Enhancing DHA-rich lipid and protein co-production by Crypthecodinium cohnii using organosolv-derived olive pomace hydrolysate

The development of sustainable biorefineries based on heterotrophic microalgae offers a promising strategy for valorizing agro-industrial residues. In this study, olive pomace (OP), a major by-product of the olive oil industry, was evaluated as a substrate for the co-production of DHA-rich lipids and protein-enriched biomass using Crypthecodinium cohnii . Culture media derived from hydrolyzed OP were used to investigate the effect of key cultivation parameters, including inoculum size, salinity, yeast extract supplementation, and pH, on biomass growth and lipid accumulation. The novelty of this work lies in providing a comprehensive downstream valorization of the remaining biomass. A stepwise refinement of these parameters enabled significant improvements in process performance, leading to biomass concentrations of 4.24 g·L⁻¹, lipid contents of 39.08 ± 0.24%, and a threefold increase in DHA yield (193 ± 7 mg·L⁻¹). The fraction of residual biomass (540–640 g·kg⁻¹) exhibited an unexpectedly high protein content (42.09 ±0.49%), offering new insights into the biochemical profile of this dinoflagellate and highlighting its potential as a valuable co-product. Overall, these results demonstrate the feasibility of using OP-derived hydrolysates as a low-cost substrate for the simultaneous production of DHA and protein-rich biomass, supporting the development of integrated and low-waste biorefinery based on C. cohnii .