Green ratio-manipulating spectrophotometric methods for simultaneous determination of empagliflozin and sitagliptin in pharmaceutical formulations

BACKGROUND: Empagliflozin (EMP) and sitagliptin (SIT) are antidiabetic agents with complementary mechanisms of action and are frequently formulated as fixed-dose combinations. However, their substantially overlapping ultraviolet absorption spectra limit the applicability of direct spectrophotometric determination in combined dosage forms. OBJECTIVE: This study aimed to develop and validate simple, rapid, and low-solvent-consumption ratio-manipulating spectrophotometric methods for simultaneous determination of EMP and SIT in combined pharmaceutical formulations, and to assess their analytical, environmental, and practical performance. METHODS: Two mathematical spectrophotometric approaches, ratio difference (RD) and first-derivative ratio spectrophotometry (1DD), were developed. In the RD method, EMP was quantified using the amplitude difference between 230 and 285 nm, whereas SIT was determined from the amplitude difference between 257 and 209 nm. In the 1DD method, the first derivative of the ratio spectra was measured at 239 nm for EMP and 250 nm for SIT, using Δλ = 8 nm and a scaling factor of 80. The methods were validated for linearity, accuracy, precision, selectivity, robustness, and sensitivity. Their environmental and practical profiles were assessed using the Carbon Footprint Reduction Index (CaFRI), Complex Modified Green Analytical Procedure Index (ComplexMoGAPI), Click Analytical Chemistry Index (CACI), Blue Applicability Grade Index (BAGI), and Whiteness Evaluation for Chemical Analysis (WECA) tools. RESULTS: Both methods were linear over concentration ranges of 2-25 µg/mL for EMP and 5-100 µg/mL for SIT. For the RD method, the LOD values were 0.326 and 0.419 µg/mL, and the LOQ values were 0.978 and 1.257 µg/mL for EMP and SIT, respectively. For the 1DD method, the corresponding LOD values were 0.324 and 0.423 µg/mL, while LOQ values were 0.972 and 1.269 µg/mL for EMP and SIT, respectively. Mean recoveries ranged from 99.18% to 100.11%, with repeatability and intermediate-precision %RSD values not exceeding 0.761% and 0.979%, respectively. The proposed methods were successfully applied to commercial combined tablets, using ethanol as the analytical solvent. Student's t- and F-tests revealed no significant difference between the proposed and reported methods at the 95% confidence level. CONCLUSION: The developed RD and 1DD methods provide rapid, separation-free, and reliable alternatives for simultaneous spectrophotometric determination of EMP and SIT. Their use of ethanol, minimal sample preparation, low solvent consumption, and favorable analytical, environmental, and practical profiles support their suitability for routine pharmaceutical quality-control analysis, particularly in laboratories with limited chromatographic resources.