阅读说明：本技术 一种磷化亚铜的应用 (Application of cuprous phosphide ) 是由 刘玲 晁代勇 董绍俊 于 2021-10-28 设计创作，主要内容包括：本发明涉及一种磷化亚铜的应用,属于磷化亚铜应用技术领域。解决了现有技术中抗菌材料抗菌性能较低,抗菌材料在使用过程中依赖光照等外界条件,以及抗菌材料的制备方法复杂,条件苛刻,成本较高的问题。本发明提供磷化亚铜作为抗菌材料的应用,该应用简便,超高效,具备广谱的杀菌效果,且在使用过程中不依赖外界辅助条件,不需要复杂、苛刻的制备方法。(The invention relates to application of cuprous phosphide, and belongs to the technical field of cuprous phosphide application. Solves the problems that the antibacterial material in the prior art has lower antibacterial performance, the antibacterial material depends on external conditions such as illumination and the like in the using process, and the preparation method of the antibacterial material is complex, the conditions are harsh and the cost is higher. The invention provides the application of cuprous phosphide as an antibacterial material, the application is simple and convenient, the application is ultra-efficient, the cuprous phosphide has a broad-spectrum sterilization effect, the application process does not depend on external auxiliary conditions, and a complex and harsh preparation method is not needed.)

1. The application of cuprous phosphide as antibacterial material.

2. Use of cuprous phosphide as an antimicrobial material according to claim 1, characterized in that cuprous phosphide is used in the form of liquid phase application or solid phase application.

3. Use of cuprous phosphide as antibacterial material according to claim 1,

the liquid phase is applied by adding cuprous phosphide into a liquid containing bacteria;

the solid phase is applied by placing cuprous phosphide on the surface of an object containing bacteria.

4. Use of cuprous phosphide as antibacterial material according to claim 2 or 3, wherein the minimum final concentration of cuprous phosphide to gram-positive bacteria with 99.9% or more inhibition rate in liquid phase application is 0.5 μ g mL^-1The minimum final concentration of cuprous phosphide for inhibiting gram-negative bacteria with the bacteriostasis rate of more than 99.9 percent is 1.5 mu g mL^-1。

5. According to claimThe application of the cuprous phosphide as the antibacterial material in the claim 2 or 3, wherein in the liquid phase application, the minimum final concentration of the cuprous phosphide for reaching the bacteriostasis rate of more than 99.9% to the natural water body is 0.8 mug mL^-1。

6. Use of cuprous phosphide as an antimicrobial material according to claim 2 or 3, wherein said solid phase is applied by preparing cuprous phosphide as a washing solution to be applied to the surface of an object containing bacteria, or preparing cuprous phosphide as a dressing to be applied to the surface of an object containing bacteria.

7. Use of cuprous phosphide as antibacterial material according to claim 6,

in the washing liquid, the concentration of cuprous phosphide is 0.5 mu g mL^-1To 40. mu.g mL^-1；

In the dressing, the dosage of the cuprous phosphide is 0.5 mu g cm^-2To 40. mu.g cm^-2。

8. Use of cuprous phosphide as antimicrobial material according to any of claims 1-3, characterized in that antimicrobial time is more than 0 min.

9. The application of cuprous phosphide as antibacterial material according to claim 8, wherein the antibacterial time for which the bacteriostatic rate is more than 99.9% is more than 20 min.

Technical Field

The invention belongs to the technical field of cuprous phosphide application, and particularly relates to application of cuprous phosphide.

Background

Bacterial diseases are infectious or infective diseases caused by bacteria. Clinically, the most effective method for treating bacterial diseases is to use antibiotics. However, the long-term and continuous use of antibiotics has led to the emergence of drug-resistant bacteria.

In order to solve this technical problem, researchers have developed a variety of nanomaterials having unique antibacterial mechanisms as novel antibacterial agents. The antibacterial application of silver nanoparticles is a successful case in the prior art. In recent years, other nanomaterials with unique physicochemical properties have also been developed for antibacterial purposes. The mechanism of action includes killing bacteria by inducing cell membrane or cell wall destruction of the bacteria by physical contact between the material and the bacteria, or mediating antibacterial application by auxiliary conditions such as light/heat. In addition, nanomaterials having enzyme-like activity (also referred to as mimetic enzymes or nanoenzymes), particularly peroxidase-like and oxidase-like materials, have been developed as novel antibacterial materials since they can catalyze the conversion of oxygen or hydrogen peroxide into active oxygen to exhibit good antibacterial properties.

These research results prove that the nano material has great potential as an antibiotic substitute, but the antibacterial materials in the prior art still have some defects. (1) Excessive release of silver ions is potentially harmful to the environment and human health, and the use of precious metals increases costs. (2) Physical sterilization is inefficient and requires higher concentrations or longer times, limiting its practical application. (3) The light/heat and other auxiliary condition mediated antibacterial application has dependence on the condition in practical use. (4) Optimal reactions of most oxidase-like and peroxidase-like mimetics require acidic conditions (pH 3-4), limiting their use in antimicrobial applications. Therefore, it remains necessary and challenging to develop new antibacterial materials as alternatives to antibiotics that can simultaneously overcome the above limitations.

Cuprous phosphide, Cu₃P, produced by the combined reaction of copper and phosphorus, is yellowish in color, brittle in texture and water-resistantThe reaction is insoluble in dilute inorganic acid and easily soluble in strong oxidizing acid to generate redox reaction. In the prior art, the application of cuprous phosphide is mainly focused on the aspects of photocatalysis, electrocatalysis, lithium ion/sodium ion batteries and the like. Wherein, the first prior art reports a Cu₃The P nanowire modified copper mesh electrode is used for inactivating pathogenic bacteria in water, and the essence of the technology is electric field sterilization, Cu₃The substance of P is used as a modifying material for improving the function of the electrode. Using Cu₃The tip nature of P affects the electric field effect and explains that the mechanism of enhancing electric field sterilization is electroporation (or electrical breakdown) (j. mater. chem.a,2018,6, 18813). The application of the cuprous phosphide as an antibacterial material is not discovered, and a related antibacterial mechanism is not reported.

The invention provides the application of cuprous phosphide as an antibacterial material, which is not based on electric field sterilization and does not need external auxiliary conditions. And further explaining the antibacterial mechanism of the material in principle, the enzyme-like property of the material can generate active oxygen substances, so that the material has the bactericidal property, simultaneously, the material mediates the glutathione depletion in bacterial cells, has the hydrolase-like activity, and increases the permeability of bacterial cell membranes, thereby increasing the antibacterial effect. The material has the sterilization characteristic that the limitation of the nano antibacterial material is overcome: that is, no metal ion is released, the use concentration is extremely low, auxiliary conditions are not required, and a wide pH condition range is effective from acidity to neutrality.

Disclosure of Invention

The invention provides an application of cuprous phosphide for solving the problems that an antibacterial material in the prior art is low in antibacterial performance, depends on external auxiliary conditions in the use process, and is complex in preparation method, harsh in conditions and high in cost.

The technical scheme adopted by the invention for solving the technical problems is as follows.

The invention provides application of cuprous phosphide as an antibacterial material.

Preferably, the cuprous phosphide is applied in the form of a liquid phase application or a solid phase application.

Preferably, the liquid phase is applied by adding cuprous phosphide into a liquid containing bacteria; the solid phase is applied by placing cuprous phosphide on the surface of an object containing bacteria.

Preferably, in the liquid phase application, the minimum final concentrations of the cuprous phosphide for inhibiting gram-positive bacteria and gram-positive bacteria are respectively 0.5 mu g mL^-1And 1.5. mu.g mL^-1。

Preferably, in the liquid phase application, the minimum final concentration of the cuprous phosphide for achieving the bacteriostasis rate of more than 99.9% to the natural water body is 0.8 mu g mL^-1。

Preferably, the solid phase is applied by preparing cuprous phosphide into a washing liquid to be coated on the surface of an object containing bacteria, or preparing cuprous phosphide into a dressing to be coated on the surface of an object containing bacteria.

Preferably, the concentration of cuprous phosphide in the washing solution is 0.5. mu.g mL^-1To 40. mu.g mL^-1(ii) a In the dressing, the dosage of the cuprous phosphide is 0.5 mu g cm^-2To 40. mu.g cm^-2。

Preferably, the antibacterial time is more than 0min, and more preferably, the antibacterial time with the bacteriostasis rate of more than 99.9 percent is more than 20 min.

Compared with the prior art, the invention has the beneficial effects that:

the invention provides the application of cuprous phosphide as an antibacterial material, the application is simple and convenient, the application is ultra-efficient, the cuprous phosphide has a broad-spectrum sterilization effect, the cuprous phosphide is not dependent on auxiliary conditions such as light/heat and the like in the using process, and a complex and harsh preparation method is not needed.

Drawings

In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, and it is apparent that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art that other drawings can be obtained according to the embodiments without inventive labor.

FIG. 1 shows Cu used in examples 1 to 19 of the present invention₃A TEM image of P;

FIG. 2 is the bookCu used in inventive examples 1 to 19₃XRD pattern of P;

FIG. 3 shows Cu of examples 1 to 2 of the present invention₃The antibacterial property detection result of P;

FIG. 4 shows Cu of examples 3 to 12 of the present invention₃The antibacterial property detection result of P;

FIG. 5 shows Cu of example 13 of the present invention₃The antibacterial property detection result of P;

FIG. 6 shows Cu of example 14 of the present invention₃The antibacterial property detection result of P;

FIG. 7 shows Cu of example 15 of the present invention₃The antibacterial property detection result of P;

FIG. 8 shows Cu of example 16 of the present invention₃The antibacterial property detection result of P;

FIG. 9 shows Cu of example 17 of the present invention₃The antibacterial property detection result of P;

FIG. 10 shows Cu of examples 18 to 19 of the present invention₃And (5) detecting the cytotoxicity of the P.

Detailed Description

For a further understanding of the invention, preferred embodiments of the invention are described below in conjunction with the detailed description, but it is to be understood that the description is intended to further illustrate the features and advantages of the invention and not to limit the claims to the invention.

The cuprous phosphide disclosed by the invention can be applied as an antibacterial material. The specific application form is not particularly limited, and the method can be suitable for various application forms of powdery antibacterial materials in the prior art, namely the application form of the cuprous phosphide can be a liquid phase or a solid phase.

The liquid phase application is preferably the addition of cuprous phosphide to the bacteria-containing liquid. In the liquid phase application, the minimum final concentration of the cuprous phosphide for reaching the gram-positive bacteria inhibition rate of more than 99.9 percent is 0.5 mu g mL^-1The minimum final concentration of cuprous phosphide for inhibiting gram-negative bacteria with the bacteriostasis rate of more than 99.9 percent is 1.5 mu g mL^-1(ii) a The minimum final concentration of the cuprous phosphide for inhibiting the bacteria rate of the natural water body to be more than 99.9 percent is 0.8 mu g mL^-1. The concentration of the seed in the liquid containing the bacteria is not particularly limited, and may be 3X 10⁶CFU mL^-1The following.

The solid phase application is preferably to place cuprous phosphide on the surface of the object containing the bacteria. Specifically, the solid phase application is that cuprous phosphide is prepared into lotion and coated on the surface of an object containing bacteria, or cuprous phosphide is prepared into dressing and coated on the surface of the object containing bacteria. The concentration of cuprous phosphide in the washing solution was 0.5. mu.g mL^-1To 40. mu.g mL^-1(ii) a The solute is distilled water. In the dressing, the dosage of the cuprous phosphide is 0.5 mu g cm^-2To 40. mu.g cm^-2。

In the invention, the antibacterial time is preferably more than 5 min; particularly preferably 20min or more; most preferably 20 min.

The terms used in the present invention generally have meanings commonly understood by those of ordinary skill in the art, unless otherwise specified. In the invention, the definition of the remarkable antibacterial action means that the antibacterial rate reaches more than 99.9 percent. The natural water body refers to a natural complex on the earth surface covered by water, oceans, rivers, lakes, marshes, glaciers and the like are all natural water bodies, and the water body not only refers to water but also comprises various substances contained in the water body, such as dissolved substances, suspended substances, aquatic organisms, bottom substances and the like. The object refers to all physical substances in nature, including human body.

In order to make those skilled in the art better understand the technical solution of the present invention, the present invention will be further described in detail with reference to the following embodiments.

In the following examples, various procedures and methods not described in detail are conventional methods well known in the art. Materials, reagents, devices, instruments, apparatuses and the like used in the following examples are commercially available unless otherwise specified. The cuprous phosphide is obtained by phosphorizing copper oxide with sodium hypophosphite at the reaction temperature of 300 ℃ for 2h, the heating rate of 5 ℃/min, and the mass ratio of the sodium hypophosphite to the copper oxide is 15: 1. Copper oxide is prepared according to the methods of the literature (Konar, S.; Kalita, H.; Puvvada, N.; Tantubay, S.; Mahto, M.K.; Biswas, S.; Pathak, A., Shapedependent catalytic activity of CuO nanostructions, journal of catalysis 2016,336, 11-22.).

The present invention is further illustrated by the following examples.

Example 1

Step one, transferring a single gram-negative bacterium escherichia coli (E.coli) colony to 20mL Luria-Bertani (LB) culture medium, culturing for 12h in a constant temperature shaking table at 37 ℃, and diluting to 3 x 10 by using sterile PBS (phosphate buffer solution) with pH 7.4⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 1.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2). Respectively treating for 0min, 5min, 10min and 20min, and diluting with sterile PBS buffer solution with pH of 7.4 to bacterial colony concentration of 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, as shown in FIG. 3.

As can be seen from FIG. 3, the concentration is 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-negative bacteria escherichia coli (E.coli) at 20min can reach 99.9%.

Example 2

Transferring single gram-positive bacterium methicillin-resistant staphylococcus aureus (MRSA) into 20mL Luria-Bertani (LB) culture medium, culturing in a constant temperature shaker at 37 ℃ for 12h, and diluting with sterile PBS buffer solution with pH 7.4 to 3 × 10⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 0.5. mu.gmL^-1Cu of (2)₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2). Respectively treating for 0min, 5min, 10min and 20min, and diluting with PBS to bacterial colony concentration of 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, as shown in FIG. 3.

As can be seen from FIG. 3, the concentration was 0.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-positive bacteria methicillin-resistant staphylococcus aureus (MRSA) at 20min can reach 99.9%.

Example 3

Step one, transferring a single gram-negative bacterium escherichia coli (E.coli) colony to 20mL Luria-Bertani (LB) culture medium, culturing for 12h in a constant temperature shaking table at 37 ℃, and diluting to 3 x 10 by using sterile PBS (phosphate buffer solution) with pH 7.4⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 1.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (b) in FIG. 1, irregular morphology is absent, particle size is around 50nm, XRD is shown as curve (b) in FIG. 2). After being treated for 0min and 20min respectively, the mixture is diluted by sterile PBS buffer solution with pH 7.4 until the colony concentration is 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration is 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-negative bacteria escherichia coli (E.coli) at 20min can reach 99.9%.

Example 4

Transferring single gram-positive bacterium methicillin-resistant staphylococcus aureus (MRSA) into 20mL Luria-Bertani (LB) culture medium, culturing in a constant temperature shaker at 37 ℃ for 12h, and diluting with sterile PBS buffer solution with pH 7.4 to 3 × 10⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 0.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (b) in FIG. 1, irregular morphology is absent, particle size is around 50nm, XRD is shown as curve (b) in FIG. 2). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration is 0.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-positive bacteria methicillin-resistant staphylococcus aureus (MRSA) at 20min can reach 99.9%.

Example 5

Step one, transferring a single gram-negative bacterium escherichia coli (E.coli) colony to 20mL Luria-Bertani (LB) culture medium, culturing for 12h in a constant temperature shaking table at 37 ℃, and diluting to 3 x 10 by using sterile PBS (phosphate buffer solution) with pH 7.4⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 1.5. mu.g mL^-1Cu of (2)₃P (morphology is shown in fig. 1 (c), spherical nanoparticles with a particle size of about 100nm, XRD is shown in fig. 2 (c)). After being treated for 0min and 20min respectively, the mixture is diluted by sterile PBS buffer solution with pH 7.4 until the colony concentration is 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration is 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-negative bacteria escherichia coli (E.coli) at 20min can reach 99.9%.

Example 6

Transferring single gram-positive bacterium methicillin-resistant staphylococcus aureus (MRSA) into 20mL Luria-Bertani (LB) culture medium, culturing in a constant temperature shaker at 37 ℃ for 12h, and diluting with sterile PBS buffer solution with pH 7.4 to 3 × 10⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 0.5. mu.g mL^-1Cu of (2)₃P (morphology is shown in fig. 1 (c), spherical nanoparticles with a particle size of about 100nm, XRD is shown in fig. 2 (c)). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

From FIG. 4 can be seenAs seen, the concentration was 0.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-positive bacteria methicillin-resistant staphylococcus aureus (MRSA) at 20min can reach 99.9%.

Example 7

Step one, transferring a single gram-negative bacterium escherichia coli (E.coli) colony to 20mL Luria-Bertani (LB) culture medium, culturing for 12h in a constant temperature shaking table at 37 ℃, and diluting to 3 x 10 by using sterile PBS (phosphate buffer solution) with pH 7.4⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 1.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (d) in figure 1, spherical nanoparticles with a particle size of about 400nm, XRD is shown as curve (d) in figure 2). After being treated for 0min and 20min respectively, the mixture is diluted by sterile PBS buffer solution with pH 7.4 until the colony concentration is 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration is 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-negative bacteria escherichia coli (E.coli) at 20min can reach 99.9%.

Example 8

Transferring single gram-positive bacterium methicillin-resistant staphylococcus aureus (MRSA) into 20mL Luria-Bertani (LB) culture medium, culturing in a constant temperature shaker at 37 ℃ for 12h, and diluting with sterile PBS buffer solution with pH 7.4 to 3 × 10⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 0.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (d) in figure 1, spherical nanoparticles with a particle size of about 400nm, XRD is shown as curve (d) in figure 2). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration was 0.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-positive bacteria methicillin-resistant staphylococcus aureus (MRSA) at 20min can reach 99.9%.

Example 9

Step one, transferring a single gram-negative bacterium escherichia coli (E.coli) colony to 20mL Luria-Bertani (LB) culture medium, culturing for 12h in a constant temperature shaking table at 37 ℃, and diluting to 3 x 10 by using sterile PBS (phosphate buffer solution) with pH 7.4⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 1.5. mu.g mL^-1Cu of (2)₃P (morphology is shown in fig. 1 (e), spherical nanoparticles with a particle size of about 30nm, XRD is shown in fig. 2 (e)). After being treated for 0min and 20min respectively, the mixture is diluted by sterile PBS buffer solution with pH 7.4 until the colony concentration is 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration is 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-negative bacteria escherichia coli (E.coli) at 20min can reach 99.9%.

Example 10

Transferring single gram-positive bacterium methicillin-resistant staphylococcus aureus (MRSA) into 20mL Luria-Bertani (LB) culture medium, culturing in a constant temperature shaker at 37 ℃ for 12h, and diluting with sterile PBS buffer solution with pH 7.4 to 3 × 10⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 0.5. mu.g mL^-1Cu of (2)₃P (morphology is shown in fig. 1 (e), spherical nanoparticles with a particle size of about 30nm, XRD is shown in fig. 2 (e)). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration was 0.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-positive bacteria methicillin-resistant staphylococcus aureus (MRSA) at 20min can reach 99.9%.

Example 11

Step one, transferring a single gram-negative bacterium escherichia coli (E.coli) colony to 20mL Luria-Bertani (LB) culture medium, culturing for 12h in a constant temperature shaking table at 37 ℃, and diluting to 3 x 10 by using sterile PBS (phosphate buffer solution) with pH 7.4⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 1.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (f) in figure 1, spherical nanoparticles with particle size of about 80nm, XRD is shown as curve (f) in figure 2). After being treated for 0min and 20min respectively, the mixture is diluted by sterile PBS buffer solution with pH 7.4 until the colony concentration is 10³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 4.

As can be seen from FIG. 4, the concentration is 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-negative bacteria escherichia coli (E.coli) at 20min can reach 99.9%.

Example 12

Transferring single gram-positive bacterium methicillin-resistant staphylococcus aureus (MRSA) into 20mL Luria-Bertani (LB) culture medium, culturing in a constant temperature shaker at 37 ℃ for 12h, and diluting with sterile PBS buffer solution with pH 7.4 to 3 × 10⁶CFU mL^-1And obtaining the bacterial solution. To the bacterial solution was added a final concentration of 0.5. mu.g mL^-1Cu of (2)₃P (morphology is shown as (f) in figure 1, spherical nanoparticles with particle size of about 80nm, XRD is shown as curve (f) in figure 2). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Finally, 100. mu.L of each bacterial dilution was spread evenly on LB agar plates, incubated at 37 ℃ for 24 hours, and the number of colonies was counted by plate counting method, the results are shown in the figure4, respectively.

As can be seen from FIG. 4, the concentration was 0.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram-positive bacteria methicillin-resistant staphylococcus aureus (MRSA) at 20min can reach 99.9%.

Examples 1 to 12 show that Cu₃P size and morphology to Cu₃The antibacterial efficiency of P has little effect.

Example 13

Mixing Cu₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2) are dispersed in the lake water sample of Chaihu, and Cu₃The final concentration of P was 0.8. mu.g mL^-1After co-culturing at 37 ℃ for 5min, 10min and 20min, 100 mul of water sample is uniformly spread on an LB agar plate, and after culturing at 37 ℃ for 24h, the number of colonies is calculated by adopting a plate counting method. The results are shown in FIG. 5.

As can be seen from FIG. 5, no Cu is present₃The blank (Control) petri dish treated with P-material had a number of different colonies, representing different bacteria in the original lake water. Based on the results of gene sequencing and annotation, the lake water contains a wide variety of microorganisms. Among the 20 most abundant genera are g __ GpIIa, g __ Exiguobacterium, g __ Bacillariophyta, g __ Acinetobacter, g __ unclassified _ Plactilaceae, g __ unclassified _ Acidirequirement, g __ Gemmobacter, g __ unclassified _ Rhizobiaes, g __ Photobacterium, g __ Brevundimonas, g __ unclassified _ Micrococcus, g __ unclassified _ Bacteria, g 8 Chlorophyta, g __ assembler, g __ Placticepus, 686g 8 Spirobacterium _ gene _ actinobacillus, g 356 Paracoccus, g __, Spirobacterium _ Schizochralski, Spirobacterium _ __, and Verbindungensis. It can be seen that the lake water sample contains a large number of gram-positive and gram-negative bacteria in addition to E.coli and MRSA. Cu₃The concentration of P was 0.8. mu.g mL^-1More than 99.9% of various bacteria can be killed within 20 min.

Example 13 shows that Cu₃The P material has broad-spectrum antibacterial performance.

Example 14

A single large intestine rodThe bacteria or MRSA were transferred to 20mL LB medium, cultured in a shaker at 37 ℃ for 12 hours, and then diluted to 3X 10 with sterile PBS buffer of pH 7.4⁶CFUmL^-1And obtaining the bacterial solution. To the bacterial solutions, 0.5. mu.g mL of final concentration was added^-1，1.0μg mL^-1，1.5μg mL^-1，2.0μg mL^-1，2.5μg mL^-1，5.0μg mL^-1，10.0μg mL^-1，20.0,μg mL^-1，40.0μg mL^-1Cu of (2)₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Each 100. mu.L of the bacterial dilution was spread evenly on LB agar plates, and after incubation at 37 ℃ for 24 hours, the number of colonies was counted by plate count, and the results are shown in FIG. 6.

As can be seen from FIG. 6, the final concentrations were 0.5. mu.g/mL or more, respectively^-1And 1.5. mu.g mL^-1Cu of (2)₃The sterilization efficiency of the P to gram positive bacteria and gram negative bacteria at 20min can reach more than 99.9 percent.

Example 14 shows Cu with a significant antibacterial effect for gram-positive and gram-negative bacteria₃The minimum concentration of P was 0.5. mu.g mL each^-1And 1.5. mu.g mL^-1Cu higher than the minimum concentration₃P also has an obvious antibacterial effect, and the antibacterial efficiency of the P reaches more than 99.9 percent. Other concentrations of Cu₃P also has antibacterial effect.

Example 15

Transferring single Escherichia coli or MRSA into 20mL LB medium, culturing in 37 deg.C constant temperature shaking table for 12h, diluting with sterile PBS buffer solution with pH 7.4 to 10CFU L^-1(also noted as 0.01CFU mL^-1),1CFU mL^-1,100CFU mL^-1,1×10⁴CFU mL^-1,1×10⁶CFUmL^-1，3×10⁶CFUmL^-1,1×10⁸CFUmL^-1A bacterial solution was obtained. To the bacterial solution was added a final concentration of 5.0. mu.g mL^-1Cu of (2)₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2). After being treated for 0min and 20min respectively, the bacterial colony concentration is diluted to 10 by PBS³CFU mL^-1And obtaining bacterial dilution. Each 100. mu.L of the bacterial dilution was spread evenly on an LB agar plate, and after culturing at 37 ℃ for 24 hours, the number of colonies was counted by plate counting, and the results are shown in FIG. 7.

As can be seen from FIG. 7, Cu₃The concentration of P material is 0.01 to 1 × 10 per ml⁸CFUmL^-1The sterilization efficiency can reach more than 99.9 percent after 20min of bacterial action.

Example 15 shows that Cu₃The antimicrobial effect of the P material is not limited to the concentration of bacteria.

Example 16

Preparation of 0.5 to 40.0. mu.g mL^-1Cu of (2)₃P mother liquor (Cu)₃P morphology is shown as figure 1 (a), spherical nanoparticles with particle size of about 35nm, XRD is shown as curve 2 (a), the solvent is distilled water), and the solution is dripped into mouse wound with wound size of about 80mm²And (6) cutting. While setting blank control group, i.e. without adding Cu³P wound natural healing group. Wound area was measured every 12h starting at 0min and the results are shown in FIG. 8, indicating Cu₃P sterilization is beneficial to wound recovery and the like.

Example 17

Mixing Cu₃P powder as dressing (Cu)₃The morphology of P is shown as (a) in figure 1, the particle size of spherical nanoparticles is about 35nm, XRD is shown as curve (a) in figure 2), and 0.5-40.0 μ g cm is taken^-2Is directly placed on the wound of a mouse, and the wound of the mouse is about 80mm²And (6) cutting. With simultaneous provision of a blank control, i.e. without Cu³P wound natural healing group of dressing. Wound area was measured every 12h starting at 0min, FIG. 9 shows the amount 1.5. mu.g cm^-2The measurement result of (2) shows Cu₃P can be used as medical dressing for wound sterilization and recovery.

Examples 16 to 17 illustrate that Cu³P nano material has medical functionAntibacterial use, in the form of a liquid or solid phase.

Example 18

MCF-7 cells (100. mu.L) in DMEM medium were seeded into 96-well plates at 5000 cells/well, one set of five wells, for a total of five sets, and Cu was added to each set of five wells₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2), Cu in each hole in a group₃The concentration of P was 0. mu.g mL^-1(ii) a Cu in each hole of a group₃The concentration of P was 2. mu.g mL^-1Cu in each hole in a group₃The concentration of P was 4. mu.g mL^-1Cu in each hole of a group₃The concentration of P was 8. mu.g mL^-1Cu in each hole of a group₃The concentration of P was 16. mu.g mL^-1After 24h incubation, CCK-8 solution was added to each well, incubation was continued for 1h, and then the cell viability was determined by measuring absorbance at 450nm with a microplate reader, the results of which are shown in fig. 10.

As can be seen from FIG. 10, Cu₃The concentration of P was 16. mu.g mL^-1There was no significant effect on the survival of MCF-7 cells.

Example 19

HepG2 cells (100. mu.L) in DMEM medium were seeded into 96-well plates at 5000 cells/well, one set of five wells, for five sets, and Cu was added to each set of five wells₃P (morphology is shown as (a) in figure 1, spherical nanoparticles with particle size of about 35nm, XRD is shown as curve (a) in figure 2), Cu in each hole in a group₃The concentration of P was 0. mu.g mL^-1(ii) a Cu in each hole of a group₃The concentration of P was 2. mu.g mL^-1Cu in each hole in a group₃The concentration of P was 4. mu.g mL^-1Cu in each hole of a group₃The concentration of P was 8. mu.g mL^-1Cu in each hole of a group₃The concentration of P was 16. mu.g mL^-1After 24h incubation, CCK-8 solution was added to each well, incubation was continued for 1h, and then the cell viability was determined by measuring absorbance at 450nm with a microplate reader, the results of which are shown in fig. 10.

As can be seen from FIG. 10, Cu₃The concentration of P was 16. mu.g mL^-1There was no significant effect on the survival of MCF-7 cells.

Examples 18 to 19 illustrate that³The P material is minimally toxic to mammalian cells.

The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.