Antimicrobial composition, preparation method therefor, and use thereof

Abstract

Disclosed is an antimicrobial composition, comprising the following components in parts by weight: 4-6 parts of vitamin C, 1-2 parts of vitamin E, 1-2 parts of angelica polysaccharides, and 1-2 parts of Ligustri lucidi fructus polysaccharides. Further disclosed are a preparation method for the antimicrobial composition and use thereof in the preparation of a drug for inhibiting bacteria and / or fungi or treating inflammation.

Description

An antibacterial composition, its preparation method and application Technical Field

[0001] This invention belongs to the field of medicine, specifically relating to an antibacterial composition, its preparation method, and its application. Background Technology

[0002] Infection is a pathological state in which pathogens (such as bacteria, viruses, and fungi) invade the body, triggering an immune system response. During infection, inflammatory factors in peripheral blood, as immunomodulatory molecules, play a crucial role in the host's defense response. The dynamic changes in inflammatory factors can reflect the intensity and characteristics of the body's response to infection, making peripheral blood testing an important tool for assessing infectious diseases. In some pathogen infections, the excessive release of inflammatory factors can lead to a "cytokine storm," subsequently triggering systemic inflammatory response syndrome (SIRS) and multiple organ dysfunction syndrome (MODS). In patients with sepsis caused by bacterial infections, the levels of TNF-α and IL-6 in peripheral blood are significantly elevated, closely related to disease severity and prognosis. Novel coronavirus infection can trigger the excessive release of pro-inflammatory factors (such as IL-6 and interferon-γ), leading to lung injury and acute respiratory distress syndrome (ARDS). Fungal infections, such as Candida albicans infection, trigger a systemic inflammatory response by activating the release of pro-inflammatory factors TNF-α and IL-1β.

[0003] Bacterial infections have become a major global public health challenge. In 2019, bacterial infections caused approximately 7.7 million deaths, accounting for 13.6% of all deaths worldwide, making it the second leading cause of death globally after ischemic heart disease. Staphylococcus aureus, Escherichia coli, Streptococcus pneumoniae, Klebsiella pneumoniae, and Pseudomonas aeruginosa are the leading pathogens causing these deaths. The types of infections caused by these bacteria include lower respiratory tract infections, bloodstream infections, and intra-abdominal infections. Antimicrobial resistance (AMR) is a growing problem, making the treatment of bacterial infections more difficult. In 2022, the World Health Organization reported high levels of resistance in bacteria causing fatal bloodstream infections, and increasing resistance to treatment in several bacteria commonly causing community infections. Bacterial infections pose a significant threat to global health, especially against the backdrop of rising antimicrobial resistance. Strengthening infection prevention, optimizing antibiotic use, improving microbial detection capabilities, and developing novel antimicrobial drugs and vaccines are crucial to mitigating the burden of bacterial infections.

[0004] However, current antibacterial agents still suffer from insufficient efficacy and stability. Many traditional antibacterial agents have a single source of active ingredient and a relatively simple formulation structure. Traditional antibacterial regimens often rely on the single use of a chemical drug or specific herbal extract, neglecting the potential synergistic effects between nutrients, antioxidants, and herbal monomers. The lack of multi-level synergistic effects in the process of active ingredients exerting their effects in vivo results in insufficient comprehensive regulatory capacity under physiological conditions, leading to less stable or significant clinical efficacy than expected. Furthermore, there may be certain side effects and safety concerns. Some commonly used antibacterial drugs are prone to adverse reactions such as gastrointestinal irritation and increased burden on the liver and kidneys with long-term use. This is because some chemical drugs have potential irritant or damaging effects on corresponding organs during metabolism and excretion. Moreover, existing antibacterial agents have relatively simple mechanisms of action and lack overall regulatory capacity. Traditional antibacterial drugs often focus on specific steps in the antibacterial response, but the infection process is complex and not limited to a single pathway. The lack of a comprehensive regulatory strategy involving multiple targets and pathways may prevent drugs from providing comprehensive and sustained relief from infection in actual use. Summary of the Invention

[0005] One object of the present invention is to provide an antibacterial composition with a sustained, mild and highly effective antibacterial effect in response to the above-mentioned technical problems.

[0006] Another object of the present invention is to provide a method for preparing the antibacterial composition.

[0007] Another object of the present invention is to provide the application of the said antibacterial composition.

[0008] To achieve the above-mentioned objectives, the present invention provides an antibacterial composition comprising, by weight, the following components: 4-6 parts of vitamin C, 1-2 parts of vitamin E, 1-2 parts of Angelica sinensis polysaccharide, and 1-2 parts of Ligustrum lucidum polysaccharide.

[0009] In a preferred embodiment, the weight ratio of vitamin C, vitamin E, angelica polysaccharide and privet polysaccharide is 5:1:1:1.

[0010] In a preferred embodiment, the antibacterial composition further includes a solvent. Preferably, the solvent is physiological saline, glucose solution, or other injectable solvent, such as water.

[0011] In a preferred embodiment, the antibacterial composition further includes a stabilizer. More preferably, the stabilizer is an antioxidant. More preferably, the antioxidant is selected from one or more of tropenoic acid, panthenol, and butylated hydroxyanisole (BHT).

[0012] On the other hand, the present invention also provides a method for preparing the antibacterial composition, the method comprising the following steps:

[0013] (1) Add Angelica sinensis polysaccharide and Ligustrum lucidum polysaccharide to water and stir to dissolve;

[0014] (2) Add vitamin C powder and stir until completely dissolved to obtain a mixed solution;

[0015] (3) Mix vitamin E with emulsifier and vegetable oil, stir evenly, and then slowly add it to the above mixed solution, stir to homogenize, so that it is evenly dispersed and forms a stable dispersion;

[0016] (4) The above dispersion is dried, pulverized and sieved to obtain the antibacterial composition.

[0017] On the other hand, the present invention also provides the use of the antibacterial composition in the preparation of a medicament for inhibiting bacteria and / or fungi.

[0018] In a preferred embodiment, the bacteria are Gram-negative or Gram-positive bacteria.

[0019] In a preferred embodiment, the bacteria are one or more of the following: Gram-negative bacteria including Escherichia coli, Shigella flexneri, Shigella sonnei, and Klebsiella pneumoniae; Gram-positive bacteria including Staphylococcus aureus; and fungi including Candida albicans.

[0020] On the other hand, the present invention also provides the use of the antibacterial composition in the preparation of medicaments for treating inflammation.

[0021] Preferably, the antibacterial composition of the present invention can be prepared in a variety of known dosage forms, including but not limited to solid dosage forms, semi-solid dosage forms, liquid dosage forms, gaseous and aerosol dosage forms, implantable dosage forms, and special dosage forms.

[0022] Preferably, the antibacterial composition of the present invention can be administered in a variety of different routes, including but not limited to oral administration, injection administration, transdermal administration, inhalation administration, topical administration, rectal administration, sublingual or oral administration, implantation administration, and administration via special routes.

[0023] Preferably, the antibacterial composition of the present invention can be administered at a dose of about 8-80 mg / kg body weight / day, etc.

[0024] In a preferred embodiment, the inflammation includes inflammation caused by immune responses, cytokine activation, or oxidative damage, such as rheumatoid arthritis, arteriosclerosis, hepatitis, and skin diseases.

[0025] Vitamin C is a water-soluble antioxidant that can directly scavenge reactive oxygen species (ROS) and inhibit the release of inflammatory factors. Vitamin C supplementation can reduce the levels of inflammatory markers C-reactive protein (CRP) and IL-6, improving various chronic inflammation-related diseases. Liposome encapsulation of vitamin C regulates the inflammatory factors IL-6, MCP-1, and MIP-1α, and has been found to significantly improve immune function. Vitamin C plays an important role in reducing pro-inflammatory factors and controlling inflammation. Vitamin C (ascorbic acid) also has certain effects in immune regulation and antibacterial activity.

[0026] Vitamin E is a fat-soluble antioxidant that protects cell membranes from oxidative damage by preventing lipid peroxidation. Vitamin E has shown significant effects in reducing TNF-α and IL-6 levels, especially under oxidative stress. Its effects on pro-inflammatory factors in patients with schizophrenia have revealed significant roles in both antioxidant and inflammatory regulation. Vitamin E has potential value in reducing postprandial inflammatory responses, particularly its ability to regulate inflammatory factors. Members of the vitamin E family have protective effects against radiation-induced inflammation by effectively mitigating radiation damage through the regulation of inflammatory factor expression. Furthermore, vitamin E can enhance the bactericidal effect of antibiotics by inhibiting the binding of bacterial lipoproteins (such as BcnA produced by Burkholderia cenocepacia).

[0027] Angelica polysaccharides are active ingredients extracted from the traditional Chinese medicine Angelica sinensis, and they have a wide range of functions. In vitro, Angelica polysaccharides show certain inhibitory effects on various tumor cell types, and the mechanism may be related to inducing apoptosis and inhibiting tumor cell proliferation. Angelica polysaccharides also have a certain hypoglycemic effect in diabetic rats, and the mechanism may be through improving pancreatic islet function and regulating insulin sensitivity. As a multifunctional drug, Angelica polysaccharides have been proven to have a wide range of pharmacological effects, including antioxidant, immunomodulatory, and hematopoietic-promoting effects. Angelica polysaccharides can significantly promote the proliferation and differentiation of hematopoietic stem cells and enhance the hematopoietic function of bone marrow.

[0028] The chemical composition of *Fructus ligustri lucidi* polysaccharides includes polysaccharides, phenolic acids, and triterpenoids. Research on these polysaccharides covers multiple effects, including anti-aging, immunomodulation, and anti-tumor activity. Specific mechanisms include free radical scavenging, enhancing immune cell function, and regulating tumor-related factors. *Fructus ligustri lucidi* polysaccharides exhibit significant anti-aging effects, inhibiting the degeneration of immune organs, enhancing immune function, and scavenging free radicals. They also have inhibitory effects on certain tumor cells, possibly involving enhanced immune function and inhibition of tumor cell proliferation. Furthermore, they can improve immunity by regulating T cell activity and enhancing macrophage function. The synergistic use of *Fructus ligustri lucidi* polysaccharides and *Cuscuta chinensis* polysaccharides can significantly enhance anti-aging effects, possibly through enhanced antioxidant capacity. Finally, *Fructus ligustri lucidi* polysaccharides can increase the antigenicity of lymphoma cell membranes, thereby enhancing the body's immune response to tumors.

[0029] By rationally combining vitamin C, vitamin E, and angelica polysaccharides and privet polysaccharides in a specific ratio, and utilizing their different bactericidal and anti-inflammatory mechanisms to create a synergistic effect, the composition of this invention achieves a complex antibacterial effect, improving the overall antibacterial and anti-inflammatory efficacy and stability, thereby obtaining better therapeutic and preventive effects in clinical applications. Experiments have shown that the composition of this invention can significantly reduce the concentration of peripheral blood inflammatory factors (such as TNF-α and IL-6) and promote peripheral blood bactericidal activity. This multi-pathway synergistic effect provides a more comprehensive bactericidal and anti-inflammatory effect compared to traditional single-drug therapy.

[0030] Compared with existing technologies, this invention improves the stability and persistence of the overall antibacterial effect. Utilizing the synergistic effect of multiple components, it enhances the comprehensive regulatory capacity of the active ingredients in vivo, resulting in improved stability and efficacy of the final formulation. Furthermore, this invention reduces the risk of potential adverse reactions and side effects. In selecting and combining ingredients, the biosafety of both traditional Chinese medicine monomers and vitamins is considered to minimize gastrointestinal, hepatic, and renal irritation and damage that may be caused by traditional chemical antibacterial drugs. Moreover, this invention further enriches the antibacterial mechanism of action and enhances overall regulatory capacity. Through the multi-stage intervention of vitamins and traditional Chinese medicine monomers, it aims to achieve a moderate balance and regulation of multiple signaling pathways in the antibacterial process, thereby obtaining a more comprehensive, lasting, and gentle antibacterial effect in clinical practice. Attached Figure Description

[0031] Figure 1 shows the colony growth of Escherichia coli.

[0032] Figure 2 shows the relative number of live bacteria in the Escherichia coli culture experiment.

[0033] Figure 3 shows the colony growth of Shigella flexneri.

[0034] Figure 4 shows the relative number of live bacteria in the Shigella flexneri culture experiment.

[0035] Figure 5 shows the concentration of IL-6 cytokine in plasma after the addition of E. coli.

[0036] Figure 6 shows the concentration of IL-6 cytokine in plasma after the addition of Shigella flexneri. Detailed Implementation

[0037] The present invention will be further described below with reference to specific embodiments. It should be understood that the following embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0038] Vitamin C is a commercially available raw material, purchased from Shanghai Beyotime Biotechnology Co., Ltd., product number ST1434.

[0039] Vitamin E is a commercially available raw material, purchased from Shanghai Beyotime Biotechnology Co., Ltd., product number S0079.

[0040] Angelica polysaccharide is a commercially available raw material purchased from Shanghai Yuanye Biotechnology Co., Ltd., product number S27815.

[0041] Privet polysaccharide is a commercially available raw material, purchased from Shanghai Ronghe Pharmaceutical Technology Development Co., Ltd., product number TDT044.

[0042] Unless otherwise specified, the reagents, instruments, etc. used in the embodiments of the present invention are all well known to those skilled in the art and can be purchased through commercial channels.

[0043] Example

[0044] 1. Preparation of antibacterial compositions (taking the preparation of solid compositions as an example)

[0045] (1) Add 2g of Angelica sinensis polysaccharide and 2g of Ligustrum lucidum polysaccharide to 200mL of purified water and stir until dissolved. If necessary, the solution can be gently heated in a water bath at 40-60℃ to accelerate the dissolution and dispersion of the polysaccharides until a homogeneous polysaccharide solution is obtained. The pH and temperature of the polysaccharide solution can be appropriately controlled according to the characteristics of the polysaccharides and the requirements of subsequent steps, usually within a neutral or slightly acidic range, such as pH 6-7.

[0046] (2) Slowly add 10g of vitamin C (L-ascorbic acid) to the above polysaccharide solution while stirring continuously until the vitamin C is completely dissolved, to obtain a clear, homogeneous polysaccharide-vitamin C mixed solution. Vitamin C is water-soluble and usually dissolves easily.

[0047] (3) Mix 2g of vitamin E with 1g of food-grade emulsifier (such as lecithin) and 1mL of vegetable oil, and disperse using high-shear stirring or ultrasonication to form a uniform emulsion. Then slowly add the mixture dropwise to the polysaccharide-vitamin C mixed solution while stirring to form a stable and dispersed emulsion system of vitamin E. Other similar methods can also be used to ensure that vitamin E is in a relatively uniform and stable dispersion state in the system. The emulsion system can be homogenized or stirred to ensure that the oil phase (vitamin E) in the system is refined and stably distributed in the aqueous phase.

[0048] (4) The uniformly dispersed liquid obtained above is spray-dried or vacuum freeze-dried to obtain a solid powder composition containing vitamin C, vitamin E, angelica polysaccharide and privet polysaccharide.

[0049] (5) The dried solid powder composition is pulverized and sieved to obtain the final product. Depending on the final purpose, it can be further processed into capsule filling, tablet compression or granular formulation.

[0050] 2. Bacterial culture

[0051] To investigate and evaluate the in vitro antibacterial efficacy of the antibacterial composition of the present invention in inhibiting bacterial growth, bacterial culture experiments were conducted.

[0052] Experimental groups: a total of four groups. (1) Blank control group: physiological saline; (2) Complex vitamin group: a complex of vitamin C and vitamin E with a weight ratio of 5:1; (3) Traditional Chinese medicine polysaccharide group: a complex of angelica polysaccharide and privet polysaccharide with a weight ratio of 1:1; (4) Combined drug group: the combination of vitamin C, vitamin E, angelica polysaccharide and privet polysaccharide prepared above, with a weight ratio of vitamin C, vitamin E, angelica polysaccharide and privet polysaccharide of 5:1:1:1.

[0053] Escherichia coli (Beina Biotechnology, catalog number BNCC336902) and Shigella flexneri (Beina Biotechnology, catalog number BNCC232380) were separately cultured in LB broth (Shanghai Beyotime Biotechnology Co., Ltd., catalog number ST163) overnight at 37°C. The bacterial concentration was then adjusted to 1.0 McFarland concentration with physiological saline to obtain bacterial suspensions.

[0054] 3. Bacterial stimulation of peripheral blood test

[0055] To investigate and evaluate the antibacterial and anti-inflammatory effects of the antibacterial composition of the present invention on peripheral blood, a bacterial stimulation peripheral blood experiment was conducted.

[0056] Peripheral blood was collected from six healthy adult subjects using glass tubes containing lithium heparin anticoagulant. 500 μL of anticoagulated whole blood was transferred to sterile, additive-free glass tubes and divided into four groups. Different combinations of the above-mentioned groups were added to each group (blank control group: the same volume of physiological saline as the anticoagulated whole blood; multivitamin group: 60 μg / ml; traditional Chinese medicine polysaccharide group: 20 μg / ml; combined medication group: 80 μg / ml), and the mixtures were thoroughly mixed.

[0057] Add 50 μL of bacterial suspension (Escherichia coli or Shigella flexneri) to the glass tube containing the above-mentioned composition and whole blood, and mix thoroughly. Incubate at 37°C for 2 hours.

[0058] Take 50 μL of whole blood containing the above composition and bacterial suspension, spread it onto a 10 cm plate containing nutrient broth (LB) solid medium (Shanghai Beyotime Biotechnology Co., Ltd., catalog number ST163), incubate overnight at 37°C, and count the number of bacterial monoclonal cells on the plate to estimate the relative number of live bacteria in the whole blood.

[0059] The experimental results are shown in Figures 1 to 4. Figure 1 shows the colony growth of *E. coli*. Figure 2 shows the relative number of viable bacteria in the *E. coli* culture experiment. Figure 3 shows the colony growth of *Shigella flexneri*. Figure 4 shows the relative number of viable bacteria in the *Shigella flexneri* culture experiment. Figures 1 to 4 show that the compound vitamin group significantly promoted peripheral blood bactericidal activity, the traditional Chinese medicine polysaccharide group had no significant effect, and the combined drug group significantly promoted peripheral blood bactericidal activity. Compared with the compound vitamin group, the combined drug group showed a more significant effect in promoting peripheral blood bactericidal activity. This indicates that the antibacterial composition of the present invention has a good bactericidal effect, especially against *E. coli* and *Shigella flexneri*. In addition, the antibacterial composition of the present invention also has a significant inhibitory effect on Gram-negative bacteria such as *Shigella sonnei* and *Klebsiella pneumoniae*, Gram-positive bacteria such as *Staphylococcus aureus*, and fungi such as *Candida albicans*, and the effect is better than that of the compound vitamin group and the traditional Chinese medicine polysaccharide group.

[0060] The concentration of IL-6 cytokine in plasma was detected using an electrochemiluminescence immunoassay on a Roche cobas e 601 electrochemiluminescence immunoassay analyzer.

[0061] The results of IL-6 cytokine concentration detection are shown in Figures 5 and 6. Figure 5 shows the plasma IL-6 cytokine concentration after the addition of *E. coli*, and Figure 6 shows the plasma IL-6 cytokine concentration after the addition of *Shigella flexneri*. The results indicate that the multivitamin group significantly inhibited the secretion of the bacterial-induced inflammatory cytokine IL-6 in peripheral blood, while the traditional Chinese medicine polysaccharide group had no significant effect. The combined treatment group significantly inhibited the secretion of the bacterial-induced inflammatory cytokine IL-6 in peripheral blood. Compared with the multivitamin group, the combined treatment group showed a more significant inhibition of the secretion of the bacterial-induced inflammatory cytokine IL-6 in peripheral blood.

[0062] It is evident that the antibacterial composition of the present invention has significantly better anti-inflammatory effects.

Claims

1. An antibacterial composition, characterized by By weight, it includes the following components: 4-6 parts vitamin C, 1-2 parts vitamin E, 1-2 parts angelica polysaccharide, and 1-2 parts privet polysaccharide.

2. The antimicrobial composition according to claim 1, characterized in that, The weight ratio of vitamin C, vitamin E, angelica polysaccharide and privet polysaccharide is 5:1:1:

1.

3. The antimicrobial composition according to claim 1, wherein The antibacterial composition also includes a solvent.

4. The antimicrobial composition according to claim 3, wherein The solvent is water.

5. The antibacterial composition according to claim 1, characterized in that, The antibacterial composition also includes a stabilizer.

6. The antibacterial composition according to claim 5, characterized in that, The stabilizer is an antioxidant.

7. The method for preparing the antibacterial composition according to any one of claims 1 to 6, characterized in that... Includes the following steps: (1) Add Angelica sinensis polysaccharide and Ligustrum lucidum polysaccharide to water and stir to dissolve; (2) Add vitamin C powder and stir until completely dissolved to obtain a mixed solution; (3) Mix vitamin E with emulsifier and vegetable oil, stir evenly, and then slowly add it to the above mixed solution, stir to homogenize, so that it is evenly dispersed and forms a stable dispersion; (4) The above dispersion is dried, pulverized and sieved to obtain the antibacterial composition.

8. Use of the antimicrobial composition according to any one of claims 1 to 6 in the preparation of a medicament for inhibiting bacteria and / or fungi.

9. The application according to claim 8, characterized in that, The bacteria are one or more of the following: Gram-negative bacteria including Escherichia coli, Shigella flexneri, Shigella sonnei, and Klebsiella pneumoniae; Gram-positive bacteria including Staphylococcus aureus; and fungi including Candida albicans.

10. Use of the antibacterial composition according to any one of claims 1 to 6 in the preparation of a medicament for treating inflammation.

Images