A biotype of Lolium rigidum Gaudin (VLR 69) shows multiple resistance to at least nine dissimilar herbicide chemistries. This biotype has enhanced metabolism to herbicides that inhibit Photosystem II, acetolactate synthase, and acetyl-coenzyme A carboxylase. The potential for malathion and piperonyl butoxide (PBO) to act as synergists for herbicides from five different chemical classes was investigated in potted plants. Of the herbicide/synergist combinations examined, PBO only synergized chlorotoluron, and malathion only synergized chlorsulfuron in this resistant biotype. The ability of PBO, malathion, 1-aminobenzotriazole (ABT), and tetcyclacis to inhibit metabolism of herbicides in vivo was also examined. ABT, PBO, and tetcyclacis inhibited metabolism of simazine and chlorotoluron, but malathion did not. Malathion alone inhibited metabolism of chlorsulfuron, ABT inhibited metabolism of diclofop, and none of these compounds affected tralkoxydim metabolism. These results suggest that at least four different herbicide-metabolizing enzymes have increased activity in this resistant biotype. In addition to enhanced metabolism, this biotype also contains a resistant form of acetyl-coenzyme A carboxylase which shows 31-, 4-, and 20-fold resistance to diclofop acid, fluazifop acid, and haloxyfop acid, respectively, but which shows no resistance to sethoxydim or tralkoxydim. Multiple resistance in this biotype of L. rigidum is clearly the result of the accumulation of several resistance mechanisms. (C) 1996 Academic Press, Inc.