Application of zinc ion chelating agent in anti-Candida albicans biofilm of azole drugs

Abstract

The invention discloses a method for enhancing Candida albicans biofilm resistance of azole drugs by using a zinc ion chelating agent, through the combined action of the zinc ion chelating agent N, N, N ', N'-tetra (2-picolyl) ethylenediamine (TPEN) and fluconazole, the activity of the Candida albicans biofilm can be reduced, and the Candida albicans biofilm can be enhanced. The early-stage adhesion process, the medium-stage yeast-to-hypha conversion process and the late-stage maturation process formed by the candida albicans biological membrane are inhibited, the structure of the candida albicans biological membrane is destroyed, then the high drug resistance of the candida albicans biological membrane to fluconazole is reversed, and the pathogenicity of the fluconazole is reduced; the problems of high drug resistance and sustained infection caused by the formation of a candida albicans biofilm are hopefully solved, and a direction is provided for the treatment of clinical fungal infection and the research and development of drugs.

Description

Technical Field

[0001] The invention relates to the field of medical technology, and in particular to application of TPEN in fluconazole anti-Candida albicans biofilm. Background Art

[0002] Candida albicans is a fungus widely found in nature and the human body, and its infection is often associated with a decrease in human resistance. Up to 80% of infections are related to the biofilm it forms. This biofilm is a complex three-dimensional structure composed of yeast spores, pseudohyphae and fungal hyphae, as well as extracellular matrix, which exhibits extremely strong resistance and tolerance to the host immune defense system and environmental stress factors. Compared with the planktonic Candida albicans, the formation of Candida albicans biofilm leads to an increase of more than 1,000 times in resistance to antifungal drugs represented by fluconazole, resulting in failure of clinical antifungal treatment. Therefore, there is an urgent need to develop new methods to overcome the azole resistance of Candida albicans caused by biofilm formation.

[0003] N,N,N',N'-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) is a zinc ion chelator with the chemical formula Because it is fat-soluble, it can pass through the cell membrane and chelate zinc ions in cells. In the biomedical field, TPEN is often used to study the role of zinc ions in cells and various biological processes, for example, to study the structure and function of biomacromolecules such as zinc metalloproteinases, chelate zinc ions in bacterial cells to produce antibacterial effects, etc. So far, there have been no reports on the use of TPEN in Candida albicans biofilms, nor has it been disclosed whether TPEN has a substantial effect in fluconazole against Candida albicans biofilms. Summary of the invention

[0004] The object of the present invention is to overcome at least one deficiency of the prior art and provide an application of a zinc ion chelator in the treatment of Candida albicans biofilm with azole drugs.

[0005] The technical solution adopted by the present invention is:

[0006] The present invention provides a method for enhancing the anti-Candida albicans biofilm activity of fluconazole (FLC) by using zinc chelator TPEN, aiming to reduce the activity of Candida albicans biofilm, inhibit the early adhesion process, the mid-term yeast to hyphae transformation process, and the late maturation process of Candida albicans biofilm growth, and destroy the Candida albicans biofilm structure.

[0007] In a first aspect, the present invention provides an application of a zinc ion chelator in the preparation of an azole drug synergist for inhibiting Candida albicans biofilm, wherein the zinc ion chelator is N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine.

[0008] In a second aspect, the present invention provides an application of a zinc ion chelator in the preparation of a drug synergist against Candida albicans biofilm, wherein the zinc ion chelator is N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine.

[0009] In a third aspect, a zinc ion chelator is used in synergistically inhibiting Candida albicans biofilm with azole drugs, wherein the zinc ion chelator is N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine.

[0010] In some examples, the azole drug is selected from fluconazole.

[0011] In some examples, the zinc ion chelator further includes N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine or a pharmaceutically acceptable solvate thereof.

[0012] In some examples, the drug is a topical agent, an oral agent, an injection agent, an implant agent, a rectal agent, a spray agent, or an inhalation agent.

[0013] In some examples, the dosage form of the drug includes but is not limited to solution, tincture, spirit, lotion, ointment, plaster, paste, oil, film, liniment, injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

[0014] In some examples, the drug further includes a pharmaceutical carrier or a pharmaceutically acceptable excipient.

[0015] In a fourth aspect, the present invention provides a composition for inhibiting Candida albicans or Candida albicans biofilm, wherein the effective components of the composition are N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine or a pharmaceutically acceptable solvate thereof and an azole drug.

[0016] The beneficial effects of the present invention are:

[0017] 1) Combination drug strategy of TPEN and fluconazole: The present invention proposes a new combination drug regimen, namely the combined use of TPEN and fluconazole, which aims to provide a more effective prevention and treatment method for Candida albicans biofilm infection. This combination drug strategy is expected to interfere with the physiological function of Candida albicans through the specific chelation of zinc ions by TPEN, while fluconazole exerts its antifungal effect and acts on fungal cells together to achieve a synergistic effect.

[0018] 2) Therapeutic mechanism targeting zinc ion dependence: In view of the key role of zinc ions in Candida albicans, the scheme of the present invention is specifically targeted at this biological characteristic. By chelating zinc ions in cells with TPEN, the transcriptional regulation and enzyme activity of the fungus are interfered with, thereby weakening the formation and maintenance of biofilms, providing a new idea for the treatment of drug-resistant biofilms.

[0019] 3) Reduce the risk of drug resistance: The scheme of the present invention is expected to reduce the development of drug resistance. Through the dual mechanism of action of TPEN and fluconazole, the possibility of fungi developing resistance to a single drug is reduced, providing a long-term and sustainable treatment plan for clinical treatment.

[0020] 4) Reduce toxic and side effects: The present invention aims to reduce the toxic and side effects that may be caused by the use of a single drug, provide a safer treatment plan, and improve the patient's treatment experience and treatment effect.

[0021] 5) Possibility of clinical application: The treatment regimen of the present invention is expected to be widely used in the treatment of fluconazole-resistant Candida albicans biofilm infections in clinical practice, and has important public health significance and market application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a graph showing the effect of TPEN concentration-dependently reducing the zinc ion concentration in Candida albicans cells in Example 1.

[0023] Figure 2 This is a graph showing the concentration-dependent inhibition of the growth of planktonic Candida albicans by TPEN in Example 2.

[0024] Figure 3 TPEN enhances the antibacterial effect of fluconazole on planktonic Candida albicans in Example 3; wherein A is a graph showing the growth inhibition of Candida albicans by TPEN combined with fluconazole in YPD medium; B is a graph showing the antibacterial effect of Candida albicans co-incubated with TPEN combined with fluconazole in RPMI 1640 medium for 24 hours and observed under a microscope; C is a graph showing the change in fluconazole sensitivity of Candida albicans detected by the disc diffusion method; and D is a statistical graph showing the diameter of the inhibition zone by the disc diffusion method.

[0025] Figure 4 This is a graph showing the inhibitory effect of TPEN on the adhesion of Candida albicans by enhancing fluconazole in the early stage of biofilm formation in Example 4.

[0026] Figure 5 This is a statistical diagram of the biomass of TPEN enhancing the inhibition of fluconazole on the adhesion of Candida albicans in the early stage of biofilm formation in Example 4.

[0027] Figure 6 This is a graph showing the inhibitory effect of TPEN on the yeast-hyphae conversion of Candida albicans in the middle stage of biofilm formation in Example 5.

[0028] Figure 7 This is a graph showing the inhibitory effect of TPEN on the maturation of Candida albicans biofilm in the late stage of biofilm formation in Example 6.

[0029] Figure 8 This is a statistical diagram of the biomass of Candida albicans biofilm maturation in which TPEN enhances the inhibition of fluconazole at the late stage of biofilm formation in Example 6.

[0030] Fig. 9 This is a diagram showing the therapeutic effect of greater wax moth systemically infected with Candida albicans biofilm in Example 8. DETAILED DESCRIPTION

[0031] The present invention will be further described below in conjunction with specific embodiments, but the present invention is not limited thereto.

[0032] The sources of experimental materials in each embodiment of the present invention are as follows:

[0033] 1. Drugs and reagents

[0034]

[0035]

[0036] 2. Instruments

[0037]

[0038] 3. Experimental strains

[0039] FLC-sensitive Candida albicans SC 5314, a common strain in international laboratories, was purchased from Hangzhou Fenghai Biotechnology Co., Ltd.

[0040] 4. Experimental Animals

[0041] Six-week-old greater wax moth larvae were purchased from Tianjin Huiyude Biotechnology Co., Ltd., with a length of 2-3 cm, a weight of 250-350 mg, good vitality, and no black spots.

[0042] Example 1 Determination of zinc ion levels in Candida albicans cells

[0043] Experimental process:

[0044] 1) Weigh 5 mg of zinc ion fluorescent probe Zinquin ethyl ester powder into a 1.5 mL centrifuge tube, add 1.206 mL DMSO to dissolve, which is a 10 mM Zinquin ethyl ester stock solution, and filter it three times with a 0.22 μm syringe filter for later use. Dilute the Zinquin ethyl ester stock solution with DMSO to a 40 μM staining working solution.

[0045] 2) Add 1×10 7albicans cells / ml, and RPMI 1640 medium without drugs was used as control.

[0046] 3) Cultivate the suspension at 37°C for 16 h at 200 rpm. Centrifuge the suspension for 10 min (3000 rpm, 4°C), discard the supernatant, and centrifuge again. Repeat the centrifugation three times.

[0047] 4) Resuspend the bacterial suspension in 40 μM Zinquin ethyl ester staining working solution and incubate at 37°C in the dark for 30 minutes. Centrifuge the bacterial suspension for 10 minutes (3000 rpm, 4°C), discard the supernatant, add physiological saline, vortex and mix, centrifuge again, and repeat centrifugation 3 times to remove the residual staining solution.

[0048] 5) Resuspend Candida albicans with physiological saline, take 200 μL of bacterial suspension into a 96-well plate, and calibrate the OD600 values ​​to be consistent. Set the excitation wavelength to 355 nm and the emission wavelength to 491 nm on the microplate reader to read the fluorescence intensity of each experimental group.

[0049] Experimental results: Figure 1 As shown in the figure, with the increase of TPEN concentration, the fluorescence intensity of zinc ions in Candida albicans cells gradually decreased, and the difference was statistically significant compared with the control group (**P<0.01), indicating that TPEN reduces the zinc ion level in Candida albicans cells by chelating zinc ions.

[0050] Example 2 Inhibitory effect of TPEN on the growth of planktonic Candida albicans

[0051] Experimental process:

[0052] 1) 1.0×10 6 cell·mL -1 Candida albicans was incubated in 5 mL of YPD liquid medium containing 20 μM, 100 μM, and 500 μM TPEN, and cultured at 37°C with continuous shaking (200 rpm). YPD liquid medium without drugs was used as a positive control.

[0053] 2) After the samples were thoroughly mixed at fixed time points (0, 1, 3, 6, 12 and 24 h), 100 μL of bacterial suspension was taken from each group, diluted in saline, and spread on YPD agar plates.

[0054] 3) After incubation at 37°C for 24–48 h, colony-forming units (CFU) were counted.

[0055] Experimental results: Figure 2As shown, in yeast amplification YPD liquid medium, as the concentration of TPEN increased, the growth of Candida albicans was significantly inhibited, indicating that TPEN inhibited the growth of planktonic Candida albicans in a concentration-dependent manner.

[0056] Example 3: Changes in the Fluconazole Sensitivity of TPEN to Planktonic Candida albicans

[0057] Experimental process:

[0058] 1. Determination of MIC and MFC of drugs against Candida albicans

[0059] 1) Take a 96-well plate and add 100 μL RPMI 1640 liquid medium and 5×10 3 CFU·mL -1 100 μL of bacterial suspension was added as a positive control. 100 μL of fluconazole working solution with different concentrations was added to wells 2-11 in descending order. 5×10 3 CFU·mL -1 The final concentrations of fluconazole in each well were 64, 32, 16, 8, 4, 2, 1, 0.5, 0.25, and 0.125 μg mL -1 The final concentration of the bacterial solution was 2.5x10 3 CFU·mL -1 ; Add 200 μL RPMI 1640 liquid culture medium to the 12th well as a negative control. After incubation at 37°C for 24-48 hours, measure the absorbance of each well at a wavelength of 600 nm. Compared with the positive control well, the minimum drug concentration corresponding to more than 80% inhibition of fungal growth is the minimum inhibitory concentration (MIC) of fluconazole.

[0060] 2) The MIC of TPEN was determined using the above method.

[0061] 3) After supplementing each well plate with TPEN at a final concentration of 0.5×MIC, the change in fluconazole MIC was determined by the above method.

[0062] 4) After incubating the 96-well plate for 48 hours, take 100 μL from each well plate and spread it on YPD solid culture medium. Continue to culture at 37°C for 24-48 hours. Determine the lowest drug concentration at which no colonies grow, which is the minimum fungicidal concentration (MFC).

[0063] 5) The growth of Candida albicans after 36 h of drug incubation was observed under an inverted microscope, and image acquisition was performed using cellSensDimension software (Olympus).

[0064] 2. Evaluation of the changes in the susceptibility of TPEN to fluconazole in Candida albicans by drug sensitivity paper diffusion method

[0065] 1) Add 5×10 6 CFU·mL -1 Candida albicans was cultured at 37°C with shaking at 200 rpm for 2 h.

[0066] 2) The bacterial suspension was inoculated on a Mueller Hinton (MH) agar plate (containing 20 g / L glucose and 0.5 μg / mL methylene blue), and a drug-sensitive paper containing 25 μg of fluconazole was placed in the center of the plate. After incubation at 37°C for 24 h, the diameter of the inhibition zone (mm) was measured using an electronic vernier caliper, and the image of the culture dish was recorded using a camera. The experiment was repeated three times.

[0067] Experimental results: As shown in Table 1, the MIC of fluconazole was 0.5 μg / ml and the MFC was > 1024 μg / ml. After supplementation with 0.5×MICTPEN, the MIC of fluconazole dropped to 0.25 μg / ml and the MFC dropped to 128 μg / ml. Figure 3 As shown in A, 1024 μg / ml fluconazole had a growth inhibitory effect on Candida albicans, the addition of 20 μM TPEN increased the growth inhibitory effect of fluconazole, and the addition of 100 μM TPEN resulted in a bactericidal effect of fluconazole within 24 h. Figure 3 As shown in Figure B, the growth of Candida albicans was observed under a microscope. Fluconazole limited the hyphal extension and colony formation of Candida albicans in a concentration-dependent manner. When TPEN was used in combination with fluconazole, the growth of Candida albicans was significantly inhibited, and hyphal extension and colony formation were further reduced. Figure 3 As shown in C and 3D, the addition of TPEN significantly increased the diameter of the fluconazole inhibition zone of Candida albicans, and the difference was statistically significant (*P<0.05). The above results show that TPEN enhances the fluconazole sensitivity of planktonic Candida albicans.

[0068] Table 1 MICs and MFCs of drugs against Candida albicans

[0069]

[0070] Example 4 Effect of TPEN on Fluconazole Inhibition of Early Adhesion of Candida albicans Biofilm

[0071] Experimental process:

[0072] 1) 1×10 6 CFU / mL of Candida albicans was resuspended in 1 ml of RPMI 1640 medium containing drugs, and medium without drugs was used as a control, and added to 24-well plates and incubated for 2 h.

[0073] 2) Remove non-adherent cells with physiological saline, fix with 1 ml / well methanol, stain with 0.1% crystal violet, rinse with double distilled water, dry at room temperature, and observe under an inverted microscope.

[0074] 3) 95% ethanol was added at 1 mL / well and incubated for 10 min until the crystal violet was completely dissolved. 200 μl of the ethanol solution containing the crystal violet was pipetted into another 96-well plate. The A570 value was read by a microplate reader to quantitatively analyze the biomass of the Candida albicans biofilm.

[0075] Experimental results: Figure 4 and Figure 5 As shown in the figure, in the early stage of biofilm formation, 1024μg / ml fluconazole had no significant inhibitory effect on the adhesion of Candida albicans, 80μM TPEN significantly inhibited the adhesion of Candida albicans, and TPEN combined with fluconazole had a better inhibitory effect on the adhesion of Candida albicans, and the difference was statistically significant compared with single drug (***P<0.001). The above results show that: in the early stage of biofilm formation, TPEN enhances the inhibitory effect of fluconazole on the adhesion of Candida albicans.

[0076] Example 5 Effect of TPEN on Fluconazole Inhibition of Yeast-Hypha Transition in Mid-stage Biofilm of Candida albicans

[0077] Experimental process:

[0078] 1) Prepare RPMI 1640 medium (containing 10% fetal bovine serum) containing different concentrations of drugs, and medium without drugs as a control, add a final concentration of 1×10 6 CFU / mL of Candida albicans was cultured with shaking at 200 rpm at 37 °C for 4 h to induce hyphae formation of Candida albicans.

[0079] 2) 1 ml of bacterial suspension was taken from each group and added to a 24-well plate, which was then placed under an inverted microscope for observation, and image acquisition was performed using cellSensDimension software (Olympus). Adobe Photoshop 25.0.0 (Adobe Systems, California, USA) was used to adjust the grayscale conversion, brightness, and contrast of the image.

[0080] Experimental results: Figure 6 As shown, both fluconazole and TPEN inhibited the budding, hyphae formation and elongation of Candida albicans in a concentration-dependent manner, and the combination of TPEN and fluconazole further enhanced the inhibitory effect on the yeast-hyphae conversion of Candida albicans.

[0081] Example 6 The role of TPEN in fluconazole inhibition of late maturation of Candida albicans biofilm

[0082] Experimental process:

[0083] 1) Adjust the concentration of Candida albicans to 1×10 in RPMI 1640 medium. 6 CFU / mL, add 1 ml of bacterial suspension to each well of a 24-well plate and incubate at 37°C for 2 h.

[0084] 2) Wash the non-adherent cells with saline, add 1 ml of RPMI 1640 medium containing drugs to each well, and use medium without drugs as a control.

[0085] 3) After incubation at 37°C for 24 h, non-adherent cells were removed with physiological saline, fixed with 1 ml / well methanol, stained with 0.1% crystal violet, rinsed with double distilled water, dried at room temperature, and observed under an inverted microscope.

[0086] 4) 95% ethanol was added at 1 mL / well and incubated for 10 min until the crystal violet was completely dissolved. 100 μl of the ethanol solution containing the crystal violet was pipetted into another 96-well plate. The A570 value was read by an ELISA reader to quantitatively analyze the biomass of the Candida albicans biofilm.

[0087] Experimental results: Figure 7 and Figure 8 As shown in the results, in the late stage of biofilm formation, co-incubation with 1024 μg / ml fluconazole caused morphological defects in the biofilm of Candida albicans, a decrease in the number of cells in the biofilm, a decrease in the cross-linking between yeast and hyphae, and a decrease in the extracellular matrix. The biomass of the biofilm was significantly different from that of the control group (***P < 0.001). TPEN combined with fluconazole further reduced the maturity of the biofilm, significantly reduced hyphae, and the biomass of the biofilm was significantly different from that of the single drug (***P < 0.001). The above results indicate that in the late stage of biofilm formation, TPEN enhances the inhibitory effect of fluconazole on the maturation of Candida albicans biofilm.

[0088] Example 7 TPEN synergistic effect of fluconazole against Candida albicans biofilm

[0089] Experimental process:

[0090] 1) Adjust the concentration of Candida albicans to 1×10 in RPMI 1640 medium. 6 CFU / mL, add 100 μl of bacterial suspension to each well of a 96-well plate and incubate at 37°C for 2 h.

[0091] 2) Non-adherent cells were removed by washing with saline, and 200 μl of RPMI 1640 medium containing drugs diluted in series was added to make the final concentrations of fluconazole and TPEN range from 0.5 to 1024 μg / ml and 1.25 μM to 160 μM, respectively. Medium without drugs was used as a control.

[0092] 3) After incubation at 37°C for 24 h, the cells were gently rinsed three times with 200 μL / well PBS, 100 μL XTT / menadione solution was added to each well, and the cells were incubated at 37°C in the dark for 2 h.

[0093] 4) 80 μL of supernatant was transferred from each well to a new 96-well cell culture plate, and the A490 nm value was read by a microplate reader to qualitatively analyze the activity of the Candida albicans biofilm.

[0094] 5) Calculate the minimum inhibitory concentration (SMIC) 80 (sessile minimal inhibitory concentration 80%), which is the drug concentration in the lowest drug concentration well when the colorimetric reading OD value is reduced by more than 80% compared with the positive control well (column 11). That is, the percentage of fungal growth inhibition = 1-(OD value of each dosing well) 490 - Negative control well OD 490 ) / (OD of positive control well 490 - Negative control well OD 490 )×100%

[0095] Experimental results: As shown in Table 2, 80 μM TPEN inhibited biofilm activity by more than 80%, so the SMIC 80 of TPEN against Candida albicans biofilm was 80 μM. As shown in Table 3, the SMIC 80 of fluconazole of Candida albicans was greater than 1024 μg / ml, and supplementation with 5 μM TPEN reduced it to 64 μg / ml, with a FICI value of 0.125, indicating that TPEN and fluconazole had a synergistic effect (SYN). As the concentration of TPEN increased, the SMIC 80 of fluconazole decreased significantly, and 40 μM TPEN reduced it to <1 μg / ml, indicating that TPEN reversed the fluconazole resistance of the biofilm. The above results show that TPEN can cooperate with fluconazole against Candida albicans biofilm and reverse the azole resistance of the biofilm.

[0096] Table 2 Effects of different concentrations of TPEN on anti-Candida albicans biofilm (mean ± SD, n = 3)

[0097]

[0098] aRelative absorbance = absorbance of experimental group / absorbance of control group. The ratio of the absorbance of A490 of Candida albicans biofilm to that of the control group measured by XTT reduction method under different concentrations of TPEN treatment.

[0099] Table 3 Evaluation of the anti-Candida albicans biofilm effect of TPEN combined with fluconazole for 24 h

[0100]

[0101] a The fractional inhibitory concentration index (FICI) method is a widely used method for evaluating the interaction of combined drugs. The calculation formula is as follows: FICI = C A / MIC A +C B / MIC B , C A With C B MIC is the minimum inhibitory concentration (MIC) of drugs A and B when they are used together to achieve the same efficacy. A With MIC B are the minimum inhibitory concentrations of drugs A and B when used alone.

[0102] b SYN, indicates synergistic effect; NI, indicates irrelevant effect. FICI ≤ 0.5 indicates synergistic effect, FICI > 4 indicates antagonistic effect, and FICI between 0.5 and 4 indicates additive or irrelevant effect.

[0103] Example 8 Effect of TPEN in Fluconazole Treatment of Candida albicans Biofilm Infection

[0104] Experimental process:

[0105] 1) 48 wax moth larvae of uniform size and good vitality were selected and randomly divided into 4 groups.

[0106] 2) Use RPMI 1640 medium to adjust the concentration of Candida albicans to 1×10 6 CFU / ml, biofilms were cultured in adherent cell dishes with a diameter of 15 cm.

[0107] 3) After 24 hours, the biofilms were collected in culture dishes, centrifuged three times (3000 g, 20 min), and the supernatant was removed. Then 5 ml of physiological saline was added to each group to prepare a biofilm suspension.

[0108] 4) Use a syringe to extract 10 μl of the biofilm suspension, inject it into the body of the greater wax moth larvae through the left front first abdominal foot, and incubate at 37°C in the dark for 2 h.

[0109] 5) Use a syringe to draw 10 μl of a solution containing 500 μg / ml fluconazole and / or 80 μM TPEN, and inject it into the body of the greater wax moth larvae through the first right anterior ventral foot. The group without drug served as the positive control group, and the group injected with only normal saline served as the blank control.

[0110] 6) The larvae were cultured at 37°C in the dark. Larvae that were immobile or unresponsive to slight touch were considered dead. The number of surviving larvae was recorded every day and observed for 4 days.

[0111] Experimental results: Fig. 9 As shown, after 4 days of hatching, compared with the 8.33% survival rate of the WT group, the survival rates of the fluconazole group and the TPEN group were significantly increased, 41.7% and 33.3%, respectively. The combination of TPEN and fluconazole further increased the survival rate of the greater wax moth larvae to 83.33%, and the difference between the groups was statistically significant (P<0.05). The above results show that TPEN can cooperate with fluconazole to treat Candida albicans biofilm infection.

[0112] The above is a further detailed description of the present invention, which should not be regarded as a limitation on the specific implementation of the present invention. For ordinary technicians in the technical field to which the present invention belongs, simple deduction or replacement without departing from the concept of the present invention is within the protection scope of the present invention.

Claims

1. An application of a zinc ion chelator in the preparation of an azole drug synergist for inhibiting Candida albicans biofilm, characterized in that: The zinc ion chelating agent is N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine.

2. Use of a zinc ion chelator in the preparation of a drug synergist for resisting Candida albicans biofilm, characterized in that: The zinc ion chelating agent is N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine.

3. An application of a zinc ion chelator in cooperating with azole drugs to inhibit Candida albicans biofilm, characterized in that: The zinc ion chelating agent is N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine.

4. The use according to claim 1 or 3, characterized in that: The azole drug is selected from fluconazole.

5. The use according to any one of claims 1 to 3, characterized in that: The zinc ion chelator also includes N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine and a pharmaceutically acceptable solvate thereof.

6. The use according to claim 2, characterized in that: The medicine is an external administration agent, an oral administration agent, an injection administration agent, an implantation administration agent, a rectal administration agent, a spray administration agent or an inhalation administration agent.

7. The use according to claim 2, characterized in that: The dosage form of the drug includes, but is not limited to, solution, tincture, spirit, lotion, ointment, plaster, paste, oil, film coating, liniment, injection, tablet, granule, capsule, pill, sustained-release agent, oral liquid preparation, powder or gel.

8. The use according to claim 2, characterized in that: The medicine also includes a drug carrier or a pharmaceutically acceptable excipient.

9. A composition for inhibiting Candida albicans or Candida albicans biofilm, characterized in that: The effective components of the composition are N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine or a pharmaceutically acceptable solvate thereof and an azole drug.

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