The official compendial USP method for the determination of Diphenoxylate HCI (DPHCI) and Atropine Sulphate (ATSO4) involves extensive sample manipulation followed by non-aqueous titration for (DPHCI) and gas chromatography for (ATSO4). Furthermore, the assay for individual tablets (content uniformity) is not specific. The proposed HPLC methodology offers substantial improvement in sensitivity, specificity and speed. The method provides simultaneous separation with minimum sample manipulation. The total elution time is less than ten minutes. The accuracy of the method was validated by comparing the results with those obtained by applying the USP XX method on commercial tablets. The specificity of the method was confirmed by the results of content uniformity of DPHCI which were more accurate than those obtained by the USP method.