The phytopathogenic fungus <i>Leptosphaeria maculans</i> can cause blackleg disease on crucifers, which results in significant yield losses. Fungal diseases involve interactions between pathogenic fungi and host plants. One aspect of these interactions is mediated by secondary metabolites produced by both fungi and host plants. Phytotoxins and elicitors as well as phytoanticipins and phytoalexins are metabolites produced by fungi and plants, respectively. This thesis describes and discusses the isolation, structure, biological activity and biosynthesis of the secondary metabolites produced by L. maculans.<p> The elicitor-toxin activity bioassay guided isolation of elicitors and phytotoxins produced by <i>L. maculans</i> in a chemically defined medium lead to the isolation of general elicitors, <i>sirodesmin PL</i> (165) and <i>deacetylsirodesmin PL</i> (166), and specific elicitors, <i>cerebrosides C</i> (14) and D (31) from minimum medium (MM) culture under standard conditions. The known phytotoxins sirodesmin PL (165) and deacetylsirodesmin PL (166) induced the production of <i>phytoalexin spirobrassinin</i> (122) in both resistant plant species (brown mustard, <i>Brassica juncea</i> cv. Cutlass) and susceptible plant species (canola, B. napus cv. Westar). A mixture of cerebrosides C (14) and D (31) induced the production of the phytoalexin rutalexin (127) in resistant plant species (brown mustard, B. juncea cv. Cutlass) but not in susceptible plant species (canola, B. napus cv. Westar). New metabolites leptomaculins A-E (267-269, 272 and 274) and deacetylleptomaculins C-E (270, 273 and 275) were isolated from elicitor-phytotoxin active fractions but did not display detectable elicitor activity or phytotoxicity after purification.<p> New metabolites maculansins A (299) and B (300), which were not detected in cultures of L. maculans incubated in MM, were isolated from cultures of <i>L. maculans</i> incubated in potato dextrose broth (PDB). Maculansins A (299) and B (300) displayed higher phytotoxicity on brown mustard than on canola and white mustard (<i>Sinapis alba cv. Ochre</i>) but did not elicit detectable production of phytoalexins in either brown mustard or canola. Metabolite 2,4-dihydroxy-3,6-dimethylbenzaldehyde (212) was produced in higher amount in cultures of L. maculans incubated in PDB than in MM and displayed strong inhibition effect on the root growth of brown mustard and canola. <i>L. maculans</i> incubated in MM amended with high concentration of NaCl produced a new metabolite, 8-hydroxynaphthalene-1-sulfate (293), and a known metabolite, bulgarein (294), which are likely involved in the self-protection. The potential intermediates involved in the biosynthesis of sirodesmin PL (165) were investigated using deuterium labeled precursors: [3,3-2H2]-L-tyrosine (251a), [3,3-2H2]O-prenyl-L-tyrosine (312a), E-[3,3,5,5,5-2H5]O-prenyl-L-tyrosine (312b), [5,5-2H2]phomamide (171a), [2,3,3-2H3]-L-serine (233d) and [5,5-2H2]cyclo-L-tyr-L-ser (252a). Intact incorporation of [5,5-2H2]phomamide (171a) into sirodesmin PL (165) suggested that leptomaculin D (272) and E (274), and deacetylleptomaculin D (273) and E (275) are not intermediates in the biosynthesis of sirodesmin PL (165). They are more likely the catabolic metabolites of sirodesmin PL (165). Phomamide (171), the intermediate in the biosynthetic pathway of sirodesmin PL (165), is likely biosynthesized by coupling of prenyl tyrosine (312) with serine (233) rather than prenylation of cyclo-L-tyr-L-ser (252). When [3,3-2H2]-L-tyrosine (251a), [3,3-2H2]O-prenyl-L-tyrosine (312a), and E-[3,3,5,5,5-2H5]O-prenyl-L-tyrosine (312b) were fed into cultures of L. maculans, a â proton exchange was detected by 1H NMR through intrinsic steric isotope effect, which occurs before the formation of phomamide (171). The biosynthesis and catabolism of sirodesmin PL (165) were proposed based on the results obtained in this work.

The phytopathogenic fungus <i>Leptosphaeria maculans</i> can cause blackleg disease on crucifers, which results in significant yield losses. Fungal diseases involve interactions between pathogenic fungi and host plants. One aspect of these interactions is mediated by secondary metabolites produced by both fungi and host plants. Phytotoxins and elicitors as well as phytoanticipins and phytoalexins are metabolites produced by fungi and plants, respectively. This thesis describes and discusses the isolation, structure, biological activity and biosynthesis of the secondary metabolites produced by L. maculans.<p> The elicitor-toxin activity bioassay guided isolation of elicitors and phytotoxins produced by <i>L. maculans</i> in a chemically defined medium lead to the isolation of general elicitors, <i>sirodesmin PL</i> (165) and <i>deacetylsirodesmin PL</i> (166), and specific elicitors, <i>cerebrosides C</i> (14) and D (31) from minimum medium (MM) culture under standard conditions. The known phytotoxins sirodesmin PL (165) and deacetylsirodesmin PL (166) induced the production of <i>phytoalexin spirobrassinin</i> (122) in both resistant plant species (brown mustard, <i>Brassica juncea</i> cv. Cutlass) and susceptible plant species (canola, B. napus cv. Westar). A mixture of cerebrosides C (14) and D (31) induced the production of the phytoalexin rutalexin (127) in resistant plant species (brown mustard, B. juncea cv. Cutlass) but not in susceptible plant species (canola, B. napus cv. Westar). New metabolites leptomaculins A-E (267-269, 272 and 274) and deacetylleptomaculins C-E (270, 273 and 275) were isolated from elicitor-phytotoxin active fractions but did not display detectable elicitor activity or phytotoxicity after purification.<p> New metabolites maculansins A (299) and B (300), which were not detected in cultures of L. maculans incubated in MM, were isolated from cultures of <i>L. maculans</i> incubated in potato dextrose broth (PDB). Maculansins A (299) and B (300) displayed higher phytotoxicity on brown mustard than on canola and white mustard (<i>Sinapis alba cv. Ochre</i>) but did not elicit detectable production of phytoalexins in either brown mustard or canola. Metabolite 2,4-dihydroxy-3,6-dimethylbenzaldehyde (212) was produced in higher amount in cultures of L. maculans incubated in PDB than in MM and displayed strong inhibition effect on the root growth of brown mustard and canola. <i>L. maculans</i> incubated in MM amended with high concentration of NaCl produced a new metabolite, 8-hydroxynaphthalene-1-sulfate (293), and a known metabolite, bulgarein (294), which are likely involved in the self-protection. The potential intermediates involved in the biosynthesis of sirodesmin PL (165) were investigated using deuterium labeled precursors: [3,3-2H2]-L-tyrosine (251a), [3,3-2H2]O-prenyl-L-tyrosine (312a), E-[3,3,5,5,5-2H5]O-prenyl-L-tyrosine (312b), [5,5-2H2]phomamide (171a), [2,3,3-2H3]-L-serine (233d) and [5,5-2H2]cyclo-L-tyr-L-ser (252a). Intact incorporation of [5,5-2H2]phomamide (171a) into sirodesmin PL (165) suggested that leptomaculin D (272) and E (274), and deacetylleptomaculin D (273) and E (275) are not intermediates in the biosynthesis of sirodesmin PL (165). They are more likely the catabolic metabolites of sirodesmin PL (165). Phomamide (171), the intermediate in the biosynthetic pathway of sirodesmin PL (165), is likely biosynthesized by coupling of prenyl tyrosine (312) with serine (233) rather than prenylation of cyclo-L-tyr-L-ser (252). When [3,3-2H2]-L-tyrosine (251a), [3,3-2H2]O-prenyl-L-tyrosine (312a), and E-[3,3,5,5,5-2H5]O-prenyl-L-tyrosine (312b) were fed into cultures of L. maculans, a â proton exchange was detected by 1H NMR through intrinsic steric isotope effect, which occurs before the formation of phomamide (171). The biosynthesis and catabolism of sirodesmin PL (165) were proposed based on the results obtained in this work.