A simple, accurate, green and selective high-performance thin-layer chromatography (HPTLC) method has been developed
and validated for the simultaneous estimation of empagliflozin, pioglitazone, and rosuvastatin in their synthetic ternary
mixture and different biological fluids. These three drugs are used for the treatment of type 2 diabetes mellitus and dyslipidemia
and have shown synergistic effects on cardiovascular outcomes. The ternary combination was separated on silica gel
TLC plates G60 F254,
utilizing a mixture of n-hexane‒ethyl acetate‒methanol‒glacial acetic acid in ratio (4.2:4:1.75:0.05,
V/V) as a developing system using ultraviolet (UV) detection at 230 nm. All experimental parameters were optimized with a
linearity range of 5‒250 ng per band for each drug, with good sensitivity and low limit of detection values reached, namely
1.72, 1.79, and 1.52 ng per band for empagliflozin, pioglitazone, and rosuvastatin, respectively. The developed method was
applied for separation of the studied drugs in their synthetic ternary mixture and different biological fluids, with good recovery
results ensuring high efficiency of the proposed approach. Eco scale, green analytical procedure index, and AGREE metric
tools were used to evaluate the greenness of the proposed method