Biological control of plant pathogens is considered as one of the green and effective technologies using beneficial microorganisms or microbial secondary metabolites against plant diseases, and so microbial natural products have played important roles in the research and development of new and green agrochemicals. To explore the potential applications for natural γ-lactam alkaloids and their derivatives, 26 γ-lactams that have flexible substituent patterns were synthesized and characterized, and their antifungal activities against eight kinds of plant pathogens belonging to oomycetes, basidiomycetes, and deuteromycetes were fully evaluated. In addition, the high potential compounds were further tested using an assay against blight of pepper to verify a practical application for controlling oomycete diseases. The potential modes of action for compound against were also investigated using microscopic technology (optical microscopy, scanning electron microscopy, and transmission electron microscopy) and label-free quantitative proteomics analysis. The results demonstrated that compound may be a potential novel fungicidal agent against oomycete diseases (EC = 4.9748 μg·mL for and EC = 5.1602 μg·mL for ) that can act on steroid biosynthesis, which can provide a certain theoretical basis for the development of natural lactam derivatives as potential antifungal agents.
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