Simultaneous determination of codeine, norcodeine and morphine in biological fluids by high-performance liquid chromatography with fluorescence detection.

A novel high-performance liquid chromatographic method for the determination of codeine, norcodeine and morphine in plasma and urine has been developed. The compounds were separated on a cyano column (15 cm x 4.6 mm, 5 microns particle size) using a mobile phase of acetonitrile-triethylamine-distilled water (4:0.1:95.9, v/v) pH 3.1 and then determined by fluorescence detection. Calibration curves in the range 5-200 ng/ml for plasma and 0.1-10 micrograms/ml for urine were linear and passed through the origin. The imprecision and inaccuracy of the assay were less than 10% and the limits of detection were 2 ng/ml for all three compounds in human plasma.