Study On The Structure And Probiotics Activity Of Konjac Glucomannan Degraded By Microwave And Enzyme Process

Konjac glucomannan is the main component of Konjac tuber.It can be fermented and utilized by intestinal microorganisms to regulate the host intestinal flora and promote intestinal immune health.It also shows great potential in improving glucose metabolism,lowering blood lipid,anti-inflammation,preventing constipation and losing weight.In this study,the purified KGM was degraded by?-mannanase enzymolysis and microwave pyrolysis to obtain a series of gradient molecular weight KGM.Then,the molecular weight and solubility of KGM oligosaccharides were determined.Then,the structural characteristics of KGM oligosaccharides were preliminarily identified by thermogravimetric analysis,X-ray diffraction analysis,Fourier infrared spectroscopy and scanning electron microscopy analysis.The conformational parameters of KGM oligosaccharides with different molecular weights were determined by high-performance gel permeation chromatography combined with light scattering technique.Then,different molecular weights of KGM were used as carbon sources,inoculated with human fecal bacteria and cultured in vitro by anaerobic fermentation.The p H value,gas production,sugar content and organic acid yield of KGM with different molecular weights under the action of intestinal flora were determined to describe the in vitro fermentation behavior of KGM.At the same time,high-throughput sequencing was combined to further study the changes of the microbial community before and after the fermentation,and establish the relationship between the fermentation behavior of different molecular weight KGM and the composition of the microbial community.Finally,low molecular weight KGM was added into the tablet candy.The production process of Konjac glucomannan oligosaccharide tablet candy was optimized through multiple single factor experiments,including drying temperature,addition amount of magnesium stearate,microcrystalline cellulose,xylitol and citric acid,and the corresponding sensory evaluation and analysis were conducted.The main research contents of this paper are as follows:(1)Taking KGM as the research object,the effects of?-mannanase enzymolysis and microwave pyrolysis on the molecular weight of KGM were investigated.Microwave heating can effectively degrade KGM.With the increase of microwave power and microwave cracking time,the molecular weight of KGM decreased significantly,and the degree of decrease of KGM molecular weight was positively correlated with power and time.The molecular weight obtained by microwave pyrolysis decreased first and then increased with the increase of KGM concentration,depending on the high speed back and forth movement of polysaccharide chains and the strong gel forming ability of KGM.KGM could be specifically hydrolyzed by?-mannanase.The molecular weight of KGM decreased with the increase of enzyme activity and enzymolysis time,and the molecular weight distribution also narrowed.With the increase of KGM concentration,the enzymatic hydrolysis was insufficient and the molecular weight also increased.A series of KGM with different molecular weights prepared by each single factor of enzymatic hydrolysis and microwave pyrolysis were screened,and the molecular weights of the enzymatic hydrolysis KGM were 1235,2537,9141,22703 and 80155 Da,respectively,which were named as EL-1,EL-2,EL-3,EL-4 and EL-5.Microwave pyrolysis KGM with molecular weights of 1235,2792,9227,27837 and 71311 Da were obtained,which were named as MP-1,MP-2,MP-3,MP-4 and MP-5,respectively.(2)The physicochemical properties,structural characteristics and solution conformation of different molecular weights of KGM were studied.The solubility of KGM decreased with the increase of molecular weight.Lowering the molecular weight of KGM will lead to its stability decline,which is more obvious in the region with lower molecular weight.In addition,the decrease of molecular weight will also lead to the destruction of some regular crystal structures in KGM,and the disappearance of crystal regions.The characteristic chemical groups of KGM did not change significantly before and after microwave pyrolysis and enzymatic hydrolysis.The results of electron microscopy showed that the linear structure of KGM was destroyed,the spherical structure gradually disappeared,and the lamellar structure gradually increased with the increase of degradation degree.EL-1,MP-1 and MP-2are mainly composed of fragmentary lamellar and clastic structures.The conformational study of the solution speculated that the small molecule region of the polysaccharide chain in EL-1,EL-2,EL-3,EL-4,MP-2,MP-3 and MP-4 had a rod-like structure,while in the small molecule region of a small amount of polymer KGM in MP-4,the polysaccharide molecule had a random linear group conformation.The conformation of a small amount of polymer region polysaccharide chains of EL-5,MP-5 and KGM in solution is spherical.(3)Based on the p H value of the fermentation solution,sugar content,gas production,OD₆₀₀ value,short chain fatty acids;SCFAs content and intestinal flora structure and diversity were analyzed to analyze the difference in the enzymatic characteristics of KGM with different molecular weights.The results of in vitro fermentation showed that the lower the molecular weight of KGM was,the more quickly it could be used by intestinal microorganisms,and the indexes of gas production,OD₆₀₀ value,sugar content and p H tended to balance in a short period of time.The total amount of organic acids in the process of fermentation increased continuously with the increase of fermentation time.Except for MP-1 and inulin groups,the acetic acid yield increased with the increase of the fermentation time,while the acetic acid yield of the other samples increased at first and then decreased during the fermentation process.Compared with the monosaccharide group,the production of butyric acid and valeric acid at the fermentation end point were significantly increased in the KGM and inulin groups with different molecular weights,and the increase of valeric acid content was related to the abnormal increase of Megasphaera.In addition,with the increase of the molecular weight of KGM oligosaccharide,the diversity of?was increased(P<0.05).Compared with the control group,there were significant differences in the structure of intestinal flora.Different molecular weights of KGM could regulate the composition of intestinal flora.In the fermentation system with different molecular weight KGM instead of carbon source,Bifidobacterium had no obvious change,but Lactobacillus significantly increased.The lower the molecular weight KGM oligosaccharide,the better the effect of promoting the proliferation of Lactobacillus.(4)Optimizing the process of Konjac glucomannan oligosaccharide pressing candy,it was found that the increase of drying temperature could reduce the water content of the material,and then reduce the hardness of the product.The increase of magnesium stearate and microcrystalline cellulose also significantly reduced the hardness of the product.The results of sensory evaluation showed that the smoothness of the sample increased with the addition of magnesium stearate.When the ratio of xylitol to citric acid is about 18.75,the best acidity and sweetness of the product is obtained.When the addition amount of magnesium stearate and microcrystalline cellulose were 2 g and 10 g,respectively,the chewability of tablet candy was the best at 60?.