1588 



PHOTOCHEMISTRY OF CHLOROPHYLL 



CHAP. 35 



10-2 M phosphate buffer (pH 7.4). It stayed in solution after precipita- 

 tion by acidification to pH 5, and its activity was found unchanged after 

 the pH was adjusted back to 7.4; but acidification to pH 4.5, or alkaliza- 

 tion to pH 9.3, destroyed the activity, which could not be brought back 

 even by immediate neutralization. The rate of methemoglobin reaction 

 in the presence of the extract had a maximum at pU 8.3 (a similar optimum, 

 pH 8.0, was found earlier for whole Stellaria chloroplasts in ferric oxalate 

 solution, while optimum pH values of about 6.5 were found with quinone 



lOOr 



c 



'x> 

 o 



o 



o 



100 

 time (sec) 



200 



Fig. 35.19F. Photochemical methemoglobin reduction (measured by oxy- 

 hemoglobin formation) by 0.4 ml. chloroplast suspension from Chenopodium 

 (1.2 g. leaves in 6 ml. 6% glucose, 0.033 M phosphate buffer, pH 7.4) (Davenport, 

 Hill, and Whatley 1952). Whale muscle hemoglobin: 0.78 X 10"* mole /I. 

 (A) crude suspension; (S) washed chloroplasts; (C) washed chloroplasts + 0.4 

 ml. first supernatant from washing; (D) washed chloroplasts + aqueous extract 

 from acetone extracted leaf material, equivalent to 0.4 ml. supernatant. 



or ferricyanide as oxidant, cf. section 5(6) below). The rate increased with 

 methemoglobin concentration, reaching saturation >1.4 X 10^* mole/ 

 1., it declined with increasing salt concentration in the extract (phosphate 

 buffer or ammonium sulfate). Chloride ions appeared to have no effect 

 on activity of the washed preparations. The reaction was inhibited by o- 

 phenanthroline and urethan (cf. section 5(d)). Attempts to fractionate the 

 — apparently proteidic — "methemoglobin-reduction-promoting" factor in 

 leaf extracts, by fractional precipitation with phosphate buffer or ammo- 

 nivmi sulfate, showed that activity was absent from the "globulin" fraction 

 precipitated at half -saturation with the salts, and concentrated in the 

 "albumin" fraction, precipitated only at complete saturation. 



Gerretsen (1951) suggested that ascorbic acid can play in chloroplast suspensions 

 (and in live cells as well) the role of a substitute reductant (replacing H2O as hydrogen 



