title: An ultra-sensitive LC-MS/MS method to determine midazolam levels in human plasma: development, validation and application to a clinical study.
abstract: OBJECTIVE
  Midazolam is a commonly used marker substrate for the in vivo assessment of CYP3A activity. Reliable pharmacokinetic assessment at sub-pharmacological doses of midazolam requires an ultra-sensitive analytical method.
  
  
  METHODS
  A new, ultra-sensitive LC-MS/MS method for the determination of midazolam in human plasma using SPE was developed and fully validated. The lowest limit of quantitation is 0.1 pg/ml with a sample volume of 500 μl.
  
  
  CONCLUSIONS
  The following parameters were validated: sensitivity, assay accuracy and precision, linearity, selectivity, and stability of midazolam at pertinent analytical and storage conditions. The validated method was utilized successfully for the sample assay during a midazolam microdosing study for the evaluation of CYP3A4 activity of a clinical candidate.
authors.affiliation: Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Mu
authors.initials: M
authors.lastname: Chen
authors.affiliation: Roche Innovation Center Shanghai, 720 Cai Lun Road, Building 5/917 Ha Lei Road, Building 6, Pudong, Shanghai 201203, China.
authors.forename: Wenzhe
authors.initials: W
authors.lastname: Lu
authors.affiliation: Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Yang
authors.initials: Y
authors.lastname: Lu
authors.affiliation: Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Lijuan
authors.initials: L
authors.lastname: Kang
authors.affiliation: Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Harry
authors.initials: H
authors.lastname: Zhao
authors.affiliation: Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Zhongping John
authors.initials: ZJ
authors.lastname: Lin
authors.affiliation: Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.
authors.forename: Weimin
authors.initials: W
authors.lastname: Wang
authors.affiliation: Roche Innovation Center Switzerland, F. Hoffmann-La Roche Ltd, Grenzacherstrasse 124, CH-4070 Basel, Switzerland.
authors.forename: Daniela
authors.initials: D
authors.lastname: Fraier
authors.affiliation: Roche Innovation Center Shanghai, 720 Cai Lun Road, Building 5/917 Ha Lei Road, Building 6, Pudong, Shanghai 201203, China.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Ottaviani
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D006801:Humans; D008874:Midazolam / Q000097:blood*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D008874:Midazolam
keywords: CYP3A4; LC–MS/MS; Phase 0 clinical study; SPE; microdosing; midazolam; ultra sensitive