Determination of 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedionein plasma by direct injection into a coupled column liquid chromatographicsystem

The chemical substance 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexane dione (NTBC) is in clinical use for the treatment of hereditary tyrosinemia type 1. In the present study, the plasma concentration of NTBC was determi ned by a coupled column liquid chromatographic method. A 20-mu l volume of plasma was diluted with phosphate buffer, pH 2, and injected into a small p recolumn (BioTrapAcid C-18) with a mobile phase containing sulfuric acid. T he precolumn was based on the restricted access principle, i.e., retention of NTBC within the lipophilic pores, while polar and large endogenous compo unds were eluted with the void volume. NTBC was transferred to the analytic al column using a mobile phase with a high content of acetonitrile. The com pound was monitored by UV detection at 278 nm. The standard curve was linea r between 0.3 and 69 mu M, and the between-day precision (RSD) was 3% (n=6 days) at 13.8 mu M and 14% (n=6 days) at 0.3 mu M NTBC in plasma. The quant itation Limit was approximately 0.3 mu M using 20 mu l of plasma. (C) 1999 Elsevier Science B.V. All rights reserved.