Olive pomace extracts obtained by different defatting methods: bioactive compounds profile and antioxidant potential in a lipid peroxidation model

The high content of unsaturated fatty acids in pet food formulations makes these products susceptible to oxidative deterioration, which can compromise quality, nutritional value, and shelf life. In this context, the growing demand for natural and sustainable preservatives in the pet food industry has encouraged the exploration of agro industrial by-products as sources of bioactive compounds. Olive pomace (OP), the main by-product of olive oil production, is rich in bioactive compounds with recognized antioxidant properties and represents a promising alternative to synthetic antioxidants. This study aimed to compare the bioactive composition and antioxidant potential of ethanolic extracts obtained from OP using pressurized liquid extraction (PLE), preceded by two distinct defatting methods: hexane extraction (HEX) and supercritical fluid extraction (SFE). OP was first defatted via HEX or SFE and subsequently subjected to PLE using absolute ethanol as the extraction solvent. The resulting extracts were concentrated under a nitrogen stream and reconstituted in a 50:50 (v/v) mixture of water and analytical-grade ethanol. The bioactive profiles of HEX-PLE and SFE-PLE were characterized by high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS), and their effects on lipid oxidation were evaluated in antioxidant-free soybean oil. The extracts were incorporated into soybean oil at 300 ppm and stored for five days. Oxidation induction time was determined using the Rancimat method (EN 14112), while thiobarbituric acid reactive substances (TBARS) were measured following oxidative induction. Commercial natural antioxidants (rosemary, green tea, and tocopherols) and the synthetic antioxidant butylated hydroxytoluene (BHT) were also evaluated at 300 ppm for comparison under the same experimental conditions. Rancimat and TBARS data were subjected to one-way analysis of variance (ANOVA) followed by Tukey’s post-hoc test (p < 0.05), while differences in bioactive compound concentrations between HEX-PLE and SFE-PLE were assessed using Student’s t-test. Neither HEX-PLE nor SFE-PLE significantly improved the oxidation induction time compared to the control (3.31 and 3.69 vs. 3.69 h; p > 0.05). In contrast, only oil added with SFE-PLE exhibited the lowest TBARS level among all treatments (77.59 µmol MDA kg⁻¹), significantly lower than the control (186.18 ± 11.66 µmol MDA kg⁻¹) and BHT (244.46 µmol MDA kg⁻¹; p < 0.05). These results indicate that the two defatting approaches yielded extracts with distinct antioxidant performance, with SFE-PLE being more effective at limiting the accumulation of secondary lipid oxidation products following oxidative induction. HPLC–MS analysis revealed significantly higher concentrations of oleacein, hydroxytyrosol, and tyrosol in SFE-PLE than HEX-PLE (p < 0.05), suggesting that their greater abundance may have contributed to SFE-PLE’s superior performance in mitigating secondary oxidation products. Overall, SFE-PLE demonstrated promising potential as a natural antioxidant source for pet food applications, particularly for reducing secondary lipid oxidation products in oils and fats.