Direct Determination of Blood Ammonia by Berthelot’s Coloraiton of Protein-free Supernatant Obtained by Folin-Wu Deproteinizaion
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A direct determination of blood ammonia circumventing the use of microdiffusion apparatus which was devised by Fujii consists in (1) deproteinization of blood by a modified Folin-Wu's procedure, (2) coloration of thus deproteinized supernatant of blood by the Berthelot reaction and (3) colorimetry. This is a simple and convenient method, but our experience disclosed several drawbacks in it. A series of experiments were carried out in our laboratory in the hope that Fujii's method might be improved. The following is the outline of our improvement. 1゜. Collection of blood sample. Blood (about 3 ml) is withdrawn with a syringe by venous puncture, transferred into a test tube containing the crystals of potassium oxalate, stirred gently to prevent coagulation, and immediately cooled by inserting into a pile of ice cubes so that release of ammonia from non-NH_3 nitrogen substances may be stopped completely. 2゜. Deproteinization. To the blood (1.0 ml) thus collected are added 10 g/dl sodium tungstate solution (0.5 ml) and 1 N sulfuric acid (0.5 ml), allowed to stand (for 15 minutes), and centrifuged to get clear supernatant. (This procedure of deproteinization insures the elimination of danger for the spontaneous increase in ammonia reading which may arise while the mixture of blood with deproteinzing reagent is allowed to stand at room temperature). 3゜. Coloration. The deproteinzed supernatant of blood (0.5 ml) is mixed with phenol-nitroprusside solution (2.0 ml) and antiformin solution (2.0 ml), mixed and incubated at 37℃ (for 3 minutes). (A stable blue coloration which is not disturbed by the co-existing non-NH_3 nitrogen substances is obtained in this way.) 4゜. The standard solutions of ammonium sulfate (1.0 ml aliquots ; 100, 400mg N/dl) are treated with deproteinizing reagents and color reagents in the same way as described above (3゜) in order to construct a calibration curve. 5゜. Colorimetry at 630 mμ.
- Yamaguchi University Graduate School of Medicineの論文
Yamaguchi University Graduate School of Medicine | 論文
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