An automatic 96-well solid phase extraction and liquid chromatography-tandem mass spectrometry method for the analysis of morphine, morphine-3-glucuronide and morphine-6-glucuronide in human plasma

A bioanalytical method using automated sample transferring, automated solid phase extraction (SPE) and liquid chromatography-tandem mass spectrometry (LC-MS-MS) was developed for morphine (MOR), and its metabolites morphine-3 -glucuronide (M3G) and morphine-6-glucuronide (M6G) in human plasma. Sample s of 0.25 ml were transferred into 96-well plate using automatic liquid han dler (Multiprobe(TM) II). Automated SPE was carried out on a 96-channel pro grammable liquid handling workstation (Quadra(TM) 96) using a C-18 sorbent. The extract was injected onto a silica column using an aqueous-organic mob ile phase. The chromatographic run time was 3.5 min per injection, with ret ention times of 1.5, 2.0 and 2.6 min for MOR, M6G, and M3G, respectively. T he detection was by monitoring MOR at m/z 286 --> 152, M6G and M3G at m/z 4 62 --> 286. The deuterated internal standards were monitored at m/z 289 --> 152 for MOR-d(3), and m/z 465 --> 289 for M6G-d(3) and M3G-d(3). The stand ard curve range was 0.5-50 ng ml(-1) for MOR, 1.0-100 ng ml(-1) for M6G, an d 10-1000 ng ml(-1) for M3G. The inter-day precision and accuracy of the qu ality control samples were <8% relative standard deviation (RSD) and <7% re lative error (RE) for MOR, <5% RSD and <2% RE for M6G, and <2% RSD and <4% RE for M3G. (C) 2002 Elsevier Science B.V. All rights reserved.