Biodegradation Of Xanthan And Bioactivities Of Xanthan-derived Oligosaccharides

Oligosaccharides have been proved to have some bioactivities such as defense response, antimicrobe,immunity reinforcement,etc.,which were degraded mainly from constituent polysaccharides of plant or pathogen cell wails.Xanthan is exopolysaccharide produced by phytopathogen Xanthomonas campestris,which was defined as a virulence factor of X. campestris inducing black rot of cruciate flower.Xanthan could not be easily degraded by most microorganisms because xanthan consists of a main chain similar to cellulose.The aim of this article is to search for a microorganism degrading xanthan to form bioactive oligosaccharides and study the special physiological activity of the formed oligosaccharides.One strain XT11 capable of degrading xanthan was isolated from soil samples.Strain XT11 should be assigned to the genus Microbacterium because the determinative taxonomic properties of strain XT11 were consistent with that of the genus Microbacterium.This agrees with the phylogenetic analysis via 16S rRNA gene sequence similarity,in which the strain XT11 is clustered in the genus Microbacteriurn.The closest genetic distance occurred between XT11 and M.hydrocarbonoxydans,the similarity of which is 97.8%.Access number of 16S rRNA gene in GenBank is DQ350882.According to the cluster analysis of the heredity relation of Microbacterium,the Microbacteriurn is divided to two cluster groups. Strain XT11 and peri-edge bacterium M.hydrocarbonoxydans is classified as a son branch. The confidence level is 85%.Strain Microbacteriurn sp.XT11 produced exoenzyme degrading glycoside linkage in the backbone of xanthan,leading to the decrease in viscosity and produce xantho-oligosaccharides.The xanthan-degrading enzyme was induced by xanthan and its glycosidic anaiogus CMC,starch and maltose.The optimal medium constituent for xanthan-degrading strain XT11 is 0.3%xanthan and 0.3%yeast extract at the initial pH7.0. The optimum culture temperature is 28℃.The glucose added to the medium promotes the cell growth of the strain XT11,but inhibits the enzyme production induced by xanthan.The carbon source sucrose,lactose,maltose,starch and starch also inhibit the induction of xanthan-degrading enzyme by xanthan.The culture time course of xanthan-degrading strain XT11 showed that the viscosity of fermentation broth was quickly descent during cell growth,while the activities of xanthan-degrading enzymes was increased gradually.The highest enzyme activity was obtained at 28-h incubation time,which is 23 U/mL.The properties of xanthan-degrading enzyme showed that the optimal reaction temperature was 40℃,the optimal pH was 6.0,the optimal concentration of substrate was 0.4%(w/v).The concentration of glucose greater than 0.06%inhibits the enzymatic reaction of xanthan-degrading enzyme.A good correlation occurred at between the viscosity decrease and reducing sugar release when xanthan was degraded by xanthan-degrading enzyme,which suggested that the backbone of molecules xanthan was degraded endogenously.This is just necessary for xantho-oligosaccharide production.Xantho-oligosaccharides with different ratio of viscosity and reducing sugars were produced from xanthan by xanthan-degrading enzyme.The xantho-oligosaccharides showed the ability to scavenging hydroxy free radicle.The activity is correlated to the concentration of oligosaccharide.The drgree of polymerization is the key factor affecting the activities of oligosaccharide.The xantho-oligosaccharide S6(the value of viscosity and reducing sugar is 40.9) has the strongest ability to discard hydroxyl free radical.Xantho-oligosaccharide could induce defense response in soybean cotyledon,which can protect plants against microbial disease.The xanthan-degrading oligosaccharides inhibit the growth of pathogen Xanthomonas campestris which is the pathogenic bacteria variety,but not inhibit other supplied pathogenic bacteria such as Esherichia coli,Erwinia carotovora,Salmonella typhi,Pseudomonas aeruginosa,Streptococcus mutans,Staphylococcus aureus and Bacillus subtilis. Xantho-oligosaccharide S4,S5(the ratio of viscosity and reducing sugar is 232.5 and 101.4 respectively) showed the great activity of bacteriostasis to pathogen Xanthomonas campestris. The xantho-oligosaccharide could inhibit the cells growth of Xanthomonas campestris and prevent the synthesis of virulence factor xanthan.Also it can inhibit partially the production of of cellulase,chitinase,pectinase and proteinase by Xanthomonas campestris.The results showed that the xantho-oligosaccharide had potential application in protection of plants against black rot of cruciferae caused by Xanthomonas campestris.