Development of a simple and sensitive HPLC-MS/MS method for determination of diazepam in human plasma and its application to a bioequivalence study.
10.12793/tcp.2017.25.4.173
- Author:
Do Hyung KIM
1
;
Ji Yoon CHO
;
Soo In CHAE
;
Bo Kyung KANG
;
Tae Gil AN
;
Wang Seob SHIM
;
Young Su NOH
;
Se Jung HWANG
;
Eun Kyoung CHUNG
;
Kyung Tae LEE
Author Information
1. Department of Life and Nanopharmaceutical Sciences, Graduate School, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemun-gu, Seoul 02447, Korea. ktlee@khu.ac.kr
- Publication Type:Original Article
- Keywords:
Liquid chromatography-tandem mass spectrometry;
human plasma;
method validation;
pharmacokinetic study;
diazepam
- MeSH:
Ammonium Compounds;
Diazepam*;
Humans*;
Liquid-Liquid Extraction;
Mass Spectrometry;
Methods*;
Plasma*;
Therapeutic Equivalency*;
Volunteers;
Voriconazole
- From:Translational and Clinical Pharmacology
2017;25(4):173-178
- CountryRepublic of Korea
- Language:English
-
Abstract:
We developed a simple, sensitive, and effective ultra-performance liquid chromatography/tandem mass spectrometry (HPLC-MS/MS) method with an electrospray ionization (ESI) interface in multiple reaction monitoring (MRM) and positive ion modes to determine diazepam concentrations in human plasma using voriconazole as an internal standard (IS). Diazepam and IS were detected at transition 285.2→193.1 and 350.2→127.1, respectively. After liquid-liquid extraction (LLE) using 1.2 ml of ethyl acetate:n-hexane (80:20, v/v), diazepam and IS were eluted on a Phenomenex Cadenza CD-C18 column (150 × 3.0 mm, 3 µm) with an isocratic mobile phase (10 mM ammonium acetate in water:methanol [5:95, v/v]) at a flow rate of 0.4 mL/min. The peak retention time was 2.32 min for diazepam and 2.01 min for IS, respectively. The lower limit of quantitation (LLOQ) was 0.5 ng/mL (S/N > 10) using 50 µL of plasma, and no interferences were observed in chromatograms. Our analytical method was fully validated and successfully applied to a bioequivalence study of two formulations of diazepam in healthy Korean volunteers.
