CH. XIII.] CREATINE. 339 



(viii.) Add 1-5 cc. of 10 per cent, soda from a burette and 

 allow the mixture to stand for 10 minutes with gentle agitation. 



(ix.) Transfer to a 100 cc. volumetric flask and wash out with 

 distilled water to make 100 cc. 



(x.) Estimate colorimetrically as in the previous exercise. 



Calculation. This is the same as in the previous exercise, proper allow- 

 ance being made for the volume of urine used. The difference between the 

 two results is the creatine, which is usually expressed in terms of creatinine. 

 To convert this to creatine it should be multiplied by 



=1-14 (log. -0567). 



404. The estimation of creatine and creatinine (Benedict).* 



Principle. The dehydration of creatine to creatinine is very rapidly 

 effected by evaporation to dryness with hydrochloric acid. A little lead is 

 added to inhibit pigment formation, the traces of hydrogen evolved preventing 

 oxidation. It is not applicable to urines containing glucose. 



Method. Into a small beaker measure that volume of urine 

 that contains 7 to 10 mgm. of creatinine. Add 10 to 20 cc. of N. 

 hydrochloric acid and a pinch or two of powdered or granulated lead. 

 Boil down over a small free flame till nearly dry and then evaporate 

 to complete dryness on a boiling water bath. Add 10 cc. of hot dis- 

 tilled water and filter through a small plug of cotton wool into a 

 narrow 25 cc. measuring cylinder. Wash out quantitatively with 

 two successive portions of about 4 cc. of hot water. Cool by 

 immersion in cold water and make the volume up to 20 cc. Measure 

 2 cc. into a 100 cc. volumetric flask, using an Ostwald pipette. Add 

 20 cc. of saturated picric acid and 1-5 cc. of a 10 per cent, solution 

 of soda that contains 5 per cent, of Rochelle salt (to prevent the 

 formation of a cloud due to traces of dissolved lead) . After standing 

 for 10 minutes with gentle agitation, make up to the mark with 

 distilled water. A standard is simultaneously prepared from I cc. 

 of the standard creatinine solution, 20 cc. of picric acid and 1-5 cc. 

 of the 10 per cent, soda containing 5 per cent, of Rochelle salt, 

 diluted to 100 cc. after standing for 10 minutes. The two solutions 

 are read as described in Ex. 402. 



* Journ. Biol. Chern., xviii., p. 191. 



