One-pot biocatalysis of potato rhamnogalacturonan and the role of its deacetylation in efficient inhibition of colon cancer cells and hydrogel mediated colon-targeted drug delivery

Abstract

Deacetylation of potato rhamnogalacturonan (PRG) by rhamnogalacturonan acetyl esterase (CtPae12B) was explored for enhanced hydrolysis of PRG by rhamnogalacturonan lyase (CtRGLf) and the effects of deacetylated PRG were studied in enhancing inhibition of colon-cancer cells and formation of colon-targeting drug delivery material. Pre-treatment of PRG with CtPae12B resulted in increased relative activity of CtRGLf. CtPae12B removed acetyl groups from both O-2 and O-3 positions of D-galactopyranosyluronic acid residues of PRG, resulting in 98 % deacetylation. PRG displayed 21.9 % degree of acetylation and 7.7 % degree of methylation. TLC and ESI-MS analysis of CtRGLf hydrolysed PRG showed unsaturated RG di-saccharide as the smallest product, with m/z 322. Deacetylated PRG-oligosaccharides displayed higher, 50 % inhibition of colon-cancer HCT-116 cells (with shrunken and globular morphology) than 35 % inhibition by acetylated PRG-oligosaccharides. FESEM and BET analysis of CtPae12B-treated PRG showed porous structure and significantly higher total surface area and pore volume than non-enzyme treated PRG. Higher drug entrapment efficiency and lower drug release rate of CtPae12B-treated PRG hydrogel (0.0033 min−1 at pH 1.2 and 0.009 min−1 at pH 7.4), than non-enzyme treated PRG hydrogel, (0.0057 min−1 at pH 1.2 and 0.02 min−1 at pH 7.4), showed it to be a potential biomaterial for sustainable colon-targeted drug delivery.