Reduction method, method for producing mannitol, reduction device, and electrode

Abstract

The present disclosure provides a reduction method which is advantageous from the viewpoint of the efficiency of reduction of NAD+ or NADP+. A reduction method according to the present disclosure includes the following steps (Ia) and (Ib). (Ia) A liquid which contains at least one selected from the group consisting of NAD+ and NADP+ is brought into contact with an electrode (1). (Ib) NAD+ or NADP+ is reduced by directly donating electrons from the electrode (1). The electrode (1) has a surface which contains at least one element that is selected from the group consisting of platinum, palladium, and nickel.

Description

Reduction method, method for producing mannitol, reduction device, and electrode

[0001] The present disclosure relates to a reduction method, a method for producing mannitol, a reduction device, and an electrode.

[0002] Conventionally, oxidized nicotinamide adenine nucleotide (NAD + ) or oxidized nicotinamide adenine dinucleotide phosphate (NADP + As a method for electrochemically reducing HCl, a method using an electron mediator is known.

[0003] For example, Patent Document 1 describes a method for inactivating allergens, which includes a reduction step in which an allergen is reduced to a reduced redox protein by donating electrons from an electrode connected to an external power source outside the reaction system. In this reduction step, electrons are donated from the electrode to an electron mediator, and then from the electron mediator to a redox molecule such as NADPH or NADH. For example, a glassy carbon electrode is used as the electrode.

[0004] For example, Patent Document 2 describes a method for measuring fructose concentration. In this method, fructose is reduced by the action of D-mannitol dehydrogenase (DMDH), and the increase in cathodic current due to a reaction in which oxidized NADH is reduced to its original reduced form in conjunction with this reaction is utilized to measure the fructose concentration. The electrode is fabricated by immobilizing DMDH on a current collector made of carbon.

[0005] International Publication No. 2021 / 023754 Japanese Patent Publication No. 63-14301

[0006] The technology described in the above patent document is + or NADP + Therefore, the present disclosure provides a method for reducing NAD + or NADP + The present invention provides a reduction method that is advantageous in terms of the efficiency of reduction.

[0007] The present disclosure relates to oxidized nicotinamide adenine nucleotide (NAD +) and oxidized nicotinamide adenine dinucleotide phosphate (NADP + contacting a liquid containing at least one selected from the group consisting of platinum, palladium, and nickel with an electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel; and directly donating electrons from the electrode to form the NAD. + or the NADP + and reducing the

[0008] The reduction method of the present disclosure is + or NADP + This is advantageous in terms of the efficiency of the reduction.

[0009] FIG. 1 is a diagram schematically showing an example of a reduction device according to an embodiment. FIG. 2 is a diagram schematically showing an example of a reduction method according to an embodiment. FIG. 3 is a diagram schematically showing another example of a reduction method according to an embodiment. FIG. 4 is a block diagram showing an example of the functional configuration of a reduction device according to an embodiment. FIG. 5 is a flowchart showing an example of a reduction method according to an embodiment. FIG. 6 is a graph showing the yield of NADH in Example 1 and Comparative Example 1. FIG. 7 is a graph showing the amount of fructose reduction in Example 2 and Comparative Example 4.

[0010] (Findings that form the basis of the present disclosure) In this specification, NAD + and NADP + In summary, NAD(P) + Reduced nicotinamide adenine nucleotide (NADH) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) are collectively referred to as NAD(P)H.

[0011] NAD(P) + The chemical reaction between NAD(P)H and NAD(P)H is expressed by the following formula (1):

[0012] NAD(P) + +H + +2e -→ NAD(P)H Formula (1) The reaction represented by the above formula (1) proceeds as a two-step reaction as shown in the following formulas (2) and (3). In formulas (2) and (3), NAD(P) ・ indicates the radical of NAD(P).

[0013] 1st stage: NAD (P) + +e - → NAD (P) ・ Formula (2) Second stage: NAD(P) ・ +e - +H + → NAD(P)H Equation (3) According to the above equation (1), NAD(P) + If electrons are donated to NAD(P), + On the other hand, the second step of the reaction shown in formula (3) requires a very large voltage. ・ As shown in formula (4), the dimerization reaction is likely to occur rapidly, and the second step reaction shown in formula (3) may not proceed and the reaction may terminate.

[0014] Dimerization reaction: NAD(P) ・ +NAD(P) ・ → NAD(P)2 Formula (4) As described in Patent Document 1, NAD(P) is converted to NAD(P) using an electron mediator. + When reducing NAD(P) + The number of molecules involved in the reduction of NAD(P) increases, making it difficult to increase the efficiency of electron transfer. + As a result of extensive research aimed at solving this problem, it was found that the efficiency of the reduction of NAD(P) is low when an electrode having a surface containing platinum is used. + We have newly discovered that NAD(P)H can be generated by donating electrons directly from an electrode to the ATP. By using such an electrode, NAD(P)H can be generated more efficiently than, for example, a carbon electrode. + It has been newly discovered that the voltage required for the reduction of NAD(P) can be lowered, and the dimerization reaction represented by the above formula (4) is easily suppressed. In addition, it is possible to reduce the voltage required for the reduction of NAD(P) from the electrode without the intervention of an electron mediator. +By donating electrons directly to NAD(P), the efficiency of electron transfer increases. + It has been newly discovered that the reduction efficiency of the present disclosure is likely to be high. Based on this new finding, the reduction method of the present disclosure has been devised.

[0015] Therefore, according to the present disclosure, NAD + or NADP + This provides an advantageous reduction method from the viewpoint of reduction efficiency.

[0016] (Embodiments of the Present Disclosure) Hereinafter, embodiments of the present disclosure will be described in detail with reference to the drawings. Note that the embodiments described below are all comprehensive or specific examples. The numerical values, shapes, materials, components, component arrangement and connection configurations, steps, and step orders shown in the following embodiments are merely examples and are not intended to limit the scope of the claims. Furthermore, among the components in the following embodiments, components that are not recited in the independent claims that represent the highest concepts are described as optional components. Furthermore, each figure is not necessarily an exact illustration. In each figure, substantially identical components are assigned the same reference numerals, and redundant explanations may be omitted or simplified. Furthermore, in this disclosure, terms indicating the relationship between elements, such as parallel and perpendicular, terms indicating the shape of elements, such as rectangular, and numerical values ​​do not only represent the strict meaning, but also include a substantially equivalent range, for example, a difference of about a few percent.

[0017] (Embodiment) Hereinafter, an embodiment will be specifically described with reference to FIGS. 1 to 5. FIG.

[0018] FIG. 1 is a diagram showing a schematic diagram of an example of a reduction device according to an embodiment. As shown in FIG. 1, the reduction device 100 includes an electrode 1, a storage chamber 4a, and a power source 20. The electrode 1 has a surface containing at least one material selected from the group consisting of platinum, palladium, and nickel. The storage chamber 4a contains NAD + and NADP +The power supply 20 is electrically connected to the electrode 1. In the reduction device 100, the NAD contained in the liquid 9a is reduced by directly donating electrons from the electrode 1. + or NADP + will be reduced.

[0019] 2 is a diagram schematically illustrating an example of a reduction method according to an embodiment. As shown in FIG. 2, the reduction device 100 can provide a reduction method including the following steps (Ia) and (Ib): (Ia) NAD + and NADP + (Ib) The electrode 1 is brought into contact with a liquid 9a containing at least one selected from the group consisting of: + or NADP + Reduce the

[0020] Since the surface of the electrode 1 contains at least one selected from the group consisting of platinum, palladium, and nickel, NAD(P) can be converted to NAD(P) by electrons directly donated from the electrode 1 without using an electron mediator. + Therefore, NAD(P) can be reduced using an electron mediator. + Compared to reducing NAD(P) + In addition, the electron transfer efficiency is increased by directly donating electrons from the electrode 1. + or NADP + Furthermore, compared to the case where an electrode without a platinum-containing surface, such as a carbon electrode, is used, NAD(P) + The voltage required for the reduction of NAD(P) tends to be low, and the dimerization reaction represented by the above formula (4) tends to be suppressed. + As shown in Figure 2, for example, the efficiency of reduction of NAD(P) + NAD(P)H is generated by the reduction of the platinum, palladium, and nickel. The platinum, palladium, and nickel that can be contained on the surface of the electrode 1 may exist as simple substances or may be contained in an alloy. The alloy may be a core-shell type alloy or a solid solution.

[0021] The surface of the electrode 1 preferably comprises platinum.

[0022] According to the above reduction method, NAD is obtained by directly donating electrons from the electrode 1. + or NADP + The liquid 9a reduces NAD only by directly donating electrons from the electrode 1. + or NADP + For this reason, the liquid 9a may be, for example, + or NADP + Even if the liquid 9a contains an electron mediator, the liquid 9a can be converted to NAD by directly donating electrons from the electrode 1. + or NADP + The operating conditions of the reduction device 100 are adjusted so that the

[0023] The electrode 1 is not limited to a specific electrode as long as it has a surface containing at least one selected from the group consisting of platinum, palladium, and nickel. For example, the electrode 1 may include a conductive substrate and a surface layer containing at least one selected from the group consisting of platinum, palladium, and nickel. In this case, examples of the conductive substrate material include a carbon material, a conductive polymer, a semiconductor material, or a metal material. Examples of carbon materials include carbon nanotubes, Ketjen Black (registered trademark), glassy carbon, graphene, fullerene, carbon fiber, carbon fabric, and carbon aerogel. Examples of conductive polymers include polyaniline, polyacetylene, polypyrrole, poly(3,4-ethylenedioxythiophene), poly(p-phenylenevinylene), polythiophene, and poly(p-phenylene sulfide). Examples of semiconductor materials include silicon, germanium, indium tin oxide (ITO), titanium oxide, copper oxide, and silver oxide. Examples of metallic materials include gold, silver, titanium, aluminum, tungsten, copper, iron, and palladium. The electrode 1 may be made of at least one selected from the group consisting of platinum, platinum alloy, palladium, palladium alloy, nickel, and nickel alloy.

[0024] In the above reduction method, NAD is obtained by directly donating electrons from the electrode 1. + or NADP + The potential difference ΔV between the electrode 1 and the reference potential (0 V) is not limited to a specific value as long as it is possible to reduce NAD. In the reduction method described above, the potential difference ΔV is adjusted to, for example, 800 mV or more. Since the surface of the electrode 1 contains platinum, the potential difference ΔV can be adjusted to a range of 800 mV or more, and NAD can be reduced by directly donating electrons from the electrode 1. + or NADP + The potential of the electrode 1 is, for example, a potential lower than the reference potential, and can be adjusted to −800 mV or less.

[0025] The potential difference ΔV is, for example, 1500 mV or less. + or NADP + The potential difference ΔV for the reduction of NAD + or NADP + Therefore, it is easy to reduce the energy required for the reduction of NAD(P). + The efficiency of reduction is likely to be higher.

[0026] The liquid 9a may further contain mannitol dehydrogenase (MDH), fructose, and at least one selected from the group consisting of NADH and NADPH. FIG. 3 is a diagram schematically illustrating another example of a reduction method according to an embodiment. As shown in FIG. 3, in this case, in the reduction device 100, fructose is converted to mannitol by MDH, and NADH or NADPH is oxidized. From another perspective, the reduction method further includes converting fructose to mannitol by mannitol dehydrogenase and oxidizing NADH or NADPH. The fructose includes, for example, D-fructose. MDH functions as a catalyst, for example, in the reaction of the following formula (5) involving NAD(P)H:

[0027] D-fructose + NAD(P)H + H + → D-mannitol + NAD(P) +As shown in formula (5), the oxidation of NAD(P)H results in NAD(P) + As described above, NAD(P) is generated by directly donating electrons from electrode 1. + is reduced to produce NAD(P)H. Therefore, as long as a predetermined voltage is applied to the electrode 1 and the liquid 9a contains fructose, the reaction shown in formula (5) occurs continuously, and most of the fructose can be converted to mannitol even if NAD(P)H is not continuously added to the liquid 9a.

[0028] Thus, for example, a method for producing fructose can be provided, which includes the reduction method described above.

[0029] Liquid 9a may contain an enzyme other than MDH that functions as a catalyst in a reaction involving NAD(P)H. In this case, liquid 9a may contain a compound other than fructose that functions as a substrate in this reaction.

[0030] The liquid 9a is, for example, NAD(P) + It is an aqueous solution containing

[0031] As shown in FIG. 1 , the reduction device 100 further includes, for example, a reference electrode 2, a counter electrode 3, a cell 4, a lid 5, terminals 6a, 6b, and 6c, leads 7a, 7b, and 7c, a stirrer 8, a control device 30, and a stirrer 40. The electrode 1, the reference electrode 2, the counter electrode 3, the cell 4, the lid 5, the terminals 6a, 6b, and 6c, and the leads 7a, 7b, and 7c constitute an electrochemical reactor 10, which is a three-electrode cell. In the electrochemical reactor 10, the electrode 1 functions as, for example, a working electrode. The electrochemical reactor 10 may also be configured as a two-electrode cell that includes the electrode 1 and the counter electrode 3 and omits the reference electrode 2.

[0032] The cell 4 has, for example, a first storage chamber 4a and a second storage chamber 4b therein. The electrode 1 and the reference electrode 2 are disposed in the first storage chamber 4a, and the counter electrode 3 is disposed in the second storage chamber 4b. The cell 4 has, for example, a connection portion 4c. The connection portion 4c connects the first storage chamber 4a and the second storage chamber 4b, and a separator 4s is disposed in the connection portion 4c. The separator 4s separates the space on the first storage chamber 4a side of the connection portion 4c from the space on the second storage chamber 4b side.

[0033] As described above, the first storage chamber 4a contains NAD(P) + A liquid 9a containing the above-mentioned compound can be accommodated in the second storage chamber 4b. This allows the liquid 9a to come into contact with the electrode 1. The second storage chamber 4b can accommodate, for example, a liquid 9b. This allows the liquid 9b to come into contact with the counter electrode 3. The liquid 9b is, for example, an electrolyte solution such as a phosphate buffer solution.

[0034] The separator 4s has, for example, ion conductivity and prevents the permeation of some of the components contained in the liquid 9a and the liquid 9b. For example, the separator 4s has, for example, ion conductivity and prevents the permeation of some of the components contained in the liquid 9a and the liquid 9b. + Furthermore, when the liquid 9a contains MDH, fructose, and NAD(P)H, the separator 4s may block the permeation of these. The separator 4s has, for example, proton conductivity. The separator 4s contains, for example, a polymer having a perfluoro side chain containing a sulfonic acid group.

[0035] The surface of the counter electrode 3 includes, for example, a conductive material. Examples of the conductive material include a carbon material, a conductive polymer, a semiconductor material, or a metal material. The conductive material is, for example, a material that does not decompose when the conductive material is oxidized. Examples of the conductive material include a carbon material, a conductive polymer, a semiconductor material, and a metal material. Examples of the carbon material include carbon nanotubes, Ketjen Black (registered trademark), glassy carbon, graphene, fullerene, carbon fiber, carbon fabric, and carbon aerogel. Examples of the conductive polymer include polyaniline, polyacetylene, polypyrrole, poly(3,4-ethylenedioxythiophene), poly(p-phenylene vinylene), polythiophene, and poly(p-phenylene sulfide). Examples of the semiconductor material include silicon, germanium, indium tin oxide (ITO), titanium oxide, copper oxide, and silver oxide. Examples of the metal material include gold, platinum, silver, titanium, aluminum, tungsten, copper, iron, and palladium. The counter electrode 3 may be a platinum electrode.

[0036] In the reduction device 100, for example, the electrode 1 functions as a cathode electrode, and the counter electrode 3 functions as an anode electrode. When a voltage is applied between the electrode 1 and the counter electrode 3, the potential of the electrode 1 is, for example, NAD(P) + The surface area of ​​the counter electrode 3 is, for example, larger than the surface area of ​​the electrode 1. With this configuration, NAD(P) + Therefore, when the liquid 9a contains fructose, a large amount of mannitol is likely to be produced in a short time.

[0037] The reference electrode 2 does not react with the components contained in the liquid 9 a and is maintained at a specific potential. The reference electrode 2 is, for example, a silver / silver chloride electrode. The potential of the reference electrode 2 is maintained at, for example, a reference potential (0 V).

[0038] In the electrochemical reactor 10, the cell 4 is closed by, for example, a lid 5. Terminals 6a, 6b, and 6c are attached to the lid 5. A lead 7a extends from the terminal 6a and electrically connects the terminal 6a to the electrode 1. A lead 7b extends from the terminal 6b and electrically connects the terminal 6b to the reference electrode 2. A lead 7c extends from the terminal 6c and electrically connects the terminal 6c to the counter electrode 3. The terminals 6a, 6b, and 6c are electrically connected to a power source 20.

[0039] The power supply 20 applies a voltage between the electrode 1 and the counter electrode 3 in accordance with a control signal output from the control device 30, for example, and adjusts the potential difference between the electrode 1 and the reference electrode 2 to a predetermined value.

[0040] The control device 30 performs information processing to control the application of voltage by the power supply 20 and the movement of a motor (not shown) of the agitator 40. The control device 30 includes, for example, a memory that stores a program for controlling the application of voltage by the power supply 20 and the movement of the motor of the agitator 40, and an arithmetic device such as a processor that reads the program and performs predetermined calculations.

[0041] In the stirring device 40, the operation of the motor is controlled in accordance with a control signal output from the control device 30. This adjusts the rotation speed and rotation time of the stirring bars 8 arranged in each of the first storage chamber 4 a and the second storage chamber 4 b.

[0042] 4 is a block diagram showing an example of the functional configuration of the reduction device 100. As shown in FIG. 4, the power supply 20 includes, for example, an acquisition unit 21, an information processing unit 22, and a voltage control unit 23.

[0043] The acquiring unit 21 acquires the control signal output from the control unit 30. The acquiring unit 21 may also acquire measurement data such as the potential of each electrode in the electrochemical reactor 10 and the value of the current flowing through the liquid 9b.

[0044] The information processing unit 22 processes the information acquired by the acquisition unit 21. For example, when the information processing unit 22 acquires a control signal from the acquisition unit 21, it outputs the acquired control signal to the voltage control unit 23. When the voltage control unit 23 starts applying a voltage to each electrode of the electrochemical reactor 10, the information processing unit 22 acquires measurement data, such as the potential of each electrode of the electrochemical reactor 10 and the value of the current flowing through the liquid 9b, acquired from the acquisition unit 21. Based on the acquired data, the information processing unit 22 calculates a voltage to be applied to the electrode 1 so as to maintain the potential difference between the electrode 1 and the reference electrode 2 at a predetermined value. Then, it outputs a control signal to the voltage control unit 23 to adjust the voltage of the electrode 1 to the calculated voltage.

[0045] The voltage control unit 23 applies a voltage to each electrode of the electrochemical reactor 10 based on the control signal output from the information processing unit 22 .

[0046] As shown in FIG. 1, the power supply 20 and the control device 30 may be separate entities, or the power supply 20 and the control device 30 may be integrally configured.

[0047] The control device 30 includes, for example, an acquisition unit 31, an information processing unit 32, a storage unit 33, and an output unit 34.

[0048] The acquisition unit 31 acquires, for example, instruction information, which is information relating to an instruction input by a user, and outputs the acquired instruction information to the information processing unit 32 .

[0049] The information processing unit 32 determines the conditions for applying a voltage to each electrode of the electrochemical reactor 10, for example, based on the instruction information acquired by the acquisition unit 31. The instruction information includes, for example, the type of enzyme, the amount of the liquid 9a, the completion time of the reaction to produce mannitol, the completion time of the reaction, etc. The instruction information may include the ratio of the target fructose concentration to the initial fructose concentration in the liquid 9a, or information corresponding to that ratio.

[0050] The information processing unit 32 may output to the output unit 34 a control signal that controls voltage application under conditions determined based on the instruction information, or may output to the output unit 34 a control signal that controls voltage application under voltage application conditions preset by the user.

[0051] The output unit 34 outputs the control signal obtained from the information processing unit 32 to the power supply 20 .

[0052] The storage unit 33 stores data such as instruction information acquired by the acquisition unit 31, computer programs executed by the control unit 30 (for example, application programs for controlling the power supply 20), and the like.

[0053] FIG. 5 is a flowchart showing an example of a reduction method. For example, liquids 9a and 9b are contained in the cell 4. Next, in step S101, conditions for terminating the reduction reaction are set. For example, these conditions are that a predetermined time has elapsed since the start of voltage application or that the current at the electrode 1 falls below a predetermined value Ie. In step S101, conditions related to the target value of the potential of the electrode 1, the target value of the voltage between the electrode 1 and the counter electrode 3, and the rotation speed of the stirring device 40 may be set. The conditions in step S101 may be set based on instruction information input to the reduction device 100. The instruction information may include, for example, the amount of the liquid 9a, the completion time of the reaction to produce mannitol, the completion time of the reaction, the ratio of the target fructose concentration to the initial fructose concentration in the liquid 9a, or information corresponding to that ratio.

[0054] Next, in step S102, application of voltage is started. For example, the control device 30 generates a control signal in accordance with a target value of the potential of the electrode 1 and a target value of the voltage between the electrode 1 and the counter electrode 3. The power supply 20 applies a voltage between the electrode 1 and the counter electrode 3 in accordance with the control signal obtained from the control device 30.

[0055] Next, in step S103, NAD(P) + is reduced to generate NAD(P)H, and fructose is converted to mannitol by the reaction of formula (5) involving NAD(P)H. MDH functions as a catalyst in this reaction.

[0056] Next, the process proceeds to step S104, where it is determined whether the condition for terminating the reduction reaction set in step S101 is met. If the result of the determination in step S104 is negative, the process proceeds to step S106, where the application of voltage between the electrode 1 and the counter electrode 3 is continued, and steps S103 and S104 are repeated.

[0057] If the determination in step S104 is affirmative, the process proceeds to step S105, where the application of voltage by the power supply 20 is terminated. Thereafter, the liquid 9a is collected, and the series of processes is completed.

[0058] Other Embodiments The method and apparatus for producing mannitol have been described based on the embodiments. However, the present disclosure is not limited to these embodiments. As long as they do not deviate from the gist of the present disclosure, various modifications that a person skilled in the art can conceive of to the embodiments and other embodiments formed by combining some of the components of the embodiments are included in the scope of the present disclosure.

[0059] (Additional Note) From the above description, the following techniques are disclosed.

[0060] (Technology 1) Oxidized nicotinamide adenine nucleotide (NAD + ) and oxidized nicotinamide adenine dinucleotide phosphate (NADP + contacting a liquid containing at least one selected from the group consisting of platinum, palladium, and nickel with an electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel; and directly donating electrons from the electrode to form the NAD. + or the NADP + and reducing the

[0061] (Technology 2) The reduction method according to Technology 1, wherein the surface contains platinum.

[0062] (Technology 3) The liquid is transferred from the electrode to the NAD + or the NADP + 3. The reduction method according to claim 1 or 2, wherein the reduction method does not contain an electron mediator that transfers electrons to the compound.

[0063] (Technology 4) The reduction method according to any one of Techniques 1 to 3, further comprising adjusting the potential difference between the electrode and a reference potential to 800 mV or more.

[0064] (Technology 5) The reduction method according to any one of Technologies 1 to 4, wherein the liquid further contains mannitol dehydrogenase, fructose, and at least one selected from the group consisting of reduced nicotinamide adenine nucleotide (NADH) and reduced nicotinamide adenine dinucleotide phosphate (NADPH), and the reduction method further comprises converting the fructose to mannitol with the mannitol dehydrogenase and oxidizing the NADH or the NADPH.

[0065] (Technology 6) A method for producing mannitol, comprising the reduction method according to Technology 5.

[0066] (Technology 7) An electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel, and an oxidized nicotinamide adenine nucleotide (NAD + ) and oxidized nicotinamide adenine dinucleotide phosphate (NADP + and a power source electrically connected to the electrodes, and the NAD is generated by directly donating electrons from the electrodes. + or the NADP + A reduction device that reduces the amount of

[0067] (Technology 8) The reduction method according to Technology 7, wherein the surface contains platinum.

[0068] (Technology 9) The reduction device according to Technology 7 or 8, wherein the liquid further contains mannitol dehydrogenase, fructose, and at least one selected from the group consisting of reduced nicotinamide adenine nucleotide (NADH) and reduced nicotinamide adenine dinucleotide phosphate (NADPH), and the reduction device converts the fructose to mannitol by the mannitol dehydrogenase and oxidizes the NADH or the NADPH.

[0069] (Technology 10) An electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel, which functions as the electrode in the reduction device according to any one of Technologies 7 to 9.

[0070] The present disclosure will be described in more detail below with reference to examples, but the present disclosure is not limited to the following examples.

[0071] [Example 1] An apparatus according to Example 1, which corresponds to the reduction apparatus 100 shown in Figure 1, was prepared. In this apparatus, a platinum electrode purchased from Tanaka Kikinzoku Kogyo Co., Ltd. was used as the working electrode according to Example 1. The electrode area of ​​the platinum electrode was 1.35 cm 2 The reference electrode was a silver-silver chloride (Ag / AgCl) electrode. The counter electrode was a coiled platinum wire with a wire diameter of 0.5 mm and a length of 23 cm. An OctoStat 30 potentiostat (Ivium Technologies BV) was used, and the working electrode, reference electrode, and counter electrode were electrically connected to the working electrode terminal, reference electrode terminal, and counter electrode terminal of the potentiostat, respectively, to form a three-electrode cell. The membrane separating the interior of the first and second storage compartments was a Nafion 117 ion-exchange membrane (MTI Corporation).

[0072] Phosphate-Buffer Saline (PBS) with a pH of 7.4 (supplied by Nippon Gene Co., Ltd.) was placed in the second chamber containing the counter electrode. A portion of the counter electrode was in contact with the PBS in the second chamber. The amount of PBS placed in the second chamber was 20 mL.

[0073] The above phosphate buffer solution was prepared separately, and NAD + The treatment solution according to Example 1 was prepared by dissolving NAD + As the NAD in the treatment solution according to Example 1, β-nicotinamide adenine dinucleotide sodium salt supplied by Sigma-Aldrich was used. +The concentration of the treatment solution according to Example 1 was 6.0 mM. 20 mL of the treatment solution according to Example 1 was contained in the first storage chamber. In the first storage chamber, a part of the working electrode and a part of the reference electrode were in contact with the treatment solution according to Example 1.

[0074] In the apparatus according to Example 1, a predetermined voltage was applied at room temperature (25°C) while stirring the treatment solution using a stirrer. This adjusted the potential of the working electrode according to Example 1 to -900 mV relative to the base potential (0 V), which is the potential of the reference electrode. The treatment solution after voltage application was sampled. 190 μL of the treatment solution obtained by sampling was mixed with 1 M acetaldehyde (provided by Fujifilm Wako Pure Chemical Industries, Ltd.) diluted with pure water, and an ADH solution obtained by dissolving alcohol dehydrogenase (ADH) (provided by Sigma-Aldrich) in the above phosphate buffer solution, resulting in an enzymatic reaction involving ADH as an enzyme. The amount of 1 M acetaldehyde was 8 μL, and the amount of ADH solution was 2 μL. In this enzymatic reaction, acetaldehyde was reduced to produce ethanol, and NADH contained in the treatment solution was oxidized. The amount of NADH in the treated solution after voltage application was quantified based on the difference between the absorbance at 340 nm of the ADH solution after the enzyme reaction and the absorbance at 340 nm of the treated solution before the enzyme reaction. The results are shown in Figure 6 and Table 1. In Figure 6, the vertical axis represents the NADH yield determined from the amount of NADH contained in the treated solution after voltage application. As shown in Figure 6, the NADH yield was 45% when voltage was applied in the device of Example 1.

[0075] [Comparative Example 1] A device according to Comparative Example 1 was prepared in the same manner as in Example 1, except that a gold electrode purchased from Atsugi Micro Co., Ltd. was used instead of the working electrode according to Example 1. The electrode area of ​​the gold electrode was 1.35 cm 2 In the same manner as in Example 1, phosphate buffer solution (PBS) was placed in the second storage chamber in which the counter electrode was placed.

[0076] NAD was added to a separately prepared phosphate buffer solution. +and methyl viologen as an electron mediator were dissolved to prepare a treatment solution according to Comparative Example 1. Methyl viologen dichloride hydrate provided by Sigma-Aldrich was used as the methyl viologen. + The concentration of the methyl viologen was 6.0 mM, and the concentration of the methyl viologen was 3.0 mM. 20 mL of the treatment solution according to Comparative Example 1 was placed in the first storage chamber. In the first storage chamber, a part of the working electrode and a part of the reference electrode were in contact with the treatment solution according to Comparative Example 1.

[0077] In the device according to Comparative Example 1, a predetermined voltage was applied at room temperature of 25°C while stirring the treatment solution using a stirrer. As a result, the potential of the working electrode according to Comparative Example 1 was adjusted to -1050 mV relative to the base potential (0 V), which is the potential of the reference electrode. The treatment solution after voltage application was sampled, and the amount of NADH in the treatment solution after voltage application was quantified in the same manner as in Example 1. The results are shown in Figure 6. As shown in Figure 6, the yield of NADH was 23% when voltage was applied in the device according to Comparative Example 1. In Comparative Example 1, NADH was converted to NADH via methyl viologen, an electron mediator. + It is believed that the amount was donated to and reduced.

[0078] Comparison between Example 1 and Comparative Example 1 shows that the NAD contained in the treatment solution according to Example 1 + The yield of NADH produced by reduction of + This is because in Example 1, NAD was produced without the aid of an electron mediator. + Since electrons are donated directly to NAD + This is thought to be because there are few types of molecules involved in the reduction of NAD, and the efficiency of electron transfer is high. + can be efficiently reduced.

[0079] [Comparative Example 2] An apparatus according to Comparative Example 2 was prepared in the same manner as in Example 1, except that a titanium electrode purchased from ESPI Metals was used instead of the working electrode according to Example 1. The electrode area of ​​the titanium electrode was 1.35 cm 2 In the same manner as in Example 1, phosphate buffer solution (PBS) was placed in the second storage chamber in which the counter electrode was placed. + The treatment solution according to Comparative Example 2 was prepared by dissolving NAD in the treatment solution according to Comparative Example 2. + The concentration of the treatment solution of Comparative Example 2 was 6.0 mM. 20 mL of the treatment solution of Comparative Example 2 was contained in the first storage chamber. In the first storage chamber, a part of the working electrode and a part of the reference electrode were in contact with the treatment solution of Comparative Example 2.

[0080] In the device according to Comparative Example 2, a predetermined voltage was applied at room temperature of 25°C while stirring the treatment solution using a stirrer. As a result, the potential of the working electrode according to Comparative Example 2 was adjusted to -900 mV, -1200 mV, and -1800 mV relative to the base potential (0 V), which is the potential of the reference electrode. The treatment solution after voltage application was sampled, and the amount of NADH in the treatment solution after voltage application was quantified in the same manner as in Example 1. The results are shown in Table 1. As shown in Table 1, when the potential of the working electrode according to Comparative Example 2 was -900 mV or -1200 mV, the yield of NADH was 0%, and + When the potential of the working electrode in Comparative Example 2 was −1800 mV, the yield of NADH was 4.2%.

[0081]

[0082] In Comparative Example 2, an electrode having a surface containing platinum was used as the working electrode. + At the potential where reduction of NAD + The reduction of NAD was not possible. + In order to cause the reduction of NAD contained in the treatment solution according to Example 1, it was necessary to adjust the potential of the working electrode to −1800 mV. + The yield of NADH produced by the reduction of was higher than the yield of NADH in Comparative Example 2 when the potential of the working electrode was adjusted to −1800 mV.

[0083] It is believed that platinum and hydrogen bond easily occur on the surface of platinum. Therefore, it is believed that a large number of hydrogen atoms exist at the interface between the electrode having a surface containing platinum and the treatment solution, compared to an electrode having a surface made of a metal other than platinum, such as titanium. Therefore, NAD(P) + The second step of the reduction reaction shown in formula (3) proceeds rapidly to give NAD(P) shown in formula (4). ・ It is thought that the dimerization reaction of NAD(P)H is suppressed and NAD(P)H is produced. Palladium and nickel, like platinum, are prone to bond with hydrogen. Therefore, even when a surface containing palladium or nickel is used, the NAD(P)H shown in formula (4) ・ On the other hand, on the surface of gold, it is difficult for hydrogen to bond with gold, so NAD(P) + Therefore, NAD(P) is not easily reduced. + Gold cannot be used as the metal constituting the surface of the electrode that comes into contact with the substrate.

[0084] [Comparative Example 3] An apparatus according to Comparative Example 3 was prepared in the same manner as in Example 1, except that a carbon electrode purchased from Nilaco Corporation was used instead of the working electrode according to Example 1. The electrode area of ​​the carbon electrode was 1.35 cm 2 In the same manner as in Example 1, phosphate buffer solution (PBS) was placed in the second storage chamber in which the counter electrode was placed. + The treatment solution according to Comparative Example 3 was prepared by dissolving NAD in the treatment solution according to Comparative Example 3. + The concentration of the treatment solution of Comparative Example 3 was 6.0 mM. 20 mL of the treatment solution of Comparative Example 3 was contained in the first storage chamber. In the first storage chamber, a part of the working electrode and a part of the reference electrode were in contact with the treatment solution of Comparative Example 3.

[0085] In the device according to Comparative Example 3, a predetermined voltage was applied at room temperature of 25°C while stirring the treatment solution using a stirrer. As a result, the potential of the working electrode according to Comparative Example 3 was adjusted to -900 mV, -1200 mV, and -1800 mV relative to the base potential (0 V), which is the potential of the reference electrode. The treatment solution after voltage application was sampled, and the amount of NADH in the treatment solution after voltage application was quantified in the same manner as in Example 1. The results are shown in Table 2.

[0086]

[0087] Example 2 The device according to Example 2 was prepared in the same manner as in Example 1. Similarly to Example 1, phosphate buffer solution (PBS) was placed in the second storage chamber in which the counter electrode was placed. A treatment solution according to Example 2 was prepared by dissolving fructose, reduced NADH, and mannitol dehydrogenase in a separately prepared phosphate buffer solution. D(-)-fructose provided by Fujifilm Wako Pure Chemical Industries, Ltd. was used as the fructose. β-nicotinamide adenine dinucleotide, reduced disodium salt hydrate provided by Sigma-Aldrich was used as the reduced NADH. Mannitol dehydrogenase (MDH) provided by Megazyme was used as the mannitol dehydrogenase. The concentration of fructose in the treatment solution according to Example 2 was 350 mM. The concentration of NADH in the treatment solution according to Example 2 was 5.6 mM. The concentration of MDH in the treatment solution according to Example 2 was 2 Units / mL. 20 mL of the treatment liquid according to Example 2 was placed in the first storage chamber. In the first storage chamber, a part of the working electrode and a part of the reference electrode were in contact with the treatment liquid according to Example 2.

[0088] In the device according to Example 2, a predetermined voltage was applied at room temperature of 25°C while stirring the treatment solution using a stirrer. As a result, the potential of the working electrode was adjusted to -900 mV relative to the base potential (0 V), which is the potential of the reference electrode. After 16 hours of voltage application, the treatment solution according to Example 2 was sampled, and the amount of fructose in the treatment solution after voltage application was measured using a Fructose Colorimetric / Fluorometric Assay Kit provided by Sigma-Aldrich. The results are shown in Figure 7. In Figure 7, the vertical axis indicates the amount of fructose reduction (reduction rate) in the treatment solution due to voltage application. As shown in Figure 7, the amount of fructose reduction in Example 2 was 46%. In Example 2, NADH produced by oxidation of NADH accompanying the decomposition of fructose was measured. + Using an electrode with a platinum surface, fructose could be efficiently reduced to produce NADH, confirming that an amount of fructose greater than the amount of NADH added (5.6 mM) could be decomposed.

[0089] [Comparative Example 4] An apparatus according to Comparative Example 4 was prepared in the same manner as in Comparative Example 1. In the apparatus according to Comparative Example 4, a gold electrode purchased from Atsugi Micro Co., Ltd. was used as the working electrode. The electrode area of ​​this gold electrode was 1.35 cm 2 As in Comparative Example 1, phosphate buffer solution (PBS) was placed in the second storage chamber in which the counter electrode was placed. A treatment solution according to Comparative Example 4 was prepared by dissolving fructose, reduced NADH, methyl viologen, and mannitol dehydrogenase in a separately prepared phosphate buffer solution. In the treatment solution according to Comparative Example 4, the fructose concentration was 350 mM, the reduced NADH concentration was 5.6 mM, and the methyl viologen concentration was 2.8 mM. The MDH concentration in the treatment solution according to Comparative Example 4 was 2 Units / mL. 20 mL of the treatment solution according to Comparative Example 4 was placed in the first storage chamber. In the first storage chamber, a portion of the working electrode and a portion of the reference electrode were in contact with the treatment solution according to Comparative Example 4.

[0090] In the device according to Comparative Example 4, a predetermined voltage was applied at room temperature of 25°C while stirring the treatment solution using a stirrer. As a result, the potential of the working electrode was adjusted to -1050 mV relative to the base potential (0 V), which is the potential of the reference electrode. After 16 hours of voltage application, the treatment solution according to Comparative Example 4 was sampled, and the amount of fructose in the treatment solution after voltage application was measured using a Fructose Colorimetric / Fluorometric Assay Kit provided by Sigma-Aldrich. The results are shown in Figure 7. As shown in Figure 7, the amount of fructose reduced in Comparative Example 4 was 44%. In Comparative Example 4, NADH produced by oxidation during the decomposition of fructose was oxidized. + Using an electrode with a platinum surface, fructose could be efficiently reduced to produce NADH, confirming that fructose in an amount equal to or greater than the amount of NADH added (5.6 mM) could be decomposed.

[0091] Comparing Example 2 and Comparative Example 4, the amount of fructose reduced in Example 2 is greater than the amount of fructose reduced in Comparative Example 4. This is because in Example 2, NAD was removed from the working electrode, which is an electrode having a surface containing platinum. + Electrons are donated directly to NAD, and NAD is more efficiently reacted with NAD than when an electron mediator is used as in Comparative Example 4. + This is thought to be because the reduction efficiency of

[0092] As described above, the NAD(P) according to the present disclosure + directly donates electrons to NAD(P) + An electrode having a platinum-containing surface capable of reducing β-glucan and a device including this electrode have been described based on the following embodiments. However, the present disclosure is not limited to these embodiments. As long as they do not deviate from the gist of the present disclosure, various modifications conceivable by a person skilled in the art to the embodiments and other embodiments constructed by combining some of the components of the embodiments are also included in the scope of the present disclosure.

[0093] According to the present disclosure, NAD(P) + directly donates electrons to NAD(P) +and a device that can decompose fructose contained in food using the electrode.

[0094] 1 Electrode 4a Storage chamber 9a Liquid 20 Power source 100 Reduction device

Claims

1. A liquid containing at least one selected from the group consisting of oxidized nicotinamide adenine dinucleotide (NAD + ), and oxidized nicotinamide adenine dinucleotide phosphate (NADP + ), and an electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel are brought into contact, and electrons are directly donated from the electrode to reduce the NAD + or the NADP + . A reduction method comprising the steps of:

2. The reduction method according to claim 1, wherein the surface contains platinum.

3. The liquid does not contain an electron mediator responsible for the transfer of electrons from the electrode to the NAD + or the NADP + The reduction method according to claim 1.

4. The reduction method according to claim 1, further comprising adjusting the potential difference between the electrode and the reference potential to 800 mV or more.

5. The liquid further contains at least one selected from the group consisting of mannitol dehydrogenase, fructose, reduced nicotinamide adenine dinucleotide (NADH), and reduced nicotinamide adenine dinucleotide phosphate (NADPH), and the mannitol dehydrogenase further converts the fructose to mannitol and oxidizes the NADH or the NADPH. The reduction method according to claim 1.

6. A method for producing mannitol, comprising the reduction method according to claim 5.

7. An electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel, a storage chamber capable of storing a liquid containing at least one selected from the group consisting of oxidized nicotinamide adenine dinucleotide (NAD + ), and oxidized nicotinamide adenine dinucleotide phosphate (NADP + ), and a power source electrically connected to the electrode, and reducing the NAD + or the NADP + by directly donating electrons from the electrode. A reducing device.

8. The reduction device according to claim 7, wherein the surface contains platinum.

9. The liquid further contains at least one selected from the group consisting of mannitol dehydrogenase, fructose, reduced nicotinamide adenine dinucleotide (NADH), and reduced nicotinamide adenine dinucleotide phosphate (NADPH), and the reduction device converts the fructose to mannitol by the mannitol dehydrogenase and oxidizes the NADH or the NADPH. The reduction device according to claim 7.

10. An electrode having a surface containing at least one selected from the group consisting of platinum, palladium, and nickel, and functioning as the electrode in the reduction device according to claim 7.

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