Precolumn-venting plug technique for direct injection of untreated blood plasma samples into reversed-phase liquid chromatography systems.

A simple column switching technique for the direct injection of untreated blood plasma samples is presented for the determination of drugs and related compounds. The system consists of injector valves, a precolumn, a three port valve and a separation column. The precolumn is used for trace enrichment and sample clean-up. Aqueous plugs are introduced on both sides of the plasma sample before it enters the precolumn. This results in a stable system, because contact between plasma proteins and the organic solvent usually present in the mobile phase is prevented. The proteins are eluted to waste with the aqueous plug fluid. The stability of the chromatographic system was studied with respect to efficiency and column back-pressure. The influence of the concentration of organic solvent in the eluent, type and particle size of the packing material, precolumn filters, pH, and ionic strength was investigated for large sample volumes. Optimal stability was obtained at low concentrations of organic solvent in the eluent, with plugs of phosphate buffer (mu = 0.1) and with a spreader at the inlet and a screen (2 microns) at the outlet of the precolumn. The precolumn was packed with 10-microns particles for trace enrichment. Under favourable conditions 40-50 large-volume (0.5-ml) plasma injections can be made into a single precolumn.