CN114702703A - Egg white hydrogel film/composite film based on one-way nanopore dehydration and preparation method thereof - Google Patents
Egg white hydrogel film/composite film based on one-way nanopore dehydration and preparation method thereof Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J5/00—Manufacture of articles or shaped materials containing macromolecular substances
- C08J5/18—Manufacture of films or sheets
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2389/00—Characterised by the use of proteins; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2479/00—Characterised by the use of macromolecular compounds obtained by reactions forming in the main chain of the macromolecule a linkage containing nitrogen with or without oxygen, or carbon only, not provided for in groups C08J2461/00 - C08J2477/00
- C08J2479/04—Polycondensates having nitrogen-containing heterocyclic rings in the main chain; Polyhydrazides; Polyamide acids or similar polyimide precursors
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Abstract
The invention relates to an egg white hydrogel film/composite film based on one-way nanopore dehydration and a preparation method thereof. The egg white or the mixture solution thereof is dehydrated in a container with a nanopore filter membrane as the bottom through a downward unidirectional nanopore to obtain transparent, fragile and water-soluble egg white bioglass, and then the egg white bioglass/composite membrane with high wet transparency, elasticity and tensile property is prepared after heat treatment; the tensile strength of the material reaches 5.8 MPa, the elongation at break is 90-110%, and the swelling ratio is 60-130%; the egg white hydrogel film/composite film is prepared into the polypyrrole-modified egg white hydrogel conductive film through polymerization reaction. Mouse fibroblast cells L929 have good survival and proliferation on the prepared egg white hydrogel membrane. The egg white hydrogel material based on one-way nanopore dehydration provided by the invention can be widely applied to the fields of medical biomaterials, 3D (three-dimensional) stents, bionic materials, drug delivery and sustained release, wearable electronic equipment and the like.
Description
Technical Field
The invention relates to an egg white hydrogel film/composite film and a conductive egg white hydrogel film formed by heat treatment of water-soluble bioglass prepared from homogeneous egg white, and a preparation method thereof, in particular to a preparation method of a flexible or conductive egg white hydrogel film/composite film/conductive film with excellent mechanical strength and good biocompatibility, belonging to the fields of biological materials, bionic materials, wearable equipment, medical tissue engineering materials and the like.
Background
Egg white, a widely high-quality and low-cost animal-derived protein, has an amino acid composition closest to that of a human body due to various physiological functions and biochemical activities, plays an important role in maintaining human health, and has been widely applied to the fields of medical treatment, food, bioengineering and the like. Egg white accounts for about 60% of the total weight of the egg, and is a yellowish, transparent and silk-sticky colloidal solution consisting of 11% of protein and 88% of water. The albumen of the egg white mainly comprises five glycoprotein such as 54 percent of egg white protein, 12 percent of ovotransferrin, 11 percent of ovomucoid, 3.5 percent of ovomucoid and 3.2 percent of lysozyme, and the like, and accounts for 84 percent of the total amount of the egg white protein. The egg white is rich in nutrition and has been used at presentBecoming one of the important materials for edible film manufacture, replacing conventional plastics and reducing the risk caused by pollution are expected. However, the mechanical property, the gel property and the solubility of the egg white protein film are poor. Therefore, a novel edible film with multiple functions, such as an egg white-glycerin film, an egg white-glycerin-lipid film, a self-crosslinking egg white bioplastic, an egg white-gelatin composite plastic or an egg white-casein bioplastic film, is developed by chemical crosslinking by utilizing disulfide bonds and sulfydryl groups rich in egg white. These edible films or food packaging films not only have a certain mechanical strength, plasticity and oxygen barrier, but also are more sensitive to factors such as humidity changes. Mixing with high polymer, active substance or biological cross-linking agent to obtain antioxidant curcumin-releasing egg white capsule, and coating TiO on egg white2Or the silver nano-particles are prepared into anti-tumor drugs or biological nano-composite hydrogel for drug slow release or delivery. The egg white and hydroxyapatite are crosslinked to prepare porous foam egg white biological ceramic suitable for soft tissue or bone growth, the porous foam egg white biological ceramic is crosslinked with dihydroxyindole to synthesize an egg white biological bionic membrane with melanin biological synthesis function, and the egg white biological bionic membrane is mixed with keratin to prepare a luminescent gold cluster composite membrane and the like.
The above researches mainly focus on synthesizing medical biomaterials, drug delivery and sustained release, green industrial products, etc. with egg white as a raw material through physical mixing and chemical crosslinking together with other materials. The technical scheme that the egg white bioglass is prepared by taking homogeneous egg white or single homogeneous egg white as a raw material and then is processed into a functional egg white hydrogel film/composite film with excellent mechanical properties through heat treatment is not reported.
Disclosure of Invention
Aiming at the defects of the existing multifunctional egg white biological composite material in the aspects of preparation technology, green processing, functionality, mechanical property and the like, the invention adopts the unidirectional nanopore dehydration technology, water molecules in egg white are subjected to unidirectional dehydration through filter membrane nanopores to cause the ordered arrangement of egg white protein molecules to form water-soluble, transparent and fragile bioglass, and provides a flexible, transparent and strong mechanical property egg white hydrogel membrane/composite membrane, a functional egg white biological composite material and a preparation method thereof through post-processing heat treatment.
The technical scheme for realizing the aim of the invention is to provide a preparation method of an egg white hydrogel membrane/composite membrane based on one-way nanopore dehydration, which comprises the steps of adding egg white or a mixture solution thereof into a mould from a sampling hole at the top of a container, wherein the mould takes a nanopore filter membrane as a container at the bottom, and sealing the sampling hole by a sealing cover; water molecules in the egg white are dehydrated downwards in a single direction through the nanopore filter membrane, and egg white bioglass is obtained on the upper surface of the nanopore filter membrane; carrying out heat treatment on the egg white bioglass at the temperature of 100-150 ℃ to obtain the egg white hydrogel film/composite film.
The egg white hydrogel film/composite film based on unidirectional nanopore dehydration is obtained by the preparation method.
The technical scheme of the invention also provides a preparation method of the conductive egg white hydrogel membrane/composite membrane based on unidirectional nanopore dehydration, which comprises the steps of adding egg white or a mixture solution thereof into a mold from a sample adding hole in the top of a container, wherein the mold takes a nanopore filter membrane as the bottom of the container, and the sample adding hole is sealed by a sealing cover; dehydrating water molecules in the egg white in a downward one-way through a nano-pore filter membrane to obtain egg white bioglass on the upper surface of the nano-pore filter membrane; carrying out heat treatment on egg white bioglass at the temperature of 100-150 ℃ to obtain an egg white hydrogel film/composite film; and carrying out polymerization reaction on the obtained egg white hydrogel film/composite film and pyrrole in an aqueous solution to obtain the polypyrrole-modified conductive egg white hydrogel film/composite film.
The conductive egg white hydrogel film/composite film based on unidirectional nanopore dehydration is obtained by the preparation method.
The egg white of the invention is poultry egg white, and comprises egg white, duck egg white, goose egg white, pigeon egg white and quail egg white. The mixture comprises protein, polysaccharide, cross-linking agent, plasticizer, medicine and pigment; the concentration of the mixture is 0.1-20% by mass. The heat treatment is to hermetically seal and wrap the egg white bioglass in an isolated manner, and adopts steaming, boiling, high-temperature baking or microwave heating.
The invention relates to a one-way nanopore dehydration mould for preparing an egg white hydrogel membrane/hydrogel composite membrane, which is a container which is processed by high polymers or metal materials and takes a nanopore filter membrane as the bottom, and the top of the container is provided with a sample adding pore and an eraser or a silica gel plug for sealing the pore; before use, deionized water can be used for testing whether the filter membrane at the bottom of the mould is closed and does not leak water. Adding polymer water solution or mixture water solution of other water soluble polymer, cross-linking agent, medicine, etc. via one small hole in the top of the container mold, and sealing the small hole with silica plug after adding sample. The nano-aperture of the filter membrane at the bottom of the mould is a dialysis membrane for intercepting 0.1-1000 kDa molecular weight or a polymer synthetic membrane with filter pores less than or equal to 50 nm. And horizontally placing the mould filled with the egg white or the mixed solution of the egg white and other substances on a mould frame, and keeping the solution in the mould horizontal. The nano-pore filter membrane at the bottom of the mould is exposed in a certain temperature and humidity environment, a high water-absorbing material or a one-way dehydration accelerator can be arranged below the nano-pore filter membrane at the bottom of the container, the one-way dehydration accelerator comprises a flowing air generating device, a constant temperature and humidity box, a negative pressure cavity and a osmotic pressure difference cavity, so that water molecules in a solution can accelerate one-way dehydration through the nano-pore filter membrane, egg white bioglass can be obtained after hours to tens of hours, and the egg white hydrogel membrane/composite membrane and the multifunctional egg white composite material thereof with high flexibility, transparency and mechanical property can be obtained after post-processing heat treatment. The egg white polymer film/hydrogel film and pyrrole are subjected to polymerization reaction in a low-temperature aqueous solution to obtain the polypyrrole-modified conductive polymer film/hydrogel film.
Compared with the prior art, the invention prepares a novel egg white hydrogel film/hydrogel composite film with flexibility, transparency and strong mechanical property and a multifunctional biological composite material thereof in an environment-friendly way by utilizing one-way nano-pore dehydration, and has the beneficial effects that:
1. the hydrogel material provided by the invention is an egg white hydrogel film/composite film which is transparent in a wet state, elastic and strong in tensile property, the tensile strength of the hydrogel material reaches 5.8 MPa, the elongation at break of the hydrogel material is 90-110%, and the swelling rate of the hydrogel material is 60-130%; the egg white hydrogel film/composite film is prepared into the polypyrrole-modified egg white hydrogel conductive film through polymerization reaction.
2. The hydrogel material provided by the invention has the characteristics of simple preparation process, environmental protection, strong mechanical property and biocompatibility; on the prepared egg white hydrogel film, mouse fibroblast L929 has good survival and proliferation, and is suitable for medical biomaterials; meanwhile, the bionic membrane can be widely applied to the fields of bionic materials, drug delivery and sustained release, wearable electronic equipment and the like.
Drawings
Fig. 1 is a schematic structural diagram of a unidirectional nanopore dehydration mold for preparing an egg white hydrogel membrane/composite membrane according to an embodiment of the present invention;
in the figure, 1, a film forming cup; 2. a nanoporous filtration membrane; 3. a fixing ring; 4. a sample application hole; 5. a sampling hole sealing plug;
FIG. 2 is a photographic image of the optical properties of egg white glass and egg white hydrogel films provided in accordance with embodiments of the present invention;
FIG. 3 is a stress versus strain graph of an egg white hydrogel film provided by an embodiment of the present invention;
FIG. 4 is a graph of the thermal performance of an egg white hydrogel film provided by an embodiment of the present invention;
FIG. 5 is a spectrum of infrared spectrum of egg white hydrogel film provided by an embodiment of the invention;
FIG. 6 is an X-ray diffraction pattern of egg white glass and two egg white hydrogel films of embodiments of the present invention;
FIG. 7 is a longitudinal section scanning electron microscope observation image of the egg white hydrogel film provided by the embodiment of the invention;
FIG. 8 is a bar graph of in vitro enzymatic stability of an egg white hydrogel membrane provided by embodiments of the present invention;
FIG. 9 is a graph showing the growth and proliferation of L929 cells on two kinds of egg white hydrogel membranes, according to the present invention;
FIG. 10 shows the growth and proliferation of L929 cells on egg white hydrogel membrane at day 5.
Detailed Description
The technical scheme of the invention is further explained by combining the drawings and the embodiment.
Example 1
This example provides the preparation of a homogeneous egg white solution required for the preparation of each example according to the invention.
Fresh eggs (Lanfei, Shanghai Dahe egg products Co., Ltd., Japan, exclusive Japan, Shanghai, China) are used for collecting egg white liquid through an egg white separator, because egg white concentrated liquid and dilute liquid are difficult to be fully mixed, the egg white can be placed in a high-speed crusher for intermittent high-speed shearing (500-5000 rpm) for 5-10 min, after egg white foam powder is removed, the rest egg white liquid is centrifuged at 5000-15000 rpm for 10-20 min, trace precipitates and a small amount of white floating substances are removed, and the collected supernatant is the egg white homogeneous liquid.
Example 2
The embodiment provides a preparation method of egg white bioglass.
Referring to fig. 1, a schematic structural diagram of a one-way nanopore dehydration mold provided in this embodiment is shown; the processing mould for dehydrating the one-way nano holes mainly comprises a film forming cup 1 with an inner cavity diameter of 30 mm or 80 mm, a nano hole filter membrane 2, a fixing ring 3, a sample adding hole 4 and a sample adding hole rubber plug 5. The circular nanopore filter membrane 2 moistened in water is screwed and fixed through the internal screw threads of the fixing ring 3, and is placed on a horizontal mould shelf, and a small amount of distilled water is added through the sample adding hole 4 by using a liquid transfer device to test whether the fixed filter membrane 2 leaks water or not. If water leaks, the operation is repeated until no water leaks.
A certain volume of homogeneous egg white liquid or a mixture solution of the egg white liquid and other molecules is added into a film forming cup 1 of a mold through a sample adding hole 4 by a pipette, and the sample adding hole 4 is blocked by a silica gel plug 5 after the sample adding is finished. Then, the whole mould shelf is placed under the environment condition of certain temperature and humidity, so that water molecules in the solution in the mould are dehydrated and dried in one way through the nano holes of the filter membrane, and a block with the area of 7.0 cm is formed after hours or tens of hours2Or 50 cm2The egg white bioglass of (a), which is a water-soluble, brittle and transparent egg white bioglass (denoted as EWG).
Example 3
The embodiment provides a preparation method of the egg white hydrogel film/composite film.
The egg white bioglass prepared by the unidirectional nanopore dehydration according to the embodiment 1 and the embodiment 2 needs to be subjected to post-processing heat treatment, and is immediately filled into a polypropylene (PP) sealing bag or wrapped and sealed by a tin-platinum paper after being taken off from a mold, and is subjected to high-temperature steam treatment for 1-4 h; or placing the mixture in a high-temperature oven at 100-140 ℃ for heat treatment for 1-4 h; namely, the egg white hydrogel film (designated as EWRM) is water-insoluble, has good mechanical strength and is soft.
Example 4
This example provides methods for testing the mechanical, structural and in vitro properties of materials provided in accordance with various embodiments of the present invention.
1. Mechanical properties
The egg white liquid is dehydrated through the unidirectional nano holes, and then is subjected to post-processing heat treatment to prepare a hydrogel film, the hydrogel film is cut into a rectangular strip-shaped egg white bioglass with a certain size in a state of being just taken off from a die with the diameter of 30 mm or 80 mm, and then is subjected to post-processing heat treatment to form the hydrogel film or the hydrogel film for testing the mechanical performance. 1 day before the mechanical property measurement, the sample strip is immersed in water at 25 ℃ for fully wetting (24 h), then the water absorption paper is taken out to absorb excessive water, and the wet film mechanical property is immediately measured. During measurement, the room temperature was maintained at about 25 ℃, and the mechanical tensile properties of the film-like samples were measured by an INSTRON 3365 universal material tester. The effective spacing is 10 mm during stretching, and when the stress-strain curve begins to drop sharply, the experiment is stopped, and data are recorded. The mean and standard deviation were calculated.
2. Swelling ratio
Cutting the egg white hydrogel film sample which is just dried to form a film into rectangular strips with the size of 3.0 multiplied by 20 mm, putting the strips into a drying oven, drying the strips at 37 ℃ overnight, and weighing the strips as M0(ii) a Then, immersing the dried sample membrane into a centrifugal tube containing excessive PBS, and placing the centrifugal tube into a biochemical incubator at 37 ℃ for heat preservation so as to enable the centrifugal tube to absorb water and swell; taking out the sample at intervals of 24 hours, wiping off the moisture on the surface of the sample by using filter paper, weighing and recording as Mn(ii) a The swelling ratio of the sample was obtained according to the following formula, and 5 samples were repeated to calculate the mean value and standard deviation (+ -SD).
Swelling ratio (%) = Mn/M0× 100 %
3. Structural analysis
About 100 mg of potassium bromide crystal is mixed with 1 mg of the above sample powder to be tested, and ground. A small amount of mixed sample powder is taken, pressed into slices, and the infrared structural characteristics of the egg white hydrogel film powder sample are detected by a Fourier transform infrared spectrometer (Nicolet 6700, Thermo Fisher, USA). Testing parameters: the scanning times are 16 times, and the resolution is 4 cm-1Spectral range of 2000-1000 cm-1. The film samples were directly measured for fourier transform infrared total reflectance spectra using an ATR accessory. Weighing 5.0 mg of the egg white hydrogel film powder sample, and performing TG, DTG and DSC analysis by using a thermogravimetry/differential thermal spectrometer (SDT 2960, TA company, USA). Setting parameters: the protective gas is nitrogen gas 100 mL/min, the temperature range is 25-800 ℃, and the temperature rise speed is 10 ℃/min. The dried sample of egg white hydrogel film was pulverized and tested with an X-ray diffractometer (X' Pert-Pro MPD) from parnacco, netherlands, Cu target, tube pressure 40 kV, tube flow 50 mA, λ = 1.5406 nm, diffraction angle (2 θ) range 5-50 °, scanning step size 0.02 °/sec, scanning speed 2 °/min.
4. Observation by scanning electron microscope
Firstly, freezing an egg white hydrogel membrane which is in swelling balance in water in liquid nitrogen at-197 ℃ for a plurality of minutes, taking out the membrane, manufacturing a sample fracture surface by using tweezers, and then continuously putting the membrane in a vacuum freeze dryer for freeze-drying; taking a thin slice sample and fixing the sample on an object stage. And (4) spraying gold on the surface for 70S, and observing the apparent morphology of the surface and the cross section or the fracture surface of the film sample under a Hitachi S-4700 cold field emission scanning electron microscope (SEM, Regulus 8230). The accelerating voltage of the scanning electron microscope is 15 kV.
Example 5
The embodiment provides a processing technology of an egg white hydrogel film prepared by unidirectional nanopore dehydration and heat treatment and the influence on mechanical properties.
1. Heat treatment method
The egg white bioglass formed by dehydration of the unidirectional nano-pores is fragile and water-soluble, and can be prepared into an egg white bioglass with certain mechanical properties only by heat treatment. Three heat treatment tests were first performed on this bioglass, and the results of the tests on the mechanical properties of the egg white hydrogel films formed therefrom are shown in table 1. The egg white bioglass is directly placed in boiling water for boiling treatment, the tensile force of the egg white bioglass is gradually reduced along with the boiling time, but the surface of an egg white hydrogel film presents a light fuzzy glass sample, which indicates that the surface hair is caused by the direct boiling of the egg white bioglass because the surface is slightly dissolved in water by the egg white contacted with the water. The egg white glass is directly placed in steam to be steamed for 5 min-90 min, a trend of rising first and then falling can also be found, and the stretching force of 20 min of steaming reaches the peak value (17.10 MPa). The effect of the final 100 ℃ heat-bake treatment group was not as good as the former two groups. These results demonstrate that the manner of heat treatment has a significant effect on both the surface and mechanical properties of the egg white hydrogel film, particularly that boiling in water causes a frosty glass-like appearance on the surface of the egg white hydrogel film.
TABLE 1 Effect of poaching, steaming and 100 deg.C Heat baking of egg white glass on the mechanical Properties of egg white hydrogel films
2. Closed water-isolation cooking treatment
The results shown in fig. 2 are the effect of heat treatment of egg white glass on the optical properties of the egg white hydrogel film. Referring to the attached figure 2, the invention provides an egg white glass (1); a hydrogel film (2) formed after boiling in water; a hydrogel film (3) formed after water isolation and boiling and a hydrogel film (4) formed by sealing and drying for 30 min at 100 ℃. Egg white bioglass (1) as shown in figure 2, prepared by unidirectional nanopore dehydration as in examples 1 and 2, is very transparent like glass, but very brittle and soluble in water. After being boiled in water for 10 min, the egg white hydrogel film (2) which is elastic, water-insoluble and good in mechanical property is formed, but the transparency is influenced, and when egg white glass is filled into a polypropylene sealing bag and boiled in a water-proof way for 10 min, the egg white hydrogel film (3) which is transparent and good in mechanical property is formed. Finally, the egg white hydrogel film (4) can be found to have stronger mechanical properties after being baked for 30 min at 100 ℃, but the color of the egg white hydrogel film can become dark and yellow. Therefore, prior to steaming, boiling or high temperature heat drying, the samples were sealed in a PP bag or wrapped with a tin foil to exclude air.
3. Temperature and time of the baking treatment
According to the analysis of the experimental results in the table 2, the egg white glass sample is pre-treated by boiling in water for 10 min and then dried at five different temperatures of 100 ℃ to 150 ℃ for 10 min. It appears that the tensile properties of the egg white hydrogel film increase with increasing temperature, reaching 13N at 130 c and 16.73N to 150 c. However, the swelling ratio property was inferior as the treatment temperature was higher as observed from the swelling ratio of the egg white hydrogel film, and the swelling ratio was only 62.01% at the maximum temperature and was less than half of that at 100 ℃. Therefore, the baking temperature should not be too high, otherwise the swelling ratio is small, and the egg white hydrogel film becomes brittle and dark in color.
TABLE 2 Effect of egg white glass high temperature heat drying treatment on stretching and swelling Properties of egg white hydrogel films
In the present example, the heat-baking treatment was performed on the egg white glass at four temperatures (100 ℃, 110 ℃, 120 ℃ and 130 ℃), and the results are shown in table 3. The 100 ℃ heat drying treatment is gradually increased along with the increase of the treatment time, the maximum value of the stretching force is increased when the treatment time is 60 min, the maximum value reaches 17.17 MPa, the elongation at break is still 90 percent, and the mechanical property is reduced when the temperature is increased. The heat drying treatment at 110 ℃ has a similar trend, but the treatment temperature is raised by 10 ℃, the treatment temperature continuously rises at a high position at the beginning, the highest tensile force value is reached at 45 min, the highest tensile force value reaches 24.85 MPa, and the elongation at break still reaches 111%. The mechanical properties are also reduced with subsequent longer treatment times. The different times of the latter two temperature treatments are similar, the time for the 120 ℃ treatment to reach the maximum value in 30 min is shorter than the time for the 130 ℃ treatment to reach the maximum value, and the time is only 20 min. From the analysis of the overall factors, the treatment time becomes shorter as the baking temperature increases when the tensile force reaches the peak, and from the analysis of the treatment temperature, the maximum value of the tensile force of the 110 ℃ treatment is the highest and then gradually decreases.
TABLE Effect of Heat-drying time from 3100 ℃ to 130 ℃ on tensile Properties of egg white hydrogel films
4. Stress-strain curve
The results are shown in fig. 3 as stress-strain curves for two egg white hydrogel films. Referring to the attached figure 3, two egg white hydrogel films obtained by respectively heating Egg White Glass (EWG) tinfoil provided by the invention in an oven at 100 ℃ and 110 ℃ for 60 min and 45 min are marked as EWHM-100 and EWHM-110. Stress-strain curves of two egg white hydrogel films (designated as EWRM-100 and EWRM-110 respectively) obtained after Egg White Glass (EWG) is dried for 60 min at 100 ℃ and 45 min at 110 ℃ respectively (figure 3). The egg white hydrogel film obtained after 110 ℃ treatment is significantly higher than the egg white hydrogel film sample treated at 100 ℃ in both strain and stress. The following thermal performance analysis, structural determination, electron microscopy, in vitro anti-proteolysis experiments and biocompatibility experiments were performed using both hydrogel membranes.
Example 5
The embodiment provides the apparent characteristics and the structural characteristics of the egg white hydrogel film prepared by unidirectional nanopore dehydration.
1. Thermal performance
FIG. 4 shows the results of thermal analysis DSC spectra of egg white glass and two egg white hydrogel films. Referring to the attached figure 4, the two egg white hydrogel films EWRM-100 and EWRM-110 are obtained by respectively carrying out heat treatment on the wrapped Egg White Glass (EWG) tinfoil for 60 min and 45 min at 100 ℃ and 110 ℃ in an oven. The thermal decomposition temperature of the water-soluble egg white glass is 314.7 ℃, and the thermal decomposition temperature of the water-soluble egg white glass is obviously increased after the water-soluble egg white glass is thermally treated into an egg white hydrogel film. From the figure it can be clearly observed that the egg white glass formed by unidirectional slow nanopore dehydration starts to absorb heat from 52.9 ℃, and the structure thereof is transformed from random coil to a-helix, and reaches a maximum value at 77.8 ℃. The heat absorption process of the two egg white hydrogel films EWHM-100 and EWHM-110 which are subjected to high-temperature treatment is continuously exothermic, random coil parts in the protein structure are fewer, and part of a-spiral structure directly begins to be converted into b-folding. The egg white glass releases heat from 77.8 ℃, the a-spiral structure begins to be converted to b-folding, the peak of the heat release is reached when the temperature is 203.0 ℃, then the heat absorption is started again, and the glass point transition temperature is 231.9 ℃. The egg white glass starts to degrade at 296.5 ℃, and reaches a degradation peak value when reaching 314.7 ℃. The two egg white hydrogel films have almost no remarkable exothermic and endothermic processes like egg white glass, and the maximum degradation peak values of EWHM-100 and EWHM-110 can be observed to be 318.7 ℃ and 320.1 ℃ respectively. The b-folding structure in the albumen is obviously increased after the egg white glass is subjected to heat treatment, and the thermal stability of the b-folding structure and the albumen is obviously higher than that of the egg white glass.
2. Infrared spectroscopy
FIG. 5 shows the results for infrared spectra of egg white glass and two egg white hydrogel films. Referring to the attached figure 5, the two egg white hydrogel films EWRM-100 and EWRM-110 are obtained by respectively placing the wrapped Egg White Glass (EWG) tinfoil in an oven at 100 ℃ and 110 ℃ and then carrying out heat treatment for 60 min and 45 min. From FIG. 5, it can be observed that the peak of the amide I band of egg white glass EWG is 1633.4 cm-1The peak value of the amide II band is 1536.5 cm-1Belonging to the random coil and partial a-helix structure, after 10 min cook-in water pretreatment, the two bands of this sample (EWHM) were clearly shifted to 1626.2 cm for amide I and amide II, respectively-1And 1527.3 cm-1While the small acromion is 1516.3 cm-1It can also be observed that this is clearly a random coil and a-helix structure shifts to the b-fold configuration; when the egg white plastic obtained by the preheating treatment is continuously subjected to high-temperature heat treatment, no matter the temperature is 100 ℃ and the sealed drying is carried out for 60 min or 110 ℃ and the sealed drying is carried out for 45 min, the amide I belt is still maintained at 1624.3 cm-1Position, and the amide II band not only appeared 1527.3 cm-1While the small shoulder signal appearing at ewmm becomes more intense 1516.3 cm-1Almost the main peak. These results demonstrate that higher temperature heat treatment shifts more of the a-helix structure to the b-fold configuration, as also demonstrated by our previous tensile experimental results.
XRD pattern
The results shown in FIG. 6 are egg white glass andx-ray diffraction pattern of egg white hydrogel film. Referring to the attached figure 6, the two egg white hydrogel films EWRM-100 and EWRM-110 are obtained by respectively carrying out heat treatment on the wrapped Egg White Glass (EWG) tinfoil for 60 min and 45 min at 100 ℃ and 110 ℃ in an oven. From the results of the X-ray diffraction analysis in FIG. 6, it is evident that egg white glass with good water solubility appears to have a diffraction peak at 19.51 deg., which is comparable to the typical amorphous structure diffraction pattern of soluble proteins, i.e., at 5 deg. to 50 deg.2qThe situation that a broad peak exists in the scattering angle range is a little different, the diffraction characteristic of the alpha-helix crystal is contained, and a diffraction peak also appears at the scattering angle of 2q of 7.68 degrees, which indicates that the formation of the alpha-helix structure is facilitated by the dehydration of the one-way nano-pores, and the infrared analysis result also indicates that the situation is different. When the transparent almost water-soluble egg white glass is subjected to high-temperature heat treatment, the diffraction pattern of the transparent almost water-soluble egg white glass is obviously changed. After being subjected to heat treatment at 100 ℃ for 60 min in a closed drying manner, the characteristics of fragility, water solubility and the like of egg white glass are changed, the egg white hydrogel film which is soft, elastic and strong in mechanical property in a wet state is formed, and the main peak appears at 21.04 degrees and small diffraction peaks appear at 29.08 degrees and 39.74 degrees; when the heat treatment temperature is increased to 110 ℃ and the sealed drying is carried out for 45 min, the diffraction peaks of the second hydrogel film EWHM-110 are obviously enhanced and appear at diffraction angles of 30.51 degrees and 30.51 degrees, and a small diffraction peak also appears at 39.74 degrees. The mechanical property, particularly the tensile property, of the alloy is stronger than that of EWHM-100, and the tensile strength and the elongation are respectively higher than 5 MPa and 110 percent elongation. The above results show that egg white is dehydrated through one-way nanopores to enable egg white protein molecules to be orderly arranged to form a crystal structure with partial a-helix characteristics, but the structure is still water-soluble, and only after high-temperature heat treatment, the protein molecular structure is converted from an a-helix structure to a b-folding structure, and finally, the egg white hydrogel film which is a new material with greatly improved mechanical properties appears.
4. Apparent characteristics
In FIG. 7, the magnification of the scanning electron micrograph is 1000. times.10 mm. Two kinds of egg white hydrogel films EWHM-100 and EWHM-110 longitudinal fracture surfaces are observed under a Hitachi S-4700 cold field emission Scanning Electron Microscope (SEM). Referring to the attached figure 7, the invention provides two egg white hydrogel films EWHM-100 (a) and EWHM-110 (b) obtained by respectively carrying out heat treatment on an Egg White Glass (EWG) tinfoil wrapped in an oven at 100 ℃ and 110 ℃ for 60 min and 45 min. The observation results are shown in the figure, no matter the egg white glass is subjected to the closed drying at 100 ℃ for 60 min or the closed drying at 110 ℃ for 45 min, the two egg white hydrogel membranes are similar in the porous network structure, and the pore size is approximately distributed between 1 mm and 10 mm. Therefore, the egg white hydrogel membrane with the network structure is expected to be very suitable for adhesion, survival and proliferation of cells.
Example 6
This example provides a preparation of polypyrrole-modified conductive egg white hydrogel film.
The egg white hydrogel film/composite film prepared by the unidirectional nanopore dehydration and the heat treatment according to the examples 1-3 is subjected to in-situ pyrrole polymerization reaction. The egg white hydrogel film was placed in 10 mL of mixed aqueous solution (2% pyrrole aqueous solution, 0.02M citric acid and 0.02M sodium sulfosalicylate), magnetically stirred for 2 hours and then 10 mL of 1.0M FeCl was added3The solution was subjected to in situ polymerization overnight at-20 ℃. And thoroughly washing the reacted egg white hydrogel membrane sample with water and ethanol for several times in turn, and mainly removing unreacted pyrrole monomers, thereby obtaining the egg white-polypyrrole modified hydrogel membrane of in-situ polymerized polypyrrole. The polypyrrole-modified dark brown hydrogel film/composite film has good conductivity and has potential application prospects in the fields of in-vivo implantation, wearable equipment and the like.
Example 7
This example provides protease resistance of an egg white hydrogel film prepared by one-way nanopore dehydration.
And preparing the five kinds of protease into aqueous solution with the concentration of 500U/mL by using PBS, adding the egg white hydrogel membrane sample, placing the aqueous solution of neutral protease, pepsin and trypsin at 37 ℃, placing the aqueous solution of alkaline protease and papain in a constant temperature shaking table at 50 ℃ for enzymolysis for 12 days by shaking (100-120 rpm), replacing fresh enzyme liquid every day, and finally determining the residual quantity of the sample. Referring to the attached figure 8, the two egg white hydrogel films EWRM-100 and EWRM-110 are obtained by respectively carrying out heat treatment on the wrapped Egg White Glass (EWG) tinfoil for 60 min and 45 min at 100 ℃ and 110 ℃ in an oven. The experimental result shows that the water-soluble egg white glass becomes two egg clear water gel films which are insoluble in water after being sealed and heated for 60 min and 45 min at 100 ℃ and 110 ℃ respectively; in general, the higher the heat treatment temperature, the more resistant the egg white hydrogel film to various proteases. However, the difference of the resistance of the two samples to various enzymes is very large, after the EWHM-110 hydrogel membrane is subjected to enzymolysis for 12 days, the pepsin is hardly degraded, the neutral protease is also rarely degraded, 58% and 72% of trypsin and papain are subjected to enzymolysis respectively, and the alkaline protease is subjected to degradation most quickly, and only 12% of the protease is left. The sequences of the resistance to degradation of the enzyme protein are pepsin, neutral protease, papain, trypsin and alkaline protease.
Example 8
This example provides biocompatibility for an egg white hydrogel film prepared by one-way nanopore dehydration.
The biocompatibility characteristics of the egg white hydrogel are known through the adhesion, the survival and the growth of mouse fibroblasts L929 on two egg white hydrogel films. In the experiment, the CCK-8 method is adopted to respectively measure the cell viability of the two egg white hydrogel films and the control group culture plate every day within 1-5 days. Referring to fig. 9, two egg white hydrogel films EWHM-100 and EWHM-110 are obtained by wrapping an Egg White Glass (EWG) tinfoil provided by an embodiment of the present invention, and then placing the wrapped Egg White Glass (EWG) tinfoil in an oven for heat treatment at 100 ℃ and 110 ℃ for 60 min and 45 min, respectively. Overall, two experimental groups of ABS450The values are very close to those of the control group, and the L929 cells almost have a linear proliferation trend. There was also no significant difference between the two sample groups, and cells were adhered, colonized and grown on the two different temperature heat treated egg white hydrogel films. It can be observed that the cells are normally grown on the control group culture plate without the egg white membrane on the 5 th day, the cells are fusiform, the two ends are slender, the cell nucleus can be observed after amplification, and some cells can not be grown and float in the culture solution to form a sphere due to the higher density. The L929 cells in the two egg white hydrogel films have clear fusiform shapes, and the nucleus and the cytoplasm of the cells are also clearClearly visible, the whole cell is full and has strong stereoscopic impression, and both ends seem to be rooted in the egg white membrane.
Referring to fig. 10, it is a graph showing the growth and proliferation of L929 cells on egg white hydrogel membrane on day 5 according to the present invention; in the figure, (a) the growth state of the L929 cells of the control group without a sample, and (b) and (c) respectively show the growth state of the L929 cells on two kinds of egg white hydrogel films EWHM-100 and EWHM-110, wherein the scale is 100 mm. See that the growth state of the L929 cells on the egg white hydrogel membrane is significantly better than the control group (fig. 10a and b). However, by day 5, it was observed that the absorbance values of the two sets of samples were about 5% to 8% lower than those of the control, and almost no difference was observed from the state of cell engraftment at day 5 (a, b, c in FIG. 10). In fact, the control group, which may have such a difference, has no relationship with the egg white membrane, which has an adsorption effect on the reactive dye during dyeing, and the dye may not be easily washed out during measurement, thus causing a slightly lower absorbance value.
Claims (10)
1. A preparation method of an egg white hydrogel film/composite film based on one-way nanopore dehydration is characterized by comprising the following steps: adding egg white or a mixture solution thereof into a mold from a sample adding hole in the top of a container, wherein the mold takes a nanopore filter membrane as the container at the bottom, and sealing the sample adding hole by using a sealing cover; water molecules in the egg white are dehydrated downwards in a single direction through the nanopore filter membrane, and egg white bioglass is obtained on the upper surface of the nanopore filter membrane; carrying out heat treatment on the egg white bioglass at the temperature of 100-150 ℃ to obtain the egg white hydrogel film/composite film.
2. The preparation method of the egg white hydrogel film/composite film based on unidirectional nanopore dehydration according to claim 1, characterized in that: the egg white is poultry egg white selected from egg white, duck egg white, goose egg white, pigeon egg white, and quail egg white.
3. The preparation method of the egg white hydrogel film/composite film based on unidirectional nanopore dehydration according to claim 1, characterized in that: the mixture comprises protein, polysaccharide, cross-linking agent, plasticizer, medicine and pigment; the concentration of the mixture is 0.1-20% by mass.
4. The preparation method of the egg white hydrogel film/composite film based on unidirectional nanopore dehydration according to claim 1, characterized in that: the heat treatment is to hermetically seal and wrap the egg white bioglass in an isolated manner, and adopts steaming, boiling, high-temperature baking or microwave heating.
5. An egg white hydrogel film/composite film based on unidirectional nanopore dehydration obtained by the preparation method of claim 1.
6. A preparation method of a conductive egg white hydrogel film/composite film based on one-way nanopore dehydration is characterized by comprising the following steps: adding egg white or a mixture solution thereof into a mold from a sample adding hole in the top of a container, wherein the mold takes a nanopore filter membrane as the bottom of the container, and the sample adding hole is sealed by a sealing cover; water molecules in the egg white are dehydrated downwards in a single direction through the nanopore filter membrane, and egg white bioglass is obtained on the upper surface of the nanopore filter membrane; carrying out heat treatment on egg white bioglass at the temperature of 100-150 ℃ to obtain an egg white hydrogel film/composite film; and carrying out polymerization reaction on the obtained egg white hydrogel film/composite film and pyrrole in an aqueous solution to obtain the polypyrrole-modified conductive egg white hydrogel film/composite film.
7. The preparation method of the conductive egg white hydrogel film/composite film based on unidirectional nanopore dehydration according to claim 6, characterized in that: the egg white is poultry egg white selected from egg white, duck egg white, goose egg white, pigeon egg white, and quail egg white.
8. The preparation method of the conductive egg white hydrogel film/composite film based on unidirectional nanopore dehydration according to claim 6, characterized in that: the mixture comprises protein, polysaccharide, enzyme, cross-linking agent, plasticizer, medicine and pigment; the concentration of the mixture is 0.1-20% by mass.
9. The preparation method of the conductive egg white hydrogel film/composite film based on unidirectional nanopore dehydration according to claim 6, characterized in that: the heat treatment is to hermetically seal and wrap the egg white bioglass in an isolated manner, and adopts steaming, boiling, high-temperature baking or microwave heating.
10. The conductive egg white hydrogel film/composite film based on unidirectional nanopore dehydration obtained by the preparation method of claim 6.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210112437.9A CN114702703B (en) | 2022-01-29 | 2022-01-29 | Egg white hydrogel film/composite film based on unidirectional nanometer Kong Tuoshui and preparation method thereof |
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2008154492A (en) * | 2006-12-22 | 2008-07-10 | Japan Tobacco Inc | Egg processed food and method for producing the same |
| KR20130014173A (en) * | 2011-07-29 | 2013-02-07 | 목포대학교산학협력단 | Manufacturing method of bio nano composite hydrogel |
| CN106620830A (en) * | 2016-12-09 | 2017-05-10 | 苏州纳贝通环境科技有限公司 | Medical tissue regeneration-promoting hydrogel dressing and preparation method thereof |
| CN112062994A (en) * | 2020-09-09 | 2020-12-11 | 苏州大学 | Egg white bioplastic, preparation method and application thereof |
| CN112401157A (en) * | 2020-10-27 | 2021-02-26 | 苏州大学 | Fresh-keeping egg white crystal powder, preparation and fresh-keeping storage method |
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Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2008154492A (en) * | 2006-12-22 | 2008-07-10 | Japan Tobacco Inc | Egg processed food and method for producing the same |
| KR20130014173A (en) * | 2011-07-29 | 2013-02-07 | 목포대학교산학협력단 | Manufacturing method of bio nano composite hydrogel |
| CN106620830A (en) * | 2016-12-09 | 2017-05-10 | 苏州纳贝通环境科技有限公司 | Medical tissue regeneration-promoting hydrogel dressing and preparation method thereof |
| CN112062994A (en) * | 2020-09-09 | 2020-12-11 | 苏州大学 | Egg white bioplastic, preparation method and application thereof |
| CN112401157A (en) * | 2020-10-27 | 2021-02-26 | 苏州大学 | Fresh-keeping egg white crystal powder, preparation and fresh-keeping storage method |
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