To develop and validate a HPTLC method for the simultaneous quantification of aegelin and skimmianine in the barks of Aegle marmelos (L.) Corr. (Rutaceae) for the first time. The HPTLC system (CAMAG, Muttanz, Switzerland) with WinCATS software under window XP was used for the analysis of constituents. Separation of each constituents were obtained with the help of HPTLC plates pre-coated with silica gel 60F-254 plates (10×10 cm2) and solvent system of toluene-ethyl acetatate (7: 3, v/v). Scanning wavelength 262 nm was used for analysis in reflection/absorption mode. The developed method was validated for linearty, LOD, LOQ, accuracy and precision. Well defined spots were obtained in the solvent system of toluene: ethyl acetate (7:3,v/v).The linearity range for aegelin and skimmianine were found to be 1-12 μg/spot with correlation coefficients of 0.996±0.0054 and 0.987±0.0046, respectively, which were indicative of good linear dependence of peak area on concentration. Limit of detection (LOD) (0.101 ng and 0.125 ng for aegelin and skimmianine), limit of quantification (LOQ) (0.639 ng and 0.736 ng for aegelin and skimmianine), accuracy (% recovery 94.45 ±0.984 for aegelin and 95.76 ± 1.103 for skimmianine), and precision (CV< 5%, for both intra-day and inter-day precisions). The method was found to be reproducible and convenient for quantitative analysis of these compounds in Aegle marmelos (L.) Corr.