CN114522635B

CN114522635B - Antibacterial microcapsule capable of controllably releasing cinnamaldehyde and preparation method thereof

Info

Publication number: CN114522635B
Application number: CN202210081044.6A
Authority: CN
Inventors: 黄强; 高晴; 何若岚; 张斌; 扶雄
Original assignee: South China University of Technology SCUT; Zhuhai Institute of Modern Industrial Innovation of South China University of Technology
Current assignee: South China University of Technology SCUT; Zhuhai Institute of Modern Industrial Innovation of South China University of Technology
Priority date: 2022-01-24
Filing date: 2022-01-24
Publication date: 2023-08-25
Anticipated expiration: 2042-01-24
Also published as: CN114522635A

Abstract

The invention discloses an antibacterial microcapsule capable of controllably releasing cinnamaldehyde and a preparation method thereof; dispersing starch in an ethanol water solution 1, and dropwise adding a NaOH solution; stirring, dripping ethanol water solution 2, and centrifuging; washing with ethanol water solution 3, dispersing, neutralizing with ethanol acid solution, centrifuging, washing with ethanol water solution 4 and absolute ethanol respectively, drying, pulverizing, and sieving to obtain hollow V-shaped starch; dispersing the extract in absolute ethyl alcohol containing cinnamaldehyde, and then adding deionized water to ensure that the mass concentration of the ethyl alcohol is 40-60%; sealing and stirring, cooling to room temperature, centrifuging, washing and drying to obtain the antibacterial microcapsule. The microcapsule prepared by the invention can efficiently encapsulate cinnamaldehyde (62.38 mg/g), has excellent controlled release effect (sustained release for 3 weeks at normal temperature) on cinnamaldehyde, and can remarkably inhibit the growth of staphylococcus aureus and escherichia coli (the diameter of a bacteriostasis zone reaches 12 mm).

Description

Antibacterial microcapsule capable of controllably releasing cinnamaldehyde and preparation method thereof

Technical Field

The invention belongs to the fields of food, medicine and chemical industry, and in particular relates to an antibacterial microcapsule capable of controllably releasing cinnamaldehyde and a preparation method thereof.

Background

In recent years, with the increasing demand for green sources of natural origin and related sustainable healthier products, essential oils from aromatic plants have received great attention. Essential oils are natural antimicrobial agents with great potential for use in the food, pharmaceutical and cosmetic industries. Cinnamaldehyde (3-phenyl-2-acraldehyde) is hydrophobic aromatic aldehyde with low molecular weight, and is a main active ingredient of cinnamon essential oil. In addition to providing cinnamon flavor, cinnamaldehyde has very excellent antibacterial, antioxidant, antifungal and anti-inflammatory activity. Its safety and effectiveness has been certified by the food additives expert committee (JECFA) of the grain and farming/world health organization. However, the efficient use of essential oils remains a significant challenge due to their high volatility and low water solubility. Some extreme conditions, such as in overheated and peracid overbased environments, may lead to loss or degradation of the active ingredient and further affect the flavor, texture and shelf life of the final product.

At present, the wall material for encapsulating cinnamaldehyde comprises cyclodextrin, emulsion, chitosan, pectin, sodium alginate and other non-starch polysaccharide. The nanoemulsion was prepared to deliver cinnamaldehyde with cinnamaldehyde stability of only 48h, and the use of a large amount of surfactant tween 20 (LWT-Food Science and Technology,2015,60 (1), 444-451) was unavoidable in the emulsion preparation process. The cinnamaldehyde is encapsulated with beta-cyclodextrin, which can only be released for 192 hours continuously, and the cost of cyclodextrin is higher than that of starch (Food Hydrocolloids,2019,90,360-366). Cinnamaldehyde microcapsules were prepared using ultrasonic-assisted cross-linking of chitosan and pectin and spray-dried to give final samples (Food Hydrocolloids,2020,99, 105316). The method has complicated steps, high raw material and equipment cost, and high temperature of 150 ℃ in the spray drying stage, thereby causing partial cinnamaldehyde loss. The development cost is low, the technology is simple and convenient, and the controllable release cinnamaldehyde microcapsule is unprecedented.

Disclosure of Invention

The invention aims to provide an antibacterial microcapsule with controllable release of cinnamaldehyde, which is simple to operate, low in cost and environment-friendly, and can release cinnamaldehyde for a long time, and a preparation method thereof.

The invention aims at realizing the following technical scheme:

a method for preparing an antibacterial microcapsule capable of controllably releasing cinnamaldehyde, comprising the following steps:

(1) Preparation of hollow V-shaped starch

(a) Dispersing starch in ethanol water solution 1 to prepare starch milk, and dripping NaOH solution; stirring, dripping ethanol water solution 2, and centrifuging;

(b) Washing and dispersing the precipitate obtained in the step (a) by using an ethanol water solution 3, neutralizing by using an ethanol acid solution, centrifuging, washing once by using an ethanol water solution 4 and absolute ethanol respectively, drying, crushing and sieving to obtain hollow V-shaped starch;

(2) Antibacterial microcapsule prepared by embedding cinnamaldehyde into hollow V-shaped starch

Dispersing the hollow V-shaped starch in absolute ethyl alcohol containing cinnamaldehyde, and then adding deionized water to ensure that the mass concentration of the ethyl alcohol is 40-60%; sealing and stirring, cooling to room temperature, centrifuging, washing and drying to obtain the antibacterial microcapsule capable of controllably releasing cinnamaldehyde.

Preferably, in the step (1), the mass fraction of starch in the starch milk in the step (a) is 3-15%;

preferably, in the step (1), the concentration of the NaOH solution in the step (a) is 2-4M; further preferably, the concentration of the NaOH solution is 2-3M;

preferably, in the step (1), the mass ratio of the starch milk, the NaOH solution and the ethanol aqueous solution 2 in the step (a) is 70-90:30-70:10-30.

Preferably, in the step (1), the dropwise adding of the NaOH solution in the step (a) is completed within 5-15 min, so that the slow dropwise adding is ensured to avoid local aggregation caused by too high speed;

preferably, in the step (1), the stirring treatment in the step (a) is performed for 20-40 min;

preferably, in the step (1), the rotational speed of the centrifugation in the step (a) is 6000-10000 g, and the time is 5-15 min.

Preferably, in the step (1), after the starch in the step (a) is dispersed in the ethanol water solution 1, stirring is carried out at a rotating speed of 50-150 rpm/min to obtain starch milk; further preferably, the stirring is carried out at a rotation speed of 100 to 150 rpm/min.

Preferably, in the step (1), the mass fraction of the ethanol in the ethanol aqueous solution 1, the ethanol aqueous solution 2 and the ethanol aqueous solution 3 is 20-60%. Further preferably, the mass fraction of the ethanol in the ethanol aqueous solution 1, the ethanol aqueous solution 2 and the ethanol aqueous solution 3 is 40-60%.

Preferably, in step (1), the number of times of washing the aqueous ethanol solution 3 in step (b) is 1 to 3;

preferably, in step (1), the hydrochloric acid solution in step (b) is hydrochloric acid dispersed in absolute ethanol so that the concentration of the hydrochloric acid is 2-4M.

Preferably, in the step (1), the temperature of the drying in the step (b) is 40-80 ℃; sieving the mixture to obtain a sieve with 100-200 meshes;

preferably, in the step (1), the mass fraction of the ethanol in the ethanol aqueous solution 4 in the step (b) is 70-90%.

Preferably, in the step (2), after deionized water is added, the mass concentration of the hollow V-shaped starch is 2-10%; further preferably, the mass concentration of the hollow V-shaped starch is 5-10%;

preferably, in the step (2), the mass ratio of the hollow V-shaped starch to the cinnamaldehyde is 2-5:1-2.

Preferably, in the step (2), the time of sealing and stirring is 0.5-2 h, and the temperature is 25-85 ℃.

Preferably, in step (1), the starch in step (a) is at least one of corn starch, potato starch, wheat starch and tapioca starch;

preferably, in the step (2), the washing is washing with absolute ethyl alcohol, and the drying is drying in an oven at 30-60 ℃.

The antibacterial microcapsule with controllable release of cinnamaldehyde prepared by the preparation method is provided.

Compared with the prior art, the invention has the advantages that:

(1) The cinnamaldehyde microcapsule prepared by the invention can efficiently embed cinnamaldehyde, and the content of the cinnamaldehyde in the compound can reach 62.38mg/g. The invention composites the hollow V-shaped starch and cinnamaldehyde in ethanol water solution at a certain temperature. The proper concentration of the ethanol can not only improve the solubility of the cinnamaldehyde in a solvent system, but also enable the starch chain to have certain mobility, and enable the starch chain to be fully contacted and compounded with the cinnamaldehyde molecules. The proper temperature increase is beneficial to increasing the Brownian motion of molecules and promoting the encapsulation of cinnamaldehyde.

(2) The cinnamaldehyde microcapsule prepared by the invention can realize long-acting release of cinnamaldehyde. Compared with the physical mixture of cinnamaldehyde, the cinnamaldehyde microcapsule prepared by the invention has obviously prolonged release time of the cinnamaldehyde under different temperature or humidity conditions. In practical application, the temperature and the humidity can be selected according to the requirement to regulate and control the release of the cinnamaldehyde, for example, a low-temperature drying and sealing environment is selected when the microcapsule is stored; if the condition of high temperature and high humidity is needed for quick release of cinnamaldehyde, the controllable release of cinnamaldehyde is realized.

(3) The cinnamaldehyde microcapsule prepared by the invention has remarkable antibacterial activity. The microcapsule with the concentration of 10mg/L can inhibit the growth of staphylococcus aureus and escherichia coli, and the inhibition effect of the concentration of 20mg/L on the staphylococcus aureus and escherichia coli is more remarkable.

(4) The raw material adopted by the invention is starch, and has the characteristics of rich resources and low cost; the compound solvent is water and ethanol, is environment-friendly, nontoxic and environment-friendly, has simple preparation process, does not need large-scale equipment, is suitable for laboratory operation, and can easily realize industrial production.

Drawings

FIG. 1 is an X-ray diffraction pattern of cinnamaldehyde microcapsules of example 1 and comparative example 1.

Detailed Description

For a better understanding of the present invention, the present invention will be further described with reference to the following examples, but the scope of the present invention is not limited to the scope expressed by examples.

Quantification of cinnamaldehyde in microcapsules:

in the examples, the content of cinnamaldehyde in the complex was quantified by gas chromatography. Specifically, 15mg of the complex was dispersed in 2mL of ethyl acetate and 1mL of deionized water, vortexed, sonicated until the complex was completely dissolved, and cinnamaldehyde was extracted into the organic phase. Gas chromatography is combined with a hydrogen Flame Ion Detector (FID) and an agilent HP-5 capillary column, and nitrogen is the carrier gas. The test conditions were: the sample injection amount is 1 mu L, and the split ratio is 20:1; the initial temperature of the chromatographic column is 80 ℃, 1min is kept, the temperature is increased to 250 ℃ at 30 ℃/min, and the temperature is kept for 2min; the temperature of the sample inlet is 250 ℃; the detector temperature was 250 ℃; the current carrying rate is 1mL/min. The corresponding peak area was converted to concentration according to the standard of cinnamaldehyde.

Antibacterial effect test of antibacterial microcapsules:

the strain E.coli ATCC25922 and staphylococcus aureus RN4220 are respectively inoculated into LB liquid culture medium, cultured at 37 ℃ and 180rpm, the OD600 value of the bacterial liquid is controlled to be slightly less than 1 by using an ultraviolet spectrophotometer, the bacterial liquid is taken out and diluted by 2 times by sterilized normal saline, 160 mu L of diluted bacterial liquid is added into 100mL of LB solid culture medium (agar content is 0.7% -1%), and the bacterial liquid is prepared into a flat plate. Taking a proper amount of starch cinnamaldehyde compound samples in the examples, respectively adding the starch cinnamaldehyde compound samples into a solid plate, setting a blank control group, placing the blank control group into a 37 ℃ incubator for culturing for 9 hours, and observing the results.

Comparative example 1

(1) Preparation of hollow V-shaped starch

10g of starch was dispersed in 70g of aqueous ethanol (40%, w/w) and 50g of 3M NaOH solution was added dropwise at a rate of 4 g/min. After stirring for 30min, 20g of ethanol aqueous solution (40%, w/w) was slowly added dropwise to the starch milk at normal temperature, and the mixture was centrifuged at 8000g for 10min;

washing the obtained precipitate with ethanol water solution (40% w/w) for 2 times, dispersing, neutralizing with 3M ethanol acid solution, centrifuging, washing with ethanol water solution (80% w/w) and absolute ethanol respectively for one time, drying the obtained starch in a 40 ℃ oven, pulverizing, and sieving to obtain hollow V-shaped starch;

2g of hollow V-shaped starch and 2g of cinnamaldehyde are fully stirred and mixed, sealed and compounded for 3 hours at 75 ℃. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

The test shows that the content of cinnamaldehyde in the obtained compound is 17.20mg/g, and the cinnamaldehyde can be slowly released for about 3 days at normal temperature. The 20mg/ml cinnamaldehyde microcapsule has a bacteriostasis zone of 3.80 mm and 3.20mm on escherichia coli and staphylococcus aureus respectively, and the bacteriostasis effect is weak, mainly because the content of the embedded cinnamaldehyde in the microcapsule is low.

Comparative example 2

(1) Preparation of hollow V-shaped starch

2g of the hollow V-starch and 2g of cinnamaldehyde were dispersed in 36g of deionized water, and the mixture was sealed and stirred at 75℃for compounding for 1 hour. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

The compound obtained by testing has the content of cinnamaldehyde of 6.24mg/g, and can release the cinnamaldehyde for about 0.5 days at normal temperature. The 20mg/ml cinnamaldehyde microcapsule has substantially no antibacterial effect on escherichia coli and staphylococcus aureus, mainly because the content of the embedded cinnamaldehyde in the microcapsule is too low.

Comparative example 3

(1) Preparation of hollow V-shaped starch

2g of the hollow V-shaped starch and 2g of cinnamaldehyde are dispersed in 36g of absolute ethanol, and the mixture is sealed and stirred at 75 ℃ for compounding for 1h. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

The test shows that the content of cinnamaldehyde in the obtained compound is 9.53mg/g, and the cinnamaldehyde can be released for about 1 day at normal temperature. The 20mg/ml cinnamaldehyde microcapsule has little antibacterial effect on escherichia coli and staphylococcus aureus, mainly because the content of the embedded cinnamaldehyde in the microcapsule is small.

Example 1

(1) Preparation of hollow V-shaped starch

2g of hollow V-starch was dispersed in absolute ethanol containing 2g of cinnamaldehyde, and then 16g of deionized water was added to make the ethanol concentration 50% (w/w). The mixture was sealed and stirred at 75 ℃ for compounding for 1h. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

Through testing, the content of cinnamaldehyde in the obtained compound is 62.38mg/g, and the cinnamaldehyde can be slowly released for 20 days at normal temperature. The inhibition zones of 10mg/ml cinnamaldehyde microcapsules on escherichia coli and staphylococcus aureus are 6.30 mm and 9.25mm respectively; the 20mg/ml cinnamaldehyde microcapsule has a bacteriostasis zone of 10.10 mm and 11.30mm on escherichia coli and staphylococcus aureus respectively. The cinnamaldehyde microcapsule can obviously inhibit the growth of escherichia coli and staphylococcus aureus.

In comparative example 1, the dry-process preparation of cinnamaldehyde microcapsules has a low embedding rate of cinnamaldehyde, and the release rate of cinnamaldehyde is too high during storage, and the inhibition effect on escherichia coli and staphylococcus aureus is poor due to the low embedding amount of cinnamaldehyde, which is unfavorable for the application of food industry. In comparative example 2, the complex environment of V-type starch and cinnamaldehyde was water, but cinnamaldehyde was poorly soluble in pure water, which was unfavorable for sufficient contact of cinnamaldehyde molecules with starch chains, thus resulting in lower cinnamaldehyde inclusion amount and no antibacterial property. In comparative example 3, the complex environment was pure absolute ethanol, which has a stronger ability to compete for cinnamaldehyde than the hydrophobic cavity of V-shaped starch due to the hydrophobicity of cinnamaldehyde molecules, so cinnamaldehyde is more prone to be dissolved in a solvent during the complex, resulting in lower cinnamaldehyde entrapment rate in the complex. Dry method, pure water or pure absolute ethyl alcohol is not beneficial to the combination of cinnamaldehyde and V-shaped starch; under the condition of the ethanol water solution with medium concentration, the starch chain has a certain degree of freedom, and cinnamaldehyde is properly dissolved in the solvent, so that more cinnamaldehyde molecules can enter the spiral cavity of the amylose.

Compared with the comparative example, the content of the wet-process embedded cinnamaldehyde in the example 1 is obviously improved, the release time is greatly improved, and the wet-process embedded cinnamaldehyde has good effect of resisting citrus of large intestine and staphylococcus aureus. The good solvent environment in the wet method is favorable for fully contacting dispersed starch and cinnamaldehyde molecules, and the ethanol with proper concentration (50%) is favorable for improving the solubility of the cinnamaldehyde and the mobility of starch chains. FIG. 1 is a graph showing the X-ray diffraction patterns of the cinnamaldehyde microcapsules of example 1 and comparative example 1, wherein the diffraction peak of the sample obtained in example 1 is sharp and the half-width is very narrow, which indicates that the crystal structure is more compact and the crystallization is more perfect, while the diffraction peak of the sample obtained in comparative example 1 is softer and the amorphous "steamed bread peak" area is larger, which indicates that the crystallization order is worse and the crystallization number is smaller. Higher temperatures increase the thermal movement of the molecules and under the toughening action of aqueous ethanol, amylose can align into a more ordered and stable structure, forming more single-helical cavities, providing more sites for binding with cinnamaldehyde molecules. Therefore, the embedding content of cinnamaldehyde is greatly improved. The crystals are orderly arranged, so that the order is high, and the cinnamaldehyde can be slowly released for a long time. The cinnamic aldehyde has high embedding content and slow release, so that the cinnamic aldehyde has excellent characteristics of resisting escherichia coli and staphylococcus aureus, thereby achieving the purposes of controllable release and antibacterial property.

Example 2

(1) Preparation of hollow V-shaped starch

2g of hollow V-starch was dispersed in absolute ethanol containing 2g of cinnamaldehyde, and then 20g of deionized water was added to make the ethanol concentration 40% (w/w). The mixture was sealed and stirred at 75 ℃ for compounding for 1h. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

The test shows that the content of cinnamaldehyde in the obtained compound is 59.48mg/g, and the cinnamaldehyde can be slowly released for about 19 days at normal temperature. The 20mg/ml cinnamaldehyde microcapsule has 9.25mm and 10.58mm of inhibition zone on coliform bacteria and staphylococcus aureus.

Example 3

(1) Preparation of hollow V-shaped starch

2g of hollow V-starch was dispersed in absolute ethanol containing 2g of cinnamaldehyde, and then 12g of deionized water was added to make the ethanol concentration 60% (w/w). The mixture was sealed and stirred at 75 ℃ for compounding for 1h. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

The test shows that the content of cinnamaldehyde in the obtained compound is 51.04mg/g, and the cinnamaldehyde can be slowly released for about 21 days at normal temperature. The 20mg/ml cinnamaldehyde microcapsule has 9.53 mm and 10.52mm inhibition zones on escherichia coli and staphylococcus aureus.

Example 4

(1) Preparation of hollow V-shaped starch

2g of hollow V-starch was dispersed in absolute ethanol containing 2g of cinnamaldehyde, and then 16g of deionized water was added to make the ethanol concentration 50% (w/w). The mixture was sealed and stirred at 85 ℃ for compounding for 1h. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

Through testing, the content of cinnamaldehyde in the obtained compound is 45.99mg/g, and the release of 60% cinnamaldehyde at normal temperature takes 17 days. The 20mg/ml cinnamaldehyde microcapsule has 9.85 mm and 11.20mm inhibition zones on escherichia coli and staphylococcus aureus.

Example 5

(1) Preparation of hollow V-shaped starch

2g of hollow V-starch was dispersed in absolute ethanol containing 2g of cinnamaldehyde, and then 16g of deionized water was added to make the ethanol concentration 50% (w/w). The mixture was sealed and stirred at 65 ℃ for compounding for 1h. After cooling to room temperature, the mixture was centrifuged, washed and dried in an oven at 40 ℃.

The test shows that the content of cinnamaldehyde in the obtained compound is 42.60mg/g, and the release of 50% cinnamaldehyde at normal temperature takes 15 days. The 20mg/ml cinnamaldehyde microcapsule has inhibition zones of 10.22 mm and 10.67mm on escherichia coli and staphylococcus aureus respectively.

The antibacterial microcapsule capable of controllably releasing the cinnamaldehyde and the preparation method thereof can effectively embed the cinnamaldehyde and realize long-acting release of the cinnamaldehyde, and have remarkable antibacterial activity; the cost is low, the preparation process is simple and convenient, large-scale equipment is not needed, and the method is suitable for laboratory operation and can easily realize industrial production.

It should be noted that, the embodiments of the present invention are not limited by the above examples, and any other changes, modifications, substitutions, combinations, and simplifications that do not depart from the spirit and principle of the present invention should be equivalent to the embodiments described above, and are included in the protection scope of the present invention.

Claims

1. A method for preparing an antimicrobial microcapsule capable of controllably releasing cinnamaldehyde, which is characterized by comprising the following steps:

(1) Preparation of hollow V-shaped starch

Dispersing the hollow V-shaped starch in absolute ethyl alcohol containing cinnamaldehyde, and then adding deionized water to ensure that the mass concentration of the ethyl alcohol is 40-60%; sealing and stirring, cooling to room temperature, centrifuging, washing and drying to obtain the antibacterial microcapsule capable of controllably releasing cinnamaldehyde; the sealing and stirring time is 0.5-2 h, and the temperature is 65-85 ℃.

2. The preparation method according to claim 1, wherein in the step (1), the mass fraction of starch in the starch milk in the step (a) is 3-15%;

the concentration of the NaOH solution is 2-4M;

the mass ratio of the starch milk to the NaOH solution to the ethanol water solution 2 is 70-90:30-70:10-30.

3. The method according to claim 1, wherein in the step (1), the dropwise addition of the NaOH solution in the step (a) is completed within 5 to 15 minutes;

the stirring treatment time is 20-40 min;

the rotational speed of the centrifugation is 6000-10000 g, and the time is 5-15 min.

4. The preparation method according to claim 1, wherein in the step (1), the starch obtained in the step (a) is dispersed in the ethanol aqueous solution 1 and stirred at a rotation speed of 50 to 150rpm/min to obtain starch milk;

the mass fraction of the ethanol in the ethanol aqueous solution 1, the ethanol aqueous solution 2 and the ethanol aqueous solution 3 is 20-60%.

5. The method according to claim 1, wherein in the step (1), the aqueous ethanol solution 3 of the step (b) is washed 1 to 3 times;

the ethanol acid solution is prepared by dispersing hydrochloric acid in absolute ethyl alcohol to ensure that the concentration of the hydrochloric acid is 2-4M.

6. The method according to claim 1, wherein in the step (1), the drying temperature in the step (b) is 40 to 80 ℃; sieving the mixture to obtain a sieve with 100-200 meshes;

the mass fraction of the ethanol in the ethanol water solution 4 is 70-90%.

7. The preparation method according to claim 1, wherein in the step (2), after deionized water is added, the mass concentration of the hollow V-shaped starch is 2-10%;

the mass ratio of the hollow V-shaped starch to the cinnamaldehyde is 2-5:1-2.

8. The method according to claim 1, wherein in step (1), the starch in step (a) is at least one of corn starch, potato starch, wheat starch and tapioca starch; in the step (2), the washing is carried out by using absolute ethyl alcohol, and the drying is carried out in an oven at 30-60 ℃.

9. The antibacterial microcapsule with controllable release of cinnamaldehyde prepared by the preparation method of any one of claims 1 to 8.