42O MISSOURI AGRICULTURAL EXP. STA. RESEARCH BULLETIN NO. 12 



Chromotographic analysis showed the presence of only one pig- 

 ment which very slowly passed through the CaCO 3 as a yellow zone. 



Experiment IV. 



Serum from Jersey Cow No. 2, to the amount ot 275 cubic 

 centimeters, was dessicated with a little more than the calculated 

 amount of plaster of Paris necessary to take up the water, and after 

 setting over night the hard mass was pulverized in a mortar. The 

 powder was moistened with 95 per cent alcohol and shaken with 

 petroleum ether (b.p.3O-5oC.) until the petroleum ether extracted 

 no more color, and then with ether until that extract was colorless. 1 

 The petroleum ether extract was concentrated to 50 c.c. and the 

 solution added to the alcohol-free ether-alcohol extract, which had 

 been concentrated to about 150 c.c. An equal volume of 4 per cent 

 alcoholic potash solution was now added to the combined ethereal 

 solutions, the ethers evaporated off and the alcoholic solution boiled 

 on the steam bath for a few minutes. The pigment was then ex- 

 tracted from the alkaline alcoholic solution in the usual way and when 

 in alcoholic solution was analyzed with respect to petroleum ether 

 and eighty to ninety per cent alcohol. Two pigments were thus 

 obtained with proportions of perhaps 95 and 5 per cent of the total. 



The petroleum ether soluble pigment gave a red colored residue 

 which dissolved instantly in carbon bisulphide with a blood-red color, 

 and showed the most beautiful absorption bands yet seen for this 

 pigment. Three bands were visible, the third band being considerably 

 fainter than the other two bands but clear and distinct. The measure- 

 ments are given in Table No. 2. 



1. The mechanism of this method of obtaining the blood serum pigment 

 is so interesting, its advantages so striking and its results so satisfactory, 

 that it is worthy of some discussion. 



It appears that the addition of just sufficient alcohol (either absolute or 

 95 per cent) to thoroughly moisten the dessicated serum liberates the main 

 lutein pigment in such a way that when the moistened mass is shaken with 

 petroleum ether the result is the same as if an 80 per cent to 90 per cent 

 alcoholic solution of the isolated pigment is shaken with petroleum ether. 

 There is the additional advantage however, that the CaSO 4 prevents the 

 formation of emulsions and holds the alcoholic solution so firmly fixed in the 

 paste that the petroleum ether can be poured away and the use of a 

 separatory funnel be entirely dispensed with. When all the pigment more 

 soluble in petroleum ether than in 80 per cent alcohol is thus extracted, 

 any pigment which remains can be readily extracted with ether which mixes 

 readily with the dilute alcohol. The pigment thus extracted can be readily 

 freed from alcohol by shaking with water leaving the last as well as the 

 first pigment extracted, in low boiling point solvents, an additional decided 

 advantage in view of the ease with which they are oxidized. It should be 

 added however that the method for extracting the pigment more soluble in 

 alcohol than in petroleum ether does not apply well for serum containing 

 much haemoglobin for in this as well as all other alcohol methods the red 

 pigment is somewhat soluble in the dilute alcohol. 



