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Somsen, Govert W.; Tak, Yvonne H.; Toraño, Javier Sastre; Jongen, Peter M.J.M.; de Jong, Gerhardus J.
Journal of Chromatography A, 05/2009, Letnik: 1216, Številka: 18Journal Article
Recently, oversulfated chondroitin sulfate (OSCS) present in certain lots of heparin was identified as the toxic contaminant responsible for severe side effects following intravenous heparin administration. The United States Pharmacopeia (USP) and European Pharmacopeia (Eur.Ph.) announced an immediate revision of their monographs for heparin sodium by adding two US Food and Drugs Administration-recommended tests for OSCS based on nuclear magnetic resonance and capillary electrophoresis (CE). However, the proposed CE method provides only partial separation of the OSCS contaminant from heparin, thereby hindering appropriate impurity profiling. Here we present an improved CE method that is especially useful for the reliable quantification of OSCS in heparin samples, but also allows determination of the common impurity dermatan sulfate (DS). Parameters such as type and concentration of background electrolyte, capillary temperature, sample concentration and injection volume were investigated and optimized. Enhancement of the OSCS–heparin separation was achieved by using high concentrations of Tris phosphate (pH 3.0) as background electrolyte. High currents and excessive Joule heating were prevented by employing fused-silica capillaries with an internal diameter of 25 μm. Good separations of OSCS, heparin and DS are obtained within 17 min. The method permits injection of relatively high heparin concentrations (up to 50 mg/ml) and large sample volumes (up to 5% of the capillary volume) allowing OSCS and DS determination in heparin down to the 0.05% and 0.5% (w/w) level, respectively. The CE method is shown to be repeatable and linear ( R 2 > 0.99) for OSCS, heparin and DS. CE analyses of OSCS-contaminated heparin samples and different heparin standards further demonstrate the utility of the method.
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