Diclofop-methyl (methyl ester of 2-[4-(2',4'-dichlorophenoxy)phenoxy]propionate; 100 micromolar) and diclofop (100 micromolar) inhibited both ATP-and PPi-dependent formation of H+ gradients by tonoplast vesicles isolated from oat (Avena sativa L., cv Dal) roots. Diclofop-methyl (1 micromolar) significantly reduced the steady-state H+ gradient generated in the presence of ATP. The ester (diclofop-methyl) was more inhibitory than the free acid (diclofop) at pH 7.4, but this relative activity was reversed at pH 5.7. Neither compound affected the rate of ATP or PPi hydrolysis by the proton-pumping enzymes. Diclofop-methyl (50, 100 micromolar), but not diclofop (100 micromolar), accelerated the decay of nonmetabolic H gradients established across vesicle membranes. Diclofop-methyl (100 micromolar) did not collapse K+ gradients across vesicle membranes. Both the (+)-and (-)-enantiomers of diclofop-methyl dissipated nonmetabolic H+ gradients established across vesicle membranes. Diclofopmethyl, but not diclofop (each 100 micromolar), accelerated the decay of H+ gradients imposed across liposomal membranes. These results show that diclofop-methyl causes a specific increase in the H permeability of tonoplast.The herbicide DM3 and its free acid form, DA, inhibit shoot and root growth and cause chlorosis and necrosis of oat and wild oat tissues (7,14,23,28). Ultrastructural damage observed in treated tissues includes breakdown of cellular membranes and destruction of cytoplasmic structure (5). Evidence indicates that they interfere with membrane-associated functions causing inhibition of mineral ion absorption (1), leakage of electrolytes from cells (7) cides is not known, presently two hypothetical mechanisms are being evaluated experimentally (10). In one model, the basis for structural and functional effects on cellular membranes is a direct interaction of the herbicide with the membrane or membrane components. A direct effect of DA on membrane function is indicated by the rapidity of depolarization of electrical potentials by DA (within 10 min [21,36]) and repolarization after removal of DA (40 s, with IAA addition [32]). The nature of the DA-membrane interaction or of the membrane components required for DA activity has not yet been determined. The second model suggests that membrane disruption is an indirect effect of herbicidal action resulting from an inhibition of the biosynthesis of membrane lipids. De novo fatty acid biosynthesis has been shown to be a sensitive site of action for DA (15,17). The I50 values for the inhibition by DA of ['4C]acetate incorporation by maize and oat chloroplasts were 0.2 and 0.1 M, respectively (15, 17). DA was more inhibitory than was DM (Iso = 10 uM) in these assays. Recent evidence indicates that the enzyme acetylCoA carboxylase is the site of inhibition by DA in this biosynthetic pathway ( 17).We have used an in vitro system of tonoplast vesicles isolated from oat root to study the possible direct action of DM and DA on membranes. With isolated membranes, effects of the h...