Preconcentration of trace amounts of methadone in human urine, plasma, saliva and sweat samples using dispersive liquid–liquid microextraction followed by high performance liquid chromatography

A simple, rapid and efficient method for the preconcentration of methadone was developed using dispersive liquid–liquid microextraction (DLLME) followed by high performance liquid chromatography with ultra violet detection (HPLC–UV). The extraction method is based on the fast injection of a mixture of extracting and disperser solvents into the aqueous solution to form a cloudy ternary component solvent (aqueous solution:extracting solvent:disperser solvent) system. The extraction parameters such as nature and volume of extracting and disperser solvents, pH of sample, and extraction time were studied for optimization. Under the optimal conditions (extracting solvent: chloroform, 250 μL; disperser solvent: methanol, 2.5 mL and pH of sample: 10.0) a linear calibration curve was obtained in the range of 0.5–5000 ng mL−1 with r2 = 0.9995. To demonstrate analytical performance, figures of merits of the proposed method in four different biological matrices (urine, plasma, saliva and sweat) spiked with methadone were investigated. The limits of detection and quantification in these matrices were ranged from 4.90 to 24.85 ng mL−1 and 16.32 to 82.75 ng mL−1, respectively. The extraction recoveries were above 97% and the preconcentration factors of methadone in distilled water, urine, plasma, saliva, and sweat samples were 196.52, 10.03, 9.93, 1.97 and 1.99, respectively. While the precision for inter-day was ≤6.43 (n = 5), it was ≤2.26 (n = 5) for intra-day assay. Finally, the method was successfully applied in the determination of methadone in the human urine, plasma, saliva and sweat samples.

► DLLME–HPLC–UV has been developed for rapid determination of methadone.
► Methadone was determined in urine, plasma, saliva and sweat samples.
► In the cases of urine and plasma samples, lower LODs were attained.
► The low sample volume makes the procedure suitable for forensic investigations.