Individual process steps optimization of Cupriavidus necator-catalyzed production of α-humulene

The global demand for the valuable sesquiterpene α-humulene is experiencing a period of sustained growth, with applications covering a diverse range of sectors, including food, fragrances, cosmetics, and pharmaceuticals. Until now, the microbial α-humulene production using Cupriavidus necator has been conducted under non-optimized process conditions, despite the enormous advantages this method offers over traditional extraction from plants or a chemical synthesis of α-humulene. In this study, the established process parameters, including the composition of the minimal medium, the L-rhamnose induction and the process temperature, were varied and optimized using shake flasks. Furthermore, the C. necator -based α-humulene production process was divided into two temperature stages. When compared to the initial conditions, these optimizations resulted in an enhanced C. necator growth and/or elevated α-humulene levels. The combination of all individual optimizations into an integrated process led to a notable increase in sesquiterpene levels, from 9.5 to 32.4 mg/L, representing a 241 % increase compared to the initial conditions without any optimizations. • Optimization of the process parameters for the α-humulene production using C. necator was successfully achieved. • The high L-rhamnose concentrations of 1 and 2 % (v/v) enhanced the final α-humulene levels. • Lowering of the cultivation temperature from 30 to 25 °C during the process increases final α-humulene levels. • Combining all individual process improvements adds up to the positive impact of each individual change. • Compared to the standard process, all individual improvements combined led to a 3.4-fold increase in final α-humulene levels.