Biomonitoring of Oxycodone and Its Metabolites in Conventional and Nonconventional Biological Matrices: A Systematic Critical Review

BACKGROUND/PURPOSE: Oxycodone is widely prescribed for moderate-to-severe pain, but is associated with a high risk of misuse and overdose. Global data indicate an increase in opioid-related hospitalizations and fatalities, highlighting the need for effective therapeutic drug monitoring (TDM). This review systematically evaluates the analytical approaches and biomonitoring of oxycodone in conventional and nonconventional matrices across clinical and medicolegal contexts. METHODS: A systematic search of PubMed and Scopus (2015-2026) identified studies reporting oxycodone detection, quantification, and pharmacokinetics in human biological matrices. Blood, plasma, urine, hair, oral fluid, meconium, and cerebrospinal fluid were collected. Data on sample preparation, analytical techniques, detection limits, and applications in therapeutic, forensic, and emergency settings were extracted. RESULTS: Plasma, whole blood, and urine were the primary matrices for oxycodone monitoring, with liquid chromatography-tandem mass spectrometry (LC-MS/MS) as the gold standard. Microsampling strategies including dried blood spots and volumetric absorptive microsampling, enable minimally invasive collection with reliable plasma correlations. Urine profiling supports the simultaneous assessment of parent drugs and metabolites, enabling the evaluation of CYP3A4 and CYP2D6 activity. Nonconventional matrices (hair, oral fluid, meconium, and cerebrospinal fluid) offer complementary information for chronic, recent, prenatal, and central nervous system exposure. Several LC-MS-based methods with validated analytical workflows and certified reference materials are currently available, providing comprehensive quantification of oxycodone and its metabolites across all matrices. These workflows often incorporate advanced extraction approaches, such as solid-phase extraction, liquid-liquid extraction, and electromembrane-based methods, which, combined with high-throughput LC-MS/MS, enhance sensitivity, specificity, and analytical sustainability. Limits of quantification across matrices range from 0.1 to 1 ng/mL, confirming the robustness of current TDM and forensic applications. CONCLUSIONS: Current evidence supports a shift toward faster analytical methodologies and the complementary use of alternative biological matrices for oxycodone biomonitoring. As no single specimen is suitable for every clinical or forensic application, matrix selection should be tailored to a specific interpretative context. Harmonization of interpretative criteria and integration with clinical, pharmacogenomic, and artificial intelligence-based approaches may further enhance oxycodone TDM.