A photosensitizer for photochemical regeneration of coenzyme nadh

Abstract

The application relates to preparation and application of a photochemical coenzyme NADH photosensitizer, and belongs to the field of biological chemical industry.The method comprises the following steps: a rhodium compound [Cp*RhCl2]2 is combined with meso-tetra(4-carboxyphenyl)porphyrin TCPP into a single molecule porphyrin photosensitizer through an amide bond and a coordination bond; the functional groups include a carboxyl group and an amino group. The photosensitizer has simple synthesis steps, mild reaction conditions, and the coenzyme NADH regeneration efficiency can reach 33.4% under light irradiation for 10 min, and has wide application prospects in the aspect of biological catalytic conversion of CO2 to produce methanol.

Description

Technical Field

[0001] This invention relates to the preparation and application of a photochemical regeneration coenzyme NADH photosensitizer, belonging to the field of biochemical engineering. Background Technology

[0002] Biocatalytic conversion of CO2 to methanol is an environmentally friendly and clean energy production process that plays a crucial role in the high-value utilization of CO2. In the biocatalytic conversion of CO2, the catalytic activity of dehydrogenase systems requires the participation of coenzymes NADH or NADPH. These coenzymes act as proton / electron carriers, binding to the dehydrogenases and directly participating in the reaction. However, the high cost and large quantities of coenzymes are detrimental to the industrial-scale production of methanol from CO2. Therefore, it is necessary to efficiently and economically regenerate and recycle these coenzymes.

[0003] Among various coenzyme regeneration technologies, photochemical regeneration of NADH / NADPH borrows the concept of photosynthesis from nature. It utilizes clean and inexpensive sunlight to induce electrons to be excited from the photosensitizer, and these excited electrons migrate from the photosensitizer to the electron medium, thus achieving coenzyme regeneration. Photochemical regeneration technology has the advantages of simple construction and green reaction processes, effectively reducing the industrial production cost of biocatalytic CO2 conversion and improving the level of the circular economy. The key to photochemical regeneration of NADH / NADPH lies in the development of highly efficient visible light-absorbing photosensitizers. The photosensitizer acts as an energy carrier and a bridge in the electron transport chain during photoexcited electron transfer; its conversion efficiency of light energy (such as conversion number and quantum efficiency) and the electron transfer efficiency with the electron medium are decisive factors for the efficient regeneration of NADH / NADPH. Currently, photosensitizers used in photochemical regeneration of NADH / NADPH include inorganic semiconductors, xanthracene organic dyes, porphyrins, and organic-inorganic hybrid materials. Among them, porphyrin photosensitizers are increasingly used in the regeneration of photochemical coenzymes NADH / NADPH due to their good absorption in the visible light region, excellent electron transport properties, and tunable photo / electric properties. In a photochemical coenzyme NADH regeneration experiment using tetra(4-carboxyphenyl)porphyrin (TCPP) as the photosensitizer and triethanolamine (TEOA) as the electron donor, the NADH yield reached 81.5% after 1 hour of illumination (Catal. Sci. Technol., 2018, 8, 2578-2587). However, the regeneration efficiency of most porphyrin photosensitizers used in current research for photochemical regeneration of NADH / NADPH is still limited by stability issues and the occurrence of side reactions (such as the generation of superoxide and singlet oxygen molecules), and its regeneration efficiency needs further improvement. Improving the photoexcited electron transport efficiency through the proximity effect can prevent side reactions and thus improve the overall regeneration efficiency. Using zinc porphyrin molecules, which have higher electronegativity, as photosensitizers resulted in higher NADH regeneration efficiency compared to metal-free porphyrin molecules (Chem. Commun., 2011, 47, 10227-10229). This is because the zinc porphyrin photosensitizer and the nitrogen atom in the electron donor TEOA are axially coupled, increasing the proximity effect between the photosensitizer and the electron donor. This not only accelerates the electron transfer efficiency between the electron donor and the photosensitizer but also promptly consumes the ground-state holes on the photosensitizer, preventing the backflow of excited electrons and reducing the occurrence of side reactions.

[0004] Previous research on the photochemical regeneration of coenzyme NADH by porphyrins has laid a solid theoretical foundation for further construction of highly efficient photosensitizers and for guiding the design and optimization of photosensitizer structures and properties. Based on this, this invention develops a single-molecule porphyrin photosensitizer for photochemically regenerating coenzyme NADH by coupling TCPP with a rhodium compound [Cp*RhCl2]2 via amide bonding. The proximity effect between the photosensitizer and the electron medium accelerates electron transfer efficiency, thereby improving the regeneration efficiency of coenzyme NADH. The photosensitizer developed in this invention has a simple synthesis procedure, mild reaction conditions, and high coenzyme NADH regeneration efficiency, showing broad application prospects in the biocatalytic conversion of CO2 to methanol. Summary of the Invention

[0005] In view of the problems existing in the prior art, the purpose of the present invention is to provide a method for preparing a photosensitizer that can be used for photochemical regeneration of coenzyme NADH.

[0006] To achieve this objective, the present invention adopts the following technical solution:

[0007] This invention provides a method for preparing a photosensitizer that can be used for photochemical regeneration of the coenzyme NADH. The method includes: bonding a rhodium compound [Cp*RhCl2]2 with TCPP via an amide bond to form a monomolecular porphyrin photosensitizer. The synthetic route is as follows:

[0008]

[0009] The photosensitizer preparation method for photochemically regenerated coenzyme NADH provided by this invention is characterized by simple synthesis steps and mild reaction conditions. The specific steps are as follows:

[0010] (1) TCPP and 2,2'-bipyridine-3-amine (BPA) were mixed in DMSO, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl) and N-hydroxysuccinimide (NHS) were added. The mixture was stirred at room temperature in the dark for 24-48 h, dialyzed in DMSO for 24 h, and stored in a clean glass bottle for later use.

[0011] (2) The product obtained in step (1) was mixed and dissolved with the rhodium compound [Cp*RhCl2]2. The mixture was stirred and reacted for 2 hours at room temperature in the dark until the solid [Cp*RhCl2]2 was completely dissolved. After the reaction was completed, the unreacted impurities were removed by dialysis with DMSO. The crude product was separated by gel permeation chromatography to obtain the pure photochemical regenerated coenzyme NADH photosensitizer.

[0012] (3) The photosensitizer product obtained in step (2) was redispersed in DMSO, EDC·HCl and NHS were added, mixed evenly, and placed on the surface of amino-modified nonwoven fabric. The reaction was carried out in a shaker at room temperature in the dark for 24-48 hours. After the reaction was completed, the nonwoven fabric covalently modified with monomolecular porphyrin photosensitizer was rinsed three times with DMSO and ultrapure water to obtain the immobilized porphyrin photosensitizer with recyclable coenzyme NADH.

[0013] Preferably, the molar ratio of TCPP and BPA in step (1) is 1:3 or 1:4.

[0014] Preferably, the molar ratio of TCPP, EDC·HCl and NHS in step (1) is 1:3:3 or 1:4:4.

[0015] Preferably, the molar ratio of BPA and [Cp*RhCl2]2 in step (2) is 2:1.

[0016] Preferably, the molar ratio of TCPP, EDC·HCl and NHS in step (3) is 1:1:1.

[0017] In this invention, a synthesized monomolecular porphyrin photosensitizer is further covalently grafted onto amino-modified nonwoven fabric. The monomolecular porphyrin photosensitizer bonded by amide bonds exhibits better stability, while simultaneously solving the problems of poor solubility, low recycling rate, and high cost associated with porphyrin-based photosensitizers. Furthermore, due to the high surface area of ​​the nonwoven fabric, NAD+ can also be achieved in the photochemical regeneration of coenzyme NADH reaction system. + Effective retention and separation of NADH.

[0018] Using a quartz photocatalytic reactor equipped with a CEL-S500 simulated daylight xenon lamp light source system, the photosensitizer molecules of this invention can effectively regenerate coenzyme NADH. After 10 minutes of illumination, the yield of coenzyme NADH is as high as 33.4%, which has potential application value.

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] (1) The photosensitive agent preparation method for photochemical regeneration of coenzyme NADH provided by the present invention utilizes the load separation performance of nonwoven fabric, the electron transfer law during photochemical regeneration of coenzyme NADH and the proximity effect to accelerate the electron transfer between the photosensitizer and the electronic medium, prevent the backflow of excited electrons, thereby improving the regeneration efficiency of coenzyme NADH.

[0021] (2) The present invention uses a covalent grafting method to bond nonwoven fabric and monomolecular porphyrin photosensitizer with amide bonds. The resulting immobilized porphyrin can exist stably in the photochemical regeneration coenzyme NADH reaction system, providing a strong guarantee for the efficient recycling of porphyrin photosensitizer.

[0022] (3) The porphyrin photosensitizer provided by the present invention has a regeneration efficiency of up to 33.4% for coenzyme NADH within 10 min, which is superior to existing porphyrin photosensitizers.

[0023] (4) The photosensitizer synthesis process provided by the present invention has simple steps, good repeatability, and mild reaction conditions, and has broad application prospects in the biocatalytic conversion of CO2 to methanol. Attached Figure Description

[0024] Figure 1 In Example 1 of the invention, step (2) prepares the compound. 1 H NMR spectrum;

[0025] Figure 2 The yield of photochemical regeneration of coenzyme NADH by the porphyrin photosensitizer prepared in Example 1 of this invention. Detailed Implementation

[0026] The technical solution of the present invention will be further illustrated below through specific embodiments. Those skilled in the art should understand that the embodiments described are merely illustrative of the present invention and should not be construed as limiting the invention.

[0027] Example 1

[0028] Step (1) Preparation of the compound: Accurately weigh TCPP (Mw = 790.77382 g / mol, 2 mM) and BPA (Mw = 171.2 g / mol, 8 mM) and dissolve them in 5 mL of DMSO. Then add EDC·HCl (Mw = 191.7 g / mol, 8 mM) and NHS (Mw = 115.09 g / mol, 8 mM) sequentially. React at room temperature in the dark for 48 h. Place the completely reacted mixture in a semi-permeable membrane with a molecular weight cutoff of 2000 Da and dialyze it in DMSO solution for 24 h. Finally, transfer it to a clean glass bottle for storage.

[0029] Step (2) Synthesis of porphyrin photosensitizer: Accurately weigh [Cp*RhCl2]2 (Mw = 618.08 g / mol, 4 mM) and add it to the above reaction solution. Continue the reaction at room temperature in the dark for 2 h until the solid [Cp*RhCl2]2 is completely dissolved to form TCPP-Rh4 complex. Then, place the above reaction solution in a semi-permeable membrane with a molecular weight cutoff of 2000 Da and dialyze for 24 h to remove unreacted impurities. The crude product is purified by gel permeation chromatography with cross-linked polyvinyl acetate gel as column packing and DMSO as eluent to obtain purple solid S1.

[0030] Example 2

[0031] Step (1) Preparation of the compound: Accurately weigh TCPP (Mw = 790.77382 g / mol, 2 mM) and BPA (Mw = 171.2 g / mol, 6 mM) and dissolve them in 5 mL of DMSO. Then add EDC·HCl (Mw = 191.7 g / mol, 6 mM) and NHS (Mw = 115.09 g / mol, 6 mM) sequentially. React at room temperature in the dark for 48 h. Place the completely reacted mixture in a semi-permeable membrane with a molecular weight cutoff of 1000 Da and dialyze it in DMSO solution for 24 h. Finally, transfer it to a clean glass bottle for storage.

[0032] Step (2) Synthesis of porphyrin photosensitizer: Accurately weigh [Cp*RhCl2]2 (Mw = 618.08 g / mol, 3 mM) and add it to the above reaction solution. Continue the reaction at room temperature in the dark for 2 h until the solid [Cp*RhCl2]2 is completely dissolved to form TCPP-Rh3 complex. Then, place the above reaction solution in a semi-permeable membrane with a molecular weight cutoff of 2000 Da and dialyze for 24 h to remove unreacted impurities.

[0033] Step (3) Preparation of immobilized porphyrin photosensitizer: EDC·HCl (Mw = 191.7 g / mol, 2 mM) and NHS (Mw = 115.09 g / mol, 2 mM) were added sequentially to the TCPP-Rh3 photosensitizer complex solution obtained in step (2), and placed on the surface of amino-modified nonwoven fabric. The reaction was carried out in a shaker for 24 h under room temperature and light-proof conditions. After the reaction was completed, the nonwoven fabric covalently modified with monomolecular porphyrin photosensitizer was washed three times with DMSO and ultrapure water, respectively, to obtain immobilized porphyrin photosensitizer S2 with recyclable coenzyme NADH.

[0034] Example 3

[0035] At room temperature, 40 mL (100 mM, pH 7.0) of buffer solution was first degassed with Ar and saturated for 1 h. Then, a solution containing 5.00 mM TEOA, 0.50 mM TCPP-Rh4, and 1.00 mM NAD was added. + The degassing buffer solution was placed in a quartz photocatalytic reactor (equipped with a CEL-S500 simulated daylight xenon lamp light source system). Ar was continuously introduced into the reaction solution, and 300 μL of the reaction solution was taken at regular intervals to measure the OD using a multifunctional microplate reader. 340 Calculate the amount of NADH generated.

[0036] After 10 minutes of light exposure, the efficiency of the TCPP-Rh4 complex in regenerating coenzyme NADH was as follows: Figure 2As shown, the TCPP-Rh4 complex photosensitizer exhibits significantly higher efficiency in photochemically regenerating coenzyme NADH than the control group TCPP. After 10 minutes of illumination, the regeneration efficiency of coenzyme NADH reaches 33.4%, which is approximately 2.06 times higher than that of the control group. This indicates that the photosensitizer provided by this invention has the function of highly efficient photochemical regeneration of coenzyme NADH and has broad application prospects in the industrial application of biocatalytic conversion of CO2 to methanol.

[0037] The applicant declares that the above description is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Those skilled in the art should understand that any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention fall within the protection and disclosure scope of the present invention.

Claims

1. A method for preparing a photosensitizer that can be used for photochemical regeneration of coenzyme NADH, the synthetic route of which is as follows: 。 2. The photosensitizer preparation method for photochemical regeneration of coenzyme NADH according to claim 1, wherein the synthesis steps are as follows: (1) TCPP and 2,2'-bipyridine-3-amine (BPA) were mixed in DMSO, and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl) and N-hydroxysuccinimide (NHS) were added. The mixture was stirred at room temperature in the dark for 24-48 h, dialyzed in DMSO for 24 h, and stored in a clean glass bottle for later use. (2) The product obtained in step (1) was mixed and dissolved with the rhodium compound [Cp*RhCl2]2. The mixture was stirred and reacted at room temperature in the dark for 2 h until the solid [Cp*RhCl2]2 was completely dissolved. After the reaction was completed, the unreacted impurities were removed by dialysis with DMSO. The crude product was separated by gel permeation chromatography to obtain the pure photochemical regenerated coenzyme NADH photosensitizer. (3) The photosensitizer product obtained in step (2) is redispersed in DMSO, EDC·HCl and NHS are added, mixed evenly, and placed on the surface of amino-modified nonwoven fabric. The reaction is carried out in a shaker at room temperature in the dark for 24~48 h. After the reaction is completed, the nonwoven fabric covalently modified with monomolecular porphyrin photosensitizer is washed three times with DMSO and ultrapure water respectively to obtain the immobilized porphyrin photosensitizer with recyclable coenzyme NADH.

3. The method for preparing photosensitizer for photochemical regeneration of coenzyme NADH according to claim 2, characterized in that: The molar ratio of TCPP and BPA in step (1) is 1:3 or 1:

4.

4. The method for preparing photosensitizer for photochemical regeneration of coenzyme NADH according to claim 2, characterized in that: The molar ratio of TCPP, EDC·HCl and NHS in step (1) is 1:3:3 or 1:4:

4.

5. The method for preparing photosensitizer for photochemical regeneration of coenzyme NADH according to claim 2, characterized in that: The molar ratio of BPA and [Cp*RhCl2]2 in step (2) is 2:

1.

6. The method for preparing photosensitizer for photochemical regeneration of coenzyme NADH according to claim 2, characterized in that: The molar ratio of TCPP, EDC·HCl and NHS in step (3) is 1:1:1.

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