In the field of Sustainable Agriculture, Agronomy and Food Science, excess nutrient discharge from livestock biogas digestate creates severe eutrophication hazards in surrounding watersheds. This empirical investigation systematically examines Anaerobic Digestate Bio-Refinery for Microalgae Chlorella vulgaris Cultivation and Lipid Accumulation through a multi-stage experimental methodology and rigorous quantitative analytical framework. Utilizing diluted anaerobic effluent cultivation, sequential batch photobioreactors, and fatty acid methyl ester gas chromatography profiling, data were gathered across multiple operational cycles and validated against established international benchmarks. The statistical and computational results reveal that Chlorella vulgaris achieved 94% ammonium removal and 88% phosphate uptake while yielding 34% intracellular lipid content suitable for biodiesel. 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 integrated livestock wastewater treatment and decentralized bioenergy production. 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.