Pleuromutilin derivative, preparation method thereof and antibacterial active composition

Abstract

The invention relates to the technical field of chemical synthesis of medicines, in particular to a pleuromutilin derivative, a preparation method thereof and an antibacterial active composition. The invention relates to a pleuromutilin derivative with a structure as shown in a formula I, or pharmaceutically and / or veterinary acceptable salt, a solvent compound, an optical isomer and a polymorphic compound of the pleuromutilin derivative. Formula I; wherein R1 is selected from one of a hydrogen atom and methyl; r2 is selected from one of a hydrogen atom and a methoxy group; and R3 is selected from one of cyclopropyl and p-fluorophenyl. According to the present invention, the quinolone drug is spliced to the pleuromutilin side chain to produce the new pleuromutilin derivative, and the obtained new compound can enhance the antibacterial activity, has significant antibacterial activity on Gram-positive bacteria and Gram-negative bacteria, and further has significant inhibition effect on MRSA and other drug-resistant bacteria so as to effectively broaden the antibacterial spectrum.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical chemical synthesis, and in particular to a pleuromutilin derivative, a preparation method thereof, and an antibacterial active composition. Background Art

[0002] The overuse of antibiotics has led to an increasingly serious problem of antibiotic resistance in pathogens, posing a severe challenge to disease prevention and control and a significant threat to human and animal health. The urgent need for antimicrobial drugs has made the use of the drug combination principle an innovative approach to addressing this problem. The drug combination principle involves combining the structures of two drugs into a single molecule, or integrating the pharmacophores of both drugs into a single molecule, known as a hybrid molecule. The newly formed hybrid molecule can either possess the properties of both drugs, enhancing their pharmacological effects while minimizing their respective toxic side effects; or it can leverage the strengths of both drugs to complement their weaknesses, exerting their respective pharmacological activities and synergistically completing the therapeutic process. Drug combination involves combining drugs with known efficacy to create new drugs. The pharmacological activity of the combined new drug can be easily predicted based on the pharmacological effects of the active pharmaceutical ingredient, providing a purposeful and foundational approach to new drug development, thereby shortening the development process and saving significant human, material, and financial resources.

[0003] Quinolones primarily act on topoisomerase IV in Gram-positive bacteria or DNA gyrase in Gram-negative bacteria, thereby interfering with DNA replication and exerting a bactericidal effect. Quinolones possess strong bactericidal properties and a broad antimicrobial spectrum, particularly effective against Enterobacter, Pseudomonas aeruginosa, Haemophilus influenzae, Neisseria gonorrhoeae, Streptococcus, Legionella, and Staphylococcus aureus. Clinically, they are effective against infections caused by sensitive bacteria in the respiratory tract, urinary tract, digestive tract, biliary tract, skin and soft tissue trauma, gynecological inflammation, and ear, nose, and eye areas. However, the widespread use of quinolones has led to an increase in drug-resistant strains, limiting their application.

[0004] Pleuromutilin is a natural compound, a broad-spectrum diterpene antibiotic produced by the culture of Pleurotus mutilus. Pleuromutilins specifically bind to the peptidyl transferase active center (PTC) of the bacterial 50S ribosome subunit, interfering with tRNA binding to the ribosome and inhibiting bacterial protein synthesis, resulting in a bactericidal effect. Pleuromutilins exhibit broad-spectrum antimicrobial activity against Gram-positive bacteria (such as Staphylococcus aureus and Streptococcus pneumoniae), mycoplasmas, and some Gram-negative bacteria (such as Escherichia coli). Pleuromutilins are clinically used to control swine panting disease, porcine contagious pleuropneumonia, chronic respiratory disease in chickens, and Mycoplasma synoviae. They also have significant efficacy against digestive tract infections such as treponemal dysentery.

[0005] Currently, only four pleuromutilin-based antimicrobial drugs have been successfully developed: tiamulin, valnemulin, retapamulin, and azamorelin. Resistance to these drugs is still rare. Therefore, developing new pleuromutilin-based antimicrobial drugs is of great significance in providing more options for treating bacterial infections in veterinary clinics. Summary of the Invention

[0006] The main purpose of the present invention is to provide a pleuromutilin derivative and its preparation method and antibacterial active composition, aiming to improve the technical problem in the prior art regarding the low number of drug-resistant bacteria of pleuromutilin-type antibacterial drugs and obtain a highly effective and long-lasting antibacterial active compound.

[0007] To achieve the above-mentioned object, the present invention provides a pleuromutilin derivative having the structure of Formula I, or a pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, or polymorph thereof; Formula I; Wherein: R1 is selected from one of hydrogen atom and methyl group; R2 is selected from one of hydrogen atom and methoxy group; R3 is selected from one of cyclopropyl group and p-fluorophenyl group.

[0008] Preferably, it has the following structure (A), (B) or (C), and has significant antibacterial activity against Gram-positive and Gram-negative bacteria, and also has a significant inhibitory effect on drug-resistant bacteria such as MRSA: Formula A; Formula B; Formula C.

[0009] Preferably, it is selected from one of the following compounds: 1-cyclopropyl-6-fluoro-7-[4-(2-{[5-({2-[(3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propancyclopenten-5-yloxy]-2-oxoethyl}thio)-1,3,4-thiadiazol-2-yl]amino}-2-oxoethyl)piperazin-1-yl]-4-oxo-1,4-dihydroquinoline-3-carboxylic acid; 1-cyclopropyl-6-fluoro-7-[4-(2-{[5-({2-[(3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propancyclopenten-5-yloxy]-2-oxoethyl}thio)-1,3,4-thiadiazol-2-yl]amino}-2-oxoethyl)-3-methylpiperazin-1-yl]-8-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid; 6-Fluoro-1-(4-fluorophenyl)-7-[4-(2-{[5-({2-[(3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propancyclopenten-5-yloxy]-2-oxoethyl}thio)-1,3,4-thiadiazol-2-yl]amino}-2-oxoethyl)piperazin-1-yl]-4-oxo-1,4-dihydroquinoline-3-carboxylic acid.

[0010] In addition, the present invention also provides a method for preparing the above-mentioned pleuromutilin derivative, or its pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, or polymorph, which has the following synthetic route: .

[0011] Preferably, the method comprises the following steps: S1: Compound 1 is used as a raw material, and in the presence of a base, a solvent is used to react with di-tert-butyl dicarbonate to produce compound 2; wherein compound 1 is ciprofloxacin, gatifloxacin or sarafloxacin; S2: Compound 2 prepared in step S1 is subjected to a substitution reaction with allyl bromide to prepare compound 3; S3: dissolving compound 3 obtained in step S2 in dichloromethane and reacting with an acid to obtain compound 4; S4: Compound 4 obtained in step S3 is dissolved in acetonitrile, and then reacted with tert-butyl bromoacetate in the presence of NN-diisopropylethylamine to obtain compound 5; S5: dissolving compound 5 obtained in step S4 in dichloromethane and reacting with an acid to obtain compound 6; S6: Compound 7-pleuromutilin was used as a raw material and reacted with p-toluenesulfonyl chloride in pyridine to generate compound 8; S7: dissolving the compound 8 obtained in step S6 in acetonitrile and iodizing with NaI in the presence of potassium carbonate to obtain compound 9; S8: reacting the compound 9 obtained in step S7 with 2-amino-5-mercapto-1,3,4-thiadiazole to obtain compound 10; S9: Compound 6 prepared in step S5 is dissolved in DMF, and reacted with compound 10 prepared in step S8 in the presence of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, NN-diisopropylethylamine, 1-hydroxybenzotriazole and 4-dimethylaminopyridine to prepare compound 11; S10: Compound 11 obtained in step S9 is dissolved in dichloromethane and reacted with tributyltin hydride in the presence of bis(triphenylphosphine)palladium chloride and water to obtain the final product 12.

[0012] The base in step S1 is one of NaHCO3, K2CO3, NaOH, KOH (or other inorganic bases), triethylamine, pyridine, piperidine, dicyclohexylamine (or other organic bases), preferably NaHCO3; the solvent is at least one of DMF, dichloromethane, chloroform, 1,2-dichloroethane (or other low-boiling halogenated hydrocarbons), ether, 1,4-dioxane, methyl tert-butyl ether (or other ether solvents), preferably DMF; the reaction temperature is 25-60°C, preferably 25°C; the molar ratio of compound 1 to di-tert-butyl dicarbonate is 1:1-3:5, preferably 1:1.2; The reaction temperature in step S2 is 40-95° C., preferably 90° C.; the molar ratio of compound 2 to allyl bromide is 1:1-1:5, preferably 1:4.5; The acid in step S3 is one of trifluoroacetic acid, HCl, H2SO4, HBr, phosphoric acid and other inorganic acids, preferably trifluoroacetic acid; the reaction temperature is 25-60°C, preferably 25°C; The reaction temperature in step S4 is 40-95° C., preferably 75° C.; the molar ratio of compound 4 to tert-butyl bromoacetate is 1:1-1:2, preferably 1:1.9; The acid in step S5 is one of trifluoroacetic acid, HCl, H2SO4, HBr, phosphoric acid and other inorganic acids, preferably trifluoroacetic acid; the reaction temperature is 25-60°C, preferably 25°C; The reaction temperature in step S6 is -5 to 5°C, preferably 0°C; the molar ratio of compound 7 to p-toluenesulfonyl chloride is 1:1 to 3:5, preferably 1:1.1; The reaction temperature in step S7 is 40-95° C., preferably 78° C.; the molar ratio of compound 8 to sodium iodide is 1:1-3:5, preferably 1:1.1; In step S8, the molar ratio of compound 9 to 2-amino-5-mercapto-1,3,4-thiadiazole is 1:1 to 3:5, preferably 1:1.1; The feeding temperature before the reaction in step S9 is -5 to 5°C, preferably 0°C; the reaction temperature is 25°C to 90°C, preferably 25°C; the molar ratio of compound 6 to 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is 1:(1.1 to 1.8), preferably 1:1.1; the molar ratio of compound 6 to NN-diisopropylethylamine is 1:(1.5 to 1.8), preferably 1:1.5; the molar ratio of compound 6 to 1-hydroxybenzotriazole is 1:(1.1 to 1.8), preferably 1:1.1; the molar ratio of compound 6 to 4-dimethylaminopyridine is 1:(0.5 to 0.8), preferably 1:0.5; the molar ratio of compound 6 to compound 10 is (1.2 to 1.5):1, preferably 1.2:1; The reaction temperature of step S10 is 10-35° C., preferably 25° C.; the molar ratio of compound 11 to bis(triphenylphosphine)palladium chloride is 1:(0.02-0.1), preferably 1:0.02.

[0013] Under the above-mentioned preferred parameter conditions, each reaction can obtain high-purity products with higher yields / yields.

[0014] In addition, the present invention further protects the use of the above-mentioned pleuromutilin derivative or a pharmaceutically and / or veterinarily acceptable solvate thereof in the preparation of an antibacterial preparation.

[0015] In addition, the present invention also provides an antimicrobial active composition comprising the above-mentioned pleuromutilin derivative and or its pharmaceutically and / or veterinarily acceptable salts, solvent compounds, optical isomers, polymorphic compounds and pharmaceutically acceptable carriers or diluents.

[0016] Preferably, the dosage form is oral or injection.

[0017] Preferably, the content of the pleuromutilin derivative or its pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, polymorph is 0.1%-99.5% (wt%), and the balance is carrier or diluent.

[0018] Compared with existing technologies, the present invention has the following beneficial effects: It covalently links two different lead compounds, producing synergistic, additive, or novel pharmacological activities in vivo. Specifically, the present invention splices a quinolone drug onto the side chain of pleuromutilin to produce a novel pleuromutilin derivative. The resulting novel compound has enhanced antibacterial activity, exhibiting significant antibacterial activity against both Gram-positive and Gram-negative bacteria, and exhibits significant inhibitory effects against drug-resistant bacteria such as MRSA, effectively broadening its antibacterial spectrum. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments of the present application. For ordinary technicians in this field, other relevant drawings can be obtained based on these drawings without paying any creative work.

[0020] Figure 1 is the compound (6A) in this embodiment. 1 HNMR spectrum; Figure 2 is the compound (6C) in this embodiment. 1 HNMR spectrum; Figure 3 is the compound (10) in this embodiment. 1 HNMR spectrum; Figure 4 is the compound (12A) in this embodiment. 1 HNMR spectrum; Figure 5 is the compound (12C) in this embodiment. 1 HNMR spectrum.

[0021] The realization of the objectives, functional features and advantages of this application will be further explained in conjunction with embodiments and with reference to the accompanying drawings. DETAILED DESCRIPTION

[0022] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of them. All other embodiments derived by persons of ordinary skill in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention.

[0023] In addition, the technical solutions between the various embodiments can be combined with each other, but they must be based on the fact that ordinary technicians in this field can implement them. When the combination of technical solutions is mutually contradictory or cannot be implemented, it should be deemed that such a combination of technical solutions does not exist and is not within the scope of protection required by the present invention.

[0024] In the present invention, "pharmaceutically and / or veterinarily acceptable salts" include salts formed with alkali metals, such as salts with inorganic bases such as sodium, potassium, magnesium, and calcium, salts with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, and fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, p-toluenesulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid, benzoic acid, malonic acid, benzenesulfonic acid, or naphthalenesulfonic acid.

[0025] In the present invention, the term "pharmaceutically and / or veterinarily acceptable solvate" refers to a hydrate or a solvate soluble in C1-C4 alcohol or other organic solvents.

[0026] The present invention provides a method for preparing the above-mentioned pleuromutilin derivative, or its pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, or polymorph, having the following synthetic route: .

[0027] In medicinal chemistry, splicing is an important method for optimizing lead compounds. The goal is to link two identical or different lead compounds or drugs together through covalent bonds to produce synergistic or additive effects in vivo, or to generate new pharmacological activities. Therefore, quinolone drugs are spliced ​​onto the side chain of pleuromutilin to produce new pleuromutilin derivatives. The hope is that these new compounds will broaden the antibacterial spectrum and be effective against drug-resistant bacteria.

[0028] The specific synthesis route includes the following steps: S1: using compound 1 as a raw material, reacting with di-tert-butyl dicarbonate in a solvent in the presence of a base to generate compound 2; wherein compound 1 is ciprofloxacin, gatifloxacin or sarafloxacin; S2: Compound 2 prepared in step S1 is subjected to a substitution reaction with allyl bromide to prepare compound 3; S3: dissolving compound 3 obtained in step S2 in dichloromethane and reacting with an acid to obtain compound 4; S4: Compound 4 obtained in step S3 is dissolved in acetonitrile, and then reacted with tert-butyl bromoacetate in the presence of NN-diisopropylethylamine to obtain compound 5; S5: dissolving compound 5 obtained in step S4 in dichloromethane and reacting with an acid to obtain compound 6; S6: Compound 7-pleuromutilin was used as a raw material and reacted with p-toluenesulfonyl chloride in pyridine to generate compound 8; S7: dissolving the compound 8 obtained in step S6 in acetonitrile and iodizing with NaI in the presence of potassium carbonate to obtain compound 9; S8: reacting the compound 9 obtained in step S7 with 2-amino-5-mercapto-1,3,4-thiadiazole to obtain compound 10; S9: Compound 6 prepared in step S5 is dissolved in DMF, and reacted with compound 10 prepared in step S8 in the presence of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, NN-diisopropylethylamine, 1-hydroxybenzotriazole and 4-dimethylaminopyridine to prepare compound 11; S10: Compound 11 obtained in step S9 is dissolved in dichloromethane and reacted with tributyltin hydride in the presence of bis(triphenylphosphine)palladium chloride and water to obtain the final product 12.

[0029] The base in step S1 is one of NaHCO3, K2CO3, NaOH, KOH (or other inorganic bases), triethylamine, pyridine, piperidine, dicyclohexylamine (or other organic bases), preferably NaHCO3; the solvent is at least one of DMF, dichloromethane, chloroform, 1,2-dichloroethane (or other low-boiling halogenated hydrocarbons), ether, 1,4-dioxane, methyl tert-butyl ether (or other ether solvents), preferably DMF; the reaction temperature is 25-60°C, preferably 25°C; the molar ratio of compound 1 to di-tert-butyl dicarbonate is 1:1-3:5, preferably 1:1.2; The reaction temperature in step S2 is 40-95° C., preferably 90° C.; the molar ratio of compound 2 to allyl bromide is 1:1-1:5, preferably 1:4.5; The acid in step S3 is one of trifluoroacetic acid, HCl, H2SO4, HBr, phosphoric acid and other inorganic acids, preferably trifluoroacetic acid; the reaction temperature is 25-60°C, preferably 25°C; The reaction temperature in step S4 is 40-95° C., preferably 75° C.; the molar ratio of compound 4 to tert-butyl bromoacetate is 1:1-1:2, preferably 1:1.9; The acid in step S5 is one of trifluoroacetic acid, HCl, H2SO4, HBr, phosphoric acid and other inorganic acids, preferably trifluoroacetic acid; the reaction temperature is 25-60°C, preferably 25°C; The reaction temperature in step S6 is -5 to 5°C, preferably 0°C; the molar ratio of compound 7 to p-toluenesulfonyl chloride is 1:1 to 3:5, preferably 1:1.1; The reaction temperature in step S7 is 40-95° C., preferably 78° C.; the molar ratio of compound 8 to sodium iodide is 1:1-3:5, preferably 1:1.1; In step S8, the molar ratio of compound 9 to 2-amino-5-mercapto-1,3,4-thiadiazole is 1:1 to 3:5, preferably 1:1.1; The feeding temperature before the reaction in step S9 is -5 to 5°C, preferably 0°C; the reaction temperature is 25°C to 90°C, preferably 25°C; the molar ratio of compound 6 to 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is 1:(1.1 to 1.8), preferably 1:1.1; the molar ratio of compound 6 to NN-diisopropylethylamine is 1:(1.5 to 1.8), preferably 1:1.5; the molar ratio of compound 6 to 1-hydroxybenzotriazole is 1:(1.1 to 1.8), preferably 1:1.1; the molar ratio of compound 6 to 4-dimethylaminopyridine is 1:(0.5 to 0.8), preferably 1:0.5; the molar ratio of compound 6 to compound 10 is (1.2 to 1.5):1, preferably 1.2:1; The reaction temperature of step S10 is 10-35° C., preferably 25° C.; the molar ratio of compound 11 to bis(triphenylphosphine)palladium chloride is 1:(0.02-0.1), preferably 1:0.02.

[0030] The technical solutions of the present invention are further described in detail below with reference to specific examples. It should be understood that the following examples are merely illustrative of the present invention and are not intended to limit the present invention. The experimental methods described in the following examples are conventional methods unless otherwise specified. The reagents and materials described are commercially available unless otherwise specified.

[0031] Example 1: Synthesis of Compound 6A (2-{4-{3-[(allyloxy)carbonyl]-1-cyclopropyl-6-fluoro-4-oxo-1,4-dihydroquinolin-7-yl}piperazin-1-yl}acetic acid) S1. To a round-bottom flask, add ciprofloxacin (5 g), sodium bicarbonate (6.27 g), and 100 mL of DMF. Stir at room temperature for 10 minutes until completely dissolved. Then, slowly add 40 mL of di-tert-butyl dicarbonate (4.025 g) in DMF. Stir at room temperature under nitrogen for 5 hours to obtain compound (2A). Proceed to the next step without further treatment.

[0032] S2. Add 6 mL of allyl bromide to the reaction mixture, raise the temperature to 90°C, and continue stirring for 24 h. After the reaction is complete, remove the solvent by rotary evaporation under reduced pressure. Dissolve the resulting solid in 150 mL of dichloromethane. Wash the resulting solution three times with water (100 mL) and twice with saturated brine (100 mL). Collect the organic phase, dry it over anhydrous sodium sulfate, and evaporate it under reduced pressure to obtain a white solid, compound (3A).

[0033] S3. Dissolve 7 g of compound (3A) in 110 mL of dichloromethane, add trifluoroacetic acid (36 mL) dropwise, and stir at room temperature for 2 h under nitrogen. Monitor the reaction progress by thin-layer chromatography. Terminate the reaction after completion, remove the solvent by rotary evaporation under reduced pressure, and then add diethyl ether (150 mL) and stir for 0.5 h. A solid precipitates, which is filtered. Wash the filter cake once with diethyl ether and dry in vacuo to obtain a white solid, i.e., compound (4A).

[0034] S4. Weigh 2 g of compound (4A) and dissolve it in acetonitrile. Add tert-butyl bromoacetate (1.54 mL) and NN-diisopropylethylamine (2 mL) respectively. Heat to 75°C and stir under nitrogen for 0.5 h. Monitor the reaction endpoint by thin-layer chromatography. After the reaction is complete, remove the solvent by rotary evaporation under reduced pressure. Dissolve the resulting viscous liquid in dichloromethane and wash twice with 1 mol / L ammonium chloride solution (100 mL) and twice with saturated brine (100 mL). Collect the organic phase, dry over anhydrous sodium sulfate, and then rotary dry under reduced pressure. Dry the resulting white solid in vacuo to obtain compound (5A).

[0035] S5. Dissolve 2 g of compound (5A) in dichloromethane (40 mL) and add trifluoroacetic acid (40 mL). Stir at room temperature overnight under nitrogen. Monitor the reaction progress by thin-layer chromatography. After completion, remove the solvent by rotary evaporation under reduced pressure. Add anhydrous ether and stir for 0.5 h to precipitate a white solid. Filter the solid, wash the filter cake once with anhydrous ether, and dry it in vacuo to obtain compound (6A) in a yield of 73.7%. MS-ESI (M+1): 430.1772.

[0036] The results of H NMR spectrum are as follows: 1 H NMR (600 MHz, DMSO-d6) δ 8.46 (s, 1H), 7.77 (d, J = 13.1hz, 1H), 7.47 (d, J = 7.3hz, 1H), 6.01 (ddt, J = 17.3, 10.4, 5.2hz, 1H), 5.46 (dq, J =17.3, 1.8hz, 1H), 5.26 (dq, J = 10.5, 1.6hz, 1H), 4.71 (dt, J = 5.2, 1.7hz,2H), 4.27 (s, 2H), 3.67 (tt, J = 7.2, 4.0hz, 2H), 3.38 (q, J = 7.0hz, 1H), 1.28 (dd, J = 7.5, 5.6hz, 2H), 1.15 – 1.07 (m, 3H).

[0037] Example 2: Synthesis of Compound 6B (2-[4-(3-[(allyloxy)carbonyl]-1-cyclopropyl-6-fluoro-8-methoxy-4-oxo-1,4-dihydroquinolin-7-yl)-2-methylpiperazin-1-yl]acetic acid) The method was the same as Example 1, except that ciprofloxacin was replaced with an equimolar amount of gatifloxacin. The resulting white solid was named Compound (6B) with a yield of 68.9%. MS-ESI (M-1): 472.1889.

[0038] The results of H NMR spectrum are as follows: 1 H NMR (600 MHz, DMSO-d6) δ 8.50 (s, 1H), 7.58 (d, J = 12.4hz, 1H), 5.45 (dq, J = 17.2, 1.8hz, 1H), 5.25 (dq, J = 10.5, 1.6hz, 1H), 4.70 (dt, J =5.1, 1.6hz, 2H), 3.42 (s, 1H), 3.36 (s, 1H), 3.33 – 3.29 (m, 3H), 3.18 (s,1H), 2.98 (p, J = 4.4hz, 2H), 1.07 (dd, J = 14.7, 5.4hz, 5H), 0.98 – 0.94 (m,2H).

[0039] Example 3: Synthesis of Compound 6C (2-[4-(3-[(allyloxy)carbonyl]-6-fluoro-1-(4-fluorophenyl)-4-oxo-1,4-dihydroquinolin-7-yl)piperazin-1-yl]acetic acid) The method was the same as Example 1, except that ciprofloxacin was replaced with an equal molar amount of sarafloxacin. The resulting white solid was designated as Compound (6C) with a yield of 65.2%. MS-ESI (M-1): 482.1531.

[0040] The results of H NMR spectrum are as follows: 1H NMR (600 MHz, DMSO-d6) δ 8.42 (s, 1H), 7.86 (d, J = 13.0hz, 1H), 7.82 – 7.74 (m, 2H), 7.57 – 7.49 (m, 2H), 6.34 (d, J = 7.2hz, 1H), 5.99 (ddt,J = 17.2, 10.5, 5.3hz, 1H), 5.43 (dq, J = 17.2, 1.8hz, 1H), 5.23 (dq, J =10.5, 1.5hz, 1H), 4.70 (dt, J = 5.3, 1.6hz, 2H), 4.18 (s, 2H), 3.43 (s, 2H).

[0041] Example 4: Synthesis of Compound 10 ((3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propanecyclopenta[8]cyclononen-5-yl 2-[(5-amino-1,3,4-thiadiazol-2-yl)thio]acetate) S6. Dissolve 10.0 g of pleuromutilin in 20 mL of pyridine and place in an ice bath; dissolve 5.6 g of p-toluenesulfonyl chloride in 10 mL of pyridine, then slowly add the above-mentioned pleuromutilin pyridine solution, stir the mixture in an ice bath for 3 h, add 50 mL of ice water and 50 mL of chloroform in turn, transfer to a separatory funnel, shake, and let it stand to separate; take the organic phase, wash it with 100 mL of sulfuric acid (4 mol / L), 100 mL of saturated sodium bicarbonate solution, and 100 mL of deionized water in turn; after washing, evaporate the organic solution under reduced pressure, add 100 mL of ether, and a large amount of white powder precipitates. Filter, wash the filter cake once with anhydrous ether, and vacuum dry to obtain compound (8).

[0042] S7. Dissolve 2.13 g of compound (8) in 81 mL of acetonitrile, add 0.66 g of anhydrous sodium iodide and 1.11 g of anhydrous potassium carbonate, and heat under reflux at 78°C for 2 h to obtain compound (9).

[0043] S8. Add 0.59 g of 2-amino-5-mercapto-1,3,4-thiadiazole to the solution of compound (9), and continue the reaction at 78°C for 3 h. The reaction solution is evaporated under reduced pressure to remove the organic solvent. Then, 50 mL of distilled water and 50 mL of chloroform are added in sequence. The mixture is placed in a separatory funnel, shaken, and allowed to stand for separation. The organic phase is collected and washed twice with 200 mL of a saturated sodium chloride aqueous solution and dried over anhydrous sodium sulfate. The organic phase is collected and rotary evaporated to dryness to obtain a mixture, which is redissolved in dichloromethane. 2 g of 100-200 mesh silica gel is added and thoroughly mixed. After the solvent is evaporated, the crude product-silica gel powder mixture is purified by column chromatography (100-200 mesh silica gel powder as the stationary phase and dichloromethane:methanol = 20:1 (V:V) as the mobile phase) to obtain compound (10) with a yield of 64.7%. MS-ESI (M+1): 494.2141.

[0044] The results of H NMR spectrum are as follows: 1 H NMR (600 MHz, Chloroform-d) δ 6.39 (dd, J = 17.4, 11.0hz, 1H), 5.71 (d, J = 8.5hz, 1H), 5.31 – 5.26 (m, 2H), 5.16 (dd, J = 17.4, 1.6hz, 1H), 3.84 (d, J = 1.7hz, 2H), 3.34 (d, J = 6.4hz, 1H), 2.32 – 2.20 (m, 2H), 2.16 (dt, J = 19.3, 9.4hz, 1H), 2.10 – 2.07 (m, 1H), 2.03 (dd, J = 16.0, 8.6hz,1H), 1.74 (dq, J = 14.5, 3.1hz, 1H), 1.67 – 1.58 (m, 2H), 1.55 – 1.40 (m,4H), 1.39 (s, 1H), 1.37 – 1.31 (m, 1H), 1.30 (d, J = 16.1hz, 1H), 1.14 (s,3H), 1.08 (dd, J = 14.1, 4.4hz, 1H), 0.86 (d, J = 7.0hz, 3H), 0.69 (d, J =7.1hz, 3H).

[0045] Example 5: Synthesis of Compound 12A (1-cyclopropyl-6-fluoro-7-[4-(2-{[5-({2-[(3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propanecyclopenten-5-yloxy]-2-oxoethyl}thio)-1,3,4-thiadiazol-2-yl]amino}-2-oxoethyl)piperazin-1-yl]-4-oxo-1,4-dihydroquinoline-3-carboxylic acid) S9. Weigh 1.9 g of compound (6A) and dissolve it in 20 mL of DMF. Then, add 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (0.95 g), NN-diisopropylethylamine (1.13 mL), and 1-hydroxybenzotriazole (0.68 g), respectively. Lower the system temperature to 0°C and maintain stirring for 1 hour to obtain reaction solution ①. Add 1.7 g of compound (10) to 20 mL of DMF and stir to dissolve to obtain reaction solution ②. Slowly add reaction solution ② dropwise to reaction solution ① at 0°C while stirring. After the addition is complete, continue stirring at the same temperature for 10 minutes. Then add 4-dimethylaminopyridine (0.26 g), slowly warm the reaction solution to room temperature, and stir under nitrogen for 12 hours. After the reaction is complete, add 400 mL of water to the reaction solution to precipitate a white solid. Filter the filter cake, wash it with a small amount of anhydrous ether, and dry it in vacuo to obtain compound (11A).

[0046] S10. Weigh 1.81 g of compound (11A) and dissolve it in dichloromethane (20 mL). Then, add bis(triphenylphosphine)palladium chloride (30 mg), followed by water (0.2 mL) and tributyltin hydride (1.4 mL). After stirring the mixture for 15 minutes, add 100 mL of anhydrous ether to precipitate a pale yellow solid. This solid was filtered, washed with a small amount of ether, and dried under vacuum. The mixture was re-dissolved in dichloromethane, and 2 g of 100-200 mesh silica gel was added and mixed thoroughly. After complete solvent evaporation, the crude product-silica gel powder mixture was purified by column chromatography (100-200 mesh silica gel as the stationary phase and dichloromethane:ethyl acetate:methanol = 2:2:1 (V:V:V) as the mobile phase) to obtain compound (12A) in a yield of 55.8%. MS-ESI (M+1): 865.3430.

[0047] The results of H NMR spectrum are as follows: 1H NMR (600 MHz, Chloroform-d) δ 14.93 (s, 1H), 8.73 (s, 1H), 7.95 (d, J = 12.7hz, 1H), 7.39 (d, J = 7.0hz, 1H), 7.26 (s, 1H), 6.40 (dd, J =17.4, 11.0hz, 1H), 5.73 (d, J = 8.5hz, 1H), 5.29 (dd, J = 11.0, 1.5hz, 1H), 5.16 (dd, J = 17.3, 1.8hz, 1H), 4.10 (qd, J = 7.1, 0.7hz, 2H), 3.99 (d, J =2.9hz, 2H), 3.57 (tt, J = 7.3, 4.0hz, 1H), 3.42 (s, 3H), 3.33 (d, J = 7.0hz,1H), 2.88 (t, J = 4.7hz, 4H), 2.25 – 2.20 (m, 1H), 2.09 – 2.06 (m, 1H), 2.02(d, J = 0.8hz, 3H), 1.74 (dq, J = 14.6, 3.2hz, 1H), 1.63 (tdd, J = 10.8, 7.7,5.3hz, 2H), 1.52 (td, J = 13.6, 13.2, 3.5hz, 2H), 1.48 – 1.41 (m, 2H), 1.39(s, 2H), 1.33 (t, J = 14.1hz, 2H), 1.25 (d, J = 0.7hz, 1H), 1.24 (d, J =0.7hz, 2H), 1.15 (s, 3H), 1.09 (dd, J = 14.1, 4.4hz, 1H), 0.86 (d, J = 7.0hz, 3H), 0.71 (d, J = 7.0hz, 3H).

[0048] Example 6: Synthesis of Compound 12B (1-cyclopropyl-6-fluoro-7-[4-(2-{[5-({2-[(3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propanecyclopenten-5-yloxy]-2-oxoethyl}thio)-1,3,4-thiadiazol-2-yl]amino}-2-oxoethyl)-3-methylpiperazin-1-yl]-8-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid) The method was the same as in Example 5, except that an equimolar amount of compound (6B) was used instead of compound (6A). The resulting white solid was designated compound (12B) with a yield of 43.2%. MS-ESI (M-1): 907.3541. H NMR spectra were as follows: 1 H NMR (600 MHz, Chloroform-d) δ 8.82 (d, J = 11.4hz, 1H), 7.89 (dd,J = 21.9, 11.9hz, 1H), 6.42 (t, J = 14.8hz, 1H), 5.76 (d, J = 7.4hz, 1H),5.32 (t, J = 11.2hz, 1H), 5.19 (dd, J = 18.5, 5.9hz, 1H), 4.78 (s, 1H), 4.02(d, J = 7.9hz, 2H), 3.82 – 3.74 (m, 4H), 3.73 (s, 2H), 3.56 – 3.49 (m, 2H),3.47 (d, J = 11.9hz, 2H), 3.44 – 3.34 (m, 4H), 2.97 – 2.93 (m, 2H), 2.93 (s,1H), 2.84 (s, 1H), 2.79 (t, J = 10.6hz, 2H), 2.31 (d, J = 6.5hz, 1H), 2.26 –2.16 (m, 2H), 2.10 (s, 1H), 2.04 (ddt, J = 20.6, 12.8, 7.1hz, 2H), 1.76 (d, J= 14.2hz, 1H), 1.65 (q, J = 10.8hz, 3H), 1.53 (d, J = 13.5hz, 1H), 1.42 (s,3H), 1.35 (dd, J = 16.2, 4.8hz, 3H), 1.23 (d, J = 7.9hz, 2H), 1.17 (s, 1H), 1.06 (s, 1H), 0.99 (dd, J = 12.3, 5.1hz, 2H), 0.73 (q, J = 8.6, 7.7hz, 4H).

[0049] Example 7: Synthesis of Compound 12C (6-fluoro-1-(4-fluorophenyl)-7-[4-(2-{[5-({2-[(3aR,4R,5R,7S,8S,9R,9aS,12R)-8-hydroxy-4,7,9,12-tetramethyl-3-oxo-7-vinyldecahydro-4,9a-propanecyclopenten-5-yloxy]-2-oxoethyl}thio)-1,3,4-thiadiazol-2-yl]amino}-2-oxoethyl)piperazin-1-yl]-4-oxo-1,4-dihydroquinoline-3-carboxylic acid) The method was the same as in Example 5, except that an equimolar amount of compound (6C) was used instead of compound (6A). The resulting white solid, designated compound (12C), was obtained in a 45.7% yield. MS-ESI (M-1): 917.3173. H NMR spectra were as follows: 1H NMR (600 MHz, Chloroform-d) δ 14.84 (s, 1H), 8.66 (s, 1H), 8.04 (d, J = 12.7hz, 1H), 7.50 (dd, J = 8.7, 4.5hz, 2H), 7.39 (t, J = 8.2hz, 2H), 6.42 (dd, J = 17.4, 11.0hz, 1H), 6.38 (d, J = 6.8hz, 1H), 5.75 (d, J = 8.5hz, 1H), 5.31 (dd, J = 11.0, 1.5hz, 1H), 5.18 (dd, J = 17.5, 1.6hz, 1H), 4.12 (q,J = 7.1hz, 1H), 4.00 (s, 2H), 3.37 (s, 3H), 3.22 – 3.17 (m, 4H), 2.79 (t, J =4.8hz, 4H), 2.33 – 2.28 (m, 1H), 2.27 – 2.21 (m, 1H), 2.18 (dd, J = 19.5,9.4hz, 1H), 2.11 – 2.09 (m, 1H), 2.05 (d, J = 4.5hz, 2H), 1.79 – 1.75 (m,1H), 1.65 (ddt, J = 13.1, 9.7, 5.2hz, 2H), 1.41 (s, 3H), 1.33 (d, J = 16.1hz,1H), 1.27 (q, J = 8.1, 7.1hz, 3H), 1.16 (s, 3H), 0.88 (d, J = 6.9hz, 3H), 0.73 (d, J = 7.0hz, 3H).

[0050] Example 8: Study on the in vitro antibacterial activity of some target compounds of the present invention 1. Test method: The minimum inhibitory concentration (MIC) of the test strains was determined by the broth two-fold dilution method. The bacterial inoculum size was 10 5 CFU / ml, each drug was tested three times for each bacterium. The test substance was dissolved with dimethyl sulfoxide and Tween 80, and then prepared into a solution with sterile distilled water or lower alcohol.

[0051] 2. Test strains: 8 laboratory strains (including 4 Gram-positive bacteria (G + ), 4 strains of Gram-negative bacteria (G -Staphylococcus aureus ATCC 29213, Methicillin-resistant Staphylococcus aureus (MRSA) USA300, Methicillin-resistant Staphylococcus aureus (MRSA) 43300, Methicillin-resistant Staphylococcus aureus (MRSA) 16183, Klebsiella pneumoniae pneumonia S11-60, Klebsiella pneumoniae pneumonia S11-62, Klebsiella pneumoniae pneumonia S11-64, Klebsiella pneumoniae pneumonia S12-14; 3. The positive control drugs were ciprofloxacin and pleuromutilin. The MIC values ​​of each compound are listed in Table 1.

[0052] Table 1 In vitro antibacterial activity data of some target compounds (MIC, μg / ml)

[0053] As shown in Table 1, the compounds of the present invention have obvious antibacterial activity. + Bacteria: The pleuromutilin derivative (compound 12A) was superior to the control drugs ciprofloxacin and pleuromutilin against the tested Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA), especially against MRSA 16183. - Bacteria: Compound 12A was superior to the control drugs ciprofloxacin and pleuromutilin against the four tested Gram-negative bacteria.

[0054] Application Example 1 An antimicrobial active premix A premix was prepared by mixing 1 part of compound (12), 1 part of polyvinyl pyrrolidone and 5 parts of starch according to conventional methods in the art.

[0055] Compared with the existing technology, the present invention connects two different lead compounds together through a covalent bond to produce a synergistic effect, an additive effect, or a new pharmacological activity in the body; the present invention splices quinolone drugs onto the side chain of pleuromutilin to produce new pleuromutilin heteroderivatives. The obtained new compounds can broaden the antibacterial spectrum and have good antibacterial activity against both Gram-positive and Gram-negative bacteria, and are effective against drug-resistant bacteria.

[0056] The above embodiments are only preferred embodiments of the present invention and are not intended to limit the patent scope of the present invention. The compounds or preparation methods mentioned in the present invention are not limited to the above ones. Therefore, the above embodiments cannot be regarded as limiting the scope of protection of the present invention. All equivalent structural transformations made using the contents of the present invention specification under the inventive concept of the present invention, or direct / indirect applications in other related technical fields are included in the patent protection scope of the present invention.

Claims

1. A pleuromutilin derivative having the structure of Formula I, or a pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, or polymorph thereof; Formula I; in: R1 is selected from a hydrogen atom and a methyl group; R2 is selected from a hydrogen atom and a methoxy group; R3 is selected from a cyclopropyl group and a p-fluorophenyl group.

2. The pleuromutilin derivative according to claim 1, or its pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, or polymorphic compound, characterized in that: Having the following structure (A), (B) or (C): Formula A; Formula B; Formula C.

3. A method for preparing the pleuromutilin derivative according to any one of claims 1 to 2, or a pharmaceutically and / or veterinarily acceptable salt, solvate, optical isomer, or polymorph thereof, characterized in that: The synthetic route is as follows: 。 4. The preparation method according to claim 3, characterized in that The steps include: S1: Compound 1 is used as a raw material, and in the presence of a base, a solvent is used to react with di-tert-butyl dicarbonate to produce compound 2; wherein compound 1 is ciprofloxacin, gatifloxacin or sarafloxacin; S2: Compound 2 prepared in step S1 is subjected to a substitution reaction with allyl bromide to prepare compound 3; S3: dissolving compound 3 obtained in step S2 in dichloromethane and reacting with an acid to obtain compound 4; S4: Compound 4 obtained in step S3 is dissolved in acetonitrile, and then reacted with tert-butyl bromoacetate in the presence of NN-diisopropylethylamine to obtain compound 5; S5: dissolving compound 5 obtained in step S4 in dichloromethane and reacting with an acid to obtain compound 6; S6: Compound 7-pleuromutilin was used as a raw material and reacted with p-toluenesulfonyl chloride in pyridine to generate compound 8; S7: dissolving the compound 8 obtained in step S6 in acetonitrile and iodizing with NaI in the presence of potassium carbonate to obtain compound 9; S8: reacting the compound 9 obtained in step S7 with 2-amino-5-mercapto-1,3,4-thiadiazole to obtain compound 10; S9: Compound 6 prepared in step S5 is dissolved in DMF, and reacted with compound 10 prepared in step S8 in the presence of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, NN-diisopropylethylamine, 1-hydroxybenzotriazole and 4-dimethylaminopyridine to prepare compound 11; S10: Compound 11 obtained in step S9 is dissolved in dichloromethane and reacted with tributyltin hydride in the presence of bis(triphenylphosphine)palladium chloride and water to obtain the final product 12.

5. The preparation method according to claim 4, characterized in that The base in step S1 is one of NaHCO3, K2CO3, NaOH, KOH, triethylamine, pyridine, piperidine, and dicyclohexylamine; the solvent is at least one of DMF, dichloromethane, chloroform, 1,2-dichloroethane, diethyl ether, 1,4-dioxane, and methyl tert-butyl ether; The acid in steps S3 and S5 is one of trifluoroacetic acid, HCl, H2SO4, HBr, and phosphoric acid.

6. The preparation method according to claim 4, characterized in that The reaction temperature in step S1 is 25 to 60° C., and the molar ratio of di-tert-butyl dicarbonate to compound 1 is 1:1 to 5:3; In step S9, the feeding temperature before the reaction is -5 to 5°C, the reaction temperature is 25°C to 90°C, the molar ratio of compound 6 to 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is 1:(1.1 to 1.8), the molar ratio of compound 6 to NN-diisopropylethylamine is 1:(1.5 to 1.8), the molar ratio of compound 6 to 1-hydroxybenzotriazole is 1:(1.1 to 1.8), the molar ratio of compound 6 to 4-dimethylaminopyridine is 1:(0.5 to 0.8), and the molar ratio of compound 6 to compound 10 is (1.2 to 1.5):

1.

7. An antimicrobial active composition, characterized in that: The invention comprises the pleuromutilin derivative according to any one of claims 1 to 2 and / or its pharmaceutically and / or veterinarily acceptable salts, solvent compounds, optical isomers, polymorphic compounds and pharmaceutically acceptable carriers or diluents.

8. The antimicrobial active composition according to claim 7, characterized in that Its dosage form is oral or injection.

9. The antimicrobial active composition according to claim 8, characterized in that The content of the pleuromutilin derivative or its pharmaceutically and / or veterinarily acceptable salt, solvent compound, optical isomer, or polymorph compound is 0.1%-99.5% (wt%), and the remainder is a carrier or diluent.

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