Highly sensitive gas chromatographic mass spectrometric method for morphine determination in plasma that is suitable for pharmacokinetic studies

A sensitive method was devised to determine morphine plasma concentrations by gas chromatography-mass spectrometry (GC-MS) using selected ion monitoring (SIM) with nalorphine as the internal standard. This method was rugged, reliable, selective and sensitive and was used to determine the morphine content of over 2000 samples. Sample preparation involved extraction of basified sample using n-butyl chloride-chloroform (5:1) and evaporation of the extract to dryness. The residue was derivatised with pentafluoropropionic anhydride, evaporated to dryness, reconstituted in 40 mu l toluene and injected onto the GC-MS. For a sample size of 1 ml, the limit of quantitation was 0.75 ng/ml (SIN ratio 10:1) and the estimated limit of detection was calculated to be 0.2 ng/ml (SIN ratio 3:1), expressed as morphine base, Precision (n=5) was 4.9% at 0.75 ng/ml, 6.8% at 1.5 ng/ml, 3.0% at 37.5 ng/ml and 2.3% at 150 ng/ml. Standard curves for the range of 0-750 ng/ml morphine in plasma were linear with all r(2) values greater than 0.99. No interfering peaks were seen for either morphine or internal standard in the blank samples. The method is suitable for pharmacokinetic studies after subclinical doses of morphine where it has been used to study morphine plasma concentrations for 6 h after a dose of only 2 mg.