Ascorbic acid (AA) and dehydroascorbic acid (DHA) are small polar molecules difficult to be retained in conventional chromatographic RP systems. Hydrophilic interaction liquid chromatography (HILIC) using Obelisk R (100 x 3.2mm, 5 microm, Sielc) analytical column and isocratic elution by ammonium acetate buffer pH 4.2 was found to be successful at this task, while other tested HILIC columns--Obelisk N (100 x 3.2mm, 5 microm, Sielc) and Luna HILIC (100 x 3.0mm, 3 microm, Phenomenex) were unsuccessful for the purposes of analysis. Charged aerosol detection (CAD) has recently become a new alternative universal detection system in HPLC, and was extremely convenient for the simultaneous analysis of AA and DHA without the need of subtraction approach and oxidation/reduction step. CAD response was found linear in defined range in spite of the fact that CAD is designated as non-linear detection method. A simple and fast HILIC-CAD method was applied for the analysis of pharmaceutical preparations containing AA. Method validation was performed including parameters of precision, accuracy, linearity, limit of detection and limit of quantitation (LOQ). The method was fast, accurate and precise for both detectors with LOQ(AA) 5 microg/ml for UV detection and 10 microg/ml for CAD, respectively. DHA was detected only by CAD within tested concentration range with LOQ(DHA) 1 microg/ml.

Department of Analytical Chemistry Faculty of Pharmacy Charles University Hradec Kralove Czech Republic

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