This study evaluated the environmental and economic impacts of substituting synthetic media with greywater for cultivating microalgae in the biofuel production process. Life cycle assessment (LCA) and technoeconomic assessment (TEA) were employed to compare the impacts of two scenarios – one containing bold’s basal (BB) media and another containing greywater as growth mediums for microalgae cultivation. Scenarios 1 and 2 mitigated 1.74 and 2.14 kg CO2 per kg of biofuel production, respectively. Substituting BB media with greywater resulted in a 16.3% reduction in energy requirements, leading to a 79.3% increase in net energy recovered. LCA findings demonstrate a reduction in all seven environmental categories. TEA reveals that, despite a 21.7% higher capital investment, scenario 2 proves more economically viable due to a 39.8% lower operating cost and additional revenue from wastewater treatment and carbon credits. The minimum selling price of biofuel dropped from Rs 73.5/kg to Rs 36.5/kg, highlighting the economic and environmental advantages of substituting BB media with greywater in microalgal biofuel production.