Alditols analyses

In a study completed during the early development of f.a.b.-m.s., both f.d. and f.a.b. were used to characterize 101 fractions containing neutral oligosaccharides isolated from human milk. Samples were examined as their peracetylated alditols. In subsequent work, the structures of two minor acidic oligosaccharides from human milk were investigated. The per-methylated derivatives were analyzed by f.a.b.-m.s., and their compositions and sequences were defined by the f.a.b. data. Methylation analysis and partial formolysis were the other principal methods used. 

Qualitative analysis of selected fractions was based on GLC separation of alditol acetates (5). Alditol acetates from uronosyl residues were prepared by methanolysis followed by reduction (4). 

Methylation analysis of PI and FIbp. The fractions were methylated according to Ciucanu and Kerek [8]. The procedure was repeated until no absorbance was detected by I.r. at 2500-3500 cm-i and the per-O-methylated polysaccharides hydrolyzed and analyzed by GLC and GLC-MS of the derived partly O-methylated alditol acetates. 

Oligosaccharides were isolated from PMII by weak acid hydrolysis and separation by SEC and HPAEC-PAD. The isolated oligosaccharides were desalted, reduced and methylated. GC-MS analysis of the partially methylated alditol acetates has been used to reveal the structure of the oligosaccharides. 

PA-1 was partially hydrolyzed with O.IM TFA (40°C, 24 h) and the three fractions (PA-l-I, PA-l-II and PA-l-III) were obtained by Bio-gel P-6. Especially, about 50% of TBA-positive material (PA-l-III) was eluted in the fraction of small oligosaccharide. Permethylated oligosaccharide-alditols from PA-l-III were analyzed by GC-EIMS, and three disaccharide-alditols (IP, 2P and 3P) were detected. EI-MS and component sugar analysis suggested that the major peak, IP was Rha-(l- 5)-Kdo-ol and the minor peaks, 2P and 3P were two epimers of Araf-( l->5)-Dha. 

The content of neutral monosaccharides was determined after an acid hydrolysis, performed as follows with 72 % sulphuric acid for 1 hr at 30°C and then, after dilution to IM sulphuric acid, for another 3 hr s at 100°C. The determination was performed by GC analysis of the prepared alditol acetates (13,14)... 

Multiple-ion monitoring is, however, of considerable value in structural studies, but only if model compounds of known structure are available for comparison. Such an approach has been used in the study of the carbohydrate structures of glycoproteins from different tissues.50 Separation of glycopeptides obtained from various tissues was performed on columns of concanavalin A-Sepharose. Structural analysis by multiple-ion monitoring of partially methylated, alditol acetates derived from the various fractions indicated that the glycopeptides were separated according to the linkage pattern of mannose (see Fig. 1). 

Methylation analysis Permethylation of intact oligosaccharides Hydrolysis of glycosidic linkages Reduction to monosaccharide alditols Acetylation of free hydroxyl groups previously involved in link-... 

Chromatographic analysis ages G.l.c. separation of substituent, partially methylated alditol acetates On-line e.i.- or c.i.-m.s. analysis G.l.c. analysis after sequential hydrolysis, reduction, and acetylation... 

With the development of new liquid phases, interest in the separation of alditol acetates has revived, and this now may be considered the most widely used method for analyzing carbohydrate mixtures, provided that reduction does not introduce an ambiguity into the analysis also, it must be borne in mind that the multiplicity of peaks obtained by trimethylsilylation of a free sugar or of its methyl glycosides may serve a useful purpose in characterization. 

Blake and Richards26 carefully examined all of the steps involved in the analysis of complex polysaccharides, and preferred to use the alditol acetates in studies on the classification,428 fractionation,427 and molecular aggregation428 of hemicelluloses and speargrass xylan.420,430... 

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