266 THE URINE. 



direction should always precede a more detailed examination (see 

 below). If a positive reaction is thus obtained, any sugar that may 

 be present is removed by fermentation. The liquid is cleared by 

 adding neutral acetate of lead, when the filtrate is examined with 

 the polarimeter. Should Irevorotation now be observed, the presence 

 of oxybutyric acid is rendered very probable. To demonstrate this 

 beyond a doubt, the liquid is evaporated to a syrup, treated with an 

 equal volume of concentrated sulphuric acid and distilled, without 

 cooling. In this manner the oxybutyric acid is decomposed with 

 the formation of w-crotonic acid, which is accordingly found in the 

 distillate. If this is present in larger amounts, it crystallizes out in 

 the distillate, when this is strongly cooled, and may be identified by 

 its melting-point, 72 C. Should smaller amounts, however, be 

 present, crystallization does not occur. In this case the distillate is 

 extracted with ether by shaking. Traces of benzoic acid and the 

 phenols are thus likewise extracted, but if then the residue of the 

 ethereal solution is washed with water other impurities are removed, 

 and the crotonic acid remains. 



Diacetic Acid. From what has been said above, it is clear that 

 every urine which contains /9-oxybutyric acid must also contain 

 diacetic acid, and in extreme cases both may be found in the blood 

 as such. On the other hand, it will also be understood that diacetic 

 acid may occur in the urine in the absence of /9-oxybutyric acid, 

 and this is indeed more common. 



Tests. As diacetic acid is rapidly decomposed on standing, it is 

 necessary that the urine should be as fresh as possible, when it is to 

 be examined in this direction. To this end, several direct tests are 

 available. 



ARNOLD'S TEST. This test is the most reliable, as it does not 

 respond to acetone and /9-oxybutyric acid, nor to the common anti- 

 pyretics, salicylic acid, or bile-pigment. Highly colored urines, 

 however, should first be filtered through animal charcoal. 



Two solutions are employed, viz., a 1 per cent, solution of sodium 

 nitrite, and a solution of para-amido-acetophenon. The latter is 

 prepared as follows : 1 gramme of the acetophenon is dissolved in 

 from 80 to 100 c.c. of distilled water, and treated drop by drop 

 with hydrochloric acid until the solution, which at first is yellow, 

 becomes entirely colorless ; an excess, however, should be carefully 

 avoided. Before using, the two solutions are mixed in the propor- 

 tion of one part of the nitrite to two of the acetophenon solution. 

 A few cubic centimeters of the urine are then treated with an equal 

 volume of the reagent, and a few drops of ammonia. All urines 

 thus give a more or less well-marked brownish-red color on shaking, 

 and in the presence of much diacetic acid an amorphous precipitate 

 of the same color is formed. If now a small amount of the colored 

 solution is treated with an excess of concentrated hydrochloric acid 

 (1012 c.c. for every 1 c.c.), a beautiful purplish-violet color de- 

 velops if diacetic acid is present. 



