Modified H-Point Standard Addition Method and Logarithmic Function for the Spectrophotometric and Spectrodensitometric Determination of Hesperidin and Diosmin in Mixtures

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Abstract
The similarity in the chemical structure of hesperidin and diosmin represents a difficulty in determining one or both of them when present together in pharmaceutical dosage forms. Different techniques were described in this article in order to determine each one of them without interference from the other or from other additives in the pharmaceutical dosage forms. The first method utilized the zero-order and first-derivative techniques for the determination of hesperidin at 290 nm in which diosmin has minimum absorbance value. The H-point standard addition method (HPSAM) was applied successfully in a concentration range from 5-40 μg ml−1, sensitivity 5μg ml−1 and detection limit 5μg. The HPSAM was also applied for the first derivative spectrophotometric determination of diosmin in the presence of hesperidin under the same concentration range, detection limit and sensitivity. The correlation coefficients(r) and regression equations for hesperidin and diosmin were found to be: 0.998, 0.994, respectively Y = 0.0226 x −0.0011 and Y = 0.2614x + 0.0643 for hesperidin and diosmin, respectively. The HPSAM was applied for the determination of hesperidin and diosmin in bulk powder mixture, in Daflon 500, and in Dioven tablets. The second method employed high performance thin layer chromatographic (HPTLC), plates for the separation of diosmin and hesperidin mixture or in pharmaceutical dosage forms, followed by spectrodensitometric measurement at 370nm. Linear relationships were obtained for hesperidin and diosmin in a concentration range from 1-10μg/spot, detection limit 0.5μg/spot and sensitivity 1μg. The range of measurements was extended to 50μg/spot utilizing the logarithmic mode. The regression equations and correlation coefficients were calculated in each case. Application of the proposed spectrodensitometric method was made to a mixture of bulk powder, and tablet dosage form with several advantages over the other published techniques. The suggested spectrophotometric, first-derivative and spectrodensitometric methods could be applied for routine analysis of mixtures of hesperidin and diosmin in the presence of closely related compounds as well as for the quality control purposes.