本文建立了一种全血样品中13种苯二氮卓类安眠镇静药物及其代谢产物地西泮、硝西泮、溴西泮、氟西泮、氯硝西泮、氟硝西泮、劳拉西泮、奥沙西泮、普拉西泮、替马西泮、7-氨基硝基西泮、7-氨基氟硝西泮和氯氮卓的超高效液相色谱-串联质谱（UPLC-MS/MS）检测方法。全血样品经过Ostro96孔磷脂过滤板提取,采用电喷雾离子源正离子（ESI+）模式和多反应检测（MRM）模式进行质谱分析,13种化合物在0.2~20 ng/mL浓度范围内均获得良好的线性,该方法的提取回收率分布在65.2%~113.9%之间,最小检测限可达0.008~0.15 ng/mL。本方法灵敏、简便、快速、高通量,适用于全血样本中痕量苯二氮卓类安眠镇静类药物的定性定量分析。

A rapid and sensitive method has been developed for simultaneous determination of 13 benzodiazepines and metabolites including diazepam, nitrazepam, bromazepam, flurazepam, clonazepam, flunitrazepam, lorazepam, oxazepam, prazepam, temazepam, 7-amino-nitragepam, 7-aminoflunitrazepam and chlordiazepam in whole blood samples by ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS/MS). The separations were carried out on an ACQULTY UPLC BEH C18 column (2.1 mm ×100 mm, 1.7 μm) at 35°C and with a constant flow rate of 0.5 mL/min and the gradient-elution consisting of acetonitrile and 5 mmoL ammonium hydrogen carbonate buffer. The MS parameters were set as capillary voltage (3 kV), desolvation gas temperature (550°C), desolvation gas flow (800 L/h), cone gas flow (150 L/h) and collision gas flow (0.15 mL/min). Ostra^TM 96-well plate was used to extract the samples. 100 μL of human whole blood containing target drugs and D5-diazepam as internal standard was transferred into the Ostra^TM 96-well plate, followed by 400 μL of 0.1% formic acid in acetonitrile. Phospholipids were removed from the whole blood via the Ostra^TM 96-well plate effectively, and the filtrate was collected under a constant pressure of 20 psi. Thereafter, the 96-well receiving plate was capped and placed on the suitable position in sample manager and was analyzed with UPLC-MS/MS system coupled with an electrospray ionization (ESI) source. The samples were measured in the mode of electraspray positive ionization (ESI+) and multiple reaction monitoring (MRM). 13 benzodiazepines and metabolites were linear in the range of 0.2~20 ng/mL with coefficients higher than 0.99. The extraction recoveries ranged from 65.2% to 113.9% for all analytes, and the limits of detection (LODs) were 0.008~0.15 ng/mL. The method presented here is validated to be simple, rapid, sensitive and accurate for the determination of trace benzodiazepines and metabolites in whole blood samples.