1562 PHOTOCHEMISTRY OF CHLOROPHYLL CHAP. 35 



4. Different Oxidants 



(a) Ferric Salts 



It will be recalled that Hill first obtained sustained photochemical oxy- 

 gen production from chloroplast preparations by adding cell extracts of 

 indefinite composition, and later by adding complex ferric salts, such as 

 ferric oxalate. Simple ferric salts, such as nitrate or perchlorate, are un- 

 suitable because of their hydrolysis in the approximately neutral solution 

 (needed for these experiments). Potassium ferricyanide was added by Hill 

 to ferric oxalate in the assumption — based on his own observations — that 

 Fe(CN)6~* ions will not take direct part in the photochemical reaction, 

 but will oxidize (in successful competition with oxygen) the ferrous oxalate, 

 produced by the reaction, back to ferric oxalate, thus enabling the photo- 

 chemical reaction to proceed until all ferricyanide is exhausted [since ferro- 

 cyanide ions — in contrast to ferrous ions — are not rapidly reoxidized to 

 the Fe(III) level by molecular oxygen]: 



light, chloroplasts 



(35.29a) 2 Fe+s oxalate + HjO ^ 2 Fe+2 oxalate + 2 H+ + >^ Oj 



dark 



(35.29b) 2 Fe+2 oxalate + 2 Fe(CN)6-3 ^^ 2 Fe(CN)o-* + 2 Fe+2 oxalate 



(35.29) 2 Fe(CN)6-3 + H2O > 2 Fe(CN)6-' + 1^ O2 + 2 H + 



Holt and French (1946), on the other hand, found that ferricyanide can 

 take part in the Hill reaction also directly, without the addition of ferric 

 oxalate. Figure 35.16 shows oxygen liberation curves recorded by them 

 with the complete Hill solution, and with the same solution minus some of 

 its constituents: (1) Hill's complete solution: 0.02 M K3Fe(CN)6, 0.01 

 M FeNH4(S04)2, 0.50 M K2C2O4, 0.02 M sucrose, 0.17 M sodium sorbitol 

 borate or 0.2 M phosphate, (2) same, minus FeNH4(S04)2; (3) same, minus 

 K2C2O4; and U) same, minus both FeNH4 (864)2 and K2C2O4. 



The figure indicates that steady liberation of oxygen at about half the 

 maximum rate observed with the complete Hill mixture can be obtained 

 with ferricyanide alone. The rate maximum at pH 7-8 was there even 

 when FeNH4 (804)2 was left out, indicating that it was not due to complex- 

 ing of Fe+' ions with OH" ions (or some other initial step of hydrolysis). 



Hill and 8carisbrick had worked with chloroplasts from Stellaria and 

 Chenopodium ; perhaps the spinach chloroplasts used by Holt and French 

 contained enough ferric oxalate [or other organic salts of Fe(III) ] to make 

 their addition unnecessary. (Liebich found 0.05% Fe in the dry matter of 

 spinach chloroplasts; cf. Vol. I, page 377; Kohman found as much as 9% 

 oxalic acid in the dry matter of spinach leaves.) 



