Characterization of Metal Sites on Reverse Phase Chromatography Column

In liquid chromatography, separations are ideally governed by analyte interactions with active sites. However, the presence of metal contaminants can result in metal-analyte interactions that can have detrimental effects on the performance of chromatography processes. Metal contamination is not always avoidable, and can be introduced as impurities in the media manufacturing process, as impurities in the feed stream, and from equipment used. Therefore understanding the metals contamination in a chromatography column, and its impact on separation performance is of interest. In this work, we characterize the adsorption behavior of the 4, 4-bipyridyl and 2, 2-bipyridyl on the metal sites of reverse phase chromatography columns. Because of its geometry, 4, 4-bipyridyl does not readily form a chelate with the metal sites, while 2, 2-bipyridyl does. In this study, we compared the adsorption strength of the compounds on metal sites vs. on the hydrophobic sites. To prevent the results from being complicated by exposed silanol sites, we compared the results with and without sulfinic acid.

Through a bi-Langmuir adsorption model, we were able to determine the population of the metal sites and estimated the adsorption strength. The simulation and parameter estimation were conducted using the CADET model. A few different metal contaminations were studied: Zn²⁺, Fe³⁺, Cu²⁺. Furthermore, we explored strategies to control the effect of metal through use of EDTA as an additive. The mathematical model also revealed whether EDTA removes the metals from the stationary phase or simply masks the metal-analyte interaction. The results of this study can be used to characterize the extent of metal contamination in a column, and therefore troubleshoot column performance issues on both analytical and preparative chromatography. Based on the data obtained through this study, we proposed a strategy to characterize column life.