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Measurement by HPLC of Midazolam and its Major Metabolite 1‐Hydroxymidazolam in Plasma of Very Premature Neonates
Author(s) -
Lee Toong Chow,
Charles Bruce
Publication year - 1996
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/(sici)1099-0801(199603)10:2<65::aid-bmc555>3.0.co;2-q
Subject(s) - chemistry , chromatography , midazolam , metabolite , high performance liquid chromatography , phosphoric acid , extraction (chemistry) , pharmacokinetics , calibration curve , detection limit , pharmacology , biochemistry , medicine , organic chemistry , sedation
A simple, selective high‐performance liquid chromatographic method with ultraviolet detection at 220nm is described for quantitation of midazolam and its primary metabolite 1‐hydroxymidazolam in 100μL plasma samples from premature infants. A mobile phase of acetonitrile:tetrahydrofuran:phosphate buffer (0.01 M , pH 6.7) (35:5:60 v/v) was pumped at 1 mL/min through a C 8 Symmetry TM (150×3.9 mm) column. Plasma (100μL) was extracted with 10% v/v isopropyl alcohol in dichloromethane containing 25 ng/mL climazolam (internal standard, IS) followed by back extraction into phosphoric acid (0.02 M ). 1‐Hydroxymidazolam, midazolam, and climazolam (IS) were eluted at 4.9, 7.4, and 8.4 min, respectively. Recoveries were >70%. Calibration curves in blank plasma were linear ( r >0.999) from 12.5 to 800 ng/mL. Within‐day and between‐day imprecision (CV%) was 1.8–6.5%, and 4.1–8.8%, respectively. Inaccuracy was 12.3%. Application of the method was demonstrated by a pharmacokinetic analysis of midazolam and 1‐hydroxymidazolam in plasma drawn from 15 premature neonates after a single intravenous dose of midazolam (0.1 mg/kg).