In the field of Sustainable Agriculture, Agronomy and Food Science, fungal fruit rot caused by Colletotrichum scovillei devastates Solanaceous vegetable production throughout tropical Asia. This empirical investigation systematically examines Novel Antifungal Lipopeptides from Endophytic Streptomyces for Anthracnose Suppression in Chili Pepper through a multi-stage experimental methodology and rigorous quantitative analytical framework. Utilizing endophyte metabolite isolation, reversed-phase HPLC fractionation, LC-MS/MS characterization, and fungal membrane permeability assays, data were gathered across multiple operational cycles and validated against established international benchmarks. The statistical and computational results reveal that purified cyclic lipopeptides inhibited fungal mycelial growth at a minimum inhibitory concentration of 8.0 microgram/mL through cell membrane lysis. Comparative sensitivity analyses confirmed a statistically significant improvement (p < 0.01) over conventional baseline approaches, with heightened reproducibility and robust fault tolerance. These comprehensive findings provide actionable theoretical insights and practical implementation guidelines for natural biopesticide formulation and eco-friendly horticultural disease control. Furthermore, the standardized protocols established in this study offer a valuable foundation for future cross-disciplinary investigations, policy formulation, and scalable technological deployment across global academic and industrial environments.