Significance of mutarotation

Many of these characteristics are discussed in the following sections. 

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Fig. 2.54 presents a two-dimensional carbon-proton shift correlation of D-lactose after mutarotational equilibration (40% a-, 60% / -D-lactose in deuterium oxide), demonstrating the good resolution of overlapping proton resonances between 3.6 and 4 ppm by means of the larger frequency dispersion of carbon-13 shifts in the second dimension. The assignment known for one nucleus - carbon-13 in this case - can be used to analyze the crowded resonances of the other nucleus. This is the significance of the two-dimensional CH shift correlation, in addition to the identification of CH bonds. For practical evaluation, the contour plot shown in Fig. 2.54(b) proves to be more useful than the stacked representation (Fig. 2.54(a)). In the case of D-lactose, selective proton decoupling between 3.6 and 4 ppm would not afford results of similiar quality. 

Mutarotation of 0.3% solutions of the freshly dissolved sugars in 12 ml of 5 mM EDTA, pH 7.4 was followed. Significant differences in mutarotation rates (AK) in the presence and absence of 100 units of bovine kidney enzyme were expressed as the ratio AK/Ksp. Differences of less than 5% in these rate constants were not considered significant. Of the 18 sugars listed, nine have been tested previously as substrates for other mammalian mutarotases with essentially the same pattern as described here. The pattern of specificity indicates that a 3-point attachment of enzyme and substrate is necessary for catalysis of mutarotation. b Data from 72). 

Examination of Figure 9.11 demonstrates an excellent separation of a-glucose from (3-glucose which points out the power of GLC for the separation of these materials as well as a significant complication in the study of carbohydrates. It is well known that solutions of some carbohydrates undergo anomerization and that an initially pure form of a sugar may result in an equilibrium mixture (mutarotation) as in Figure 9.12. 

The question of the detailed composition of sugar solutions in various solvents together with the chemistry of the mutarotation reaction has continued to attract attention through the years. In their classical review, Mutarotation of Sugars in Solution, completed in 1969, Isbell and Pig-man have cited references to more than 320 papers and books (5, 6). The appearance of this volume justifies Isbells faith that carbohydrate chemistry continues to have a significant impact on the development of new concepts permeating all branches of chemistry and biochemistry (7). ... 

The numerical values of the time constants summarized in Table 1 confirm that there is no particular time for significant mutarotation to take place in rapid on-line analyses. If glucose uptake is anomerically specific, as has been shown by Benthin et al. [23], analytical results obtained with a glucose oxidase method must be corrected accordingly. The same holds true for analyses of rapid biological transients and for any spiking method (if the spiking solution happens to be freshly prepared). 

A small amount of each anomer was successfully obtained as crystals for mutarotation studies. The mutarotation curves are normal and, from this result, it was concluded that no significant proportions of the furanoses or of the aldehydo form appear during the mutarotation. This result contrasts with the complex mutarotation exhibited by )8-D-ribopyranose." The mutarotation of 5-thio-a-D-ribopyranose is strongly dependent on pH it shows a period of half-change of 10 hours at pH —4.4, and —7 minutes at pH —6.6. 

Mutarotation has limited, rather diagnostic, significance in food chemistry and technology. Practical use of this reaction is demonstrated in milk powder manufacture. Evaporation of milk at a rate lower than mutarotation of lactose yields a product with less a-lactose isomer, which crystallizes in prism- or pyramid-like form. Fast milk evaporation gives an amorphous mixture of a- and [3-lactose (5.7b). 

The observation that mutarotation of monosaccharides is retarded significantly by such solvents as l r,i f-dimethylformamide and methyl sulfoxide induced Kuhn and coworkers to investigate the pyranose-furanose interconversion more closely. Indeed, the rrii mechanism (normal a-/3 interconversion) is completely suppressed in these solvents, and the Isbell conversion (the rri2 mechanism) can be followed according to the equation ... 

Numerous workers have examined the rate constants for the mutaro-tation of D-glucose in the presence of acetic acid and sodium acetate in an attempt to ascertain whether equation 9 or 10 applies. Although the two equations differ widely, it is not easy to distinguish experimentally between them. It is probable that the concerted mechanism is of no significance in the mutarotation of sugars in aqueous solution, with the possible exception of reactions catalyzed by the water molecule, as will be discussed later. In solvents of low dielectric constant, the formation of ionic intermediates becomes less favored, and the concerted mechanism may apply. Some of the methods that have been used for studying these reactiqns and for determining the catalytic coefficients will be considered next. 

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