Graduate Thesis Or Dissertation
 

A proposed mechanism for diuron-induced phytotoxicity

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  • Previous research has shown that specific carotenoid pigments function through oxidation-reduction reactions to protect chlorophyll from photooxidation in the presence of reduced nicotinamide adenine dinucleotide phosphate (NADP). The hypothesis that diuron (3-(3, 4- d chloropheny1)-1, 1-dimethylurea) initiates phytotoxicity by inhibition of NADPH synthesis was examined. All experiments were conducted in vitro using functional chloroplasts isolated from spinach (Spinacia oleracea, L.) leaves. The initial experiment indicated that light was necessary to express diuron toxicity. After 8 hr of illumination, chlorophyll concentration in the diuron treatments was reduced to 10% of the concentration in the control treatment. The addition of an artificial electron donor system (ascorbate DPIP) overcame the effects of diuron and maintained chlorophyll concentrations at higher levels than in the control treatment after 14 hr of chloroplast illumination. Ascorbate + DPIP protected chlorophyll against diuron only in functional caloroplasts, indicating that an intact enzyme system is necessary for the protective action. Ascorbate + DPIP provided protection in the presence of methylamine HC1, an inhibitor of photophosphiorylation, which suggests that a lack of adenosine triphosphate (ATP) is not the cause of diuron toxicity. Kinetic studies showed that carotenoid pigments began to degrade more quickly than chlorophyll pigments in the presence of diuron which tends to support the proposal that carotenoid pigments are functioning naturally to protect chlorophyll from photooxidation. Diuron toxicity, as measured by chlorophyll degradation was much greater when ratios of chlorophyll to diuron molecules were 200:1 or lower, than at higher ratios of 300:1 to 800:1. These results support a previous conclusion that diuron acts at a photosynthetic reaction center comprised of approximately 250 chlorophyll molecules. These data also support a previous proposal that phytotoxicity from certain herbicides which inhibit the Hill reaction is directly related to chlorophyll concentration. Diuron is a potent inhibitor of the Hill reaction thus preventing the formation of NADPH. Experimental evidence gained in this study support the hypothesis that specific carotenoid pigments function in the presence of reduced NADP to protect photosensitized chlorophyll from becoming potential oxidants. Therefore it is proposed that diuron induces phytotoxicity by limiting NADPH, thus catalyzing photosensitized oxidation reactions which are lethal to the cell. In greenhouse studies, diuron toxicity to red kidney bean plants (Phaseolus vulgaris L.) was not overcome by the addition of ascorbate or mercapto ethanol in combination with DPIP. Both ascorbate and mercapto ethanol were toxic to plant tissue. Because of the toxic symptoms noted, we postulated that the electron donors were rapidly metabolized, forming secondary toxic compounds before reaching the chloroplast system. Therefore, they were unable to function as electron donors in preventing diuron toxicity.
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