THE MITOCHONDRIAL CONSTITUENTS OF PROTOPLASM. 



91 



(2) Janus green C (Farbwerke Hoechst Company), dimethylsafraninazodi- 

 methvlanilinchloride : 



N 



(CH4),N \y\/\y N = N 



Fio. F. 



N (CH,), 



\y 



(3) Janus green B (Farbwerke Hoechst Company), diethylsafraninazodi- 

 methylanilinchloride : 



N 



/^/\/\ 



(aH,),N 



N = N 



N(CH,), 



\y 



Fig. G. 



The specificity of the reaction is shown by the fact that only tlie latter, janus 

 green B, the one originally recommended by Michaelis, will stain mitochondria, 

 though the others differ only in the substitution of an H., or (CH,,)-, in the place of 

 the (C2H5)2 group. The presence of the diethyl group in the safranin molecule is 

 evidently the determining factor. Compounds containing two ethyl groups are 

 more basic than those containing two methyl groups, and the compounds con- 

 taining two methyl groups are, in turn, more basic than those with two hydrogen 

 atoms alone, so that there is a decrease in basicity as we pass from the diethyl to 

 the dimethyl and to the dihydrogen. This may well e.xplain the difference in the 

 behavior of these janus greens toward mitochondria. One would expect to find 

 that janus green CJ, with the two methyl groups, would color the mitochondria 

 better than the janus green of Grtibler, which possesses only the H., group, but I 

 have failed to detect any difference between the two. Both of them occasionally 

 stain mitochondria, together with other cell structures, when used in relatively 

 high concentrations. Nevertheless, the difference in the actions of the janus green 

 B with the two ethyl groups and the other two dyes, together with the difference 

 in basicity between them, would suggest that the dye actually combines chemically 

 with the mitochondria and that the staining is not simply a process of selective 

 absorption. Our evidence, however, is too scanty to permit us to arrive at any 

 conclusion. The poor results obtained with some samples of the janus green are 

 probably due to admixtures of the first and second varieties. 



