Optimization of Saponification Process for Quantification of Individual and Total Tocopherols and Phytosterols in Canola Oil Refining By‐Products

ABSTRACT In this study, saponification variables were optimized using a face‐centred central composite design‐response surface methodology. The variables included reaction temperature (50°C–120°C), reaction time (30–180 min), KOH concentration (0.5–3 M) and pyrogallol concentration (0–1% w/v). Using this approach, individual and total tocopherols and phytosterols in canola oil refining by‐products, including soapstock, oil from spent bleaching earth (SBE), and deodorizer distillate, were quantified, and effects of variables were evaluated. The obtained results suggest that the non‐saponified extract in distillate presented a substantial level of tocopherols and phytosterols and their levels increased after saponification by ~60% and ~20%, respectively. Quadratic models were significant (model p < 0.001) for all responses with good fit ( R 2 ≥ 0.91) and predictability. Furthermore, the effects of variables on individual and total tocopherols and phytosterols were explained and potential mechanisms were described. For tocopherols, pyrogallol generally exerted a protective effect, whereas stronger alkali and prolonged reaction time tended to negatively affect tocopherols, with γ‐tocopherol showing greater reduction compared to α‐tocopherol in some matrices. For phytosterols, temperature and time generally increased phytosterols, while losses during hot alkaline conditions and liquid–liquid extraction could offset gains depending on the by‐products. The optimized conditions were experimentally validated and the co‐optimized condition (120°C, 180 min, 0.5 M KOH, 1% w/v pyrogallol) for distillate yielded ~12.7 g/kg total tocopherols and ~108.9 g/kg total phytosterols. These conditions provide a practical basis for matrix‐specific saponification and reliable quantification of tocopherols and phytosterols in canola oil refining by‐products.