Bioanalytical Method Development, Validation, and Quantification ofDutasteride in Rat Plasma by Reverse Phase HPLC

Introduction: Dutasteride, a 5α-reductase inhibitor, is widely prescribed for the treatment of benign prostatic hyperplasia. Although most analytical studies on dutasteride have utilized LCMS/MS techniques, there’s a timely need for a simple, accurate, and reliable alternative method. The objective of this study was to develop and validate a reverse-phase high-performance liquid chromatography (RP-HPLC) method for the quantification of dutasteride in rat plasma, intended for pharmacokinetic and bioavailability studies. Methods: A robust RP-HPLC method was developed and validated in accordance with ICH Q2(R2) guidelines. Plasma samples were processed using protein precipitation for extraction. Chromatographic separation was achieved on a Syncronis C18 column (250 mm × 4.6 mm, 5 µm), employing an isocratic mobile phase of methanol and water (80:20, v/v) at a flow rate of 1 mL/min. Dutasteride was eluted at a retention time of 3.191 minutes with a total run time of 6 minutes. Results: The method exhibited excellent linearity in the concentration range of 50–250 µg/mL, with a correlation coefficient (r²) of ≥ 0.9992. The lower limit of quantification (LOQ) was determined to be 6.39 µg/mL. Accuracy ranged between 90% and 110%, while inter-day precision was less than 2% relative standard deviation (RSD). The extraction recovery of dutasteride was greater than 90%, confirming method efficiency and robustness. Discussion: The validated method demonstrated high sensitivity, precision, and reproducibility, meeting all standard bioanalytical method validation requirements. Compared to LC-MS/MS, the RP-HPLC method offers a simpler, cost-effective, and efficient alternative for the routine quantification of dutasteride in plasma samples. Conclusion: A validated, efficient, and sensitive RP-HPLC method was successfully developed for the determination of dutasteride in rat plasma. The method is suitable for routine bioanalytical applications, including pharmacokinetic and bioavailability studies.