PROTAMINS 53 



13. The Protamins. — Kossel, to whose investigations we 

 owe our comparatively extensive knowledge of these, "the sim- 

 plest proteins," prepares them in the following manner (12). 



The minced ripe testicles of herrings (yielding clupein), salmon 

 (yielding salmin) or sturgeons (yielding sturin) are shaken up in 

 water, whereby a suspension of spermatozoa is obtained. This 

 fluid is coagulated with acetic acid and the precipitate washed 

 with alcohol and ether 'and dried. About 100 grams of the dry 

 mass is then shaken up in 500 cc. of a 1 per cent solution of 

 H2SO4 for one-half hour and filtered; this extraction is repeated 

 six times and the extracts are mixed. The filtered extract is 

 precipitated with three times its mass of alcohol, the fluid is 

 syphoned off and the precipitate is dissolved in hot water and 

 again precipitated with alcohol. 



This precipitate of protamin sulphate is dissolved in about one 

 and one-half litres of hot water and the solution is allowed to 

 cool, when a small part of the sulphate separates out as a yellow 

 or brownish colored oil. This least soluble portion of the pro- 

 tamin sulphate is rejected. The supernatant fluid is collected 

 and evaporated to a small volume when the greater part of the 

 protamin sulphate separates out as an oil. The mixture is placed 

 in a separatory funnel and the oil drawn off. 



This preparation may be freed from the last traces of associated 

 nucleic acid in the foflowing manner: It is dissolved in hot 

 water and then precipitated with sodium picrate. This precipi- 

 tate is well washed with water and then freed from picric acid 

 by shaking it up with ether in the presence of an excess of H2SO4. 

 The protamin sulphate is precipitated out of the sulphuric acid 

 solution by the addition of alcohol. This precipitation should be 

 repeated. The protamin sulphate should now come down as a 

 loose white precipitate. If the precipitate has a glutinous ap- 

 pearance, the solution in water and precipitation by alcohol 

 should be repeated. Finally, the precipitate is dried at 110 to 

 120° C. 



According to Kossel, clupein and salmin are identical, the 

 formula of the sulphate being a multiple of C30H57N17O6, 2 H2SO4. 

 The most probable formula for sturin sulphate is, according to 

 the same author, 4 CseHegNigOr + 11 H2SO4. 



These substances are strong bases; when one precipitates the 

 H2SO4 out of a solution of clupein sulphate by the addition of 



