Methylation of polysaccharides

Most early publications on bacterial polysaccharides were concerned with impure products and poorly-described organisms. Many more recent papers are of limited value also, due to low yields, lack of characterization of products and arbitrary interpretations of data. Low yields of methylated polysaccharides may be due to degradation of the bacterial polysaccharide during methylation, or to degradation of the hydrolytic products of the methylated polysaccharide (to form methyl levulinate, etc.46). The great importance of (a) complete methylation of polysaccharide products prior to structural determination by hydrolysis and (6) quantitative identification of the hydrolytic products, has been emphasized previously. Other difficulties in end group analysis have been discussed recently.7... 

Methylation of polysaccharides is carried out in the presence of strong bases. Sulfate groups, depending on their position, may be split to give anhydro sugars or remain intact (see above, Section IV.3.c). Splitting is possible, but only if the spatial position of sulfate is favorable for intramolecular attack by alkoxide according to the... 

Methylation of polysaccharides is now speedily and efficiently achieved by treating them in dimethyl sulfoxide with sodium methyl-sulfinylmethanide and iodomethane.1783 It is well known that strong bases, and, in particular, the base mentioned, cause a /3-elimination... 

Hakomori methylation of polysaccharides containing uronic acid residues results in their complete esterification. The ester group in the esterified residue (A in 12) is electron-withdrawing, allowing the hydrogen attached to C5 to become sufficiently acidic that on treatment with base it is eliminated giving the 4,5-unsaturated residue (B in 12). If the uronic acid is 4-linked, the substituent at C-4 is eliminated (Lindberg et al., 1975). 

O-Methylation is of outstanding importance in stmctural polysaccharide chemistry. A novel method for the methylation of polysaccharides using microwave irradiation was described by Singh et al. (2003). Seed gum from Cyamopsis tetragonol-obus (Guar) was fiilly methylated with dimethyl sulfate and sodium hydroxide using 100% microwave power in 4 min with 68% yield. The completely methylated seed gum was further hydrolyzed by 70% formic add followed by 0.5N H SO under full microwave power. 

In 1947, L-rhamnose was first recognized by Stacey as a constituent of Pneumococcus Type II specific polysaccharide. This finding was confirmed, in 1952, by Kabat et al. and in 1955 again by Stacey when 2,4- and 2,5-di-O-methyl-L-rhamnose were synthesized and the former was shown to be identical with a di-O-methylrhamnose, obtained by hydrolysis of the methylated polysaccharide. This result indicated a pyranose ring structure for the rhamnose units in the polysaccharide. Announcement of the identification of D-arabinofuranose as a constituent of a polysaccharide from M. tuberculosis aroused considerable interest. The L-enantiomer had been found extensively in polysaccharides, but reports of the natural occurrence of D-arabinose had been comparatively rare. To have available reference compounds for comparison with degradation products of polysaccharides, syntheses of derivatives (particularly methyl ethers) of both d- and L-arabinose were reported in 1947. 

Kvemheim, A.L. (1987). Methylation analysis of polysaccharides with butyllithium in dimethyl sulfoxide. Acta Chem. Scand. Ser. B41,150-152. 

Reeves and Goebel72 have shown that hydrolysis of the reduced methylated capsular polysaccharide of Type III pneumococcus yields 2,3,6-trimethyl-D-glucose and the anomeric forms of methyl 2,4-dimethyl-D-glucoside. The cellobiuronic acid units in the polysaccharide are thus linked through position 3 of the D-glucuronic acid residue, probably by /3-D-linkages. That is, the polysaccharide contains alternate 1,3-and l,4-/3-D-linkages. 

The isolation of 2,3-dimethyl-D-glucose from the hydrolysis products of certain methylated polysaccharides has been an important factor in assigning structures to these polysaccharides. From trimethyl-starch it has been recovered in about 3% yield, together with 2,3,4,6-tetra-methyl- and 2,3,6-trimethyl-D-glucopyranose, and arises from the points of linkage of the repeating chains of the amylopectin component.67,69,70 From a dimethyl-starch the yield is considerably higher (75%).71 Other sources are the methylated capsular polysaccharide of Rhizobium radici-... 

Catalytic reduction and methanolysis of the methylated capsular polysaccharide of Type III pneumococcus gives a mixture of methyl 2,3,6-trimethyl- and 2,4-dimethyl-a/3-D-glucopyranosides. The latter, which arises from glucuronic acid units in the polysaccharide, can be separated into crystalline a- and 6-isomers identical with synthetic specimens.84,88... 

Methylation of a purified alkaline-soluble polysaccharide from New Zealand flax is accomplished by subjecting it to five successive methyla-... 

The aforementioned deuterated derivatives were prepared by way of reduction of a ketone, aldehyde, or ester with sodium borodeu-teride, or by deuteroboration of an alkene. An interesting reaction, perhaps eventually applicable to direct deuteration of polysaccharides, was reported by Koch and Stuart413 and by them and their coworkers,41b who found that treatment of methyl a-D-glucopyranoside with Raney nickel catalyst in deuterium oxide results in exchange of protons attached to C-2, C-3, C-4, and C-6. In other compounds, some protons of CHOH groups are not replaced, but the spectra may nevertheless be interpreted with the aid of a- and /3-deuterium effects. 

Our original approach to polysaccharide C-13 n.m.r. spectral analysis consisted of making a minimum number of hypotheses about expected structure-to-spectra relationships (8). By then comparing spectra to known structure for a series of D-glucans, we attempted to establish the validity of these hypotheses and to establish how diverse a structural difference could be accommodated The hypotheses were as follows. Firstly, that each polymer could be considered as an assembly of independent saccharide monomers. Secondly, that these hypothetical saccharide monomers would be 0 alkylated (0 -methylated) in the same positions as the actual saccharide linked residues (it had previously been established that 0-methylation of any a-D-glucopyranosyl carbon atom position resulted in a down-field displacement of vlO p.p.m. for the associated resonance). Thirdly, that each differently substituted residue would have a completely different set of chemical shift values for each carbon atom position (different from the unsubstituted saccharide) but that only the carbon atom positions involved in inter-saccharide linkages would have A6 greater that 1 p.p.m. And, fourthly, that the hypothetical 0-alkylated residues would contribute resonances to the total spectrum proportional to their mole ratio in the polymers. 

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