RADIOMETRIC METHOD FOR MEASUREMENT OF L-ASPARAGINASE AT CONCENTRATIONS BELOW 1X10-5I.U./ML - TECHNIQUE AND APPLICATION

A radiometric method for the measurement of low levels of L-asparaginase activity (EC 3.5.1.1) was devised. This technique uses a protracted incubation at 37.degree. C to magnify amidohydrolytic activity. During this time, in most of the cases examined, L-[U-14C]asparagine is hydrolyzed to L-[U-14C]aspartic acid in a linear way; the L-[U-14C]aspartic acid so generated is transaminated with .alpha.-ketoglutaric acid by L-glutamic acid oxaloacetate transaminase (EC 2.6.1.1) and the [U-14C]oxaloacetic acid so formed is .beta.-decarboxylated by Zn2+ at pH 5.0. Using this procedure, low levels of L-asparaginase were detected in the serum of the chicken, horse and ox. Maleimide, which inhibits mammalian L-asparaginase without affecting the enzymes from bacterial sources, was used to discount the possibility that bacterial contamination of mammalian samples was responsible for the activity seen. When a survey was conducted of the distribution of L-asparaginase in the organs of Mus musculus, testis was found to contain surprisingly high levels of the analogous hydrolase from liver. The applicability of this technique to the measurement of feeble activities of L-asparaginase leeched from Dacron tubing to which L-asparaginase was covalently bound, was demonstrated.