Graphene-magnetite as magnetic solid phase adsorbent for extraction of 4-hydroxybenzoic acid and 3,4-dihydroxybenzoic acid in stingless bee honey

Abstract

Stingless bee honeys are rich in secondary metabolites such as free phenolic acids which can be easily absorbed into the body. Trace amount of the phenolic acids had made the analysis difficult, hence sample pretreatment is crucial. In this work, a graphene-magnetite composite (G-Fe3O4) was synthesized and assessed as an adsorbent for magnetic solid phase extraction (MSPE) of two phenolic acids namely 4-hydroxybenzoic acid (4-HB) and 3,4-dihydroxybenzoic acid (3,4-DHB) from honey samples prior to analysis using high performance liquid chromatography with ultraviolet-visible detector (HPLC-UV/Vis). Characterizations of G-Fe3O4 were performed using Fourier transform infrared spectroscopy (FTIR), low vacuum scanning electron microscopy (LVSEM) and nitrogen adsorption analysis. Several MSPE parameters affecting the extraction of these two phenolic acids were optimized. Optimum MSPE conditions were 50 mg of G-Fe3O4 adsorbent, vortex rotational speed of 1600 rpm, 5 min extraction time, 30 mL sample volume at pH 0.5, 200 μL methanol as desorption solvent (5 min sonication assisted) and 5% w/v NaCl salt. Matrix-matched calibration was used for the analysis of the two phenolic acids from several honey samples. Calibration graphs were linear in the range 1–50 μg/g (R2 = 0.9997) for 4-HB and 3–50 μg/g (R2 = 0.9996) for 3,4-DHB. The limit of detection (LOD = 3S/N) calculated for 4-HB and 3,4-DHB was 0.08 μg/g and 0.14 μg/g, respectively. Good relative recoveries (72.6-110.6%) were obtained for both phenolic acids from honey samples with RSD < 6.0% (n = 3). The developed G-Fe3O4 MSPE method was applied to the analysis of both phenolic acids in honey samples from Johor Bahru, Johor. Two Trigona spp. honey samples (H1 and H2) and a commercial honey sample (H3) were used in this study. The amount of 4-HB and 3,4-DHB in H1 sample were 0.14 ± 0.9 μg/g and 0.67 ± 1.7 μg/g honey, respectively. H2 sample showed slightly higher amount of both phenolic acids (0.47 ± 3.1 μg/g for 4-HB and 1.61 ± 2.3 μg/g for 3,4-DHB). The amount of 4-HB and 3,4-DHB extracted from H3 sample was below the LOD of the developed method. The developed G-Fe3O4 MSPE method offered is simple, environmental friendly and efficient for extraction of phenolic acids from honey samples.