Comparative Analysis of Phenolic Acid Profiles, Antioxidant Capacity, and Antimicrobial Activity of Honeys from Different Botanical and Geographical Origins

Abstract

The composition of honey is complex and can vary depending on its botanical and geographical origin. This study evaluated the phenolic acid profile (PAP), antioxidant activity, and antimicrobial properties of honey samples from different geographical origins, such as Romania (nine types: multiflower, thyme, manna, hawthorn, black locust, linden, rapeseed, mint, and pasture), Malaysia (Tualang), New Zealand (Manuka), and Spain (chestnut). Twelve different varieties of honey were analyzed, one sample for each type, and all analyses were performed in triplicate. Phenolic acids were extracted by solid-phase extraction (SPE) using C18 cartridges, followed by high-performance liquid chromatography with diode-array detection (HPLC-DAD) analysis at 310 nm. Antioxidant activity was assessed using the 2,2′-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assays. Antimicrobial activity was evaluated by agar diffusion and broth microdilution to determine the minimum inhibitory concentration (MIC), while microbial anti-adherence activity was assessed using the crystal violet staining assay to determine the minimum anti-adherence inhibitory concentration (MAIC). The tested microorganisms included the reference strains Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Enterococcus faecalis ATCC 29212, and Candida albicans ATCC 10231, as well as two C. albicans isolates (24114 and 5329). Several phenolic acids, including chlorogenic, caffeic, ferulic, trans-cinnamic, 3-O-methylgallic, p-coumaric, and syringic acids, were identified at varying concentrations across the analyzed honey samples. Rapeseed honey exhibited the highest overall concentration of phenolic acids. Among the honey samples analyzed, chestnut honey had the highest concentration of cinnamic acid, mint honey the highest concentration of ferulic acid, and multiflower honey the highest concentration of syringic acid. Tualang honey exhibited the highest antioxidant activity by both methods, reaching 2.103 mg TE/g honey for DPPH and 1.701 mg TE/g honey for ABTS. Tualang honey also exhibited the strongest antimicrobial activity against C. albicans and P. aeruginosa, whereas multifloral honey was the most active against E. faecalis. The analyzed honey extracts differed in their phenolic acid composition and biological activities, including antioxidant, antimicrobial, and anti-adherence effects of microorganisms on substrates. While associations were observed between selected phenolic acids and some of the evaluated biological activities, the overall biological properties of honey are more likely to arise from the combined contribution of multiple bioactive constituents than from individual compounds alone. The novelty of this study lies in the comparative assessment of the PAP obtained following SPE, and the biological activities of different types of honey from various botanical and geographical sources.