CN103304417A - Preparation method and application of amphiphatic copolymer modified chitosan compound - Google Patents

Abstract

The invention discloses a preparation method of amphiphatic copolymer modified chitosan compound. The method comprises the following steps: firstly using a mixture of lactic acid and L-malic acid as initial raw materials to directly prepare polylactic acid and malic acid oligomer by polycondensation, and preparing a series of copolymers in different hydrophilic degrees by changing the proportion of the lactic acid and L-malic acid (respectively as 1:1, 1:2, 1:3, 1:5, 1:8 and 1:10), respectively compounding the copolymers with different components with the water-soluble chitosan in mixing to obtain various compounds with different components; adding 0.8-2.4% of polyethylene glycol diglycidyl ether in the compound solution as the crosslinking agent, reacting for 6 hours at 70 DEG C in the water solution, and crosslinking the chitson in the compound to prepare the compound hydrogel; meanwhile, selecting the compound with low crosslinking degree to tiling on a glass plate, drying at 45 DEG C to form the film. The gel and the film can be used as a drug sustained release carrier, a wound dressing and a medical abherent.

Description

A kind of preparation and application of amphiphilic copolymer modified chitosan mixture

Technical field

This research relates to the preparation of the synthetic and chitosan complexes of a kind of lactic acid and oxysuccinic acid amphiphilic copolymer, belongs to the bio-medical material technical field.

Background technology

Chitosan is the deacetylated product of chitin, has the effect that strengthens the macrophage phagocytic ability, promotes wound healing, is a kind of excellent property, the great medical material of development potentiality.Chitosan gel rubber is treated skin injury as wound dressing, and following effect is arranged: (1) antibacterial, can promote wound healing, and reduce paralysed trace and form; (2) anastalsis.Can also stop the formation of fibrinogen bundle in the time of hemostasis.(3) biological barrier effect.Aquagel has the wound surface of isolation and anti-microbial effect as the postoperative release agent, can effectively stop the bonding of wound face, avoids second operation.Poly(lactic acid) and polymalic acid are special aliphatic polyesters, have excellent biological compatibility and degradation property, and its degraded product can participate in tricarboxylic acid cycle in the body and be absorbed by organism.In addition, polymalic acid has pendant carboxy group, modifies easily and introduces other functional groups or small-molecule drug, in fields such as biological medicine, biomaterials great application prospect is arranged.Above-mentioned two kinds of superpolymer all are biocompatibility and degradable materials, and are cheap, and safety non-toxic is fit to human body and uses.

The hydrogel of the Preparation of Chitosan of one-component flowability when low crosslinking degree is bigger, and mechanical strength is not enough, influences use properties.The film of the Preparation of Chitosan of one-component is crisp, and biodegradation period is oversize, and unfavorable eliminating is external.The used linking agent of preparation chitosan gel rubber generally has glutaraldehyde, formaldehyde, epoxy chloropropane etc., and these cross-linker molecules have toxicity in various degree, remain in gel inside can't remove, and have increased the potential danger of product.Too high glutaraldehyde content can increase the cytotoxicity of gel, and color burn and gel are crisp hard, is unfavorable for the bonding and extension at wound surface.Cross low glutaraldehyde content and then cause crosslinking degree not enough, influence result of use.

Therefore the present invention intends with a kind of amphiphilic copolymer chitosan being carried out modification, and preparation is based on the mixture of chitosan.Use a kind of macromolecule crosslink agent cross-linking then, prepare aquagel and film product respectively.By the modification that amphiphilic copolymer participates in, in mixture, introduce suitable hydrophobicity composition (poly(lactic acid)), so that the mechanical property of strongthener.Polymalic acid contains side chain carboxyl group, easily with contain amino chitosan and carry out compoundly, form uniform mixture material.Use macromolecule crosslink agent polyoxyethylene glycol bisglycidyl ether with crosslinking complexes at last, compared with the result with the small molecules linking agent, gained toughness of material and kindliness increase, and have improved the mechanical property of chitosan, shorten degradation cycle in vivo, be suitable for human body more.

Summary of the invention

1. be that the starting raw material direct condensation prepares poly(lactic acid) and oxysuccinic acid oligopolymer with lactic acid and L MALIC ACID mixture, the ratio of change lactic acid and L MALIC ACID (be respectively 1: 1,1: 2,1: 3,1: 5, profit was 1: 10 in 1: 8) the different multipolymer of a series of hydrophilicities of preparation.

2. will form different multipolymers and mix with water-soluble chitosan respectively and carry out compoundly, obtain based on the different mixture of components chitosan, various.In complex solution, add 0.8-2.4% polyoxyethylene glycol bisglycidyl ether and make linking agent and in the aqueous solution, react, make chitosan crosslinked in the mixture, preparation mixture hydrogel.

3. the mixture of selecting simultaneously to contain low cross-linking agent (0.1-0.5%) is tiled on the sheet glass, suitably forms film behind the heat drying.

Advantage of the present invention:

1. the modification that participates in by amphiphilic copolymer is introduced suitable hydrophobicity composition (poly(lactic acid)) in mixture, so that the mechanical property of strongthener reduces mobile.

2. the polymalic acid of certain ingredients contains side chain carboxyl group in the multipolymer, easily with contain amino chitosan and carry out compoundly, form uniform mixture material.

3. the multipolymer participation is compound, has weakened the intersegmental hydrogen bond action of chitosan molecule chain, has shortened biodegradation period, is conducive to use at human body.

With macromolecule crosslink agent with crosslinking complexes, compared with the result with the small molecules linking agent, gained toughness of material and kindliness increase, with making the people feel comfortable in vivo.Avoided the faint toxicity brought with linking agents such as glutaraldehyde;

5. raw materials used chitosan, poly(lactic acid) and polymalic acid all are biocompatibility and the good material of degradation property, and chitosan has anti-microbial effect.All preparation process need not any organic solvent, environmental friendliness.This gel and film product can be used as many-sided purposes such as slow releasing carrier of medication, wound dressings and medical stick agent.

Description of drawings

The preparation synoptic diagram of the multipolymer of Fig. 1 lactic acid/oxysuccinic acid and chitosan complexes

The infrared spectrogram of Fig. 2 lactic acid/oxysuccinic acid multipolymer (1: 2)

Fig. 3 lactic acid/oxysuccinic acid multipolymer and chitosan complexes polyoxyethylene glycol bisglycidyl ether crosslinking reaction formula

The cytotoxicity experiment Photomicrograph result of Fig. 4 chitosan complexes.A: negative control sample result B: chitosan complexes sample result.

Embodiment:

Embodiment 1: the preparation of lactic acid and oxysuccinic acid amphiphilic copolymer:

In 100 milliliters of round-bottomed flasks, add 5 gram lactic acid and 10 gram L MALIC ACID mixtures and 0.10 gram zinc lactate crystal, slowly heating up is heated to 110 ℃ of reactions 1 hour, connects the water pump decompression and continues reaction 14 hours, obtains white solid amphiphilic copolymer product.

Embodiment 2: in 100 milliliters of round-bottomed flasks, changing the ratio of L MALIC ACID and lactic acid, is 1: 1 according to lactic acid and L MALIC ACID weight ratio respectively, 1: 2,1: 3,1: 5,1: 8 and 1: 10, take the conditions of similarity reaction among the embodiment 1, can obtain a series of multipolymers.

Embodiment 3: the preparation of amphiphilic copolymer and chitosan complexes:

Add water 80 milliliters of soluble chitosan 2.5 grams and distilled water in 250 ml beakers, soaking and stirring is until forming even chitosan solution.Be that 1: 2 multipolymer is mixed with 3.0% the aqueous solution with lactic acid and L MALIC ACID component ratio, under whipped state, 12 milliliters of this solution slowly splashed into chitosan aqueous solution and form mixture.

Embodiment 4: be made into 3.0% the aqueous solution with the multipolymer of different lactic acid and L MALIC ACID component ratio (1: 1,1: 2,1: 3,1: 5,1: 8 and 1: 10), form mixture with chitosan aqueous solution respectively.

Embodiment 5: the preparation of chitosan complexes hydrogel:

In 100 milliliters of three-necked bottles; add gained amphiphilic copolymer and chitosan complexes 50 grams among the embodiment 3; add the polyoxyethylene glycol bisglycidyl ether according to the weight ratio with chitosan for the 0.8-2.4% weight ratio; stir; under nitrogen protection, reacted 6 hours in 70 ℃ of aqueous solution, form hydrogel.

Embodiment 6: the preparation of chitosan complexes film:

In 100 ml beakers, add gained amphiphilic copolymer and chitosan complexes solution 30 grams among the embodiment 3, add the polyoxyethylene glycol bisglycidyl ether according to the weight ratio 0.1-0.5% with chitosan, after stirring, be tiled on the sheet glass vacuum drying film forming under 45 ℃ of conditions.

Embodiment 7: the chitosan complexes hydrogel is as the application of pharmaceutical carrier:

With chitosan complexes hydrogel lyophilize among the embodiment 5, take by weighing the 0.1g Ibuprofen BP/EP then and be dissolved in 15ml N, in the dinethylformamide, put into xerogel 0.1g again, constant temperature was placed 48 hours for 37 ℃ ± 0.5 time.Then gel is taken out, the Ibuprofen BP/EP with dehydrated alcohol flush away gel surface is dried to constant weight in the cryogenic vacuum baking oven.Take by weighing 50mg medicine carrying sample and place the 200mL release medium respectively, discharge at 37 ℃ ± 0.5 time constant temperature, certain interval of time sampling liquid 4mL centrifugally goes its stillness of night, and the fresh medium of additional equal volume.Measure absorbancy at the 264nm place with ultraviolet spectrophotometer, press the typical curve regression equation calculation and discharge Ibuprofen BP/EP content in the liquid.

Embodiment 8: the chitosan complexes film is as the application of pharmaceutical carrier:

Take by weighing that film sample 0.10g places the 200mL release medium among the embodiment 6, discharge at 37 ℃ ± 0.5 time constant temperature.Operation steps is with embodiment 7.

Embodiment 9: the chitosan complexes hydrogel is used for medical dressing:

Add water 40 milliliters of soluble chitosan 2 grams and distilled water in 100 ml beakers, soaking and stirring is until forming even chitosan solution.Be that 1: 2 multipolymer is mixed with 3.0% the aqueous solution with lactic acid and L MALIC ACID component ratio, under whipped state, 8 milliliters of this solution slowly splashed into chitosan aqueous solution and form mixture.Add polyoxyethylene glycol bisglycidyl ether 0.15 gram then in the mixture, pour into after stirring a mould (10 * 8 * 0.5cm), insert in the thermostat container 70 ° until forming the sheet gel, this gel can be used as wound dressings.

Embodiment 10: the cytotoxicity experiment of chitosan complexes:

By vitro culture human microvascular endothelial cell (mvec) (HMEC-1cells), adopt the cytotoxicity of srb assay test chitosan complexes.With cell culture fluid 10%MCDB the cell of logarithmic phase being made into concentration is 6 * 10 ³The cell suspending liquid of individual/mL ^[79], every hole 150 μ L are inoculated in 3 96 well culture plates, place 37 ℃, 5%CO ₂Cultivate 24h in the incubator.Original fluid in the every hole of sucking-off, every hole adds 150 μ L, negative controls (10%MCDB substratum), positive control solution (0.64% phenol substratum), sample sets (being diluted to the sample of 0.1g/L with the 10%MCDB nutrient solution), continue to place 37 ℃, 5%CO ₂Cultivate in the incubator, respectively at respectively taking out a board test on the the 1st, 2,3 day.Observe, estimate cell growth condition by inverted microscope after taking out culture plate.Calculate the relative appreciation rate of cell by srb assay: pour out nutrient solution, every hole adds 200 μ L 10%TCA stationary liquids, at 4 ℃ of fixing 40min.Abandon stationary liquid, deionized water wash dries.Every hole adds 100 μ L 0.4%SRB staining fluids, 37 ℃ of dyeing 30min.Abandon staining fluid, deionized water wash dries.Every hole adds 150 μ L 10mmol/L Tris, and (every hole is surveyed 3 times, and each 60s gets average) shaken at the 540nm place on microplate reader, measures the absorbancy 0D value in every hole.Calculate the relative appreciation rate of cell (RGR), assess sample toxic grade according to following formula.

The relative appreciation rate of cell (RGR) calculation formula:

$\mathrm{RGR}=\frac{{\mathrm{OD}}_{\mathrm{expenmental\; group}}}{{\mathrm{OD}}_{\mathrm{negarive\; group}}}\×100\%$

The cytotoxicity grade is zero level as a result.

Claims (7)

1. the oligopolymer of a lactic acid and L MALIC ACID and synthetic, it is characterized in that lactic acid and L MALIC ACID were respectively 1: 1 according to weight ratio, 1: 2,1: 3,1: 5, the mixed of 1: 8 and 1: 10, heating under vacuum to 130 ℃ direct condensation obtained the multipolymer of lactic acid and L MALIC ACID in 15 hours under catalytic condition, it is amphiphilic that product has hydrophilic, has infrared spectrogram shown in Figure 3.

2. the synthetic 0.1-0.8% of the being to use zinc lactate of the described multipolymer of claim 1 is made catalyzer.

3. claim 1 a gained lactic acid and the multipolymer of L MALIC ACID and the mixture of chitosan is characterized in that the amphiphilic copolymer of lactic acid and oxysuccinic acid and water-soluble chitosan are compounded to form in the aqueous solution.The weight ratio of multipolymer and chitosan was respectively 1: 5, and 1: 10,1: 15,1: 20.

4. chitosan complexes hydrogel; it is characterized in that the compound water solution of amphiphilic copolymer and chitosan is made linking agent with the polyoxyethylene glycol bisglycidyl ether in the aforesaid right requirement 2; react formation after 6 hours in 70 ℃ of aqueous solution under nitrogen protection, the reaction synoptic diagram as shown in Figure 2.The consumption control of linking agent is being 0.8-2.4%.

5. chitosan complexes water film, it is characterized in that the compound water solution of amphiphilic copolymer and chitosan is made linking agent with the polyoxyethylene glycol bisglycidyl ether in the aforesaid right requirement 2, consumption by linking agent is that 0.1-0.5% mixes, be tiled on the sheet glass vacuum drying film forming under 45 ℃ of conditions.

In claim 3 and the claim 4 chitosan complexes hydrogel and film as the application of drug release carrier.

7. the chitosan complexes hydrogel is used for medical dressing in the claim 3.

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