Reactive oxygen species and granule proteases produced by neutrophils contribute to the pathogenesis of inflammatory diseases. In this study, a cellular model in isolated human neutrophils was established to elucidate the anti-inflammatory functions of 16-hydroxycleroda-3,13(14)E-dien-15-oic acid (PL3S), a clerodane diterpenoid from Formosan Polyalthia longifolia var. pendula. PL3S significantly inhibited the generation of superoxide anion and the release of elastase in formyl-l-methionyl-l-leucyl-l-phenylalanine (FMLP)-activated human neutrophils in a concentration-dependent fashion with IC50 values of 3.06+/-0.20 and 3.30+/-0.48 microM, respectively. PL3S did not affect cAMP-dependent pathway, and the inhibitory effect of PL3S was not reversed by protein kinase A inhibitor. PL3S did not display antioxidant or superoxide anion-scavenging ability, and it failed to alter the subcellular NADPH oxidase activity. PL3S concentration-dependently inhibited calcium mobilization caused by FMLP but not thapsigargin. Furthermore, PL3S attenuated the FMLP-induced protein kinase B (AKT) and p38 mitogen-activated protein kinase phosphorylation. However, PL3S had no effect on FMLP-induced phosphorylation of extracellular regulated kinase and c-Jun N-terminal kinase. In summary, these results indicate that the suppressive effects of PL3S on human neutrophil respiratory burst and degranulation are at least partly mediated by inhibition of calcium, AKT, and p38 signaling pathways.