COMBINED REVERSED-PHASE AND ANION-EXCHANGE SOLID-PHASE EXTRACTION FORTHE ASSAY OF THE NEUROPROTECTANT NBQX IN HUMAN PLASMA AND URINE

A combination of reversed phase and anion exchange solid phase extract ion (SPE) was investigated for the liquid chromatographic assay of the neuroprotectant agent NBQX in human plasma and urine. Reversed phase SPE alone using a variety of phases failed to yield sufficiently pure extracts for the assay of plasma in the low ng/mL concentration range using UV detection. However, using Bond Elut Certify IL(TM) SPE column s containing a mixture of reversed phase and anion exchange functional ities markedly improved the purity of plasma extracts. Furthermore, a change of detector wavelength from UV (294 nm) to the visible region ( 380 nm) removed some minor interfering peaks originating from plasma. Using optimized SPE conditions with an extraction recovery of 92.3% an d a high performance liquid chromatographic procedure with Lichrospher (TM) C18 as the stationary phase, the lower limit of quantitation in h uman plasma was 2 ng/mL (corresponding to 0.75 ng injected on-column). Intra-assay coefficients of variation ranged from 9.6% at 2 ng/mL to 1.3% at 10 ng/mL. A similar SPE procedure was applied to human urine w ith acceptable recovery (88.3%), but an analytical column of different selectivity (Chromspher(TM) B C18) was necessary in order to avoid in terference from the urine. The limit of quantitation for the urine ass ay was 25 ng/mL, and the intra-assay precision ranged from 4.6% at 25 ng/mL to 2.2% at 500 ng/mL.