CN107875984A - The method and its application of freezing preparation of biological active material/water soluble drug microcapsules - Google Patents
The method and its application of freezing preparation of biological active material/water soluble drug microcapsules Download PDFInfo
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J13/00—Colloid chemistry, e.g. the production of colloidal materials or their solutions, not otherwise provided for; Making microcapsules or microballoons
- B01J13/02—Making microcapsules or microballoons
- B01J13/025—Applications of microcapsules not provided for in other subclasses
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61J—CONTAINERS SPECIALLY ADAPTED FOR MEDICAL OR PHARMACEUTICAL PURPOSES; DEVICES OR METHODS SPECIALLY ADAPTED FOR BRINGING PHARMACEUTICAL PRODUCTS INTO PARTICULAR PHYSICAL OR ADMINISTERING FORMS; DEVICES FOR ADMINISTERING FOOD OR MEDICINES ORALLY; BABY COMFORTERS; DEVICES FOR RECEIVING SPITTLE
- A61J3/00—Devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms
- A61J3/07—Devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms into the form of capsules or similar small containers for oral use
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/5005—Wall or coating material
- A61K9/5015—Organic compounds, e.g. fats, sugars
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/5089—Processes
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J13/00—Colloid chemistry, e.g. the production of colloidal materials or their solutions, not otherwise provided for; Making microcapsules or microballoons
- B01J13/02—Making microcapsules or microballoons
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J13/00—Colloid chemistry, e.g. the production of colloidal materials or their solutions, not otherwise provided for; Making microcapsules or microballoons
- B01J13/02—Making microcapsules or microballoons
- B01J13/04—Making microcapsules or microballoons by physical processes, e.g. drying, spraying
Abstract
The present invention relates to a kind of method of freezing preparation of biological active material/water soluble drug microcapsules, comprise the following steps：Water-soluble bioactive material/water soluble drug is dissolved in lytic agent, it is stored refrigerated, obtain bioactivity/water soluble drug aqueous solution；Organic solvent, biocompatible surfactants, bioactivity/water soluble drug aqueous solution are stirred mixing, form stable microemulsion system；Microemulsion is contained into closed container, and the freezen protective in the case where being stirred vigorously effect；The addition inorganic cation lipid materials reacted while stirring into the emulsion freezed；Reaction is taken out sample and is freeze-dried after terminating, obtain bioactivity/water soluble drug microcapsules.Obtained bioactive substance/water soluble drug microcapsules are as the application on biology sensor or slow releasing pharmaceutical induction system.The medicine encapsulation of the present invention is good, and biological active component is stable, and cost is relatively low.
Description
Technical field
The present invention relates to biology for nano-hollow material preparation, drug delivery, biosensor technology field, more particularly to
A kind of method and its application of freezing preparation of biological active material/water soluble drug microcapsules.
Background technology
Because China's preparation research lags for a long time, the preparation of conventional formulation and low-level repetition is more, slow-release controlled-release preparation
It is less, therefore China should increase the exploitation dynamics of slow-release controlled-release technology, strive using the conventional formulation in kind in several years
It is transformed into sustained-release and controlled release preparation and strengthens targeting preparation research, increase high potency drugs preparation new varieties strengthens compound sustained-released, controlled release system
Agent, liquid slow-release, the research and development paces of controlled release preparation.
Microcapsules technology refer to scattered solid matter, drop or gas to be completely enclosed within one layer of dense film formed it is micro-
The method of capsule.Usual dense film is made up of natural or synthetic high polymer material, degradation material, the microcapsules shape containing solid
Shape is typically same with solid phase, and the shape of the microcapsules containing liquid or gas is generally spherical.Recent years is with the micro- glue of macromolecule
The progress of capsule technology, microcapsules are applied to the technical field at high tip, such as medical treatment, medicine, biochemistry and e-mail
The fields such as breath.
In order to treat various diseases, people need to often take various medicines, and medicine is chemicals influential on physiological function
Matter.But some medicines can have some shortcomings when taking, therefore drug microcapsule can be overcome to these shortcomings.Microcapsule membrane
With the release for changing material in physical appearance and property, and extension and control film, storage stability is improved, by immiscible
The effects such as composition isolation.Microcapsules are divided into impermeable microcapsules and semi-transparent microcapsules according to the difference of its capsule material, impermeable microcapsules
Capsule material is relatively thick, and suitable internal environment is provided for core, and core and external environment condition are kept apart, and prevents it with outer
The material of portion's environment is reacted, and when release is needed, capsule material is destroyed with the methods of pressurization, heating or radiation.It is semi-transparent micro-
Capsule need not destroy capsule material when in use, and the small-molecule substance in core or external environment condition can freely pass through capsule
Material, and balance is rapidly achieved, thus the function of sustained release and control release can be reached.For semi-transparent microcapsules, the table of microcapsules
The physical properties such as face structure, film thickness, porous and core distribution are to determine the principal element of core sustained release performance.It is main special
Point is that the stability for increasing medicine avoids first pass effect from delaying to discharge, and the character for reducing toxic side effect change medicine and thing material is covered
Smell and excitant.
Since Micro-Encapsulation Technique comes out, its new method prepared, new technology are always just the master of many workers
Want one of research direction.At present, chemical method, Physical and physical-chemical process three major types preparation method have been primarily formed.However, absolutely
Most of microcapsules prepare methods due to the limitation of reaction condition be required for by higher temperature (>37 DEG C) handled, such as spray
Seasoning, macromolecule interfacial polymerization etc..Too high temperature can destroy the activity of bioactive substance or the drug effect of medicine, no
Beneficial to the preparation of Biofunctional material.The characteristics of bioactive materials, is that its bioactivity work(will not be destroyed under cryogenic conditions
Can, be advantageous to the storage and transport of bioactive functions, therefore carry out bioactive substance microcapsules solidification at low ambient temperatures
Meet the requirement of productivity dynamics.
The content of the invention
It is micro- that the technical problems to be solved by the invention are to provide a kind of freezing preparation of biological active material/water soluble drug
The method and its application of capsule, to overcome above-mentioned deficiency of the prior art.
The technical scheme that the present invention solves above-mentioned technical problem is as follows：A kind of freezing preparation of biological active material/water-soluble
The method of property drug microcapsule, specifically comprises the following steps：
S100, water-soluble bioactive material/water soluble drug is dissolved in lytic agent, it is stored refrigerated, obtain biological work
Property/the water soluble drug aqueous solution；
S200, organic solvent, biocompatible surfactants, bioactivity/water soluble drug aqueous solution stirred
Mixing is mixed, forms stable microemulsion system；
S300, by microemulsion contain into closed container, and be stirred vigorously effect under freezen protective；
S400, the addition inorganic cation lipid materials reacted while stirring into the emulsion freezed；
S500, reaction take out sample after terminating, and sample are freeze-dried, and obtain the micro- glue of bioactivity/water soluble drug
Capsule.
On the basis of above-mentioned technical proposal, the present invention can also do following improvement.
Further, in the step S100, prepare bioactivity/water soluble drug aqueous solution and specifically include following steps：
S110, water-soluble bioactive material/water soluble drug is subjected to Bio-Pretreatment, Bio-Pretreatment process includes
Sterilizing, removal of impurities, deoxygenation, purification, freezing；
S120, aseptically, is dissolved in 0 DEG C to 4 DEG C by water-soluble bioactive material/water soluble drug after processing
Lytic agent in, be made into corresponding bioactivity/water soluble drug aqueous solution；
S130, the bioactivity prepared/water soluble drug aqueous solution is positioned over it is stored refrigerated in less than 4 DEG C of environment.
Further, in the step S200, by organic solvent, biocompatible surfactants, bioactivity/water solubility
When pharmaceutical aqueous solution is stirred mixing, it must be carried out in 0 DEG C to 4 DEG C of environment.
Further had the beneficial effect that using above-mentioned：1st, the mobility and dissolubility of more than 0 DEG C guarantee water, less than 2,4 DEG C
That many medicines and biomaterial store the temperature for keeping activity for a long time, as a kind of needs of vaccine, anticarcinogen below 4 DEG C it is cold
Hide and preserve, its activity can be destroyed higher than 4 DEG C.
Further, the organic solvent is nontoxic low-freezing organic solvent not soluble in water, including hexamethylene, just oneself
Alkane, atoleine, ethyl acetate, dichloromethane.
Further, the quality of lytic agent is the 1% to 60% of organic solvent quality in microemulsion system, biocompatible surfaces
The quality of activating agent is the 1% to 30% of lytic agent quality, and the quality of bioactive substance/water soluble drug is the matter of lytic agent
The 1% to 30% of amount.
Further had the beneficial effect that using above-mentioned：Ensure that organic solvent content is more than dissolving agent content, form stabilization
Water-in-oil system.
Further, in the step S300, the speed of stirring is 200r/min to 1500r/min.
Further had the beneficial effect that using above-mentioned：Ensure in dispersiveness and water-in-oil system, it is micro- that drop particle diameter is less than 2
Rice, so as to which the small ice drop particle diameter formed in law system is freezed is less than 2 microns.
Further, microemulsion, need to be by 1 DEG C/min to 5 DEG C/min rate of temperature fall at 0 DEG C to -80 DEG C when being freezed
At a temperature of, freeze 2h to 148h.
Further, the biocompatible surfactants include octylame, span-80, cetyl trimethylammonium bromide,
Neopelex, lauryl sodium sulfate, A- alkene sulfonates, alkylsulfonate, A- sulfo groups monocarboxylic acid and its derivative
Thing, aliphatic acid sulfoalkyl are cruel, aliphatic acid sulfoalkyl acid amides, butanedioic acid double cruel sulfonate, sulfuric acid, gelatin, alkyl glycerylethers
One or more of combinations in sulfonate, sucrose ester and phosphate ester salt.
Further, the water-soluble bioactive material/water soluble drug includes but is not limited to：Glucose oxidase, receive
Beans kinases, mescenchymal stem cell, DNA vaccine for foot-and-mouth disease, methyl-glyoxal, glycyrrhizic acid, mono methoxy polyethylene glycol one poly- third are handed over
Cruel second hands over cruel, injection hepatitis B vaccine, beta-galactosidase gene DNA vaccination, human hepatoma cell strain, VeroE6 cells, coloured glaze
Saptoril, alkali protease, bovine serum albumin(BSA), dexamethasone phosphate, secretory antibody CD137 and antibody OX40 hybridoma
Cell, Naproxenic acid, hydrogenated palm oil, lactobacillic acid, alliin, 1- pyridines -4- (3- amidos) pyrimidine piperazine, 4- (the chloro- 4- of 3-
Methoxybenzene amido) -6- iodine quinazoline, captopril, cell, biology enzyme, BCG vaccine, inactivation SARS vaccines, Tilapia mossambica hammer
Bioactivity, the drug substances such as water-soluble enzyme, bacterium, medicine, the viruses such as bacteria vaccine, enteropathogenic E. Coli vaccine, astaxanthin.
Further, bioactivity/water soluble drug Microcapsules Size size is 10nm to 1000nm, and microcapsules wall thickness is
1nm to 100nm.
The beneficial effects of the invention are as follows：
1) the characteristics of phase transformation congeals into ice occurs using water droplet low temperature, the advantages of combining soft or hard template, has
Nano-hollow material pattern, size, reaction are controllable, and template easily removes；
2) be advantageous to protect divided active component as template using aqueous phase；
3) reaction condition is gentle, is advantageous to technique amplification；
4) reaction protects the activity of biomaterial, is reflected in the progress of ice water drops surface, protect the biology of inside without heating
Material；
5) whole technique has simple to operate, and green, medicine encapsulation is good, and biological active component is stable, cost compared with
It is low.
Obtained bioactive substance/water soluble drug microcapsules are as biology sensor or slow releasing pharmaceutical induction system
On application.
Brief description of the drawings
The TEM figures of the hollow titanium dioxide parcel astaxanthins of Fig. 1.
The XRD of the hollow titanium dioxide parcel bovine serum albumin(BSA)s of Fig. 2.
The TEM figures of Fig. 3 hollow silicon dioxides parcel hepatitis B vaccine.
Fig. 4 hollow silicon dioxides wrap up captopril XRD spectrum.
The IR figures of the hollow titanium dioxide parcel glucose oxidases of Fig. 5.
Embodiment
The principle and feature of the present invention are described below in conjunction with accompanying drawing, the given examples are served only to explain the present invention, and
It is non-to be used to limit the scope of the present invention.
The present invention is the characteristics of phase-state change occurs in building-up process using template, to combine soft template method, die
The advantages of plate method, the method that have devised freezing preparation of biological active material/water soluble drug microcapsules.This method can be used
In fields such as biology enzyme solidification, cell curing, viral solidification, drug deliveries, for preparing bioactive enzyme microcapsules, DNA epidemic diseases
The bioactive substance microcapsules such as seedling microcapsules, viral microcapsules, cell microcapsules, water soluble drug microcapsules, can be used for
Manufacture the biosystems such as enzyme sensor, water soluble drug delivery system, cancer cell sensor, viral sensor, vaccine sensor
Manufacture.
Embodiment one：
The method that freezing prepares mesenchymal stem cells bio-microcapsule, specifically comprises the following steps：
S100, prepare cell suspension：It is 6 × 106/ml to configure Tissue Culture Dish inner cell density, is added in every ware thin
Born of the same parents digestive juice 10ml, 5min to 10min is digested, add DMEM culture mediums 20ml and terminate digestion, be collected in centrifuge tube, use cell
After survival rate and analysis of accounts instrument meter number, 3min is centrifuged under 2000rpm/min speed, goes supernatant liquid to obtain purifying thin
Born of the same parents, add sterile saline and be diluted to required concentration, cell suspension is made, by 0.1ml mesenchymal stem cells suspension in 4 DEG C of bars
It is dissolved under part in 5mL distilled water；
S200,2ml octylames, 50ml n-hexanes be added in the solution in step S100, stirred, it is micro- to form W/O
Emulsion；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 with 3 DEG C/min
DEG C, at -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 800r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
48h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica bag
Wrap up in mesenchymal stem cells bio-microcapsule.
Embodiment two：
The method that freezing prepares anticarcinogen 1- pyridines -4- (3- amidos) pyrimidine piperazine microcapsules, specifically includes following step
Suddenly：
S100, by 0.1g water-soluble anticancer medicine 1- pyridines -4- (3- amidos) pyrimidine piperazines (MA2BC) under the conditions of 4 DEG C it is molten
In 5mL distilled water；
S200,2mlSPan80,50ml ethyl acetate is added in the solution in step S100, stirred, form W/
O microemulsions；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 with 4 DEG C/min
DEG C, and at -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 1200r/min；
The a certain amount of fourth fat ester of metatitanic acid four is added in S400, the emulsion freezed into step S300, it is anti-to continue freezing stirring
Answer 105h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain titanium dioxide bag
Wrap up in 1- pyridines -4- (3- amidos) pyrimidine piperazine microcapsules.
As shown in figure 4, embodiment three：
The method that freezing prepares captopril drug microcapsule, specifically comprises the following steps：
S100, captopril bulk drug is dissolved in deionized water under the conditions of 4 DEG C, is made into certain density material liquid；
S200,2mlSPan80,50ml atoleine is added in the solution in step S100, stirred, form W/
O microemulsions；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -20 with 2 DEG C/min
DEG C, at -20 DEG C of temperature, freezing stirring 12 hours, mixing speed 1150r/min；
A certain amount of Aluminate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction 120h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain aluminum oxide parcel
Captopril drug microcapsule.
Fig. 4 is that hollow silicon dioxide wraps up captopril XRD spectrum, and the implication expressed by the figure is：Peak type is amorphous
The characteristic peak of Silica-coated captopril.
Example IV：
The method that freezing prepares targeting anticarcinogen 4- (3- chloro-4-methoxies anilino-) -6- iodine quinazoline microcapsules, tool
Body comprises the following steps：
S100,0.1g4- (3- chloro-4-methoxies anilino-) -6- iodine quinazoline bulk drugs are dissolved under the conditions of 4 DEG C
Ionized water, it is made into certain density material liquid；
S200,2mlSPan80,50ml atoleine is added in the solution in step S100, stirred, form W/
O microemulsions；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -20 with 1 DEG C/min
DEG C, and at -20 DEG C of temperature, freezing stirring 12 hours, mixing speed 950r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
86h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silicon oxide-wrapped
4- (3- chloro-4-methoxies anilino-) -6- iodine quinazoline drug microcapsules.
As shown in figure 3, embodiment five：
The method that freezing prepares hepatitis B vaccine microcapsules, specifically comprises the following steps：
S100,0.1g injections hepatitis B vaccine (abbreviation hepatitis B vaccine) is dissolved in deionized water under the conditions of 4 DEG C, carried out
Certain density material liquid is made into after aseptic process；
S200,2mlSPan80,50ml pentane is added in the solution in step S100, stirred, form W/O
Microemulsion；
S300, by microemulsion be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -20 DEG C with 2 DEG C/min,
At -20 DEG C of temperature, freezing stirring 12 hours, mixing speed 1000r/min；
A certain amount of tetraethyl orthosilicate, which is added, in S400, the emulsion freezed into step S300 is added to the emulsion freezed
In, continue to freeze stirring reaction 64h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silicon oxide-wrapped
Hepatitis B vaccine drug microcapsule.
Fig. 3 is the TEM figures that hollow silicon dioxide wraps up hepatitis B vaccine, and the implication expressed by the figure is：Silicon dioxide hollow
Spherolite footpath is 30-100nm, and hepatitis B vaccine is wrapped in inside hollow silica ball, wall thickness 3-5nm.
Embodiment six：
The method that freezing prepares inactivation SARS pharmaceutical vaccine microcapsules, specifically comprises the following steps：
The inactivation SARS-CoV solution for taking 0.1ml to purify under S100, sterile working is dissolved in deionized water under the conditions of 4 DEG C,
It is made into certain density material liquid；
S200,2mlSPan80,50ml pentane is added in the solution in step S100, stirred, form W/O
Microemulsion；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 with 3 DEG C/min
DEG C, at -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 1050r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, is added to the breast freezed
In liquid, continue to freeze stirring reaction 52h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silicon oxide-wrapped
Inactivation SARS pharmaceutical vaccine microcapsules.
As shown in figure 1, embodiment seven：
The method that freezing prepares astaxanthin microcapsules, specifically comprises the following steps：
Take 0.1g astaxanthins to be dissolved in deionized water under the conditions of 4 DEG C under S100, the operation of sterile lucifuge, be made into certain density
Material liquid；
S200,2ml emulsifying agents (sucrose ester), 50ml ethyl acetate is added in the solution in step S100, stirring is equal
It is even, form W/O microemulsions；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 with 2 DEG C/min
DEG C, and at -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 850r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continue to freeze stirring reaction
48h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silicon oxide-wrapped
Astaxanthin microcapsules.
Fig. 1 is the TEM figures that hollow titanium dioxide wraps up astaxanthin, and the implication expressed by the figure is：Titanium dioxide hollow ball
Particle diameter is 300-600 nanometers, and freshwater shrimp element is wrapped in inside titanium dioxide hollow ball, wall thickness 100-200nm.
Embodiment eight：
The method that freezing prepares BCG vaccine bio-microcapsule, specifically comprises the following steps：
S100, BCG vaccine freezing dry powder agent 0.1g is dissolved in 5% sodium bicarbonate solution under 4 DEG C of aseptic conditions, be made into
Certain density material liquid；
S200,2ml emulsifying agents (sucrose ester), 50ml ethyl acetate is added in the solution in step S100, stirring is equal
It is even, form W/O microemulsions；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 DEG C with 5 DEG C/min
At -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 350r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
72h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silicon oxide-wrapped
Astaxanthin microcapsules.
As shown in figure 5, embodiment nine：
The method that freezing prepares glucose oxidase microcapsules, specifically comprises the following steps：
Glucose oxidase 0.1g is dissolved in deionized water under S100, sterile working, is made into certain density material liquid；
S200,2ml octylames, 50ml pentanes be added in the solution in step S100, stirred, it is micro- to form W/O
Emulsion；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 with 5 DEG C/min
DEG C, at -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 1200r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
106h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica solidification
Glucose oxidase microcapsules.
Fig. 5 is the IR figures that hollow titanium dioxide wraps up glucose oxidase, and the implication expressed by the figure is：IR collection of illustrative plates is Portugal
The characteristic peak of grape carbohydrate oxidase and the characteristic peak of titanium dioxide.
Embodiment ten：
The method that freezing prepares Nattokinase microcapsules, specifically comprises the following steps：
To pass through under S100, sterile working zymotic fluid caused by deep fermentation second day with 5000rpm, 15min from
The heart is separated, and obtains supernatant, then obtained supernatant is filtered, and be can obtain filtrate and is dissolved in deionized water, is made into certain
The Nattokinase proferment feed liquid of concentration；
S200,2g gelatin, 50ml dichloromethane be added in the solution in step S100, stirred, it is micro- to form W/O
Emulsion；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 DEG C with 5 DEG C/min
At -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 450r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
110h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica solidification
Nattokinase microcapsules.
As shown in Fig. 2 embodiment 11：
The method that freezing prepares bovine serum albumin microcapsules, specifically comprises the following steps：
Bovine serum albumen solution is dissolved in deionized water under S100, sterile working, is made into certain density cow's serum egg
White material liquid；
S200,2g gelatin, 50ml atoleines be added in the solution in step S100, stirred, it is micro- to form W/O
Emulsion；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -20 DEG C with 5 DEG C/min
At -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 450r/min；
A certain amount of tetraethyl orthosilicate, which is added, in S400, the emulsion freezed into step S300 is added to the emulsion freezed
In, continue to freeze stirring reaction 144h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica solidification
Bovine serum albumin microcapsules.
Fig. 2 is the XRD that hollow titanium dioxide wraps up bovine serum albumin(BSA), and the implication expressed by the figure is：Peak type is nothing
The titanium dioxide that shapes wraps up the characteristic peak of bovine serum albumin(BSA).
Embodiment 12：
The method that freezing prepares beta-galactosidase gene DNA vaccination microcapsules, specifically comprises the following steps：
Beta-galactosidase gene DNA vaccination solution is dissolved in deionized water under S100, sterile working, is made into certain dense
The beta-galactosidase gene DNA vaccination material liquid of degree；
S200,2g gelatin, 50ml n-hexanes be added in the solution in step S100, stirred, form W/O micro emulsions
Liquid；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 DEG C with 5 DEG C/min
At -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 550r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
72h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica solidification
Beta-galactosidase gene DNA vaccination microcapsules.
Embodiment 13：
The method that freezing prepares Tilapia mossambica hammer bacteria microcapsule, specifically comprises the following steps：
S100, Tilapia mossambica Streptococcusagalactiae SIP prokaryotic expression engineering bacteria SIP-pET32a (+) inoculations are trained in LB liquid
To support in base, 37 DEG C of shaken cultivations to OD600 are 0.5 to 0.6, add derivant ITPG, 25 DEG C of induced expressions 6h, 4000r/min,
15min is centrifuged, collects thalline, ultrasonic disruption bacterium solution, sip protein is collected and purifies standby, and sip protein is dissolved in deionization
In water, certain density sip protein material liquid is made into；
S200,2gSPan80,50ml n-hexane is added in the solution in step S100, stirred, it is micro- to form W/O
Emulsion；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -30 DEG C with 5 DEG C/min
At -30 DEG C of temperature, freezing stirring 12 hours, mixing speed 700r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
72h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica solidification
Tilapia mossambica hammer bacteria microcapsule.
Embodiment 14：
The method that freezing prepares enteropathogenic E. Coli microcapsule vaccine, specifically comprises the following steps：
S100, Escherichia coli whole cell vaccine is dissolved in deionized water it is made into suitable concentration；
S200,2g gelatin, 50ml n-hexanes be added in the solution in step S100, stirred, form W/O micro emulsions
Liquid；
S300, by W/O microemulsions be mounted in closed container in be put into cryostat pot or refrigerator, be cooled to -40 DEG C with 5 DEG C/min
At -40 DEG C of temperature, freezing stirring 12 hours, mixing speed 800r/min；
A certain amount of tetraethyl orthosilicate is added in S400, the emulsion freezed into step S300, continues to freeze stirring reaction
72h；
S500, reaction take out sample after terminating, ethanol washing centrifugation, be then freeze-dried sample, obtain silica solidification
Enteropathogenic E. Coli microcapsule vaccine.
Although embodiments of the invention have been shown and described above, it is to be understood that above-described embodiment is example
Property, it is impossible to limitation of the present invention is interpreted as, one of ordinary skill in the art within the scope of the invention can be to above-mentioned
Embodiment is changed, changed, replacing and modification.
Claims (10)
- A kind of 1. method of freezing preparation of biological active material/water soluble drug microcapsules, it is characterised in that specifically include as Lower step：S100, water-soluble bioactive material/water soluble drug is dissolved in lytic agent, it is stored refrigerated, obtain bioactivity/water The soluble drug aqueous solution；S200, organic solvent, biocompatible surfactants, bioactivity/water soluble drug aqueous solution is stirred it is mixed Close, form stable microemulsion system；S300, by microemulsion contain into closed container, and be stirred vigorously effect under freezen protective；S400, the addition inorganic cation lipid materials reacted while stirring into the emulsion freezed；S500, reaction take out sample after terminating, and sample are freeze-dried, and obtain bioactivity/water soluble drug microcapsules.
- 2. a kind of method of freezing preparation of biological active material/water soluble drug microcapsules according to claim 1, its It is characterised by, in the step S100, prepares bioactivity/water soluble drug aqueous solution and specifically include following steps：S110, water-soluble bioactive material/water soluble drug is carried out to Bio-Pretreatment, Bio-Pretreatment process includes going out Bacterium, removal of impurities, deoxygenation, purification, freezing；S120, aseptically, water-soluble bioactive material/water soluble drug after processing is dissolved in 0 DEG C to 4 DEG C molten Solve in agent, be made into corresponding bioactivity/water soluble drug aqueous solution；S130, the bioactivity prepared/water soluble drug aqueous solution is positioned over it is stored refrigerated in less than 4 DEG C of environment.
- 3. a kind of method of freezing preparation of biological active material/water soluble drug microcapsules according to claim 2, its It is characterised by, in the step S200, by organic solvent, biocompatible surfactants, bioactivity/water soluble drug water When solution is stirred mixing, it must be carried out in 0 DEG C to 4 DEG C of environment.
- 4. a kind of method of freezing preparation of biological active material/water soluble drug microcapsules according to claim 3, its It is characterised by, the organic solvent is nontoxic low-freezing organic solvent not soluble in water, including hexamethylene, n-hexane, liquid Paraffin, ethyl acetate, dichloromethane.
- 5. a kind of method of freezing preparation of biological active material/water soluble drug microcapsules according to claim 4, its It is characterised by, the quality of lytic agent is the 1% to 60% of organic solvent quality in microemulsion system, biocompatible surfactants Quality be the 1% to 30% of lytic agent quality, the quality of bioactive substance/water soluble drug is the quality of lytic agent 1% to 30%.
- 6. a kind of method of freezing preparation of biological active material/water soluble drug microcapsules according to claim 1, its It is characterised by, in the step S300, the speed of stirring is 200r/min to 1500r/min.
- 7. a kind of method of freezing preparation of biological active material/water soluble drug microcapsules according to claim 6, its It is characterised by, microemulsion, need to be by temperature of 1 DEG C/min to the 5 DEG C/min rate of temperature fall at 0 DEG C to -80 DEG C when being freezed Under, freeze 2h to 148h.
- A kind of 8. freezing preparation of biological active material/water soluble drug microcapsules according to claim any one of 1-7 Method, it is characterised in that the biocompatible surfactants include octylame, span-80, cetyl trimethyl bromination Ammonium, neopelex, lauryl sodium sulfate, A- alkene sulfonates, alkylsulfonate, A- sulfo groups monocarboxylic acid and its spread out Biology, aliphatic acid sulfoalkyl are cruel, aliphatic acid sulfoalkyl acid amides, butanedioic acid double cruel sulfonate, sulfuric acid, gelatin, alkyl glycerols One or more of combinations in ether sulfonate, sucrose ester and phosphate ester salt.
- A kind of 9. freezing preparation of biological active material/water soluble drug microcapsules according to claim any one of 1-7 Method, it is characterised in that the water-soluble bioactive material/water soluble drug includes but is not limited to：Glucose oxidase, Nattokinase, mescenchymal stem cell, DNA vaccine for foot-and-mouth disease, methyl-glyoxal, glycyrrhizic acid, mono methoxy polyethylene glycol one poly- third Hand over cruel second hand over cruel, injection hepatitis B vaccine, beta-galactosidase gene DNA vaccination, human hepatoma cell strain, VeroE6 cells, The hybridization of rilanipidic acid, alkali protease, bovine serum albumin(BSA), dexamethasone phosphate, secretory antibody CD137 and antibody OX40 Oncocyte, Naproxenic acid, hydrogenated palm oil, lactobacillic acid, alliin, 1- pyridines -4- (3- amidos) pyrimidine piperazine, (3- is chloro- by 4- 4- methoxybenzenes amido) -6- iodine quinazoline, captopril, cell, biology enzyme, BCG vaccine, inactivation SARS vaccines, Tilapia mossambica chain The bioactivity such as water-soluble enzyme, bacterium, medicine, the viruses such as coccus vaccine, enteropathogenic E. Coli vaccine, astaxanthin, medicine thing Matter.
- 10. using bioactive substance made from any one of claim 1-9/water soluble drug microcapsules as biology sensor Or the application in slow releasing pharmaceutical induction system.
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|---|---|---|---|---|
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102525940A (en) * | 2011-11-30 | 2012-07-04 | 江苏大学 | Inorganic/organic composite protein/polypeptide medicament sustained release microspheres and preparation method thereof |
| CN102703417A (en) * | 2012-06-26 | 2012-10-03 | 东南大学 | Preparation method of cell-loaded biological microcapsules |
| US20130017229A1 (en) * | 2009-12-18 | 2013-01-17 | President And Fellows Of Harvard College | Active scaffolds for on-demand drug and cell delivery |
| CN103239756A (en) * | 2012-02-03 | 2013-08-14 | 佛山兰赛特生物科技有限公司 | Biological active microsphere and preparation method thereof |
-
2017
- 2017-11-16 CN CN201711134607.9A patent/CN107875984A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20130017229A1 (en) * | 2009-12-18 | 2013-01-17 | President And Fellows Of Harvard College | Active scaffolds for on-demand drug and cell delivery |
| CN102525940A (en) * | 2011-11-30 | 2012-07-04 | 江苏大学 | Inorganic/organic composite protein/polypeptide medicament sustained release microspheres and preparation method thereof |
| CN103239756A (en) * | 2012-02-03 | 2013-08-14 | 佛山兰赛特生物科技有限公司 | Biological active microsphere and preparation method thereof |
| CN102703417A (en) * | 2012-06-26 | 2012-10-03 | 东南大学 | Preparation method of cell-loaded biological microcapsules |
Non-Patent Citations (1)
| Title |
|---|
| 陈震: "冰冻辅助反向微乳法制备纳米TiO2和SiO2空心材料及其应用", 《常州大学硕士学位论文》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111205321A (en) * | 2020-03-08 | 2020-05-29 | 博露(厦门)生物股份有限公司 | Preparation and application of anti-coronavirus astaxanthin derivative |
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