Correlation between volatile oxidation products and inflammatory markers in docosahexaenoic acid: Insights from OPLS-DA and predictive modeling

DHA with varying degrees of oxidation was obtained by the Schaal oven acceleration method. Then, the temporal changes in free radicals, volatile compounds, and carbonyl compounds in oxidized DHA was analyzed using HS-SPME-GC-MS, the content of which were increased with the oxidation time. RAW264.7 macrophages were used to evaluate the effects of DHA oxidation products on the oxidative stress, gene and protein expression of inflammatory factors at different oxidation times. With the increase of oxidation time, oxidized DHA stimulated cells with significantly higher levels of reactive oxygen species (ROS), nitric oxide (NO) and malondialdehyde (MDA), while superoxide dismutase (SOD) activity was significantly inhibited. It also stimulated a significant increase in the expression of tumor necrosis factor-α (TNF-α), interleukin (IL-1β) and inducible nitric oxide synthase (iNOS), which exacerbated the cellular inflammatory response. Data on DHA oxidation products were analyzed using OPLS-DA. The compound (E,E)-2,4-heptadienal was identified as a potential differential metabolite capable of distinguishing among six oxidation time points. Furthermore, the pro-inflammatory effects of oxidized DHA were accurately predicted using partial least squares regression (PLSR) and support vector regression (SVR) models. This study established a correlation between volatile oxidation products and their potential pro-inflammatory effects, which were readily detectable through high-throughput assays. It thereby provided a rapid and reliable method for assessing the predicted bioactivity of DHA at varying levels of oxidation.