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The aim of this study was to assess free fatty acids&#8217; (FAs) <i>ex vivo</i> anti-/proinflammatory capabilities and their influence on inflammatory gene expression and H₂O₂ production by human peripheral blood mononuclear cells (PBMCs). Anthropometric and clinical measurements were performed in 26 participants with metabolic syndrome. Isolated PBMCs were incubated <i>ex vivo</i> for 2 h with several free fatty acids&#8212;palmitic, oleic, &#945;-linolenic, &#947;-linolenic, arachidonic and docosahexaenoic at 50 &#956;M, and lipopolysaccharide (LPS) alone or in combination. H₂O₂ production and IL6, NF&#954;B, TLR2, TNF&#945;, and COX-2 gene expressions were determined. Palmitic, &#947;-linolenic, and arachidonic acids showed minor effects on inflammatory gene expression, whereas oleic, &#945;-linolenic, and docosahexaenoic acids reduced proinflammatory gene expression in LPS-stimulated PBMCs. Arachidonic and &#945;-linolenic acids treatment enhanced LPS-stimulated H₂O₂ production by PBMCs, while palmitic, oleic, &#947;-linolenic, and docosahexaenoic acids did not exert significant effects. Oleic, &#945;-linolenic, and docosahexaenoic acids induced anti-inflammatory responses in PBMCs. Arachidonic and &#945;-linolenic acids enhanced the oxidative status of LPS-stimulated PBMCs. In conclusion, PBMC <i>ex vivo</i> assays are useful to assess the anti-/proinflammatory and redox-modulatory effects of fatty acids or other food bioactive compounds.