474 X. BILE PIGMENT FORMATION, ETC. 



lie 5-10 mn closer to the red than those of the choleglobin compounds 

 from hemoglobin. Since a similar difference is known to exist between 

 analogous myohemoglobin and hemoglobin compounds, it seems 

 unlikely that the globin part of the molecule is altered. This holds 

 only for the initial stage of the reaction; later, alterations of the 

 globin — and particularly of the type of linkage between globin and 

 prosthetic groups — occur, accompanied by denaturation of the 

 globin. Holden (1317) suggests that the protein of choleglobin may 

 be no longer unaltered globin; his evidence is based, however, on the 

 properties and mechanism of formation of a green compound, which 

 is only spectroscopically similar to choleglobin, but which belongs 

 to a different class of pigments (c/. under Section 6.). 



4.2.2. Spectrophotometric Analysis. The solutions obtained by 

 the coupled oxidation of hemoglobin and ascorbic acid contain hemo- 

 globin, oxyhemoglobin, hemiglobin, and ferro- and ferricholeglobin. 

 Nevertheless, a spectrophotometric analysis was worked out by 

 Lemberg and co-workers {1710). By means of carbon monoxide, 

 alkali, and dithionite the hemoglobin derivatives are converted into 

 carbon monoxide-denatured globin hemochrome, and the choleglobin 

 derivatives into carbon monoxide-denatured globin cholehemochrome. 

 By measurement of the absorptions at 630 and 570 myu the concen- 

 tration of total hemoglobin and choleglobin compounds can be found. 

 The extinction coefficients of the carboxycholehemochrome were 

 obtained from measurements on almost pure denatured globin 

 cholehemochrome (green pigment), as well as on mixtures of chole- 

 globin and hemoglobin in the initial stages of the reaction, from 

 the ratio: 



+ AeesomM/ — A €570 mi* 



Since this ratio remains constant during the first stage of the reaction, 

 it can be assumed that no other pigments are formed. 



In the later stages, however, the ratio decreases — indicating the 

 disappearance of hemoglobin with conversion into other pigments. 

 Under the conditions of the analysis, both biliverdin and verdoheme 

 compounds would be transformed by alkaline dithionite into yellow 

 compounds which do not absorb strongly at either 630 or 570 m/x. 



4.3. Structure of the Prosthetic Group 



The structure of the prosthetic group of choleglobin is not yet 

 clear. It is not identical with verdoheme, as shown by the difference 



