Removal effect of NH₄⁺-N and NO₂⁻-N by Chlorella vulgaris and the assimilation pathway of NO₂⁻-N

Since the non-ionic ammonia and ionic ammonia can be converted into each other, in aquaculture, ammonia nitrogen (NH₄⁺-N) and nitrite nitrogen (NO₂⁻-N) are the key factors that affect the growth and development of aquatic animals. Therefore, the removal of NH₄⁺-N and NO₂⁻-N in aquaculture water is of great significance to ensure the health of aquaculture animals. As we all know, microorganisms and algae play important roles in maintaining the ecological balance of aquaculture ponds. It has been proved that Chlorella had the ability of purifying aquaculture water, with different Chlorella species different removal efficiency on NH₄⁺-N and NO₂⁻-N. Chlorella vulgaris is widely used in aquaculture, however, we know little about the removal effect on nitrogen nutrient especially NO₂⁻-N by C. vulgaris. Given the shortage of effective ecological control measures in aquaculture, it is of great significance to illustrate the removal effect of NH₄⁺- N and NO₂⁻-N by C. vulgaris and related influencing factors. Aiming to evaluate the application prospect of C. vulgaris in purifying NH₄⁺-N and NO₂⁻-N in water, in the present study, C. vulgaris was taken as the research object and feed wastewater was taken as the culture medium. We firstly detected the cell density of C. vulgaris and the temporal variations of NH₄⁺-N and NO₂⁻-N in water under aeration, light, combined light and aeration conditions. Then we analyzed the effects of time (X₁), light intensity (X₂) or initial C. vulgaris density (X₃) on the removal rates of NH₄⁺-N and NO₂⁻-N (Y). Finally, we evaluated the removal efficiency of NH₄⁺-N, NO₂⁻-N and NO₃⁻-N from water by C. vulgaris, and we analyzed the potential pathway of NO₂⁻-N assimilation by C. vulgaris. The results showed that C. vulgaris could remove NH₄⁺-N, NO₂⁻-N and NO₃⁻-N significantly under suitable light conditions. The NH₄⁺- N removal rate reached up to at 18 000 lx (96.23%), and NO₂⁻-N removal rate reached up to 99.19% at 9 000 lx. The initial density of C. vulgaris at 2.5×10⁵ cells/mL had the highest removal rates for NH₄⁺-N and NO₂⁻-N, accounting 94.92% and 99.05%, respectively. The regression equation of NH₄⁺-N and NO₂⁻-N removal rates with treatment time and light intensity was as follows: Y_NH4⁺-N=1.189X₁+5.79×10⁻⁴X₂+24.158 (R²=0.664), Y_NO2⁻-N=1.562X₁+1.909×10⁻³X₂−26.078 (R²=0.762). The regression equation of NH₄⁺-N and NO₂⁻-N removal rates with treatment time and initial C. vulgaris density was as follows: Y_NH4⁺-N =0.888X₁+1.02×10⁻⁵X₃+32.555 (R²=0.408), Y_NO2⁻-N =1.746X₁+1.64×10⁻⁵X₃−17.250 (R²=0.613). The order of nitrogen removal by C. vulgaris was NH₄⁺-N>NO₃⁻-N>NO₂⁻-N, and the activity of nitrite reductase in C. vulgaris at NH₄⁺-N decline stage was significantly lower than that at NO₂⁻-N decline stage. In conclusion, C. vulgaris can significantly reduce the contents of NH₄⁺-N and NO₂⁻-N in water, and NO₂⁻-N may be reduced to NH₄⁺-N by intracellular nitrite reductase and assimilated by C. vulgaris. These results provide scientific basis for in-situ bioremediation of aquaculture waters.