CN114652716A - Application of dimetridazole in preparation of anti-beta-lactam drug-resistant escherichia coli drugs - Google Patents
Application of dimetridazole in preparation of anti-beta-lactam drug-resistant escherichia coli drugs Download PDFInfo
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- CN114652716A CN114652716A CN202210332764.5A CN202210332764A CN114652716A CN 114652716 A CN114652716 A CN 114652716A CN 202210332764 A CN202210332764 A CN 202210332764A CN 114652716 A CN114652716 A CN 114652716A
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
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- A61K31/00—Medicinal preparations containing organic active ingredients
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- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/54—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one sulfur as the ring hetero atoms, e.g. sulthiame
- A61K31/542—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one sulfur as the ring hetero atoms, e.g. sulthiame ortho- or peri-condensed with heterocyclic ring systems
- A61K31/545—Compounds containing 5-thia-1-azabicyclo [4.2.0] octane ring systems, i.e. compounds containing a ring system of the formula:, e.g. cephalosporins, cefaclor, or cephalexine
- A61K31/546—Compounds containing 5-thia-1-azabicyclo [4.2.0] octane ring systems, i.e. compounds containing a ring system of the formula:, e.g. cephalosporins, cefaclor, or cephalexine containing further heterocyclic rings, e.g. cephalothin
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
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Abstract
The invention relates to the field of biological medicine, in particular to the field of prevention and treatment of animal-derived bacterial diseases. The invention discloses application of dimetridazole in preparation of a beta-lactam drug resistance escherichia coli resistant drug. The invention also provides a pharmaceutical composition for resisting the escherichia coli with the beta-lactam drug resistance, which comprises dimetridazole and cephalosporin antibiotics such as cefotaxime. The dimetridazole and cefotaxime combined application has a synergistic effect, can improve the sensitivity of antibiotics and reduce the using amount of the antibiotics, achieves the aim of better synergistic antibacterial, and simultaneously provides technical support for veterinary clinical treatment of diseases caused by beta-lactam drug-resistant escherichia coli.
Description
Technical Field
The invention relates to the field of biological medicine, in particular to the field of prevention and treatment of animal-derived bacterial diseases.
Background
Escherichia coli is a main pathogenic bacterium causing urinary tract infection and blood infection, and after the Escherichia coli disease is caused, drug therapy is mostly adopted, but due to improper use or abuse of antibacterial drugs, the treatment cost is increased, high drug resistance of human and animals is caused, and the serious threat to public health is caused. In the last 10 years, the detection rate of beta-lactam drug-resistant escherichia coli is continuously increased in the global scope, the mortality rate of infection caused by the beta-lactam drug-resistant escherichia coli is higher, the beta-lactam drug-resistant escherichia coli is separated from livestock, poultry, wastewater and the environment such as chicken, duck, pig and cattle, drug-resistant strains can exist in the environment and water body for a long time, the risk of human infection is increased possibly, and the beta-lactam drug-resistant escherichia coli poses serious threat to global public health. Colistin is the last line of defense for treating gram-negative bacteria, however, there are reports of colistin resistance in recent years, and the treatment for infections caused by beta-lactam resistant bacteria is in a situation of no medicine cure, so that the development of new effective medicines or the search of new pharmaceutical compositions for solving the problem of drug resistance is urgently needed.
Disclosure of Invention
The invention aims to provide the application of dimetridazole in preparing a drug for resisting escherichia coli with beta-lactam drug resistance.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides an application of dimetridazole in preparation of a beta-lactam drug resistance escherichia coli resistant drug.
Preferably, the concentration of the dimetridazole is 2-2000 mug/mL.
The invention also provides a pharmaceutical composition for resisting the beta-lactam drug resistance escherichia coli, which comprises dimetridazole and cephalosporin antibiotics.
Preferably, the cephalosporin antibiotics include cefotaxime.
Preferably, the concentration of the dimetridazole in the pharmaceutical composition is 120-130 mug/mL, and the concentration of the cefotaxime is 60-70 mug/mL.
Preferably, the concentration of dimetridazole in the pharmaceutical composition is 60-65 mug/mL, and the concentration of cefotaxime is 125-130 mug/mL.
Preferably, the concentration of dimetridazole in the pharmaceutical composition is 30-35 mug/mL, and the concentration of cefotaxime is 125-130 mug/mL.
By adopting the technical scheme, the invention has the following beneficial effects:
the dimetridazole and cefotaxime are used together, the combination can improve the sensitivity of antibiotics and reduce the dosage of the antibiotics, and the composition has a synergistic effect. The dimetridazole with the main effect of expelling parasites is used as a synergist of cephalosporin antibiotics, and the dimetridazole has a synergistic effect. The invention provides technical support for the veterinary clinical treatment of diseases caused by beta-lactam drug-resistant escherichia coli.
Drawings
FIG. 1 shows the MIC measurements of dimetridazole and cefotaxime;
FIG. 2 shows the results of the combined use of dimetridazole and cefotaxime.
Detailed Description
The invention provides an application of dimetridazole in preparation of a beta-lactam drug resistance escherichia coli resistant drug.
In the invention, the dimetridazole solution takes a TSA culture medium as a solvent and dimetridazole as a solute; the concentration of the dimetridazole solution is 2-2000 mug/mL, and the preferable concentration is 3.9-2000 mug/mL.
The invention also provides a pharmaceutical composition for resisting the beta-lactam drug resistance escherichia coli.
In the invention, the pharmaceutical composition comprises an anti-beta-lactam drug-resistant escherichia coli pharmaceutical composition.
In the present invention, the cephalosporin antibiotics include cefotaxime.
In a specific embodiment of the invention, the concentration of dimetridazole in the pharmaceutical composition is 120-130 μ g/mL, the concentration of cefotaxime is 60-70 μ g/mL, and more preferably, the concentration of dimetridazole is 125 μ g/mL, and the concentration of cefotaxime is 64 μ g/mL.
In another embodiment of the invention, the concentration of dimetridazole in the pharmaceutical composition is 60-65 μ g/mL, the concentration of cefotaxime is 125-130 μ g/mL, and more preferably, the concentration of dimetridazole is 62.5 μ g/mL, and the concentration of cefotaxime is 128 μ g/mL.
In another specific embodiment of the invention, the concentration of dimetridazole in the pharmaceutical composition is 30-35 μ g/mL, the concentration of cefotaxime is 125-130 μ g/mL, and further preferably, the concentration of dimetridazole is 31.25 μ g/mL, and the concentration of cefotaxime is 128 μ g/mL.
The technical solutions provided by the present invention are described in detail below with reference to examples, but they should not be construed as limiting the scope of the present invention.
Description of the Experimental materials
Experimental strains: escherichia coli BNCC358242, purchased from North Nay organisms.
Preparation of a cefotaxime solution of 1024 μ g/mL: accurately weighing 10.24mg of cefotaxime, adding into 10ml of LTSB culture medium, and shaking to dissolve.
Preparation of a dimetridazole solution of 2000 mug/mL: accurately weighing 20.00mg of dimetridazole, adding the dimetridazole into a mixed solution of 5mLTSB culture medium and 5mLNaOH, and carrying out ultrasonic treatment for 4 hours at the working frequency of 40 KHz.
Example 1: respectively determining Minimum Inhibitory Concentration (MIC) of cefotaxime and dimetridazole
1) Taking out a frozen escherichia coli drug-resistant strain at minus 80 ℃, inoculating a disposable inoculating loop dipped bacterial liquid into a TSA culture medium, carrying out overnight culture at 37 ℃, selecting a single colony to be inoculated into a TSB culture medium, carrying out shake culture at 37 ℃ and 200rpm for 10-16 h, adjusting the turbidity to 0.5 McLeod by using a bacterial turbidimeter to keep the bacterial concentration at 1 x 108CFU/mL。
2) Respectively measuring MIC of cefotaxime and dimetridazole by trace broth dilution method, and diluting the obtained bacterial solution by 100 times to make its concentration be 1 × 106CFU/mL。
3) And (3) selecting one row in a 96-well plate, and sequentially adding 100 mu L of cefotaxime solution and 100 mu L of bacterial liquid diluted according to the multiple proportion into the 1 st to 10 th wells, wherein the concentrations of the cefotaxime solution after the multiple dilution are respectively 1024 mu g/mL, 512 mu g/mL, 256 mu g/mL, 128 mu g/mL, 64 mu g/mL, 32 mu g/mL, 16 mu g/mL, 8 mu g/mL, 4 mu g/mL and 2 mu g/mL, the 11 th well is a positive control group, 200 mu L of bacterial liquid is added, the 12 th well is a negative control group, and 200 mu L of TSB culture medium is added. Culturing at 37 deg.C for 12-16 h, observing result, and determining OD600；
And selecting another row in a 96-well plate, sequentially adding 100 mu L dimetridazole solution and 100 mu L bacterial liquid diluted according to the multiple ratio into the 1 st to 10 th wells, wherein the concentration of the dimetridazole solution after the multiple ratio dilution is respectively 2000 mu g/mL, 1000 mu g/mL, 500 mu g/mL, 250 mu g/mL, 125 mu g/mL, 62.5 mu g/mL, 31.3 mu g/mL, 15.6 mu g/mL, 7.8 mu g/mL and 3.9 mu g/mL, the 11 th well is a positive control group, 200 mu L bacterial liquid is added, the 12 th well is a negative control group, and 200 mu L TSB culture medium is added. Culturing at 37 deg.C for 12-16 h, observing result, and determining OD600。
As a result:
OD measured by cefotaxime solution under different concentrations600As shown in table 1:
TABLE 1 OD of treated bacteria solutions of cefotaxime in different concentrations600
OD measured by dimetridazole solution under different concentrations600As shown in table 2:
TABLE 2 OD of bacteria solution treated with dimetridazole solution of different concentrations600
When two tables are considered together, the MIC values of the available imidazoxime and dimetridazole are respectively as follows: MICCefotaxime＝512μg/mL，MICDimetridazole＝500μg/mL。
Example 2: determination of minimum inhibitory concentration of cefotaxime and dimetridazole
According to the results of the above determination, the minimum inhibitory concentration values of the combination were determined by the checkerboard method in 96-well plates, 50. mu.L of cefotaxime solution was added to each well, the concentrations of the drugs were 1MIC, 1/2MIC, 1/4MIC, 1/8MIC, 1/16MIC, 1/32MIC in this order, 50. mu.L of dimetridazole solution was added, the concentrations of the drugs were 2MIC, 1MIC, 1/2MIC, 1/4MIC, 1/8MIC, 1/16MIC, 1/32MIC in this order, and then 100. mu.L of 1 × 10 MIC was added6CFU/mL bacterial dilution, and set up only containing 200 u L bacterial dilution of the positive control and only containing 200L TSB medium negative control. Culturing at 37 ℃ for 12-16 h, observing the result, and determining OD600And calculating FIC (bacteriostatic concentration fraction index), and evaluating the combined medication result according to the FIC index.
The FIC index is calculated as follows:
if the FIC is less than or equal to 0.5, a synergistic effect is shown; if FIC is more than 0.5 and less than or equal to 1, an additive effect is shown; if 1< FIC <2, then it appears to be an irrelevant effect: if the FIC is more than or equal to 2, antagonism is shown, and the smaller the FIC index is, the stronger the synergistic effect of the two medicines is.
OD measured when the two drugs are used in combination600As shown in table 3:
TABLE 3 OD of bacterial liquid in combination of two drugs600
(1-6 cefotaxime with 1MIC, 1/2MIC, 1/4MIC, 1/8MIC, 1/16MIC and 1/32MIC in sequence; Dimeidazole with 2MIC, 1MIC, 1/2MIC, 1/4MIC, 1/8MIC, 1/16MIC and 1/32MIC in sequence from A to G)
As can be seen from the above examples, the present invention provides the inhibitory effect of dimetridazole and cefotaxime against the drug-resistant Escherichia coli at different concentrations, wherein MIC was foundDimetridazole＝500μg/mL，MICCefotaxime512 μ g/mL. The invention also provides OD for the drug-resistant escherichia coli when the dimetridazole and the cefotaxime are used together600The combination of dimetridazole and cefotaxime is found to have obvious synergistic antibacterial effect on multidrug resistant Escherichia coli isolates.
When the FIC index of the two medicines in combination is calculated, three combinations of FIC is found to be less than 0.5: 1/4MIC dimetridazole is combined with 1/8MIC cefotaxime, and the FIC index is 125/500+64/512 which is 0.375; 1/8MIC dimetridazole is combined with 1/4MIC cefotaxime, and the FIC index is 62.5/500+128/512 which is 0.375; 1/16MIC dimetridazole is combined with 1/4MIC cefotaxime, and the FIC index is 31.25/500+128/512 which is 0.3125. The best antibacterial effect is comprehensively considered, and the best effect is achieved when 1/16MIC dimetridazole and 1/4MIC cefotaxime are used together.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Claims (7)
1. Application of dimetridazole in preparing medicines for resisting beta-lactam drug-resistant Escherichia coli is provided.
2. The use according to claim 1, wherein the concentration of dimetridazole is 2-2000 μ g/mL.
3. The pharmaceutical composition for resisting the beta-lactam drug resistance escherichia coli is characterized by comprising dimetridazole and cephalosporin antibiotics.
4. The pharmaceutical composition of claim 3, wherein the cephalosporin antibiotic comprises cefotaxime.
5. The pharmaceutical composition for resisting the beta-lactam drug resistance Escherichia coli according to claim 4, wherein the concentration of dimetridazole in the pharmaceutical composition is 120-130 μ g/mL, and the concentration of cefotaxime in the pharmaceutical composition is 60-70 μ g/mL.
6. The pharmaceutical composition for resisting the beta-lactam drug resistance Escherichia coli according to claim 4, wherein the concentration of dimetridazole in the pharmaceutical composition is 60-65 μ g/mL, and the concentration of cefotaxime in the pharmaceutical composition is 125-130 μ g/mL.
7. The pharmaceutical composition for resisting the beta-lactam drug resistance Escherichia coli according to claim 4, wherein the concentration of dimetridazole in the pharmaceutical composition is 30-35 μ g/mL, and the concentration of cefotaxime in the pharmaceutical composition is 125-130 μ g/mL.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103127130A (en) * | 2013-03-08 | 2013-06-05 | 海南卫康制药(潜山)有限公司 | Cefotaxime sodium pharmaceutical composition powder injection |
| CN103432162A (en) * | 2013-08-30 | 2013-12-11 | 天津生机集团股份有限公司 | Composition for treating poultry intestinal bacteria and anaerobic bacteria mixing infection and preparation method thereof |
| CN108066338A (en) * | 2017-12-26 | 2018-05-25 | 金华智济药物科技合伙企业（有限合伙） | New antibiotic composition when prevention and treatment aerobic bacteria and anaerobic bacteria mixed infection and preparation method thereof |
| CN108096198A (en) * | 2017-12-26 | 2018-06-01 | 金华智济药物科技合伙企业（有限合伙） | Novel antibacterial pharmaceutical composition when prevention and treatment aerobic bacteria and anaerobic bacteria mixed infection and preparation method thereof |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103127130A (en) * | 2013-03-08 | 2013-06-05 | 海南卫康制药(潜山)有限公司 | Cefotaxime sodium pharmaceutical composition powder injection |
| CN103432162A (en) * | 2013-08-30 | 2013-12-11 | 天津生机集团股份有限公司 | Composition for treating poultry intestinal bacteria and anaerobic bacteria mixing infection and preparation method thereof |
| CN108066338A (en) * | 2017-12-26 | 2018-05-25 | 金华智济药物科技合伙企业（有限合伙） | New antibiotic composition when prevention and treatment aerobic bacteria and anaerobic bacteria mixed infection and preparation method thereof |
| CN108096198A (en) * | 2017-12-26 | 2018-06-01 | 金华智济药物科技合伙企业（有限合伙） | Novel antibacterial pharmaceutical composition when prevention and treatment aerobic bacteria and anaerobic bacteria mixed infection and preparation method thereof |
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