TRANSAMINASE ACTIVITY IN HUMAN BLOOD

Abstract

Enzymatic transamination consists of the enzyme catalyzed reversible transfer of the alpha amino nitrogen of an amino acid to an alpha-keto acid with the synthesis of a second amino acid and a second alpha-keto acid. Enzymes catalyzing different transamination reactions are found widely distributed in animal tissues and have been shown to change in activity in some tissues during disease (1-3). These observations prompted the present study to determine if transaminase activity could be demonstrated in human serum and blood cellular elements and, if so, to study any variations in activity of this enzyme in the blood of normal and diseased man. METHODS AND MATERIALS The two transaminases found most active in animal tissues are: 1. "Glutamic-oxalacetic transaminase" Aspartate + alpha-keto glutarate = glutamate + oxalacetate 2. "Glutamic-pyruvic transaminase" Alanine + alpha-keto glutarate = glutamate + pyruvate When aspartate or alanine are incubated with alphaketo glutarate and a source of enzyme, the rate of production of glutamate may be taken as a measure of transaminase activity. The amount of glutamate produced after a given incubation period under standardized conditions was measured by quantitative paper chromatographic analysis (4). One-tenth molar solutions of I-aspartate, I-alanine, and alpha-keto glutarate were prepared in 0.06 M phosphate buffer and the pH of the solutions adjusted to pH 7.6. For serum transaminase determinations, 0.5 ml. of clear, non-hemolyzed serum, 1.5 ml. of 0.06 M phosphate buffer, pH 7.6, and 0.5 ml. of either the alanine or aspartate solutions were incubated for ten minutes at 37°C. At this time, 0.5 ml. of the alpha-keto glutarate solution was added and the incubation continued for 18 hours. For whole blood hemolysate transaminase determinations, equal volumes of blood and distilled water were shaken together for ten minutes, 1.0 ml. of the hemolysate was added to 1.0 ml. of the phosphate buffer, and the substrates added as above. The time of incubation of the hemolysate substrate mixture was three hours.