A validated, uncomplicated, and widely applicable HPLC-UV technique for quantifying cimetidine, famotidine, nizatidine, and ranitidine in human urine is elucidated. This represents the inaugural singular HPLC method documented for scrutinizing all four H2 antagonists in human biological specimens. Furthermore, this technique was employed for assessing ranitidine and its byproducts in human urine. All calibration plots exhibited strong linear correlation (r 2 > 0.9960) across the experimental ranges. The methodology demonstrated commendable precision and accuracy, manifesting overall intra- and inter-day fluctuations of 0.2–13.6% and 0.2–12.1%, respectively. The separation of ranitidine and its metabolites through this analysis notably enhanced resolution, precision, and accuracy in comparison to prior methodologies. Subsequently, the analysis was effectively implemented in a human volunteer investigation utilizing ranitidine as the prototype compound.