**4. Summary and Conclusions**

In this study we reported the development of a continuously operated amine transaminase-catalyzed reaction, which is based on the integration of a reactive crystallization step for the in situ-removal of the product amine as a product salt. The presented concept involves the use of a membrane reactor, which retains the whole cell biocatalyst and two separate vessels for the application and collection of the donor salt (saturator) and product salt (crystallizer). The saturator provides a constant concentration of the required donor amine salt isopropylammonium 3,3-diphenylpropionate simultaneous to the crystallizer that collects the product amine salt (*S*)-1-(3-methoxyphenyl)ethylammonium 3,3-diphenylpropionate.

In conclusion, the shown triple vessel concept with its central membrane reactor and final crystallizer allows to overcome the very unfavorable chemical reaction equilibrium of the amine transaminase-catalyzed reaction in a continuously operated vessel concept. A fully stoichiometric reaction was achieved, which is not obtained in classical reaction concepts using isopropylamine as donor amine. The spatial separation of biocatalyst, saturator and crystallizer allows a full control of these components, including its separate removal and recycling after usage. The shown concept achieves very high product purity by the integrated crystallization step with only very few downstream-processing steps. The application of the membrane reactor provides a localization of the biocatalyst and prevents the use of potentially harmful immobilization techniques. The herein achieved space-time-yield of 1.2 g/(L·d) is directly correlated with the applied biocatalyst activity, which will increase in parallel with higher biocatalyst loadings. Future studies will also target the optimization of the shown concept in order to improve process productivity in such a continuous reaction mode. This primarily includes

techniques to prevent the undesired nucleation by an optimized reactor design and the use of purified enzyme within the biocatalyst chamber.

**Author Contributions:** Conceptualization, J.v.L. and D.H.; methodology, D.H., P.K. and E.T.; formal analysis, D.H. and J.v.L.; investigation, D.H. and J.v.L.; writing—original draft preparation, D.H.; writing—review and editing, J.v.L. and E.T.; visualization, D.H. and J.v.L.; supervision, J.v.L.; project administration, J.v.L.; funding acquisition, J.v.L. All authors have read and agreed to the published version of the manuscript.

**Funding:** Funding by German Research Foundation (DFG, grant LA 4183/1-1), the Central Innovation Program SME of the Federal Ministry for Economic Affairs and Energy (ZIM, grant number 16KN073233) and the Leibniz ScienceCampus Phosphorus Research Rostock (grant CryPhos), is gratefully acknowledged.

**Acknowledgments:** The authors thank Hubert Bahl and Ralf-Jörg Fischer for their continuous support in microbiology, Dirk Michalik and Heike Borgwaldt for their assistance with NMR measurements, Martin Köckerling and Florian Schröder for their assistance with XRPD-measurements and Sandra Diederich and Rike Thomsen for technical and experimental support. We acknowledge financial support by Deutsche Forschungsgemeinschaft and Universität Rostock within the funding programme Open Access Publishing.

**Conflicts of Interest:** The authors declare no conflict of interest.
