The current study aimed to develop and validate a reverse-phase high-performance liquid chromatographic (RP-HPLC) method that is quick, easy, and sensitive for the simultaneous measurement of tretinoin and clindamycin phosphate in lipid-based semisolid pharmaceutical formulations, particularly microgel. Due to the difficulties involved in co-analysing a hydrophilic and a lipophilic drug inside the same matrix, a strong analytical technique was necessary for precise and repeatable quantification. Chromatographic separation was accomplished using a Phenomenex C-18 column (150 mm × 4.6 mm, 5 µm) and an isocratic mobile phase consisting of 0.01 N phosphate buffer and acetonitrile (20:80, v/v) at a flow rate of 1.0 mL/min. Clindamycin phosphate and tretinoin were detected at wavelengths of 210 and 353 nm, respectively. Standard calibration curves were created across a concentration range of 2–50 µg/mL for both analytes. The limits of detection (LOD) and quantification (LOQ) for clindamycin phosphate and tretinoin were 6.78 µg/mL and 20.56 µg/mL, respectively, and 0.14 µg/mL and 0.42 µg/mL, demonstrating the method's good linearity (R² = 0.99). With %RSD values constantly below 2%, recovery trials at three concentration levels (70%, 100%, and 120%) showed mean accuracy ranging from 98.18% to 102.93% for clindamycin phosphate and 116.80% to 123.36% for tretinoin. These results validated the accuracy, precision, and repeatability of the approach. This method effectively verified the entrapment efficiency of the drug-loaded microgel, meeting ICH requirements, and is suitable for regular quality control of combination formulations containing tretinoin and clindamycin.