Densitometric HPTLC Method for the Simultaneous Determination of Novel 1-Coumarinyl-1,2,3-Triazole Derivatives

HPTLC-densitometric techniques show considerable promise as alternative chromatographic models for determining partitioning parameters in complex mixtures comprising inorganic, organic, and biomolecular components. In this study, five newly synthesized 1-coumarinyl-1,2,3-triazole derivatives were effectively separated using silica gel 60 F254 plates as the stationary phase. Three mobile phase systems, chloroform: methanol (9:1 v/v), hexane: ethyl acetate (3:2 v/v), and hexane: ethyl acetate (1:1 v/v) were employed, resulting in well-resolved compounds with Rf values ranging from 0.27 to 0.66. Validation of the developed methods was carried out in compliance with ICH standards. Linearity was observed across the range of 50–500 ng/band, with correlation coefficient (R²) values between 0.9923 and 0.9991, indicating excellent linearity. The high selectivity and specificity of the methods ensured the accurate separation of the synthesized compounds in their pure form, free from any interference by starting materials or intermediates. Limits of detection (LOD) for the target compounds ranged from 17.5 to 47.8 ng/spot, while limits of quantification (LOQ) were 52.6 to 144.8 ng/spot using spectrophotometric detection and 47.5 to 96.8 ng/spot using spectrofluorometric detection. These findings highlight the applicability of the developed HPTLC methods for both qualitative and quantitative analysis of this compound series, even at concentrations as low as 10⁻⁹ g in biological fluids..