CN102824332A - Preparation method of antimicrobial peptide slow-release microcapsules - Google Patents
Preparation method of antimicrobial peptide slow-release microcapsules Download PDFInfo
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Abstract
The invention discloses a preparation method of antimicrobial peptide slow-release microcapsules and relates to a preparation method of microcapsules. The invention solves the problems that a preparation mode of the existing chitosan/sodium alginate microcapsules is only suitable for laboratories or the small-scale production, the industrial large-scale production is difficult to form and the existing antimicrobial peptide is short of an effectively slow-release envelope. The preparation method comprises the following steps of: 1, uniformly mixing aqueous solution of sodium alginate and chitosan acid solution by a high-speed homogeneous machine to form slow-release particles; and 2, adding the antimicrobial peptide and the slow-release particles into a medical mixer together to carry out embedding to obtain antimicrobial peptide particles and then carrying out beta-cyclodextrin embedding on the antimicrobial peptide particles to obtain the antimicrobial peptide slow-release microcapsules. The preparation method can be used for producing an antimicrobial peptide medicament and has a very large development space in the field of medicine.
Description
Technical field
The present invention relates to a kind of method for preparing of microcapsule.
Background technology
Microcapsule technology is called microcapsule embedded technology again, or the microcapsule granulating technique, is solid, liquids and gases material are embedded in the same microencapsulation, becomes a kind of technology of solid particle.Usually call " wall material " or " coating " constituting the microcapsule shell material, call " the capsule heart " perhaps " core " etc. wrapping in the microcapsule inner material.Core can be solid, liquid or gas.
Usually, roughly be divided three classes Microencapsulation Method chemical method, physics method and physical-chemical process.Wherein chemical method comprises interfacial polymerization, situ aggregation method, orifice-freezing method, envelope method etc.; The physics method comprises a pot bag method, air suspension, nebulization, squeezing and pressing method etc.; Physical-chemical process comprises complex coacervation, single coacervation, oil-phase separating method, complex phase emulsion method, powder bed method etc.
The most commonly used at present is to be the microcapsule of embedded material preparation with chitosan, sodium alginate.Chitosan is dissolved in acetic acid solution, adds a certain amount of calcium chloride, and regulate the pH value of chitosan solution with NaOH solution; Other takes by weighing in the sodium alginate adding distilled water, and it is fully dissolved, and adds core then, stirs with Glass rod.Aqueous solution with core and sodium alginate slowly splashes in chitosan-calcium chloride solution at last, and the magnetic stirring apparatus stirring at low speed is collected microcapsule, drying for standby.It is even that this method need slowly dropwise splash into the aqueous solution of core and sodium alginate the peplos rate and the parcel that could guarantee core in the microcapsule in chitosan-calcium chloride solution; But this preparation method only is suitable for laboratory or small-scale production, is difficult to form industrialization large-scale production.
Antibacterial peptide has broad-spectrum bactericidal action, and comprising especially has tangible inhibitory or killing effect to Resistant strain to gram positive bacteria and gram negative bacteria.For example, the sarcotoxin I that is separated to from sarcophagid just is enough to kill shigella, bacillus megaterium, micrococcus luteus, corynebacterium, Proteus sonnei and streptomycin is produced chemical sproof escherichia coli when concentration is 0.3 μ g/mL.Moriein (silk silkworm antibacterial peptide) all has the obvious suppression effect to 13 kinds of antibacterials such as e. coli jm109, bacillus subtilis, staphylococcus aureuses, and their minimum lethal concentration all is lower than 0.9 μ M.Sarcophagid antibacterial peptide SarCotoxinIA can suppress the growth of some harmful intestinal; Especially the most obvious to the colon bacillus 0157 action effect that often causes alimentary toxicosis; And it is to intravital symbiotic bacteria of normal person and not damaged; Therefore can be used to change the composition of intestinal bacteria group, thus enhances human body body constitution.Antibacterial peptide mainly is through changing the permeability of film to the antibacterial lethal effect; Make thalline dead outward because of the cell Dissolve things inside rushes down; Perhaps antibacterial peptide passes film and combines with DNA, RNA; Cause bacterial death fast, this sterilization mode is difficult for making antibacterial to produce chemical sproof mutant strain, is expected to be developed as antibacterials of new generation.But existing antibacterial peptide also lacks a kind of release-controlled coated effectively.
Summary of the invention
The present invention will solve existing chitosan/sodium alginate micro gel capsule preparation method and only be suitable for laboratory or small-scale production, is difficult to form industrialization large-scale production and existing antibacterial peptide also lacks release-controlled coated effectively problem.
The antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with sodium alginate aqueous solution and chitosan acid solution with high speed homogenizer mixing, form slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are together added carry out embedding in the medical mixer, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule is carried out the beta-schardinger dextrin-embedding, promptly obtain the antibacterial peptide slow-release microcapsule.
Method for preparing of the present invention is not added calcium chloride in preparing chitosan acid solution process, can not cause toxic residue, and avoided calcium chloride in the preparation process to by the influence of embedding antibacterial peptide, its activity is not lost.A large amount of primary amino radicals is arranged on the chitosan molecule chain, a large amount of carboxyls is arranged on the strand of sodium alginate, can attract to form polyelectrolyte film by positive and negative charge.
Method for preparing of the present invention has been revised the prescription and the technology of slow-release microcapsule, and adopts high speed homogenizer and medical mixer to carry out embedding, be fit to large-scale industrialized production, and the antibacterial peptide embedding is even.Carry out the beta-schardinger dextrin-embedding and not only can the antibacterial peptide envelop rate be brought up to more than 90%, and the slow-release microcapsule of the present invention's preparation can more effectively carry out the stable sustained-release of antibacterial peptide.
The antibacterial peptide slow-release microcapsule mode of appearance rounding of method for preparing preparation of the present invention, even, the dried particle diameter 1.2～3mm of antibacterial peptide slow-release microcapsule.The antibacterial peptide diameter of particle is about 0.35～0.65mm; Dried antibacterial peptide diameter of particle is 0.1～0.2mm.
The antibacterial peptide slow-release microcapsule of method for preparing preparation of the present invention does not develop immunity to drugs, and the antibacterial peptide envelop rate is up to more than 90%, but large-scale industrialized production.
The specific embodiment
The specific embodiment one: this embodiment antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with sodium alginate aqueous solution and chitosan acid solution with high speed homogenizer mixing, form slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are together added carry out embedding in the medical mixer, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule is carried out the beta-schardinger dextrin-embedding, promptly obtain the antibacterial peptide slow-release microcapsule.
The specific embodiment two: the difference of this embodiment and the specific embodiment one is: in the step 1 in the chitosan acid solution chitosan mass concentration be that 0.25% (W/V)～1.5% (W/V), deacetylation are 82%～85%, chitosan is 3.3 * 10 by molecular weight
5～4.6 * 10
5High molecular weight chitosan and molecular weight be 0.72 * 10
5～1.19 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.3～0.8 in the chitosan acid solution.Other step and parameter are identical with embodiment one.
The chitosan acid solutions needs >=0.25% (W/V) in this embodiment, and the chitosan acid solutions raises, the controlled enhancing of microcapsule.
Glucose adds as plasticizer in this embodiment, can improve slow-release microcapsule intensity more than 5%.
Low-molecular weight chitoglycan prepared microcapsule granule strength is high, and the microcapsule granule thickness of its preparation is higher than high molecular weight chitosan, and anti-swelliong power is stronger, thereby mechanical strength is stronger; But the chitosan molecule amount is low excessively, and its toughness and elasticity are relatively poor relatively; And the degree of depth of chitosan entering sodium alginate is the same, and the chitosan that molecular weight is big more and the binding site number of sodium alginate are just many more, and the ability of between sodium alginate, building bridge is just strong more, and the intensity that forms microcapsule is just high more, and controlled release is just strong relatively more.Ratio among the present invention between reasonable proportioning low-molecular weight chitoglycan and the high molecular weight chitosan all is significantly improved the strength and toughness of microcapsule granule and elasticity.The intensity of antibacterial peptide slow-release microcapsule of the present invention reaches more than 74.95, and toughness and elasticity reach more than 58.42; Antibacterial peptide and slow-releasing granules carry out the envelop rate 70.26 of embedding.And the release of antibacterial peptide in the microcapsule granule of the mixing Preparation of Chitosan that contains high molecular weight chitosan and low-molecular weight chitoglycan is slow, and effectively slow release reaches the release medicine that continues steady in a long-term.Experiment shows that antibacterial peptide is slower than the microcapsule granule of simple high molecular weight chitosan preparation at the rate of release that the present invention mixes in the microcapsule granule of Preparation of Chitosan; 12 hours antibacterial peptide burst sizes of antibacterial peptide slow-release microcapsule of the present invention are 70%, are 85% and add 12 hours antibacterial peptide burst sizes of simple polymer chitosan.
The specific embodiment three: this embodiment with the specific embodiment one or two difference is: the mass concentration of sodium alginate is 1.5%～2.0% in the step 1 sodium alginate aqueous solution.Other step and parameter are identical with embodiment one or two.
The specific embodiment four: this embodiment with the specific embodiment one, two or three difference is: regulating pH value in step 1 sodium alginate aqueous solution and the chitosan acid solution blending process is 5.2～5.7.Other step and parameter are identical with embodiment one, two or three.
It is the highest to regulate pH value in this embodiment method for preparing step 1 sodium alginate aqueous solution and the chitosan acid solution blending process and be the antibacterial peptide slow-release microcapsule intensity that 5.5 embodiments prepare.And the high more slow-release microcapsule of pH value of reaction system discharges proteic speed and accelerates thereupon in the experiment proof preparation process, and rate of release increases with the reduction of protein molecular weight.
Show in in-vitro simulated (gastric juice, intestinal juice) test and zoology test: this microcapsule can adhere to crack-free more than 48 hours on the mouse small intestine surface, thereby guarantees that medicine can fully be absorbed by small intestinal.Under the outer stripping experiment condition of microencapsulated insecticide object; Discharge 85% in 12 hours inherent simulated intestinal fluids; And in simulated gastric fluid, only discharge 15%, thus guaranteeing entrapped drug not by stomach acids destroy, this microcapsule is after drying under the preparation process condition; Its original strength character and controlled release properties can be kept, solid preparation can be used for.
The specific embodiment five: this embodiment with the difference of one of specific embodiment one to four is: sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution.Other step and parameter are identical with one of embodiment one to four.
The specific embodiment six: this embodiment with the difference of one of specific embodiment one to five is: the particle diameter of the slow-releasing granules that step 1 forms is 0.35～0.65mm, and the particle diameter after the oven dry is 0.1～0.2mm.Other step and parameter are identical with one of embodiment one to five.
The specific embodiment seven: this embodiment with the difference of one of specific embodiment one to six is: the mass ratio of antibacterial peptide and slow-releasing granules is (3.3～8.6) in the step 2: (2～4); Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD; The relative molecular weight of HMW antibacterial peptide is 40～50KD.Other step and parameter are identical with one of embodiment one to six.
Antibacterial peptide is selected low-molecular-weight and two kinds of antibacterial peptides of HMW for use in this embodiment, because different its rate of release of molecular weight are different, antibiotic drug effect is therefore continuity also, reaches to continue steadily to discharge effectively.
The acquisition of HMW antibacterial peptide: with the fly larvae of deepfreeze,, filter impurity such as maggot fur, extract the HMW antibacterial peptide with salting out method then with organizing pulverizer to pulverize.
The acquisition of low-molecular-weight antibacterial peptide: with bromelain or papain (4000U/g) decomposing macromolecular amount antibacterial peptide, HMW antibacterial peptide concentration is that the consumption of 60～100g/L, bromelain or papain is that 1.5% (mass fraction), pH value are 7, hydrolysis temperature is that 45 ℃, enzymolysis time are 80～200 minutes.
The specific embodiment eight: this embodiment with the difference of one of specific embodiment one to seven is: the beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with extruding granulator or rotation granulator then; The mass ratio of beta-schardinger dextrin-and starch is (0.05～0.1): (99.95～99.9), the heavy amount of beta-schardinger dextrin-and starch are 6.9～17.2 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 1.2～3mm.Other step and parameter are identical with one of embodiment one to seven.
The specific embodiment nine: this embodiment with the difference of one of specific embodiment one to eight is: the particle diameter of antibacterial peptide microgranule is 0.35～0.65mm in the step 2.Other step and parameter are identical with one of embodiment one to eight.
The specific embodiment ten: this embodiment with the difference of one of specific embodiment one to nine is: the solvent of chitosan acid solution is that volumetric concentration is 0.1%～5% organic acid or mineral acid in the step 1; Solvent is preferably acetic acid.Other step and parameter are identical with one of embodiment one to nine.
The specific embodiment 11: this embodiment with the difference of one of specific embodiment one to ten is: the rotating speed of step 1 high speed homogenizer is 8000～20000r/min.Other step and parameter are identical with one of embodiment one to ten.
The specific embodiment 12: this embodiment antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, be that 1.5%～2.0% sodium alginate aqueous solution and mass concentration are that 0.25% (W/V)～1.5% (W/V), deacetylation are that 82%～85% chitosan acid solution is with high speed homogenizer mixing with mass concentration; And to regulate pH value be 5.5, forms slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are (3.3～8.6) in mass ratio: the ratio of (2～4) together adds carries out embedding in the medical mixer, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule is carried out the beta-schardinger dextrin-embedding, promptly obtains the antibacterial peptide slow-release microcapsule.
A kind of chitosan of step is 3.3 * 10 by molecular weight
5～4.6 * 10
5High molecular weight chitosan and molecular weight be 0.72 * 10
5～1.19 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.3～0.8 in the chitosan acid solution;
The solvent of chitosan acid solution is that volumetric concentration is 0.1%～5% organic acid or mineral acid in the step 1;
Sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution;
Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide in the step 2, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD, and the relative molecular weight of HMW antibacterial peptide is 40～50KD;
The beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with extruding granulator or rotation granulator then; The mass ratio of beta-schardinger dextrin-and starch is (0.05～0.1): (99.95～99.9), the heavy amount of beta-schardinger dextrin-and starch are 6.9～17.2 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 1.2～3mm.
The rotating speed of step 1 high speed homogenizer is 8000～20000r/min.
The particle diameter of antibacterial peptide microgranule is 0.35～0.65mm in this embodiment method step two.
The antibacterial peptide slow-release microcapsule antibacterial peptide envelop rate of this embodiment method preparation is up to more than 90%, but large-scale industrialized production.
The antibacterial peptide slow-release microcapsule of this embodiment method preparation does not develop immunity to drugs, and can select different antibacterial peptides for use to different controlling objects.For shigella, bacillus megaterium, micrococcus luteus, corynebacterium, Proteus sonnei, streptomycin is produced chemical sproof escherichia coli, escherichia coli JM 109, bacillus subtilis, staphylococcus aureus, harmful intestinal (especially to often causing the colon bacillus 0157 of alimentary toxicosis), kills Fusarium spp. Fusarium moniliforme, kills Fusarium spp. Fusarium oxysporium, the mankind and plant pathogenic fungi excellent inhibition and killing effect are all arranged.And adopt antibacterial peptide to intravital symbiotic bacteria of normal person and not damaged, and therefore can be used to change the composition of intestinal bacteria group, thus enhances human body body constitution.
The antibacterial peptide slow-release microcapsule zoopery of this embodiment preparation
The antibacterial peptide slow-release microcapsule zoopery of this embodiment preparation shows that the antibacterial peptide slow-release microcapsule can adhere to crack-free more than 48 hours on the mouse small intestine surface, thereby guarantees that medicine can fully be absorbed by small intestinal.Under the external stripping experiment condition of antibacterial peptide slow-release microcapsule medicine, discharge 85% in 12 hours inherent simulated intestinal fluids, and in simulated gastric fluid, only discharge 15%, thereby guarantee that entrapped drug is not by stomach acids destroy.The antibacterial peptide slow-release microcapsule can keep its original strength character and controlled release properties after drying under the preparation process condition, can be used for solid preparation.
The antibacterial peptide slow-release microcapsule release medium pH value experiment of this embodiment preparation
The antibacterial peptide slow-release microcapsule release medium pH value experiment of this embodiment preparation shows that antibacterial peptide slow-release microcapsule Release Performance in acidity or alkaline medium is all good, but has the pH dependency, and rate of releasing drug is than fast in alkaline medium in acid medium.In acid medium, alkali-soluble ALG (sodium alginate) is insoluble skeleton, and acid soluble Cts (chitosan) is the solubility gel; Otherwise in alkaline medium then.Medicine discharges through dissolving diffusion and the corrosion of capsule material.Along with macromolecule degraded in vivo, the structure of composite membrane of this embodiment antibacterial peptide slow-release microcapsule becomes loose gradually, makes to include the resistance minimizing that antibacterial peptide therefrom dissolves and spreads, and the antibacterial peptide rate of release is accelerated.Just to reduce caused rate of releasing drug slack-off when consistent with content of dispersion when the quickening of antibacterial peptide rate of release; The long time constant of antibacterial peptide discharges and is achieved, thereby this embodiment antibacterial peptide slow-release microcapsule all demonstrates good Release Performance under acid and alkali condition.
The intensity of the little glue of antibacterial peptide slow release of this embodiment preparation is 75.02, and toughness and elasticity are 58.42.
The specific embodiment 13: this embodiment antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with mass concentration be 1.6% sodium alginate aqueous solution and mass concentration be 1.0% (W/V), deacetylation be 82% chitosan acid solution with high speed homogenizer mixing, and to regulate pH value be 5.5, forms slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are 3.3. in mass ratio: 4 ratio together adds carries out embedding in the medical mixer, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule is carried out the beta-schardinger dextrin-embedding, promptly obtains the antibacterial peptide slow-release microcapsule.
A kind of chitosan of step is 3.5 * 10 by molecular weight
5～4.5 * 10
5High molecular weight chitosan and molecular weight be 0.75 * 10
5～1.15 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.4 in the chitosan acid solution;
The solvent of chitosan acid solution is that volumetric concentration is 0.2% acetic acid in the step 1;
Sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution;
Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide in the step 2, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD, and the relative molecular weight of HMW antibacterial peptide is 40～50KD;
The beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with extruding granulator then; The mass ratio of beta-schardinger dextrin-and starch is 0.05: 99.95, and the heavy amount of beta-schardinger dextrin-and starch is 17 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 2.5mm～3mm.
The rotating speed of step 1 high speed homogenizer is 10000r/min.
The intensity of the little glue of antibacterial peptide slow release of this embodiment preparation is 75.23, and toughness and elasticity are 59.17.
The antibacterial peptide slow-release microcapsule antibacterial peptide envelop rate of this embodiment method preparation is 96.5%, but large-scale industrialized production.
This embodiment antibacterial peptide separates acquisition from sarcophagid, so the antibacterial peptide slow-release microcapsule that obtains of this embodiment can kill shigella, bacillus megaterium, micrococcus luteus, corynebacterium, Proteus sonnei effectively and streptomycin is produced chemical sproof escherichia coli.
The specific embodiment 14: this embodiment antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with mass concentration be 1.6% sodium alginate aqueous solution and mass concentration be 0.8% (W/V), deacetylation be 85% chitosan acid solution with high speed homogenizer mixing, and to regulate pH value be 5.5, forms slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are that 4: 3.5 ratio together adds and carries out embedding in the medical mixer in mass ratio, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule are carried out the beta-schardinger dextrin-embedding, promptly obtain the antibacterial peptide slow-release microcapsule.
A kind of chitosan of step is 3.8 * 10 by molecular weight
5～4.2 * 10
5High molecular weight chitosan and molecular weight be 0.80 * 10
5～1.10 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.5 in the chitosan acid solution;
The solvent of chitosan acid solution is that volumetric concentration is 1% hydrochloric acid in the step 1;
Sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution;
Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide in the step 2, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD, and the relative molecular weight of HMW antibacterial peptide is 40～50KD;
The beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with rotation granulator then; The mass ratio of beta-schardinger dextrin-and starch is 0.07: 99.93, and the heavy amount of beta-schardinger dextrin-and starch is 15 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 2.2mm～2.5mm.
The rotating speed of step 1 high speed homogenizer is 12000r/min.
The intensity of the little glue of antibacterial peptide slow release of this embodiment preparation is 75.88, and toughness and elasticity are 59.02.
The antibacterial peptide slow-release microcapsule antibacterial peptide envelop rate of this embodiment method preparation is 97.3%, but large-scale industrialized production.
This embodiment antibacterial peptide is a silk silkworm antibacterial peptide, so the antibacterial peptide slow-release microcapsule that this embodiment obtains all has the obvious suppression effect to 13 kinds of antibacterials such as escherichia coli JM 109, bacillus subtilis, staphylococcus aureuses.
The specific embodiment 15: this embodiment antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with mass concentration be 2.0% sodium alginate aqueous solution and mass concentration be 1.2% (W/V), deacetylation be 84% chitosan acid solution with high speed homogenizer mixing, and to regulate pH value be 5.5, forms slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are that 6: 3 ratio together adds and carries out embedding in the medical mixer in mass ratio, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule are carried out the beta-schardinger dextrin-embedding, promptly obtain the antibacterial peptide slow-release microcapsule.
A kind of chitosan of step is 3.7 * 10 by molecular weight
5～4.1 * 10
5High molecular weight chitosan and molecular weight be 0.90 * 10
5～1.18 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.6 in the chitosan acid solution;
The solvent of chitosan acid solution is that volumetric concentration is 3% acetic acid in the step 1;
Sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution;
Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide in the step 2, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD, and the relative molecular weight of HMW antibacterial peptide is 40～50KD;
The beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with extruding granulator then; The mass ratio of beta-schardinger dextrin-and starch is 0.09: 99.91, and the heavy amount of beta-schardinger dextrin-and starch is 12 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 1.8mm～2mm.
The rotating speed of step 1 high speed homogenizer is 15000r/min.
The intensity of the little glue of antibacterial peptide slow release of this embodiment preparation is 74.53, and toughness and elasticity are 62.52.
The antibacterial peptide slow-release microcapsule antibacterial peptide envelop rate of this embodiment method preparation is 97.3%, but large-scale industrialized production.
The specific embodiment 16: this embodiment antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with mass concentration be 1.8% sodium alginate aqueous solution and mass concentration be 1.3% (W/V), deacetylation be 84% chitosan acid solution with high speed homogenizer mixing, and to regulate pH value be 5.5, forms slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are that 8: 2 ratio together adds and carries out embedding in the medical mixer in mass ratio, and to regulate pH value be 5.5, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule are carried out the beta-schardinger dextrin-embedding, promptly obtain the antibacterial peptide slow-release microcapsule.
A kind of chitosan of step is 3.5 * 10 by molecular weight
5～4.0 * 10
5High molecular weight chitosan and molecular weight be 1.00 * 10
5～1.10 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.7 in the chitosan acid solution;
The solvent of chitosan acid solution is that volumetric concentration is 4% acetic acid in the step 1;
Sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution;
Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide in the step 2, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD, and the relative molecular weight of HMW antibacterial peptide is 40～50KD;
The beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with rotation granulator then; The mass ratio of beta-schardinger dextrin-and starch is 0.1: 99.9, and the heavy amount of beta-schardinger dextrin-and starch is 8 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 1.4mm～1.8mm.
The rotating speed of step 1 high speed homogenizer is 18000r/min.
The intensity of the little glue of antibacterial peptide slow release of this embodiment preparation is 76.58, and toughness and elasticity are 59.02.
The antibacterial peptide slow-release microcapsule antibacterial peptide envelop rate of this embodiment method preparation is 97.3%, but large-scale industrialized production.
Claims (10)
1. the method for preparing of antibacterial peptide slow-release microcapsule is characterized in that the antibacterial peptide slow-release microcapsule prepares according to the following steps:
One, with sodium alginate aqueous solution and chitosan acid solution with high speed homogenizer mixing, form slow-releasing granules;
Two, antibacterial peptide and slow-releasing granules are together added carry out embedding in the medical mixer, obtain the antibacterial peptide microgranule, then the antibacterial peptide microgranule is carried out the beta-schardinger dextrin-embedding, promptly obtain the antibacterial peptide slow-release microcapsule.
2. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 1; It is characterized in that in the step 1 that the chitosan mass concentration is that 0.25% (W/V)～1.5% (W/V), deacetylation are 82%～85% in the chitosan acid solution, chitosan is 3.3 * 10 by molecular weight
5～4.6 * 10
5High molecular weight chitosan and molecular weight be 0.72 * 10
5～1.19 * 10
5Low-molecular weight chitoglycan form by 1: 1 mass ratio; And adding glucose in the chitosan acid solution, the mol ratio of chitosan and glucose is 0.3～0.8 in the chitosan acid solution.
3. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 2, the mass concentration that it is characterized in that sodium alginate in the step 1 sodium alginate aqueous solution is 1.5%～2.0%.
4. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 3 is characterized in that regulating pH value in step 1 sodium alginate aqueous solution and the chitosan acid solution blending process is 5.2～5.7.
5. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 4 is characterized in that sodium alginate is 4: 6 with the chitosan mass ratio in step 1 sodium alginate aqueous solution and the chitosan acid solution.
6. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 5 is characterized in that the particle diameter of the slow-releasing granules that step 1 forms is 0.35～0.65mm, and the particle diameter after the oven dry is 0.1～0.2mm.
7. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 6; The mass ratio that it is characterized in that antibacterial peptide and slow-releasing granules in the step 2 is (3.3～8.6): (2～4); Antibacterial peptide is made up of by 1: 2 mass ratio low-molecular-weight antibacterial peptide and HMW antibacterial peptide, and the relative molecular weight of low-molecular-weight antibacterial peptide is 6.2～17.2KD; The relative molecular weight of HMW antibacterial peptide is 40～50KD.
8. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 7; It is characterized in that the beta-schardinger dextrin-embedding is carried out according to the following steps in the step 2: in the antibacterial peptide microgranule, add beta-schardinger dextrin-and starch, carry out the beta-schardinger dextrin-embedding with extruding granulator or rotation granulator then; The mass ratio of beta-schardinger dextrin-and starch is (0.05～0.1): (99.95～99.9), the heavy amount of beta-schardinger dextrin-and starch are 6.9～17.2 times of antibacterial peptide particle mass; The particle diameter of the antibacterial peptide slow-release microcapsule that obtains after the beta-schardinger dextrin-embedding is 1.2～3mm.
9. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 7, the particle diameter that it is characterized in that antibacterial peptide microgranule in the step 2 is 0.35～0.65mm.
10. the method for preparing of antibacterial peptide slow-release microcapsule according to claim 9, the solvent that it is characterized in that chitosan acid solution in the step 1 are that volumetric concentration is 0.1%～5% organic acid or mineral acid.
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