66 VITAMINS A AND CAROTENES 



The chemistry and biochemistry of vitamin A 2 have only lately been 

 clarified, with its purification and the preparation of crystalline derivatives 

 by Shantz,!'' the accurate estimation of its biological activity by Shantz 

 and Brinkman,!^ and the total synthesis of vitamin A2 and retinene2 by 

 Farrar et al}"^- ^^ The latter research has established the structures shown in 

 Fig. 15. The presence of a second double bond in the ring had been sug- 

 gested earlier by Gray and Cawley^^ and by Morton et al}^ Other properties 

 of vitamin A2 and retinene2 are listed in Table XVI. 



Retinene2, like retinenei, is easily prepared from the corresponding vita- 

 min A by chromatographic oxidation on manganese dioxide.^*' ^^ We have 

 found it best in this process to use only about 0.3 g. of manganese dioxide 

 to oxidize 10 mg. of vitamin A2, half as much therefore as is used in oxidiz- 

 ing vitamin Ai.'® 



D. THE ALCOHOL DEHYDROGENASE SYSTEM 



Whereas the usual solutions of rhodopsin yield retinene and opsin as the 

 end products of bleaching, in the retina retinene is reduced quantitatively 

 to vitamin A. 



This process is catalyzed by a soluble enzyme system, to which we first 

 gave the name retinene reductase. It consists of an enzyme protein or apo- 

 enzyme, working together with the coenzyme cozymase, coenzyme I or DPN. 

 The latter is one of the major coenzymes of biological oxidation and reduc- 

 tion, the substance which in yeast reduces acetaldehyde to ethyl alcohol 

 and in muscle reduces pyruvic acid to lactic acid. In the retina it transfers 

 two hydrogen atoms to the aldehyde group of retinene, reducing it to the 

 alcohol group of vitamin A:-^ 



retinene reductase or 

 alcohol dehydrogenase 



C19H27CHO -1- DPN-H2 ■ C19H27CH2OH + DPN 



Retinene -|- dihydrocozymase Vitamin A -\- cozymase 



In the porphyropsin system the same enzyme reduces retineneo to vita- 

 min A2. The enzyme proteins of both systems are interchangeable, and the 

 coenzyme is identical in both.-^ 



This enzyme system is readily assembled in solution. The protein is 

 readily extracted with buffer solutions from homogenized retinas or from 

 suspensions of the outer segments of the rods. Reduced cozymase is pre- 

 pared by Ohlmeyer's procedure.^*^ As substrate, retinenei or retinene2 is 



26 E. LeB Gray and J. D. Cawley, J. Biol. Chem. 134, 397 (1940). 



26 R. A. Morton, M. K. Salah, and A. L. St.ubbs, Nature 159, 744 (1947). 



" G. Wald, Biochim. et Biophiis. Acta 4, 215 (1950). 



28 G. Wald and R. IIul)l)ard, ./. Gen. Physiol. 32, 367 (1948-1949). 



2i' G. Wald, Science 109, 482 (1949); Biochim. et Biophys. Acta 4, 215 (1950). 



3" P. Ohlmeyer, Biochem. Z. 297, 66 (1938). 



