Metabolism of cruciferous chemical defenses by plant pathogenic fungi

Plants produce complex mixtures of secondary metabolites to defend themselves from pathogens. Among these defenses are metabolites produced de novo, phytoalexins, and constitutive metabolites, phytoanticipins. As a counter-attack, pathogenic fungi are able to transform such plant defenses utilizing detoxifying enzymes. This thesis investigates the metabolism of two important cruciferous phytoalexins (brassinin (33) and camalexin (39)) by the phytopathogenic fungus Botrytis cinerea and the metabolism of cruciferous phytoanticipins (glucosinolates and derivatives) by three economically important fungi of crucifers Alternaria brassicicola, Rhizoctonia solani and Sclerotinia sclerotiorum to investigate their role in cruciferous defense. In the first part of this thesis, the transformations of brassinin (33) and camalexin (39) by B. cinerea were investigated. During these studies a number of new metabolites were isolated, their chemical structures were determined using spectroscopic techniques, and further confirmed by synthesis. Camalexin (39) was transformed via oxidative degradation and brassinin (33) was hydrolyzed to indoly-3-methanamine (49). The metabolic products did not show detectable antifungal activity against B. cinerea, which indicated that these transformations were detoxification processes. Camalexin (39) was found to be more antifungal than brassinin (33). In the second part of this thesis, the metabolism of glucobrassicin (86), 1-methoxyglucobrassicin (87), 4-methoxyglucobrassicin (90), phenylglucosinolate (65), and benzylglucosinolate (66), the corresponding desulfoglucosinolates and derivatives by three fungal pathogens (A. brassicicola, R. solani and S. sclerotiorum) was investigated and their antifungal activity against the same pathogens was tested. Aryl iii glucosinolates 65 and 66 were metabolized by A. brassicicola but not by R. solani or S. sclerotiorum, whereas indolylglucosinolates were not metabolized by any pathogen. Indolyl desulfoglucosinolates (159 and 233) were transformed by R. solani and S. sclerotiorum to the corresponding carboxylic acids and indolyl acetonitriles 40, 102, and 103 were also metabolized to the corresponding carboxylic acids by all pathogens. None of the glucosinolates or their desulfo derivatives showed antifungal activity, but some of their metabolites showed low to very high antifungal activities. Among these metabolites, diindolyl-3-methane (113) showed the highest antifungal activity, and benzyl isothiocyanate (170) showed higher inhibitory effect against R. solani and S. sclerotiorum, but did not inhibit the growth of A. brassicicola. The cell-free extracts of A. brassicicola, R. solani, and S. sclerotiorum were tested for myrosinase activity against several glucosinolates. The cell-free extracts of mycelia of A. brassicicola displayed higher myrosinase activity for sinigrin (131), phenyl and benzyl glucosinolates 65 and 66, but lower activities for glucobrassicin (86) and 1-methoxyglucobrassicin (87); no myrosinase activity was detected in mycelia of either R. solani or S. sclerotiorum