Chemical Analysis of Polyphenolic Content and Antioxidant Screening of 17 African Propolis Samples Using RP-HPLC and Spectroscopy

Abstract

Background and objectives: Propolis is a resinous material produced by honeybees. Its chemical composition is highly complex and varies significantly depending on geographic region and season. This intrinsic variability presents challenges to the standardization and quality control of propolis. This study aimed to evaluate the chemical composition, total phenolic content, and antioxidant potential of propolis collected from seventeen geographical regions across Africa.

Methods: A reverse-phase high-performance liquid chromatography (RP-HPLC) method coupled with a photodiode array detector (PDA) was used for analysis of propolis samples. The flavonoid and phenolic contents of the samples were determined using colorimetric and Folin-Ciocalteu methods. Antioxidant capacity was assessed using the 2,2-diphenyl-1-picrylhydrazyl assay.

Results: Five flavonoids (naringenin, pinocembrin, galangin, chrysin, and quercetin), one flavonoid glycoside (rutin), six phenolic acids (caffeic acid, p- coumaric acid, cinnamic acid, chlorogenic acid, ferulic acid, and gallic acid), and an aromatic ester - caffeic acid phenethyl ester were simultaneously detected and quantified using RP-HPLC with an ACE-5 C18 column (250 mm × 4.6 mm i.d., 5 µm) and PDA detector. The reference standards showed good linearity with regression coefficients (R²) ranging from 0.96 to 0.99. For precision, repeatability, and stability studies, the relative standard deviation for all reference standards was below 2.5%. The 2,2-diphenyl-1-picrylhydrazyl assay yielded EC₅₀ values ranging from 17.6 ± 0.39 to 0.16 ± 0.001 mg/mL.

Conclusions: RP- HPLC method for the simultaneous quantification of thirteen reference standards will serve as a reliable tool for the standardization and quality evaluation of propolis. The flavonoid and phenolic contents are key contributors to the antioxidant activity of propolis and reflect local plant biodiversity and bee-plant interactions within the ecosystem.

Keywords

African propolis / Flavonoids / Standardization / Quality control / Apiceuticals / Honeybees / High-performance chromatography

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