Determination of 7-aminoflunitrazepam in urine by dispersive liquid-liquid microextraction with liquid chromatography-electrospray-tandem mass spectrometry.
Dispersive liquid-liquid microextraction (DLLME) and liquid chromatography-electrospray-tandem mass spectrometry (LC-ES-MS/MS) procedure was presented for the extraction and determination of 7-aminoflunitrazepam (7-aminoFM2), a biomarker of the hypnotic flunitrazepam (FM2) in urine sample. The method was based on the formation of tiny droplets of an organic extractant in the sample solution using water-immiscible organic solvent [dichloromethane (DCM), an extractant] dissolved in water-miscible organic dispersive solvent [isopropyl alcohol (IPA)]. First, 7-aminoFM2 from basified urine sample was extracted into the dispersed DCM droplets. The extracting organic phase was separated by centrifuging and the sedimented phase was transferred into a 300 microl vial insert and evaporated to dryness. The residue was reconstituted in 30 microl mobile phase (20:80, acetonitrile:water). An aliquot of 20 microl as injected into LC-ES-MS/MS. Various parameters affecting the extraction efficiency (type and volume of extraction and dispersive solvent, effect of alkali and salt) were evaluated. Under optimum conditions, precision, linearity (correlation coefficient, r(2)=0.988 over the concentration range of 0.05-2.5 ng/ml), detection limit (0.025 ng/ml) and enrichment factor (20) had been obtained. To our knowledge, DLLME was applied to urine sample for the first time.
Melwanki
Mahaveer B
MB
Department of Chemistry, Soochow University, Taipei, Taiwan.
Chen
Wei-Shan
WS
Bai
Hsin-Yu
HY
Lin
Tzuen-Yeuan
TY
Fuh
Ming-Ren
MR
D014151:Anti-Anxiety Agents; D002853:Chromatography, Liquid; D005445:Flunitrazepam / Q000031:analogs & derivatives* / Q000652:urine; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D014151:Anti-Anxiety Agents; D005445:Flunitrazepam; C036343:7-aminoflunitrazepam