Biocatalytically Active Surfaces by Additive Manufacturing

To fulfil the demands of future industry, smart reactors (sustainable, multipurpose, artificial intelligent, resilient, transferable) are essential. Additive manufacturing is a key technology, when aiming for a flexible and tailor-made reactor set-up in a fast and cost-effective manner. When applied in packed bed reactors, these structures can be designed in a heat and mass transfer enhancing manner, thus, increasing the efficiency of the entire process. The focus of the present work is the development of an environmentally friendly immobilization platform, as well as a rational and tailored reactor design using smart process technology. Here, the goal is to implement improved immobilization strategies for autonomously operated bioprocesses, using three approaches: 1) additively manufactured smart hydrogels with encapsulated enzymes; 2) improved enzyme carriers by 3D-printing using smart materials for autonomous process control; 3) universal, 3D-printable immobilization platform fabricated from spider silk fusion proteins, enabling a holistically sustainable immobilization process without the use of harsh chemicals.
Additively manufactured PNiPAm hydrogels were processed with encapsulated enzymes (Est2, Esterase 2 from Alicyclobacillus acidocaldarius, and CalB, Candida antarctica lipase B) and tested for their applicability as universal immobilization matrix, reaching residual activities of 22 and 51 %. The proof-of-concept in aqueous and organic media was successful and the applied hydrogels showed one third of the initial activity after four subsequent runs. In addition, diffusion limitation was present but reduced by application of a rotating bed reactor. Secondly, surface-modification by controlled polymerization on 3D-printed structures was demonstrated. The applicability in aqueous and organic environment is given, with enhanced activities of the Est2 on PA12-PAA structures. Maximum activity was determined for a polymer synthesis time of 2 h, where a 6-fold increased amount of enzyme was detected in contrast to untreated PA12.
In addition, the polymer layer thickness and the swelling behavior of the respective polymer have a significant influence on the enzyme activity and applicability in autonomous process control. In the last part of this study, the application of a fully sustainable immobilization approach was investigated. The biochemical characterization revealed improved stabilities of the spider silk fusion protein compared to the wildtype. Additionally, the self-assembly leads to trivial processing of the hydrogel and is stable under the applied reaction conditions. No leaching was detectable, and the recyclability study was terminated after 10 runs, with 58 % residual activity.