Simultaneous determination of dorzolamide and timolol by first-order derivative UV spectroscopy in simulated biological fluid for in vitro drug release testing

Abstract

Dorzolamide hydrochloride and timolol maleate is a well-established fixed-dose combination for the treatment of glaucoma worldwide. The utilization of simulated biological fluids can give a superior understanding of the release mechanisms and practicable in vivo nature of a dosage form that can improve the predictive potential of in vitro drug release testing. No method has been reported so far for the simultaneous estimation of dorzolamide and timolol in simulated tear fluid. In the present study, a simple, precise, and accurate first-order derivative ultraviolet spectrophotometric method for the routine analysis of dorzolamide and timolol in simulated tear fluid is proposed for in vitro drug release testing. The developed method was validated as per International Conference on Harmonization guidelines Q2 (R1). First-order derivative UV spectrophotometry was successfully applied to separate the overlapping peaks of dorzolamide and timolol by measuring peak amplitude at 251.80 nm and absorbance at 295.00 nm, respectively. The method was found to be accurate and precise, with a recovery range of 98.0 –101.0% and low relative standard deviations (<2.0%). The developed method was successfully applied for in vitro drug release testing of in-house in situ gel and marketed eye drops containing dorzolamide and timolol. Various mathematical models were adopted to fit the in vitro drug release profile. It was observed that the drug release of both drugs from the in situ gel and the marketed solution were best fitted by the Higuchi and first-order kinetic models, respectively. Moreover, the analysis of variance (ANOVA) provision was used for the validation of results. Overall, with the advantages of simple and fast operations, as well as reliability, the proposed method offers an ideal platform for routine analysis as compared to other high-cost and time-consuming chromatographic techniques. Having access to such a robust method will encourage the use of simulated tear fluid for in vitro drug release testing of ocular products and help to predict the in vivo performances of ophthalmic preparations.