A Green UPLC-UV Method for L-Ascorbic Acid Determination Based on a Biodegradable Chelating Agent and Synergistic Hydrophobic-Electrostatic Interactions

Abstract Background Ascorbic acid (Vitamin C) is the most widely consumed dietary supplement, which necessitates extensive quantitative testing of these products for batch release and post-market surveillance. However, most methods for ascorbic acid determination rely on hazardous reagents. It is therefore essential to adopt a green analytical chemistry (GAC) approach to evaluate and improve these methods, thereby mitigating their adverse environmental impact of analytical activities. Objective This study aimed to develop a green ultra-performance liquid chromatography (UPLC) method for determining L-ascorbic acid in different types of dietary supplements, using biodegradable reagents and minimizing organic solvent consumption. Method A phosphate solution (pH 3.0) containing 5 mmol/L oxalic acid (OA) was used for sample preparation. A 2-µL aliquot of the ascorbic acid extract was injected into a UPLC system. Separation was performed on an AX-C18 column (2.1 mm i.d. × 100 mm, 1.9 µm) with a mobile phase of 0.4% (v/v) phosphoric acid in water. Isocratic elution was carried out at a flow rate of 0.2 mL/min for 3 minutes. Ascorbic acid was detected at 245 nm and quantitated against standard solutions of L-ascorbic acid. Results OA demonstrated effectiveness in stabilizing ascorbic acid comparable to that of conventional chelators, while offering superior environmental sustainability. When coupled with a UPLC system, this method achieved a low limit of quantitation (0.02 μg/mL) and excellent recoveries (98.3%-101.2%). Analytical Eco-Scale and AGREE metrics were used to evaluate the greenness of this method, indicating its excellent environmental performance. Conclusions This newly developed method demonstrated robust performance across diverse sample matrices, providing an eco-friendly and highly efficient approach for determining vitamin C in dietary supplements and health foods fortified with L-ascorbic acid. Highlights Minimized organic solvent usage, faster turnaround time, and improved environmental compatibility make this a precise, efficient, and green method for vitamin C determination.