Compound TU-BDN, its preparation method and application in preparing drugs for preventing or treating toxoplasma infection

Abstract

The application provides a compound TU-BDN, a preparation method thereof and application of the compound in preparation of a medicine for preventing or treating toxoplasma infection. 20 H 14 N4O5S, the application discloses that TU-BDN has significant inhibitory activity on intracellular parasitic protozoa toxoplasma, has small toxic side effects, and can expand a new way for effective use of TU-BDN. Vero cells infected with toxoplasma are used as an in-vitro experimental model to verify the anti-toxoplasma activity and cell safety of the compound TU-BDN, and the results show that the half effective inhibitory concentration of TU-BDN on toxoplasma is 3.231 micromoles, TU-BDN can significantly inhibit the proliferation of toxoplasma in host cells, the half cytotoxic concentration of TU-BDN on Vero cells is 105.8 micromoles, and the safety index SI is 32.75. Compared with the existing medicines for treating toxoplasma infection, TU-BDN has the problems of large side effects and poor effect, the effective inhibitory concentration of TU-BDN is much smaller than the cytotoxicity, TU-BDN has significant effects on prevention and treatment of toxoplasma infection, and TU-BDN can be widely used for preparation of a medicine for preventing or treating toxoplasma infection.

Description

Technical Field

[0001] This invention belongs to the field of parasitic disease prevention and control technology, specifically relating to a compound TU-BDN, its preparation method, and its application in the preparation of drugs for the prevention or treatment of Toxoplasma gondii infection. Background Technology

[0002] Toxoplasmosis is an obligate intracellular parasitic protozoan disease caused by Toxoplasma gondii. Toxoplasma gondii infects almost all warm-blooded animals and humans, and even some cold-blooded animals, residing within all nucleated cells of the animal's body. Statistics show that most human toxoplasmosis infections are asymptomatic, but infection in infants and immunocompromised patients (such as AIDS patients, organ transplant recipients, and those with malignant tumors) can lead to severe or fatal illness. Toxoplasmosis infection is also a significant cause of miscarriage, stillbirth, and other reproductive disorders in pregnant animals and women, and is one of the mandatory prenatal screening tests in my country.

[0003] Currently, clinical treatment of toxoplasmosis still relies on chemotherapy. Due to the complexity of the Toxoplasma gondii life cycle, the diversity of its pathogenesis, and the differences in its biological characteristics, there are currently no preventative or specific drug treatments. Although the combination of pyrimethamine and sulfadiazine is currently the gold standard for treating toxoplasmosis in clinical practice, this treatment method is often accompanied by serious side effects, and it is not a complete cure, has a high relapse rate, and a high treatment failure rate.

[0004] Therefore, developing a safe and effective anti-Toxoplasma gondii drug is an urgent problem to be solved and also has broad market prospects. In view of this, this invention is proposed. Summary of the Invention

[0005] To address the problems of existing technologies, such as drug resistance, significant toxic side effects, and poor efficacy despite large dosages in preventing or treating toxoplasmosis, this invention specifically provides the compound TU-BDN, its preparation method, and its application in the preparation of drugs for the prevention or treatment of toxoplasmosis, specifically including:

[0006] One of the objectives of this invention is to provide a compound TU-BDN with the molecular formula C 20 H 14 N4O5S, chemical structural formula is:

[0007] .

[0008] A second objective of this invention is to provide a method for preparing the compound TU-BDN, comprising the following steps:

[0009] Add 3,5-dinitrobenzoyl chloride dropwise to a dry acetone solution of potassium thiocyanate. After the addition is complete, heat to reflux under nitrogen protection and maintain reflux for 1.5-2.5 h. After cooling the resulting reaction solution to room temperature, add 2-aminobiphenyl, heat to reflux again, and maintain reflux for 8-10 h. After cooling to room temperature, pour into crushed ice, allow to stand to precipitate, filter, and dry to obtain crude product. Recrystallize the crude product from ethanol to obtain TU-BDN.

[0010] In a preferred embodiment, the molar ratio of potassium thiocyanate to 3,5-dinitrobenzoyl chloride is 6:(3-5); preferably, the molar ratio of potassium thiocyanate to 3,5-dinitrobenzoyl chloride is 3:2.

[0011] In a preferred embodiment, 8-10 mL of dry acetone is used for every 1 mmol of potassium thiocyanate; preferably, 9.1 mL of dry acetone is used for every 1 mmol of potassium thiocyanate.

[0012] In a preferred embodiment, the molar ratio of potassium thiocyanate to 2-aminobiphenyl is 6:(3-5); preferably, the molar ratio of potassium thiocyanate to 2-aminobiphenyl is 3:2.

[0013] In a preferred embodiment, the reflux process following the addition of 2-aminobiphenyl is monitored by TLC until the reaction is complete; preferably, the developing solvent for TLC monitoring is n-hexane and ethyl acetate in a volume ratio of 4:1.

[0014] In a preferred embodiment, the volume of the crushed ice is 2-4 times the volume of the reaction product; the standing time is 0.5-3 h; the drying can be carried out using conventional methods known to those skilled in the art, such as drying at 40-60℃ for 4-8 h; the ethanol recrystallization uses anhydrous ethanol, and the amount of anhydrous ethanol used is 3-8 times the mass of the crude product.

[0015] The third objective of this invention is to provide the application of compound TU-BDN in the preparation of drugs for the prevention or treatment of toxoplasmosis infection.

[0016] In a preferred embodiment, the drug for preventing or treating Toxoplasma gondii infection is a drug that inhibits the ability of Toxoplasma gondii to lyse cells.

[0017] In a preferred embodiment, the drug for preventing or treating toxoplasmosis infection is a drug that inhibits the invasion and proliferation of Toxoplasma gondii.

[0018] In a preferred embodiment, the TU-BDN has a 50% inhibitory effect on Toxoplasma gondii at 3.231 μM.

[0019] A fourth objective of this invention is to provide a composition for the prevention or treatment of toxoplasmosis infection, wherein the active ingredient of the composition includes TU-BDN.

[0020] In a preferred embodiment, the composition for preventing or treating Toxoplasma gondii infection is: a composition that inhibits the ability of Toxoplasma gondii to lyse cells, a composition that inhibits the invasion and proliferation of Toxoplasma gondii, or a composition that eliminates Toxoplasma gondii parasites from the body of humans or non-human animals; preferably, the non-human animals include, but are not limited to, pigs, cattle, sheep, horses, dogs, cats, chickens, etc., which are susceptible to Toxoplasma gondii.

[0021] In a preferred embodiment, the composition further includes one or more pharmaceutically acceptable carriers, the carriers comprising one or more of the following: diluents, wetting agents, binders, disintegrants, lubricants, flavor and color modifiers, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, preservatives, pH adjusters, surfactants, excipients, fillers, and synergists.

[0022] In a preferred embodiment, the diluent includes substances such as starch, sucrose, cellulose, and inorganic salts; the wetting agent includes substances such as water and ethanol; the binder includes substances such as starch paste, dextrin, sugar, cellulose derivatives, gelatin, povidone, and polyethylene glycol; the disintegrant includes substances such as starch, sodium carboxymethyl starch, low-substituted hydroxypropyl cellulose, sodium bicarboxymethyl cellulose, and surfactants; the lubricant includes substances such as talc, calcium stearate, magnesium stearate, magnesium dodecyl sulfate, and polyethylene glycol; the flavoring and coloring agents include substances such as pigments, sweeteners, flavorings, and gelling agents; the solvent includes substances such as water, glycerin, and ethanol; and the solubilizer includes substances such as... The ingredients include: Tween compounds, maltose compounds, sulfates, sulfonates, etc.; the cosolvents include organic acids (such as citric acid) and their salts, inorganic salts, polyethylene glycol, etc.; the emulsifiers include Span compounds, glycerol fatty acid esters, gum arabic, gelatin, agar, sodium alginate, etc.; the antioxidants include sulfites, ascorbic acid, gallic acid and their salts, etc.; the metal complexing agents include disodium ethylenediaminetetraacetate, polycarboxylic acid compounds, etc.; the preservatives include parabens, quaternary ammonium compounds, chlorhexidine acetate, etc.; the pH adjusters include hydrochloric acid, tartaric acid, acetic acid, sodium hydroxide, sodium bicarbonate, ethylenediamine, meglumine, phosphates, citrates, etc.

[0023] In this invention, the dosage forms of the composition include, but are not limited to, oral, topical, suppository, and sterile injectable solutions in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, and sprays. It is understood that the TU-BDN-containing compositions of this invention can be administered in various ways depending on the different excipients and dosage forms.

[0024] Compared with the prior art, the technical solution of the present invention has the following advantages:

[0025] 1. Through long-term research, the inventors discovered that the compound TU-BDN has a significant inhibitory effect on the intracellular parasitic protozoan Toxoplasma gondii, and has few toxic side effects, which can expand new avenues for the effective utilization of TU-BDN.

[0026] 2. The inventors have demonstrated through extensive experiments that the compound TU-BDN has significant preventive or therapeutic effects against Toxoplasma gondii infection: using Vero cells infected with Toxoplasma gondii as an experimental model, EC50 was measured... 50 The results showed that the compound TU-BDN had half the inhibitory effect on Toxoplasma gondii at 3.231 μM, and that a small amount of application could achieve a highly effective anti-Toxoplasma gondii infection effect.

[0027] 3. Further experiments have confirmed that compound TU-BDN, as an anti-Toxoplasma gondii drug, also exhibits low toxicity: Cytotoxicity of TU-BDN against Vero monkey kidney cells was detected using the CCK8 assay, and the results showed that even at a maximum dose of 105.8 μM, TU-BDN showed no cytotoxicity against Vero cells. Therefore, compared to existing drugs for treating Toxoplasma gondii infection, which suffer from significant side effects and poor efficacy, the compound TU-BDN provided in this invention has an effective inhibitory concentration far lower than its cytotoxicity, making it highly effective, low-toxicity, and safe for preparing drugs to prevent or treat Toxoplasma gondii infection.

[0028] 4. Through extensive experimental research, this invention has also discovered that the anti-Toxoplasma gondii effect of the compound TU-BDN is mainly achieved by inhibiting its invasion and intracellular proliferation. Furthermore, the effect of TU-BDN on Toxoplasma gondii is dose-dependent, indicating that the higher the concentration, the better the anti-parasitic effect. Attached Figure Description

[0029] To more clearly illustrate the technical solutions in the specific embodiments of this application or the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of this application. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0030] Figure 1 This is a graph showing the toxicity test results of TU-BDN on Vero cells in Example 1 of the present invention;

[0031] Figure 2 This is a graph showing the inhibitory effect of TU-BDN on Toxoplasma gondii in Example 2 of the present invention.

[0032] Figure 3The figures show the plaque phagocytic results of TU-BDN and the control group in Example 3 of this invention;

[0033] Figure 4 This is a statistical graph showing the anti-proliferative effect of TU-BDN and the control group on intracellular RH in Example 4 of the present invention;

[0034] Figure 5 This is a graph showing the anti-invasion effect of TU-BDN and the control group against extracellular RH-type Toxoplasma gondii in Example 4 of this invention. Detailed Implementation

[0035] To enable those skilled in the art to better understand the present invention, the present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments. However, it should be understood that the scope of protection of the present invention is not limited to the specific embodiments. Based on the embodiments in this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0036] Unless otherwise specified, the technical means used in this invention are conventional means well known to those skilled in the art. All raw materials, reagents, instruments, and equipment used in this invention can be purchased commercially or prepared using existing methods. Unless otherwise specified, all reagents used in this invention are of analytical grade. In the embodiments of this invention, the room temperature is 20-30 °C. The Vero cells and HFF cells used in this invention were purchased from the ATCC cell bank. In the embodiments of this invention, the complete culture medium used is: high-glucose DMEM medium containing 10% fetal bovine serum, supplemented with 100 U / mL penicillin and 100 μg / mL streptomycin.

[0037] In this embodiment of the invention, the preparation method of compound TU-BDN includes the following steps:

[0038] Add freshly prepared 3,5-dinitrobenzoyl chloride (1.1 mmol) dropwise to a freshly prepared potassium thiocyanate (1.65 mmol) and dry acetone (15 mL) solution. After the addition is complete, heat to 56 °C under nitrogen protection and reflux for 2 h to obtain a turbid solution. Cool to room temperature. Add 2-aminobiphenyl (1.1 mmol) to the reaction solution, heat again to 56 °C, and reflux for 8–10 h. Monitor by TLC (hexane and ethyl acetate, volume ratio 4:1) until the starting material spots disappear. Cool to room temperature and pour into crushed ice twice the volume of the reaction product. After standing for 1 hour, a precipitate formed. The solid was collected, filtered, and dried to constant weight to obtain the crude product. Three times the mass of the crude product in anhydrous ethanol was added, and the mixture was heated to reflux (approximately 78°C) to completely dissolve the solid. The mixture was then allowed to cool naturally to room temperature for recrystallization to obtain the product N-([1,1'-biphenyl]-2-ylaminothiocarbonyl)-3,5-dinitrobenzamide, i.e., TU-BDN. The molecular formula of TU-BDN is C2. 20 H 14 N4O5S, chemical structural formula is:

[0039] .

[0040] Example 1: Study on the toxicity of TU-BDN to Vero cells

[0041] 1. Experimental Procedure

[0042] Cytotoxicity assay: The cytotoxicity of TU-BDN to Vero cells was determined using the CCK-8 assay.

[0043] African green monkey kidney (Vero) cell suspension was seeded into 96-well cell culture plates, approximately 100 μL (5000 cells) per well. The culture plates were then pre-cultured in an incubator for 18 h (37℃, 5% CO2, saturated humidity) to allow the cells to adhere.

[0044] The study included a blank control group, a control group, and an experimental group. The blank control group was not seeded with cells, but only received complete culture medium containing 0.1% DMSO. Both the control and experimental groups were seeded with Vero cells, with the control group receiving complete culture medium containing 0.1% DMSO. The experimental groups had nine concentration gradients of TU-BDN: 2 μM, 4 μM, 8 μM, 16 μM, 31.25 μM, 62.5 μM, 125 μM, 250 μM, and 500 μM. The final DMSO concentration for all three groups was 0.1%. The original culture medium in each well was gently aspirated, and the corresponding culture medium or drug-containing medium was added to each group, with a final volume of 100 μL per well. Each group was divided into three replicates.

[0045] The culture plates were incubated at 37℃, 5% CO2, and saturated humidity for 24 h. After incubation, 10 μL of CCK-8 reagent was added to each well, and the plates were incubated in the dark for 2 h. The absorbance (OD) at 450 nm was measured using a microplate reader. Cell viability was calculated using the following formula. The independent experiments were repeated three times, and the half-maximal cytotoxic concentration (MCC) was obtained by fitting a dose-response curve. 50 .

[0046] Cell viability (%) = (OD) 实验组 -OD 空白组 ) / (OD 对照组 -OD 空白组） ×100%

[0047] 2. Experimental Results

[0048] like Figure 1 The graph shows the toxicity test results of TU-BDN on Vero cells. The vertical axis represents cell viability, with the 0.1% DMSO control group as 100% baseline. The horizontal axis represents the administered concentration of compound TU-BDN. As the concentration of TU-BDN increases, cell viability decreases in a dose-dependent manner. When the concentration reaches 105.8 μM, the cell viability drops to 50%, which is the half-maximal cytotoxic concentration (MCC) of compound TU-BDN on Vero cells. 50 The concentration was 105.8 μM, indicating that TU-BDN has advantages such as low toxicity and safety to host cells within the effective concentration range.

[0049] Example 2: Inhibitory effect of TU-BDN on Toxoplasma gondii

[0050] 1. Experimental Procedure

[0051] This experiment included a control group (DMSO group) and an experimental group (TU-BDN treatment group), both inoculated with Vero cells and Toxoplasma gondii tachyzoites. Specific procedures included:

[0052] African green monkey kidney (Vero) cell suspension was seeded into 96-well cell culture plates, approximately 100 μL (5000 cells) per well. The culture plates were then pre-cultured in an incubator for 18 h (37℃, 5% CO2, saturated humidity) to allow the cells to adhere.

[0053] Fresh, viable Luciferase tachyzoites were harvested from Vero cells, counted using a hemocytometer, and 3 × 10⁻⁶ cells were collected. 5 One tachyzoite was inoculated into Vero cells (multiple of infection MOI = 60:1) and incubated at 37°C, 5% CO2, and saturated humidity for 2 hours to allow the tachyzoites to complete their invasion. The original culture medium in the wells was gently aspirated and the cells were gently washed twice with sterile PBS to remove any uninvaded free tachyzoites.

[0054] The control group was given complete culture medium containing 0.1% DMSO; the experimental group was prepared with nine concentration gradients of compound TU-BDN: 0.1 μM, 0.3 μM, 1 μM, 3 μM, 10 μM, 30 μM, 50 μM, 75 μM, and 100 μM. The final concentration of DMSO in both the control and experimental groups was 0.1%, with a final volume of 200 μL per well and three replicates per group.

[0055] After incubating the culture plates at 37℃, 5% CO2, and saturated humidity for 24 h, the culture medium was gently aspirated from each well, and 100 μL of cell lysis buffer was added to each well. After lysis at room temperature for 5 min, the lysis buffer was aspirated and transferred to a 1 mL EP tube. The plates were centrifuged at 12000 r / min for 5 min, and 100 μL of the supernatant was added to a 96-well microplate. Then, under dark conditions, 100 μL of luciferase substrate working solution was added to each well, and the mixture was stirred. The luminescence intensity (RLU) was measured using a chemiluminescence analyzer. The inhibition rate of Toxoplasma gondii proliferation by compound TU-BDN was calculated using the following formula. The independent experiments were repeated three times, and the half-maximal effective concentration (EC50) was obtained by fitting a dose-response curve. 50 .

[0056] Inhibition rate % = (RLU) DMSO -RLU 处理组 ) / RLU DMSO ×100%.

[0057] 2. Experimental Results

[0058] Figure 2 The graph shows the inhibitory effect of TU-BDN on Toxoplasma gondii, with the vertical axis representing the Toxoplasma gondii proliferation inhibition rate and the horizontal axis representing the administered concentration of compound TU-BDN. The results show that TU-BDN has a half-maximal effective concentration (EC50) against Toxoplasma gondii proliferation. 50 The concentration of TU-BDN was 3.231 μM, and its inhibitory effect on Toxoplasma gondii was dose-dependent, with higher concentrations showing stronger inhibitory effects, indicating that TU-BDN has a good inhibitory effect on Toxoplasma gondii. Meanwhile, the safety index SI (SI=CC) was [not specified in the original text]. 50 / EC 50 The concentration of TU-BDN reached 32.75, indicating that TU-BDN has strong selectivity for parasites and low host toxicity, and has excellent potential for the development of anti-Toxoplasma gondii drugs.

[0059] Example 3: The effect of TU-BDN on inhibiting intracellular Toxoplasma gondii plaque formation.

[0060] 1. Experimental Procedure

[0061] Human foreskin fibroblasts (HFFs), a classic cell model for Toxoplasma gondii research, were seeded into 12-well cell culture plates and cultured at 37°C, 5% CO2, and saturated humidity until the cells were 100% confluent. The cells were then cultured statically for another 24 hours to form a monolayer of cells with complete contact inhibition.

[0062] Tachyzoites of freshly lysed HFF monolayers of Toxoplasma gondii type I (RH) were collected, and 150 live tachyzoites were inoculated into each well of HFF monolayers (MOI approximately 1:600). The culture plates were divided into control and experimental groups, with 6 replicates in each group. The control group was given complete medium containing 0.1% DMSO, while the experimental group was given complete medium containing 15 μM compound TU-BDN and 0.1% DMSO. The final volume of medium in each well was fixed at 1 mL, and the final concentration of DMSO in both groups was 0.1%.

[0063] After incubating the culture plate at 37℃, 5% CO2, and saturated humidity for 7 days, the culture medium in the wells was gently aspirated, and the plate was washed twice with sterile PBS. 1 mL of 4% paraformaldehyde was added to each well, and the plate was fixed at room temperature for 30 min. The fixative was then aspirated, and the plate was washed three times with PBS. 1 mL of 0.5% crystal violet staining solution was added to each well, and the plate was stained at room temperature for 20 min. The staining solution was then aspirated, and the plate was gently washed with sterile water until no excess staining solution remained. After air drying, the plaque morphology and plaque area were recorded by microscopic photography.

[0064] 2. Experimental Results

[0065] like Figure 3 Under the same magnification microscope, larger plaques were observed in the DMSO negative control group, which were significantly different from the plaques formed in the TU-BDN group, indicating that TU-BDN can significantly inhibit the growth of Toxoplasma gondii in host cells and plaque formation.

[0066] Example 4: Antiproliferative effect of TU-BDN on intracellular Toxoplasma gondii type I strain (RH)

[0067] 1. Experimental Procedure

[0068] Proliferation experiment: Collect 1×10 5 One RH tachyzoite was inoculated into a 12-well plate covered with a monolayer of HFF cells. After 4 hours of parasite invasion, the culture medium was changed and the cells were divided into two groups: the control group was given complete medium containing 0.1% DMSO, and the experimental group was given complete medium containing 15 μM (5 times EC). 50 The compound TU-BDN and 0.1% DMSO were used in complete culture medium, with the final concentration of DMSO in both groups being 0.1%.

[0069] After culturing at 37℃, 5% CO2, and saturated humidity for 24 h, the original culture medium was gently aspirated and the cells were gently washed twice with sterile PBS. 1 mL of 4% paraformaldehyde was added to each well and the cells were fixed at room temperature for 15 min. The fixative was aspirated, the cells were washed three times with PBS, and 0.1% Triton X-100 was added for permeation at room temperature for 10 min. The permeation solution was aspirated, the cells were washed three times with PBS, and 1% BSA / PBS was added for blocking at room temperature for 30 min.

[0070] After blocking, rabbit-derived anti-Toxoplasma gondii surface protein GAP45 primary antibody (1:300 diluted in 1% BSA / PBS) was added and incubated overnight at 4°C. The next day, the primary antibody was discarded, the cells were washed three times with PBS, and FITC-labeled goat anti-rabbit fluorescent secondary antibody (1:100 diluted in 1% BSA / PBS) was added and incubated at room temperature in the dark for 1 h. The secondary antibody was discarded, the cells were washed three times with PBS, and DAPI working solution (1 μg / mL) was added and stained at room temperature in the dark for 5 min to label the host cell nuclei.

[0071] Observation under a fluorescence microscope: multiple fields of view were randomly selected, covering the upper, lower, left, right, and center positions, and the number of Toxoplasma gondii tachyzoites in 100 vacuoles with worms was counted. The independent experiment was repeated 3 times.

[0072] Invasion experiment: Invasion was performed using 12-well culture plates filled with HFF cells, and 1×10⁶ cells with uniform viability were collected. 5 One RH tachyzoite was resuspended in complete medium containing 0.1% DMSO (control group) and complete medium containing 15 μM of compound TU-BDN and 0.1% DMSO (experimental group), respectively, and added to the corresponding wells simultaneously. The final concentration of DMSO in both groups was 0.1%.

[0073] After culturing in an incubator at 37℃, 5% CO2, and saturated humidity for 30 min, immediately remove the culture medium and gently wash three times with sterile PBS to thoroughly remove any uninvaded tachyzoites (observe under a microscope to ensure all extracellular tachyzoites are clean). Replace with fresh complete culture medium and continue culturing for 20 h. Then, proceed with the fixation, permeabilization, blocking, and IFA staining steps described above for the proliferation experiment (nucleus staining with primary antibody GAP45, secondary antibody FITC, and DAPI).

[0074] During fluorescence microscopy, 8-10 fields of view were randomly selected (covering the upper, lower, left, right, and center positions) to acquire images of Toxoplasma gondii tachyzoites and host cell nuclei.

[0075] The number of vacuoles containing parasites and the number of host cells containing tachyzoites in each image were counted. The antiproliferative effects of TU-BDN expression and the control group on intracellular RH tachyzoites were analyzed. Figure 4 As shown.

[0076] The Toxoplasma gondii invasion rate was calculated as the number of vacuoles carrying the parasite divided by the number of host cells. The average value of all images was taken as the invasion rate for that group. The anti-invasion effects of TU-BDN and the control group against extracellular RH-type Toxoplasma gondii were investigated. Figure 5 As shown.

[0077] 2. Experimental Results

[0078] After Toxoplasma gondii invades a host cell, it forms vacuolar vesicles within which tachyzoites proliferate via endogenous binary fission. Therefore, the number of tachyzoites in these vacuolar vesicles within the same timeframe after cell invasion reflects the proliferative capacity of the parasite. Statistical results from proliferation experiments showed that, compared to the control group (containing 0.1% DMSO), the experimental group containing 15 μM TU-BDN had a significantly lower number of tachyzoites within the vacuolar vesicles. Invasion experiments showed that the invasion rate of the TU-BDN experimental group was significantly lower than that of the control group. (The text then abruptly shifts to a discussion of TU-BDN and its associated CC...) 50 =105.8 μM, EC 50 With a concentration of 3.231 μM, a safety index (SI) of 32.75, and a dosage of 15 μM within the safe range of no significant toxicity to host cells, TU-BDN can significantly inhibit the invasion and intracellular proliferation of RH strain Toxoplasma gondii, further verifying its low-toxicity and highly effective anti-Toxoplasma gondii activity.

[0079] The foregoing description of specific exemplary embodiments of the invention is for illustrative and explanatory purposes. These descriptions are not intended to limit the invention to the precise forms disclosed, and it will be apparent that many changes and variations can be made in accordance with the foregoing teachings. The exemplary embodiments were chosen and described in order to explain the specific principles of the invention and its practical application, thereby enabling those skilled in the art to implement and utilize various different exemplary embodiments of the invention, as well as various different choices and variations. The scope of the invention is intended to be defined by the claims and their equivalents.

Claims

1. A compound TU-BDN, characterized in that, The molecular formula is C 20 H 14 N4O5S, chemical structural formula is: 。 2. The method for preparing the compound TU-BDN as described in claim 1, characterized in that, Includes the following steps: Add 3,5-dinitrobenzoyl chloride dropwise to a dry acetone solution of potassium thiocyanate. After the addition is complete, heat to reflux under nitrogen protection and maintain reflux for 1.5-2.5 h. After cooling the resulting reaction solution to room temperature, add 2-aminobiphenyl, heat to reflux again, and maintain reflux for 8-10 h. After cooling to room temperature, pour into crushed ice, allow to stand to precipitate, filter, and dry to obtain crude product. Recrystallize the crude product from ethanol to obtain TU-BDN.

3. The method for preparing the compound TU-BDN as described in claim 2, characterized in that, The molar ratio of potassium thiocyanate to 3,5-dinitrobenzoyl chloride is 6:(3-5); 9.1 mL of dry acetone is used for every 1 mmol of potassium thiocyanate.

4. The method for preparing the compound TU-BDN as described in claim 2, characterized in that, The molar ratio of potassium thiocyanate to 2-aminobiphenyl is 6:(3-5).

5. The use of the compound TU-BDN as described in claim 1 in the preparation of a medicament for the prevention or treatment of toxoplasmosis infection.

6. The use of the compound TU-BDN as described in claim 5 in the preparation of a medicament for the prevention or treatment of toxoplasmosis infection, characterized in that, The drugs mentioned for preventing or treating Toxoplasma gondii infection are drugs that inhibit the ability of Toxoplasma gondii to lyse cells.

7. The use of the compound TU-BDN as described in claim 5 in the preparation of a medicament for the prevention or treatment of toxoplasmosis infection, characterized in that, The drugs mentioned for preventing or treating toxoplasmosis infection are drugs that inhibit the invasion and proliferation of Toxoplasma gondii.

8. The use of the compound TU-BDN as described in claim 5 in the preparation of a medicament for the prevention or treatment of toxoplasmosis infection, characterized in that, The TU-BDN at 3.231 μM showed a 50% inhibitory effect on Toxoplasma gondii.

9. A composition for the prevention or treatment of toxoplasmosis infection, characterized in that, The active ingredient of the composition includes the compound TU-BDN as described in claim 1.

10. The composition for preventing or treating toxoplasmosis infection as described in claim 9, characterized in that, The composition further includes one or more pharmaceutically acceptable carriers, said carriers comprising one or more of the following: diluents, wetting agents, binders, disintegrants, lubricants, flavor and color modifiers, solvents, solubilizers, cosolvents, emulsifiers, antioxidants, metal complexing agents, preservatives, pH adjusters, surfactants, excipients, fillers, and synergists.

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