Assessing prebiotic effects of resistant starch on modulating gut microbiota with an in vivo animal model and an in vitro semi-continuous fermentation model.

摘要

The objective of our study is to understand the underlying mechanisms of resistant starch (RS) in improving human health from two correlated aspects, modulating gut microbiota and producing SCFA. We hypothesized that when using an in vivo animal model and an in vitro fermentation model, resistant starch consumption would shift gut microbiota pattern towards favoring RS fermentation, especially SCFA production.;We used PCR-DGGE (Denaturing Gradient Gel Electrophoresis) to analyze colonic microbiota with bacteria universal 16S rRNA gene primers. Our results showed two RSs induced significant shifts of colonic microbiota compared to control digestible starch-fed rats. Moreover, differential bacterial patterns were observed between two RSs such as the specific enrichment of putative Bifidobacterium pseudolongum by HA7-SA, but not by HA7. More importantly, a significant correlation was observed between gut microbiota patterns and ACF numbers developed in AOM treated animals. Further analysis of starch fermentation-associated bacteria groups showed similar shifts of Clostridium cluster IV by two RSs with increased putative Ruminococcus bromii. By contrast, the bacterial pattern of Clostridium cluster XIVa showed correlation with ACF number. Our results suggested that gut microbiota patterns modulated by RS were related to differential efficacy of HA7 and HA7-SA in decreasing colonic carcinogenesis.;We established an in vitro semi-continuous anaerobic incubation model to compare fermentability, i.e. SCFA production, of four high amylose starches: HAV, HAVI, HAVII and GEMS-067 from four maize lines with different genetic background and amylose contents (55%, 65%, 70% and 70% respectively). This was done to evaluate prebiotic effects of resistant starch on modulating gut microbiota hypothetically towards favoring RS fermentation. We observed significantly decreased pH, increased gas production, increased butyrate and total SCFA concentration in incubations with the four SR compared to blank BHI medium control starting at wk 1. There was no difference between SR. Molar proportions of butyrate was increased by SR with decreased acetate proportion, both of which achieved stability starting at wk 2. Additionally, propionate concentration was only increased by SRV at the end of the 3-wk incubation compared to BHI medium, but not by other SR. Large inter-individual variation was observed in the proportional increase of butyrate by SR compared to blank BHI medium control. No significant difference was found between lean and overweight individuals from fermentation indicators measured. We concluded that a stable long-term semi-continuous in vitro fermentation model was established to simulate carbohydrate fermentation in human lower gut. We also showed significant increase of butyrate production by RS fermentation with human fecal microbiota.;The modulation of microbiota by amylase digestion residues of HAV, VI and VII in our semi-continuous fermentation model was further analyzed with PCR-DGGE by examining total bacterial pattern and that of RS fermentation associated bacteria groups, including Clostridium cluster IV, XIVa and Bacteroides fragilis group. We hypothesized that bacterial patterns obtained after 3 wk fermentation will be selected by three SR incubation and lean/overweight microbiota as well. Our results showed that total bacterial patterns were shifted by three SR incubation with fecal microbiota from 30 donors at the end of 3-wk fermentation compared to control BHI medium. Moreover, bacterial pattern in SRV fermentation samples differed from that of SRVI and SRVII, which shared a certain degree of similarity. However, bacterial pattern of Clostridium cluster IV and XIVa in SRVI fermentation samples differed from SRV and SRVII, which shared similar patterns, whereas no shifts were observed by any SR in Bacteroides fragilis group bacterial pattern compared to blank BHI medium control. Most important, we observed the putative Ruminococcus bromii was specifically selected during SR incubation by microbiota from lean individuals, but not by microbiota from overweight and obese individuals. We concluded that our in vitro semi-continuous fermentation model can be used to assess prebiotic effects of RS by simulating long-term RS consumption in humans. (Abstract shortened by UMI.)