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Abstract
The aims of this work were to isolate and characterize phenolic constituents of Georgia-grown blackberry (Rubus spp.) cultivars (i.e., Navaho, Kiowa, and Ouachita) and to examine their antioxidant and anti-inflammatory capacities. Phenolic compounds were identified and quantified using HPLC and MS techniques. Column chromatography on Amberlite XAD-16 was employed to recover polyphenolic extracts, which were further fractionated on Sephadex LH-20. ESI-MS studies confirmed the presence of cyanidin-3-O-glucoside as the predominant anthocyanin in all the cultivars; cyanidin-3-O-rutinoside and cyanidin-3-O-xyloylrutinoside were also present in the Kiowa cultivar. Ellagitannin isomers; i.e., lambertianin A and lambertianin C were confirmed in all blackberry cultivars with pedunculagin and lambertianin D also being detected in Navaho and Kiowa blackberries, respectively. Total phenolics content (TPC) and total anthocyanins content were determined spectrophotometrically using the classical Folin-Ciocalteu assay and a pH differential method, respectively. Antioxidant capacities were assessed by the FRAP and TEAC assays; values for the preparations ranged from 26.12.7 to 178.16.9 mmol Fe2+ equivalents/100g of fraction (d.w.) and 13.60.7 to 79.40.9 mol Trolox equivalents/g of fraction (d.w.), respectively. All fractions collected inhibited the formation of advanced glycation endproducts (AGEs); the % inhibition of AGEs ranged from 20.71.9 to 79.42.2%. Significant positive correlations (p<0.05) were determined for all fractions based on the TPC vs the FRAP and TEAC assays as well as % inhibition of protein glycation. The anti-inflammatory effects of low- and high-molecular-weight phenolic fractions (LMPF and HMPF, respectively) from blackberries were examined by assessing the effects of edema and polymorphonuclear (PMN) leukocyte infiltration 24 h after topical application of an irritant, 12-O-tetradecanoylphorbol-13-acetate (TPA), using the mouse ear model. The mouse ears of treatment groups were applied a solution of the LMPF, HMPF, or indomethacin, a non-steroidal anti-inflammatory drug (NSAID), 30 min after TPA application. The NSAID, LMPF, and HMPF (from all three cultivars) significantly (p<0.05) reduced TPA-induced injury when compared to the TPA-positive control group. Myeloperoxidase (MPO) activity, a biomarker for PMN leukocyte infiltration, was also significantly (p<0.05) reduced for treatment groups. Inhibition of both edema and MPO activity indicates marked anti-inflammatory activities arising from the phenolic constituents in the blackberries.