Effects Of Probiotic Fermentation On Immunological Activity,Structural Characteristics Of Water-Soluble Polysaccharides From Carrot

Carrot is an Umbelliferae herb with rich nutritional value,and has a reputation of“small ginseng”.Probiotics are a class of bioactive microorganisms that are beneficial to the body.In our preliminary work,the probiotics NCU 116 from and carrot were successfully combined to develop a new type of fermented carrot juice.At the same time,it was found that probiotic fermentation had certain effects on the nutrient composition of carrot juice and the functional activity of carrot polysaccharide.In this paper,water-soluble polysaccharides were extracted from the carrot juice before and after probiotic fermentation,which were WSP-n and WSP-p respectively.The immunomodulatory activities and basic structural characteristics of the two polysaccharides were studied and compared.The main contents are summarized as follows:?1?Water-soluble polysaccharides?WSP-n and WSP-p?of carrot juice before and after fermentation of probiotics were extracted by water extraction and alcohol precipitation.A mouse model of immunosuppression induced by cyclophosphamide was established to compare the effects of two polysaccharides on immune function in mice.The results showed that both WSP-n and WSP-p could increase the thymus index of immunosuppressed mice,slow down the spleen enlargement,enhance the proliferation of spleen lymphocytes,and alleviate the disorder of mouse T cell subsets caused by cyclophosphamide.Administration of WSP-n and WSP-p increased the levels of immunoglobulins in cytokines and serum in spleen tissues.The combined results showed that WSP-p improved the immune-suppressed mice more strongly than that of WSP-n.?2?The mouse model of immunosuppression induced by cyclophosphamide was established to compare the different effects of WSP-n and WSP-p on intestinal health of immunosuppressed mice,by investigating mucosal injury,intestinal mucosal immunity and intestinal metabolism.The results showed that WSP-p could repair the pathological morphology of small intestine in immunosuppressed mice more strongly than that of WSP-n.WSP-p could significantly increase the number of goblet cells and mucin area in immunosuppressed mice in a dose-dependent manner.Both of WSP-n and WSP-p had no significant effect on mouse CD4⁺T cells level,while medium dose WSP-n could significantly increase CD8⁺T cell content.The high-dose WSP-p group had significant differences in the recovery level of mouse fecal water content compared with the model group,and the effect was better.?3?The WSP-n and WSP-p were separated and purified to obtain three fractions,respectively.The first two fractions both from the two polysaccharides were mainly neutral sugars,while the third one was mainly acidic sugars.Basic structure analysis of the obtained six component polysaccharides showed that the infrared spectra of the purified polysaccharide of carrot was similar before and after fermentation,and all of them exhibited typical absorption peaks of carbohydrate characteristics.The relative molecular weight and polydispersity index of purified polysaccharide in WSP-n were larger than WSP-p.The results of monosaccharide composition showed that the monosaccharide species of purified polysaccharide in WSP-n and WSP-p was consistent,but the content was different.Microscopic morphology of each purified component in WSP-n and WSP-p was different.It indicated that the fermentation of probiotics had a slight effect on the structural characteristics of carrot polysaccharide.?4?Due to the limitations of the monosaccharide composition hydrolysis method,this paper proposed a new hydrolysis method and optimizes the hydrolysis conditions.After the first step of hydrolysis by TFA,TFA was removed and then the second hydrolysis was carried out by H₂SO₄,which released as much uronic acid as possible while reducing the loss of neutral sugar.TFA concentration,hydrolyze time and temperature were selected for optimization of the first hydrolysis step.The hydrolysis conditions of the first step were optimized by single factor test and response surface experiment.The second step H₂SO₄ hydrolysis conditions were optimized under the previous work of our group.Taking carrot polysaccharide as an example,both of the traditional one-step hydrolysis method and the two-step hydrolysis method were used to hydrolyze it.It was found that the total amount of monosaccharides in the carrot obtained by the two-step hydrolysis method was18.87%,higher than that of the one-step hydrolysis.