OBJECTIVES: Of several enzymes metabolizing xenobiotics, cytochrome P450 (CYP) and peroxidase enzymes seem to be most important. One of the major challenges in studies investigating metabolism of xenobiotics is to resolve which of these two groups of enzymes is predominant to metabolize individual xenobiotic compounds. Utilization of selective inhibitors of CYP and peroxidase enzymes might be a useful tool to identify the contribution of these enzymes to metabolism of xenobiotics in samples, where both types of enzymes are present. The aim of this study was to investigate specificities of several known CYP inhibitors to these enzymes; whether they inhibit only the CYP enzymes and do not inhibit peroxidases. METHODS: Since the oxidation of o-anisidine catalyzed by a model peroxidase used, horseradish peroxidase (HRP), is a two-substrate reaction, the inhibition potential of tested chemicals was studied with respect to both peroxidase substrates, o-anisidine and hydrogen peroxide. Initial velocities of o-anisidine oxidation by HRP under various conditions were determined spectrophotometrically. RESULTS: The CYP inhibitors metyrapone, troleandomycine, disulfiram, sulfaphenazole, quinidine and 1-aminobenzotriazole do not inhibit o-anisidine oxidation catalyzed by HRP. In contrast, ketoconazole, diethyldithiocarbamate, ellipticine, α-naphtoflavone, proadifen SKF525A, piperonylbutoxide, were found to inhibit not only the CYPs, but also the HRP-mediated oxidation of o-anisidine. Interestingly, α-naphtoflavone inhibits oxidation of o-anisidine by HRP with respect to H2O2, but not with respect to o-anisidine. Diethyldithiocarbamate is the most potent peroxidase inhibitor of o-anisidine oxidation with Ki with respect to o-anisidine of 10 μM and Ki with respect to H2O2 of 60 μM, being even the better peroxidase inhibitor than the classical "peroxidase inhibitor" - propyl gallate (Ki with respect to o-anisidine of 60 μM and Ki with respect to H2O2 of 750 μM). CONCLUSIONS: The results of the present study demonstrate that 1-aminobenzotriazole, a potent inhibitor of various CYP enzymes, seems to be the best candidate suitable for utilization in studies evaluating participation of CYP enzymes in metabolism of xenobiotics in various complex biological materials containing both CYP and peroxidase enzymes. Moreover, precaution to prevent misinterpretation of results is necessary in cases when proadifen SKF525A, piperonylbutoxide, diethyldithiocarbamate, ketoconazole, α-naphtoflavone and ellipticine are used in similar studies (as CYP inhibitors in various complex biological materials containing both CYP and peroxidase enzymes), since these chemicals can except of CYP enzymes inhibit also peroxidase-mediated reactions.

• MeSH
• aktivace enzymů účinky léků   MeSH
• benzoflavony chemie   farmakologie   MeSH
• chinidin chemie   farmakologie   MeSH
• disulfiram chemie   farmakologie   MeSH
• dithiokarb chemie   farmakologie   MeSH
• elipticiny chemie   farmakologie   MeSH
• inhibitory enzymů chemie   farmakologie   MeSH
• ketokonazol chemie   farmakologie   MeSH
• křenová peroxidasa antagonisté a inhibitory   metabolismus   MeSH
• lidé MeSH
• metyrapon chemie   farmakologie   MeSH
• piperonylbutoxid chemie   farmakologie   MeSH
• proadifen chemie   farmakologie   MeSH
• substrátová specifita účinky léků   MeSH
• sulfafenazol chemie   farmakologie   MeSH
• systém (enzymů) cytochromů P-450 antagonisté a inhibitory   MeSH
• triazoly chemie   farmakologie   MeSH
• troleandomycin chemie   farmakologie   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

The major aim of this work is to demonstrate the applicability of micellar electrokinetic capillary chromatography with SDS based pseudostationary phase for the screening of cytochrome P450 inhibitors. In contrast with the other capillary electrophoresis modes the cytochrome P450 reaction mixture thus could be used for the analysis without any pre-treatment. Cytochrome P450 2C9, one of the most important isoforms in human liver, was chosen as a model example for this study in combination with diclofenac as a probe substrate. The inhibitory effect on the given cytochrome P450 reaction was evaluated for two inhibitors with different inhibition potential - strong inhibitor sulfaphenazole and moderate inhibitor ketoconazole. As a result 50% inhibitory concentrations IC(50) and inhibition constants K(i) were evaluated; their values for both inhibitors were in a good agreement with the literature data determined by different methods.

• MeSH
• aromatické hydroxylasy antagonisté a inhibitory   chemie   MeSH
• chromatografie micelární elektrokinetická kapilární metody   MeSH
• diklofenak farmakologie   chemie   MeSH
• financování organizované MeSH
• ketokonazol farmakologie   chemie   MeSH
• lidé MeSH
• molekulární struktura MeSH
• reprodukovatelnost výsledků MeSH
• sulfafenazol farmakologie   chemie   MeSH
• systém (enzymů) cytochromů P-450 antagonisté a inhibitory   chemie   MeSH
• Check Tag
• lidé MeSH