two staining solutions can be made identical if different batches of 

 stain are used is to make them up on the basis of the weight of 

 actual dye present in the stain used. This can be done only if the 

 manufacturer has co-operated to the extent of printing the actual 

 dye content of each batch of stain on the container in which it is 

 sold. This is not yet commonly done; but the Commission is 

 issuing its certification only to batches of stain on which the total 

 dye content is stated. In this way it is hoped that eventually all 

 stains on the market will be so labeled; and then when staining 

 formulae are readjusted so as to call for definite quantities of actual 

 dye, the preparation of staining solutions will be put on a more 

 scientific basis. 



CHAPTER III 



THE SPECTROPHOTOMETRIC ANALYSIS OF DYES. 



DYES are extremely difficult to analyze by chemical methods. 

 Their chemistry is in many cases obscure, they often differ 

 from one another only in the way the chemical groups are 

 combined together; different dyes may react alike to all known 

 chemical tests and differ so slightly in solubility that it is difficult 

 to distinguish one from another. For all these reasons it proves 

 that spectrophotometric methods offer decided advantages in the 

 examination of dyes over methods of chemical analysis, both in 

 respect to general utility and in regard to convenience of appli- 

 cation. 



When a ray of light passes thru a prism it is resolved, as is well 

 known, into many rays differing from each other in wave length and 

 in color. Now the color of any substance arises from the selective 

 absorption or reflection of definite parts of the visible spectrum, as 

 light passes thru or is reflected from this substance. In the visible 

 spectrum is included light of wave lengths intermediate between 

 about 400 and 725 millimicrons. (The millimicron, denoted by the 

 symbol ijlijl, is one millionth of a millimeter in length.) The color 

 of the light in the spectrum varies, with increasing wave length, 

 from violet to red, appearing blue at about 450^ijLt, green at about 

 500/i^i, yellow at about 550/x/x, and orange at about 600/xju, as shown 

 in Fig. 1. The color of light which reaches the eye after trans- 

 mission thru or reflection from a colored substance is comple- 

 mentary to the color of the light absorbed by that substance. A 

 violet dye, for example, appears violet because of its predominent 

 absorption of yellow light. The complementary colors correspond- 

 ing to the various parts of the spectrum are also shown in Fig. 1 

 beneath the colors of the spectrum. 



The color of substances is ordinarily of complex origin, depending 



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