Sensitive assay for midazolam and its metabolite 1'-hydroxymidazolam in human plasma by capillary high-performance liquid chromatography.
A sensitive high-performance liquid chromatographic method is described for the quantification of midazolam and 1'-hydroxymidazolam in human plasma. Sample (1 ml plasma) preparation involved a simple solvent extraction step with a recovery of approximately 90% for both compounds. An aliquot of the dissolved residue was injected onto a 3 microm capillary C18 column (150 mm x 0.8 mm I.D.). A gradient elution was used. The initial mobile phase composition (phosphate buffer-acetonitrile, 65:35) was maintained during 16 min and was then changed linearly during a 1-min period to phosphate buffer-acetonitrile, 40:60. The flow-rate of the mobile phase was 16 microl/min and the eluate was monitored by UV detection. The limits of quantification for midazolam and 1'-hydroxymidazolam were 1 ng/ml and 0.5 ng/ml, respectively. The applicability of the method was demonstrated by studying the pharmacokinetics of midazolam, and its major metabolite 1'-hydroxymidazolam, in human volunteers following i.v. bolus administration of a subtherapeutic midazolam dose (40 microg/kg).
Laboratory of Pharmacokinetics (FATC 7355), School of Pharmacy, Catholic University of Louvain, Brussels, Belgium.
S L
SL
Eeckhoudt
J P
JP
Desager
Y
Y
Horsmans
A J
AJ
De Winne
R K
RK
Verbeeck
D001189:Aryl Hydrocarbon Hydroxylases; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D051544:Cytochrome P-450 CYP3A; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000008:administration & dosage; D005069:Evaluation Studies as Topic; D006801:Humans; D008874:Midazolam / Q000031:analogs & derivatives* / Q000097:blood* / Q000493:pharmacokinetics; D010089:Oxidoreductases, N-Demethylating / Q000037:antagonists & inhibitors / Q000378:metabolism; D012293:Rifampin / Q000008:administration & dosage; D012680:Sensitivity and Specificity
D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System; C040081:1-hydroxymethylmidazolam; D001189:Aryl Hydrocarbon Hydroxylases; D051544:Cytochrome P-450 CYP3A; D010089:Oxidoreductases, N-Demethylating; D008874:Midazolam; D012293:Rifampin