CN111733482A

CN111733482A - Preparation method and application of chitin nanofiber

Info

Publication number: CN111733482A
Application number: CN202010704075.3A
Authority: CN
Inventors: 李振亚; 尹新明; 苏丽娟
Original assignee: Henan Agricultural University
Current assignee: Henan Agricultural University
Priority date: 2020-07-16
Filing date: 2020-07-21
Publication date: 2020-10-02
Anticipated expiration: 2040-07-21
Also published as: CN111733482B

Abstract

The invention belongs to the field of new material preparation, and particularly relates to a preparation method and application of chitin nanofiber. Diluting with concentrated acid, adding chitin powder, heating for acidolysis, centrifuging, washing, resuspending, precipitating, and ultrasonic treating to obtain aqueous dispersion of chitin nanofiber, or lyophilizing to obtain solid powder. The chitin material is decomposed into microfibril in acid solution and then is ultrasonically crushed into nanometer fiber. The preparation method is simple, mild, efficient and controllable, reduces the using amount of acid liquor and the energy consumption of preparation, and simultaneously, the nano-fibers are uniform, good in dispersibility and suspension property, higher in charged performance and remarkable in nano-material characteristics. The size of the chitin nanofiber can be controllably adjusted by adjusting the ultrasonic crushing condition; the controllable adjustment of the surface electric quantity of the chitin nano-fiber is realized by adjusting the temperature and the acidolysis ratio.

Description

Preparation method and application of chitin nanofiber

Technical Field

The invention belongs to the field of new material preparation, and particularly relates to a preparation method and application of chitin nanofiber.

Background

Chitin is a natural high molecular polymer which only has positive charges in nature and is widely present in exoskeletons of crustaceans, tissues and organs of mollusks, cell walls of fungi and the like. Chitin exists in fiber shape, and the structural unit N-acetylglucosamine is connected through hydrogen bonds, and has alpha, beta and gamma 3 configurations. Among them, alpha-chitin is the most common, has high crystallinity and stable physical properties, and is derived from shrimp shells. The chitin has the advantages of no toxicity, biodegradability and biocompatibility, can improve plant growth, induce plant resistance and improve crop yield, and is applied to crop pest control, seed coating and fruit and vegetable fresh keeping. At present, in biomedicine, the nano-scale chitin particles have large specific surface area and high charge density, can carry drug particles to transport across membranes, protect drug macromolecules from being degraded, and can control the release of drugs or macromolecules on a target. In addition, the nanochitin can effectively interact with cell membranes and enter cells in an endocytosis mode, so that the bioavailability of the medicine is improved, the metabolism of the medicine is influenced, and the release of the medicine is slowed down.

The chitin nanoparticles currently used for research applications are prepared by acidolysis, oxidation, mechanical milling, and in addition, ultrasonic decomposition enables chitin fibrils/rods with dimensions in the micron range to be obtained.

The method comprises the steps of slowly destroying O-bonds in a chitin structure unit by high-concentration strong acid under high-temperature heating, cutting off a main chain structure unit by the high-concentration acid liquor to obtain chitin particles, gradually reducing the size of particle fragments through a plurality of repeated cyclic acidolysis-collection-acidolysis processes, and removing acid ions through dialysis ion exchange.

When the oxidation method is used for preparing the chitin nano-fibers, the C-6 position in the chitin structure can be specifically oxidized in hypochlorous acid liquid, so that a main chain is broken and decomposed into broken particles, the shape, the size, the dispersity and the surface charge characteristics of the nano-fibers prepared by the method are closely related to the content and the proportion of hypochlorous acid, but only fiber particles with the length of micron order can be prepared generally, and the real nano-chitin can be obtained by additional reprocessing.

When the nano chitin is prepared by a grinding method, high-energy-consumption precise instruments such as a high-pressure water spray machine, a grinding machine or a high-speed stirring machine are needed, the preparation efficiency and the yield are low, the relative equipment cost and the energy-consumption material cost are high, and the method is not suitable for large-scale preparation and production.

At present, many progress has been made on the preparation of nanochitin in China, patent No. CN105861478A discloses a method for preparing nanochitin based on an acidolysis method, which can obtain nanoscale chitin particles through strong acid acidolysis, vigorous mechanical stirring and repeated dialysis for many times, patent No. CN106868631A discloses a method for hydrolyzing chitin through organic binary acid and obtaining nanochitin through homogenization treatment, patent No. CN108864445A discloses a method for preparing nanochitin dispersion through mixing chitin acid solution with a cross-linking agent, freeze-thawing treatment and mechanical treatment, and patent No. CN106520741A discloses a method for preparing nanochitin with high surface activity by using an acidolysis method and an enzymolysis method.

The ultrasonic wave can form high-energy chemical acting force through cavitation at a water/air interface, including forming an energy core in water, the energy core grows and then collapses, along with the rapid generation of temperature, pressure and heating/cooling, the repeated continuous process provides an energy source for breaking intermolecular hydrogen bonds among chitin fibers, so that the broken main chain is continuously dissociated to form nano fibers with smaller particles, but the method is long in time consumption and extremely low in preparation efficiency when being used alone, the charge characteristics of the prepared nano particles are not obvious, and the chemical activity and the biological activity of the nano particles are low, so that the method is not suitable for the fields of biology, medicine and new materials.

However, these methods also show disadvantages such as low preparation efficiency and high preparation cost, and it often takes several days from the chitin raw material to the preparation of nanoparticles, and the preparation process is complicated with large consumption of materials, reagents and energy, and some systems react violently, inevitably increasing the preparation cost.

But proper ultrasonication can effectively reduce the particle size of chitin nanoparticles, and the principle lies in that: chitin is heated correspondingly or contacts with oxidant in acid liquor or oxidizing liquid to form micro-fiber or nano-fiber, the fibers can be rapidly aggregated to form chitin gel through hydrogen bond and hydrophobic effect, during the process of forming and aggregating the fibers to form gel, intermolecular hydrogen bond of the fibers is weakened, and the lower energy input is enough to destroy intermolecular force of the chitin. The applicant participates in patent No. CN105638663A and discloses a method for preparing a nano chitin aqueous suspension agent by using a hydrolysis method to be applied to wheat aphid control. At present, although a lot of researches are carried out on the preparation method of the nano chitin, the researches on how to improve the preparation technology and effectively improve the preparation efficiency are less.

Disclosure of Invention

The invention provides a preparation method and application of chitin nanofibers, and solves the technical problem that the length and the charged property of the existing chitin fibers are uncontrollable and unadjustable.

The technical scheme of the invention is realized as follows:

a preparation method of chitin nano-fibers comprises the following steps:

(1) preparing low-concentration inorganic acid solution, heating in water bath or oil bath, adding chitin powder, stirring at low speed for full acidolysis to obtain acidolysis suspension;

(2) centrifuging the acidolysis suspension prepared in the step (1) at a high speed, taking the precipitate, washing the precipitate with water, and dispersing the precipitate in water again to obtain a dispersion liquid;

(3) and carrying out ultrasonic crushing on the dispersion liquid to obtain the chitin nano-fiber aqueous suspension.

The inorganic acid solution in the step (1) is sulfuric acid or hydrochloric acid, and the concentration of the inorganic acid solution is 1-3 mol/L.

The heating temperature in the step (1) is 30-70 ℃.

The heating temperature is 50-70 ℃.

The proportion of the chitin powder to the inorganic acid solution in the step (1) is 1 g: (50-500) mL.

The proportion of the precipitate to the water in the step (2) is 1 g: (50-200) mL.

The corresponding ultrasonic crushing conditions per 100g of the dispersion in the step (3) are as follows: the ultrasonic crushing power is 200-400W, the ultrasonic crushing frequency is 20-25KHz, the ultrasonic crushing time is 15-90min, and the temperature of the dispersion liquid is not higher than 40 ℃ during ultrasonic treatment.

The chitin nanofiber prepared by the method has the length of 10-900nm and the width of 5-10nm, and has obvious positive charge property.

The chitin nano-fiber is applied to preparation of agricultural insect and disease prevention, medicine transmission and new medicine carrier reagents.

The preparation principle of the application is as follows: the chitin main chain structure is damaged by acid liquor, residual chain microfibers with different sizes are decomposed/degraded, the microfiber structure is dissociated by using energy provided by ultrasonic waves after ultrasonic crushing, intermolecular hydrogen bonds in the microfibers are damaged, and finally the nanofiber is prepared. The whole method not only improves the preparation efficiency, but also reduces the energy consumption and the acid liquor consumption. In addition, due to the advantage of ultrasonic crushing, the nano-fibers have higher consistency in particle size, and due to the fact that a large amount of ammonium radicals are exposed due to the separation of chitin side chain acetyl in an acidic environment, the nano-fibers have obvious electrification, so that the physical/chemical properties are obviously improved, the biological application performance is obviously improved, and the nano-fibers are suitable for the aspects of agricultural insect prevention and disease resistance, medicine transmission and application of new medicine carriers.

The invention has the following beneficial effects:

1. the chitin nano-fiber aqueous dispersion with uniform particle size, good dispersity, obvious nano-characteristic and good particle charging performance can be obtained by simple acidolysis and ultrasonic crushing in one step.

2. The preparation method has mild preparation conditions, the preparation temperature is 60-90 ℃, the requirement on the acid resistance of instruments and equipment is low, the risk of the preparation process is reduced, and the controllability of the operation is improved. The preparation process is simple and high in efficiency, only needs one-step acidolysis process and one-step ultrasonic crushing process, the acidolysis time is 2-4 h, and the ultrasonic crushing time is 30-60 min, so that the preparation method is suitable for large-scale preparation and production.

3. The chitin nanofiber prepared by the method is small in particle size, good in uniform dispersion performance, high in particle surface electric quantity and remarkable in nano characteristic. The size of the chitin nano-fiber can be controllably adjusted by adjusting the ultrasonic crushing condition; the controllable adjustment of the surface electric quantity of the chitin nano-fiber is realized by adjusting the temperature and the acidolysis ratio.

Drawings

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

FIG. 1 is a TEM image of chitin nanofibers according to example 2 of the present invention.

FIG. 2 is a graph showing the effective particle size and particle size distribution of chitin nanofibers in an aqueous dispersion according to example 1 of the present invention.

FIG. 3 is a graph showing the effective particle size and particle size distribution of chitin nanofibers in an aqueous dispersion according to example 2 of the present invention.

FIG. 4 is a graph showing the effective particle size and particle size distribution of chitin nanofibers in an aqueous dispersion according to example 3 of the present invention.

FIG. 5 is the electromotive force distribution graph of chitin nanofibers in water dispersion in example 1 of the present invention.

FIG. 6 is the electromotive force distribution graph of chitin nanofibers in water dispersion in example 2 of the present invention.

FIG. 7 is a graph showing the electromotive force machine distribution of chitin nanofibers in aqueous dispersion in example 3 of the present invention.

Detailed Description

The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments of the present invention, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be obtained by a person skilled in the art without inventive effort based on the embodiments of the present invention, are within the scope of the present invention.

The invention provides a preparation method of chitin nano-fibers, wherein the experimental operation needs attention as follows: diluting concentrated hydrochloric acid with water to the required low concentration, transferring 100mL each and heating in a three-neck round-bottom flask to the required temperature, weighing 4 groups of chitin powder respectively, adding the diluted acid solution, and slowly stirring. After reaching the preset acidolysis time, the mixed solution is introduced into a centrifuge bottle, cooled to 4 ℃ by ice water bath, then subjected to high-speed centrifugation, collected and precipitated, washed with water for 2 times and resuspended for dispersion. And (3) carrying out ultrasonic treatment on the sediment after the re-suspension and dispersion according to preset power and time, monitoring the temperature of the dispersion liquid to be not higher than 40 ℃ in the ultrasonic process to obtain the aqueous dispersion of the chitin nano-fibers, and directly using or freeze-drying the aqueous dispersion of the chitin nano-fibers according to tests and requirements to obtain a solid. The following are specific examples carried out by the inventors.

Example 1

The invention provides a preparation method of chitin nano-fibers, which comprises the following steps:

(1) preparing 50mL of 1 mol/L inorganic acid solution, heating the inorganic acid solution in a water bath or oil bath at 50 ℃, adding 1g of chitin powder, and stirring the mixture at a low speed for 2 hours until the mixture is sufficiently acidolyzed to obtain acidolysis suspension;

(2) centrifuging the acidolysis suspension prepared in the step (1) at a high speed, washing the precipitate with water for 2 times, and dispersing the precipitate in 50mL of water again to obtain a dispersion solution;

(3) sonicating the dispersion under the corresponding sonication conditions per 100g of dispersion: the ultrasonic crushing power is 200W, the ultrasonic crushing frequency is 20KHz, the ultrasonic crushing time is 30min, and the temperature of the dispersion liquid is kept at 0 ℃ during ultrasonic treatment to obtain the chitin nanofiber aqueous suspension.

Fig. 2 is a graph showing the particle size and the distribution of the particle size of the chitin nanofibers obtained in example 1 in an aqueous dispersion, from which it can be seen that the prepared nanofibers have a length of 55 to 900nm, 90% of the fibers have a size in the range of 100 to 500nm, the particle size distribution is concentrated, and the dispersibility index is 0.246, which shows that the prepared chitin nanofibers have better dispersibility and stability.

Fig. 5 is a graph showing the zeta potential and the distribution of the zeta potential of the chitin nanofibers prepared in example 1, from which it can be seen that the prepared nanofibers have a concentrated zeta potential distribution with an average zeta potential of 17.6mV, showing good charging properties of the chitin nanofibers.

Example 2

(1) preparing 100mL of 2 mol/L inorganic acid solution, heating the inorganic acid solution in a water bath or oil bath at 60 ℃, adding 1g of chitin powder, and stirring the mixture at a low speed for 3 hours until the mixture is sufficiently acidolyzed to obtain acidolysis suspension;

(2) centrifuging the acidolysis suspension prepared in the step (1) at a high speed, washing the precipitate with water for 1 time, and dispersing the precipitate in 100mL of water again to obtain a dispersion solution;

(3) sonicating the dispersion under the corresponding sonication conditions per 100g of dispersion: the ultrasonic crushing power is 200W, the ultrasonic crushing frequency is 20KHz, the ultrasonic crushing time is 30min, and the temperature of the dispersion liquid is kept at 0 ℃ during ultrasonic treatment to obtain the chitin nano-fibers.

Fig. 1 is a transmission electron microscope image of the chitin nanofiber obtained in this example 2, from which chitin nanoparticles are obtained in fibrous form, with obvious morphological features, complete structure of the fiber, good dispersibility, and no aggregation or polymerization. Chitin nanofibers are predominantly centered within 100 to 200nm in length and 5 to 10nm in width, representing significant nanofiber characteristics.

Fig. 3 is a graph showing the particle size and particle size distribution of the chitin nanofibers obtained in example 2 in an aqueous dispersion, from which it can be seen that the length of the prepared nanofibers is 45 to 500nm, 90% of the fibers have a size in the range of 70 to 300nm, the particle size distribution is concentrated, and the dispersibility index is 0.216, which illustrates that the prepared chitin nanofibers have good dispersibility and stability.

Fig. 6 is a graph showing the electromotive potential and distribution of the chitin nanofibers prepared in this example 2, and it can be seen that the nanofibers prepared have a concentrated electromotive potential distribution with an average electromotive potential of 51.8mV, showing good charging properties of the chitin nanofibers.

Example 3

(1) preparing 200mL of 3 mol/L inorganic acid solution, heating the solution in a water bath or oil bath at 70 ℃, adding 1g of chitin powder, and stirring the solution at a low speed for 5 hours until full acidolysis is performed to obtain acidolysis suspension;

(2) centrifuging the acidolysis suspension prepared in the step (1) at a high speed, washing the precipitate with water for 2 times, and dispersing the precipitate in 200mL of water again to obtain a dispersion solution;

(3) sonicating the dispersion under the corresponding sonication conditions per 100g of dispersion: the ultrasonic crushing power is 400W, the ultrasonic crushing frequency is 25KHz, the ultrasonic crushing time is 40min, and the temperature of the dispersion liquid is kept at 0 ℃ during ultrasonic treatment to obtain the chitin nanofiber aqueous suspension.

Fig. 4 is a graph showing the particle size and particle size distribution of the chitin nanofibers obtained in example 3 in an aqueous dispersion, from which it can be seen that the prepared nanofibers have a length of 10 to 50nm, 95% of the fibers have a size in the range of 10 to 30nm, the particle size distribution is concentrated, and the dispersibility index is 0.195, which indicates that the prepared chitin nanofibers have good dispersibility and stability.

Fig. 7 is a graph showing the electromotive potential and distribution of the chitin nanofibers fabricated in this example 3, from which it can be seen that the nanofibers fabricated have a concentrated electromotive potential distribution with an average electromotive potential of 53.5mV, showing good charging properties of the chitin nanofibers.

Example 4

(1) preparing 500mL of 2 mol/L inorganic acid solution, heating the inorganic acid solution in a water bath or oil bath at 50 ℃, adding 1g of chitin powder, and stirring the mixture at a low speed for 5 hours until the mixture is sufficiently acidolyzed to obtain acidolysis suspension;

(2) centrifuging the acidolysis suspension prepared in the step (1) at a high speed, washing the precipitate with water for 2 times, and dispersing the precipitate in 500mL of water again to obtain a dispersion solution;

(3) sonicating the dispersion under the corresponding sonication conditions per 100g of dispersion: the ultrasonic crushing power is 400W, the ultrasonic crushing frequency is 23KHz, the ultrasonic crushing time is 30min, and the temperature of the dispersion liquid is kept at 0 ℃ during ultrasonic treatment to obtain the chitin nano aqueous suspension.

Examples of the effects of the invention

Table 1 shows the properties of the chitin fibers prepared in examples 1-3:

table 1 example preparation parameter levels and chitin nanofibrils characterization comparison

From the data analysis of the invention and table 1, it can be found that increasing the concentration and amount of the acid solution and increasing the power of the ultrasonic disruption can achieve the purpose of effectively reducing the particle size of the prepared nanochitin.

Increasing the acid concentration and amount is effective in reducing the particle size because chitin microfibers contain 10 or more parallel bundle units in their structure and constitute the chitin backbone. Hydrochloric acid, as a stable chemical agent, can destroy the-O-bonds between the main chain structures of the nanochitin, and particularly under the heating condition, increasing the acid liquor ratio can accelerate the bond breakage and the parallel beam unit peeling, thereby reducing the particle size.

The particle size can also be effectively reduced by increasing the power of ultrasonic crushing, and the mechanism is as follows: chitin is heated/contacted with an oxidizing agent in an acid solution to form microfibers or nanofibers, which can rapidly aggregate through hydrogen bonding and hydrophobic interactions to form chitin gels, which are themselves a very rapid process. Furthermore, at the same time as this process of fiber formation and aggregation to form a gel, intermolecular hydrogen bonding of the fibers is weakened, where the lower energy input is sufficient to break the intermolecular forces of chitin. The ultrasonic wave can form high-energy chemical acting force through cavitation at water/air interface, including forming the energy nuclear in aqueous, the energy nuclear grows, takes place to collapse in succession, along with produce temperature, pressure and heating/cooling fast, this repeated continuation process provides the energy source for breaking chitin fibrous intermolecular hydrogen bond for the main chain that breaks continues to dissociate, forms the nanofiber of smaller granule, and ultrasonic power is bigger, and the energy that provides is bigger, and the dissociation between chitin fibrous molecule is more on unit length, and nanofiber length is shorter, thereby effective particle size reduces.

In addition, the charge quantity of the prepared nanochitin fibers is positively correlated with the amount of the acid liquor, the amounts of the acid liquor in examples 1, 2 and 3 are respectively 0.05 mol, 0.2 mol and 0.6 mol, and the average electromotive potentials of the corresponding nanochitins are respectively 17.6mV, 51.8mV and 53.5 mV. Under the conditions of heating and acid liquor, the chitosan structure is deacetylated to expose amino, protonation is carried out under the condition that the acidity is lower than the pKa of the chitosan structure, then the chitosan structure is converted into ammonium radicals, positive charges are shown in a water dispersion phase, and the average electromotive potential is an important nano characteristic of a nanochitin aqueous suspension.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.

Claims

1. The preparation method of the chitin nano-fiber is characterized by comprising the following steps:

2. The method of claim 1, wherein the chitin nanofibers are prepared by: the inorganic acid solution in the step (1) is sulfuric acid or hydrochloric acid, and the concentration of the inorganic acid solution is 1-3 mol/L.

3. The method of claim 2, wherein the chitin nanofibers are prepared by: the heating temperature in the step (1) is 30-70 ℃.

4. The method of claim 3, wherein the chitin nanofibers are prepared by: the heating temperature is 50-70 ℃.

5. The method of claim 2, wherein the chitin nanofibers are prepared by: the proportion of the chitin powder to the inorganic acid solution in the step (1) is 1 g: (50-500) mL.

6. The method of claim 1, wherein the chitin nanofibers are prepared by: the proportion of the precipitate to the water in the step (2) is 1 g: (50-200) mL.

7. The method for preparing chitin nanofibers according to claim 1, wherein the conditions of the corresponding ultrasonication per 100g of the dispersion in step (3) are: the ultrasonic crushing power is 200-400W, the ultrasonic crushing frequency is 20-25KHz, the ultrasonic crushing time is 15-90min, and the temperature of the dispersion liquid is not higher than 40 ℃ during ultrasonic treatment.

8. Chitin nanofibrils prepared according to the method of any of claims 1 to 7, characterized in that: the chitin nanofiber has the length of 10-900nm and the width of 5-10nm, and has obvious positive charge property.

9. The use of the chitin nanofibers of claim 8 in the preparation of agricultural insect and disease resistant, pharmaceutical delivery and new drug carrier agents.