Conclusion and Outlook
The world-wide growing demand for natural flavors poses a chance for enzyme-based production processes to be implemented to specialty and fine chemicals industry. In vitro multi-enzyme cascade reactions are a promising part of such novel, enzyme-based approaches, in that they present a chance for process integration. Within our research group, a three-enzyme cascade reaction sequence with integrated cofactor regeneration and integrated intermediate extraction in a two-phase system has been successfully established and scaled up for application in a miniplant at the example of the ester cinnamyl cinnamate. Due to their high complexity compared to conventional chemical synthesis routes, however, such enzyme-based processes are difficult to mathematically describe. On the other hand, mathematical modeling is an important prerequisite for the industrial application of novel production processes.
Therefore, in this contribution the authors introduce an Aspen Custom Modeler® - implemented mathematical model to describe the aforementioned complex biotechnological production process for a miniplant-scale reactor setup. Here, sub-models for each thermodynamic phase and for the integrated extraction step respectively are connected through differential mass balance equations, resulting in one holistic mathematical model describing the cascade reaction. In this study, successful validation of the model with independent experimental data obtained from the miniplant is presented for the first time. Using this validated model, multi-objective mathematical optimization runs were performed with the in-house-developed optimization tool Adv:PO. As a result, the authors can now identify optimal process operating windows for any given multi-objective function within the cascade reaction. The Pareto-optimal dependency of cofactor concentration on cinnamyl cinnamate space-time yield is analyzed and shown in this work as a representative example for such optimization results, obtaining a suitable cofactor concentration range of 1.4 mmol/l to 4.2 mmol/l.
Future research will now further investigate the optimal operating settings for the miniplant-scale production process. Specifically, integration of the product separation step will be the focus of our work: promising results for cinnamyl cinnamate separation from the product mixture were already achieved using annular chromatography. Incorporating the downstream process into the mathematical model for process optimization will show the full potential of this novel process specifically, and the benefits of applying mathematical modeling and optimization to biotechnology in general.