404 ' PHYSIOLOGICAL CHEMISTRY. 



lect this amount of distillate. Extract the distillate three times 1 

 with ether in a separatory funnel, evaporate the ether and heat 

 the residue at 160 C. for a few minutes to remove volatile fatty 

 acids. Dissolve the residue in 50 c.c. of water, filter and titrate 

 this aqueous solution of crotonic acid with -f^ sodium hydroxide 

 solution, using phenolphthalein as indicator. 



Calculation. One c.c. of y^- sodium hydroxide solution equals 

 0.0086 gram of crotonic acid, i part of crotonic acid equals 1.21 

 part of /?-oxybutyric acid, and i c.c. of T \ sodium hydroxide solu- 

 tion equals 0.01041 gram of /?-oxybutyric acid. To compute the 

 quantity of /?-oxybutyric acid, in grams, multiply the number of 

 cubic centimeters of T %- sodium hydroxide solution used by 0.01041. 



4. Bergell's Method. Render 100-300 c.c. of sugar-free 2 urine 

 slightly alkaline with sodium carbonate, evaporate the alkaline urine 

 to a syrup on a water-bath, cool the syrup, rub it up with syrupy 

 phosphoric acid (being careful to keep the mixture cool), 20-30 

 grams of finely pulverized, anhydrous cupric sulphate and 20-25 

 grams of fine sand. Mix the mass thoroughly, place it in a paper 

 extraction thimble 3 and extract the dry mixture with ether in a 

 Soxhlet apparatus (Fig. 125, page 410). Evaporate the ether, dis- 

 solve the residue in about 25 c.c. of water, decolorize the fluid with 

 animal charcoal, if necessary, and determine the content of /?-oxy- 

 butyric acid by a polarization test. 



5. Boekelman and Bouma's Method. Place 25 c.c. of urine in 

 a flask, add 25 c.c. of 12 per cent sodium hydroxide and 25 c.c. of 

 benzoyl chloride, stopper the flask and shake it vigorously for three 

 minutes under cold water. Remove the clear fluid by means of a 

 pipette, filter it and subject it to a polarization test. Through the 

 action of the benzoyl chloride all the laevo-rotatory substances ex- 

 cept /?-oxybutyric acid will have been removed and the laevo-rotation 

 now exhibited by the urine will be due entirely to that acid. 



XVIII. Acidity. 



Folin's Method. The total acidity of urine may be determined 

 as follows: Place 25 c.c. of urine in a 200 c.c. Erlenmeyer flask 

 and add 15-20 grams of finely pulverized potassium oxalate and 

 1-2 drops of a i per cent phenolphthalein solution to the fluid. 



1 Shaffer has recently called attention to the fact that it is extremely difficult 

 to extract all of the crotonic acid if but three extractions are made. 



2 If sugar is present it must be removed by fermentation. 



3 The Schleicher and Schull fat-free extraction thimble is very satisfactory. 



