Galactose chromatography

Acid hydrolysis of the basic hexasaccharide yielded the disaccharide 31. Its mobility in paper chromatography lay between those of the corresponding (1 — 3)-linked (from S10A) and /3-(l — 6)-linked (from S29) isomers, which is why it was assumed to be (1 — 4)-linked. The (1 — 2)-linked isomer was excluded, as the D-galactose residue that is part of 31 carries a D-galactopyranosyl group linked to 0-2 in the original polysaccharide, as will be discussed. 

A better method has been described by Schwarz (S6). Washed red blood cells are lysed, precipitated with trichloroacetic acid below 0°C and the supernatant quickly neutralized. Speed and low temperatures are necessary to prevent hydrolysis of galactose-l-phosphate which is very sensitive to acid. Barium acetate and ethanol are added, and the precipitated barium salt of galactose-l-phosphate washed with 80% ethanol. The barium salt is then hydrolyzed by heating with dilute hydrochloric acid, acid and salts removed with mixed ion-exchange resins, and the galactose estimated by paper chromatography as described above. It is probably better to avoid the Amberlite MB-1 resin used by Schwarz and, instead, to use a weak base resin mixture, such as Amberlite MB-4. Recovery of added galactose-l-phosphate should be determined simultaneously. 

Some workers have performed hydrolyses in an autoclave, but it has been shown that 0.5 M sulfuric acid at 120° degrades 33% of L-arabinose and 22% of D-galactose in two hours.75 Such methods are, therefore, only suitable for qualitative analyses, unless accurate corrections are made. Similarly, 90% formic acid has been found76 to decompose 48% of D-xylose and 36% of D-galactose in 20 hours at 100°. Gas-liquid chromatography has been used to examine the products formed by the acid degradation of sugars.77... 

Analysis of glycosaminoglycan hydrolyzates is often complicated by the presence of basic and acidic glycoses, in addition to neutral monosaccharides. Adsorption chromatography on silica gel was used to classify such a hydrolyzate, and the neutral fraction was shown to contain D-glucose, D-galactose, and D-mannose.288 Separation on ion-exchange resins may also be used.51,149 The resolution of the acidic and basic fractions is discussed in Sections IX (p. 71) and X (p. 78). 

---