Exploration of the Physiological Response Characteristics of Four Chlorella Strains to Ammonia Nitrogen Concentration in the Culture System

To address the scarcity of high-quality, ammonia-tolerant microalgae strains for the conversion and utilization of high-concentration ammonia nitrogen wastewater such as biogas slurry, four strains of Chlorella (FACHB-5, FACHB-11, GB-Z3, and LZ-29) were selected as research subjects. The strains were cultured for 10 days in modified BG-11 media with a gradient of total ammonia nitrogen concentrations (TAN: 50, 100, 150, 200, and 250 mg·L^-1, designated as TAN50 to TAN250). Physiological response characteristics-including biomass, ammonia nitrogen removal rate, chlorophyll a content, protein content, and activities of nitrogen assimilation enzymes glutamine synthetase (GS) and glutamate synthase (GOGAT) - were determined to clarify the differences in high-concentration ammonia nitrogen tolerance among the four strains, aiming to screen for a high-concentration ammonia nitrogen-tolerant strain. The results show that FACHB-5, FACHB-11, and GB-Z3 exhibited stable photosynthetic and growth activities only under TAN50-TAN100 or TAN50-TAN150, beyond which their activities were significantly inhibited as the ammonia nitrogen concentration increased. In contrast, LZ-29 maintained stable photosynthetic and growth activities across all TAN concentrations. On the 10th day of cultivation, its chlorophyll a content increased by 4.40-8.99, 3.34-7.74, and 3.36-15.48 mg·g^-1 compared to FACHB-5, FACHB-11, and GB-Z3, respectively. Under the TAN250 condition, the physiological responses of the four strains diverged further. By day 10, GB-Z3 died, while the biomass of FACHB-5 and FACHB-11 (1.38 and 1.60 g·L^-1) was significantly lower than that of LZ-29 (2.02 g·L^-1), indicating that the ammonia nitrogen tolerance of LZ-29 was significantly stronger than that of FACHB-5, FACHB-11, and GB-Z3. Concurrently, the protein content, ammonia nitrogen removal rate, and assimilation rate of LZ-29 were significantly higher than those of FACHB-5 and FACHB-11. Compared to FACHB-5 and FACHB-11, the protein content of LZ-29 (51.99%) increased by 14.54 and 15.14 percentage points, the ammonia nitrogen removal rate (81.25%) increased by 18.61 and 14.41 percentage points, and the ammonia nitrogen assimilation rate (83.92%) increased by 11.44 and 8.14 percentage points, respectively, demonstrating that LZ-29 possesses a more efficient capacity for ammonia nitrogen-to-protein conversion. Further analysis of the nitrogen assimilation enzyme activities revealed that the GS and GOGAT activities of FACHB-5, FACHB-11, and GB-Z3 exhibited a trend of initially increasing and then decreasing with rising TAN concentrations, peaking at TAN100-TAN150 before being significantly inhibited. Conversely, the GS and GOGAT activities of LZ-29 continuously increased with rising TAN concentrations, reaching 3.62 and 20.45 U·mg^-1 prot at TAN250, respectively. This indicates that LZ-29 can adapt to high-concentration ammonia nitrogen environments through an adaptive upregulation of nitrogen assimilation enzyme activities to enhance ammonia nitrogen uptake and conversion. Therefore, LZ-29 is a high-concentration ammonia nitrogen-tolerant strain with substantial potential for advancing the construction of collaborative technologies for the efficient treatment and resource utilization of high-concentration ammonia nitrogen wastewater such as biogas slurry.