12 BULLETIN 433, U. S. DEPAETMENT OF AGEICULTUEE. 



Fat. — For the determination of fat, the dried residue from the moisture deter- 

 mination was crushed and exhausted with absolute ether in a Soxhlet apparatus 

 for a minimum of 16 hours. After distilling the ether, the fatty residue was 

 dried and weighed in the usual manner. 



xUh. — Three to five grams of the fresh material were weighed into a tared 

 platinum dish and ignited in an electric muffle furnace at a temperature not 

 exceeding 600° C. In this way a light, fluffy, gray ash was obtained without 

 fusing. Preliminary work showed this method to give as accurate results as 

 either the calcium acetate or the charring and extraction method, and to be 

 more convenient. 



Total nitrogen was determined by the Kjeldahl-Gunning method, using 

 3-5 grams of the original material. 



Total phosphorus. — ^\Vith a view to equalizing any possible errors inherent 

 in the methods used, so that the ratio of the inorganic to the total phosphorus 

 might be obtained with the maximum of accuracy, the same method was used 

 for the final estimation of total phosphorus as was employed in the inorganic 

 phosphorus determination. After 6 to ^10 grams of the finely ground material 

 had been digested with sulphuric and nitric acids by the well-known Neumann 

 method, the phosphorus was precipitated from the diluted and neutralized 

 solution by a considerable excess of magnesia mixture and a large excess of 

 ammonia. The magnesium ammonium phosphate was then filtered off, dissolved 

 in dilute nitric acid, and reprecipitated and weighed as ammonium phosphomol- 

 ybdate by the method of Lorenz (1901), (c. f. Neubauer and Liicker, 1912), 

 with observance of all analytical precautions. The results are reported in 

 terms of percentages of the elementary phosphorus. 



Ammonia was determined by the well-known aeration method of Folin. 

 The apparatus consisted essentially of a Folin absorption cylinder containing 

 dilute sulphuric acid for the washing of the incoming air, a Hopkins safety 

 bulb for removing any sulphuric acid spray from the air current, a 16-ounce 

 bottle of tall form for the reception of the sample, a 250 c. c. Erlenmeyer 

 flask for safety purposes, and a Folin absorption cylinder containing 10 c. c. 

 of tenth-normal sulphuric acid in which the ammonia was received. The above 

 were all connected in series in the order named, and the receiver was con- 

 nected with a vacuum system. 



About 20 grams of the ground meat were weighed accurately into the tall 

 bottle, suspended in the smallest amount of water practicable, and treated with 

 50 c. c. of a solution containing 4 per cent sodium carbonate, 0.1 per cent of 

 sodium fluorid, and sodium chlorid ^ to saturation. There was also added 25 c. c, 

 of pure ethyl alcohol, which was replaced as occasion required to prevent 

 foaming. The bottle was then connected in the series and a strong air current 

 was drawn through the apparatus for a period of at least four hours. The 

 excess of standard acid was estimated by titration, carminic acid being added 

 as an indicator. Preliminary experiments had shown that a four-hour aeration 

 period was more than sufficient. 



Results are reported in terms of percentages of elementary nitrogen. 



Preparation of extract. — Previous investigators who have determined the 

 soluble constituents of muscular tissue have, as a rule, used water as a solvent, 

 although solutions of various salts, of varying degrees of concentration, have 

 been used to some extent. Taking all of the facts concerning the nature of the 

 constituents of muscular tissue into consideration, it was concluded that the use 

 of a solvent isotonic with the muscle sei-uui would throw the most light upon 



1 Folin, O. Journal of Biological Chemistry, v. 8, p. 497. 



