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INTRODUCTION For the analysis of a sugar mixture it is generally necessary to have as many different analytical processes as there are sugars. These methods should be selected in such a way that the most varied properties of the sugars are represented. A consideration of some of these properties follows. Total sugar.-In a relatively few instances, total sugar can be determined in a mixture by densimetric, refractometric, or desiccation methods. Obviously, such methods can be applied only to solutions uncontaminated with nonsugars. The densities and refractive indices of the pure constituents must be known, and in most instances it must be assumed that the properties of the mixture are in linear relation with those of the constituents. In crude materials these methods are more frequently used for the determination of total dry substance than for total sugars and thus are contributory to the determination of purity. Polarizing power.-This property can be expressed in terms of specific rotation for a monochromatic wave length or as the rotation per gram in 100 ml, in terms of the saccharimetric scale, or as the ratio of the specific rotation of the sugar in question to that of sucrose. For many sugars the specific rotation varies appreciably with concentration, and the tendency at present is to select the concentration of total substance rather than the partial concentration of each sugar in assigning the value of the specific rotation for the calculation. The specific rotations of the sugars are tabulated on page 563. Reducing power.-For the purposes of calculation, it is convenient to state the reducing powers of the various sugars in terms of that of dextrose. The "reducing ratio" may be defined as the ratio of weights of dextrose to the sugar in question which produce the same weight of reduced copper. Thus by the Allihn method, 240 mg of levulose is required to reduce the same weight of copper as 219.5 mg of dextrose. The reducing ratio of levulose is then 0.915. In the study of starch. and its scission products, maltose is frequently used as the unit of reducing power. In some cases levulose is taken as the sugar of unit reducing power. Usually the reducing ratios of dextrose to the various sugars vary somewhat with the concentration of sugar. Thus Jackson [1], using the Munson and Walker method, found that the reducing power of levulose varied from 0.912 for 89 mg of copper to 0.937 for 375 mg of copper. With the Allihn method, the ratios appear to be more nearly independent of concentration. Browne [2] found the following reducing ratios for the Allihn method: Levulose 0.915; arabinose, 1.032; xylose, 0.983; galactose, 0.898; invert sugar, 0.958. The ratios for the more common sugars can, for the Munson and Walker method, be calculated from table 78, p. 564. The most comprehensive table of reducing ratios, including both common and unusual sugars, is that of Isbell, Pigman, and Frush (table 23, p. 190), who employed a modification of the Scales method. 2. SELECTIVE METHODS (a) RECAPITULATION OF PREVIOUSLY DESCRIBED METHODS Many of the methods of analysis described in detail in previous chapters are selective for the respective sugars and are directly applicable to their determination when they occur in admixture with other sugars. Thus the Clerget method is selective for sucrose and raffinose and is particularly free from analytical error when invertase is used as the inverting agent. Levulose is selectively determined by the modified Nyns procedure, as described on page 203, but in this case corrections must be applied for the slight reduction by other sugars. Aldoses can be distinguished from ketoses by oxidation to aldonic acids in mildly alkaline solution (p. 208). Monoses can with fair approximation be determined in the presence of reducing disaccharides by the modified Barfoed procedure (p. 206). Pentoses and pentosans upon distillation in the presence of hydrochloric acid yield furfural, which can be estimated as described on page 241. These selective methods, as well as the general methods described above, can be combined in a great variety of ways for the analysis. of sugar mixtures. It is usually true, however, that the application of the selective methods to actual mixtures must be made with caution, since frequently unexpected complications are encountered. It is for this reason that in the following pages will be described only those applications which have been thoroughly studied. (b) INVERT SUGAR BY POLARIZATION AT TWO TEMPERATURES By polarization of a solution at two widely separated temperatures, invert sugar or levulose can be determined selectively in the presence of other sugars, since the levulose constituent of invert sugar possesses a high temperature coefficient, whereas the rotatory power of dextrose is independent of temperature. Browne, however, [3, p. 298], points out that 1.5 g of arabinose, 3.0 g of galactose, 7.0 g of maltose, 9.0 g of lactose, or 50 g of sucrose, in 100 ml produces approximately the same change of polarization with change of temperature as 1 g of levulose or 2 g of invert sugar. In some instances, the partial change of rotation of these sugars with temperature can be applied as a correction. Gubbe's [4] comprehensive formulas for the specific rotation of invert sugar can be solved for the change of its rotation with change of temperature or for the temperature of complete inactivation. This inactivation is due to the fact that since the rotatory power of levulose diminishes with increasing temperature, while that of dextrose remains constant, there is some temperature at which the two rotations are equal and opposite. Gubbe's formulas [a]2-19.657-0.0361C [a] = [a] +0.3246 (t-20)-0.00021 (t-20)2 (53) (54) indicate that the specific rotation at 20° C is variable with concentration but that the temperature coefficient is independent of concentration. The temperature of inactivation therefore is not a constant but a function of concentration. Browne [3] has calculated that the temperature of inactivation varies with concentration of invert sugar from 83.2° C for 2 g to 90.2° C for 60 g in 100 ml. For general purposes, 87° C is usually taken as the temperature of optical inactivity of invert sugar. On the other hand, the temperature coefficient of the rotation of invert sugar is 0.0180° S for each gram in 100 ml. regardless of concentration. Hence invert sugar without any assumption of a temperature of inactivation can be determined by P'-P 0.0180 (t'-t) = grams in 100 ml, (55) in which P' is the saccharimetric reading at t' and P the reading at t in a 200-mm column. Obviously, any pair of temperatures sufficiently separated will serve for the determination. The suggestion seems valid that the method of determination by temperature coefficient is more reliable than that of polarization at the temperature of inactivation. Not only is there considerable experimental difficulty in accurately maintaining a temperature of 87° C, but there remains the uncertainty that inactivity has been attained. The method of temperature coefficient is free from these uncertainties, since any pair of temperatures will serve, provided they are accurately observed. After the temperature coefficient has been determined, the invert sugar is calculated by formula 55. If it is desired to determine the other constituent of the mixture, the rotation of the invert_sugar at 20° C (P20) can be substituted into the low-temperature polarization by reference to table 76, p. 563, which is computed from the Gubbe specific rotation formula 53, in which C refers to the concentration of total solids in 100 ml. The third column gives the rotation which each gram of invert sugar contributes to the total rotation at various concentrations of sugar. Note that these values are expressed in saccharimeter degrees. P20 so found can be transformed to P, if necessary, by formula 55. The rotation of invert sugar is deducted algebraically from the observed polarization, leaving a remainder which represents the rotation of the second constituent. If there is an excess of dextrose, it can be calculated quantitatively by dividing its rotation by that of 1 g of dextrose at the concentration of total sugais in the mixture. The rotation of dextrose can be selected from table 74, p. 562. Frequently the second constituent of the mixture is commercial glucose. This product as manufactured in this country is a liquid of density varying from 41° to 45° Baumé, and has a specific rotation varying from 100° to 125°. Obviously no exact determination is possible by means of polariscopic measurement, but if a specific rotation of say 108° is arbitrarily assumed, a measure of the constituent is obtained by dividing the observed rotation (corrected by deducting the rotation of the invert sugar) by 0.1600, the polarization of 1 g of the liquid product. The analyst should always state the specific rotation which is assumed for the purpose of calculation. Any other probable specific rotation can be assumed for the purposes of this calculation, and the appropriate divisor can be found in table 28, which is taken from Browne's Handbook of Sugar Analysis [3]. TABLE 28.-Rotatory power of commercial glucose (c) LEVULOSE BY POLARIZATION AT TWO TEMPERATURES By the procedure outlined in the previous section, levulose can be determined by polarization at two temperatures. The change of rotation of 1 g of levulose per degree change of temperature should be exactly twice as great as that of invert sugar, or 0.036. Wiley [5] gives the value 0.0357. The average computed value of five previous investigations [3] is 0.0362, the difference between the extreme values being about 20 percent. On the other hand, Jackson and Mathews [6] in an extended investigation found experimentally between 20° and about 70° C a coefficient of 0.03441. The value was found to be independent of concentration between 3 and 18 g of levulose in 100 ml. There is thus an outstanding discrepancy of 4.5 percent between the older values of the temperature coefficient and the recent experimental determination of Jackson and Mathews. The lower value of the coefficient has been closely verified by Lothrop [7], who found in a limited number of experiments between 20° and 70° C a mean value of 0.0341. By application of the Jackson and Mathews coefficient, 0.03441, to the analysis of levulose in honey, Lothrop found a close agreement with the levulose percentage as determined by chemical methods. This important coefficient requires further investigation. Not only is it divergent from Wiley's value, but it is inconsistent with the invert-sugar coefficient, 0.018, which should be exactly half that of levulose. It is apparent that the coefficient remains constant with varying concentration of sugar, but to what extent it is constant between different temperature intervals is at present undetermined. |