CN101773813B - Method for preparing hollow micro-capsules on basis of specific interaction between concanavalin and glycogen - Google Patents
Method for preparing hollow micro-capsules on basis of specific interaction between concanavalin and glycogen Download PDFInfo
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- CN101773813B CN101773813B CN2010101229240A CN201010122924A CN101773813B CN 101773813 B CN101773813 B CN 101773813B CN 2010101229240 A CN2010101229240 A CN 2010101229240A CN 201010122924 A CN201010122924 A CN 201010122924A CN 101773813 B CN101773813 B CN 101773813B
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Abstract
The invention discloses a method for preparing hollow micro-capsules on the basis of the specific interaction between concanavalin and glycogen. The method based on the specific recognition of the glucose components in concanavalin and glycogen comprises the following steps: forming a multi-layer membrane on the surfaces of carbonate colloidal particles by using the layer-by-layer assembly method; and further removing the carbonate colloidal particles by using the EDTA (ethylene diamine tetraacetic acid) disodium complex method to obtain the hollow micro-capsules. The micro-capsule prepared by the invention is capable of producing the specific response to specific monosaccharide and polysaccharide substances under the condition of the physiological pH value and has the advantages of low material cost, good biocompatibility, good biodegradability and the like. Therefore, the invention has good application prospect in the field of biomedical science.
Description
Technical field
The present invention relates to a kind of method for preparing hollow microcapsule, especially prepare the method for hollow microcapsule based on concanavalin and the special interaction of glycogen.
Background technology
Microcapsules are by film forming matter the space in the capsule and the capsule external space to be kept apart formed specific three dimensional structure, and its shape is based on sphere, also can be oval, square or rectangular, polygonal and other is various irregularly shaped.Tradition microcapsules size is generally micron to the millimeter level, wall thickness be sub-micron to the hundreds of micron, cyst wall is made up of natural or synthetic macromolecular material usually, also but inorganic compound.Microcapsules all have crucial application in fields such as food, medicine, cosmetics, bio-medical.
The preparation method of microcapsules has a lot.According to the principle that cyst wall forms, the traditional preparation process technology of microcapsules cardinal principle can be divided three classes: the physical method of the chemical method of utilization reaction generation cyst wall, the physico-chemical process that utilizes the formation cyst wall that is separated and utilization machinery or other physical action formation cyst wall.Recent study personnel have developed many new microcapsule preparation methods again, as template assembling, matrix polymerization, surface grafting polymerization, dispersin polymerization etc., this wherein based on the microcapsules that utilize layer-by-layer preparation on the colloidal particles surface owing to have that the material selection is various, structure is accurately controlled, easily give advantage such as capsule intelligent response and obtained broad research.Environment pH value, ion concentration, temperature, laser, magnetic field etc. have intelligent response to external world to utilize the capsule of this method preparation, but realize that the required stimulus of above response is difficult to realize mostly in human body, this has brought challenge in the application prospect in bio-medical field for these intelligent capsule systems, and preparation has the intelligent microcapsules of response then can address this problem preferably to carbohydrate especially glucose.Existing minority mainly contains two classes about the report of the microcapsules of glucose responding.One class is based on the interaction of phenyl boric acid group and carbohydrate, but the glucose responding of this type of capsule mostly needs in pH＞9 o'clock could to show apparent in viewly, yet is difficult to find such pH environment in the human body.Thereby another kind of is to utilize glucose oxidase oxidizing glucose molecule to cause the pH value of local microenvironment to change to realize the glucose responding of capsule with the principle that influences the capsule permeability, but the costing an arm and a leg and inactivation very easily of its used raw material-enzyme.Utilize the microcapsules of the special interaction of agglutinin (as concanavalin)-polysaccharide (as glycogen) preparation then to have the biocompatibility that under the physiological pH value, has glucose responding, raw material with degradability is good, the cheap advantage such as stablize of cost of material, can remedy the deficiency of reporting the capsule system preferably, the gained capsule also has response for the polysaccharide molecule based on glucose unit simultaneously, and above advantage makes these microcapsules in the bio-medical field good prospects for application be arranged.
Summary of the invention
The purpose of this invention is to provide and a kind ofly prepare the method for hollow microcapsule, to realize capsule stimuli responsive to glucose and LMD under the physiological pH value based on concanavalin and the special interaction of glycogen.
The method for preparing hollow microcapsule based on concanavalin and the special interaction of glycogen of the present invention, be to adopt concanavalin that will contain the glucose binding site and the glycogen that constitutes by glucose unit to carry out layer assembly on the colloidal particles surface, obtain behind the polymer multi-layer film colloidal particles to be removed, be hollow microcapsule based on concanavalin and the special interaction preparation of glycogen thereby obtain cyst wall.Specifically may further comprise the steps:
1) be that 7.1-7.8, concentration are to add CaCl in the tris-HCI buffer of 0.025-0.05mol/L toward the pH value
2With MnCl
2, make and contain 1mmol/L Mn in the buffer solution
2+With 1mmol/L Ca
2+, with this buffer solution be solvent respectively compound concentration be the concanavalin solution of 0.2mg/ml and the glycogen solution that concentration is 1mg/ml;
2) be that 4mg/ml, pH value are 7 and contain in the polyethyleneimine: amine aqueous solution of 0.5M NaCl toward concentration, add 5-25mg carbonate colloidal particle, the vibration absorption centrifugal washing in back by every milliliter of polyethyleneimine: amine aqueous solution;
3) with step 2) the gained particle is scattered in the concanavalin solution of step 1), vibration absorption and be the tris-HCI buffer centrifuge washing of 0.025-0.05mol/L with concentration obtains the particle that surface-assembled has one deck concanavalin molecule;
4) step 3) gained particle is scattered in the glycogen solution of step 1), vibration absorption and be the tris-HCI buffer centrifuge washing of 0.025-0.05mol/L with concentration obtains further assembling on concanavalin molecular layer surface the particle of one deck glycogen molecule;
5) at step 4) gained particle surface repeating step 3) and 4), assembling concanavalin layer and glycogen layer are until the required assembling number of plies, the gained particle is scattered in the disodium ethylene diamine tetra-acetic acid solution that concentration is 0.1mol/L, colloidal particle is removed in vibration, obtains cyst wall and is the hollow microcapsule based on concanavalin and the special interaction structure of glycogen.
Among the present invention, said carbonate is calcium carbonate or manganese carbonate.
Among the present invention, the said assembling number of plies is generally the 3-15 layer.
The beneficial effect of the inventive method is:
The present invention utilizes the special interaction between the glucose unit contained in glucose binding site on the concanavalin and the glycogen molecule to prepare hollow microcapsule.The gained capsule has under the physiological pH value glucose and glucan has the biocompatibility of stimulating responsive, raw material and degradability is good, the cheap advantage such as stable of cost of material, in the bio-medical field good prospects for application is arranged.
Description of drawings
Fig. 1 a) has assembled the optical microscope photograph of 12 layers microcapsules under pH 7.8 conditions, b) has assembled 12 layers the dried stereoscan photograph of microcapsules under pH 7.8 conditions.
Assembled the optical microscope photograph of 12 layers microcapsules under Fig. 2 pH 7.1 conditions.
Assembled the fluorescence microscope photo of 12 layers microcapsules under Fig. 3 pH 7.8 conditions.
Assembled the stereoscan photograph of 4 layers microcapsules under Fig. 4 pH 7.8 conditions.
Assembled the stereoscan photograph of 6 layers microcapsules under Fig. 5 pH 7.8 conditions.
Fig. 6 is the fluorescence microscope photo before having assembled 12 layers microcapsules under pH 7.8 conditions and mixing with glucose solution a); B) assemble 12 layers microcapsules under pH 7.8 conditions and mixed fluorescence microscope photo after 20 seconds with glucose solution.
Assemble 12 layers microcapsules under Fig. 7 pH 7.8 conditions and mixed fluorescence microscope photo after 10 minutes with the dextran solution of molecular weight 20,000.
Assemble 12 layers microcapsules under Fig. 8 pH 7.8 conditions and mixed fluorescence microscope photo after 12 minutes with the dextran solution of molecular weight 40,000.
The specific embodiment
Further specify the present invention below in conjunction with example, but these examples are not used for limiting the present invention.
Example 1
1) in trishydroxymethylaminomethane-hydrochloric acid (Tris-HCl) buffer solution that the pH value is 7.8, concentration is 0.05mol/L, adds CaCl
2With MnCl
2, make and contain 1mmol/L Mn in the buffer solution
2+With 1mmol/L Ca
2+, with this buffer solution be solvent respectively compound concentration be the concanavalin solution of 0.2mg/ml and the glycogen solution that concentration is 1mg/ml;
2) be that 4mg/ml, pH value are 7 and contain in the polyethyleneimine: amine aqueous solution of 0.5M NaCl toward 4ml concentration, add 40mg calcium carbonate colloidal particle, the vibration absorption centrifugal washing in back;
3) with step 2) the gained particle is scattered in the concanavalin solution of step 1), and vibration absorption and be the tris-HCI buffer centrifuge washing of 0.025mol/L with concentration obtains the particle that surface-assembled has one deck concanavalin molecule;
4) step 3) gained particle is scattered in the glycogen solution of step 1), vibration absorption and be the tris-HCI buffer centrifuge washing of 0.025mol/L with concentration obtains further assembling on concanavalin molecular layer surface the particle of one deck glycogen molecule;
5) at step 4) gained particle surface repeating step 3) and 4), assembling concanavalin layer and glycogen layer are until being assembled to 12 layers, the gained particle is scattered in the disodium ethylene diamine tetra-acetic acid solution that concentration is 0.1mol/L, colloidal particle is removed in vibration, obtains the hollow microcapsule that cyst wall makes up for the special interaction based on concanavalin-glycogen.Optical microscope photograph under the gained microcapsules hygrometric state is seen Fig. 1 a, and the stereoscan photograph under the dry state is seen Fig. 1 b.
Example 2
Step is with example 1, but the middle in steps pH of buffer condition of institute replaces 7.8 with 7.1.Optical microscope photograph under the gained microcapsules hygrometric state is seen Fig. 2.
Example 3
Step is with example 1, but the common concanavalin of concanavalin replacement of in step 3), using fluorescein isocyanates (FITC) mark.Fluorescence microscope photo under the gained microcapsules hygrometric state is seen Fig. 3.
Example 4
Step is with example 1, but step 2) middle with manganese carbonate colloidal particle replacement calcium carbonate colloidal particle.
Example 5
Step is with example 1, but step 2) in 4ml polyethyleneimine: amine aqueous solution, add the 20mg calcium carbonate particle.
Example 6
Step is with example 1, but the assembling number of plies of capsule-wall is 4 layers in the step 5).The dried stereoscan photograph of gained microcapsules is seen Fig. 4.
Example 7
Step is with example 1, but the assembling number of plies of capsule-wall is 6 layers in the step 5).The dried stereoscan photograph of gained microcapsules is seen Fig. 5.
Example 8
(solvent is that concentration is 0.025mol/L, pH 7.4 and contains 1mmol/L Mn with example 3 gained microcapsule suspensions and equal-volume 20mg/ml glucose solution
2+, Ca
2+The Tris-HCl buffer solution) mix, fluorescence microscope monitoring microcapsules pattern down changes, the capsule pattern is shown in Fig. 6 a before mixing, mix after 20 seconds the capsule pattern shown in Fig. 6 b, as seen capsule stimulates glucose response has taken place, and is showing as the violent contraction on the size and the crumple of cyst wall on the pattern.
Example 9
(solvent is that concentration is 0.025mol/L, pH 7.4 and contains 1mmol/L Mn with the dextran solution of example 3 gained microcapsule suspensions and equal-volume 100mg/ml molecular weight 20,000
2+, Ca
2+The Tris-HCl buffer solution) mix, fluorescence microscope monitoring microcapsules pattern down changes, mix after 10 minutes the capsule pattern as shown in Figure 7, visible capsule is that 20,000 glucan stimulates response has taken place to molecular weight, shows as size and taken place obviously to reduce and the cyst wall crumple on pattern.
Example 10
(solvent is that concentration is 0.025mol/L, pH 7.4 and contains 1mmol/L Mn with the dextran solution of example 3 gained microcapsule suspensions and 100mg/ml molecular weight 40,000
2+, Ca
2+The Tris-HCl buffer solution) mixed in 1: 2 by volume, fluorescence microscope monitoring microcapsules pattern down changes, mix after 12 minutes the capsule pattern as shown in Figure 8, as seen capsule is that 40,000 glucan stimulates response has taken place to molecular weight, shows as size on pattern reducing and the cyst wall crumple to a certain degree taken place.
Claims (2)
1. one kind prepares the method for hollow microcapsule based on concanavalin and the special interaction of glycogen, and this method may further comprise the steps:
1) be that 7.1-7.8, concentration are to add CaCl in the tris-HCI buffer of 0.025-0.05mol/L toward the pH value
2With MnCl
2, make and contain 1mmol/L Mn in the buffer solution
2+With 1mmol/L Ca
2+, with this buffer solution be solvent respectively compound concentration be the concanavalin solution of 0.2mg/ml and the glycogen solution that concentration is 1mg/ml;
2) be that 4mg/ml, pH value are 7 and contain in the polyethyleneimine: amine aqueous solution of 0.5M NaCl toward concentration, add 5-25mg carbonate colloidal particle, the vibration absorption centrifugal washing in back by every milliliter of polyethyleneimine: amine aqueous solution;
3) with step 2) the gained particle is scattered in the concanavalin solution of step 1), vibration absorption and be the tris-HCI buffer centrifuge washing of 0.025-0.05mol/L with concentration obtains the particle that surface-assembled has one deck concanavalin molecule;
4) step 3) gained particle is scattered in the glycogen solution of step 1), vibration absorption and be the tris-HCI buffer centrifuge washing of 0.025-0.05mol/L with concentration obtains further assembling on concanavalin molecular layer surface the particle of one deck glycogen molecule;
5) at step 4) gained particle surface repeating step 3) and 4), assembling concanavalin layer and glycogen layer are until the required assembling number of plies, the gained particle is scattered in the disodium ethylene diamine tetra-acetic acid solution that concentration is 0.1mol/L, colloidal particle is removed in vibration, obtains cyst wall and is the hollow microcapsule based on concanavalin and the special interaction structure of glycogen;
Above-mentioned carbonate is calcium carbonate or manganese carbonate.
2. according to claim 1ly prepare the method for hollow microcapsule, it is characterized in that the said assembling number of plies is the 3-15 layer based on concanavalin and the special interaction of glycogen.
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