CN107126425A - A kind of tanshinone IIA PEG PLGA PEG nanoparticles and preparation method thereof - Google Patents
A kind of tanshinone IIA PEG PLGA PEG nanoparticles and preparation method thereof Download PDFInfo
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Abstract
The present invention relates to a kind of tanshinone IIA PEG PLGA PEG nanoparticles and preparation method thereof.Carrier material used in the present invention is PEG PLGA PEG triblock copolymer compounds, the medicine of package-contained is tanshinone IIA, the nm of particle size range 100~400 of nanoparticle, the drugloading rate 10%~40% of tanshinone IIA in nanoparticle, envelop rate 50%~90%, the tanshinone IIA PEG PLGA PEG nanoparticles that the present invention is provided, preparation technology is simple, obtained drug-carrying nanometer particle size is homogeneous, and surface is smooth.The drugloading rate and envelop rate to medicine can be effectively improved simultaneously, hydrophily, biocompatibility and the bioavilability of tanshinone IIA can be improved, effective Drug controlled release, extend the circulating half-life of medicine in vivo, for being had very important significance in treatment ischemic cerebral apoplexy with cardiovascular and cerebrovascular disease.
Description
Technical field
The present invention relates to pharmaceutical technology field, it is related to a kind of preparation for containing tanshinone IIA PEG-PLGA-PEG nanoparticles
Method and purposes.
Background technology
Tanshinone IIA（Tanshinone IIA）It is the clinical conventional blood-activating and stasis-removing red sage root for diterpene quinone
One of principle active component, belong to fat-soluble medicine, be soluble in organic solvent, be a kind of natural anti-oxidation medicine.It is red
Ginseng ketone IIA has extensive pharmacological action, and tanshinone IIA can be by protecting vascular endothelial cell, reducing internal lipid peroxidation
The various ways such as matter, immunological regulation, play a part of improving atherosclerosis.Clinical research finds that tanshinone IIA also has
Suppress cardiac muscle cell apoptosis effect, played a key role in the development for the treatment of angiocardiopathy；Tanshinone IIA is not only in treatment
It is widely used in terms of angiocardiopathy, also has for cerebral central nervous system damage caused by hypoxic-ischemic and significantly control
Therapeutic effect.Pharmacological experiment is it has been proved that the anti-inflammatory of tanshinone IIA, suppress Apoptosis, anti-oxidant and scavenging activated oxygen
Effect, can be for treatment cerebral ischemia etc. disease and effect clearly.
Although tanshinone IIA has extensive pharmacological activity in terms of many brain diseases are treated, in medicament research and development
Also many problems are not resolved, such as tanshinone IIA is insoluble in water, and water soluble preparation is made and has difficulties, and blood
Circulating half-life is short, bioavilability is low, across poor permeability of blood-brain barrier etc., therefore limits tanshinone IIA in clinic
Application.At present, structural modification is mainly carried out to tanshinone IIA by sulfonated reaction, is converted into water miscible tanshinone
IIA sulfonic acid sodium injections, but be due to tanshinone IIA sodium sulfonate half-life short in blood, fast is respectively mutually 26 with slow phase
Min and 108 min, it is therefore necessary to use larger dose（160 mg/250 ml/ days）Carry out drip-feed【Shao Hesheng, Jing Xinan,
Yin Lanqin, Gu Jianfeng, Zhao Minmin.The research of distribution excretion and metabolism of the tanshinone IIA sodium sulfonate in rat body,《Chinese patent drug
Research》1979,2：8-12】.In addition, sulfonate sodium is ionic compound, it is more difficult to by using lipid and protein component as base
The biofilm structure system of plinth so that the utilization rate of tanshinone IIA is not high, considerably reduces the therapeutic effect of tanshinone IIA,
And side effect of the repetitively administered to normal organ tissue is larger.
With the fast development of nanometer technology, diameter is less than having been obtained extensively in biomedical sector for 100 nm nano materials
General application, especially in fields such as medical imaging, medical diagnosis on disease, drug targeting are delivered, cancer is precisely treated, gene transfections.
The treatment of cranial vascular disease, due to the obstruction of blood-brain barrier, about 98% hydrophilic small molecules and most macromoleculars are all
It can not pass through, make drug molecule be difficult to reach affected area, limit the clinical efficacy to cranial vascular disease.And nano-medicament carrier
Due to the characteristics such as excellent biocompatibility, modifiability, one of the focus studied as brain targeting drug delivery system.Research hair
It is existing, chemotherapeutic agent, polypeptide, pharmaceutical grade protein are loaded using nano medicament carrying system, slow-release controlled-release and target administration can be realized
Purpose so that this kind of medicine will not be degraded and remove in a short period of time, so as to reduce drug administration by injection frequency, improve and suffer from
The compliance of person simultaneously, by it is selectively targeted it is ligand modified after, can carrying medicaments molecule pass through blood-brain barrier, significantly improve medicine
Thing reaches the purpose of precisely treatment brain diseases in the valid density at cerebral tissue change position.Reduce other non lesion positions
Drug concentration, reduces the toxic side effect to body.
Poly(D,L-lactide-co-glycolide (poly (lactic-co-glycolic acid), PLGA) is by lactic acid
A kind of function that (lactic acid, LA) and hydroxyacetic acid (glycolic acid, GA) are polymerized according to different proportion
Property macromolecule organic material, passes through food and medicine Surveillance Authority (FDA) certification in the U.S..At present, have been widely used for
Prepare artificial conduit, slow releasing carrier of medication (microballoon, nanoparticle, micropill), the system such as implants and film【Acharya S,
Sahoo S K. PLGA nanoparticles containing various anticancer agents and tumour
delivery by EPR effect. Adv Drug Deliv Rev. 2011, 63 (3): 170-183.；Chen Y C,
Liu D Z, Liu J J, et al. Development of terbinafine solid lipid nanoparticles
as a topical delivery system. Int J Nanomedicine. 2012, 7: 4409-4418.】.As receiving
Rice pharmaceutical carrier PLGA has advantages below：1. it is polymerized, is synthesized by lactic acid (LA) and hydroxyacetic acid (or glycolic, GA)
Technical maturity；2. there is controllable particle diameter, nanoparticle decentralization is small；3. there is good biological degradability and bio-compatible
Property, non-immunogenicity；PLGA copolymers catabolite final in human body is carbon dioxide (CO2) and water (H2O), and
It can be excreted by normally giving birth to old metabolism, therefore there is no excitant and toxicity to the body of people.4. medicine can be realized
Long-time sustained release（Several weeks or several months）, medicine frequency is reduced, patient's compliance is improved；5. PLGA nanometers after modification
Grain, suitable for embedding bioactive molecule such as albumen, gene DNA, vaccine etc.；6. after PLGA surface modification aglucons, medicine can be achieved
The targeted delivery of thing【Cho H J, Yoon I S, Yoon H Y, et al. Polyethylene glycol-
conjugated hyaluronic acid-ceramide self-assembled nanoparticles for targeted
delivery of doxorubicin. Biomaterials. 2012, 33 (4): 1190-1200.；Collins M
NBirkinshaw C. Hyaluronic acid based scaffolds for tissue engineering-A
review. Carbohydr Polym. 2013, 92 (2): 1262-1279.】.Utilize nano-carrier PEG-PLGA-PEG
Tanshinone IIA is contained, the low problem of hydrophily, biocompatibility, bioavilability of tanshinone IIA can be improved.Nanoparticle
The hydrophily on surface will influence the speed that phagocyte swallows to it with lipophilicity, thus extend tanshinone IIA in vivo
Blood circulatory half-life need to increase the hydrophily on PLGA surfaces.Because PLGA block crystallinity is stronger, and there is hydrophobicity, so easily
Removed by RES, it is difficult to which medicine is reached into targeting moiety.And PEG-PLGA-PEG triblock copolymerization thing has hydrophily, and it is biological
Compatibility is good, non-toxic, antigenicity and immunogenicity, soluble in water and many organic solvents.By with PLGA copolymerizations
PEG-PLGA-PEG, can combine PEG and PLGA advantage, effectively delay RES removing, extension body circulation time.
The content of the invention
For the technical problem of above-mentioned tanshinone IIA conventional formulation, the red sage root is contained it is an object of the invention to provide one kind
The preparation method of ketone IIA PEG-PLGA-PEG nanoparticles, this method process is simple, favorable reproducibility.What profit was prepared in this way
Tanshinone IIA PEG-PLGA-PEG nanoparticles, size is homogeneous, good with obvious chondritic, stability, drugloading rate and bag
Envelope rate is higher, good dispersion, and the circulating half-life of tanshinone IIA drug-carrying nanometer particle in vivo is obviously prolonged, and is difficult clear by RES
Remove.
The present invention provides one kind and contains tanshinone IIA PEG-PLGA-PEG nanoparticles：It is to use triblock copolymer PEG-
PLGA-PEG contains lipophilic drugs tanshinone IIA, after freeze-drying, obtains that property is stable, size is homogeneous, and add water redissolution
The good drug-carrying nanometer particle of dissolubility afterwards.
Described triblock copolymer PEG-PLGA-PEG nano carrier materials, its molecular weight is 10000~60000；Its
In, PLGA content is 40%~96%, and molecular weight is 2000~30000, the ratio of glycolide and lactide during synthesis PLGA
Example is 1/100~99/100；Wherein PEG content is 4%~60%, and molecular weight is 1000~5000.
The mass fraction of described tanshinone IIA is 80%~98%；The type of emulsifying agent is：Tween 80, polyvinyl alcohol
（PVA）, PLURONICS F87, soybean lecithin or lecithin one or more of mixture therein, in interior aqueous phase and outer aqueous phase
Mass fraction be 0.1%~10%；The scope of described nanometer particle size is the load of tanshinone IIA in 100~400 nm, nanoparticle
Dose 10%~35%, envelop rate 50%~90%.
A kind of preparation method for containing tanshinone IIA PEG-PLGA-PEG nanoparticles, technique is as follows:
（1）Tanshinone IIA and PEG-PLGA-PEG are dissolved in mixed organic solvents as oil phase, by emulsifying agent be dissolved in from
Sub- water is used as interior aqueous phase；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In interior aqueous phase be slowly dropped in organic phase,
It is ultrasonically formed colostrum；
（3）Emulsifying agent is dissolved in deionized water and is used as outer aqueous phase（Emulsifying agent used is same in outer aqueous phase and interior aqueous phase
Emulsifying agent）, under conditions of ice-water bath stirring, use syringe aspiration step（2）In colostrum, deeply outer aqueous phase liquid level under,
Ultrasonic vibration formation emulsion is slowly added dropwise；
（4）The organic solvent in emulsion is removed by the way of stirring, it is anti-with deionized water by nanometer suspension solution high speed centrifugation
Ultrasonic disperse is cleaned again, is freeze-dried with after filtering with microporous membrane, you can obtain carrying red sage root IIA PEG-PLGA-PEG nanometers
Grain.
Step（1）The amount of the middle tanshinone IIA for adding prescribed dose is 1mg/ml~20mg/ml, and mixing used is organic
Solvent is any two kinds of mixing in dichloromethane, chloroform, acetone, ethyl acetate, absolute ethyl alcohol, and the ratio of mixing is 1:1
~1:20, the PEG-PLGA-PEG of addition amount are the mg/ml of 0.5 mg/ml~50.
Step（3）The ratio of middle colostrum and outer aqueous phase is 1：1~1:30, the speed of dropwise addition is 10~30 drops/per minute, is stirred
The speed mixed is the r/min of 100 r/min~700.
In a kind of described preparation method for containing tanshinone IIA PEG-PLGA-PEG nanoparticles, ultrasonic power is
100 w~500 w, ultrasonic mode is 10 s of s intervals of ultrasonic 2 s~60~30 s, the min of continuous ultrasound 5~40.
In a kind of described preparation method for containing tanshinone IIA PEG-PLGA-PEG nanoparticles, stirring volatilization is organic molten
The time of agent is 4~6 h, and ultracentrifugal rotating speed used is the r/min of 10000 r/min~16000, and centrifugation time is 20
The min of min~60.
It is an advantage of the invention that：
1. prepared by the present invention contains tanshinone IIA, the carrier material diblock copolymer PEG-PLGA-PEG of use is a kind of
Toxicity is low, cost is low, the preferable high polymer material of biological degradability, when the PLGA in copolymer degrades in human body, degraded production
Thing may participate in metabolism, and ultimately form carbon dioxide (CO2) and water (H2), O it will not assemble in vivo.Therefore, bio-compatible
Good, the non-immunogenicity of property, it is safe；
2. hydroxyacetic acid in PLGA in carrier material（GA）Ratio with lactic acid (LA) is 1/100~99/100, and degradation capability reaches
To most preferably can slowly degrade, control medicine slowly discharges, and can give full play to the pharmacological action of tanshinone IIA；PEG hydrophilic radicals
Opsonic action of the blood to nanoparticle can be reduced, so as to extend the blood halflife of medicine；And without using secondary solvent, can be with
Avoid producing other normal structure organs side effect, convenient drug administration increases patient adaptability；
3.PEG-PLGA-PEG is amphipathic nature polyalcohol, and its nanoparticle constituted can effectively be dissolved in water after freeze-drying
Solution, its inner hydrophobic region can provide space for tanshinone IIA, improve the dissolubility of tanshinone IIA, change it in body
Interior distribution, improves bioavilability.
Brief description of the drawings
Fig. 1 is the tanshinone IIA PEG-PLGA-PEG nanoparticle scanning electron microscope (SEM) photographs that embodiment 1 is prepared.
Fig. 2 is the tanshinone IIA PEG-PLGA-PEG nanoparticle particle size distribution figures that embodiment 1 is prepared.
Fig. 3 is the high-efficient liquid phase chromatogram of tanshinone IIA.
Fig. 4 is the tanshinone IIA PEG-PLGA-PEG nanoparticle In-vitro release curves that embodiment 1 is prepared.
Fig. 5 is the tanshinone IIA PEG-PLGA-PEG nanoparticles group that embodiment 1 is prepared and clinical tanshinone IIA note
Penetrate the blood concentration-time curve of agent group.
Embodiment
The invention will be further described with reference to embodiments, but the invention is not limited in these embodiments.
Embodiment 1 contains the preparation of tanshinone IIA PEG-PLGA-PEG nanoparticles
（1）By 2mg tanshinone IIAs and 30 mg PEG-PLGA-PEG ultrasonic dissolutions in 2 ml acetone and dichloromethane（Acetone：Two
Chloromethanes=1:9）In the mixed solvent as oil phase, prepare mass fraction and be used as water for 0.5% poloxamer F188 solution
Phase；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In the μ L of interior aqueous phase 30 with 10 ~ 30 drop/it is per minute
Speed is added drop-wise in organic phase, ultrasonic 5 s intervals 5 s, continuous ultrasound 10min formation colostrum；
（3）Under conditions of ice-water bath stirring, syringe aspiration step is used（2）In colostrum, by the way of being added dropwise under liquid level,
It is added drop-wise to 10 ~ 30 drops/speed per minute in the poloxamer F188 solution of 20 ml 0.5%（Colostrum emulsion ratio is 1:10）, ultrasound
5 s interval 5 s, the min of continuous ultrasound 20 formation emulsions；
（4）5h is stirred at room temperature, the organic solvent in emulsion is removed, and the min of 15000 r/min high speed centrifugations 30 outwells supernatant
Liquid, the nanometer suspension solution for carrying tanshinone can be obtained by cleaning ultrasonic disperse repeatedly with deionized water, with 0.22 μm of miillpore filter
The medicine not wrapped up is filtered to remove, is freeze-dried, you can obtains carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles, measures
The average grain diameter of nanoparticle is 158nm, and envelop rate is 90%, and drugloading rate is 35%.
Embodiment 2 contains the preparation of tanshinone IIA PEG-PLGA-PEG nanoparticles
（1）By 4mg tanshinone IIAs and 30 mg PEG-PLGA-PEG ultrasonic dissolutions in 2 ml acetone and dichloromethane（Acetone：Two
Chloromethanes=1:8）In the mixed solvent as oil phase, prepare mass fraction and be used as aqueous phase for 0.5% Tween-80 solution；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In the μ L of interior aqueous phase 30 with 10 ~ 30 drop/it is per minute
Speed is added drop-wise in organic phase, ultrasonic 5 s intervals 5 s, continuous ultrasound 10min formation colostrum；
（3）Under conditions of ice-water bath stirring, syringe aspiration step is used（2）In colostrum, by the way of being added dropwise under liquid level,
It is added drop-wise to 10 ~ 30 drops/speed per minute in the Tween-80 solution of 20 ml 0.5%, 5 s of ultrasonic 5 s intervals, continuous ultrasound 20
Min formation emulsions；
（4）5h is stirred at room temperature, the organic solvent in emulsion is removed, and the min of 15000 r/min high speed centrifugations 30 outwells supernatant
Liquid, the nanometer suspension solution for carrying tanshinone can be obtained by cleaning ultrasonic disperse repeatedly with deionized water, with 0.22 μm of miillpore filter
The medicine not wrapped up is filtered to remove, is freeze-dried, you can carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles are obtained.Measure
Average grain diameter is 180nm, and drugloading rate is 27%, and envelop rate is 81%.
Embodiment 3 contains the preparation of tanshinone IIA PEG-PLGA-PEG nanoparticles
（1）By 8mg tanshinone IIAs and 30 mg PEG-PLGA-PEG ultrasonic dissolutions in 2 ml acetone and dichloromethane（Acetone：Two
Chloromethanes=1:8）In the mixed solvent as oil phase, prepare mass fraction and be used as interior aqueous phase for 0.5% PVA solution；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In 30 μ L interior aqueous phase with 10 ~ 30 drop/it is per minute
Speed is added drop-wise in organic phase, ultrasonic 5 s intervals 5 s, continuous ultrasound 10min formation colostrum；
（3）Under conditions of ice-water bath stirring, syringe aspiration step is used（2）In colostrum, by the way of being added dropwise under liquid level,
It is added drop-wise to 10 ~ 30 drops/speed per minute in the PVA solutions of 20 ml 0.5%（Colostrum emulsion ratio is 1:10）, ultrasonic 5 s intervals 5
S, the min of continuous ultrasound 20 formation emulsion；
（4）5h is stirred at room temperature, the organic solvent in emulsion is removed, and the min of 15000 r/min high speed centrifugations 30 outwells supernatant
Liquid, the nanometer suspension solution for carrying tanshinone can be obtained by cleaning ultrasonic disperse repeatedly with deionized water, with 0.22 μm of miillpore filter
The medicine not wrapped up is filtered to remove, is freeze-dried, you can obtains carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles, measures
The average grain diameter of nanoparticle is 210nm, and drugloading rate is 17%, and envelop rate is 74%.
Embodiment 4 contains the preparation of tanshinone IIA PEG-PLGA-PEG nanoparticles
（1）By 8mg tanshinone IIAs and 40 mg PEG-PLGA-PEG ultrasonic dissolutions in 2 ml ethyl acetate and dichloromethane（Second
Acetoacetic ester：Dichloromethane=1:8）In the mixed solvent as oil phase, prepare mass fraction and be used as aqueous phase for 1% PVA solution；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In the μ L of interior aqueous phase 30 with 10 ~ 30 drop/it is per minute
Speed is added drop-wise in organic phase, ultrasonic 5 s intervals 5 s, continuous ultrasound 10min formation colostrum；
（3）Under conditions of ice-water bath stirring, syringe aspiration step is used（2）In colostrum, by the way of being added dropwise under liquid level,
It is added drop-wise to 10 ~ 30 drops/speed per minute in 30 ml % PVA solution（Colostrum emulsion ratio is 1:20）, ultrasonic 5 s intervals 5
S, the min of continuous ultrasound 20 formation emulsion；
（4）5h is stirred at room temperature, the organic solvent in emulsion is removed, and the min of 15000 r/min high speed centrifugations 30 outwells supernatant
Liquid, the nanometer suspension solution for carrying tanshinone can be obtained by cleaning ultrasonic disperse repeatedly with deionized water, use 0.45um miillpore filters
The medicine not wrapped up is filtered to remove, is freeze-dried, you can obtains carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles, measures
The average grain diameter of nanoparticle is 240nm, and drugloading rate is 12%, and envelop rate is 61%.
Embodiment 5 contains the preparation of tanshinone IIA PEG-PLGA-PEG nanoparticles
（1）By 2mg tanshinone IIAs and 20 mg PEG-PLGA-PEG ultrasonic dissolutions in 2 ml absolute ethyl alcohols and dichloromethane（Nothing
Water-ethanol：Dichloromethane=1:10）In the mixed solvent as oil phase, prepare mass fraction and make for 1.0% Tween-80 solution
For aqueous phase；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In the μ L of interior aqueous phase 30 with 10 ~ 30 drop/it is per minute
Speed is added drop-wise in organic phase, ultrasonic 5 s intervals 5 s, continuous ultrasound 10min formation colostrum；
（3）Under conditions of ice-water bath stirring, syringe aspiration step is used（2）In colostrum, by the way of being added dropwise under liquid level,
It is added drop-wise to 10 ~ 30 drops/speed per minute in 1.0% Tween-80 solution（Colostrum emulsion ratio is 1:5）, ultrasonic 5 s intervals 5
S, the min of continuous ultrasound 20 formation emulsion；
（4）5h is stirred at room temperature, the organic solvent in emulsion is removed, and the min of 15000 r/min high speed centrifugations 30 outwells supernatant
Liquid, the nanometer suspension solution for carrying tanshinone can be obtained by cleaning ultrasonic disperse repeatedly with deionized water, with 0.22 μm of miillpore filter
The medicine not wrapped up is filtered to remove, is freeze-dried, you can obtaining carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles will survey
The average grain diameter for obtaining nanoparticle is 196nm, and drugloading rate is 20%, and envelop rate is 71%.
Embodiment 6 contains the preparation of tanshinone IIA PEG-PLGA-PEG nanoparticles
（1）By 10mg tanshinone IIAs and 60 mg PEG-PLGA-PEG ultrasonic dissolutions in 2 ml acetone and dichloromethane（Acetone：
Dichloromethane=1:5）In the mixed solvent as oil phase, prepare mass fraction and be used as aqueous phase for 1.5% Tween-80 solution；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In the μ L of interior aqueous phase 30 with 10 ~ 30 drop/it is per minute
Speed is added drop-wise in organic phase, ultrasonic 5 s intervals 5 s, continuous ultrasound 10min formation colostrum；
（3）Under conditions of ice-water bath stirring, syringe aspiration step is used（2）In colostrum, by the way of being added dropwise under liquid level,
It is added drop-wise to 10 ~ 30 drops/speed per minute in 40ml 1.5% Tween-80 solution（Colostrum emulsion ratio is 1:20）, ultrasonic 5 s
Intermittently 5 s, the min of continuous ultrasound 20 formation emulsion；
（4）5h is stirred at room temperature, the organic solvent in emulsion is removed, and the min of 15000 r/min high speed centrifugations 30 outwells supernatant
Liquid, the nanometer suspension solution for carrying tanshinone can be obtained by cleaning ultrasonic disperse repeatedly with deionized water, with 0.22 μm of miillpore filter
The medicine not wrapped up is filtered to remove, is freeze-dried, you can obtaining carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles will survey
It is 260 nm to obtain average grain diameter, and drugloading rate is 10%, and envelop rate is 53%.
The drugloading rate of the tanshinone IIA PEG-PLGA-PEG nanoparticles of embodiment 7 and the measure of envelop rate
The content of tanshinone IIA is determined using high performance liquid chromatography:Chromatographic column：Waters C18 chromatographic columns（250 mm ×4.6
Mm, 5 μm）, mobile phase：Methanol：Water=75: 25（v/v）Column temperature：25 DEG C of flow velocitys：1.0 ml/min ultraviolet detection wavelength：268
~270 nm, sample size：20 μL；
The tanshinone IIA standard solution that concentration is 1,5,10.0,25.0,50,80,100 μ g/ml is taken respectively, according to chromatostrip
Part is tested, and tanshinone IIA concentration is fitted with peak area, regression equation is set up.Precision weighs 2 mg freezed
Tanshinone IIA PEG-PLGA-PEG nanoparticles, are placed in 10 ml volumetric flasks, with shaking up constant volume after methanol dilution.0.22 μm micro-
After the membrane filtration of hole, the 20 μ L every time of sample introduction 3 times.Detected with HPLC, determine peak area, according to standard curve regression equation, calculated
The content of tanshinone IIA., can (tanshinone IIA high-efficient liquid phase chromatogram be such as in the hope of drugloading rate and envelop rate according to below equation
Shown in Fig. 3), envelop rate %=（Total dose of contained dose/input in nano-carrier）× 100%, drugloading rate %=（Nanometer is carried
Total dose of contained dose/input in body）×100%.
Embodiment 8:Tanshinone IIA PEG-PLGA-PEG nanoparticles mode of appearance, zate current potentials, particle size and distribution
Measure.
Embodiment 9:The mode of appearance of nanoparticle (prepared by embodiment 2) is observed using SEM, such as Fig. 1 institutes
Show, the particle diameter distribution and zeta current potentials of nanoparticle can be determined using laser particle size analyzer, as shown in Figure 2.
The measure of the tanshinone IIA PEG-PLGA-PEG nanoparticle tablets in vitro of embodiment 10.
Tanshinone IIA PEG-PLGA-PEG nanoparticles after precision weighing tanshinone IIA bulk drug and freeze-drying are each
8mg, is respectively placed in the PBS of the mol/L of 5 mL 0.01 pH=7.4, places into 50 mL centrifuge tube, be immediately placed on
In 37 ± 1 DEG C of water bath with thermostatic control shaking tables, 100 r/min are at the uniform velocity stirred, and keep sink conditions.Respectively at 0,15min, 30min,
The mL of point in time sampling 1 as defined in 1h, 2h, 4h, 8h, 12h, 24h, 48h, 72h, while supplementing the fresh medium of equivalent isothermal.
After sample centrifuges 25 min through 15 000 r/min, supernatant is taken out with after 0.22 μm of filtering with microporous membrane, the first for the amount of doubling
The μ L of sample introduction 20 after alcohol dilution, record peak area, substitute into standard curve, calculate the concentration liquid of tanshinone IIA in solution, computational rules
Medicine in time adds up release rate.Each 3 parts of sample operation repetitive, draws external with the average value of preparation to the time
Cumulative release profile（As shown in Figure 4）.
Embodiment 11:Tanshinone IIA PEG-PLGA-PEG nanoparticle rat Internal pharmacokinetics property is investigated.
Using healthy male SD rat, two groups are randomly divided into（n = 10）, every group 5, blank tanshinone IIA（10mg /
kg）It is suspended in carboxymethylcellulose sodium solution, PEG-PLGA-PEG nanoparticles (contain TIIA 10mg/kg, embodiment 1
Prepare) it is scattered in physiological saline, respectively by tail vein injection, respectively at after administration（0,5,15,30 and 45 minute; 1,
2,4,6,8,12nd, 24 and 36 hours）, blood sample is collected by tail vein（0.5ml）, 4000r/min centrifugation l0min, take supernatant (blood
200 μ L clearly), add 500 μ L methanol, and vortex 3min, 7000r/min centrifugation l0min takes supernatant to cross 0.22 μm of micropore filter
Film, using the concentration of tanshinone IIA in Syrups by HPLC filtrate, calculates the concentration of tanshinone IIA in blood, draws PEG-
PLGA-PEG nanoparticles group and the clinical time graph (as shown in Figure 5) of tanshinone IIA injection group blood concentration one.
Claims (10)
1. a kind of tanshinone IIA PEG-PLGA-PEG nanoparticles, it is characterised in that：Described nanoparticle comprising tanshinone IIA,
PEG-PLGA-PEG triblock copolymerization compound, emulsifying agent, water for injection.
2. a kind of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 1, it is characterised in that be to use PEG-
PLGA-PEG triblock copolymers compound loads lipophilic drugs tanshinone IIA, described PEG-PLGA- as carrier material
The molecular weight of PEG triblock copolymer compounds is 10000~60000；Wherein, PLGA content is 40%~96%, and molecular weight is
The ratio of glycolide and lactide is 1/100~99/100 during 2000~30000, synthesis PLGA；Wherein PEG content is
4%~60%, molecular weight is 1000~5000.
3. a kind of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 1, it is characterised in that described one
Tanshinone IIA PEG-PLGA-PEG nanoparticles are planted, the purity of tanshinone IIA used is 80%~98%；Described nanometer particle size
Scope be 100~400 nm, the drugloading rate 10%~35% of tanshinone IIA, envelop rate 50%~90% in nanoparticle.
4. a kind of preparation method of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 1, its feature exists
In:（1）Tanshinone IIA and PEG-PLGA-PEG are dissolved in mixed organic solvents as oil phase, by emulsifying agent be dissolved in from
Sub- water is used as interior aqueous phase；
（2）Under conditions of ice-water bath stirring, syringe aspiration step is used（1）In interior aqueous phase be slowly dropped in organic phase,
It is ultrasonically formed colostrum；
（3）Emulsifying agent is dissolved in deionized water and is used as outer aqueous phase（Emulsifying agent used is same in outer aqueous phase and interior aqueous phase
Emulsifying agent）, under conditions of ice-water bath stirring, use syringe aspiration step（2）In colostrum, deeply outer aqueous phase liquid level under,
It is slowly added dropwise and is ultrasonically formed emulsion；
（4）Stirring removes the organic solvent in emulsion, and by nanometer suspension solution high speed centrifugation, ultrasound is cleaned repeatedly with deionized water
It is scattered, it is freeze-dried with after filtering with microporous membrane, you can obtain carrying red sage root ketone IIA PEG-PLGA-PEG nanoparticles.
5. a kind of preparation method of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 3, its feature exists
In:Step（1）The amount of the middle tanshinone IIA for adding prescribed dose is 1mg/ml~20mg/ml, mixed organic solvents used
For dichloromethane, chloroform, acetone, ethyl acetate, any two kinds of mixing in absolute ethyl alcohol, the ratio of mixing is 1:1~1:
20, the PEG-PLGA-PEG of addition amount are the mg/ml of 0.5 mg/ml~50.
6. a kind of preparation method of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 3, its feature exists
In:Emulsifier type used is Tween-80, polyvinyl alcohol（PVA）, PLURONICS F87, soybean lecithin or lecithin are wherein
One or more of mixtures, in interior aqueous phase and outer aqueous phase the mass fraction of emulsifying agent be 0.1%~10%.
7. a kind of preparation method of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 3, its feature exists
In:Step（2）In in the ratio of aqueous phase and organic phase be 1：1~1:30, the speed of dropwise addition is 10~30 drops/per minute, stirring
Speed be the r/min of 100 r/min~700.
8. a kind of preparation method of tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 3, its feature exists
In:Step（3）The ratio of middle colostrum and outer aqueous phase is 1：1~1:30, the speed of dropwise addition is 10~30 drops/per minute, stirring
Speed is the r/min of 100 r/min~700.
9. a kind of preparation method for containing tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 3, it is special
Levy and be:The power of ultrasound is the w of 100 w~500, and ultrasonic mode is the s of 10 s of s intervals of ultrasonic 2 s~60~30, continuously
5~40 min of ultrasound.
10. a kind of preparation method for containing tanshinone IIA PEG-PLGA-PEG nanoparticles according to claim 3, it is special
Levy and be:The time for stirring volatile organic solvent is 4~6 h；Ultracentrifugal rotating speed used is 10000 r/min~16000
R/min, centrifugation time is the min of 20 min~60；Miillpore filter used is 0.22 μm.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108159396A (en) * | 2018-01-26 | 2018-06-15 | 广州加原医药科技有限公司 | A kind of phycocyanin nanometer formulation and preparation method thereof |
| CN109091464A (en) * | 2018-07-24 | 2018-12-28 | 华南理工大学 | A kind of preparation method of the amphiphilic dispersed nano pharmaceutical carrier of PLGA-PEG |
| EA032386B1 (en) * | 2018-03-19 | 2019-05-31 | Общество С Ограниченной Ответственностью "Марлин Биотех" | Method for delivery of nuclear acids to the heart by means of plga nanoparticles |
| CN114941182A (en) * | 2022-05-25 | 2022-08-26 | 华南理工大学 | PEG-PLGA block copolymer self-assembly nanofiber and preparation method thereof |
Citations (3)
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| CN1608675A (en) * | 2003-10-22 | 2005-04-27 | 四川大学 | Medicine-carrying nanometer polymer particle and its prepn and use |
| CN101810862A (en) * | 2009-02-20 | 2010-08-25 | 北京大学 | Tanshinone IIA oral administration polymer micelle composition |
| CN103083250A (en) * | 2013-01-22 | 2013-05-08 | 上海中医药大学附属曙光医院 | Tanshinone IIA-polyactic acid/hydroxyacetic acid microsphere and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1608675A (en) * | 2003-10-22 | 2005-04-27 | 四川大学 | Medicine-carrying nanometer polymer particle and its prepn and use |
| CN101810862A (en) * | 2009-02-20 | 2010-08-25 | 北京大学 | Tanshinone IIA oral administration polymer micelle composition |
| CN103083250A (en) * | 2013-01-22 | 2013-05-08 | 上海中医药大学附属曙光医院 | Tanshinone IIA-polyactic acid/hydroxyacetic acid microsphere and preparation method thereof |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108159396A (en) * | 2018-01-26 | 2018-06-15 | 广州加原医药科技有限公司 | A kind of phycocyanin nanometer formulation and preparation method thereof |
| EA032386B1 (en) * | 2018-03-19 | 2019-05-31 | Общество С Ограниченной Ответственностью "Марлин Биотех" | Method for delivery of nuclear acids to the heart by means of plga nanoparticles |
| CN109091464A (en) * | 2018-07-24 | 2018-12-28 | 华南理工大学 | A kind of preparation method of the amphiphilic dispersed nano pharmaceutical carrier of PLGA-PEG |
| CN114941182A (en) * | 2022-05-25 | 2022-08-26 | 华南理工大学 | PEG-PLGA block copolymer self-assembly nanofiber and preparation method thereof |
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