Steric Inhibition by Butyryl Groups on Gut Microbial Amylases Significantly Impacts In Vitro Fecal Fermentation of Butyrylated Starch

Colonic starch fermentation requires the assistance of gut microbial amylases (GMAs). However, it remains unknown whether chemically substituted butyryl groups induce a steric inhibitory effect on GMAs analogous to that observed with pancreatic amylase during the intestinal digestion of butyrylated starch. In this study, we investigated the in vitro fermentation and enzymatic hydrolysis performance of three types of butyrylated starch. The results showed that the esterolysis of butyryl groups was a rate-limiting process, and the fermentation of butyrylated starch was partially inhibited by steric inhibition of the butyryl groups on GMAs. X-ray photoelectron spectroscopy (XPS) results further confirmed the negative correlation between the fermentation rate of starch and the relative content of butyryl groups accessible to the gut microbiota. Moreover, analyses of the enzymatic characteristics and the resulting hydrolysate composition demonstrated that a relatively high multiple attack degree (MAD) made GMAs more susceptible to steric inhibition by butyryl groups, thus producing more malto-oligosaccharides, which are preferred by butyrate-producing bacteria. These findings provide important insights into the fermentation behavior of butyrylated starch from the perspective of interactions between microbial amylases and starch.