434 MISSOURI AGRICULTURAL EXP. STA. RESEARCH BULLETIN NO. 12 



quite dry, all crystals of (NH 4 ) 2 SO 4 were removed with a spatula 

 and the protein dissolved in about 75 cubic centimeters of water, in 

 which it was readily soluble. This deep yellow solution was neutral 

 in reaction. It contained a small amount of (NH 4 ) 2 SO 4 , the amount 

 of which was determined quantitatively. (NH 4 ) 2 SO 4 was added 

 to a portion of this solution to bring the concentration up to a normal 

 solution. An equal volume of saturated (NH t ) 2 SO., solution was 

 added to the remainder, giving a solution between 3 and 4 normal and 

 one similar to the one whose coagulation temperature was observed 

 above. The coagulation temperature of the pigmented protein of these 

 two solutions was carefully studied. Both solutions were found to 

 contain a small amount of colorless protein which coagulated between 

 65 and 75 C. This was filtered off, the nitrate retaining its original 

 yellow color. This nitrate was then studied further. 



The 3-4 normal or one half saturated (NH 4 ) 2 SO 4 solution acted 

 in a manner identical with the solution whose study is recorded above. 

 The first opalescence appeared between 79 and 80 C., and complete 

 coagulation did not take place until the temperature was raised to 

 86 C. 



It was not found possible to cause a clear coagulation of the 

 pigmented protein in the neutral normal solution of (NH 4 ).. SO 4 even 

 when the temperature was raised to 90 C. Opalescence began, how- 

 ever, between 82 and 82.5 C. Coagulation was readily obtained 

 when the solution was heated to 89 C. in the presence of a very little 

 HC1. (3 drops of a */2 normal HC1 solution were added to 10 c. c. of 

 solution.) 



The large amount of evidence which has now been submitted in 

 regard to the transportation of the carotin in the blood serum will 

 justify but one conclusion, namely that the carotin exists in the blood 

 in conjugation with one of the proteins. The evidence will also justify 

 the conclusion that the protein with which the carotin is combined is 

 an albumin. 



Summarizing the evidence, we have shown that the carotin car- 

 rying protein is precipitated from its solution in the serum or from its 

 aqueous solution, on complete saturation only with ammonium sulphate, 

 or by saturation with magnesium sulphate only in one per cent acetic 

 acid solution, or by heating its half saturated ammonium sulphate 

 solution to 86 C. ; the protein may also be coagulated by alcohol, or 

 by boilding its solution in the presence of acetic acid. As in all salting 

 out methods for the precipitation of proteins, the pigmented protein 

 is readily soluble in water after being thrown down by ammonium 



