The mode of action of four photosynthetic inhibitor herbicides, bromacil, CMU ioxynil Na and metribuzin, was studied by following the sequence of events which occurred during the treatment of flax cotyledon leaves. The involvement of light was determined by treating leaves in the dark and at 0.2, 5.25 and 30 w/m2. CO2 fixation was rapidly inhibited by the four herbicides at all light intensities, Chloroplasts isolated from these leaves still showed FeCH reduction for some time after the inhibition of CO2 fixation, due to a reversibility of herbicide binding. SiMo reduction decreased at a slower rate than FeCN reduction in bromacil, CMU and metribuzin treatments and this was an indicator of PS2 integrity. SiMo reduction in chloroplasts isolated from ioxymil Na treated leaves however decreased at a similar rate to FeCH reduction. P31 activity monitored by ascorbate photo-oxidation was increased initially in bromacil, CMH and metribuzin treatments in the light, prior to decreasing at a much slower rate than SiNo reduction. With ioxynil Na no stimulation in PS2 activity was apparent. A herbicide induced chlorosis occurred more rapidly with ioxynil Na. Carotene breakdown preceded xanthophyll and chlorophyll breakdown in ioxynil Na and CMU treatments in the light. The presence of 1o2 quenchers in the incubation medium, or herbicide treatment under anaerobic conditions, decreased or prevented pigment breakdown. The ultrastructural changes were identical in bromacil, CMU and metribuzin treatments. In the light there was a marked swelling of the chloroplast preceding disruption and swelling of the thylakoids, prior to chloroplast and tonoplast rupture. Ioxynil Na caused a rapid swelling of the thylakoids resulting in the appearance of many vesicles in the stroma. Ethane formation, an indication of membrane damage, was apparent once chloroplast disruption was initiated and increased as chloroplast and cellular degradation continued. The mode of action of bromacil, CMU and metribuzin appeared identical, however ioxynil Na acted by a slightly different mechanism due possibly to a breakdown to iodine. Light was a major factor in the action of all four herbicides as toxicity occurred more rapidly at the higher light intensities. A scheme summarizing the role of light and subsequent biochemical events leading to plant cell death is presented. The effects of the four herbicides were also compared with isolated chloroplasts. The order of inhibition of FeCH reduction was metribuzin bromacil ioxynil Na CMU. SiMo reduction was partially insensitive to the herbicides, although it was completely inhibited by high concentrations of ioxynil Na. In trypsin treated chloroplasts FeCH reduction was insensitive to bromacil, CMU and metribuzin and partially insensitive to ioxynil Na. A proteinaceous binding site is proposed between Q and plastoquinone. Ioxynil Na is suggested to bind to a trypsin unaffected site prior to the first SiMo acceptance site.
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