Antibacterial cotton fabric based on thymol and method for its preparation

Abstract

The application discloses a thymol-based antibacterial cotton fabric and a preparation method thereof. A thymol small molecule ATRP initiator (Thymol-Br) is prepared through an acylation reaction of thymol; the thymol small molecule ATRP initiator is combined with a hydrophilic monomer and a silane coupling agent to prepare a thymol antibacterial copolymer Thymol-P(DMAEMA-r-MPS) through atom transfer radical polymerization (ATRP); and finally, the copolymer is grafted on the surface of the cotton fabric through a thermal curing reaction to form the antibacterial cotton fabric. The application improves the use stability of thymol by polymer modification of the natural product thymol, and the prepared antibacterial cotton fabric can kill drug-resistant bacteria, and has excellent and long-acting antibacterial effect.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of antibacterial surfaces, and particularly relates to a thymol-based antibacterial cotton fabric and a preparation method thereof. BACKGROUND

[0002] Globally, bacterial infection is still a serious threat to public health, causing huge economic losses to human society every year. Although the use of broad-spectrum antibiotics has saved millions of lives, due to the high adaptability of bacteria, the overuse and misuse of traditional antibiotics have led to an increasing increase in antibiotic resistance. According to statistics, more than 700,000 people die every year due to antibiotic resistance. The development of new antibiotics requires huge costs and a long cycle, and most importantly, even if new antibiotics are developed, the threat of drug-resistant bacteria cannot be solved. Therefore, it is urgent to reduce the use of antibiotics and develop new effective antibacterial agents with different antibacterial mechanisms from antibiotics.

[0003] Natural products play a very important role in the process of drug discovery and development, and many antibacterial agents are natural products or semi-synthetic variants. The broad-spectrum antibacterial activity of essential oils has been widely recognized, and among them, the phenolic natural essential oil component shows good antibacterial activity, and the best active one is thymol. The antibacterial mechanism of thymol is attributed to the change of cell membrane permeability, which causes important cell contents to leak and leads to bacterial death. However, thymol has some use limitations, such as poor stability, poor water solubility, and volatile characteristics. If directly added to the substrate, it is difficult to ensure that thymol can stably play a role; on the other hand, due to the fast release speed of essential oils, it cannot meet the requirements of long-term antibacterial infection of wounds in the medical field. In order to improve the use limitations, researchers have compensated for the defects of thymol by using emulsion stabilization systems to improve the use range and performance of thymol. However, the emulsion stabilization system is always a slow-release system, which can only improve but not completely solve the defects of low-temperature volatility and the like. SUMMARY

[0004] In order to solve the shortcomings and deficiencies of the prior art, the purpose of the present application is to provide a thymol-based antibacterial cotton fabric with excellent antibacterial effect and long-acting, which weakens the influence of volatility and water solubility of natural product small molecule thymol through polymer modification, improves the use stability, and expands the application range and use field.

[0005] Another purpose of the present application is to provide a preparation method of a thymol-based antibacterial cotton fabric, and the purpose of the present application is achieved by the following technical scheme:

[0006] The application discloses a thymol-based antibacterial cotton fabric, and relates to the technical field of antibacterial cotton fabrics.

[0007] The reaction molar ratio of the thymol-based ATRP initiator (Thymol-Br), the hydrophilic monomer and the silane coupling agent is 1:15:3-1:20:5, and preferably 1:20:4.

[0008] The application discloses a preparation method of the thymol-based antibacterial cotton fabric.

[0009] (1) thymol is dissolved in an organic solvent, and then reacts with an acyl halide derivative in an alkaline environment; after a period of reaction, purification and drying, a thymol-based ATRP initiator (Thymol-Br) is obtained;

[0010] (2) the thymol-based ATRP initiator (Thymol-Br), a hydrophilic monomer and a silane coupling agent are used as raw materials to perform an ATRP reaction under a catalytic system; after the reaction is completed, the reaction is stopped through air, the product is purified through a neutral alumina column after elution, and the thymol-based antibacterial copolymer Thymol-P (DMAEMA-r-MPS) is prepared after precipitation, standing and drying;

[0011] (3) the thymol-based antibacterial copolymer Thymol-P (DMAEMA-r-MPS) obtained in step (2) is weighed, 200 mL of a solvent is added to form a mixed solution with a concentration of 20-40 mg / mL; cotton fabric is immersed and coated for 5-10 minutes, and then the cotton fabric is taken out and dried and cured; after curing, the cotton fabric is washed with an organic solvent for three times, and then vacuum dried to obtain the antibacterial cotton fabric TP@CF.

[0012] In step (1), the acyl halide derivative is one of 2-bromoisobutyryl bromide, 2-bromopropionyl bromide, bromoacetyl bromide and 2-bromobutyryl bromide.

[0013] In step (1), the organic solvent is dichloromethane.

[0014] In step (1), the reaction molar ratio of the thymol and the acyl halide derivative is 1:1.5-1:3.

[0015] In step (1), the reaction time is 4-12 hours, and the reaction temperature is 20-30℃.

[0016] In step (1), the drying process is drying in a vacuum drying box at 40℃ for 12 hours.

[0017] In step (2), the hydrophilic monomer is N,N-dimethylaminoethyl methacrylate (DMAEMA),

[0018] 3-(dimethylamino) methyl propyl methacrylate, methacryloxyethyl trimethyl ammonium chloride, wherein the silane coupling agent is one of γ-methacryloxypropyl trimethoxysilane (KH570), methacryloxypropyl triethoxysilane, and methacryloxypropyl triisopropoxysilane.

[0019] In step (2), the molar ratio of the thymol small molecule ATRP initiator (Thymol-Br) to the catalytic system is 1:1-1:2, and the catalytic system is cuprous bromide / PMDETA or cuprous chloride / Me6Tren.

[0020] In step (2), the reaction temperature is 40-50℃, and the reaction time is 6-24 hours.

[0021] In step (2), the precipitation process is precipitation in anhydrous ice n-hexane, the standing temperature is -20-0℃, and the drying process is drying in a vacuum drying box at 50℃ for 6 hours.

[0022] In step (3), the drying and curing process is curing at 120℃ in a vacuum drying box for 2 hours.

[0023] In step (3), the solvent is toluene, and the organic solvent is tetrahydrofuran, methanol, or acetone.

[0024] Compared with the prior art, the present application has the following advantages and beneficial effects:

[0025] (1) The present application modifies the natural product small molecule thymol by polymerization, which reduces the influence of its volatility and water solubility, improves its use stability, expands its application range and use field, and is more green and healthy.

[0026] (2) The present application first synthesizes a thymol-based antibacterial cotton fabric surface, and the preparation method is simple and efficient, has a wide application range, and has excellent and long-acting antibacterial effect on cotton fabric, and can kill drug-resistant bacteria.

[0027] (3) The present application uses ATRP method to prepare polymers, and can obtain natural product antibacterial polymers with controllable molecular weight. Attached Figure Description

[0028] Figure 1 This is a schematic diagram of the synthesis of the small molecule ATRP initiator (Thymol-Br) of thymol.

[0029] Figure 2 This is a schematic diagram of the synthesis of the thymol antibacterial copolymer Thymol-P (DMAEMA-r-MPS).

[0030] Figure 3 This is the NMR spectrum of the small molecule ATRP initiator (Thymol-Br) of thymol.

[0031] Figure 4 This is the NMR spectrum of the thymol antibacterial copolymer Thymol-P (DMAEMA-r-MPS).

[0032] Figure 5 This is a flowchart of the preparation process for antibacterial cotton fabric (TP@CF).

[0033] Figure 6 These are scanning electron microscope images of the antibacterial cotton fabric (TP@CF) prepared in the embodiments of this application, the cotton fabric (MP@CF) prepared in the comparative example of this application, and the cotton fabric (Pristine CF) without any treatment.

[0034] Figure 7 These are images showing the antibacterial results of the oscillation method for the antibacterial cotton fabric (TP@CF) prepared in the embodiments of this application, the antibacterial cotton fabric (MP@CF) prepared in the comparative example of this application, and the cotton fabric (Pristine CF) without any treatment.

[0035] Figure 8 This is a diagram showing the inhibition zones of three bacteria against the antibacterial cotton fabric TP@CF.

[0036] Figure 9 This is a graph showing the bactericidal rates of antimicrobial cotton fabric (TP@CF) and standard antimicrobial cotton fabric (AAA Class) against three types of bacteria.

[0037] Figure 10 The diagram shows the bactericidal kinetics of the antibacterial cotton fabric (TP@CF) prepared in the embodiments of this application, the antibacterial cotton fabric (MP@CF) prepared in the comparative example of this application, and the cotton fabric (Pristine CF) without any treatment. Detailed Implementation

[0038] The present invention will be further described in detail below with reference to embodiments, but the embodiments of the present invention are not limited thereto. All raw materials involved in the present invention can be purchased directly from the market. For process parameters not specifically specified, conventional techniques can be referred to.

[0039] Preparation of thymol small molecule ATRP initiator (Thymol-Br)

[0040] Synthesis of thymol small molecule ATRP initiator: build a reflux device, 10 grams of thymol, 16.28 grams of 4-dimethylamino pyridine were dissolved in 200 mL of dichloromethane, after stirring and dissolving, 18.4 mL of triethylamine was added into a 500 mL three-necked flask in turn, then the system was mechanically stirred under the condition of ice water bath and N2 protection, 16.4 mL of 2-bromoisobutyryl bromide mixed solution diluted with dichloromethane was added dropwise, after the addition was completed, the ice water bath was removed, and the reaction was continued at room temperature for 4-12 hours. After the reaction was completed, the white solid bromate generated in the reaction was first removed by filtration, the filtrate was concentrated by rotary evaporation, and the aqueous solution was extracted with ether several times, then the upper organic phase was collected after standing and separating. The separated product was washed with 0.1 mol / L hydrochloric acid solution and saturated sodium bicarbonate solution in turn. The upper liquid obtained by extraction was dehydrated with anhydrous magnesium sulfate overnight, filtered, and concentrated by rotary evaporation to obtain the crude product. The crude product was purified by silica gel column, and the eluent was petroleum ether / ethyl acetate=98:2 (V / V). The purified product was concentrated by rotary evaporation to obtain the final product. The product was placed in a vacuum drying oven and treated at 40°C for 12 hours to obtain a light yellow transparent liquid product with a yield of 95%.

[0041] Preparation of thymol antibacterial copolymer Thymol-P(DMAEMA-r-MPS)

[0042] Check the airtightness of the Schlenk reaction tube, and after checking that there is no error, add 1.0 gram of thymol small molecule ATRP initiator, 11.21 mL of dimethylaminoethyl methacrylate, 3.17 mL of γ-methacryloyloxypropyl trimethoxysilane, 1.39 mL of N,N,N',N,'N"-pentamethyldiethylenetriamine, 15 mL of anhydrous toluene into the Schlenk reaction tube in turn under nitrogen environment, and perform freeze-thaw degassing treatment. Then, 0.95 grams of purified cuprous bromide powder is added under nitrogen environment, and freeze-thaw degassing cycle is performed again for three times to create an oxygen-free environment for the ATRP system. Finally, the Schlenk reaction tube under nitrogen environment is placed in a 40°C oil bath for heating and stirring for 12 hours. After the reaction is completed, air is introduced under ice water conditions for quenching. After quenching, the product is purified by a slightly shorter neutral alumina column with anhydrous tetrahydrofuran as the eluent. The purified product obtained after concentration by rotary evaporation is precipitated dropwise in anhydrous ice n-hexane, and the precipitated product is placed in a low temperature environment and the supernatant is removed. Finally, it is placed in a vacuum drying oven and treated at 50°C for 6 hours to obtain the thymol antibacterial copolymer.

[0043] Preparation of antibacterial cotton fabric TP@CF

[0044] A certain amount of thymol antibacterial copolymer Thymol-P(DMAEMA-r-MPS) was taken into a 500 mL round-bottom flask, 200 mL of toluene was added to form a solution with a concentration of 20-40 mg / mL. After 5-10 minutes of immersion coating, the cotton fabric was taken out and placed in a vacuum drying oven at 120°C for 2 hours. After curing, the cotton fabric was washed with tetrahydrofuran, methanol, and acetone in turn three times, and then vacuum dried to obtain the antibacterial cotton fabric TP@CF.

[0045] Preparation of antibacterial cotton fabric MP@CF of Comparative Example 1

[0046] The airtightness of the Schlenk reaction tube was checked, and after no error was found, 1.0 grams of 2-bromoisobutyrate (MBiB) initiator, 11.21 mL of dimethylaminoethyl methacrylate, 3.17 mL of γ-methacryloyloxypropyl trimethoxysilane, 1.39 mL of N,N,N',N,'N"-pentamethyldiethylenetriamine, 15 mL of anhydrous toluene were sequentially added to the Schlenk reaction tube under a nitrogen environment, and freeze-thaw degassing was performed. Then, 0.95 grams of purified cuprous bromide powder was added under a nitrogen environment, and freeze-thaw degassing was performed again for three cycles to create an oxygen-free environment for the ATRP system. Finally, the Schlenk reaction tube under a nitrogen environment was placed in a 40°C oil bath for heating and stirring for 12 hours. After the reaction was completed, air was introduced under ice water conditions for quenching. After quenching, the product was purified by passing through a short neutral alumina column using anhydrous tetrahydrofuran as the eluent. After concentration by rotary evaporation, the purified product was dropped into anhydrous ice-n-hexane drop by drop, and the precipitated product was placed in a low-temperature environment and the supernatant was removed. Finally, it was placed in a vacuum drying oven and treated at 50°C for 6 hours to obtain the P(DMAEMA-r-MPS) copolymer.

[0047] A certain amount of P(DMAEMA-r-MPS) antibacterial copolymer was taken into a 500 mL round-bottom flask, 200 mL of toluene was added to form a solution with a concentration of 20-40 mg / mL. After 5-10 minutes of immersion coating, the cotton fabric was taken out and placed in a vacuum drying oven at 120°C for 2 hours. After curing, the cotton fabric was washed with tetrahydrofuran, methanol, and acetone in turn three times, and then vacuum dried to obtain the antibacterial cotton fabric MP@CF.

[0048] Experiment 1: Oscillation method

[0049] The oscillation method refers to the FZ / T73023-2006 standard. The oscillation method can be applied to samples with low water absorption requirements, fibrous, powdery, hairy or feather, uneven fabrics, etc., and is especially suitable for determining the antibacterial performance of non-overflow type fabrics. The specific operation of this experiment is as follows: 1) Staphylococcus aureus (S. aureus), Escherichia coli (E. coli), and methicillin-resistant Staphylococcus aureus (MRSA) were inoculated into LB broth culture medium and incubated in a constant temperature shaker at 200 rpm and 37°C overnight; 2) 50 μL of each of the three groups of bacterial suspension cultured to the logarithmic phase (1×10 8 CFU mL -1 ) overnight was transferred to PBS buffer salt solution, and 0.1 g of antibacterial cotton fabric TP@CF, antibacterial cotton fabric MP@CF, and untreated cotton fabric (Pristine CF) was added to each group, and the total volume of the mixed culture solution was 20 mL; 3) The mixed culture solution was incubated in a constant temperature shaker at 200 rpm and 37°C; 4) After 24 hours of incubation, 100 μL of the mixed culture solution was coated, and after the surface of the culture dish was free of water particles, the culture dish was inverted and incubated in a constant temperature incubator at 37°C for 24 hours; 5) The bacterial growth was observed.

[0050] Figure 7 The survival of the three bacteria in the three cotton fabrics after 24 hours of co-culture is shown, which is characterized by the spread plate method. Figure 7 It can be found that the two functionalized cotton fabrics TP@CF and MP@CF have excellent antibacterial and bactericidal effects on gram-positive bacteria (S. aureus), and can achieve a bactericidal rate of 98% after 24 hours of co-culture.

[0051] The drug-resistant bacteria (MRSA) are more difficult to be completely killed than the gram-positive bacteria (S. aureus), but TP@CF still exhibits excellent bactericidal performance. The results of gram-negative bacteria (E. coli) show a polarized phenomenon, although the antibacterial effect is slightly insufficient compared to gram-positive bacteria (S. aureus), but the gram-negative bacteria (E. coli) in the TP@CF group are mostly killed after 24 hours of co-culture; the results of the MP@CF group are quite different, and the experimental results show that MP@CF has no killing effect on gram-negative bacteria (E. coli), and the plate is covered with more dense gram-negative bacteria (E. coli) after 24 hours of co-culture. In addition, based on the FZ / T73023-2006 standard, TP@CF still exhibits excellent antibacterial performance after 70 washes, indicating that TP@CF has durable performance.

[0052] Experiment 2 Zone of Inhibition (ZOI)

[0053] The inhibition zone test referenced the Kirby-Bauer method

[98] . The inhibition zone test, also known as the diffusion method, is essentially a method that uses the diffusion of drugs in an agar plate to inhibit or kill surrounding bacteria. It is a test that can be used to determine whether an antibacterial surface is spill-type. The specific operation of this experiment is as follows: 1) Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus (MRSA) were inoculated into LB broth culture medium and cultured overnight in a constant temperature shaker at 200 rpm and 37°C; 2) the overnight culture was taken to the mid-log phase (1×10 8 CFU mL -1 1) Spread 100 μL of each of the three bacterial suspensions onto a solid culture medium; 2) Place antibacterial cotton fabric TP@CF (1 cm × 1 cm) flat on the solid culture medium and gently press it with tweezers to ensure close contact between the cotton fabric and the solid culture medium; 3) Cover with sealing film and incubate in a constant temperature incubator for 24 hours. Take it out and observe whether there is an inhibition zone and calculate the size of the inhibition zone.

[0054] Figure 8 The results of the inhibition zone test on three types of bacteria using antibacterial cotton fabric TP@CF were obtained from... Figure 8 It can be seen that after 24 hours of incubation in a constant temperature incubator, no inhibition zones were produced in the three bacterial experimental groups of the antibacterial cotton fabric TP@CF, indicating that the antibacterial mode of TP@CF is not release sterilization, but contact sterilization, and TP@CF is not an overflow cotton fabric.

[0055] Experiment 3 Oscillation-Time Sterilization Experiment

[0056] Referring to the oscillation method and time-based sterilization curves, the applicant designed an oscillation-time sterilization experiment for cotton fabrics. This experiment effectively combines the intuitive results of oscillation plate coating with the kinetic study of time-based sterilization. The specific experimental procedures are as follows: 1) Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus (MRSA) were inoculated into LB broth and cultured overnight in a constant temperature shaker at 200 rpm and 37°C; 2) The overnight culture was collected at mid-log (1×10⁻⁶). 8 CFU mL -1Three groups of 50 μL each of the three bacterial suspensions were transferred to PBS buffer solution. At the same time, 0.1 g each of antibacterial cotton fabric TP@CF, antibacterial cotton fabric MP@CF, and untreated cotton fabric (PristineCF) were added to each group, and the total volume of the mixed culture medium was 20 mL. 3) The mixed culture medium was placed in a constant temperature shaker at 200 rpm and 37℃. 4) At certain time intervals (6 time points of 2, 4, 6, 8, 10 and 24 hours in this experiment), 100 μL of the mixed culture medium was taken and plated. After there were no water droplets on the surface of the culture dish, it was covered with sealing film and inverted in a constant temperature incubator at 37℃ for 24 hours. 5) The bacterial growth was observed.

[0057] Depend on Figure 10 Data on bactericidal kinetics were obtained. TP@CF showed good bactericidal effects against all three types of bacteria, but it was clearly found that TP@CF was far more efficient at killing Gram-positive bacteria (S. aureus) than Gram-negative bacteria (E. coli). In the S. aureus and MRSA experimental groups, TP@CF began to show bactericidal effects within 4 hours, and was able to kill most of the bacteria within 8 hours. In the E. coli experimental group, although TP@CF also began to show bactericidal effects within 4 hours, its bactericidal efficiency was not comparable to that of the other two bacteria. Even after 24 hours of co-culture, although most of the E. coli had been killed, the bacteria were not completely eradicated. As for the MP@CF group, although it showed good bactericidal performance against Gram-positive bacteria (S. aureus) and drug-resistant bacteria (MRSA), its bactericidal effect and efficiency were slightly inferior to those of the TP@CF group. Most importantly, the results of the oscillation-time sterilization experiment show that the MP@CF group did not even have an inhibitory effect on E. coli.

Claims

1. A method for preparing thymol-based antibacterial cotton fabric, characterized by: The preparation method comprises the following steps: (1) thymol is dissolved in an organic solvent, and then reacts with an acyl halide derivative in an alkaline environment for a period of time, and then purified and dried to obtain a thymol small molecule ATRP initiator; (2) thymol small molecule ATRP initiator, hydrophilic monomer and silane coupling agent are used as raw materials to perform ATRP reaction under a catalytic system; after the reaction is completed, the reaction is stopped by air, and then the product is purified by a neutral alumina column, precipitated and dried to obtain thymol-based antibacterial copolymer Thymol-P; (3) the thymol-based antibacterial copolymer Thymol-P obtained in step (2) is weighed, 200 mL of solvent is added to form a mixed solution with a concentration of 20-40 mg / mL, and then cotton fabric is immersed and coated for 5-10 minutes, and then taken out for drying and curing; after curing, the cotton fabric is washed with an organic solvent three times, and then vacuum dried to obtain antibacterial cotton fabric TP@CF; In step (1), the acyl halide derivative is one of 2-bromoisobutyryl bromide, 2-bromopropionyl bromide, bromoacetyl bromide and 2-bromobutyryl bromide; the molar ratio of thymol to acyl halide derivative is 1:1.5-1:3; and the reaction time is 4-12 hours. In step (2), the molar ratio of thymol small molecule ATRP initiator, hydrophilic monomer and silane coupling agent is 1:15:3-1:20:5; the hydrophilic monomer is one of N,N-dimethylaminoethyl methacrylate, 3-(dimethylamino) methyl methacrylate and methacryloyloxyethyl trimethylammonium chloride; and the silane coupling agent is one of γ-methacryloyloxypropyl trimethoxysilane, methacryloyloxypropyl triethoxysilane and methacryloyloxypropyl triisopropoxysilane. In step (3), the drying and curing process is 120℃ curing for 2 hours in a vacuum drying oven.

2. The preparation method of the antibacterial cotton fabric based on thymol according to claim 1, wherein in step (2), the molar ratio of thymol small molecule ATRP initiator, hydrophilic monomer and silane coupling agent is 1:20:

4.

3. The preparation method of the antibacterial cotton fabric based on thymol according to claim 1, wherein in step (1), the organic solvent is dichloromethane.

4. The preparation method of the antibacterial cotton fabric based on thymol according to claim 1, wherein in step (1), the reaction temperature is 20-30℃; In step (1), the drying process is 12 hours of drying in a vacuum drying oven at 40℃.

5. The preparation method of the antibacterial cotton fabric based on thymol according to claim 1, wherein in step (2), the molar ratio of thymol small molecule ATRP initiator and the catalytic system is 1:1-1:2; and the catalytic system is cuprous bromide / PMDETA or cuprous chloride / Me6Tren. ​ ​ ​ ​ 6. The method for preparing thymol-based antibacterial cotton fabric according to claim 1, characterized in that: In step (2), the reaction temperature is 40-50°C, and the reaction time is 6-24 hours. In step (2), the precipitation process is precipitation in anhydrous ice n-hexane, the standing temperature is -20-0°C, and the drying process is drying in a vacuum drying oven at 50°C for 6 hours.

7. The method for preparing thymol-based antibacterial cotton fabric according to claim 1, characterized in that: In step (3), the solvent is toluene, and the organic solvent is tetrahydrofuran, methanol or acetone.

8. The thymol-based antibacterial cotton fabric prepared by the method of any one of claims 1-7.

Images