Clobazam and norclobazam quantitation in serum by capillary gas chromatography with electron-capture detection

Abstract

We have developed a simple, rapid method for determination of the antiepileptic drug clobazam and its major active metabolite, norclobazam in serum. Serum (200 microL) made alkaline with sodium borate buffer is extracted with toluene. After evaporation of the organic layer and reconstitution with toluene, the extract is analyzed within 5 min by capillary gas chromatography (Hewlett Packard HP 5890A GC; 63Ni electron-capture detector; HP-5 column). Linear calibrations for clobazam and norclobazam (clobazam: r = 0.999; norclobazam: r = 0.997) have permitted automated integrator calculation of results. Imprecision and accuracy were evaluated using an in-house control. Intra- and interassay coefficients of variation (CV) were 6.2% (n = 10) and 9.8% (n = 21), respectively for clobazam and 5.7% (n = 10) and 6.3% (n = 21), respectively for norclobazam. The mean concentrations for clobazam and norclobazam in the control were 3.5 mumol/L and 8.7 mumol/L respectively, representing 106% and 100% of the weighed-in values. Simplicity, specificity, sensitivity, and rapidity are attributes that make this assay suitable for monitoring of clobazam and norclobazam in serum.