High-throughput determination of barbiturates in human plasma using on-line column-switching ultra-fast liquid chromatography-tandem mass spectrometry
High-throughput determination of barbiturates in human plasma using on-line column-switching ultra-fast liquid chromatography-tandem mass spectrometry
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DOI：
10.1007/s11419-012-0155-4
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发表时间：
2013-01-01
影响因子：
2.2
通讯作者：
Sato, Keizo
中科院分区：
文献类型：
--
作者：
Lee, Xiao-Pen;Kumazawa, Takeshi;Sato, Keizo
开发了一种通过在线柱切换超快液相色谱-串联质谱 (MS-MS) 测定人血浆中八种巴比妥类药物（巴比妥、阿洛巴比妥、苯巴比妥、环巴比妥、异戊巴比妥、司可巴比妥、硫喷妥钠和硫戊醛）的高通量方法。将掺有八种巴比妥酸盐和 5-(4-甲基苯基)-5-苯基乙内酰脲（内标）的血浆样品（100 μl）用 300 μl 13.3 mM 乙酸铵/乙腈（33:67，v/v）稀释。离心和过滤后，将澄清的上清液直接注入萃取柱（Oasis HLB 柱）。以下过程是完全自动化的。通过反冲萃取柱洗脱保留在萃取柱上的分析物，并通过柱切换将其引入分析柱（Phenomenex Onyx 整体式 C-18 柱，100 mm x 4.6 mm 内径）。通过负离子大气压化学电离的多重反应监测进行定量。在 3 分钟（包括萃取时间）的分析时间内，八种药物均实现了良好的峰分离和峰形。所有添加到血浆中的药物的回收率为 80-93%。八种药物的回归方程在血浆 10-5000 ng/ml 范围内表现出良好的线性，检测限范围为 1.0 至 10 ng/ml。定量下限和上限分别为10-50 ng/ml和5000 ng/ml。 Intraday and interday coefficients of variation for all the drugs were not > 9.1 %.定量准确度为92.0-108%。该方法成功应用于测定志愿者口服后人血浆中异戊巴比妥的水平。
A high-throughput method was developed for determinations of eight barbiturates (barbital, allobarbital, phenobarbital, cyclobarbital, amobarbital, secobarbital, thiopental, and thiamylal) in human plasma by on-line column-switching ultra-fast liquid chromatography-tandem mass spectrometry (MS-MS). Plasma samples (100 mu l) spiked with the eight barbiturates and 5-(4-methylphenyl)-5-phenylhydantoin (internal standard) were diluted with 300 mu l of 13.3 mM ammonium acetate/acetonitrile (33:67, v/v). After centrifugation and filtration, the clear supernatant was injected directly onto the extraction column (Oasis HLB cartridge column). The following procedure was fully automated. The analytes retained on the extraction column were eluted by backflushing of the extraction column and introduced onto the analytical column (Phenomenex Onyx monolithic C-18 column, 100 mm x 4.6 mm i.d.) by column switching. Quantification was performed by multiple reaction monitoring with negative-ion atmospheric pressure chemical ionization. Good peak separation and peak shapes of the eight drugs were achieved within an analysis time of 3 min, including the extraction time. All drugs spiked into plasma showed recoveries of 80-93 %. The regression equations for the eight drugs showed excellent linearities in the range of 10-5000 ng/ml of plasma, and the limits of detection ranged from 1.0 to 10 ng/ml. The lower and upper limits of quantitation were 10-50 ng/ml and 5000 ng/ml, respectively. Intraday and interday coefficients of variation for all the drugs were not > 9.1 %. The accuracies of quantitation were 92.0-108 %. The method was successfully applied to determination of the level of amobarbital in human plasma after its oral administration to a volunteer.