596 PRACTICAL ORGANIC AND BIO-CHEMISTRY 



(1) Standard Acetone Solution. 



Folin and Denis have found, like Marriott, that freshly distilled solu- 

 tions give more turbidity than undistilled. In order to avoid continual 

 distillations for the preparation of the standard solution, they dilute a stronger 

 solution with ^N sulphuric acid and thus obtain a standard which keeps for 

 several weeks. It is prepared by diluting 2 c.c. of pure acetone, prepared 

 from its bisulphite compound, with 500 c.c. of water, distilling and collecting the 

 distillate in 100 c.c. of *25N sulphuric acid. The volume is diluted with acid 

 to about 1000 c.c. and its acetone content determined by titration with iodine 

 and thiosulphate (p. 593). The calculated quantity of this solution is diluted 

 with '25N acid so that 10 c.c. contain '5 mgm. Further standards are pre- 

 pared by taking this quantity, distilling and diluting with acid so that 10 c.c. 

 of the distilled diluted solution contain *5 mgm. 



(2) Acetone Estimation. 



0*5-5 c - c - f urine (sufficient to give '5 mgm. acetone) are put in the 

 test tube with i c.c. of 10 percent, sulphuric acid and the acetone aspirated 

 into 10 c.c. of 2 per cent, sodium bisulphite solution. 1 The aeration for -2 

 mgm. is complete in 10 minutes with a good water supply. 



(3) Acetone + Aceto- Acetic Acid Estimation. 



Fresh urine generally contains 2-10 times more aceto-acetic acid than 

 acetone, so that the sample must be diluted so as to contain '3-7 mgm. of 

 total acetone. 



The suitable quantity of urine is put into the test tube with i c.c. of 10 

 per cent, sulphuric acid and is heated in a boiling water-bath, the acetone 

 being collected as above. An extremely slow air current is used for the first 

 10 minutes and is then increased, but need not be rapid, for another 5 

 minutes. 



(4) Comparison with the Standard Solution. 



The solutions are transferred to a 100 c.c. measuring flask, diluted to 50 

 or 60 c.c., 15 c.c. of Scott-Wilson reagent are added, made up to 100 c.c. 

 and mixed. At the same time 10 c.c. of standard acetone solution (-5 mgm.) 

 are added to 10 c.c. of the same bisulphite solution 2 in a TOO c.c. flask and 

 diluted to 50 or 60 c.c. ; 15 c.c. of reagent are added, made up to 100 c.c. 

 and mixed. The turbidities are compared in a Duboscq colorimeter and are 

 directly proportional (i mgm. acetone = i'8 mgm. aceto-acetic acid). 



Estimation in Blood (Marriott). 



The blood is drawn by puncture; 1-2 c.c. in cases of acidosis, 5 c.c. 

 in cases with little or no acidosis. It is run into a weighed 50 c.c. conical 

 flask containing 20 c.c. of i per cent, potassium oxalate. The flask is weighed 

 and the increase gives the amount of blood. Sufficient colloidal iron solution 

 is added to bring the volume to 50 c.c. and the solutions are mixed; '5 gm. 

 of sodium sulphate is added, the flask is stoppered and well shaken to dissolve 

 the sulphate and break up the mass of precipitate. The thick liquid is 

 put into a centrifuge tube and centrifuged for about 2 minutes. The clear 

 liquid is poured off through a filter paper and used for the estimation. 



This procedure is similar to that used by Gardner and Maclean for 

 glucose estimations in blood (p. 588) and the same quantities and manipulation 

 could be equally well taken. 



The acetone determination is made in 15 or 30 c.c. of the filtrate by 

 distillation with 30 c.c. of sulphuric acid of sp. gr. 1*59, redistilled with per- 

 oxide and sodium hydroxide as in the general method (p. 593). This gives 

 pre- formed acetone + acetone from aceto-acetic acid. By aeration the pre- 

 luniK-d acetone only is obtained. 



The clear liquid free from proteins could be used for the acetone 

 estimations by the Folin and Denis procedure, 



1 This dilute solution does not keep for more than a week, but is easily prepared when 

 required. Dilute bisulphite retains the acetone perfectly ; if water be used, the air current 

 must be much slower. 



* This must be used to obtain strictly comparable turbidities, 



