Separation And Characterization Of Water-soluble Polysaccharide From Cinnamomum Camphora Seed Kernel And Its Application In Yogurt

Polysaccharide is a kind of biological macromolecular compound with very complex chemical structure,which has a variety of potential pharmacological activities,such as antibacterial,anticancer,lipid-lowering,hypoglycemic,antioxidant,anti-inflammatory,immune regulation and regulation of intestinal microorganisms.In addition,polysaccharides have no side effects that also can be used in food and medicine fields instead of synthetic drugs.As a new plant resource,Cinnamomum camphora seed kernel(CCSK)can be found everywhere in the south part of Yangtze River in China,and its resource is very rich.At present,researches about CCSK mainly focus on the extraction techniques,functional properties and physiological activities of Cinnamomum camphora seed kernel oil(CCSKO)and protein,but there are few studies about 17.90% carbohydrate in CCSK.As one of the main technologies for extracting CCSKO,aqueous enzymatic method can not only obtain high quality oil,but also retain the structure and function of protein,polysaccharide,polyphenols,saponins and other substances in byproducts.Therefore,in this study,the process wastewater from the aqueous enzymatic method extraction of CCSKO was used as raw material,the separation,purification,structural characterization and proliferation activity of Cinnamomum camphora seed kernel polysaccharide(CCSKP)on probiotics in vitro were studied,and CCSKP was then used as a functional factor in yogurt products.1.The crude polysaccharide(CCSKP)of CCSK was obtained by deproteinizing,depigmentation,ethanol precipitation and freeze drying,using the process wastewater of extracting CCSKO by aqueous enzymatic method as the main raw material.CCSKP1-A was further purified by DEAE-52 cellulose and dexan Sephadex G-50 gel column.The total sugar content and protein content of CCSKP were 64.7±1.27% and 11.33±1.12%,indicating that CCSKP may be a natural polysaccharide-protein complex.The total sugar content of CCSKP1-A was 89.50±0.46%,and no uronic acid content was detected,so CCSKP1-A may be a high purity neutral polysaccharide.The structural characteristics of CCSKP and CCSKP1-A were analyzed by HPGC,IC,FTIR and SEM.The results showed that CCSKP was mainly composed of arabinose(0.4%),glucosamine hydrochloride(0.3%),galactose(1.8%),glucose(91.1%)and mannose(6.5%).The average molecular weight of CCSKP was 14080 Da,7934Da and 5361 Da,respectively.It is a polydisperse polysaccharide-protein complex with rough surface morphology.The average molecular weight of CCSKP1-A was 4219 Da,which was mainly composed of arabinose(0.3%),glucosamine hydrochloride(0.4%),galactose(1.6%),glucose(88.3%)and mannose(9.4%).CCSKP1-A may be a relatively uniform neutral polysaccharide with smooth surface structure.2.The possible structure of CCSKP1-A was preliminarily predicted by methylation experiment combined with one-dimensional and two-dimensional NMR analysis techniques.The results showed that the main chain of CCSKP1-A containing 1,4-linked-α-D-Glcp and1,4,6-linked-α-D-Glcp residues,α-D-Glcp,α-D Manp and β-D-Glcp terminal residues mainly through O-4/O-6 links in 1,4-linked-α-D-Glcp and 1,4,6-linked-α-D-Glcp or 1,3,4-linked-β-DGlcp residues.3.The maximum degree of hydrolysis of CCSKP and CCSKP1-A in simulated gastric fluid was 4.66±0.13% and 1.69±0.03%,respectively,and the maximum degree of resistance toα-amylase hydrolysis was 5.90±0.13% and 5.74±0.23%,respectively.The results indicated that CCSKP and CCSKP1-A had good anti-digestibility.In addition,the in vitro proliferation effect of Cinnamomum camphora seed kernel polysaccharides on probiotics was investigated.The results showed that Lactobacillus acidophilus,Bifidobacterium longum,Bifidobacterium infantile and Bifidobacterium adolescentis could all effectively utilize CCSKP and CCSKP1-A and promote their own growth and reproduction,and the growth reached the peak after culture for 32 ～ 36 h,corresponding produced the short-chain fatty acids(SCFAs),mainly acetic acid and lactic acid,with the most acetic acid content.These results indicated that CCSKP and CCSKP1-A can be used as potential prebiotics to promote the growth and propagation of probiotics,and the culture-medium containing CCSKP has better adaptability to probiotics.4.CCSKP was then used as functional factor to prepare yogurt products.The results showed that the optimal formula of CCSKP yogurt was as follows: fermentation time 9h,inoculum amount 0.12%,adding amount of CCSKP 0.10%,adding amount of white sugar 5%,the resulting CCSKP yogurt was milky white,exquisite taste,sweet and sour flavor rich.