Green Synthesis of Biogenic Gold Nanoparticles Using Crassula Ovata and Bryophyllum Pinnatum Leaf Extracts: Characterization and In-Vitro Evaluation of Antioxidant, Cytotoxic and Anti-Inflammatory Activities

Abstract

Ethanolic leaf extracts of Crassula ovata and Bryophyllum pinnatum were used as reducing and capping systems for biogenic gold nanoparticles, and the products were carried through three unrelated in vitro endpoints under one protocol. Extract was reacted with 1 mM chloroauric acid at a 1:9 extract-to-salt ratio (pH 6.0, 70 °C, 1 h, 24 h dark maturation) to give CO-AuNPs and BP-AuNPs, and a 1:1 (w/w) physical mixture of the two purified powders (MIX-AuNPs) was carried as a combination arm. Both batches gave a single surface plasmon resonance band, at 531.87 ± 0.45 nm for CO-AuNPs and 535.70 ± 0.50 nm for BP-AuNPs, with no band in the solvent, salt or extract controls. Hydrodynamic diameter was 69.13 ± 1.25 and 63.03 ± 1.20 nm, polydispersity index 0.228 ± 0.009 and 0.205 ± 0.007, and zeta potential −25.37 ± 0.65 and −30.33 ± 0.85 mV. Diffraction peaks indexed to face-centred cubic gold with mean crystallite sizes of 25.83 ± 3.04 and 19.22 ± 2.04 nm; EDX returned 76.91 and 79.02 wt% gold with carbon and oxygen signals consistent with an adsorbed phytochemical layer. Cumulative gold-equivalent release over 24 h in phosphate-buffered saline pH 7.4 reached 79.23 ± 2.85% and 85.27 ± 2.80%. The potency order was the same on every endpoint: MIX-AuNPs > BP-AuNPs > CO-AuNPs > BP-EXT > CO-EXT. DPPH IC50 fell from 143.11 µg/mL for the C. ovata extract to 45.42 µg/mL for MIX-AuNPs, against 10.34 µg/mL for ascorbic acid. At 24 h the MTT IC50 for MIX-AuNP was 110.15 µg/mL (MCF-7), 122.25 µg/mL (HCT116) and 142.68 µg/mL (A549). In lipopolysaccharide-stimulated RAW 264.7 macrophages held above 84% viability, MIX-AuNPs lowered TNF-α by 77.55 ± 5.11% and IL-6 by 79.55 ± 5.12% at 50 µg/mL. After Holm adjustment the mixture was not significantly superior to either individual system at matched concentrations, so the combination result is reported as enhanced activity and not synergy.