Study On The Fermentation Characteristics Of Resistant Starches And Their Regulation Mechanism On Gut Microbiota

Imbalances in the human gut microbiome have been tied to a wide range of gastrointestinal and systemic conditions affecting host health.Dietary fiber is considered a key element in shaping the beneficial host-microbe symbiosis.Starch is not only the most common carbohydrate in our daily life,but also an important source of human energy.In particular,resistant starch has attracted much attention due to its beneficial regulation of the gut microbiome contributing to a reduction in the risk of colon-related diseases.In this thesis,the starch degradation mechanism and changes in microbial community composition and functionality occurring during in vitro fecal fermentation of A-and B-type polymorphic starches were explored.To this end,in vitro batch fermentations were performed focusing on differences between native A and B-type starches,as well as,modifications in the degree of cross-linking or starch encapsulation with chitosan.In follow-up experiments,the inter-individual variability in the response to chitosan encapsulated starch microspheres was characterized in vitro and the physiological effects of the microspheres were explored in an in vivo mouse model.The main objectives and findings were:1.The in vitro fecal fermentation characteristics of typical A-(normal maize,waxy maize starch)and B-type(high amylose maize,potato)polymorphic starches were investigated.Changes in the granular starch structure(SEM,XRD,SEC),as well as,the microbial community composition and metabolic activity(gas production,SCFA)were evaluated over the course of fermentation.Marked differences in the fermentation rate,microbiota composition and microbial metabolite production were observed between A and B-type starches.Concurrent with the increased relative abundance of Roseburia,Blautia,and Lachnospiraceae,the butyrate production supported by B type high amylose maize and potato starches(ca.38 m M)was significantly higher than that of waxy and normal maize A type starches(23 m M for waxy maize starch and 33 m M for normal maize starch).A-type polymorphic starches,on the other hand,displayed a faster fermentation rate and promoted Megamonas.X-ray diffraction and size-exclusion chromatography of the A and B-type starch residues revealed a similar “side-byside” fermentation pattern.The “exo-pitting” fermentation pattern was also revealed through the SEM image changes during the fermentation time course.2.A series of low,medium,and highly cross-linked starches(CL-5,CL-8,CL-12)were fabricated to evaluate the effect of traditional chemical crosslinking on the fermentation characteristics of high amylose maize starch.Cross-linking decreased the gas production of high amylose maize starch throughout fermentation.The butyrate production rate of crosslinked starches gradually decreased with an increasing cross-linking degree.Certain beneficial gut microbes belonging to the Blautia and Clostridiales genera were promoted by starches with low and medium cross-linking degrees,whereas HAMS with a high cross-linking degree increased the relative abundance of Bacteroides uniformis and Ruminococcus bromii.3.We prepared a series of starch-entrapped microspheres with a variable chitosan to starch ratio(CS:S=2:1,CS:S=1:2,CS:S=1:6,CS:S=1:12)by means of electro-spraying and investigated the fermentability by human fecal microbiota in an in vitro batch system.Starch encapsulation controlled the microbial gas production,and the fermentation rate gradually decreased with increasing chitosan proportions.Moreover,the starch and chitosan composites displayed a synergistic effect on the gut microbiota composition.Beneficial gut microbes belonging to the genera Roseburia,Lachnospiraceae and Clostridiales were promoted by all the microspheres.The abundance of the aforementioned health-promoting taxa reached a maximum in chitosan:starch microspheres with a 1:6(w/w)ratio(CS:S=1:6).*4.The inter-individual variability in the response of fecal microbiota obtained from six healthy individuals to starch-entrapped microspheres(CS:S=1:6)was studied using 48 h in vitro batch incubations.A slow fermentation rate was observed in all the donors.Microspheres displayed a significantly lower damaged and higher intact cell counts in comparison to native starch after48 hours of fermentation.Moreover,MS fermentation selectively stimulated the propionate or butyrate production and promoted beneficial species including Bifidobacterium,Bacteroides and Agathobacter in a donor-dependent way.5.We studied in vivo dietary interventions with starch-entrapped microspheres(MS,CS:S=1:6)and high amylose maize starch(HAMS)in mice suffering from high-fat diet induced metabolic disorder and gut microbiome dysbiosis.MS more efficiently controlled body and adipose tissue weight gain compared to HAMS.Furthermore,MS significantly reduced the blood glucose,insulin,lipid and pro-inflammatory cytokine levels.The altered gut microbiota composition favoring Streptococcaceae,bacilli,Firmicutes and unclassified Clostridiales was predicted to promote fatty acid,pantothenate and Co A biosynthesis.In line with this,elevated fecal SCFA,in particular,propionate concentrations were observed in MS-fed mice.