The interaction of retinoids and flavonoids with phospholipases A2 (PLA2) was studied to assess the mechanism of inhibition. Retinoids, such as retinal, retinol, retinoic acid and retinol acetate, and flavonoids, such as quercetin, rutin, morin and sciadopitysin, inhibit Ca2+-dependent PLA2 activity of human synovial fluid (HSF) in vitro in a dose-dependent fashion; ID20S ranged from 2-8 microM. Retinal inhibited neutral active Ca2+-dependent PLA2S from human platelets, human plasma, human polymorphonuclear leukocytes and Naja mossambica mossambica venom in a dose-dependent manner while quercetin inhibits extracellular PLA2 activities of human plasma, HSF and N. m. mossambica venom in a dose-dependent manner but not PLA2 activity derived from human platelets and polymorphomonuclear leukocytes. Inhibition of PLA2 activity by both flavonoid and retinoids were independent of Ca2+ or Na+. Increasing substrate concentration (9-144 nmols) relieved the inhibition of HSF-PLA2 activity by quercetin indicating probable interaction with the substrate. The inhibition by retinal is independent of substrate concentration suggesting that inhibition by retinal is probably due to direct interaction with the enzyme. both retinal and quercetin quenched the relative fluorescent intensity of N. m. mossambica PLA2 and in a dose-dependent manner in the same concentration range at which they inhibit in vitro PLA2 activity. Retinal and quercetin shift the thermotropic phase transition of distearoylphosphatidylethanolamine (DSPE) liposomes. Both compounds broadened the transition peak, shifted the Tm to lower temperature, and decreased enthalpy significantly. These findings indicate that inhibition of non-pancreatic human PLA2S by retinoids and flavonoids can be mediated by interaction with enzyme and/or substrate.