Dual hydrocarbon activation strategy for synthesis of novel non-natural amino acids and polypeptides

Abstract

The invention discloses application of a dual hydrocarbon activation strategy to synthesis of novel non-natural amino acids and polypeptides, belonging to the technical field of organic synthesis. The method comprises the following steps: by taking a lysine derivative and acrolein as starting substrates, reacting for 2 hours at the temperature of 40 DEG C under the action of a rhodium catalyst to obtain a series of non-natural amino acids and polypeptides. The method has the advantages that reaction conditions are mild, raw materials are easy to prepare, only one-step reaction is needed, and experimental operation is simple; antibacterial experiments show that the novel non-natural amino acids and polypeptides have good antibacterial activity on fungi in agriculture and forestry. # imgabs0 #

Description

Technical Field

[0001] The invention belongs to the technical field of organic synthesis, and in particular relates to a dual carbon-hydrogen activation strategy for the synthesis of novel non-natural amino acids and polypeptides. Background Art

[0002] Due to the advantages of high efficiency and specificity, peptides are becoming important drugs. Since natural peptides face problems of metabolic instability and low membrane permeability, and non-natural peptides often enhance biological activity and improve drug metabolism kinetics, the development of effective peptide late modification methods plays an important role in medicinal chemistry and chemical biology. Among them, chemical and position-selective peptide late modification is particularly important, because it can achieve precise regulation of the biochemical and physicochemical properties of peptides by fine-tuning the peptide structure. At present, the methods of peptide late modification are mainly limited to one-step coupling reactions, which limits the complexity and diversity of peptides. For example, in 2014, Yu Jinquan et al. used palladium-catalyzed C(sp 3 )-H activation strategy was used for the arylation reaction of alanine in peptides (J.Am.Chem.Soc.2014,136,16940-16946); in 2017, Ackermann et al. used ruthenium-catalyzed C(sp 2 )-H activation strategy was used for the arylation reaction of tryptophan in peptides (Angew.Chem.Int.Ed.2017,56,1576-1580); in 2018, Wang Huan et al. used palladium-catalyzed C(sp 2 The )-H activation strategy is used for the alkenylation reaction of phenylalanine in polypeptides (Angew. Chem. Int. Ed. 2018, 57, 13912-13916). Therefore, it is of great significance to develop complex tandem reactions for the efficient synthesis of new non-natural polypeptides. Summary of the invention

[0003] Purpose of the invention: In view of the above technical problems existing in the prior art, the present invention aims to provide a dual carbon-hydrogen activation strategy for the synthesis of novel non-natural amino acids and peptides, using readily available lysine derivatives and acrolein to synthesize non-natural amino acids and peptides in one step.

[0004] Technical solution: To achieve the above purpose, the technical solution adopted by the present invention is:

[0005] The dual carbon-hydrogen activation strategy is used for the synthesis of novel non-natural amino acids and peptides, wherein the novel non-natural amino acids and peptides are compounds having the following structural formula:

[0006]

[0007] Among them, R 1One selected from Boc, Fmoc, aspartic acid, serine and naproxen, R 2 One selected from hydrogen, methyl and ethyl. The method is characterized in that lysine derivatives, acrolein, rhodium catalyst, silver hexafluoroantimonate and 2,4,6-trimethylbenzoic acid are dissolved in an organic solvent and reacted at 40°C for 2 hours. After the reaction is completed, the reaction system is separated and purified to obtain novel non-natural amino acids and polypeptides. The reaction formula is as follows:

[0008]

[0009] The dual carbon-hydrogen activation strategy is used for the synthesis of novel non-natural amino acids and peptides, the molar ratio of lysine derivatives to acrolein is 1:3, the rhodium catalyst is a pentamethylcyclopentadienyl rhodium catalyst, and the molar ratio of lysine derivatives to rhodium catalyst is 1:0.05.

[0010] The dual carbon-hydrogen activation strategy is used for the synthesis of novel non-natural amino acids and polypeptides. The steps of separation and purification of the reaction system are as follows: dichloromethane is added to the reaction system for dilution, followed by concentration under reduced pressure, and the obtained concentrate is separated and purified by column chromatography on silica gel, the eluent is a mixture of petroleum ether and ethyl acetate in sequence, the eluent is collected and the solvent is evaporated to obtain the target product.

[0011] Compared with the prior art, the present invention has the following beneficial effects:

[0012] (1) The synthesis method of the present invention has mild reaction conditions, readily available raw materials, simple experimental operation, and can produce non-natural amino acids and polypeptides in one step.

[0013] (2) The obtained non-natural amino acids and peptides have good antibacterial activity against agricultural and forestry fungi. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 The NMR spectrum of compound 3a prepared in Example;

[0015] Figure 2 The NMR spectrum of compound 3b prepared in Example;

[0016] Figure 3 The NMR spectrum of compound 3c prepared in Example;

[0017] Figure 4 The NMR spectrum of compound 3d prepared in Example;

[0018] Figure 5 This is the NMR spectrum of compound 3e prepared in Example;

[0019] Figure 6The NMR spectrum of compound 3f prepared in Example; Figure 7 Attached is the abstract. DETAILED DESCRIPTION

[0020] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific implementation methods of the present invention are described in detail below in conjunction with the embodiments of the specification.

[0021] Example 1

[0022] The reaction equation is as follows:

[0023]

[0024] Lysine derivative 1a (0.1mmol, 36.4mg), acrolein 2 (0.3mmol, 20μL), [RhCp*Cl2]2 (0.005mmol, 3.1mg), silver hexafluoroantimonate (0.02mmol, 6.9mg), 2,4,6-trimethylbenzoic acid (0.2mmol, 32.8mg) and 1,2-dichloroethane (1mL) were added to a 10mL pressure tube equipped with a magnetic bar and reacted at 40°C for 2h. The reaction mixture was monitored by TLC until the raw material 1a was consumed. After the reaction was completed, the reaction was diluted with dichloromethane, concentrated in vacuo, and purified by column chromatography to obtain product 3a (33.0mg, 72%).

[0025] Characterization data: 1 Ⅱ NMR (400MHz, CDCl3) δ9.78 (s, 1H), 7.25-7.19 (m, 1H), 7.11 (dd, J = 7.7, 2.2Hz, 1H), 6.92 (dd, J = 7.3, 2.3Hz, 1H), 5.88 (dd, J = 7.3, 2.4Hz, 1H), 5.78-5.69 (m, 1H), 5.15(s, 1H), 4.28(s, 1H), 4.09(s, 1H), 3.72(s, 3H), 3.27(s, 2H), 3.09-2 .88(m, 4H), 2.69(s, 1H), 1.85(s, 1H), 1.75-1.64(m, 3H), 1.44-1.38(m, 11H). 13 C NMR (101MHz, CDCl3) δ202.0, 173.1, 169.2, 155.3, 144.9, 141.7, 133.0, 129.8, 129.1, 128.6, 123.8, 121.1, 79.7, 53.2, 52.2, 47.6, 46.0, 32.3, 31.4, 28.2, 28.0, 27.0, 22.5.

[0026] Embodiment 2:

[0027] The reaction equation is as follows:

[0028]

[0029] Lysine derivative 1b (0.1mmol, 37.8mg), acrolein 2 (0.3mmol, 20μL), [RhCp*Cl2]2 (0.005mmol, 3.1mg), silver hexafluoroantimonate (0.02mmol, 6.9mg), 2,4,6-trimethylbenzoic acid (0.2mmol, 32.8mg) and 1,2-dichloroethane (1mL) were added to a 10mL pressure tube equipped with a magnetic bar and reacted at 40°C for 2h. The reaction mixture was monitored by TLC until the raw material 1b was consumed. After the reaction was completed, the reaction was diluted with dichloromethane, concentrated in vacuo, and purified by column chromatography to obtain product 3b (35.4mg, 75%).

[0030] Characterization data: 1 H NMR (400MHz, CDCl3) δ9.81 (s, 1H), 6.94 (s, 1H), 6.76 (s, 1H), 5.89 (d, J = 7.4Hz, 1H), 5.74 (q, J = 7.4Hz, 1H), 5.13 (s, 1H), 4.30 (s, 1H), 4.11 (s, 1H) , 3.75 (s, 3H), 3.26 (s, 2H), 3.09-2.87 (m, 4H), 2.68 (d, J=17.5Hz, 1H), 2 .30 (s, 3H), 1.86 (s, 1H), 1.70 (q, J=8.1, 7.6Hz, 3H), 1.47-1.39 (m, 11H). 13 C NMR (101MHz, CDCl3) δ202.2, 173.2, 169.2, 155.4, 144.9, 141.8, 140.1, 130.2, 129.5, 129 .3, 124.7, 120.9, 79.8, 53.3, 52.2, 48.0, 46.2, 32.2, 31.5, 28.2, 28.1, 27.1, 22.5, 21.0.

[0031] Embodiment 3:

[0032] The reaction equation is as follows:

[0033]

[0034] Lysine derivative 1c (0.1mmol, 51.4mg), acrolein 2 (0.3mmol, 20μL), [RhCp*Cl2]2 (0.005mmol, 3.1mg), silver hexafluoroantimonate (0.02mmol, 6.9mg), 2,4,6-trimethylbenzoic acid (0.2mmol, 32.8mg) and 1,2-dichloroethane (1mL) were added to a 10mL pressure tube equipped with a magnetic bar and reacted at 40°C for 2h. The reaction mixture was monitored by TLC until the raw material 1c was consumed. After the reaction was completed, the reaction was diluted with dichloromethane, concentrated in vacuo, and purified by column chromatography to obtain the product 3c (39.8mg, 68%).

[0035] Characterization data: 1 H NMR (400MHz, CDCl3) δ9.80 (s, 1H), 7.77 (d, J = 7.6Hz, 2H), 7.60 (d, J = 7.5Hz, 2H), 7.41 (t, J = 7.8Hz, 2H), 7.32 (d, J=7.7Hz, 2H), 6.95 (s, 1H), 6.74 (d, J=8.8Hz, 1H), 5.89 (d, J=7.4Hz, 1H), 5.75 (t, J=7.7Hz, 1H), 5.67-5.48 (m, 1H ), 4.40 (q, J = 6.1Hz, 3H), 4.16 (d, J = 36.9Hz, 2H), 3.77 (s, 3H), 3.34 (d, J = 2.8Hz, 2H), 3.29 (s, 1H), 3.10-2.89 (m, 3H), 2.58 (q, J=7.7Hz, 3H), 1.94 (s, 1H), 1.80-1.63 (m, 3H), 1.48 (dd, J=16.5, 8.2Hz, 2H), 1.22 (d, J=7.7Hz, 3H). 13 C NMR (151MHz, CDCl3) δ202.3, 172.9, 169.3, 156.0, 146.3, 146.0, 145.0, 143.8, 143.7, 142.9, 141.8, 141.2, 130.4, 129.2, 128.4, 128.3, 127.6, 127.0, 125.1, 123.4, 121.0, 119.89, 119.87, 104.1, 66.9, 53.7, 52.4, 47.1, 46.1, 34.2, 31.5, 29.4, 28.3, 28.0, 27.1, 22.4, 15.0.

[0036] Embodiment 4:

[0037] The reaction equation is as follows:

[0038]

[0039] Lysine derivative 1d (0.1mmol, 60.6mg), acrolein 2 (0.3mmol, 20μL), [RhCp*Cl2]2 (0.005mmol, 3.1mg), silver hexafluoroantimonate (0.02mmol, 6.9mg), 2,4,6-trimethylbenzoic acid (0.2mmol, 32.8mg) and 1,2-dichloroethane (1mL) were added to a 10mL pressure tube equipped with a magnetic bar and reacted at 40°C for 2h. The reaction mixture was monitored by TLC until the raw material 1d was consumed. After the reaction was completed, the reaction was diluted with dichloromethane, concentrated in vacuo, and purified by column chromatography to obtain product 3d (41.5mg, 59%).

[0040] Characterization data: 1 H NMR (400MHz, CDCl3) δ9.81 (s, 1H), 7.24 (t, J = 7.9Hz, 1H), 7.13 (d, J = 7.5Hz, 2H), 6.94 (d, J =7.6Hz, 1H), 5.90 (d, J = 7.3Hz, 1H), 5.77 (t, J = 7.6Hz, 2H), 4.60-4.47 (m, 2H), 4.15-4.00 (m , 1H), 3.74 (s, 3H), 3.28 (q, J=19.2, 16.2Hz, 2H), 3.12-2.84 (m, 5H), 2.72 (q, J=12.6Hz, 1H ), 2.61 (dd, J=17.3, 6.2Hz, 1H), 1.99-1.83 (m, 2H), 1.80-1.61 (m, 4H), 1.48-1.44 (m, 18H). 13 C NMR (101MHz, CDCl3) δ202.1, 172.3, 171.3, 170.8, 169.2, 155.5, 145.0, 141.8, 133.1, 130.0, 129.2, 128. 7, 123.9, 121.2, 81.7, 80.3, 52.4, 52.2, 50.6, 47.7, 46.2, 37.4, 31.9, 31.5, 28.3, 28.0, 27.2, 22.7, 22.5.

[0041] Embodiment 5:

[0042] The reaction equation is as follows:

[0043]

[0044] Lysine derivative 1e (0.1mmol, 54.2mg), acrolein 2 (0.3mmol, 20μL), [RhCp*Cl2]2 (0.005mmol, 3.1mg), silver hexafluoroantimonate (0.02mmol, 6.9mg), 2,4,6-trimethylbenzoic acid (0.2mmol, 32.8mg) and 1,2-dichloroethane (1mL) were added to a 10mL pressure tube equipped with a magnetic bar and reacted at 40°C for 2h. The reaction mixture was monitored by TLC until the raw material 1e was consumed. After the reaction was completed, the reaction was diluted with dichloromethane, concentrated in vacuo, and purified by column chromatography to obtain the product 3e (36.0mg, 57%).

[0045] Characterization data: 1 H NMR (400MHz, CDCl3) δ9.79 (s, 1H), 7.33 (d, J = 8.6Hz, 5H), 7.27-7.21 (m, 1H), 7.13 (d, J = 7.7Hz, 2H), 6.94 (d, J=7.5Hz, 1H), 5.85 (d, J=7.5Hz, 1H), 5.80-5.70 (m, 1H), 5.47 (s, 1H), 4.61 (d, J=7.9Hz, 1H), 4.57 (s, 2H), 4.32 (s, 1H), 4.04 (s, 1H), 3.92 (s, 1H), 3.74 (s, 3H), 3.62-3.56 (m, 1H), 3.23 (d, J=30.9Hz, 2H), 3.0 1(t,J=23.2Hz, 4H), 2.71(s, 1H), 1.92(s, 1H), 1.67(s, 3H), 1.47(d,J=2.7Hz, 9H), 1.38(d,J=7.8Hz, 2H). 13 CNMR (101MHz, CDCl3) δ202.1, 172.3, 170.2, 169.2, 145.0, 141.7, 137.4, 133.0, 129.9, 129.2, 128.7, 128.4, 1 27.9, 127.7, 123.9, 121.2, 80.2, 73.4, 69.9, 53.8, 52.3, 52.2, 47.5, 46.1, 32.0, 31.4, 28.2, 28.0, 27.1, 22.4.

[0046] Embodiment 6:

[0047] The reaction equation is as follows:

[0048]

[0049] Lysine derivative 1f (0.1mmol, 47.6mg), acrolein 2 (0.3mmol, 20μL), [RhCp*Cl2]2 (0.005mmol, 3.1mg), silver hexafluoroantimonate (0.02mmol, 6.9mg), 2,4,6-trimethylbenzoic acid (0.2mmol, 32.8mg) and 1,2-dichloroethane (1mL) were added to a 10mL pressure tube equipped with a magnetic bar and reacted at 40°C for 2h. The reaction mixture was monitored by TLC until the raw material 1f was consumed. After the reaction was completed, the reaction was diluted with dichloromethane, concentrated in vacuo, and purified by column chromatography to obtain the product 3f (42.8mg, 75%).

[0050] Characterization data: 1 H NMR (400MHz, CDCl3) δ9.78 (s, 1H), 7.69 (dd, J = 20.3, 10.7Hz, 3H), 7.38 (d, J = 7.7Hz, 1H), 7.25 (d, J = 7.8Hz, 1H), 7.15 (d, J=15.9Hz, 3H), 6.95 (d, J=7.5Hz, 1H), 6.29-6.06 (m, 1H), 5.86 (d, J=7.5Hz, 1H), 5.76 (d, J=7.6Hz, 1H), 4.56 (s, 1 H), 4.10 (d, J=40.0Hz, 1H), 3.93 (s, 3H), 3.75 (d, J=10.7Hz, 1H), 3.64 (s, 3H), 3.25 (d, J=15.2Hz, 2H), 3.10 (d, J=10. 9Hz, 1H), 3.05-2.86 (m, 3H), 2.70 (s, 1H), 1.88 (s, 1H), 1.73-1.57 (m, 4H), 1.52 (d, J=6.6Hz, 1H), 1.40-1.24 (m, 3H). 13 C NMR (101MHz, CDCl3) δ202.0, 174.2, 172.6, 169.4, 157.6, 145.0, 141.7, 136.1, 133.7, 133.1, 130.0, 129.2, 129.0, 128.9, 128.7, 127.3, 126.2, 126.1, 123.9, 121.5, 119.0, 105.6, 55.3, 52.2, 47.4, 46.7, 46.0, 31.4, 29.3, 27.9, 27.1, 22.4, 18.5.

[0051] Embodiment 6:

[0052] The compounds 3c-3f prepared in Examples 3-6 were subjected to biological activity test experiments, and the specific process is as follows:

[0053] The plant fungi used in this experiment are strains preserved at 4℃ in the laboratory, including rapeseed sclerotinia, apple rot, golden shell cystspore and sugarcane pineapple pathogen. The culture medium used is potato agar glucose medium (PDA for short). PDA culture medium formula: 200g potato (peeled), 20g glucose, 15g agar, 1000mL distilled water, preparation method: wash and peel the potatoes, weigh 200g and cut into small pieces, add water and boil (boil for 20-30 minutes, can be pierced by a glass rod), filter into a beaker with eight layers of gauze, add 15-20g agar according to experimental needs, add 20g glucose, stir evenly, cool slightly after fully dissolved, add water to 1000mL, sterilize at 121℃ for 15 minutes after packaging, cool and set aside.

[0054] Experimental method: Growth rate method was used.

[0055] (1) First, culture the two plant fungi on a PDA plate at 25°C for about 3-6 days for later use;

[0056] (2) Heat and melt the PDA medium, cool to 45-50°C, add 250 μL of 10 g / L test compound 3c3f to prepare a medium containing 50 mg / L drug solution, and pour into culture dishes to cool. Boscalid is used as a positive control.

[0057] (3) Using sterile techniques, use a cork puncher to punch a circular bacterial cake (0.50 cm in diameter) at the edge of the hyphae of each strain after 6 days of culture (the growth conditions should be as consistent as possible), then use an inoculation needle to pick it up to the center of the drug-containing plate, and then place the culture dish upside down in an incubator (28°C) for culture;

[0058] (4) Observe and measure the growth of mycelium at different times after treatment, measure the diameter using the cross method, process the data, and calculate the inhibition rate;

[0059] (5) Inhibition rate (%) = (control mycelium diameter - treated mycelium diameter) / (control mycelium diameter - 0.5) × 100;

[0060] (6) Each treatment was repeated 3 times.

[0061] The test results are shown in Table 1.

[0062] Table 1 Test results of the inhibitory activity of novel non-natural amino acids and peptides against agricultural and forestry pathogenic fungi

[0063]

[0064] The results of the bactericidal activity determination of the experimental groups 3c-3f and the control agent boscalid are shown in Table 1. From the results in Table 1, it can be seen that at a concentration of 50 mg / L, compounds 3c-3f showed different degrees of antibacterial activity against four plant fungi, among which compounds 3d and 3e could effectively inhibit the growth of rapeseed sclerotinia, with inhibition rates of 43.2% and 41.3%, respectively; 3f could effectively inhibit the growth of apple rot pathogen and golden shell cystspore, with inhibition rates of 42.5% and 51.2%, respectively.

[0065] The novel non-natural amino acids and polypeptides of the present invention have obvious structural differences and distinct chemical structure characteristics, and show good effects in preventing and controlling rapeseed sclerotinia and golden shell cystspores; and can be used to prevent and control fungal diseases of agricultural or forestry plants. The preparation method of the compound is simple, the yield is high, and the product has stable properties.

Claims

1. The dual carbon-hydrogen activation strategy is used for the synthesis of novel unnatural amino acids and peptides, characterized by Lysine derivatives, acrolein, rhodium catalyst, silver hexafluoroantimonate, and 2,4,6-trimethylbenzoic acid were dissolved in an organic solvent and reacted at 40°C for 2 hours. After the reaction, the reaction system was separated and purified to obtain novel non-natural amino acids and peptides.

2. The preparation method according to claim 1, characterized in that The novel non-natural amino acids and polypeptides are compounds having the following structural formula: Among them, R 1 One selected from Boc, Fmoc, aspartic acid, serine and naproxen, R 2 One selected from the group consisting of hydrogen, methyl and ethyl.

3. The preparation method according to claim 1, characterized in that The molar ratio of the lysine derivative to the acrolein is 1:

3.

4. The preparation method according to claim 1, characterized in that The molar ratio of the lysine derivative to the rhodium catalyst is 1:0.

05.

5. The preparation method according to claim 1, characterized in that The rhodium catalyst is a pentamethylcyclopentadienyl rhodium catalyst.

6. The preparation method according to claim 1, characterized in that The steps of separating and purifying the reaction system are as follows: dichloromethane is added to the reaction system for dilution, followed by concentration under reduced pressure, and the obtained concentrate is separated and purified by column chromatography on silica gel, the eluent is a mixture of n-hexane and ethyl acetate in sequence, the eluent is collected and the solvent is evaporated to obtain novel non-natural amino acids and polypeptides.

7. The preparation method according to claim 1, characterized in that The obtained non-natural amino acids and peptides have good antibacterial activity against agricultural and forestry fungi.

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