METHOD FOR IDENTIFICATION OF AZO DYES 243 colors present in any unknown will be related to that of the standard dyes on the paper. Thus, by means of careful selection of solvents, the need for the other 25 compounds as standards is avoided (this is, in fact, very con- venient since many of these dyes are not easily available). The method is long and slow, since sometimes as many as five successive elutions are required. These disadvantages are balanced by a considerable increase in accuracy since errors in the Rs values are insignificant, since subjective factors are greatly reduced, and since conditions become more reproducible. In substance, the method should not lead to errors in interpretation and is not influenced appreciably by laboratory conditions. Procedure and Results. The chromatographic technique is that de- scribed for the short method (with alkaline and acid chromatograms being observed before drying). Standard solutions and unknowns are spotted separately on the starting line, but in at least one of the points it is con- venient to place standards and unknowns together. The flow sheet is given in Table 4, comment on which we believe un- necessary. The color of the spots is considered only in the final stages since unknowns are frequently mixtures of dyes which may be super- imposed in some solvents. Auxiliary colors added in small proportions for shading purposes must always be considered and should not be confused with principal coloring matters. The positions of auxiliary dyes given in Table 3 are similar to the positions in the solvents employed in this method with minor variations: The spectrum in saline solvents is closely related to that in solvent 2, in two-phase solvents to that in solvent 1, in acid solvents to that in solvent 3 and in alkaline solvents to that in solvent 4. Interpretation of the results is always easy if the unknown contains a single color. If two or more dyes are present in unknowns, the interpre- tation is also easy after the dyes are separated in the final stages of the analysis. If several spots appear in the first chromatograms, the interpre- tation becomes more complex, but, even in this case, it is possible, in many instances, to follow the analysis for each spot without interference from the others. If this is not possible, the dyes may be isolated and recovered as follows: several drops or a band of test solution instead of a single drop are spotted along the starting line after which a new chromatogram is developed with the same solvent system. The colored zones are then cut transversely, and the dye is recovered by elution with water. From this solution, acidified with tartaric acid, the color is dyed on wool, stripped with dilute ammonia and concentrated for chromatography. The indicated analysis is followed then with each recovered fraction. Although some of the dyes discussed in this manuscript are on the U.S.A. certifiable list, the samples used in this work were of British, German and

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