Chromatographic Determination of Metallic Impurities in Reversed-Phase HPLC Columns

Abstract

A new test procedure is proposed for the characterization of packed RP columns with respect to their metal content. It is based on the peak asymmetry of 2,2‘-bipyridyl, which can be directly correlated to the metal content of the stationary phase. Since an unbuffered eluent (methanol/water, 49:51 w/w) is used, the influence of the simultaneously present silanol groups on peak asymmetry is assessed via 4,4‘-bipyridyl, which exhibits no chelating activity. A metal factor, MF, is introduced, which is the ratio of the peak asymmetries of 2,2‘-bipyridyl and 4,4‘-bipyridyl multiplied by 100. The superiority of this test compared to others is demonstrated via various commercially available columns. It is also demonstrated that columns continuously collect metal ions from the column components and the instrument hardware. Stainless steel nets and frits are the principal source of the metallic contamination. But even in totally metal-free LC systems, the stationary phase collects metal ions from the HPLC grade eluent components.