CN114350590A - Ion response microcapsule and preparation method and application thereof - Google Patents
Ion response microcapsule and preparation method and application thereof Download PDFInfo
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- CN114350590A CN114350590A CN202111601958.2A CN202111601958A CN114350590A CN 114350590 A CN114350590 A CN 114350590A CN 202111601958 A CN202111601958 A CN 202111601958A CN 114350590 A CN114350590 A CN 114350590A
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Abstract
The invention belongs to the technical field of tissue engineering, and particularly relates to an ion response microcapsule, a preparation method and application thereof+、K+、Cl‑Plasma of monovalent ions and Ga2+、Mg2+The microcapsule membrane can keep complete shape and does not swell in the environment of high-valence cations, and the microcapsule membrane can be used in HCO3‑、CO3 2‑、HPO4 2‑、H2PO4‑The microcapsule membrane can swell and even completely dissolve in plasma environment to release the contents such as cells in the microcapsule, and the swelling reaction condition of the microcapsule membrane is mild, such as HCO3‑The ion concentration is 10.0-30.0mM, and the swelling can occur at pH7.2-7.4, without affecting the activity and function of the cells in the microcapsule. The microcapsule core is a tissue-like microenvironment composed of sodium alginate and extracellular matrixes such as collagen, hyaluronic acid and fibronectin, and is beneficial to cell proliferation and activity improvement. The ion response is therefore very smallThe capsule has wide application prospect in the fields of cell culture and cell therapy.
Description
Technical Field
The invention belongs to the technical field of tissue engineering, and particularly relates to an ion response microcapsule as well as a preparation method and application thereof.
Background
The three-dimensional culture of cells can better support cell proliferation, survival and maintenance of intrinsic characteristics by providing a microenvironment which is closer to the in vivo survival conditions. At present, the three-dimensional cultured cells are widely applied to the fields of stem cell clinical medicine, regenerative medicine research and the like, and show good effects in a plurality of in vitro experiments, preclinical animal research and clinical trial research. The three-dimensional cell culture is mainly realized by cell-cell aggregation or adhesion between cells and biological materials, and the following culture methods are mainly adopted: (1) the culture method without biological materials comprises the following steps: the cells are limited in a limited space by the space limitation or the gravity influence, the contact among the cells is increased, the cells are aggregated into balls by adhesion molecules, and the cells are cultured in the form of the cell balls, mainly comprising the culture methods of a hanging drop method, a liquid drop method, a low adsorption culture plate method, a concave pore plate method and the like. These culture methods can control the size of the cell pellet more precisely, but are only suitable for research due to the large workload and high cost. Scale culture and clinical transformation are difficult to realize. (2) Three-dimensional scaffold culture: the three-dimensional culture scaffold obtained by crosslinking hydrophilic polymer, copolymer or monomer macromolecule cultures cells, and promotes cell proliferation and secretion by adding bioactive factors to improve the treatment effect, but is not beneficial to scale preparation and cell culture due to the special structural characteristics of the scaffold. (3) And (3) microcarrier culture: the cell growth micro-carrier comprises a solid micro-carrier and a porous micro-carrier, and cells grow on the surface of the spherical micro-carrier or inside a pore channel to form a cell aggregate. The porous microcarrier not only provides a supporting condition for cell growth to promote cell adhesion, but also can provide high-efficiency mass transfer to improve cell activity, and finally obtains a large number of cells with high activity by effectively simulating a microenvironment for cell in vivo growth. However, the cell harvest requires digestion, and the cell balls are digested into loose single cells or small cell aggregates, so that the three-dimensional cell biological characters cannot be maintained. The above three-dimensional cell culture methods all have significant drawbacks and do not allow for the production of large quantities of three-dimensionally grown cells.
The microcapsule culture is a good three-dimensional cell culture method, cells can form cell balls in the microcapsules due to the aggregation effect of microcapsule membranes, and the good permeability of the microcapsule membranes can provide sufficient nutrients for the cells, so that the three-dimensional cells with good cell activity and functions are obtained. Meanwhile, the microencapsulation technology can also be used for large-scale cell culture, and the cells in the capsule can be prevented from being subjected to the action of mechanical shearing force in the reactor due to the protection effect of the biological microcapsule, so that the damage to the cells is reduced, the oxygen supply efficiency can be improved, and higher cell density can be obtained.
Although a large number of three-dimensional cells with good activity and function can be obtained by microencapsulation culture, the microcapsule membrane is difficult to remove under mild conditions due to high strength, and can be dissolved only under conditions of strong acid or strong alkali (such as pH >8), so that the cell activity cannot be ensured, and the application of the microcapsule technology in three-dimensional cell treatment is limited. The common microcapsules for encapsulating cells at present comprise AC (sodium alginate-chitosan) microcapsules, AP (sodium alginate-alpha-polylysine) microcapsules, AO (sodium alginate-polyornithine) microcapsules and the like, and the microcapsules can obtain three-dimensional cells with good activity, but because the microcapsule membrane has high strength, the microcapsule membrane is difficult to remove under mild conditions, the microcapsule membrane can be dissolved under conditions of strong acid or strong base (such as pH >8), the influence on the cell activity is large, and the cells cannot be harvested after the culture is finished, so that the application in three-dimensional cell treatment is limited. And the alpha-polylysine and the polyornithine are exclusive products of SIGMA company, are expensive and cannot be popularized and applied in a large scale.
At present, no similar microcapsule exists in the research, the capsule membrane can be dissociated under the physiological condition, and cells with good activity are harvested.
Disclosure of Invention
In order to make up for the defects of the prior art, the invention provides the ion response microcapsule, the preparation method and the application thereof, so as to effectively solve the technical problems mentioned in the background technology. The invention aims to develop an ion response microcapsule which has good capsule membrane stability in the cell culture stage, has a protection effect on cells, can obtain 3D cells with good activity, can dissociate the capsule membrane under physiological conditions after the culture is finished, and can harvest the internal cells for cell therapy or other applications.
The above purpose of the invention is realized by the following technical scheme:
an ion response microcapsule comprises a microcapsule membrane and a microcapsule inner core, wherein the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine; the microcapsule core is a tissue-like microenvironment formed by extracellular matrix components and is in a sol or gel state.
Preferably, the extracellular matrix components include sodium alginate, collagen, hyaluronic acid and fibronectin.
A preparation method of an ion response microcapsule comprises the following steps:
(1) adding sodium alginate, collagen, hyaluronic acid and fibronectin into normal saline, stirring at high speed, and dissolving to form a mixed solution;
(2) dripping the mixed solution into 0.1M calcium chloride solution by electrostatic liquid drop method for calcification for 30min to form calcium alginate gel beads;
(3) adding calcium alginate gel beads into a mixed solution of epsilon-polylysine and chitosan, wherein the volume ratio of the calcium alginate gel beads to the mixed solution is 1:10, and carrying out shaking reaction on a universal shaking table for 10min to form a polyelectrolyte microcapsule membrane;
(4) adding the polyelectrolyte microcapsule membrane into an alpha-polylysine solution, wherein the volume ratio of calcium alginate gel beads to the alpha-polylysine solution is 1:10, and carrying out shaking reaction for 10min by using a universal shaking table;
(5) and (4) adding the microcapsule prepared in the step (4) into a 55.0mM sodium citrate solution to liquefy the microcapsule core, and washing with normal saline for 3 times to obtain the liquefied core microcapsule.
Preferably, the sodium alginate is a low-viscosity material, the molecular weight is 80.0-200.0 kD, the G/M ratio is 1.0-3.0, the solution concentration is 1.2-3.0 wt%, and the solution viscosity is 40.0-80.6 mPa & s.
Preferably, the chitosan is water-soluble chitosan, such as chitosan hydrochloride, chitosan quaternary ammonium salt and the like, the molecular weight is 20.0-50.0 kD, the deacetylation degree is 80.0-100.0%, and the solution concentration is 0.1-1.0 wt%.
Preferably, the molecular weight of the epsilon-polylysine is 10.0-30.0 kD, and the solution concentration is 0.01-0.1 wt%.
Preferably, the molecular weight of the alpha-polylysine is 10.0-30.0 kD, and the solution concentration is 0.01-0.1 wt%.
Preferably, the concentration of the collagen, hyaluronic acid and fibronectin extracellular matrix component is 0.1-1.0 wt%.
The invention also provides an application of the ion response microcapsule in cell culture and cell treatment.
Compared with the prior art, the invention has the beneficial effects that:
(1) the technical scheme provided by the invention develops a novel ion response microcapsule with ion response characteristic, and the microcapsule membrane is coated with Na+、K+、Cl-Plasma of monovalent ions and Ga2+、Mg2+Can be kept intact in high valence cation environment without swelling, and can be used in HCO3-、CO3 2-、HPO4 2-、H2PO4-The microcapsule membrane can swell and even completely dissolve in plasma environment to release the contents such as cells in the microcapsule, and the swelling reaction condition of the microcapsule membrane is mild, such as HCO3-The ion concentration is 10.0-30.0mM, the swelling can occur when the pH is 7.2-7.4, and the activity of cells in the microcapsule is not influenced.
(2) The shell of the ion response microcapsule provided by the invention is prepared by crosslinking sodium alginate with chitosan, epsilon-polylysine and alpha-polylysine. The hydrolysis constants pKb of chitosan and epsilon-polylysine are around 7.0 and 7.5, while HCO3 -The pKb of the micromolecule acid radical ion is close to the hydrolysis constant of the chitosan pKb, so the micromolecule acid radical ion is similar to H+Similar binding capacity in HCO3-H binding by competition in the environment of micromolecular acid radical ions+To influence the amino group in the moleculeThe seed formation affects the complex reaction with sodium alginate molecules, and the microcapsule membrane is dissolved. And the pKb of the alpha-polylysine is between 4.0 and 5.0, and H+The binding capacity of the compound is also far stronger than that of HCO3 -Isosmall acid radical ion, HCO3 -The small molecular acid radical ions do not influence the protonation of the sodium alginate and the complex reaction of the sodium alginate and the small molecular acid radical ions. The strength of the microcapsule membrane in different ionic environments can be regulated and controlled by regulating different crosslinking degrees of sodium alginate, chitosan and alpha-polylysine, so that the microcapsule has an ionic response characteristic. Thus by reducing HCO during the cell culture phase3 -The small molecule acid radical ion concentration can keep the microcapsule membrane intact, is favorable for obtaining cell growth, and obtaining three-dimensional cell mass, and HCO in the environment is increased in the treatment stage3 -The small molecular acid radical ion concentration can swell the microcapsule membrane, increase the permeability, release the growth factors and cells in the microcapsule and obtain good treatment effect.
(3) The ion response microcapsule core provided by the invention is a similar organized microenvironment consisting of sodium alginate and extracellular matrixes such as collagen, hyaluronic acid, fibronectin and the like, and is beneficial to cell proliferation and activity improvement.
(4) Sodium alginate, collagen and other positively charged growth factors with negative charge, such as basic fibroblast growth factor bFGF, etc. and has slow releasing effect.
(5) The technical scheme provided by the invention has the advantages of simple production process flow operation, and easy process amplification and industrial production.
Therefore, the ion response microcapsule has wide application prospect in the fields of cell culture and cell therapy.
Drawings
FIG. 1 is a diagram of a variation of microencapsulated cell culture;
FIG. 2 shows the ion response performance of microcapsule for encapsulating cells, wherein a is NaHCO3Result after 2min of ion reaction, b is NaHCO3Ion reaction for 6 h.
Detailed Description
The invention is described in more detail below with reference to specific examples, without limiting the scope of the invention. Unless otherwise specified, the experimental methods adopted by the invention are all conventional methods, and experimental equipment, materials, reagents and the like used in the experimental method can be obtained from commercial sources.
The technical key point of the invention is to select proper microcapsule preparation materials and preparation methods to obtain the microcapsules with the ion response characteristics. The microcapsule ion response performance is related to the property of the preparation material, the preparation method and the preparation parameters, which is also the key technical point of the protection of the invention.
Example 1
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
(1) weighing a certain mass of sodium alginate, collagen, hyaluronic acid and fibronectin, adding the sodium alginate, the collagen, the hyaluronic acid and the fibronectin into 10mL of physiological saline, and dissolving the mixture at a magnetic stirring rotating speed of 800 rpm;
(2) dripping the mixed solution into 100mL of 0.1M calcium chloride solution by electrostatic liquid drop method to calcify for 30min to form calcium alginate gel beads with the volume of about 5 mL;
(3) adding the calcium alginate gel beads into 50mL of chitosan and epsilon-polylysine mixed solution, and reacting for 10min by using a universal shaking table to form a polyelectrolyte microcapsule membrane;
(4) adding the microcapsules into 50mL of alpha-polylysine solution, and oscillating and reacting for 10min by using a universal shaking table;
(5) the microcapsule core is liquefied by adding the microcapsule into 50mL of 55.0mM sodium citrate solution, and the liquefied core microcapsule is obtained by washing 3 times with physiological saline.
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 1
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan and epsilon-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
(1) weighing a certain mass of sodium alginate, collagen, hyaluronic acid and fibronectin, adding the sodium alginate, the collagen, the hyaluronic acid and the fibronectin into 10mL of physiological saline, and dissolving the mixture by magnetic stirring at 800 rpm;
(2) dripping the mixed solution into 100mL of 0.1M calcium chloride solution by electrostatic liquid drop method to calcify for 30min to form calcium alginate gel beads with the volume of about 5 mL;
(3) adding the calcium alginate gel beads into 50mL of chitosan and epsilon-polylysine mixed solution, and carrying out oscillation reaction for 10min by using a universal shaking table to form a polyelectrolyte microcapsule membrane;
(4) the microcapsule core is liquefied by adding the microcapsule into 50mL of 55.0mM sodium citrate solution, and the liquefied core microcapsule is obtained by washing 3 times with physiological saline.
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: concentration 0.05 wt%, molecular weight 12.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: the concentration is 0.5 wt%;
(3) epsilon-polylysine: the concentration was 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 2
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for the preparation
(1) Sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: concentration of 0.05 wt%, molecular weight of 12.0 kD;
(4) α -polylysine: concentration 0.03 wt%, molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: the concentration was 0.03 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 3
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.08 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 4
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan and epsilon-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: concentration 1.2 wt%, viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 5
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: a molecular weight of 18.0 kD;
(5) collagen, hyaluronic acid, fibronectin: the concentrations were each 0.2 wt%.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.03 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 6
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: the molecular weight is 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 7
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.08 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 8
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by calcium alginate hydrogel through electrostatic complexing and crosslinking of alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
(1) weighing a certain amount of sodium alginate, collagen, hyaluronic acid and fibronectin, adding into 10mL of physiological saline, and magnetically stirring at 800rpm to completely dissolve;
(2) dripping the mixed solution into 0.1M calcium chloride solution with the concentration of 100mL by an electrostatic liquid drop method for calcification for 30min to form calcium alginate gel beads, wherein the volume of the gel beads is about 5 mL;
(3) adding calcium alginate gel beads into 50mL of alpha-polylysine solution, and carrying out oscillation reaction for 10min by using a universal shaking table to form a polyelectrolyte microcapsule membrane;
(4) the microcapsule core is liquefied by adding the microcapsule into 50mL of 55.0mM sodium citrate solution, and the liquefied core microcapsule is obtained by washing 3 times with physiological saline.
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) α -polylysine: 0.03 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 9
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by calcium alginate hydrogel through electrostatic complexing and crosslinking of alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method of the ion response microcapsule comprises the following steps:
same comparative example 8
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) α -polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Comparative example 10
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by calcium alginate hydrogel through electrostatic complexing and crosslinking of alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 8
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: a concentration of 1.2% by weight, a viscosity of 80.6 mPas, a molecular weight of 123.0kD, a G/M ratio of 1.57;
(2) α -polylysine: at a concentration of 0.08 wt%, and a molecular weight of 18.0 kD;
(3) collagen, hyaluronic acid, fibronectin: the concentrations were each 0.2 wt%.
4. Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) α -polylysine: 0.08 wt% and a molecular weight of 18.0 kD.
5. The properties of the microcapsule are as follows: the particle size was 230. mu.m.
Example 2
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 11
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan and epsilon-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 12
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties and concentrations of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) sodium alginate: 2.0 wt%, collagen 0.2 wt%, hyaluronic acid 0.2 wt%, fibronectin 0.2 wt%;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.03 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 13
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.08 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 14
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan and epsilon-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 15
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.03 wt%.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: concentration of 1.0 wt%, molecular weight of 28.0kD, degree of deacetylation of 95.0%;
(3) epsilon-polylysine: the concentration is 0.05 wt%, and the molecular weight is 12.0 kD;
(4) α -polylysine: at a concentration of 0.03 wt%, and a molecular weight of 18.0 kD.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 16
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate and 0.2 wt% of collagen; 0.2 wt% of hyaluronic acid; 0.2 wt% fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.05 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 17
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same as example 1
3. The properties of the materials used are as follows:
(1) sodium alginate: a concentration of 2.0% by weight, a viscosity of 137.4 mPas, a molecular weight of 123.0kD, a G/M ratio of 1.57;
(2) chitosan hydrochloride: concentration of 1.0 wt%, molecular weight of 28.0kD, degree of deacetylation of 95.0%;
(3) epsilon-polylysine: the concentration is 0.05 wt%, and the molecular weight is 12.0 kD;
(4) α -polylysine: at a concentration of 0.08 wt%, and a molecular weight of 18.0 kD;
(5) collagen, hyaluronic acid, fibronectin: the concentrations were each 0.2 wt%.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 1.0 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.08 wt%.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 18
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by calcium alginate hydrogel through electrostatic complexing and crosslinking of alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 8
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: a concentration of 2.0% by weight, a viscosity of 137.4 mPas, a molecular weight of 123.0kD, a G/M ratio of 1.57;
(2) α -polylysine: at a concentration of 0.03 wt%, a molecular weight of 18.0 kD;
(3) collagen, hyaluronic acid, fibronectin: the concentrations were each 0.2 wt%.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) α -polylysine: at a concentration of 0.03 wt%, and a molecular weight of 18.0 kD.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 19
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by calcium alginate hydrogel through electrostatic complexing and crosslinking of alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 8
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 137.4 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) α -polylysine: molecular weight 18.0 kD.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) α -polylysine: concentration 0.05 wt%, molecular weight 18.0 kD.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Comparative example 20
1. The present embodiment provides an ion-responsive microcapsule, which comprises the following components:
(1) the microcapsule membrane is formed by calcium alginate hydrogel through electrostatic complexing and crosslinking of alpha-polylysine;
(2) the microcapsule core is a quasi-organized microenvironment composed of sodium alginate and extracellular matrix components such as collagen, hyaluronic acid, fibronectin, etc., and is in sol or gel state.
2. The preparation method comprises the following steps:
same comparative example 8
3. The properties of the materials used for preparation are as follows:
(1) sodium alginate: a concentration of 2.0% by weight, a viscosity of 137.4 mPas, a molecular weight of 123.0kD, a G/M ratio of 1.57;
(2) α -polylysine: at a concentration of 0.08 wt%, and a molecular weight of 18.0 kD;
(3) collagen, hyaluronic acid, fibronectin: the concentrations were each 0.2 wt%.
4. Preparation of solution concentration:
(1) mixing the solution: 2.0 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) α -polylysine: the concentration is 0.08 wt%, and the molecular weight is 18.0 kD.
5. The properties of the microcapsule are as follows: the particle size was 420 μm.
Example 3
Experimental investigation of microcapsule ion response performance
1. Preparing a reagent:
(1) preparing a DMEM/F12 culture solution: adding 1 bottle of DMEM/F12 culture solution solid powder into 1L of distilled water, and stirring for dissolving;
(2) preparing an ion response reaction solution: get the aboveDMEM/F12 medium 100.0mL, quantitative NaHCO was added3Powder, preparing ion response liquid, NaHCO3The final concentrations were 10.0mM, 20.0mM and 30.0 mM.
2. The experimental method comprises the following steps:
(1) and (3) adding 5.0mL of ion response reaction liquid into a 15.0mL centrifuge tube, then adding 100-200 test microcapsules into the centrifuge tube, and slowly oscillating the universal shaking table to ensure that the microcapsules are fully contacted with the ion response reaction liquid.
(2) The microcapsule state was observed under a microscope at regular intervals.
3. The experimental results are as follows:
TABLE 1 Small particle size microcapsules ion response Properties
TABLE 2 ion response Performance of large particle size microcapsules
From the experimental results of Table 1 and Table 2, the microcapsules have ion response performance with NaHCO in the culture solution3The concentration increases and the microcapsule strength decreases, the microcapsule prepared in example 1 is in NaHCO3The form of the culture solution of DMEM/F12 with the concentration of less than 10.0mM is well maintained, no swelling phenomenon occurs, and the culture solution of DMEM/F12 with the concentration of 20.0mM of NaHCO does not contain any swelling phenomenon3The DMEM/F12 culture solution has microcapsule swelling and volume increasing, but the microcapsule is not broken when NaHCO is used3At an increase in concentration to 30.0mM, the microcapsules not only swell but also rupture until the microcapsule membrane is completely dissolved. According to the experimental result of the comparative example, the strength of the microcapsule is closely related to the concentration of the crosslinking reagent, the influence of the chitosan concentration on the strength of the microcapsule is small, the chitosan concentration is increased, and the improvement degree of the strength of the microcapsule is not obvious; the microcapsule strength is closely related to the concentration of the alpha-polylysine, and the microcapsule strength is obviously improved along with the increase of the concentration of the alpha-polylysine. The strength of the microcapsules is also closely related to the particle size,small particle size microcapsules have higher strength. The experimental result also shows that the single alpha-polylysine cross-linked microcapsule has higher strength, and the analysis reason is that the steric hindrance of the alpha-polylysine molecular chain is smaller, and the polyelectrolyte membrane structure formed by the electrostatic complexation reaction with the calcium alginate gel is more compact, so the polyelectrolyte membrane has higher strength. Chitosan has larger steric hindrance, a polyelectrolyte membrane with a loose structure can be formed by electrostatic complexation reaction with calcium alginate gel, and then when the chitosan reacts with alpha-polylysine, most reaction sites are occupied and only a cross-linking reaction can be performed in the loose space structure, the formed polyelectrolyte complex membrane structure is certainly not compact than the alpha-polylysine complex membrane, so the microcapsule strength is not high but the single alpha-polylysine complex membrane has high strength.
Example 4
Experimental study of ion response microcapsule cell culture
1. Preparation of microcapsule for encapsulating cells:
(1) weighing a certain amount of sodium alginate, collagen, hyaluronic acid and fibronectin, adding into 10mL of physiological saline, and dissolving by magnetic stirring (800 rpm);
(2) the mixed solution and the cell density are 106After being mixed with Chinese hamster ovary cells CHO cells, the mixture is dripped into 100mL of 0.1M calcium chloride solution by an electrostatic liquid drop method to be calcified for 30min to form calcium chloride gel beads, and the volume of the gel beads is about 5 mL;
(3) adding the calcium alginate gel beads into 50mL of chitosan and epsilon-polylysine mixed solution, and carrying out oscillation reaction for 10min by using a universal shaking table to form a polyelectrolyte microcapsule membrane;
(4) adding the microcapsules into 50mL of alpha-polylysine solution, and oscillating and reacting for 10min by using a universal shaking table;
(5) the microcapsules are added into 50mL of 55.0mM sodium citrate solution to liquefy the inner cores of the microcapsules, and the microcapsules are washed for 3 times by normal saline to obtain the microcapsules for encapsulating cells.
The properties of the materials used for preparation are as follows:
(1) sodium alginate: viscosity 80.6 mPas, molecular weight 123.0kD, G/M ratio 1.57;
(2) chitosan hydrochloride: the molecular weight is 28.0kD, and the deacetylation degree is 95.0 percent;
(3) epsilon-polylysine: molecular weight 12.0 kD;
(4) α -polylysine: molecular weight 18.0 kD.
Preparation of solution concentration:
(1) mixing the solution: 1.2 wt% of sodium alginate, 0.2 wt% of collagen, 0.2 wt% of hyaluronic acid and 0.2 wt% of fibronectin;
(2) chitosan hydrochloride: 0.5 wt%;
(3) epsilon-polylysine: 0.05 wt%;
(4) α -polylysine: 0.05 wt%. The properties of the microcapsule are as follows: the particle size was 240. mu.m.
2. Culturing the microencapsulated cells:
(1) preparing a DMEM/F12 culture solution: adding 1 bottle of DMEM/F12 culture solution solid powder into 1.0L of distilled water, stirring for dissolving, adding HEPES (hydroxyethyl piperazine ethanesulfonic acid) as a buffer solution, wherein the final concentration of HEPES is 20 mM;
(2) adding microencapsulated cells into the above DMEM/F12 culture solution, and subpackaging into 24-well plates, each well containing 2.0mL of culture solution, incubator at 37 deg.C and 5% CO2Culturing under the condition.
(3) The growth state of the cells is observed under a microscope for periodic sampling.
3. Examination of the ion response performance of the microcapsule:
(1) preparing an ion response reaction solution: 10.0mL of the DMEM/F12 culture solution was added with a fixed amount of NaHCO3Powder, preparing ion response liquid, NaHCO3The final concentration was 30.0 mM.
(2) And adding 5.0mL of ion response reaction liquid into a 15.0mL centrifuge tube, adding 50-100 test microencapsulated cells into the centrifuge tube, and slowly oscillating by using a universal shaking table to ensure that the microcapsules are fully contacted with the ion response liquid.
(3) The microencapsulated cell state was observed under a microscope.
4. The experimental results are as follows:
from FIGS. 1 and 2, it can be seen that the CHO cells were active well in the microcapsules, and that the cells formed larger fine particles by the day 13 of the cultureThe cells are clustered, and the microcapsule has higher strength in buffer solution containing HEPES, the shape is kept intact, and no swelling occurs. The microcapsules were added to a solution containing 30.0mM NaHCO3The microcapsule swells after 2min, the volume of the microcapsule gradually increases, and after 6h, the microcapsule membrane is completely dissolved, and cells are released from the microcapsule into the culture solution.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Claims (9)
1. An ion response microcapsule is characterized by comprising a microcapsule membrane and a microcapsule inner core, wherein the microcapsule membrane is formed by electrostatic complexing and crosslinking of calcium alginate hydrogel through chitosan, epsilon-polylysine and alpha-polylysine; the microcapsule core is a tissue-like microenvironment formed by extracellular matrix components and is in a sol or gel state.
2. An ion-responsive microcapsule according to claim 1 in which the extracellular matrix components comprise sodium alginate, collagen, hyaluronic acid and fibronectin.
3. A preparation method of an ion response microcapsule is characterized by comprising the following steps:
(1) adding sodium alginate, collagen, hyaluronic acid and fibronectin into normal saline, stirring at high speed, and dissolving to form a mixed solution;
(2) dripping the mixed solution into 0.1M calcium chloride solution by electrostatic liquid drop method for calcification for 30min to form calcium alginate gel beads;
(3) adding calcium alginate gel beads into a mixed solution of epsilon-polylysine and chitosan, wherein the volume ratio of the calcium alginate gel beads to the mixed solution is 1:10, and carrying out shaking reaction on a universal shaking table for 10min to form a polyelectrolyte microcapsule membrane;
(4) adding the polyelectrolyte microcapsule membrane into an alpha-polylysine solution, wherein the volume ratio of calcium alginate gel beads to the alpha-polylysine solution is 1:10, and carrying out shaking reaction for 10min by using a universal shaking table;
(5) and (4) adding the microcapsule prepared in the step (4) into a 55.0mM sodium citrate solution to liquefy the microcapsule core, and washing with normal saline for 3 times to obtain the liquefied core microcapsule.
4. The method for preparing an ion-responsive microcapsule according to claim 3, wherein the sodium alginate in step (1) is a low viscosity material having a molecular weight of 80.0 to 200.0kD, a G/M ratio of 1.0 to 3.0, a solution concentration of 1.2 to 3.0 wt%, and a solution viscosity of 40.0 to 80.6 mPas.
5. The method of claim 3, wherein the chitosan of step (3) is water-soluble chitosan, has a molecular weight of 20.0 to 50.0kD, a degree of deacetylation of 80.0 to 100.0%, and a solution concentration of 0.1 to 1.0 wt%.
6. The method for preparing an ion-responsive microcapsule according to claim 3, wherein the molecular weight of epsilon-polylysine in step (3) is 10.0 to 30.0kD, and the solution concentration is 0.01 to 0.1 wt%.
7. The method of claim 3, wherein the alpha-polylysine of step (4) has a molecular weight of 10.0 to 30.0kD and a solution concentration of 0.01 to 0.1 wt%.
8. The method of claim 3, wherein the concentration of the collagen, hyaluronic acid, fibronectin extracellular matrix component in step (1) is 0.1-1.0 wt%.
9. The application of the ion response microcapsule is characterized in that the ion response microcapsule is applied to cell culture and cell therapy.
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| CN1616657A (en) * | 2003-11-10 | 2005-05-18 | 中国科学院大连化学物理研究所 | Method for preparing fixed yeast cell micro capsule |
| CN102051354A (en) * | 2009-11-04 | 2011-05-11 | 中国科学院大连化学物理研究所 | Microcapsule encapsulated with filamentous scaffolds as well as preparation method and applications thereof |
| CN102101036A (en) * | 2009-12-18 | 2011-06-22 | 中国科学院大连化学物理研究所 | Alginate-epsilon-polylysine microcapsules and preparation and application thereof |
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| CN1616657A (en) * | 2003-11-10 | 2005-05-18 | 中国科学院大连化学物理研究所 | Method for preparing fixed yeast cell micro capsule |
| CN102051354A (en) * | 2009-11-04 | 2011-05-11 | 中国科学院大连化学物理研究所 | Microcapsule encapsulated with filamentous scaffolds as well as preparation method and applications thereof |
| CN102101036A (en) * | 2009-12-18 | 2011-06-22 | 中国科学院大连化学物理研究所 | Alginate-epsilon-polylysine microcapsules and preparation and application thereof |
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