Alpha-MG-based xanthone NO donor compound as well as synthesis method and application of alpha-MG-based xanthone NO donor compound

Abstract

The invention provides an alpha-MG-based xanthone NO donor compound, a synthesis method and application, and belongs to the technical field of biological medicines.The synthesis method comprises the steps that alpha-MG serves as a starting raw material, symmetric and asymmetric modification is conducted on the C3 site or / and the C6 site of a xanthone alpha-MG structure, an NO donor is introduced, and the alpha-MG-based xanthone NO donor compound is obtained. And synthesizing the xanthone NO donor compound based on the alpha-MG. The alpha-MG-based xanthone NO donor compound is used as a sensitizing agent for resisting gram-positive bacteria or serving as a clinical drug-resistant strain MRSA. The alpha-MG-based xanthone NO donor compound disclosed by the invention has the advantages of relatively strong in-vivo and in-vitro antibacterial activity, difficulty in drug resistance and the like, can effectively remove biofilm virulence factors of gram-positive bacteria such as MRSA, is an active compound for treating staphylococcus aureus which is a main pathogen of cow mastitis, and has a broad application prospect. The synthesis method is simple, short in route, clean in process and easy for large-scale industrialization.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and relates to an α-MG xanthone NO donor compound, a synthesis method and an application thereof, and specifically relates to a novel α-mangostin xanthone NO donor compound, a synthesis method and an application thereof to resist MRSA and other bacteria. Background Art

[0002] Cow mastitis seriously threatens the healthy development of the dairy industry. Studies have shown that among the 220 million dairy cows in the world, about one-third suffer from various types of mastitis, which is the main cause of economic losses in the dairy farming industry. The economic losses caused by cow mastitis worldwide are as high as 35 billion US dollars each year. Although the causes of the disease are complex, it is mainly caused by bacteria such as Staphylococcus aureus. The current incidence rate is on the rise. The long-term and large-scale use of antibiotics in prevention and control will not only lead to the production of drug-resistant strains, but also cause antibiotic residues in dairy products, thereby endangering human health. In addition, biofilm formation increases the drug resistance of pathogenic bacteria. According to statistics, two-thirds of microbial infections currently originate from biofilm formation, which enhances their pathogenicity and drug resistance. Therefore, in response to the current economic losses caused by cow mastitis and the human health problems caused by it, it is urgent to develop new Gram-positive drugs such as Staphylococcus aureus and enhancers that reduce drug resistance caused by biofilm.

[0003] Natural antimicrobial agents refer to a class of compounds with antimicrobial activity extracted from certain plants or animals. For example, natural antimicrobial peptides from animals, chitosan derivatives extracted from shrimp shells and crab shells, and extracts from Andrographis paniculata, Houttuynia cordata, mangosteen peels, garlic and mint. Natural biological antimicrobial agents have the advantages of good biocompatibility, low toxicity, and a wide range of sources. However, the shortcomings of natural biological antimicrobial agents, such as weak antimicrobial activity, short antimicrobial duration, and poor drugability, limit its application prospects. This patent uses the main active ingredient of the Chinese medicine mangosteen, the xanthone α-mangostin (α-MG), as the skeleton structure for research. α-Mangostin is recognized as a xanthone compound with strong pharmacological activity, including antibacterial, anti-inflammatory, antifungal, antioxidant, and antiallergic activities. At the same time, the xanthone core structure of α-mangostin has the property of naturally targeting cell membranes, which has inspired many scientists to use it as a lead compound for structural modification. Based on the antibacterial effect of gas molecule NO and its activity in inhibiting biofilm formation and eliminating existing biofilms (Identification of a novel nitric oxide donor peptide with dual biofilm removal and antibacterial activity for intervention in device-related infections, Medicinal Chemistry, 2020, 63:9127-9135), and the ability of the aromatic nitro group to control the release of NO, a stronger anti-Gram-positive bacterial effect than the lead compound was obtained and the drug resistance caused by biofilm was eliminated, including methicillin-resistant Staphylococcus aureus (MRSA) exhibiting strong antibacterial activity (design and synthesis of amphiphilic xanthone membrane-targeted antibacterial agents with improved membrane selectivity, Medicinal Chemistry, 2013, 56, 2359-2373). It has been reported that simple sugars such as mannose are introduced into the 6-hydroxyl group of α-mangostin, which solves the poor water solubility of the lead compound and improves the targeted antibacterial activity of α-mangostin sugar compounds (xanthone esters and sugar derivatives and their preparation and antibacterial use, CN115260146B). However, there has been no report on xanthone NO donor derivatives for drug-resistant colonies and bacterial communities with biofilms causing pathogenic infections and antibiotic resistance. The present invention uses xanthone α-MG as a lead, adopts a symmetrical or asymmetrical method, introduces an aromatic NO donor skeleton, rationally restructures the α-MG chemical space, has better antibacterial activity, and uses α-MG as a drug target carrier to cooperate with NO to clear biofilms and avoid drug resistance. A series of xanthone NO donor derivatives with good inhibitory activity on Gram-positive bacteria and reduction of Gram-positive bacterial biofilms are obtained. It can better solve the problem of MRSA-induced mastitis or biofilm infection in dairy cows. Summary of the invention

[0004] The purpose of the present invention is to overcome the shortcomings of the above-mentioned prior art, and to provide a new antibacterial NO donor compound based on α-MG xanthone, a synthesis method and application. Specifically, α-MG is used as a lead compound, and the symmetry and asymmetry of the C3 position or / and C6 position on the xanthone α-MG structure are modified, and NO donors are introduced to synthesize a series of NO donor compounds, and antibacterial activity and anti-biofilm activity screening are performed to obtain a new α-MG xanthone NO donor compound with good antibacterial activity and inhibition and removal of biofilm. The NO donor compound described in the present invention is uniquely designed, which is conducive to improving the drugability of the molecule and the targeted controlled release of the NO donor; the conditions in the synthesis process are mild, the reaction reagents are easily available, and the reaction route is short; the α-MG xanthone NO donor compound has the ability to target cell wall biosynthesis and damage cell membrane structure, resulting in enhanced permeability, outflow of contents, blocking biofilm formation, and removing mature biofilms, and can be used as a new broad-spectrum antibacterial drug and biofilm resistance enhancement agent.

[0005] To achieve the above purpose, the technical solution adopted by the present invention is:

[0006] A NO donor compound based on α-MG xanthone, wherein the general structural formula of the NO donor compound based on α-MG xanthone is:

[0007]

[0008] Wherein, R1 and R2 are independent H or NO donors, and at least one of R1 and R2 is a NO donor;

[0009] The NO donor is WR3, W is X, O or CO, and R3 is n = 0, 1 or 2;

[0010] When R4 is NO2, R5, R6 or R7 are each independently H, CH3, CF3 or X; when R5 is NO2, R4, R6 or R7 are each independently H, CH3, CF3 or X; when R6 is NO2, R4, R5 or R7 are each independently H, CH3, CF3 or X; when R7 is NO2, R4, R5 or R6 are each independently H, CH3, CF3 or X; X is F, Cl, Br or I;

[0011] Specifically, the structural formula of the α-MG xanthone NO donor compound is:

[0012]

[0013] Among them, R1 and R2 are independent NO donors respectively.

[0014] Furthermore, R1 and R2 are each independently any of the following structural formulas:

[0015]

[0016] Furthermore, R1 and R2 are each independently any of the following structural formulas:

[0017]

[0018] Furthermore, the structural formula of the α-MG xanthone NO donor compound is:

[0019]

[0020] Wherein, R1 is any of the following structural formulas:

[0021]

[0022] Further, R1 is any of the following structural formulas:

[0023]

[0024] A method for synthesizing the above-mentioned α-MG-based xanthone NO donor compound, the synthesis method is to use α-MG as a starting material, perform symmetric and asymmetric modifications on the C3 position and / or C6 position of the xanthone α-MG structure, introduce an NO donor, and synthesize the α-MG-based xanthone NO donor compound; the specific chemical reaction equation is:

[0025]

[0026] Furthermore, the synthesis method is to take α-MG and an acyl halide derivative, a halogenated hydrocarbon or an alcohol derivative, and carry out a substitution reaction under alkaline conditions to obtain the α-MG-based xanthone NO donor compound.

[0027] Furthermore, the synthesis method comprises the following steps:

[0028] S1, mixing α-MG, anhydrous solvent and anhydrous alkali reagent to obtain solution A;

[0029] S2, taking one of an anhydrous solvent containing an acyl halide derivative, an anhydrous solvent containing a halogenated hydrocarbon, and an anhydrous solvent containing an alcohol derivative, slowly dripping it into solution A for preliminary reaction, then heating to reflux for reaction, and after the reaction is completed, concentrating to obtain a semi-solid residue;

[0030] S3, taking the semi-solid residue and recrystallizing it with an organic solvent to obtain the α-MG xanthone-based NO donor compound.

[0031] Further, the acyl halide derivative is R3COX, the halogenated hydrocarbon is R3X, and the alcohol derivative is R3OH;

[0032] Among them, R3 is n = 0, 1 or 2;

[0033] When R4 is NO2, R5, R6 or R7 are each independently H, CH3, CF3 or X;

[0034] When R5 is NO2, R4, R6 or R7 are each independently H, CH3, CF3 or X;

[0035] When R6 is NO2, R4, R5 or R7 are each independently H, CH3, CF3 or X;

[0036] When R7 is NO2, R4, R5 or R6 are each independently H, CH3, CF3 or X;

[0037] X is F, Cl, Br or I.

[0038] Further, the anhydrous alkaline reagent is potassium carbonate, triethylamine, hydroxylamine, sodium hydroxide, potassium hydroxide, cesium carbonate, N,N-diisopropylethylamine, ammonia water or calcium hydroxide;

[0039] The anhydrous solvent is tetrahydrofuran, 1,4-dioxane, acetone, NN-dimethylformamide or acetonitrile;

[0040] The pre-reaction temperature is room temperature and the time is 30 to 60 minutes;

[0041] The reflux temperature is 45 to 65°C, preferably 50°C, and the reflux time is 1 to 5 hours;

[0042] The organic solvent used for recrystallization is at least one of toluene, xylene, benzene, n-hexane and cyclohexane, preferably toluene and n-hexane.

[0043] A use of the above-mentioned α-MG xanthone-based NO donor compound in the preparation of a sensitizer against Gram-positive bacteria or as a clinical drug-resistant strain MRSA.

[0044] Furthermore, the α-MG xanthone-based NO donor compound is used to inhibit or eliminate Gram-positive bacteria and / or Gram-negative biofilms;

[0045] Furthermore, the α-MG xanthone-based NO donor compound is used to reduce or even replace antibiotics in the breeding industry, including pastures; or as a drug for treating diseases such as cow mastitis; or as a molecular probe for revealing the antibacterial or drug resistance-reducing mechanism of xanthone compounds; or for inhibiting or removing Gram-positive bacterial biofilms in medical devices.

[0046] The beneficial effects of the α-MG xanthone-based NO donor compound, synthesis method and application of the present invention are:

[0047] The α-MG xanthone NO donor compound of the present invention has the advantages of strong antibacterial activity in vivo and in vitro, low drug resistance, etc., and can effectively remove the biofilm virulence factors of Gram-positive bacteria such as MRSA, and is an active compound for treating Staphylococcus aureus, the main pathogen of bovine mastitis;

[0048] The α-MG xanthone-based NO donor compound disclosed in the present invention focuses on its drug-like properties and the reduction of drug resistance caused by the formation of biofilms by bacterial communities, which is achieved by strategies such as symmetrically or asymmetrically introducing aromatic NO donor fragments;

[0049] The α-MG xanthone-based NO donor compound disclosed by the present invention uses an α-MG precursor as a starting material and is obtained through a combination of different chemical reactions. The overall synthesis conditions are mild, the reaction reagents are readily available, the reaction route is short, the selectivity is strong, the yield is high, the reaction process is green and clean, it is environmentally friendly, and it is easy to mass produce.

[0050] The mechanism of action of the α-MG xanthone NO donor compound involved in the present invention has a strong ability to depolarize cell membranes, increase the permeability of cell membranes, and destroy the integrity of cell membranes, resulting in enhanced permeability and outflow of contents, thereby inhibiting the activity of Gram-positive bacteria, and at the same time controlling the release of NO gas to inhibit the formation of Gram-positive bacterial biofilms and eliminate mature biofilms. Therefore, on the one hand, it is used to fight Gram-positive bacteria including MRSA or as a sensitizer for clinical drug-resistant strains MRSA, and by inhibiting and removing Gram-positive bacteria or negative biofilms, it has a synergistic effect to treat diseases such as mastitis caused by Staphylococcus aureus in dairy cows, thereby reducing antibiotics in the breeding industry and even replacing antibiotics to increase new inputs; on the other hand, it can be used as a molecular probe to reveal the antibacterial or drug resistance reduction mechanism of xanthone compounds, and to inhibit and remove Gram-positive bacterial biofilms such as medical devices.

[0051] All α-MG xanthone-based NO donor compounds disclosed in the present invention have good activity in inhibiting Gram-positive bacteria including MRSA or removing the biofilm formation of Gram-positive bacteria;

[0052] The antibacterial mechanism of all α-MG xanthone NO donor compounds involved in the present invention is unique, which is to inhibit the synthesis of bacterial cell walls and destroy the cell membrane structure while controlling the release of NO, inhibit the formation of Gram-positive bacterial biofilm and remove mature biofilm, thereby achieving antibacterial and reducing drug resistance caused by biofilm;

[0053] All α-MG xanthone-based NO donor compounds involved in the present invention can be used for preventing and controlling cow mastitis, and reducing or delaying the drug resistance of Staphylococcus aureus to veterinary clinical antibiotics. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] Figure 1 It is a transmission electron microscopy image of the removal of Staphylococcus aureus biofilm based on α-MG xanthone NO donor compound 3 in Example 6 of the present invention. DETAILED DESCRIPTION

[0055] The present invention is described in detail below with reference to examples. The following examples are implemented on the premise of the technical solution of the present invention, using α-MG as a precursor, acylating or etherifying it, and preparing α-mangostin xanthone-based NO donor compounds with different substitution positions, and provide detailed implementation methods and specific operation processes, which will help those skilled in the art to further understand the present invention, but do not limit the invention in any form. It should be pointed out that for those skilled in the art, without departing from the concept of the present invention, several adjustments and improvements can be made, which all belong to the protection scope of the present invention.

[0056] Example 1 Synthesis of NO Donor Compounds Based on Acylation of C3 or C6 Positions α-MG Xanthone

[0057] 1. Synthesis of α-MG-based xanthone NO donor compound 1

[0058] S1. Preparation of solution A and solution B

[0059] In a dry flat-bottom flask, α-MG (1 mmol), 30 mL of acetone and 3 mL of triethylamine (pH 10) were added to form solution A.

[0060] Solution B was prepared by adding 3,5-dinitrobenzoyl chloride (1 mmol) and mixing with 20 mL of tetrahydrofuran (THF) in a dry separatory funnel.

[0061] S2. Substitution reaction

[0062] Under nitrogen protection, solution B was gradually dripped into solution A, and the mixture was stirred at room temperature for a pre-reaction of 30 minutes. The resulting mixture was refluxed at 50°C while continuously stirred. After the reaction was completed using a TLC monitor, the reflux was stopped, and the mixture was evaporated and concentrated under vacuum to obtain a semi-solid residue. The specific chemical reaction equation is as follows:

[0063]

[0064] Among them, the structural formula of R1 is

[0065] S3, post-processing

[0066] Saturated sodium bicarbonate solution was added to the semi-solid residue, gently stirred for 10 minutes, filtered, washed with distilled water, and the solid product was collected. The obtained solid product was added to toluene for recrystallization to obtain 63.46 mg of light yellow solid 1, which is the α-MG xanthone-based NO donor compound 1, with a yield of 41%. The NMR image of the α-MG xanthone-based NO donor compound 1 is: 1 HNMR(400MHz, CDCl3)δ=13.75(s,1H),9.71(s,3H),7.20(s,1H),6.72(s,1H),6.89(s,1H),5.27-5.12( m,2H),4.15-4.04(m,3H),3.80(s,3H),3.57-3.55(d,2H),1.84(s,3H),1.70(s,3H),1.58-1.56(d,6H). 13 C NMR(100MHz, CDCl3)δ=182.50,161.35,160.71,156.01,155.58,153.83,153.70,149.00,132.88,132.53,129.94, 127.27,125.88,121.46,115.84,114.21,109.36,105.12,99.88,62.65,60.62,26.74,25.65,22.46,17.99,14.30.

[0067] 2. Synthesis of α-MG-based xanthone NO donor compounds 2 to 14

[0068] According to the method in step 1 of synthesizing α-MG xanthone-based NO donor compound 1, α-MG xanthone-based NO donor compounds 2 to 14 are synthesized. The process steps, parameters and raw materials are basically the same. The only difference is the type of acyl chloride derivatives. The acyl chloride derivative in the process of synthesizing α-MG xanthone-based NO donor compound 1 is 3,5-dinitrobenzoyl chloride, while the acyl chloride derivatives in the process of synthesizing α-MG xanthone-based NO donor compounds 2 to 14 are 4-nitrobenzyl bromide, 2-chloro-4-nitrobenzoyl chloride, 4-nitrobenzoyl chloride, 4-methyl-3-nitrobenzoyl chloride, 3-nitrobenzoyl chloride, 2- Trifluoromethyl-4-nitrobenzoyl chloride, 2-methyl-4-nitrobenzoyl chloride, 4-chloro-3-nitrobenzoyl chloride, 2-nitrobenzoyl chloride, 2-chlorobenzoyl chloride, 2,4-dichlorobenzoyl chloride, 2-chloro-4-dimethylaminobenzoyl chloride, benzoyl chloride, after obtaining the corresponding crude product (i.e., semi-solid residue), all adopt the same post-treatment method, i.e., recrystallization from toluene, to obtain light yellow solids 2 to 14 (i.e., α-MG xanthone-based NO donor compounds 2 to 14), with yields of 46%, 46%, 46%, 43%, 57%, 51%, 47%, 41%, 38%, 61%, 63%, 55%, 70% respectively. It can be seen that the yield of the method for synthesizing α-MG xanthone-based NO donor compounds of the present invention is 30 to 70%.

[0069] The chemical reaction equation for synthesizing α-MG-based xanthone NO donor compounds 2 to 14 is:

[0070]

[0071] Among them, the structural formulas corresponding to R1 in α-MG xanthone NO donor compounds 2 to 14 are:

[0072]

[0074] The yield of α-MG xanthone NO donor compound 2 is 46%, and the morphology is light yellow solid. The NMR image is: 1H NMR (400MHz, CDCl3) δ6.68(s,1H),6.64(s,1H),6.14-6.07(m,2H),5.47(t,J=12.0Hz,2H),5.37(t,12.0Hz,2H),5.24(t,J=8.0,1H),5.13 (t,J=8.0,1H),4.65-4.61(m,4H),4.22-3.98(m,2H),3.52(s,3H),3.33-3.22(m,2H),2.47(s,3H),1.84(s,3H),1.76(s,3H),1.68(s,6H). 13 C NMR (100MHz, CDCl3) δ175.23,168.32,161.23,157.11,155.28,155.20,147.20,146.16,139.77,133.54,132.17,130.11,127 .23,125.12,121.12,119.21,118.15,115.21,108.11,99.23,97.05,69.36,26.04,25.85,25.71,23.17,20.78,18.31,17.55.

[0075] The yield of α-MG xanthone NO donor compound 3 is 46%, and the morphology is light yellow solid. The NMR image is as follows: 1 H NMR (400MHz, CDCl3) δ6.70 (s, 1H), 6.65 (s, 1H), 6.32-6.21 (m, 2H), 5.63 (t, J= 16.0Hz,2H),5.12(t,J=16.0Hz,2H),5.06(t,J=4.0Hz,1H),5.02(t,J=4.0Hz, 1H),4.76-4.70(m,4H),4.20(d,2H),3.82(d,J=4.0Hz,3H),3.38(d,J=44.0Hz ,2H),3.08(m,1H),1.82(s,3H),1.74(s,3H),1.65(s,6H),1.43(s,3H),1.41(s 3H). 13C NMR (100MHz, CDCl3) δ177.70,175.40,161.52,155.96,155.72,154.25,150.38,144.05,138.13,132.20,131.88,130.90,123.80,122.36,1 18.24,118.11,114.28,112.68,98.95,98.87,96.81,96.72,69.68,6 0.76,34.30,34.12,25.98,23.20,19.16,18.47,18.16,17.99,17.96.

[0076] The yield of α-MG xanthone NO donor compound 4 is 46%, and the morphology is light yellow solid. The NMR image is as follows: 1 H NMR (400MHz, CDCl3) δ13.70(s,1H),6.74(s,1H),6.60(s,1H),6.17 -5.98(m,1H),5.77(d,J=12.0Hz,1H),5.56(d,J=12.0Hz,1H),5.20(t,J=8.0Hz,1H),5.18(t,J=8.0Hz,1H),4.60(d,J= 4.0Hz,2H),4.13-4.09(m,2H),3.90(s,3H),3.32(d,J=8.0Hz,2H),2.34(s,3H),1.85(s,3H),1.78(s,3H),1.68(s,6H). 13 C NMR (100MHz, CDCl3) δ182.88,168.66,161.24,157.66,157.64,154.70,144.52,136.72,132.26,132.10,123.16,122. 86,121.90,121.50,116.10,112.00,107.22,100.26,100.08,99.56,98.43,69.72,60.92,26.40,26.05,25.85,22.45.

[0077] The yield of α-MG xanthone NO donor compound 5 is 43%, and the morphology is light yellow solid. The NMR image is as follows: 1H NMR(400MHz, CDCl3)δ13.70(s,1H),6.74(s,1H),6.60(s,1H),6.24-6.16(m,1H) ,5.52(d,J=12.0Hz,1H),5.38(d,J=12.0Hz,1H),5.24(t,J=8.0Hz,1H),5.12(t,J =8.0Hz,1H),4.70(d,J=4.0Hz,2H),4.16-4.10(m,2H),3.62(s,3H),3.20(d,J=8. 0Hz,2H),2.86-2.82(m,1H),1.86(s,3H),1.74(s,3H),1.68(s,6H),1.36(s,3H). 13 C NMR (100MHz, CDCl3) δ183.68,172.68,162.80,156.32,154.20,152.20,148.20,136.20,132.22,131.90,131.82,123.01,1 22.90,121.60,121.46,118.56,116.22,112.18,106.94,100.92,70.52,60.92,34.32,25.90,25.70,22.14,18.19,17.86.

[0078] The yield of α-MG xanthone NO donor compound 6 was 57%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR (400MHz, CDCl3) δ7.20(s,1H),6.52(s,2H),6.21(s,1H),5.28 -5.10(m,2H),4.22(s,2H),3.76(s,3H),3.28(d,2H),2.50(s,3H),1.80(s,3H),1.76(s,3H),1.68(s,3H). 13 CNMR(100MHz, CDCl3)δ176.22,171.11,159.84,159.82,154.80,153.22,148.14,142.80,137.23,131.59,123.65,12 1.24,118.63,114.24,109.15,101.48,100.71,61.90,61.87,25.82,22.88,21.52,18.32,17.92.TOF-MS,m / z:[M+Na + ],calcd for C 26 H 28 NaO7 +,475.1727,found:475.1738.

[0079] The yield of α-MG xanthone NO donor compound 7 was 51%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR (400MHz, CDCl3) δ7.16(s,1H),6.52(s,1H),6.42(s,2H),5.22(t,J=4.0Hz,1H),5.14(t,J=4.0Hz,1H),4.12(d,J=8.0Hz,2H) ,3.80(s,3H),3.60(d,J=8.0Hz,2H),3.10(m,1H),1.80(s,3H),1.76(s,3H),1.70(s,3H),1.66(s,3H),1.45(s,3H),1.43(s,3H). 13 CNMR (100MHz, CDCl3) δ176.76,171.28,158.82,154.65,153.86,147.36,141.76,136.16,132.96,122.86,120.36 ,117.56,113.32,108.32,100.42,100.44,99.54,67.16,60.86,33.55,25.46,24.83,21.76,17.80,17.30,17.06.

[0080] The yield of α-MG xanthone NO donor compound 8 was 47%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR (400MHz, CDCl3) δ13.62(s,1H),6.78(s,1H),6.59(s,1H),5.26-5.14(m,2H),4.05(d,J=4.0 Hz,2H),3.78(s,3H),3.33(d,J=4.0Hz,2H),2.34(s,3H),1.83(s,3H),1.78(s,3H),1.68(s,6H). 13 C NMR (100MHz, CDCl3) δ182.62,168.93,161.14,156.05,155.30,154.47,153.76,143.01,137.40,132.36,132.22,1 23.09,121.66,116.07,112.27,107.02,101.80,100.30,62.09,26.73,25.93,25.86,22.44,21.22,18.32,17.96.

[0081] The yield of α-MG xanthone NO donor compound 9 was 41%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR (400MHz, CDCl3) δ13.62(s,1H),6.73(s,1H),6.61(s,1H),6.55(s,1H),5.22(t,J=4.0Hz,1H),5.15(d,J=4.0Hz,1H),4.03 (d,J=4.0Hz,1H),3.76(s,3H),3.30(d,J=4.0Hz,2H),2.88-2.86(m,1H),1.84(s,3H),1.74(s,3H),1.68(s,6H),1.36(s,3H). 13 C NMR (100MHz, CDCl3) δ182.42,175.11,160.95,155.80,155.74,155.14,152.10,148.20,142.82,137.24,132. 22,122.30,121.55,116.66,112.22,106.56,101.20,99.80,61.80,34.20,26.26,25.22,23.22,18.82,17.88.

[0082] The yield of α-MG xanthone NO donor compound 10 was 38%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR (400MHz, CDCl3) δ13.70(s,1H),6.66(s,1H),6.24(s,1H),5.50(t,1H),5.44(t,J=8.0Hz,1H),4.24( d,J=4.0Hz,2H),3.96(s,3H),3.62(d,J=4.0Hz,2H),2.62(s,3H),2.04(s,6H),1.86(s,3H),1.80(s,3H). 13 C NMR (100MHz, CDCl3) δ183.06,168.26,162.16,160.20,154.60,148.80,146.52,139.14,135.02,132.20,1 23.10,121.64,117.08,110.49,109.28,103.84,93.35,61.82,26.24,25.96,21.55,21.06,18.31,18.02.

[0083] The yield of α-MG xanthone NO donor compound 11 was 61%, and the morphology was light yellow solid. The NMR image was as follows: 1H NMR (400MHz, CDCl3) δ13.62(s,1H),6.80(s,1H),6.58(s,1H),5.30(t,J=8.0Hz,1H),5.20(t,J=8.0Hz,1H),4.06(d,J=4.0Hz,2H),3 .80(s,3H),3.50(d,J=4.0Hz,2H),2.90-2.84(m,1H),1.82(s,3H),1.72(s,3H),1.70(s,3H),1.62(s,3H),1.48(s,3H),1.32(s,3H). 13 C NMR (100MHz, CDCl3) δ182.88,175.22,161.32,156.16,155.28,154.22,152.32,141.38,136.40,132.38,12 2.86,121.80,121.76,116.74,112.22,106.10,101.78,100.26,62.23,34.60,26.74,25.95,25.87,18.03.

[0084] The yield of α-MG xanthone NO donor compound 12 was 63%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR(400MHz, CDCl3)δ=13.52(s,1H),8.32(d,1H),7.60(s,1H),7.22(d,1H),7.18(s,1H),6.70(s,1H),5.26(d,J =12.5Hz,2H),4.36–4.08(m,3H),3.78(s,3H),3.74(d,2H),2.06(s,1H),1.86(s,6H),1.72(s,3H),1.70(s,3H). 13 C NMR(100MHz, CDCl3)δ=181.18,162.06,160.62,154.82,153.36,148.30,142.42,139.82,138.20,136.09,134.64,132.16,131.76, 127.38,126.24,122.18,121.52,117.20,110.54,109.22,103.70,93.20,62.22,60.42,26.42,25.78,21.42,18.16,17.84,14.14.

[0085] The yield of α-MG xanthone NO donor compound 13 was 55%, and the morphology was light yellow solid. The NMR image was as follows:1 H NMR (400MHz, CDCl3) δ13.78(s,1H),6.66(s,1H),6.22(s,1H),5.32-5.20(m,2H),4.12(d,J=4.0Hz,2H) ,4.08(t,J=6.4Hz,2H),3.80(s,3H),3.72(t,J=6.4Hz,2H),3.44(d,J=4.0Hz,2H),1.94-1.40(m,20H). 13 C NMR (100MHz, CDCl3) δ182.16,161.66,158.74,156.66,154.05,144.12,137.36,135.22,123.24,122 .76,111.76,108.86,103.77,98.90,93.16,62.97,61.00,32.61,28.96,27.21,26.86,25.54,18.56.

[0086] The yield of α-MG xanthone NO donor compound 14 was 70%, and the morphology was light yellow solid. The NMR image was as follows: 1 H NMR (400MHz, CDCl3) δ13.50(s,1H),6.86(s,1H),6.32(s,1H),5.30-5.20(m,2H),4.20(d,J=6.8Hz,2H),4.18(d,J =8.0Hz,2H),4.02(d,J=8.0Hz,2H),3.68(s,3H),3.62(t,J=6.8Hz,4H),3.32(d,J=6.8Hz,2H),1.78-1.52(m,30H). 13 C NMR (100MHz, CDCl3) δ182.82,162.30,160.86,158.52,155.78,155.22,144.24,138.18,134.76,124.52,123.36,122.38,111.96, 103.92,98.86,89.42,68.79,64.11,63.28,62.66,61.22,60.84,32.76,29.20,28.00,26.04,25.68,22.28,18.32,18.26,17.96.

[0087] Example 2 Minimum Inhibitory Concentration Test of α-MG and α-MG-based Xanthone NO Donor Compounds

[0088] 1) Experimental principle

[0089] This experiment is based on the growth of bacteria in MH (Mueller-Hinton Broth) culture medium in a 96-well plate. After a period of time, white precipitate will be produced due to the large amount, which makes it easy to distinguish whether there is bacterial growth. The antibacterial agent is diluted in multiples and added to the bacterial suspension. After culturing for a period of time, the last well without precipitation is the minimum inhibitory concentration (MIC).

[0090] 2) Experimental methods

[0091] According to the NCCLS experimental operation requirements, the strains used include Staphylococcus aureus ATCC 29213, Staphylococcus aureus ATCC6538, MRSA2 and Escherichia coli ATCC 25922. The strains were revived and cultured to the logarithmic growth phase for use. Use a sterile pipette to take the bacterial solution from the bacterial culture and drip it into the 96-well plate. The amount of bacterial solution dripped depends on the experimental needs and the concentration of the bacterial solution. α-MG and α-MG-based xanthone NO donor compounds 1 to 14 were dissolved in dimethyl sulfoxide to 2 mg / mL, and then 25.6 μL was added to the first column, and then MH (B) medium was added to 200 μL, 2-fold dilution, and diluted to the 10th column in turn. The control drug vancomycin (i.e., positive control) was treated in the same dissolution, addition, and 2-fold dilution method, and cultured at 37°C for 18 to 24 hours and the results were observed. The minimum inhibitory concentration of the control drug should be within the reference range, and the experimental results of the same batch are considered reliable.

[0092] 3) Experimental results

[0093] The antibacterial activity of α-MG and its derivatives against standard strains and multidrug-resistant strains of Escherichia coli and Staphylococcus aureus was tested by microdilution method, and the results are shown in Table 1.

[0094] Table 1 Minimum inhibitory concentrations of α-MG and α-MG-based xanthone NO donor compounds 1 to 14 against bacteria

[0095]

[0096]

[0097] Note: 1 to 14 in Table 1 represent NO donor compounds 1 to 14 based on α-MG xanthone.

[0098] 4) Experimental conclusion

[0099] From the data in Table 1, it can be seen that the lead compound α-MG and α-MG-based xanthone NO donor compounds 1 to 14 have antibacterial activity against Gram-positive bacteria and clinical isolate MRSA2, among which α-MG-based xanthone NO donor compounds 1, 3 to 10 have stronger antibacterial activity. Example 3 NO release based on α-MG xanthone NO donor compounds

[0100] 1) Experimental principle

[0101] The basic principle of NO detection is to indirectly determine the NO content by measuring the NO concentration in the body or the substances produced by chemical reactions. NO is a highly reactive gas molecule that is difficult to detect directly, so it is usually detected by measuring its metabolites or using specific chemical reactions. Chemical colorimetry is used to detect NO by measuring its metabolites in the body. For example, NO in human serum can generate nitrite (NO2 - ) and nitrate (NO3 - ). A purple-red complex is generated through chemical reaction, and the change rate of absorbance is measured to indirectly determine the concentration of NO.

[0102] 2) Experimental methods

[0103] The NO level produced by α-MG xanthone-based NO donor compounds 1 to 14 was determined by colorimetry according to the manufacturer's instructions using a nitrite colorimetric assay kit (Beyotime, China). Briefly, Staphylococcus aureus ATCC 6538 or MASA2 were cultured at a density of approximately 1 × 10 8 CFU / mL, treated with α-MG xanthone NO donor compounds 1 to 14, cultured at 37°C for 20 min, and then lysed by ice bath ultrasonication at a ratio of 1:5 for 5 min. Subsequently, the bacterial lysate was harvested, mixed with Griess reagent at 37°C for 10 min, and then measured at 540 nm with a microplate reader. Bacteria treated with 0.4% DMSO in the culture medium were used as negative controls for the background level of nitrite production (i.e., solvent controls), and different concentrations of sodium nitrite were prepared as positive controls. Each experiment was repeated at least three times. The experimental results are shown in Table 2.

[0104] Table 2 Amount of NO produced by α-MG-based xanthone NO donor compounds 1-14 in Staphylococcus aureus ATCC6538 and MASA2

[0105]

[0106] Note: 1 to 14 in Table 2 represent α-MG xanthone-based NO donor compounds 1 to 14 respectively.

[0107] 3) Experimental results

[0108] From the data in Table 2, it can be seen that the amount of NO produced by α-MG xanthone-based NO donor compounds 1, 3-10 in Staphylococcus aureus ATCC 6538 and MASA2 is higher than that of the solvent control (no compound added) to varying degrees, among which the release amount of α-MG xanthone-based NO donor compound 3 is the highest.

[0109] Example 4 Inhibition of Biofilm by α-MG and α-MG-based Xanthones NO Donor Compounds

[0110] 1) Experimental principle

[0111] Biomembrane crystal violet quantification is a commonly used biological experimental method used to determine the permeability of biological membranes such as cell membranes, mitochondria, and endoplasmic reticulum. This method is based on crystal violet dye, which can bind to biological membranes and diffuse into the interior of cells through the cell membrane. After staining, the permeability of the biological membrane can be indirectly reflected by measuring the degree of absorption of the dye in the cell. The absorbance (OD value) of the stained cells can be measured by a spectrophotometer. The absorbance is proportional to the concentration of crystal violet in the cell, and the concentration of crystal violet in the cell is related to the permeability of the biological membrane. Therefore, by measuring the absorbance, the permeability of the biological membrane can be indirectly calculated.

[0112] 2) Experimental methods

[0113] LB medium (3 mL) was inoculated with an overnight bacterial culture of Staphylococcus aureus ATCC 6538 and cultured at 37°C until the culture reached the mid-logarithmic phase, and the bacterial suspension was diluted 1:100 and set aside. The biofilm inhibition test was performed by adding the bacterial suspension (100 μL) to a 96-well plate containing 100 μL of different concentrations (0 to 128 μg / mL) of α-MG and α-MG-based xanthone NO donor compounds 1 to 14.

[0114] The 96-well plate was placed in an incubator at 37°C for 12 to 16 hours. After completion, the bacteria were removed from the 96-well plate, washed three times with phosphate-buffered saline (1×PBS), then 0.1% crystal violet was added, placed at room temperature for 15 to 20 minutes, and then the crystal violet was removed and washed three times with PBS buffer. Finally, ethanol was added to the 96-well plate to dissolve the crystal violet adsorbed on the plate, and the 96-well plate was placed on a zymometer to read the absorbance value at 595 nm. Compared with the untreated control (no drug group), the minimum polymer concentration that resulted in a 90% reduction in biofilm will be reported as the minimum inhibitory biofilm concentration of biomass. Each experiment was repeated at least three times. Biofilm inhibition rate (%) = (OD control-OD sample) / OD control × 100%.

[0115] The removal experiment was first incubated in a 96-well plate at 37°C for 12 h to obtain a mature biofilm. After removing the planktonic cells, the biofilm was treated with different concentrations of α-MG xanthone-based NO donor compounds 1 to 14 at 37°C for 12 h. The other steps were similar to the inhibition experiment. As described above, the samples were analyzed for biofilm biomass, and the biofilm concentration and biofilm biomass removal rate were calculated. Biofilm eradication rate (%) = (OD control-OD sample) / OD control × 100%.

[0116] 3) Experimental results

[0117] The inhibitory effects of α-MG xanthone-based NO donor compounds 1 to 14 on the biofilm of Staphylococcus aureus ATCC 6538 were measured by the crystal violet method. The results are shown in Table 3.

[0118] Table 3 Minimum inhibitory concentrations of α-MG-based xanthone NO donor compounds 1 to 14 on bacterial biofilm

[0119]

[0120] Note: 1 to 14 in Table 3 represent α-MG xanthone-based NO donor compounds 1 to 14 respectively.

[0121] 4) Experimental conclusion

[0122] From the data in Table 3, it can be seen that among the inhibitory effects of α-MG xanthone-based NO donor compounds 1, 3 to 10 on the biofilm of the listed bacterial communities, α-MG xanthone-based NO donor compound 3 has the best effect.

[0123] Example 5 Detection of hemolytic activity of α-MG and α-MG-based xanthone NO donor compounds

[0124] 1) Experimental principle

[0125] The hemolysis test is a commonly used experimental method to detect the activity of blood antibodies. Its principle is based on the phenomenon that red blood cells are lysed when exposed to specific antibodies.

[0126] 2) Experimental methods

[0127] 4% rabbit erythrocytes (100 μL) were added to a 96-well plate containing 100 μL of different concentrations (0 to 128 μg / ml) of α-MG and α-MG-based xanthone NO donor compounds 1 to 14, with 1% Triton-X 100 as a positive control and PBS buffer as a negative control. The 96-well plate was incubated in a thermostat at 37°C and 60 rpm for 1 h. After incubation, the 96-well plate was centrifuged at a centrifugal force of 1000 g for 3 min. The supernatant (100 μL) was taken and the corresponding absorbance of each well was measured at 540 nm. Each experimental data was repeated three times.

[0128] 3) Experimental results

[0129] The experimental results are shown in Table 4. The α-MG-based xanthone NO donor compounds 1 to 14 were not hemolytic at a concentration of 128 μg / mL.

[0130] Table 4 Hemolytic activity of α-MG and α-MG based xanthone NO donor compounds 1 to 14

[0131]

[0132]

[0133] 4) Experimental conclusion

[0134] α-MG-based xanthone NO donor compounds 1, 3-10 have good biofilm removal activity and are not prone to causing erythrocyte hemolysis.

[0135] Example 6 Observation of the effect of α-MG xanthone-based NO donor compounds on bacterial morphology under transmission electron microscopy

[0136] 1) Experimental principle

[0137] The morphology of the sample tissue can be observed based on a transmission electron microscope. Ultrathin sections with a thickness of 10 to 100 nm are prepared by ultrathin sectioning technology. Changes in bacterial morphology after incubation with the α-MG xanthone-based NO donor compound 3 are observed under a transmission electron microscope.

[0138] 2) Experimental methods

[0139] Staphylococcus aureus ATCC6538 revived to the logarithmic growth phase was incubated with different concentrations of α-MG xanthone-based NO donor compound 3 for 2 hours, the culture medium was discarded by centrifugation, washed 2-3 times with PBS buffer, fixed with 2.5% glutaraldehyde, dehydrated, sprayed with gold, observed under transmission electron microscope, and the thickness of cell wall was measured, which was used as the drug group. The group without any drug was used as the blank group.

[0140] 3) Experimental results

[0141] Staphylococcus aureus was incubated with α-MG xanthone NO donor compound 3 and compared with the blank group. The results are as follows: Figure 1 As shown. It can be seen that the biofilm in the blank group (without any drug) is obvious, while the biofilm in the drug-treated group is obviously removed. Figure 1 The left picture is the transmission electron micrograph of the blank group, and the right picture is the transmission electron micrograph of Staphylococcus aureus ATCC6538 incubated with α-MG xanthone-based NO donor compound 3 at a concentration of 32 μg / mL and after a series of treatments (i.e., the drug-treated group).

[0142] 4) Experimental conclusion

[0143] It can be seen from the experimental results that the α-MG xanthone-based NO donor compound 3 can effectively remove the Gram-positive bacterial biofilm after acting on bacteria for a certain period of time.

[0144] Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

Claims

1. A NO donor compound based on α-MG xanthone, characterized in that: The general structural formula of the α-MG xanthone NO donor compound is: Wherein, R1 and R2 are independent H or NO donors, and at least one of R1 and R2 is a NO donor; The NO donor is WR3, W is X, O or CO, and R3 is n = 0, 1 or 2; When R4 is NO2, R5, R6 or R7 are each independently H, CH3, CF3 or X; When R5 is NO2, R4, R6 or R7 are each independently H, CH3, CF3 or X; When R6 is NO2, R4, R5 or R7 are each independently H, CH3, CF3 or X; When R7 is NO2, R4, R5 or R6 are each independently H, CH3, CF3 or X; X is F, Cl, Br or I.

2. The α-MG xanthone-based NO donor compound according to claim 1, characterized in that: R1 and R2 are each independently any of the following structural formulas:

3. The α-MG xanthone-based NO donor compound according to claim 1 or 2, characterized in that: The structural formula of the α-MG xanthone NO donor compound is: Wherein, R1 is any of the following structural formulas:

4. A method for synthesizing an α-MG xanthone NO donor compound according to any one of claims 1 to 3, characterized in that: The synthesis method uses α-MG as a starting material, performs symmetric and asymmetric modification on the C3 position and / or C6 position of the xanthone α-MG structure, introduces an NO donor, and synthesizes the α-MG-based xanthone NO donor compound.

5. The method for synthesizing an α-MG xanthone NO donor compound according to claim 4, characterized in that: The synthesis method is to take α-MG and an acyl halide derivative, a halogenated hydrocarbon or an alcohol derivative, and carry out a substitution reaction under alkaline conditions to obtain the α-MG-based xanthone NO donor compound.

6. The method for synthesizing an α-MG xanthone NO donor compound according to claim 5, characterized in that: The synthesis method comprises the following steps: S1, mixing α-MG, anhydrous solvent and anhydrous alkali reagent to obtain solution A; S2, taking one of an anhydrous solvent containing an acyl halide derivative, an anhydrous solvent containing a halogenated hydrocarbon, and an anhydrous solvent containing an alcohol derivative, slowly dripping it into solution A for preliminary reaction, then heating to reflux for reaction, and after the reaction is completed, concentrating to obtain a semi-solid residue; S3, taking the semi-solid residue and recrystallizing it with an organic solvent to obtain the α-MG xanthone-based NO donor compound.

7. The method for synthesizing an α-MG xanthone NO donor compound according to claim 5 or 6, characterized in that: The acyl halide derivative is R3COX, the halogenated hydrocarbon is R3X, and the alcohol derivative is R3OH; Among them, R3 is n = 0, 1 or 2; When R4 is NO2, R5, R6 or R7 are each independently H, CH3, CF3 or X; When R5 is NO2, R4, R6 or R7 are each independently H, CH3, CF3 or X; When R6 is NO2, R4, R5 or R7 are each independently H, CH3, CF3 or X; When R7 is NO2, R4, R5 or R6 are each independently H, CH3, CF3 or X; X is F, Cl, Br or I.

8. The method for synthesizing an α-MG xanthone NO donor compound according to claim 6, characterized in that: The anhydrous alkaline reagent is potassium carbonate, triethylamine, hydroxylamine, sodium hydroxide, potassium hydroxide, cesium carbonate, N,N-diisopropylethylamine, ammonia water or calcium hydroxide; The anhydrous solvent is tetrahydrofuran, 1,4-dioxane, acetone, NN-dimethylformamide or acetonitrile; The pre-reaction temperature is room temperature and the time is 30 to 60 minutes; The reflux temperature is 45-65°C and the reflux time is 1-5h; The organic solvent used for recrystallization is at least one of toluene, xylene, benzene, n-hexane and cyclohexane.

9. Use of the α-MG xanthone-based NO donor compound according to any one of claims 1 to 3 in the preparation of a sensitizer against Gram-positive bacteria or as a clinical drug-resistant strain MRSA.

10. The use of the α-MG xanthone NO donor compound according to claim 9, characterized in that: The α-MG xanthone-based NO donor compound is used for inhibiting or eliminating Gram-positive bacteria and / or Gram-negative biofilms.

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