Simultaneous Quantification of Citicoline and Tyrosine by Green HPTLC-Densitometry: Application to Pharmaceutical Formulations and Biological Fluids

Citicoline (CTN) and Tyrosine (TSN) are co-formulated in fixed-dose therapy, leveraging their synergistic effects to enhance cognitive function and mental clarity. This study presents the development of a high-performance thin-layer chromatography (HPTLC) method for the simultaneous determination of Citicoline (CTN) and Tyrosine (TSN) in a pharmaceutical capsules, plasma, and urine. A green HPTLC system composed of phosphate buffer pH = 4 and methanol (70:30 v/v) was optimized and applied on silica gel TLC plates (G60 F254). (CTN) and (TSN) were quantified within linear ranges of 0.05‒1.25 µg/band and 0.12‒2.5 µg/band, respectively, using UV detection at 256 nm. The proposed HPTLC method was statistically validated according to ICH in terms of linearity, precision, accuracy, specificity, robustness, detection limits, with LOQ values of 0.043 µg/band for (CTN) and 0.107 µg/band for (TSN). Application of this method extended to pharmaceutical formulations, spiked human plasma, and urine, confirming peak identity and purity. A statistical comparison with published analytical techniques revealed no significant differences, highlighting the reliability of this approach.To assess environmental impact of the advised method, the eco-scale, Green Analytical Procedure Index (GAPI),AGREE metric and Blue Analytical Greenness Index (BAGI).tools were employed, demonstrating the sustainability of this method. High recovery rates further validate its efficacy, positioning it as a green, accurate, and reproducible alternative for routine analysis of (CTN) and (TSN).