CN102807677A - CA-PLGA-TPGS (Cholic Acid-Poly-Lactic-co-Glycolic Acid copolymer-Tocopherol Polyethylene Glycol Succinate) star copolymer as well as preparation method and application thereof - Google Patents
CA-PLGA-TPGS (Cholic Acid-Poly-Lactic-co-Glycolic Acid copolymer-Tocopherol Polyethylene Glycol Succinate) star copolymer as well as preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a CA-PLGA-TPGS (Cholic Acid-Poly-Lactic-co-Glycolic Acid copolymer-Tocopherol Polyethylene Glycol Succinate) star copolymer as well as a preparation method and an application thereof. A structural formula of the CA-PLGA-TPGS star segmented copolymer is as shown in a formula 1. The preparation method comprises the steps as follows: 1) preparing a CA-PLGA; 2) carrying out carboxylation on a TPGS; and 3) preparing the CA-PLGA-TPGS star copolymer. The CA-PLGA-TPGS star copolymer has the advantages of good biocompatibility and biodegradability and is applied to the field of pharmaceutical preparations and tissue engineering, in particular to the field of pharmaceutical preparations for anti-tumor treatment or anti-cardiovascular diseases.
Description
Technical field
The present invention relates to a kind of CA-PLGA-TPGS star copolymer and preparation method thereof and application.
Background technology
Biodegradable polymer material since its in vivo excellent biological compatibility and degradability as drug administration system carrier be widely used in target slow-release, controlled release is studied.Poly (glycolide-lactide) multipolymer (PLGA) is a main solid support material of getting permission the slow controlled release medicament of clinical application at present in the world; Obtain by rac-Lactide and NSC 403079 copolymerization according to a certain ratio; It has the advantage of two kinds of materials concurrently, can well remedy the deficiency of two kinds of materials simultaneously again.PLGA has excellent biological compatibility, biodegradability and tensile strength, in human non-toxic, do not have accumulation, so it has very tempting application prospect and high commercial value at biomedical sector.
Vitamin E TPGS (TPGS) is the abbreviation of polyoxyethylene glycol VE-succinate (D-α-tocopherol polyethylene glycol 1000succinate); It is the soluble derivative of vitamin E; Carboxyl and polyoxyethylene glycol esterification by VE-succinate (TOS) form; Molecular structure is as follows, loaded " USP ".
TPGS has been widely used in pharmaceutical prepn, the foods and cosmetics research; TPGS is flaxen waxy solid; Intimate tasteless, it is a kind of amphiphile, amphiphilic molecule, and a hydrophilic polar head and a hydrophobic aliphatic carbon chain afterbody molecule are arranged; Can be water-soluble, also can be dissolved in most of polar organic solvents.Cholic acid (Cholic Acid; CA) be a kind of main elementary bile acide; It is as emulsifying agent biology synthetic biomolecules on mammiferous lung in the digestive process of fat; But have a hard steroid skeleton structure and four reactive groups, have good Bc in vivo.Its molecular structural formula is as follows:
Than single linear polymeric, the high molecular polymer of the design shape improvement that brought performances such as many physics, chemistry, biology in advance such as star-like grade.Compare with the linear polymeric of same molecular amount, star-like polymer has littler hydrodynamic volume, and the viscosity in solution is little, in blood, is difficult for being blocked, and medicine is had higher encapsulation rate and drug loading.(Mohammad R N, et al.Polymer 52 (2011) 2799 and Giovanni M, et al.Macromol.Rapid Commun.25 (2004) 1139).
Summary of the invention
The purpose of this invention is to provide a kind of star-like biodegradable CA-PLGA-TPGS star copolymer and preparation method thereof.
CA-PLGA-TPGS star copolymer provided by the present invention, its structural formula is suc as formula shown in the I:
Wherein: m=5-260, n=1-200.
TPGS described in the formula I represents the group shown in the formula II:
Wherein: p=23-115.
Prepare the method for above-mentioned CA-PLGA-TPGS star copolymer, comprise the steps:
(1) preparation of CA-PLGA: according to feed molar per-cent; With the lactide monomer of 2%-95%, the glycolide monomer of 2%-95% and the cholic acid initiator of 0.1%-20% is raw material; Under the condition of anhydrous and oxygen-free and catalyzer existence, carry out polyreaction, obtain the CA-PLGA multipolymer;
(2) TPGS's is carboxylated: according to feed molar per-cent; With the TPGS (polyoxyethylene glycol VE-succinate) of 5%-90%, the Succinic anhydried (SA) of 5%-90% or the catalyzer of maleic anhydride and 5%-50% is that raw material reacts; Reaction finishes the back reaction solution that obtains is precipitated with precipitation agent; Filter, will precipitate drying, promptly obtain carboxylated polyoxyethylene glycol VE-succinate (CTPGS);
(3) preparation of CA-PLGA-TPGS star-type polymer: CA-PLGA, CTPGS and dehydrating condensation agent are reacted under the effect of catalyzer; Reaction finishes the back reaction solution that obtains is precipitated with precipitation agent; Filter; To precipitate drying, promptly obtain star-like amphipathic nature block polymer CA-PLGA-TPGS; The mol ratio of said CTPGS and CA-PLGA is 1-15.
Wherein, catalyzer described in the step 1) is selected from following at least a: stannous octoate, stannous iso caprylate, organic guanidine (like acetic acid tetramethyl-dibutyl guanidine), metallic zinc, tributyltin chloride, ferric acetyl acetonade, zinc lactate, nano zine oxide, taurine, ethanol iron, n-propyl alcohol iron, Virahol iron and propyl carbinol iron.Said catalyst consumption is the 0.01%-1% of said monomer molar total amount.
In the step 1), the temperature of reaction of said polyreaction is 120-180 ℃, and the reaction times is 6-24 hour.The number-average molecular weight Mn of the CA-PLGA multipolymer that is obtained is 3000-50000.
Step 2) catalyzer described in be selected from following a) and b) in any one or a) and b) according to mol ratio (0.1-2): 1 mixture formed: a) pyridine, 2-picoline, 4-picoline or 4-Dimethylamino pyridine (DMAP); B) triethylamine quadrol, triethylene diamine or tetrem alkene triamine.Step 2) reaction solvent of reaction can be dioxane, methylene dichloride, toluene, THF or N, dinethylformamide (DMF) etc. described in.Quantity of solvent is 2-20 a times of reactant quality.
Step 2) temperature of reaction of reaction is 5-60 ℃ described in, reacts to be 8-48 hour.Said precipitation agent can be ether, normal heptane, sherwood oil or their any mixture.Carry out drying under the vacuum condition to being deposited in, the exsiccant temperature is 30-50 ℃, and the time is 12-48h.
In the step 3), said dehydrating condensation agent can be N, and N-NSC 57182 (DCC) or 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide (EDC), its add-on are 1-15 times of CA-PLGA molar weight.
Said catalyzer is pyridine, 2-picoline, 4-picoline or 4-Dimethylamino pyridine (DMAP), and its add-on is 0.01-10 a times of CA-PLGA molar weight.
The reaction solvent that reacts described in the step 3) can be methylene dichloride (DCM), toluene, N, and dinethylformamide (DMF), dioxane or THF, its consumption are 2-20 times of reactant quality.
The temperature of reaction of reacting described in the step 3) is 5-60 ℃, reacts to be 8-48 hour.Said precipitation agent can be ether, normal heptane, sherwood oil, methyl alcohol, ethanol, the perhaps mixed solvent of these solvents.Carry out drying under the vacuum condition to being deposited in, the exsiccant temperature is 30-50 ℃, and the time is 12-48h.
The molecular weight of the star-like amphipathic nature block polymer CA-PLGA-TPGS that step 3) obtained is 5000-80000.
A further object of the present invention provides the application of above-mentioned CA-PLGA-TPGS star copolymer.
This CA-PLGA-TPGS star copolymer can be used as pharmaceutical excipient on the one hand and in pharmaceutical formulations (like drug-carrying nanometer particle, medicine carrying gel and medicine carrying microballoons etc.), uses; Also can be used as tissue engineering material on the other hand and be applied to field of tissue engineering technology (for example artificial organs, sutures and intravascular stent etc.).
CA-PLGA-TPGS star copolymer of the present invention also is specially adapted to the pharmaceutical excipient of antitumor drug and cardiovascular disease resistant pharmaceutical prepn.
The method that the present invention prepares the CA-PLGA-TPGS star copolymer is simple, and is pollution-free.The multipolymer that is obtained has excellent biological compatibility, biodegradability, is a kind of up-and-coming material.
Description of drawings
Fig. 1 is the synthetic synoptic diagram of star-like amphipathic nature block polymer CA-PLGA-TPGS.
Fig. 2 be multipolymer CA-PLGA and CA-PLGA-TPGS nucleus magnetic resonance figure (
1HNMR).
Fig. 3 is the gel permeation chromatography figure (GPC) of multipolymer CA-PLGA and CA-PLGA-TPGS.
Fig. 4 is the thermogravimetric analysis curve (TGA) of multipolymer CA-PLGA, CTPGS, CA-PLGA-TPGS.
Fig. 5 is field emission scanning electron microscope collection of illustrative plates (FESEM) collection of illustrative plates of carrying docetaxel CA-PLGA-TPGS nanoparticle.
Fig. 6 detects the particle diameter and the size distribution result of medicine carrying CA-PLGA-TPGS nanoparticle for laser particle analyzer.
Fig. 7 measures the result of medicine carrying CA-PLGA-TPGS nanoparticle for zeta potential instrument.
Fig. 8 characterizes the result of Docetaxel crystal and carrying docetaxel PLGA and CA-PLGA-TPGS nanoparticle for DSC.
Fig. 9 is the vitro drug release curve of carrying docetaxel PLGA and CA-PLGA-TPGS nanoparticle.
Figure 10 is carrying docetaxel CA-PLGA-TPGS nanoparticle and the cell viability experimental result of blank CA-PLGA-TPGS nanoparticle (the identical nanoparticle suspension concentration with drug-carrying nanometer particle) to the MCF-7 cell.
Figure 11 is carry the MCF-7 cell that tonka bean camphor-6 nanoparticle 37 ℃ under hatch 4 hour of laser confocal scanning electron microscope observation with the preparation of CA-PLGA-TPGS copolymer material.Nucleus is dyed blueness with DAPI, and it is green carrying tonka bean camphor-6 nano particle, and respectively through EGFP passage and DAPI passage observation of cell picked-up situation: figure A is the situation of observing through the DAPI passage; Figure B is the situation of observing through the EGFP passage; Figure C is through the EGFP passage with through the result after the doubling of the image of DAPI passage observation.
Figure 12 is for carrying the Electronic Speculum figure of taxol CA-PLGA-TPGS microball prepn.
Embodiment
The present invention will be described through specific embodiment below, but the present invention is not limited thereto.
Experimental technique described in the following embodiment like no specified otherwise, is ordinary method; Said reagent and material like no specified otherwise, all can obtain from commercial sources.
The preparation method of CA-PLGA-TPGS star copolymer comprises the steps:
(1) by the mole percentage composition get 95% lactide monomer, 4% glycolide monomer and 1% cholic acid (CA) is raw material, puts into polymerizing pipe, adds 0.5% acetic acid tetramethyl-dibutyl guanidine of monomer molar amount; Vacuumize inflated with nitrogen, triplicate; Polymerizing pipe tube sealing when vacuum tightness is higher than 70pa 145 ℃ of heating, reacted 12 hours; Promptly obtaining number-average molecular weight is 32000 (m=140, multipolymer CA-PLGA n=8).
(2) get 30% TPGS (Mn=1500) by mole per-cent, 40% Succinic anhydried (SA), 30% catalyzer 4-Dimethylamino pyridine (DMAP) and triethylamine, wherein the molar ratio of DMAP and triethylamine is 1: 1.Add the solvent dioxane that is equivalent to 10 times of quality of raw material; 30 ℃ of temperature of reaction; React and after 24 hours be precipitated out product, precipitation agent is that ether and methyl alcohol volume ratio are 5: 1 mixed solvent, filters; 40 ℃ of vacuum-drying 24h will be deposited in, carboxylated polyethylene glycol 1000 vitamin E succinic acid ester (CTPGS) can be obtained.
(3) preparation of CA-PLGA-TPGS star-type polymer.Reactant CA-PLGA and CTPGS, the molar ratio of CTPGS are 5 times of CA-PLGA; N, N-NSC 57182 (DCC) is a dehydrating condensation agent, add-on is 3 times of molar weight of CA-PLGA; The 4-Dimethylamino pyridine is a catalyzer, and catalytic amount is 0.2 times of molar weight of CA-PLGA.The methylene dichloride that is equivalent to 10 times of quality of raw material is a reaction solvent.30 ℃ of temperature of reaction; Behind the reaction 24h be precipitated out product, precipitation agent is that sherwood oil and methyl alcohol volume ratio are 2: 1 mixed solvent, filters; To be deposited in 40 ℃ of vacuum-drying 24h; Can obtain molecular weight is 36580 (m=140, n=8, star-like amphipathic nature block polymer CA-PLGA-TPGS p=23).
The preparation method of CA-PLGA-TPGS star copolymer comprises the steps:
(1) take by weighing the lactide monomer of 17.28g (0.12mol), glycolide monomer and 0.54 (0.0013mol) cholic acid (CA) of 4.64g (0.04mol) is a raw material, puts into polymerizing pipe, add 1-2 and drip stannous iso caprylate as catalyzer; Vacuumize inflated with nitrogen, triplicate; With alcohol blast burner to the polymerizing pipe tube sealing; 145 ℃ of reacting by heating 12 hours, promptly obtaining number-average molecular weight was 21307 (m=78, multipolymer CA-PLGA n=26).
(2) by quality be the TPGS (Mn=1500) of 3g (2mmol), 0.26g (2.6mmol) Succinic anhydried, 0.348g (2.8mmol) 4-Dimethylamino pyridine (DMAP) and 0.256g (2.5mmol) triethylamine.50ml (51g) dioxane is as reaction solvent, and 30 ℃ of temperature of reaction are reacted and after 24 hours are precipitated out product; Precipitation agent is an ether; Filter, will be deposited in 40 ℃ of vacuum-drying 24h, can obtain carboxylated polyethylene glycol 1000 vitamin E succinic acid ester (CTPGS).
(3) preparation of CA-PLGA-TPGS star-type polymer.Reactant CA-PLGA and CTPGS are respectively 3.5g (0.16mmol) and 1.1g (0.68mmol).Dehydrating condensation agent N, N-NSC 57182 (DCC) 0.125g (0.6mmol), catalyzer 4-Dimethylamino pyridine (DMAP) 0.015g (0.12mmol).The methylene dichloride of 50ml is a reaction solvent.30 ℃ of temperature of reaction; Behind the reaction 24h be precipitated out product, precipitation agent is that anhydrous diethyl ether and methyl alcohol volume ratio are 2: 1 mixed solvent, filters; To be deposited in 40 ℃ of vacuum-drying 24h; Can obtain number-average molecular weight is 26198 (m=78, n=26, star-like amphipathic nature block polymer CA-PLGA-TPGS p=23).Shown in Figure 1 is the building-up reactions formula of the star-like amphipathic nature block polymer of CA-PLGA-TPGS.
Proton nmr spectra (
1HNMR), gel permeation chromatography (GPC) and thermogravimetric analysis (TGA) result prove that star-like amphipathic nature block polymer CA-PLGA-TPGS synthesizes successfully.Fig. 2 is the hydrogen nuclear magnetic resonance spectrogram of polymkeric substance CA-PLGA and CA-PLGA-TPGS.Methyl (CH in the rac-Lactide in the PLGA segment
3) signal of hydrogen and methyne (CH) hydrogen appears at 1.62ppm (a peak) respectively and 5.21ppm (b peak) locates, and is arranged in the methylene radical (CH that fignal center that 4.62-4.82ppm (c peak) locates then belongs to NSC 403079
2) proton.Methylene radical (CH in the TPGS polyoxyethylene glycol
2) absorption peak appear at 3.65ppm (h peak) and locate, this is the characteristic peak of TPGS.4.41ppm located to occur to represent the PLGA segment terminal connect hydroxyl-the CH-peak, show rac-Lactide and NSC 403079 open loop formation polymer segment; Script connects three hydroxyls-CH-peak and moves on to 3.80ppm from 3.21ppm on the CA molecule, show that hydroxyl and the second rac-Lactide on the CA reacts, and changes the ester bond structure into.Proton nmr spectra (
1HNMR) analytical results also explains that the CA-PLGA-TPGS multipolymer synthesizes successfully.Fig. 3 is the gel permeation chromatography figure (GPC) of polymkeric substance CA-PLGA and CA-PLGA-TPGS.As can be seen from the figure, polymkeric substance CA-PLGA and CA-PLGA-TPGS are unimodal, explain that these two polymkeric substance successfully synthesize and have a good monodispersity.This reaction product of unimodal explanation of polymkeric substance CA-PLGA is not the physical mixed of glycolide monomer, lactide monomer and cholic acid initiator; This reaction product of unimodal explanation of polymkeric substance CA-PLGA-TPGS is not the physical mixed of CA-PLGA and TPGS.Wherein, the appearance time of CA-PLGA is 20.5min, and the CA-PLGA-TPGS multipolymer appearance time be 19.3min.Detect peak among the GPC result of multipolymer CA-PLGA-TPGS less than TPGS, and only occur one unimodal.The number-average molecular weight Mn that draws multipolymer CA-PLGA through GPC result is 21307, and multipolymer CA-PLGA-TPGS number-average molecular weight Mn is 26198.Molecular weight through polymkeric substance CA-PLGA and CA-PLGA-TPGS also can draw TPGS and successfully be coupled to CA-PLGA and form amphipathic nature block polymer.Fig. 4 is carboxylated TPGS, the thermogravimetric analysis curve (TGA) of polymkeric substance CA-PLGA and CA-PLGA-TPGS.What TGA detected is with variation of temperature, and composition causes the situation of weight change in the sample because of evaporation cracking etc.As can be seen from the figure; Carboxylated TPGS only has a flex point at 300-420 ℃; CA-PLGA has only located a flex point at 250-360 ℃, and the figure of CA-PLGA-TPGS has three flex points, and a composition in each flex point representation polymer is weightless because of heating; The 250-340 ℃ of weightless peak with 330-420 ℃ is respectively partly to be caused by PLGA in the polymkeric substance and TPGS, and wherein the weightless temperature of PGA and PLA overlaps to some extent.The thermogravimetric analysis detected result proves that further multipolymer CA-PLGA-TPGS synthesizes successfully.
Embodiment 3, preparation CA-PLGA-TPGS star copolymer
The preparation method of CA-PLGA-TPGS star copolymer comprises the steps:
(1) by the mole percentage composition get 75% lactide monomer, 10% glycolide monomer and 15% cholic acid (CA) is raw material, puts into polymerizing pipe, adds the feed molar amount and be 0.5% catalyzer tributyltin chloride; Vacuumize, inflated with nitrogen repeats 3 times; Tube sealing under vacuum state; 155 ℃ of following frit reactions 12 hours, promptly obtaining number-average molecular weight was 4600 (m=19, multipolymer CA-PLGA n=3).
(2) get 40% TPGS (Mn=1500) by mole per-cent, 45% maleic anhydride, 15% catalyzer pyridine and diethylenetriamine, wherein the molar ratio of pyridine and diethylenetriamine is 1: 1.Add the solvent toluene be equivalent to 8 times of quality of raw material, 25 ℃ of temperature of reaction are reacted and after 40 hours are precipitated out product; Precipitation agent is the ice anhydrous diethyl ether; Filter, will be deposited in 40 ℃ of vacuum-drying 24h, can obtain carboxylated polyethylene glycol 1000 vitamin E succinic acid ester (CTPGS).
(3) preparation of CA-PLGA-TPGS star-type polymer.Reactant CA-PLGA and CTPGS, the molar ratio of CTPGS are 4 times of CA-PLGA.N, N-NSC 57182 (DCC) is a dehydrating condensation agent, add-on is 2 times of the molar weights of CA-PLGA.The 4-Dimethylamino pyridine is a catalyzer, and catalytic amount is 0.3 times of the molar weight of CA-PLGA.Add the N that is equivalent to 8 times of quality of raw material, dinethylformamide (DMF) is a reaction solvent.25 ℃ of temperature of reaction behind the reaction 40h are precipitated out product, and precipitation agent is a sherwood oil, filters, and will precipitate and 40 ℃ of vacuum-drying 24h, and can obtain number-average molecular weight is 9250 (m=19, n=3, star-like amphipathic nature block polymer CA-PLGA-TPGS p=23).
The CA-PLGA-TPGS nanoparticle of embodiment 4, preparation carrying docetaxel (Docetaxel)
Utilize ultrasonic emulsification/solvent evaporation method to prepare carrying docetaxel (Docetaxel) CA-PLGA-TPGS nanoparticle.The preparation method is following: accurately take by weighing CA-PLGA-TPGS multipolymer and a certain amount of Docetaxel powder of 200mg embodiment 2 preparations, be dissolved in the 12ml methylene dichloride.Under agitation condition, the massfraction that this solution is joined 240ml is in the 0.03%TPGS aqueous solution.Under condition of ice bath, disperse 120s with the 25w power ultrasonic, form emulsion oil-in-water, organic solvent is removed in the decompression volatilization.The centrifugal 15min of 20000rpm is with deionized water wash three times, to remove TPGS and free Docetaxel medicine.The gained deposition is resuspended in the 10ml deionized water, and lyophilize gets carrying docetaxel CA-PLGA-TPGS nanoparticle product.Utilization prepares carrying docetaxel PLGA nanoparticle with quadrat method.The ESEM result of carrying docetaxel CA-PLGA-TPGS nanoparticle sees Fig. 5, and the nanoparticle size is than homogeneous, smooth in appearance, and spherical in shape, particle diameter is greatly about about 200nm.As shown in Figure 6, the nanoparticle narrower particle size distribution, particle diameter about about 200nm, has further confirmed the observations of ESEM greatly.As shown in Figure 7, the Zeta potential of nanoparticle is about-20mV, and the absolute value of surface charge is than higher, and repulsive interaction is stronger between the particle, thereby stable at the disperse phase camber.As shown in table 1, using 0.03%TPGS to prepare drug loading as emulsifying agent is 10% polyene-containing taxol nanoparticle, and encapsulation rate is reached near 100%.As shown in Figure 8, Docetaxel crystalline endotherm(ic)peak is at 173 ℃, and carrying docetaxel PLGA and CA-PLGA-TPGS nanoparticle are not all seen Docetaxel crystalline endotherm(ic)peak, explain that Docetaxel is wrapped in the nanoparticle, and its endotherm(ic)peak has disappeared.As shown in Figure 9, carrying docetaxel PLGA has similar release profiles with the CA-PLGA-TPGS nanoparticle, is two phase release characteristics and follows initial " burst effect ".CA-PLGA-TPGS nanoparticle release rate is faster, is prone to satisfy clinical requirement.Shown in figure 10, the blank CA-PLGA-TPGS nanoparticle of medicine carrying does not have excellent biological compatibility, because it does not all have tangible toxicity to the MCF-7 cell under different nanoparticle suspension concentration; And carrying docetaxel CA-PLGA-TPGS nanoparticle has significant cytotoxicity.In addition, MTT experimental result explanation carrying docetaxel CA-PLGA-TPGS nanoparticle has time and concentration dependent to the toxicity of MCF-7 cell.
Table 1. prepares in the polyene-containing taxol nanoparticle process with the CA-PLGA-TPGS copolymer material, and drug loading and emulsifying agent are to the influence of particle diameter, Zeta potential and the encapsulation rate of nanoparticle
The CA-PLGA-TPGS nanoparticle of tonka bean camphor-6 is carried in embodiment 5, preparation
Utilize ultrasonic emulsification/solvent evaporation method preparation to carry the CA-PLGA-TPGS nanoparticle of tonka bean camphor-6.The preparation method is following: accurately take by weighing the CA-PLGA-TPGS star copolymer of 100mg embodiment 2 preparations and the tonka bean camphor-6 of 12mg, be dissolved in the 8ml methylene dichloride.Under agitation condition, this solution is joined in the 0.03%TPGS aqueous solution of 120ml.Under condition of ice bath, disperse 120s with the 25w power ultrasonic, form emulsion oil-in-water, organic solvent is removed in the decompression volatilization.The centrifugal 15min of 20000rpm is with deionized water wash three times, to remove de-emulsifier TPGS and free tonka bean camphor-6.The gained deposition is resuspended in the 10ml deionized water, and lyophilize must be carried the CA-PLGA-TPGS nanoparticle product of tonka bean camphor-6.The nanoparticle particle diameter is about 220 nanometers.Utilize the nuclear staining of DAPI pair cell, can in the cellular uptake experiment, confirm to carry the position of tonka bean camphor-6 nanoparticle in cell through the location of pair cell nuclear.Figure 11 is to the picked-up result of carrying tonka bean camphor-6 nano particle by the CA-PLGA-TPGS preparation with laser confocal scanning electron microscope observation MCF-7 cell.As can be seen from the figure, only after hatching 2h with cell, nanoparticle is just absorbed by cell.Merge the figure C that obtains from figure A with figure B and can know to such an extent that see, greeny nanoparticle great majority are arranged in tenuigenin, tightly surround the nucleus that is blue.
Taxol (Paclitaxel) CA-PLGA-TPGS microballoon is carried in embodiment 6, preparation
Utilize the solvent evaporation method preparation to carry taxol (Paclitaxel) CA-PLGA-TPGS microballoon.The preparation method is following: accurately take by weighing the CA-PLGA-TPGS multipolymer of 150mg embodiment 2 preparations and the taxol powder of 50mg, be dissolved in the 16ml methylene dichloride.Under agitation condition, this solution is joined in 0.2% Z 150PH (PVA) aqueous solution of 500ml.The 800rpm rotating speed stirred two hours down, formed emulsion oil-in-water, and 500rpm rotating speed volatilization down spends the night.The centrifugal 15min of 2500rpm is with deionized water wash three times, to remove de-emulsifier PVA and free taxol drug.The gained deposition is resuspended in the 10ml deionized water, and lyophilize must be carried taxol CA-PLGA-TPGS microball prepn.Shown in figure 12, carry taxol CA-PLGA-TPGS microsphere features smooth surface, particle diameter is about 15 microns.The drug loading that carries taxol CA-PLGA-TPGS microballoon is about 25%, and encapsulation rate is more than 80%.
Claims (10)
1. CA-PLGA-TPGS star block copolymer, its structural formula is suc as formula shown in the I:
Wherein: m=5-260, n=1-200;
TPGS represents the group shown in the formula II:
Wherein: p=23-115.
2. CA-PLGA-TPGS star block copolymer according to claim 1 is characterized in that: said CA-PLGA-TPGS star block copolymer prepares according to arbitrary said method among the claim 3-7.
3. the method for preparing the star block copolymer of CA-PLGA-TPGS described in the claim 1; Comprise the steps: the preparation of (1) CA-PLGA: according to feed molar per-cent; With the lactide monomer of 2%-95%, the glycolide monomer of 2%-95% and the cholic acid initiator of 0.1%-20% is raw material; Under the condition of anhydrous and oxygen-free and catalyzer existence, carry out polyreaction, obtain the CA-PLGA multipolymer;
(2) TPGS's is carboxylated: according to feed molar per-cent; With the polyoxyethylene glycol VE-succinate of 5%-90%, the Succinic anhydried of 5%-90% or the catalyzer of maleic anhydride and 5%-50% is that raw material reacts; Reaction finishes the back reaction solution that obtains is precipitated with precipitation agent; Filter, will precipitate drying, obtain carboxylated polyoxyethylene glycol VE-succinate;
(3) preparation of CA-PLGA-TPGS star-type polymer: CA-PLGA, CTPGS and dehydrating condensation agent are reacted under the effect of catalyzer; Reaction finishes the back reaction solution that obtains is precipitated with precipitation agent; Filter, will precipitate drying, obtain star block copolymer CA-PLGA-TPGS; Wherein, the mol ratio of said CTPGS and CA-PLGA is 1-15.
4. method according to claim 3 is characterized in that: catalyzer described in the step 1) is selected from following at least a: stannous octoate, stannous iso caprylate, organic guanidine, metallic zinc, tributyltin chloride, ferric acetyl acetonade, zinc lactate, nano zine oxide, taurine, ethanol iron, n-propyl alcohol iron, Virahol iron and propyl carbinol iron; Said catalyst consumption is the 0.01%-1% of said monomer molar amount;
In the step 1), the temperature of reaction of said polyreaction is 120-180 ℃, and the reaction times is 6-24 hour.
5. according to claim 3 or 4 described methods; It is characterized in that: said method also is included in step 2 after the step 1)) preceding step of the CA-PLGA multipolymer that obtains being carried out purifying; Method is following: CA-PLGA multipolymer bullion is dissolved in methylene dichloride or the ETHYLE ACETATE, adds methyl alcohol or sherwood oil and make CA-PLGA multipolymer deposition, filter; To be deposited in 30-50 ℃ of vacuum-drying, promptly obtain the CA-PLGA multipolymer of purifying.
6. according to each described method among the claim 3-5, it is characterized in that: step 2) described in catalyzer be selected from following a) and b) in any one or a) and b) according to mol ratio 0.1-2: 1 mixture formed: a) pyridine, 2-picoline, 4-picoline or 4-Dimethylamino pyridine (DMAP); B) triethylamine quadrol, triethylene diamine or tetrem alkene triamine;
Step 2) reaction solvent of reaction is selected from following at least a described in: dioxane, methylene dichloride, toluene, THF and N, dinethylformamide, its consumption are 2-20 times of raw material total mass;
Step 2) temperature of reaction of reaction is 5-60 ℃ described in, reacts to be 8-48 hour;
Step 2) precipitation agent described in is selected from following at least a: ether, normal heptane and sherwood oil;
Step 2) in said being deposited in carried out drying under the vacuum condition, the exsiccant temperature is 30-50 ℃, and the time is 12-48h.
7. according to each described method among the claim 3-6; It is characterized in that: in the step 3); Said dehydrating condensation agent is N, N-NSC 57182 or 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide, and its consumption is 1-15 a times of said CA-PLGA molar weight;
Catalyzer described in the step 3) is pyridine, 2-picoline, 4-picoline or 4-Dimethylamino pyridine, and its add-on is 0.01-10 a times of said CA-PLGA molar weight;
The reaction solvent that reacts described in the step 3) is selected from following at least a: methylene dichloride, toluene, N, dinethylformamide, dioxane and THF, its consumption are 2-20 times of raw material total mass;
The temperature of reaction of reacting described in the step 3) is 5-60 ℃, reacts to be 8-48 hour;
Precipitation agent described in the step 3) is selected from following at least a: ether, normal heptane, sherwood oil, methyl alcohol and ethanol; In the step 3) said being deposited in carried out drying under the vacuum condition, the exsiccant temperature is 30-50 ℃, and the time is 12-48h.
8. claim 1 or 2 described CA-PLGA-TPGS star block copolymers are as the application of pharmaceutical excipient in pharmaceutical formulations.
9. application according to claim 8 is characterized in that: said pharmaceutical prepn is a microparticle formulation, preferred drug-carrying nanometer particle, medicine carrying gel or medicine carrying microballoons; Said pharmaceutical prepn is anti-tumor medicinal preparation and/or cardiovascular disease resistant pharmaceutical prepn.
10. claim 1 or 2 described CA-PLGA-TPGS star block copolymers are as the application of tissue engineering material.
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| CN103073730A (en) * | 2012-12-27 | 2013-05-01 | 清华大学深圳研究生院 | CA-(PLA-ran-PLC-b-PEC) periodic copolymer and preparation method and application thereof |
| CN103159959A (en) * | 2013-03-07 | 2013-06-19 | 清华大学深圳研究生院 | M-PLGA-TPGS star type amphiphilic copolymer and preparation method and application |
| CN110964204A (en) * | 2018-09-29 | 2020-04-07 | 南方医科大学 | PLGA microspheres of charge-loaded positive micelle/insulin compound and preparation method thereof |
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| CN102030891A (en) * | 2010-11-09 | 2011-04-27 | 清华大学深圳研究生院 | TPGS-b-(PLA-ran-PGA) copolymer and preparation method and application thereof |
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| CN102030891A (en) * | 2010-11-09 | 2011-04-27 | 清华大学深圳研究生院 | TPGS-b-(PLA-ran-PGA) copolymer and preparation method and application thereof |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103073730A (en) * | 2012-12-27 | 2013-05-01 | 清华大学深圳研究生院 | CA-(PLA-ran-PLC-b-PEC) periodic copolymer and preparation method and application thereof |
| CN103073730B (en) * | 2012-12-27 | 2015-07-22 | 清华大学深圳研究生院 | CA-(PLA-ran-PLC-b-PEC) periodic copolymer and preparation method and application thereof |
| CN103159959A (en) * | 2013-03-07 | 2013-06-19 | 清华大学深圳研究生院 | M-PLGA-TPGS star type amphiphilic copolymer and preparation method and application |
| CN110964204A (en) * | 2018-09-29 | 2020-04-07 | 南方医科大学 | PLGA microspheres of charge-loaded positive micelle/insulin compound and preparation method thereof |
| CN110964204B (en) * | 2018-09-29 | 2021-03-12 | 南方医科大学 | PLGA microspheres of charge-loaded positive micelle/insulin compound and preparation method thereof |
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