CHAPTER IV 



THE SPECTROPHOTOMETRIC AND 

 CHROMATOGRAPHIC ANALYSIS OF DYES 



CHEMICAL methods alone are inadequate in the analysis of 

 many dyes. Not only is the detailed chemistry of some dyes 

 obscure, but the reactions are often complicated by adulterat- 

 ing dyestuffs in such a manner as to preclude entire reliance on 

 rigorous chemical methods. Often a slight change in the arrange- 

 ment of atoms within the molecule will make a marked change in 

 the nature of the dye, while such a minor change in structure is not 

 always readily detected by chemical means alone. Hence the ad- 

 visability of employing certain physico-chemical characteristics as 

 displayed by the spectrophotometer in the study of dyes. So im- 

 portant is this study that a discussion of the principles involved is 

 necessary here. 



The absorptive spectra obtained by the spectrophotometer are 

 usually characteristic of any particular dye. Quantitative as well 

 as qualitative data may be obtained by the spectrophotometer; 

 and from the combined results nearly all dyes, even though differ- 

 ing from each other only in very minor particulars of chemical 

 structure may be easily differentiated. This method is not only 

 rapid but is also convenient. 



The method depends upon the fact that any colored substance 

 absor})s light of certain definite wave lengths and transmits or re- 

 flects the rest. The absorption spectrum is essentially the inverse 

 of that which is reflected or transmitted. Therefore the color of 

 light which reaches the eye after transmission through or reflection 

 from a colored substance is complementary to the color of light 

 absorbed by that substance. A violet dye, for example, appears 

 that color because of its predominant absorption of greenish yel- 

 low light. The absorption maximum is quite characteristic of any 

 dye; any two dyes having the same absorption curve (a somewhat 

 rare occurrence) are of essentially the same color. 



The wave length is ordinarily measured in millimicrons (m/^), 

 or Angstrom units (A) which are O.lm^t. The visible spectrum 

 })egins at around 400 m/x in the violet and runs to about 750m/x in 

 the red. The wave lengths of the six most important colors, and 

 their complementary colors are given in Table 1. 



A variety of terms and units have come into common usage in 

 connection with the practice of spectrophotometry and numerous 

 symbols to represent them are employed. Since there is some con- 

 fusion concerning the symbols used, the relationships between the 

 various symbols and units and definitions of the terms most fre- 

 quently encountered in the literature are given in Table 2. 



The color of substances is ordinarily of complex origin, depending 



