[Identification of estazolam, alprazolam and triazolam in human urine by LC/MSn].
OBJECTIVE
To investigate the fragmentation behavior of triazolobenzodiazepines and to develop a specific, sensitive and rapid LC/MSn assay for simultaneous determination of estazolam, alprazolam and triazolam in human urine.


METHODS
After oral administration of a single 4 mg dose of the drugs to each of three healthy volunteers, urine samples were purified by solid-phase extraction, and then injected into an ODS column (150 mm x 4.6 mm) with a mobile phase of methanol-water (8:2) for LC/MSn analysis. The structures of estazolam, alprazolam and triazolam in human urine were identified by direct comparison of the observed mass spectra and the chromatographic retention time with those of the reference substance. The mass spectrometer (Finnigan LCQ) was operated in positive mode and in two scan modes including SIM and full scan MS/MS mode. The obtained mass spectra was analyzed assisted with the software Mass Frontier 1.0 for their fragmentation pathways.


RESULTS
The full scan MS/MS spectra of each compound gave characteristic fragment ions of [M + H - N2]+ and [M + H - Cl]+. The detection limit was below 0.5 ng.mL-1 for estazolam, alprazolam and triazolam in human urine.


CONCLUSIONS
The method is useful in forensic and clinical toxicology in which unequivocal identification of eatazolam, alprazolam and triazolam is desired.
Gu
Jing-kai
JK
Laboratory of Drug Metabolism, College of Life Science, Jilin University, Changchun 130023, China. gujingkai@163.net
Xia
Rong
R
Zhong
Da-fang
DF
Sun
Lu
L
D000525:Alprazolam / Q000652:urine*; D014151:Anti-Anxiety Agents / Q000652:urine*; D002853:Chromatography, Liquid; D004949:Estazolam / Q000652:urine*; D006801:Humans; D008297:Male; D021241:Spectrometry, Mass, Electrospray Ionization; D014229:Triazolam / Q000652:urine*
D014151:Anti-Anxiety Agents; D014229:Triazolam; D004949:Estazolam; D000525:Alprazolam