30 



Mr. E. G. Young. On the Optical Rotatory 



experiments with both CH 3 COOHand H2SO4 in an endeavour to determine the 

 optimum P;i conditions. It soon became evident that any amorphous precipi- 

 tate which was formed before crystallisation never became crystalline. This 

 is in sharp contrast with the behaviour of ovalbumin. The conditions for the 

 deposition are spread over a much wider range of concentration both of 

 hydrogen ions and of ammonium sulphate. Crystals, moreover, will continue 

 to form as very long narrow needles for several weeks after their first 

 appearance. 



When the first crop of crystals has been obtained, centrifuged off and 

 placed in distilled water, there is always a considerable residue, although no 

 amorphous material can be seen amongst the crystals. The nature of this 

 residue has been investigated and is discussed in a later section. In order to 

 discover whether the insoluble residue remaining after the first crystallisation 

 could be eliminated by previous washing of the serum with ether, this experi- 

 ment was tried on a sample of serum with complete success. The crystals 

 formed with greater ease, probably due to the lowering of the surface tension 

 at the interfaces of growing crystal and mother liquor. The angles of the 

 crystals were very sharp and the size was uniform ; furthermore, the crystals 

 dissolved rapidly in distilled water, leaving only a minute trace of insoluble 

 matter. 



The procedure which I have found successful for obtaining a large uniform 

 yield of crystals and for reciystallisation is as follows. 



The serum is washed with an equal volume of ether in several small 

 quantities by the use of a separating funnel. Serum globulin is removed by 

 addition of an equal volume of saturated (NH 4 ) 2 S04 solution. The precipi- 

 tate is filtered off after 4 or 5 hours. The orange-yellow filtrate is slowly 

 acidified by means of acetic acid (10 per cent ) or sulphuric acid, N/3, to the 

 point of the first trace of turbidity, the solution being constantly stirred. 

 This point corresponds to a Ph value of 6'0, and a pigment change from red 

 to yellow somewhat precedes it. The cloudiness will gradually become more 

 dense as further masses of crystals separate. After two hours the acidity is 

 further increased by the addition of about half as much acid as previously 

 added. Crystals will be found to form with much greater ease once their 

 deposition has been induced at a low hydrion concentration. After another 

 two hours a third portion of acid is added, equal to the second. This should 

 establish the Ph of 4 - 9 to 5*1, below which it is not advisable to go. The 

 yield of crystalline material may be further increased by addition of more 

 (NH 4 ) 2 S0 4 solution, a few cubic centimetres at a time (1 c.c. per 100 c.c. of 

 fluid) until precipitation ceases. The crystals so separated go into solution 

 practically without residue, but the solution is strongly pigmented. Crystalli- 



