Demethylhalaman-enoxacin conjugate as well as preparation method and application thereof

Abstract

The invention relates to a norhalal-enoxacin conjugate as well as a preparation method and application thereof. The norhalal-enoxacin conjugate comprises a norhalal-enoxacin coupling compound, or a stereoisomer, a prodrug, a crystal form, a pharmaceutically acceptable salt, a pharmaceutically acceptable ester or a pharmaceutically acceptable solvate selected from the compound. Compared with enoxacin, kanamycin, streptomycin, lincomycin and azithromycin, the norharmann-enoxacin conjugate disclosed by the invention has a smaller MIC (Minimum Inhibitory Concentration) value, which shows that the norharmann-enoxacin conjugate has a better anti-staphylococcus aureus effect.

Description

Technical Field

[0001] The present invention relates to the technical field of medicines, in particular to a norharman-enoxacin conjugate and a preparation method and application thereof. Background Art

[0002] Currently, the main anti-Staphylococcus aureus drugs include penicillins, vancomycins, aminoglycosides, tetracyclines, quinolones, lincosamides, and macrolides. However, drug resistance in S. aureus is becoming increasingly common, and the drug dosages required to treat infections are increasing. Infections caused by this bacterium pose a serious threat to human health, necessitating the development of new drugs for the treatment of S. aureus infections. Summary of the Invention

[0003] In view of the shortcomings of the existing technology, the first object of the present invention is to provide a norharman-enoxacin conjugate, which has a good anti-Staphylococcus aureus effect.

[0004] The second object of the present invention is to provide a method for preparing a norharman-enoxacin conjugate, which has the advantages of simple steps and high yield.

[0005] The third object of the present invention is to provide an application of a norharman-enoxacin conjugate, which has the advantage of treating Staphylococcus aureus infection.

[0006] To achieve the above first object, the present invention provides the following technical solutions: A norharman-enoxacin conjugate selected from the compound represented by the following formula (I), or selected from the stereoisomers, prodrugs, crystal forms, pharmaceutically acceptable salts, pharmaceutically acceptable esters or pharmaceutically acceptable solvates of the compound; .

[0007] Specifically, in the "norharman-enoxacin conjugate" of the present invention, The specific meaning of the "stereoisomer" is that when the chemical name is specifically designated as (R)- or (S)-isomer, it should be understood that the main configuration is (R)-isomer or (S)-isomer respectively; any asymmetric carbon atom may exist in (R)-, (S)- or (R, S)-configuration, preferably in (R)- or (S)-configuration; The term "crystal form" specifically refers to the crystal structure of a substance. During crystallization, a substance may be affected by various factors, which may change the intramolecular or intermolecular bonding pattern, resulting in different arrangements of molecules or atoms in the crystal lattice space, thus forming different crystal structures. The compounds of the present invention may exist in one crystal structure or in multiple crystal structures, i.e., they may have "polymorphism." The compounds of the present invention may exist in different crystal forms. The specific meaning of the "pharmaceutically acceptable salt" refers to a salt formed by an acidic functional group (such as a carboxyl group) present in the compound provided by the present invention and a suitable inorganic or organic cation (base), or a salt formed by a basic functional group (such as an amino group) present in the compound provided by the present invention and a suitable inorganic or organic anion (acid); non-limiting examples include quaternary ammonium salts, acetates, adipates, camphorates, alginate, citrate, aspartate, benzoate, benzenesulfonate, maleate, hydrogen sulfate, one of butyrate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, 2-hydroxyethanesulfonate (isothiosulfate), lactate, methanesulfonate, hydroiodide, nicotinate, 2-naphthalenesulfonate, oxalate, palmitate, pectinate, persulfate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, phosphate, 3-phenylpropionate, glutamate, bicarbonate, p-toluenesulfonate, and undecanoate; Wherein, the quaternary ammonium salt is composed of C1~C6 alkyl halide, dialkyl sulfate, C 12 ~C 18 Alkyl halides and C6~C 10 One of the aryl halides is prepared by quaternization; the alkyl group of the C1~C6 alkyl halide is one of methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, tert-butyl and isobutyl, and the halide is one of chloride, bromide and iodide; the dialkyl sulfate is one of dimethyl sulfate, diethyl sulfate, dibutyl sulfate and dipentyl sulfate; the C 12 ~C 18 The alkyl group of the alkyl halide is one of dodecyl, decyl, tetradecyl and octadecyl, and the halide is one of chloride, bromide and iodide; the C6~C 10 The aryl halide is one of benzyl bromide and phenethyl bromide; Meanwhile, the acid in the salt is one of hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, oxalic acid, maleic acid, succinic acid and citric acid; The term "pharmaceutically acceptable ester" specifically refers to an ester formed between a carboxyl group present in the compounds provided by the present invention and a suitable alcohol; non-limiting examples include formates, acetates, propionates, butyrates, acrylates, ethylsuccinates, stearic acid esters, or palmitates; such esters can undergo hydrolysis in the presence of an acid or base to generate the corresponding acid or alcohol; The specific meaning of the "pharmaceutically acceptable solvate" refers to a compound that exists in combination with a certain solvent molecule. "Solvate" or "solvate" can be used interchangeably; the combination can include a stoichiometric amount of a certain solvent, such as a monohydrate or a dihydrate, or can include any amount of water; for example, methanol or ethanol can form an "alcoholate", which can also be stoichiometric or non-stoichiometric. The term "solvate" as used herein refers to a solid form, that is, a compound in a solution of a solvent, although it can be solvated, it is not a solvate as the term is used herein.

[0008] To achieve the above second purpose, the present invention provides the following technical solutions: A method for preparing a norharman-enoxacin conjugate, the route of the preparation method is as follows: ; The preparation method comprises the following steps: S1 is subjected to a cyclization reaction between L-tryptophan methyl ester hydrochloride and formaldehyde in an organic solvent, and after the reaction is completed, the compound of the above formula (1) is obtained by post-treatment; S2 In an organic solvent, the compound of formula (1) is subjected to oxidation reaction under the action of triethylamine (Et3N) and trichloroisocyanuric acid (TCCA), and after the reaction is completed, the compound of formula (2) is obtained by post-treatment; S3: In an organic solvent, the compound of formula (2) is subjected to a hydrolysis reaction under the action of sodium hydroxide (NaOH), and after the reaction is completed, the compound of formula (3) is obtained by post-treatment; S4 In an organic solvent, the compound of the above formula (3) and the compound of the above formula (II) undergo an amine-acid condensation reaction in the presence of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI), 1-hydroxybenzotriazole (HoBt), and triethylamine (Et3N). After the reaction is completed, the compound of the above formula (I) is obtained by post-treatment.

[0009] Furthermore, in the S1, L-tryptophan methyl ester hydrochloride is dissolved in methanol, and formaldehyde solution is added to carry out cyclization reaction. The mixture is heated to 80°C and refluxed for 6 hours. After rotary evaporation to obtain a concentrated solution, 5 times the volume of methyl tert-butyl ether is added, and a large amount of gray solid is precipitated. The solid is stirred for recrystallization, filtered and dried to obtain the compound of the above formula (1).

[0010] Furthermore, in S2, the compound of formula (1) and N,N-dimethylformamide were stirred in an ice-salt bath at -20°C for 5 minutes, and then triethylamine and trichloroisocyanuric acid solution were added. The reaction was continued for 0.5 hours, and then in ice water at 0°C for 2 hours. The reaction solution was transferred to ice water and the pH was adjusted to 9 with triethylamine. During this period, a yellow-white solid was precipitated, which was filtered to obtain the compound of formula (2).

[0011] Furthermore, in S3, the compound of formula (2), methanol and sodium hydroxide solution are mixed, and the oil bath is heated to 80°C. The mixed solution changes from a muddy yellow suspension to a clear solution, and the mixture is reacted for 2 hours. After cooling, the pH is adjusted to 3-4 with hydrochloric acid, during which a white solid precipitates. The mixture is filtered to obtain the compound of formula (3).

[0012] Furthermore, in the S4, the compound of the above formula (3) is dissolved in N,N-dimethylformamide, and then 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride, 1-hydroxybenzotriazole and triethylamine are added in sequence, and stirred at room temperature for 0.5 h. Then, the compound of the above formula (II) is added, and the mixture is reacted at 50° C. for 3 h. Dichloromethane and water are added for extraction 3 times, and the organic phase is back-extracted with saturated brine 5 times. The organic phase is dried over anhydrous sodium sulfate, filtered and eluted by spin drying to obtain the compound of the above formula (I).

[0013] To achieve the third objective, the present invention provides the following technical solutions: A desmethylharman-enoxacin conjugate is used in the preparation of a drug for resisting Staphylococcus aureus infection.

[0014] Specifically, in the "drug" of the present invention, The drug can be introduced into the body by injection, spraying, nasal drops, eye drops, penetration, absorption, physical or chemical mediation methods such as muscle, intradermal, subcutaneous, intravenous, mucosal tissue; or can be introduced into the body after being mixed or encapsulated with other substances; When necessary, one or more pharmaceutically acceptable carriers may be added to the drug. Such carriers include conventional diluents, excipients, fillers, binders, wetting agents, disintegrants, absorption enhancers, surfactants, adsorption carriers, lubricants, and the like. The drug can be prepared in various forms, such as injections, suspensions, powders, tablets, and granules. The various dosage forms described above can be prepared according to conventional methods in the pharmaceutical field.

[0015] Furthermore, the Staphylococcus aureus is Staphylococcus aureus ATCC25923.

[0016] Furthermore, the active ingredient of the drug is a compound represented by the above formula (I), or is selected from stereoisomers, prodrugs, crystal forms, pharmaceutically acceptable salts, pharmaceutically acceptable esters or pharmaceutically acceptable solvates of the compound.

[0017] In summary, the beneficial technical effects of the present invention are: The norharman-enoxacin conjugate of the present invention has a smaller MIC value than enoxacin, kanamycin, streptomycin, lincomycin and azithromycin, indicating that it has a better anti-Staphylococcus aureus effect; The preparation method of the present invention has simple steps, convenient operation, easy industrial production, high yield, and is suitable for large-scale production applications; The norharman-enoxacin conjugate of the present invention can be used to prepare drugs for treating Staphylococcus aureus infections. The conjugate also has low toxicity and good biocompatibility, ensuring its safe application in the medical field. Experimental verification shows that the norharman-enoxacin conjugate provided by the present invention exhibits significant anti-Staphylococcus aureus activity and is expected to become a new drug for treating Staphylococcus aureus infections, providing a new treatment option for clinical practice. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 This is the hydrogen nuclear magnetic resonance spectrum of the compound of formula (1) prepared in Example 3 of the present invention.

[0019] Figure 2 This is the carbon NMR spectrum of the compound of formula (1) obtained in Example 3 of the present invention.

[0020] Figure 3 This is a high-resolution mass spectrum of the compound of formula (1) obtained in Example 3 of the present invention.

[0021] Figure 4 This is the hydrogen nuclear magnetic resonance spectrum of the compound of formula (2) prepared in Example 3 of the present invention.

[0022] Figure 5 This is the carbon NMR spectrum of the compound of formula (2) obtained in Example 3 of the present invention.

[0023] Figure 6 This is a high-resolution mass spectrum of the compound of formula (2) obtained in Example 3 of the present invention.

[0024] Figure 7 This is the hydrogen nuclear magnetic resonance spectrum of the compound of formula (3) prepared in Example 3 of the present invention.

[0025] Figure 8 This is the carbon NMR spectrum of the compound of formula (3) prepared in Example 3 of the present invention.

[0026] Figure 9 This is a high-resolution mass spectrum of the compound of formula (3) obtained in Example 3 of the present invention.

[0027] Figure 10 This is the hydrogen nuclear magnetic resonance spectrum of the compound of formula (I) prepared in Example 3 of the present invention.

[0028] Figure 11 This is the carbon NMR spectrum of the compound of formula (I) prepared in Example 3 of the present invention.

[0029] Figure 12 This is a high-resolution mass spectrum of the compound of formula (I) prepared in Example 3 of the present invention. DETAILED DESCRIPTION

[0030] In order to make the technical means, creative features, objectives and functions achieved by the present invention clearer and easier to understand, the present invention is further explained below with reference to the accompanying drawings and specific implementation methods.

[0031] Example 1: A norharman-enoxacin conjugate disclosed in the present invention, selected from the compound represented by the following formula (I), or selected from the stereoisomers, prodrugs, crystal forms, pharmaceutically acceptable salts, pharmaceutically acceptable esters or pharmaceutically acceptable solvates of the compound; .

[0032] Example 2: A method for preparing a norharman-enoxacin conjugate disclosed in the present invention. The preparation method is as follows: ; The preparation method comprises the following steps, S1 is subjected to a cyclization reaction between L-tryptophan methyl ester hydrochloride and formaldehyde in an organic solvent, and after the reaction is completed, the compound of the above formula (1) is obtained by post-treatment; S2 In an organic solvent, the compound of formula (1) is subjected to oxidation reaction under the action of triethylamine (Et3N) and trichloroisocyanuric acid (TCCA), and after the reaction is completed, the compound of formula (2) is obtained by post-treatment; S3: In an organic solvent, the compound of formula (2) is subjected to a hydrolysis reaction under the action of sodium hydroxide (NaOH), and after the reaction is completed, the compound of formula (3) is obtained by post-treatment; S4 In an organic solvent, the compound of the above formula (3) and the compound of the above formula (II) undergo an amine-acid condensation reaction in the presence of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI), 1-hydroxybenzotriazole (HoBt), and triethylamine (Et3N). After the reaction is completed, the compound of the above formula (I) is obtained by post-treatment.

[0033] Example 3: A method for preparing a norharman-enoxacin conjugate disclosed in the present invention, which is different from Example 2 in that the specific implementation of S1 is: Pour L-tryptophan methyl ester hydrochloride (5g, 19.6mmol, 1.0eq) and 100mL of methanol solution into a 250mL eggplant flask, add a magnetic stirrer to stir until the raw materials are fully dissolved, then add 37% formaldehyde solution (2.2mL, 21.6mmol, 1.1eq), heat to 80℃ and reflux for 6h, monitor the reaction until complete, and rotary evaporate the solvent in the bottle until 10~15mL of solution remains. The rotary evaporation process is accompanied by the precipitation of some solids. Add 5 times the volume of methyl tert-butyl ether (60mL) to the rotary evaporation concentrate, and a large amount of gray solid precipitates. Stir on the stirrer for 10min to ensure sufficient recrystallization of the product. Finally filter and dry, refer to Figures 1-3 , 4.55 g of the off-white compound of formula (1) was obtained, with a yield of 87.3%.

[0034] 1 H NMR (400 MHz, DMSO-d6) δ 11.12 (s, 1H), 7.49 (d, J = 7.8 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.12 (t, J = 7.6 Hz, 1H), 7.03 (t, J = 7.4 Hz, 1H), 4.64 (dd, J = 10.3, 5.3 Hz, 1H), 4.41 (s, 2H), 3.84 (s, 3H), 3.06 (dd, J =16.1, 10.3 Hz, 1H); 13 C NMR (126 MHz, DMSO-d6) δ 169.50, 136.72, 126.87,126.19, 122.19, 119.50, 118.35, 111.89, 104.71, 54.12, 53.52, 22.39; HRMS(ESI) calcd for C13H14N2O2, [M+H]+: m / z 231.1134, found: 231.1130. Example 4: A method for preparing a norharman-enoxacin conjugate disclosed in the present invention is different from Example 3 in that S2 is specifically implemented by adding the compound of formula (1) (3.0 g, 11.2 mmol, 1.0 eq) and 60 mL of N,N-dimethylformamide (DMF) solvent to a 250 mL eggplant-shaped flask, stirring for 5 min in an ice-salt bath at -20°C, and then adding triethylamine (3.92 mL, 28.2 mmol, 2.5 eq). ; Then, weigh trichloroisocyanuric acid (2.62g, 11.2mmol, 1.0eq) at room temperature and place it in a 20mL sample bottle. Add 10mL of DMF solvent and shake thoroughly to dissolve it to obtain a clear solution. After the sample bottle is also placed at -20℃ and cooled for 10min, the DMF solution of trichloroisocyanuric acid (TCCA) is slowly added to the reaction bottle with a syringe. Keep the reaction under this condition for 0.5h, change to 0℃ ice water condition and react for 2h. The reaction is completed by spot plate detection. The reaction solution is poured into ice water with a volume of 10 times that of DMF, and the pH is adjusted to 9 with an appropriate amount of triethylamine. During this period, a yellow-white solid precipitates. Figures 4-6 Finally, the mixture was filtered and washed with water to obtain 2.07 g of the yellow-white compound of formula (2) with a yield of 81.5%.

[0035] 1 H NMR (400 MHz, DMSO-d6) δ 12.09 (s, 1H), 8.98 (s, 1H), 8.93 (s, 1H), 8.41 (d, J = 7.9 Hz, 1H), 7.68 (d, J = 8.2 Hz, 1H), 7.62 (d, J = 8.0 Hz, 1H),7.32 (t, J = 7.4 Hz, 1H), 3.92 (s, 3H); 13 C NMR (126 MHz, DMSO-d6) δ 166.52,141.44, 137.92, 136.95, 134.22, 129.16, 127.96, 122.69, 121.37, 120.70,118.11, 112.87, 52.41; HRMS (ESI) calcd for C13H10N2O2, [M+Na]+: m / z249.0640, found: 249.0649. Example 5: A method for preparing a norharman-enoxacin conjugate disclosed in the present invention. The difference from Example 4 is that the specific implementation of S3 is to weigh the compound of formula (2) (0.5g) into a 100mL eggplant-shaped bottle, and add 10mL of methanol and a 2M NaOH mixed solution (v / v=1:1). The system is initially a muddy yellow suspension. The reaction bottle is placed in an oil bath. After the temperature is slowly raised to 80°C, the system changes from turbidity to clarity. The reaction is continued for 2h. After the reaction solution is cooled, the pH is adjusted to 3-4 with 1M HCl. A large amount of white solid will precipitate. Figures 7-9 Finally, the product was filtered and washed with water to obtain 30.35 g of off-white reaction product with a yield of 81.5%.

[0036] 1 H NMR (500 MHz, DMSO-d6) δ 12.10 (s, 1H), 8.97 (s, 1H), 8.93 (s, 1H), 8.41 (d, J = 7.9 Hz, 1H), 7.68 (d, J = 8.3 Hz, 1H), 7.61 (t, J = 7.5 Hz, 1H),7.32 (t, J = 7.4 Hz, 1H); 13 C NMR (126 MHz, DMSO-d6) δ 167.21, 141.57, 137.86,137.54, 133.59, 129.24, 128.36, 122.73, 121.35, 120.69, 117.77, 112.87; HRMS(ESI) calcd for C13H14N2O2, [M+Na]+: m / z 235.0483, found: 235.0486. Example 6: A method for preparing a norharman-enoxacin conjugate disclosed in the present invention is different from Example 5 in that S4 is specifically implemented as follows: weigh the compound of formula (3) (100 mg, 0.47 mmol, 1.0 eq) in a 100 mL eggplant flask, add a magnetic stirrer and 20 mL of DMF solvent, stir to fully dissolve it, then add 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI) (108 mg, 0.57 mmol, 1.2 eq) and 1-hydroxybenzotriazole (HoBt) (96 mg, 0.71 mmol, 1.5 eq), triethylamine (130 μL, 0.94 mmol, 2 eq) in sequence, stir the mixture at room temperature for 0.5 h, then add the compound of formula (II) (enoxacin, 151 mg, 0.47 mmol, 1.0 eq), and gradually heat the system to 50°C. The reaction is terminated after plate monitoring for 3 h. Add dichloromethane and water to the system and extract 3 times, retain the organic phase, and then back-extract 5 times with saturated brine, dry the organic phase with anhydrous sodium sulfate, filter and spin dry, and then load the column, using dichloromethane and methanol as eluents, refer to Figures 10-12 , 125 mg of the compound of formula (I) was collected, with a yield of 52%.

[0037] 1 H NMR (500 MHz, DMSO-d6) δ 15.31 (s, 1H), 11.89 (s, 1H), 8.96 (s,1H), 8.90 (s, 1H), 8.52 (s, 1H), 8.33 (d, J = 7.8 Hz, 1H), 8.07 (d, J = 13.3Hz, 1H), 7.64 (d, J = 8.3 Hz, 1H), 7.59 (t, J = 7.5 Hz, 1H), 7.29 (t, J = 7.4Hz, 1H), 4.48 (q, J = 7.1 Hz, 2H), 3.98 (d, J = 8.6 Hz, 2H), 3.90 (t, J = 9.2Hz, 6H), 1.37 (t, J = 7.0 Hz, 3H); 13C NMR (126 MHz, DMSO-d6) δ 176.82,168.48, 166.32, 150.43, 148.16, 145.25, 142.87, 141.45, 136.57, 132.45,129.08, 128.46, HRMS (ESI) calcd for C27H23FN6O4,[M+Na] + : m / z 537.1663, found:537.1682. Example 7: Use of a norharman-enoxacin conjugate disclosed in the present invention in the preparation of drugs for resisting Staphylococcus aureus infection.

[0038] To verify the activity of the compound of formula (I) against Staphylococcus aureus ATCC25923 infection, the following test was performed.

[0039] 1) Preparation of culture medium: LB medium: Take 10 g / L NaCl, 10 g / L peptone, 5 g / L yeast powder, and water as solvent, mix well, adjust the pH to about 7.2 ± 0.2 with 1 mol / L NaOH solution at 25 ° C, divide into 250 mL conical flasks, and incubate at 121 ° C, 1 × 10 5 Sterilize at 4°C for 20 min and set aside.

[0040] LB agar medium: Take 10 g / L NaCl, 10 g / L peptone, 5 g / L yeast powder, and water as solvent, mix well, then add 1.5-2.0% agar powder, adjust the pH to about 7.2±0.2 with 1 mol / L NaOH solution at 25°C, and heat at 121°C, 1×10 5 Sterilize under Pa for 20 min and pour into plates for later use.

[0041] 2) Preparation of bacterial suspension: Staphylococcus aureus ATCC25923 strain (provided by Nanjing Medical University) stored at -80℃ was thawed at room temperature and streaked onto LB agar medium. It was cultured at 37℃ for 24 hours. A well-developed single colony was taken and inoculated again. It was cultured at 37℃ for 24 hours to ensure that the strain was in the growth phase. A single colony was selected and inoculated into LB medium. It was cultured at 37℃ and 220 rpm for 16 hours. The 10% lysate was diluted with fresh LB medium. 6The Staphylococcus aureus solution was obtained by folding, and the concentration was determined to be 1.5×10 9 CFU / mL.

[0042] 3) Prepare a solution of the compound of formula (I) to be tested: The compound of formula (I) prepared in Example 6, enoxacin, kanamycin, streptomycin, lincomycin and azithromycin were prepared into 1 mg / mL stock solutions using sterile DMSO as solvent and stored in a -20°C refrigerator for future use.

[0043] Determination of the minimum inhibitory concentration of compound I against Staphylococcus aureus: Prepare a sterile 96-well plate for antibacterial activity testing. Add the stock solution to the second column of the 96-well plate (flat bottom). Add 2 μL of the stock solution to the second row of the second column. Each group of three replicates. Add 200 μL of Staphylococcus aureus solution (1.5×10 9 CFU / mL), add 100μL of bacterial solution to columns 3 to 10, repeatedly pipette and beat each well in column 2 five times to mix, draw 100μL from each well and add it to column 3, mix, draw 100μL again and add it to column 4, and so on, draw 100μL from column 9 and discard. Column 10 is the growth control well, add 200μL of bacterial solution, and add 200μL of blank culture medium to the outermost circle of the 96-well plate. Make the final concentration of the compound determination be 10, 5, 2.5, 1.25, 0.63, 0.31, 0.16, 0.078μg / mL of dilution. Place the prepared 96-well plates in a 37°C constant temperature incubator and culture for 18~20h, then measure the OD with a microplate reader. 600 The results were recorded. All experiments were repeated three times.

[0044] Using enoxacin as a positive control drug for the detection of Staphylococcus aureus, the above step a was repeated to make the final concentration of enoxacin in the dilution solution be 10, 5, 2.5, 1.25, 0.63, 0.31, 0.16, and 0.078 μg / mL.

[0045] Use kanamycin, streptomycin, lincomycin and azithromycin as positive control drugs when detecting Staphylococcus aureus. Repeat the above step a so that the final concentration of each positive drug is 128, 64, 32, 16, 8, 4, 2, and 1 μg / mL dilution.

[0046] Table 1 MIC value (μg / mL) Compound of formula (I) Enoxacin Kanamycin Streptomycin Lincomycin Azithromycin S. aureus ATCC25923 1.25 2.5 4 16 2 8 As shown in Table 1, the compound represented by Formula I provided by the present invention has excellent anti-Staphylococcus aureus effect.

[0047] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not limiting. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the purpose and scope of the technical solutions of the present invention, which should all be included in the scope of the claims of the present invention.

Claims

1. A norharman-enoxacin conjugate, characterized in that: A compound selected from the following formula (I), or a stereoisomer, prodrug, crystal form, pharmaceutically acceptable salt, pharmaceutically acceptable ester or pharmaceutically acceptable solvate of the compound; 。 2. A norharman-enoxacin conjugate according to claim 1, characterized in that: The salt is one of quaternary ammonium salt, acetate, adipate, camphorate, alginate, citrate, aspartate, benzoate, benzenesulfonate, maleate, bisulfate, butyrate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, 2-hydroxyethanesulfonate (isothiosulfate), lactate, methanesulfonate, hydroiodide, nicotinate, 2-naphthalenesulfonate, oxalate, palmitate, pectinate, persulfate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, phosphate, 3-phenylpropionate, glutamate, bicarbonate, p-toluenesulfonate and undecanoate.

3. The method for preparing a norharman-enoxacin conjugate according to any one of claims 1 to 2, characterized in that: The route of the preparation method is as follows, ； The preparation method comprises the following steps: S1 is subjected to a cyclization reaction between L-tryptophan methyl ester hydrochloride and formaldehyde in an organic solvent, and after the reaction is completed, the compound of the above formula (1) is obtained by post-treatment; S2: In an organic solvent, the compound of formula (1) is subjected to oxidation reaction under the action of triethylamine and trichloroisocyanuric acid, and after the reaction is completed, the compound of formula (2) is obtained by post-treatment; S3: In an organic solvent, the compound of formula (2) is subjected to a hydrolysis reaction under the action of sodium hydroxide, and after the reaction is completed, the compound of formula (3) is obtained by post-treatment; S4 In an organic solvent, the compound of the above formula (3) and the compound of the above formula (II) undergo an amine-acid condensation reaction in the presence of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride, 1-hydroxybenzotriazole and triethylamine. After the reaction is completed, the compound of the above formula (I) is obtained by post-treatment.

4. The method for preparing a norharman-enoxacin conjugate according to claim 3, wherein: In the S1, L-tryptophan methyl ester hydrochloride is dissolved in methanol, and formaldehyde solution is added to carry out cyclization reaction. The mixture is heated to 80°C and refluxed for 6 hours. After rotary evaporation to obtain a concentrated solution, 5 times the volume of methyl tert-butyl ether is added. A large amount of gray solid is precipitated, which is stirred for recrystallization, filtered and dried to obtain the compound of formula (1).

5. The method for preparing a norharman-enoxacin conjugate according to claim 3, wherein: In S2, the compound of formula (1) and N,N-dimethylformamide are stirred in an ice-salt bath at -20°C for 5 minutes, and then triethylamine and trichloroisocyanuric acid solution are added. The reaction is continued for 0.5 hours, and then in ice water at 0°C for 2 hours. The reaction solution is transferred to ice water and the pH is adjusted to 9 with triethylamine. During this period, a yellow-white solid is precipitated, which is filtered to obtain the compound of formula (2).

6. The method for preparing a norharman-enoxacin conjugate according to claim 3, wherein: In the S3, the compound of formula (2), methanol and sodium hydroxide solution are mixed, and the oil bath is heated to 80°C. The mixed solution changes from a muddy yellow suspension to a clear state. The mixture is reacted for 2 hours. After cooling, the pH is adjusted to 3-4 with hydrochloric acid. During this period, a white solid precipitates, which is filtered to obtain the compound of formula (3).

7. The method for preparing a norharman-enoxacin conjugate according to claim 3, wherein: In the S4, the compound of the above formula (3) is dissolved in N,N-dimethylformamide, and then 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride, 1-hydroxybenzotriazole and triethylamine are added in sequence. The mixture is stirred at room temperature for 0.5 h, and then the compound of the above formula (II) is added. The mixture is reacted at 50° C. for 3 h. Dichloromethane and water are added for extraction 3 times. The organic phase is back-extracted with saturated brine 5 times. The organic phase is dried over anhydrous sodium sulfate, filtered and eluted by rotary evaporation to obtain the compound of the above formula (I).

8. Use of the norharman-enoxacin conjugate according to any one of claims 1 to 2 in the preparation of a drug for resisting Staphylococcus aureus infection.

9. The use of a norharman-enoxacin conjugate according to claim 8, characterized in that: The Staphylococcus aureus is Staphylococcus aureus S. aureus ATCC25923.

10. The use of a norharman-enoxacin conjugate according to claim 8, characterized in that: The active ingredient of the drug is the compound represented by the above formula (I), or is selected from the stereoisomers, prodrugs, crystal forms, pharmaceutically acceptable salts, pharmaceutically acceptable esters or pharmaceutically acceptable solvates of the compound.

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