Hexosamines chromatography

IonPac KC-811 column separated the labile compounds N-acetylneuraminic acid and N-glycolylneuraminic acid released by mild acid hydrolysis of bovine vitronectin.245 Sialic acid is extremely labile to conditions of handling and must be released by mild acid hydrolysis.246 Derivatization with phe-nylisothiocyanate and separation by reversed phase chromatography was found useful in analysis of hexosamines from gastric mucosa.247 A review on separation of sugars and other carbohydrates which covers many important aspects is available.248... 

Yamashina and coworkers18 72 isolated, on Dowex 50, the total aspartamidoglycan from a proteolytic digest of ovalbumin. It had mean D-mannose hexosamine L-asparagine ratios of 5 3 1. Purified a-D-mannosidase liberated exactly half the D-mannose from the glycopeptide. The residue was subjected to chromatography on Bio-Gel... 

Hexosamines. Hexosamines, in paper chromatography, are, like the sugar acids, sensitive to the pH of the developing solvent. Partridge... 

Neutral sugars and hexosamines were estimated after Dowex-50 (H+)-catalyzed hydrolysis by gas liquid chromatography of glycitol acetate derivatives (26,27). N-Acetylneuraminic acid was measured by a periodate-resorcTrToT"procedure (28). 

If a protein does not contain hexosamines, one other test for the presence of neutral sugars should be made. This can be done by paper chromatography (Carsten and Pierce 1963 Spiro 1965) or gas chromatography ( 2.13.3) after appropriate hydrolysis ( 2.13.2), or a simple colorimetric reaction, such as the orcinol method (below), may be used. 

Hexosamines, hexosami-nitols and amine-containing di- and tri-saccharides Reduction NaBH, Af-de-acetylation anion exchange chromatography, borate-citrate buffers Donald [239]... 

The linkage of the hexosamines to 2-deoxystreptamine was indicated to be at positions 4 and 6 by the periodate oxidation of kanamycin, in which only the 2-deoxystreptamine survived. Conclusive evidence was afforded by the isolation of di-A -acetyl-2-deoxy-5-0-methyl-streptamine from the hydrolysis products of methylated tetra-Af-acetylkanamycin. In addition, the isolation, from this hydrolyzate, of 3-acetamido-3-deoxy-2,4,6-tri-0-methyl-a-D-glucose demonstrated that kanosamine is present as the pyranoid form in kanamycin. Earlier evidence that both hexosamines are present in kanamycin as pyranoid forms had been obtained from periodate-oxidation studies on tetra-iV-acetylkanamycin. At pH 2.5, this derivative rapidly consumes 2 moles of periodate per mole, with the formation of 1 mole of formic acid and no formaldehyde. Examination by paper chromatography of the products of hydrolysis of the reaction mixture by strong acid showed the presence of 2-deoxystreptamine and kanosamine but not of 6-amino-6-deoxy-D-glucose. 

Quigley ME and Englyst HN (1992) Determination of neutral sugars and hexosamines by high-performance liquid chromatography with pulsed amperometric detection. Analyst 117 1715-1718. 

Honda, S. Konishi, T. Suzuki, S. Takahashi, M. Kakehi, 20. K. Ganno, S. Automated analysis of hexosamines by high-petfonnance liquid chromatography with photometric and fluorimetric postcolumn labeling using 2-cyanoacetamide. 21. Anal. Biochem. 1983,134,483. 

Sensitive monitoring of hexosamines in high-performance liquid chromatography by fluorimetric postcolumn labelling using the 2,4-pentanedione-fomialdehyde system. 22. J. Chromatogr. 1983,281, 340. 

M boric acid, 0.05 M NaCl and 0.05 M sodium tetraborate) and dried prior to chromatography in solvent XIX or XX. The total time required for the two-dimensional chromatography is 6—8 hrs. Table 202 gives the hJS -values for these hexosamines. 

Because of the large number of substances which interfere (321) with the hexosamine color determination, a preliminary separation from contaminating substances, either by ion-exchange (284, 323), or by paper chromatography (324), should be made. 

The j8-acetamidodeoxyhexosidase present in normal human urine has been purified by affinity chromatography the enzyme is a glycoprotein containing residues of sialic acid, D-galactose, o-glucose, D-mannose, and unidentified hexosamines. ... 

---