A facile enzymatic technique for the estimation of nanomolar concentrations of N-phosphonacetyl-l-aspartic acid in plasma

A sensitive radiometric enzyme-inhibition assay is described for the determination of N-phosphonacetyl-l acid (PALA, NSC-224,131) in plasma. The assay is based on the inhibition by PALA, at low concentrations of the competitive co-substrate carbamyl phosphate, of L-aspartate transcarbamylase prepared from mouse spleen. The assay responds with good reproducibility and in proportional way to concentrations of PALA over the range of 2-20 nM; as such, it represents a 100-fold increase in sensitivity over existing methodologies. The applicability and pertinence of this assay to long-term pharmacokinetic studies with PALA is demonstrated by the one month persistence of nanomolar concentrations of PALA in the plasma of mice treated with a single dose of the drug.