Determination of flavonoids in pharmaceutical preparations using Terbium sensitized fluorescence method

Background and the Purpose of the Study: The aim of this study was development and validation of a simple, rapid and sensitive spectrofluorimetric method for determination of total flavonoids in two topical formulations of Calendula officinalis, Ziziphus Spina-christi and an oral drop of Hypiran perforatum L. The proposed method is based on the formation of terbium (Tb3+)-flavonoids (quercetin as a reference standard) complex at pH 7.0, which has fluorescence intensely with maximum emission at 545 nm when excited at 310 nm. Method: For ointments masses of topical formulations were weighed and added to ethanol-aqueous buffer (pH 10.0) and the resulting mixtures were shaken and then two phases were separated by centrifugation. Aqueous phases were filtered and then diluted with water. For Hypiran drops an appropriate portion was diluted with ethanol and then aliquots of sample or standard solutions were determined according to the experimental procedure. Results: Under the optimum conditions, total concentrations of flavonoids (as quercetin equivalent) in three tested formulations were found to be 0.204 mg/g (for Dermatin cream), 0.476 mg/g (for Calendula ointment) and 13.50 mu g/ml (for Hypiran drops). Analytical recoveries from samples spiked with different amounts of quercetin were 96.1-104.0 % with RSD % of less than 3.5. Conclusion: The proposed method which requires a simple dissolution step without any matrix interferences provided high sensitivity and selectivity and was easily applied to determine total flavonoids in real samples of three investigated formulations with excellent reproducibility.