Separations of sugars

Dowex 2-X8 1.2 0.75 Strongly basic (but less basic than Dowex 1 type) anion exchanger with S-DVB matrix for deionization of carbohydrates and separation of sugars, sugar alcohols, and glycosides. 

In sugar mills a rapid separation of sugar crystals from molasses is achieved through the use of massecuite and centrifugal force. The motor drives a basket full of molasses which undergoes repeated cycles of operation, i.e. 

Fig. 1 (A) Chromatographic separation of sugars. Track 1 fructose, 2 sucrose, 3 glucose, 4 mixture of the substances in tracks 1-3, 5 mixture of substances in tracks 1-3 and 6, 6 Fructo-oligosaccharides, 7 1-kestose, 8 mixture of glucose, maltose, maltotriose and maltotetraose. (B) Absorption scan of track 5 with 200 ng each substance per chromatogram zone 1 = fructosyl-nystose, 2 = nystose, 3 = 1-kestose, 4 = fructose, 5 = sucrose, 6 = glucose.

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... 

The application of the SMB-technique to the downstream processing of biotechnological products requires some specific changes to meet the special demands of bioproduct isolation. Some exemplary applications are given including separations of sugars, proteins, monoclonal antibodies, ionic molecules and optical isomers and for desalting. 

Although the isolation and identification of new disaccharides, tri-saccharides and tetrasaccharides and their derivatives, either by acid hydrolysis or by controlled oxidative degradation, " would be of great help in these studies it would appear to be worth while to develop other indirect methods of approach involving the use of enzymes capable of effecting scission at specific points in the molecular complex. Better methods for the quantitative separation of sugars and their derivatives are in the process of development and it is not unlikely that in the near future it will be possible to derive formulas not only for plant gums but for the many related complex polysaccharides. 

Only model studies have so far been reported, and the method has not yet been applied to the analysis of sugar mixtures, with the exceptions noted.48,49 The recovery of glucose-14C trifluoroacetate from a column of Carbowax 20 M or SF-96 has been shown to be 5 to 14% and 3 to 5%, respectively.208 Table III (p. 112) records further details of the separation of sugars and their derivatives as trifluoroacetates. 

Besides bonded phases that incorporate linear alkyl chains with 8 or 18 carbon atoms and that can be utilised between pH 2 and pH 13, there are polar stationary phases with linear chains incorporating aminopropyl, cyanopropyl, benzyl or mixed moieties. For example, aminoalkyl chains can be used for the separation of sugars. 

An alternative approach, which additionally favors the separation of sugars from interfering compounds, is the precolumn formation of the derivatives in this case trace sugar detection is sometimes possible, as nmol or pmol orders are approached. Examples include the use of dan-sylhydrazine (with either fluorimetric or colorimetric detection) (39) and l-phenyl-3-methyl-5-pyrazolone (UV absorption or electrochemical detection) (40). 

In most cases, the separation of alcohols, usually methanol, ethanol, and glycerol, is carried out contemporaneously with the separation of sugars and organic acids, and almost always the desire is to quantify all these analytes. It is seen, therefore, that the mobile phase is often an aqueous acid solution, even though only water may be used (5,9). Sulphuric acid is the one most frequently used, although phosphoric acid is preferred by some, since it is less corrosive on the components of the HPLC system (10). The concentration of sulphuric acid normally varies between 0.004 N and 0.01 N or more. The choice of acid may, however, be dictated by other considerations. This is the case, for example, with the use of a conductivity detector, which requires an appropriate conductivity suppressor system. If such a device is not available for a particular... 

Since the discovery of the reaction of phenylhydrazine with sugars to form osazones by Fischer,35 many investigations have utilized this reaction in the identification and separation of sugars, but the precise structures of osazones and their mode of formation are still in doubt.34 There is no doubt, however, that the initial reaction involves the formation of a hydrazone, which then reacts further to yield eventually a f)u-l,2-hydrazone or osazone. 

Many different solvent developers have been used in the separation of sugars and related compounds. Three of these, phenol-water, collidine-water, and 1-butanol-acetic acid-water,27 also commonly employed in the resolution of amino acid mixtures, are widely used. Other commonly used solvent developers are 1-butanol-ammonia-water, 1-butanol-ethanol-water,27 1-butanol-pyridine-water,61 ethyl acetate-acetic acid-water, and ethyl acetate-pyridine-water.26... 

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