The first step in determining vitamins A and E includes sample preparation. In the first step, an internal standard solution is added. During the precipitation, higher molecular substances are removed. After centrifugation, the supernatant is used for injection into the HPLC system.
The separation via HPLC follows an isocratic method at 30 °C using a “reversed-phase“ column; one run lasts 15 minutes. A UV detector performs the detection at two different wavelengths (Vitamin A: 325 nm; Vitamin E: 300 nm). The quantification is performed with the delivered standard solution; the concentration is calculated by integrating the peak areas/heights in the internal standard calibration mode.