Effects of Cultivation Conditions and Bean Curd (Tofu) Wastewater Application on ALA Accumulation in Chlorella sp. L166 and Its Mutant C-12

Alpha-linolenic acid (ALA) is an essential omega-3 fatty acid and a vital component in food applications. In this study, we investigated a range of physicochemical culture conditions—including pH, temperature, and carbon source—to evaluate biomass and ALA accumulation in Chlorella sp. L166 and its mutant, C-12. The study aimed to identify favorable culture conditions and evaluate the feasibility of using diluted bean curd (tofu) wastewater as a low-cost medium. Under mixotrophic cultivation, ALA content was determined via GC-MS, and the removal efficiencies of total nitrogen (TN), total phosphorus (TP), and chemical oxygen demand (COD) were simultaneously monitored. The results showed that L166 achieved its highest ALA accumulation at pH 6.0 and 23 °C with maltose. C-12 exhibited appropriate ALA accumulation at pH 7.0 and 23 °C with maltose and reached its maximum biomass at pH 8.0 and 25 °C with glucose. After 8 days of cultivation in threefold-diluted tofu wastewater, C-12’s ALA content reached 6.1 mg/g, significantly higher than that observed in BG11 medium. Meanwhile, both strains removed 81.2–83.2% of TN, 35.7–36.0% of TP, and 42.6–43.5% of COD. This study provides preliminary data on the effects of culture conditions on microalgal ALA production, highlighting the potential for future practical applications of C-12.