CN110051649A - Supercritical CO2The method that expansion solutions crystallization prepares lipid nano particle - Google Patents

Supercritical CO2The method that expansion solutions crystallization prepares lipid nano particle Download PDF

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CN110051649A
CN110051649A CN201810057007.5A CN201810057007A CN110051649A CN 110051649 A CN110051649 A CN 110051649A CN 201810057007 A CN201810057007 A CN 201810057007A CN 110051649 A CN110051649 A CN 110051649A
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lipid
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文震
何红霞
王帅
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Shenzhen University
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    • 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
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    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
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Abstract

The present invention provides a kind of supercritical CO2The method that expansion solutions crystallization prepares lipid nano particle, the method comprising the steps of: the lipid, stabilizer, model drug of formula ratio being dissolved in solvent and form lipid soln, is passed through supercritical CO2Lipid soln is expanded under the conditions of the pressure of setting, temperature, is quickly sprayed into aqueous media by nozzle, forms lipid nano particle suspension, CO through crystallisation by cooling2The residual solvent in lipid nano particle suspension is dissolved and removed, lipid nano particle is obtained.The crystal form feature and transformation of crystal that this method passes through control lipid, the crystallization process for inhibiting lipid to change from alpha-crystal form to beta crystal, the drug generated by lipid transformation of crystal is efficiently avoided to squeeze out and nanoparticle reuniting effect, be conducive to improve nanoparticle load drug stabilisation, and nanoscale is kept to be uniformly distributed, it is supercritical CO2Lipid nano particle assembling provides a new way in medium.

Description

Supercritical CO2The method that expansion solutions crystallization prepares lipid nano particle
Technical field
The invention belongs to field of chemical engineering, and in particular to supercritical CO2Expansion solutions crystallization prepares lipid nano particle Method.
Background technique
Lipid nano particle (solid lipid nanosparticles, SLNs) is meant with biodegradable high-melting fat Matter is nanoparticle made of framework material, is extremely promising newtype drug transmitting carrier.Lipid nano particle the advantage is that Toxicity is low, good biocompatibility, preparation safety, reduces or reduce the damage of drug normal tissue, is particularly suitable for drug administration by injection.
But there is also defects for lipid nano particle: lipid is in crystallization process, perfection of the lipid experience from α-crystal form to-beta crystal Crystallization process is easy to squeeze out drug from lattice, causes to carry medicine instability problem.Meanwhile lipid is during transformation of crystal It generates lipid sur to expose, mutually fusion is easy between lipid nanometer, causes agglomeration, generate size distribution unevenness, carry medicine The problems such as low, stability is poor is measured, application of the lipid nano particle in pharmaceutical carrier is limited.
The preparation method of lipid nano particle has film dispersion method, film-ultrasonic dispersion, fusion method, freeze-thaw method, injection Method, reverse phase evaporation, multi-emulsion method etc., and traditional preparation methods are difficult to control accurately the crystallization process of lipid.Therefore, one kind is found It is capable of the new method of accuracy controlling lipid nano particle crystallization, inhibits conversion of the lipid from alpha-crystal form to beta crystal, is to improve lipid to receive The load drug stabilisation of the grain of rice, and keep the equally distributed key of nanoscale.
Supercritical fluid is a kind of environmental-friendly medium, has been used for the preparation such as liposome.Chinese patent " a kind of nanometer of leaf Sour liposome and its supercritical carbon dioxide preparation method " (CN201310727241.1), " supercritical carbon dioxide prepares nanometer The method of arbutin liposome " (CN201310607897.X), takes supercritical extract/incubation method to prepare liposome, emphasis It is that liposomal particle size and dissolvent residual control.Inventor is prepared for atractylone rouge using improved super-critical rapid expansion technology Plastid (atractylone lipidosome and preparation method thereof, CN 20091018872.6), overcritical microemulsion technology is prepared for nano-lipid Body (method that supercritical fluid CO 2 prepares nano liposomes, CN 201010297180.6).
Lipid nano particle is different from liposome property, and the cerium sulphate crystal of lipid nano particle is the pharmaceutically stable for determining nanoparticle The key of property.On the basis of early-stage study, inventor is by CO2Expansion solutions combine with lipid, are lipid nano particle Assembling provide a new way.Detailed process are as follows: by CO2It is passed through in lipid soln, expands lipid soln, destroy lipid Lattice arrangement reduces lipid crystallinity, promotes lipid crystal form amorphization.Supercritical liq is after rapid expanding depressurizes, then leads to It crosses nozzle atomization to be injected into aqueous media, lipid crystallisation solidification, obtains the lipid nano particle of good dispersion, even particle size distribution Son.
Compare conventional method, supercritical CO2The advantage that crystallization prepares lipid nano particle is the fine of lipid crystallization process Regulation.By adjusting supercritical CO2Parameter controls the expansion process of lipid, further controls the crystal form feature and crystal form of lipid Conversion inhibits crystallization process from alpha-crystal form to beta crystal that change from of lipid, efficiently avoids generating because of lipid transformation of crystal Drug is squeezed out with nanoparticle reuniting effect, is conducive to improve nanoparticle load drug stabilisation, and nano particle size is kept to be uniformly distributed.From And it is supercritical CO2Lipid nano particle assembling in medium provides a new way, novelty with technology and application and It is creative.
Summary of the invention
It is an object of that present invention to provide a kind of methods for preparing lipid nano particle.
Purpose according to the present invention, specifically, this method utilizes supercritical CO2Lipid soln is expanded, inhibits lipid by α The crystallization process that crystal form changes to beta crystal avoids the drug generated by lipid transformation of crystal extrusion and nanoparticle reuniting effect, Drug discharges and nanoparticle reunion shortcoming suddenly in lipid nano particle to solve prior art preparation.System of the present invention The method of standby lipid nano particle, comprising the following steps:
1) lipid of formula ratio, stabilizer, model drug are dissolved in solvent and form lipid soln, be placed at overcritical In reaction kettle, it is passed through supercritical CO2It is expanded under the conditions of the pressure of setting, temperature；
2) by supercritical CO2The lipid soln of expansion is quickly sprayed by nozzle to overcritical precipitating kettle by certain speed In interior aqueous media, lipid nano particle suspension is formed through crystallisation by cooling；
3) by CO2Overcritical precipitating kettle is continued through, the residual solvent in lipid nano particle suspension is dissolved and remove, is received Collect lipid nano particle.
The lipid is glycerin monostearate, and the stabilizer is phosphatidyl choline；The solvent is ethyl alcohol；The mould Type drug is resveratrol.
The lipid soln formula ratio are as follows: the mass percent 0.80% of lipid, the mass percent 0.20% of stabilizer, Model drug mass percent 0.10%.
The bulbs of pressure are 8~14MPa, preferably 10~13MPa.
The expansion temperature is 35~65 DEG C, preferably 45~60 DEG C.
The overcritical lipid soln is sprayed to overcritical precipitating kettle with the flow velocity of 5L/min.
Pressure is 5MPa in the precipitating kettle, and the aqueous media temperature is 2 DEG C, volume 50mL, and stirring rate is 1000r/min。
The supercritical CO2Flow velocity is 2L/min.
The lipid nano particle encapsulation rate 80% or so prepared under the preferred processing condition, drugloading rate are put down 7% or so Equal granularity 100nm or so, narrow particle size distribution and it is uniform.
Compared with prior art, the features and advantages of the invention are:
The application is by CO2Expansion lipid soln prepares lipid nano particle, is compared to conventional method advantage: (1) adjusting Supercritical CO2Parameter controls the expansion process of lipid, further controls the crystal form feature and transformation of crystal of lipid, realizes lipid The precise controlling of nanoparticle crystallization is conducive to improve load drug stabilisation；(2) the rapid expanding mass transfer characteristics of supercritical fluid (such as high diffusivity degree and low viscosity) is conducive to nanoscale, the dispersion of even-grained lipid nano particle, precipitation and collection；3) rouge Matter nanoparticle is assembled in supercritical CO2It is completed in medium, the solvent used is with CO2It flows out, no solvent residue in nanoparticle, tool There is environmental-friendly feature.
Detailed description of the invention
Fig. 1 display present invention prepares a kind of flow chart of embodiment of the method for solid lipid nano granule, wherein 1-CO2 Steel cylinder；2- cold；3- high-pressure pump 1；4- reaction kettle；5- nozzle；6- precipitating kettle；7,9- collector；8- separating still；10- flow Meter；V1-V10- valve；H1-H4- heat exchanger；
Fig. 2 shows the surface topography scanning electron microscope (SEM) photograph (SEM) of lipid nano particle prepared according to the methods of the invention, wherein 1- cream Change/solvent evaporated method preparation lipid nano particle (EE-RES-SLNs)；Lipid nano particle (the SC- of 2- super-critical crystallization preparation RES-SLNs)；
Fig. 3 shows microstructure transmission electron microscope (TEM) figure of lipid nano particle prepared according to the methods of the invention；Wherein 1- cream Change/solvent evaporated method preparation lipid nano particle (EE-RES-SLNs)；Lipid nano particle (the SC- of 2- super-critical crystallization preparation RES-SLNs)；
Fig. 4 shows the particle size distribution figure of lipid nano particle prepared according to the methods of the invention；Wherein 1- emulsification/solvent evaporated method The lipid nano particle (EE-RES-SLNs) of preparation；The lipid nano particle (SC-RES-SLNs) of 2- super-critical crystallization preparation；
Fig. 5 shows the X-ray diffracting spectrum (XRD) of lipid nano particle prepared according to the methods of the invention；The wherein white Chenopodiaceae of 1- Reed alcohol (RES)；2- glycerin monostearate (GMS)；3- phosphatidyl choline (PC)；4- physical mixture (PMIX)；5- emulsification/molten The resveratrol lipid nano particle (EE-RES-SLNs) of agent evaporation preparation；The resveratrol lipid of 6- super-critical crystallization preparation is received The grain of rice (SC-RES-SLNs)
Fig. 6 show lipid nano particle prepared according to the methods of the invention X-ray diffracting spectrum (angle of diffraction: 15~ 30°)；Wherein 1- glycerin monostearate (GMS)；2- physical mixture (PMIX)；The white black false hellebore of 3- emulsification/solvent evaporation preparation Alcohol lipid nano particle (EE-RES-SLNs)；The resveratrol lipid nano particle (SC-RES-SLNs) of 4- super-critical crystallization preparation
Specific embodiment
To better understand the essence of the present invention, following embodiment is used for the technology contents that the present invention will be described in detail, but It should not be construed as limiting the scope of the invention.
The preparation and measurement of 1 lipid nano particle of embodiment
1. the preparation of lipid nano particle
Experimental provision: supercritical reaction apparatus is manufactured and designed, reaction kettle by South China Science & Engineering University's modern chemical industry technique center Capacity is 100cm3, collection kettle capacity is 200cm3, maximum receiving pressure 35MPa, maximum temperature 368K.
Preparation method: in one embodiment, the method that the present invention prepares lipid nano particle can be in supercritical CO2Instead It answers in device and completes, flow chart is as shown in Figure 1.Specific steps are as follows: by 80mg tristerin, 80mg phosphatidyl choline, 10mg resveratrol is dissolved in 11.3mL ethyl alcohol, forms lipid ethanol solution.Lipid soln is injected into CO2Fluid reaction kettle 4 In, sealed cans.
Open CO2Steel cylinder, CO2Gas enters reaction kettle 4 after being pressurizeed by high-pressure metering pump 3 after cold 2 is cooled to liquid, The temperature of reaction kettle is controlled by heat exchanger H2.Close valve V3, Open valve V2, CO2Into reaction kettle gas circuit, in the bulbs of pressure 12MPa, expansion temperature are to be incubated for 1h to lipid soln at 55 DEG C.Quick release reactor pressure, by CO2The lipid soln of expansion With CO2Precipitating kettle 6, the crystallization of lipid quick emulsification are injected into the flow velocity of 5L/min through nozzle 5, emulsion droplet is formed by curing in ice water Lipid nano particle suspension.The pressure of precipitating kettle 6 is set as 5MPa, and aqueous media temperature is 2 DEG C, volume 50mL, stirring speed Rate is 1000r/min.After course of injection, 5MPa pressure is kept to continue to be passed through into deposition kettle with the flow velocity of 2L/min CO2, make the ethyl alcohol in nanoparticle suspension with CO2Precipitating kettle is flowed out, parses and recycles into separating still 8.1h is extracted, keeps ethyl alcohol complete It is complete to remove.CO2And after the metering of spinner flowmeter 10, CO is reentered2Gas circuit is recycled later through cold 2 is cooling.Reaction After the completion, lipid nano particle suspension made of being collected in collector 7.
Lipid nano particle is prepared using classical emulsification/solvent evaporation simultaneously.The lipid soln 15mL with formula is taken, with note Lipid soln is slowly injected into the deionized water stirred at 50mL 1000r/min by about 1 drop/5s speed by emitter dropwise In.Continue to stir water phase under 75 DEG C of constant temperature to ethanol evaporation, and water phase is concentrated to 20mL, obtains translucent nanoemulsion.It will It is cooled and solidified in the ice water that nanoemulsion stirs under being rapidly added to 30mL 1000r/min, continues to stir 2h in 2 DEG C of constant temperature, i.e., Obtain lipid nano particle suspension.
2. the drug carrying ability of solid lipid nano granule measures
Resveratrol gross mass measures in lipid nano particle: utilizing ALPHA1-2 Lpplus freeze drier (Germany Christ company) by lipid nano particle suspension, for 24 hours, lipid is made in vacuum freeze drying under the conditions of cryogenic temperature is -50 DEG C Nanoparticle powder.The above-mentioned lipid nano particle powder of 10mg is taken, is dispersed in the nanoparticle that 1mg/mL is made in 10mL distilled water again Suspension.It takes 1mL nanosuspension to be placed in 50mL volumetric flask, the ethanol solution demulsification of 10mL20%TritonX-100 is added, And scale is settled to dehydrated alcohol.Using RF-5000 sepectrophotofluorometer (Japanese Shimadzu Corporation) three-dimensional fluorescence spectrum method Resveratrol gross mass is measured, determination condition is excitation wavelength lambda ex=290nm and emission wavelength lambda em=380nm.
Wherein WAlwaysIt is in lipid nano particle comprising resveratrol gross mass (mg), WIt is freeNot to be encapsulated in lipid nano particle Free resveratrol quality (mg), WNanoparticleIt is lipid nano particle quality
3. dissolvent residual detects
Using Agilent 7694E headspace sampling/6890 gas chromatographic detection residual ethanol solvent of Agilent.5mL lipid is received Grain of rice suspension is placed in sample injection bottle, is placed in head-space sampler, with N after the ethyl alcohol volatilization in sample2Into chromatography. Head space condition: 80 DEG C of equilibrium temperature, equilibration time 30min, sampling volume 2mL.GC conditions: Agilent DB-624 (30m × 0.32mm × 1.80 μm) chromatographic column, 180 DEG C of injector temperature, 250 DEG C of fid detector temperature, chromatographic column program liter Temperature: 40 DEG C (10min) → (50 DEG C/min) → 120 DEG C (5min), N2Flow velocity 2.0mL/min.
4. the characterization of solid lipid nano granule
Microstructure characterization method: taking lipid nano particle suspension 1 to drip, and drop is on copper platform after 100 times of dilution, vacuum refrigeration Metal spraying is conductive after drying, utilizes Hitachis-3400N scanning electron microscope (SEM) (Hitachi, Japan) scanning electron microscope pair Lipid nano particle configuration of surface is observed and is taken pictures.Lipid nano particle 1 is taken to drip simultaneously, drop is on copper mesh after 100 times of dilution, benefit Lipid nano particle micromorphology is observed and taken pictures with JEM-1230 transmission electron microscope (TEM) (Japan Electronics Corporation).
Particle size distribution measuring: taking lipid nano particle one to drip, and after 100 times of dilution, Malven Zetasizer Nano ZS is dynamic The particle diameter distribution and polydispersity coefficient of state laser nano particles distribution instrument (Malven instrument company, Britain) measurement lipid nano particle (PDI)。
5. data processing
Data statistic analysis is carried out to data using SPSS software, data are the average value of three groups of parallel laboratory tests, by mean value ± standard deviation (Means ± SD) indicates.Using the significant difference between t check analysis data group, P < 0.05: the level of signifiance, P < 0.01: extremely significant level.
6 embodiment results
It is as shown in table 1 by lipid nano particle physicochemical property manufactured in the present embodiment.
The lipid nano particle physicochemical property of 1 distinct methods of table preparation
*P < 0.05 is compared, significant difference with emulsification/solvent evaporation group
**P < 0.01 is compared, extremely significant sex differernce with emulsification/solvent evaporation group
Emulsification/solvent evaporation preparation lipid nano particle Residual ethanol is 0.22 ± 0.03%, and super-critical crystallization system It is standby to completely remove dissolvent residual in nanoparticle.Super-critical crystallization and emulsification/solvent evaporate the nanoparticle of two kinds of distinct methods preparation Drug carrying ability and size distribution have significant difference (p < 0.01and p < 0.05), super-critical crystallization preparation nanometer Grain encapsulation rate reaches 84.71 ± 0.89%, and drugloading rate is 7.91 ± 0.08%, and average grain diameter 106.51 ± 1.06nm, PDI are 0.146±0.009。
The surface topography and content microstructure of the prepared lipid nano particle of super-critical crystallization, emulsification/solvent evaporation and As shown in Fig. 2 (SEM), Fig. 4 (TEM).The lipid nano particle granular size of emulsion dispersion preparation is uneven, and agglomeration.It is super In a highly dispersed state, the allusion quotation of lipid nano particle is presented in clear smooth spherical morphology to the lipid nano particle of critical crystal preparation Type microscopic feature.
Super-critical crystallization, emulsification/solvent evaporation prepared lipid nano particle partial size and size distribution are as shown in Figure 4. Emulsification/solvent evaporation preparation nanoparticle average grain diameter is larger (178.64 ± 3.69nm), particle diameter distribution not enough uniformly (PDI 0.362 ± 0.031), and there is 5 μm or so of larger particle, display portion nanoparticle has been reunited.Super-critical crystallization preparation Nanoparticle average grain diameter at 100nm or so (106.51 ± 1.06nm, p < 0.01), particle diameter distribution is in unimodal normal distribution (PDI 0.163 ± 0.014, p < 0.05), display nanoparticle are uniformly distributed in nanoscale.
The crystalline characteristics of 2 lipid nano particle of embodiment
The present embodiment is investigated super-critical crystallization process and is controlled the crystal form of lipid nano particle.
Using D8ADVANCE X-ray diffraction analyzer (XRD, German Bruker company) to the white black false hellebore of composition prescription Alcohol (RES), glycerin monostearate (GMS), soybean lecithin (PC), resveratrol/glycerin monostearate/soybean lecithin Physical mixture (PMIX), emulsification described in embodiment 1/solvent evaporation preparation lipid nano particle (EE-RES-SLNs), super Lipid nano particle prepared by critical crystal (SC-RES-SLNs) sample is analyzed.The above-mentioned sample of 10mg respectively, in high pressure 50KV, Guan Liu 1000A, scanning speed are 2 ° of min-1, scanning range (2 θ) is 5~60 °, carries out XRD determining.
The present embodiment is to the Crystalline form analysis of lipid nano particle as shown in figure 5, wherein 15~30 ° of area diffraction feature such as Fig. 6 It is shown.There are α-crystalline state, β '-crystalline state, β-crystalline state polytropism for lipid.5.72 ° of GMS diffracted absorption peak, 7.54 °, At 19.56 °, 20.52 °, 22.92 °, 23.72 °, 19.56 °, 22.92 ° be beta crystal characteristic diffraction peak, 20.52 °, 23.72 ° For β ' crystal form characteristic diffraction peak, show that lipid using beta crystal as primary crystallization, and contains a small amount of β ' crystal form.EE-RES-SLNs exists 19.56 ° and 22.92 ° (beta-crystal), 20.52 ° of diffracted absorption peak positions and tristerin with 23.72 ° (β '-crystal form) Similar, the total diffracted intensity of crystal form weakens, and shows that RES-SLNs lipid includes that β and β ' crystal form crystallize.Super-critical crystallization preparation is received The total diffracted intensity of grain of rice SC-RES-SLNs further declines, and shows that becoming for unsettingization (amorphization) occurs in lipid Gesture.In addition to there is the characteristic absorption peak of β and β ' crystal form, occurs α-state crystal form characteristic peak at 21.16 °, show some lipid With α-state crystal form crystallization.
In addition, resveratrol characteristic diffraction peak be (6.66 °, 13.26 °, 16.26 °, 19.16 °, 20.20 °, 22.24 °, 23.47 °, 28.22 °), there is the X- characteristic diffraction peak of lipid, resveratrol in physical mixture, shows that drug is not supported on In lipid.After forming SC-RES-FLNs, resveratrol X- characteristic diffraction peak disappears, and shows that drug is complete in the form of amorphous state It is supported in lipid nano particle.
Super-critical crystallization utilizes CO2High osmosis, expand lipid soln, and quick release pressure, the bulbs of pressure and heavy Barometric gradient between analysis pressure leads to larger degree of supersaturation, destroys the lattice of lipid beta crystal neatly arranged, and lipid tends to Irregular lattice arrangement, produces lattice defect.Lipid lattice diffracted intensity is significantly lower, promotes lipid that unsettingization is presented (amorphization) trend.Lattice defect and its α-state crystal form characteristic peak occur, and show lipid from α-crystal form to β-crystalline substance The perfect crystallization process of type conversion is suppressed, and lipid mixes crystal form crystallization with α, β with β ', is avoided lipid perfection crystallization and is made At drug be extruded the defect and lipid interface fusion phenomenon of lattice, be conducive to improve and carry drug stabilisation, and keep nanometer Size distribution.
Influence of 3 bulbs of pressure of embodiment to lipid nano particle drug carrying ability
The present embodiment investigates influence of the bulbs of pressure to the drug carrying ability of lipid nano particle.Controlling expansion temperature is 50 DEG C, Lipid is expanded in the case where being pre-expanded 8~14 MPa of pressure.Lipid soln is with CO2It is injected into buffer solution with 5L/min, collects institute The lipid nano particle suspension of preparation.Lipid nano particle encapsulation rate and drugloading rate are measured by 1 method of embodiment, as a result such as 1 institute of table Show.
Lipid nano particle drug carrying ability under the different bulbs of pressure of table 1
The load performance of lipid nano particle depends on the interaction of two factors: the degrees of expansion and drug of lipid drop In CO2Middle solubility.As pressure lower (8~12MPa), lipid drop degrees of expansion increases, and penetrates into lipid lattice ability and increases By force, the deformation extent of lattice increases, and the lattice defect space for accommodating drug increases, and drugloading rate and encapsulation rate improve.And pressure liter Height is to 13MPa or more, and resveratrol is in CO2Middle solubility increases, and more free drugs are dissolved in CO2In, rather than it is dispersed in rouge In matter carrier, drug carrying ability is caused to decline instead.Therefore, there are one compared with ratio of greater inequality example for the bulbs of pressure, when the bulbs of pressure are 10~ 12MP can effectively improve drug carrying ability.
4 bulbs of pressure of embodiment influence the size distribution of lipid nano particle
This example investigates influence of the bulbs of pressure to the size distribution of lipid nano particle.Using the identical parameter of embodiment 3, press 1 method of embodiment detects the size distribution of lipid nano particle, and the results are shown in Table 2.
This example demonstrates that the volume of lipid drop increases with the increase of the bulbs of pressure.When quick injection releases stress When, under the barometric gradient between the bulbs of pressure and precipitating pressure, the lipid drop of expansion is shrunk to smaller particle automatically To balance inside and outside differential pressure.So droplet retracts degree is bigger when barometric gradient is bigger, i.e., precipitating pressure fixing when, the bulbs of pressure Higher, partial size is smaller.But with pressure rise to 13MPa or more, this influence tends to become smaller.It is examined in conjunction with drug carrying ability Consider, the bulbs of pressure form there are a more excellent section, i.e. 10~13MPa of the bulbs of pressure lipid nano particle, and lipid nano particle carries Pharmacological property can be higher, and average grain diameter is smaller.
Influence of 5 expansion temperature of embodiment to lipid nano particle drug carrying ability
Lipid nano particle drug carrying ability under the different expansion temperatures of table 3
This example investigates influence of the expansion temperature to lipid nano particle drug carrying ability.The control bulbs of pressure are 12MPa, are being expanded Lipid is expanded at 35~65 DEG C of temperature.Lipid soln is with CO2It is injected into buffer solution with 5L/min, collects prepared lipid Nanoparticle suspension.Lipid nano particle encapsulation rate and drugloading rate are measured by the method for embodiment, the results are shown in Table 3.
This example explanation, lipid nano particle encapsulation rate and drugloading rate increase with temperature and are increased.When temperature be lower than 40 DEG C, rouge Matter lattice arrangement is more close, and small amount resveratrol is supported in lipid lattice.When temperature continues to increase, lipid expands, rouge Mass flow denaturation increases, and lattice arrangement is more loose, can accommodate more drug, shows as drug carrying ability raising.And temperature increases To 60 DEG C (close to GMS fusing point), lipid lattice deformability degree is smaller, and the capacity for loading resveratrol increases trend and slows down.Therefore, It is preferred that expansion temperature is 45~65 DEG C, nanoparticle load performance is higher.
Influence of 6 expansion temperature of embodiment to the size distribution of lipid nano particle
This example investigates influence of the expansion temperature to lipid nano particle size distribution.Using the identical parameter of embodiment 5 and side Method, measurement prepare lipid nano particle size distribution, and the results are shown in Table 4.
Lipid nano particle granularity under the different expansion temperatures of table 4
Temperature depends on lipid drop degrees of expansion to grain diameter influence and particle encounter interacts.When temperature 35~55 DEG C, influence of the temperature to lipid liquid drip expansion is occupied an leading position, and as temperature increases, lipid fluidity increases, lattice arrangement ratio It is more at random, lipid crystallization is increased with α-crystal form crystal area proportion, the perfect crystallization process that lipid is converted from α-crystal form to beta-crystal by To inhibition, lipid particle caused by the exposure of the nanoparticle lipid sur as caused by crystal transfer mutually merges effect and is suppressed, Be conducive to reduce particle diameter.And when temperature is higher than 60 DEG C, lipid nano particle between particle caused by molecular thermalmotion because colliding Probability enhancing, influence have been more than the influence of lipid crystal transfer, and the lipid particle of formation mutually collides and forms aggregate, nanometer The partial size and PDI of particle increase.Comprehensively consider influence of the temperature to drug carrying ability of embodiment 5, preferable temperature is 45~60 DEG C.
Examples 1 to 7 shows supercritical CO2Expansion lipid soln crystallization technique provides a kind of lipid nano particle assembling Method realizes the finely regulating to lipid crystallization process, the crystallization process for inhibiting lipid to change from alpha-crystal form to beta crystal, effectively Ground avoids the drug extrusion generated by lipid transformation of crystal and nanoparticle reuniting effect, and it is steady to be conducive to raising nanoparticle load medicine It is qualitative, and nano particle size is kept to be uniformly distributed.Totally apparently, in 10~13MPa of the selected bulbs of pressure, expansion temperature is 45~60 At DEG C, the nanoparticle encapsulation rate 80% or so of super-critical crystallization preparation, average particle size 100nm or so, particle diameter distribution is uniform, reaches Quality standard of the Chinese Pharmacopoeia about Nano medication.

Claims (8)

1. a kind of supercritical CO2The method that expansion solutions crystallization prepares lipid nano particle, the method comprising the steps of:
1) lipid of formula ratio, stabilizer, model drug are dissolved in solvent and form lipid soln, be placed at supercritical reaction In kettle, it is passed through supercritical CO2It is expanded under the conditions of the pressure of setting, temperature；
2) by supercritical CO2The lipid soln of expansion is quickly sprayed by nozzle to the water in overcritical precipitating kettle by certain speed In phase medium, lipid nano particle suspension is formed through crystallisation by cooling；
3) by CO2Overcritical precipitating kettle is continued through, the residual solvent in lipid nano particle suspension is dissolved and remove, collects rouge Matter nanoparticle.
2. the stabilizer is phosphatidyl according to the method described in claim 1, wherein the lipid is glycerin monostearate Choline；The model drug is resveratrol, and the solvent is ethyl alcohol.
3. according to the method described in claim 1, the lipid soln formula ratio are as follows: the mass percent 0.80% of lipid, surely Determine the mass percent 0.20% of agent, model drug mass percent 0.10%.
4. according to the method described in claim 1, the bulbs of pressure are 8~14MPa, preferably 10~13MPa.
5. according to the method described in claim 1, the expansion temperature is 35~65 DEG C, preferably 45~60 DEG C.
6. according to the method described in claim 1, the overcritical lipid soln is sprayed with the flow velocity of 5L/min to described heavy It analyses in kettle.
7. pressure is 5MPa in the precipitating kettle, the aqueous media temperature is 2 DEG C, volume 50mL, and stirring rate is 1000r/min。
8. the supercritical CO2Flow velocity is 2L/min.
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