Increasing restrictions on synthetic fungicides and pathogen resistance necessitate eco-friendly alternatives. This research investigated the antimicrobial potential of specialized metabolites from Capsicum chinense 'Bhut Jolokia', examining how genetic mutations affecting pungency influence the overall phytochemical profile and biological activity against economically important soil-borne pathogens. Hydromethanolic extracts were prepared from the pungent cv. C-449 (capsaicinoid-rich) and its non-pungent mutant C-449_mutant (capsinoid-rich). Gas chromatography–mass spectrometry (GC-MS) was employed to distinguish chemical profiles between metabolite extracts. Antifungal screening utilized standardized agar dilution techniques against six major horticultural pathogens (Botrytis cinerea, Fusarium equiseti, Macrophomina phaseolina, Neocosmospora falciformis, N. keratoplastica, and Sclerotinia sclerotiorum). Additionally, ex situ protective assays were conducted on cucumber slices artificially inoculated with S. sclerotiorum to validate real-world efficacy. GC-MS revealed that C-449 extract was dominated by capsaicinoids (30.64%: capsaicin 19.94%, dihydrocapsaicin 10.70%) and fatty acids (20.30%). C-449_mutant lacked intact capsinoids, showing degradation products (hydrolysis: e.g., 3-hydroxy-4-methoxybenzoic acid; thermolysis: e.g., guaiacols) and fatty acids (24.74%). Both extracts demonstrated broad-spectrum antifungal activity, with the capsaicinoid-rich extract achieving superior pathogen control (complete inhibition at 250-750 μg/mL) compared to the capsinoid variant (375-1000 μg/mL). S. sclerotiorum showed the highest sensitivity while N. keratoplastica exhibited the greatest resistance. Both natural extracts outperformed azoxystrobin and fosetyl-Al in comparative assessments. Given its commercial viability advantages (eliminating worker protection concerns and crop pungency risks), the non-pungent extract was selected for ex situ validation. In cucumber slice assays, it provided complete protection against S. sclerotiorum at 1125 μg/mL, while lower concentrations failed to prevent colonization. This study establishes C. chinense extracts, especially the low-pungency variant, as viable candidates for sustainable disease management, addressing both efficacy and critical application constraints for modern horticulture.