240 GEORGE H. HOGEBOOM AND WALTER C. SCHNEIDER 



tatively the enzyme activities characteristic of the latter particles. In the 

 face of these observations, it is apparent that the hypothesis that micro- 

 somes are artifacts derived from mitochondria must involve some rather 

 quaint assumptions. 



4. The Soluble Fraction 



After the removal from the liver homogenate of nuclei, mitochondria, 

 and microsomes, there remains a supernatant accounting for about 40% 

 of the original total nitrogen. Most of the nitrogen of this "soluble fraction" 

 of the cell is in the form of proteins in solution. In addition, lipids are present 

 that migrate centripetally in the centrifuge,^^'^*^ but there is apparently 

 little other material that is not in true solution. Although it has been as- 

 sumed for operational purposes that the soluble fraction represents the "cell 

 sap," the possibility must also be recognized that it may include sub- 

 stances extracted from the particulate elements of the cell. The latter con- 

 sideration imparts a definite risk to any unequivocal conclusions concerning 

 the cytochemical significance of biochemical analyses of the supernatant. 

 Nevertheless, in view of evidence indicating that soluble proteins are not 

 extracted under proper conditions from such structures as nuclei*^ -^^ and 

 mitochondria (see above) , it is probable that analyses of the soluble fraction 

 are in many instances significant from the cytochemical standpoint. As in 

 studies of the biochemical properties of the other isolated cell constituents, 

 however, all conclusions must be carefully weighed in terms of the concen- 

 tration and recovery obtained. 



The presence in the supernatant of a large number of proteins is demon- 

 strated by the complicated patterns obtained in electrophoretic studies by 

 Sorof and Cohen'^-'^^^ and Gjessing et al.^^* That a correspondingly large 

 number of enzymes should have been found in the fraction is, of course, not 

 surprising. Perhaps the best example of the role of the soluble fraction in a 

 metabolic process of major importance is found in the studies of LePage and 

 Schneider' ^^ of anaerobic glycolysis. These investigators followed the rate 

 of lactic acid formation in an anaerobic system including glucose, hexose 

 diphosphate, pyruvate, ATP, and DPN, and found that over 50% of the 

 activity of rabbit liver homogenates could be recovered in the soluble frac- 

 tion. The other cell fractions showed little or no activity by themselves 

 but were capable of stimulating the activity of the supernatant. Although 

 the complexity of the reaction is such that the rate-limiting component is 

 not known, the implication of this investigation is that the supernatant 

 contains most of the enzymes involved. The enhancement of lactic acid 



"2 S. Sorof and P. P. Cohen, J. Biol. Chem. 190, 303 (1951). 

 i9» S. Sorof and P. P. Cohen, J. Biol. Chem. 190, 311 (1951). 

 13" E. C. Gjessing, C. S. Floyd, and A. Chanutin, J. Biol. Chem. 188, 155 (1951). 



