & inhibitors ; genetics ; metabolism ; Cytochrome P-450 CYP2E1 ; genetics ; metabolism ; Cytochrome P-450 CYP2E1 Inhibitors ; Cytochrome P-450 CYP3A ; genetics ; metabolism ; Cytochrome P-450 CYP3A Inhibitors ; Cytochrome P-450 Enzyme Inhibitors ; Cytochrome P-450 Enzyme System ; genetics ; metabolism ; Enzyme Activation ; drug effects ; Flavonoids ; pharmacology ; Humans ; RNA, Messenger ; genetics ; metabolism

To study the enzyme kinetics of ligustilide metabolism and the effects of selective CYP450 inhibitors on the metabolism of ligustilide in liver microsomes of rat, a LC-MS method was established for quantitative analysis of ligustilide in liver microsomes incubation system with nitrendipine as internal standard. The determination m/z for ligustilide was 173, and for nitrendipine, 315. An optimum incubation system was found and various selective CYP inhibitors were used to investigate their inhibitory effects on the metabolism of ligustilide. The results showed that enzyme kinetics of ligustilide could be significantly inhibited by ketoconazole, trimethoprim and a-naphthoflavon but scarcely inhibited by omeprazole, 4-methylpyrazole and quinidine. Therefore, CYP3A4, CYP2C9 and CYP1A2 are the major isoenzyme participated in in vitro metabolism of ligustilide.
4-Butyrolactone ; analogs & derivatives ; metabolism ; Animals ; Benzoflavones ; pharmacology ; Cytochrome P-450 CYP1A2 ; Cytochrome P-450 CYP3A ; Cytochrome P-450 Enzyme Inhibitors ; Cytochrome P-450 Enzyme System ; Cytochromes ; antagonists & inhibitors ; Ketoconazole ; pharmacology ; Kinetics ; Male ; Microsomes, Liver ; metabolism ; Rats ; Rats, Sprague-Dawley ; Trimethoprim ; pharmacology

BACKGROUND AND PURPOSE: To compare the efficacy and safety of antiplatelet agents for the secondary prevention of ischemic stroke based on cytochrome P450 2C19 (CYP2C19) polymorphisms. METHODS: This study was a prospective, multicenter, randomized, parallel-group, open-label, blind genotype trial. First time non-cardiogenic ischemic stroke patients were enrolled and screened within 30 days. Participants were randomized to receive either triflusal or clopidogrel for secondary stroke prevention. The primary outcome was the time from randomization to first recurrent ischemic stroke or hemorrhagic stroke. RESULTS: The required sample size was 1,080 but only 784 (73%) participants were recruited. In patients with a poor CYP2C19 genotype for clopidogrel metabolism (n=484), the risk of recurrent stroke among those who received triflusal treatment was 2.9% per year, which was not significantly different from those who received clopidogrel treatment (2.2% per year; hazard ratio [HR], 1.23; 95% confidence interval [CI], 0.60–2.53). In the clopidogrel treatment group (n=393), 38% had good genotypes and 62% poor genotypes for clopidogrel metabolism. The risk of recurrent stroke in patients with a good CYP2C19 genotype was 1.6% per year, which was not significantly different from those with a poor genotype (2.2% per year; HR, 0.69; 95% CI, 0.26–1.79). CONCLUSIONS: Whilst there were no significant differences between the treatment groups in the rates of stroke recurrence, major vascular events, or coronary revascularization, the efficacy of antiplatelet agents for the secondary prevention of stroke according to CYP2C19 genotype status remains unclear.
Cytochrome P-450 CYP2C19 ; Cytochrome P-450 Enzyme System* ; Cytochromes* ; Genotype ; Humans ; Metabolism ; Platelet Aggregation Inhibitors ; Prospective Studies ; Random Allocation ; Recurrence ; Sample Size ; Secondary Prevention* ; Stroke*

The paper is aimed to study the metabolic characteristics of osthol (Ost) in isolated hepatocytes of rat to identify which isoforms of CYP450 were responsible for Ost metabolism in vitro. The concentration of Ost in isolated hepatocytes incubation system was determined by HPLC-UV. The effects of incubation time, substrate concentration and hepatocytes amount on the metabolic characteristics of Ost were investigated. CYP2C8 inhibitor quercetin (Que), CYP2C9 inhibitor sulfaphenazole (Sul), CYP2D6 inhibitor yohimbine (Yoh), CYP3A4 inhibitor troleandomycin (Tro) and CYP450 inducer rifampicin (Rif) were used to investigate their effects on the metabolism of Ost. The metabolism of Ost in isolated rat hepatocytes showed an enzymatic kinetic characteristics. Rif induced Ost elimination in rat hepatocytes; Yoh, Sul, Que did not have effects on Ost metabolism in vitro. Between 0-200 micromol x L(-1), Tro inhibited Ost metabolism in a concentration-dependent manner. CYP3A4 is the enzyme metabolizing Ost in vitro; CYP2C8, CYP2C9 and CYP2D6 did not involve in Ost metabolism in rat hepatocytes.
Animals ; Cells, Cultured ; Cnidium ; chemistry ; Coumarins ; isolation & purification ; metabolism ; Cytochrome P-450 CYP2D6 Inhibitors ; Cytochrome P-450 CYP3A ; Cytochrome P-450 Enzyme Inhibitors ; Cytochrome P-450 Enzyme System ; Hepatocytes ; metabolism ; Male ; Plants, Medicinal ; chemistry ; Quercetin ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Rifampin ; pharmacology ; Sulfaphenazole ; pharmacology ; Troleandomycin ; administration & dosage ; pharmacology ; Yohimbine ; pharmacology

Rats were continuously given different doses of water extract of Polygonum multiflorum (1, 10 g x kg(-1)) for 7 days to prepare liver microsomes. Cocktail in vitro incubation approach and Real-time quantitative PCR technology were used to observe the effect of water extract of P. multiflorum on CYP450 enzymatic activities and mRNA expressions in rat liver. Compared with the blank control group, both 1, 10 g x kg(-1) water extract of P. multiflorum treated groups showed significant inhibitions in CYP2E1 enzymatic activities and mRNA expressions (enzymatic activities of CYP2E1, P < 0.01; mRNA expression of CYP2E1, P < 0.05 in 1 g x kg(-1) group, P < 0.01 in 10 g x kg(-1) group). They revealed a significant increase in the enzymatic activity of CYP3A1 (P < 0.01), but without significant change in mRNA expressions. The 10 g x kg(-1) group showed a significant inhibition in CYP1A2 enzymatic activities and mRNA expressions in rat livers (P < 0.01).
Animals ; Cytochrome P-450 CYP1A2 ; genetics ; metabolism ; Cytochrome P-450 CYP2E1 ; genetics ; metabolism ; Cytochrome P-450 Enzyme Inhibitors ; administration & dosage ; isolation & purification ; Drugs, Chinese Herbal ; administration & dosage ; isolation & purification ; Liver ; drug effects ; enzymology ; Male ; Microsomes, Liver ; drug effects ; enzymology ; Polygonum ; chemistry ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley

Mesaconitine was incubated with rat liver microsomes in vitro. The metabolites of mesaconitine in rat liver microsomes were identified by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method with high resolution power. A typical reaction mixture of 100 mol L-1 Tris-HCI buffer (pH 7.4) containing 0.5 gL-1 microsomal protein and 50 micro molL-1 mesaconitine was prepared. The above reaction mixture was divided into six groups, and the volume of each group was 200 micro L. The incubation mixture was pre-incubated at 37 degrees C for 2 min and the reactions were initiated by adding NADPH generating system. After 90 min incubation at 37 degrees C, 200 micro L of acetonitrile was added to each group to stop the reaction. The metabolites of mesaconitine were investigated by UPLC-MS/MS method. Mesaconitine and 6 metabolites M1-M6 were found in the incubation system. The structures were characterized according to the data from MS/MS spectra and literatures. The metabolic reactions of mesaconitine in rat liver microsomes included the demethylation, deacetylation, dehydrogenation and hydroxylation. The major metabolic pathways of mesaconitine in rat liver microsomes were determined by UPLC-MS/MS on multiple reaction monitoring (MRM) mode combined with specific inhibitors of cytochrome P450 (CYP) isoforms, including alpha-naphthoflavone (CYP1A2), quinine (CYP2D), diethyldithiocarbamate (CYP2E1), ketoconazole (CYP3A) and sulfaphenazole (CYP2C), separately. Mesaconitine was mainly metabolized by CYP3A. CYP2C and CYP2D were also more important CYP isoforms for the metabolism reactions of mesaconitine, but CYP1A2 and CYP2E1 haven't any contribution to MA metabolism in rat liver microsomes.
Aconitine ; analogs & derivatives ; metabolism ; Animals ; Chromatography, High Pressure Liquid ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 CYP3A Inhibitors ; Cytochrome P-450 Enzyme Inhibitors ; Cytochrome P-450 Enzyme System ; metabolism ; Enzyme Inhibitors ; pharmacology ; Ketoconazole ; pharmacology ; Male ; Metabolic Networks and Pathways ; Microsomes, Liver ; enzymology ; metabolism ; Quinine ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Sulfaphenazole ; pharmacology ; Tandem Mass Spectrometry

Inhibition or/and induction of CYP3A4 are the major mechanisms underlying the common clinical drug-drug interactions, which has been gained attention in new drug discovery and development as well as clinical practice. Quantitative prediction of drug-drug interactions at the early stage of drug development is advantageous for reducing the cost and duration of development and providing more information for the later clinical studies. The review summarizes the update progress on quantitative prediction of in vivo drug-drug interactions derived from models based on in vitro inhibition or/and induction for CYP3A4.
Area Under Curve ; Cytochrome P-450 CYP3A ; metabolism ; Cytochrome P-450 CYP3A Inhibitors ; Drug Interactions ; Enzyme Inhibitors ; pharmacology ; Humans ; Intestines ; metabolism ; Liver ; metabolism ; Models, Biological ; Pharmaceutical Preparations ; administration & dosage ; metabolism