Optimization of Nostoc sp. biomass and protein composition in high-density cultivator based on DoE

High-density (HD) cultivation systems are a valuable alternative to improve the productivity of phototrophic microorganisms. They are designed to address the major challenges such as low cell densities, light attenuation, and inefficient gas exchange, offering innovative solutions. This work investigates the HD cultivation of the nitrogen-fixing cyanobacterium Nostoc sp. PCC 7120 under diazotrophic conditions, employing a hydrophobic membrane system for efficient CO 2 and N 2 transfer. A design of experiment (DoE) approach is applied to evaluate the influence of key operating variables, including light intensity, photoperiod, and phosphorus concentration, on biomass productivity (P X ) and nitrogen fixation (P N ). Mathematical models, describing the interactions between these process variables and growth responses are developed and validated, enabling the optimization of cultivation conditions to maximise productivity. Remarkable biomass productivity of 1.53 g X L −1 d −1 was obtained in this system, under diazotrophic conditions. P X was largely affected by light intensity, which should be supplied continuously, aligning with the ability of heterocystous species like Nostoc sp. of simultaneously perform photosynthesis and N-fixation. Additionally, phosphorus concentration was a critical parameter that must be precisely controlled to prevent substrate limitation or inhibition, with an optimal value of 34.1 mg P L −1 . The produced biomass was further characterised with a focus on its nitrogen-rich molecules, particularly proteins, and nutritional value, assessed through essential amino acid content. The results showed that protein content was improved (53.4 %) at 280 μmol m −2 s −1 , 25.5 mg P L −1 , and 18 h light period, with essential amino acid ranging between 11 % and 15 %. • HD cultivation is remarkably efficient for N 2 supply to diazo-phototrophic species. • DoE-derived models are efficient in representing N fixation and biomass growth for Nostoc . • Optimized conditions led to a remarkable productivity of 1.53 g X L −1 d −1 biomass under diazotrophic conditions. • Under 280 μmol m −2 s −1 , 25.5 mg P L −1 , and 18 h light period a protein content of 53.4 % was obtained. • Production of high-value biomass with 11–15 % essential amino acid content was achieved.