BIOANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF ANTIHYPERTENSIVE DRUGS IN RABBIT PLASMA USING RP-UPLC: A COMPREHENSIVE REVIEW

V. S. Mani Kumar Pinaka*, Rajiv Kukkar

ABSTRACT

Bioanalytical method development is an essential component of pharmaceutical research for the reliable quantification of therapeutic agents in biological matrices. Reverse-phase ultra-performance liquid chromatography (RP-UPLC) provides rapid separation, high chromatographic efficiency, reduced solvent consumption, and improved analytical throughput compared with conventional chromatographic approaches. This review discusses the development and validation of RP-UPLC methods for the determination of antihypertensive drugs in rabbit plasma. Particular emphasis is placed on biological sample preparation, protein precipitation, liquid–liquid extraction, solid-phase extraction, chromatographic optimization, internal-standard selection, detector selection, and validation according to ICH M10 requirements. Critical validation parameters including selectivity, specificity, calibration model, sensitivity, accuracy, precision, recovery, matrix effect, carryover, dilution integrity, and stability are discussed. Bioanalytical methods must demonstrate fitness for their intended purpose and generate reliable concentration data suitable for pharmacokinetic, toxicokinetic, bioavailability, bioequivalence, and other preclinical investigations (ICH, 2022). Recent literature indicates increasing use of chromatographic and hyphenated techniques for quantitative bioanalysis of antihypertensive agents, with particular attention to matrix effects, extraction efficiency, sensitivity, and regulatory compliance (Javid et al., 2025). RP-UPLC therefore represents a valuable analytical platform for rapid and reproducible bioanalysis of antihypertensive drugs in rabbit plasma.

Keywords: RP-UPLC, Bioanalytical Method Development, Method Validation, Antihypertensive Drugs, Rabbit Plasma, Pharmacokinetics, Matrix Effect, Recovery, ICH M10.