CN103961713A - Application of mesoporous silica based controlled release material modified by chitosan - Google Patents
Application of mesoporous silica based controlled release material modified by chitosan Download PDFInfo
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- CN103961713A CN103961713A CN201410188283.7A CN201410188283A CN103961713A CN 103961713 A CN103961713 A CN 103961713A CN 201410188283 A CN201410188283 A CN 201410188283A CN 103961713 A CN103961713 A CN 103961713A
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Abstract
The invention relates to an application of a mesoporous silica based controlled release material modified by a chitosan. The material comprises a mesoporous silica nanoparticle of a compound or group capable of producing a free radical, the compound or group of the free radical loaded in a mesoporous channel of the mesoporous silica nanoparticle, and a chitosanpolyelectrolytes membrane coating the mesoporous silica nanoparticle, wherein the compound or group capable of producing the free radical is loaded in the mesoporous channel. The material responds to the release speed of the compound or group capable of producing the free radical, loaded by a pH control material.
Description
Technical field
The present invention relates to a kind of slow controlled-release material and application thereof, be specifically related to the slow controlled-release material of a kind of chitosan-modified mesoporous silicon-dioxide-substrate and application thereof, belong to nano biological medical material technical field.
Background technology
Reactive oxygen free radical (ROS) is a class active specy, comprises the reduzate of oxygen molecule, as: singlet oxygen (
1o
2), ultra-oxygen anion free radical (O
2-), hydrogen peroxide (H
2o
2), hydroxyl radical free radical (HO), and the product of Ionization Potential of C-Centered Radicals and oxygen molecule, as: peroxy radical (ROO), alkoxy free group (RO) and organic hydroperoxide thing (ROOH) etc.Because free radical has chemical-biological activities, and can pass freely through cell membrane, thereby free radical has the characteristic of killing cell.Quantity research shows greatly, the oxidative damage of biomacromolecule in excessive free radical meeting trigger cell, and the function of meeting Profilin matter, destroys lipid, and causes DNA chain interruption and then trigger cell death.By radiation-actuate, produce free radical (radiotherapy) and be proved to be a kind of effective treatment of cancer means.Another method is exactly by drug-induced generation free radical (chemotherapy).The anticarcinogen that some are common, as camptothecine, amycin, vincristine, the toxic action of arsenic trioxide and bleomycin etc. is all relevant with free radical.How can discharge free radical for a long time around at tumor tissues will be a kind of mode of effective treatment cancer.
Peroxide is the compound that a class contains peroxide bridge (O-O), and general formula is R-O-O-R'.Peroxide bridge in these molecules (O-O) has very strong oxidability, fracture easily occurs and generate two free radical RO, is therefore widely used in plastics and rubber industry as radical initiator.These initiators can produce rapidly free radical under the stimulation of light/heat.Just because of the characteristic of this generation free radical of peroxide, we consider use peroxide as a kind of radical source the free radical therapy for cancer.But peroxide itself is as radical initiator, very sensitive to light and heat, and all unstable in a lot of solvents, and this just makes peroxide be difficult to clinical treatment use.Because these peroxide have probably decomposed when also not arriving lesions position, produced free radical, thereby normal cell has been caused to major injury.In addition, a lot of peroxide, especially some organic peroxides, in water, dissolubility is very low, and this has just limited their further oral and subcutaneous injection application.How can improve the stability of these peroxide, improving their dissolubility in water and further realize transporting of controllability will be a very promising research field, but but have not been reported.
It may be the thinking addressing this problem that peroxide is packed into a kind of carrier, and the report of having seen is used liposome [VB Patel, AN Misra, YS Marfatia, Drug Dev Ind Pharm.2001,27,863-9], microsphere [M Jelvehgari, MR Siahi-Shadbad, S Azarmi, Int J Pharm.2006,308,124-32; RC Wester, R Patel, S Nacht, J AmAcad Dermatol.1991,24,720-6], polymer [J Shim, HS Kang, WS Park, J Control Release.2004,97,477-84] and Liposome nanaparticle [A Dingler, RP Blum, H Niehus, J Microencapsul.1999,16,751-67] load and transport peroxide.But these organic carriers, because self chemical stability is limited, adds that medicine efficiency of loading is low, thereby have limited their clinical practice.Therefore, in the urgent need to developing, a kind ofly there is higher chemistry and heat stability, and there is the free radical delivery systme of high biocompatibility and degradability.
Summary of the invention
The present invention is intended to overcome the defect that existing free radical delivery systme exists, and the invention provides the slow controlled-release material of a kind of chitosan-modified mesoporous silicon-dioxide-substrate and application thereof.
The invention provides the slow controlled-release material of a kind of chitosan-modified mesoporous silicon-dioxide-substrate, described material can load can be produced the compound of free radical or the mesoporous silicon dioxide nano particle of group, load on compound or the group of the free radical in the sub-mesopore orbit of mesoporous silicon dioxide nano particle by mesopore orbit, and the chitosan polyelectrolyte membranes composition that is coated described mesoporous silicon dioxide nano particle.
Preferably, compound or the group of generation free radical comprise peroxide or the group that contains peroxide bridge.
Preferably, produce in the compound of free radical or mesopore orbit that group was loaded into or was connected to mesoporous silicon-dioxide-substrate material and/or mesoporous surface.
Preferably, free radical refers to peroxide, the inner oxide series of intermediate products in course of reaction, comprises the highly active intermediate product that has that exists and do not exist with radical form with radical form.
Preferably, intermediate product comprises at least one in peroxy radical, alkoxy radical, alkyl diradical, organic hydroperoxide thing.
The present invention also provides a kind of above-mentioned application of stating material, the chitosan polyelectrolyte membranes that described material can produce the compound of free radical or the mesoporous silicon dioxide nano particle of group and coated described mesoporous silicon dioxide nano particle by mesopore orbit load forms, and described material response pH value is controlled the compound that can produce free radical of material load or the rate of release of group.
Preferably, the compound that can produce free radical of described material load or the release rate of group under pH=6.5 condition are greater than the release rate under pH=7.4 condition.
Beneficial effect of the present invention:
The pore volume of mesoporous silicon oxide is large, useful load is large, chitosan polyelectrolyte membranes is under the condition of neutrality and alkalescence (simulation normal cell), because the protonated degree of amino on its surface is low, the effect of intermolecular hydrogen bonding is greater than the effect of electrostatic repulsion, strand is reunited, become orderly coherent condition, effectively stop compound or the group of loading to discharge, and under the condition of acid (simulation tumor cell), the protonated degree of amino on its surface is high, electrostatic repulsion is greater than intermolecular hydrogen bonding, with water effect, become collosol state, compound or the group of load can be discharged, reach near target slow-release effect pH response of the present invention free radical delivery systme (pH=6.5) generation free radical tumor tissues responsively, and in not release of normal structure place (pH=7.4), thereby avoided the damaging action of radical pair normal structure.Therefore, there is great pharmaceutical sector application prospect and value.
Accompanying drawing explanation
Fig. 1 is the transmission electron micrograph (TEM) of hollow mesoporous silicon dioxide nano particle (HMSNs) prepared in an embodiment of the invention;
Fig. 2 is the Fourier transform infrared spectroscopy figure (FTIR) of the hollow mesoporous silicon dioxide nano complex that is loaded with benzoyl peroxide (BPO) (BPO@HMSNs) prepared in an embodiment of the invention;
Fig. 3 is the ultraviolet/visible absorption spectra of the release liquid of the BPO@HMSNs-Cs-2 for preparing in an embodiment of the invention.
The specific embodiment
Below in conjunction with accompanying drawing and the following specific embodiment, further illustrate the present invention, should be understood that following embodiment and/or accompanying drawing are only for the present invention is described, and unrestricted the present invention.
The invention discloses a kind of application of pH response free radical delivery systme of mesoporous silicon-dioxide-substrate, belong to nano biological medical material technical field.Described free radical system is to utilize mesoporous silicon-dioxide-substrate material for carrier, floods to load can produce the compound of free radical or modify by chemical group the group that keyed jointing can produce free radical by physics.It is the chitosan polyelectrolyte by or keyed jointing pH response coated at meso-porous titanium dioxide silicon face that described pH response discharges, and the pH response that realizes the free radical to producing discharges.The free radical discharging can be attacked tumor tissues, and then causes malignant cell dead.The present invention not only preparation technology is simple, and can realize free radical in tumor tissues targeting, the release of pH response around, thereby significantly reduces conventional medicament for the damage of normal structure, thereby has great physiological significance and potential applicability in clinical practice.
The pH response free radical delivery systme that the invention provides a kind of mesoporous silicon-dioxide-substrate, described system utilizes mesoporous silicon-dioxide-substrate material for carrier, by load free-radical generating compound (group), controlled release free radical.
Described mesoporous silicon-dioxide-substrate material, has mesoporous silicon oxide or the hollow mesopore silicon dioxide nano material of mesopore orbit.A large amount of loadings that the present invention utilizes cavity structure that Metaporous silicon dioxide material is huge or abundant mesopore orbit to realize free-radical generating compound, or utilize the abundant silicon hydroxyl of silica surface, realize with a large amount of keys of free-radical generating group and connecting, realize the payload to free-radical generating compound.
Described free-radical generating compound (group), the peroxide or the group that contain peroxide bridge (O-O).
The preparation method of described free radical system, free-radical generating compound (group) be loaded into mesoporous silicon-dioxide-substrate material or be connected in the mesopore orbit of mesoporous silicon-dioxide-substrate material and (or) mesoporous surface.
In the preparation method of described free radical system, the carrying method of free-radical generating compound (group) is physics dipping or chemical group keyed jointing.
Described pH response free radical delivery systme, mesoporous silicon oxide area load has the chitosan polyelectrolyte of pH response.
The chitosan polyelectrolyte of described pH response, chitosan polyelectrolyte is wrapped by or is connected to meso-porous titanium dioxide silicon face.
Described controlled free radical delivery systme, the free radical producing refers to peroxide, the inner oxide series of intermediate products in course of reaction, comprises the highly active intermediate product that has that exists and do not exist with radical form with radical form.
Described peroxide, the intermediate product of inner oxide, comprise peroxy radical (ROO), alkoxy radical (RO), alkyl diradical (R) and organic hydroperoxide thing (ROOH) etc.
Described controlled free radical delivery systme, the purposes of the pH response free radical delivery systme of described any one mesoporous silicon-dioxide-substrate is for the damage of tumor tissues and killing of tumor cell.
In the last few years, inorganic mesoporous silicon dioxide nano carrier is owing to having higher chemistry and heat stability, and there is high biocompatibility and degradability, be more and more used to the aspects such as catalysis, bio-imaging, medicine/gene transport, thereby be subject to paying close attention to widely.Wherein, hollow mesoporous silicon dioxide nano particle (HMSNs) especially receives publicity, and this is mainly because hollow mesoporous silicon dioxide nano particle has larger specific surface area, pore volume, there is huge cavity structure, thereby can improve the efficiency of loading of medicine.Meanwhile, the siliceous hydroxyl of the rich surface of these mesopore silicon dioxide nano materials, can the various functional groups of keyed jointing, thus the controllability drug release ability of giving the various stimuli responsives of HMSNs.In addition, utilize nanoparticle originally in tumor tissues EPR effect around, can realize nanoparticle and transport for tumor tissues targeting around.It is a kind of for loading the good carrier material of this unsettled peroxide that this just makes mesopore silicon dioxide nano material become.
Free-radical generating compound of the present invention is the organic peroxides such as benzoyl peroxide, arteannuin, tert-butyl hydroperoxide.
The preparation method of free radical delivery systme of the present invention, comprises the steps:
Steps A) free-radical generating compound (group) by physics, flood in the mesopore orbit that is loaded into mesoporous silicon-dioxide-substrate material or is connected to mesoporous silicon-dioxide-substrate material by chemical group and (or) mesoporous surface;
Step B) pass through physisorption or chemical bonding mode at the chitosan polyeletrolyte of mesoporous silicon-dioxide-substrate material surface or mesopore orbit internal load pH response, the effectively coated and follow-up pH response that realizes the compound that loads discharges;
Steps A) and step B) order according to concrete loading substance and chitosan loaded mode, adjust to some extent.
The pH response free radical delivery systme of mesoporous silicon-dioxide-substrate of the present invention utilize nanoparticle this in tumor tissues EPR effect around, can realize nanoparticle and transport for the targeting of tumor tissues.The preparation method of free radical delivery systme of the present invention is simple and easy to do.PH response free radical delivery systme of the present invention can be responsively near tumor tissues (pH=6.5) produce free radical, and in not release of normal structure place (pH=7.4), thereby avoided the damaging action of radical pair normal structure.Therefore, there is great pharmaceutical sector application prospect and value.
Fig. 1 is the transmission electron micrograph (TEM) of hollow mesoporous silicon dioxide nano particle (HMSNs) prepared in an embodiment of the invention;
Fig. 2 is the Fourier transform infrared spectroscopy figure (FTIR) of the hollow mesoporous silicon dioxide nano complex that is loaded with benzoyl peroxide (BPO) (BPO@HMSNs) prepared in an embodiment of the invention;
Fig. 3 is the ultraviolet/visible absorption spectra of the release liquid of the BPO@HMSNs-Cs-2 for preparing in an embodiment of the invention.
Further exemplify embodiment below to describe the present invention in detail.Should understand equally; following examples are only used to further illustrate the present invention; can not be interpreted as limiting the scope of the invention, some nonessential improvement that those skilled in the art's foregoing according to the present invention is made and adjustment all belong to protection scope of the present invention.The technological parameter that following example is concrete etc. is only also an example in OK range, and those skilled in the art can be done in suitable scope and be selected by explanation herein, and do not really want to be defined in the below concrete numerical value of example.
Embodiment 1
Hollow mesoporous silicon dioxide nano particle (BPO@HMSNs-Cs-1) of preparation encasement polysaccharide (Cs) and loading benzoyl peroxide (BPO).
Steps A) prepare hollow mesoporous silicon dioxide nano particle (HMSNs)
First, utilize stober method to prepare solid silica nanosphere.Afterwards, the solid silica nanosphere (200mg) preparing is distributed in 40mL deionized water, and ultrasonic 15 minutes to disperse nanoparticle wherein.During this period, by 300mg cetyl trimethyl ammonium bromide (C
16tAB) be distributed to 60mL deionized water, in 60mL ethanol and 1.3mL ammonia, under room temperature, mix.Afterwards, above-mentioned solid silica nanosphere suspension after ultrasonic is joined to C
16in TAB mixed liquor and ultrasonic 1 hour.Afterwards, in said mixture, add fast 0.53mL ethyl orthosilicate (TEOS) and continue stirring 7 hours.After reaction finishes, the white depositions obtaining is centrifugal.Afterwards, these precipitate are re-dispersed into 0.4M Na
2cO
3in aqueous solution, and stir 13 hours at 50 ℃ of condition lower magnetic forces.The final white depositions obtaining is dried through centrifugal, alcohol wash final vacuum.Finally, by ion exchange, remove the surfactant C in prepared hollow mesoporous silicon oxide duct
16tAB: the dried sample (0.5g) of above-mentioned acquisition is distributed in the mixed liquor that contains 95% ethanol water and 0.2g ammonium nitrate, this mixture is stirred 5 hours at 60 ℃ of lower magnetic forces, product is dried through centrifugal, alcohol wash final vacuum, gets final product and to obtain hollow mesoporous silicon dioxide nano particle (HMSNs).
Fig. 1 is the transmission electron micrograph (TEM) of hollow mesoporous silicon dioxide nano particle (HMSNs) prepared of the present embodiment.As seen from Figure 1: prepared hollow mesoporous silica nanospheres is regular spherical structure, and diameter is about 350nm.Prepared preparing spherical SiO 2 is hollow and possesses mesopore orbit clearly, and mesoporous wall thickness is about 80nm.
Step B) benzoyl peroxide (BPO) being packed into hollow mesoporous silicon dioxide nano particle (HMSNs) adopts simple physics dipping method to load benzoyl peroxide (BPO).First by by steps A) HMSNs that obtains is distributed in BPO acetone soln, by this suspension stirring at normal temperature 12 hours under dark surrounds.Afterwards, the suspension obtaining is centrifugal, with a small amount of ethanol, wash away the BPO molecule of surface adsorption, vacuum drying at room temperature, can obtain the HMSNs that has loaded BPO afterwards, is labeled as BPO HMSNs.
Fig. 2 is the Fourier transform infrared spectroscopy figure of hollow mesoporous silicon dioxide nano particle (BPO@HMSNs) of the loading BPO for preparing of the present embodiment.Compare with simple HMSNs, in BPO@HMSNs, occurred being positioned at 1760,1228,1000 and 706cm
-1the characteristic peak of the BPO at place, these characteristic peaks correspond respectively to the ν C=O (1760cm of benzoyl peroxide
-1), ν C=C (1600～1450cm
-1), δ C-H (706cm
-1) and ν C-O (1228 and 1000cm
-1), thereby proved the successful loading of BPO.
Step C) BPO@HMSNs being carried out to surperficial chitosan (Cs) is coated
0.6g chitosan is joined in the acetic acid aqueous solution of 100mL10% to stirring at normal temperature 24 hours, obtains 0.6% chitosan colloidal sol, then with sodium hydroxide by the pH regulator to 6.0 of solution.By step B) in the BPO@HMSNs that makes be distributed in above-mentioned chitosan colloidal sol, slowly stir 24 hours.Afterwards, after centrifugal, washing, be drying to obtain coated mesoporous silicon dioxide nano particle that is loaded with BPO of chitosan (Cs), be labeled as BPO@HMSNs-Cs-1.
With surface prepared by transmission electron microscope (TEM) observation post, be coated with chitosan and be mounted with the mesoporous silicon dioxide nano complex (BPO@HMSNs-Cs-1) of BPO.TEM result shows, the surface of HMSNs has one deck organic molecule to cover, formed nano-complex is with respect to HMSNs before, and particle size slightly increases because of chitosan coated, TEM clearly surface chitosan coated in the success on nano-complex surface.
Embodiment 2
Prepare keyed jointing chitosan (Cs) and load hollow mesoporous silicon dioxide nano particle (BPO@HMSNs-Cs-2) of benzoyl peroxide (BPO) and the BPO release evaluation of pH response.
Steps A) prepare hollow mesoporous silicon dioxide nano particle (HMSNs)
According to the steps A of embodiment 1) prepare HMSNs;
Step B) γ-GPTMS being carried out in hollow mesoporous silicon dioxide nano particle (HMSNs) surface modifies
Utilize the feature of the siliceous hydroxyl of HMSNs rich surface, HMSNs is reacted with silane coupler γ-glycidoxypropyltrimethoxy silane (γ-GPTMS), make finishing one deck γ-GPTMS molecule of HMSNs, become and take the HMSNs (being labeled as HMSNs-G) that epoxy alkyl is end group, follow-up chitosan-modified to facilitate;
Step C) pack benzoyl peroxide (BPO) into HMSNs-G
Adopt simple physics dipping method benzoyl peroxide (BPO) molecular diffusion to be entered to the mesopore orbit of HMSNs-G.By step B) HMSNs-G that obtains is distributed in BPO acetone soln, and under dark surrounds, stirring at normal temperature is 12 hours.Afterwards, the suspension obtaining is centrifugal, with a small amount of ethanol, wash away the BPO molecule of surface adsorption, vacuum drying at room temperature, can obtain the HMSNs-G that has loaded BPO afterwards, is labeled as BPO HMSNs-G;
Step D) at BPO@HMSNs-G surface keyed jointing chitosan (Cs)
BPO@HMSNs-G surface keyed jointing a large amount of epoxy alkyls, the epoxy alkyl-amino additive reaction of the amino that utilizes epoxy alkyl and chitosan under acidic catalyst, is keyed to BPO@HMSNs-G by Cs surperficial.0.6g chitosan is joined in the acetic acid aqueous solution of 100mL10% to stirring at normal temperature 24 hours, obtains 0.6% chitosan colloidal sol, then with sodium hydroxide by the pH regulator to 5.0 of solution.By step C) the BPO@HMSNs-G that obtains is distributed in above-mentioned chitosan colloidal sol, and under dark condition, stirring at normal temperature is 12 hours, and standing 6 hours, the pH to 7.4 of regulator solution afterwards.Through centrifugal, washing, after dry, can obtain surperficial keyed jointing chitosan (Cs) and load the silicon dioxide nanocomposite of BPO, be labeled as: BPO@HMSNs-Cs-2;
The BPO@HMSNs-Cs-2 obtaining is carried out to energy dispersion spectrum (EDS) analysis and find, compare with HMSNs, in BPO@HMSNs-Cs-2, occurred the absworption peak of obvious C element, the successful keyed jointing of Cs has been described;
Step e) evaluate the BPO release of the pH response of BPO@HMSNs-Cs-2
BPO@HMSNs-Cs-2 is distributed in the PBS solution of different pH to detect the release conditions of BPO in prepared BPO@HMSNs-Cs-2.Selecting the PBS solution of pH7.4 and 6.5 is in order to simulate respectively the pathological tissues environment (pH ≈ 6.5) such as normal tissue cell environment (pH ≈ 7.4) and Inflamed tissue and cancerous cell.By mixed liquor as for jiggling on the shaking table under room temperature.At specific interval, mixed liquor is centrifugal, and from supernatant, take out 3mL and measure for ultraviolet/visible absorption spectra.In this simultaneously, in the mixed liquor after centrifugal, add the release medium that 3mL is fresh, mixed liquor, after mixing, continues as on shaking table, for follow-up drug release analysis.In order to measure the content of BPO in obtained solution, taken out release medium, after ethanol dilution, is carried out to ultraviolet/visible absorption spectra determination and analysis by the alcoholic solution of acquisition.Every part of test is done three times, and the test result of three times is averaged.
Fig. 3 is the ultraviolet/visible absorption spectra of the release liquid of BPO@HMSNs-Cs-2.As can be seen from the figure, in release medium, BPO has an obvious absworption peak at 234nm place, and this is consistent with the absworption peak (λ max=234nm) of BPO ethanol water, and this BPO discharging with regard to explanation has kept good stability.
The cumulative release curve of the BPO that analysis obtains according to ultraviolet/visible absorption spectra finds that the release of BPO in BPO@HMSNs-Cs-2 has obvious pH response.Release amount of medicine in the PBS environment that burst size in the PBS of pH7.4 environment will be starkly lower than at pH6.5.This huge release difference is mainly caused by following reason: amino ionization state and pH in chitosan have very large relation, in the release medium in pH=7.4, chitosan molecule presents orderly reunion state, and the network structure that chitosan subsides has effectively stoped BPO to discharge from BPO HMSNs-Cs-2.Compare with the limited release under pH7.4 condition, in the PBS of pH6.5 solution, the rate of release of BPO increases to some extent, and this is mainly that chitosan molecule due to HMSNs surface flexibility and the expansion behavior in different pH environment causes.Under the microenvironment in acid (pH6.5), the amino of chitosan occurs protonated, further, counter ion counterionsl gegenions in PBS solution can be at chitosan polyelectrolyte surface aggregation to supplement unnecessary electric charge, this enrichment meeting causes the increase of solution osmotic pressure, the osmotic pressure of this increase makes more hydrone from solution, enter chitosan polyelectrolyte layer, result, chitosan layer expands becomes a kind of loose porous network structure, and become a kind of mixed and disorderly flexibility reunion state from orderly reunion state before, this is just released the mesopore orbit of HMSNs surface part, and then cause the BPO of follow-up acceleration to discharge.Analysis based on above, we can infer, chitosan has been given the controllability release characteristics of a kind of pH response of HMSNs carrier, that is to say, near pathological tissues, BPO accelerated release in vitro.But, the leakage that substantially can not produce BPO under the microenvironment of normal structure.BPO release characteristics under this pH response has increased the therapeutic efficiency of BPO and has reduced the toxic and side effects of its normal tissue.
Embodiment 3
Prepare mesoporous silicon dioxide nano particle (MSNs-OOH) that hydrogen peroxide is modified
Steps A) prepare mesoporous silicon dioxide nano particle (MSNs)
By 0.28g sodium hydroxide (NaOH) and 1.0g cetyl trimethyl ammonium bromide (C
16tAB) be distributed in 480mL deionized water, and at 80 ℃ vigorous stirring 1 hour.Afterwards 5.0mL ethyl orthosilicate (TEOS) is added in above-mentioned mixed liquor, and continue to stir 2 hours at 80 ℃.After reaction finishes, the white depositions obtaining is centrifugal, with a large amount of ethanol, wash, afterwards vacuum lyophilization.Finally, by ion exchange, remove the surfactant C in prepared mesoporous silicon oxide duct
16tAB: the dried sample (0.5g) of above-mentioned acquisition is distributed in the mixed liquor that contains 95% ethanol water and 0.2g ammonium nitrate, mixture is stirred 5 hours at 60 ℃ of lower magnetic forces, product, through centrifugal, alcohol wash, vacuum drying, gets final product to obtain mesoporous silicon dioxide nano particle (MSNs);
Step B) prepare mesoporous silicon dioxide nano particle (MSNs-OOH) that hydrogen peroxide is modified
Utilize the strong oxidizing property of sulphuric acid and hydrogen peroxide mixture, thereby make mesoporous silicon surface that peroxidization occur, obtain mesoporous silicon dioxide nano particle that hydrogen peroxide is modified.In the ice bath of 0 ℃, by by steps A) MSNs that makes is distributed in the hydrogen peroxide mixed liquor of 65% sulphuric acid and 30%.The suspension obtaining, after ultrasonic 30 minutes, is continued to magnetic agitation 4 hours.Obtained suspension is centrifugal, and with a large amount of washings, until the pH of solution reaches 7 left and right, finally by the white depositions room temperature vacuum drying obtaining, can obtain mesoporous silicon dioxide nano particle that hydrogen peroxide is modified, be labeled as MSNs-OOH.
Embodiment 4
Mesoporous silicon dioxide nano particle (MSNs-OO (the CH for preparing tert-butyl hydroperoxide
3)
3) and free-radical generating evaluation
A) prepare mesoporous silicon dioxide nano particle (MSNs)
According to the steps A of embodiment 3) prepare MSNs;
Step B) prepare mesoporous silicon dioxide nano particle (MSNs-Cl) of surface chlorination
There is a large amount of silicon hydroxyls, thionyl chloride (SOCl in mesoporous silicon dioxide nano particle (MSNs) surface
2) be the chlorinating agent of groups such as silicon hydroxyl conventional in organic synthesis.First use SOCl
2mSNs is carried out to chlorination.By steps A) the dry MSNs (2.0g) that makes is distributed to 90mL SOCl
2in 100mL chloroform mixed liquor, after ultrasonic 30 minutes, under magnetic agitation, reflux 24 hours.After reaction finishes, by the product sucking filtration obtaining, with chloroform and acetone, wash one time to remove unnecessary SOCl
2, vacuum drying afterwards, can obtain mesoporous silicon dioxide nano particle (being labeled as MSNs-Cl) of surface chlorination.The MSNs-Cl of gained is transferred to inflated with nitrogen protection in glove box, for follow-up peroxidating, modifies;
Step C) prepare the mesoporous silicon dioxide nano particle (MSNs-OO (CH of surperficial tert-butyl hydroperoxide
3)
3)
By step B) MSNs-Cl that makes is distributed in 90mL dioxane and 12mL tert-butyl hydroperoxide (TBHP) mixed liquor, and adds wherein 0.3g NaHCO
3, reactant mixture stirring reaction 12 hours in dry nitrogen atmosphere at 20 ℃.After reaction finishes, suspension through sucking filtration, methanol wash, afterwards at room temperature vacuum drying, can obtain mesoporous silicon dioxide nano particle of surperficial tert-butyl hydroperoxide, be labeled as MSNs-OO (CH
3)
3;
Step D) evaluate MSNs-OO (CH
3)
3produce the characteristic of free radical
Use fluorescence experiments to detect MSNs-OO (CH
3)
3the free radical generating.In order to detect MSNs-OO (CH
3)
3the characteristic that produces free radical, cell is giving MSNs-OO (CH
3)
3cultivate after 4 hours, we,, to probe 2 ' 7 '-diacetyl dichlorofluorescein (DCFH-DA) that has added a kind of non-fluorescence in cell culture medium, after further cultivating 30 minutes, carry out fluorescence microscope.Experimental result finds that cell is giving MSNs-OO (CH
3)
3after cultivation, occurred obvious green DCF fluorescence, this just illustrates the generation that has oxygen-derived free radicals in incubation.By contrast, in blank group cell, substantially can't see fluorescence.Constructed MSNs-OO (CH has been described
3)
3can effectively realize the release of free radical.
Embodiment 5
Hollow mesoporous silicon dioxide nano particle (ART@HMSNs-Cs-1) of preparation encasement polysaccharide (Cs) and loading arteannuin (ART)
Steps A) prepare hollow mesoporous silicon dioxide nano particle (HMSNs)
According to the steps A of case study on implementation 1) prepare HMSNs;
Step B) arteannuin (ART) is packed into hollow mesoporous silicon dioxide nano particle
Adopt simple physics dipping method to load arteannuin (ART).First by steps A) HMSNs that obtains is distributed in ART alcoholic solution, and under dark surrounds, stirring at normal temperature is 12 hours.Afterwards, the suspension obtaining is centrifugal, with a small amount of washing, remove the ART molecule of surface adsorption, vacuum drying at room temperature, can obtain the HMSNs that has loaded ART afterwards, is labeled as ART HMSNs;
Step C) ART@HMSNs being carried out to surperficial chitosan (Cs) is coated
0.6g chitosan is joined in the acetic acid aqueous solution of 100mL10% to stirring at normal temperature 24 hours, obtains 0.6% chitosan colloidal sol, then with sodium hydroxide by the pH regulator to 6.0 of solution.Then by step B) in the ART@HMSNs that makes be distributed in above-mentioned chitosan colloidal sol, slowly stir 24 hours.Afterwards, centrifugal, wash, be drying to obtain coated mesoporous silicon dioxide nano particle (being labeled as ART@HMSNs-Cs-1) that is loaded with ART of chitosan (Cs).
The HMSNs making and ART HMSNs-Cs-1 are carried out to nitrogen and wash desorption isotherm analysis.Analysis result discovery, simple HMSNs presents typical Langmuir IV type isothermal line, is relatively pressing (P/P
0) between 0.3-0.4, have an obvious capillary tube to shrink.By contrast, after ART loading and further Chitosan-coated, in ART@HMSNs-Cs-1 sample, substantially can't detect the variation of relative pressure.We infer, chitosan molecule may be adsorbed in the surface, duct of mesoporous silicon oxide, and mesopore orbit is got clogged.This has also indirectly proved that Cs is coated in the success of mesoporous silicon nanoparticle surface.
Embodiment 6
The Cytotoxic evaluation of the free radical of preparing hollow mesoporous silicon dioxide nano particle (ART@HMSNs-Cs-2) of keyed jointing chitosan (Cs) and loading arteannuin (ART) and producing
Steps A) and step B) with the steps A of embodiment 2) and step B);
Step C) pack arteannuin (ART) into HMSNs-G
Adopt simple physics dipping method arteannuin (ART) molecular diffusion to be entered to the mesopore orbit of HMSNs-G.By step B) HMSNs-G that obtains is distributed in ART alcoholic solution, and under dark surrounds, stirring at normal temperature is 12 hours.Afterwards, the suspension obtaining is centrifugal, with a small amount of washing, remove the ART molecule of surface adsorption, vacuum drying at room temperature, can obtain the HMSNs-G that has loaded ART afterwards, is labeled as ART HMSNs-G;
Step D) at ART@HMSNs-G surface keyed jointing chitosan (Cs)
ART@HMSNs-G surface keyed jointing a large amount of epoxy alkyls, the epoxy alkyl-amino additive reaction of the amino that utilizes epoxy alkyl and chitosan under acidic catalyst, is keyed to ART@HMSNs-G by Cs surperficial.0.6g chitosan is joined in the acetic acid aqueous solution of 100mL10% to stirring at normal temperature 24 hours, obtains 0.6% chitosan colloidal sol, then with sodium hydroxide by the pH regulator to 5.0 of solution.By step C) the ART@HMSNs-G that obtains is distributed in above-mentioned chitosan colloidal sol, and under dark condition, stirring at normal temperature is 12 hours, and standing 6 hours, the pH to 7.4 of regulator solution afterwards.Through centrifugal, washing, after dry, can obtain surperficial keyed jointing chitosan (Cs) and load the silicon dioxide nanocomposite of ART, be labeled as: ART@HMSNs-Cs-2.
Step e) observe the cytotoxic effect of ART@HMSNs-Cs-2
1. cell culture:
Mankind mastopathy cell ZR75-30 comes from Chinese Academy of Sciences's cell bank, and cultivate in the RPMI-1640 of pH7.4 culture medium, during cultivation, in culture medium, add green grass or young crops-streptomycin mixed liquor of 10% heat-inactivated hyclone (FBS) and 1%, and be placed on and contain 5.0%CO
237 ℃ of constant incubators in cultivate.All cell experiments all carry out at exponential phase cells;
2. pair cell gives ART@HMSNs-Cs-2 cultivation
ZR75-30 cell is after the digestion of 0.25% pancreatin solution is collected, and bed board, in 96 orifice plates, and is containing 5.0%CO
237 ℃ of constant incubators in cultivate.After 24 hours, outwell initial culture medium, and change the new culture medium that contains finite concentration ART@HMSNs-Cs-2 into, and continue to be placed in the time that constant incubator is cultivated appointment.After cultivation finishes, discard the culture medium that contains powder body, with PBS gently wash cell 2 times, use inverted microscope to observe the variation of cellular morphology before and after ART@HMSNs-Cs-2 medication;
The cellular morphology electronic photo result that cell is given after ART@HMSNs-Cs-2 cultivates shows: the ZR75-30 cell of blank group, after 24 hours cultivate, presents good fusiformis pattern and has good adherent property.By contrast, after giving ART@cultivation in HMSNs-Cs-224 hour, cell presents obvious multinucleation phenomenon, and occurs a large amount of vacuoles in cell, and most cells presents circle, and cell adsorptivity reduces.Illustrated that our prepared nano-complex can be successfully by cytophagy, and shown the damaging action to tumor cell.
PH response free radical delivery systme of the present invention can be responsively near tumor tissues (pH=6.5) produce free radical, and in not release of normal structure place (pH=7.4), thereby avoided the damaging action of radical pair normal structure.Therefore, there is great pharmaceutical sector application prospect and value.
Claims (7)
1. a chitosan-modified mesoporous silicon-dioxide-substrate delays controlled-release material, it is characterized in that, described material can load can be produced the compound of free radical or the mesoporous silicon dioxide nano particle of group, load on compound or the group of the free radical in the sub-mesopore orbit of mesoporous silicon dioxide nano particle by mesopore orbit, and the chitosan polyelectrolyte membranes composition that is coated described mesoporous silicon dioxide nano particle.
2. material according to claim 1, is characterized in that, the compound or the group that produce free radical comprise peroxide or the group that contains peroxide bridge.
3. material according to claim 1 and 2, is characterized in that, in the compound of generation free radical or the mesopore orbit that group was loaded into or was connected to mesoporous silicon-dioxide-substrate material and/or mesoporous surface.
4. according to arbitrary described material in claim 1-3, it is characterized in that, free radical refers to peroxide, the inner oxide series of intermediate products in course of reaction, comprises the highly active intermediate product that has that exists and do not exist with radical form with radical form.
5. material according to claim 4, is characterized in that, intermediate product comprises at least one in peroxy radical, alkoxy radical, alkyl diradical, organic hydroperoxide thing.
6. an application for the arbitrary described material of claim 1-5, is characterized in that, described material response pH value is controlled the compound that can produce free radical of material load or the rate of release of group.
7. application according to claim 6, is characterized in that, the compound of the generation free radical of described material load or the release rate of group under pH=6.5 condition are greater than the release rate under pH=7.4 condition.
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| CN112494421A (en) * | 2020-12-23 | 2021-03-16 | 华中科技大学 | Slow-release soluble microneedle, preparation method and application |
| CN114028585A (en) * | 2021-12-14 | 2022-02-11 | 中南大学 | Dihydroartemisinin-loaded nano carrier and preparation method and application thereof |
| CN115506157A (en) * | 2022-10-19 | 2022-12-23 | 卡尔美体育用品有限公司 | Antibacterial warm-keeping down jacket fabric and preparation method thereof |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109276753A (en) * | 2018-11-30 | 2019-01-29 | 上海交通大学医学院附属第九人民医院 | Load the medical titanium alloy bracket and preparation method thereof of the mesopore bioactive glass modification of potassium ferrate |
| CN112494421A (en) * | 2020-12-23 | 2021-03-16 | 华中科技大学 | Slow-release soluble microneedle, preparation method and application |
| CN114028585A (en) * | 2021-12-14 | 2022-02-11 | 中南大学 | Dihydroartemisinin-loaded nano carrier and preparation method and application thereof |
| CN114028585B (en) * | 2021-12-14 | 2023-02-21 | 中南大学 | Dihydroartemisinin-loaded nano carrier and preparation method and application thereof |
| CN115506157A (en) * | 2022-10-19 | 2022-12-23 | 卡尔美体育用品有限公司 | Antibacterial warm-keeping down jacket fabric and preparation method thereof |
| CN115506157B (en) * | 2022-10-19 | 2024-02-13 | 卡尔美体育用品有限公司 | Antibacterial warm down jacket fabric and preparation method thereof |
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