Australian Canola Oil - the influence of varietal traits, processing and storage on phytosterols, tocopherols and carotenoids

Since its development, Brassica napus (canola, or low erucic acid and low glucosinolate rapeseed) has become a substantial commodity crop in Australia, and in many parts of the world. This research explored the behaviours of several classes of health-beneficial bioactive compounds, phytosterols, tocopherols and carotenoids in canola seed and oil, as affected by genetics, processing and storage conditions. Firstly, the development of a rapid analytical method for the simultaneous determination of phytosterols, tocopherols and carotenoids was achieved, by use of high performance liquid chromatography, and diode array detection, coupled with tandem mass spectrometry (HPLC-DAD-MS/MS). The use of mass spectrometry offered enhanced selectivity to enable the determination of both individual free and esterified sterols without the need for saponification. Furthermore, the use of normal phase liquid chromatography allowed for the analysis of intact lipids, greatly reducing the sample preparation time, and presenting new knowledge concerning the determination of bioactives in lipid matrices. In an investigation of the effects of genetics and growing environment on concentrations of phytosterols, tocopherols, and carotenoids, significant genotype (G), and environmental (E) effects were observed for all analytes. G x E effects were also observed for the majority of analytes, with the exception of brassicasterol, indicating bioactive enhancement of canola seed and oil is achievable via seed breeding. Positive correlations between β- carotene and lutein (r = 0.62, p < 0.01), and between free phytosterols and their corresponding esterified phytosterols (r = 0.43, 0.79, and 0.79 for β-sitosterol, campesterol and brassicasterol, respectively, all p < 0.01), illustrated the potential for symbiotic breeding - to breed for one analyte, with associated increases to others. A multiphase experimental design was developed and applied, for the first time to conduct a multi-stage laboratory process. The effects of various canola oil processing techniques currently used within Australia were investigated, by determining concentrations of phytosterols, tocopherols and carotenoids in seed and oil samples taken at varying stages along each process. Results indicated improved retention of bioactives compared to previous studies, indicating increased preservation with modern processing plant techniques. Extraction and seed preprocessing techniques were shown to enhance all bioactives considerably, which warrants further investigation. A storage trial was conducted to investigate bioactive behaviour in stored seed and oil over 10 months. The oil was extracted using both expellerpressing and solvent extraction, to provide a novel comparison of the storage of minimally refined oils with different extraction techniques. The expeller press oil was shown to have higher concentrations of bioactives, and was more stable than the solvent extracted oil. Although some degradation occurred, results indicated that all bioactives were well preserved, and oxidation was minimal, in seed stored at < 40 °C for the entire 10 month period, and in extracted oils when stored at –18 °C and 4 °C for 6 months. Collectively, the studies present new knowledge on the behaviour of phytosterols, tocopherols and carotenoids in canola seed and oil during seed growth, seed storage, oil processing and oil storage. With this knowledge, biofortification of canola oil is possible, however further research needs to be undertaken to utilise these results in a commercial setting.