Antibacterial and antioxidant digallium triselenide nanosheet as well as preparation method and application thereof

Abstract

The invention relates to an antibacterial and antioxidant digallium triselenide nanosheet and a preparation method and application thereof, and is characterized in that block digallium triselenide is used as a raw material, and the nanoscale digallium triselenide nanosheet is prepared through a one-step liquid phase ultrasonic stripping method; the radial size of the nano-scale gallium triselenide nanosheet is 10 to 100 nm, and the thickness of the nanosheet layer is 2 to 5 nm; the antibacterial and antioxidant digallium triselenide nanosheet prepared by the invention has the advantages of simple synthesis, clear structure, adjustable size and the like, and the material preparation method is simple, mild in reaction condition and suitable for large-scale production; the digallium triselenide nanosheet has efficient antibacterial activity, has a low-dosage broad-spectrum antibacterial effect of 10-25 [mu] g / mL, and is not easy to induce bacterial drug resistance, and compared with blocky digallium triselenide, the antibacterial performance is improved by nearly 800 times; in addition, the gallium triselenide nanosheet can remove active oxygen and reduce the oxidative stress level of cells, so that the cell protection function is achieved.

Description

Technical Field

[0001] The invention relates to the technical field of gallium triselenide nanosheets, and in particular to an antibacterial and antioxidant gallium triselenide nanosheet, a preparation method and application thereof. Background Art

[0002] Antibiotic resistance is one of the most serious threats to global public health. According to the World Health Organization, if not controlled, infections based on resistant bacteria are expected to cause 350 million deaths by 2050. Therefore, it is urgent to develop effective strategies to deal with the increasingly serious problem of bacterial resistance.

[0003] For a long time, the development of new antibiotics has been considered the most direct means to combat bacterial resistance. However, the development of new antibiotics is far behind the growth rate of resistant bacteria. Therefore, it is difficult to cope with the proliferation and spread of resistant bacteria by relying solely on the development of antibiotics. Nano antibacterial materials provide a new opportunity to solve the problem of bacterial resistance due to their controllable physical and chemical properties, good biocompatibility, and tissue and cell penetration.

[0004] Iron and sulfur are important elements that constitute many types of biological molecules. Iron / sulfur-containing enzymes are involved in a series of physiological activities such as pathogen oxidative stress defense, electron transfer, and DNA synthesis. In recent years, there have been reports of the use of gallium and selenium, which are close to the atomic radius of iron and sulfur, to construct nano-antibacterial materials, exerting a "Trojan horse" effect, so that bacteria cannot distinguish these materials from the essential iron and sulfur elements of the body after ingesting them, thereby causing metabolic disorders in bacteria and ultimately inducing bacterial death. In addition, selenium, as a special trace element, can also act as an antioxidant to resist damage caused by oxidative stress. At present, the development of materials with both antibacterial and antioxidant functions is still in the exploratory stage. Making full use of the characteristics of the element to construct such functional integrated nanomaterials based on gallium selenide shows great application potential. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide an antibacterial and antioxidant gallium triselenide nanosheet, a preparation method and application thereof, so as to avoid the disadvantages of existing metal-based antibacterial materials such as low antibacterial efficiency, difficulty in functional integration and poor biocompatibility.

[0006] To solve the above technical problems, the technical solution of the present invention is: an antibacterial and antioxidant gallium triselenide nanosheet, the innovation of which lies in: using bulk gallium triselenide as raw material, nano-scale gallium triselenide nanosheets are prepared by a one-step liquid phase ultrasonic exfoliation method; the nano-scale gallium triselenide nanosheets have a radial size of 10-100nm and a sheet thickness of 2-5nm.

[0007] A method for preparing antibacterial and antioxidant gallium selenide nanosheets, the innovation of which lies in: the specific preparation method is as follows:

[0008] Using bulk gallium triselenide as raw material, nanometer-scale gallium triselenide nanosheets were prepared by a one-step liquid phase ultrasonic exfoliation method.

[0009] Furthermore, bulk gallium triselenide is mixed with a dispersant to obtain a dispersion; the dispersion is ultrasonically treated to obtain a gallium triselenide nanosheet dispersion, which is then centrifugally purified to obtain gallium triselenide nanosheets.

[0010] Further, the dispersant includes any one of water, N-methylpyrrolidone, isopropanol, and ethanol, or a combination of two or more thereof; and / or, the mass concentration of gallium triselenide in the dispersion is 2-100 mg / mL; and / or, the ultrasonic device used in the ultrasonic treatment includes at least any one of a cell crusher and an ultrasonic cleaner; and / or, the temperature of the ultrasonic treatment is -20-25°C; and / or, the time of the ultrasonic treatment is 0.5h-24h; and / or, the ultrasonic power of the ultrasonic treatment is 100-1000W; and / or, the centrifugal purification includes: first removing large particles in the gallium triselenide nanosheet dispersion by low-speed centrifugation at 500-10000rpm, and then purifying by high-speed centrifugation at 15000-20000rpm.

[0011] The invention discloses an application of an antibacterial and antioxidant gallium triselenide nanosheet, wherein the innovation point is that the antibacterial and antioxidant gallium triselenide nanosheet is applied in antibacterial, antioxidant or cell protection.

[0012] The advantages of the present invention are:

[0013] 1) The antibacterial and antioxidant gallium triselenide nanosheets prepared by the present invention have the advantages of simple synthesis, clear structure, adjustable size, etc., and the material preparation method is simple, the reaction conditions are mild and suitable for large-scale production; the gallium triselenide nanosheets have high antibacterial activity, have a low-dose broad-spectrum antibacterial effect of 10-25 μg / mL, and are not easy to induce bacterial resistance. Compared with bulk gallium triselenide, the antibacterial performance is improved by nearly 800 times; in addition, the gallium triselenide nanosheets can remove active oxygen and reduce the level of cellular oxidative stress, thereby exerting a cell protection function. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments.

[0015] Figure 1 This is a transmission electron microscope image of gallium triselenide nanosheets prepared using water as a dispersant in Example 1 of the present invention;

[0016] Figure 2This is a transmission electron microscope image of gallium triselenide nanosheets prepared using isopropyl alcohol as a dispersant in Example 3 of the present invention;

[0017] Figure 3 This is a statistical diagram of the particle size of gallium triselenide nanosheets prepared using isopropyl alcohol as a dispersant in Example 3 of the present invention;

[0018] Figure 4 This is an atomic force microscope image of gallium triselenide nanosheets prepared using isopropanol as a dispersant in Example 3 of the present invention (the inserted image is the corresponding height statistics graph);

[0019] Figure 5a , 5b 5c and 5c are mapping images of gallium triselenide nanosheets prepared using isopropanol as a dispersant in Example 3 of the present invention;

[0020] Figure 6 The X-ray crystal diffraction pattern of gallium triselenide nanosheets prepared using isopropanol as a dispersant in Example 3 of the present invention;

[0021] Figure 7 This is a transmission electron microscope image of gallium triselenide nanosheets prepared using N-methylpyrrolidone as a dispersant in Example 10 of the present invention;

[0022] Figure 8 This is a transmission electron microscope image of gallium triselenide nanosheets prepared using ethanol as a dispersant in Example 11 of the present invention;

[0023] Figure 9a and Figure 9b The picture of the gallium triselenide nanosheets obtained in Example 3 removing Staphylococcus aureus and Escherichia coli from the agar plate in Example 12 and the corresponding antibacterial statistical result bar graph;

[0024] Fig.10a and Fig.10b A photograph of an agar plate showing the removal of methicillin-resistant Staphylococcus aureus by the gallium triselenide nanosheets obtained in Example 3 in Example 13 and a bar graph showing the corresponding antibacterial statistical results;

[0025] Fig.11a and Fig.11b Agar plate photograph of the bulk gallium triselenide material (16 mg / mL) in Example 14 for removing Staphylococcus aureus and Escherichia coli and a corresponding bar graph of antibacterial statistical results;

[0026] Fig.12 This is a statistical result diagram of the drug resistance of bacteria caused by gallium triselenide nanosheets and gentamicin obtained in Example 3 in Example 15 in Escherichia coli cultured for 14 generations;

[0027] Fig.13A bar graph showing the removal of ABTS· by gallium triselenide nanosheets obtained in Example 3 at different concentrations in Example 16 (the inserted picture is the corresponding solution picture);

[0028] Fig.14 A bar graph showing the scavenging of DPPH· by gallium triselenide nanosheets obtained in Example 3 at different concentrations in Example 17 (the inserted picture is the corresponding solution picture);

[0029] Fig.15 A bar graph showing the OH removal of gallium triselenide nanosheets obtained in Example 3 at different concentrations in Example 18 (the inserted picture is the corresponding solution picture);

[0030] Fig.16 A bar graph showing the removal of O2-· by gallium triselenide nanosheets obtained in Example 3 at different concentrations in Example 19 (the inserted picture is the corresponding solution picture);

[0031] Fig.17 This is a bar graph of the cytotoxicity of HaCaT cells in Example 21 incubated with different concentrations of F127-GaSe nanosheets for 24 hours;

[0032] Fig.18a and Fig.18b This is an inverted fluorescence microscope image and a bar graph of the corresponding fluorescence intensity statistical results for evaluating the intracellular reactive oxygen levels of HaCaT cells under different treatment conditions in Example 22.

[0033] Fig.19 This is a bar graph of the cytotoxicity of 1.3 mM hydrogen peroxide and different concentrations of F127-gallium triselenide nanosheets in Example 22. DETAILED DESCRIPTION

[0034] In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Generally, the components of the embodiments of the present invention described and shown in the drawings here can be arranged and designed in various different configurations.

[0035] Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the invention claimed for protection, but merely represents selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0036] Embodiment 1:

[0037] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0038] (1) First, 10 mg of bulk gallium triselenide powder was weighed and dispersed in ultrapure water to a final concentration of 2 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated for 0.5 h at a power of 300 W. During the treatment, a 0°C coolant was used to maintain the reaction temperature.

[0039] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 10000 rpm for 10 min, and then the supernatant was collected to obtain gallium triselenide nanosheets with a size of about 10 nm; Figure 1 , which is a transmission electron microscope morphology image of the gallium triselenide nanosheet material obtained in this example, it can be seen that gallium triselenide is a sheet-like nanomaterial.

[0040] Embodiment 2:

[0041] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0042] (1) First, 10 mg of bulk gallium triselenide powder was weighed and dispersed in ultrapure water to a final concentration of 2 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated for 3 h at a power of 100 W. During the treatment, a 0°C coolant was used to maintain the reaction temperature.

[0043] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 10,000 rpm for 10 min, and then the supernatant was collected to obtain gallium triselenide nanosheets with a size of about 10 nm.

[0044] Embodiment 3:

[0045] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0046] (1) First, 10 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 2 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 300 W for 3 h. During the treatment, the reaction temperature was maintained at 0°C with a coolant.

[0047] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 5000 rpm for 10 min, and then the supernatant was collected and centrifuged at 20000 rpm in a high-speed centrifuge. After washing three times with isopropanol, the gallium triselenide nanosheets were purified; Figure 2 As shown, the morphology of the gallium triselenide nanosheet obtained in this example is observed by transmission electron microscopy; the particle size statistical analysis shows that its size is about 22nm, which can be seen in Figure 3; Then, the height was characterized by atomic force microscopy, which showed that its height was about 3.8nm, see Figure 4 , the inserted figure is the corresponding height statistical diagram, proving that this condition can synthesize a gallium triselenide sheet material; in order to further confirm the distribution of elements on the material, an analysis was performed through Mapping, such as Figure 5a 5b is the HADDF image of the material, 5b is the mapping of the element gallium, and 5c is the mapping of the element selenium. The results show that the two elements gallium and selenium are evenly distributed on the surface of the material. Subsequently, the crystal structure was characterized by X-ray crystal diffractometer. Figure 6 , and the results showed that its crystal structure is consistent with that of bulk gallium triselenide.

[0048] Embodiment 4:

[0049] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0050] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 300 W for 3 h. During the treatment, the reaction temperature was maintained at 0°C with a coolant.

[0051] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 5000 rpm for 10 min. The supernatant was then collected and centrifuged at 20000 rpm in a high-speed centrifuge. After washing three times with isopropanol, gallium triselenide nanosheets were obtained after purification.

[0052] Embodiment 5:

[0053] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0054] (1) First, 500 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 100 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 300 W for 3 h. During the treatment, the reaction temperature was maintained at 0°C with a coolant.

[0055] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 5000 rpm for 10 min. The supernatant was then collected and centrifuged at 20000 rpm in a high-speed centrifuge. After washing three times with isopropanol, gallium triselenide nanosheets were obtained after purification.

[0056] Embodiment 6:

[0057] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0058] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 300 W for 3 h. During the treatment, the reaction temperature was maintained at 0°C with a coolant.

[0059] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 5000 rpm for 10 min. The supernatant was then collected and centrifuged at 17000 rpm in a high-speed centrifuge. After washing three times with isopropanol, gallium triselenide nanosheets were obtained after purification.

[0060] Embodiment 7:

[0061] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0062] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 300 W for 3 h. During the treatment, the reaction temperature was maintained at 0°C with a coolant.

[0063] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 5000 rpm for 10 min. The supernatant was then collected and centrifuged at 15000 rpm in a high-speed centrifuge. After washing three times with isopropanol, gallium triselenide nanosheets were obtained after purification.

[0064] Embodiment 8:

[0065] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0066] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 600 W for 1 h. During the treatment, the reaction temperature was maintained at -25°C with a coolant.

[0067] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 2000 rpm for 10 min. The supernatant was then collected and centrifuged at 20000 rpm in a high-speed centrifuge. After washing three times with isopropanol, gallium triselenide nanosheets were obtained after purification.

[0068] Embodiment 9:

[0069] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0070] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in isopropanol to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 1000 W for 1 h. During the treatment, the reaction temperature was maintained at -25°C with a coolant.

[0071] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 500 rpm for 10 min. The supernatant was then collected and centrifuged at 20,000 rpm in a high-speed centrifuge. After washing three times with isopropanol, gallium triselenide nanosheets were obtained after purification.

[0072] Embodiment 10:

[0073] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0074] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in N-methylpyrrolidone to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 600 W for 3 h. During the treatment, the reaction temperature was maintained at -25°C with a coolant.

[0075] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 2000 rpm for 10 min, and then the supernatant was collected and centrifuged at 20000 rpm in a high-speed centrifuge. After washing three times with isopropanol, the gallium triselenide nanosheets were obtained after purification. Figure 7 As shown, it can be observed through transmission electron microscopy that a gallium triselenide nanosheet can also be obtained.

[0076] Embodiment 11:

[0077] The preparation method of gallium triselenide nanosheets with antibacterial and antioxidant functions is as follows:

[0078] (1) First, 250 mg of bulk gallium triselenide powder was weighed and dispersed in ethanol to a final concentration of 50 mg / mL. Then, it was placed in a cell crusher and ultrasonically treated at 400 W for 5 h. During the treatment, a -10°C coolant was used to maintain the reaction temperature.

[0079] (2) After the ultrasonic treatment, the sample solution was placed in a centrifuge and centrifuged at 5000 rpm for 10 min, and then the supernatant was collected and centrifuged at 20000 rpm in a high-speed centrifuge. After washing three times with ethanol, the gallium triselenide nanosheets were purified. Figure 8 As shown, it can be observed through transmission electron microscopy that a smaller gallium triselenide nanosheet can be obtained.

[0080] Example 12: Antibacterial performance test on Staphylococcus aureus and Escherichia coli

[0081] The inventors of this case tested the antibacterial performance of the gallium triselenide nanosheets obtained in Example 3 against Staphylococcus aureus and Escherichia coli: the experiment was divided into 5 groups (blank group, 10μg group, 15μg group, 20μg group, 25μg group). The specific implementation steps are as follows:

[0082] Step (1) Blank group: 50 μL of saline was added to 450 μL of bacterial suspension, and the final bacterial concentration was 10 8 CFU / mL;

[0083] Step (2) Experimental group: 50 μL of GaSe nanosheets of different concentrations were added to 450 μL of bacterial suspension, with a final bacterial concentration of 10 8 CFU / mL;

[0084] Step (3) The mixed solution prepared above was placed in a constant temperature shaker and incubated at 37°C for 2 h, and then the bacterial survival rate was evaluated by the plate method, such as Figure 9a The following is a photo of gallium triselenide nanosheets removing Staphylococcus aureus and Escherichia coli from agar plates. Figure 9b This is a bar graph of the statistical results of bacterial survival rates. The results show that the survival rates of both bacteria decrease with increasing concentrations. When the material concentration is 25 μg / mL, the survival rates of Staphylococcus aureus and Escherichia coli are only 1.4% and 2.7%, respectively.

[0085] Example 13: Antibacterial performance test of methicillin-resistant Staphylococcus aureus

[0086] The inventors of this case tested the antibacterial performance of the gallium triselenide nanosheets obtained in Example 3 against methicillin-resistant Staphylococcus aureus: the experiment was divided into 5 groups (blank group, 10μg group, 15μg group, 20μg group, 25μg group). The specific implementation steps are as follows:

[0087] Step (1) Blank group: 50 μL of saline was added to 450 μL of bacterial suspension, and the final bacterial concentration was 10 8 CFU / mL;

[0088] Step (2) Experimental group: 50 μL of GaSe nanosheets of different concentrations were added to 450 μL of bacterial suspension, with a final bacterial concentration of 10 8 CFU / mL;

[0089] Step (3) The mixed solution prepared above was placed in a constant temperature shaker and incubated at 37°C for 2 h, and then the bacterial survival rate was evaluated by the plate method, such as Fig.10a The picture shown is a photo of gallium triselenide nanosheets removing methicillin-resistant Staphylococcus aureus from an agar plate. Fig.10bThe bacterial survival rate statistical results are shown in the bar graph. The results show that the bacterial survival rate decreases with increasing concentration. When the material concentration is 25μg / mL, the bacterial survival rate is only 1.6%.

[0090] Example 14: Antibacterial performance improvement test

[0091] The inventors of this case tested the antibacterial performance of the gallium triselenide nanosheets obtained in Example 3 by using Staphylococcus aureus and Escherichia coli to compare with the bulk gallium triselenide. The experiment was divided into two groups (blank group and 16 mg / mL bulk gallium triselenide group). The specific implementation steps are as follows:

[0092] Step (1) Blank group: 50 μL of saline was added to 450 μL of bacterial suspension, and the final bacterial concentration was 10 8 CFU / mL;

[0093] Step (2) Experimental group: Take 50 μL of 160 mg / mL bulk gallium triselenide solution and add it to 450 μL of bacterial suspension. The final bacterial concentration is 10 8 CFU / mL;

[0094] Step (3) The mixed solution prepared above was placed in a constant temperature shaker and incubated at 37°C for 2 h, and then the bacterial survival rate was evaluated by the plate method, such as Fig.11a The figure shows an agar plate photo of the bulk gallium triselenide material (16 mg / mL) removing Staphylococcus aureus and Escherichia coli. Fig.11b This is a bar chart of the statistical results of bacterial survival rate. The results show that when the material concentration is 16 mg / mL, the survival rates of Staphylococcus aureus and Escherichia coli are 1.4% and 26.9%, respectively.

[0095] Example 15: Drug resistance and antibacterial performance test

[0096] The inventors of this case tested the gallium triselenide nanosheets obtained in Example 3 with Escherichia coli to see whether the gallium triselenide nanosheets were easy to induce the development of bacterial resistance compared with the antibiotic gentamicin. Simply put, the gallium triselenide nanosheets and gentamicin were incubated with Escherichia coli respectively, and the surviving bacteria were selected for continued subculture, and then the antibacterial test was continued, and this cycle was repeated 14 times. The results showed that the antibacterial concentration of gentamicin was increased by 8 times, while the gallium triselenide nanosheets still maintained a relatively stable antibacterial rate at the original concentration, such as Fig.12 Shown is the statistical results of bacterial resistance.

[0097] Example 16: Clearing ABTS Performance Evaluation

[0098] The inventors of this case evaluated the antioxidant properties of the gallium triselenide nanosheets obtained in Example 3 using ABTS·. First, the gallium triselenide nanosheet dispersion and the ABTS· working solution were mixed at a volume ratio of 1:4 and blown evenly. At this time, the working concentrations of the gallium triselenide nanosheets were 2, 4, 6, 8, 10, and 12 μg / mL, respectively. The mixed solution was added to a 96-well plate at a volume of 100 μL per well, and repeated 3 times. The 96-well plate was placed in a dark environment to react for 30 minutes, and then the absorbance of the mixture was measured at 734 nm with an enzyme reader. The results showed that the removal efficiency of ABTS· gradually increased with the increase of the material concentration. When the material concentration was 12 μg / mL, the removal rate reached 94%. Fig.13 A bar graph showing the removal of ABTS· by GaSe nanosheets (the inset is the corresponding solution image).

[0099] Example 17: DPPH removal performance evaluation

[0100] The inventors of this case evaluated the antioxidant properties of the gallium triselenide nanosheets obtained in Example 3 by DPPH·. Using 95% ethanol as a solvent, a 0.12 mg / mL DPPH· solution was prepared. The DPPH· solution and the gallium triselenide nanosheet dispersion were mixed in equal volumes and blown evenly. At this time, the working concentrations of the gallium triselenide nanosheets were 2, 4, 6, 8, and 10 μg / mL, respectively. The results showed that the scavenging efficiency of DPPH· gradually increased with the increase of the material concentration. When the material concentration was 10 μg / mL, the scavenging rate reached 68.4%. Please refer to Fig.14 , is a bar graph of DPPH· scavenging by GaSe nanosheets (the inserted image is the corresponding solution image).

[0101] Example 18: OH scavenging performance evaluation

[0102] The inventors of this case evaluated the antioxidant properties of the gallium triselenide nanosheets obtained in Example 3 by OH·. First, FeSO4 was dissolved in an acetic acid-sodium acetate buffer solution with a pH of 4 to a final concentration of 4mM, and then H2O2 was diluted to 40mM with deionized water. Finally, a 1mM 3,3',5,5'-tetramethylbenzidine solution with dimethyl sulfoxide as a solvent was prepared. Next, the AA-CDs dispersion and FeSO4 solution, H2O2 solution, and TMB solution were mixed in a volume ratio of 1:1:1:1 and blown evenly. At this time, the working concentrations of AA-CDs were 5, 10, 15, 20, 25, and 30μg / mL, respectively. The mixed solution was added to a 96-well plate at a volume of 100μL per well, and repeated 3 times. The 96-well plate was placed in a dark environment for reaction for 30 minutes, and then the absorbance of the mixed solution at 652nm was measured. The results showed that the OH· removal efficiency gradually increased with the increase of material concentration. When the material concentration was 20 μg / mL, the removal rate reached 68.6%. Fig.15 Shown is a bar graph of OH removal by GaSe nanosheets (the inserted image is the corresponding solution image).

[0103] Example 19: O2- Removal Performance Evaluation

[0104] The inventors of this case evaluated the antioxidant properties of the gallium triselenide nanosheets obtained in Example 3 by O2-·. The gallium triselenide nanosheet dispersion was mixed with the β-NADH solution and the NBT solution in a volume ratio of 1:1:1 (100 μL each), blown evenly, and finally 100 μL of PMS was added to start the reaction. At this time, the working concentrations of AA-CDs were 10, 20, 30, 40, 50, and 60 μg / mL, respectively. The mixed solution was added to a 96-well plate at a volume of 100 μL per well, and repeated 3 times. The 96-well plate was placed in a dark environment for reaction for 30 minutes, and then the absorbance of the mixture was measured at 560 nm with an enzyme reader. The results showed that with the increase of the material concentration, the O2-· removal efficiency gradually increased. When the material concentration was 60 μg / mL, the removal rate reached 69.5. Fig.16 Shown is a bar graph of O2-· scavenging by GaSe nanosheets (the inserted image is the corresponding solution image).

[0105] Example 20: Surface modification of Pluronic F127

[0106] The inventors of this case can further expand the application field of the material by surface modification of the gallium triselenide nanosheets obtained in Example 3 with the polymer Pluronic F127. Specifically, a volume of 5 mL of an isopropanol solution with a concentration of 500 mg / mL Pluronic F127 is evenly mixed with 30 mg of gallium triselenide nanosheets, and then the solution is added dropwise to ultrapure water and stirred for 2 hours. Subsequently, Pluronic F127-modified gallium triselenide nanosheets are obtained by centrifugation, which are called F127-gallium triselenide sheet materials.

[0107] Example 21: Biocompatibility Assessment

[0108] The inventors of this case evaluated the cytotoxicity of the F127-GaSe nanosheets obtained in Example 20 by human skin keratinocytes (HaCaT) cells. After the cells in good condition were incubated with cell culture medium containing different concentrations of GaSe nanosheets for 24 hours, the cytotoxicity of the material was evaluated by the CCK-8 method. The results showed that the cell survival rate was higher than 90%. Fig.17 Shown is a bar graph of the cytotoxicity of HaCaT cells incubated with different concentrations of F127-GaSe nanosheets for 24 hours.

[0109] Example 22: Assessment of Cellular Oxidative Protection

[0110] The inventors of this case evaluated the F127-GaSe nanosheets obtained in Example 20 by using a hydrogen peroxide-induced cell oxidative stress model to evaluate the scavenging activity of GaSe nanosheets on intracellular reactive oxygen species and the protective effect on oxidative stress cells. First, HaCaT cells were seeded into a confocal microplate, and the culture medium was removed after culturing for 24 hours for subsequent evaluation experiments. The specific experimental steps are as follows:

[0111] First, the experiment was divided into three groups according to the different treatment methods: the first group added fresh culture medium, the second group added culture medium containing 1.3mM hydrogen peroxide, and the third group added culture medium containing 1.3mM hydrogen peroxide and 40μg / mL gallium triselenide nanosheets. After the group treatment was completed, the cells were placed in a cell culture incubator for 6 hours, then the culture medium was removed and DCFH-DA (an intracellular reactive oxygen fluorescence indicator) was added, and finally observed under a fluorescence microscope. The results showed that the green fluorescence in the cells of the hydrogen peroxide group was the strongest (indicating the highest level of reactive oxygen), the fluorescence of the simple culture medium group was the weakest, and the fluorescence intensity of the cells in the material group was significantly lower than that of the hydrogen peroxide group, proving that the material has an excellent ability to remove intracellular reactive oxygen. Fig.18a and Fig.18b Inverted fluorescence microscopy images and corresponding fluorescence intensity statistical results bar graphs are used to evaluate the intracellular reactive oxygen species levels in HaCaT cells under different treatment conditions.

[0112] In view of the good ability of gallium triselenide nanosheets to remove reactive oxygen species in cells, the present invention further evaluated the protective effect of different concentrations of materials on oxidative stress cells by CCK-8 method. The results showed that with the increase of the concentration of gallium triselenide nanosheets, the cell survival rate gradually increased. When the concentration was 50μg / mL, the cell survival rate was 75%, which was 30% higher than the blank group (0μg / mL). Fig.19 The figure is a bar graph showing the cytotoxicity of 1.3 mM hydrogen peroxide and different concentrations of F127-GaSe nanosheets.

[0113] Those skilled in the art should understand that the present invention is not limited to the above-mentioned embodiments. The above-mentioned embodiments and descriptions are only for explaining the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention may be subject to various changes and improvements, and these changes and improvements shall all fall within the scope of the present invention claimed for protection.

Claims

1. An antibacterial and antioxidant gallium selenide nanosheet, characterized in that: Using bulk gallium triselenide as a raw material, nanometer-scale gallium triselenide nanosheets are prepared by a one-step liquid phase ultrasonic exfoliation method; the nanometer-scale gallium triselenide nanosheets have a radial size of 10-100nm and a sheet thickness of 2-5nm.

2. A method for preparing antibacterial and antioxidant gallium selenide nanosheets, characterized in that: The specific preparation method is as follows: Using bulk gallium triselenide as raw material, nanometer-scale gallium triselenide nanosheets were prepared by a one-step liquid phase ultrasonic exfoliation method.

3. The method for preparing the antibacterial and antioxidant gallium selenide nanosheets according to claim 2, characterized in that: Mixing bulk gallium triselenide with a dispersant to prepare a dispersion; The dispersion liquid is subjected to ultrasonic treatment to obtain a gallium triselenide nanosheet dispersion liquid, and then centrifuged and purified to obtain gallium triselenide nanosheets.

4. The method for preparing the antibacterial and antioxidant gallium selenide nanosheets according to claim 3, characterized in that: The dispersant includes any one of water, N-methylpyrrolidone, isopropanol, and ethanol, or a combination of two or more thereof; And / or, the mass concentration of gallium triselenide in the dispersion is 2-100 mg / mL; And / or, the ultrasonic device used in the ultrasonic treatment includes at least any one of a cell disruptor and an ultrasonic cleaner; And / or, the temperature of the ultrasonic treatment is -20-25°C; And / or, the ultrasonic treatment time is 0.5h-24h; And / or, the ultrasonic power of the ultrasonic treatment is 100-1000W; And / or, the centrifugal purification comprises: firstly removing large particles in the dispersion of gallium triselenide nanosheets by low-speed centrifugation at 500-10000 rpm, and then purifying by high-speed centrifugation at 15000-20000 rpm.

5. An application of an antibacterial and antioxidant gallium selenide nanosheet, characterized in that: The antibacterial and antioxidant gallium triselenide nanosheet is used in antibacterial, antioxidant or cell protection.

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