Pectin from coffee Pulp: extraction, characterization, and bioactivities

Abstract

Coffee pulp, a significant by-product of coffee processing in Thailand, represents a promising source of value-added compounds. This study investigates the physicochemical characteristics and bioactive properties, specifically the prebiotic and anticancer potential, of pectin derived from coffee pulp (CP). Pectin was isolated using a double extraction method and subsequently modified enzymatically (SFD). Comprehensive characterization involved thermogravimetric analysis (TGA) for purity, Fourier-transform infrared spectroscopy (FTIR) for the degree of esterification (%DE), gel permeation chromatography (GPC) for molecular weight, and high-performance anion-exchange chromatography (HPAEC-PAD) for sugar composition and structure. Bioactivity assessments included the DPPH assay for antioxidant potential, in vitro growth studies with Pediococcus sp. (PE), Lactococcus lactis (LL), and Lactobacillus acidophilus (LA) to determine Prebiotic Index and Activity Scores, and the CCK-8 assay for cytotoxicity against SW480 colon cancer and CCD 841 CoN normal colon cells, with apoptosis confirmed via Annexin V-FITC/PI staining. Results were benchmarked against commercial prebiotics (inulin), pectins (HMP, L20, L40), and 5- Fluorouracil (5-FU). The extraction yielded 7.99% low methoxy pectin (LMP) with 4.76% DE, 68.05% purity, a weight-average molecular weight (Mw) of 68.6 KDa, and a linear structure (CP). Enzymatic modification yielded SFD, which retained LMP characteristics (0.00% DE) but exhibited increased branching and a significantly

reduced Mw of 1.10 KDa. SF demonstrated the highest antioxidant potential (IC50 of 101.33 µg/mL), surpassing commercial pectins which failed to reach 50% inhibition. Furthermore, CP significantly promoted the growth of all tested lactic acid bacteria, showing higher prebiotic index (PE: 0.95, LL: 3.88, LA: 3.73) and activity scores (PE: 0.25, LL: 0.40, LA: 0.72) than commercial pectins. Notably, only SFD exhibited significant anticancer activity, selectively inhibiting SW480 cell proliferation (IC50 of 3.40 mg/mL) more effectively than normal CCD 841 CoN cells (IC50 of 11.13 mg/mL), while unmodified CP showed no effect. SFD induced late-stage apoptosis in 19.80 ± 7.51% of SW480 cells, compared to 10.68 ± 1.82% in controls and 25.97 ± 3.51% with 5-FU. These findings highlight that coffee pulp pectin is a promising natural prebiotic for food applications and gut health support. Moreover, its enzymatically modified form (MCPP) shows potential as a selective anticancer agent, warranting further investigation for therapeutic applications.