Preparation Of Manno-oligosaccharides By Enzymatic Hydrolysis And Its Function

As a degradation product of Konjac glucomanna(KGM),manno-oligosaccharides(MOS)not only retain the functional characteristics of KGM as a kind of dietary fiber,but also improve the action effect of KGM for its larger viscosity.Moreover,MOS,a kind of functional oligosaccharides,can improve enteric microorganism environment,increase body immunity,and reduce blood glucose and blood fat.Although there have been a-KGM was degraded to MOS by hydrolysis with β-mannosidase.The optimal degradation condition was explored,the quality inspection of degradation product was conducted,and in vivo and in vitro experiments were carried out to verify its function.The results are stated as below:1.MOS sample that Mw≤2000 Da was obtained by controlling single variables,such as degradation,pH value,and reaction time.On the basis of single factor optimization,the orthogonal experiment of a four-factor(reaction temperature,pH value,enzyme dose,reaction time)three-level was carried out,and the optimal reaction conditions were obtained: the degradation temperature of 50℃,pH value of 5.5,reaction for 2 h,and enzyme dose of 150 U/g,and the yield of target MOS was 65.72%.2.The product of MOS was tested according to relevant national standards,and the results showed that the water content was 6.74%,ash content was 13.63%,the pH value of 10 g/L sample was 6.08,acid non-soluble substance content was 0.37%,protein content was 0.081%,total bacterial count was 690 cfu/g,heavy metal of Hg content was0.0034 mg/kg,Pb content was 0.48 mg/kg,As content was 0.311 mg/kg and E.coli was not existed.3.Experiment was done to analyze how the MOS effect the growth of microorganisms in vitro.The result shows that under the condition of pure culture,MOS can promote the growth of both Lac.johnsonii and E.Coli.,while under the condition of co-culture,MOS can promote the growth of Lac.johnsonii,but inhibit the growth of E.Coli.Compared with the control group(glucose as the carbon source),the addition of MOS significantly decreased the PH value of fermentation broth.These indicate that MOS inhibited the proliferation of E.coli by promoting the production of lactic acid by Lac.johnsonii.4.Animal experiment.The carbon clearance test showed that the carbon clearance phagocytic index of high dose group mice(1.5 g/kg ?bw MOS)increased significantlycompared with the control group(distilled water)(P < 0.05).In the delayed type hypersensitivity test,the swelling degree of the middle dose group(1 g/kg?bw MOS)and the high dose group was higher than that of the control group,and the difference was significant(P<0.05).Compared with the control group,the thymus index of the high dose group increased significantly(P<0.05),and the sleen index increased highly significantly(P<0.01).Moreover,the experiment on cytokines and antibodies in serum were detected.The results showed that the content of IgA in low dose group(0.5 g/kg?bw MOS)showed the highest level,and there is a significant difference compared with the control group(P< 0.01);while IgG showed the highest level in high dosage group,with significant difference(P<0.05),IFN-γ of middle dosage group showed significant difference(P<0.05).and that of high dose group showed very significant difference(P<0.01).5.High-throughput sequencing was performed using DNA which is extracting from fresh excreta of mice cecum.It is found that the abundance and diversity of enteric microorganism in high dosage group are higher than those of control group.Moreover the contents of some beneficial microflora,such as Lactobacillaceae,Streptococcus faecalis and Roseburia were higher than those in the control group.