ABSTRACT
For analysis of apolar PS oxidation products (ASOPs), the approach described above is not suitable. Due to the LC clean-up, apolar oxidation products such as ketoenes and ketodienes, which elute before the main sterol peak, are not collected and thus not detected by GC-FID. Oxidation products such as the hydroperoxides are not suitable for quantification by GC due to their thermal instability. To detect these apolar and polar PS oxidation products and to investigate fast options to quantify their levels a new method was developed based on NPLC with evaporative light scattering detection (ELSD). This approach would circumvent the time-consuming LC sample clean-up and TMS derivatization. The NPLC approach provides a good separation between the different types of oxidation products owing to the differences in polarity. However, unlike the GC approach, there is no separation between the different PS oxidation products of one type, e.g., 7-hydroxy-PS. The detection limit that can be achieved is limited by the huge
response for the unoxidized PS. Nevertheless, a good signal-to-noise ratio for both apolar and polar oxidation was observed (Fig. 18); the limit of detection for individual components was estimated at 5mg/kg. This is still 50-fold higher than for the GC method, which was also reflected in the results when the same sample was analyzed for the concentration levels of the polar oxidation products by both the HPLC-GC-FID and the HPLC-ELSD approach. The latter technique gave systematically lower values because components below 5mg/kg were not detected. These usually are compounds like 25dihydroxy-PS and/or oxidation products of PS present at relatively low levels, e.g., brassicasterol and avenasterol. The main advantages of this approach remain speed of analysis and the ability to quantify apolar oxidation products.
