Application of idebenone in preparation of antibacterial product

Abstract

The invention discloses an application of idebenone in preparation of an antibacterial product. According to the invention, the in-vitro antibacterial activity and antibacterial spectrum of idebenone on clinical common strains are researched, and the research result shows that idebenone has an inhibiting effect on the growth of staphylococcus aureus. When the concentration of idebenone is within the range of 32-128 mu g / mL, growth of drug-resistant and sensitive staphylococcus aureus can be remarkably inhibited. Therefore, idebenone can be used for treating staphylococcus aureus infection and can be used for preparing antibacterial products.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine, and specifically relates to use of idebenone in preparing antibacterial products. Background Art

[0002] Staphylococcus aureus is a common Gram-positive conditional pathogen that often causes community or hospital-acquired infections. The pathogenic factors of Staphylococcus aureus include a variety of toxins and invasive enzymes, which can cause various types of infections, such as empyema, pneumonia, skin and soft tissue infections, implant-related infections and sepsis, etc., and severe cases can lead to death. In the 2024 WHO's bacterial priority pathogen alert list, methicillin-resistant Staphylococcus aureus (MRSA) is a high-priority pathogen. The outbreaks and hospital infections caused by MRSA not only increase the medical burden, but also seriously threaten human health. It is one of the difficult problems that the medical community still needs to solve.

[0003] According to the 2022 monitoring results of the China Antimicrobial Surveillance Network (CHINET), Staphylococcus aureus in my country has ranked first in the isolation rate of Gram-positive bacteria. Due to the abuse of antimicrobial drugs, the number of strains of drug-resistant Staphylococcus aureus has increased significantly. Among them, MRSA is only effective against vancomycin. It has stronger drug resistance and higher mortality rate, and its clinical detection rate is on the rise. In the 1980s, methicillin-resistant Staphylococcus aureus has become the main pathogen of nosocomial infections. In recent years, vancomycin-resistant Staphylococcus aureus has also appeared in Japan and the United States.

[0004] Lycopene has been shown to have antioxidant activity. By eliminating free radicals, it can inhibit the release of α-hemolysin and lactate dehydrogenase by Staphylococcus aureus, thereby inhibiting the pathogenicity of Staphylococcus aureus. 10 It is derived from human body synthesis and external intake. It has been proven to have antioxidant and free radical scavenging capabilities and can inhibit the growth of Staphylococcus aureus. As an antioxidant, idebenone also has the characteristics of limiting the formation of free radicals and is often used to treat moderate Alzheimer's disease and dementia caused by cerebrovascular diseases.

[0005] Currently, the effect of idebenone in inhibiting Staphylococcus aureus has not been reported. Summary of the invention

[0006] Purpose of the invention: The purpose of the present invention is to provide a use of idebenone in the preparation of antibacterial products. The present invention studies the in vitro antibacterial activity and antibacterial spectrum of idebenone against common clinical strains, and the results show that idebenone has an inhibitory effect on the growth of Staphylococcus aureus.

[0007] The present invention has found that idebenone concentrations in the range of 32 to 128 μg / mL can significantly inhibit the growth of drug-resistant and sensitive Staphylococcus aureus. The drug concentration can reach a peak in the blood 15 minutes after oral administration, and has no long-term accumulation effect in the host. It is safe and well tolerated at a dose of up to 2250 mg / d. Therefore, idebenone can be used to treat Staphylococcus aureus infections and can be used to prepare antibacterial products.

[0008] Technical solution: The purpose of the present invention is achieved through the following technical solution:

[0009] The invention provides use of idebenone in preparing an antibacterial product.

[0010] The antibacterial products include antibacterial drugs, antibacterial daily chemicals, and food additives.

[0011] The antibacterial drug comprises idebenone and a pharmaceutically acceptable carrier or excipient.

[0012] The auxiliary materials include one or more of emulsifiers, disintegrants, wetting agents, diluents, antioxidants, preservatives or adhesives.

[0013] The emulsifier is selected from at least one of Tweens, Spans, glycerol fatty acid esters, pectin, agar, sodium alginate or silicon dioxide.

[0014] The disintegrant is selected from at least one of starch, sodium carboxymethyl starch, cross-linked polyvinyl pyrrolidone, low-substituted hydroxypropyl cellulose or cross-linked polyvinyl pyrrolidone.

[0015] The wetting agent is selected from at least one of water and ethanol.

[0016] The diluent is selected from at least one of starch, sugar, cellulose or inorganic salts.

[0017] The antioxidant is selected from at least one of ascorbic acid, sulfite, bisulfite, gallic acid and lipids thereof.

[0018] The preservative is selected from at least one of benzoic acid and its salts, sorbic acid and its salts or parabens.

[0019] The binder is selected from at least one of starch slurry, sodium carboxymethyl cellulose, povidone, hydroxypropyl cellulose, methyl cellulose or ethyl cellulose.

[0020] The dosage form of the antibacterial drug is injection, tablet, capsule, oral liquid, microparticle or ointment.

[0021] The antibacterial daily chemical product is a facial cleanser, hand soap, hand cream, cosmetics or a women's care lotion.

[0022] The drug of the present invention can be administered in various known ways, such as oral administration, injection, transdermal administration, etc. The drug of the present invention can be administered alone or in combination with other drugs. The oral composition can be any oral acceptable dosage form, including but not limited to tablets, capsules, oral solutions and microparticles.

[0023] Sterile injectable compositions can be formulated according to techniques known in the art using suitable dispersants or wetting agents and suspending agents. Pharmaceutically acceptable carriers and solvents that can be used include water, sodium chloride solution, and the like.

[0024] The medicine of the present invention can be made into common preparations, and can also be made into sustained-release preparations, controlled-release preparations, targeted preparations and various microparticle drug delivery systems.

[0025] The idebenone of the present invention can be used as the sole active ingredient for preparing antibacterial products, or can be used in combination with other active ingredients for preparing antibacterial products.

[0026] Actual dosage levels of the active ingredients in the medicaments of the present invention may be varied to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient. The selected dosage level will depend upon a variety of factors, including route of administration, time of administration, rate of excretion, duration of treatment, other drugs, compounds, and / or materials used in combination with idebenone, age, sex, weight, general health and past medical history of the patient being treated, and similar factors well known in the medical arts.

[0027] The antibacterial product of the present invention is a product for inhibiting Staphylococcus aureus.

[0028] The idebenone significantly inhibits the growth of drug-resistant and sensitive Staphylococcus aureus in the range of 32 to 128 μg / mL.

[0029] Beneficial effects:

[0030] The idebenone of the present invention can be used as an active ingredient for treating Staphylococcus aureus infection, which opens up new uses of idebenone and provides a new option for preparing antibacterial products. The idebenone of the present invention has an inhibitory effect on the growth of Staphylococcus aureus and can therefore be used for preparing antibacterial products. DETAILED DESCRIPTION

[0031] The technical solution of the present invention is described in detail below through specific embodiments, but the protection scope of the present invention is not limited to the embodiments.

[0032] If no specific techniques or conditions are specified in the examples, the techniques or conditions described in the literature in the field or the product instructions are used. If no manufacturer is specified for the reagents or instruments used, they are all conventional products that can be purchased through regular channels.

[0033] The experimental methods in the following examples are conventional methods unless otherwise specified. The experimental materials used in the following examples are commercially available products unless otherwise specified.

[0034] Idebenone purity ≥98%, CAS number: 58186-27-9.

[0035] Example 1 Idebenone in vitro antibacterial activity test

[0036] 1. Experimental Materials and Instruments

[0037] 35℃ constant temperature incubator, sterile cotton swabs, Thermo Fisher turbidimeter and turbidimetric tubes, prepared LB broth medium (Shanghai Bowei, prepared according to the product instructions), pipette and matching pipette tip, 50mL centrifuge tube, electronic balance, drug sensitivity plate, pipette, and biological safety cabinet.

[0038] The Mérieux Vitek 2XL fully automatic microbial identification and drug sensitivity analyzer was used to identify drug-resistant and sensitive strains from 113 strains of clinical sample isolates. The clinical sample isolates in this experiment were from the First People's Hospital of Huai'an City. Table 1 shows the classification of the 113 clinical sample isolates:

[0039] Table 1 Classification of 113 strains isolated from clinical samples

[0040]

[0041] 2. Experimental methods and steps

[0042] 1. Strain recovery

[0043] (1) Disinfect the biosafety cabinet with ultraviolet light for 0.5 hours before the experiment;

[0044] (2) Take the clinical strain to be tested out of the -80°C freezer;

[0045] (3) Use a sterile inoculating loop to take the frozen bacterial mass in the test tube and inoculate it onto a Columbia blood agar plate;

[0046] (4) The inoculated blood plate was incubated at 35±2℃ for 24 hours.

[0047] 2. Strain Purification

[0048] (1) Disinfect the biosafety cabinet with ultraviolet light for 0.5 hours before the experiment;

[0049] (2) taking out the clinical strain to be tested that has been revived in the incubator;

[0050] (3) Use a disposable sterile inoculation loop to pick up a single colony and continue to inoculate it onto a new Columbia blood agar plate;

[0051] (4) The inoculated blood plate was incubated at 35±2℃ for 24 hours.

[0052] 3. Preparation of drug sensitivity test plate

[0053] (1) Dissolve 20.48 mg of idebenone in 0.4 mL of dimethyl sulfoxide (DMSO) to prepare a liquid with a concentration of 51.2 mg / mL, store it in a 10 mL centrifuge tube, and number the mother solution;

[0054] (2) Add 0.2 mL of the mother solution to a centrifuge tube containing 40 mL of LB broth to dilute the idebenone concentration to 256 μg / mL, number 12;

[0055] (3) Take 10 50 mL centrifuge tubes and add 20 mL of LB broth medium to each tube, numbering them from 2 to 11;

[0056] (4) Pipette 20 mL of liquid from centrifuge tube No. 12, add it to centrifuge tube No. 11, and mix thoroughly to make the concentration reach 128 μg / mL;

[0057] (5) Serial dilutions were performed according to the above method to make the final drug concentrations 64, 32, 16, 8, 4, 2, 1, 0.5, and 0.25 μg / mL, respectively;

[0058] (6) Use a pipette to draw 100 μL of each concentration of drug solution and add it to the corresponding wells of the 96-well plate. Add 100 μL of LB broth culture medium to the first column as a blank control. The concentration settings of the drug sensitivity test plate are shown in Table 2:

[0059] Table 2 Drug sensitivity test plate concentration settings (unit: μg / mL)

[0060] 1 column 2 columns 3 columns 4 columns 5 columns 6 columns 7 columns 8 columns 9 columns 10 columns 11 columns 12 columns A 0 0.25 0.5 1 2 4 8 16 32 64 128 256 B 0 0.25 0.5 1 2 4 8 16 32 64 128 256 C 0 0.25 0.5 1 2 4 8 16 32 64 128 256 D 0 0.25 0.5 1 2 4 8 16 32 64 128 256 E 0 0.25 0.5 1 2 4 8 16 32 64 128 256 F 0 0.25 0.5 1 2 4 8 16 32 64 128 256 G 0 0.25 0.5 1 2 4 8 16 32 64 128 256 H 0 0.25 0.5 1 2 4 8 16 32 64 128 256

[0061] (7) Since DMSO at high concentrations can also inhibit bacterial growth, in order to eliminate the interference of DMSO on the interpretation of experimental results, the same method was used to prepare DMSO dilutions, whose concentrations were consistent with the DMSO content in the wells shown in Table 2, namely the DMSO control plate compared with idebenone.

[0062] 4. In vitro antibacterial activity and antibacterial spectrum test of idebenone

[0063] (1) Preparation of bacterial suspension: Use a micropipette to take an appropriate amount of physiological saline and add it to a turbidimetric tube. Pick several pure colonies from the plate of the strain to be tested (purified blood plate), grind and mix them on the inner wall of the turbidimetric tube, and use a turbidimeter to prepare a bacterial suspension with a turbidity of 0.5 for later use;

[0064] (2) Inoculation: Use a micropipette to take 20 μL of the bacterial suspension prepared in step (1) and add it to 3.2 mL of LB broth and mix well;

[0065] (3) Sample addition: Add the bacterial solution obtained in step (2) to the above-mentioned drug-sensitive plate and DMSO control plate using a micropipette at a volume of 100 μL per well, and cover the 96-well plate with drugs and bacteria with a cover plate. The drug concentrations of each well in the drug-sensitive plate after adding the bacterial solution are shown in Table 3.

[0066] (4) Adding samples to the control plate: Similar to step (3), add bacterial solution of equal concentration to each well of the DMSO control plate in sequence.

[0067] Table 3 Drug concentrations of drug sensitivity plate and control plate (unit: μg / mL)

[0068] 1 column 2 columns 3 columns 4 columns 5 columns 6 columns 7 columns 8 columns 9 columns 10 columns 11 columns 12 columns A 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 B 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 C 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 D 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 E 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 F 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 G 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128 H 0 0.125 0.25 0.5 1 2 4 8 16 32 64 128

[0069] (5) Cultivation: Place the above-mentioned drug-sensitive plates and control plates in a constant temperature incubator at 35±2°C for 24 hours.

[0070] Turbidity in the drug-coated wells was considered to indicate bacterial growth, and the lowest coating concentration corresponding to no turbidity was the minimum inhibitory concentration (MIC value) of the drug.

[0071] 3. Experimental Results

[0072] The MIC distribution results of idebenone against five common clinical bacteria are shown in Table 4:

[0073] Table 4 MIC distribution of idebenone against five common clinical bacteria (μg / mL)

[0074]

[0075] Note: MIC: minimum inhibitory concentration; MIC 50 and MIC 90 : MIC required for idebenone to inhibit the growth of 50% and 90% of the tested bacteria respectively; MIC GM: Geometric mean of the minimum inhibitory concentration of idebenone for each strain

[0076] The MIC distribution results of idebenone against Staphylococcus aureus are shown in Table 5:

[0077] Table 5 MIC distribution of idebenone against Staphylococcus aureus (μg / mL)

[0078]

[0079] Note: MIC: minimum inhibitory concentration; MIC 50 and MIC 90 : MIC required for idebenone to inhibit the growth of 50% and 90% of the tested bacteria respectively; MIC GM: Geometric mean of the minimum inhibitory concentration of idebenone for each strain

[0080] From the results in Table 4 and Table 5, we can find that:

[0081] 1. Sensitivity of idebenone to Staphylococcus aureus:

[0082] Among 113 strains of S. aureus , 92.5% of clinical strains were sensitive to idebenone;

[0083] Among the sensitive S. aureus , 95.8% were sensitive to idebenone;

[0084] Among the resistant Staphylococcus aureus, 83.4% were sensitive to idebenone;

[0085] Other strains were not sensitive to idebenone.

[0086] 2. The influence of DMSO, the solvent of idebenone:

[0087] DMSO is used as a solvent for idebenone. In the drug sensitivity control plate, the growth of strains in the wells corresponding to each concentration was not significantly different from that in the negative control wells (without drug and DMSO), and all had bacterial growth. This indicates that DMSO does not affect the sensitivity detection of idebenone.

[0088] Since most of the Staphylococcus aureus were sensitive to idebenone, it can be inferred that idebenone has an inhibitory effect on the growth of Staphylococcus aureus. These findings are very important for the potential application of idebenone in the clinical treatment of Staphylococcus aureus infections. This suggests that idebenone may be an effective antibacterial agent, especially for those Staphylococcus aureus that are resistant to traditional antibiotics.

[0089] As described above, although the present invention has been shown and described with reference to specific preferred embodiments, it should not be construed as limiting the present invention itself. Various changes in form and details may be made without departing from the spirit and scope of the present invention as defined in the appended claims.

Claims

1. Use of idebenone in the preparation of antibacterial products.

2. The use according to claim 1, characterized in that The antibacterial products include antibacterial drugs, antibacterial daily chemicals, and food additives.

3. The use according to claim 2, characterized in that: The dosage form of the antibacterial drug is injection, tablet, capsule, oral liquid, microparticle or ointment.

4. The use according to claim 2, characterized in that: The antibacterial daily chemical product is a facial cleanser, hand soap, hand cream, cosmetics or a women's care lotion.

5. The use according to claim 1, characterized in that: Idebenone is used as the sole active ingredient in the preparation of antibacterial products.

6. The use according to claim 1, characterized in that: Idebenone is used in combination with other active ingredients to prepare antibacterial products.

7. The use according to claim 1, characterized in that The antibacterial product is a product that inhibits Staphylococcus aureus.

8. The use according to claim 1, characterized in that The idebenone significantly inhibits the growth of drug-resistant and sensitive Staphylococcus aureus in the range of 32 to 128 μg / mL.

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