IV. DETERMINATION 87 



These are some of the problems that lie immediately ahead. They 

 scarcely touch as yd upon the most important problem of all — the general 

 role of \itainin A in ccilular metal)olism. Yet the beginninji; that has been 

 made with \itaniin A nu'tal)olism in the study of visual systems seems now 

 to lead in several directions l)eyond the retina; and it offers some promise 

 of bringing us to this issue also. 



IV. Determination 



H. H. INHOFFEN arul II. POMMER 



A. PHYSICAL AND CHEMICAL METHODS 



1. General Considerations 



A clear distinction must be drawn between determinations on vegetable 

 materials which contain only provitamins A, and determinations on animal 

 materials where both vitamin A and the provitamins are analyzed. A 

 series of color reactions is in use to demonstrate the presence of vitamin A 

 as well as of the provitamins; some of them are also practicable for quanti- 

 tative estimations. The best-known and most widely used reagent is the 

 solution of antimony trichloride in pure chloroform. This Carr-Price re- 

 action^ is very sensitive and gives a blue color with provitamins A and 

 vitamins A. But it must be borne in mind that all carotenoids and some 

 synthetic polyene compounds show the same color reaction. ^"^ Thus, the 

 appearance of a blue color cannot be regarded always as a proof, and it is 

 therefore necessary to determine at least the maximum of the light absorp- 

 tion of the blue color which for vitamin Ai is near 617 m^, for vitamin A2 

 near 693 m/i, and for /3-carotene near 590 m/u.^ Recently, F. D. Collins^^ 

 has published that for /3-carotene another maximum of absorption lies near 

 1020 m/i. As this blue color follows the Lambert-Beer law, it can also be 

 used for the determination of the quantitative ratios. As is well known, all 

 spectrophotometric methods of determination are based on this law. Its 

 equation is 



log:p = kcd = E 



in which /o represents the intensity of the incident light. Id is the intensity 

 of the emerging light (monochromatic), k is the extinction coefficient per 



' F. H. Carr and E. A. Price, Biochem. ./. 20, 497 (192G) . 



'* P. Karrer and E. Jucker, Carotinoide. Verlag Birkhiiuser, Basel, 1948. 



* R. A. Morton, J. Phann. and Pharmacol. 2, 129 (1950). 



»" F. D. Collins, Nature 165, 818 (1950). 



