136 COLORATION IN LEIPTINOTARSA. 



be obtained in permanent condition. For example, if the black spots on the 

 pronotum of decemlineata be treated with HNO3 until the color is reduced to 

 a reddish brown, and is then washed and the acid neutralized, it will remain 

 reddish brown for as long a time as the state productive of the color persists. 

 In the case of brown species, like zetterstedti, I have not been able to produce 

 black or a darker color by chemical processes, although lighter colors can be 

 easily obtained. In these beetles the blacks are not real black, but are dense 

 deposits of a dark-brown pigment which appears black by reflected light, and 

 in zetterstedti the deposit of this brown pigment is not dense enough to allow 

 of the development of a darker color. This deep-brown pigment seems to 

 represent the end of a series of compounds beyond which no known chemxical 

 or natural process seems to be able to produce any that are darker. It repre- 

 sents the extreme of specialization of these pigments, but not necessarily the 

 extrem.e of their chemical comiplexity. 



Bottler, working on the hair of animals and on silk, has shown these pig- 

 ments to be azo compounds, and in insects they belong to this same series. I 

 have lately made a second examination of them in Leptinotarsa, and I find 

 that my present results confirm entirely the views expressed in my 1903 

 paper. I have previously shown that these colors are soluble in alcohol. If 

 now an alcoholic solution of the cuticula pigments of L. decemlineata be 

 made acid with HCl, and put into a test tube with some mild reducing agent, 

 as metallic tin, the solution passes through a series of color changes like those 

 described above, finally becoming colorless. Any mild oxidizing agent will, 

 on the other hand, produce the reverse series of color changes and restore the 

 original color. This same series of color changes is found in the case of the 

 azo compounds, of the hydrazo, azo, diazo, oxyazo, and amidoazo compounds 

 series. That these colors are azo compounds can be further proven by the 

 test with H2SO4, or with the glacial acetic-acid test. Thus, if a solution of 

 the cuticula color of decemlineata in alcohol be added to a small amount of 

 H2SO4, and then if NH^OH be added slowdy and the whole shaken, the solu- 

 tion will acquire a rose color, the depth of color depending upon the degree 

 of dilution. These tests are reliable and indicate clearly the presence of azo 

 pigments. 



V/e are safe, then, in concluding that these cuticula pigments are azo com- 

 pounds, but whether they are azo, diazo, oxyazo, or amidoazo is not known. 

 Their color and color reactions suggest a similarity to the oxyazo compounds, 

 but this similarity is possibly only superficial. 



DEVELOPMENT OF CUTICULA COLORS. 



The first study of the development of these colors was made by myself 

 in 1900, when I advanced the view that they are secretions elaborated by 

 the hypodermis and poured out upon the surface of the cuticula. Later I 



