224 Dctcction of Aceto-Acetic Acid [Jan. 



If such a urine (B) first be boiled iinder a reflux condenser in 

 presence of a little acetic, oxalic or sulfuric acid, and the Le Nobel 

 test aiDplied, the test gives either a negative result or the response 

 is much diminished in intensity. As these urines contained aceto- 

 acetic acid, which would be destroyed by heat, it was evident that the 

 previous presence of this acid in the urine could account for the 

 anomahes observed. That this was so was estabhshed in the fol- 

 lowing way. 



The urine was acidified with oxalic acid, saturated with sodium 

 Chlorid and rendered acetone-free by aspiration for an hour with a 

 current of air, as in the Folin method of estimating acetone. At 

 the end of that time a determination of free acetone, by the Folin 

 method, showed that none was present, although the residual urine 

 responded to the Le Nobel test with undiminished intensity, and 

 the test became negative when the liquid was boiled for fifteen 

 minutes under a reflux cond.enser. 



Aceto-acetic acid, however, is stated in the literature to give a 

 faint reddish-brown or orange-red coloration with sodium nitroprus- 

 sid and ammonia — unchanged by the addition of acid. To deter- 

 mine this point, asolutionof aceto-acetic acid was madebyhydrolyz- 

 ing ethyl aceto-acetate with the theoretical quantity of potassium 

 hydroxid in the cold for twenty-four hours. This hydrolyzed Solu- 

 tion was found to respond to the Le Nobel test exactly as the urine 

 of an acidosis patient. The test became negative on boiling the 

 Solution under a reflux condenser, and was unaff ected by the removal 

 of the free acetone. 



In consequence of these facts the authors have no hesitation in 

 saying that aceto-acetic acid of itself responds to the Le Nobel test 

 and that, in the great majority of cases, a positive result when the 

 Le Nobel test is applied to a urine indicates aceto-acetic acid and 

 not acetone. On comparing the delicacy of the Le Nobel test for 

 aceto-acetic acid in urine, with the ferric chlorid test, the authors 

 found that the former will just detect about one part of aceto- 

 acetic acid in 30,000 parts of urine, while the latter fails at i part 

 in 7,000. The limit of detection of aceto-acetic acid in water by 

 the Le Nobel test is over i part in 80,000. 



