222 



G. WALD 



VOL. 4 (1950) 



0.3 



0.1 



300 20 40 60 80 400 20 40 60 00 



Wavelength- mu. 



of Ohlmeyer (1938); the substrate, synthetic retinenci; 

 and the apoenzyme, contained in a clear, almost colourless 

 extract of homogenized frog or cattle retinas. When these 

 three components are mixed and incubated for 1-2 hours 

 at room temperature, the retinencj is quantitatively 

 reduced to vitamin Aj (Fig. 6, upper half). 



Fig. 6. The action of frog retinene reductase on synthetic retinene^ 

 and retinencj. Each of the experimental mixtures included a 

 synthetic retinene dissolved in 1% digitonin, 0.7 mg of reduced 

 cozymase per ml, 5.5 mg of nicotinamide per ml, and extracts of 

 homogenized frog retinas in m/30 phosphate buffer, pn 6.81. The 

 controls differed only in that the retinal extracts were replaced 

 with either the same extract which had been boiled for Yo minute 

 (upper figure) or with the phosphate buffer alone (lower figure). 

 The enzyme and control mixtures were incubated together for 

 2 hours at 23° C. Methanol was added to each to a concentration 

 of 60%, and they were extracted with hexane. The spectra of the 

 hexane extracts are shown. Those from the controls (solid circles) 

 show the spectra of the unaltered retinenes; those from the enzyme 

 mixtures (open circles) show complete conversion to the corre- 

 sponding vitamins A. 



RETINENE2 AND VITAMIN A.; SPECIFICITY OF RETINENE REDUCTASE 



In the rods of freshwater fishes, cyclostomes and certain amphibia, rhodopsin is 

 replaced by the purple, light-sensitive porphyropsin. This takes part in a retinal cycle 

 identical in form with the rhodopsin system, but based upon the new carotenoids, 

 retinenCo and vitamin A_^ (Wald, 1937; 1945-46): 



Porphyropsin 

 (522 m^) 



\ 



\light 



\Russet intermediates 



\ \ 

 \ ^ 

 Retinencj -|- protein 



Vitamin A^ -\- protein 



I 

 (345-350 m/n in petroleum ether) 

 (-f SbClg > 692-696 m/u) 



(384 m/t in petroleum ether) 

 {+ SbCl3 > 705 m//) 



The structure of vitamin Ag is still uncertain. It seems clear, however, that like A, 

 it is a primary alcohol; and that retinene.j, as emerges from experiments of Morton et al. 

 and from those discussed below, is in all probability its aldehyde. 



Morton, Sal ah, and Stubbs (1946) reported that when solutions of vitamin Ao 

 in petroleum ether are let stand in the cold over solid manganese dioxide, the vitamin 

 is replaced by a product resembling retinene^ in spectrum and antimony chloride reac- 

 tion. They found that this product forms, as does retinenCi, a 2-4-dinitrophenyl- 

 hydrazone, indicating the presence of a carbonyl group. That this substance possesses 

 a conjugated carbonyl group is shown also b}^ a large displacement of its spectrum in 

 References p. 228. 



