CN110551709A - nano cellulose based 3D printed lactic acid bacteria embedding material and preparation method and application thereof - Google Patents
nano cellulose based 3D printed lactic acid bacteria embedding material and preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a nano cellulose-based 3D printed lactic acid bacteria embedding material and a preparation method and application thereof. The method comprises the following steps: adding sodium alginate into the nano-cellulose solution, uniformly mixing, and centrifuging to obtain a mixed solution; adding chitosan into water, mixing, and adding glacial acetic acid to obtain chitosan solution; adding lactobacillus powder into the chitosan solution, and uniformly mixing to obtain chitosan-lactobacillus suspension; 3D printing is carried out on the mixed solution serving as an outer layer material and the chitosan-lactobacillus suspension serving as an inner layer material to obtain a printed product; and spraying a calcium chloride solution on the surface of the printed product, curing, and carrying out cold drying to obtain the lactobacillus embedding material for the nanocellulose-based 3D printing. According to the lactobacillus embedding material for the nanocellulose-based 3D printing, nanocellulose and chitosan are used as carriers to fix lactobacillus, efficient continuous circulating fermentation of yoghourt can be achieved, and the problems of difficult preservation, low utilization rate and high cost of the lactobacillus can be solved.
Description
Technical Field
The invention belongs to the field of food fermentation, and particularly relates to a nano cellulose-based 3D printed lactic acid bacteria embedding material, and a preparation method and application thereof.
Background
At present, the development of the yoghourt industry in China is full, and the yoghourt is favored by consumers with unique taste, flavor and high nutritive value. Lactic acid bacteria used for fermenting the yoghourt are human intestinal probiotics, and are widely developed and applied in the fields of food, medicine and the like. Because the wet lactic acid bacteria have short preservation period, the wet lactic acid bacteria are usually prepared into a dry state by a vacuum freeze drying technology so as to prolong the preservation time. However, the phase transition process of lyophilization can result in microbial cell loss, death, or inactivation of certain enzyme proteins. Therefore, it is required to develop a good protective agent for lactic acid bacteria to minimize damage, death, etc. of lactic acid bacteria and to improve the survival rate thereof. The traditional fermentation of lactic acid drinks, yoghourt and the like is to directly add dry or wet lactic acid bacteria, and is one-time addition fermentation, so that the strain consumption is large, the operation is complex and the cost is high.
3D printing technology has outstanding advantages in the accurate design and construction of material structures. The composite material with the hollow micro-pipeline structure is accurately constructed by utilizing the coaxial 3D printing technology, and the hollow micro-structure in the material can provide conditions for storage, activation and proliferation of lactic acid bacteria.
the nano-cellulose is a natural polysaccharide, has excellent shear thinning performance, is suitable for being used as a substrate of 3D ink, is a novel dietary fiber, and has a health-care effect on the gastrointestinal tract of a human body. However, 3D printing inks containing only nanocellulose have insufficient viscoelasticity.
Disclosure of Invention
in order to overcome the defects in the prior art, the invention aims to provide a nano cellulose-based 3D printed lactic acid bacteria embedding material, and a preparation method and application thereof.
The invention provides a nano-cellulose-based 3D printed lactic acid bacteria embedding material, and particularly relates to a nano-cellulose-based 3D printed lactic acid bacteria embedding material which is non-toxic and can provide a microenvironment for preservation, activation and proliferation of strains.
the invention also aims to provide a preparation method of the nano cellulose based 3D printed lactic acid bacteria embedding material. The preparation method comprises the steps of preparing nanocellulose-based coaxial 3D printing outer-layer ink by using nanocellulose as a matrix of the outer-layer printing ink and sodium alginate as a rheological reinforcing agent; the inner layer of the coaxial 3D printing adopts lactobacillus suspension dispersed in chitosan solution matrix; and then preparing a nanocellulose-based lactic acid bacteria embedded composite material with a hollow micro-pipeline by a coaxial 3D printing technology, namely the nanocellulose-based 3D printed lactic acid bacteria embedded material.
According to the preparation method of the nano cellulose based 3D printed lactobacillus embedding material, provided by the invention, an immobilization technology is used, the mobility of lactobacillus is limited by a physical or chemical method, and the activity is ensured, so that the technology opens up a new way for strain preservation. The invention combines the immobilization technology and the continuous culture technology, and develops a material which combines the continuous fermentation technology and the lactic acid bacteria preservation technology as a novel yoghourt starter to realize the continuous inoculation and fermentation of the yoghourt.
The purpose of the invention is realized by at least one of the following technical solutions.
The invention provides a preparation method of a nano cellulose based 3D printed lactobacillus embedding material, which adopts a mixed system of nano cellulose and sodium alginate as outer layer printing ink; adopting lactobacillus suspension dispersed in chitosan solution as inner layer printing solution; the nano cellulose-based 3D printing lactic acid bacteria embedding material which is nontoxic and can provide a microenvironment for preservation, activation and proliferation of strains is prepared through coaxial 3D printing.
The invention provides a nano-cellulose-based lactic acid bacteria embedding material for 3D printing, which is a nano-cellulose-based lactic acid bacteria embedding material for continuous fermentation of yoghourt.
the invention provides a nanocellulose-based 3D printing lactic acid bacteria embedding material, and a preparation method and application thereof. The invention provides a lactobacillus embedding material with a hollow microstructure prepared by a coaxial 3D printing technology, wherein the outer layer adopts nano-cellulose as a matrix, sodium alginate as a rheological reinforcing agent, and the inner layer is lactobacillus dispersion liquid under a chitosan matrix. The material takes the nano-cellulose and the chitosan as carriers to fix the lactic acid bacteria, realizes high-efficiency continuous circulating fermentation of the yoghourt, and can solve the problems of difficult preservation, low utilization rate and high cost of the lactic acid bacteria.
The invention provides a preparation method of a nano cellulose based 3D printed lactic acid bacteria embedding material, which comprises the following steps:
(1) Adding sodium alginate into the nano-cellulose solution under stirring, uniformly mixing, filling into a printing container, and centrifuging to remove bubbles to obtain a mixed solution (mixed system of nano-cellulose and sodium alginate);
(2) Adding chitosan into pure water at room temperature, mixing uniformly, adding glacial acetic acid (fully dissolving chitosan in water), and mixing uniformly to obtain a chitosan solution;
(3) Adding lactobacillus powder into the chitosan solution obtained in the step (2) under the condition of stirring, and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) Performing coaxial 3D printing by taking the mixed solution obtained in the step (1) as an outer layer material and the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material to obtain a 3D printing product (wet nanocellulose-based 3D printing lactobacillus embedding material);
(5) And (3) uniformly spraying a calcium chloride solution on the surface of the 3D printed product in the step (4), carrying out curing treatment, and then carrying out freeze drying to obtain the nano cellulose based 3D printed lactic acid bacteria embedding material.
further, the mass-volume ratio concentration of the nano-cellulose solution in the step (1) is 0.03-0.10 g/mL; the mass ratio of the sodium alginate to the nano cellulose solution is 1:10-1: 100.
Preferably, the sodium alginate is sodium alginate dry powder, and the purity level of the sodium alginate is analytically pure.
further, the centrifugation rate of the centrifugation treatment in the step (1) is 2000-8000rpm, and the centrifugation treatment time is 1-10 min.
Further, the mass-to-volume ratio of the chitosan to the water in the step (2) is 1-10: 1 g/L; the volume of the glacial acetic acid is 0.5-2.0% (volume fraction) of the volume of the chitosan solution.
Further, the mass ratio of the lactobacillus powder and the chitosan solution in the step (3) is 1:5-1: 100.
Further, the printing pressure of the coaxial 3D printing in the step (4) is 0.1-0.5MPa, and the size of the needle head of the coaxial 3D printing is 30G/21G-18G/14G; the printing speed of the outer layer is 1-10 mm/s; the inner layer printing speed is 1-5 mL/h.
further, the 3D printed product obtained in the step (4) does not need to be dried, and the calcium chloride solution is directly and uniformly sprayed on the 3D printed product in the step (5) to carry out curing treatment.
Further, the mass-volume ratio concentration of the calcium chloride solution in the step (5) is 0.01-0.1 g/mL; the mass-to-volume ratio of the 3D printing product (wet weight) to the calcium chloride solution is 1-5: 1 g/mL.
further, the time of the curing treatment in the step (5) is 5-50 s.
The invention provides a nano cellulose base 3D printed lactic acid bacteria embedding material prepared by the preparation method.
The nano cellulose-based 3D printed lactobacillus embedding material provided by the invention has excellent 3D printing performance and can effectively embed, fix and preserve lactobacillus. The nano cellulose-based 3D printed lactic acid bacteria embedding material provided by the invention can preserve lactic acid bacteria for more than one year at the temperature of 4-25 ℃.
The nano cellulose-based 3D printed lactic acid bacteria embedding material provided by the invention can be applied to yoghourt fermentation, especially the continuous fermentation process of yoghourt.
The nanocellulose-based 3D printing lactic acid bacteria embedding material for continuous fermentation of yogurt, which is prepared by the preparation method of any one of the above, has a good fixing and embedding effect on lactic acid bacteria.
According to the invention, a coaxial 3D printing technology is utilized, natural polysaccharide cellulose and chitosan are used as carriers to fix lactic acid bacteria, and the nano cellulose-based lactic acid bacteria embedding material with a hollow micro pipeline is prepared, and can provide a favorable microenvironment for the fixation, activation and proliferation of lactic acid bacteria. Thereby realizing high-efficiency continuous circulating fermentation of the yoghourt and solving the problems of difficult preservation, low utilization rate and high cost of the lactobacillus. The invention innovatively develops a novel food fermentation material and expands the application of the 3D printing technology in the field of food fermentation.
In order to meet the requirement of the coaxial printing outer layer material, the sodium alginate and the cellulose nanofiber are compounded to improve the property of the ink. If uniform dispersion of the lactic acid bacteria in the hollow micro-channel is to be achieved while entrapping the lactic acid bacteria, the inner layer printing solution used needs to have a certain consistency. In the preparation method provided by the invention, the chitosan solution can provide proper consistency, can be decomposed into chitosan oligosaccharide with gastrointestinal tract health care function under an acidic condition, and the slow release of the chitosan in the inner layer of the printing material can inhibit the excessive fermentation of the yoghourt in the continuous fermentation process. Thus, chitosan is suitable as a substrate for the inner printing solution.
compared with the prior art, the invention has the following advantages and beneficial effects:
(1) According to the preparation method provided by the invention, the adopted raw materials are nano-cellulose, sodium alginate and chitosan, and the preparation method has the advantages of wide source, low cost, safety and no toxicity; the nano cellulose-based 3D printed lactic acid bacteria embedding material provided by the invention has stable performance, simple preparation process, wide raw material source and low energy consumption, and is beneficial to large-scale industrial production;
(2) According to the preparation method provided by the invention, the coaxial 3D printing technology is utilized to design and construct the hollow micro-channels in the lactobacillus embedded composite material, so that the nano cellulose-based 3D printed lactobacillus embedded material is obtained, the material can provide structural conditions for the fixation, activation and proliferation of lactobacillus, and the material has the advantage of accurately and efficiently designing and constructing the internal structure of the material;
(3) the nano cellulose-based 3D printed lactobacillus embedding material provided by the invention can effectively embed, fix and preserve lactobacillus, and the lactobacillus can be preserved in the material for more than one year at 4-25 ℃;
(4) After the nano cellulose-based 3D printed lactobacillus embedding material provided by the invention is fermented, the inner layer chitosan in the embedding material (composite material) can be degraded into functional factors such as chitosan oligosaccharide and the like to be compounded with fermented yoghurt, and the formed compounded yoghurt has a health care effect on gastrointestinal tracts;
(5) if the nano-cellulose-based 3D-printed lactic acid bacteria embedding material provided by the invention is applied to the continuous fermentation process of the yoghourt, taking the volume of a fermentation system as 0.5-10L as an example, the nano-cellulose-based 3D-printed lactic acid bacteria embedding material provided by the invention can be added by only 1-10g, so that the continuous inoculation fermentation of the yoghourt can be realized, the yoghourt fermentation cost can be reduced, the probability of mixed bacteria pollution is reduced, and the fermentation efficiency is improved.
Drawings
Fig. 1 is a scanning electron microscope image of the nanocellulose-based 3D-printed lactic acid bacteria embedding material obtained in example 3.
FIG. 2 is a scanning electron microscope image of the cross section of the hollow micro-channel of the dry sample of the nanocellulose-based 3D-printed lactic acid bacteria embedding material obtained in example 3.
FIG. 3 is a scanning electron microscope image of the surface of a dried sample of the nanocellulose-based 3D-printed lactic acid bacteria embedding material obtained in example 3.
Detailed Description
The following examples are presented to further illustrate the practice of the invention, but the practice and protection of the invention is not limited thereto. It is noted that the processes described below, if not specifically described in detail, are all realizable or understandable by those skilled in the art with reference to the prior art. The reagents or apparatus used are not indicated to the manufacturer, and are considered to be conventional products available by commercial purchase.
example 1
A preparation method of a nano cellulose based 3D printed lactic acid bacteria embedding material comprises the following steps:
(1) Under the condition of room temperature, adding 0.2g of sodium alginate powder into 20g of nano-cellulose solution with the concentration of 0.03g/mL, uniformly mixing, then filling into a 3D printing needle cylinder, carrying out centrifugal treatment to eliminate bubbles, wherein the speed of the centrifugal treatment is 2000rpm, and the time of the centrifugal treatment is 1min, so as to obtain a mixed solution;
(2) Adding 0.01g of chitosan into 10mL of pure water, uniformly stirring to uniformly disperse the chitosan in the water, then adding 0.05mL of glacial acetic acid, uniformly mixing to completely dissolve the chitosan to obtain a chitosan solution;
(3) Adding 2g of lactobacillus powder into the chitosan solution in the step (2), and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) Taking the mixed solution obtained in the step (1) as an outer layer material, taking the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material, and carrying out coaxial 3D printing under the pressure of 0.1MPa, at the inner layer printing rate of 1mL/h, at the outer layer printing rate of 1mm/s and at the printing needle model of 30G/21G to obtain a 3D printing product;
(5) Uniformly spraying a calcium chloride solution with the concentration of 0.01g/mL on the surface of the 3D printing product in the step (4), wherein the mass-to-volume ratio of the 3D printing product (wet weight) to the calcium chloride solution is 1: 1g/mL, carrying out curing treatment for 5s, and then carrying out freeze drying to obtain the nanocellulose-based 3D printed lactic acid bacteria embedding material.
the effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 1 is similar to that of example 3, and the nano cellulose based 3D printed lactic acid bacteria embedding material also has a good fixing effect on lactic acid bacteria, and the lactic acid bacteria are attached to the printing material, and the shapes of the bacteria are full and complete, as shown in figure 1. The effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 1 is similar to that of example 3, the shape is the same, the fidelity is high, the formability is good, the weight is light, the storage and the transportation are easy, a hollow micro-channel with a complete structure is formed inside the material, the lactic acid bacteria are effectively fixed, embedded and stored by using the structure of the hollow micro-channel, and the damage to the thallus structure of the lactic acid bacteria is avoided in the process, which can be seen in fig. 2. The effect of the nano cellulose-based 3D printed lactic acid bacteria embedding material prepared in the embodiment 1 is similar to that of the embodiment 3, the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material is also compact, certain protection can be provided for strains in the storage process, the pollution of mixed bacteria and bacteriophage is reduced, a plurality of micropores are distributed on the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material, the nano cellulose-based 3D printed lactic acid bacteria embedding material can be used as contact points of lactic acid bacteria and raw milk during continuous fermentation of yoghourt.
Example 2
a preparation method of a nano cellulose based 3D printed lactic acid bacteria embedding material comprises the following steps:
(1) Under the condition of room temperature, adding 0.3g of sodium alginate powder into 20g of nano-cellulose solution with the concentration of 0.04g/mL, uniformly mixing, then filling into a 3D printing needle cylinder, carrying out centrifugal treatment to eliminate bubbles, wherein the speed of the centrifugal treatment is 3000rpm, and the time of the centrifugal treatment is 3min, so as to obtain a mixed solution;
(2) Adding 0.03g of chitosan into 10mL of pure water, uniformly stirring to uniformly disperse the chitosan in the water, then adding 0.07mL of glacial acetic acid, uniformly mixing to completely dissolve the chitosan to obtain a chitosan solution;
(3) Adding 1.5g of lactobacillus powder into the chitosan solution in the step (2), and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) Taking the mixed solution obtained in the step (1) as an outer layer material, taking the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material, and carrying out coaxial 3D printing, wherein the pressure of the coaxial 3D printing is 0.2MPa, the printing speed of the inner layer is 2mL/h, the printing speed of the outer layer is 3mm/s, and the type of a printing needle is 25G/18G, so as to obtain a 3D printing product;
(5) Uniformly spraying a calcium chloride solution with the concentration of 0.03g/mL on the surface of the 3D printing product in the step (4), wherein the mass-to-volume ratio of the 3D printing product (wet weight) to the calcium chloride solution is 2: 1g/mL, carrying out curing treatment for 10s, and then carrying out freeze drying to obtain the nanocellulose-based 3D printed lactic acid bacteria embedding material.
The effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 2 is similar to that of example 3, and the nano cellulose based 3D printed lactic acid bacteria embedding material also has a good fixing effect on lactic acid bacteria, and the lactic acid bacteria are attached to the printing material, and the shapes of the bacteria are full and complete, as shown in figure 1. The effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 2 is similar to that of example 3, the shape is the same, the fidelity is high, the formability is good, the weight is light, the storage and the transportation are easy, a hollow micro-channel with a complete structure is formed inside the material, the lactic acid bacteria are effectively fixed, embedded and stored by using the structure of the hollow micro-channel, and the damage to the thallus structure of the lactic acid bacteria is avoided in the process, which can be seen in fig. 2. The effect of the nano cellulose-based 3D printed lactic acid bacteria embedding material prepared in the embodiment 2 is similar to that of the embodiment 3, the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material is also compact, certain protection can be provided for strains in the storage process, the pollution of mixed bacteria and bacteriophage is reduced, a plurality of micropores are distributed on the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material, the nano cellulose-based 3D printed lactic acid bacteria embedding material can be used as contact points of lactic acid bacteria and raw milk during continuous fermentation of yoghourt.
example 3
A preparation method of a nano cellulose based 3D printed lactic acid bacteria embedding material comprises the following steps:
(1) Under the condition of room temperature, adding 0.4g of sodium alginate powder into 20g of nano-cellulose solution with the concentration of 0.05g/mL, uniformly mixing, then filling into a 3D printing needle cylinder, carrying out centrifugal treatment to eliminate bubbles, wherein the speed of the centrifugal treatment is 4000rpm, and the time of the centrifugal treatment is 5min, so as to obtain a mixed solution;
(2) Adding 0.05g of chitosan into 10mL of pure water, uniformly stirring to uniformly disperse the chitosan in the water, then adding 0.1mL of glacial acetic acid, uniformly mixing to completely dissolve the chitosan to obtain a chitosan solution;
(3) adding 1.0g of lactobacillus powder into the chitosan solution in the step (2), and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) Taking the mixed solution obtained in the step (1) as an outer layer material, taking the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material, and carrying out coaxial 3D printing, wherein the pressure of the coaxial 3D printing is 0.3MPa, the printing speed of the inner layer is 3mL/h, the printing speed of the outer layer is 4mm/s, and the type of a printing needle is 22G/19G, so as to obtain a 3D printing product;
(5) Uniformly spraying a calcium chloride solution with the concentration of 0.04g/mL on the surface of the 3D printing product in the step (4), wherein the mass-to-volume ratio of the 3D printing product (wet weight) to the calcium chloride solution is 3: 1g/mL, carrying out curing treatment for 20s, and then carrying out freeze drying to obtain the nanocellulose-based 3D printed lactic acid bacteria embedding material.
The nanocellulose-based 3D printing lactic acid bacteria embedding material prepared in example 3 has a good fixing effect on streptococcus thermophilus, as shown in an electron microscope image of figure 1, lactic acid bacteria are attached to the printing material, and the shapes of the cells are full and complete. The nanocellulose-based 3D printing lactic acid bacteria embedding material for continuous fermentation of the yogurt prepared by the method (namely the nanocellulose-based 3D printing lactic acid bacteria embedding material) is high in form fidelity, good in formability, light in weight and easy to store and transport. As shown in fig. 2, a hollow micro channel with a complete structure is formed inside the nanocellulose-based 3D-printed lactobacillus embedding material provided in example 3, and in combination with fig. 1, the nanocellulose-based 3D-printed lactobacillus embedding material provided in example 3 effectively utilizes the hollow micro channel structure to fix, embed and preserve the lactobacillus, and does not damage the thallus structure of the lactobacillus in the process. As shown in fig. 3, the surface of the nano cellulose-based 3D-printed lactic acid bacteria embedding material provided in example 3 is relatively dense, and can provide a certain protection for the strains during storage, thereby reducing the contamination of infectious microbes and bacteriophage. Meanwhile, the surface of the lactobacillus embedding material printed by the nanocellulose-based 3D printing provided by the embodiment 3 is distributed with a plurality of micropores, and the micropores can be used as contact points of lactobacillus and raw milk during continuous fermentation of yoghourt, so that continuous inoculation and fermentation are facilitated.
example 4
A preparation method of a nano cellulose based 3D printed lactic acid bacteria embedding material comprises the following steps:
(1) Under the condition of room temperature, adding 1.0g of sodium alginate powder into 20g of nano-cellulose solution with the concentration of 0.06g/mL, uniformly mixing, then filling into a 3D printing needle cylinder, carrying out centrifugal treatment to eliminate bubbles, wherein the speed of the centrifugal treatment is 6000rpm, and the time of the centrifugal treatment is 8min, thus obtaining a mixed solution;
(2) adding 0.08g of chitosan into 10mL of pure water, uniformly stirring to uniformly disperse the chitosan in the water, then adding 0.15mL of glacial acetic acid, uniformly mixing to completely dissolve the chitosan to obtain a chitosan solution;
(3) adding 0.5g of lactobacillus powder into the chitosan solution in the step (2), and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) taking the mixed solution obtained in the step (1) as an outer layer material, taking the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material, and carrying out coaxial 3D printing, wherein the pressure of the coaxial 3D printing is 0.4MPa, the printing speed of the inner layer is 4mL/h, the printing speed of the outer layer is 6mm/s, and the type of a printing needle is 19G/15G, so as to obtain a 3D printing product;
(5) uniformly spraying a calcium chloride solution with the concentration of 0.05g/mL on the surface of the 3D printing product in the step (4), wherein the mass-to-volume ratio of the 3D printing product (wet weight) to the calcium chloride solution is 4: 1g/mL, carrying out curing treatment for 30s, and then carrying out freeze drying to obtain the nanocellulose-based 3D printed lactic acid bacteria embedding material.
the effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 4 is similar to that of example 3, and the nano cellulose based 3D printed lactic acid bacteria embedding material also has a good fixing effect on lactic acid bacteria, and the lactic acid bacteria are attached to the printing material, and the shapes of the bacteria are full and complete, as shown in figure 1. The effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 4 is similar to that of example 3, the shape is the same, the fidelity is high, the formability is good, the weight is light, the storage and the transportation are easy, a hollow micro-channel with a complete structure is formed inside the material, the lactic acid bacteria are effectively fixed, embedded and stored by using the structure of the hollow micro-channel, and the damage to the thallus structure of the lactic acid bacteria is avoided in the process, which can be seen in fig. 2. The effect of the nano cellulose-based 3D printed lactic acid bacteria embedding material prepared in the embodiment 4 is similar to that of the embodiment 3, the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material is also compact, certain protection can be provided for strains in the storage process, the pollution of mixed bacteria and bacteriophage is reduced, a plurality of micropores are distributed on the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material, the nano cellulose-based 3D printed lactic acid bacteria embedding material can be used as contact points of lactic acid bacteria and raw milk during continuous fermentation of yoghourt.
Example 5
a preparation method of a nano cellulose based 3D printed lactic acid bacteria embedding material comprises the following steps:
(1) Under the condition of room temperature, adding 2.0g of sodium alginate powder into 20g of nano-cellulose solution with the concentration of 0.10g/mL, uniformly mixing, then filling into a 3D printing needle cylinder, carrying out centrifugal treatment to eliminate bubbles, wherein the speed of the centrifugal treatment is 8000rpm, and the time of the centrifugal treatment is 10min, thus obtaining a mixed solution;
(2) adding 0.10g of chitosan into 10mL of pure water, uniformly stirring to uniformly disperse the chitosan in the water, then adding 0.2mL of glacial acetic acid, uniformly mixing to completely dissolve the chitosan to obtain a chitosan solution;
(3) adding 0.1g of lactobacillus powder into the chitosan solution in the step (2), and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) taking the mixed solution obtained in the step (1) as an outer layer material, taking the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material, and carrying out coaxial 3D printing, wherein the pressure of the coaxial 3D printing is 0.5MPa, the printing speed of the inner layer is 5mL/h, the printing speed of the outer layer is 10mm/s, and the type of a printing needle is 18G/14G, so as to obtain a 3D printing product;
(5) Uniformly spraying a calcium chloride solution with the concentration of 0.1g/mL on the surface of the 3D printing product in the step (4), wherein the mass-to-volume ratio of the 3D printing product (wet weight) to the calcium chloride solution is 5: 1g/mL, carrying out curing treatment for 50s, and then carrying out freeze drying to obtain the nanocellulose-based 3D printed lactic acid bacteria embedding material.
The effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 5 is similar to that of example 3, and the nano cellulose based 3D printed lactic acid bacteria embedding material also has a good fixing effect on lactic acid bacteria, and the lactic acid bacteria are attached to the printing material, and the shapes of the bacteria are full and complete, as shown in figure 1. The effect of the nano cellulose based 3D printed lactic acid bacteria embedding material prepared in example 5 is similar to that of example 3, the shape is the same, the fidelity is high, the formability is good, the weight is light, the storage and the transportation are easy, a hollow micro-channel with a complete structure is formed inside the material, the lactic acid bacteria are effectively fixed, embedded and stored by using the structure of the hollow micro-channel, and the damage to the thallus structure of the lactic acid bacteria is avoided in the process, which can be seen in fig. 2. The effect of the nano cellulose-based 3D printed lactic acid bacteria embedding material prepared in the embodiment 5 is similar to that of the embodiment 3, the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material is also compact, certain protection can be provided for strains in the storage process, the pollution of mixed bacteria and bacteriophage is reduced, a plurality of micropores are distributed on the surface of the nano cellulose-based 3D printed lactic acid bacteria embedding material, the nano cellulose-based 3D printed lactic acid bacteria embedding material can be used as contact points of lactic acid bacteria and raw milk during continuous fermentation of yoghourt.
The above examples are only preferred embodiments of the present invention, which are intended to be illustrative and not limiting, and those skilled in the art should understand that they can make various changes, substitutions and alterations without departing from the spirit and scope of the invention.
Claims (10)
1. The preparation method of the lactobacillus embedding material for nanocellulose-based 3D printing is characterized by comprising the following steps:
(1) adding sodium alginate into the nano-cellulose solution, uniformly mixing, and then carrying out centrifugal treatment to eliminate bubbles to obtain a mixed solution;
(2) Adding chitosan into water, uniformly mixing, then adding glacial acetic acid, and uniformly mixing to obtain a chitosan solution;
(3) adding lactobacillus powder into the chitosan solution in the step (2), and uniformly mixing to obtain chitosan-lactobacillus suspension;
(4) performing coaxial 3D printing by taking the mixed solution obtained in the step (1) as an outer layer material and the chitosan-lactobacillus suspension obtained in the step (3) as an inner layer material to obtain a 3D printing product;
(5) And (3) uniformly spraying a calcium chloride solution on the surface of the 3D printed product in the step (4), carrying out curing treatment, and then carrying out freeze drying to obtain the nano cellulose based 3D printed lactic acid bacteria embedding material.
2. the preparation method according to claim 1, wherein the mass-to-volume concentration of the nanocellulose solution of step (1) is 0.03-0.10 g/mL; the mass ratio of the sodium alginate to the nano cellulose solution is 1:10-1: 100.
3. the method as claimed in claim 1, wherein the centrifugation in step (1) is carried out at a centrifugation rate of 2000-8000rpm for 1-10 min.
4. The method according to claim 1, wherein the mass-to-volume ratio of the chitosan to the water in the step (2) is 1-10: 1 g/L; the volume of the glacial acetic acid is 0.5-2.0% of the volume of the chitosan solution.
5. The preparation method according to claim 1, wherein the mass ratio of the lactobacillus powder to the chitosan solution in the step (3) is 1:5-1: 100.
6. the preparation method according to claim 1, wherein the printing pressure of the coaxial 3D printing in the step (4) is 0.1-0.5MPa, and the needle size of the coaxial 3D printing is 30G/21G-18G/14G; the printing speed of the outer layer is 1-10 mm/s; the inner layer printing speed is 1-5 mL/h.
7. the preparation method according to claim 1, wherein the mass-to-volume concentration of the calcium chloride solution in the step (5) is 0.01 to 0.1 g/mL; the mass-to-volume ratio of the 3D printing product to the calcium chloride solution is 1-5: 1 g/mL.
8. the production method according to claim 1, wherein the curing treatment time of step (5) is 5 to 50 seconds.
9. A nanocellulose-based 3D-printed lactic acid bacteria embedding material prepared by the preparation method of any one of claims 1 to 8.
10. use of the nanocellulose-based 3D printed lactic acid bacteria embedding material of claim 9 in yogurt fermentation.
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