Method and device for inhibiting bacterial biofilm by driving nano-magnetic particles with magnetic field
By using a magnetic field to drive nanoparticles, the movement of nanoparticles under the influence of a magnetic field, in conjunction with vancomycin, effectively inhibits bacterial biofilms, solving the problem of eliminating drug-resistant strains with traditional antibiotics, and is applicable to the field of wound healing.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- GENERAL HOSPITAL OF THE NORTHERN WAR ZONE OF THE CHINESE PEOPLES LIBERATION ARMY
- Filing Date
- 2024-11-21
- Publication Date
- 2026-05-26
AI Technical Summary
Existing technologies are ineffective at removing bacterial biofilms, especially against drug-resistant strains, where traditional antibiotics are not very effective at inhibiting bacterial growth.
The method employs magnetic field-driven nanoparticles. By adding nanoparticles and vancomycin at a level lower than that that inhibits bacterial biofilm formation to the bacterial culture medium, and generating a uniform magnetic field in a power source with a coil, the movement of the nanoparticles is controlled to inhibit the formation of bacterial biofilm.
It achieves effective elimination of drug-resistant bacteria. The magnetic field-driven nanoparticle device is small in size and easy to use, making it suitable for wound healing and solving the problem of drug resistance caused by traditional antibiotics.
Smart Images
Figure CN122075684A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of biomedical technology, specifically relating to a method and apparatus for inhibiting bacterial biofilms by magnetically driven nanoparticles. Background Technology
[0002] Bacterial resistance is the phenomenon of bacteria becoming insensitive to antimicrobial drugs, a specific manifestation of bacterial survival processes. Bacterial resistance can be divided into endogenous resistance and acquired resistance. Endogenous resistance refers to the natural resistance to a particular antimicrobial drug due to the bacteria's unique structure. Acquired resistance mainly occurs through two pathways: chromosomal mutation or horizontal transfer of exogenous mobile genetic elements. These pathways lead to drug inactivation / alteration, changes in drug binding sites / targets, reduced intracellular drug accumulation, and biofilm formation, preventing the bacteria from being inhibited or killed by drugs, thus resulting in resistance.
[0003] Bacterial biofilms are large, aggregated bacterial films formed when microbial colonies adhere to the surface of an object they come into contact with. To resist external environmental pressures (especially the killing effect of disinfectants and antibacterial drugs), they enclose themselves within the film. Compared to individual, dispersed, planktonic bacteria, this is a unique form of bacterial survival. Therefore, bacterial biofilms serve as an excellent haven for bacteria, preventing antibiotic damage. With the development of microbiology, immunology, molecular biology, materials science, and mathematics, the intricate structure of biofilms has been gradually revealed, and their role in promoting bacterial resistance to drugs has gained significant attention in medicine. At the same time, the enormous harm that bacterial biofilms pose to human society is increasingly attracting concern.
[0004] Currently, the methods for preventing and controlling bacterial biofilms include: (1) physical removal methods, which mainly include mechanical cleaning, ultrasonic cleaning and electric shock; in daily life, mechanical removal of bacterial biofilms is often used, such as brushing teeth to clean dental plaque by friction, but this method is not suitable for the treatment of bacterial biofilms in organisms. (2) Nanocarriers are a submicroscopic drug delivery system at the nanoscale; by encapsulating drugs in them, the release rate can be adjusted, their distribution in the body can be changed, the permeability of biofilms can be increased and the bioavailability can be improved, but the stability and biosafety of this method need to be strictly evaluated.
[0005] In recent years, with the advancement of nanotechnology, especially the development of nanomaterial carriers, nanomaterials have been increasingly widely used in the fields of medicine and healthcare. Nanomaterials as carriers have been approved for clinical use, including the treatment of infections and tumors. Among various types of nanoparticles, iron oxide-based nanoparticles have been widely applied in various fields. Under the guidance of a magnetic field, magnetic nanoparticles can move in a regular pattern within biological fluids. They can also be used to load drugs, with the magnetic field regulating the trajectory of the nanoparticles within the body to ultimately achieve targeted drug delivery. Therefore, there is an urgent need for a strategy to utilize magnetic fields to guide the movement of iron nanoparticles, thereby achieving the effect of inhibiting pathogens. Summary of the Invention
[0006] This invention addresses the problem of antibiotic resistance in existing technologies by providing a method and apparatus for inhibiting bacterial biofilms using magnetic field-driven nanoparticles. This method achieves antibacterial effects through physical and mechanical means and can more effectively eliminate drug-resistant bacteria compared to traditional antibiotics.
[0007] The technical solution adopted in this invention is as follows: the method for inhibiting bacterial biofilms by magnetically driven nanoparticles includes the following steps:
[0008] Step 1: Add nano-magnetic particles to the culture medium containing bacteria;
[0009] Step 2: Add vancomycin to the bacterial culture medium described in Step 1 at a level lower than the bacterial biofilm inhibitory level.
[0010] Step 3: Place the bacterial culture medium described in Step 2 into a coil equipped with a power source;
[0011] Step 4: Turn on the power supply from Step 3, adjust the power parameters to generate a uniform magnetic field in the coil, and allow the magnetic field to intervene for 18 hours. This magnetic field can control the movement of the nano-magnetic particles and inhibit the formation of bacterial biofilms.
[0012] In step one, the magnetic core diameter of the added nanomagnetic particles is 20-50 nm, and the hydrated particle size is 80-150 nm.
[0013] In step four, the magnetic field strength of the uniform magnetic field generated by the coil is 3-10 mT, and the frequency is 1500 Hz.
[0014] In step four, the magnetic field strength waveform of the uniform magnetic field generated by the coil includes a sinusoidal magnetic field and a pulsed magnetic field.
[0015] In step one, the bacteria in the culture medium include Staphylococcus aureus.
[0016] In step two, the concentration of vancomycin added to the bacterial culture medium is 6.25 ug / ml.
[0017] The device used in the above-mentioned method of inhibiting bacterial biofilm by driving nanomagnetic particles with magnetic field includes a power source capable of providing a uniform magnetic field with a magnetic field strength of 3 to 10 mT and a frequency of 1500 Hz, and a coil made of wire wound into a spiral shape.
[0018] The beneficial effects of this invention are as follows: This method of inhibiting bacterial biofilms using magnetically driven nanoparticles, through the physical and mechanical movement of the nanoparticles driven by a magnetic field and in synergistic effect with drug administration, can better inhibit bacterial biofilms and is more suitable for eliminating drug-resistant bacteria. Simultaneously, the designed device for inhibiting bacterial biofilms using magnetically driven nanoparticles is small in size, easy to use, and highly practical. Compared with traditional antibiotic antibacterial methods, this technology can be combined with superparamagnetic nanoparticles for application in wound healing to solve the problem of drug resistance in existing antibiotics, and has significant commercial value. Attached Figure Description
[0019] Figure 1 This is a flowchart of the present invention.
[0020] Figure 2 This is a photograph of the experimental results of the inhibition of Staphylococcus aureus biofilm by the sinusoidal magnetic field-induced nanomagnetic beads of the present invention (Example 1).
[0021] Figure 3 These are photographs of the experimental results of the inhibition of Staphylococcus aureus biofilm by sinusoidal magnetic field nanobeads with different magnetic field strengths according to the present invention (Example 2).
[0022] Figure 4 These are photographs of the experimental results of the inhibition of Staphylococcus aureus biofilm by different magnetic field-induced nanobeads according to the present invention (Example 3).
[0023] Figure 5 This is a schematic diagram of the device used in the method of inhibiting bacterial biofilm by magnetic field-driven nanoparticles according to the present invention.
[0024] The numbers in the diagram are explained as follows: 1. Power supply, 2. Coil. Detailed Implementation
[0025] The bacterial resistance crisis is becoming increasingly severe, posing a major global public health problem and seriously endangering human and animal health and the ecological environment. Data from the China Antimicrobial Resistance Surveillance Network (CHINET) and the National Bloodstream Infection Antimicrobial Resistance Surveillance (BRICS) report show that Gram-negative bacteria are the predominant pathogens causing bloodstream infections, with an increasing trend year by year. Furthermore, the detection rate of drug-resistant bacteria is also increasing annually. Vancomycin-resistant Enterococci and carbapenem-resistant Enterobacteriaceae are at low prevalence levels, while carbapenem-resistant Klebsiella pneumoniae is at a high prevalence level. The prevalence of multidrug-resistant and extensively drug-resistant bacteria poses a serious challenge to clinical anti-infective treatment and imposes a heavy economic burden on patients.
[0026] To address the problem of antibiotic resistance in traditional antibiotics, this method involves adding superparamagnetic nanobeads to the bacterial solution and then using a uniform magnetic field to intervene in the movement of the nanobeads. This magnetic field drives the nanobeads to undergo physical and mechanical movement, thereby inhibiting bacterial biofilms and making it more suitable for eliminating drug-resistant bacteria.
[0027] The specific steps of this invention are described in detail. This method for inhibiting bacterial biofilms using magnetically driven nanoparticles includes:
[0028] Step one involves adding nano-magnetic particles to a culture medium containing bacteria, including Staphylococcus aureus. The added nano-magnetic particles have a magnetic core diameter of 20–50 nm and a hydrated particle size of 80–150 nm.
[0029] Step two, add vancomycin at a concentration lower than the amount that inhibits bacterial biofilm formation to the bacterial culture medium from step one. The concentration of vancomycin is 6.25 ug / ml.
[0030] Step 3: Place the culture medium containing bacteria from Step 2 into a coil equipped with a power source.
[0031] Step four: Turn on the power supply used in step three to control the coil to release the magnetic field, and adjust the power supply parameters to make the coil generate a uniform magnetic field; the magnetic field strength of the uniform magnetic field generated by the coil is 3-10 mT, and the frequency is 1500 Hz. Moreover, the waveform of the uniform magnetic field strength generated by the coil includes a sinusoidal magnetic field and a pulsed magnetic field.
[0032] The magnetic field intervention lasted for 18 hours. This magnetic field could control the movement of nanomagnetic particles and inhibit the formation of bacterial biofilms.
[0033] The reagents developed for this method of inhibiting bacterial biofilms using magnetically driven nanoparticles include superparamagnetic nanobeads, vancomycin, and sterile saline.
[0034] The device used in this method of inhibiting bacterial biofilm by driving nanomagnetic particles with magnetic field includes a power supply 1 capable of providing a uniform magnetic field with a magnetic field strength of 3 to 10 mT and a frequency of 1500 Hz, and a coil 2 made of wire wound into a spiral shape.
[0035] Example 1: Evaluation of the inhibitory effect of sinusoidal magnetic field (magnetic field strength 8mT, frequency 1500Hz) induced by magnetic nanobeads on Staphylococcus aureus biofilm:
[0036] ① Take two 8-well microplates and divide them into experimental group 1 and experimental group 2. Add 50 μL of culture medium, 10 μL of bacterial culture, 10 μL of vancomycin (concentration 0.0625 μg / μL), and 10 μL of magnetic nanobeads sequentially to each group. Then add another 50 μL of culture medium, and top up the liquid volume with sterile water. Finally, seal the plates with sterile membrane. Incubate experimental group 1 at room temperature for 18 hours, and incubate experimental group 2 at room temperature for 18 hours under a magnetic field current of 5 A and a frequency of 1500 Hz (e.g., ...). Figure 2 (As shown).
[0037] ② Remove the liquid from the wells, wash the plate twice with sterile water, add 150 μL of methanol to each well, and incubate for 30 min.
[0038] ③ Remove the methanol, wait for the remaining methanol in the well to evaporate completely, add 150 μL of crystal violet solution, and incubate for 30 min.
[0039] ④ Remove the crystal violet solution, wash the ELISA plate three times with sterile water, add 30% glacial acetic acid solution, let stand for 30 minutes, and then place it in an ELISA reader to detect the OD value (absorbance) at a wavelength of 510 nm.
[0040] Experimental results:
[0041] Table 1. Inhibitory effect of sinusoidal magnetic field-induced nanomagnetic beads on Staphylococcus aureus biofilm (OD value)
[0042]
[0043] Antibacterial efficiency = (Positive control value (with bacteria only) - Antibacterial experimental group (magnetic field, magnetic beads, with drug, with bacteria)) / Positive control value (with bacteria only)
[0044] Conclusion: Magnetic nanobeads had no inhibitory effect on Staphylococcus aureus biofilm under the influence of a magnetic field. However, under the influence of vancomycin concentrations of 0.0625 ug / ul and 10ul, the magnetic nanobeads induced by this magnetic field condition exhibited a significant inhibitory effect on Staphylococcus aureus biofilm, with an antibacterial efficiency of 92.54%.
[0045] Example 2: Evaluation of the inhibitory effect of sinusoidal magnetic field induction of nanomagnetic beads with different magnetic field strengths on Staphylococcus aureus biofilm:
[0046] ① Take 6 eight-well microplates and divide them into control and experimental groups. Add 50 μL of culture medium, 10 μL of bacterial culture, 10 μL of vancomycin (concentration 0.0625 μg / μL), and 10 μL of magnetic nanobeads sequentially to each group. Then add another 50 μL of culture medium, and bring the liquid volume to a balance with sterile water. Finally, seal with sterile membrane. Experimental group 1 is cultured at room temperature for 18 hours. The control group is not subjected to a magnetic field. The experimental groups are cultured at room temperature for 18 hours under magnetic field conditions of 1 mT, 2 mT, 4 mT, 6 mT, and 8 mT at a frequency of 1500 Hz (e.g., ...). Figure 3 (As shown).
[0047] ② Remove the liquid from the wells, wash the plate twice with sterile water, add 150 μL of methanol to each well, and incubate for 30 min.
[0048] ③ Remove the methanol, wait for the remaining methanol in the well to evaporate completely, add 150 μL of crystal violet solution, and incubate for 30 min.
[0049] ④ Remove the crystal violet solution, wash the ELISA plate three times with sterile water, add 30% glacial acetic acid solution, let stand for 30 minutes, and then place it in an ELISA reader to detect the OD value at a wavelength of 510 nm.
[0050] Table 2. OD values of the inhibitory effect of sinusoidal magnetic field-induced nanomagnetic beads on Staphylococcus aureus biofilm at different magnetic field intensities.
[0051]
[0052] Antibacterial efficiency = (Positive control value (with bacteria only) - Antibacterial experimental group (magnetic field, magnetic beads, with drug, with bacteria)) / Positive control value (with bacteria only)
[0053] The antibacterial rate at a field strength of 1 mT is 26.48%.
[0054] At a field strength of 2mT, the antibacterial rate is 20.50%.
[0055] At a field strength of 4mT, the antibacterial rate is 64.52%.
[0056] At a field strength of 6 mT, the antibacterial rate is 80.95%.
[0057] At a field strength of 8 mT, the antibacterial rate is 77.67%.
[0058] Conclusion: AC magnetic fields with field strengths of 6 mT and 8 mT have the best inhibitory effect on biofilms.
[0059] Example 3: Evaluation of the inhibitory effect of different magnetic field-induced nanomagnetic beads on Staphylococcus aureus biofilm:
[0060] ① Take four 8-well microplates and divide them into a control group and an experimental group. Add 50 μL of culture medium, 10 μL of bacterial culture, 10 μL of vancomycin (concentration 0.0625 μg / μL), and 10 μL of magnetic nanobeads sequentially to each group. Then add another 50 μL of culture medium, and bring the liquid volume to a balance with sterile water. Finally, seal the plates with sterile membrane. Experimental group 1 was cultured at room temperature for 18 hours. The control group was not subjected to a magnetic field. The experimental groups were cultured at room temperature for 18 hours under static magnetic field (field strength: 6 mT), pulsed magnetic field (field strength: 6 mT, frequency: 1500 Hz), and alternating magnetic field (field strength: 6 mT, frequency: 1500 Hz), respectively. Figure 4 (As shown).
[0061] ② Remove the liquid from the wells, wash the plate twice with sterile water, add 150 μL of methanol to each well, and incubate for 30 min.
[0062] ③ Remove the methanol, wait for the remaining methanol in the well to evaporate completely, add 150 μL of crystal violet solution, and incubate for 30 min.
[0063] ④ Remove the crystal violet solution, wash the ELISA plate three times with sterile water, add 30% glacial acetic acid solution, let stand for 30 minutes, and then place it in an ELISA reader to detect the OD value at a wavelength of 560 nm.
[0064] Experimental results:
[0065] Table 3. OD values of the inhibitory effects of different magnetic field-induced nanomagnetic beads on Staphylococcus aureus biofilm.
[0066]
[0067] Antibacterial efficiency = (Positive control value (with bacteria only) - Antibacterial experimental group (magnetic field, magnetic beads, with drug, with bacteria)) / Positive control value (with bacteria only)
[0068] Static magnetic field antibacterial rate = -108.1%
[0069] Pulsed magnetic field antibacterial rate = 31%
[0070] The antibacterial rate of alternating magnetic fields is 86.6%.
[0071] Conclusion: A 6mT alternating magnetic field has the best inhibitory effect on biofilms.
[0072] In addition, the present invention provides a method for developing magnetically driven nanoparticle reagents, including:
[0073] Step 1: Take 25 μL of magnetic beads with a particle size of 30 nm, add 2 ml of sterile saline, and mix well.
[0074] Step 2: Add 1 mg of vancomycin to 1 ml of sterile saline and mix well.
[0075] Step 3: After filtration through a 0.45µm filter membrane, dispense the contents into sterile test tubes for later use.
[0076] Furthermore, the device used in the method for inhibiting bacterial biofilm by magnetic field-driven nanoparticles provided by the present invention includes a power supply 1 for controlling the release of magnetic field by the coil and a coil 2 made of wire wound into a spiral shape. The power supply 1 can make the coil 2 generate a uniform magnetic field with a magnetic field strength of 3 to 10 mT and a frequency of 1500 Hz.
Claims
1. A method for inhibiting bacterial biofilms using magnetically driven nanoparticles, characterized in that, Includes the following steps: Step 1: Add nano-magnetic particles to the culture medium containing bacteria; Step 2: Add vancomycin to the bacterial culture medium described in Step 1 at a level lower than the amount that inhibits bacterial biofilm formation. Step 3: Place the bacterial culture medium described in Step 2 into a coil equipped with a power source; Step 4: Turn on the power supply from Step 3, adjust the power parameters to generate a uniform magnetic field in the coil, and allow the magnetic field to intervene for 18 hours. This magnetic field can control the movement of the nano-magnetic particles and inhibit the formation of bacterial biofilms.
2. The method for inhibiting bacterial biofilms using magnetically driven nanoparticles according to claim 1, characterized in that: In step one, the magnetic core diameter of the added nanomagnetic particles is 20-50 nm, and the hydrated particle size is 80-150 nm.
3. The method for inhibiting bacterial biofilms using magnetically driven nanoparticles according to claim 1, characterized in that: In step four, the magnetic field strength of the uniform magnetic field generated by the coil is 3-10 mT, and the frequency is 1500 Hz.
4. The method for inhibiting bacterial biofilms using magnetically driven nanoparticles according to claim 1, characterized in that: In step four, the magnetic field strength waveform of the uniform magnetic field generated by the coil includes a sinusoidal magnetic field and a pulsed magnetic field.
5. The method for inhibiting bacterial biofilms using magnetically driven nanoparticles according to claim 1, characterized in that: In step one, the bacteria in the culture medium include Staphylococcus aureus.
6. The method for inhibiting bacterial biofilms using magnetically driven nanoparticles according to claim 1, characterized in that: In step two, the concentration of vancomycin added to the bacterial culture medium is 6.25 ug / ml.
7. The apparatus used in the method for inhibiting bacterial biofilm by magnetically driven nanoparticles according to claim 1 includes a power supply capable of providing a uniform magnetic field with a magnetic field strength of 3 to 10 mT and a frequency of 1500 Hz, and a coil made of wire wound into a spiral shape.