484 W. D. Bonner, jr. 



From the foregoing it may be readily appreciated that in the tissues of 

 higher plants the complexities of the cytochrome components are not found 

 in the c and a components but in the number and diversity of the cytochromes 

 b. It is the complexity of the b components of plant tissues that gives to them 

 the characteristic visual absorption pattern on reduction, a broad band in the 

 green portion of the spectrum, a band very reminiscent of the absorption 

 spectrum one can observe in well perfused liver slices. The complexity of the 

 number of b components in plant tissues immediately raises the question as 

 to their reahty. Is it possible that there are as many as five b components ? 

 Since, on the basis of visual spectroscopy, there have been described since 

 1951, five different b components, is there a possibility that the use of more 

 refined techniques will reveal even more ? It is of the utmost importance to 

 answer these questions and also to understand the role that each of the cyto- 

 chromes b plays in electron transport to oxygen and the role, if any, of those 

 components not directly involved in the main transport chain. It is possible 

 that some cytochromes are involved in photosynthesis and in the reactions 

 of photomorphogenesis. All these questions are closely coupled to the 

 structure and function of the various inclusions found in the cytoplasm of 

 plant cells. Therefore, it is of great importance to know the location of each 

 of these different cytochromes b, should they turn out to be real. We can 

 begin to answer some of the above questions and try to point a way to the 

 efforts which will have to be invested in an attempt to answer the others. 



EXPERIMENTAL 



During the past months we have carried out a systematic investigation of 

 the cytochromes of plant tissues with two primary objectives, viz., (a) what 

 are the cytochrome b components of plant tissues and where are they located 

 in the cell? {b) are there species variation in the cytochrome composition 

 of the higher plant tissues ? This investigation has been carried out using 

 intact tissue as well as various cell-free preparations derived from them. 



The cytochrome components have been dehneated through the use of the 

 divided beam spectrophotometer developed by Chance (1957), and adapted 

 for low temperature work by Estabrook (1956). A full discussion of the 

 methodology is given by Estabrook (this volume, p. 436). The reaction cell, 

 illustrated in Fig. 1, was made according to the modification of Yocum and 

 Bonner (unpublished work) who have shown that the glycerol and devitrifi- 

 cation procedure of Keihn and Hartree (1949) need not be adhered to. Good 

 spectra can be obtained by freezing in aqueous solution provided a con- 

 centrated suspension of particulate matter is used. This procedure lends 

 itself well to the study of frozen steady states. It should be emphasized that 

 the absorption maxima obtained at low temperature in aqueous solution 

 are shifted to slightly longer wave-lengths when compared to those obtained 

 in 50 % glycerol at the same temperature. 



