A comparative green analytical approach based on capillary zone electrophoresis and RP-HPLC for 5-fluorouracil analysis in pharmaceutical formulations

Abstract Two complementary analytical methods based on capillary electrophoresis (CE) and Reversed-phase high-performance liquid chromatography (RP-HPLC) were developed and validated for the quantitative determination of 5-fluorouracil (5-FU) in the pharmaceutical formulation VERRUTOL®. CE separation was achieved using a 50 mM borate buffer (pH 9) under an applied voltage of 30 kV, allowing migration of 5-FU within approximately 5.5 min with stable electroosmotic flow and controlled current. The RP-LC method enabled efficient chromatographic separation with symmetrical peak characteristics and short run time using a simplified mobile phase system. In both methods, complete baseline resolution of 5-FU from salicylic acid and formulation excipients was obtained. The methods were validated in accordance with ICH guidelines. Excellent linearity was observed within the studied concentration ranges, with correlation coefficients ( R 2 ) exceeding 0.999. Precision (intra- and inter-day) and accuracy were within acceptable limits. The calculated limits of detection (LOD) and quantification (LOQ) demonstrated adequate sensitivity for routine quality control purposes, confirming the capability of both techniques to quantify low levels of 5-FU reliably. The environmental, sustainability, and analytical performance of the proposed methods were rigorously evaluated using established assessment tools. The CE and RP-LC methods demonstrated strong environmental and sustainability profiles, as indicated by an AGREE score of 0.76 and 0.69, a BAGI index of 82.5 and 80, and analytical whiteness values of 90.7 and 80.6. These results confirm their adherence to green analytical chemistry principles and their practical applicability. Compared with previously reported procedures, the proposed methods offer reduced analysis time, simplified experimental conditions, and improved sustainability. In particular, the CE method minimizes reagent consumption and organic solvent use, supporting the principles of green analytical chemistry. Both methods are suitable for routine pharmaceutical quality control, providing reliable, selective, and efficient determination of 5-FU in dosage forms.