BLOOD ANALYSIS 287 



of a saturated solution of potassium carbonate, and drive off the ammonia by 

 aspiration into another tube containing 15 c.c. of hundredth-normal hydrochloric 

 or sulphuric acid. Titrate the excess of acid with hundredth-normal sodium 

 hydroxide or potassium hydroxide, 1 using methyl red or alizarin as indicator. 

 The aspiration apparatus of Meyers (see Fig. 90) may be used. 



Calculations. Each cubic centimeter of acid neutralized by the ammonia 

 during aspiration indicates o.oi gram of urea per 100 c.c. of blood, or 0.00467 

 gram of urea nitrogen per 100 c.c. of blood. In case the blood should be one of 

 the rare samples containing over 0.15 per cent of urea, all the acid will be neu- 

 tralized, and it will be necessary to repeat the determinations, using in the deter- 

 mination only i c.c. of blood. Fresh blood contains so little ammonia that it 

 may be disregarded. For further discussion of the urease method see Chapter 

 XXVII. 



2. Sugar, (a) Benedict 2 Modification of the Method of Lewis 

 and Benedict. 3 Principle. The red color obtained by heating a 

 glucose solution with picric acid and sodium carbonate is employed 

 as the basis of the colorimetric determination. The blood protein 

 is removed by precipitation with picric acid. 



Procedure. Two c.c. of blood are aspirated through a hypodermic needle 4 

 and a piece of rubber tubing into an Ostwald pipette, a little powdered potassium 

 oxalate in the tip of the pipette preventing clotting. The blood is drawn up a 

 little above the mark and the end of the pipette is closed with the finger. After 

 the rubber tubing and needle are disconnected, the blood is allowed to flow back 

 to the mark and is discharged at once into a 25 c.c. volumetric flask, or into a 

 large test-tube graduated at 12.5 c.c. and at 23 c.c. The pipette is twice rinsed 

 with distilled water, these washings being added to the blood. The contents 

 of the flask are shaken to insure thorough mixing and a consequent laking or 

 hemolysis of the blood, which is practically complete after a minute or two. 

 A solution of sodium picrate and picric acid 6 is added to the 25 c.c. mark (using 

 a few drops of alcohol to dispel foam if necessary) and the mixture thoroughly 

 shaken. After a' minute or two (or longer) the mixture is poured upon a dry 

 filter, and the clear filtrate collected in a dry beaker. Exactly 8 c.c. of the filtrate 

 are measured into a large test-tube bearing graduations at the 12.5 c.c. and 25 c.c. 

 mark, and i c.c. of 20 per cent (anhydrous) sodium carbonate solution is added. 

 The tube is plugged with cotton and immersed in boiling water for 10 minutes. 6 

 It is then removed, and the contents are cooled under running water and diluted 



1 Rose and Coleman (Biochem. Bull., 3, 411, 1914) suggest the colorimetric determina- 

 tion of the ammonia. . 



2 Benedict: Jour. Biol. Chem., 34, 203, 1918. 



3 Lewis and Benedict: Jour. Biol. Chem., 20, 61, 1915. For other modifications see 

 Pearce: Jour. Biol. Chem., 22, 525, 1915, and Myers & Bailey: Jour. Biol. Chem., 24, 147, 

 1916. 



4 It may be more convenient to draw about 5 c.c. of blood directly into a test-tube 

 containing a little finely powdered potassium oxalate and removing 2 c.c. portions of this 

 with the Ostwald pipette. 



5 To prepare the picrate-picric acid solution, place 36 gm. of dry powdered picric acid 

 in a liter flask or stoppered cylinder, add 500 c.c. of i per cent sodium hydroxide solution, 

 and 400 c.c. of hot water. Shake occasionally until dissolved. Cool arid dilute to i liter. 



6 Longer heating up to half an hour makes no change in the color. 



