| Exo-polysaccharide (EPS) of bacteria is a favorable source for the preparation of new-type oligosaccharides with special bioactivities. Generally, EPS of microorganism was composed of disaccharide to octasaccharide repeating-units. The repeating-units consist of two to four monosaccharides, some of the sugar chain structures containing characteristic functional groups, such as acetyl group, acetone acid or uronic acid. So the composition and structure of EPS are very diversiform. By means of enzyme hydrolysis technology, oligosaccharides with novel biological activity can be produced. In this work, we tried to produce EPS from Klebsiella K13, and used bacteriophage-borne glycanase as tool enzyme to obtain new oligosaccharides which may possess some novel biological activities and application potential in the area of prebiotics and anti- harmful bacteria factors.At the beginning of this work, the medium components and culture conditions for the production of EPS from Klebsiella K13 are optimized. Results showed that EPS yield reached 4.59g/ L, improved 73.9% compared with control.Biological properties of P13 were studied. Results showed that the virus particle approximated to round; 23nm in size. The nuclear acid type of the phage is ssRNA and longer than 15000bp. The plaque on the solid plate is round and hyaline, around of which is halo, formed by the released glycanase from bacteriophage. The phage titer is 2.0×10~9 pfu/ml, and the stability, latency and burst period to infect host cell are good. The reaction conditions of glycanase degrading exo-polysaccharide are optimized. Results were showed as follows: reaction temperature, 48℃; concentration of EPS, 1%; the initial pH, 6.5; reaction time, 2 hours; concentration of glycanase, 4 times of stock solution.To evaluate the prebiotic effect and bacteriostasis of these oligosaccharides, static batch culture fermentation in vitro was applied to examine the effect of KOS (oligosaccharides from Klebsiella K13 ), choosing other oligosaccharides as control. In comparison with the other tested samples, PI value of KOS was the highest, gas production was very low; SCFA yield of KOS was medium, but having many types. It indicated that KOS can be a good prebiotic. In addition, KOS had a marked inhibition effect on E. coilO157, but opposite to Salmonella and Lactobacillus. Growth-promoting effect of KOS on Lactobacillus is useful to us. At present, there are few reports on the research of biological activity of oligosaccharides from bacterial EPS. This study enriches the application prospect of KOS and other oligosaccharides.To study the structure of oligosaccharides, EPS was degraded by bacteriophage–borne glycanase. Enzyme hydrolysis components were separated and purified by passing through Bio-Gel P4 column. Molecular weights of the purified fractions were determined by ESI-CID-MS. Assistance of the information from GC, IR and NMR of EPS and oligosaccharides, the structures were analyzed. So the structure of oligosaccharides could be primarily illustrated. A conclusion could be drawn that the bacteriophage-borne glycanase could be used as a good tool for the production of novel oligosaccharides from bacterial EPS.
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