The spectra of the NBRC 0005 and 0622 mannans high throughput chemical screening had no and small signals at 4.788 ppm, respectively. However, the NBRC 103857 mannan showed a significantly large signal. The signal at 5.172 ppm, cross peak 6, which corresponds to the 1,2 linked mannose residue substituted by the 1,2 linked mannose residue, was also present in parallel with the signal at 4.788 ppm. These results indicate that the NBRC 103857 mannan contains a large amount of 1,2 linked mannose residues. Instead, the signal at around 5.28 ppm, cross peak 4, corresponding to the intermediary 1,2 linked mannose residue, was very small compared to the other two strain mannans. The signal at around 5.56 ppm, cross peak 1, which corresponds to a phosphodiesterified 1,2 linked mannobiose residue, is present in the NBRC 0005 and 0622 mannans. On the other hand, the NBRC 103857 mannan has a signal at 5.457 ppm, cross peak 2, indicates that the mannan contains mannose as the phosphodiesterified form instead of the 1,2 linked mannobiose. The 1H NMR signal of the mannan from S. cerevisiae is apparently different from the other three C. glabrata mannans. The signal at 5.432 ppm, cross peak 3, indicates the presence of the 1,3 linked mannobiose as the phosphodiesterified form instead of mannose or 1,2 linked mannobiose. Furthermore, it is apparent from the DQF COSY that the S. cerevisiae mannan contains a large amount of the 1,3 linkedmannose residue at the non reducing terminal of the side chains based on the presence of cross peaks 7, 12 and 16 and does not contain the 1,2 linked mannose residue due to the absence of cross peaks 6, 14 and 15. Phosphodiesterified oligosaccharides and O linked oligosaccharides in the mannans The phosphodiester linkage in the mannan was selectively hydrolyzed using 10 mM HCl at 100 for 1 h. The released carbohydrate from the NBRC 0005 and 0622 mannans was biose and that from the NBRC 103857 mannan was mannose.
The 1H NMR spectrum of biose was completely the same as that of 1,2 linked mannobiose obtained from the mannan of C. albicans. The O linked oligosaccharides in the mannans were eliminated by treatment with 0.5 M NaBH4/0.1 M NaOH at 25 for 18 h. As shown in Fig. 3B, oligosaccharides up to Bicalutamide Calutide tetraose were obtained from the three strain mannans. The 1H NMR spectra indicated that these oligosaccharides consisted of 1,2 linked mannose residues, and from the signal at 5.147 ppm, it is apparent that only tetraose contained the 1,3 linked mannose residue at the non reducing terminal. Acetolysis of the mannans To determine the side chain structure of the N linked mannan moiety, we subjected these mannans to mild acetolysis. Figs. 4A, B, and C show the elution profile of the acetolysis products from the NBRC 0622, 103857, and S. cerevisiae mannans, respectively. From the NBRC 0622 mannan, Man4 and Man3 were obtained as the main oligosaccharides. On the other hand, from the NBRC 103857 mannan, Man3 was the largest one of the main oligosaccharides. Furthermore, a small amount of the longer oligosaccharides, Man5 to Man7, were obtained from the three mannans. These 1H NMR spectra are shown in Fig. 5. The result indicated that Man2 and Man3 from the three mannans consisted of 1,2 linked mannose residues except for Man3 from the NBRC.