Influence of mobile phase conditions on the clean-up effect of restricted-access media precolumns for plasma samples injected in a column-switching system

Various mobile phases including phosphate buffer, pure water and five kinds of biological buffers (pH around 7) were systematically studied in terms of their ability to clean-up plasma matrix on precolumns of restricted-access media in a column-switching system. The necessary washing time, buffer pH, type and content of organic modifier were evaluated with respect to plasma elution profiles on restricted-access media precolumns. The influence of different mobile phases on the recovery of plasma matrix from alkyl-diol silica precolumns was studied by means of a scanning spectrophotometer. Our results show that phosphate buffer near physiological pH with small amounts of 2-propanol or acetonitrile was preferable for direct injection of large plasma volumes (500 mu L). More than 93 % of the proteins in a plasma matrix can be recovered within 3 min from the alkyl-diol silica C-18 column (25 m x 4 mm I.D.) as measured at 280 nm for all selected mobile phases except for tris (hydroxymethyl) aminomethane buffer, from which only about 88 % was obtained.