488 Messrs. J. A. Gardner and F. W. Fox. Origin and 



We determined, therefore, to study the autolysis of various organs, and in 

 this paper give an account of our experiments on liver and spleen. 



Method. 



The whole fresh organ, freed as completely as possible from adhering tissue 

 and fat, was finely minced in a mincing machine, pulped in a mortar, and 

 divided into the required number of approximately equal portions, which 

 were then accurately weighed. One portion was analysed at once and the 

 others mixed, in some cases, with sufficient toluene, and in other cases with 

 a 2 per cent, solution of sodium fluoride to prevent bacterial growth. They 

 were then placed in sterile flasks in the incubator at 37° C. and allowed to 

 autolyse for definite periods. The autolysed tissue was not examined 

 bacteriologically, but the smell indicated that no appreciable putrefaction 

 had taken place. 



Extraction of Fat. 



This is the most important stage of the process, and one in which, in our 

 'experience, error is most likely to occur. The pulped tissue — fresh or auto- 

 lysed — was mixed with excess of alcohol and the fluid portion then drained 

 off. The solid matter was placed in a paper thimble and extracted with 

 alcohol for several days, the material being taken out at intervals and 

 re-ground. Finally the extraction was completed by means of ether. 



The alcoholic fluids were evaporated and the residue thoroughly extracted 

 with ether. The ethereal extract was added to the above and made to known 

 volume. An aliquot portion was evaporated to obtain the total ether extract, 

 and, when desired, to estimate the free cholesterol. 



The extraction was carried out in a flask or matrass with a very long wide 

 neck. The paper thimble was placed in an ordinary Soxhlet tube with 

 syphon, suspended by a platinum wire in the neck of the flask. Above this 

 was suspended a closely wound glass spiral tube through which cold water 

 circulated, and which served as a very efficient condenser. By varying the 

 height of the Soxhlet tube and the condenser the apparatus could be adapted - 

 for use with various solvents, and the temperature of the actual extracting 

 solvent could be varied within considerable limits. Another important 

 advantage was that all corks and joints were dispensed with, and the 

 apparatus was easy to manipulate and clean. 



Hydrolysis of the Fat. 

 The fat was hydrolysed in ethereal solution with a large excess of an 

 alcoholic solution of sodium ethoxide. as described in previous papers, but as 

 the esters of cholesterol are said to be difficult to hydrolyse in the cold by 



