CN116747221B - Antibacterial composition and preparation method and application thereof - Google Patents

Abstract

The invention belongs to the field of antibacterial technology, and specifically relates to an antibacterial composition and its preparation method and application. The antibacterial composition of the present invention includes the following components by weight: 8 to 32 parts of antibiotics and 312 to 625 parts of shikimic acid. The antibacterial composition provided by the invention uses the natural Chinese medicine extract shikimic acid combined with antibiotics to inhibit or kill drug-resistant bacteria, which can significantly reduce the dosage of antibiotics, reduce adverse reactions and side effects, and improve the safety of medication. Moreover, shikimic acid and antibiotics can It produces obvious synergistic effect and has a significant inhibitory effect on drug-resistant bacteria such as Staphylococcus aureus or Escherichia coli. And the examples show that the present invention has screened out shikimic acid, a traditional Chinese medicine extract that has a strong antibacterial effect on methicillin-resistant Staphylococcus aureus in vitro. When shikimic acid and ceftiofur sodium are used in combination, the drug can be used as soon as possible. It kills methicillin-resistant Staphylococcus aureus within 2 hours, and the effect is significantly better than that of shikimic acid and ceftiofur sodium alone.

Description

Antibacterial composition and preparation method and application thereof

Technical field

The invention belongs to the field of antibacterial technology, and specifically relates to an antibacterial composition and its preparation method and application.

Background technique

Staphylococcus aureus is a bacterium that causes a series of diseases such as endocarditis, bacteremia, pneumonia, and skin and soft tissue infections. Currently, it is mainly treated with penicillins, cephalosporins, erythromycin, lincomycin, Antibiotic treatment such as clindamycin kills it. However, with the overuse of antibiotics, bacteria have developed various resistance mechanisms under huge selection pressure, and these resistance mechanisms eventually led to the emergence of superbugs.

Methicillin-resistant Staphylococcus aureus (MRSA) is a common multidrug-resistant and highly virulent pathogenic bacteria among Staphylococcus aureus. Since its first discovery in the United Kingdom in 1961, MRSA has spread throughout the world and has become one of the most important pathogens causing community-acquired and healthcare-associated infections. The spread of drug-resistant bacteria not only causes a sharp increase in human mortality, but also threatens economic animals such as cattle, sheep, pigs, and chickens, causing huge economic losses to agricultural production. In the post-antibiotic era, the rate of research and development of antibacterial drugs is far slower than the rate of emergence and spread of bacterial resistance. In this regard, combined drugs have been proposed to combat drug-resistant bacteria.

As a type of natural medicine, traditional Chinese medicine contains a large number of potential antibacterial adjuvants. These compounds have wide distribution, novel structures, and good biological safety, but their direct bactericidal activity is weak and rarely exert pressure on bacteria. Therefore, The frequency of resistance genes in bacterial populations can be controlled to a certain extent. The combined use of traditional Chinese medicine extracts and antibiotics can effectively combat drug-resistant bacteria, but antibiotics still play the main killing role, which can lead to some adverse reactions and side effects, and is not very safe.

Therefore, there is an urgent need for an antibacterial composition that can reduce the dosage of antibiotics or reduce the adverse reactions and side effects caused by antibiotics.

Contents of the invention

In view of this, the object of the present invention is to provide an antibacterial composition that combines shikimic acid, a traditional Chinese medicine extract with the same origin as medicine and food, in combination with antibiotics to inhibit or kill drug-resistant bacteria, which can significantly reduce the dosage of antibiotics and reduce adverse reactions and side effects. It improves the safety of medication, and shikimic acid can produce obvious synergistic effects with antibiotics, and has a significant inhibitory effect on drug-resistant bacteria such as Staphylococcus aureus or Escherichia coli.

In order to achieve the above objects, the present invention provides the following technical solutions:

The invention provides an antibacterial composition, which includes the following components by weight: 8 to 32 parts of antibiotics and 312 to 625 parts of shikimic acid.

Preferably, the antibiotics include β-lactam antibiotics.

Preferably, the β-lactam antibiotic includes penicillin, ampicillin, amoxicillin or ceftiofur.

Preferably, the ceftiofur includes ceftiofur sodium.

The present invention provides a method for preparing the antibacterial composition described in the above technical solution, which includes: mixing antibiotics and shikimic acid to obtain the antibacterial composition.

The present invention also provides the application of the antibacterial composition described in the above technical solution in the preparation of antibacterial drugs.

Preferably, the bacteria include drug-resistant bacteria.

Preferably, the drug-resistant bacteria include Staphylococcus aureus and/or Escherichia coli.

Preferably, the Staphylococcus aureus includes methicillin-resistant Staphylococcus aureus.

The present invention also provides an antibacterial drug, the active ingredients of which include the antibacterial composition described in the above technical solution.

Beneficial effects:

The invention provides an antibacterial composition, including antibiotics and shikimic acid. As a natural traditional Chinese medicine extract, shikimic acid can produce significant synergistic effects when combined with antibiotics, reduce the dosage of antibiotics, reduce adverse reactions and side effects, and improve It is safe to use and can have a significant inhibitory effect on drug-resistant bacteria such as Staphylococcus aureus or Escherichia coli. According to the records of the examples, the present invention starts from the extracts of traditional Chinese medicine homologous to food and medicine, and through the preparation of growth curves and broth micro-dilution checkerboard method tests, it is screened out for the strong antibacterial effect on methicillin-resistant Staphylococcus aureus in vitro. The traditional Chinese medicine extract monomer - shikimic acid, when combined with ceftiofur sodium, can kill methicillin-resistant Staphylococcus aureus (MRSA) within 2 hours at the fastest, and the effect is significantly better than shikimic acid and ceftiofur sodium are administered separately.

The antibacterial composition provided by the invention can be used to prepare drugs with antibacterial effects, and is expected to provide a reference for the development of new auxiliary drugs for ceftiofur sodium to treat severe infectious diseases, improve its therapeutic effectiveness, and solve the increasingly serious problem of ceftiofur sodium. The problem of drug resistance provides new research ideas and R&D directions.

Description of the drawings

In order to explain the embodiments of the present invention or the technical solutions in the prior art more clearly, the drawings needed to be used in the embodiments will be briefly introduced below.

Figure 1 is a bactericidal curve diagram of the combination of 1/4MIC ceftiofur sodium and SA at different concentrations in Example 1;

Figure 2 is a bactericidal curve diagram of the combination of 1/8MIC ceftiofur sodium and SA at different concentrations in Example 1;

Figure 3 is a graph showing the effects of different concentrations of SA on cell survival rate in Example 2;

Figure 4 is a graph showing the effects of different treatments on the survival rate of the MRSA-infected mouse model in Example 3;

Figure 5 is a graph showing the results of the bacterial load in the liver tissue of the MRSA-infected mouse model after different treatments at 24h, 48h and 72h in Example 4;

Figure 6 is a graph showing the results of the spleen tissue bacterial load of the MRSA infected mouse model after different treatments at 24h, 48h and 72h in Example 4;

Figure 7 is a graph showing the results of bacterial load in the kidney tissue of the MRSA mouse model after different treatments at 24h, 48h and 72h in Example 4;

Figure 8 is a graph showing the survival of mice after administration of SA doses of 150, 100 and 75 mg/kg mouse body weight respectively in Example 5;

Figure 9 is a diagram of the liver tissue sections of the MRSA-infected mouse model after different treatments in Example 6, the control group (upper left), the MRSA injection group (upper right), the CF injection group (lower left), and the CF+SA injection group (lower right);

Figure 10 is a diagram of the spleen tissue section of the MRSA-infected mouse model after different treatments in Example 7, the control group (upper left), the MRSA injection group (upper right), the CF injection group (lower left), and the CF+SA injection group (lower right);

Figure 11 shows kidney tissue sections of the MRSA-infected mouse model after different treatments in Example 8, including the control group (upper left), MRSA injection group (upper right), CF injection group (lower left), and CF+SA injection group (lower right).

Detailed ways

The invention provides an antibacterial composition, which includes the following components by weight: 8 to 32 parts of antibiotics and 312 to 625 parts of shikimic acid.

In parts by weight, the antibacterial composition of the present invention includes 8 to 32 parts of antibiotics, preferably 8 to 16 parts, and more preferably 8 parts. The antibiotics of the present invention preferably include β-lactam antibiotics; the β-lactam antibiotics preferably include penicillin, ampicillin, amoxicillin or ceftiofur, more preferably ceftiofur, more preferably ceftiofur sodium. The present invention has no special restrictions on the source of the antibiotics, and conventional commercially available products can be used.

Based on the weight parts of the antibiotics, the antibacterial composition of the present invention includes 312 to 625 parts of shikimic acid, preferably 400 to 625 parts, more preferably 500 to 625 parts, and more preferably 625 parts. The shikimic acid of the present invention is preferably star anise extract shikimic acid. The shikimic acid of the present invention improves the antibacterial efficacy of β-lactam antibiotics and plays a synergistic effect. The present invention has no special limitation on the source of the star anise extract shikimic acid, and it can be either conventional commercially available products or prepared by yourself. As in a specific embodiment of the present invention, the shikimic acid was purchased from Sichuan Hengrui Tongda Biotechnology Co., Ltd., and the product number is 04020149.

The antibacterial composition provided by the invention uses shikimic acid combined with antibiotics, which can produce significant synergistic effects, reduce the dosage of antibiotics, reduce adverse reactions and side effects, improve the safety of medication, and is resistant to Staphylococcus aureus or Escherichia coli. Bacteria can produce significant inhibitory effects.

The present invention provides a method for preparing the antibacterial composition described in the above technical solution, which includes: mixing the antibiotic and shikimic acid to obtain the antibacterial composition. The present invention has no special limitation on the mixing method, and it is enough to use conventional mixing methods in this field to mix evenly.

The present invention also provides the application of the antibacterial composition described in the above technical solution in the preparation of antibacterial drugs. In the present invention, the bacteria preferably include drug-resistant bacteria; the drug-resistant bacteria in the invention preferably include Staphylococcus aureus and/or Escherichia coli, further preferably Staphylococcus aureus, and more preferably methicillin-resistant aureus staphylococcus. The methicillin-resistant Staphylococcus aureus of the present invention is preferably the methicillin-resistant Staphylococcus aureus standard bacterium ATCC33591.

The present invention also provides an antibacterial drug, the active ingredients of which include the antibacterial composition described in the above technical solution.

The antibacterial composition provided by the invention can be used to prepare drugs with antibacterial effects, and is expected to provide a reference for the development of new auxiliary drugs for the treatment of severe infectious diseases with ceftiofur sodium, improve its therapeutic benefits, and solve the increasingly serious resistance to ceftiofur sodium. Drug issues provide new research ideas and R&D directions.

In order to further illustrate the present invention, the antibacterial composition provided by the present invention is described in detail below in conjunction with the accompanying drawings and examples, but they should not be understood as limiting the protection scope of the present invention.

Example 1

Antibacterial effects of shikimic acid SA and β-lactam antibiotics on MRSA standard strain ATCC33591

Determination of the minimum inhibitory concentration (MIC) of a single drug

Taking the MRSA standard strain ATCC33591 as the research object, the MIC value of each single drug was determined using the broth microdilution method recommended by the American Clinical Laboratory Standards Committee (CLSI).

Preparation of bacterial suspension: Inoculate the recovered ATCC33591 with streaks on brain heart infusion agar. After culturing at 37°C for 24 hours, single clone colonies were picked out in MH broth and cultured with shaking at 37°C for 8 hours. Use blank MH to dilute the cultured bacterial liquid until the turbidity reaches 0.5 McFarland specific turbidity (bacterial liquid concentration is about 10 ⁸ CFU/mL), and then dilute the bacterial liquid 1000 times to serve as the test bacterial liquid, with a final concentration of 10 ⁵ CFU /mL.

Broth microdilution method detection: Dilute the stock solution of the test drugs (shikimic acid SA, penicillin, ampicillin, amoxicillin and ceftiofur sodium) to a certain high concentration, and dilute it to 5 mg/mL and 2.5 mg/mL. mL, 1.25mg/mL, 0.625mg/mL, 0.3125mg/mL and so on for 10 concentrations (well 1 to 10), 100 μL per well, and add the above bacterial suspension into wells 1-10 (for To prevent high-concentration drugs from contaminating low-concentration drugs, the order of adding samples is from low concentration to high concentration). Two sets of controls are set up in each experiment: the 11th hole is the test bacterial liquid control, which is the positive control, and the 12th hole is the broth blank control, which is the negative control. After culturing for 24 hours at a constant temperature of 37°C, the results were observed.

Result judgment: The minimum concentration at which no bacterial growth is observed with the naked eye is the MIC of the drug. If the positive control has bacterial growth and the negative control is still clear, the result can be judged to be valid. Otherwise, it is invalid and must be repeated. Make two parallel rows of each medicine and repeat twice. The MIC test results of shikimic acid (SA), penicillin (Penicillin), ampicillin (Ampicillin), amoxicillin (Amoxicillin) and ceftiofur sodium (CF) are shown in Table 1.

Combination drug experiment

The combined drug susceptibility test was conducted using the checkerboard method. Based on the MIC test results of shikimate SA and each β-lactam antibiotic single drug, the broth dilution checkerboard method was used to design the two drugs at 2 times, 1 times, 1/2 times, and 1 /4 times, 1/8 times and 1/16 times MIC for 6×6 combination. Shikimic acid and each antibiotic were diluted with MH broth to a specific drug concentration and added to each well of a 96-well plate. In addition, the individual MIC of two single drugs, the negative (only broth was added) and the positive well (only broth and MRSA standard strain ATCC33591 bacterial solution were added) were made for control. Prepare the bacterial suspension according to the previous method for preparing the bacterial suspension, add it to each well of the 96-well plate, and place it in a 37°C constant temperature incubator. Observe and record the results after 24 hours to obtain shikimic acid SA and each The respective combined MICs of β-lactam antibiotics under the combined effect are calculated according to the formula FICI = (MIC of drug A in combination/MIC of drug A alone) + (MIC of drug B in combination/MIC of drug B alone) Calculate FICI. Staphylococcus aureus ATCC29213 was used as the quality control strain in each experiment. The results are shown in Table 1.

Table 1 The anti-MRSA effect of SA combined with β-lactam antibiotics

Note: FICI≤0.5 is synergy; 0.5<FICI<1 is additive; 1<FICI<2 is irrelevant effect; FICI>2 is antagonism.

As can be seen from Table 1, after measurement, the minimum inhibitory concentration of shikimic acid SA against the MRSA standard strain ATCC33591 is 5 mg/mL. In addition, SA also exhibits synergistic effects with other β-lactam antibiotics.

Bacterial growth curve under the combined effect of SA and ceftiofur sodium

Preparation of antibacterial drugs: Taking the MRSA standard strain ATCC33591 as the research object, based on the single drug minimum inhibitory concentration (MIC) test results, the ceftiofur sodium and shikimic acid stock solutions were diluted with MH broth to the following concentrations: ceftiofur Ceftiofur sodium 1/4MIC, ceftiofur sodium 1/8MIC, shikimic acid 1/8MIC, shikimic acid 1/16MIC, ceftiofur sodium 1/4MIC+shikimic acid 1/8MIC, ceftiofur sodium 1/4MIC+shikimic acid 1/ 16MIC, ceftiofur sodium shikimic acid 1/8MIC+shikimic acid 1/8MIC, ceftiofur sodium 1/8MIC+shikimic acid 1/16MIC.

Preparation of bacterial suspension: Use a sterile rod to pick a single colony of the MRSA standard strain ATCC33591, inoculate it into a McFarland turbidity tube containing sterile PBS, and use a McFarland turbidimeter to adjust the turbidity to 0.5 McFarland turbidity. The bacterial concentration is approximately 10 ⁸ CFU/mL; then dilute the bacterial liquid 1000 times to serve as the test bacterial liquid, with a final concentration of 10 ⁵ CFU/mL.

Growth curve drawing: Set up three parallel groups of the above antibacterial drug combinations and add them to a sterile 96-well plate respectively. The volume of each group in the 96-well plate is 100 μL. Finally, add 100 μL of the above bacterial suspension to each well, cover the 96-well plate, and place it in a microplate reader with a constant temperature of 37°C for culture. Within 0-24 hours, measure the OD value of the bacterial solution at a wavelength of 600 nm on a microplate reader every hour (24 time points). Plot the bacterial growth curve between the obtained OD ₆₀₀ value and the corresponding time. The results are shown in Figures 1 and 2.

The results in Figure 1 show that the use of 1/8 MIC SA alone has almost no antibacterial effect; the combined use of 1/4 MIC ceftiofur sodium and 1/16 MIC SA significantly reduced the number of Staphylococcus aureus within the first 12 hours, and The effect continues after 18 hours; and treatment with 1/4MIC of ceftiofur sodium and 1/8MIC of SA can completely kill Staphylococcus aureus ATCC33591 within 2 hours.

The results in Figure 2 show that the use of 1/8 MIC SA alone has almost no antibacterial effect; the combined use of 1/8 MIC ceftiofur sodium and 1/16 MIC SA significantly reduced the number of Staphylococcus aureus within the first 8 hours, and The effect continues after 10 hours; and treatment with 1/8 MIC of SA and 1/8 MIC of ceftiofur sodium can completely kill Staphylococcus aureus ATCC33591 within 2 hours.

Example 2

Toxicity experiment of SA alone on bovine mammary epithelial cells (BMECs)

Experimental purpose: to verify the toxicity of SA to BMECs and determine the in vivo safety of the drug.

experimental method:

(1) Inoculate BMECs into a 96-well plate at a density of 100,000 cells/mL, 100 μL/well, and culture for 24 hours at 37°C in a 5% CO ₂ incubator.

(2) When the cells grow to about 80%, divide the 96 wells into 8 groups, and add 2.5 mg/mL, 1.875 mg/mL, 1.25 mg/mL, 0.625 mg/mL, 0.313 mg/mL, and 0.156 mg respectively. /mL, 0.078mg/mL, 0mg/mL SA culture medium (culture medium composition: 89% 1640, 10% fetal bovine serum, 1% penicillin-streptomycin solution), 100μL/well, culture for 24h.

(3) Add 10 μL of CCK-8 reagent to each well and incubate for 1 to 4 hours. 450nm detection wavelength, 600nm reference wavelength.

(4) Each group has 6 replicate wells and a blank control group (no cells, only drugs and CCK-8)

The calculation formula for cell survival rate is as follows:

Cell viability (%) = (OD _Drug - OD _Blank )/(OD _Control - OD _Blank ) × 100%.

OD _Drug : drug (SA) group, OD _Blank : blank control group (plus SA and CCK-8), OD _Control : no drug (SA) group; the results are shown in Table 2 and Figure 3.

Table 2 Cell survival rate results under different concentrations of SA

As can be seen from Table 2 and Figure 3, compared with the unmedicated control group after each concentration of SA treatment, SA ≤ 0.625 mg/mL showed no significant effect on cell viability.

Example 3

MRSA high concentration lethal model

After the mice were adaptively fed for one week, 50 BALB/c female mice aged 6 to 8 weeks were randomly divided into 5 groups with 10 mice in each group. Group 1 was the blank control group, and each mouse in the other four groups was intraperitoneally injected with 0.5 mL of MRSA bacterial suspension (final concentration: 7×10 ⁸ CFUs/mouse). 2 hours after bacterial infection, the mice in the blank control group were injected with 300 μL of PBS buffer (pH value of 7.4), and the mice in the other four groups were injected intraperitoneally with 300 μL of PBS buffer (pH value of 7.4), which was recorded as the MRSA group; 300 μL SA (injected at 50 mg/kg according to the mouse body weight) was recorded as the SA group; 300 μL of ceftiofur sodium (injected at 5 mg/kg according to the mouse body weight) was recorded as the CF group; 300 μL of SA+ceftiofur sodium (based on the mouse body weight) The body weight of mice was injected with 50 mg/kg + 5 mg/kg), which was recorded as the SA + CF group. After antibiotic treatment, the survival rate of each group of mice within 96 hours was calculated. The results are shown in Table 2 and Figure 4.

Table 2 Survival rate of MRSA mouse model after different treatments (%)

As can be seen from Table 2 and Figure 4, within 48 hours after infection, the mortality rate of mice was extremely high, and the mortality rate in each group of the MRSA group (without drug addition), SA group, and ceftiofur sodium group reached 20% to 50%. , while the combination group had a 100% survival rate, which was significantly higher than other groups, indicating that the combination of SA and ceftiofur sodium can significantly improve the survival rate of bacteremia-infected mice.

Example 4

MRSA low concentration bacteremia model

After the mice were adaptively fed for one week, 42 BALB/c female mice aged 6 to 8 weeks were randomly divided into 6 groups with 7 mice in each group. Each mouse was injected intraperitoneally with 0.5 mL of MRSA bacterial suspension (final concentration: 1×10 ⁸ CFUs/mouse) to establish a low-concentration MRSA bacteremia model. 2 hours after bacterial infection, mice in the six groups were intraperitoneally injected with 300 μL of PBS buffer (pH value 7.4), which was recorded as the MRAS group; 300 μL of SA (50 mg/kg injected according to the mouse body weight), which was recorded as the SA group; 300 μL Ceftiofur sodium (5 mg/kg injected according to the mouse body weight) was recorded as the CF group; 300 μL of SA+ceftiofur sodium (50 mg/kg+5 mg/kg injected according to the mouse body weight) was recorded as the CF+SA1 group; 300 μL of SA + ceftiofur sodium (injection of 75 mg/kg + 5 mg/kg based on mouse body weight), recorded as CF + SA2 group; 300 μL of SA + ceftiofur sodium (injection of 100 mg/kg + 5 mg/kg based on mouse body weight) , recorded as CF+SA3 group. At the 24h, 48h, and 72h time points, the liver, spleen, and kidney tissues of the mice in each group were collected and counted using plate colony technology. The specific steps were: The surviving mice in each group were killed at the 24h, 48h, and 72h time points, and the remaining Weigh the liver, spleen, and kidney tissues and organs, homogenize them, and then dilute them 10 times with PBS into a series of concentrations: 10 ^-1 , 10 ^-2 , 10 ^-3, etc., select 100 μL of PBS solution at the appropriate dilution concentration , spread it evenly on the MH agar plate with a sterile rod, incubate at 37°C for 24 hours and observe the results. Select the plate with 30 to 300 colonies for colony counting. The results are shown in Tables 3 to 5 and Figures 5 to 7 .

Table 3 Colony count results (CFU) of liver tissue at 24h, 48h, and 72h time points

Table 4 Colony count results (CFU) of spleen tissue at 24h, 48h, and 72h time points

MRSA CF SA CF+SA1 CF+SA2 CF+SA3 24 hours 1.2×10 ⁷ 9.1×10 ⁵ 7.5×10 ⁵ 6.0×10 ⁵ 6.9×10 ⁵ 4.4×10 ⁴ 48h 2.2×10 ⁶ 2.9×10 ⁵ 2.5×10 ⁴ 2.8×10 ⁴ 1.5×10 ⁴ 4.6×10 ³ 72h 4.0×10 ⁵ 5.7×10 ⁴ 7.0×10 ⁴ 2.1×10 ⁴ 8.4×10 ³ 6.7×10 ³

Table 5 Colony count results (CFU) of spleen tissue at 24h, 48h, and 72h time points

MRSA CF SA CF+SA1 CF+SA2 CF+SA3 24 hours 1.6×10 ⁷ 6.6×10 ⁵ 3.6×10 ⁶ 1.0×10 ⁵ 8.3×10 ⁴ 2.0×10 ⁴ 48h 3.0×10 ⁷ 1.2×10 ⁶ 2.9×10 ⁶ 1.5×10 ⁵ 2.2×10 ⁴ 2.2×10 ⁴ 72h 4.3×10 ⁵ 6.7×10 ⁴ 7.0×10 ⁴ 1.0×10 ⁴ 1.7×10 ⁴ 1.0×10 ⁴

It can be seen from Tables 2 to 4 and Figures 5 to 7 that the combination group can significantly reduce the bacterial load in liver, spleen and kidney tissues compared with the MRSA group, and has a good therapeutic effect in a dose-dependent manner. The combined group was generally better than the SA group and ceftiofur sodium group, and it was inferred that this may be the killing effect of the combined mechanism of the two.

Example 5

acute toxicity test

The acute toxicity test in mice was conducted in accordance with the "Guiding Principles for Research on Acute Toxicity of Chemical Drugs in China" (H-GPT1-1) and the European Center for Chemical Ecotoxicology and Toxicology. Six female BALB/c mice in each group (6-8 weeks old, weight 18-20g) were injected intraperitoneally with shikimic acid SA at doses of 150 mg/kg, 100 mg/kg and 75 mg/kg according to the mouse body weight. Mice were injected with PBS as a control group (Control). The poisoning symptoms, abnormal behaviors and survival conditions of mice in each group were observed for 72 hours. The results are shown in Figure 8.

As can be seen from Figure 8, mice in each group showed no symptoms of poisoning, abnormal behavior, or death within 72 hours, indicating that 150 mg/kg had no effect on mice and shikimic acid has good safety.

Example 6

Histopathological examination

After the mice were adaptively fed for one week, 28 BALB/c female mice aged 6 to 8 weeks were randomly divided into 4 groups with 7 mice in each group. One group was the blank control group, and each mouse in the other three groups was injected intraperitoneally with 0.5 mL of MRSA bacterial suspension (final concentration: 1×10 ⁸ CFUs/mouse). 2 hours after bacterial infection, mice in the three groups were intraperitoneally injected with 300 μL of PBS buffer (pH value 7.4), recorded as the MRSA group; 300 μL of ceftiofur sodium (5 mg/kg based on mouse body weight), recorded as CF group; 300 μL of SA + ceftiofur sodium (injection of 75 mg/kg + 5 mg/kg according to the mouse body weight) was recorded as the SA + CF group. The dosage and method of administration were the same as those in the lethality experiment, and administration was repeated 12 hours later. After 24 hours, the mice were sacrificed by cervical dislocation, and the liver, spleen, and kidneys of the mice were removed aseptically and fixed in 4% formaldehyde. The liver, spleen and kidney tissues were dehydrated, paraffin embedded, sectioned, HE stained and other procedures in sequence, and then photographed and recorded under an optical microscope. The results are shown in Figure 9-11.

As can be seen from Figure 9, the infection group showed pathological changes in the liver, such as high edema and balloon-like degeneration of liver cells, and a large number of small vacuoles in the cytoplasm; the treatment group achieved some relief; Figure 10 shows that the spleen of the infection group showed megakaryocytes and The pathological changes of neutrophilia were effectively alleviated in the treatment group. Figure 11 shows that the renal tissue of the infection group showed pathological changes such as a large amount of homogeneous red-stained liquid in the lumen, and the lesions in the treatment group were effectively alleviated. Especially the effect of the combined treatment group was more obvious.

It can be seen from the above examples that the antibacterial composition provided by the present invention uses the natural Chinese medicine extract shikimic acid combined with antibiotics to treat and kill drug-resistant bacteria, significantly reducing the amount of antibiotics used, and while killing drug-resistant bacteria, it also There is no obvious impact on the liver, spleen, kidney and other organs and tissues, with few side effects and high safety. When used in combination at an appropriate concentration, it can significantly inhibit and kill methicillin-resistant Staphylococcus aureus (MRSA) in a short period of time. killing effect.

Although the above embodiments describe the present invention in detail, they are only part of the embodiments of the present invention, not all embodiments. People can also obtain other embodiments based on this embodiment without any inventive step. These embodiments All belong to the protection scope of the present invention.

Claims (6)

1. An antibacterial composition, characterized in that it includes the following components by weight: 8 to 32 parts of antibiotics and 312 to 625 parts of shikimic acid; the antibiotics are β-lactam antibiotics; the β-lactam Antibiotics are penicillin, ampicillin, amoxicillin or ceftiofur sodium. 2. The method for preparing an antibacterial composition according to claim 1, characterized by comprising: mixing antibiotics and shikimic acid to obtain the antibacterial composition. 3. The application of the antibacterial composition according to claim 1 or the antibacterial composition obtained by the preparation method according to claim 2 in the preparation of antibacterial drugs, the bacteria being drug-resistant bacteria; the drug-resistant bacteria being Grapevine aureus cocci. 4. The application according to claim 3, characterized in that the Staphylococcus aureus is methicillin-resistant Staphylococcus aureus. 5. The application according to claim 4, characterized in that the methicillin-resistant Staphylococcus aureus is the methicillin-resistant Staphylococcus aureus standard strain ATCC33591. 6. An antibacterial drug, characterized in that the active ingredient of the antibacterial drug is the antibacterial composition of claim 1 or the antibacterial composition obtained by the preparation method of claim 2.