Optimization of Green Sample Preparation for the Determination of Hydroxycinnamic Acids in Multi-Floral Honey Using Response Surface Methodology

Abstract

Featured Application: The present study provides practical guidelines for the solubilization of hydroxycinnamic acids from honey for their chromatographic determination. The proposed green preparation step can be used for honey hydroxycinnamic fingerprinting for different botanical and/or geographical origins. In addition, it can be also utilized for the evaluation of diverse biological effects of hydroxycinnamic acids present in honey. Hydroxycinnamic acids are an essential phenolic group in honey that are related with its diverse health effects. Furthermore, they can be considered as potential biomarkers for botanical and geographical origin. The extraction of hydroxycinnamic acids from honey requires complicated extraction techniques due to their low contents and matrix particularity. The goal of the present work was to develop a green sample preparation method for the solubilization of hydroxycinnamic acids in honey samples. Thus, a Box–Behnken design has been used to investigate the effect of four factors: (i) percentage of ethanol (60–90%, v/v), (ii) temperature (30–60 °C), (iii) solvent-honey ratio (10–30 mL g−1), and (iv) sonication time (10–40 min) on the contents of caffeic, chlorogenic, and ferulic acids. Based on the desirability index, the ultrasound-assisted mixing of honey with 60.0% (v/v) ethanol at a solvent-honey ratio of 10 mL g−1 for a period of 17.8 min at a temperature of 54.6 °C resulted in the maximum solubilization of hydroxycinnamates. Subsequently, the greenness of developed method was assessed and applied successfully on the analysis of honeys. The present study reports for first time the hydroxycinnamic acid fingerprint of multi-floral honey produced in Cyprus.