Preparation method of 1, 2, 3-triazole compound and application of 1, 2, 3-triazole compound in antifungal field

Abstract

The invention discloses a preparation method of a 1, 2, 3-triazole compound and application of the 1, 2, 3-triazole compound in the antifungal field, and relates to the technical field of antifungal drugs, and the key point of the technical scheme is that the structural general formula of the compound is # imgabs0 #, R group represents multiple dihalogen substituents, and the halogen is F, Cl or Br. The invention also provides application and a preparation method of the compound, the compound has relatively strong inhibitory activity on deep fungi, and compared with currently used antifungal drugs clinically, the compound has the advantages of low toxicity, high efficiency, simple and convenient preparation method and relatively high yield, and can be suitable for preparing efficient antifungal drugs.

Description

Technical Field

[0001] The present invention relates to the technical field of antifungal drugs, and more specifically, to a preparation method of 1,2,3-triazole compounds and their application in the antifungal field. Background Art

[0002] In the past three decades, due to the extensive use of broad-spectrum antibiotics in clinical practice, the increase in the number of patients undergoing cancer radiotherapy, chemotherapy and organ transplantation, the widespread use of corticosteroids and immunosuppressants, and the prevalence of AIDS, the number of patients with low or absent immune function has continued to increase, and deep fungal infections have gradually become an infectious disease that seriously threatens human life and health.

[0003] Although the research and development of antifungal drugs has a long history and has made great achievements, there are still few types of antifungal drugs used in clinical practice. Their disadvantages are that they have large adverse reactions, most of them only have antibacterial effects, and drug-resistant strains continue to appear, and the needs of clinical treatment cannot be met. At present, our urgent task is to develop a new generation of highly effective, low-toxic, and broad-spectrum antifungal drugs.

[0004] Existing antifungal drugs mainly include azoles that act on lanosterol 14a-demethylase, allylamines that act on squalene epoxidase, and lipopeptides that act on cell walls and -1,3-β-glucan synthase. Among them, azole antifungal drugs are the most widely used and most popular drugs in clinical research. However, there has been no report on the antifungal activity of 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(disubstituted benzyl)methylamino]}-2-ol compounds.

[0005] Therefore, the present invention aims to provide a method for preparing a 1,2,3-triazole compound and its application in the antifungal field, so as to solve the above problems. Summary of the invention

[0006] The purpose of the present invention is to provide a preparation method of 1,2,3-triazole compounds and their application in the antifungal field. The 1,2,3-triazole compounds of the present invention have relatively strong inhibitory activity against deep fungi, and compared with the antifungal drugs currently used in clinical practice, have the advantages of low toxicity, high efficiency, simple preparation method and relatively high yield, and can be suitable for preparing highly effective antifungal drugs.

[0007] The above technical purpose of the present invention is achieved by the following technical scheme: 1,2,3-triazole compound, the general structural formula of the compound is:

[0008]

[0009] Wherein, the R group represents various dihalogen substituents, and the halogen is F, Cl or Br.

[0010] The present invention is further configured that: the compound includes a pharmaceutically acceptable salt.

[0011] The present invention is further configured as follows: the pharmaceutically acceptable salt is acetate, oxalate, bisulfate, citrate, hydrochloride, sulfate, hydrobromide or methanesulfonate.

[0012] The present invention also provides a pharmaceutical composition comprising a 1,2,3-triazole compound and a pharmaceutical carrier.

[0013] The present invention also provides a method for preparing a 1,2,3-triazole compound, comprising the following steps:

[0014] S1. Dissolve 2-(2,4-difluorophenyl)-2-[(1H-1,2,4-triazol-1-yl)methyl]oxirane methanesulfonate, ethylamine and triethylamine in anhydrous ethanol, heat and stir to reflux, and continue the reaction for 6 hours to obtain key compound II;

[0015] S2, dissolving the key compound II in anhydrous DMF, adding anhydrous potassium carbonate, potassium iodide, and propyne bromide, stirring at room temperature overnight to obtain compound III;

[0016] S3. Dissolve compound III in DMSO, add various disubstituted benzyl bromides and sodium azide reaction solution, prepare a catalyst by dissolving copper sulfate pentahydrate and sodium ascorbate in water, add the catalyst to the reaction solution, stir at room temperature for 1 hour, and obtain the target compound A1-11.

[0017] The present invention also provides an application of a 1,2,3-triazole compound in the antifungal field.

[0018] The present invention also provides an application of a 1,2,3-triazole compound in the preparation of a therapeutic antifungal drug.

[0019] In summary, the present invention has the following beneficial effects:

[0020] 1. Compared with the antifungal drugs currently used in clinical practice, the compounds of the present invention have the advantages of high efficiency and low toxicity, and have strong inhibitory activity against deep-seated fungi. Therefore, they can be used to prepare highly effective antifungal drugs;

[0021] 2. The preparation method of the compound of the present invention has high yield and is simple, and the prepared compound has relatively good antifungal effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is a synthetic route diagram of the target compound of type A in the embodiments of the present invention. DETAILED DESCRIPTION

[0023] The following is combined with Figure 1 The present invention is described in further detail.

[0024] Example 1: 1,2,3-triazole compounds, the general structural formula of the compound is:

[0025]

[0026] The R group represents a variety of dihalogen substituents, and the halogen is F, Cl or Br.

[0027] The preparation reaction route of 1,2,3-triazole compounds in this embodiment is as follows:

[0028] a) 2-(2,4-difluorophenyl)-2-[(1H-1,2,4-triazol-1-yl)methyl]oxirane methanesulfonate, ethylamine and triethylamine were dissolved in anhydrous ethanol, heated and stirred to reflux, and the reaction was continued for 6 hours to obtain the key compound II;

[0029] b) Dissolve the key compound II in anhydrous DMF, add anhydrous potassium carbonate, potassium iodide, and propyne bromide, and stir at room temperature overnight to obtain compound III;

[0030] c) Compound III was dissolved in DMSO, various disubstituted benzyl bromides and sodium azide reaction solutions were added, copper sulfate pentahydrate and sodium ascorbate were dissolved in water to prepare a catalyst, added to the reaction solution, and stirred at room temperature for 1 h to obtain the target compound A1-11.

[0031] The synthetic route of target compounds of type A is as follows Figure 1 As shown, specifically, the preparation steps of the target compound A1-11 are as follows:

[0032] 1. Preparation of Compound II

[0033] Select a 500ml round-bottom flask, add compound I (21g, 63.05mmol), 250ml of anhydrous ethanol, 10ml of ethylamine, and 20ml of triethylamine in sequence, heat to 100°C in an oil bath, and stir magnetically for 6 hours. TLC monitoring indicates that the reaction is complete. After the reaction solution is cooled, use a rotary evaporator to evaporate the solvent, use a 500ml separatory funnel, extract and wash the reaction solution twice with 200ml of ethyl acetate and 200ml of water, collect the ethyl acetate layer with a 500ml conical flask, add anhydrous sodium sulfate, let it stand, and dry. Use a rotary evaporator to evaporate ethyl acetate to obtain oily compound II (15.6g, 55.26mmol), with a yield of 87.7%.

[0034] 2. Preparation of Compound III

[0035] A 250ml eggplant-shaped bottle was selected, and compound II (14.00g, 49.59mmol), 50ml N,N-dimethylformamide, propargyl bromide (11.31g, 95.07mmol), anhydrous potassium carbonate (13.14g, 95.07mmol), potassium iodide (823mg, 4.96mmol) were added in sequence, and magnetic stirring was carried out at room temperature for 6 hours. After TLC monitoring indicated that the reaction was completed, the reaction solution was extracted and washed twice with 200ml ethyl acetate and 200ml water using a 500ml separatory funnel, and the ethyl acetate layer was collected with a 500ml conical flask, and anhydrous sodium sulfate was added to stand and dry. The ethyl acetate was evaporated with a rotary evaporator to obtain a crude compound, and then subjected to column chromatography [the mobile phase was petroleum ether / ethyl acetate (V / V=2 / 1→1 / 1] to obtain a solid compound 3 (11.72g, 36.61mmol), with a yield of 73.8%.

[0036] 3. Preparation of target compound A1:

[0037] Select a 25ml eggplant-shaped bottle, add sodium azide (130mg, 2.00mmol), 3,4-difluorobenzyl bromide (207mg, 1mmol), 10ml of dimethyl sulfoxide in sequence, and stir magnetically at room temperature for 6-8 hours. After TLC monitoring indicates that the reaction of benzyl bromide is complete, add compound 3 (160mg, 0.50mmol), sodium ascorbate (20mg, 0.1mmol), copper sulfate pentahydrate (25mg, 0.1mmol), 1ml of purified water, and stir magnetically at room temperature for 15 minutes. After TLC monitoring indicates that the reaction of compound 3 is complete, add 10ml of dilute ammonia water to the reaction solution. Use a 250ml separatory funnel to extract and wash the reaction solution twice with 100ml ethyl acetate and 100ml water, collect the ethyl acetate layer, extract twice with dilute hydrochloric acid, separate the aqueous layer with a 250ml conical flask, add anhydrous sodium carbonate to the aqueous layer to adjust the pH to about 7, and then extract twice with 100ml ethyl acetate. After separating the ethyl acetate layer, add anhydrous sodium sulfate, let stand, dry for 4 hours, filter, and remove the solvent under reduced pressure to obtain solid compound A1 (146mg, 0.30mmol) with a yield of 59.7%.

[0038] 4. Preparation of target compound A2: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(2,6-difluorobenzyl)methylamino]}-2-propanol:

[0039] Using 2,6-difluorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0040] 5. Preparation of target compound A3: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(3,5-difluorobenzyl)methylamino]}-2-propanol:

[0041] Using 3,5-difluorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0042] 6. Preparation of target compound A4: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(2,4-difluorobenzyl)methylamino]}-2-propanol:

[0043] Using 2,4-difluorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0044] 7. Preparation of target compound A5: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(2,5-difluorobenzyl)methylamino]}-2-propanol:

[0045] Using 2,5-difluorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0046] 8. Preparation of target compound A6: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(3,4-dichlorobenzyl)methylamino]}-2-propanol:

[0047] Using 3,4-dichlorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0048] 9. Preparation of target compound A7: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(2,3-dichlorobenzyl)methylamino]}-2-propanol:

[0049] Using 2,3-dichlorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0050] 10. Preparation of target compound A8: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(3,5-dichlorobenzyl)methylamino]}-2-propanol:

[0051] Using 3,5-dichlorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0052] 11. Preparation of target compound A9: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(2,5-dichlorobenzyl)methylamino]}-2-propanol:

[0053] Using 2,5-dichlorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0054] 12. Preparation of target compound A10: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(3-chloro, 4-fluorobenzyl)methylamino]}-2-propanol:

[0055] Using 3-chloro, 4-fluorobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0056] 13. Preparation of target compound A11: 1-(1H-1,2,4-triazol-1-yl)-2-(2,4-difluorophenyl)-3-{N-ethyl-N-[1H-1,2,3-triazol-4-yl-1-(3,5-dibromobenzyl)methylamino]}-2-propanol:

[0057] Using 3,5-dibromobenzyl bromide as raw material, the specific preparation method is the same as the preparation of compound A1.

[0058] The reagents used in the examples of the present invention are all commercially available analytically pure.

[0059] The chemical structure, yield, molecular formula and H NMR spectrum data of the above prepared compounds are shown in Table 1:

[0060] Table 1 Chemical structure, molecular formula and H NMR spectrum of preferred compounds

[0061]

[0062]

[0063]

[0064]

[0065] Example 2: Pharmacological Experiments of 1,2,3-Triazole Compounds

[0066] In this example, a conventional in vitro antibacterial experimental method was used.

[0067] 1. Experimental Materials

[0068] (1) Experimental strains:

[0069] This experiment selected the following three common human pathogenic fungal strains as screening objects.

[0070] ATCC standard strains: Candida albicans SC5314, Cryptococcus neoformans h99.

[0071] Clinical strain: Aspergillus fumigatus 7544 was provided by the New Drug Research Center of the Department of Pharmacy, Naval Medical University. It was collected from clinical samples and identified by morphology and biochemistry.

[0072] (2) Culture medium:

[0073] RPMI 1640 culture medium

[47] : RPMI1640 (Gibco BRL) 10 g, NaHCO3 2.0 g, morpholinepropanesulfonic acid (MOPS) (Sigma) 34.5 g (0.165 M), add 800 ml of triple-distilled water to dissolve, adjust the pH to 7.0 (25°C) with 1N NaOH, make up to 1000 ml, filter and sterilize, and store at 4°C.

[0074] Sandburg Dextrose Agar Medium (SDA): 10 g of peptone, 40 g of glucose, and 18 g of agar are dissolved in 900 ml of triple-distilled water, and 50 ml of 2 mg / ml chloramphenicol aqueous solution is added to adjust the pH to 7.0, and the volume is made up to 1000 ml. After high-pressure sterilization, it is stored at 4°C.

[0075] YEPD culture medium: 10g yeast extract, 20g peptone, 20g glucose, add 900ml triple-distilled water to dissolve, add 50ml 2mg / ml chloramphenicol aqueous solution, make up to 1000ml, and store at 4℃ after high-pressure sterilization.

[0076] (3) Drug testing:

[0077] Fluconazole, ketoconazole, and voriconazole were provided by the New Drug Research Center of the School of Pharmacy, Second Military Medical University. DMSO was domestically produced and of analytical grade.

[0078] (4) Instruments: Water-insulated electric constant temperature incubator (Shanghai Yuejin Medical Device Factory); THZ-82A desktop constant temperature oscillator (Shanghai Yuejin Medical Device Factory); Multiskan MK3 enzyme label analyzer (Labsystems Dragon)

[0079] 2. Experimental methods

[0080] (1) Preparation of bacterial solution:

[0081] Before the experiment, a small amount of spherical fungi such as Cryptococcus neoformans, Candida albicans and Candida parapsilosis were picked from the SDA medium stored at 4°C using an inoculation loop and inoculated onto the YPD medium. The fungi were grown at 30°C for one day to allow the fungi to be in the late exponential growth phase. The fungi were added to 1 ml of YEPD culture medium and reactivated using the above method. After 16 hours, the fungi were counted using a hemocytometer and the concentration of the culture medium was adjusted to 1×10 3 ~5×10 3 Pieces / ml.

[0082] The filamentous fungi were inoculated onto the SDA slant, and the subcutaneous tissue fungi and systemic fungi were cultured at 35°C for one week. After each fungus was activated twice, an appropriate amount of RPMI1640 culture medium was added to the SDA slant, and the colonies were blown with a pipette to make the fungal spores free in the RPMI1640 culture medium. After the culture medium was counted with a hemocytometer, the spore concentration was adjusted to 1×10 3 ~5×10 3 Pieces / ml.

[0083] (2) Preparation of drug solution:

[0084] The tested drugs were prepared into 64 g·L-1 solutions with DMSO and stored at -20°C. Before the experiment, the drug solutions were taken out and placed in a 35°C incubator for melting.

[0085] (3) Preparation of drug sensitivity plate:

[0086] Take a sterile 96-well plate, add 200l of RPMI 1640 to well 1 of each row as a blank control; add 100l of freshly prepared bacterial solution to wells 3 to 12; and add 200l of test compound solution to well 2. Make 10-fold dilutions in wells 2 to 11, so that the final drug concentrations in each well are 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25 and 0.125 mg·L-1, respectively. Add 100l of bacterial solution to wells 2 to 12, and the DMSO content in each well is less than 1%; well 12 does not contain drugs as a positive control. Each drug sensitivity plate is incubated at 35°C.

[0087] (4) MIC value determination:

[0088] Candida, Cryptococcus neoformans and filamentous fungi were cultured at 30°C for 24h, 72h and one week, respectively. The drug concentration in the lowest well where the OD value decreased by more than 80% compared with the positive control well was observed as MIC80 (drug concentration when fungal growth was inhibited by 80%).

[0089] When the MIC80 value of the drug exceeds the measured concentration range, it is counted according to the following method: when the MIC80 value is higher than the maximum concentration of 64 mg·L-1, it is counted as ">64 mg·L-1"; when the MIC50 value is the lowest concentration or below the lowest concentration, no distinction is made and it is counted as "≤0.125 mg·L-1".

[0090] The above experiments were performed in parallel 2 to 3 times. The MIC80 values ​​were accepted only when they could be accurately repeated or differed by only one concentration, and the higher concentration was used as the MIC80 value. [; When the MIC80 values ​​differed by more than two concentrations, the experiment had to be repeated until it met the requirements.

[0091] 3. Experimental results

[0092] In vitro antifungal activity of the compounds to be screened:

[0093] In this example, the CLSI-recommended micro-liquid dilution method was used to detect the minimum inhibitory concentration (MIC) values ​​of the compounds to be screened against three common clinical pathogenic fungi. The experimental data are shown in Table 1.

[0094] Quality Control:

[0095] During the experiment, the MIC fluctuation range of the quality control drug fluconazole did not exceed 1 drug gradient concentration, which was within the MIC standard range of the quality control strains published by CLSI-M27A3 and CLSI-M38A2, indicating that the result data in this example are reliable.

[0096] The results of in vitro antifungal experiments are shown in Table 2:

[0097] Table 2 In vitro activity of a series of compounds against common pathogenic fungi (MIC μg / ml)

[0098]

[0099]

[0100] Among them, FCZ. Fluconazole, VCZ. Voriconazole.

[0101] It can be seen from this that the compounds of the present invention and their salts can be used to prepare antifungal drugs.

[0102] This specific embodiment is merely an explanation of the present invention and is not a limitation of the present invention. After reading this specification, those skilled in the art may make non-creative modifications to the present embodiment as needed. However, as long as they are within the scope of the claims of the present invention, they are protected by the patent law.

Claims

1. 1,2,3-triazole compounds characterized by: The general structural formula of the compound is: Wherein, the R group represents various dihalogen substituents, and the halogen is F, Cl or Br.

2. The 1,2,3-triazole compound according to claim 1, characterized in that: The compounds include pharmaceutically acceptable salts.

3. The 1,2,3-triazole compound according to claim 2, characterized in that: The pharmaceutically acceptable salt is acetate, oxalate, bisulfate, citrate, hydrochloride, sulfate, hydrobromide or methanesulfonate.

4. A pharmaceutical composition characterized by: The invention comprises the 1,2,3-triazole compound as described in any one of claims 1 to 3 and a pharmaceutically acceptable carrier.

5. The method for preparing a 1,2,3-triazole compound according to any one of claims 1 to 3, characterized in that: The following steps are involved: S1. Dissolve 2-(2,4-difluorophenyl)-2-[(1H-1,2,4-triazol-1-yl)methyl]oxirane methanesulfonate, ethylamine and triethylamine in anhydrous ethanol, heat and stir to reflux, and continue the reaction for 6 hours to obtain key compound II; S2, dissolving the key compound II in anhydrous DMF, adding anhydrous potassium carbonate, potassium iodide, and propyne bromide, stirring at room temperature overnight to obtain compound III; S3. Dissolve compound III in DMSO, add various disubstituted benzyl bromides and sodium azide reaction solution, prepare a catalyst by dissolving copper sulfate pentahydrate and sodium ascorbate in water, add the catalyst to the reaction solution, stir at room temperature for 1 hour, and obtain the target compound A1-11.

6. Use of the 1,2,3-triazole compound according to any one of claims 1 to 3 in the antifungal field.

7. Use of the 1,2,3-triazole compound according to any one of claims 1 to 3 in the preparation of antifungal drugs.

Images