阅读说明：本技术 纳米纤维膜及其运用及纳米纤维膜杀菌液及其使用方法 (Nanofiber membrane and application thereof, nanofiber membrane sterilization liquid and application method thereof ) 是由 姚凯勇 于 2021-08-19 设计创作，主要内容包括：本申请涉及新型抗菌材料领域,具体涉及纳米纤维膜杀菌液的制备及其使用方法,其中纳米纤维膜通过壳聚糖和聚乙二醇的醋酸-水溶液经静电纺丝制备得到。本申请中的纳米纤维膜抗菌液通过上述纳米纤维膜和乙酸共同配置溶液得到,可以在肉制品表面形成保护膜,进而提供长效的肉制品保鲜效果。(The application relates to the field of novel antibacterial materials, in particular to preparation of a nano-fiber membrane sterilization liquid and a using method thereof, wherein the nano-fiber membrane is prepared by performing electrostatic spinning on acetic acid-water solution of chitosan and polyethylene glycol. The nanofiber membrane antibacterial liquid is obtained by preparing a solution through the nanofiber membrane and acetic acid, a protective film can be formed on the surface of a meat product, and a long-acting meat product fresh-keeping effect is further provided.)

1. The nanofiber membrane is characterized by being prepared by electrostatic spinning of a mixed component I, wherein the mixed component I comprises the following components:

the concentration of the chitosan in the mixed component I is 50-150 g/L;

the concentration of the polyethylene glycol in the mixed component I is 2-10 g/L;

acetic acid, wherein the total mass fraction of the acetic acid in the mixed components is 30-60%;

and (3) water.

2. The nanofiber membrane as claimed in claim 1, wherein the mixed component I further comprises an antibacterial auxiliary agent with a mass fraction not more than 4%, and the antibacterial auxiliary agent is at least one of polyphenol or plant essential oil.

3. The nanofiber membrane as claimed in claim 2, wherein the mixed component I further comprises a surfactant with a mass fraction of not more than 0.6%.

4. The nanofiber membrane according to claim 3, wherein the surfactant is a food grade amphoteric surfactant.

5. The nanofiber membrane as claimed in claim 1, wherein the mixed component I further comprises 0.1-0.3% of soybean polysaccharide by mass fraction.

6. The nanofiber membrane as claimed in claim 1, wherein the polyethylene glycol has a number average molecular weight of 1000-1500 Da, and the chitosan has a molecular weight of 80-2000 KDa.

7. The nanofiber membrane as claimed in claim 1, wherein in the electrospinning process, the voltage is 10-15 KV, the solution flow rate is 0.3-0.5 mL/h, the distance from the needle to the receiving plate is 13-16 cm, and the inner diameter of the needle is 0.05-0.06 mm.

8. Use of the nanofiber membrane as claimed in any one of claims 1 to 8, characterized in that it is used in the field of preservation of meat products.

9. The nanofiber membrane sterilization liquid is characterized by comprising the following components:

acetic acid, wherein the volume fraction of the acetic acid in the nano fiber membrane sterilization liquid is 1-5%;

the nanofiber membrane as claimed in any one of claims 1 to 7, wherein the concentration of the nanofiber membrane bactericidal solution is 5 to 50 g/L;

the balance being water.

10. The use method of the nanofiber membrane sterilization liquid is characterized in that the nanofiber membrane sterilization liquid as claimed in claim 9 is used according to the proportion of 100-200 mu L/cm²The coating is applied to the surface of a sample to be coated, and then the sample is placed in a sterile environment at the temperature of 20-25 ℃ for 2-4 hours.

Technical Field

The application relates to the field of antibacterial materials, in particular to preparation of a nanofiber membrane sterilization liquid and a using method thereof.

Background

The chilled fresh poultry meat is one of the mainstream meat products in China, and the safety and the quality of the chilled fresh poultry meat are important factors influencing the health of people. In the process of processing, storing and selling the cold fresh poultry meat, the bacteria such as pseudomonas, lactobacillus, enterobacter, suicidella, enterococcus and the like can cause the putrefaction and the deterioration of the cold fresh poultry meat and influence the quality and the sanitation of the cold fresh poultry meat.

At present, for cold fresh poultry meat, fresh keeping is mainly carried out in the following way:

1. preserving with antibiotics. However, the use of antibiotics is liable to cause the bacteria to develop drug resistance, and the popularization and application of the antibiotics are limited.

2. The low-temperature storage mode is adopted, generally, when the storage temperature is lower than 0 ℃, the taste of the meat is adversely affected, and when the storage temperature is higher than 0 ℃, the meat is difficult to store for a long time, generally, the storage time of the poultry meat is 5-6 days at 4 ℃.

3. In addition, meat products are preserved by radiation, smoking and other methods, and the methods have adverse effects on the taste of meat.

Disclosure of Invention

In order to eliminate the use of antibiotics in the process of meat preservation and enable meat to have longer preservation time and better taste, the application provides a nanofiber membrane and application thereof, and a nanofiber membrane sterilization solution and a using method thereof, firstly, the application provides the nanofiber membrane which is prepared by mixing a component I through electrostatic spinning, wherein the component I comprises the following components:

the concentration of the chitosan in the mixed component I is 50-150 g/L;

the concentration of the polyethylene glycol in the mixed component I is 2-10 g/L;

acetic acid, wherein the total mass fraction of the acetic acid in the mixed components is 30-60%;

and (3) water.

In the technical scheme, chitosan and polyethylene glycol are used as main components, and through electrostatic spinning, the formed chitosan system nanofiber membrane has good antibacterial performance on one hand, and the polyethylene glycol can play a reinforcing effect in the chitosan system nanofiber membrane, so that the nanofiber membrane is more complete, and a better membrane covering effect and an antibacterial effect are further formed. In an acetic acid system, the structure of the nanofiber membrane is favorably formed, so that the obtained nanofiber membrane has a good antibacterial effect.

At present, the antibacterial agents of nanofiber membranes are not used and popularized in the field of food preservation, and related contents are rarely researched. The nanofiber membrane prepared by the method has good antibacterial performance, can be applied to the preservation of meat products, particularly chilled poultry meat, and has good market popularization value and application prospect.

Optionally, the mixed component i further comprises an antibacterial auxiliary agent with a mass fraction of not more than 4%, wherein the antibacterial auxiliary agent is at least one of polyphenol or plant essential oil.

The antibacterial auxiliary agent can be doped in the structure of the nanofiber membrane in the electrostatic spinning process, so that the antibacterial effect of the prepared nanofiber membrane is further improved, and the nanofiber membrane has a better effect when being applied to meat product antibiosis.

Optionally, the mixed component I further comprises a surfactant with the mass fraction not more than 0.6%.

In the technical scheme, the surfactant is beneficial to improving the stability and the dispersibility of the nanofiber membrane, so that the nanofiber membrane has better film forming property and adhesion capacity when being applied to meat products, and the antibacterial effect of the nanofiber membrane on the meat products is further improved.

Optionally, the surfactant is a food grade amphoteric surfactant.

By adopting the amphoteric surfactant, the formed nanofiber membrane has better stability, stronger membrane forming capability on the surface of a sample and stronger sterilization effect.

Optionally, the mixed component I also comprises soybean polysaccharide with the mass fraction of 0.1-0.3%.

The soybean polysaccharide further improves the adhesion capability and the film forming capability of the nanofiber membrane, so that the nanofiber membrane has better performance and better use effect when being applied to antibacterial practice.

Optionally, the number average molecular weight of the polyethylene glycol is 1000-1500 Da, and the molecular weight of the chitosan is 80-2000 KDa.

Experiments prove that the polyethylene glycol and chitosan molecules in the molecular weight range can well form the overall structure of the nanofiber membrane, the structure is stable, the dispersibility in the water phase is good, and when the antibacterial nano-fiber membrane is applied to the surface of a meat product, the antibacterial nano-fiber membrane has a good antibacterial effect and has no obvious influence on the taste of the meat product.

Optionally, in the electrostatic spinning process, the voltage is 10-15 KV, the solution flow rate is 0.3-0.5 mL/h, the distance from the needle head to the receiving plate is 13-16 cm, and the inner diameter of the needle head is 0.05-0.06 mm.

By adopting the parameters, the nanofiber membrane can be well prepared, and has good antibacterial performance and adhesion performance when being applied to the field of meat products.

Secondly, the application of the nanofiber membrane is further provided, and the nanofiber membrane is applied to the field of meat product preservation.

In the field of meat product preservation, the following requirements are placed on the antibacterial agent: 1. is non-toxic and edible; 2. has good antibacterial effect; 3. the processing is convenient; 4. has no obvious negative influence on the mouthfeel of the meat product. When the nanofiber membrane is applied to the field of meat product sterilization and preservation, the advantages can be taken into consideration, and therefore the nanofiber membrane has a good practical application prospect.

Thirdly, the application also provides a nanofiber membrane sterilization liquid which comprises the following components:

acetic acid, wherein the volume fraction of the acetic acid in the nano fiber membrane sterilization liquid is 1-5%;

the concentration of the nanofiber membrane in the nanofiber membrane sterilization solution is 5-50 g/L;

the balance being water.

In the technical scheme, the prepared nanofiber membrane antibacterial solution has a strong sterilization effect by taking an acetic acid aqueous solution as a basic component. The acidic condition provided by the acetic acid is helpful for promoting the antibacterial ability of the nanofiber membrane, and further stronger antibacterial property is realized.

Finally, the application provides a using method of the nanofiber membrane antibacterial solution, wherein the nanofiber membrane antibacterial solution is used according to the proportion of 100-200 mu L/cm²The coating is applied to the surface of a sample to be coated, and then the sample is placed in a sterile environment at the temperature of 20-25 ℃ for 2-4 hours.

In the technical scheme, after the nano antibacterial liquid kills bacteria on the surface of the meat product, a layer of antibacterial protective film can be formed, so that the bacteria on the meat product can be killed and killed for a long time, and the meat product can be stored for a longer time.

In summary, the present application includes at least one of the following advantages:

1. in the application, the nanofiber membrane is obtained by performing electrostatic spinning on chitosan and polyethylene glycol, can be applied to the field of meat product sterilization and preservation, has better sterilization performance, and meanwhile, eliminates the use of antibiotics, and has better commercial application prospect.

2. In the further arrangement of the application, the antibacterial performance of the nanofiber membrane is further improved by adding the antibacterial auxiliary agent.

3. In the application, the nanofiber membrane is prepared into the nanofiber membrane antibacterial liquid in an acetic acid solution, and the antibacterial performance of the nanofiber membrane is further improved by compounding with acetic acid.

4. In the application, the application method of the nanofiber membrane is further provided, and the nanofiber membrane can be smeared and then kept still and dried in an aseptic environment, so that a complete antibacterial membrane can be formed on the surface of a sample, and a better antibacterial effect is provided.

Detailed Description

The present application is further described in detail in connection with the following examples.

The preservation of meat products has always been an important problem to be overcome in the food field. Whether the antibiotic treatment is carried out by adopting antibiotics, or the surface treatment (such as irradiation, smoking and the like) is carried out on the meat product, or the pickling, drying and other treatments are carried out on the meat product, the method has respective limitations.

In the market, after the meat product is placed on a shelf, if the meat product is not sold as soon as possible, the meat product can be stale after being placed for one to two days. At higher temperatures, rancidity and the like can occur even within two days, the main reason for this being the growth of bacteria in the meat products. Although the antibiotic has a good effect, the antibiotic is easy to cause drug resistance of bacteria and has certain adverse effect on human bodies.

The nano antibacterial agent of chitosan is a new direction for the antibacterial of meat products, and has no application examples in the field at present, and related researches are less. The purpose of the present application is to provide a meat product which has little effect on meat quality,

in the following examples, the parameters of a portion of the feedstock are shown in Table 1.

TABLE 1 Material parameter Table

Examples A1-A17, nanofiber membranes, were prepared by mixing component I and electrospinning. The specific formulation of blend component I is shown in Table 2.

Component summary of component I of Table 2, examples A1 to A17

Wherein the number average molecular weight of the chitosan is 1000KDa, and the number average molecular weight of the polyethylene glycol is 1200. The surfactant is rhamnolipid. Rhamnolipid is an anionic surfactant. The antibacterial auxiliary agent is cinnamon essential oil.

The composition I can be prepared by pre-preparing chitosan-acetic acid solution and polyethylene glycol-acetic acid solution with specific concentrations, and then performing electrostatic spinning by using the parameters shown in Table 3.

Electrostatic spinning parameters in Table 3, examples A1-A17

Parameter(s)	Voltage of	Flow rate of solution	Needle head toDistance of needle plate	Inside diameter of needle
					Unit of	KV	mL/h	cm	mm
Data of	12	0.5	15	0.05

Example a18, a nanofiber membrane, differs from example a11 in that the surfactant is dodecyl dimethyl betaine.

Example a19, a nanofiber membrane, differs from example a11 in that the surfactant is sorbitan monooleate.

Example a20, a nanofiber membrane, differs from example a11 in that the surfactant is sucrose monolaurate.

Example a21, a nanofiber membrane, differs from example a17 in that the surfactant is dodecyl dimethyl betaine.

Examples a 22-a 35, nanofiber membranes, differ from example a18 in that the molecular weights of the polyethylene glycol and chitosan used are specified in table 4.

TABLE 4 number average molecular weights of polyethylene glycol and chitosan in examples A22-A35

Examples a 36-a 37, nanofiber membranes, differ from example a33 in that the specific parameters are shown in table 5.

Electrostatic spinning parameters in Table 5 and examples A36-A37

Parameter(s)	Voltage of	Flow rate of solution	Needle to needle plate distance	Inside diameter of needle
					Unit of	KV	mL/h	cm	mm
Example A36	10	0.3	13	0.05
					Example A37	15	0.5	16	0.06

Examples A38-41, the difference between nanofiber membrane and example A9 is that equal mass of lavender essential oil, thyme essential oil, tea polyphenol or tannin are respectively used as antibacterial auxiliary agent.

On the basis of the nanofiber membrane, the following examples are further provided:

examples B1-B341, nanofiber membrane bactericidal solution and methods of use, nanofiber membrane bactericidal solution preparation methods were as follows: acetic acid aqueous solution with volume fraction of 1% was prepared, and the nanofiber membranes of examples a1 to a37 were dissolved in the acetic acid aqueous solution at a concentration of 5g/mL, and uniformly mixed to obtain nanofiber membrane antibacterial solution.

When in use, the nano-fiber sterilizing solution is added at the concentration of 100 mu L/cm²The amount of the sample is coated on the surface of the duck meat block, and the duck meat block is placed for 4 hours at 20 ℃ in an aseptic environment to obtain a corresponding sample 1-41.

Examples B42 to 50, the difference between the nanofiber membrane and example 1 is that the amounts of the nanofiber membrane and acetic acid used are specifically shown in table 6.

Table 6, nanofiber membranes of examples B42-B50 and amounts of acetic acid used

Examples	Nanofiber Membrane concentration (g/L)	Acetic acid volume fraction (%)
			Example B42	3	1
Example B43	10	1
			Example B44	20	1
Example B45	50	1
			Example B46	80	1
Example B47	20	0.5
			Example B48	20	2
Example B49	20	5
			Example B50	20	8

In examples B42 to B50, samples 42 to 50 were obtained by the same experimental method as in examples B1 to B41.

Example B51 differs from example B33 in that the nanofiber membrane concentration was 20g/L and the volume fraction of acetic acid was 2%, using the same method of use, sample 51 was obtained.

In addition, in example B51, the use method was adjusted to obtain samples shown in table 7.

TABLE 7 methods of use of samples 52-58

The comparison is carried out by the following comparative examples.

Comparative example 1, which is a sterilizing solution, is different from example B1 in that the nanofiber membrane is replaced with chitosan of equal mass. The molecular weight of chitosan is 1200 KDa. Sample 1' was obtained using the same method as example B1.

A control sample was additionally prepared, and treated in the same manner using 50. mu.g/L kanamycin to obtain sample 2'.

The following experiments were conducted on samples 1 to 58 and samples 1 'and 2' to verify the effectiveness thereof.

Experiment 1, for each sample, different strains were inoculated on the surface of the sample, and the concentration of the bacterial liquid was 10⁷cfu/mL, the dosage of the bacterial liquid is 20 mu L/cm². After inoculation, the sample is placed in an environment with the temperature of 4 ℃ for 24 hours, and then the surface is cleaned by sterile normal saline. Then, 1g of the sample was weighed in a sterile environment, pulverized in 10mL of sterile physiological saline, mixed well, diluted to 100mL with physiological saline, and cultured by pipetting 100. mu.L of the plate medium. After 24h of incubation, the surface of the dish was observed as the area of coverage of the colony on the surface of the culture name. + + + + + + represents 100% of plaque overlay; + + + + represents 60-90% of the plaque overlay; + represents 20-60% of plaque overlay plate; + represents plaque number within 10; -represents a sterile plaque growth.

Meanwhile, another sample is taken and placed for 120h, then the bacteria liquid is coated, and after the sample is placed for 24h, the steps are carried out in the same way.

Experiment 2, each sample is placed in a ventilated non-sterile environment at 20 ℃, the environment humidity is controlled to be 50-60, the samples are observed once every 12 hours, and the time for the samples to start to decay is recorded.

First, the results of experiments performed on samples 1 to 17 and samples 1 'and 2' are shown in table 8.

TABLE 8 Experimental results of samples 1-17 and samples 1 'and 2' in experiment 1 and experiment 2

According to the experiment, the scheme in the application can basically play the antibacterial effect similar to kanamycin, and can effectively prolong the shelf life of the duck meat. And the adopted components, such as chitosan, plant essential oil and the like, are components which are not easy to form drug resistance, and have small harm to the health of human bodies. Compared with the sample 1', the technical scheme in the application adopts the method to prepare the chitosan into the nanofiber membrane, so that the film forming property and the adhesion property of the chitosan on the surface of duck meat are greatly improved, and the chitosan has a good antibacterial effect in both short-term and long-term dimensions.

In samples 8-10, the antibacterial auxiliary agent is added, and the result shows that the antibacterial auxiliary agent can further improve the antibacterial capability of the nanofiber membrane, and the antibacterial capability is increased along with the increase of the addition amount of the antibacterial auxiliary agent. But when the mass fraction of the antibacterial auxiliary agent reaches 4%, the concentration of the antibacterial auxiliary agent is continuously increased, and the overall antibacterial performance is not further improved. On the basis of increasing the antibacterial auxiliary agent, the addition of the surfactant is beneficial to further improving the adhesive force of the nanofiber membrane on the surface of the meat product, so that the antibacterial performance is further improved within a long-time dimension. In addition, in samples 14-16, soybean polysaccharide is added and doped in the nanofiber membrane, so that stronger adhesion capability and better antibacterial effect can be provided, the formed membrane structure is more stable and is not easy to damage, and further better antibacterial effect is provided.

Further, experiments 1 to 2 were performed on the samples 18 to 35, and the results are shown in table 9. Since all of the measurements were negative after 24h of standing, the experimental results here were omitted, and only a portion of 120h of standing was shown.

TABLE 9 Experimental results of samples 18-35 in experiment 1 and experiment 2

In the samples, the types of the surfactants in the samples 18-21 are adjusted, and in the sample 18, the amphoteric surfactant is adopted, and has a better antibacterial effect compared with a nonionic surfactant or a cationic surfactant, and probably because the amphoteric surfactant can play a role in destroying charges on the surfaces of strains for different strains, the killing effect can be better realized. The molecular weights of polyethylene glycol and chitosan were adjusted in samples 22-35. Generally, the larger the molecular weight of the both, the better the antibacterial effect, but when the molecular weight of chitosan or polyethylene glycol is too large, the overall viscosity is increased, and the feeling in the mouth is significantly affected.

Further, experiments 1 and 2 were performed on the samples 36 to 58, and the experimental results are shown in table 10.

TABLE 10 Experimental results of samples 36-58 in experiment 1 and experiment 2

In the samples 38-41, different antibacterial auxiliary agents are used, and different antibacterial auxiliary agents have different antibacterial effects on different bacteria, but can play a role in improving the antibacterial effect, can be selected according to actual needs during use, and can also be used in a mixed mode. In samples 42-50, the amounts of the nanofiber membrane and acetic acid are adjusted, the antibacterial effect of the nanofiber membrane is better when the amount of the nanofiber membrane is larger, but the improvement trend basically has an effect in the concentration of less than 50g/L, and after the concentration of the nanofiber membrane is further improved, the antibacterial performance is not obviously influenced, but the taste and the cost are obviously influenced. The excessive consumption of the acetic acid can cause the film forming performance of the nano-fiber film on the surface of the meat product to be poor, influence the long-acting antibacterial effect and also have obvious influence on the taste of the meat product.

In samples 52-58, the amount of the antibacterial liquid of the nanofiber membrane is adjusted, the antibacterial effect is improved along with the increase of the amount of the antibacterial liquid of the nanofiber membrane, but when the amount reaches 200 muL/cm²The highest antibacterial effect is achieved, and the subsequent continuous increase of the dosage of the nano antibacterial liquid has no obvious influence on the whole antibacterial effect.

In addition, experiment 3 was performed on part of samples 1 to 58 to verify the meat quality, and the experimental method was as follows: experiment 3, 1 to 58 samples were coated with an antibacterial solution, left for 5 days, and then boiled in distilled water and tasted. Meanwhile, a piece of duck meat is taken and placed at the temperature of minus 20 ℃, after being placed for 5 days, the duck meat is taken out, boiled in distilled water and tasted, and the sample X is marked. The mouth feel of each sample was scored and the results are shown in table 11.

Each sample was divided into 10 full points, and when the taste of the meat was similar to that of the untreated fresh meat, the meat was regarded as full points by the taster, and the meat was measured 10 times and averaged.

TABLE 11 sample mouthfeel List

Sample numbering	Score the points	Sample numbering	Score the points
				X	8.6	35	8.9
1	9.1	42	9.4
				2	9.1	43	9.3
3	9.3	44	9.3
				4	9.2	45	9.2
5	9.0	46	9.0
				6	8.8	52	9.4
7	9.0	53	9.3
				12	9.2	54	9.3
17	9.3	55	9.2
				28	8.7	56	9.2
33	9.3	57	8.9

According to the experiment, the antibacterial liquid is used for treating the duck meat, the influence on the taste of the duck meat is small, the taste of the duck meat can be preserved as much as possible compared with the treatment methods such as smoking, radiation and pickling, and the antibacterial liquid has a good practical value.

The present embodiment is only for explaining the present application, and it is not limited to the present application, and those skilled in the art can make modifications of the present embodiment without inventive contribution as needed after reading the present specification, but all of them are protected by patent law within the scope of the claims of the present application.