Influence of Season, Hive Position, Extraction Method and Storage Temperature on Polyphenols and Antioxidant Activity of Croatian Honey

The aim of our study was to compare the composition of polyphenolic compounds between the Croatian acacia (Robinia pseudoacacia L.) and chestnut (Castanea sativa Mill.) honey from several aspects: production season, hive position (on the edge and in the middle of a series of hives), part of the hive (small or normal extension), and honey extraction method (centrifuging or draining honey combs). Additionally, in acacia honey, we also monitored the influence of different storage temperatures (room temperature (RT) and 4 °C) on the content of phenolic compounds. To separate, identify and quantify individual flavonoids and phenolic acids from the honey, we used the HPLC method. The total polyphenols and antioxidant activity of the samples, their antimicrobial activity and their elemental content were also measured. The significant influence of the season, hive position, and extraction method on the total identified phenolic compounds, phenolic acids, flavonoids, total phenols and antioxidant activity was detected in almost all the acacia and chestnut honey samples. Chestnut honey from 2013 had more total phenolics (TPs) and antioxidant capacity (FRAP) than chestnut from 2014 and 2015. Honey collected from smaller extensions of hives had significantly higher TPs and FRAP compared to normal hive extensions. Centrifugation reduced the TPs and FRAP in most cases, but not always uniformly. Storage at RT led to the predominance of gallic, p-coumaric and benzoic acid in acacia honey, while storage at 4 °C maintained p-coumaric acid as the dominant phenolic acid. Flavonoids, particularly pinobanksin in acacia honey and hesperetin/pinobanksin in chestnut honey, were less affected by the storage conditions compared to phenolic acids. The non-centrifuged chestnut sample from 2015 showed the lowest MIC values against the most tested pathogenic bacteria. All the honey samples showed an extremely low concentration of heavy metals and relatively high concentrations of potassium and calcium.