Agnieszka Augustyniak, Elżbieta Skrzydlewska

Zasady doboru warunków elucji gradientowej i ich wykorzystanie do oznaczania leków
2006-08-16

Rules for gradient elution conditions selection and their use in drug determination. Gradient elution is widely applied in analytical chromatography to reduce the separation time and to improve the separation of mixtures by varying solvent strength during the elution process. The first part of the paper deals with the general issues concerning the selection of conditions in gradient elution, in which the following additional variables can be selected: the shape of gradient, eluent composition at the beginning and at the end of analysis, flow-rate and program time (defined as gradient volume Vg) as well as parameters of the column. Nowadays HPLC systems enable to apply various kinds of gradients shapes: linear (multi-linear), stepwise and nonlinear (concave and convex). In the case of acids/bases analytes, whose retention strongly depend on pH of the eluent, pH gradient RP-HPLC method is used, whose principles and application have been described in the second part of the paper. It can be noted that pH gradient provides relatively narrow, symmetrical peaks of approximately the same width, with minimized tailing. The decreased peak tailing might be due to the peak compression. The beneficial effect in determination of some drugs characterised by acid-base properties, such as opipramol results from the use of combined pH/organic gradient, which is realized by linear changes of pH, accompanied by linear increase of organic modifier content in the mobile phase.