ESTIMATION OF CREATININE. 697 



acid the violet or purple-red color passes into' blue, which soon becomes pale 

 and finally a pale yellow color. It differs from the creatinine in color and 

 the absorption band as well as in that the creatinine reaction requires more 

 sodium nitroprusside. 



The best method for preparing creatinine is the following, suggested 

 by FoLiN. 1 The creatinine is first precipitated as the double picrate 

 of creatinine and potassium by means of picric acid according to JAFFE'S 

 method, and then this precipitate, while still moist, is decomposed by 

 KHCOs and water. The solution, which contains the creatinine besides 

 potassium carbonate and small amounts of impurities, is neutralized 

 with sulphuric acid and the sulphate precipitated by alcohol. The 

 creatinine is now converted into the double zinc-chloride salt and this 

 last treated with moist lead hydroxide. After the removal of the lead, 

 the solution contains a mixture of creatinine and creatine, which last is 

 completely transformed into creatinine by heating for forty-eight hours 

 with normal sulphuric acid. After exact neutralization with barium- 

 hydroxide solution it is concentrated to the point of crystallization. 



According to recent work of FOLIN and BLANCK the creatinine-zinc chloride 

 can be dissolved in warm 10 per cent sulphuric acid when creatinine-zinc alum 

 (CJIrNsO^SCXZnSO^ 8H 2 is obtained and from this the creatinine can be 

 obtained by decomposing with barium acetate and removing the zinc by H 2 S. 

 Creatine can, according to FOLIN and DENIS, 2 be transformed into creatinine by 

 heating in an autoclave for 3 hours under a pressure of 4-5 kg. per qcm. 



The quantitative estimation of creatinine used to be performed accord- 

 ing to NEUBAUER'S method for the preparation of creatinine, or more 

 simply by SALKOWSKi's 3 modification of this method. As this method 

 is now seldom used we refer the reader to other hand-books. 



FOLTN 4 has suggested a colorimetric method for determining creatinine 

 which is based upon JAFFE'S picric-acid reaction and is as follows: 10 

 cc. of the urine are treated in a graduated flask of 500 cc. capacity 

 with 15 cc. of a 1.2 per cent solution of picric acid and 5 cc. of a 10 

 per cent NaOH solution. After shaking and allowing to stand for five 

 minutes it is diluted with water to 500 cc. and mixed. This solution 

 is now compared in a DUBOSCQ colorimeter Jwith a 1/2 normal potassium- 

 dicromate solution. The latter solution has in a layer 8 mm. thick 

 exactly the same intensity of color as a layer 8.1 mm. thick of a solution 

 of 10 milligrams creatinine after the addition of 15 cc. picric-acid solu- 

 tion and 5 cc. NaOH solution and dilution to 500 cc. The calculations 

 are simple. For example, in case the urine tested in a layer 7.2 mm. 

 thick has the same color as the dichromate solution in a layer 8 mm. 

 thick, then the quantity of creatinine in 10 cc. of the urine will be 



8 1 

 = =^X10, or 11.25 milligrams. This method has been tried by many 



authorities and found to be trustworthy. 



1 Zeitschr. f. physiol. Chem., 41. 



2 Folin and Blanck, Journ. of biol. Chem., 8, with Denis, ibid., 8. 



3 Zeitschr. f. physiol. Chem., 10 and 14. 



., 41. 



