Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
1248366 | TrAC Trends in Analytical Chemistry | 2011 | 10 Pages |
This review highlights the basis of gradient reversed-phase high-performance liquid chromatography (RP-HPLC) of ionogenic analytes.We describe pH-gradient RP-HPLC in strict theoretical terms, with examples of experiments to provide improvements in analyte separations and peak shapes, and an original method of pKa determination.Finally, we present the concept of the combined pH/organic-modifier gradient mode of RP-HPLC and illustrate it with applications. It allows optimization of separation of ionogenic analytes, along with a method for determining their biorelevant physico-chemical parameters [e.g., hydrophobicity (log kw) and acidity (pKa)]. The method is applicable to drugs and other xenobiotic mixtures, including individual analytes of interest assayed in biological fluids.