Application of linoleic acid in preparation of medicine for improving sensitivity of gram-positive bacteria to antibiotics

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to application of linoleic acid in preparation of a medicine for improving sensitivity of gram-positive bacteria to antibiotics. It is found for the first time that combination of linoleic acid and antibiotics can significantly improve the sensitivity of gram-positive bacteria to antibiotics; in-vivo research data shows that the combination of linoleic acid and antibiotics not only can improve the survival rate of mice infected with gram-positive bacteria, but also can improve the ability of mice to remove gram-positive bacteria; linoleic acid and antibiotics are further prepared into the anti-infection composition, on one hand, the remarkable clinical anti-infection effect can be achieved, and on the other hand, the post-effect of the antibiotics can be prolonged; moreover, the linoleic acid is a necessary fatty acid in nutrition of human bodies and animals, is high in nutritional value, is widely applied to food and medicines, and is high in safety.

Description

[0001] This application is a divisional application of the invention patent application with application date of July 30, 2024, application number 2024110312484, and invention name “Application of linoleic acid in the preparation of drugs for increasing the sensitivity of Gram-positive bacteria to antibiotics”. Technical Field

[0002] The present invention belongs to the field of biomedicine technology and more specifically relates to the use of linoleic acid in the preparation of a drug for increasing the sensitivity of Gram-positive bacteria to antibiotics. Background Art

[0003] Staphylococcus aureus ( Staphyloccocus aureus Rosenbach ) is a common clinical pathogen that often causes community and hospital infections, and the infections it causes are second only to Escherichia coli. Reasonable antibiotic intervention is still an effective means of treating Staphylococcus aureus infections. β-lactam drugs represented by penicillin and cephalosporin are one of the important drugs for the clinical treatment of Staphylococcus aureus infections. However, due to the widespread use of β-lactam drugs in clinical practice, especially methicillin-resistant Staphylococcus aureus ( methicillin-resistant staphylococcus aureus The increase in the detection rate of MRSA and its multidrug-resistant characteristics have greatly limited the clinical efficacy of β-lactam drugs.

[0004] Therefore, in the current situation where the drug resistance of Gram-positive bacteria including MRSA is becoming increasingly serious and the development of new antimicrobial drugs is becoming increasingly difficult, finding effective antimicrobial drug enhancers and restoring the sensitivity of multidrug-resistant bacteria including MRSA to existing key antimicrobial drugs through reasonable combination drug strategies are of great significance for improving the clinical efficacy of antimicrobial drugs and delaying the development of drug resistance. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to overcome the defects and shortcomings that Gram-positive bacteria have developed resistance to existing antibiotics and the research and development of new antibacterial drugs is becoming increasingly difficult. The present invention provides the use of linoleic acid in the preparation of antibiotic synergists. The antibiotics can not only improve the survival rate of the body against infected Gram-positive bacteria, but also improve the body's ability to eliminate Gram-positive bacteria.

[0006] The purpose of the present invention is to provide an application of linoleic acid in combination with antibiotics in the preparation of a drug for increasing the sensitivity of Gram-positive bacteria to antibiotics.

[0007] Another object of the present invention is to provide the use of linoleic acid in combination with antibiotics in the preparation of a medicament for preventing and / or treating Gram-positive bacterial infection.

[0008] Another object of the present invention is to provide a drug for increasing the sensitivity of Gram-positive bacteria to antibiotics.

[0009] The above-mentioned purpose of the present invention is achieved through the following technical solutions: The present invention provides an application of protected linoleic acid in the preparation of an antibiotic synergist, wherein the antibiotic is selected from cephalosporins, aminoglycosides, quinolones, tetracyclines, carbapenems, glycopeptides, macrolides, sulfonamides, penicillins or pharmaceutically acceptable salts thereof, or a compound preparation containing one or more of the above antibiotics; and the penicillins do not include ampicillin.

[0010] Linoleic acid is an important unsaturated fatty acid and an essential fatty acid for the human body. It has the potential to lower cholesterol, soften blood vessels, and promote microcirculation, potentially preventing various cardiovascular and cerebrovascular diseases. The inventors' team has discovered for the first time that the combination of linoleic acid and the aforementioned antibiotics can significantly increase the sensitivity of Gram-positive bacteria to antibiotics. In vivo data show that combining linoleic acid with the aforementioned antibiotics (such as cefoperazone sodium and sulbactam sodium) not only improves the survival rate of Gram-positive bacteria but also enhances the body's ability to eliminate them. Combining linoleic acid with antibiotics to create an anti-infective composition can achieve significant clinical anti-infective efficacy and prolong the antibiotic's post-antibiotic effect (PAE), which persists after exposure to an antibiotic, even after the antibiotic's serum concentration has dropped below the minimum inhibitory concentration or has disappeared). Furthermore, linoleic acid is an essential fatty acid in human and animal nutrition, boasting high nutritional value. It is widely used in food and pharmaceuticals and is highly safe.

[0011] Furthermore, the linoleic acid increases the sensitivity of Gram-positive bacteria to antibiotics.

[0012] The present invention discloses an application of protected linoleic acid in combination with antibiotics in the preparation of a drug for increasing the sensitivity of Gram-positive bacteria to antibiotics, wherein the antibiotics are selected from cephalosporins, aminoglycosides, quinolones, tetracyclines, carbapenems, glycopeptides, macrolides, sulfonamides, penicillins or pharmaceutically acceptable salts thereof, or a compound preparation containing one or more of the antibiotics; and the penicillins do not include ampicillin.

[0013] The present invention also protects the use of linoleic acid in combination with antibiotics in the preparation of a medicament for preventing and / or treating Gram-positive bacterial infection, wherein the antibiotics are selected from cephalosporins, aminoglycosides, quinolones, tetracyclines, carbapenems, glycopeptides, macrolides, sulfonamides, penicillins or pharmaceutically acceptable salts thereof, or a compound preparation containing one or more of the above antibiotics; and the penicillins do not include ampicillin.

[0014] Preferably, the antibiotic is selected from cefoperazone sodium and sulbactam sodium (cephalosporins + β-lactamase inhibitors, i.e., a combination preparation), cefotaxime (cephalosporins), cefoperazone (cephalosporins), amikacin (aminoglycoside), tobramycin (aminoglycoside), gentamicin (aminoglycoside), ofloxacin (quinolone), levofloxacin (quinolone), tetracycline (tetracyclines), meropenem (carbapenem), vancomycin (glycopeptide), azithromycin (macrolide), sulfadiazine (sulfonamide) or amoxicillin sodium and clavulanate potassium (penicillin + β-lactamase inhibitor, i.e., a combination preparation).

[0015] Furthermore, the cefoperazone sodium and sulbactam sodium is a compound preparation of cefoperazone and sulbactam, wherein both cefoperazone and sulbactam exist in the form of sodium salts; further, in the compound preparation, the mass ratio of cefoperazone to sulbactam is 1:1.

[0016] Furthermore, the amoxicillin sodium clavulanate potassium is a compound preparation of amoxicillin and clavulanic acid, wherein amoxicillin exists in the form of a sodium salt, and clavulanate potassium exists in the form of a potassium salt; further, in the compound preparation, the mass ratio of amoxicillin to clavulanic acid is (2-7):1.

[0017] Preferably, in the compound preparation, the mass ratio of amoxicillin to clavulanic acid is 2:1, 4:1 or 7:1.

[0018] Furthermore, the Gram-positive bacteria are selected from one or more of Staphylococcus aureus, Streptococcus iniae, Streptococcus agalactiae, Streptococcus pyogenes, Bacillus subtilis, and Enterococcus faecalis.

[0019] Furthermore, the Staphylococcus aureus includes sensitive bacteria and drug-resistant bacteria.

[0020] Furthermore, the sensitive bacteria is Methicillin-Sensitive Staphylococcus Aureus (MSSA), and the resistant bacteria is Methicillin-Resistant Staphylococcus Aureus (MRSA).

[0021] Furthermore, the linoleic acid acts as an antibiotic synergist to enhance the ability of antibiotics to kill Gram-positive bacteria.

[0022] Furthermore, the linoleic acid acts as an antibiotic synergist to enhance the ability of antibiotics to eliminate Gram-positive bacteria.

[0023] Furthermore, the linoleic acid acts as an antibiotic synergist to enhance the body's resistance to Gram-positive bacteria infection.

[0024] Furthermore, the linoleic acid acts as an antibiotic synergist to increase the amount of antibiotics that enter cells. Exogenous addition of linoleic acid can increase the permeability of the bacterial membrane of Gram-positive bacteria, thereby increasing the amount of antibiotics that enter the cells and thus promoting the sensitivity of bacteria to antibiotics.

[0025] Furthermore, the linoleic acid acts as an antibiotic synergist to delay the after-effects of Gram-positive bacteria on antibiotics.

[0026] Furthermore, the linoleic acid acts as an antibiotic synergist to delay the after-effects of Gram-positive bacteria on cefoperazone sodium and sulbactam sodium.

[0027] The present invention also protects a drug for increasing the sensitivity of Gram-positive bacteria to antibiotics, comprising an effective amount of linoleic acid and an antibiotic; the antibiotic is selected from cephalosporins, aminoglycosides, quinolones, tetracyclines, carbapenems, glycopeptides, macrolides, sulfonamides, penicillins or pharmaceutically acceptable salts thereof, or a compound preparation containing one or more of the antibiotics; and the penicillins do not include ampicillin.

[0028] Preferably, the mixing ratio of linoleic acid to antibiotic is 1:(2-40,000) g / mol, which means that 2-40,000 g of antibiotic is used in combination with 1 mol of linoleic acid.

[0029] More preferably, the mixing ratio of the linoleic acid and the antibiotic is 1:(50-40000) g / mol.

[0030] Compared with the prior art, the present invention has the following beneficial effects: the present invention discovers for the first time that the combination of linoleic acid and antibiotics significantly increases the sensitivity of Gram-positive bacteria to antibiotics; in vivo research data show that the combination of linoleic acid and antibiotics can not only increase the survival rate of mice infected with Gram-positive bacteria, but also improve the mice's ability to eliminate Gram-positive bacteria; further, linoleic acid and antibiotics are prepared into an anti-infection composition, which can achieve significant clinical anti-infection effects on the one hand, and prolong the clinical effects of antibiotics on the other hand; and linoleic acid is an essential fatty acid in human and animal nutrition, has high nutritional value, has been widely used in food and medicine, and is highly safe. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a diagram showing the results of PCR amplification of clinical Staphylococcus aureus genes in Example 1.

[0032] Figure 2 Statistical graph showing the results of linoleic acid killing the clinically sensitive Staphylococcus aureus MSSA 2 (A) and the drug-resistant Staphylococcus aureus MRSA 7 (B) in Example 2, as well as the linoleic acid concentration effect of the bactericidal effect.

[0033] Figure 3 Statistical graph showing the time effect of linoleic acid in killing clinical Staphylococcus aureus MRSA 7 (A) and MSSA 2 (B) in Example 2.

[0034] Figure 4 This is a statistical chart showing the results of linoleic acid killing multiple strains of clinically sensitive Staphylococcus aureus (A) and resistant bacteria (B) in Example 2.

[0035] Figure 5 This is a statistical graph showing the results of Example 3 showing that linoleic acid can improve the survival rate (A) and clearance ability (B) of mice infected with clinical Staphylococcus aureus.

[0036] Figure 6 This is a statistical graph showing the results of Example 4 showing that linoleic acid can increase the sensitivity of clinical Staphylococcus aureus (A) and other Gram-positive bacteria (B) to cefoperazone sodium and sulbactam sodium, and can increase the sensitivity of Staphylococcus aureus to other antibiotics (C).

[0037] Figure 7 Statistical graphs of the effects of linoleic acid concentration (A), antibiotic concentration (B) and time gradient (C) on linoleic acid improving the sensitivity of clinical Staphylococcus aureus to cefoperazone sodium and sulbactam sodium in Example 5.

[0038] Figure 8 This is a statistical graph showing the results of Example 6 showing that linoleic acid synergistically with cefoperazone sodium and sulbactam sodium can improve the survival rate (A) and clearance ability (B) of mice infected with clinical Staphylococcus aureus.

[0039] Figure 9 This is a statistical graph showing the results of improving the permeability of clinically sensitive Staphylococcus aureus MSSA (A) and resistant Staphylococcus aureus MRSA (B) after adding linoleic acid in Example 7.

[0040] Figure 10 This is a statistical chart showing the results of increasing the antibiotic content in clinical Staphylococcus aureus after adding linoleic acid in Example 7.

[0041] Figure 11 This is a statistical chart showing the results of Example 8 in which linoleic acid can delay the post-treatment effect of cefoperazone sodium and sulbactam sodium on Staphylococcus aureus. DETAILED DESCRIPTION

[0042] The present invention will be further described below with reference to the accompanying drawings and specific examples, but the examples do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0043] Unless otherwise specified, the reagents and materials used in the following examples were commercially available.

[0044] Example 1 Analysis of clinical antibiotic resistance of Staphylococcus aureus Nine strains of sensitive Staphylococcus aureus ( Staphylococcu saureus , MSSA) and 20 strains of methicillin-resistant Staphylococcus aureus ( Methicillin-resistant Staphylococcu saureus , MRSA), named MSSA 1 to 9 and MRSA 1 to 20.

[0045] 1.1 Determination of drug resistance of clinical strains Twenty-nine clinical Staphylococcus aureus strains were cultured overnight, inoculated at a 1:100 ratio, and then incubated to an OD600 of 0.5. The strains were then diluted 100-fold and the minimum inhibitory concentrations (MICs) of these strains to daptomycin (DAP), vancomycin (VAN), cefoperazone sodium and sulbactam sodium (SCF), and oxacillin (OX) were determined according to the Clinical and Laboratory Standards Institute (CLSI) microbial susceptibility testing protocol. The results are shown in Table 1.

[0046] Table 1 Minimum inhibitory concentration (MIC) of four antibiotics against 29 strains of Staphylococcus aureus

[0047] Note: For OX, MIC ≤ 2 indicates sensitive bacteria; for DAP: MIC ≤ 1 indicates sensitive bacteria; for VAN: MIC ≤ 2 indicates sensitive bacteria.

[0048] As shown in Table 1, the above 20 MRSA strains were all resistant to the four antibiotics tested, which also showed that these 20 strains were not only methicillin-resistant but also multidrug-resistant (Table A in Table 1), while 9 MRSA strains were sensitive to all four antibiotics (Table B in Table 1).

[0049] 1.2 Genetic identification of clinical strains femB (651bp) is an intrinsic gene of Staphylococcus aureus. mecA (310bp) is a unique gene of methicillin-resistant strains. PVL is an exotoxin produced by Staphylococcus aureus - leukocidin, which damages the body's defense barrier and immune response by destroying white blood cells and phagocytes.

[0050] Clinical strains were cultured overnight, and the three genes of each strain were amplified by colony PCR (the amplification primer sequences are shown in Table 2), and the amplified products were detected by gel electrophoresis. Figure 1 As can be seen from the figure, for MRSA strains, each resistant strain can amplify these three genes, indicating that it is indeed Staphylococcus aureus, and is a methicillin-resistant strain, and is toxic. For sensitive strains, each strain can amplify femB , but it cannot be expanded mecA , indicating that these strains are indeed Staphylococcus aureus and are sensitive bacteria. However, some strains have exotoxin genes, while others do not.

[0051] Table 2 Primer sequences

[0052] Example 2 Linoleic acid can kill Staphylococcus aureus from clinical sources 2.1 Strain sample preparation Single colonies of clinical bacteria were picked from solid LB plates, inoculated into 5 mL of LB liquid medium, and cultured at 37°C and 200 rpm for 16 h. The bacterial suspension was collected and centrifuged at 8000 rpm for 5 min. The supernatant was removed and the cells were washed with an equal volume of 0.85% saline. Finally, the cells were suspended in 1× M9 minimal medium (containing 10 mM acetate and 1 M CaCl2). The OD value of the bacterial suspension was adjusted to 0.2 and 5 mL was aliquoted into test tubes for subsequent experiments.

[0053] 2.2 Linoleic acid can kill clinical Staphylococcus aureus and has a linoleic acid concentration gradient effect Prepare samples of clinically sensitive strain MSSA 2 and clinically drug-resistant strain MRSA 7 according to 2.1 in Example 2. Linoleic acid was added to the test tubes to give final concentrations of 0, 0.01, 0.05, 0.1, 0.5, 1, 5, and 10 mM, respectively. Three biological replicates were performed for each concentration. Culture was performed at 37°C, 200 rpm, for 10 hours. The number of viable bacteria was detected using a plate, and then the survival rate of bacteria at different linoleic acid concentrations was calculated. The calculation formula is: survival rate (%) = (number of viable bacteria after adding linoleic acid / number of viable bacteria without adding linoleic acid) × 100%. The results are shown in Figure 2 As can be seen from the figure, the survival rate of bacteria, whether sensitive or resistant, is significantly reduced after adding linoleic acid. And as the concentration of linoleic acid increases, the bactericidal efficiency gradually increases. The specific situation is: For clinically sensitive bacteria MSSA 2 (such as Figure 2 As shown in Figure A), when 0.01mM linoleic acid is added, it can play a better killing role, and the bactericidal effect is increased by 8.07 times compared with the control group (the survival rate dropped to 12.39%); when the concentration of linoleic acid gradually increases from 0.05mM to 10mM, the bactericidal efficiency increases by 19.91 times, 129.58 times, 709.17 times, 1925.93 times, 4681.57 times and 9577.32 times respectively (the survival rate also drops from 5.02% to 0.01%).

[0054] For clinical drug-resistant bacteria MRSA 7 (such as Figure 2 As shown in Figure B), when the concentration of linoleic acid was 0.01mM, the bactericidal effect was also improved by 7.14 times compared with the control group (the survival rate dropped to 14.00%); similarly, when the concentration of linoleic acid gradually increased from 0.05mM to 10mM, the bactericidal efficiency increased by 15 times, 39.47 times, 230.77 times, 1257.24 times, 2675.16 times and 4199.48 times respectively (the survival rate also dropped from 6.67% to 0.02%).

[0055] At a 1mM concentration of linoleic acid, the bactericidal efficiency against sensitive and resistant bacteria increased by 1925.93-fold and 1257.24-fold, respectively. Increasing the linoleic acid concentration further increased the bactericidal efficiency. However, due to its acidic nature, adding too high a concentration can affect the pH of the incubation system. Therefore, 1mM linoleic acid was selected for subsequent studies.

[0056] This result shows that linoleic acid can kill clinical Staphylococcus aureus, including sensitive strains and resistant strains, and has a linoleic acid concentration gradient effect.

[0057] 2.3 Linoleic acid kills clinical Staphylococcus aureus in a time-dependent manner MSSA 2 and MRSA 7 samples were prepared according to 2.1 of Example 2. 1 mM linoleic acid was added to each test tube. No linoleic acid was used as a control. Live bacteria were counted on plates at different times and the survival rate was calculated to study the relationship between bactericidal efficiency and time.

[0058] The results are as follows Figure 3 As shown in the figure, after adding linoleic acid, the bacterial count of clinical bacteria MSSA 2 and MRSA7 did not decrease within 2 hours, and then the bacterial count decreased significantly as time went on, showing a time effect. The specific situation is as follows: For MRSA 7 ( Figure 3 For example, the bacterial count decreased by 15 times in 4 hours, and by 137.67-13641.57 times in 6-12 hours. Figure 3 For example, in Figure B, the number of bacteria decreased by 12.8 times in 4 hours, and the reduction in the number of bacteria ranged from 216.15 to 16818.73 times in 6-12 hours.

[0059] These results demonstrate that linoleic acid's ability to kill clinical Staphylococcus aureus is time-dependent. Linoleic acid can achieve significant killing effects against clinical Staphylococcus aureus after just 10 hours. For ease of use, subsequent experiments will use a 10-hour bactericidal duration for linoleic acid.

[0060] 2.4 Linoleic acid is widely used to kill Staphylococcus aureus in clinical practice Prepare the remaining 8 sensitive and 19 resistant bacterial strains according to 2.1 of Example 2. Each bacterial strain was divided into two groups: an M9 control group and a 1 mM linoleic acid test group. After incubation at 37°C, 200 rpm, viable bacteria were counted and the survival rate of each strain after the addition of linoleic acid was calculated. Three biological replicates were performed for each strain.

[0061] See the results Figure 4 As can be seen from the figure, linoleic acid can kill all clinically sensitive bacteria and drug-resistant bacteria, but the degree of sensitivity improvement varies for different strains. The specific description is as follows: To 8 sensitive bacteria ( Figure 4 Figure A in the figure shows that, except for the MSSA1 strain, the bactericidal multiple after adding linoleic acid was only 81.97 times, the bactericidal multiples of the other strains ranged from 1925.93 to 98846.78 times.

[0062] For 19 drug-resistant strains ( Figure 4(Figure B in the figure) The sterilization multiples ranged from 7.39 to 14918.69. Three strains (11, 12, and 15) had sterilization multiples below 100 times; five strains (8, 13, 14, 16, and 17) had sterilization multiples greater than 100 times; and 11 strains (1-6, 9, 10, and 18-20) had sterilization multiples greater than 1000 times.

[0063] Example 3 Linoleic acid can improve the resistance of mice to clinical Staphylococcus aureus infection 3.1 Linoleic acid can improve the survival rate of mice against clinical Staphylococcus aureus infection Balb / c mice (6-8 weeks, approximately 20 g, half male and half female) were divided into three groups, 16 in each group: a normal saline control group, an antibiotic treatment group, and a linoleic acid test group. The challenge strain was the drug-resistant MRSA 7, and the infection dose was 1.2 × 10 8 CFU were measured by intraperitoneal injection. One hour after bacterial infection, mice in the treatment group were intravenously administered cefoperazone sodium and sulbactam sodium at a dose of 200 mg / kg; mice in the experimental group were intraperitoneally administered linoleic acid at a dose of 100 mg / kg. Mouse mortality was observed and recorded daily, and the survival rate of each group was calculated daily for a total of 7 days. The calculation formula was: Survival rate = number of surviving mice per group / total number of mice per group × 100%.

[0064] The experimental results are shown in Figure 5 As shown in Figure A, in the saline control group, 68% died on the first day, and all died on the second day. In the antibiotic-treated group, 31% died on the first day, 62% died on the second day, and no further deaths occurred, with a survival rate of 7%. In the linoleic acid-treated group, no deaths occurred on the first day, 7% died on the second day, and then stabilized, with no further deaths, for a survival rate of 93%. Compared with the control group, the protection rate of mice injected with linoleic acid increased by 93%, and compared with the antibiotic-treated group, the protection rate of mice injected with linoleic acid increased by 86%.

[0065] The results show that when mice are infected with methicillin-resistant bacteria, linoleic acid can significantly improve the mice's resistance to methicillin-resistant bacteria infection when antibiotic treatment is basically ineffective.

[0066] 3.2 Linoleic acid can improve the clearance of clinical Staphylococcus aureus infection in mice Balb / c mice (6-8 weeks old, approximately 20 g, half male and half female) were divided into two groups, 6 in each group, a control group and a linoleic acid test group. The challenge strain was methicillin-resistant Staphylococcus aureus MRSA 7, and the infection dose was 1.4×10 6CFU were determined by intraperitoneal injection. One hour after bacterial infection, linoleic acid was administered intraperitoneally at a dose of 100 mg / kg. Six hours later, equal weights of viscera (liver, spleen, and kidneys) were collected and thoroughly ground. The samples were then diluted and plated to count the bacterial count within each organ.

[0067] See the results Figure 5 As shown in Figure B, when mice were infected with drug-resistant bacteria, linoleic acid treatment reduced bacterial loads in the spleen, kidney, and liver by 1,563-fold, 1,009-fold, and 4,419-fold, respectively. These results indicate that linoleic acid significantly improves the clearance of clinical Staphylococcus aureus infections in mice.

[0068] Example 4 Linoleic acid can increase the sensitivity of Gram-positive bacteria to antibiotics 4.1 Linoleic acid can increase the sensitivity of clinical Staphylococcus aureus to cefoperazone sodium and sulbactam sodium The results of 2.4 in Example 2 showed that linoleic acid had a killing effect on clinically derived Staphylococcus aureus. However, it was also found that the bactericidal effect varied among different strains, with some strains showing a killing effect of less than 1000-fold. Given that bacterial infections are still currently treated with antibiotics, can linoleic acid, in combination with antibiotics, increase the sensitivity of strains to antibiotics? To this end, a bactericidal test was conducted on 14 bacterial strains, including MRSA 8, 11-17, and MSSA 1, all of which had a bactericidal effect of less than 1000-fold with linoleic acid in 2.4 in Example 2. Linoleic acid was also used to kill 14 strains, including MSSA2, MSSA8, MRSA4, 9, and 18, for which the bactericidal effect of linoleic acid was greater than 1000-fold.

[0069] The above strain samples were prepared according to 2.1 of Example 2. Each strain was divided into four groups: an M9 control group, an antibiotic group, a linoleic acid group, and a linoleic acid + antibiotic group. The antibiotics used were cefoperazone sodium and sulbactam sodium, and the concentrations used for each strain were shown in Table 3. The linoleic acid concentration was 1 mM. After incubation at 37°C and 200 rpm for 10 hours, the viable bacterial counts were determined using a plate, and the survival rates of the bacteria in the different treatment groups were calculated. The calculation formula was: Survival rate (%) = (viable bacterial count after addition of linoleic acid and / or antibiotics / viable bacterial count in the control group) × 100%.

[0070] See the results Figure 6As shown in Figure A, antibiotics alone had no or very weak bactericidal effect on these strains, with a bactericidal multiple ranging from only 1.07-2.57. However, the addition of linoleic acid to the antibiotics significantly improved the sensitivity of all strains to the antibiotics. For strains where the bactericidal effect of linoleic acid alone was less than 1000-fold, the bactericidal multiple increased by 7.88-282.44-fold compared to the original bactericidal multiple. Furthermore, for strains where the bactericidal effect of linoleic acid alone was already greater than 1000-fold, the bactericidal multiple increased by 1.35-4-fold compared to the original bactericidal multiple.

[0071] Table 3 Concentrations of cefoperazone sodium and sulbactam sodium used in the experiment

[0072] 4.2 Linoleic acid can increase the sensitivity of other Gram-positive bacteria to cefoperazone sodium and sulbactam sodium Samples of Streptococcus iniae, Streptococcus agalactiae, Streptococcus pyogenes, Bacillus subtilis, and Enterococcus faecalis were prepared according to 2.1 of Example 2. The antibiotic resistance (MIC) of these bacteria is shown in Table 4. Each strain was divided into four groups: an M9 control group, an antibiotic group, a linoleic acid group, and a linoleic acid + antibiotic group. The antibiotics used were cefoperazone sodium and sulbactam sodium, and the concentrations used for each strain were: 300 μg / mL for Streptococcus iniae, 50 μg / mL for Streptococcus agalactiae, 400 μg / mL for Streptococcus pyogenes, 200 μg / mL for Bacillus subtilis, and 400 μg / mL for Enterococcus faecalis. The linoleic acid concentration was 0.01 mM. After incubation at 37°C and 200 rpm for 10 hours, the viable cell count was determined using a plate, and the survival rate of the bacteria in the different treatment groups was calculated. The calculation formula is: Survival rate (%) = (viable cell count after addition of linoleic acid and / or antibiotics / viable cell count in the control group) × 100%.

[0073] Table 4 Bacterial resistance detection

[0074] Note: For Streptococcus dolphins, Streptococcus agalactiae, and Streptococcus pyogenes: Penicillins ≤0.25 is sensitive, and there is no standard for resistance and intermediate levels; Cephalosporins ≤0.25 is sensitive, and there is no standard for resistance and intermediate levels; Carbapenems ≤0.5 is sensitive, and there is no standard for resistance and intermediate levels; Tetracycline: ≤2 is sensitive, 4 is intermediate, and ≥8 is resistant; Levofloxacin: ≤2 is sensitive, 4 is intermediate, and ≥8 is resistant; Vancomycin ≤1 is sensitive, and there is no standard for resistance and intermediate levels.

[0075] Enterococcus faecalis: Penicillins: ≤8 is sensitive, no intermediate values ​​have been reported, and ≥16 is resistant; Tetracyclines: ≤4 is sensitive, 8 is intermediate, and ≥16 is resistant; Levofloxacins: ≤2 is sensitive, 4 is intermediate, and ≥8 is resistant; Vancomycin: ≤4 is sensitive, 8-16 is intermediate, and ≥32 is resistant.

[0076] Bacillus subtilis: Penicillins ≤8 are sensitive, 8-16 are intermediate, and ≥32 are resistant; cephalosporins ≤4 are sensitive, 8 are intermediate, and ≥16 are resistant; Tobramycin ≤4 are sensitive, 8 are intermediate, and ≥16 are resistant; Tetracycline: ≤4 are sensitive, 8 are intermediate, and ≥16 are resistant; Levofloxacin: ≤0.5 are sensitive, 1 is intermediate, and ≥2 are resistant.

[0077] R: resistant; I: intermediate; S: sensitive.

[0078] As shown in Table 4, the Gram-positive bacteria listed above are resistant to most of the antibiotics tested and are multidrug-resistant bacteria.

[0079] Depend on Figure 6 As shown in Figure B, antibiotics alone had no or very weak bactericidal effect on these strains, with kill rates ranging from only 1.1 to 1.69. However, the addition of linoleic acid significantly improved the sensitivity of all strains to the antibiotics, with kill rates increasing by 315.58, 3.71, 2254.49, 77.45, and 284.59 for Streptococcus iniae, Streptococcus agalactiae, Streptococcus pyogenes, Bacillus subtilis, and Enterococcus faecalis, respectively.

[0080] 4.3 Linoleic acid can increase the sensitivity of clinical Staphylococcus aureus to other antibiotics Clinically resistant MRSA 9 samples were prepared according to Section 2.1 of Example 2 and divided into four groups: an M9 control group, a linoleic acid group, an antibiotic group, and a linoleic acid + antibiotic group. The antibiotic concentrations used in the experiment were: vancomycin at 100 μg / mL, and all other antibiotics at 100 μg / mL; the linoleic acid concentration was 0.01 mM. After incubation at 37°C and 200 rpm for 10 hours, the viable bacterial count was determined using a plate, and the survival rate of the bacteria in each treatment group was calculated. The calculation formula was: Survival rate (%) = (viable bacterial count after addition of linoleic acid and / or antibiotics / viable bacterial count in the control group) × 100%.

[0081] See the results Figure 6As shown in Figure C, when antibiotics were added alone, bacterial survival rates ranged from 67.24-98.27%, with the exception of vancomycin, where the survival rate was 44.82%. When linoleic acid was added alone, the bacterial survival rate was 70%. However, when linoleic acid was added to the antibiotics, the bacterial sensitivity to antibiotics was increased by 2.82-2826 times, with the exception of polymyxin B, clindamycin, and ampicillin.

[0082] These results show that when linoleic acid is used in combination with cefoperazone sodium and sulbactam sodium, or other antibiotics such as tetracycline, meropenem, amikacin, levofloxacin, tobramycin, gentamicin, vancomycin, cefotaxime, azithromycin, amoxicillin sodium and clavulanate potassium and sulfadiazine, the bactericidal ability of antibiotics against these bacteria is enhanced, indicating that linoleic acid can not only significantly increase the sensitivity of clinical Staphylococcus aureus to cefoperazone sodium and sulbactam sodium, but also increase the sensitivity of Gram-positive bacteria such as Streptococcus, Bacillus subtilis and Coprococcus to cefoperazone sodium and sulbactam sodium and other antibiotics including tetracycline, meropenem, amikacin, levofloxacin, tobramycin, gentamicin, vancomycin, cefotaxime, azithromycin, amoxicillin sodium and clavulanate potassium and sulfadiazine.

[0083] Example 5 Linoleic acid improves the sensitivity of clinical Staphylococcus aureus to cefoperazone sodium and sulbactam sodium, which is affected by linoleic acid concentration, antibiotic concentration and time. Taking Staphylococcus aureus as the representative of Gram-positive bacteria and cefoperazone sodium and sulbactam sodium as the representatives of antibiotics, we further explored the effect of combined use of linoleic acid and antibiotics.

[0084] 5.1 Linoleic acid concentration effect According to 2.1 in Example 2, clinical drug-resistant strain MRSA 12 samples were prepared and divided into 7 groups: an antibiotic control group (200 μg / mL cefoperazone sodium and sulbactam sodium), and 6 experimental groups: linoleic acid was added to 200 μg / mL cefoperazone sodium and sulbactam sodium, respectively, to make the final concentrations of 0.01, 0.05, 0.1, 0.5, 1 and 5 mM, respectively. Three biological replicates were performed for each concentration. The cells were cultured at 37°C and 200 rpm for 10 hours, and the number of viable bacteria was detected by plate and then converted into Log10 values. A decrease of 1 Log10, i.e., a decrease of 1 order of magnitude, indicates that the number of bacteria has decreased by 10 times, and a decrease of 2 Log10, i.e., a decrease of 2 orders of magnitude, indicates that the number of bacteria has decreased by 100 times. The results are shown in FIG. Figure 7 As shown in Figure A, the number of surviving bacteria decreases after adding linoleic acid in addition to antibiotics. The decrease in surviving bacteria becomes more pronounced as the concentration of linoleic acid increases.

[0085] The specific results are as follows: Adding 0.01 mM linoleic acid to antibiotics significantly improved antibiotic sensitivity, reducing the number of surviving bacteria by nearly 0.5 log (0.5 orders of magnitude) compared to the control group (where antibiotics were added alone), a reduction of approximately 5-fold. When the linoleic acid concentration was gradually increased from 0.05 mM to 5 mM, the number of surviving bacteria decreased by 1.32, 1.97, 2.86, 3.28, and 3.73 orders of magnitude, respectively, representing a decrease in bacterial counts from a dozen to over a thousand-fold.

[0086] 5.2 Antibiotic Concentration Effect Clinically resistant MRSA 12 samples were prepared according to 2.1 of Example 2 and divided into 6 groups: a linoleic acid control group (1 mM) and 5 experimental groups: 1 mM linoleic acid was added to which cefoperazone sodium and sulbactam sodium were added, to final concentrations of 10, 50, 100, 200, and 400 μg / mL, respectively. Three biological replicates were performed for each concentration. The cells were cultured at 37°C, 200 rpm, for 10 hours. The viable counts were determined by plate counting and then converted to Log10 values. Results are shown in Table 1. Figure 7 As shown in Figure B, the number of surviving bacteria decreases after adding antibiotics to linoleic acid. The decrease in surviving bacteria becomes more pronounced as the concentration of antibiotics increases.

[0087] The specific results are as follows: Adding 10 μg / mL of antibiotics to linoleic acid significantly improved antibiotic sensitivity, reducing the number of surviving bacteria by 0.59 logs compared to the control group (linoleic acid alone), a reduction of approximately sixfold. When the antibiotic concentration was gradually increased from 50 μg / mL to 400 μg / mL, the number of surviving bacteria decreased by 1.76, 3.24, 4.15, and 4.61 logs, respectively, representing a decrease in bacterial counts from nearly 200-fold to over 10,000-fold.

[0088] 5.3 Time Effect According to 2.1 of Example 2, clinical drug-resistant strain MRSA 12 samples were prepared and divided into three groups: an M9 control group, an antibiotic control group (200 μg / mL cefoperazone sodium and sulbactam sodium), and a linoleic acid (1 mM) + antibiotic (200 μg / mL cefoperazone sodium and sulbactam sodium) test group. The cells were cultured at 37°C and 200 rpm. Samples were taken every 2 hours for 12 hours. The viable counts were determined using plate counts and converted to Log10 values. Three biological replicates were performed for each test sample. Results are shown in Table 1. Figure 7As shown in Figure C, the bacterial count in the M9 control group remained essentially stable over 12 hours. Over time, the bacterial counts in the antibiotic and linoleic acid + antibiotic groups gradually decreased. Importantly, at the same time point, the number of surviving bacteria in the linoleic acid + antibiotic group was significantly lower than that in the antibiotic group. Furthermore, the decrease in the number of surviving bacteria became even more pronounced over time.

[0089] The specific results are as follows: After 2 hours, the number of surviving bacteria in the linoleic acid + antibiotic group decreased by 0.67 orders of magnitude, or about 7 times, compared to the surviving bacteria in the antibiotic group. From 4 to 12 hours, the number of surviving bacteria in the linoleic acid + antibiotic group decreased by 1.21 to 2.96 orders of magnitude, or about 10 to 1,000 times.

[0090] The above test results show that linoleic acid can cooperate with cefoperazone sodium and sulbactam sodium to improve the sensitivity of resistant Staphylococcus aureus to cefoperazone sodium and sulbactam sodium, and has antibiotic concentration and linoleic acid concentration effects, as well as time effects.

[0091] Example 6 Linoleic acid synergistically combines cefoperazone sodium and sulbactam sodium to improve the resistance of mice to clinical Staphylococcus aureus infection 6.1 Linoleic acid synergistically combines cefoperazone sodium and sulbactam sodium to improve the survival rate of mice infected with Staphylococcus aureus Taking Staphylococcus aureus as the representative of Gram-positive bacteria and cefoperazone sodium and sulbactam sodium as the representatives of antibiotics, we further explored the effect of combined use of linoleic acid and antibiotics.

[0092] Balb / c mice (6-8 weeks, approximately 20 g, half male and half female) were divided into four groups, 16 in each group: a normal saline control group, an antibiotic treatment group, a linoleic acid test group, and an antibiotic + linoleic acid test group. The challenge strain was methicillin-resistant MRSA 12, and the infection dose was 1.2 × 10 8 CFU were measured by intraperitoneal injection. One hour after bacterial infection, mice in the antibiotic treatment group received 200 mg / kg of cefoperazone sodium and sulbactam sodium via intravenous injection. Mice in the linoleic acid experimental group received 200 mg / kg of linoleic acid via intraperitoneal injection. Mice in the antibiotic + linoleic acid experimental group received antibiotics and linoleic acid according to their respective injection methods and dosages. Mice were observed and recorded daily for mortality, and the survival rate of each group was calculated daily for a total of 7 days. The calculation formula was: Survival rate (%) = number of surviving mice per group / total number of mice per group × 100%.

[0093] The experimental results are shown in Figure 8As shown in Figure A, in the saline control group, 81% died on the first day, and all died on the second day. In the antibiotic group, 31% died on the first day, 62% died on the second day, and no further deaths occurred, with a survival rate of 7%. In the linoleic acid group, 7% died on the first day, 62% died on the second day, and 7% died on the third day, but then stabilized and no further deaths occurred, with a survival rate of 24%. In the antibiotic plus linoleic acid group, no deaths occurred on the first day, 13% died on the second day, and then stabilized and no further deaths occurred, with a survival rate of 87%.

[0094] The results showed that the protection rate of mice in the linoleic acid group increased by 24% compared to the saline control group and by 17% compared to the antibiotic treatment group. The protection rate of mice in the antibiotic + linoleic acid group increased by 63% compared to the linoleic acid group and by 80% compared to the antibiotic treatment group. This suggests that when mice are infected with methicillin-resistant bacteria, linoleic acid can improve their resistance to methicillin-resistant infection to a certain extent, even when antibiotic treatment is largely ineffective. However, the combination of linoleic acid and antibiotics significantly improves the resistance of mice to methicillin-resistant infection.

[0095] 6.2 Linoleic acid synergistically combines cefoperazone sodium and sulbactam sodium to improve clearance of clinical Staphylococcus aureus infection in mice Balb / c mice (6-8 weeks old, approximately 20 g, half male and half female) were divided into four groups, each with 6 mice: a normal saline control group, an antibiotic treatment group, a linoleic acid test group, and an antibiotic + linoleic acid test group. The challenge strain was methicillin-resistant Staphylococcus aureus MRSA 12, with an infection dose of 1.4 × 10 6 CFU were determined by intraperitoneal injection. One hour after bacterial infection, mice in the antibiotic group received 200 mg / kg of cefoperazone sodium and sulbactam sodium via intravenous injection. Mice in the linoleic acid group received 200 mg / kg of linoleic acid via intraperitoneal injection. Mice in the antibiotic + linoleic acid group received antibiotics and linoleic acid according to their respective injection methods and doses. Six hours later, equal weights of viscera (liver, spleen, and kidneys) were thoroughly ground, diluted, plated, and counted.

[0096] See the results Figure 8Panel B shows that when mice were infected with drug-resistant bacteria, antibiotic treatment reduced bacterial loads in the spleen, kidney, and liver by 55-fold, 40-fold, and 33-fold, respectively. Linoleic acid treatment reduced bacterial loads in the spleen, kidney, and liver by 20-fold, 8-fold, and 17-fold, respectively. Linoleic acid and antibiotic treatment reduced bacterial loads in the spleen, kidney, and liver by 136-fold, 111-fold, and 97-fold, respectively, compared to the antibiotic-treated group. Compared to the linoleic acid treatment group, bacterial loads decreased by 51-fold, 22-fold, and 50-fold, respectively. These results demonstrate that the combination of linoleic acid and antibiotics significantly enhances the clearance of clinical Staphylococcus aureus infection in mice.

[0097] Example 7: Linoleic acid increases bacterial membrane permeability, allowing antibiotics to enter the cell and enhance sensitivity to antibiotics. Taking Staphylococcus aureus as the representative of Gram-positive bacteria and cefoperazone sodium and sulbactam sodium as the representatives of antibiotics, we further explored the mechanism by which the combined use of linoleic acid and antibiotics improves the efficacy of the drugs.

[0098] 7.1 Exogenous linoleic acid can increase bacterial membrane permeability Prepare bacteria according to 2.1 in Example 2, with a total of 9 strains of MSSA and 10 strains of MRSA. Each bacterial strain was divided into 2 groups, one as a control group and one as a linoleic acid (1 mM) experimental group. Incubate at 37°C 200 rpm on a shaker for 10 hours. Then, take 100 μL and add it to 900 μL of M9, add 2 μL of 2.5 mM SYTO9 dye. Incubate at 37°C 200 rpm for 45 minutes, detect fluorescence by flow cytometry, and determine changes in membrane permeability by comparing the fluorescence intensity of the experimental group with that of the control group. The results are shown in Figure 2. Figure 9 The membrane permeability of both MSSA and MRSA was enhanced after exogenous addition of linoleic acid, indicating that linoleic acid can increase the membrane permeability of bacteria.

[0099] 7.2 Exogenous linoleic acid can promote the entry of antibiotics into bacterial cells The method of detecting the inhibitory effect of antibiotics on microorganisms and calculating the activity (potency) of antibiotics is used to determine the concentration of antibiotics entering bacterial cells.

[0100] Sample preparation for measuring intracellular antibiotic concentrations: MRSA strain 7 samples were prepared according to Example 2.1 and divided into three groups: a control group without substance or antibiotics, a cefoperazone sodium and sulbactam sodium group, and a cefoperazone sodium and sulbactam sodium + linoleic acid group. Three biological replicates were performed in each group. The cells were incubated at 37°C and 200 rpm for 10 hours, then harvested by centrifugation and washed multiple times to remove residual antibiotics from the culture medium. The cells were suspended in 1× M9 minimal medium (containing 10 mM acetate and 1 M CaCl2), adjusted to an OD value of 1.0, and centrifuged to collect 10 mL of the culture medium. After adding 350 μL of ultra-disruption solution containing 2% SDS and ultrasonically disrupting the solution (ultra-disruption was performed on ice, ultra-disruption power was 35%, ultrasonication was performed for 2 seconds and then stopped for 3 seconds, and ultrasonic disruption time was 25 minutes), the supernatant was collected by centrifugation and filtered to remove any residual bacteria in the supernatant. The ultra-disruption solutions of the control group, cefoperazone sodium and sulbactam sodium group, and cefoperazone sodium and sulbactam sodium + linoleic acid group were obtained, respectively.

[0101] Preparation of test strains: Staphylococcus aureus ATCC17978 was cultured overnight to saturation, diluted with 1×M9 minimal medium (containing 10 mM acetate, 1 M CaCl2) to an OD600 of 0.2, and then diluted 10,000 times.

[0102] To create a standard curve: Take 100 μL of the prepared test strain dilution and add 20 μL of cefoperazone sodium and sulbactam sodium to final concentrations of 0, 10, 20, 40, 60, and 80 ng / mL, respectively. Mix the samples thoroughly and incubate them on a shaker at 37°C at 200 rpm for 10 hours. Count the viable bacteria using a plate. Draw a standard curve with bacterial cell count as the y-axis and antibiotic concentration as the x-axis.

[0103] Determination of intracellular antibiotic concentrations: Take 100 μL of the prepared test strain dilution and add 20 μL of the prepared bacterial intracellular antibiotic sample (i.e., super-ruptured liquid from the control group, super-ruptured liquid from the cefoperazone sodium and sulbactam sodium group, and super-ruptured liquid from the cefoperazone sodium and sulbactam sodium + linoleic acid group). Mix thoroughly and incubate at 37°C, 200 rpm, for 10 hours. Viable bacterial counts were determined using a plate. The antibiotic concentration in the sample was then calculated using a standard curve comparing cefoperazone sodium and sulbactam sodium to bacterial counts.

[0104] See the results Figure 10 As shown in the figure, adding linoleic acid to the antibiotics significantly increased the amount of antibiotics that entered the cells. Compared to adding the antibiotics alone, the amount of antibiotics that entered the cells increased by 9.8 times after adding linoleic acid. This result suggests that linoleic acid can increase the amount of antibiotics that enter the bacterial cells.

[0105] The above test results show that after adding linoleic acid exogenously, the cell membrane permeability of the bacteria increases, and the amount of antibiotics entering the intracellular bacteria increases. This indicates that linoleic acid increases the permeability of the bacterial membrane, thereby increasing the intracellular content of antibiotics, thereby promoting the sensitivity of bacteria to antibiotics.

[0106] Example 8 Linoleic acid can delay the post-antibiotic effect (PAE) of cefoperazone sodium and sulbactam sodium Taking Staphylococcus aureus as a representative of gram-positive bacteria and cefoperazone sodium and sulbactam sodium as a representative of antibiotics, the effect of linoleic acid combined with antibiotics was further explored.

[0107] Taking three clinical strains MSSA 2, MRSA 7 and MRSA 12 as representatives. Single bacterial colony was picked and inoculated in a 250 mL conical flask containing 50 mL LB and incubated at 37°C, 200 rpm overnight. Then the bacteria were transferred to 5 mL LB test tubes at a ratio of 1:100. When the bacteria grew to OD 600 0.2, the bacteria were collected by centrifugation and washed with physiological saline three times. The bacteria were resuspended with 5 mL MHB, and 500 μL of the above bacterial solution was added to a test tube containing 4.5 mL MHB. Each strain was divided into 5 groups: MHB control group, 1 × MIC SCF, 1 × MIC SCF + 1 mM linoleic acid, 2 × MIC SCF, 2 × MIC SCF + 1 mM linoleic acid. Each treatment had 3 biological replicates. Incubate at 37°C, 200 rpm for 2 hours. Take 100 μL of the above bacterial solution and dilute it 1000 times, then take 100 μL and add it to a 4.9 mL MHB test tube. The diluted bacterial solution was incubated at 37°C, 200 rpm. At 0, 1, 2, 4, 6, 8, 12 hours, 100 μL of the bacterial solution was taken and diluted, and then the viable bacterial count was detected by plating. All strains had 3 biological replicates. Then take the time point as the horizontal coordinate, and the average value of the logarithm of the number of colonies corresponding to it as the vertical coordinate, to construct the bacterial growth curve.

[0108] Then calculate the PAE by the bacterial growth curve, the formula is as follows: PAE = T-C, where T is the time required for the viable bacterial count in the test culture to increase by 1 Log10 CFU observed immediately after dilution, and C is the corresponding time of the control group without exposure to antibiotics. For this test, the PAE of SCF alone (i.e. PAE SCF ), the PAE of SCF + linoleic acid (PAE SCF+亚油酸 ), by comparing PAE SCF and PAE SCF+亚油酸 , to determine whether the exogenous addition of linoleic acid will prolong the PAE of the antibiotic SCF.

[0109] The growth curve results were drawn as Figure 11 . Through further calculation, the PAE of the three strains after treatment with cefoperazone sodium and sulbactam sodium alone and linoleic acid + cefoperazone sodium and sulbactam sodium under the two conditions of 1×MIC and 2×MIC were obtained. The PAE of the cefoperazone sodium and sulbactam sodium antibiotic group, i.e., the SCF group, in the table is the result of comparison between the group with the addition of antibiotics and the control group; the PAE after treatment with linoleic acid + cefoperazone sodium and sulbactam sodium, i.e., SCF + linoleic acid, in the table is the result of comparison between the addition of linoleic acid + cefoperazone sodium and sulbactam sodium and the addition of antibiotics alone. The results are summarized in Table 5. The specific situation is detailed as follows: Table 5 PAE of SCF and SCF+linoleic acid for 2 hours against three strains of Staphylococcus aureus (hours)

[0110] For MSSA 2 strains, the PAEs of cefoperazone sodium and sulbactam sodium at 1×MIC and 2×MIC were 0.15 and 2.11 hours, respectively. The PAEs of SCF+linoleic acid were prolonged compared with those of the antibiotic group, by 2.01 and 2.82 hours, respectively.

[0111] For MRSA 7, the PAEs of cefoperazone sodium and sulbactam sodium at 1×MIC and 2×MIC were 1.71 and 3.51 hours, respectively. The PAEs of SCF+linoleic acid were prolonged compared with those of the antibiotic group, by 3.44 and 4.06 hours, respectively.

[0112] For MRSA 12 strains, the PAEs of cefoperazone sodium and sulbactam sodium at 1×MIC and 2×MIC were 0.96 and 1.04 hours, respectively. The PAEs of SCF+linoleic acid were prolonged compared with those of the antibiotic group, by 1.52 and 2.37 hours, respectively.

[0113] Overall, linoleic acid can prolong the PAE of SCF, and the extension time varies among different strains.

[0114] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. The application of linoleic acid in the preparation of antibiotic synergists, characterized in that: The antibiotic is selected from quinolone antibiotics or pharmaceutically acceptable salts thereof or a compound preparation containing one or more of the antibiotics.

2. The application according to claim 1, characterized in that The linoleic acid increases the sensitivity of Gram-positive bacteria to antibiotics.

3. The use of linoleic acid in combination with antibiotics in the preparation of a drug for increasing the sensitivity of Gram-positive bacteria to antibiotics, characterized in that: The antibiotic is selected from quinolone antibiotics or pharmaceutically acceptable salts thereof or a compound preparation containing one or more of the antibiotics.

4. Use of linoleic acid in combination with antibiotics in the preparation of a medicament for preventing and / or treating Gram-positive bacterial infection, characterized in that: The antibiotic is selected from quinolone antibiotics or pharmaceutically acceptable salts thereof or a compound preparation containing one or more of the antibiotics.

5. The use according to any one of claims 1 to 4, characterized in that: The antibiotic is selected from ofloxacin or levofloxacin.

6. The use according to any one of claims 2 to 4, characterized in that: The Gram-positive bacteria are selected from one or more of Staphylococcus aureus, Streptococcus iniae, Streptococcus agalactiae, Streptococcus pyogenes, Bacillus subtilis, and Enterococcus faecalis.

7. A drug for increasing the sensitivity of Gram-positive bacteria to antibiotics, characterized in that: Contains effective amounts of linoleic acid and antibiotics; the antibiotics are selected from quinolone antibiotics or pharmaceutically acceptable salts thereof or compound preparations containing one or more of the antibiotics.

8. The medicine according to claim 7, characterized in that The antibiotic is selected from ofloxacin or levofloxacin.

9. The drug according to claim 7 or 8, characterized in that The mixing ratio of the linoleic acid and the antibiotic is 1: (2-40000) g / mol.

10. The drug according to claim 9, characterized in that The mixing ratio of the linoleic acid and the antibiotic is 1: (50-40000) g / mol.

Images