E. E. Nelson and C. W. Greene 49 



washed with 95 per cent alcohol and transferred to a Gooch 

 crucible. The crucible was placed in a Greene's (1909) modified 

 Soxhlet apparatus and extracted continuously with 95 per cent 

 alcohol for 12 hours. This was followed with absolute alcohol 

 for 12 hours, and then ether for from 18 to 24 hours. After the 

 alcohol-ether extraction was complete the residue was transferred 

 to a weighing vial, dried to constant weight at 105C., and weighed. 

 This fraction appears in the tables as the protein residue. 



The preliminary ether extract, the ether from the Soxhlet, and 

 the ether-soluble material obtained from the alcohol-water extracts 

 as described below, were evaporated and dried at 50 C. and 

 75 mm. of mercury in a vacuum oven. The oily material was 

 dissolved in dry ether, filtered into weighing vials, and dried 

 to constant weight in the vacuum oven. This is the lipoid frac- 

 tion of the tables. At this point there was generally a small 

 amount of material which was insoluble in the ether. This was 

 dissolved in water and added to the alcohol-water extracts as 

 described below. 



The alcohol and water extracts were combined and evaporated 

 to dryness on the water bath. The dry residue was then extracted 

 with ether, and the ether-soluble material added to that from the 

 preliminary ether extraction and the Soxhlet extraction as men- 

 tioned in the preceding paragraph. The residue insoluble in 

 ether was taken up with water and made up to 250 cc. This 

 solution contains the extractives. 100 cc. were evaporated' to 

 dryness in a platinum shell, weighed, ashed, and weighed again. 

 The water-soluble solids, and the ash of the water-soluble solids 

 were obtained from these figures. 



Total nitrogen of the extractives was determined in an aliquot 

 by Gulick's modification of the Folin-Farmer colorimetric method 

 (1914). 



Creatine was determined in 10 cc. aliquots by dehydrolysis 

 and calculated as creatinine (Folin, 1914). The 10 cc. samples 

 were evaporated to dryness on the water bath with 10 cc. of 

 N HC1 and a bit of metallic lead, and the residue taken up quan- 

 titatively with hot water and washed into a 25 cc. volumetric 

 flask. 10 cc. of saturated picric acid and 1 cc. of 10 per cent 

 NaOH were added, the mixture was cooled at the tap, and allowed 

 to stand. The necessary amount of standard creatinine solution 



