Development of a fully integrated falling film microreactor for gas-liquid-solid biotransformation with surface immobilized O2-dependent enzyme

Juan Manuel Bolivar Bolivar, Christina E M Krämer, Birgit Ungerböck, Torsten Mayr, Bernd Nidetzky

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Microstructured flow reactors are powerful tools for the development of multiphase biocatalytic transformations. To expand their current application also to O2-dependent enzymatic conversions, we have implemented a fully integrated falling film microreactor that provides controllable countercurrent gas-liquid phase contacting in a multi-channel microstructured reaction plate. Advanced non-invasive optical sensing is applied to measure liquid-phase oxygen concentrations in both in- and out-flow as well as directly in the microchannels (width: 600μm; depth: 200μm). Protein-surface interactions are designed for direct immobilization of catalyst on microchannel walls. Target enzyme (here: d-amino acid oxidase) is fused to the positively charged mini-protein Zbasic2 and the channel surface contains a negatively charged γ-Al2O3 wash-coat layer. Non-covalent wall attachment of the chimeric Zbasic2_oxidase resulted in fully reversible enzyme immobilization with fairly uniform surface coverage and near complete retention of biological activity. The falling film at different gas and liquid flow rates as well as reactor inclination angles was shown to be mostly wavy laminar. The calculated film thickness was in the range 0.5-1.3×10-4m. Direct O2 concentration measurements at the channel surface demonstrated that the liquid side mass transfer coefficient (KL) for O2 governed the overall gas/liquid/solid mass transfer and that the O2 transfer rate (≥0.75mM·s-1) vastly exceeded the maximum enzymatic reaction rate in a wide range of conditions. A value of 7.5 (±0.5)s-1 was determined for the overall mass transfer coefficient KLa, comprising a KL of about 7×10-5m·s-1 and a specific surface area of up to 105m-1.

LanguageEnglish
JournalBiotechnology and Bioengineering
DOIs
StatusPublished - 2016

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Biotransformation
Enzymes
Gases
Immobilization
Oxidoreductases
Liquids
Mass transfer
Microchannels
Enzyme immobilization
Membrane Proteins
Proteins
Oxygen
Amino Acids
Bioactivity
Specific surface area
Reaction rates
Film thickness
Amino acids
Flow rate
Catalysts

Keywords

  • Falling-film microreactor
  • Gas/liquid/solid biotransformation
  • Immobilized oxidase
  • Microchannel
  • Oxygen mass transfer
  • Z-binding module

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

Cite this

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title = "Development of a fully integrated falling film microreactor for gas-liquid-solid biotransformation with surface immobilized O2-dependent enzyme",
abstract = "Microstructured flow reactors are powerful tools for the development of multiphase biocatalytic transformations. To expand their current application also to O2-dependent enzymatic conversions, we have implemented a fully integrated falling film microreactor that provides controllable countercurrent gas-liquid phase contacting in a multi-channel microstructured reaction plate. Advanced non-invasive optical sensing is applied to measure liquid-phase oxygen concentrations in both in- and out-flow as well as directly in the microchannels (width: 600μm; depth: 200μm). Protein-surface interactions are designed for direct immobilization of catalyst on microchannel walls. Target enzyme (here: d-amino acid oxidase) is fused to the positively charged mini-protein Zbasic2 and the channel surface contains a negatively charged γ-Al2O3 wash-coat layer. Non-covalent wall attachment of the chimeric Zbasic2_oxidase resulted in fully reversible enzyme immobilization with fairly uniform surface coverage and near complete retention of biological activity. The falling film at different gas and liquid flow rates as well as reactor inclination angles was shown to be mostly wavy laminar. The calculated film thickness was in the range 0.5-1.3×10-4m. Direct O2 concentration measurements at the channel surface demonstrated that the liquid side mass transfer coefficient (KL) for O2 governed the overall gas/liquid/solid mass transfer and that the O2 transfer rate (≥0.75mM·s-1) vastly exceeded the maximum enzymatic reaction rate in a wide range of conditions. A value of 7.5 (±0.5)s-1 was determined for the overall mass transfer coefficient KLa, comprising a KL of about 7×10-5m·s-1 and a specific surface area of up to 105m-1.",
keywords = "Falling-film microreactor, Gas/liquid/solid biotransformation, Immobilized oxidase, Microchannel, Oxygen mass transfer, Z-binding module",
author = "{Bolivar Bolivar}, {Juan Manuel} and Kr{\"a}mer, {Christina E M} and Birgit Ungerb{\"o}ck and Torsten Mayr and Bernd Nidetzky",
year = "2016",
doi = "10.1002/bit.25969",
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journal = "Biotechnology and Bioengineering",
issn = "0006-3592",
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T1 - Development of a fully integrated falling film microreactor for gas-liquid-solid biotransformation with surface immobilized O2-dependent enzyme

AU - Bolivar Bolivar,Juan Manuel

AU - Krämer,Christina E M

AU - Ungerböck,Birgit

AU - Mayr,Torsten

AU - Nidetzky,Bernd

PY - 2016

Y1 - 2016

N2 - Microstructured flow reactors are powerful tools for the development of multiphase biocatalytic transformations. To expand their current application also to O2-dependent enzymatic conversions, we have implemented a fully integrated falling film microreactor that provides controllable countercurrent gas-liquid phase contacting in a multi-channel microstructured reaction plate. Advanced non-invasive optical sensing is applied to measure liquid-phase oxygen concentrations in both in- and out-flow as well as directly in the microchannels (width: 600μm; depth: 200μm). Protein-surface interactions are designed for direct immobilization of catalyst on microchannel walls. Target enzyme (here: d-amino acid oxidase) is fused to the positively charged mini-protein Zbasic2 and the channel surface contains a negatively charged γ-Al2O3 wash-coat layer. Non-covalent wall attachment of the chimeric Zbasic2_oxidase resulted in fully reversible enzyme immobilization with fairly uniform surface coverage and near complete retention of biological activity. The falling film at different gas and liquid flow rates as well as reactor inclination angles was shown to be mostly wavy laminar. The calculated film thickness was in the range 0.5-1.3×10-4m. Direct O2 concentration measurements at the channel surface demonstrated that the liquid side mass transfer coefficient (KL) for O2 governed the overall gas/liquid/solid mass transfer and that the O2 transfer rate (≥0.75mM·s-1) vastly exceeded the maximum enzymatic reaction rate in a wide range of conditions. A value of 7.5 (±0.5)s-1 was determined for the overall mass transfer coefficient KLa, comprising a KL of about 7×10-5m·s-1 and a specific surface area of up to 105m-1.

AB - Microstructured flow reactors are powerful tools for the development of multiphase biocatalytic transformations. To expand their current application also to O2-dependent enzymatic conversions, we have implemented a fully integrated falling film microreactor that provides controllable countercurrent gas-liquid phase contacting in a multi-channel microstructured reaction plate. Advanced non-invasive optical sensing is applied to measure liquid-phase oxygen concentrations in both in- and out-flow as well as directly in the microchannels (width: 600μm; depth: 200μm). Protein-surface interactions are designed for direct immobilization of catalyst on microchannel walls. Target enzyme (here: d-amino acid oxidase) is fused to the positively charged mini-protein Zbasic2 and the channel surface contains a negatively charged γ-Al2O3 wash-coat layer. Non-covalent wall attachment of the chimeric Zbasic2_oxidase resulted in fully reversible enzyme immobilization with fairly uniform surface coverage and near complete retention of biological activity. The falling film at different gas and liquid flow rates as well as reactor inclination angles was shown to be mostly wavy laminar. The calculated film thickness was in the range 0.5-1.3×10-4m. Direct O2 concentration measurements at the channel surface demonstrated that the liquid side mass transfer coefficient (KL) for O2 governed the overall gas/liquid/solid mass transfer and that the O2 transfer rate (≥0.75mM·s-1) vastly exceeded the maximum enzymatic reaction rate in a wide range of conditions. A value of 7.5 (±0.5)s-1 was determined for the overall mass transfer coefficient KLa, comprising a KL of about 7×10-5m·s-1 and a specific surface area of up to 105m-1.

KW - Falling-film microreactor

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KW - Immobilized oxidase

KW - Microchannel

KW - Oxygen mass transfer

KW - Z-binding module

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JO - Biotechnology and Bioengineering

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SN - 0006-3592

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