Simultaneous determination and quantification of 12 benzodiazepines in serum or whole blood using UPLC/MS/MS.
The benzodiazepines are a large, commonly prescribed family of psychoactive drugs. We describe a method permitting the simultaneous detection and quantification of 12 benzodiazepines in serum using ultra-performance liquid chromatography (UPLC) coupled with tandem mass spectrometry (MS/MS). Analytes included alprazolam, temazepam, oxazepam, nordiazepam, clonazepam, lorazepam, diazepam, chlordiazepoxide, midazolam, flunitrazepam, 7-aminoclonazepam, and 7-aminoflunitrazepam. Sample pretreatment is simple consisting of protein precipitation using cold acetonitrile (ACN) mixed with the deuterated internal standards. Samples were capped and vortexed for 5 min to ensure maximum precipitation. Following a 5-min centrifugation period, 400 microL of the supernatant was transferred to a clean tube and evaporated down under nitrogen. Samples were reconstituted in 200 microL of a deionized water:ACN (80:20) mixture and transferred to appropriate vials for analysis. Chromatographic run time was 7.5 min, and the 12 analytes were quantified using multiple reaction monitoring (MRM) and 6-point calibration curves constructed for each analyte at concentrations covering a clinically significant range.
Gunn
Josh
J
AIT Laboratories, Indianapolis, IN, USA.
Kriger
Scott
S
Terrell
Andrea R
AR
D000525:Alprazolam / Q000097:blood; D001569:Benzodiazepines / Q000097:blood*; D002707:Chlordiazepoxide / Q000097:blood; D002853:Chromatography, Liquid / Q000379:methods*; D002998:Clonazepam / Q000031:analogs & derivatives / Q000097:blood; D005445:Flunitrazepam / Q000031:analogs & derivatives / Q000097:blood; D006801:Humans; D008140:Lorazepam / Q000097:blood; D008874:Midazolam / Q000097:blood; D003708:Nordazepam / Q000097:blood; D010076:Oxazepam / Q000097:blood; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*
D001569:Benzodiazepines; C020860:7-aminoclonazepam; D002998:Clonazepam; D005445:Flunitrazepam; D003708:Nordazepam; D010076:Oxazepam; D002707:Chlordiazepoxide; C036343:7-aminoflunitrazepam; D008140:Lorazepam; D008874:Midazolam; D000525:Alprazolam