Pyridine sulfonamide iridium complexes, methods of making, and high selectivity catalytic regeneration of nadh

Abstract

This invention relates to the field of organometallic complex catalysis, specifically disclosing an organometallic iridium complex capable of efficiently and selectively catalyzing the regeneration of coenzyme NADH and its preparation method. The organometallic iridium complex is a novel pentamethylcyclopentadienyl (Cp*)pyridinesulfonamide iridium complex. The advantage of this invention is that this organometallic iridium complex can efficiently and selectively catalyze the oxidation of coenzyme NAD. + Low-temperature hydrogenation converts it into reduced coenzyme NADH, and it exhibits good biocompatibility.

Description

Technical Field

[0001] This invention relates to the fields of organometallic complex preparation and catalytic chemistry, specifically to the design and preparation methods of organometallic iridium complexes, and to the application of organometallic iridium complexes in the catalytic regeneration synthesis of the reduced coenzyme nicotinamide adenine dinucleotide (NADH). Background Technology

[0002] Currently, biocompatible organometallic catalysts, as an emerging frontier in chemical biology, have shown significant application value in disease treatment, bioimaging and diagnosis, and biochemical synergistic catalytic preparation. Biochemical cascade catalytic systems based on organometallic catalysts and bioenzymes represent a promising and unique synthetic method that combines the advantages of both chemical and biocatalysis. However, establishing biochemical cascade catalytic systems faces two main challenges: (1) developing the coenzyme nicotinamide adenine dinucleotide (NAD). + (2) Solving the incompatibility problem between chemical catalysts and corresponding enzymes. (NADH) High-efficiency chemical catalytic regeneration catalyst; + NADH plays a crucial role in biological metabolism and is closely related to human health. Therefore, researchers have developed various methods to mimic NAD. + The intercatalytic transformation of NADH includes electrocatalysis, photocatalysis, chemical catalysis, and metal-organic catalysis. Among these, metal-organic catalysis has become an important catalytic approach due to its versatility, ease of modification, and tolerance to water and oxygen. Extensive research on various biocompatible metal-organic catalytic reactions has also contributed to the understanding of NAD... + The efficient regeneration of / NADH is guaranteed.

[0003] Among various metal-organic catalysts, iridium-organic catalysts have been shown to be active in complex biological environments. Previous studies have shown that pyridine amide-type iridium complexes exhibit outstanding catalytic performance for formic acid decomposition and aldehyde reduction, mainly due to the strong electron-donating effect of the N⁻ anion. Pyridine amide-iridium complexes have been shown to exhibit excellent catalytic activity for the regeneration of the reduced coenzyme nicotinamide adenine dinucleotide (NADH) under physiological conditions, even surpassing some related biological enzymes (such as formic acid dehydrogenase). However, existing highly active metal-iridium complex catalysts for NADH regeneration still exhibit low selectivity for NADH, thus greatly limiting the application of highly active metal-iridium complex catalysts in chemo-biological co-catalysis.

[0004] Based on this, the present invention synthesizes a novel pentamethylcyclopentadienyl (Cp*)pyridinesulfonamide iridium complex by adjusting the electronegativity of the ligand. This complex not only exhibits excellent catalytic performance for the regeneration of coenzyme NADH, but its catalytic activity for other enzyme substrates (α-ketoglutarate, benzaldehyde, and trimethyl-p-benzoquinone (TMBQ)) is significantly limited. Therefore, based on the excellent selective catalytic regeneration performance of the invented pyridinesulfonamide iridium complex for coenzyme NADH and its good biocompatibility, it can be applied to construct more superior chemo-enzyme synergistic catalytic systems, demonstrating excellent chemo-biological synergistic preparation performance. Summary of the Invention

[0005] To address the low selectivity of existing high-activity iridium metal complex catalysts for NADH regeneration and their limitations in chemical-biological synergistic catalysis applications, the first objective of this invention is to provide a novel organometallic iridium complex that is structurally stable, biocompatible, and exhibits high efficiency and selectivity in catalyzing NADH regeneration. The second objective of this invention is to provide a simple synthetic route and a high-yield method for preparing the aforementioned iridium metal complex.

[0006] To achieve the first objective mentioned above, the present invention adopts the following technical solution:

[0007] An organometallic iridium complex capable of highly selectively catalyzing the regeneration of coenzyme NADH, wherein the iridium complex can efficiently catalyze the oxidation of coenzyme NAD. + The hydrogenated and reduced coenzyme NADH exhibits significantly lower catalytic activity against other enzyme substrates (α-ketoglutarate, benzaldehyde, and trimethyl-p-benzoquinone (TMBQ)). The iridium complex is characterized by being a novel pentamethylcyclopentadienyl (Cp*)pyridinesulfonamide iridium complex, with the structural formula shown in Formula 1.

[0008] To achieve the second objective mentioned above, the present invention adopts the following technical solution:

[0009] A method for preparing an organometallic iridium complex of Formula 1 capable of highly selectively catalyzing the regeneration of coenzyme NADH, characterized by comprising the following steps:

[0010] Step 1: Dissolve the raw material 2-mercaptopyridine in dichloromethane solvent, cool to -10-5 degrees Celsius, add mixed acid concentrated hydrochloric acid-hypochlorous acid or concentrated sulfuric acid-hypochlorous acid to carry out sulfonation reaction. After the reaction is completed, the mixture is extracted with dichloromethane and dried with anhydrous sodium sulfate to obtain the activated intermediate 2-pyridinesulfonyl chloride.

[0011] Step 2: Mix the intermediate 2-pyridinesulfonyl chloride obtained in Step 1 with various substituted aniline compounds and react them in dry dichloromethane, acetonitrile, or tetrahydrofuran solvent for 10 to 30 hours.

[0012] Step 3: Add a certain amount of dichloromethane to the reaction solution in Step 2, wash three times with a 1.0 mol / L phosphoric acid solution, wash three times with deionized water, dry with anhydrous sodium sulfate, and then distill under pressure to obtain the crude product 2-pyridinesulfonamide ligand.

[0013] Step 4: The crude 2-pyridine sulfonamide ligand obtained in Step 3 was separated using a silica gel normal-phase column with dichloromethane / petroleum ether as the mobile phase to obtain pure 2-pyridine sulfonamide ligand with a yield of 70%~90%.

[0014] Step 5: Dissolve the pure 2-pyridinesulfonamide ligand, pentamethylcyclopentadiene dichloroiridium dimer and ammonium hexafluorophosphate obtained in step 4 in anhydrous ethanol and reflux for 10 to 24 hours under nitrogen protection.

[0015] Step 6: Cool the reaction solution from Step 5 to room temperature, filter, wash three times with ethanol, and collect the solid, which is the pure pyridinesulfonamide type iridium complex shown in Formula 1.

[0016] Preferably, in step 2, the molar ratio of the intermediate 2-pyridinesulfonyl chloride to various substituted aniline compounds is 1:1 to 10.

[0017] Preferably, in step 4, the volume ratio of the mobile phase dichloromethane to petroleum ether is 1:0.1~10.

[0018] Preferably, in step 5, the molar ratio of the 2-pyridinesulfonamide ligand to the pentamethylcyclopentadiene iridium dichloride dimer and ammonium hexafluorophosphate is 0.5~5 : 1 : 2~10.

[0019] The advantages of this invention are:

[0020] The organometallic iridium complex provided by this invention has a stable structure, which is beneficial for long-term storage at room temperature, in air, and in humid environments.

[0021] (2) The organometallic iridium complex provided by the present invention can efficiently and selectively catalyze the oxidation of coenzyme NAD. + Low-temperature hydrogenation converts it into reduced coenzyme NADH, but its catalytic activity against other enzyme substrates (α-ketoglutarate, benzaldehyde, and trimethyl-p-benzoquinone (TMBQ)) is significantly lower. This makes it advantageous to combine it with related biological dehydrogenases to construct an excellent chemical-enzyme synergistic catalytic system, which can be applied to the preparation of special biochemicals such as L-glutamic acid and trimethyl-p-benzoquinone. Attached Figure Description

[0022] Figure 1 This is a synthetic route diagram of the iridium complex shown in Formula 1;

[0023] Figure 2 The iridium complex shown in Formula 1 catalyzes the oxidation of coenzyme NAD. + A schematic diagram of the hydrogenation reaction to prepare reduced coenzyme NADH, and a schematic diagram comparing the yields of NADH regeneration catalyzed by 2-pyridinebenzamide iridium complex 5.

[0024] Figure 3 This is a schematic diagram of the reaction of iridium complex catalyzing the hydrogenation of benzaldehyde to benzyl alcohol, the hydrogenation of trimethyl-p-benzoquinone to trimethylhydroquinone, and the hydrogenation of α-ketoglutarate to L-glutamic acid, as shown in Formula 1. It is also a schematic diagram comparing the yields of benzyl alcohol, trimethylhydroquinone, and L-glutamic acid catalyzed by the hydrogenation of iridium complex 5 with that of 2-pyridinebenzamide.

[0025] Figure 4 In the presence of various biomolecules, iridium complex 1a of formula 1 exhibits its effect on NAD. + A schematic diagram comparing the yields of reduced coenzyme NADH prepared by hydrogenation.

[0026] Figure 5 This is a schematic diagram comparing the yields of L-glutamic acid produced by constructing a chemical-enzyme cascade catalytic system by combining the iridium complex 1a-f of formula 1 with L-glutamic acid dehydrogenase GLDH. Implementation

[0027] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments.

[0028] Structure of novel pyridinesulfonamide type iridium complex

[0029] In the highly efficient and selective iridium complex structure for catalyzing the regeneration of coenzyme NADH provided by this invention, the two nitrogen atoms in the ligand N-(4-substituted phenyl)pyridine-2-sulfonamide coordinate with metallic iridium to form a stable five-membered ring structure. The metallic iridium further reacts with pentamethylcyclopentadienyl (Cp*) and chloride ions (Cl... - A stable sandwich-structured iridium complex is formed through single coordination, as shown in Formula 1.

[0030] The present invention provides a novel method for preparing pyridine sulfonamide-type iridium complexes.

[0031] A method for preparing an organometallic iridium complex of Formula 1 capable of highly selectively catalyzing the regeneration of coenzyme NADH, characterized by comprising the following steps:

[0032] The present invention provides a method for preparing organometallic iridium complexes with high selectivity for catalyzing the regeneration of coenzyme NADH, as shown in Formula 1. The specific synthetic route is attached. Figure 1 As shown.

[0033] Example 1: A method for preparing N-phenylpyridine-2-sulfonamide pentamethylcyclopentadiene (Cp*) iridium chloride complex 1a, specifically including the following steps:

[0034] Step 1: Dissolve 1.0 g of 2-mercaptopyridine in 20 mL of dichloromethane solvent, cool to -10-5 degrees Celsius, add 20 mL of concentrated hydrochloric acid-hypochlorous acid (volume ratio 1:1) to carry out sulfonation reaction. After reacting for 10 to 30 minutes, extract the mixture with 100 mL of dichloromethane and dry with 10 g of anhydrous sodium sulfate to obtain the activated intermediate 2-pyridinesulfonyl chloride.

[0035] Step 2: The obtained intermediate 2-pyridinesulfonyl chloride (about 9.0 mmol) was mixed with aniline (1.67 g, 18.0 mmol) and reacted in dry dichloromethane, acetonitrile or tetrahydrofuran solvent for 24 hours;

[0036] Step 3: Add 100-200 mL of dichloromethane to the reaction solution, wash three times with 50 mL of 1.0 mol / L phosphoric acid solution, wash three times with 50 mL of deionized water, dry with 50 g of anhydrous sodium sulfate, and distill under pressure to obtain the intermediate crude product N-phenylpyridine-2-sulfonamide.

[0037] Step 4: The crude intermediate product N-phenylpyridine-2-sulfonamide obtained in Step 3 was separated using a silica gel normal-phase column with dichloromethane / petroleum ether = 1:5 as the mobile phase to obtain 1.79 g (7.67 mmol) of pure N-phenylpyridine-2-sulfonamide, with a yield of 85.2%.

[0038] Step 5: Dissolve the pure ligand N-phenylpyridine-2-sulfonamide (58.5 mg, 0.25 mmol), pentamethylcyclopentadiene dichloroiridium dimer Ir-dimer (100 mg, 0.125 mmol), and ammonium hexafluorophosphate (81.5 mg, 0.5 mmol) obtained in Step 4 in 20 mL of anhydrous ethanol, and reflux at 80 °C for 24 hours under nitrogen protection;

[0039] Step 6: Cool the reaction solution from Step 5 to room temperature, filter, wash three times with 20 mL of ethanol, and collect the solid as pure iridium complex 1a 56.322 mg (0.095 mmol), yield 75.6%.

[0040] The nuclear magnetic resonance (NMR) spectra (H1N, C1N, and mass spectra) of the intermediate product obtained in step 4 are as follows:

[0041] 1 H NMR (400 MHz, CDCl3) δ 8.81 – 8.67 (m, 1H), 8.04 (s, 1H), 7.90 (d, J = 7.8 Hz, 1H), 7.81 (td, J = 7.7, 1.7 Hz, 1H), 7.46 (ddd, J = 7.5, 4.7, 1.0Hz, 1H), 7.21 (d, J = 4.0 Hz, 4H), 7.13 – 7.02 (m, 1H);

[0042] 13 C NMR (101 MHz, CDCl3) δ 156.18 (s), 150.10 (s), 138.05 (s), 136.10 (s), 129.22 (s), 127.00 (s), 125.68 (s), 123.16 (s), 122.69 (s);

[0043] MS (ESI): m / z cacld for ([M+Na + ] + C 11 H 10 N2NaO2S 257.0361; found257.0362.

[0044] The 1H NMR, 1C NMR, and mass spectrometry data of the iridium complex 1a obtained in step 6 are as follows:

[0045] 1 H NMR (600 MHz, DMSO) δ 8.76 (d, J = 5.4 Hz, 1H), 8.27 (td, J = 7.8, 1.1 Hz, 1H), 7.93 (d, J = 7.8 Hz, 1H), 7.85 – 7.78 (m, 1H), 7.56 (d, J = 7.8Hz, 2H), 7.17 (t, J = 7.8 Hz, 2H), 6.91 (t, J = 7.3 Hz, 1H), 1.37 (s, 15H);

[0046] 13 C NMR (151 MHz, DMSO) δ 157.25 (s), 153.81 (s), 145.69 (s), 142.08(s), 129.49 (s), 128.36 (s), 124.71 (s), 122.14 (s), 121.05 (s), 87.35(s),8.55(s);

[0047] MS (ESI): m / z cacld for ([M-Cl - ] + C 21 H 24 IrN2O2S 561.1188; found561.1206.

[0048] Example 2: Preparation method of N-(4-carboxyphenyl)pyridine-2-sulfonamide pentamethylcyclopentadiene (Cp*) chloroiridium complex 1b. The specific steps for preparing iridium complex 1a are basically the same, except that...

[0049] Replace the aniline described in step 2 with 4-carboxyaniline (2.47 g, 18.0 mmol).

[0050] The N-phenylpyridine-2-sulfonamide ligands described in steps 3, 4, and 5 are respectively replaced with N-(4-carboxyphenyl)pyridine-2-sulfonamide;

[0051] The pure ligand N-(4-carboxyphenyl)pyridine-2-sulfonamide obtained in step 4 had a yield of 88.3%.

[0052] The yield of iridium complex 1b described in step 6 was 80.2%.

[0053] The 1H NMR, 1C NMR, and mass spectrometry data of the iridium complex 1b obtained in step 6 are as follows:

[0054] 1 H NMR (600 MHz, DMSO) δ 12.49 (s, 1H), 8.79 (d, J = 5.3 Hz, 1H), 8.29 (td, J = 7.8, 1.3 Hz, 1H), 7.97 (d, J= 7.7 Hz, 1H), 7.86 – 7.83 (m,1H), 7.78 – 7.73 (m, 2H), 7.65 – 7.60 (m, 2H), 1.39 (s, 15H);

[0055] 13 C NMR (151 MHz, DMSO) δ 167.70 (s), 156.89 (s), 153.96 (s), 150.51(s), 142.27 (s), 129.99 (s), 129.83 (s), 123.68 (s), 123.57 (s), 121.40 (s), 87.63 (s), 87.63 (s), 8.61 (s), 8.61 (s);

[0056] MS (ESI): m / z cacld for ([M-Cl - ] + C 22 H 24 IrN2O4S 605.1086; found605.1094.

[0057] Example 3: The preparation method of N-(4-carboxyphenyl)pyridine-2-sulfonamide pentamethylcyclopentadiene (Cp*) chloroiridium complex 1c is basically the same as that for the preparation of iridium complex 1a, except that...

[0058] Replace the aniline mentioned in step 2 with 4-cyanoaniline (2.12 g, 18.0 mmol);

[0059] The N-phenylpyridine-2-sulfonamide ligands mentioned in steps 3, 4, and 5 are respectively replaced with N-(4-cyanophenyl)pyridine-2-sulfonamide;

[0060] The pure ligands N-(4-cyanophenyl)pyridine-2-sulfonamide and N-(4-methoxyphenyl)pyridine-2-sulfonamide obtained in step 4 had a yield of 84.6%.

[0061] The yield of iridium complex 1c described in step 6 was 73.6%.

[0062] The 1C NMR spectrum, 1H NMR spectrum, and mass spectrometry data of the iridium complex obtained in step 6 are as follows:

[0063] 1 H NMR (600 MHz, DMSO) δ 8.80 (d, J= 5.4 Hz, 1H), 8.30 (dd, J =11.3, 4.3 Hz, 1H), 7.99 (d, J = 7.8 Hz, 1H), 7.86 (dd, J = 9.6, 3.5 Hz, 1H), 7.67 (d, J = 8.7 Hz, 2H), 7.63 (d, J = 8.7 Hz, 2H), 1.40 (s, 15H);

[0064] 13 C NMR (151 MHz, DMSO) δ 156.63 (s), 154.04 (s), 150.75 (s), 142.39 (s), 132.74 (s), 130.04 (s), 124.34 (s), 121.57 (s), 120.09 (s), 102.93 (s), 87.80 (s), 8.64 (s);

[0065] MS (ESI): m / z cacld for ([M-Cl - ] + C 23 H 23 IrN3O2S 586.1140; found586.1151.

[0066] Example 4: Preparation method of N-(4-carboxyphenyl)pyridine-2-sulfonamide pentamethylcyclopentadiene (Cp*) chloroiridium complex 1d. The specific steps for preparing iridium complex 1a are basically the same, except that...

[0067] Replace the aniline mentioned in step 2 with 4-methoxyaniline (2.21 g, 18.0 mmol).

[0068] The N-phenylpyridine-2-sulfonamide ligands described in steps 3, 4, and 5 are respectively replaced with N-(4-methoxyphenyl)pyridine-2-sulfonamide;

[0069] The yield of the pure ligand N-(4-methoxyphenyl)pyridine-2-sulfonamide obtained in step 4 was 89.4%.

[0070] The yield of the iridium complex 1d described in step 6 was 68.6%.

[0071] The 1-day proton, carbon, and mass spectrometry data of the iridium complex obtained in step 6 are as follows:

[0072] 1 H NMR (600 MHz, DMSO) δ 8.74 (d, J = 5.4 Hz, 1H), 8.25 (td, J = 7.8, 1.0 Hz, 1H), 7.90 (d, J = 7.8 Hz, 1H), 7.84 – 7.75 (m, 1H), 7.46 (d, J = 8.9Hz, 2H), 6.77 (d, J = 8.9 Hz, 2H), 3.70 (s, 3H), 1.38 (s, 15H);

[0073] 13 C NMR (151 MHz, DMSO) δ 157.26 (s), 155.04 (s), 153.74 (s), 141.99 (s), 138.51 (s), 129.33 (s), 126.05 (s), 120.88 (s), 113.65 (s), 87.27 (s), 55.59 (s), 8.57 (s);

[0074] MS (ESI): m / z cacld for ([M-Cl - ] + C 22 H 26 IrN2O3S 591.1293; found591.1312.

[0075] Example 5: Preparation method of N-(4-carboxyphenyl)pyridine-2-sulfonamide pentamethylcyclopentadiene (Cp*) chloroiridium complex 1e. The specific steps for preparing iridium complex 1a are basically the same, except that...

[0076] Replace the aniline mentioned in step 2 with 4-methylaniline (1.93 18.0 mmol);

[0077] The N-phenylpyridine-2-sulfonamide ligand described in steps 3, 4, and 5 is replaced with N-(4-methylphenyl)pyridine-2-sulfonamide;

[0078] The yield of the pure ligand N-(4-methylphenyl)pyridine-2-sulfonamide obtained in step 4 was 90.1%.

[0079] The yield of the iridium complex 1e described in step 6 was 70.4%.

[0080] The 1H NMR, 1C NMR, and mass spectrometry data of the iridium complex obtained in step 6 are as follows:

[0081] 1 H NMR (600 MHz, DMSO) δ 8.75 (d, J = 5.3 Hz, 1H), 8.26 (td, J = 7.8, 1.2 Hz, 1H), 7.91 (d, J = 7.8 Hz, 1H), 7.82 – 7.78 (m, 1H), 7.44 (d, J = 8.3Hz, 2H), 6.98 (d, J = 8.2 Hz, 2H), 2.23 (s, 3H), 1.38 (s, 15H);

[0082] 13 C NMR (151 MHz, DMSO) δ 157.29 (s), 153.76 (s), 142.95 (s), 142.02(s), 130.93 (s), 129.40 (d, J = 3.9 Hz), 128.81 (s), 124.94 (s), 124.75 (s), 120.95 (s), 87.28 (s), 20.87 (s), 8.57 (s);

[0083] MS (ESI): m / z cacld for ([M-Cl - ] + C 22 H 26 IrN2O2S 575.1344; found575.1366.

[0084] Example 6: Preparation method of N-(4-carboxyphenyl)pyridine-2-sulfonamide pentamethylcyclopentadiene (Cp*) chloroiridium complex 1f. The specific steps for preparing iridium complex 1a are basically the same, except that...

[0085] Replace the aniline mentioned in step 2 with 4-chloroaniline (2.29 g, 18.0 mmol);

[0086] The N-phenylpyridine-2-sulfonamide ligands described in steps 3, 4, and 5 are respectively replaced with N-(4-chlorophenyl)pyridine-2-sulfonamide;

[0087] The yield of the pure ligand N-(4-chlorophenyl)pyridine-2-sulfonamide obtained in step 4 was 87.2%.

[0088] The yield of the iridium complex 1f described in step 6 was 78.4%.

[0089] The 1f NMR (1H NMR), C NMR, and mass spectrometry data of the iridium complex obtained in step 6 are as follows:

[0090] 1 H NMR (600 MHz, DMSO) δ 8.77 (d, J = 5.4 Hz, 1H), 8.28 (td, J = 7.7, 0.8 Hz, 1H), 7.94 (d, J = 7.8 Hz, 1H), 7.82 (t, J = 6.2 Hz, 1H), 7.55 (d, J =8.8 Hz, 2H), 7.23 (d, J = 8.8 Hz, 2H), 1.39 (s, 15H);

[0091] 13 C NMR (151 MHz, DMSO) δ 156.93 (s), 153.88 (s), 144.74 (s), 142.19(s), 129.66 (s), 128.24 (s), 126.09 (s), 125.84 (s), 121.18 (s), 87.49(s),8.61(s);

[0092] MS (ESI): m / z cacld for ([M-Cl - ] + C 21 H 23 ClIrN2O2S 595.0798; found595.0802.

[0093] As can be seen from the above, the method for preparing iridium complexes provided by the present invention has a simple synthetic route, high yield, and is easy to obtain iridium complexes in large quantities.

[0094] The novel pyridine sulfonamide-type iridium complex provided by this invention has the following characteristics and performance for catalyzing the regeneration of coenzyme NADH.

[0095] Example 7: Catalytic activity of the provided iridium complex for the regeneration of coenzyme NADH

[0096] To evaluate the catalytic regeneration performance of the prepared complex 1a-f on the coenzyme NADH, and to compare it with the previously reported highly active 2-pyridinebenzamide-iridium complex 5, NAD... + The reaction was carried out at 37°C for 0.5 hours under the following conditions: HCOONa concentration of 8 mmol / L, HCOONa concentration of 0.1 mol / L, and iridium complex content of 0.1 mol%. (See attached diagram.) Figure 2 As shown, 2-pyridinebenzamide iridium complex 5 and complexes 1a-f shown in Formula 1 all exhibited excellent NADH regeneration performance. The NADH yield of 2-pyridinebenzamide iridium complex 5 was 68.2%, while the NADH yields of 1a-f under the same conditions reached 77.3%, 77.8%, 73.32%, 74.14%, 68.2%, and 50.3%, respectively. The difference in catalytic activity of the iridium complexes with different substitutions can be attributed to the difference in electronegativity of the complexes themselves.

[0097] Example 8: Selectivity of the provided iridium complex for catalyzing the regeneration of coenzyme NADH

[0098] The synergistic effect between chemical catalysts and enzymes is crucial for establishing chemo-enzyme cascade catalytic systems. To achieve this, the catalyst needs to maintain low catalytic activity towards the substrate but provide highly selective catalytic ability for NADH regeneration. Therefore, this invention investigated the direct catalytic hydrogenation performance of iridium complexes 1a-f on various dehydrogenase-catalyzed substrates, including benzaldehyde, trimethyl-p-benzoquinone, and α-ketoglutaric acid, and compared them with complex 5. The reaction was carried out in phosphate buffer (0.2 M, pH = 7.4) at a substrate concentration of 30 mmol / L, an HCOONa concentration of 100 mmol / L, and an iridium complex content of 0.1 mol%, at 37°C for 1 hour. (See attached...) Figure 3 As shown in Figure A, the hydrogenation performance of 2-pyridinebenzamide iridium complex 5 on three different substrates is significantly higher than that of the iridium complex provided in this invention. The yield of benzyl alcohol prepared by hydrogenation of benzaldehyde by 2-pyridinebenzamide iridium complex 5 is as high as 98.5%, while the highest yield of benzyl alcohol prepared by hydrogenation of benzene by the iridium complex provided in this invention is only 57.9%. With the same substituents, the benzyl alcohol yield of iridium complex 1a provided in this invention is only 38.2%. Under the same conditions, the yield of hydroquinone TMHQ prepared by hydrogenation of trimethyl-p-benzoquinone (TMBQ) by 2-pyridinebenzamide iridium complex 5 reaches 61.5% (see Appendix). Figure 3 B), while the iridium complexes 1a-f provided by this invention all have a TMHQ yield of less than 30.0%. Further, as... Figure 3As shown in C, the 2-pyridinebenzamide iridium complex 5 catalyzes the preparation of L-glutamic acid from α-ketoglutarate with a yield of 1.0%, while the iridium complex 1a-f provided by the present invention hardly produces L-glutamic acid.

[0099] These results indicate that, compared with 2-pyridinebenzamide iridium complex 5, the iridium complex provided by this invention has better selectivity for catalyzing the regeneration of coenzyme NADH and is more suitable for the construction of efficient chemical-enzyme cascade catalytic systems.

[0100] Example 9: Biocompatibility of the provided iridium complex

[0101] Establishing a robust chemo-enzyme cascade catalytic system requires biocompatibility of the chemical catalyst; therefore, this invention investigates the compatibility of the prepared iridium complex with various biomolecules. The compatibility of the invented iridium complex 1a with NAD+ was examined in the presence of various biomolecules (including amino acids, nucleobases, proteins, glucose, and salts). + Catalytic performance of hydrogenation. In a phosphate buffer solution (0.2 M, pH = 7.4), containing a certain concentration of various biomolecules, NAD... + The reaction was carried out at 37°C for 0.5 hours under the following conditions: HCOONa concentration of 8 mmol / L, HCOONa concentration of 0.1 mol / L, and iridium complex content of 0.1 mol%. (See attached diagram.) Figure 4 As shown, adenine (Ade), glucose (GLu), sodium acetate (AcONa), oxidized glutathione (GSSG), and L-glutamate dehydrogenase (GLDH) have almost no effect on the catalytic NADH regeneration performance of iridium complex 1a. However, biomolecules containing free sulfhydryl groups, such as reduced glutathione (GSH), cysteine ​​(Cys), captopril (CAP), bovine serum albumin (BSA), and dehydrogenase (ADH), significantly reduce the activity of the iridium complex.

[0102] These results are similar to those of previously reported iridium complexes, indicating that the iridium complexes provided by this invention have good compatibility with most biomolecules, but poor inhibition of thiol groups.

[0103] Application of the novel pyridine sulfonamide-type iridium complex provided by this invention in a chemical-bioenzyme synergistic catalytic system

[0104] Example 9: The provided iridium complex and L-glutamate dehydrogenase GLDH co-catalyze the preparation of L-glutamate dehydrogenase.

[0105] Based on the high activity and selectivity of the provided iridium complex for catalytic NADH regeneration, this invention combines the iridium complex 1a-f with L-glutamate dehydrogenase GLDH to construct a highly efficient chemical-enzyme cascade catalytic system for the production of L-glutamate. The system is prepared in phosphate buffer solution (0.2 M, pH = 7.4) containing 10.0 U GLDH and 1.0 mmol / L NAD. + The reaction was carried out at 37°C for 24 hours under the following conditions: 0.1 mol / L HCOONa, 50.0 mmol / L (NH4)2SO4, and an iridium complex content of 0.1 mol%. (See attached image.) Figure 5 As shown, when only iridium complexes are used to catalyze the hydrogenation of α-ketoglutarate to prepare L-glutamic acid, the yield of L-glutamic acid is less than 5.0%. However, under the same conditions, the yield of L-glutamic acid is significantly increased to 50.0% when using a cascade catalytic system composed of iridium complexes and GLDH. Among them, the cascade catalytic system composed of complex 1e shows the greatest improvement in the yield of L-glutamic acid, increasing it from 0.66% to 62.8%.

[0106] The above results indicate that the iridium complex provided by this invention has good NADH regeneration activity, good biocompatibility, and efficient hydrogenation ability of α-ketoglutarate. It can also be combined with L-glutamate dehydrogenase GLDH to construct an excellent chemical-enzyme cascade catalytic system for the efficient preparation of L-glutamate.

Claims

1. A pyridinesulfonamide iridium complex, characterized in that, The structural formula of the pyridine sulfonamide iridium complex is as follows: ； Formula 1, Where X = H, -COOH, -CN, -OCH3, -CH3, Cl.

2. The method for preparing the pyridinesulfonamide iridium complex according to claim 1, characterized in that, The preparation steps of pyridinebenzenesulfonamide type iridium complex are as follows: Dissolve 1.0 g of 9.0 mmol of 2-mercaptopyridine in 10-50 mL of dichloromethane solvent, cool to -10-5°C, and add 20-100 mL of concentrated hydrochloric acid to hypochlorous acid in a ratio of 1:1-10 or concentrated sulfuric acid to hypochlorous acid in a ratio of 1:1-10 to carry out a sulfonation reaction. After reacting for 10 to 30 minutes, the mixture is extracted with 50-100 mL of dichloromethane and dried with 10-20 g of anhydrous sodium sulfate to obtain the activated intermediate 2-pyridinesulfonyl chloride. The obtained intermediate 2-pyridinesulfonyl chloride was mixed with various substituted aniline compounds and reacted in dry dichloromethane, acetonitrile, or tetrahydrofuran solvent for 10 to 30 hours. Add 100-200 mL of dichloromethane to the reaction solution, wash three times with 50 mL of 1.0 mol / L phosphoric acid solution, wash three times with 50 mL of deionized water, dry with 20-50 g of anhydrous sodium sulfate, and distill under reduced pressure to obtain the crude product 2-pyridinesulfonamide ligand. The crude 2-pyridine sulfonamide ligand was separated using a silica gel normal-phase column with dichloromethane / petroleum ether = 1-10:0.1-10 as the mobile phase to obtain pure 2-pyridine sulfonamide ligands in yields of 70%–90%. 0.25 mmol of pure 2-pyridinesulfonamide ligand, 0.125 mmol of [Cp IrCl2]2 Ir-dimer and 0.5 mmol ammonium hexafluorophosphate were dissolved in 10-20 mL of anhydrous ethanol and refluxed at 80°C for 10 to 24 hours under nitrogen protection. Cool the reaction solution from step (5) to room temperature, filter, wash three times with 20 mL of ethanol, and collect the solid, which is the pure pyridinebenzenesulfonamide type iridium complex shown in Formula 1.

3. The application of the pyridine sulfonamide iridium complex according to claim 1 in the highly selective catalytic regeneration of NADH, characterized in that, The regeneration conditions for coenzyme NADH by the iridium complex are as follows: (1) Dissolve 1.5 mmol of oxidized coenzyme nicotinamide adenine dinucleotide (NAD+) in 5.0-20.0 mL of deionized water, then add 15 mmol of formic acid or sodium formate or ammonium formate or potassium formate or calcium formate or zinc formate, and 1.5-150 μmol of the iridium complex, and then react the mixture at 0-50 degrees Celsius for 1-20 hours; (2) After the reaction is complete, add 5.0~20.0 mL of acetonitrile to the reaction solution to precipitate the product, and then filter to obtain the crude product; (3) The crude product was redissolved in 2.0-5.0 mL of deionized water and further separated and purified by reversed-phase column chromatography. During purification, 0.05 M triethylammonium bicarbonate TEAB buffer containing 0-30% acetonitrile was used as the eluent. The crude product was collected and the residual TEAB was removed by vacuum distillation with methanol. The obtained product was further dissolved in ultrapure water and lyophilized to obtain pure target NADH with a yield of 60-85%.

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