Physicochemical Characteristics, Antioxidant Properties, and Identification of Bioactive Compounds in Australian Stingless Bee Honey Using High-Performance Thin-Layer Chromatography

Abstract

This study investigates the physiochemical properties, chemical composition, and antioxidant activity of Australian stingless bee honey blends from two bee species, Tetragonula carbonaria and Tetragonula hockingsi, harvested in Burpengary East, Queensland at different times of the year. The moisture content of the honey samples ranged from 26.5% to 30.0%, total soluble solids from 70.0 to 73.5° Brix, and pH from 3.57 to 4.19. The main sugars identified were trehalulose (13.9 to 30.3 g/100 g), fructose (12.9 to 32.3 g/100 g), and glucose (4.80 to 23.7 g/100 g). The total phenolic content (TPC), measured using the Folin–Ciocalteu assay, ranged from 26.1 to 58.6 mg of gallic acid equivalents/100 g. The antioxidant activity was investigated with the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, with values ranging from 1.39 to 6.08 mmol of Trolox equivalents/kg. Antioxidant constituents were determined using a High-Performance Thin-Layer Chromatography (HPTLC)-DPPH assay. The HPTLC-DPPH analysis revealed that honey samples collected in May 2022 contained the highest number of antioxidant compounds. Some constituents were identified using an HPTLC-derived database and also quantified utilising HPTLC analysis. Lumichrome was present in all honey samples, while luteolin and kaempferide were detected only in some. Kaempferol or isorhamnetin was also found to be present, although a definitive distinction between these two chemically closely related compounds could not be made by HPTLC analysis. The results showed that honey produced by Tetragonula hockingsi and Tetragonula carbonaria shares similar properties and composition when harvested at the same time, with only minor differences in moisture, fructose, and glucose content.