Determination of fentanyl in biological and water samples using single-drop liquid–liquid–liquid microextraction coupled with high-performance liquid chromatography Original Research Article

A single-drop liquid–liquid–liquid microextraction (LLLME) method coupled with high-performance liquid chromatography (HPLC) was developed for the determination of fentanyl in biological (plasma and urine) and wastewater samples. Fentanyl is a potent synthetic narcotic analgesic administered in the form of a transdermal patch for the management of chronic pain. Fentanyl was extracted from 0.01 M NaOH solution (donor phase) into a thin layer of organic phase (100 μL), then back-extracted into 5 μL of the acidic acceptor microdrop (1 × 10−3 M HClO4) immersed in the organic membrane from the tip of a 25-μL HPLC syringe. After the extraction, the microdrop was withdrawn into the syringe and injected directly into a HPLC system for analysis. The parameters influencing the extraction efficiency including the organic solvent and its volume, acceptor microdrop volume, composition of the donor and acceptor phases, stirring rate, temperature, salt addition and pre- and back-extraction times were investigated and optimized. At the most appropriate conditions (100 μL of n-octane, 3.6 mL of the donor phase maintained at 0.01 M NaOH, 5 μL of 1 × 10−3 M HClO4 as the acceptor microdrop, stirring rate of 1000 rpm for pre-extraction and 700 rpm for back-extraction, 30 °C, no salt addition, 30 min for pre-extraction and 20 min for back-extraction), an enrichment factor (EF) of 355 was obtained. The limit of detection (LOD) was 0.1 ng mL−1 (based on S/N = 3) and intra- and inter-day relative standard deviations less than 9% were obtained. The calibration graph was linear within the range of 0.5–1000 ng mL−1 with the correlation coefficient (r) of 0.9999. Finally, the feasibility of the proposed method was evaluated by extraction and determination of fentanyl in plasma, urine and wastewater samples and satisfactory results were obtained.