Biocatalysis: A sustainable approach to chemical synthesis

Biocatalysis is the use of enzymes and occasionally other biological molecules to catalyze chemical reactions. It is used in a wide range of applications, including the production of chemicals, pharmaceuticals, flavoring agents, or food and beverages.

One of the main advantages of biocatalysis is its ability to highly selectively catalyze reactions under mild conditions, such as at moderate temperatures and pH values. This makes it a more environmentally friendly and intrinsically safer alternative than traditional chemical synthesis.

In addition to its environmental benefits, biocatalysis can also offer economic advantages, such as lower production costs and the ability to synthesize complex molecules that are difficult to access by other methods. These factors make biocatalysis an attractive option for many industries.

Some specific examples for applications of biocatalysis include the production of enzymes for laundry detergents, the synthesis of pharmaceuticals, and the production of biofuels.

 

Biocatalysis, the future of sustainable chemistry

Enzymes display remarkable chemo-, regio-, and stereoselectivity. The identification of a suitable enzyme that converts the mandated substrate to the desired product isomer, allows to significantly shorten reaction pathways compared to traditional multistep synthesis. Notably, protection and deprotection strategies can often be omitted, and difficult isomer separation may be avoided.

The mild conditions and modest temperatures employed in enzymatic reactions, in combination with the highly selective catalysis, typically result in few to no significant side reactions.

Additionally, environmental and toxicological benefits can be reaped by avoiding pathways that employ metal catalysts and hazardous reagents.

 

Exploring enzymes types and their applications

Enzymes are divided into seven classes according to the type of reaction they catalyze.

EC 1 - Oxidoreductases catalyze oxidation-reduction reactions, hence involving the transfer of electrons from one molecule to another.

EC 2 - Transferases catalyze the transfer of certain groups from one molecule to another.

EC 3 - Hydrolases catalyze the hydrolysis of various bonds.

EC 4 - Lyases catalyze the cleavage of various bonds by means other than hydrolysis, often forming a double bond or a new ring structure.

EC 5 - Isomerases catalyze the rearrangement of atoms within a molecule to form a structural isomer.

EC 6 - Ligases catalyze the formation of a new chemical bond, facilitated by the concommitant consumption of a high-energy cosubstrate.

EC 7 - Translocases facilitate the movement of ions or molecules across membranes or their separation within membranes.

 

Factors that influence the performance of enzymes

The activity and stability of enzymes are governed by several factors, including temperature, pH, solvent type or buffer conditions, water activity, immobilization, stirring, and reaction parameters. Understanding these factors allows us to optimize enzyme-catalyzed reactions and to improve their efficiency and effectiveness.

Temperature
Enzymes are subject to two competing processes: On the one hand an increase in temperature raises the catalytic turnover according to Arrhenius, and on the other hand it accelerates the breakdown of the enzyme. The combined result is a bell-shaped temperature-activity curve, i.e., enzymes have a specific optimal temperature with peak productivity flanked by diminishing returns at both higher and lower temperatures.

Enzymes are frequently engineered to shift the temperature range of optimal activity and to thereby make them more resistant to denaturation. The desired shift in temperature can be both to higher and lower temperatures to either allow for reactions at elevated temperatures or to ensure high activity even at low temperatures. Enzymes can also be engineered to allow for a broadened substrate scope or non-natural substrates and products. Finally, engineering may confer stability in non-conventional media.

pH
Similar to temperature dependence, enzymes show peak activity within a narrow pH range. When moving away from the optimal pH, the activity is diminished and finally lost. Consequently, some form of pH control is often necessary.

Solvent/buffer
Choice of buffer and buffer strength may provide pH control and stability in aqueous systems and consequently also determine activity and stability. In non-conventional solvents, pH effects are defined by the interplay of physicochemical properties of substrates, products, and the reaction medium. This may go so far as to render these effects completely void. However, solvents themselves may have a strong impact on enzyme stability and activity. If necessary, process analytical technology (PAT) and titration allow for active pH control in any case. Choice of solvent also strongly affects the enzymatic activity.

Water activity
Enzymes require a minimum amount of water to sustain their catalytic function. This is typically due to the need for structurally-bound water and becomes important for reactions in non-aqueous media. However, the share of water required is frequently less than 1% of the total reaction medium. As a consequence - and perhaps counter-intuitively - a small amount of water is also added to reactions that from a thermodynamic perspective should ideally be 'water-free' (i.e., condensation reactions).

Immobilization
Immobilization of free enzymes onto solid materials (enzyme carriers) often provides stabilization and thereby prolonged lifetime of the enzyme. However, it also affects enzymatic activity whereby an increase is the more desirable yet less prevalent result. The success of the immobilization is strongly defined by the compatibility between enzyme and enzyme carrier material, and trials with several different materials are advisable. Arguably, the most common enzyme carrier materials are synthetic polymers (resins), but inorganic materials such as glass, natural fibers, etc. are also used and bio-based organic alternatives gain traction as well. The final choice is typically based on overall catalytic productivity.

Inherently connected to the enzyme carrier material is the method of attachment. While some enzymes, notably lipases, may be attached non-covalently by hydrophobic interactions, covalent attachment is often needed. The chemistries used hereby are versatile and frequently react nucleophilic free amines or thiols from the enzyme surface with electrophilic reactive groups (epoxides, aldehydes) on the carrier surface. The exact choice of attachment strategy will definitely affect the catalytic properties of the obtained heterogenous biocatalyst preparation.

Read more in our insight 'Introduction to Enzyme Immobilization'.

Mixing
In the case of industrially preferred immobilized enzymes, the method of mixing determines the longevity of the heterogeneous catalyst. For batch reactors, the rotating bed reactor offers almost complete protection against the grinding effects that standard stirred tank reactors suffer from. As a result, extended recycling becomes feasible and convenient.

 

 

A rotating bed reactor (RBR) is designed to prevent disintegration of solid phase particles by retaining them inside a rotating cylinder during the reaction. This minimizes solid phase debris even at high speeds, which would normally occur due to mechanical forces when using stirred tank reactors. Due to this characteristic of the RBR, reaction rates can be improved by increasing the stirring speed without simultaneously shortening the lifespan of the solid particles. This ensures easy in-line monitoring of the process, as well as a more efficient recycling of the solid phase, making the RBR a cost and time efficient alternative to conventional methods for immobilized enzyme biocatalysis.

Reaction parameters
Perhaps more related to the efficiency of the enzymatic reaction than the stability of the enzyme are parameters such as reagent stoichiometry, gas introduction method, and the minimization of temperature or concentration gradients. Proper mixing ensures the necessary mass transfer of all substrates to the enzyme and is crucial especially in three-phase reactions where saturation with gaseous reagents has to be considered as well. Here too, the RBR provides important advantages over stirred tank reactors.

 

Exploring the rapid development of biocatalysis

Don't miss out on the exciting developments in biocatalysis! With engineered enzymes that are more stable, more active, and capable of working in a wider range of conditions, biocatalysis is becoming increasingly accessible and easy to use. And with a growing range of free and immobilized enzymes available on the market, now is the perfect time to start exploring the benefits of biocatalysis for yourself. - Join the biocatalysis revolution today!

Recommended reading

Application L1701
A novel hierarchically structured siliceous packing to boost the performance of rotating bed enzymatic reactors

Katarzyna Szymańska, Klaudia Odrozek, Aurelia Zniszczoł, Wojciech Pudło, and Andrzej B. Jarzębski Chem. Eng. J., 2017, 315, pp. 18-24.

Application 1014
Biocatalysis by immobilized enzymes in a rotating bed reactor

Time lapse video showing how straightforward it is to use immobilized enzymes in a rotating bed reactor. A substrate giving a yellow coloured product was used to follow the reaction progress of an ester hydrolysis by an immobilized lipase. This substrate is commonly used to screen and characterize lipases.

Application L1704
Biocatalysis engineering: the big picture

Roger A. Sheldon and Pedro C. Pereira Chem. Soc. Rev., 2017, 46(10), pp. 2678-2691.

Application 1023
Biocatalysis in rotating bed reactors - from screening to production

Enzyme screening is an important step in the development of a biocatalytic application. The behaviour of the biocatalyst is often hard to predict, meaning that different combinations of materials and reaction conditions need to be tested.

Application L2204
Design of a green chemoenzymatic cascade for scalable synthesis of bio-based styrene alternatives

Philipp Petermeier, Jan Philipp Bittner, Simon Müller, Emil Byström, and Selin Kara Green Chemistry, 2022, 24(18), pp. 6889-6899.

Application 1003
Improving reactions in emulsions using a rotating bed reactor

When working with an emulsion (and particularly with a heterogeneous catalyst) the mass transfer between the phases is critical. Insufficient mixing leads to lower interfacial area per volume, and in turn to poor mass transfer across the phases.

Application L1301
Efficient biocatalysis with immobilized enzymes or encapsulated whole cell microorganism by using the SpinChem reactor system

Hendrik Mallin, Jan Muschiol, Emil Byström, and Uwe T. Bornscheuer ChemCatChem, 2013, 5, pp. 3529-3532.

Application 1002
Efficient synthesis of chiral lactones by encapsulated cells in a rotating bed reactor

Whole cell biocatalysis is powerful, but not straightforward. One way of utilizing whole cells is to encapsulate them in a matrix such as alginate to make them easier to separate from a reaction mixture. However, alginate beads are not mechanically stable enough to be packed into columns and are easily destroyed in stirred tank reactors (STR). This makes enzyme recycling ineffective, at the same time as mass transfer limitations may prevail.

Application 1028
Enzyme immobilization screening using rotating bed reactors

Finding the optimal chemistry and solid-phase material for immobilization of enzymes relies heavily on trial and error. The right resin will ensure satisfactory immobilization yield, as well as high activity and stability of the enzyme.

Application L2213
Impact of critical parameters influencing enzymatic production of structured lipids using response surface methodology with water activity control

Ariana Causevic, Eimantas Gladkauskas, Kim Olofsson, Patrick Adlercreutz, and Carl Grey Biochem. Eng. J., 2022, 187, 108610.

Application L1903
L-Asparaginase production in rotating bed reactor from Rhizopus microsporus IBBL-2 using immobilized Ca-alginate beads

Anup Ashok and Santhosh Kumar Devarai 3 Biotech, 2019, 9(9), 349.

Application L1402
Lipase catalyzed regioselective lactamization as a key step in the synthesis of N-Boc (2R)-1,4-oxazepane-2-carboxylic acid

Carl-Johan Aurell, Staffan Karlsson, Fritiof Pontén, and Søren M. Andersen Org. Process Res. Dev., 2014, 18(9), pp. 1116-1119.

Application L1602
Modularized biocatalysis: Immobilization of whole cells for preparative applications in microaqueous organic solvents

Jochen Wachtmeister, Philip Mennicken, Andreas Hunold, and Dörte Rother ChemCatChem, 2016, 8, pp. 607-614.

Application L2117
Multi‐enzyme cascade reaction in a miniplant two‐phase‐system: Model validation and mathematical optimization

Jens Johannsen, Francesca Meyer, Claudia Engelmann, Andreas Liese, Georg Fieg, Paul Bubenheim, and Thomas Waluga AIChE J., 2021, 67(4), e17158.

Application L2102
New frontiers in enzyme immobilisation: Robust biocatalysts for a circular bio-based economy

Roger A. Sheldon, Alessandra Basso, and Dean Brady Chem. Soc. Rev., 2021, 50(10), pp. 5850-5862.

Application L2110
Process design of a continuous biotransformation with in situ product removal by cloud point extraction

Oliver Fellechner and Irina Smirnova Can. J. Chem. Eng., 2021, 99, pp. 1035-1049.

Application L2112
Production of hydroxytyrosol rich extract from Olea europaea leaf with enhanced biological activity using immobilized enzyme reactors

Alexandra V. Chatzikonstantinou, Αrchontoula Giannakopoulou, Stamatia Spyrou, Yannis V. Simos, Vassiliki G. Kontogianni, Dimitrios Peschos, Petros Katapodis, Angeliki C. Polydera, and Haralambos Stamatis Environ. Sci. Pollut. Res., 2022, 29, pp. 29624-29637.

Application L2115
Production of recombinant choline oxidase and its application in betaine production

S. Lokesha, Y. S. Ravi Kumar, P. S. Sujan Ganapathy, Prashant Gaur, and H. M. Arjun 3 Biotech, 2021, 11, 410.

Application L2119
Pseudomonas taiwanensis biofilms for continuous conversion of cyclohexanone in drip flow and rotating bed reactors

Ingeborg Heuschkel, Selina Hanisch, Daniel C. Volke, Erik Löfgren, Anna Hoschek, Pablo I. Nikel, Rohan Karande, and Katja Bühler Eng. Life Sci., 2021, 21(3-4), pp. 258-269.

Application L1703
Reaction engineering of biocatalytic (S)-naproxen synthesis integrating in-line process monitoring by Raman spectroscopy

M. Aßmann, A. Stöbener, C. Mügge, S. K. Gaßmeyer, L. Hilterhaus, R. Kourist, A. Liese, and S. Kara React. Chem. Eng., 2017, 2(4), pp. 531-540.

Application L1606
Recent advances in whole cell biocatalysis techniques bridging from investigative to industrial scale

Jochen Wachtmeister and Dörte Rother Curr. Opin. Biotechnol., 2016, 42, pp. 169-177.

Application 9001
Recycling biocatalysts with rotating bed reactors in EasyMax systems

Investigating reactions can easily grow from an idea into very time-consuming projects, but the upside of properly understanding the reaction is great. The choice of equipment has a very large impact on the efforts required. The rotating bed reactor is a tool that unlocks the full potential of your Mettler-Toledo EasyMax™ 102 Advanced synthesis workstation for this development.

Application L1705
Role of biocatalysis in sustainable chemistry

Roger A. Sheldon and John M. Woodley Chem. Rev., 2018, 118(2), pp. 801-838.

Application 1032
Rotating bed reactor for immobilized enzymatic reactions

This case study presents a lipase-mediated stereoselective acetylation of a racemic amine in a rotating bed reactor.

Application 1025
Soft alginate beads used in a rotating bed reactor

Stirred vessels tend to damage soft heterogeneous catalysts, like enzymes immobilized in agarose or alginate beads, with activity loss and tedious workup as consequence. In a fixed bed reactor, these materials are easily compressed by the pressure gradient, leading to a loss of flow rate. Overcoming these challenges opens up the possibility to use biocatalysis as a tool for greener processes and more sustainable manufacturing.

Application 9002
Screening of Immobilized Enzymes – Fast and Convenient Reaction Optimization

The stable reaction environment in the EasyMax™ 102 Advanced synthesis workstation and the high flow rates through the SpinChem® RBR allowed for quick and convenient screening of different immobilized lipases to find the enzyme most suitable for further reaction optimization.

Application 1027
Screening of immobilized lipases using rotating bed reactors

The SpinChem rotating bed reactor (RBR) has been proven to be a time and labor-efficient tool in the screening of biocatalysts. Here, we present the quick simultaneous screening of six different immobilized lipases for the esterification of lauric acid to propyl laurate using our pre-packed MagRBR lipase screening kit.

Application 1044
Simple scale-up using flexible reactors

Research and development quickly takes new directions, and the requirements on a laboratory may vary with every new project. Limiting yourself to equipment with a narrow scope of conditions and applications may become expensive, since new equipment must be acquired for anything out of scope. With budgets quickly consumed by other projects, the need for new equipment may mean significant delays and a reduced capability to take on emerging opportunities.

Application 1009
Multistep synthesis or simultaneous extraction simplified in a rotating bed reactor

The synthesis of products, such as active pharmaceutical ingredients (APIs), often involves multiple steps using heterogeneous catalysts or adsorbents. Thus, the simultaneous use of multiple solid phases either during synthesis or downstream processing is frequently highly advantageous.

Application 1040
The importance of baffles in a reactor vessel

Can you use a rotating bed reactor (RBR) in any type of vessel? - Absolutely! Would the performance be higher with baffles in the vessel? - Definitely!

Application L2001
Using SpinChem rotating bed reactor technology for immobilized enzymatic reactions: A case study

Subhash Pithani, Staffan Karlsson, Hans Emtenäs, and Christopher T. Öberg Org. Process Res. Dev., 2019, 23(9), pp. 1926-1931.

Application L1706
Whole-cell cascade biotransformations for one-pot multistep organic synthesis

Shuke Wu and Zhi Li ChemCatChem, 2018, 10(10), pp. 2164-2178.

Application 1050
Lipase-catalyzed hydrolysis in 750 L using a rotating bed reactor

Biocatalysis offers many benefits in the production of chemicals and active pharmaceutical ingredients. One major challenge has been the deployment of immobilized enzymes in an efficient way on large scale. The rotating bed reactor offers a convenient way to scale a biocatalytic process.

Application 1051
How the loading of solids influences reaction speed

Sometimes you don’t want to pack the entire rotating bed reactor full with your solid-phase material. Fully loading might simply be wasteful, or you may want to experiment with your reaction conditions. But how does the amount of solids in the rotating bed reactor influence the reaction performance? Can you use only 10% of the full capacity?

Application L2202
Asymmetric hydrogenation of C = C bonds in a SpinChem reactor by immobilized old yellow enzyme and glucose dehydrogenase

Teng Ma, Weixi Kong, Yunting Liu, Hao Zhao, Yaping Ouyang, Jing Gao, Liya Zhou, and Yanjun Jiang Appl. Biochem. Biotechnol., 2022, 194, pp. 4999–5016.

Application L2207
Lipase catalyzed green epoxidation of oleic acid using ultrasound as a process intensification method

Adriana Freites Aguilera, Pontus Lindroos, Jani Rahkila, Mark Martinez Klimov, Pasi Tolvanen, and Tapio Salmi Chem. Eng. Process. Process Intensif., 2022, 174, 108882.

Application L2212
Epoxidation of tall oil fatty acids and tall oil fatty acids methyl esters using the SpinChem® rotating bed reactor

Krzysztof Polaczek, Eliza Kaulina, Ralfs Pomilovskis, Anda Fridrihsone, and Mikelis Kirpluks J. Polym. Environ., 2022, 30, pp. 4774–4786.

Application L2203
Peroxygenase-driven ethylbenzene hydroxylation in a rotating bed reactor

Markus Hobisch, Piera De Santis, Simona Serban, Alessandra Basso, Emil Byström, and Selin Kara Org. Process Res. Dev., 2022, 26(9), pp. 2761-2765.

Application L2301
Comparison of four immobilization methods for different transaminases

Tobias Heinks, Nicolai Montua, Michelle Teune, Jan Liedtke, Matthias Höhne, Uwe T. Bornscheuer, and Gabriele Fischer von Mollard Catalysts, 2023, 13(2), 300.

Application L2302
Fatty acid epoxidation on enzymes: Experimental study and modeling of batch and semibatch operation

Wilhelm Wikström, Adriana Freites Aguilera, Pasi Tolvanen, Robert Lassfolk, Ananias Medina, Kari Eränen, and Tapio Salmi Ind. Eng. Chem. Res., 2023, 62(23), pp. 9169-9187.

Application L2303
Amination of a green solvent via immobilized biocatalysis for the synthesis of Nemtabrutinib

Christopher K. Prier, Karla Camacho Soto, Jacob H. Forstater, Nadine Kuhl, Jeffrey T. Kuethe, Wai Ling Cheung-Lee, Michael J. Di Maso, Claire M. Eberle, Shane T. Grosser, Hsing-I Ho, Erik Hoyt, Anne Maguire, Kevin M. Maloney, Amanda Makarewicz, Jonathan P. McMullen, Jeffrey C. Moore, Grant S. Murphy, Karthik Narsimhan, Weilan Pan, Nelo R. Rivera, Anumita Saha-Shah, David A. Thaisrivongs, Deeptak Verma, Adeya Wyatt, and Daniel Zewge ACS Catal., 2023, 13(12), pp. 7707-7714.

Application L2120
Rotating bed reactor packed with heterofunctional structured silica-supported lipase. Developing an effective system for the organic solvent and aqueous phase reactions

Daria Kowalczykiewicz, Katarzyna Szymańska, Danuta Gillner, and Andrzej B. Jarzębsk Microporous Mesoporous Mater., 2021, 312, 110789.

Application L2003
Pharmaceutical industry perspectives on flow chemocatalysis and biocatalysis

Laura Leemans Martin, Theo Peschke, Francesco Venturoni, and Serena Mostarda Curr. Opin. Green Sustainable Chem., 2020, 25, 100350.

Application L2104
Compartmentalization in biocatalysis

Robert Kourist and Javier González‐Sabín In: Biocatalysis for Practitioners: Techniques, Reactions and Applications

Application L2111
Streamlining design, engineering, and applications of enzymes for sustainable biocatalysis

Roger A. Sheldon and Dean Brady ACS Sustainable Chem. Eng., 2021, 9(24), pp. 8032–8052.

Application L2114
Practical multienzymatic transformations: Combining enzymes for the one‐pot synthesis of organic molecules in a straightforward manner

Jesús Albarrán‐Velo, Sergio González‐Granda, Marina López‐Agudo, and Vicente Gotor‐Fernández In: Biocatalysis for Practitioners: Techniques, Reactions and Applications

Application L2008
Probing batch and continuous flow reactions in organic solvents: Granulicella tundricola hydroxynitrile lyase (GtHNL)

José Coloma, Yann Guiavarc'h, Peter-Leon Hagedoorn, and Ulf Hanefeld Catal. Sci. Technol., 2020, 10(11), pp. 3613-3621.

Application L2304
Boosting the catalytic performance of a marine yeast in a SpinChem® reactor for the synthesis of perillyl alcohol

Silvia Donzella, Concetta Compagno, Francesco Molinari, Francesca Paradisi, and Martina Letizia React. Chem. Eng., 2023, 8(12), pp. 2963-2966.

Application L2305
Enzymatic synthesis of ascorbyl palmitate in a rotating bed reactor

Jessica Holtheuer, Luigi Tavernini, Claudia Bernal, Oscar Romero, Carminna Ottone, and Lorena Wilson Molecules, 2023, 28(2), 644.

Application L2306
Selective peroxygenase-catalysed oxidation of toluene derivatives to benzaldehydes

Yutong Wang, Niklas Teetz, Dirk Holtmann, Miguel Alcalde, Jacob M. A. van Hengst, Xiaoxiao Liu, Mengfan Wang, Wei Qi, Wuyuan Zhang, and Frank Hollmann ChemCatChem, 2023, 15(13), e202300645.

Application L2208
Process intensification in oxidative biocatalysis

Guillem Vernet, Markus Hobisch, and Selin Kara Curr. Opin. Green Sustainable Chem., 2022, 38, 100692.

Application L2310
Large scale production of vanillin using an eugenol oxidase from Nocardioides sp. YR527

Daniel Eggerichs, Kathrin Zilske, and Dirk Tischler Mol. Catal., 2023, 546, 113277.

Application L2311
New enzymatic reactor designs: From enzymatic batch to 3D microreactors and monoliths

Kim Shortall, Katarzyna Szymańska, Cristina Carucci, Tewfik Soulimane, and Edmond Magner In: Biocatalyst Immobilization, Foundations and Applications, 2022

Application 1069
Cellulose-bead immobilized enzymes as biodegradable and renewable catalysts

Environmentally benign and safe synthesis is enabled by highly active biocatalysts. To bolster economic and ecological aspects, catalyst reuse is essential and achieved by heterogenization of otherwise soluble enzymes onto solid supports. Here, this is demonstrated on novel renewable and non-polluting cellulose beads.

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