Introduction: There are so many promising plant based chemical constituents are present which act as alternative therapy for the control of diabetes. But due to lack of its proper quality control parameters they are not widely used. The US FDA patent is approved for fixed dosage combination of Mangiferin (MF) and Berberin HCl (BER) as antidiabetic herbal formulation. Objective: The literature review suggested that no UV spectroscopic method been reported in the literature review for the combination formulation of MF and BER. Methodology: Absorbance ratio method uses the ratio of absorbances at two selected wavelengths, one which is an iso-absorptive point and other being the λ-max of one of the two components. From the overlay spectra of two drugs, it is evident that MF and BER show an iso-absorptive point at 317 nm. The second wavelength used is 257 nm, which is the λ-max of MF. Methanol was used as a solvent in this method. The method was validated with respect to linearity, accuracy, precision as per the International conference on harmonisation (ICH) guidelines. Results: The drug response with respect to absorbance was linear over the concentration range 5-30 μg/ml for MF and 10-60 μg/ml for BER. The percentage recovery of MF and BER as found to be 100.00% and 100.07% respectively. Conclusion: The method can be successfully employed for the simultaneous determination of MF and BER in pharmaceutical formulations. The developed method is validated as per ICH guideline Q2(R1).