Brewer's spent grain and crude glycerol: Sustainable substrates for 2-phenylethanol production by Yarrowia lipolytica.

• Microbial fermentation for de novo production of the aroma molecule 2-phenylethanol. • Valorizing brewer's spent grain and crude glycerol to produce value-added compounds. • Use of Golden Gate Assembly for Yarrowia lipolytica strain manipulation. • Transformation of yeast with amyloglucosidase gene for starch hydrolysis. • Optimization of culture medium composition through central composite design. The development of natural, efficient, and cost-effective methods for producing the aromatic alcohol 2-phenylethanol (2-PE) is necessary. 2-PE, characterized by a rosy scent, is widely used in the cosmetic, pharmaceutical, and food industries. It is commonly utilized as a flavoring agent in desserts, baked goods, and beverages, enhancing the overall flavor with fruity, honeyed, creamy, and caramelized notes. In this study, the optimization of 2-PE production through fermentation of the yeast Yarrowia lipolytica on brewer's spent grain (BSG) was examined using a central composite design. The impact of crude glycerol (CG) and yeast extract (YE) on 2-PE production were investigated. A prerequisite for utilizing BSG as a substrate is the hydrolysis of its complex components, primarily starch. This was achieved by transforming the amyloglucosidase (AMG) gene into the genome of Y. lipolytica . Fermentations were carried out in 500 mL baffled Erlenmeyer flasks with a working volume of 100 mL. Optimally, 1.52 g/L of 2-PE was produced after 168 h fermentation in a medium containing 2.87 g/L yeast extract and 45.9 g/L crude glycerol in BSG.