In this article, we provide mass data on released fatty acids in neutrophils stimulated with A23187 in the presence and absence of phorbol 12-myristate 13-acetate (PMA). A23187 alone caused a highly selective accumulation of non-esterified arachidonic acid (AA) relative to other saturated (palmitic and stearic acids) and unsaturated fatty acids (oleic and linoleic acids). However, the absolute levels of non-esterified arachidonic acid, oleic acid and linoleic acid were significantly increased (by twofold) in response to the action of A23187 in the presence of PMA. We also noted a further twofold increase in the non-esterified arachidonic acid in the presence of BW755C, a dual inhibitor of cyclooxygenase and lipoxygenases, with no apparent change in the levels of oleic and linoleic acids and other saturated fatty acids. These results suggest that PMA potentiates not only the release of AA but also the release of oleic and linoleic acids from neutrophil phosphoglycerides. This is possibly due to the activation of non-specific phospholipase A2 (PLA2). Our data also suggest a possible role for the 5-lipoxygenase metabolites in PMA-induced synergism with regard to the release of AA through AA-specific and PMA-sensitive PLA2. Protein kinase C appears, therefore, to play a role in the regulation of both AA-specific and non-specific PLA2 in human neutrophils.