GC-TOF/MS-based metabolomic strategy for combined toxicity effects of deoxynivalenol and zearalenone on murine macrophage ANA-1 cells

• GC-TOF/MS based metabolomics strategy was conducted to study the combined mycotoxic effects of DON and ZEN.
• DON may inhibit the estrogenic effects of ZEN.
• The combined effect of “DON + ZEN” may exacerbate toxicity in the pentose phosphate pathway.
• Palmitic acid metabolism is likely a new pathway effected by the combination, “DON + ZEN”.

The actual health risk from exposure to combined mycotoxins is unknown, and few studies have focused on changes to cellular biological systems (e.g., metabolomics) caused by combined mycotoxic effects. To evaluate the combined mycotoxic effects of deoxynivalenol (DON) and zearalenone (ZEN) on the level of cellular biological systems, gas chromatographic, time-of-flight mass spectroscopy (GC-TOF/MS) of the complete murine macrophage ANA-1 cell metabolome was implemented in this study. Using optimized chromatography and mass spectrometry parameters, the metabolites detected by GC-TOF/MS were identified and processed using multivariate statistical analysis, including principal component analysis (PCA) and orthogonal projection on latent-structures discriminant analysis (OPLS-DA). The metabolite sets were screened for further pathway analysis under rules of t-test (P) value < 0.05, VIP value > 1, and similarity value > 500. The mainly interfered metabolism pathways were categorized into two dominant types: amino acid metabolism and glycometabolism. Four metabolites, palmitic acid, 1-monopalmitin, ribose-5-phosphate and 2-deoxy-D-galactose, occur only under combined “DON + ZEN” treatment, indicating abnormal metabolism in ANA-1 cells. The metabolic state of ANA-1 cells under induction by combined “DON + ZEN” illustrates that DON may inhibit the estrogenic effects of ZEN. Thus, the combined effect of “DON + ZEN” may exacerbate toxicity in the pentose phosphate pathway, while palmitic acid metabolism is likely a new pathway effected by the combination, “DON + ZEN.”