The signals at 4.783 and 5.161 ppm of Man3 Everolimus mTOR inhibitor from the NBRC 103857 mannan corresponds to the non reducing terminal1,2 linked mannose and 1,2 substituted 1,2 linked mannose residues, respectively, therefore, we determined the structure to be Man 2Man 2Man. The 1H NMR assignment results are shown in Table 1. The presence of the signal at 5.145 ppm in the two Man4 indicated that they contained an 1,3 linked mannose residue at the non reducing terminal. On the other hand, the long oligosaccharides from Man5 to Man7 contain characteristic 1H NMR signals at 5.08 5.11 ppm, which seem to correspond to the 1,6 linked mannose residues substituted by 1,2 linked ones. Furthermore, the signal at 4.783 ppm, which corresponds to non reducing terminal 1,2 linked mannose residue, was also present in Man5 and Man6 from the NBRC 103857 mannan. The 1H NMR spectra of the oligosaccharides obtained from the NBRC 0005 mannan by the acetolysis was basically the same as that of the NBRC 0622 mannan. Methylation analysis There are several reports about the presence of Man 2Man 2Man 2Man 6Man 2Man 2Man 2Man or related oligosaccharides in the long side chain of the mannans from C. krusei and C. glabrata. Therefore, in order to determine the side chain structure, a methylation analysis of the long oligosaccharides from Man5 to Man7 obtained by acetolysis was carried out. As shown in Table 2, these long oligosaccharides from the NBRC 0622 mannan, 103857 mannan, and S. cerevisiae mannan contained a 3,4 di O methyl 1,2,5,6 tetra O acetyl mannitol, which corresponds to a 2,6 di O substituted mannose residue. In the preceding study, we found the presence of a 3,6 branched mannose residue in the side chain of the C.
albicans and C. guilliermondii mannans. However, it is apparent that these C. glabrata mannans contain no such branched side chain. Furthermore the absence of the 6 O substituted mannose residue and the presence of about two moles of the non reducing terminal mannose residues in one mole of each long oligosaccharide suggested that these originated from two neighboring sidechains connected by the backbone 1,6 linkage. Structure of 1,6 linkage containing oligosaccharides In order to confirm the position of the 2,6 di O substituted mannose residue, we analyzed the long oligosaccharides Man6 from the NBRC 0622 mannan, Man5 from the NBRC 103857 mannan, and Man5 from the S. cerevisiae mannan by DQF COSY. The 1,6 branched mannopentaose, Sk Man5, from the S. kluyveri mannan was used as the control. The boxed regions in Fig. 6 show the H 1 H 2 correlated cross peaks of the mannose residues of the oligosaccharides. Cross peak 4 indicates an intermedial 1,2 linked mannose residue in the linear oligosaccharide. On the other hand, cross peak 5 indicates an Vinorelbine 71486-22-1 upfield shifted 1,2 linked mannose residue by a steric effect due to the presence of the 6 O substituted mannose residue at the intermediary position of the oligosaccharide. The presence of cross peak 5 in Sk Man5 indicated that the 1,6 linked mannose residue was connected to the third mannose residue from the reducing terminal to make a branched structure. In Man5, Man6, and Man7 from the C. glabrata and S. cerevisiae mannans, the presence of cross peak 8 or 10.