Cytochroines Cooled in Liquid Nitrogen 



449 



resides on the red side (long wavelength side) of the major band in these b- 

 type pigments, whereas it is on the blue side of the spectrum with cytochrome 

 c. Also of interest is the fact that the major absorption band of cytochromes 

 /?!, b^ and b^, that is the band of highest extinction, has its absorption maxi- 

 mum at about 553 m/*, approximately the same as that seen for cytochrome 

 c\ (Fig. 10). The alkaline haemochromes, however, differ between cytochromes 



1^ 



400 



500 



550 



Fig. 13. The pigments of rat liver microsomes. Microsomes were diluted in 

 0-1 M phosphate buffer, pH 7-4 and then a few crystals of Na2So04 were added. 

 The sample was then diluted with an equal volume of glycerol. The resulting 

 spectrum (solid line curve) shows the double a-band of cytochrome b^. A similar 

 sample reduced by NajSoOi was treated by bubbling with CO to give the dashed 

 line curve. Optical depth was 1 mm. Condition II. 



of Z)2 and b^ type and the cytochrome c\ type, the latter having the maximum 

 of the reduced alkaline haemochrome at 549 mfx. 



Of principal interest is the type of spectrum one obtains with cytochrome 

 65 of liver microsomes. The spectra of the reduced, and CO binding pigments 

 of the rat liver microsomes are presented in Fig. 13. In addition to the split 

 of the a and /i bands one sees that the addition of CO causes the appearance 

 of a large absorption band at about 448 m/^, confirming the previous observa- 

 tion of Khngenberg (1958). The split of the a- and /5-bands does not appear 

 to be influenced by the presence of CO. 



Of interest are two contradictory experiments concerning the interpre- 

 tation of the split of the a-band of cytochrome ^5 into two distinct absorption 

 bands. In an attempt to resolve the question of whether this represents the 

 contribution of two haematin components (cf. Chaix, Petit, Monier and 

 Zajdela (1958)) or one, the spectral properties of cytochrome b^ purified by 



