ED. Clark and L. H. Almy 487 
in a tall Folin aeration cylinder. 25 gm. of ground flesh were added and 
the mixture was shaken until the material was entirely disintegrated. 
50 ec. of alcohol were next added to prevent foaming, and 5 ce. of kero- 
sene later if necessary. The mixture was finally treated just previous to 
aeration with 10 ce. of 10 per cent sodium hydroxide. solution. The am- 
monia was absorbed in dilute sulfuric acid solution (10 cc. of 0.1 n sulfuric 
acid in 100 ec. of water) in 12 ounce bottles. At the end of a 43 hour aera- 
tion period, the solutions in the absorbing bottle were titrated back with 
0.1 N sodium hydroxide solution, using as WEA two drops of 1 per 
cent alcoholic solution of alizarin red. : 
Gross Fat Extraction. 
In order to obtain the fish oil in a condition as nearly unchanged as 
possible, care was constantly exercised to prevent overheating of the 
material. The ground flesh was placed in 6 inch shallow Petri dishes in 
sufficient quantity to form thin layers when pressed down on the bottoms _ 
of the dishes with a spatula. The water was driven off at 50°C. in a’ 
vacuum oven. The dried residue was ground, placed in a large paper ex- - 
traction thimble, and extracted with low boiling petroleum ether (40- 
50°C.) in a Soxhlet extractor. Aftera 12 hour extraction, the solvent was _ 
evaporated somewhat, the mixture filtered through fine filter paper, the _ 
remainder of the ether carefully evaporated, and the oil finally dried for | 
3 hours at 50°C. in a vacuum oven. 
Determination of Fat Constants. 
In some fish the amount of fat obtained was so small that but 
very few of the fat constants could be determined, due to lack of | 
material, and when such a condition existed usually only the : 
iodine number was determined. 
Specific Gravity.—The specific gravity was determined in a Sprengel or 
Ostwald pyknometer at 25°C., compared with water at the same tem- 
perature, and the constants were based on weights in air. 
« Index of Refraction.—This was determined at 30°C. with an Abbe re- 
fractometer. 
Chem., 1908, v, 71. Steel, M., An improvement of the Folin method for 
.the determination of urinary ammonia nitrogen, ibid., 1910, viii, 365. 
Shulansky, J., and Gies, W. J., Studies of aeration methods for the de- 
termination of ammonium nitrogen. The ammonium nitrogen in beef, 
Biochem. Bull., 1913, iii, 45. Clark, E. D., Report on meat and fish, 1915, 
J. Assn. Agric. Chem., 1917, ii, 229. 
THE JOURNAL OF BIOLOGICAL CHEMISTRY, VOL. XXXIII, NO. 3 
