176 THE ESTIMATION OF PROTEINS. 



tion after precipitation of the casein and albumin ; this consists 

 chiefly of the unprecipitated portions of casein and albumin. 

 It may be estimated by precipitation as copper salt by Ritt- 

 hausen's method, as described above ; by precipitation by tannin 

 and estimation of the nitrogen in the precipitate by KjeldahFs 

 method ; or by evaporation of. the solution and estimation of 

 the nitrogen. 



Sebelein's Method, though more tedious, is preferable to the 

 above ; to 10 grammes of milk, 20 c.c. of a saturated solution of 

 magnesium sulphate are added. Solid magnesium sulphate in 

 the form of powder is then added in small quantities at a time 

 till no more is dissolved. The solution is allowed to stand for 

 twelve hours and filtered ; the precipitate is washed four or five 

 times with a saturated solution of magnesium sulphate, an 

 operation which takes some time. The filter and its contents 

 are dropped into a Kjeldahl digestion flask, 30 c.c. of sulphuric 

 acid added, and the nitrogen estimated, as previously described ; 

 an increased volume of soda solution to neutralise the 30 c.c. 

 of acid used must be employed. The nitrogen multiplied by 

 6-39 will give the weight of casein. 



The magnesium sulphate must be free from sodium sulphate, 

 commercial " Epsom salts " sometimes containing this impurity. 

 If distinct acidity be developed in the milk, this should be 

 neutralised previous to the addition of magnesium sulphate. 



The albumin is separated by diluting the filtrate and precipi- 

 tating by the addition of tannin, or phospho-tungstic acid ; the 

 precipitate is collected on a filter, and the nitrogen therein esti- 

 mated by Kjeldahl's method. The albumin may be estimated 

 less exactly by boiling the filtrate after dilution and addition of a 

 small quantity of acetic acid ; it is collected on a tared filter and 

 weighed as such. 



In order to avoid the tedious washing with a saturated solu- 

 tion of magnesium sulphate, Leffmann and Beam take a larger 

 quantity of milk (say 20 grammes), dilute with twice its bulk of 

 saturated magnesium sulphate solution, add powdered magnesium 

 sulphate till saturated, and make up to a definite volume with 

 saturated magnesium sulphate solution in a graduated cylinder. 

 The solution is allowed to stand and the lower clear portion is 

 removed by a pipette ; this is filtered and an aliquot portion 

 taken ; the albumin is estimated in this, as directed above. 

 The casein is determined by subtracting the albumin nitrogen 

 from the total nitrogen and multiplying the difference by 6-39. 



Sodium chloride, to which a little calcium chloride has been 

 added, can be substituted for magnesium sulphate ; the preci- 

 pitate is less easy to treat, owing to the formation of hydrogen 

 chloride on heating the precipitate with sulphuric acid. 



