مرکز منطقه ای اطلاع رساني علوم و فناوري -

چكيده لاتين :

Determination of the kinetics of mechanism-based enzyme inactivation (MBI) involves a pre-incubation followed by an incubation stage. To minimise inhibition during the second stage it is recommended to dilute the concentration of inhibitor and to use a high concentration of probe substrate (e.g. 10 fold > Km). The aim of this study was to quantify the impact of changing the study design. Two different dilutions of the pre-incubation samples (1.25 and 4 times) and different concentrations of the probe substrate dextromethorphan (5, 10 and 20 |nM) were used to determine the kinetic constants describing the MBI of CYP2D6 by methylenedioxymetham-phetamine. Changing the concentration of dextromethorphan had no significant effect on either KI or kinact values (P> 0.05). This is expected when the substrate concentration is well in excess of its Km. KI and kinact values increased as expected (3.81 to 12.92 (Mu)M (P< 0.01) and 0.22 to 0.29 min-1 (P< 0.02), respectively) as the dilution was increased. These data provide evidence that study design is important in characterising MBI.