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enzymatic system for sustainable conversion of CO2 to formate
Catalysis Communications 136 (2020) 105903
Contents lists available at ScienceDirect
Catalysis Communications journal homepage: www.elsevier.com/locate/catcom
Short communication
A coupled photocatalytic/enzymatic system for sustainable conversion of CO2 to formate
T
⁎
FengJuan Gua, YanZi Wanga, ZiHui Menga, WenFang Liua, , LiYuan Qiub a b
School of Chemistry and Chemical Engineering, Beijing Institute of Technology, Liangxiang Higher Education Park, Fangshan District, Beijing 102488, PR China Systems Engineering Research Institute, Haidian District, Beijing 100094, PR China
A R T I C LE I N FO
A B S T R A C T
Keywords: CO2 reduction Photo-enzymatic catalysis Formate Cofactor regeneration
Enzymatic conversion of CO2 into high-added chemicals is advantageous in mild reaction conditions and high specificity, for which enzyme immobilization and cofactor regeneration are often necessary. Although many efforts have been made, unfortunately, the turnover number (TN) of cofactor does not exceed 100 (~1 usually). Here, an integrated process of formate synthesis from CO2 catalyzed by hollow fiber membrane immobilized enzyme succeeded with a simple UV/TiO2 photocatalytic coenzyme regeneration was explored. By optimizing the operating conditions and NADH concentration, TN reached 125 after 4.5 h. This process can be extended to the production of methanol and other similar systems.
1. Introduction The increasing concerns on CO2 conversion into important feedstock in the chemical industries has arisen due to the negative environmental prospects of fossil fuels use [1]. As a potential alternative of traditional heterogeneous catalysis, enzymatic-catalyzed CO2 hydrogenation to produce methanol with the assistant of the reduced nicotinamide adenine dinucleotide (NADH) has received much attention for its high specificity as well as mild reaction conditions [2–4], in which the step from CO2 to formate is critical for the cascade reaction [5]. Nevertheless, the practical application of dehydrogenases is quite limited due to their high costs and the stoichiometric consumption of cofactors [6]. For which, both enzyme immobilization and coenzyme regeneration are necessary. Many efforts have been made for the immobilization of dehydrogenase(s) involved. They were encapsulated [7–10], absorbed [11,12] or covalently bonded [4,13–15] to different matrices, in which the sol-gel particles were often used. Although these strategies are effective in improving the stability or reactivity of the biocatalysts, most of them are unsuitable to continuous manipulation. In our previous work [16], a hollow fiber membrane (HFM) micro-reactor was developed for the conversion of CO2 to formate by combining hydrophobic polymer HFMs as gas distributors and the enzyme-bearing polyethylene (PE) HFMs in the same module, which was proved to enable higher catalytic efficiency than usual aeration mode. HFM-attached formate dehydrogenase (FDH) showed excellent reusability and storage stability
⁎
as well. For coenzyme regeneration, photochemical way is clean and sustainable, in which organic dyes are the most often used. Kim et al. [17] used eosin Y (EY) as a photosensitizer and cobaloxime as a co-catalyst to regenerate cofactor for the enzymatic conversion of CO2 to formate, and a yield of 28% was achieved after 4 h (TN of NADH was 0.28). Yadav et al. [18] employed chemically converted graphene coupled isatin-substituted porphyrin (CCG-IP) for cofactor regeneration in the production of methanol from CO2 in the presence of free enzymes, regrettably, the yield of methanol after 1 h was only 8.41%. Ji et al. [19] encapsulated enzymes into hollow nanofiber, with EY and [Cp⁎Rh(bpy) (H2O)]2+ for NADH regeneration, and the yield of methanol reached 90.6% after illumination for 10 h, corresponding a TN of 0.906. They also constructed 5, 10, 15, 20-tetrakis (4-carboxyphenyl) porphyrin (TCPP)/EY/Rh supramolecular assemblies to enhance the electron transfer between photosensitizer and Rh mediator, however, the yield of methanol was only 22.8%, corresponding to a TN value of 0.228 [20]. Besides a lower TN value (~1), organic dyes are prone to degradation as photoreaction proceeds [21]. Semiconductor materials usually have much higher thermal and chemical stability. Two kinds of delicate photocatalysts including WS2/ g-C3N4 [22] and BiFeO3 [23] have been used for cofactor regeneration in the enzymatic systhesis of methanol from CO2, and a higher TN of 16.7 was acquired after 10 h in the former. However, the biocatalysts are in a free state in both cases, which are labile and difficult to be recycled.
Corresponding author. E-mail address: [email protected] (W. Liu).
https://doi.org/10.1016/j.catcom.2019.105903 Received 26 September 2019; Received in revised form 1 November 2019; Accepted 8 December 2019 Available online 09 December 2019 1566-7367/ © 2019 Elsevier B.V. All rights reserved.
Catalysis Communications 136 (2020) 105903
F. Gu, et al.
Scheme 1. Schematic illustration of a coupled photocatalytic/enzymatic system for sustainable CO2 conversion. The reaction solution associated with TiO2 particles was circulated between FDH-attached HFM reactor and photocatalytic unit by the peristaltic pump.
2.3. Enzymatic reaction
Compared with organic dyes or doped/modified semiconductor, pristine TiO2 as a classic photocatalyst is much more stable and cheaper, and has excellent catalytic properties under ultraviolet light [24]. In present work, a high-efficiency and sustainable production of formate from CO2 was achieved by creatively coupling an enzymebearing HFM reactor with a simple UV/TiO2 photocatalytic coenzyme regeneration unit (see Scheme 1). The operating conditions and NADH concentration were optimized, and the effect of electron donor was investigated. At last, the photo-enzyme catalysis system was also compared with a single photocatalytic or enzymatic system.
For comparison with the photo-enzyme catalysis, a single enzymatic CO2 reduction reaction (see Eq. (1)) without cofactor regeneration was conducted in the same membrane module bearing FDH. Differently, no photocatalyst and relevant components were included and only NADH was dissolved in an EDTA-NaOH buffer solution (pH 7.0), which was circulated with the peristaltic pump, and a higher NADH concentration of 5 mM was used in order to obtain a detectable amount of product. FDH
CO2 + NADH + H+ ⇌ HCOOH + NAD+
(1)
2. Experimental
3. Results and discussion
2.1. Photo-enzyme catalysis
3.1. Bioactivity assay of regenerated NADH
The chemistry in the photo-enzymatic synthesis of formate from CO2 is shown as Scheme S1. The photocatalytic reaction was performed at ambient temperature in a quartz vessel with magnetic stirring. Typically, 30 mg TiO2 powder was added to 20 mL EDTA-NaOH buffer solution (pH 7.0), stirred for 10 min and then treated with ultrasound for 15 min. Next, NADH (0.01–2 mM) and [Cp*Rh(bpy)(H2O)]2+ (0.25 mM) were successively loaded into the suspension. The mixture was stirred and purged with nitrogen in the dark for 15 min before the system was exposed to two 23 W UV lamps with the major emission wavelength of 365 nm. As the enzymatic-catalysis reactor, the FDH-loaded HFM module (Fig. S1) was placed in a 37 °C water bath, and then connected in series with the peristaltic pump and the photocatalytic unit (see Scheme 1), thereby the reaction mixture containing TiO2 particles circulated between them once the pump was started up. CO2 was continuously fed into the HFM reactor at a speed of 3 mL min−1 adjusted with a mass flow meter (Alicat, 0–20 SCCM, with the accuracy of ± 0.5%).
NADH regeneration is vital for a continuous enzymatic production of formate from CO2 due to its stoichiometric consumption. However, in the process of regeneration, biologically inactive 1,6-NADH and the dimers of NAD+ may be formed under the action of the intermediate radical [25]. Therefore, [Cp*Rh(bpy)(H2O)]2+ was often used as a cocatalyst for the selective production of biologically active 1,4-NADH, and its effectiveness has been validated in our previous work [21,24]. In spite of this, the bioactivity assay of regenerated NADH in specific application is still necessary. After being illuminated for 3 h, the regenerated NADH was added in the FDH-attached HFM reactor. The result was compared with the system using fresh NADH (Fig. S3). It can be seen that the reaction kinetic curves for the both systems nearly overlap each other, implying that the regenerated NADH has comparable bioactivity to fresh that and it is competent to assist the enzymatic synthesis of formate from CO2. On this basis, a photo-enzyme coupled system with in-situ cofactor regeneration was studied. 3.2. Photo-enzyme catalysis of CO2
2.2. Photocatalytic reaction
3.2.1. Effect of gas velocity Due to the high diffusivity of gas molecules, gas velocity generally has a relatively small effect on the overall mass transfer within the membrane reactor [26], while it will affect the conversion rate of CO2 [27]. To this end, with a certain liquid velocity (40 mL min−1), the effect of gas velocity on the synthesis of formate in the photo-enzyme catalysis system was studied in a range of 0.5–8 mL min−1 and the results are shown in Fig. 1. From Fig. 1, it can be found that the initial reaction rate and the yield of formate first increased and then decreased with gas velocity increased. The maximum initial reaction rate and yield were acquired
As control, a single photocatalytic reaction was performed at ambient temperature in a quartz vessel with magnetic stirring. The ingredients were the same as the photo-enzyme catalysis except for those involved in the enzymatic reaction. First, 7.5 mg TiO2 powder and 0.22 mL aqueous solution of [Cp*Rh(bpy)(H2O)]2+ (5 mM) was added to 5 mL EDTA-NaOH buffer solution (pH 7.0) to make a final concentration of 1.5 mg mL−1 and 0.25 mM. Then, the suspension was stirred for 10 min and treated with ultrasound for 15 min in the shield of light. During the reaction, CO2 was continuously fed into the reactor via a glass tube at a speed of 3 mL min−1. 2
Catalysis Communications 136 (2020) 105903
F. Gu, et al.
Fig. 1. Effect of gas velocity on (a) the production-time curve and (b) the initial reaction rate and yield. The liquid velocity was 40 mL min−1 and NADH concentration was 2 mM.
at a gas velocity of 3 mL min−1, which was 1.04 mM h−1 and 81.7%. Under this condition, the production of formate increased with the time within 4.5 h. When gas velocity was higher or lower, the production of formate first increased with the time and then the reaction reached the equilibrium after 3 h (Fig. 1a), at the same time, the initial reaction rate and the yield of formate decreased (Fig. 1b). It can be attributed to the reduced residence time in the membrane reactor and contact time between the substrate CO2 and enzyme molecules on the membrane surface in the former and less gas-liquid interfaces in the latter [26].
rate and the yield of formate increased first and then decreased, and the optimal liquid velocity was 40 mL min−1. With a lower liquid velocity, on one hand, TiO2 nanoparticles tend to deposit in the pipeline, so that the regeneration performance of the photocatalytic unit deteriorates; on the other hand, there is a stagnant liquid film on the surface of the enzymatic membrane, and the diffusion of CO2 through the liquid-membrane interface was a rate limiting step. Therefore, with the increase in liquid velocity, the mass transfer in the liquid phase gradually transforms into the reaction kinetics restriction mechanism [28].
3.2.2. Effect of liquid velocity During continuous operation, liquid velocity plays a crucial role in reducing the thickness of the boundary layer and promoting mass transfer between the interface and the body region [27]. Thus, the influence of liquid velocity on the coupling reaction was examined at a certain gas velocity (3 mL min−1) and the results are displayed in Fig. 2. As can be seen, with liquid velocity increased, the initial reaction
3.2.3. Effect of NADH concentration In the enzymatic reduction, the hydrogen protons need to be delivered to the active sites of oxidoreductase by coenzyme [29]. One mole of NADH is consumed to produce one mole of formate. In current work, NADH was regenerated when the reaction mixture was transported into the photocatalytic cell, thereby a constant supply was 3
Catalysis Communications 136 (2020) 105903
F. Gu, et al.
Fig. 2. Effect of liquid velocity on (a) the production-time curve and (b) the initial reaction rate and yield. The gas velocity was 3 mL min−1 and NADH concentration was 2 mM.
efficient and sustainable for CO2 conversion, as a result, formate can be rapidly produced even at a very low loading amount of NADH.
secured. Fig. 3a illustrates the effect of NADH concentration on the production of formate. At all concentrations investigated, the production of formate increased nearly linearly with the prolonged time. When NADH concentration was decreased, the initial reaction rate decreased, whereas, the yield of formate increased (Fig. 3b), indicating an upgraded utilization efficiency of cofactor. A comparison of TN and TOF values with research status was made (Table S1). In this work, when NADH concentration was 1 mM, TN and TOF was 1.32 and 0.293 h−1, respectively, higher than the system regenerated by glutamate dehydrogenase [30], lower than that using WS2/g-C3N4 [22]. When NADH concentration was 0.5 mM, TN and TOF was 3.08 and 0.684 h−1, respectively, obviously higher than most of the reported values [17,18,20,31], although free enzymes were used in these systems. When NADH concentration was 0.1 mM, TN and TOF was lower than the system of TCPP [32]. Furtherly, when NADH concentration was decreased to 0.05 or 0.01 mM, TN could be upgraded to 23.8 or 125, proving that the photo-enzyme integrated system is highly
3.2.4. Effect of electron donor EDTA was chosen as the electron donor in current work because of its stronger complexing ability with TiO2 and thus better properties for NADH regeneration than others such as TEOA and H2O [24]. However, the high oxidizing potential of TiO2 generates holes that can lead to EDTA break down into small molecules like CO2 [29]. In order to confirm that the source of formate was mainly from carbon dioxide rather than EDTA, different electron donors (TEOA/H2O) were investigated and the results are illustrated in Fig. S4. It can be found that the reaction rate in the system using TEOA was a little lower than that using EDTA, and the production of formate after 4.5 h in the former was 81.8% of the latter, revealing that a significant amount of formate can be produced when EDTA was replaced with TEOA. When no additional electron donor was used except for water, 4
Catalysis Communications 136 (2020) 105903
F. Gu, et al.
Fig. 3. Effect of NADH concentration on (a) the production-time curve and (b) the initial reaction rate and yield. The liquid velocity was 40 mL min−1 and gas velocity was 3 mL min−1.
photocatalytic system. In order to confirm the action of cofactor regeneration system, a single enzyme-catalyzed reaction without cofactor regeneration was conducted under the same conditions except for a higher NADH concentration. Wihout the integration of the photocatalytic unit, the initial reaction rate of the enzyme-catalyzed system was only 0.140 mM h−1 and the production of formate after 4.5 h was 0.535 mM, both of which were far lower than the photo-enzyme coupling system and higher than the photocatalytic system. It can be concluded that cofactor regeneration is vital for propelling the enzymatic reaction towards the conversion of CO2 and the combination of photocatalysis produces a strong synergistic effect on the enzymatic reaction. It is also noted that the reaction rate in the system without liquid circulation (Fresh NADH in Fig. S3) in HFM reactor was obvious lower than the same system with liquid circulation (Biocatalysis in Fig. S4). The initial reaction rate was 0.082 and 0.140 mM h−1, respectively. With liquid circulation, the reaction was accelerated by a factor of 1.71, revealing that this strategy is effective in intensifying the mass transfer
the kinetic curve of the photo-enzymatic coupled system was almost the same as that of single enzymatic conversion (Fig. S4). Considering that a higher NADH concentration (5 mM) was used in the latter than the former (2 mM), the above result proves that NADH is regenerated in the photo-enzymatic coupled system that strengthens the overall reaction, even with a weaker electron donor, and the source of formate is from carbon dioxide. 3.3. Comparison with other catalytic methods As well known, the semiconductor material also catalyzes the fixation of CO2 in the photocatalytic system. Therefore, a single photocatalytic reduction of CO2 was executed and the production of formate was detected as the control. As shown in Fig. S4, the production of formate increased slowly with the time and was only 0.184 mM after 4.5 h, and the initial reaction rate was 0.0513 mM h−1. For the photoenzyme catalysis system, these values were 1.634 mM and 1.04 mM h−1, respectively, which were 8.88 and 20.3 times of the 5
Catalysis Communications 136 (2020) 105903
F. Gu, et al.
in the liquid.
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4. Conclusions In conclusion, the integration of a simple UV/TiO2 photocatalytic coenzyme regeneration system with a FDH-bearing HFM reactor is successful for the high-efficient and sustainable production of formate from CO2, in which the initial reaction rate is separately 20.3 and 7.43 times of that in a single photocatalytic or enzyme-catalyzed system, and the production of formate after 4.5 h is separately 8.88 and 3.05 times. Special ventilation and liquid circulation mode greatly accelerate the reaction. With instant regeneration, a high production of formate can be obtained even at a very low loading amount of NADH. TN and TOF are obviously higher than most of the reported values, indicating an elevated utilization efficiency of cofactor. Continued efforts to apply these strategies in the production of methanol and other similar biotransformations may well prove important as the process intensification concept continues to develop. Declaration of Competing Interest The authors declare no conflict of interest. Acknowledgements This work is financially supported by the Beijing Municipal Natural Science Foundation (grant number 2172050). Appendix A. Supplementary data Supplementary data to this article can be found online at https:// doi.org/10.1016/j.catcom.2019.105903. References [1] Q. Liu, L. Wu, R. Jackstell, M. Beller, Nat. Commun. 6 (2015) 5933–5947, https:// doi.org/10.1038/ncomms6933. [2] R. Obert, B.C. Dave, J. Am. Chem. Soc. 121 (1999) 12192–12193, https://doi.org/ 10.1021/ja991899r. [3] S.W. Xu, Y. Lu, J. Li, Z.Y. Jiang, H. Wu, Ind. Eng. Chem. Res. 45 (2006) 4567–4573, https://doi.org/10.1021/ie051407l. [4] E.Z. Bilal, D. Dustin, P. Wang, Biotechnol. Bioeng. 99 (2008) 508–514, https://doi. org/10.1002/bit.21584. [5] W.F. Liu, Y.H. Hou, B.X. Hou, Z.P. Zhao, Chin. J. Chem. Eng. 22 (2014) 1328–1332, https://doi.org/10.1016/j.cjche.2014.09.026.
6
Contents lists available at ScienceDirect
Catalysis Communications journal homepage: www.elsevier.com/locate/catcom
Short communication
A coupled photocatalytic/enzymatic system for sustainable conversion of CO2 to formate
T
⁎
FengJuan Gua, YanZi Wanga, ZiHui Menga, WenFang Liua, , LiYuan Qiub a b
School of Chemistry and Chemical Engineering, Beijing Institute of Technology, Liangxiang Higher Education Park, Fangshan District, Beijing 102488, PR China Systems Engineering Research Institute, Haidian District, Beijing 100094, PR China
A R T I C LE I N FO
A B S T R A C T
Keywords: CO2 reduction Photo-enzymatic catalysis Formate Cofactor regeneration
Enzymatic conversion of CO2 into high-added chemicals is advantageous in mild reaction conditions and high specificity, for which enzyme immobilization and cofactor regeneration are often necessary. Although many efforts have been made, unfortunately, the turnover number (TN) of cofactor does not exceed 100 (~1 usually). Here, an integrated process of formate synthesis from CO2 catalyzed by hollow fiber membrane immobilized enzyme succeeded with a simple UV/TiO2 photocatalytic coenzyme regeneration was explored. By optimizing the operating conditions and NADH concentration, TN reached 125 after 4.5 h. This process can be extended to the production of methanol and other similar systems.
1. Introduction The increasing concerns on CO2 conversion into important feedstock in the chemical industries has arisen due to the negative environmental prospects of fossil fuels use [1]. As a potential alternative of traditional heterogeneous catalysis, enzymatic-catalyzed CO2 hydrogenation to produce methanol with the assistant of the reduced nicotinamide adenine dinucleotide (NADH) has received much attention for its high specificity as well as mild reaction conditions [2–4], in which the step from CO2 to formate is critical for the cascade reaction [5]. Nevertheless, the practical application of dehydrogenases is quite limited due to their high costs and the stoichiometric consumption of cofactors [6]. For which, both enzyme immobilization and coenzyme regeneration are necessary. Many efforts have been made for the immobilization of dehydrogenase(s) involved. They were encapsulated [7–10], absorbed [11,12] or covalently bonded [4,13–15] to different matrices, in which the sol-gel particles were often used. Although these strategies are effective in improving the stability or reactivity of the biocatalysts, most of them are unsuitable to continuous manipulation. In our previous work [16], a hollow fiber membrane (HFM) micro-reactor was developed for the conversion of CO2 to formate by combining hydrophobic polymer HFMs as gas distributors and the enzyme-bearing polyethylene (PE) HFMs in the same module, which was proved to enable higher catalytic efficiency than usual aeration mode. HFM-attached formate dehydrogenase (FDH) showed excellent reusability and storage stability
⁎
as well. For coenzyme regeneration, photochemical way is clean and sustainable, in which organic dyes are the most often used. Kim et al. [17] used eosin Y (EY) as a photosensitizer and cobaloxime as a co-catalyst to regenerate cofactor for the enzymatic conversion of CO2 to formate, and a yield of 28% was achieved after 4 h (TN of NADH was 0.28). Yadav et al. [18] employed chemically converted graphene coupled isatin-substituted porphyrin (CCG-IP) for cofactor regeneration in the production of methanol from CO2 in the presence of free enzymes, regrettably, the yield of methanol after 1 h was only 8.41%. Ji et al. [19] encapsulated enzymes into hollow nanofiber, with EY and [Cp⁎Rh(bpy) (H2O)]2+ for NADH regeneration, and the yield of methanol reached 90.6% after illumination for 10 h, corresponding a TN of 0.906. They also constructed 5, 10, 15, 20-tetrakis (4-carboxyphenyl) porphyrin (TCPP)/EY/Rh supramolecular assemblies to enhance the electron transfer between photosensitizer and Rh mediator, however, the yield of methanol was only 22.8%, corresponding to a TN value of 0.228 [20]. Besides a lower TN value (~1), organic dyes are prone to degradation as photoreaction proceeds [21]. Semiconductor materials usually have much higher thermal and chemical stability. Two kinds of delicate photocatalysts including WS2/ g-C3N4 [22] and BiFeO3 [23] have been used for cofactor regeneration in the enzymatic systhesis of methanol from CO2, and a higher TN of 16.7 was acquired after 10 h in the former. However, the biocatalysts are in a free state in both cases, which are labile and difficult to be recycled.
Corresponding author. E-mail address: [email protected] (W. Liu).
https://doi.org/10.1016/j.catcom.2019.105903 Received 26 September 2019; Received in revised form 1 November 2019; Accepted 8 December 2019 Available online 09 December 2019 1566-7367/ © 2019 Elsevier B.V. All rights reserved.
Catalysis Communications 136 (2020) 105903
F. Gu, et al.
Scheme 1. Schematic illustration of a coupled photocatalytic/enzymatic system for sustainable CO2 conversion. The reaction solution associated with TiO2 particles was circulated between FDH-attached HFM reactor and photocatalytic unit by the peristaltic pump.
2.3. Enzymatic reaction
Compared with organic dyes or doped/modified semiconductor, pristine TiO2 as a classic photocatalyst is much more stable and cheaper, and has excellent catalytic properties under ultraviolet light [24]. In present work, a high-efficiency and sustainable production of formate from CO2 was achieved by creatively coupling an enzymebearing HFM reactor with a simple UV/TiO2 photocatalytic coenzyme regeneration unit (see Scheme 1). The operating conditions and NADH concentration were optimized, and the effect of electron donor was investigated. At last, the photo-enzyme catalysis system was also compared with a single photocatalytic or enzymatic system.
For comparison with the photo-enzyme catalysis, a single enzymatic CO2 reduction reaction (see Eq. (1)) without cofactor regeneration was conducted in the same membrane module bearing FDH. Differently, no photocatalyst and relevant components were included and only NADH was dissolved in an EDTA-NaOH buffer solution (pH 7.0), which was circulated with the peristaltic pump, and a higher NADH concentration of 5 mM was used in order to obtain a detectable amount of product. FDH
CO2 + NADH + H+ ⇌ HCOOH + NAD+
(1)
2. Experimental
3. Results and discussion
2.1. Photo-enzyme catalysis
3.1. Bioactivity assay of regenerated NADH
The chemistry in the photo-enzymatic synthesis of formate from CO2 is shown as Scheme S1. The photocatalytic reaction was performed at ambient temperature in a quartz vessel with magnetic stirring. Typically, 30 mg TiO2 powder was added to 20 mL EDTA-NaOH buffer solution (pH 7.0), stirred for 10 min and then treated with ultrasound for 15 min. Next, NADH (0.01–2 mM) and [Cp*Rh(bpy)(H2O)]2+ (0.25 mM) were successively loaded into the suspension. The mixture was stirred and purged with nitrogen in the dark for 15 min before the system was exposed to two 23 W UV lamps with the major emission wavelength of 365 nm. As the enzymatic-catalysis reactor, the FDH-loaded HFM module (Fig. S1) was placed in a 37 °C water bath, and then connected in series with the peristaltic pump and the photocatalytic unit (see Scheme 1), thereby the reaction mixture containing TiO2 particles circulated between them once the pump was started up. CO2 was continuously fed into the HFM reactor at a speed of 3 mL min−1 adjusted with a mass flow meter (Alicat, 0–20 SCCM, with the accuracy of ± 0.5%).
NADH regeneration is vital for a continuous enzymatic production of formate from CO2 due to its stoichiometric consumption. However, in the process of regeneration, biologically inactive 1,6-NADH and the dimers of NAD+ may be formed under the action of the intermediate radical [25]. Therefore, [Cp*Rh(bpy)(H2O)]2+ was often used as a cocatalyst for the selective production of biologically active 1,4-NADH, and its effectiveness has been validated in our previous work [21,24]. In spite of this, the bioactivity assay of regenerated NADH in specific application is still necessary. After being illuminated for 3 h, the regenerated NADH was added in the FDH-attached HFM reactor. The result was compared with the system using fresh NADH (Fig. S3). It can be seen that the reaction kinetic curves for the both systems nearly overlap each other, implying that the regenerated NADH has comparable bioactivity to fresh that and it is competent to assist the enzymatic synthesis of formate from CO2. On this basis, a photo-enzyme coupled system with in-situ cofactor regeneration was studied. 3.2. Photo-enzyme catalysis of CO2
2.2. Photocatalytic reaction
3.2.1. Effect of gas velocity Due to the high diffusivity of gas molecules, gas velocity generally has a relatively small effect on the overall mass transfer within the membrane reactor [26], while it will affect the conversion rate of CO2 [27]. To this end, with a certain liquid velocity (40 mL min−1), the effect of gas velocity on the synthesis of formate in the photo-enzyme catalysis system was studied in a range of 0.5–8 mL min−1 and the results are shown in Fig. 1. From Fig. 1, it can be found that the initial reaction rate and the yield of formate first increased and then decreased with gas velocity increased. The maximum initial reaction rate and yield were acquired
As control, a single photocatalytic reaction was performed at ambient temperature in a quartz vessel with magnetic stirring. The ingredients were the same as the photo-enzyme catalysis except for those involved in the enzymatic reaction. First, 7.5 mg TiO2 powder and 0.22 mL aqueous solution of [Cp*Rh(bpy)(H2O)]2+ (5 mM) was added to 5 mL EDTA-NaOH buffer solution (pH 7.0) to make a final concentration of 1.5 mg mL−1 and 0.25 mM. Then, the suspension was stirred for 10 min and treated with ultrasound for 15 min in the shield of light. During the reaction, CO2 was continuously fed into the reactor via a glass tube at a speed of 3 mL min−1. 2
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Fig. 1. Effect of gas velocity on (a) the production-time curve and (b) the initial reaction rate and yield. The liquid velocity was 40 mL min−1 and NADH concentration was 2 mM.
at a gas velocity of 3 mL min−1, which was 1.04 mM h−1 and 81.7%. Under this condition, the production of formate increased with the time within 4.5 h. When gas velocity was higher or lower, the production of formate first increased with the time and then the reaction reached the equilibrium after 3 h (Fig. 1a), at the same time, the initial reaction rate and the yield of formate decreased (Fig. 1b). It can be attributed to the reduced residence time in the membrane reactor and contact time between the substrate CO2 and enzyme molecules on the membrane surface in the former and less gas-liquid interfaces in the latter [26].
rate and the yield of formate increased first and then decreased, and the optimal liquid velocity was 40 mL min−1. With a lower liquid velocity, on one hand, TiO2 nanoparticles tend to deposit in the pipeline, so that the regeneration performance of the photocatalytic unit deteriorates; on the other hand, there is a stagnant liquid film on the surface of the enzymatic membrane, and the diffusion of CO2 through the liquid-membrane interface was a rate limiting step. Therefore, with the increase in liquid velocity, the mass transfer in the liquid phase gradually transforms into the reaction kinetics restriction mechanism [28].
3.2.2. Effect of liquid velocity During continuous operation, liquid velocity plays a crucial role in reducing the thickness of the boundary layer and promoting mass transfer between the interface and the body region [27]. Thus, the influence of liquid velocity on the coupling reaction was examined at a certain gas velocity (3 mL min−1) and the results are displayed in Fig. 2. As can be seen, with liquid velocity increased, the initial reaction
3.2.3. Effect of NADH concentration In the enzymatic reduction, the hydrogen protons need to be delivered to the active sites of oxidoreductase by coenzyme [29]. One mole of NADH is consumed to produce one mole of formate. In current work, NADH was regenerated when the reaction mixture was transported into the photocatalytic cell, thereby a constant supply was 3
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Fig. 2. Effect of liquid velocity on (a) the production-time curve and (b) the initial reaction rate and yield. The gas velocity was 3 mL min−1 and NADH concentration was 2 mM.
efficient and sustainable for CO2 conversion, as a result, formate can be rapidly produced even at a very low loading amount of NADH.
secured. Fig. 3a illustrates the effect of NADH concentration on the production of formate. At all concentrations investigated, the production of formate increased nearly linearly with the prolonged time. When NADH concentration was decreased, the initial reaction rate decreased, whereas, the yield of formate increased (Fig. 3b), indicating an upgraded utilization efficiency of cofactor. A comparison of TN and TOF values with research status was made (Table S1). In this work, when NADH concentration was 1 mM, TN and TOF was 1.32 and 0.293 h−1, respectively, higher than the system regenerated by glutamate dehydrogenase [30], lower than that using WS2/g-C3N4 [22]. When NADH concentration was 0.5 mM, TN and TOF was 3.08 and 0.684 h−1, respectively, obviously higher than most of the reported values [17,18,20,31], although free enzymes were used in these systems. When NADH concentration was 0.1 mM, TN and TOF was lower than the system of TCPP [32]. Furtherly, when NADH concentration was decreased to 0.05 or 0.01 mM, TN could be upgraded to 23.8 or 125, proving that the photo-enzyme integrated system is highly
3.2.4. Effect of electron donor EDTA was chosen as the electron donor in current work because of its stronger complexing ability with TiO2 and thus better properties for NADH regeneration than others such as TEOA and H2O [24]. However, the high oxidizing potential of TiO2 generates holes that can lead to EDTA break down into small molecules like CO2 [29]. In order to confirm that the source of formate was mainly from carbon dioxide rather than EDTA, different electron donors (TEOA/H2O) were investigated and the results are illustrated in Fig. S4. It can be found that the reaction rate in the system using TEOA was a little lower than that using EDTA, and the production of formate after 4.5 h in the former was 81.8% of the latter, revealing that a significant amount of formate can be produced when EDTA was replaced with TEOA. When no additional electron donor was used except for water, 4
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Fig. 3. Effect of NADH concentration on (a) the production-time curve and (b) the initial reaction rate and yield. The liquid velocity was 40 mL min−1 and gas velocity was 3 mL min−1.
photocatalytic system. In order to confirm the action of cofactor regeneration system, a single enzyme-catalyzed reaction without cofactor regeneration was conducted under the same conditions except for a higher NADH concentration. Wihout the integration of the photocatalytic unit, the initial reaction rate of the enzyme-catalyzed system was only 0.140 mM h−1 and the production of formate after 4.5 h was 0.535 mM, both of which were far lower than the photo-enzyme coupling system and higher than the photocatalytic system. It can be concluded that cofactor regeneration is vital for propelling the enzymatic reaction towards the conversion of CO2 and the combination of photocatalysis produces a strong synergistic effect on the enzymatic reaction. It is also noted that the reaction rate in the system without liquid circulation (Fresh NADH in Fig. S3) in HFM reactor was obvious lower than the same system with liquid circulation (Biocatalysis in Fig. S4). The initial reaction rate was 0.082 and 0.140 mM h−1, respectively. With liquid circulation, the reaction was accelerated by a factor of 1.71, revealing that this strategy is effective in intensifying the mass transfer
the kinetic curve of the photo-enzymatic coupled system was almost the same as that of single enzymatic conversion (Fig. S4). Considering that a higher NADH concentration (5 mM) was used in the latter than the former (2 mM), the above result proves that NADH is regenerated in the photo-enzymatic coupled system that strengthens the overall reaction, even with a weaker electron donor, and the source of formate is from carbon dioxide. 3.3. Comparison with other catalytic methods As well known, the semiconductor material also catalyzes the fixation of CO2 in the photocatalytic system. Therefore, a single photocatalytic reduction of CO2 was executed and the production of formate was detected as the control. As shown in Fig. S4, the production of formate increased slowly with the time and was only 0.184 mM after 4.5 h, and the initial reaction rate was 0.0513 mM h−1. For the photoenzyme catalysis system, these values were 1.634 mM and 1.04 mM h−1, respectively, which were 8.88 and 20.3 times of the 5
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in the liquid.
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4. Conclusions In conclusion, the integration of a simple UV/TiO2 photocatalytic coenzyme regeneration system with a FDH-bearing HFM reactor is successful for the high-efficient and sustainable production of formate from CO2, in which the initial reaction rate is separately 20.3 and 7.43 times of that in a single photocatalytic or enzyme-catalyzed system, and the production of formate after 4.5 h is separately 8.88 and 3.05 times. Special ventilation and liquid circulation mode greatly accelerate the reaction. With instant regeneration, a high production of formate can be obtained even at a very low loading amount of NADH. TN and TOF are obviously higher than most of the reported values, indicating an elevated utilization efficiency of cofactor. Continued efforts to apply these strategies in the production of methanol and other similar biotransformations may well prove important as the process intensification concept continues to develop. Declaration of Competing Interest The authors declare no conflict of interest. Acknowledgements This work is financially supported by the Beijing Municipal Natural Science Foundation (grant number 2172050). Appendix A. Supplementary data Supplementary data to this article can be found online at https:// doi.org/10.1016/j.catcom.2019.105903. References [1] Q. Liu, L. Wu, R. Jackstell, M. Beller, Nat. Commun. 6 (2015) 5933–5947, https:// doi.org/10.1038/ncomms6933. [2] R. Obert, B.C. Dave, J. Am. Chem. Soc. 121 (1999) 12192–12193, https://doi.org/ 10.1021/ja991899r. [3] S.W. Xu, Y. Lu, J. Li, Z.Y. Jiang, H. Wu, Ind. Eng. Chem. Res. 45 (2006) 4567–4573, https://doi.org/10.1021/ie051407l. [4] E.Z. Bilal, D. Dustin, P. Wang, Biotechnol. Bioeng. 99 (2008) 508–514, https://doi. org/10.1002/bit.21584. [5] W.F. Liu, Y.H. Hou, B.X. Hou, Z.P. Zhao, Chin. J. Chem. Eng. 22 (2014) 1328–1332, https://doi.org/10.1016/j.cjche.2014.09.026.
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