Microbial Bioprocess for Extracellular Squalene Production and Formulation of Nanoemulsions

Sustainable production of natural resources requires a clean manufacturing process with clean raw materials due to the risk of environmental contaminations. Microbial cell factories have enabled the production of natural products including hydrophobic chemicals. However, the microbial cells limit the production titer because of the limited intracellular storage capacity and no transport across the cell membrane. Herein, we engineered Corynebacterium glutamicum to produce hydrophobic squalene as a model product. Notably, squalene was secreted in the defined culture medium without in situ extractions. As a result, the secreted squalene was related to the glycolipid association with synthesized squalene in the cell envelope and a facilitated secretion occurred during cell division. Protein engineering and pathway engineering of C. glutamicum enhanced the total squalene (tSQ) production, where fed-batch fermentation resulted in 1406 ± 16 mg/L tSQ production (32% tSQ in the medium). Promoting extraction with dodecane overlay, bio-squalene nanoemulsions were formulated and the creaming effects were analyzed for cosmetic application. Combining metabolic pathway engineering and facilitated secretion in microbial cells will be a good strategy to produce sustainable hydrophobic chemicals.