Oligosaccharides purification

Towbin, H., et al. (1988). Chromogenic Labeling of Milk Oligosaccharides Purification by Affinity Chromatography and Structure Determination, Anal. Biochem. 173 1-9. 

Nobre C, Teixeira JA, Rodrigues LR. Fructo-oligosaccharides purification from a fermentative broth using an activated charcoal column. New Biotechnol 2012 29(3) 395—401. 

To allow all culture productiou to be coutrolled, a method for rapid analysis is required. Prior to development of an LC-MS method, the analysis was both complex and time-consuming, involving the purification of a relatively large amount of the antibody using affinity chromatography, enzymatic release, and subsequent derivatizafion of the oligosaccharides and their analysis by using capillary electrophoresis. 

After the release of the oligosaccharides, they must be purified by a variety of methods before structural analyses can be undertaken. Ion-ex-change chromatography, gel-filtration chromatography, or some type of electrophoresis is usually used for the purification. At this point, structural analyses may begin. 

Zopf, D.A., Smith, D.F., Drzeniek, Z., Tsai, C.-M., and Ginsburg, V. (1978a) Affinity purification of antibodies using oligosaccharide-phenethylamine derivatives coupled to Sepharose. In Methods in Enzymology, (V. Ginsburg, ed.), Vol. 50, pp. 171-175. Academic Press, New York. 

Malto-oligosaccharide aldonolactones react with ethylenediamine to give Ar-(2-aminoethyl)aldonamides (113-115), which have been successfully grafted onto carriers via amide linkages. The malto-oligosaccharides were produced by degradation of amylose with alpha amylase. After purification of the oligosaccharides, they were converted into the lactones by hypoiodite or electrolytic oxidation. 

For the determination of the phosphate group a new procedure was established (64a) using a two-step deacylation of the rough form LPS (E. coli F515 and S. minnesota R595) and h.p.l.c. purification of the intact core-backbone oligosaccharide. Data obtained by H-, 13C-, and 31P-n.m.r. spectroscopy as well as f.a.b.-m.s. analysis unequivocally showed the presence of 4 -phos-phate at GlcN (II). A similar procedure was also applied to E. coli J5 LPS (66a). 

Oligosaccharides and glycoconjugates in living cells often exist as closely related mixtures. Their isolation from natural sources in homogeneous form is therefore very difficult, involving tedious purification and difficult characterization. This sequence of steps tends to result in low yields. This difficult situation presents chemical synthesis with a major opportunity to positively affect progress in the biochemical understanding of the processes described above.4... 

Glycal assembly on a solid support eliminates the repetitive purifications usually associated with oligosaccharide synthesis. As a method, it has a certain generality as it does not require any specific enzymes or complex starting materials. Both natural and nonnatural sugars may be used in the constructions. 

Cleavage Purification Scheme 7.1 Strategies for solid-phase oligosaccharide synthesis. 

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