Alkali metal cation doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD (P) H and nicotinamide artificial coenzyme, and preparation method and application thereof

Abstract

The invention discloses an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD (P) H and nicotinamide artificial coenzymes and a preparation method and application thereof.The preparation method comprises the steps that firstly, a carbon-nitrogen precursor and alkali metal chloride are ground in a mortar, deionized water is added for mixing after grinding, the evenly-mixed material is transferred to a porcelain boat, a ceramic cover is covered, and the carbon nitride microsphere catalyst is obtained; the preparation method comprises the following steps: putting the raw materials into a tubular furnace, slowly heating and roasting in an N2 atmosphere, after roasting is finished, grinding the obtained solid product into powder, washing with ionized water for multiple times until the pH value of a mixed solution is 7-8, then centrifuging the mixture to obtain a solid, adding the solid into a beaker, then adding deionized water for ultrasonic treatment, and finally centrifuging the material after ultrasonic treatment is finished, thereby obtaining the high-purity titanium dioxide. And drying the obtained solid to obtain the carbon-nitrogen catalyst to be prepared, and the prepared carbon-nitrogen catalyst has a good utilization effect on visible light which is dominant in a solar spectrum, and has good NAD (P) H and nicotinamide artificial coenzyme regeneration performance.

Description

Technical Field

[0001] The present invention belongs to the technical field of material preparation, and particularly relates to an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, and a preparation method and application thereof. Background Art

[0002] Nicotinamide cofactors (NAD(P) / NAD(P)H), as a general proton / electron carrier, play an important role in biological oxidation-reduction reactions in vivo and in vitro. For example, NAD(P) + / NAD(P)H is a regulator that controls cell metabolism in vivo. At the same time, reactions catalyzed by redox enzymes in vitro usually require redox cofactors such as NAD(P)H to provide the supply of electrons and protons. These cofactors are expensive and need to be continuously regenerated to have practical significance. Nicotinamide cofactor-dependent redox enzymes are used in a wide range of industrial and biotechnological applications. In such enzyme catalysis, nicotinamide cofactors play an important role in electron and energy transfer. However, these cofactors show poor stability to extreme conditions such as high temperature, organic solvents, and acidic / alkaline pH, which limits the potential of redox enzymes in industrial applications. Therefore, the development of artificial cofactors and redox enzymes favorable for artificial cofactors becomes particularly important, which is beneficial to the development of in vitro enzyme synthesis systems in terms of availability and cost. Therefore, aiming at the characteristics that traditional enzyme-catalyzed NAD(P)H regeneration can only target natural coenzymes and utilize chemical energy, the development of catalysts that expand from natural coenzymes to artificial coenzymes and catalytic systems driven by renewable energy rather than chemical energy has practical application significance.

[0003] Therefore, the present invention proposes an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, and a preparation method and application thereof. Summary of the Invention

[0004] In view of the problems existing in the prior art, the purpose of the present invention is to provide an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, and a preparation method and application thereof, which have the characteristics of good catalytic effect, simple preparation method, wide raw material source, and safety and environmental protection.

[0005] To achieve the above purpose, the technical solution of the present invention is as follows: The present invention proposes a preparation method for an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, comprising the following steps: 1) Grind the carbon-nitrogen precursor and the corresponding alkali metal chloride in a mortar, add deionized water for mixing after grinding, and set aside after mixing evenly; 2) Transfer the uniformly mixed materials in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and slowly heat it for calcination under a N2 atmosphere. After the calcination is completed, grind the obtained solid product into powder for standby; 3) Wash the powder obtained in step 2) with ionized water multiple times to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7-8. Then, centrifuge the mixture to obtain a solid; 4) Add the solid obtained in step 3) to a beaker, then add deionized water for ultrasonic treatment. After the ultrasonic treatment is completed, finally centrifuge the materials, dry and encapsulate the obtained solid, which is the alkali metal cation-doped carbon nitride microsphere catalyst to be prepared.

[0006] Furthermore, in step 1), the carbon-nitrogen precursor is melamine, dicyandiamide or melamine, preferably melamine; the alkali metal chloride is lithium chloride, sodium chloride, potassium chloride, rubidium chloride or cesium chloride, preferably cesium chloride.

[0007] Furthermore, in step 1), the volume ratio of deionized water to the mass of the carbon-nitrogen precursor and the mass of the metal chloride is 1:1-5:0.5-4, preferably 1:3:2. The volume unit is mL and the mass unit is g.

[0008] Furthermore, in step 2), the heating rate is 1.5-2.5 °C / h, preferably 2.2 °C / h; the calcination temperature is 450-600 °C, preferably 550 °C; the calcination time is 3-5 h, preferably 4 h.

[0009] Furthermore, in step 4), the ultrasonic time is 0-36 h, preferably 24 h, the drying temperature is 60-110 °C, preferably 70 °C, and the drying time is 10-26 h, preferably 24 h.

[0010] The present invention provides an alkali metal cation-doped carbon nitride microsphere catalyst prepared by the above method.

[0011] The present invention also provides an application of the alkali metal cation-doped carbon nitride microsphere catalyst in photocatalytic regeneration of cofactors.

[0012] Furthermore, it includes the following steps: First, add the carbon nitride catalyst and triethanolamine to the sodium phosphate buffer solution for ultrasonic dispersion treatment, and then add and the oxidized coenzyme to obtain a mixed solution. Then, transfer the mixed solution to a photocatalytic reaction kettle, stir it under sealed conditions, and introduce nitrogen. React under the irradiation of a xenon lamp light source to carry out the photocatalytic regeneration of the coenzyme.

[0013] Further, the mass ratio of the carbon nitride catalyst to the volume of the sodium phosphate buffer solution is 1:0.5 - 2, preferably 1:1, where the mass unit is g and the volume unit is mL; the volume ratio of triethanolamine to the sodium phosphate buffer solution is 0.1 - 0.5:1, preferably 0.15:1, and the volume unit is mL; The molar ratio to the oxidized coenzyme is 0.2:3.

[0014] Further, the nitrogen gas flow rate is 1 - 30 mL / min, preferably 10 mL / min; the xenon lamp wavelength is 300 - 800 nm, preferably in the visible light band with a wavelength of 420 - 800 nm.

[0015] Principle of the present invention: In the present invention, through the modification of the carbon nitride material with alkali metal cations of different radii, the doping ions embedded inside the modified carbonized carbon material and a large number of cyano and hydroxyl functional groups formed on the surface achieve the effect of adjusting the intrinsic energy level structure of the material, enabling the prepared carbon nitride catalyst to have good utilization of visible light that dominates the solar spectrum.

[0016] Compared with the prior art, the beneficial effects of the present invention are as follows: 1) Firstly, in the present invention, the carbon nitride precursor and the corresponding alkali metal chloride are ground in a mortar, then deionized water is added and mixed. After mixing evenly, it is set aside. The evenly mixed material is transferred to a porcelain boat, covered with a ceramic lid, and placed in a tubular furnace. It is slowly heated and calcined in an N2 atmosphere. After the calcination is completed, the obtained solid product is ground into powder and set aside. The obtained powder is washed multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Subsequently, the mixture is centrifuged to obtain a solid. The obtained solid is added to a beaker, and then deionized water is added for ultrasonic treatment. After the ultrasonic treatment is completed, finally, the material is centrifuged, and the obtained solid is dried and encapsulated, which is the carbon nitride catalyst to be prepared. The prepared carbon nitride catalyst has good utilization of visible light that dominates the solar spectrum, thus having excellent NAD(P)H and nicotinamide-based artificial coenzyme regeneration performance; 2) The present invention has the characteristics of simple method, wide source of raw materials, the prepared catalyst does not contain precious metals, good catalytic effect, and safety and environmental protection. Description of the drawings

[0017] Figure 1 It is the SEM diagram of the carbon nitride microsphere catalyst prepared in Examples 1 - 12 of the present invention; Figure 2 It is the FT-IR diagram of the carbon nitride microsphere catalyst prepared in Examples 1 - 8 of the present invention; Figure 3 It is the Zeta-potential diagram of the carbon nitride microsphere catalyst prepared in Examples 1 - 12 of the present invention; Figure 4 Performance comparison chart of the regenerated NADH of the carbon nitride microsphere catalysts prepared in Examples 1-8 of the present invention; Figure 5 Performance comparison chart of the regenerated NADH of the carbon nitride microsphere catalysts prepared in Examples 9-12 of the present invention; Figure 6 Performance comparison chart of the catalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes in Examples I-1 to I-6 of the present invention; Figure 7 Time-reduced coenzyme production comparison chart of Example I-7 of the present invention; Figure 8 Time-reduced coenzyme production comparison chart of Example I-8 of the present invention. Detailed implementation mode

[0018] The present invention will be further described below in conjunction with the accompanying drawings and specific embodiments, but the scope protected by the present invention is not limited to the described scope.

[0019] Example 1

[0020] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for the photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, comprising the following steps: 1) Weigh 12 g of cesium chloride and 3 g of melamine, place them in a mortar and grind them. After grinding, add 2 mL of deionized water and mix. After mixing evenly, set aside; 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N2 atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder and set aside; 3) Wash the powder obtained in step 2) multiple times with ionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7-8, and then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat it for 24 h. After completion, centrifuge to obtain a precipitate, and finally place the solid precipitate in a 70 °C vacuum oven and dry it for 24 h. The final alkali metal cation-doped carbon nitride microsphere catalyst 1 is obtained.

[0021] Example 2

[0022] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for the photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, comprising the following steps: 1) Weigh 12 g of rubidium chloride and 3 g of melamine, place them in a mortar and grind. After grinding, add 2 mL of deionized water and mix. After mixing evenly, set aside. 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h. After the calcination is completed, grind the obtained solid product into powder and set aside. 3) Wash the powder obtained in step 2) several times with ionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Subsequently, centrifuge the mixture to obtain a solid. 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to collect the precipitate, and finally place the solid precipitate in a 70 °C vacuum oven and dry for 24 h. Obtain the final alkali metal cation-doped carbon nitride microsphere catalyst 2.

[0023] Example 3

[0024] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, comprising the following steps: 1) Weigh 12 g of potassium chloride and 3 g of melamine, place them in a mortar and grind. After grinding, add 2 mL of deionized water and mix. After mixing evenly, set aside. 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h. After the calcination is completed, grind the obtained solid product into powder and set aside. 3) Wash the powder obtained in step 2) several times with ionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Subsequently, centrifuge the mixture to obtain a solid. 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to collect the precipitate, and finally place the solid precipitate in a 70 °C vacuum oven and dry for 24 h. Obtain the final alkali metal cation-doped carbon nitride microsphere catalyst 3.

[0025] Example 4

[0026] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, comprising the following steps: 1) Weigh 12 g of sodium chloride and 3 g of melamine, place them in a mortar and grind. After grinding, add 2 mL of deionized water and mix well. Set aside after mixing evenly; 2) Transfer the material ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, put it into a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder and set aside; 3) Wash the powder obtained in step 2) multiple times with ionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to collect the precipitate, and finally place the solid precipitate in a 70 °C vacuum oven and dry for 24 h. Obtain the final alkali metal cation-doped carbon nitride microsphere catalyst 4.

[0027] Example 5

[0028] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, comprising the following steps: 1) Weigh 12 g of lithium chloride and 3 g of melamine, place them in a mortar and grind. After grinding, add 2 mL of deionized water and mix well. Set aside after mixing evenly; 2) Transfer the material ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, put it into a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder and set aside; 3) Wash the powder obtained in step 2) multiple times with ionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to collect the precipitate, and finally place the solid precipitate in a 70 °C vacuum oven and dry for 24 h. Obtain the final alkali metal cation-doped carbon nitride microsphere catalyst 5.

[0029] Example 6

[0030] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, comprising the following steps: 1) Weigh 3 g of melamine, place it in a mortar and grind. After grinding, add 2 mL of deionized water and mix well. Set aside after mixing evenly; 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder for standby; 3) Wash the powder obtained in step 2) multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8, and then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to obtain the precipitate, and finally place the solid precipitate in a vacuum oven at 70 °C and dry for 24 h. The final alkali metal cation-doped carbon nitride microsphere catalyst 6 is obtained.

[0031] Example 7

[0032] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, comprising the following steps: 1) Weigh 12 g of cesium chloride and 3 g of monocyanamide, place them in a mortar and grind, add 2 mL of deionized water after grinding, mix evenly and set aside; 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder for standby; 3) Wash the powder obtained in step 2) multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8, and then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to obtain the precipitate, and finally place the solid precipitate in a vacuum oven at 70 °C and dry for 24 h. The final alkali metal cation-doped carbon nitride microsphere catalyst 7 is obtained.

[0033] Example 8

[0034] A preparation method of a cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, comprising the following steps: 1) Weigh 12 g of cesium chloride and 3 g of dicyandiamide, place them in a mortar and grind, add 2 mL of deionized water after grinding, mix evenly and set aside; 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder for standby; 3) Wash the powder obtained in step 2) multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8, and then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to take the precipitate, and finally place the solid precipitate in a vacuum oven at 70 °C and dry for 24 h. The final alkali metal cation-doped carbon nitride microsphere catalyst 8 is obtained.

[0035] Example 9

[0036] A preparation method of a cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, comprising the following steps: 1) Weigh 12 g of cesium chloride and 3 g of melamine, place them in a mortar and grind, add 4 mL of deionized water after grinding, mix evenly and set aside; 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder for standby; 3) Wash the powder obtained in step 2) multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8, and then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 24 h. After completion, centrifuge to take the precipitate, and finally place the solid precipitate in a vacuum oven at 70 °C and dry for 24 h. The final alkali metal cation-doped carbon nitride microsphere catalyst 9 is obtained.

[0037] Example 10

[0038] 1) Weigh 12 g of cesium chloride and 3 g of melamine, place them in a mortar and grind, add 2 mL of deionized water after grinding, mix evenly and set aside; 2) Transfer the materials ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and under a N₂ atmosphere, heat it to 550 °C at a heating rate of 2.2 °C / h and calcine for 4 h; after the calcination is completed, grind the obtained solid product into powder for standby; 3) Wash the powder obtained in step 2) several times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Then centrifuge the mixture to obtain a solid. 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 0 h. After completion, centrifuge to collect the precipitate. Finally, place the solid precipitate in a vacuum oven at 100 °C and dry for 24 h. Obtain the final alkali metal cation-doped carbon nitride microsphere catalyst 10.

[0039] Example 11

[0040] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, comprising the following steps: 1) Weigh 12 g of cesium chloride and 3 g of melamine, place them in a mortar and grind. After grinding, add 2 mL of deionized water and mix. Set aside after mixing evenly. 2) Transfer the material ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, put it into a tube furnace, and heat it to 550 °C at a heating rate of 2.2 °C / h under a N2 atmosphere and calcine for 4 h. After the calcination is completed, grind the obtained solid product into powder for later use. 3) Wash the powder obtained in step 2) several times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7 - 8. Then centrifuge the mixture to obtain a solid. 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat for 12 h. After completion, centrifuge to collect the precipitate. Finally, place the solid precipitate in a vacuum oven at 100 °C and dry for 24 h. Obtain the final alkali metal cation-doped carbon nitride microsphere catalyst 11.

[0041] Example 12

[0042] A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes, comprising the following steps: 1) Weigh 12 g of cesium chloride and 3 g of melamine, place them in a mortar and grind. After grinding, add 2 mL of deionized water and mix. Set aside after mixing evenly. 2) Transfer the material ground and mixed evenly in step 1) to a porcelain boat, cover it with a ceramic lid, put it into a tube furnace, and heat it to 550 °C at a heating rate of 2.2 °C / h under a N2 atmosphere and calcine for 4 h. After the calcination is completed, grind the obtained solid product into powder for later use. 3) Wash the powder obtained in step 2) multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7-8, and then centrifuge the mixture to obtain a solid; 4) Transfer the solid obtained in step 3) to a beaker, add 200 mL of deionized water and ultrasonically treat it for 36 h. After completion, centrifuge to collect the precipitate, and finally place the solid precipitate in a vacuum oven at 100 °C and dry it for 24 h. The final alkali metal cation-doped carbon nitride microsphere catalyst 12 is obtained.

[0043] Test the above-prepared alkali metal cation-doped carbon nitride catalyst. The test method is as follows: First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 60 mg of NAD + (3 mM) to obtain a mixed solution. Then transfer the mixed solution to a photocatalytic reactor, stir it under sealed conditions, and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat it in a 37 °C water bath. Then use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light with a wavelength greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of the coenzyme.

[0044] Take the reaction solution every 5 min to detect the concentration of NADH. Take 1 mL of the centrifuged supernatant, then dilute it with 2 mL of phosphate buffer solution, and detect the absorbance at a wavelength of 340 nm. Compare it with the standard curve to obtain the relationship between the concentration of NADH and the absorbance.

[0045] The calculation formula is: .

[0046] From Figure 1 the scanning electron microscope (SEM) images, it can be seen that the morphological structures of the alkali metal cation-doped carbon nitride catalysts 1 - alkali metal cation-doped carbon nitride 12 all show irregular particles and flakes, and the alkali metal cations with different radii result in different degrees of microsphere morphology of the catalysts. The addition of water during the grinding process of the catalyst and the influence of the ultrasonic time both have an impact on whether the catalyst produces microspheres.

[0047] Furthermore, from Figure 2 the Fourier transform infrared (FT-IR) spectrogram, it can be seen that the graphitic carbon nitride material modified with a larger ionic radius is the key factor for the alkali metal cation-doped carbon nitride microsphere catalyst to have a high coenzyme reproduction yield. Carbon nitride catalyst 6 is the original unmodified graphitic carbon nitride, without a cyano peak (2178 cm-1 ), and hydroxyl peaks (3200 cm -1 -3600 cm -1 , 1150 cm -1 , 1000 cm -1 ). There is a basic heptazine ring structure (1200 cm -1 -1800 cm -1 ). As the radius of the doped alkali metal cations increases, a large number of cyano groups (2178 cm -1 ) and hydroxyl peaks (3200 cm -1 -3600 cm -1 , 1150 cm -1 , 1000 cm -‍1 ) appear, indicating the introduction of a large number of these two types of functional group structures. The heptazine ring structure is consistent with that of the graphitic carbon nitride material, indicating that the heptazine ring structure is not damaged. As the ultrasonic time prolongs, the amounts of cyano and hydroxyl groups also gradually increase. After ultrasonic treatment for more than 24 h, the excessive content of cyano and hydroxyl groups leads to the destruction of the overall carbon-nitrogen structure, reducing the visible light response and the ability to regenerate coenzymes. Therefore, ultrasonic treatment for 24 h with the alkali metal cation-doped carbon nitride microsphere catalyst 1 modified by Cs ions with the largest ionic radius is the best.

[0048] It can be seen from the zeta-potential diagram ( Figure 3 ) that due to the incorporation of alkali metal cations into the carbon nitride matrix, when the ions are dispersed in the solution, the surface cations tend to dissolve in water, forming colloidal particles with a negative zeta-potential. The negative potential in the solution is related to the radius of the doped alkali metal cations. The larger the ionic radius, the more negative the zeta-potential, which can quickly adsorb the reactants onto the catalyst surface and is more conducive to the progress of the coenzyme regeneration process. The alkali metal cation-doped carbon nitride microsphere catalyst 1 modified by cesium chloride with the largest ionic radius is the best, with the most negative zeta-potential at -18.0 mV, showing the best NADH regeneration performance.

[0049] The experiment on the photocatalytic regeneration of NADH by the alkali metal cation-doped carbon nitride microsphere catalyst ( Figure 4 ) shows that as the radius of the doped alkali metal cations increases, the concentration of NADH regeneration increases significantly, and the carbon-nitrogen catalyst 1 has the highest NADH yield. As the ultrasonic time prolongs, the concentration of NADH regeneration increases accordingly. After ultrasonic treatment for more than 24 h, the NADH concentration will decrease again. Therefore, ultrasonic treatment for 24 h is the best.

[0050] From the comparison chart of the NADH yields of Figure 4 and Figure 5 , it can be seen that the catalysts prepared from the precursors of monocyanamide and dicyandiamide can both regenerate NADH, but the effect is not as good as that of melamine (comparisonFigure 4 The carbonitrogen catalyst 1). Alkali metal chlorides can be lithium chloride, sodium chloride, potassium chloride and rubidium chloride, all of which have the effect of regenerating NADH, but their effects are not as high as that of cesium chloride. When preparing the catalyst, too high or too low water consumption, ultrasonic treatment time, washing times, drying temperature, and drying time will all reduce the concentration of catalytic NADH. The optimal yield is obtained using the preferred conditions.

[0051] Compare the application conditions of the catalyst in Example 1 Example I-1 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 60 mg of NADP + (3 mM) to obtain a mixed solution. Then transfer the mixed solution to a photocatalytic reactor, stir under sealed conditions, and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat in a 37 °C water bath. Then use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light with a wavelength greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADPH.

[0052] Example I-2 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 26 mg of nicotinamide-based artificial coenzyme NAD + -m1 (3 mM) to obtain a mixed solution. Then transfer the mixed solution to a photocatalytic reactor, stir under sealed conditions, and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat in a 37 °C water bath. Then use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light with a wavelength greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH-m1.

[0053] Example I-3 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 27 mg of nicotinamide-based artificial coenzyme NAD +-m2 (3 mM) to obtain a mixed solution. Then, transfer the mixed solution to a photocatalytic reactor, stir it under sealed conditions, and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat it in a 37 °C water bath. Then, use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light with a wavelength greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH-m2.

[0054] Example I-4 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 29 mg of nicotinamide-based artificial coenzyme NAD + -m3 (3 mM) to obtain a mixed solution. Then, transfer the mixed solution to a photocatalytic reactor, stir it under sealed conditions, and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat it in a 37 °C water bath. Then, use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light with a wavelength greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH-m3.

[0055] Example I-5 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 30 mg of nicotinamide-based artificial coenzyme NAD + -m4 (3 mM) to obtain a mixed solution. Then, transfer the mixed solution to a photocatalytic reactor, stir it under sealed conditions, and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat it in a 37 °C water bath. Then, use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light with a wavelength greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH-m4.

[0056] Example I-6 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add 2.6 mg (0.2 mM) and 32 mg of nicotinamide-based artificial coenzyme NAD +-m5 (3 mM) to obtain a mixed solution, and then transfer the mixed solution to a photocatalytic reactor. Stir under sealed conditions and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat in a 45 °C water bath. Then use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH-m5.

[0057] Example I-7 The reaction is carried out at 37 °C. The reaction system consists of glucose (100 mM), NAD(P) + or other nicotinamide-based artificial coenzymes (3 mM), glucose dehydrogenase (GDH), and 30 ml of sodium phosphate buffer. Transfer the mixed solution to a 500 mL photocatalytic reactor, add a magnetic stir bar for magnetic stirring, seal, and bubble with nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat in a 37 °C water bath. Measure the absorbance value with a UV-visible spectrophotometer.

[0058] Example I-8 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to sodium phosphate buffer, and then add 2.6 mg (0.2 mM) and 60 mg of nicotinamide-based artificial coenzyme NAD + (3 mM) to obtain a mixed solution. Then transfer the mixed solution to a photocatalytic reactor. Stir under sealed conditions and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat in a 25 °C water bath. Then use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH.

[0059] Example I-9 First, add 30 mg of carbon nitride catalyst and 4.5 mL of triethanolamine to sodium phosphate buffer, and then add 2.6 mg (0.2 mM) and 60 mg, NAD + (3 mM) to obtain a mixed solution. Then transfer the mixed solution to a photocatalytic reactor. Stir under sealed conditions and introduce nitrogen at a rate of 10 mL / min. Pre-inflate for 30 min and heat in a 45 °C water bath. Then use a 300 w xenon light source, take a visible light filter with a cut-off wavelength of 420 nm, control the light wavelength output by the xenon light source to visible light greater than 420 nm, and vertically irradiate the photocatalytic reactor to carry out the photocatalytic regeneration reaction of NADH.

[0060] It can be seen from Figure 6 Examples (I-1) - (I-6) that NADPH and other nicotinamide-based artificial coenzymes can all be photocatalytically regenerated, and the regeneration concentration is comparable to that of NADH. As substitutes for natural cofactors, synthetic nicotinamide cofactor analogs are inexpensive, have good chemical stability, and even improve the biocatalytic efficiency. These synthetic artificial coenzymes can operate more complex reactions with various other redox enzymes. Photocatalytic regeneration of artificial coenzymes will promote the use of redox biocatalysts in chemical production at low cost, thus more widely changing the use of redox enzymes in industrial biocatalysis.

[0061] From Figure 7 it can be seen from Example I-7 that when GDH is used as the regenerating enzyme, the regeneration efficiency of NAD + and NADP + is slightly higher than the photocatalytic regeneration efficiency. However, for nicotinamide-based artificial coenzymes, the regeneration efficiency of GDH can be ignored. In contrast, the photocatalytic method can regenerate all artificial coenzymes with a relatively high efficiency, demonstrating the advantages of this catalytic system in practical applications.

[0062] From Figure 8 it can be seen from Examples I-8 and I-9 that increasing or decreasing the temperature will reduce the regeneration concentration of NADH (compared with Figure 4 Catalyst 1 in), so 37 °C is selected as the reaction temperature.

Claims

1. A preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes, characterized in that It includes the following steps: 1) Grind the carbonitrogen precursor and the corresponding alkali metal chloride in a mortar. After grinding, add deionized water and mix. After mixing evenly, set aside for later use; 2) Transfer the evenly mixed material in step 1) to a porcelain boat, cover it with a ceramic lid, place it in a tube furnace, and slowly heat it for roasting in a N2 atmosphere. After the roasting is completed, grind the obtained solid product into powder for later use; 3) Wash the powder obtained in step 2) multiple times with deionized water to wash away the residual water-soluble chloride salts until the pH value of the mixed solution is 7-8. Then, centrifuge the mixture to obtain a solid; 4) Add the solid obtained in step 3) to a beaker, then add deionized water for ultrasonic treatment. After the ultrasonic treatment is completed, finally centrifuge the material, dry the obtained solid and encapsulate it, which is the alkali metal cation-doped carbon nitride microsphere catalyst to be prepared.

2. The preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes according to claim 1, characterized in that In step 1), the carbonitrogen precursor is monocyanamide, dicyandiamide or melamine; the alkali metal chloride is lithium chloride, sodium chloride, potassium chloride, rubidium chloride or cesium chloride.

3. The preparation method of an alkali metal cation-doped graphitic carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide-based artificial coenzymes according to claim 1, characterized in that In step 1), the volume ratio of deionized water to the mass of the carbonitrogen precursor and the mass of the metal chloride is 1:1 - 5:0.5 - 4, where the volume unit is mL and the mass unit is g.

4. The preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes according to claim 1, characterized in that In step 2), the heating rate is 1.5 - 2.5 °C / h; the roasting temperature is 450 - 600 °C; the roasting time is 3 - 5 h.

5. The preparation method of an alkali metal cation-doped carbon nitride microsphere catalyst for photocatalytic regeneration of NAD(P)H and nicotinamide artificial coenzymes according to claim 1, characterized in that In step 4), the ultrasonic time is 0 - 36 h, the drying temperature is 60 - 110 °C, and the drying time is 10 - 26 h.

6. An alkali metal cation-doped carbon nitride microsphere catalyst prepared by the method according to any one of claims 1 - 5.

7. Application of the alkali metal cation-doped carbon nitride microsphere catalyst according to claim 6 in photocatalytic regeneration of cofactors.

8. The application according to claim 7, wherein It includes the following steps: First, add the carbon nitride catalyst and triethanolamine to the sodium phosphate buffer solution, perform ultrasonic dispersion treatment, and then add and oxidized coenzyme to obtain a mixed solution. Then, transfer the mixed solution to a photocatalytic reactor, stir under sealed conditions, and introduce nitrogen. React under the irradiation of a xenon lamp source to carry out the photocatalytic regeneration of the coenzyme.

9. The application according to claim 8, wherein The ratio of the mass of the carbon nitride catalyst to the volume of the sodium phosphate buffer solution is 1:0.5 - 2, where the mass unit is g and the volume unit is mL; the volume ratio of triethanolamine to the sodium phosphate buffer solution is 0.1 - 0.5:1, where the volume unit is mL; The molar ratio to the oxidized coenzyme is 0.2:

3.

10. The application according to claim 8, characterized in that The nitrogen gas flow rate is 1 - 30 mL / min; the wavelength of the xenon lamp is 300 - 800 nm.

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