VOL. 4 (1950) 



RETINENES AND VITAMINS A 



219 



outer segment, unlike the whole retina, the 

 retinene^ which results is not converted to 

 vitamin Aj. The isolated outer limb lacks 

 some component of the system which per- 

 forms this conversion. 



It does not help this situation to sus- 

 pend outer limbs in the presence of intact 

 retinas. But if whole retinas are ground up 

 in Ringer solution or phosphate buffer, 

 though in the process almost all the outer 

 segments are detached from other struc- 

 tures, the suspension which results does 

 convert its retinenci efficiently to vitamin 

 Aj. The crushing of the retinal cells relea- 

 ses substances which promote this process 

 in the outer limbs. 



If such a retinal brei is centrifuged at 

 high speed and the clear, colourless superna- 

 tant solution is poured off, the solid residue 

 — which retains all the rhodopsin — has lost 

 the power to reduce retinenci. It regains this 

 capacity on re-adding to it the supernatant. 

 Fuithermore, if one suspends isolated rod 

 outer segments in such a water extract of 

 crushed retinas, they now reduce their 

 retinenci to vitamin A^. The retinal extract 

 supplies whatever the isolated outer limb 



Fig. 2. Rod outer segments of the frog, sus- 

 pended in Ringer solution. Magnification 

 about 500 diameters. The longitudinal stria- 

 tions which can be seen in most of the outer 

 limbs are characteristic of fresh preparations, 

 and probably are evidence of a fibrillar struc- 

 ture. Later, cross-striations appear, and even- 

 tually dominate the structure; the first of 

 these also are visible in the photograph (From 



W.\LD .\ND HUBB.\RD, I948-49). 



lacks for performing this conversion (Fig. 3). 



The water-soluble factor concerned with this process did not seem to involve a pro- 

 tein. It was relatively heat-stable, retaining most of its activity after boiling for as 

 long as seven minutes. Also the ease and completeness with which it left the retinal 

 tissue in a single extraction suggested that it was made up of small and relatively 

 simple molecules — perhaps a coenzyme, 01 a hydrogen-donating substrate. 



Now one would expect an enzyme protein to be rela- 

 tively specific ; and since retinenci is found only in retinas, 



Fig. 3. Rod outer limbs suspended in a water extract of retina 

 convert retinene^ to vitamin A^; washed retinal tissue is inactive. 

 Isolated rod outer limbs were frozen-dried and preextracted with 

 petroleum ether in the dark. Whole retinal tissue was ground, 

 extracted with neutral phosphate buffer, and the outer limb 

 material was suspended in the extract. Both this suspension and 

 the washed retinal tissue were irradiated, incubated, and extracted 

 with hexane. Spectra of the antimony chloride tests of these 

 extracts are shown. That from the washed retinal tissue displays 

 the band of unchanged retinenej (curve b); while the outer limb 

 preparation suspended in retinal washings has converted its 

 retinenei entirely to vitamin A^ (curve a). (From W.\ld and 



HUBB.^RD, 1948-49). 



700 



600 

 Wavelength- mu. 



References p. 228. 



