Evaluation of analytical procedures for urinary codeine and morphine measurements.

Different analytical procedures for quantitation of total morphine and codeine in urine were evaluated. Hydrolysis of urine with hydrochloric acid and four different enzymes was compared. In order to effect complete hydrolysis of glucuronide conjugates, hydrolysis with 6.5M HCl containing bisulphite and heating at 100 degrees C for 20 min was required. Results using this hydrolysis procedure agreed well with results from direct analysis of free and conjugated forms by liquid chromatography. Use of more dilute HCl but at a higher temperature was not sufficient. Some enzymatic procedures appeared to give effective hydrolysis with a higher variability, but these procedures were more time-consuming. There was a good quantitative agreement between FID gas chromatography and ion-trap gas chromatography/mass spectrometry. The validated analytical procedure was applied to authentic patient samples.