LCGC North America - March 2010 - (Page 234)
234 LCGC NORTH AMERICA VOLUME 28 NUMBER 3 MARCH 2010 www.chromatographyonline.com A Global Approach to HPLC Column Selection Using Reversed-Phase and HILIC Modes: What To Try When C18 Doesn’t Work Experimental data are presented to help understand high performance liquid chromatography column retention in a way that should make column selection and method development proceed faster when initial results with C18 columns are disappointing. Three stationary phases having widely different surface properties are profiled with aqueousorganic mobile phases using a series of simple probe compounds that also represent a wide range of chemical characteristics. From these global results, it is possible to draw conclusions about the applicability of certain phase types toward classes of compounds and offer general rules for selecting a column when C18 does not work. Such an approach can replace the trial-and-error method and does not require an advanced understanding of retention mechanisms or complex mathematical treatments. This approach should be particularly helpful when trying to separate polar or ionic compounds that are not easily retained on traditional reversed-phase columns. Merlin K. L. Bicking* and Richard A. Henry† *ACCTA, Inc., St. Paul, Minnesota †Consultant, State College, Pennsylvania Direct correspondence to: mbicking@accta.com. queous–organic mobile phases (for example, water–acetonitrile) have become preferred for high performance liquid chromatography (HPLC) analysis due to their wide availability, high quality, low toxicity, and compatibility with popular detection methods. Selecting the optimum column to work with aqueous mobile phases is an important part of HPLC method development. Given the vast number of choices and the lack of chemical detail about many column products, many analysts do not take time to explore column options in any systematic way. Most prefer to simply use familiar columns having traditional C18 or closely related phases. Published column selection guides and other resources are available; however, they tend to be biased toward specific products or involve mathematical treatments that can be too complex and timeconsuming for the busy analyst. Recommendations for column selection have appeared in the literature and virtually every LC textbook. Early column selection schemes were uncomplicated, because there were relatively few column choices. However, as the number of columns and separations has grown, it has become difficult to organize the information for use in developing new methods. Historically, HPLC applications have highlighted success stories for individual columns. If one has different compounds, success depends upon the similarity of the analyte to the compounds in the published application. If one could not acquire the same column or preferred to evaluate another brand,
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