Determination of clozapine and its major metabolites in human plasma and red blood cells by high-performance liquid chromatography with ultraviolet absorbance detection.
An isocratic high-performance liquid chromatographic (HPLC) method with UV absorbance detection is described for the quantification of clozapine (8-chloro-11-(4'-methyl)piperazino-5H-dibenzo[b,e]-1,4-diazepine) and its two major metabolites in plasma and red blood cells (RBCs). The method involves sample clean-up by liquid-liquid extraction with ethyl acetate. The organic phase was back-extracted with 0.1 M hydrochloric acid. Loxapine served as the internal standard. The analytes were separated by HPLC on a Kromasil Ultrabase C18 analytical column (5 microns particle size; 250 x 4.6 mm I.D.) using acetonitrile-phosphate buffer pH 7.0 (48:52, v/v) as eluent and were measured by UV absorbance detection at 254 nm. The limits of quantiation were 20 ng/ml for clozapine and N-desmethylclozapine and 30 ng/ml for clozapine N-oxide. Recovery from plasma or RBCs proved to be higher than 62%. Precision, expressed as % C.V., was in the range 0.6-15%. Accuracy ranged from 96 to 105%. The method's ability to quantify clozapine and two major metabolites simultaneously with precision, accuracy and sensitivity makes it useful in therapeutic drug monitoring.
Guitton
C
C
Laboratoire de Pharmacocinétique, Hôpital Carémeau, Nîmes, France.
Kinowski
J M
JM
Aznar
R
R
Bressolle
F
F
D014150:Antipsychotic Agents / Q000097:blood*; D002851:Chromatography, High Pressure Liquid; D003024:Clozapine / Q000031:analogs & derivatives / Q000097:blood*; D004355:Drug Stability; D004912:Erythrocytes / Q000737:chemistry*; D006801:Humans; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet
D014150:Antipsychotic Agents; C058272:norclozapine; D003024:Clozapine; C079149:clozapine N-oxide