title: Liquid chromatography-tandem mass spectrometry for detection of low concentrations of 21 benzodiazepines, metabolites, and analogs in urine: method with forensic applications.
abstract: BACKGROUND
  Commonly used methods for detecting benzodiazepines (BZPs) and BZP-like substances, such as zolpidem and zopiclone, may not detect low concentrations of these drugs. We developed a liquid chromatographic-tandem mass spectrometric method for identifying these drugs and their relevant metabolites.
  
  
  METHODS
  We extracted BZPs from urine by solid-phase extraction with a mixed-mode phase (OASIS HLB cartridges). Chromatographic separation was performed with a Waters XTerra MS C18 [150 x 2.1 mm (i.d.); bead size, 5 microm] reversed-phase column with deuterated analogs of the analytes as internal standards (IS). Detection was performed with a triple-quadruple mass spectrometer that monitored 2 specific transitions per compound in the electrospray, positive-ion selected-reaction monitoring mode. We tested this technique on urine samples from 12 healthy volunteers and 1 forensic sample obtained in a case of alleged drug-facilitated sexual assault.
  
  
  RESULTS
  Chromatographic separation was achieved within 18 min. The linear dynamic ranges extended from 0.02 or 0.1 microg/L (depending on the drug or metabolite) to 50 microg/L. Extraction recovery (range) was 77%-110%. Limits of detection were < or = 0.05 microg/L. No ion suppression was seen except for alprazolam, for which baseline decreased by almost 20%. In the forensic urine sample, the method detected alprazolam (3.5 microg/L) and its characteristic metabolite, alpha-hydroxyalprazolam (0.17 microg/L).
  
  
  CONCLUSIONS
  This method measured low concentrations of BZPs and BZP-like substances and might be useful for analyses of urine in suspected drug-facilitated sexual assault cases.
authors.affiliation: Department of Pharmacology-Toxicology, Limoges University Hospital, Limoges, France.
authors.forename: Oscar
authors.initials: O
authors.lastname: Quintela
authors.forename: François-Ludovic
authors.initials: FL
authors.lastname: Sauvage
authors.forename: Fabienne
authors.initials: F
authors.lastname: Charvier
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Gaulier
authors.forename: Gérard
authors.initials: G
authors.lastname: Lachâtre
authors.forename: Pierre
authors.initials: P
authors.lastname: Marquet
mesh_terms: D053961:Azabicyclo Compounds; D001569:Benzodiazepines / Q000378:metabolism* / Q000652:urine*; D002851:Chromatography, High Pressure Liquid; D005554:Forensic Medicine / Q000379:methods; D006801:Humans; D006993:Hypnotics and Sedatives / Q000378:metabolism* / Q000652:urine*; D010879:Piperazines / Q000378:metabolism / Q000652:urine; D011725:Pyridines / Q000378:metabolism / Q000652:urine; D021241:Spectrometry, Mass, Electrospray Ionization; D015813:Substance Abuse Detection / Q000379:methods*; D000077334:Zolpidem
chemical_list: D053961:Azabicyclo Compounds; D006993:Hypnotics and Sedatives; D010879:Piperazines; D011725:Pyridines; C515050:zopiclone; D001569:Benzodiazepines; D000077334:Zolpidem