CN104274833A - Preparation of chitosan and malic acid and lactic acid copolymer composite drug carrier - Google Patents
Preparation of chitosan and malic acid and lactic acid copolymer composite drug carrier Download PDFInfo
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- CN104274833A CN104274833A CN201310282080.XA CN201310282080A CN104274833A CN 104274833 A CN104274833 A CN 104274833A CN 201310282080 A CN201310282080 A CN 201310282080A CN 104274833 A CN104274833 A CN 104274833A
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Abstract
The invention discloses a method for preparation of a nano micelle as a drug carrier by compositing chitosan and a malic acid and lactic acid copolymer, and belongs to the technical field of biological materials and release. Firstly, the malic acid and lactic acid copolymer (Poly (MA-Co-LA)) is obtained by copolycondensation reaction of lactic acid and malic acid under the catalytic action of stannous chloride; then the malic acid and lactic acid copolymer (Poly (MA-Co-LA)) is composited with the chitosan, and the chitosan and malic acid-lactic acid copolymer polyelectrolyte composite nanoparticles are formed by self-assembly by electrostatic interaction between two polyelectrolytes. The drug doxorubicin hydrochloride is loaded onto the nanoparticles for release control, and the carrier has excellent drug loading and controlled release effects. The e chitosan and malic acid-lactic acid copolymer nano micelle prepared by the method is stablein structure, simple in synthesis, good in biocompatibility, free of cell toxicity, and larger in accumulated release amount.
Description
Technical field
The present invention relates to the preparation of a kind of biodegradable chitosan and malic acid and lactic acid copolymer combination drug carrier, belong to biomaterial and slow release method field.
Background technology
Compound polyelectrolyte (Polyelectrolyte Complex, is abbreviated as PEC) is mixed by the polyelectrolyte with opposite charges, namely combined by Coulomb force between polycation and polyanion and formed.Its purposes mainly contains 2 points: the property 1. utilizing its finished product, as a kind of materials'use.2. utilize some special effects produced in poly-anions and canons interaction process, thus be applied to the aspects such as flocculation, absorption.Compound polyelectrolyte is as a kind of material, and its most important performance is the hydrophilic of height and still can keeps good mechanical performance in the swollen state.That is familiar with compound polyelectrolyte along with people deepens continuously, and compound polyelectrolyte will obtain applying more and more widely.
Chitosan is a kind of natural polycation polysaccharide obtained by chitin deacetylase base, dissolves in acidic aqueous solution, nontoxic, nonirritant, without sensitization, without mutagenic action, has good biocompatibility and biological degradability.Chitosan as pharmaceutical carrier can Drug controlled release, prolong drug curative effect, reduce poisonous side effect of medicine, improve the permeability of hydrophobic drug cell membrane and the stability of medicine and change route of administration, greatly can also strengthen the target administration ability of preparation.Due to biocompatibility and the degradability of its uniqueness, and wide material sources, technology of preparing is ripe, and operation is simple and easy, tasteless, heat-resisting, and the advantages such as low price, are widely used in medicine and other fields at present.
Polylactic acid and polymalic acid all have good biocompatibility and degradability, and its catabolite can participate in tricarboxylic acid cycle in body and be absorbed by organism.
First the present invention prepares the random copolymer of lactic acid and malic acid, utilizes the pendant carboxy group in polymalic acid construction unit and the amino in chitosan molecule to carry out compound, then defines nanoparticle by self assembly in aqueous.This nanoparticle as the carrier of anticancer drugs, doxorubicin, can carry out pH Co ntrolled release.
Summary of the invention
The invention provides the preparation method of a kind of chitosan and malic acid and lactic acid random copolymer complex nanometer granule.First malic acid and lactic acid random copolymer has been prepared, then compound action amino in the carboxyl of suspended side chain in polymalic acid construction unit in this random copolymer and chitosan molecule is utilized, be prepared into polyelectrolyte nanoparticle, and be used as load and the Co ntrolled release of amycin.
Beneficial effect of the present invention:
1. the complex nano-micelle that prepared by the present invention can be used in art of pharmacy, and due to the existence of group multiple on chitosan, make nanoparticle have good biocompatibility, chitosan has certain antibacterial action.
2. utilize carboxyl and the chitosan compound of polymalic acid construction unit suspended side chain in random copolymer, Compound Degree is high, and two kinds of segments are evenly distributed, and is conducive to the stable of product property and controls.Polymalic acid has certain self-catalysis Degradation simultaneously, suitably can accelerate the degradation speed of product, remove fast in body.Introducing polylactic acid chain segment is to increase the certain hydrophobicity of complex, gives the mechanical strength that composite nanoparticle is certain, strengthens serviceability.
3. the size of complex nanometer granule forms relevant with the chain link of polylactic acid in copolymer (PLA) and polymalic acid (PMA), can be regulated by rate of charge during synthesis.
4. preparation process of the present invention need not any organic solvent, and environmental friendliness, step is easy, product good biocompatibility.
Accompanying drawing explanation
The infrared spectrogram of Fig. 1 chitosan and malic acid and lactic acid copolymer complex.A in figure, b, c, d, e represent the copolymer that malic acid and lactic acid molar ratio are 1: 0,1: 0.5,1: 1,1: 2,1: 3 respectively.
The nuclear magnetic spectrogram of Fig. 2 chitosan and malic acid and lactic acid copolymer complex.A in figure, b, c, d, e represent the copolymer that malic acid and lactic acid molar ratio are 1: 0,1: 0.5,1: 1,1: 2,1: 3 respectively.
The Viscosity Analysis of Fig. 3 chitosan and malic acid and lactic acid copolymer complex solution.A in figure, b, c, d, e represent the copolymer that malic acid and lactic acid molar ratio are 1: 0,1: 0.5,1: 1,1: 2,1: 3 respectively.
The scanning electron microscope of Fig. 4 chitosan and malic acid and lactic acid copolymer complex nano-micelle.A in figure, b, c, d, e represent copolymer and the nanoparticle that is combined into of chitosan that malic acid and lactic acid molar ratio are 1: 0,1: 0.5,1: 1,1: 2,1: 3 respectively.
TEM photo before and after Fig. 5 chitosan and malic acid and lactic acid copolymer complex nano-micelle medicine carrying.A in figure represent malic acid and lactic acid molar ratio be 1: 1 copolymer and chitosan complexes medicine carrying before nanoparticle, b represent malic acid and lactic acid molar ratio be 1: 1 copolymer and chitosan complexes medicine carrying after nanoparticle.
The drug release patterns of Fig. 6 chitosan and malic acid and lactic acid copolymer complex nano-micelle.A in figure, b, c, represent copolymer and the release profiles of chitosan complexes medicine-carried nano particles in the PBS buffer of pH5.8, pH7.4 and pH8.0 that malic acid and lactic acid molar ratio are 1: 1 respectively.
Detailed description of the invention:
Embodiment 1: the purification of chitosan: take 3g chitosan and be dissolved in the acetum of 1% (m/v) of 200mL, stir.The chitosan solution be dissolved in acetic acid is slowly instilled in 1% (m/v) sodium hydroxide solution, magnetic agitation, drip and continue to stir 2h, filter out precipitation, with deionized water wash, be filtered to filtrate in neutral.The a small amount of soak with ethanol 3h of precipitation after last filtration, filter, vacuum drying is to constant weight.Obtaining chitosan grind into powder, to be placed in exsiccator for subsequent use.
Embodiment 2: the preparation of malic acid/lactic acid copolymer: take malic acid, the lactic acid that mol ratio is respectively 1: 0,1: 0.5,1: 1,1: 2 and 1: 3 and join in the mono-neck flask of 250mL respectively, add catalyst stannous chloride 0.2g respectively, flask is placed in oil bath pan and connects condensing unit.With oil bath raised temperature to 110 DEG C, react after two hours, raised temperature to 135 DEG C also under reduced pressure (keeps pressure at 8-9kPa) and continues reaction 8 hours, obtains malic acid and lactic acid copolymer.
Embodiment 3: the preparation of micelle: the malic acid of the different proportion of preparation, lactic acid copolymer are dissolved in the weak solution that ultra-pure water is made into 0.3% respectively, prepare the chitosan solution of 0.3% in addition, getting a certain amount of copolymer solution under magnetic agitation condition respectively is slowly added drop-wise in a certain amount of chitosan solution, both control mass ratio, abundant stirring makes its mix homogeneously, forms complex solution.
Embodiment 4: the preparation of carrier micelle: the chitosan solution 10mL of preparation 0.3%, add 1mg doxorubicin hydrochloride, after ultrasonic 1h, magnetic agitation 1h makes its mix homogeneously.Get the copolymer solution of the different proportion of 20mL0.3% respectively, dropwise add in above-mentioned system, products therefrom is red and has obvious Tyndall phenomenon.
Embodiment 5 Fig. 1 is the infrared spectrum of each polymer, measures polymer architecture, scanning wavelength scope 500 ~ 4000cm with Fourier infrared spectrograph (FTLA2000-104, ABB BOMEN Corporation)
-1, resolution is 4cm
-1.As seen from Figure 1, the copolymer characteristic absworption peak of different monomers rate of charge is substantially identical.2800-3500cm
-1wide and the loose peak interior in a big way at place belongs to the stretching vibration peak of-OH and-COOH; 2900-2800cm
-1several small peaks at place; 1740cm
-1place is the stretching vibration peak of C=O, 1711cm
-1place is the stretching vibration peak of C=O in carboxyl.Illustrate that lactic acid and malic acid there occurs copolyreaction.1400cm
-1place is for being methylene-CH
2with the bending vibration of-CH on methine.950cm
-1place absworption peak be OH...O face outside deformation vibration, the existence of carboxyl can be confirmed.
Embodiment 6 Fig. 2 is the nuclear magnetic spectrogram of each polymer, uses
1h-NMR nuclear-magnetism (Mercury-Vx400, Varian Corporation) measures the structure of copolymer, with heavy water (D
20) for solvent, tetramethylsilane are internal standard substance.As seen from Figure 2, the multiplet of about 5.52ppm is the proton peak of the methine CH in malic acid construction unit; The multiplet of about 2.81-3.24ppm is the methylene CH in malic acid construction unit
2proton peak; The multiplet of about 5.27ppm is the proton peak of the methine CH in lactic acid construction unit; The multiplet of about 1.48ppm is the methyl CH in lactic acid construction unit
3proton peak.
Embodiment 7 Fig. 3 is the viscogram of each polymer, and as seen from the figure, along with the rising of copolymer concentration, apparent viscosity increases gradually.In the copolymer of different monomeric charge ratio, along with the increase of lactic acid monomer content, viscosity, all in increase tendency, may be because the ratio of viscosities polymalic acid of polylactic acid is large, lactic acid monomer rate of charge causes polylactic acid chain segment in copolymer longer when increasing, so viscosity is larger.
Embodiment 8 Fig. 4 to be malic acid and lactic acid molar ratio be 1: 0,1: 0.5,1: 1,1: 2,1: 3 copolymer and the nanoparticle that is combined into of chitosan.Composite surface pattern is with the S-4800 type scanning electronic microscope observation of Japanese HITACHI company.As can be seen from the figure, the nanoparticle of preparation is spherical in shape, nanoparticle has obvious shell one nuclear structure, dark parts material is made up of jointly the hydrophobic part of copolymer and chitosan in the middle of nanoparticle, ambient color is divided into the hydrophilic segment of copolymer to form compared with superficial part, the copolymer nano particle diameter size that all energy and chitosan compound obtain of different monomers mol ratio is about 300nm, and particle size distribution is comparatively homogeneous.But the nanoparticle Electronic Speculum figure and the particle size distribution that examine gained when finding that polymer monomers mol ratio is 1: 1 are more afterwards relatively slightly good, favorable dispersibility, nanoparticle becomes more regular spherical, and experimental result repeatability is slightly good.So have employed in the experiment of nanoparticle load amycin monomer mole ratio be 1: 1 copolymer and chitosan compound to have leisure white glue bundle.
TEM photo before and after the polymer of embodiment 9 Fig. 5 is monomer mole ratio when being 1: 1 gained and chitosan composite micelle medicine carrying, as seen from the figure, after medicine carrying, nanometer particle size increases, doxorubicin hydrochloride is wrapped in core and the surface of nanoparticle, the dissolubility of amycin is caused to be deteriorated after HCL on doxorubicin hydrochloride and the amino on chitosan react, multiple Doxorubicin molecules hydrophobic part is reunited together, causes larger stain as shown in the figure.Nanoparticle almost spherical, mode of appearance is full.
Embodiment 10 Fig. 6 is the release profiles of medicine carrying complex.Carrier micelle ADD-05 ultra-pure water is diluted half, precision measures 5mL carrier micelle in bag filter respectively, be placed in respectively 250mL medium be pH5.8,7.4 and 8.0 PBS, controlling bath temperature is (37+0.5) DEG C, magnetic agitation at a slow speed, sample 4mL at regular intervals and add the fresh dissolution medium of the identical temperature of same volume, by the concentration of UV spectrophotometer measuring sample Chinese medicine.As seen from the figure, release in first two hours is slower, in the PBS buffer system of PH5.8, PH7.4 and PH8.0, release rate is respectively 15.17%, 12.28% and 11.30%, after 30 hours, release rate is respectively 76.07%, 68.01% and 59.63%, after 34h, release tends towards stability, and drug release shows slow-releasing.
Claims (5)
1. the preparation of a malic acid and lactic acid random copolymer, it is characterized in that taking malic acid and the lactic acid that mol ratio is respectively 1: 0,1: 0.5,1: 1,1: 2 and 1: 3, join respectively in the mono-neck flask of 250mL, add stannous chloride 0.2g respectively, flask is placed in oil bath pan and heats.Raised temperature to 110 DEG C, reacts two hours follow-up high-temperatures to 135 of continuing rising DEG C and under reduced pressure (keeps pressure at 8-9kPa) and react 8 hours again, obtain malic acid and lactic acid copolymer.The molecular weight of copolymer is respectively 1070,1175,1406,1706,2143, and it is random state that the construction unit ratio of malic acid and lactic acid is respectively 1: 0,1: 0.50,1: 1.25,1: 2.18,1: 3.55 two kind of construction unit.
2. the preparation of a chitosan and malic acid and lactic acid random copolymer composite nanoparticle, it is characterized in that using malic acid described in claim 1 and lactic acid random copolymer as polyanion, using chitosan as polycation, by the amino generation electrostatic interaction of positively charged on carboxyl electronegative on copolymer and chitosan, the complex of generating chitosan and malic acid and lactic acid copolymer, and self assembly forms nanoparticle in water, the particle diameter of nanoparticle is about 300nm.
3. the preparation method of chitosan described in claim 2 and malic acid and lactic acid random copolymer composite nanoparticle and condition, above-mentioned gained malic acid and lactic acid random copolymer is it is characterized in that to be dissolved in tetrahydrofuran solution, with petroleum ether and diethyl ether solution mixing, washing, vacuum drying.In addition chitosan is dissolved in acetum, slowly instills in sodium hydroxide solution after stirring, magnetic agitation, centrifugation, with deionized water washing sediment, is filtered to filtrate in neutral, the a small amount of soak with ethanol of precipitation after last filtration, filter, vacuum drying is to constant weight.Again malic acid and lactic acid copolymer, chitosan are dissolved in ultra-pure water respectively separately, under magnetic stirring aqueous copolymers solution are dropwise added drop-wise in chitosan aqueous solution, carboxyl (-COO electronegative on copolymer chain
-) with chitosan chain on positively charged amino (-NH
3 +) between generate complex nano-micelle by electrostatic interaction.
4. malic acid according to claim 2 and the preferred mass ratio prepared as lactic acid copolymer Poly (MA-co-LA), it is characterized in that the mass ratio of MA and LA is 1: 0 ~ 1: 3, in this proportion, the nano-micelle formed is more stable, copolymer hydrophobic segment is longer, and micelle particle diameter is larger.
5. malic acid according to claim 2 and lactic acid copolymer-chitosan complexes nano-micelle are the application of Thermosensitive Material Used for Controlled Releasing of Medicine.The chitosan solution 10mL of preparation 0.3%, add 1mg doxorubicin hydrochloride, after ultrasonic 1h, magnetic agitation 1h makes its mix homogeneously.Get the copolymer solution of the different proportion of 20mL0.3% respectively, dropwise add in above-mentioned system, products therefrom is red and has obvious Tyndall phenomenon.The complex nanometer granule of gained is placed in bag filter and discharges 80 hours in the PBS buffer system of PH5.8, PH7.4 and PH8.0.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104721140A (en) * | 2015-03-27 | 2015-06-24 | 上海大学 | Preparation method of amphipathic ionic drug loading vesicle with targeted controllable drug release and embolic effect |
| CN107163255A (en) * | 2017-04-21 | 2017-09-15 | 四川大学 | Redox and the random graft type pharmaceutical carrier and method of pH double-responses |
| CN107880272A (en) * | 2017-11-27 | 2018-04-06 | 安徽雪郎生物科技股份有限公司 | A kind of β polymalic acids chitosan oligosaccharide compound salt and its preparation method and application |
| CN112500575A (en) * | 2020-12-08 | 2021-03-16 | 江南大学 | Modified chitosan microgel and preparation method and application thereof |
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| JP2005187671A (en) * | 2003-12-26 | 2005-07-14 | National Institute For Materials Science | Method for producing polymalic acid copolymer |
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| JP2005187671A (en) * | 2003-12-26 | 2005-07-14 | National Institute For Materials Science | Method for producing polymalic acid copolymer |
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| Title |
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| 戴征展: ""聚L-谷氨酸—壳聚糖聚电解质复合物及药物缓释初步研究"", 《中国优秀硕士学位论文全文数据库》 * |
Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104721140A (en) * | 2015-03-27 | 2015-06-24 | 上海大学 | Preparation method of amphipathic ionic drug loading vesicle with targeted controllable drug release and embolic effect |
| CN104721140B (en) * | 2015-03-27 | 2019-04-02 | 上海大学 | The preparation method that amphiphilic ions type drug holding theca with the controllable drug release of targeting and embolism effect steeps |
| CN107163255A (en) * | 2017-04-21 | 2017-09-15 | 四川大学 | Redox and the random graft type pharmaceutical carrier and method of pH double-responses |
| CN107163255B (en) * | 2017-04-21 | 2020-10-20 | 四川大学 | Redox and pH dual-response random-graft type drug carrier and method |
| CN107880272A (en) * | 2017-11-27 | 2018-04-06 | 安徽雪郎生物科技股份有限公司 | A kind of β polymalic acids chitosan oligosaccharide compound salt and its preparation method and application |
| CN107880272B (en) * | 2017-11-27 | 2020-06-05 | 安徽雪郎生物科技股份有限公司 | β -polymalic acid chitosan oligosaccharide compound salt and preparation method and application thereof |
| CN112500575A (en) * | 2020-12-08 | 2021-03-16 | 江南大学 | Modified chitosan microgel and preparation method and application thereof |
| CN112500575B (en) * | 2020-12-08 | 2021-08-10 | 江南大学 | Modified chitosan microgel and preparation method and application thereof |
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Application publication date: 20150114 |