Design of Dynamic Experiments (DoDE) to optimize the lutein production by Coccomyxa onubensis in a novel high-cell density reactor

Lutein, a carotenoid involved in light harvesting and photoprotection, has demonstrated significant therapeutic benefits, particularly for ocular health. This study aims to optimize biomass and lutein production from an extremophilic microalga Coccomyxa onubensis in a fed-batch system. C. onubensis was cultivated in a membrane high-cell-density photobioreactor. Design of Dynamic Experiments was employed to optimize dynamic cultivation factors, including time-varying light intensity profiles (80–800 μmol∙ m⁻²∙ s⁻¹), nitrogen feeding strategies (60–600 mg_N∙ L⁻¹), and three constant CO₂ levels (0.04 %, 1 %, and 10 %). Biomass productivity peaked at 1.28 ± 0.23 g∙ L⁻¹∙ d⁻¹ under 6.5 % CO₂, with gradual light increase (peak 800 μmol photons∙ m⁻² ∙ s⁻¹), and moderately high nitrogen (499 mg∙ L⁻¹ over 5 days). Conversely, lutein productivity reached 3.32 ± 0.58 mg∙ L⁻¹∙ d⁻¹ under moderate light intensity (peak 414 μmol photons∙ m⁻²∙ s⁻¹) and limited nitrogen (217 mg_N∙ L⁻¹ over 1 day) under 10 % CO₂. This work provides optimized and validated cultivation strategies for impactful time varying factors, implementable at industrial level. DoDE proved to be valuable tool for multifactorial optimization and interpretation of complex interactions, significantly enhancing biomass and lutein production under acidophilic, high cell density conditions where bacterial contamination can be minimized.