LC-MS/MS methods for synthetic opioids in urine: analytical and clinical evaluation

Opioids are potent analgesic compounds that exert their effects on the central nervous system and are widely used in clinical practice; however, their increasing misuse and associated health risks necessitate reliable and sensitive analytical monitoring. Urine remains the primary biological matrix for assessing compliance and detecting illicit opioid use. This review critically evaluates liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods for the determination of buprenorphine, methadone, tramadol, fentanyl, and pethidine in urine. Across the reviewed studies, diverse sample preparation strategies were employed, with calibration ranges typically reported at the ng/mL level. Limits of detection ranged from 0.005 to 10 ng/mL, while limits of quantification varied between 0.025 and 208 ng/mL. Analysis times ranged between approximately 2.5 and 24 min, reflecting differences in analytical throughput and chromatographic resolution. Sample preparation strategies, including solid-phase extraction, liquid-liquid extraction, and microextraction techniques, were critically compared in terms of recovery, matrix effects, and efficiency. Chromatographic separation predominantly relied on C18 columns with mobile phases containing formic acid, ammonium formate, or ammonium bicarbonate, and all methods applied positive electrospray ionization. Importantly, analytical trade-offs between opioid-specific assays and broader multi-analyte LC-MS/MS workflows were identified, particularly regarding sensitivity, metabolite coverage, and robustness against matrix effects. Overall, LC-MS/MS remains the most reliable confirmatory approach for urinary opioid analysis; however, challenges persist in the harmonization of analytical protocols, differentiation of structurally similar analogs, and standardization of matrix-effect evaluation. Future developments should focus on harmonization and integration of advanced analytical strategies to enhance clinical and forensic applicability.