Natural antibacterial nanocellulose based on apocynum venetum and preparation method thereof

Abstract

The invention discloses natural antibacterial nanocellulose based on apocynum venetum and a preparation method of the natural antibacterial nanocellulose. The preparation method comprises the following steps: adding sodium bromide, 2, 2, 6, 6-tetramethylpiperidine oxide and sodium hypochlorite into a buffer solution, adjusting the pH value to prepare a precursor solution, adding apocynum venetum, carrying out high-temperature stirring reaction, sequentially filtering, washing and centrifuging until a neutral precipitate is obtained, adding deionized water, and carrying out crushing treatment to obtain a nano-cellulose suspension; and finally freeze-drying to obtain the nano cellulose. The nano cellulose has the advantages of high surface ratio, good biocompatibility, thermal stability, degradability, antibacterial property and the like, the preparation method is simple, the cost is low, the nano cellulose shows excellent properties in the aspects of antibacterial or mechanical properties and the like, and the nano cellulose is a biomass nano material with wide prospects in the fields of biomedicine and environmental protection.

Description

Technical Field

[0001] The invention belongs to the technical field of preparation of nanomaterials and relates to a natural antibacterial nanocellulose based on apocynum venetum and a preparation method thereof. Background Art

[0002] Cellulose, as one of the most abundant renewable resources, comes from various organisms such as plants (trees), microorganisms (algae, fungi, bacteria) and animals (cysts). Cellulose is a polysaccharide composed of multiple linear (1,4) linked β-D-glucan chains, each of which has three active hydroxyl groups. In addition, due to its highly crystalline polysaccharide structure, wide availability, excellent biodegradability, outstanding biocompatibility and good mechanical properties, the preparation of various functional materials has attracted widespread attention from researchers. Currently, cellulose and its derivatives have been widely used in various fields such as biomedicine, bioelectronics, energy, textiles and papermaking.

[0003] The current technologies for preparing nanocellulose based on Apocynum venetum have certain defects and shortcomings. For example, the nanocellulose prepared by mechanical methods will have uneven size distribution and high energy consumption of the equipment; the strong acid hydrolysis method will discharge a large amount of acidic waste liquid, which will damage the environment, and some cellulose will be degraded; although the enzymatic hydrolysis method is environmentally friendly, it has a slow reaction rate, a long cycle, and is significantly affected by factors such as temperature and pH, resulting in low production efficiency and high cost. Summary of the Invention

[0004] In view of this, in order to solve the problems existing in the background technology, the present invention provides a method for preparing nanocellulose by oxidation of Apocynum venetum raw material.

[0005] The preparation method of the present invention is simple and quick, and the prepared nanocellulose has good biocompatibility, antibacterial properties, and high surface area ratio, and has broad prospects in the fields of aerogels, hydrogels, and biomedicine.

[0006] The technical solution adopted by the present invention comprises the following steps:

[0007] (1) Preparation of precursor solution:

[0008] Sodium bromide, 2,2,6,6-tetramethylpiperidinyl oxide and sodium hypochlorite are added to a buffer solution and the pH value is adjusted to prepare a precursor solution, and then apocynum venetum is added to the precursor solution for high temperature stirring reaction;

[0009] (2) Filtration and centrifugation:

[0010] The solution prepared in step (1) is filtered, washed and centrifuged in sequence until a neutral precipitate is obtained;

[0011] (3) Ultrasonic dispersion:

[0012] The precipitate obtained in step (2) was placed in a beaker and deionized water was added, and the precipitate was placed in an ultrasonic cell crusher for crushing to obtain a nanocellulose suspension;

[0013] (4) Freeze-drying and grinding:

[0014] The nanocellulose suspension treated in step (4) is freeze-dried for two days to obtain nanocellulose.

[0015] The step (1) is specifically as follows:

[0016] First, sodium carbonate, sodium bicarbonate and ultrapure water are mixed to form a buffer solution;

[0017] Then, sodium bromide and 2,2,6,6-tetramethylpiperidinium oxide (Tempo) are added to the buffer solution, and the pH value of the buffer solution is adjusted to 9-11 by adding sodium hydroxide, preferably set to 10±0.5, and the above solution is stirred until it becomes clear to form a clear solution;

[0018] Then, a sodium hypochlorite solution is added to the clear solution to obtain a precursor solution;

[0019] Finally, the apocynum venetum is chopped and added into the precursor solution. The apocynum venetum will gradually dissolve and react in the precursor solution, and the mixture is stirred at 30-60° C. for 4-8 hours.

[0020] In the step (1), the mass volume ratio of sodium carbonate, sodium bicarbonate, sodium bromide, 2,2,6,6-tetramethylpiperidinium oxide, sodium hypochlorite solution, and ultrapure water is 0.9-1g:0.1-0.2g:0.1-0.15g:0.015-0.02g:10-20mL:90-110mL.

[0021] In the buffer solution, the mass ratio of sodium carbonate to sodium bicarbonate is (9-10):(1-2).

[0022] In the step (1), the aspect ratio of the chopped apocynum venetum is required to be greater than 50, so that the apocynum venetum can be more conducive to the dissolution reaction in the precursor solution.

[0023] The step (2) is specifically as follows:

[0024] The solution prepared in step (1) is filtered to remove the supernatant to obtain a precipitate, and the precipitate is washed and then added with deionized water and centrifuged repeatedly until the solution of the precipitate with deionized water reaches neutrality.

[0025] Repeated operation refers to repeatedly washing and centrifuging the precipitate.

[0026] In the step (3), the ratio of the precipitate to the deionized water is set so that the mass concentration of the precipitate in the nanocellulose suspension is 1:5;

[0027] In the step (3), the ultrasonic cell disruptor is operated at 550 to 650 W with a working cycle of 2 to 3 seconds on and 2 to 3 seconds off for 10 to 30 minutes to obtain a nanocellulose suspension.

[0028] In the step (4), the nanocellulose suspension is poured into a culture dish and placed in a refrigerator for 10 to 14 hours, taken out and then placed in a cold dryer for 1 to 2 days, and finally ground completely to obtain nanocellulose.

[0029] In the step (2), before filtering the precipitated product of the reaction, a terminator is added to terminate the oxidation reaction, and the terminator is an ethanol solution with a volume fraction of 90-98%.

[0030] The present invention adopts an innovative alkaline dissolution treatment for Apocynum venetum, rather than the strong acid dissolution treatment of the prior art, which can avoid excessive degradation of some cellulose, bring high surface charge and functionalization potential, and achieve advantages such as excellent mechanical properties and intact cellulose structure.

[0031] The invention prepares a buffer solution of sodium carbonate and sodium bicarbonate; then adds sodium bromide and Tempo (2,2,6,6-tetramethylpiperidinyl oxide) into the buffer solution; finally, puts apocynum venetum into the mixed solution and continuously stirs it under certain temperature conditions to prepare nanocellulose with a high surface area ratio and antibacterial properties.

[0032] The present invention has the following beneficial effects:

[0033] The invention controls the particle size of nano cellulose extracted from apocynum venetum by adjusting the temperature and time of different water bath heating.

[0034] The invention realizes different aspect ratios and carboxyl contents of nanocellulose by adding different amounts of sodium hypochlorite to oxidize primary alcohol hydroxyl groups in cellulose molecules and converting them into carboxyl groups.

[0035] The nanocellulose synthesized under alkaline conditions has a uniform and adjustable diameter distribution, a diameter of 10-30 nm, a high specific surface area and good antibacterial properties.

[0036] In summary, the present invention can control the diameter of nanocellulose by varying the content of sodium hypochlorite solution, and varying the oxidation temperature and time. The preparation method is simple, the reaction conditions are mild, and it is easy to operate. The nanocellulose produced by the method of the present invention appears as a pale white powder with a needle-like structure at the nanoscale and a controllable diameter. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 、 3 , 5 are scanning electron microscope images of nanocellulose prepared in Examples 1, 3, and 5 respectively

[0038] Figure 2 、 4 , 6 are the diameter distribution diagrams of the nanocellulose prepared in Examples 1, 3, and 5, respectively.

[0039] Figure 7 、 8 The antibacterial test diagrams of nanocellulose prepared in Examples 2 and 4 are

[0040] Figure 9 This is the infrared spectrum of the nanocellulose prepared in Comparative Example 6.

[0041] Figure 10 1 and 2 are scanning electron microscope images of nanocellulose prepared in comparative examples 1 and 2.

[0042] Figure 11 , 13 are the diameter distribution diagrams of the nanocellulose prepared in comparative examples 1 and 2, respectively. DETAILED DESCRIPTION

[0043] The present invention is further described below by way of examples.

[0044] The embodiments of the present invention are as follows:

[0045] Example 1:

[0046] 0.35 g of Apocynum venetum was chopped, 0.85 g of sodium carbonate, 0.1 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.012 g of Tempo and 0.1 g of sodium bromide were added to the buffer solution, sodium hydroxide was added to adjust the pH value of the buffer solution to 9, and the solution was stirred until a clear solution was obtained. Then, 8 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 40 ° C for 4 h to react.

[0047] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 6000 rpm / min. Finally, 3 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2-3 seconds on and 2-3 seconds off for 12 minutes to obtain a nanocellulose suspension with a concentration of 16%;

[0048] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0049] The diameter of the nanocellulose in Example 1 is as follows. Figure 1 , as shown in Figure 2, it can be seen that the diameter of the treated sample reaches about 25nm.

[0050] Example 2:

[0051] 0.5 g of Apocynum venetum was chopped, 0.9 g of sodium carbonate, 0.1 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.015 g of Tempo and 0.12 g of sodium bromide were added to the buffer solution, sodium hydroxide was added to adjust the pH value of the buffer solution to 9, and the solution was stirred until a clear solution was obtained. Then, 9 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 45 ° C for 4.5 h to react.

[0052] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 6500 rpm / min. Finally, 3.5 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2-3 seconds on and 2-3 seconds off for 15 minutes to obtain a nanocellulose suspension with a concentration of 14%;

[0053] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0054] The antibacterial effect of the nanocellulose in Example 2 is as follows Figure 9 As shown in the figure, it can be seen that the antibacterial rate of the treated sample against S. aureus is about 85% at 40 mg / mL, and the antibacterial rate against E. coli is about 40%.

[0055] Example 3:

[0056] 0.5 g of Apocynum venetum was chopped, 0.92 g of sodium carbonate, 0.12 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.016 g of Tempo and 0.11 g of sodium bromide were added to the buffer solution, sodium hydroxide was added to adjust the pH value of the buffer solution to 9.5, and the solution was stirred until a clear solution was obtained. Then, 9.5 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 50 ° C for 5 h to react.

[0057] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 7000 rpm / min. Finally, 4 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2 to 3 seconds on and 2 to 3 seconds off for 18 minutes to obtain a nanocellulose suspension with a concentration of 12.5%;

[0058] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0059] The diameter of the nanocellulose in Example 3 is as follows. Figure 3 ,4, it can be seen in the figure that the diameter of the treated sample reaches about 20nm.

[0060] Example 4:

[0061] 0.5 g of Apocynum venetum was chopped, 0.93 g of sodium carbonate, 0.11 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.017 g of Tempo and 0.12 g of sodium bromide were added to the buffer solution, sodium hydroxide solution was added to adjust the pH value of the buffer solution to 10, and the solution was stirred until a clear solution was obtained. Then, 10 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 55 ° C for 5.5 h to react.

[0062] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 7500 rpm / min. Finally, 4.5 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2-3 seconds on and 2-3 seconds off for 20 minutes to obtain a nanocellulose suspension with a concentration of 14%;

[0063] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0064] The antibacterial effect of the nanocellulose in Example 4 is as follows Figure 10As shown in the figure, it can be seen that the antibacterial rate of the treated sample against S. aureus is about 90% at 80 mg / mL, and the antibacterial rate against E. coli is about 75%.

[0065] Example 5:

[0066] 0.5 g of Apocynum venetum was chopped, 0.94 g of sodium carbonate, 0.12 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.018 g of Tempo and 0.13 g of sodium bromide were added to the buffer solution, sodium hydroxide solution was added to adjust the pH value of the buffer solution to 10, and the solution was stirred until a clear solution was obtained. Then, 11 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 60 ° C for 6 h to react.

[0067] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 8000 rpm / min. Finally, 5 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2 to 3 seconds and a rest period of 2 to 3 seconds for 22 minutes to obtain a nanocellulose suspension with a concentration of 10%;

[0068] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0069] The diameter of the nanocellulose in Example 5 is as follows. Figure 5 ,6, it can be seen in the figure that the diameter of the treated sample reaches about 28nm.

[0070] Example 6:

[0071] 0.5 g of Apocynum venetum was chopped, 0.95 g of sodium carbonate, 0.13 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.02 g of Tempo and 0.14 g of sodium bromide were added to the buffer solution, sodium hydroxide was added to adjust the pH value of the buffer solution to 11, and the solution was stirred until a clear solution was obtained. Then, 12 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 65 ° C for 6.5 h to react.

[0072] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 9000 rpm / min. Finally, 6 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2-3 seconds on and 2-3 seconds off for 24 minutes to obtain a nanocellulose suspension with a concentration of 8.3%;

[0073] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0074] The FITR of the nanocellulose in Example 6 is as follows: Figure 11 As shown in the figure, it can be seen that the carboxylate groups were successfully introduced into the treated sample, and the -1 There is an asymmetric stretching vibration peak of carboxylate.

[0075] Comparative Example 1:

[0076] 0.5 g of Apocynum venetum was chopped, 0.94 g of sodium carbonate, 0.12 g of sodium bicarbonate and 100 mL of deionized water were mixed to form a buffer solution, 0.018 g of Tempo and 0.13 g of sodium bromide were added to the buffer solution, sodium hydroxide solution was added to adjust the pH value of the buffer solution to 10, and the solution was stirred until a clear solution was obtained. Then, 11 mL of sodium hypochlorite solution was added to form a precursor solution, and Apocynum venetum was placed in the precursor solution and heated at 60 ° C for 10 h to react.

[0077] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 8000 rpm / min. Finally, 5 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2 to 3 seconds and a rest period of 2 to 3 seconds for 22 minutes to obtain a nanocellulose suspension with a concentration of 10%;

[0078] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0079] The diameter of the nanocellulose in comparative example 1 is as follows: Figure 7 ,8, it can be seen in the figure that the diameter of the treated sample reaches about 31nm.

[0080] Comparative Example 2:

[0081] 0.5 g of Apocynum venetum was chopped and put into a 4 M hydrochloric acid solution with a solid-liquid ratio of 1:15. The solution was treated in a water bath at 80° C. at a speed of 500 rpm for 4 h.

[0082] After the reaction, the supernatant was removed to obtain a precipitate, which was repeatedly washed with deionized water and centrifuged until neutral at a centrifuge speed of 8000 rpm / min. Finally, 5 mL of deionized water was added and the mixture was placed in an ultrasonic cell disruptor at 500 W with a working cycle of 2 to 3 s and a rest period of 2 to 3 s for 22 minutes to obtain a nanocellulose suspension with a concentration of 10%.

[0083] The nanocellulose suspension was placed in a refrigerator and frozen for 12 hours. After being taken out, it was placed in a cold dryer and freeze-dried for two days. Finally, it was completely ground to obtain nanocellulose.

[0084] The diameter of the nanocellulose in comparative example 2 is as follows: Figure 12 , as shown in Figure 13, it can be seen that the diameter of the sample after treatment can only reach about 18.07μm.

[0085] The nanocellulose produced by the present invention exhibits advantages such as a high surface area ratio, good biocompatibility, thermal stability, degradability, and antibacterial properties. Furthermore, the preparation method is simple and cost-effective, and the product exhibits excellent antibacterial and mechanical properties, making it a promising biomass nanomaterial in the fields of biomedicine and environmental protection.

[0086] The above specific embodiments are used to illustrate the present invention rather than to limit the present invention. Any modifications and changes made to the present invention within the spirit of the present invention and the protection scope of the claims shall fall within the protection scope of the present invention.

[0087] The above description is only a preferred embodiment of the present invention. Therefore, any equivalent changes or modifications made according to the structure, characteristics and principles described in the scope of the patent application of the present invention are included in the scope of the patent application of the present invention.

Claims

1. A method for preparing natural antibacterial nanocellulose based on Apocynum venetum, characterized in that: The steps of this method are as follows: (1) Preparation of precursor solution: Sodium bromide, 2,2,6,6-tetramethylpiperidinyl oxide and sodium hypochlorite are added to a buffer solution and the pH value is adjusted to prepare a precursor solution, and then apocynum venetum is added to the precursor solution for high temperature stirring reaction; (2) Filtration and centrifugation: The solution prepared in step (1) is filtered, washed and centrifuged in sequence until a neutral precipitate is obtained; (3) Ultrasonic dispersion: The precipitate obtained in step (2) is added with deionized water and placed in an ultrasonic cell crusher for crushing to obtain a nanocellulose suspension; (4) Freeze-drying and grinding: The nanocellulose suspension treated in step (4) is freeze-dried to obtain nanocellulose.

2. The method for preparing a natural antibacterial nanocellulose based on Apocynum venetum according to claim 1, characterized in that: The step (1) is specifically as follows: First, sodium carbonate, sodium bicarbonate and ultrapure water are mixed to form a buffer solution; Then, sodium bromide and 2,2,6,6-tetramethylpiperidinium oxide are added to the buffer solution, the pH value of the buffer solution is adjusted to 9-11 by adding sodium hydroxide, and the above solution is stirred until it becomes clear to form a clear solution; Then, a sodium hypochlorite solution is added to the clear solution to obtain a precursor solution; Finally, the apocynum venetum is chopped and added into the precursor solution, and stirred at 30-60° C. for 4-8 hours.

3. The method for preparing a natural antibacterial nanocellulose based on Apocynum venetum according to claim 1 or 2, characterized in that: In the step (1), the mass volume ratio of sodium carbonate, sodium bicarbonate, sodium bromide, 2,2,6,6-tetramethylpiperidinium oxide, sodium hypochlorite solution, and ultrapure water is 0.9-1g:0.1-0.2g:0.1-0.15g:0.015-0.02g:10-20mL:90-110mL.

4. The method for preparing a natural antibacterial nanocellulose based on Apocynum venetum according to claim 1, characterized in that: In the step (1), the aspect ratio of the chopped apocynum venetum is greater than 50, so that the apocynum venetum can be more conducive to the dissolution reaction in the precursor solution.

5. The method for preparing a natural antibacterial nanocellulose based on Apocynum venetum according to claim 1, characterized in that: The step (2) is specifically as follows: The solution prepared in step (1) is filtered to remove the supernatant to obtain a precipitate, and the precipitate is washed and then added with deionized water and centrifuged repeatedly until the solution of the precipitate with deionized water reaches neutrality.

6. The method for preparing a natural antibacterial nanocellulose based on Apocynum venetum according to claim 1, characterized in that: In the step (3), the ratio of the precipitate to the deionized water is set so that the mass concentration of the precipitate in the nanocellulose suspension is 1:5; In the step (3), the ultrasonic cell disruptor is operated at 550 to 650 W with a working cycle of 2 to 3 seconds on and 2 to 3 seconds off for 10 to 30 minutes to obtain a nanocellulose suspension.

7. The method for preparing a natural antibacterial nanocellulose based on Apocynum venetum according to claim 1, characterized in that: In the step (4), the nanocellulose suspension is poured into a culture dish and placed in a refrigerator for 10 to 14 hours, taken out and then placed in a cold dryer for 1 to 2 days, and finally ground completely to obtain nanocellulose.

8. The method for preparing high surface area ratio nanocellulose according to claim 1, wherein: In the step (2), before filtering the precipitate, a terminator is added to terminate the reaction, and the terminator is an ethanol solution with a volume fraction of 90-98%.

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