vni METHOD OF PREPARING ALBUMIN- CRYSTALS 325 



half-saturated ammonium-sulphate solutions according to the method 

 invented by Hofmeister 1 and improved by Hopkins and Pinkus. 2 

 Hopkins' most recent instructions 3 for the preparation of crystalline 

 egg-albumin are as follows : 



Procure newly-laid eggs and collect the egg-white. Measure it 

 carefully, and add exactly the same amount of a saturated ammonium- 

 sulphate solution. Beat the two together till the whole mass forms 

 a stiff froth, and let it stand overnight. 



Filter off the precipitated globulins and mucoids, and to the clear 

 filtrate add very gradually, under constant stirring, a solution of 10 

 per cent glacial acetic acid till a slightly milky permanent precipitate 

 is formed. To litmus paper the .mixture by this time will be slightly 

 acid. Now add to each 100 ccm. of this milky mixture 1 ccm. of 

 the 10 per cent glacial acetic acid, when a bulky amorphous pre- 

 cipitate is formed, which in the course of five hours becomes crystalline. 

 To obtain the full yield of crystals (at least 60 grms. per litre) let the 

 mixture stand till next day. 



Pure crystals are obtained thus : Filter off the precipitate, and 

 wash it in three changes of half -saturated solution of ammonium 

 sulphate containing 1 per 1000 of glacial acetic acid. Dissolve the 

 crystals in a minimal quantity of water ; add very slowly, stirring 

 gently all the while, a saturated solution of ammonium sulphate till 

 a distinct precipitate is formed ; then add, in addition, for each litre of 

 the solution, 2 ccm. of saturated ammonium-sulphate solution. As a 

 rule, the albumin will have recrystallised in twenty-four hours. Should 

 the crystals, however, not form readily, agitate the vessel containing 

 the solution gently, but do not shake violently, as mechanical coagula- 

 tion is apt to occur. 



To remove the ammonium sulphate, wash the crystals repeatedly 

 with a completely saturated solution of pure sodium chloride contain- 

 ing 1 per cent acetic acid. 



Krieger 4 recommends for the preparation of serum-albumin crystals 

 sulphuric acid saturated with ammonium sulphate instead of acetic acid. 



These methods for preparing albumin crystals have been used 

 extensively during the last few years for the preparation of albumins, 

 for hsemocyanin, 5 haemoglobin, 6 and phyco-erythin. 7 



1 F. Hofmeister, Zeitschr.f. Physiol. Chem. 14. 163 (1889) ; 16. 187 (1891). 



2 F. G. Hopkins and S. W. Pinkus, Journ. of Physiol. 23. 130 (1898). 

 :! Hopkins, ibid. 25. 306 (1900). 



4 H. T. Krieger, Kristallinische Eiweissstoffe, med. Dissertation, Strassburg, 1899. 



5 M. Henze, Zeitschr. fur physiol. -Ohemie, 33. 370 (1901). 

 " F. N. Schulz, ibid. 24. 449 (1898). 



7 Molisch, Eotanikerzeitung, 1894, p. 177 ; 1895, p. 131. 



