Gasometric determination of urea with urease

Urea is changed by urease into ammonium carbonate. The ammonia is commonly determined as a measure of the urea. However, as shown by Partos 1 and by Mirkin, 2 one can also estimate the urea from the CO₂ formed. The manometric blood gas apparatus of Van Slyke and Neill 3 is particularly adapted to this determination, because of the wide range over which it yields accurate results.

Blood. The entire estimation can be carried out in the apparatus, as the blood proteins protect the urease from inactivation by the mercury. For a micro-determination, 0.2 cc. of blood and 1 cc. of 0.02 N lactic acid are placed in the apparatus, which is evacuated and shaken 1 minute to remove preformed CO₂. Five-tenths cc. of 10 per cent urease solution 4 (Squibb) and 0.1 cc. (3 drops) of CO₂ - free 0.3 M Na₂ HPO₄ solution are then added. Mercury in the chamber is lowered and raised again, to bring the urease into contact with the blood solution wetting the walls. The mixture is allowed to stand 5 minutes or more for decomposition of the urea. Three drops of N/1 lactic acid are then added, plus enough water to bring the total solution to 2 cc. The CO₂ is then extracted by 1 1/2 minute shaking, and determined, as described by Van Slyke and Neill for micro CO₂ determination in 0.2 cc. of blood. A concentration of 1 millimol of CO₂ per liter of blood is equivalent to 1 millimol or 60 mg. of urea, per liter.

For 1 cc. portions of blood, 1 cc. of 0.1 N lactic acid is used to remove preformed CO,. The amount 0.3 M Na, HPO, is increased five-fold to 0.5 cc.; the enzyme to 0.5 cc. of 20 per cent urease; and after digestion 0.5 cc. of 1 N lactic acid is added.