Cycloastragenol derivative with activity of resisting methicillin-resistant staphylococcus aureus and preparation method of cycloastragenol derivative

Abstract

The invention discloses cycloastragenol derivatives with activity of resisting methicillin-resistant staphylococcus aureus and a preparation method of the cycloastragenol derivatives. According to the invention, cycloastragenol is used as a matrix, and is optimized by a chemical synthesis means to obtain a series of novel cycloastragenol derivatives. Experiments for resisting methicillin-resistant staphylococcus aureus prove that the series of compounds have good activity for resisting methicillin-resistant staphylococcus aureus, can be used as active ingredients of antibacterial drugs, and have relatively wide application in development of novel antibiotics.

Description

Technical Field

[0001] The present invention belongs to the field of medicine, specifically cycloastragenol derivatives with antibacterial activity and their preparation methods, and specifically relates to the design and synthesis of cycloastragenol derivatives and the application of the obtained cycloastragenol derivatives in the preparation of antibacterial drugs. Background Art

[0002] Staphylococcus aureus belongs to the genus Staphylococcus and is a Gram-positive bacterium. Patients infected with Staphylococcus aureus will experience symptoms such as nausea, vomiting, spastic pain in the upper and middle abdomen, and diarrhea. Among them, vomiting is the most prominent and is an important pathogen causing animal infections. In recent years, due to the abuse of antibiotics, more and more bacteria have evolved from drug-sensitive strains to superbugs with resistance to multiple antibiotics. Therefore, there is an urgent need to develop new antibacterial drugs to solve the infection of multi-drug resistant bacteria.

[0003] Astragalus membranaceus, as a traditional Chinese herbal medicine, has the effects of enhancing the body's immune function, strengthening the heart and reducing blood pressure, lowering blood sugar, diuresis, anti-aging, and anti-fatigue. Cycloastragenol is the aglycone of astragaloside IV and belongs to tetracyclic triterpenoid compounds. It is the only known telomerase activator discovered so far and can also delay cell aging, with anti-inflammatory, antioxidant, antibacterial and other effects. Compared with astragaloside IV, cycloastragenol has higher liposolubility and enhanced membrane penetration ability. However, cycloastragenol has poor water solubility and very poor metabolic stability. By structurally modifying cycloastragenol, its solubility and pharmacokinetic properties can be improved. The presence of multiple hydroxyl groups in the structure of cycloastragenol provides convenience for its structural modification. By modifying its hydroxyl groups, its metabolic stability may be improved. At the same time, introducing functional groups that can form salts, such as amino groups, will greatly improve the water solubility of its derivatives. Summary of the Invention

[0004] The present invention provides a series of cycloastragenol derivatives with anti-methicillin-resistant Staphylococcus aureus activity and their preparation methods. The series of cycloastragenol derivatives with anti-methicillin-resistant Staphylococcus aureus activity provided by the present invention are compounds with structural formulas A1 - A7 and B1 - B7.

[0005]

[0006]

[0007] The above cycloastragenol derivatives are prepared through the following steps:

[0008] Preparation method of compounds A1 - A7:

[0009] 1) Dissolve cyclococcidiol in dichloromethane, add the corresponding amino acid, a condensing agent, and 4-dimethylaminopyridine at room temperature. After reacting at room temperature for 24 hours, add saturated ammonium chloride aqueous solution to the reaction solution, extract with dichloromethane, and concentrate to obtain a crude product.

[0010] 2) The crude product is purified by silica gel column to obtain the condensation product.

[0011] 3) Dissolve the condensation product in an organic solvent, add a hydrogen chloride to obtain an organic solvent solution, stir at room temperature until white solid is produced, and filter to obtain the desired target products A1 - A7.

[0012] The amino acids described in step 1) of the above method are: N-tert-butoxycarbonyl-L-phenylalanine, (2S,3S)-2-((tert-butoxycarbonyl)amino)-3-methylpentanoic acid, (S)-2-((tert-butoxycarbonyl)amino)propanoic acid, N-tert-butoxycarbonyl-glycine, tert-butoxycarbonyl-β-alanine, N-tert-butoxycarbonyl-L-methionine, tert-butoxycarbonyl-D-leucine.

[0013] The silica gel described in step 2) of the above method is 300 - 400 mesh silica gel, and the mobile phase is dichloromethane and methanol, and the ratio of the two is dichloromethane / methanol = 20 / 1;

[0014] The hydrogen chloride organic solvent solution described in step 3) of the above method is hydrogen chloride - methanol, hydrogen chloride - dioxane, hydrogen chloride - ethyl acetate, and preferably hydrogen chloride - dioxane.

[0015] Preparation method of compounds B1 - B7:

[0016] 1) Dissolve cyclococcidiol in dichloromethane, add the corresponding organic acid, a condensing agent, and 4-dimethylaminopyridine at room temperature. After reacting at room temperature for 24 hours, add saturated ammonium chloride aqueous solution to the reaction solution, extract with dichloromethane, and concentrate to obtain a crude product.

[0017] 2) The crude product is purified by silica gel column to obtain the condensation product.

[0018] The organic acids described in step 1) of the above method are: 4-phenoxybenzoic acid, o-phenylbenzoic acid, 2,2,3,3-tetramethylcyclopropanecarboxylic acid, 2-(4-isobutylphenyl)propanoic acid, m-fluorobenzoic acid, m-methylbenzoic acid, o-methylbenzoic acid; a condensing agent is dicyclohexylcarbodiimide, diisopropylcarbodiimide or 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide, and preferably diisopropylcarbodiimide.

[0019] The silica gel described in step 2) of the above method is 300 - 400 mesh silica gel, and the mobile phase is dichloromethane and methanol, and the ratio of the two is dichloromethane / methanol = 20 / 1;

[0020] By structurally modifying cyclocophorol, a series of cyclocophorol derivatives have been obtained. The antibacterial activity of the obtained cyclocophorol derivatives has been greatly improved compared with that of cyclocophorol. The present invention provides a series of cyclocophorol derivatives with antibacterial activity and provides a preparation method thereof. The obtained cyclocophorol derivatives have been confirmed to have good antibacterial activity through experiments against methicillin-resistant Staphylococcus aureus and can be used as active ingredients of antibacterial drugs, showing broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 It is the test result of the antibacterial activity of cyclocophorol derivatives;

[0022] Figure 2 It is the hydrogen spectrum of cyclocophorol derivatives. DETAILED DESCRIPTION OF THE INVENTION

[0023] Example 1:

[0024] Synthesize compound A1: Dissolve cyclocophorol (200 mg) in dichloromethane (4 mL), add N-Boc-L-phenylalanine (212 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract three times with dichloromethane (30 mL * 3), combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. Dissolve the obtained compound in ethyl acetate (5 mL), add 5 mL of hydrogen chloride-dioxane, stir at room temperature for 2 hours and then filter to collect the filter cake to obtain the target product A1 217.3 mg (yield = 80.50%): 1 H NMR (400 MHz, Methanol-d 4)δ 7.39 - 7.36 (m, 2H), 7.34 - 7.30 (m, 3H), 4.73 - 4.63 (m, 2H), 4.38 - 4.33 (m, 1H), 3.78 - 3.74 (m, 1H), 3.49 - 3.42 (m, 1H), 3.38 - 3.34 (m, 1H), 3.21 (s, 1H), 3.14 (br.s, 1H), 2.65 - 2.57 (m, 1H), 2.36 (d, J = 7.76 Hz, 1H), 2.13 - 1.98 (m, 4H), 1.95 (d, J = 9.7 Hz, 1H), 1.87 - 1.79 (m, 2H), 1.73 - 1.60 (m, 5H), 1.50 - 1.37 (m, 4H), 1.28 - 1.25 (m, 6H), 1.21 (s, 3H), 1.19 (s, 1H), 1.14 - 1.08 (m, 6H), 1.05 (s, 3H), 1.02 - 0.97 (m, 6H), 0.62 - 0.55 (m, 1H), 0.46 - 0.41 (m, 1H).

[0025] Synthesis of Compound A2: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and ((2S,3S)-2-((tert-butoxycarbonyl)amino)-3-methylpentanoic acid (185.0 mg), diisopropylcarbodiimide (124 μL), and 4-dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added, and the mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried, and concentrated to obtain the crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. The obtained compound was dissolved in ethyl acetate (5 mL), 5 mL of hydrogen chloride - dioxane was added, and the mixture was stirred at room temperature for 2 hours and then filtered. The filter cake was collected to obtain the target product A2 112.1 mg (yield = 43.70%): 1 H NMR (400 MHz, Methanol-d 4)δ4.77 - 4.71(m, 1H), 4.69 - 4.63(m, 1H), 4.05 - 3.99(m, 1H), 3.78 - 3.73(m, 1H), 3.49(t, J = 10.1Hz, 1H), 3.21(s, 1H), 2.61(q, J = 9.76Hz, 1H), 2.36(d, J = 7.72Hz, 1H), 2.08 - 1.94(m, 5H), 1.88 - 1.80(m, 2H), 1.76 - 1.54(m, 7H), 1.49(d, J = 9.3Hz, 2H), 1.45 - 1.37(m, 3H), 1.28(s, 3H), 1.26(s, 3H), 1.21(s, 3H), 1.19(s, 6H), 1.13(s, 3H), 1.11 - 1.08(m, 6H), 1.05 - 0.97(m, 8H), 0.95 - 0.80(m, 1H), 0.64 - 0.58(m, 1H), 0.48 - 0.43(m, 1H). Synthesis of Compound A3: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and (S)-2-((tert-butoxycarbonyl)amino)propanoic acid (151.4 mg), diisopropylcarbodiimide (124 μL), and 4-dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added, and the mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried, and concentrated to obtain a crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. The obtained compound was dissolved in ethyl acetate (5 mL), and 5 mL of hydrogen chloride-dioxane was added. After stirring at room temperature for 2 hours, the mixture was filtered, and the filter cake was collected to obtain the target product A3 102.4 mg (yield = 42.75%): 1 H NMR(400MHz, M ethanol-d 4)δ4.76 - 4.63(m, 2H), 4.18 - 3.91(m, 2H), 3.79 - 3.72(m, 1H), 3.52 - 3.41(m, 1H), 2.66 - 2.57(m, 1H), 2.41 - 2.35(m, 1H), 2.10 - 1.94(m, 5H), 1.83(d, J = 12.7Hz, 2H), 1.78 - 1.71(m, 2H), 1.70 - 1.62(m, 3H), 1.58(d, J = 7.4Hz, 2H), 1.49(d, J = 9.1Hz, 2H), 1.44 - 1.38(m, 2H), 1.29 - 1.25(m, 5H), 1.22(s, 3H), 1.20 - 1.16(m, 3H), 1.13(s, 3H), 1.10(s, 6H), 1.05 - 0.99(m, 5H), 0.93(s, 3H), 0.65 - 0.58(m, 1H), 0.48 - 0.40(m, 1H).

[0026] Synthesis of Compound A4: Dissolve cyclococcidiol (200 mg) in dichloromethane (4 mL), add (N - tert - butoxycarbonyl - glycine (140.1 mg), diisopropylcarbodiimide (124 μL) and 4 - dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract with dichloromethane three times (30 mL * 3). Combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. Dissolve the obtained compound in ethyl acetate (5 mL), add 5 mL of hydrogen chloride - dioxane, stir at room temperature for 2 hours and then filter. Collect the filter cake to obtain the target product A4 45.8 mg (yield = 20.61%): 1 H NMR (400 MHz, Methanol - d 4)δ4.76 - 4.72(m, 1H), 4.69 - 4.64(m, 1H), 3.88 - 3.83(m, 2H), 3.82 - 3.73(m, 2H), 3.51 - 3.45(m, 1H), 2.63 - 2.58(m, 1H), 2.37(d, J = 7.8Hz, 1H), 2.10 - 1.94(m, 5H), 1.85 - 1.80(m, 2H), 1.76 - 1.70(m, 2H), 1.69 - 1.59(m, 4H), 1.52 - 1.46(m, 2H), 1.45 - 1.36(m, 3H), 1.35 - 1.31(m, 1H), 1.28(s, 3H), 1.26(s, 3H) 1.22(s, 3H), 1.17(s, 3H), 1.12(s, 3H), 1.09(s, 3H), 1.01(s, 3H), 0.95 - 0.92(m, 1H), 0.64 - 0.58(m, 1H), 0.47 - 0.42(m, 1H).

[0027] Synthesis of Compound A5: Dissolve cyclococcidiol (200 mg) in dichloromethane (4 mL), add tert-butoxycarbonyl-β-alanine (151 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract three times with dichloromethane (30 mL * 3), combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. Dissolve the obtained compound in ethyl acetate (5 mL), add 5 mL of hydrogen chloride-dioxane, stir at room temperature for 2 hours and then filter. Collect the filter cake to obtain the target product A5 160.6 mg (yield = 67.42%): 1 H NMR(400MHz, Methanol-d 4)δ4.69 - 4.62 (m, 2H), 3.82 - 3.71 (m, 2H), 3.69 - 3.56 (m, 1H), 3.51 - 3.44 (m, 1H), 3.24 - 3.17 (m, 2H), 2.80 - 2.71 (m, 2H), 2.61 (q, J = 10.7 Hz, 1H), 2.36 (d, J = 7.4 Hz, 1H), 2.07 - 1.99 (m, 3H), 1.95 (d, J = 10.5 Hz, 1H), 1.86 - 1.78 (m, 2H), 1.53 - 1.35 (m, 5H), 1.31 - 1.27 (m, 5H), 1.26 (s, 3H), 1.21 (s, 3H), 1.15 (s, 3H), 1.13 (s, 3H), 1.07 (s, 3H), 1.01 (s, 3H), 0.94 - 0.84 (m, 1H), 0.62 - 0.58 (m, 1H), 0.45 - 0.41 (m, 1H).

[0028] Synthesis of Compound A6: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and then N - tert - butyloxycarbonyl - L - methionine (199.5 mg), diisopropylcarbodiimide (124 μL), and 4 - dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added. The mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried, and concentrated to obtain the crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. The obtained compound was dissolved in ethyl acetate (5 mL), and 5 mL of hydrogen chloride - dioxane was added. After stirring at room temperature for 2 hours, the mixture was filtered, and the filter cake was collected to obtain the target product A6 146.5 mg (yield = 55.68%): 1 H NMR (400 MHz, Methanol - d 4)δ4.81 - 4.77(m, 1H), 4.73 - 4.68(m, 1H), 4.28(s, 1H), 3.85 - 3.77(m, 2H) 3.75 - 3.68(m, 1H), 3.56 - 3.50(m, 1H) 2.80 - 2.73(m, 2H), 2.66(q, J=11.3, 10.7Hz, 2H), 2.41(d, J=7.8Hz, 1H), 2.37 - 2.32(m, 1H), 2.26 - 2.22(m, 1H), 2.18(s, 3H), 2.11 - 2.05(m, 3H), 2.03 - 1.98(m, 1H), 1.95 - 1.84(m, 3H) 1.79 - 1.65(m, 6H), 1.59 - 1.51(m, 3H), 1.49 - 1.42(m, 3H), 1.39 - 1.36(m, 1H), 1.33(s, 3H), 1.31(s, 3H), 1.26(s, 3H), 1.23(s, 3H), 1.18(s, 3H), 1.16(s,, 3H), 1.06(s, 3H), 1.02 - 0.88(m, 1H), 0.68 - 0.63(m, 1H), 0.53 - 0.48(m, 1H).

[0029] Synthesis of Compound A7: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and then tert - butoxycarbonyl - D - leucine (185.0 mg), diisopropylcarbodiimide (124 μL) and 4 - dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added. The mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried and concentrated to obtain the crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product. The obtained compound was dissolved in ethyl acetate (5 mL), and 5 mL of hydrogen chloride - dioxane was added. After stirring at room temperature for 2 hours, it was filtered, and the filter cake was collected to obtain the target product A7 161.6 mg (yield = 63.22%): 1 H NMR(400MHz, Methanol - d 4)δ 4.71 (d, J = 12.1 Hz, 1H), 4.68 - 4.62 (m, 1H), 4.07 - 4.01 (m, 1H), 3.80 - 3.72 (m, 1H), 3.69 - 3.64 (m, 1H), 3.49 (t, J = 10.1 Hz, 1H), 2.61 (q, J = 10.8 Hz, 1H), 2.37 (d, J = 7.2 Hz, 1H), 2.07 - 1.93 (m, 4H), 1.86 - 1.78 (m, 4H), 1.73 - 1.61 (m, 6H), 1.49 (d, J = 8.9 Hz, 2H), 1.39 (d, J = 11.1 Hz, 3H), 1.35 - 1.28 (m, 6H), 1.26 (s, 3H), 1.22 (s, 3H), 1.19 (s, 3H), 1.12 (s, 3H), 1.10 (s, 3H), 1.05 - 0.99 (m, 10H), 0.95 - 0.85 (m, 1H), 0.65 - 0.59 (m, 1H), 0.48 - 0.43 (m, 1H).

[0030] Synthesis of Compound B1: Dissolve cyclocophorol (200 mg) in dichloromethane (4 mL), add 4-phenoxybenzoic acid (171.38 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract with dichloromethane three times (30 mL * 3), combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B1 168.3 mg (yield = 61.25%): 11H NMR (400 MHz, Chloroform-d) δ 8.02 (s, 1H), 8.00 (s, 1H), 7.38 (t, J = 7.7 Hz, 2H), 7.18 (t, J = 7.4 Hz, 1H), 7.06 (s, 1H), 7.04 (s, 1H), 7.00 (s, 1H), 6.98 (s, 1H), 4.82 - 4.77 (m, 1H), 4.69 (q, J = 7.4 Hz, 1H), 3.75 (t, J = 7.1 Hz, 1H), 3.59 - 3.51 (m, 1H), 2.59 (q, J = 10.6 Hz, 1H), 2.35 (d, J = 7.8 Hz, 1H), 2.05 - 1.96 (m, 5H), 1.95 - 1.91 (m, 1H), 1.83 - 1.78 (m, 1H), 1.74 - 1.67 (m, 3H), 1.65 - 1.57 (m, 3H), 1.54 - 1.44 (m, 3H), 1.43 - 1.41 (m, 1H), 1.38 (d, J = 11.4 Hz, 1H), 1.29 (s, 3H), 1.27 (s, 3H), 1.26 - 1.24 (m, 1H), 1.22 (s, 3H), 1.19 (s, 3H), 1.17 (s, 3H), 1.14 - 1.12 (m, 4H), 0.97 (s, 3H), 0.58 (d, J = 4.4 Hz, 1H), 0.42 (d, J = 4.4 Hz, 1H).

[0031] Synthesis of Compound B2: Dissolve cyclococcidiol (200 mg) in dichloromethane (4 mL), add o-phenylbenzoic acid (158.6 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract three times with dichloromethane (30 mL * 3), combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B2, 71.7 mg (yield = 26.72%): 11H NMR (400 MHz, Chloroform-d) δ 7.82 (dd, J = 7.8, 1.4 Hz, 1H), 7.51 (td, J = 7.5, 1.4 Hz, 1H), 7.41 (dd, J = 7.7, 1.4 Hz, 1H), 7.39 - 7.37 (m, 1H), 7.36 - 7.33 (m, 3H), 7.32 - 7.31 (m, 1H), 7.31 - 7.29 (m, 1H), 4.67 (q, J = 7.4 Hz, 1H), 7.61 (dd, J = 11.7, 4.5 Hz, 1H), 3.74 (t, J = 7.8, 1H), 3.44 (td, J = 9.8, 3.4 Hz, 1H), 2.57 (q, J = 10.6 Hz, 1H), 2.32 (d, J = 7.9 Hz, 1H), 2.02 - 1.97 (m, 3H), 1.96 - 1.92 (m, 1H), 1.76 - 1.71 (m, 2H), 1.62 - 1.56 (m, 3H), 1.43 - 1.37 (m, 3H), 1.36 - 1.32 (m, 3H), 1.29 - 1.28 (m, 4H), 1.25 (s, 3H), 1.23 (s, 3H), 1.21 (s, 3H), 1.13 (s, 3H), 1.02 (s, 3H), 0.93 (s, 3H), 0.90 - 0.86 (m, 1H), 0.78 (s, 3H), 0.46 (d, J = 4.5 Hz, 1H), 0.32 (d, J = 4.4 Hz, 1H).

[0032] Synthesis of Compound B3: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and 2,2,3,3 - tetramethylcyclopropanecarboxylic acid (113.8 mg), diisopropylcarbodiimide (124 μL), and 4 - dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added. The mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried, and concentrated to obtain the crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B3, 70.4 mg (yield = 28.63%). 11H NMR (400 MHz, Chloroform-d) δ 4.68 (q, J = 7.3 Hz, 1H), 4.56 (q, J = 4.8 Hz, 1H), 3.74 (t, J = 8.0 Hz, 1H), 3.54 - 3.47 (m, 1H), 2.57 (q, J = 10.7 Hz, 1H), 2.33 (d, J = 7.8 Hz, 1H), 2.02 - 1.98 (m, 2H), 1.91 - 1.84 (m, 1H), 1.79 - 1.64 (m, 2H), 1.62 - 1.54 (m, 5H), 1.45 - 1.42 (m, 2H), 1.41 - 1.39 (m, 1H), 1.37 - 1.35 (m, 1H), 1.34 - 1.31 (m, 1H), 1.29 (s, 3H), 1.26 - 1.24 (m, 7H), 1.22 (s, 3H), 1.21 (s, 3H), 1.18 (s, 3H), 1.17 (s, 3H), 1.13 (s, 3H), 1.12 (s, 3H), 1.05 (s, 3H), 0.95 (s, 3H), 0.88 - 0.84 (m, 2H), 0.53 (d, J = 4.4 Hz, 1H), 0.38 (d, J = 4.4 Hz, 1H).

[0033] Synthesis of Compound B4: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and 2-(4-isobutylphenyl)propanoic acid (165.0 mg), diisopropylcarbodiimide (124 μL), and 4-dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added, and the mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried, and concentrated to obtain the crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B4, 202.6 mg (yield = 74.65%): 11H NMR (400 MHz, Chloroform-d) δ 7.21 (s, 1H), 7.19 (s, 1H), 7.08 (s, 1H), 7.06 (s, 1H), 4.67 (q, J = 7.2 Hz, 1H), 4.54 - 4.47 (m, 1H), 4.24 (br.s, 1H), 3.83 - 3.77 (m, 1H), 3.76 - 3.72 (m, 1H), 3.69 - 3.65 (m, 1H), 3.47 - 3.39 (m, 1H), 2.56 (q, J = 10.5 Hz, 1H), 2.44 (s, 1H), 2.42 (s, 1H), 2.32 (d, J = 7.8 Hz, 1H), 2.01 - 1.94 (m, 3H), 1.85 - 1.80 (m, 1H), 1.78 - 1.71 (m, 2H), 1.64 - 1.56 (m, 4H), 1.52 - 1.46 (m, 4H), 1.41 - 1.34 (m, 3H), 1.28 (s, 3H), 1.24 - 1.22 (m, 2H), 1.21 - 1.19 (m, 2H), 1.1 (s, 3H), 1.13 (s, 6H), 1.05 - 1.02 (m, 1H), 0.97 - 0.95 (m, 1H), 0.92 (s, 3H), 0.89 (s, 3H), 0.89 - 0.88 (m, 1H), 0.87 (s, 3H), 0.86 - 0.85 (m, 2H), 0.82 - 0.79 (m, 2H), 0.49 (d, J = 0.44 Hz, 1H), 0.35 (d, J = 4.4 Hz, 1H).

[0034] Synthesis of Compound B5: Dissolve cyclococcidiol (200 mg) in dichloromethane (4 mL), add m-fluorobenzoic acid (112.1 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract three times with dichloromethane (30 mL * 3), combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B5 206.5 mg (yield = 84.24%): 11H NMR (400 MHz, Chloroform-d) δ 7.82 (d, J = 7.7 Hz, 1H), 7.72 - 7.67 (m, 1H), 7.43 - 7.36 (m, 1H), 7.26 - 7.20 (m, 1H), 4.82 - 4.78 (m, 1H), 4.69 (q, J = 7.2 Hz, 1H), 4.36 (br.s, 1H), 3.76 - 3.71 (m, 1H), 3.58 - 3.40 (m, 1H), 2.60 (q, J = 10.7 Hz, 1H), 2.45 (br.s, 1H), 2.35 (d, J = 7.8 Hz, 1H), 2.08 - 1.96 (m, 5H), 1.94 - 1.90 (m, 1H), 1.80 (dd, J = 12.4, 4.6 Hz, 1H), 1.74 - 1.68 (m, 2H), 1.68 - 1.62 (m, 2H), 1.62 - 1.55 (m, 2H), 1.51 (d, J = 9.6 Hz, 1H), 1.48 - 1.41 (m, 3H), 1.41 - 1.35 (m, 1H), 1.28 (s, 3H), 1.26 (s, 3H), 1.20 (s, 3H), 1.18 (s, 6H), 1.12 (s, 3H), 0.96 (s, 3H), 0.57 (d, J = 4.4 Hz, 1H), 0.42 (d, J = 4.4 Hz, 1H).

[0035] Synthesis of Compound B6: Dissolve cycloastragenol (200 mg) in dichloromethane (4 mL), add m-toluic acid (108.9 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg). After reacting at room temperature for 24 hours, add 30 mL of saturated ammonium chloride aqueous solution, extract three times with dichloromethane (30 mL * 3), combine the organic phases, wash with saturated brine, dry and concentrate to obtain the crude product. The crude product is purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B6 176.3 mg (yield = 72.39%). 11H NMR (400 MHz, Chloroform-d) δ 7.87 - 7.83 (m, 2H), 7.37 - 7.30 (m, 2H), 4.85 - 4.79 (m, 1H), 4.70 (q, J = 7.3 Hz, 1H), 3.75 (t, J = 7.1 Hz, 1H), 3.60 - 3.52 (m, 1H), 2.59 (q, J = 10.6 Hz, 1H), 2.40 (s, 3H), 2.35 (d, J = 7.8 Hz, 1H), 2.05 - 1.97 (m, 4H), 1.96 - 1.91 (m, 1H), 1.82 (dd, J = 12.5, 4.4 Hz, 1H), 1.74 - 1.69 (m, 2H), 1.69 - 1.58 (m, 4H), 1.53 (s, 1H), 1.50 (s, 1H), 1.49 - 1.45 (m, 1H), 1.44 - 1.41 (m, 1H), 1.41 - 1.36 (m, 1H), 1.30 (s, 3H), 1.28 (s, 3H), 1.26 - 1.24 (m, 1H), 1.23 (s, 3H), 1.20 (s, 6H), 1.14 (s, 3H), 1.13 - 1.12 (m, 1H), 0.9 (s, 3H), 0.91 - 0.79 (m, 1H), 0.59 (d, J = 4.5 Hz, 1H), 0.43 (d, J = 4.4 Hz, 1H).

[0036] Synthesis of Compound B7: Cycloastragenol (200 mg) was dissolved in dichloromethane (4 mL), and then o-toluic acid (108.92 mg), diisopropylcarbodiimide (124 μL) and 4-dimethylaminopyridine (24 mg) were added. After reacting at room temperature for 24 hours, 30 mL of saturated ammonium chloride aqueous solution was added. The mixture was extracted with dichloromethane three times (30 mL * 3). The combined organic phases were washed with saturated brine, dried and concentrated to obtain the crude product. The crude product was purified by silica gel column (DCM / MeOH = 20 / 1) to obtain the target product B7 104.1 mg (yield = 42.75%). 11H NMR (400 MHz, Chloroform-d) δ 7.92 (d, J = 7.1 Hz, 1H), 7.41 - 7.36 (m, 1H), 7.26 - 7.21 (m, 2H), 4.85 - 4.80 (m, 1H), 4.70 (q, J = 7.3 Hz, 1H), 3.75 (t, J = 7.1 Hz, 1H), 3.58 - 3.53 (m, 1H), 2.61 (s, 3H), 2.35 (d, J = 7.8 Hz, 1H), 2.05 - 1.96 (m, 5H), 1.95 - 1.92 (m, 1H), 1.83 - 1.74 (m, 2H), 1.73 - 1.59 (m, 6H), 1.52 (d, J = 9.6 Hz, 1H), 1.47 - 1.45 (m, 1H), 1.43 - 1.40 (m, 1H), 1.30 (s, 3H), 1.27 (s, 3H), 1.25 (s, 3H), 1.23 (s, 3H), 1.22 (s, 3H), 1.17 (s, 3H), 1.14 (s, 3H), 0.97 (s, 3H), 0.89 - 0.85 (m, 1H), 0.58 (d, J = 4.5 Hz, 1H), 0.42 (d, J = 4.5 Hz, 1H).

[0037] Example 2:

[0038] Activity test of the compound of the present invention:

[0039] 1. Experimental materials

[0040] 1.1 Preparation of the drug: Cycloastragenol derivative, prepared by the preparation method described in Example 1. Immediately before use, it was prepared into a stock solution of 3.2 mM with cell-grade DMSO and then diluted to the required concentration with LB broth. Ceftazidime (CAZ) was used as a positive control.

[0041] 1.2 Preparation of the culture medium

[0042] Preparation of solid medium: Weigh a certain amount of LB agar solid powder as needed, dissolve it in distilled water, sterilize it under high temperature and high pressure conditions at 121 °C for 30 minutes, transfer it to a sterile petri dish while it is hot, and store it in a 4 °C refrigerator after natural cooling and solidification.

[0043] Preparation of liquid medium: Weigh a certain amount of LB broth solid powder as needed, dissolve it in distilled water, sterilize it under high temperature and high pressure conditions at 121 °C for 30 minutes, cool it to room temperature, and store it in a 4 °C refrigerator for later use.

[0044] 2. Experimental method

[0045] 2.1 Bacterial strain activation: Use the streak plate method to inoculate the experimental strain (Staphylococcus aureus ATCC43300) onto an LB solid medium. Invert the medium and place it in a constant temperature incubator at 37°C for 24 hours of incubation.

[0046] 2.2 Bacterial strain subculture: Pick 3 - 5 colonies with the same morphology from the LB solid medium using an inoculation loop and inoculate them into a shaking flask containing 4 - 5 mL of LB liquid medium. Place the shaking flask in a shaker at 37°C and 150 rpm to culture the bacteria until the logarithmic growth phase (6 - 8 hours) for standby.

[0047] 2.3 Bacterial suspension preparation: Prepare a 0.5 MCF bacterial suspension of the test bacteria using LB broth medium, containing approximately 1 - 5*10 6 CFU / mL.

[0048] 2.4 Drug dilution: In columns 2 - 11 of a 96 - well plate, add 100 μL of LB medium to each well. In the first column of wells, add 190 μL of LB medium. Take 10 μL of the drug and add it to the wells in the first column. Use a pipette to blow and mix up and down for two - fold serial dilution. Starting from the first column, perform serial dilution successively backward to make the drug concentrations in each well decrease. The 11th column serves as the growth control, containing only medium and bacterial suspension, without drugs. The 12th column is the blank control, containing only medium, without adding bacterial suspension and drugs.

[0049]

[0050]

[0051] 2.5 Bacterial inoculation: Add 100 μL of the prepared bacterial suspension to each well except the blank control well, so that the final bacterial amount in each well is approximately 5*10 5 CFU / mL

[0052] 2.6 Bacterial culture: Cover the 96 - well plate and place it in an incubator at 37°C for 24 hours of incubation.

[0053] 2.7 Result observation: Use an enzyme - linked immunosorbent assay (ELISA) reader to detect the absorbance at a wavelength of 600 nm to judge the growth of bacteria. The lowest drug concentration that completely inhibits the growth of bacteria is the minimum inhibitory concentration (MIC).

[0054] 3. Experimental results: The results show that the growth inhibitory effect of some cyclocophorol derivatives on methicillin - resistant Staphylococcus aureus is significantly higher than that of the cyclocophorol control group. It can be seen that the cyclocophorol derivatives prepared in Example 1 have a relatively obvious inhibitory effect on the growth of methicillin - resistant Staphylococcus aureus, indicating that this series of derivatives has good antibacterial effects. The results are shown in Figure 1 。

Claims

1. A cycloastragenol derivative having anti-methicillin-resistant Staphylococcus aureus activity, characterized in that: Cycloastragenol (CAG) derivatives (A1-A7, B1-B7) or pharmaceutically acceptable salts thereof obtained by using cycloastragenol (CAG) as a parent and connecting amino acids or other organic acids to its 3-hydroxyl group in the form of an ester group are used in the development of new antibiotic adjuvants.

2. The preparation method of compound A1-A7 as claimed in claim 1 is as follows: the first step is condensation: dissolving cycloastragenol in dichloromethane, adding the corresponding amino acid, a condensing agent and 4-dimethylaminopyridine at room temperature, reacting at room temperature for 24 hours, adding saturated ammonium chloride aqueous solution to the reaction solution, extracting with dichloromethane, concentrating to obtain a crude product, and purifying the crude product with a silica gel column to obtain a condensation product. The second step is deprotection to form a salt: dissolving the condensation product in an organic solvent, adding an organic solvent solution of hydrogen chloride, stirring at room temperature until a white solid is produced, and filtering to obtain the desired target product A1-A7.

3. The preparation method of compound B1-B7 as claimed in claim 1 is: dissolving cycloastragenol in dichloromethane, adding the corresponding organic acid, a condensing agent and 4-dimethylaminopyridine at room temperature, reacting at room temperature for 24 hours, adding saturated ammonium chloride aqueous solution to the reaction solution, extracting with dichloromethane, concentrating to obtain a crude product, and purifying the crude product by silica gel column to obtain a condensation product.

4. The method for preparing compounds A1-A7 and B1-B7 according to claims 1 and 2, characterized in that: The condensing agent is dicyclohexylcarbodiimide, diisopropylcarbodiimide or 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide.

5. The method for preparing compounds A1-A7 according to claim 1, characterized in that: The amino acids are N-tert-butoxycarbonyl-L-phenylalanine, (2S,3S)-2-((tert-butoxycarbonyl)amino)-3-methylpentanoic acid, (S)-2-((tert-butoxycarbonyl)amino)propionic acid, N-tert-butoxycarbonyl-glycine, tert-butoxycarbonyl-β-alanine, N-tert-butoxycarbonyl-L-methionine, and tert-butoxycarbonyl-D-leucine.

6. The method for preparing compound B1-B7 according to claim 2, characterized in that: The organic acid is 4-phenoxybenzoic acid, o-phenylbenzoic acid, 2,2,3,3-tetramethylcyclopropanecarboxylic acid, 2-(4-isobutylphenyl)propionic acid, m-fluorobenzoic acid, m-toluic acid and o-toluic acid.

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