Preparation of propolis extractions

Propolis samples were collected from the south of Turkey (Hatay region). Propolis samples were frozen at −24°C and a grinder was used to break them into small pieces (Delonghi Kg49, Hampshire, UK). Two grams of propolis were shaken with 20 ml 70% ethanol/water (v/v) using a shaker for 1 h, followed by ultra-sonication (Bandelin Sonorex RK100, Berlin, Germany) for 30 min. The mixture was filtered using filter papers (Watman No: 1, Buckinghamshire, UK) to remove wax and bee parts. Finally, the filtrate was placed into glass tubes (tared tubes) and evaporated using a vacuum centrifuge (Jouan, RC 10-10). The tared tubes were weighed after evaporation of propolis solution to determine the amount of the dry propolis extract (DPE), which was stored at −24°C until HPLC-DAD (High-Performance Liquid Chromatography-Diode Array Detector) analysis.