Research ArticleBIOCHEMISTRY

Defect-induced activity enhancement of enzyme-encapsulated metal-organic frameworks revealed in microfluidic gradient mixing synthesis

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Science Advances  29 Jan 2020:
Vol. 6, no. 5, eaax5785
DOI: 10.1126/sciadv.aax5785
  • Fig. 1 Schematics of microfluidic laminar flow synthesis and bulk solution synthesis of enzyme-MOF composites.

    (A) Synthesis of enzyme-MOF composites in a microfluidic laminar flow. Zinc nitrate, 2-MeIM, and enzymes were introduced into the three inlets; enzyme-MOF composites were formed in the microchannel via diffusion-based gradient mixing. (B) Synthesis of enzyme-MOF composites in bulk solution. The reactants were added into the beaker and mixed through stirring.

  • Fig. 2 Microfluidic flow synthesis of Cyt c–MOF composites with different mixing schemes.

    (A to C) Bright-field images of as-synthesized Cyt c–MOF composites produced with different mixing schemes (marked on the images) on microfluidic chip. Scale bars, 0.5 mm. (D) Scanning electron microscope (SEM) image of Cyt c–MOF composites prepared with mixing scheme 1. Scale bar, 500 nm. (E) Transmission electron microscope (TEM) and energy-dispersive spectroscopy (EDS) elemental mapping images of Cyt c–MOF composites prepared with mixing scheme 1. Scale bar, 500 nm. (F) SEM image of the enzyme-MOF composites from bulk solution synthesis presented regular dodecahedron morphology, the same as pure MOFs. Scale bar, 500 nm. (G) Thermal gravimetric analysis (TGA) curves of pure MOFs and the Cyt c–MOF composites in air. (H) X-ray powder diffraction (XRD) patterns of pure MOFs and the Cyt c–MOF composites obtained by bulk solution synthesis and microfluidic synthesis using scheme 1.

  • Fig. 3 Investigation on the gradient mixing for the synthesis of enzyme-MOF composites in the microchannel.

    (A) Molar ratio between 2-MeIM and Zn2+ at the positions where the enzyme has a maximum probability of diffusion into the growing MOF crystals alongside the microchannel in three mixing schemes. The area in orange color indicates the common range of concentration ratios in bulk solution synthesis of enzyme-MOF composites. (B) Simulation results (shown as heat map) of the molar ratio between 2-MeIM and Zn2+ alongside the microchannel where all three reactants meet up. The dashed lines alongside the microchannel reflect the position where the enzyme has a maximum probability of diffusion into the growing MOF crystals. (C) Relative activities of native glucose oxidase (GOx) and GOx-MOFs synthesized in microchannel with mixing scheme 1 and GOx-MOFs synthesized in bulk solution with ratios of 2-MeIM to zinc nitrate ranging from 50 to 100, respectively.

  • Fig. 4 Structural feature and catalytic performance of microfluidic flow synthesized enzyme-MOF composites.

    (A) Fourier-transformed (FT)–EXAFS spectra of GOx-MOF composites synthesized by bulk solution synthesis (red) and microfluidic flow synthesis (blue). a.u., arbitrary units. (B) Pore size distributions calculated using density functional theory (DFT) method. GOx-MOF composites were prepared through microfluidic flow synthesis using scheme 1 (drawn on the left main ordinate axis, marked in blue). As controls, bulk solution–synthesized GOx-MOF composites were prepared with ratios between 2-MeIM and zinc nitrate ranging from 40 to 100 (drawn on the right minor ordinate axis, marked in red). (C and D) Relative activity of free enzymes, microfluidic flow–synthesized enzyme-MOF composites, and bulk solution–synthesized enzyme-MOF composites. All the enzymatic assays were carried out at the same protein concentration, normalized using the activity of free enzyme in solution as 100%. (E) Relative enzyme activity (versus time) of HRP-adsorbed MOFs, free HRP, microfluidic-synthesized HRP-MOFs, and bulk solution–synthesized HRP-MOF composites after trypsin digestion. (F) Thermal stability of free native HRP and HRP-MOF composites at 60° and 70°C, respectively.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/6/5/eaax5785/DC1

    Section S1. Materials

    Section S2. Fabrication of PDMS-based microfluidic chips

    Section S3. Theory and numerical simulation

    Section S4. Microfluidic flow synthesis of enzyme-MOF composites

    Section S5. Bulk solution synthesis of enzyme-MOF composites

    Section S6. Zeta-sizer analysis of Cyt c–MOF composites

    Section S7. TEM and EDS analysis

    Section S8. Powder XRD of enzyme-MOFs and pattern analysis

    Section S9. SEM analysis of pure MOFs and enzyme-MOF composites

    Section S10. Measurements of the loading amount of enzymes in MOFs

    Section S11. Determination of the activity of enzyme-MOF composites

    Section S12. XAFS spectra of bulk solution and microfluidic flow–synthesized ZIF-8 crystal

    Section S13. Bulk solution synthesis of enzyme-MOF composites of varied reactants ratio and characterization results in contrast with microfluidic flow synthesis

    Section S14. Investigation of the effect of flow rate on activity of enzyme-MOF composites

    Section S15. Spatially controlled synthesis of Au NP–MOF composites

    Section S16. Stability of enzyme-MOF composites at high temperature and in the presence of protease digestion

    Section S17. Recyclability and long-term stability of enzyme-MOFs

    Section S18. The influence of flow rate on defect properties

    Section S19. Numerical calculation of substrate diffusion in enzyme-MOFs

    Fig. S1. Pictures of the microfluidic chip for synthesis of enzyme-MOF composites.

    Fig. S2. Numerical simulation of velocity distribution in the microchannel.

    Fig. S3. Numerical simulation results of concentration distribution of reactants in different positions in the microchannel.

    Fig. S4. Three on-chip mixing schemes and corresponding simulation results of laminar flow diffusive mixing for the synthesis of enzyme-MOF composites.

    Fig. S5. Simulation results of laminar flow diffusive mixing for the synthesis of enzyme-MOF composites using mixing order scheme 1 with tuned flow rates.

    Fig. S6. The real image of the setup for microfluidic flow synthesis of enzyme-MOF composites.

    Fig. S7. Zetasizer analysis of Cyt c–MOF composites prepared by microfluidic synthesis with scheme 1.

    Fig. S8. TEM and EDS mapping images of Cyt c–MOF composites prepared on microfluidic chips with different mixing schemes.

    Fig. S9. XRD patterns of pure MOFs and the Cyt c–MOF composites obtained using scheme 1 on microfluidic chip with the indexation of peaks.

    Fig. S10. SEM images of pure MOFs and enzyme-MOF composites.

    Fig. S11. TGA curves of GOx- and HRP-MOF composites in air.

    Fig. S12. Relative enzyme activity of Cyt c–MOF composites prepared by microfluidic synthesis with different mixing schemes.

    Fig. S13. Relative enzyme activity of different enzymes before and after lyophilization process.

    Fig. S14. Comparison of catalytic activity between different constituents of enzyme-MOF composites and their combination.

    Fig. S15. EXAFS spectra and feff fit curve of bulk solution–synthesized and microchannel-synthesized MOFs.

    Fig. S16. N2 adsorption-desorption isotherm of the MOF particles prepared by microfluidic synthesis (scheme 1) and bulk solution synthesis with different metal/ligand ratios.

    Fig. S17. Investigation of the effect of flow rate on activity of HRP-MOF composites.

    Fig. S18. HAADF-STEM images of Au NP–MOF composites prepared by bulk solution synthesis and microfluidic flow synthesis.

    Fig. S19. The long-term stability of microfluidic and bulk solution synthesized HRP-MOF composites.

    Fig. S20. The reusability of HRP-adsorbed MOFs and microfluidic- and bulk solution–synthesized HRP-MOF composites.

    Fig. S21. Pore size distributions of MOF particles calculated using DFT method.

    Fig. S22. Schematic figure of the size of substrate and pore size of enzyme-MOF composites prepared by microfluidic synthesis and bulk solution synthesis.

    Fig. S23. Numerical simulation result of substrate diffusion into enzyme-MOF composites.

    Table S1. Literature data of diffusion coefficient.

    Table S2. Data obtained from fitting XAFS spectra to pure ZIF-8 crystal feff path.

    Table S3. Data of ICP-OES and elemental analysis of MOFs prepared by bulk solution synthesis and microfluidic synthesis (scheme 1).

    Table S4. Pore volume data of MOFs synthesized by bulk solution synthesis and microfluidic synthesis (scheme 1).

    References (4958)

  • Supplementary Materials

    This PDF file includes:

    • Section S1. Materials
    • Section S2. Fabrication of PDMS-based microfluidic chips
    • Section S3. Theory and numerical simulation
    • Section S4. Microfluidic flow synthesis of enzyme-MOF composites
    • Section S5. Bulk solution synthesis of enzyme-MOF composites
    • Section S6. Zeta-sizer analysis of Cyt c–MOF composites
    • Section S7. TEM and EDS analysis
    • Section S8. Powder XRD of enzyme-MOFs and pattern analysis
    • Section S9. SEM analysis of pure MOFs and enzyme-MOF composites
    • Section S10. Measurements of the loading amount of enzymes in MOFs
    • Section S11. Determination of the activity of enzyme-MOF composites
    • Section S12. XAFS spectra of bulk solution and microfluidic flow–synthesized ZIF-8 crystal
    • Section S13. Bulk solution synthesis of enzyme-MOF composites of varied reactants ratio and characterization results in contrast with microfluidic flow synthesis
    • Section S14. Investigation of the effect of flow rate on activity of enzyme-MOF composites
    • Section S15. Spatially controlled synthesis of Au NP–MOF composites
    • Section S16. Stability of enzyme-MOF composites at high temperature and in the presence of protease digestion
    • Section S17. Recyclability and long-term stability of enzyme-MOFs
    • Section S18. The influence of flow rate on defect properties
    • Section S19. Numerical calculation of substrate diffusion in enzyme-MOFs
    • Fig. S1. Pictures of the microfluidic chip for synthesis of enzyme-MOF composites.
    • Fig. S2. Numerical simulation of velocity distribution in the microchannel.
    • Fig. S3. Numerical simulation results of concentration distribution of reactants in different positions in the microchannel.
    • Fig. S4. Three on-chip mixing schemes and corresponding simulation results of laminar flow diffusive mixing for the synthesis of enzyme-MOF composites.
    • Fig. S5. Simulation results of laminar flow diffusive mixing for the synthesis of enzyme-MOF composites using mixing order scheme 1 with tuned flow rates.
    • Fig. S6. The real image of the setup for microfluidic flow synthesis of enzyme-MOF composites.
    • Fig. S7. Zetasizer analysis of Cyt c–MOF composites prepared by microfluidic synthesis with scheme 1.
    • Fig. S8. TEM and EDS mapping images of Cyt c–MOF composites prepared on microfluidic chips with different mixing schemes.
    • Fig. S9. XRD patterns of pure MOFs and the Cyt c–MOF composites obtained using scheme 1 on microfluidic chip with the indexation of peaks.
    • Fig. S10. SEM images of pure MOFs and enzyme-MOF composites.
    • Fig. S11. TGA curves of GOx- and HRP-MOF composites in air.
    • Fig. S12. Relative enzyme activity of Cyt c–MOF composites prepared by microfluidic synthesis with different mixing schemes.
    • Fig. S13. Relative enzyme activity of different enzymes before and after lyophilization process.
    • Fig. S14. Comparison of catalytic activity between different constituents of enzyme-MOF composites and their combination.
    • Fig. S15. EXAFS spectra and feff fit curve of bulk solution–synthesized and microchannel-synthesized MOFs.
    • Fig. S16. N2 adsorption-desorption isotherm of the MOF particles prepared by microfluidic synthesis (scheme 1) and bulk solution synthesis with different metal/ligand ratios.
    • Fig. S17. Investigation of the effect of flow rate on activity of HRP-MOF composites.
    • Fig. S18. HAADF-STEM images of Au NP–MOF composites prepared by bulk solution synthesis and microfluidic flow synthesis.
    • Fig. S19. The long-term stability of microfluidic and bulk solution synthesized HRP-MOF composites.
    • Fig. S20. The reusability of HRP-adsorbed MOFs and microfluidic- and bulk solution–synthesized HRP-MOF composites.
    • Fig. S21. Pore size distributions of MOF particles calculated using DFT method.
    • Fig. S22. Schematic figure of the size of substrate and pore size of enzyme-MOF composites prepared by microfluidic synthesis and bulk solution synthesis.
    • Fig. S23. Numerical simulation result of substrate diffusion into enzyme-MOF composites.
    • Table S1. Literature data of diffusion coefficient.
    • Table S2. Data obtained from fitting XAFS spectra to pure ZIF-8 crystal feff path.
    • Table S3. Data of ICP-OES and elemental analysis of MOFs prepared by bulk solution synthesis and microfluidic synthesis (scheme 1).
    • Table S4. Pore volume data of MOFs synthesized by bulk solution synthesis and microfluidic synthesis (scheme 1).
    • References (4958)

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