Isolation, purification and structural elucidation of EPS-I, an extracellular polysaccharide from Enterococcus durans

The rough crystal of exopolysaccharide produced by Enterococcus durans, a strain of lactic acid bacteria screened from the intestine of a cock in our laboratory, was purified by CM-cellulose column chromatography, DEAE-Sephadex A-25 ionexchange and Sephadex G-100 gel chromatography to give EPS-I. The EPS-I was eluted as a single peak in HPLC analysis, indicating the homogeneity of EPS-I and free from low-molecular-weight polysaccharides. The molecular weight of the EPS-I was determined as 42 000 by the light scattering method. The result of its elemental analysis was C 41.08% and H 7.23% without the elements of N, P and S. Monosaccharide analysis showed that it was composed of Glc and Man in a molar ratio of 4:1. Sugar composition analysis, methylation analysis and H-1 and C-13 NMR spectroscopy revealed that the EPS-I was composed of pentasaccharide repeating units. The sequence of sugar residues was determined by using two-dimensional NMR, including heteronuclear multiple-bond correlation(HMBC) and nuclear overhauser effect spectroscopy (NOESY). The structure of the pentasaccharide repeating unit of EPS-I was given, a new excellular polysaccharide from lactic acid bacterium compared with other EPSs was reported.