Barbatic acid, a lichen-derived depside, inhibited oxygen evolution in spinach thylakoid membranes. It also affected parameters of Chl fluorescence, (Fm'-F)/F and Fv/Fm. Using specific donors and acceptors of electrons, we found two sites of inhibition in the PS II complex. The primary site, which is responsible for the inhibition of oxygen evolution, is at the reducing side of QA, possibly at QB. The other site is at the oxidizing side of P680 but not in the oxygen evolving complex, suggesting Yz as the target. At both sites, irreversible binding of the depside to the targets seems to be responsible for the inhibitions. Among the 8 lichen acids compared, barbatic acid was the most potent inhibitor for both the reducing site and oxidizing site.