Rat small intestine extract as a source of mammalian α- and β-glycosidases to study polyphenol bioaccessibility and deglycosylation in vitro: A case study with matrix-devoid and matrix-defined apple fractions

Most polyphenols are glycosylated, affecting their uptake, metabolism, and biological activity. However, the attached sugar must be removed before absorption and functionality can take place. Yet, despite the biological and chemical implications of polyphenol (de-)glycosylation, most in vitro digestion assays omit the utilization of intestinal brush border α- and/or β-glycosidases to study polyphenol bioaccessibility and deglycosylation. This study investigated the effect of rat small intestine extract (RSIE) as an affordable source of mammalian α- and β-glycosidases in different food matrices: matrix-devoid whole apple extract, whole apple, apple juice, and apple pomace. Using the INFOGEST 2.0 model, transepithelial polyphenol absorption, UHPLC-ESI-QTOF-MS/MS, and the inclusion of RSIE at the 15 U*mL^-1 maltase activity reported in the human epithelium, the role of RSIE in polyphenol bioaccessibility and deglycosylation was explored. Moreover, the effect of the plant cell wall (PCW) matrix on the role of RSIE was mechanistically investigated by comparing whole apple (or pomace) with their respective extracts. 36 glycosylated polyphenols were identified, including 33 β-O-glycosides and 3 α-O-glycosides. The content of bioaccessible polyphenol β-O-glycosides and α-O-glycosides was significantly lower (p < 0.001) when RSIE was present, which resulted in a concomitant generation of the aglycone forms (phloretin, quercetin, ferulic acid, caffeic acid and p-coumaric acid). However, the concentration of aglycones was much lower than the reduction in the concentration of glycosylated polyphenols, strongly suggesting that polyphenols bind to RSIE. Matrix-devoid whole apple extract, or pomace extract, exhibited higher polyphenol bioaccessibility than whole apple or pomace, likely due to reduced interactions between polyphenols and the food matrix. Importantly, these differences in bioaccessibility diminished with RSIE, suggesting that RSIE α-glycosidases cleaved α-glucans and disrupted the PCW structure.