Composition of thiazolidinyl urea compound and polymyxin and application of composition in preparation of salmonella infection resisting medicine

Abstract

The invention belongs to the technical field of biological medicines, and discloses a composition of a thiazolidinyl urea compound and polymyxin and application of the composition to preparation of salmonella infection resisting medicines. According to a sterilization curve obtained after the thiazolidinyl urea compound and the polymyxin are combined for use, the 24-hour bacterium loading amount is reduced by more than 100 times compared with that of a single drug, and a good synergistic sterilization effect is shown when the thiazolidinyl urea compound and the polymyxin are combined for use. The thiazolidinyl urea compound is simple in source and synthesis, the dosage of polymyxin can be remarkably reduced, the possibility that the polymyxin generates drug resistance is reduced under the condition that a better sterilizing effect is achieved, and the thiazolidinyl urea compound has a better prospect in application of the polymyxin combined drug to the salmonella infection resisting drug.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine and discloses a composition of a thiozolidinyl urea compound and polymyxin and application of the composition in preparing a drug for resisting salmonella infection. Background Art

[0002] Salmonella is a Gram-negative bacterium that causes foodborne illness when humans or farmed animals consume food contaminated with Salmonella, which can cause local gastroenteritis to systemic diseases. The drug resistance of Salmonella has become a threat to global public health. With the increasing rate of Salmonella resistance in clinical tests and the limited supply of new antibiotics, polymyxin has become the last line of defense for antibiotic treatment of multidrug-resistant Salmonella.

[0003] Since the 1950s, polymyxins have been used as antibiotics to combat clinical infections caused by Gram-negative bacteria in humans and veterinary settings. However, due to the clinical toxicity of polymyxins, their use rate decreased significantly in the 1980s. However, as the harm of drug-resistant bacteria has intensified, polymyxins have been used again as a last resort for the treatment of multidrug-resistant bacteria. Since 2016, when the plasmid-mediated mobile polymyxin resistance gene (mcr-1) was reported, polymyxin resistance has become an emerging public health threat worldwide. Polymyxins have been discontinued in the veterinary field in my country and have effectively reduced the detection rate of polymyxin-resistant Escherichia coli. However, polymyxins are still very important antibiotics in clinical treatment. The problem of polymyxin resistance based on Enterobacteriaceae is quite important and prominent, so it is urgent to screen effective polymyxin enhancers to treat polymyxin-resistant Enterobacteriaceae infections. By artificially synthesizing natural compounds that have synergistic effects with polymyxins through groups with antibacterial activity, the sensitivity of polymyxin-resistant Enterobacteriaceae to polymyxins can be effectively restored. This is also one of the strategies to effectively explore alternative methods for antibiotics and solve the problem of bacterial resistance. Summary of the invention

[0004] The technical problem to be solved by the present invention is to make up for the deficiencies in the prior art. First, a composition of a thiozolidinyl urea compound and polymyxin is provided. The present invention screens a specific compound in combination with polymyxin and finds that the specific compound can be used as a polymyxin synergist to synergistically promote the bactericidal effect.

[0005] Another object of the present invention is to provide the use of the above composition in preparing anti-Salmonella infection drugs.

[0006] The above technical objectives are achieved through the following solutions:

[0007] A composition, consisting of a thiozolidinyl urea compound and polymyxin; the thiozolidinyl urea compound has the following structural formula:

[0008]

[0009] Preferably, the polymyxin is colistin.

[0010] The present application also found that thiozolidinyl urea compounds can synergistically enhance the effect of polymyxin in resisting Salmonella infection.

[0011] Based on this, the present application also claims to protect the use of thiozolidinyl urea compounds as polymyxin enhancers.

[0012] Through experiments, it was found that the composition composed of thiozolidinyl urea compounds and polymyxin can be used in the preparation of anti-Salmonella infection, and has a significant bactericidal effect, which is stronger than that of a single polymyxin.

[0013] Furthermore, the Salmonella is ATCC14028 and polymyxin-resistant Salmonella.

[0014] Furthermore, when used in combination, the concentration of the thiozolidinyl urea compound is 20-30 mg / L, and the concentration of polymyxin is 32-64 mg / L.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] The present invention finds through the sterilization curve after the thiozolidinyl urea compound is used in combination with polymyxin that the 24h bacterial load is reduced by more than 100 times compared with the single drug, showing that the thiozolidinyl urea compound and polymyxin have a good synergistic sterilization effect when used in combination. The thiozolidinyl urea compound is simple in source and synthesis, can significantly reduce the dosage of polymyxin, and reduces the possibility of polymyxin resistance while achieving a better sterilization effect, and has a good prospect in the application of drugs for the combined use of polymyxin to resist Salmonella infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 Checkerboard heat map and in vitro killing curves of thiozolidinyl urea compounds against Salmonella. DETAILED DESCRIPTION

[0018] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the 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.

[0019] Unless otherwise specified, the experimental methods used in the examples of the present invention are all conventional methods; the materials, reagents, etc. used are reagents and materials that can be obtained from commercial channels unless otherwise specified.

[0020] The polymyxins used in the following examples are all polymyxin E.

[0021] The synthesis of thiozolidinyl urea compounds can be directly referred to patent: CN112047889A, and its structural characterization is as follows:

[0022] 248.6Hz,1C),111.1,49.0,19.5(2C).HRMS(ESI):m / z[M+H] + Calculate for C 11 H 12 F2N2S: 243.0762, found: 243.079.

[0023] Example 1 Synergistic effect of thiozolidinyl urea compounds and polymyxin against Salmonella in vitro

[0024] 1. Cool the autoclaved LPM broth with a pH of 5.8 and set aside. The standard strain of Salmonella ATCC14028 is stored in the laboratory.

[0025] 2. Preparation before the test:

[0026] (1) The standard strain of Salmonella ATCC14028 was inoculated onto an LB agar plate and cultured to an appropriate size.

[0027] (2) Weigh an appropriate amount of thiozolidinyl urea compound and dissolve it in dimethyl sulfoxide to make the final drug concentration 5000 mg / L, vortex mix, filter and set aside. Polymyxin is prepared into a stock solution with a concentration of 5120 mg / L according to CLSI and set aside.

[0028] (3) The standard strain of Salmonella ATCC14028 was inoculated onto an LB agar plate and cultured to an appropriate size.

[0029] 3. MIC and FICI of thiozolidinyl urea compounds and polymyxin against Salmonella

[0030] (1) The standard strain of Salmonella ATCC14028 was inoculated into 4 mL of LB broth and shaken at 37°C and 180 rpm until the logarithmic growth phase;

[0031] (2) Use the prepared LPM broth to dilute the shaken bacteria to 100 times, about 10 6 CFU / mL, reserve;

[0032] (3) Take a sterile 96-well plate, add 180 μL of LPM broth medium to column 1, and add 100 μL of LPM broth medium to columns 2-11;

[0033] (4) Then add 20 μL of drug to the first column, mix by pipetting, and then pipette 100 μL into the second well, and so on, pipette 100 μL into the tenth well and discard;

[0034] (5) Add 100 μL of diluted bacterial solution to columns 1 to 11 and 200 μL of LPM broth to column 12;

[0035] (6) The 96-well plate was placed in a 37°C incubator for 16 h, and the absorbance of the bacterial culture at 600 nm was measured by an ELISA reader. The MIC results are shown in Table 1.

[0036] Table 1

[0037]

[0038] 4. In vitro bactericidal curves of thiozolidinyl urea compounds and polymyxin against Salmonella

[0039] (1) Inoculate the standard strain of Salmonella ATCC14028 and the polymyxin-resistant Salmonella (17ES) in 4 mL LB broth test tubes and shake at 37°C and 180 rpm until the logarithmic growth phase;

[0040] (2) Use the prepared LPM broth to dilute the shaken bacterial solution 10 times to make the bacterial count about 10 7 CFU / mL, reserve;

[0041] (3) Add 25 mg / L thiozolidinyl urea compounds or polymyxin with a certain antibacterial concentration in combination or alone to the test tube containing LPM broth, then add 0.4 mL of the diluted bacterial solution to make the total system 4 mL. Vortex thoroughly to mix, and set up a blank control group as quality control. Except for not adding drugs, the other conditions of the control group are the same as those of the experimental group.

[0042] (4) At 0, 3, 6, 9, and 24 hours of culture, 100 μL of bacterial solution was drawn from each tube and added into a 2 mL centrifuge tube containing 900 μL of 0.85% saline for gradient dilution. 25 μL of the diluted bacterial solution was dropped onto the LB agar plate. The LB agar plate was then placed in a 37°C incubator for 18 hours and counted. The experimental results were statistically analyzed after three biological replicates. The number of colonies at each time point was counted and plotted into a sterilization curve.

[0043] Results: By Figure 1From the in vitro bactericidal curve, it can be seen that the bacterial load after the combination of thiozolidinyl urea compounds and polymyxin was reduced by more than 100 times compared with that of single drug in 24 hours, indicating that the combination of thiozolidinyl urea compounds and polymyxin has a better bactericidal effect.

[0044] Obviously, the above embodiments of the present invention are only examples for clearly illustrating the technical solution of the present invention, and are not intended to limit the specific implementation methods of the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the claims of the present invention shall be included in the protection scope of the claims of the present invention.

Claims

1. A composition, characterized in that It is composed of a thiozolidinyl urea compound and polymyxin; the structural formula of the thiozolidinyl urea compound is as follows:

2. The composition according to claim 1, characterized in that Polymyxin is polymyxin E.

3. The use of thiozolidinyl urea compounds in the preparation of polymyxin synergists, characterized in that: The structural formula of the thiozolidinyl urea compound is as follows:

4. Use of the composition according to claim 1 or 2 in the preparation of a drug for resisting Salmonella infection.

5. The use according to claim 4, characterized in that: The Salmonella is ATCC14028 and polymyxin-resistant Salmonella.

6. The use according to claim 4, characterized in that: The concentration of the thiozolidinyl urea compound is 20-30 mg / L, and the concentration of polymyxin is 32-64 mg / L.

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