The structure-activity relationship of 18-carbon fatty acids (C(18) FAs) on human neutrophil functions and their underlying mechanism were investigated. C(18) unsaturated (U)FAs potently inhibited superoxide anion production, elastase release, and Ca(2+) mobilization at concentrations of <10 microM in formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP)-activated human neutrophils. However, neither saturated FA nor esterified UFAs inhibited these neutrophil functions. The inhibitory potencies of C(18) UFAs decreased in the following order: C(18):1 > C(18):2 > C(18):3 > C(18):4. Notably, the potency of attenuating Ca(2+) mobilization was closely correlated with decreasing cellular responses. The inhibitions of Ca(2+) mobilization by C(18) UFAs were not altered in a Ca(2+)-containing Na(+)-deprived medium. Significantly, C(18) UFAs increased the activities of plasma membrane Ca(2+)-ATPase (PMCA) in neutrophils and isolated cell membranes. In contrast, C(18) UFAs failed to alter either the cAMP level or phosphodiesterase activity. Moreover, C(18) UFAs did not reduce extracellular Ba(2+) entry in FMLP- and thapsigargin-activated neutrophils. In summary, the inhibition of neutrophil functions by C(18) UFAs is attributed to the blockade of Ca(2+) mobilization through modulation of PMCA. We also suggest that both the free carboxy group and the number of double bonds of the C(18) UFA structure are critical to providing the potent anti-inflammatory properties in human neutrophils.