Abstract
This study aimed to develop a reliable method for profiling reducing sugars in honey using capillary zone electrophoresis with laser-induced fluorescence detection (CZE-LIF). Reducing sugars were derivatized with 8-aminopyrene-1,3,6-trisulfonic acid (APTS) in the presence of 2-picoline borane, a safer alternative to sodium cyanoborohydride. Key parameters influencing the derivatization efficiency-temperature, pH, incubation time, and reagent concentrations-were systematically optimized. The highest labeling efficiency for glucose, mannose, and maltose was achieved at 50 °C in 0.5 M citric acid with 0.1 M APTS, while fructose showed low reactivity due to its ketose structure. To reduce the background signal from excess reagents, three cleanup strategies were evaluated. Liquid-liquid extraction with ethyl acetate effectively removed unreacted APTS without significant analyte loss, whereas solid-phase extraction and microextraction caused substantial losses of hydrophilic sugars. The method showed good linearity (0.5-10 mM, R(2) > 0.994), precision (RSD 0.81-13.73%), and accuracy (recoveries 93.47-119.75%). Stability studies indicated that sugar standards should be stored at -20 °C. The method was successfully applied to the analysis of four nectar honeys-rapeseed, acacia, phacelia, and dandelion-revealing differences in glucose and fructose content related to botanical origin. The results confirm the suitability of CZE-LIF for sensitive and selective carbohydrate analyses in complex food matrices.