Sustainable in situ extraction of microalgae-derived terpenoids using functionalized silica microparticles

Chlamydomonas reinhardtii is a green microalga that has been genetically engineered to produce heterologous terpenoid metabolites. While this green biochemical approach for producing high-value chemicals holds tremendous potential, the current in situ extraction methods, utilizing traditional solvents such as dodecane and perfluorinated chemicals, present separation challenges, unfavorable economics, and risk of cell toxicity. Here, we develop a low-cost solvent-free approach based on silanized silica particles to fill the technology gap. We determine the feasibility and specificity of three differently coated (C11, C16, C18) silica particles to extract nine terpenoid metabolites from C. reinhardtii cultures during cultivation. The results reveal that the extraction efficiencies of functionalized particles, particularly those coated with C18, demonstrated high potential as an alternative to traditional extraction methods. While the extraction efficiencies of coated microparticles are for most compounds lower than those observed for dodecane-based extractions, it holds higher potential for larger-scale cultivations where the use of dodecane is restricted due to its flammability and tendency to form emulsions. In addition, the present study provides insights into the optimization of particle-to-culture volume ratio, particle saturation, and the required number of product elution steps, and proves the feasibility of an upscaled extraction of the sesquiterpenoid patchoulol as a representative metabolite in 5 L hanging bag reactors. This study paves the way for a circular bioprocess where product capture with traditional solvent overlays is impractical due to the formation of emulsions, and at the same time allows for an easy product recovery (solid-liquid separation) and reuse of functionalized particles over numerous cycles.