A Simple and Rapid HPLC-UV Method for Acetaminophen Quantification in Human Plasma: Enhancing Clinical Monitoring in Overdose Scenarios.

BACKGROUND: A rapid and cost-effective HPLC-UV method was developed and validated for the quantification of acetaminophen in human plasma. OBJECTIVE: To develop and validate a simple and rapid assay for acetaminophen quantification in human plasma, aimed at strengthening clinical monitoring, particularly in overdose scenarios. METHODS: The method utilizes isocratic elution with ultraviolet detection and a simple one-step protein precipitation using acetonitrile. This approach yields a short run time of approximately 8 minutes and requires a minimal injection volume of 20 μL. The assay provides a broad dynamic range (2–250 μg/mL), covering both therapeutic and toxic concentrations. RESULTS: The calibration curve exhibited excellent linearity (R² = 0.999), and method validation demonstrated acceptable precision (CV ≤ 15% for quality control samples and ≤ 20% at the lower limit of quantification) and accuracy (within ±15%, or ±20% at the LLOQ). Its clinical applicability was confirmed by successfully quantifying acetaminophen in 55 plasma samples from healthy volunteers receiving a therapeutic oral dose of a 1000 mg. Although no internal standard was used, the method’s rigorous validation and minimal matrix effects support its omission, in accordance with FDA, EMA, and ICH guidelines. CONCLUSION: This method offers a reliable and accessible alternative to more complex analytical techniques, with significant potential to support patient care in both therapeutic drug monitoring and overdose management. Further validation in overdose settings is required and will be pursued as part of future work.