Ecofriendly single-step HPLC and TLC methods for concurrent analysis of ternary antifungal mixture in their...

Two accurate, sensitive, and selective methods for simultaneous determination of miconazole nitrate (MIC), nystatin (NYS), and metronidazole (MET) in pure state or drug product were established and verified. First, RP-HPLC-DAD was designed. Separation was accomplished using a ZOBRAX Eclipse Plus RP-C₈ column that was running under an isocratic elution of methanol: 0.05% aqueous solution of sodium dodecyl sulphate (40: 60 v/v), with a flow rate that was regulated at 0.8 mL/min. The column temperature was adjusted at 25 °C and diode array detector was monitored at 220 nm. The linearity range of the proposed method was achieved at the concentration of 5-50, 4-50, and 4-40 µg/mL and the attained retention time for the studied drugs was 2.52, 3.52 and 4.99 min for MIC, NYS, and MET, correspondingly. Second, a TLC-densitometric approach was used to resolve the three compounds. Resolution of the three cited drugs was carried out using TLC aluminum plates pre-coated with 0.25 mm silica gel 60 F₂₅₄. A developing solvent comprised ethyl acetate: toluene: methanol: triethyl amine: formic acid (3: 1: 7: 0.3: 0.1 by volume) (pH = 5.5) was utilized and scanning of the resolved bands at 215 nm. Linearity of the developed TLC method was evaluated and evident to be 0.4-2, 0.4-2.2, and 0.4-2 μg/band for MIC, NYS, and MET, in that order. The suggested chromatographic methods were verified according to ICH directives. The findings of the developed chromatographic procedures were statistically compared with the results of the reported ones using student's t-test and F-test. Furthermore, two green assessment tools evaluated the indicated methods' level of greenness (GAPI and AGREE).