Optimization of Heterotrophic Culture Conditions for Lipid Accumulation in Cyclotella cryptica and Analysis of the Underlying Metabolic Regulation Mechanism

To investigate the optimal conditions for lipid accumulation in heterotrophically cultured Cyclotella cryptica, we initially conducted autotrophic and heterotrophic single-factor comparative experiments on sodium nitrate (NaNO₃), sodium dihydrogen phosphate dihydrate (NaH₂PO₄·2H₂O), sodium silicate (Na₂SiO₃) concentrations in F/2 medium and determined the cultivation temperature. Using autotrophic optimization results as controls, response surface methodology (RSM) was employed to optimize the cultivation conditions based on the three factors (sodium nitrate concentration, sodium silicate concentration, and temperature) that had the greatest impact on biomass and lipid yield. Combined with transcriptome analysis, the difference in lipid metabolism between the two culture methods was compared. Combined with transcriptome analysis, the difference in lipid metabolism between the two cultivation methods was compared. Single-factor analysis indicated the identical optimal parameters for heterotrophic and autotrophic cultivation, sodium nitrate 149.6 mg/L, sodium dihydrogen phosphate dihydrate 11.44 mg/L, sodium silicate 41.75 mg/L and 25.0 ℃. However, RSM optimization revealed the distinct heterotrophic optimum condition, sodium nitrate 145.7 mg/L, sodium silicate 43.0 mg/L and 26.0 ℃, different from that of autotrophic optimum condition, nitrate 147.2 mg/L, sodium silicate 43.0 mg/L and 24.2 ℃. Under optimized heterotrophic conditions, biomass and total lipid productivity increased by 25.64% and 51.53%, respectively, in comparison with autotrophic cultivation. Transcriptomic analysis demonstrated that the expression of genes associated with the glycolysis, pentose phosphate pathway, fatty acid synthesis and long-chain fatty acid elongation pathways was upregulated under heterotrophy while that of genes functioning in tricarboxylic acid cycle and fatty acid degradation was downregulated. These metabolic shifts confirmed that heterotrophic cultivation significantly enhanced lipid accumulation in C. cryptica. Our findings provided a theoretical basis for large-scale culture and lipid production of C. cryptica.