APPENDIX TO CHAPTER IV CHOLINE 151 



and extracting with anhydrous alcohol, which leaves betaine hydro- 

 chloride undissolved. The method may be shortened by omitting the 

 second precipitation with phosphotungstic acid, and in place of it 

 precipitating the filtrate from arginine at once with mercuric chloride 

 (after removal of the silver). It is also possible to combine Stanek's 

 process with mercuric chloride precipitation. 



The properties of trigonelline are similar to those of betaine and 

 the separation from choline is effected in the same way. According 

 to Schulze 3-4 per cent, of these bases escape precipitation with phos- 

 photungstic acid. In alcoholic solution 5 per cent, of the trigonelline 

 and choline escaped precipitation by mercuric chloride, but in the case 

 of betaine the loss was more than double this amount, so that it is 

 advisable to concentrate the filtrate. 



Stanek's Method for the Estimation of Choline and Betaine. 



The method [1905, 1906, i, 2] is based on the fact that betaine, 

 being a very weak base, is set free from its salts by sodium bicarbonate, 

 while choline is not. It is carried out as follows : To 25-40 c.c. of the 

 aqueous solution, containing at most 5 per cent, of the mixed hydro- 

 chlorides of choline and betaine, sodium or potassium bicarbonate is 

 added to make 5 per cent, and then a solution of 153 grm. of iodine 

 and 100 grm. of potassium iodide in 200 grm. of water is added until 

 precipitation is complete ; the precipitate consists of brown choline 

 ennea-iodide and soon becomes crystalline. It is collected on a paper 

 disk in a Gooch crucible, washed with water and transferred to a 

 Kjeldahl flask for nitrogen determination. If desired, the choline may 

 instead be recovered from the periodide by adding finely divided 

 (" molecular ") copper (see p. 122), boiling with cupric chloride and 

 copper and, after filtration, treating the filtrate with hydrogen sulphide. 

 The solution then contains the choline as hydrochloride. 



The betaine is estimated by concentrating the filtrate which passed 

 through the Gooch crucible to 25 c.c. and adding enough sulphuric 

 acid to make 10 per cent. ; the solution is then saturated with sodium 

 chloride, and the betaine is now precipitated with the potassium 

 tri-iodide solution (already used for choline). After standing for three 

 hours the precipitated betaine per-iodide is collected, washed five 

 times with 5 c.c. of saturated sodium chloride and transferred to a 

 Kjeldahl flask in which its nitrogen content is determined. 



For the estimation of choline (and betaine) in plants Stanek ex- 

 tracts the air dry material with 96 per cent, alcohol which is distilled 

 off; the aqueous residue is boiled with baryta and the barium is re- 



