Routine determination of morphine, morphine 3-beta-D-glucuronide and morphine 6-beta-D-glucuronide in human serum by liquid chromatography coupled toelectrospray mass spectrometry

A robust liquid chromatographic mass spectrometric method capable of quanti fying morphine, morphine 3-beta-D-glucuronide and morphine 6-beta-D-glucuro nide down to 1.0 ng/ml, 5.0 ng/ml and 2.0 ng/ml respectively in human serum is presented. The method was validated over linear ranges of 1.0 to 20.0 n g/ml for morphine, 5.0 to 500.0 ng/ml for morphine 3-beta-D-glucuronide and 2.0 to 100.0 ng/ml for morphine 6-beta-D-glucuronide using deuterated morp hine as internal standard. In tandem mass spectrometry conditions, the prod uct ions of morphine-3-glucuronide and morphine-6-glucuronide were the ion mit corresponding to the morphine moiety. By contrast morphine which presen ted numerous product ions after collision did not allowed a tandem methodol ogy. Compounds were extracted on 100 mg C-18 columns and analysed on the PE Sciex API 300 system equipped with a C-18 column and electrospray ionisati on interface. The interrun precision of quality controls (1.0, 2.0, 10.0, 2 0.0 ng/ml for morphine, 5.0, 10.0, 250.0, 500.0 ng/ml for morphine 3-beta-D -glucuronide and 2.0, 4.0, 50.0, 100.0 ng/ml for morphine 6-beta-D-glucuron ide) was less than or equal to 9.3% and accuracy was between 97.9 and 109.8 % for each analyte. Sample stabilities in biological matrix were also inves tigated. This method has been applied to pharmacokinetic analysis of morphi ne, morphine 3-beta-D-glucuronide and morphine 6-beta-D-glucuronide in huma n serum. (C) 1999 Elsevier Science BN. All rights reserved.