II. CHEMISTRY 7 



According to Coiijard^^ treatment of tissue sections ^\dth reineckate pre- 

 cipitates choline reineckate as birefringent crystals which are readily seen 

 with a polarizing microscope. Keenan^^ has described microscopic proce- 

 dures for the quantitative detection of traces of choline as the reineckate 

 and as the chloroplatinate. 



The periodide separation is generally considered to be one of the most 

 sensitive methods of precipitating choline. Griess and Harrow-^ had utilized 

 the insolubility of the periodide to isolate choline as early as 1885. In 1896 

 Florence^^ described a medico-legal test for semen stains based upon the 

 typical crystals formed when this material was treated with iodine in po- 

 tassium iodide solution. Bocarius^" isolated the typical Florence crystals 

 and proved by chemical identification that choline was the substance which 

 gave the insoluble periodide. Booth*^ estimated that in aqueous solutions 

 potassium triiodide gives a precipitate with choline at a dilution of about 

 1:50,000. Stanek*^ studied the chemical composition of the choline perio- 

 dide precipitate and the conditions under which it is formed. A detailed 

 study of the periodide procedure for the isolation and subsequent estima- 

 tion of choline was made by Kiesel.** 



Choline may also be precipitated from water with phosphotungstic, 

 silicotungstic, and phosphomolybdic acids. ^^' ^'* Ackerman^^ used dipicryl- 

 amine as a precipitant, the choline salt being only slightly soluble in water 

 (0.02 g. in 100 ml. of water at 20°). The low solubility of the salt permitted 

 the separation of choline from betaine and aminoethanol. SchoorP^ pub- 

 lished descriptions and enlarged micrographs of the double salts of choline 

 hydrochloride with the following reagents: platinum chloride, sodium gold 

 chloride, mercuric chloride, mercuric iodide, potassium bismuth iodide, 

 picric acid, and picrolonic acid. 



Several combined water-soluble forms of choline have been isolated from 

 biological materials. In 1929 Dale and Dudley'^ succeeded in isolating 

 acetylcholine from an extract of horse spleen in sufficient quantities for 

 chemical identification. Since that time the acetylcholine in tissues has 



26 R. Coujard, Compt. rend. soc. biol. 142, 15 (1948). 



" G. L. Keenan, /. Assoc. Offic. Agr. Chemists 26, 96 (1943). 



28 P. Griess and G. Harrow, Ber. 18, 717 (1885). 



"A. Florence, Arch. Anthropol. II, 11 (1896). 



30 N. Bocarius, Hoppe-Seyler's Z. physiol. Chem. 34, 339 (1901). 



31 F. J. Booth, Biochem. J. 29, 2064 (1935). 



32 V. Stanek, Hoppe-Seyler's Z. physiol. Chem. 46, 280 (1905); 47, 83 (1906); 48, 334 

 (1906). 



" A. Kiesel, Hoppe-Seyler's Z. physiol. Chem. 53, 215 (1907). 



3" L. Lematte, G. Bionot, E. Kahane, and M. Kahane, Compt. rend. 191, 1130 (1930). 



35 D. Ackermann, Hoppe-Seyler's Z. physiol. Chem. 281, 197 (1944). 



36 N. Schoorl, Pharm. Weekhlad. 55, 363 (1918). 



3^ H. H. Dale and H. W. Dudley, J. Physiol. (London) 68, 97 (1929). 



