Specific methods for the determination of radioactivity in D-(-)-3-hydroxybutyrate in blood plasma

Two simple, high-yield rapid methods with good reproducibility are described, which permit the determination of radioactivity in plasma D-(-)-3-hydroxybutyrate. The compound is converted to acetoacetate, using a modified enzymatic method. In procedure 1, acetoacetate is reacted with 2,4-dinitrophenylhydrazine; the resulting hydrazone is oxidised by means of a sample oxidiser, and the product 14CO2 is collected in scintillation liquid and counted. In procedure 2, a Conway microdiffusion unit is applied. The acetoacetate is decarboxylated to acetone in the presence of o-phenylenediamine, and the acetone is then diffused into semicarbazide solution. This solution, containing the semicarbazone derivative of labelled acetone, is transferred to liquid scintillation and counted. In both procedures the radioactivity is measured simultaneously in a separate sample which was not subjected to the enzymatic conversion of D-(-)-3-hydroxybutyrate. The difference in radioactivity between the two samples is attributed to labelled (D-(-)-3-hydroxybutyrate.