CN108743566B

CN108743566B - Preparation method and application of PHBV/rosiglitazone sustained-release membrane

Info

Publication number: CN108743566B
Application number: CN201810501378.8A
Authority: CN
Inventors: 段宣初; 刘海蓉; 张凤; 黄宇婷; 刘可; 潘政
Original assignee: Hunan University; Second Xiangya Hospital of Central South University
Current assignee: Hunan University; Second Xiangya Hospital of Central South University
Priority date: 2018-05-23
Filing date: 2018-05-23
Publication date: 2021-02-02
Anticipated expiration: 2038-05-23
Also published as: CN108743566A

Abstract

The invention discloses a preparation method and application of a PHBV/rosiglitazone sustained-release membrane, belonging to the technical field of medicines, and the PHBV/rosiglitazone sustained-release membrane is prepared into PHBV-TCM/DMF silk fluid with the mass volume ratio of 5% for standby after mixing PHBV powder, chloroform and dimethylformamide. Dissolving rosiglitazone in a dimethyl sulfoxide solution to prepare RSG/DMSO solutions with different concentration gradients. And then adding the mixture into spinning solution, preparing the spinning solution with different final concentrations, transferring the spinning solution into a glass syringe, and performing electrostatic spinning to form a rosiglitazone/PHBV slow-release membrane, wherein the rosiglitazone/PHBV slow-release membrane is used for resisting glaucoma filterability surgery, is safe, effective and durable in resisting scar formation after glaucoma surgery, improves the long-term success rate of glaucoma surgery, and has very important clinical significance.

Description

Preparation method and application of PHBV/rosiglitazone sustained-release membrane

Technical Field

The invention belongs to the technical field of medicines, and relates to a preparation method and application of a PHBV/rosiglitazone sustained-release membrane.

Background

Glaucoma is a neurodegenerative disease, is mainly characterized by progressive visual deterioration and visual field loss, is the leading cause of irreversible blindness in the world, and is also one of the blinding eye diseases which can be prevented and controlled. The ultimate goals of glaucoma treatment are primarily to control intraocular pressure and protect the optic nerve. Methods for controlling intraocular pressure mainly include drug, laser, and surgical treatment. The operation treatment mode is Glaucoma Filtration Surgery (GFS), which is also the main means for treating glaucoma at present, however, excessive proliferation of the postoperative Tenon's fibroblast cells (HTFs) leads to the formation of filter channel scarring, so that the failure rate of GFS after operation for 1 year is up to 15%, and antimetabolites, hormones, cyclosporine a and anti-vascular endothelial growth factors studied at home and abroad have obtained a certain curative effect, but it is difficult to effectively control rosiglitazone to control filter channel scarring for a long time. Meanwhile, the medicines cannot be applied individually, and often cause a series of damages to eye tissues due to overhigh concentration or overlong acting time, such as superficial corneal vascularization, bleb leakage, low-intraocular-pressure macular degeneration, endophthalmitis and the like. Therefore, the finding of a more effective, safer and more accurate medicament for resisting the scar after glaucoma operation has very important clinical significance.

PHBV is a biological polyester which is produced by adding 3-hydroxy glutaric acid in the process of synthesizing PHB by gram bacteria by using starch as a raw material. It is produced by bacteria that convert the feed to PHBV for storage and use as energy. When the accumulation reaches a certain degree, people use steam to crack the bacteria and collect purified PHBV. Compared with traditional high polymer materials such as PP and PGA, PHBV can be decomposed by bacteria under proper conditions, and the decomposition products are water and carbon dioxide. However, since the mechanical properties of PHBV are degraded by secondary crystallization at room temperature, PHBV cannot be directly used as a packaging material instead of PP or the like. PHBV has been used in the fields of packaging materials, filtration and separation and the like, and is less used in the medical field.

Rosiglitazone (RSG) acts as a class of PPAR γ agonists and inhibits tissue inflammation and fibrotic progression in a variety of diseases. In recent 10 years, under the funding of national science foundation (item numbers: 81170843, 81670859), the research of in vivo and in vitro experiments proves that rosiglitazone can effectively inhibit the scarring of the filter passage and a plurality of related SCI papers are published. Research on a specific regulation mechanism and a signal path of the rosiglitazone for inhibiting the scarring is still in progress, the existing research result proves that the rosiglitazone can regulate the scarring process through an autophagy-related path, and two related articles are drafted and submitted. Although the rosiglitazone prepared into eye drops with the concentration of 0.5 percent or subconjunctival injection has definite anti-scarring effect and few side effects, the medicine has short action time and needs repeated medication. In order to make it possible for rosiglitazone to be actually used in clinical practice, the present invention provides a new scientific problem based on the earlier research, and it is expected to find a more suitable ocular drug sustained release system (DDS) to increase the drug concentration of rosiglitazone in tissues and prolong the maintenance time thereof, so that the drug effect is more durable and the side effect is lower.

Disclosure of Invention

The invention aims to provide a preparation method and application of a PHBV/rosiglitazone sustained-release membrane which is more effective, safer and more accurate.

The specific technical scheme is as follows:

a preparation method of a PHBV/rosiglitazone sustained release membrane comprises the following steps:

step 1, weighing PHBV powder, dissolving the PHBV powder into chloroform TCM solution, magnetically stirring the solution at room temperature, ultrasonically dissolving the solution for 5 to 10 minutes at 50 to 70 ℃ until the solution is colorless and transparent, adding dimethylformamide DMF, and preparing PHBV-TCM/DMF spinning solution with the mass-volume ratio of 5 percent for later use;

step 2, dissolving rosiglitazone in a DMSO solution, preparing an RSG/DMSO solution with the concentration range of 0.5-50mg/ml, adding the RSG/DMSO solution into the previous spinning solution, preparing final spinning solutions with different concentrations, then transferring the final spinning solutions into a glass syringe, fixing the final spinning solutions into a clamping groove of an injection pump, connecting the positive electrode of a high-voltage power supply with a dull-ground metal needle at the front section of the syringe, connecting the negative electrode of the high-voltage power supply with an aluminum plate coated with aluminum foil paper, keeping the distance between the needle and a receiver at 10-15cm, adjusting the voltage of the high-voltage power supply to be 15KV, adjusting the propelling speed of the injection pump to be 5-8ml/h, controlling the ambient temperature to be 20-30 ℃ and the humidity to be;

and 3, after spinning is finished, uncovering the rosiglitazone/PHBV slow-release membrane on the aluminum foil paper, and drying for 3 days at room temperature to remove residual solvent.

Further, in the step 1, the PHBV powder is weighed to have a mass of 500mg-1 g.

Further, in step 1, the volume of chloroform TCM solution is 9ml to 18 ml.

Further, in step 1, dimethylformamide DMF was added in a volume of 1ml to 2 ml.

The PHBV/rosiglitazone sustained-release membrane is applied to the preparation process of the medicament for resisting the scar after glaucoma operation.

Compared with the prior art, the invention has the beneficial effects that:

the earlier stage research of the invention finds that the rosiglitazone has better effect on inhibiting the formation of the cicatrization, and in order to continuously inhibit the cicatrization, the rosiglitazone is prepared into a slow-release medicament which is more suitable for clinical patients, relieves the pain of the patients and improves the success rate of the operation.

The effective, safer and more accurate anti-glaucoma postoperative scar medicine has very important clinical significance.

The rosiglitazone/PHBV sustained-release membrane has good effect of continuously resisting the scar formation after glaucoma filtration, and has little side effect. Establishing a rosiglitazone/PHBV slow release fiber with uniform and fine diameter through electrostatic spinning, and finally forming a rosiglitazone/PHBV slow release film on a receiving plate; through in vitro release experiments, in vitro degradation experiments, cell experiments and in vivo animal experiments, the release mechanism, in vivo drug release behaviors, intraocular drug distribution and the like of the rosiglitazone/PHBV sustained-release membrane are discussed from multiple aspects, and meanwhile, the action time of the continuous effective anti-fibrosis of the rosiglitazone/PHBV is determined, and the effectiveness and the safety of the rosiglitazone/PHBV are verified. Lays a solid theoretical foundation for clinical popularization and application.

The method has the characteristics of more effectiveness, safety and accuracy, and is suitable for popularization and application.

Drawings

FIG. 1 is a diagram of a rosiglitazone/PHBV membrane provided in an embodiment of the present invention.

FIG. 2 shows an optical microscope photograph of a rosiglitazone/PHBV film after electrostatic spinning according to an embodiment of the present invention.

FIG. 3 is a block diagram of a rosiglitazone/PHBV sustained release film according to an embodiment of the present invention.

FIG. 4 shows the in vitro release and degradation experiments of the rosiglitazone/PHBV sustained release film provided by the embodiment of the present invention: and cutting each group of the slow release membranes into the slow release membranes with the same size, immersing the slow release membranes into physiological saline, and taking each time point to perform related detection graphs.

FIG. 5 is a graph showing the degree of fibrosis of HTFs induced by TGF- β 1 stimulation under the action of different concentrations of rosiglitazone/PHBV sustained release membranes.

Figure 6 in vivo experiments (animal experiments) provided by the present invention: establishing a rabbit eye GFS operation model, and verifying a pharmacokinetic and safety diagram of the rabbit eye GFS operation model through the intervention of a rosiglitazone/PHBV sustained-release membrane.

FIG. 7 shows the state of the rosiglitazone/PHBV sustained release film with different concentrations under a scanning electron microscope.

In the figure: the left side shows the surface morphology of each group under the 500-fold electron microscope, and the right side shows the surface morphology of each group under the 2,000-fold electron microscope. The final concentration of RSG in each group is as follows: control group (0mg/ml RSG), group A (0.05mg/ml RSG), group B (0.5mg/ml RSG), and group C (5mg/ml RSG).

FIG. 8 is a graph of the in vitro release curve of rosiglitazone/PHBV sustained release films with different concentrations provided by the embodiment of the present invention.

FIG. 9 is a graph showing the in vitro degradation behavior of various groups of rosiglitazone/PHBV sustained release membranes with different concentrations provided by the embodiment of the present invention.

Detailed Description

The technical solution of the present invention will be described in further detail with reference to specific embodiments.

rosiglitazone/PHBV release, degradation, in vivo experiments:

in vivo experiments, rosiglitazone/PHBV membrane was placed at the surgical site, i.e. under the conjunctiva, and the anti-scarring effect was observed.

Such as the rosiglitazone/PHBV membrane shown in FIG. 1. FIG. 2 is an optical microscope photograph of rosiglitazone/PHBV film after electrostatic spinning.

The invention is further described below with reference to specific assays.

1. Preparation of rosiglitazone/PHBV sustained-release membrane with different concentrations

By using an electrostatic spinning method, the spinning voltage, the advancing speed of an injection pump, the distance between a syringe needle and a receiving plate, the ambient temperature and the humidity are adjusted to prepare the rosiglitazone/PHBV sustained-release membranes with different final concentrations, namely 0mg/ml (control group), 0.05mg/ml (group A), 0.5mg/ml (group B) and 5mg/ml (group C). The optimum parameters are found to eject small, uniform diameter, fine fibers to form a uniform fiber film.

2. In vitro Release assay

High Performance Liquid Chromatography (HPLC), which is a high pressure liquid chromatography, is a sample analysis method developed on the basis of the classical liquid chromatography and having higher efficiency, higher speed and higher sensitivity. The release condition of the rosiglitazone/PHBV sustained-release membrane developed in the project along with the prolonging of time in vitro can be rapidly and efficiently detected by HPLC, and the cumulative release curve of the rosiglitazone is drawn. According to the in vitro release rosiglitazone curve of the rosiglitazone/PHBV sustained-release membrane, the time, peak and duration of the starting release are searched.

3. In vitro degradation experiments

The rosiglitazone/PHBV sustained release membranes with different concentrations, namely a control group (O mg/ml), a group A (0.05mg/ml), a group B (0.5mg/ml) and a group C (5mg/ml), are soaked in physiological saline, stored in a constant temperature incubator at 37 ℃, the pH value of the physiological saline solution is measured every week, the change of the pH value of each group along with the time is compared, and a relevant curve is drawn. Meanwhile, the surface morphology of each group of slow release membranes degraded within 1 month to half a year is observed through SEM. And (3) comparing the degradation speed of the slow release membranes with different concentrations and the pH value of a degradation product, and checking the influence of the rosiglitazone on the degradation of the PHBV.

4. In vitro studies (cell experiments)

Human subconjunctival Tenon's fibroblastic cells (HTFs) are subjected to primary culture, and an HTFs cell model and a cell model induced by TGF-beta 1 are established. Inoculating cells on the surface of a rosiglitazone/PHBV slow release membrane sterilized by ethylene oxide, placing the cells in a pore plate, culturing for 12h, 24h, 36h, 48h and 72h, observing the form of HTFs cells on the surface of the slow release membrane by a Scanning Electron Microscope (SEM), and observing and detecting the relative number of the cells by a laser confocal scanning microscope; cell counting kit-8(CCK8) is used for detecting the cell proliferation conditions of the HTFs on the surface of the rosiglitazone/PHBV with different concentrations, and immune fluorescence is used for detecting the expression conditions of related fibrin such as SP1, Collagen1, alpha-SMA, CTGF and the like. After the cells are collected, detecting related fibrin and mRNA expression conditions of various groups of SP1, Collagen1, alpha-SMA, CTGF and the like by a q-PCR and Western Blot detection method; and judging the proliferation change and the differentiation level to MFs of HTFs in the rosiglitazone/PHBV sustained-release membrane groups with different concentrations after the induction of TGF-beta 1, thereby further confirming the anti-fibrosis effect of the rosiglitazone.

5. In vivo study (animal experiments)

Establishing a rabbit eye glaucoma filterability operation (GFS) animal model, respectively placing rosiglitazone/PHBV sustained-release membranes with different concentrations and proper sizes (the sizes are most suitable for suturing the conjunctiva without shrinking the membrane and being slightly larger than a filtering bleb) below the scleral flap, above the scleral flap and below the scleral flap in the operation, and recording intraocular pressure fluctuation, the filtering bleb condition, side effects and the rosiglitazone drug concentration in aqueous humor within 1 month after the operation. Detecting the fibrosis of tissues (including conjunctiva, Tenon's capsule and sclera tissues) in a filtering bleb area at 1 week, 2 weeks, 4 weeks, 6 weeks, 8 weeks and 12 weeks after operation, detecting the fibrosis expression degree of each tissue by HE staining and massson staining, detecting the expression degree of each fibrosis index of SP1, Collagen1, alpha-SMA and CTGF by immunohistochemistry, and detecting the expression degree of each Collagen gene and protein by q-PCR and WB. Finding out the most suitable concentration, size and placement position of the rosiglitazone/PHBV sustained-release membrane and further proving the anti-fibrosis effect of the rosiglitazone.

The invention is further described with reference to specific examples.

1) Preparation of rosiglitazone/PHBV sustained-release membrane

Mixing PHBV powder, chloroform (TCM) and Dimethylformamide (DMF), and preparing into PHBV-TCM/DMF (9/1) spinning solution with mass-volume ratio of 5% for later use. Dissolving rosiglitazone in a dimethyl sulfoxide (DMSO) solution to prepare RSG/DMSO solutions with different concentration gradients. In order to find the most suitable RSG concentration, the RSG is prepared according to the following step-type concentration ratio: i.e., 0mg/ml (control group), 0.5mg/ml (group A), 5mg/ml (group B), 50mg/ml (group C). Then adding the mixture into spinning solution to prepare spinning solution with different final concentrations. And transferring the uniformly mixed spinning solution into a glass syringe for electrostatic spinning. After spinning, the rosiglitazone/PHBV slow release film on the aluminum foil paper is uncovered and dried for 3 days at room temperature to remove the residual solvent. And carrying out subsequent tests on the rosiglitazone/PHBV sustained-release membrane, finding out the optimal gradient concentration, and carrying out a concentration test near the gradient concentration to find out the optimal rosiglitazone concentration.

2) In vitro Release assay

The prepared rosiglitazone/PHBV sustained-release membranes with each concentration, namely a control group (Omg/ml), a group A (0.05mg/ml), a group B (0.5mg/ml) and a group C (5mg/ml), are dried for three days in a ventilated and dark place, after the solvent is volatilized, three fiber sustained-release membranes with the same size (6 x 6cm) are cut out from each group, each group of sustained-release membranes are immersed into appropriate physiological saline (sustained-release medium) with the same amount, the membranes are placed in an environment with the temperature of 37 ℃, the sustained-release medium with the same amount is taken at each time point, the concentration of the sustained-release medium is measured by using a high performance liquid chromatography according to a rosiglitazone standard substance, and the average value of the repeated three times of each group is taken. And detecting the release curve of each group of rosiglitazone sustained release films.

3) In vitro degradation experiments

The prepared rosiglitazone/PHBV slow release film is dried for three days at room temperature by ventilation and air drying; cutting out three sustained-release membranes with the same size from each of the control group, group A, group B and group C, measuring the pH value of normal saline, placing the sustained-release membranes in the normal saline with the same amount, and placing the membranes in an incubator at 37 deg.C. Every other week, the solution was changed and its pH was measured to detect the pH of each group of metabolites. The surface morphology of each group of the sustained-release membranes degraded in vitro for 1 month to half a year is observed by using SEM, and compared with the morphology change of undegraded sustained-release membranes.

4) In vitro experiments (cell experiments)

(1) Isolation and culture of human subconjunctival Tenon's follicle fibroblasts (HTFs): approved by the hospital committee of theory, a small amount of conjunctiva and subconjunctival tissues are taken out from the strabismus children operation, transferred into a sterile culture dish filled with a DMEM culture medium within half an hour, separated and sheared into conjunctival sac tissues, and the conjunctival sac tissues are sheared into tissue blocks with smaller thickness by using an ophthalmic scissors; adding 2.5g/L trypsin, then placing in an incubator at 37 ℃ for digestion for 10-20min, adding DMEM to stop digestion, blowing by using a gun head, removing tissue blocks by using a 200-mesh filter screen, collecting filtered liquid, centrifuging for 5-10min at 1000r/min, taking precipitates to be resuspended in a DMEM complete culture medium containing 10% FBS and 1% streptomycin, carrying out subsequent culture and generation after 2-5 days when cells adhere to the wall, and carrying out subsequent experiments by taking cells from the 3 rd generation to the 8 th generation.

(2) And (3) safety detection of the rosiglitazone/PHBV sustained-release membrane: after being sterilized by ethylene oxide, each group of rosiglitazone/PHBV sustained-release membranes with different concentrations are cut into proper sizes and placed in a pore plate, and HTFs are inoculated on the pore plate. Detecting cell proliferation and toxicity by using CCK 8; and observing the form of the HTFs growing on the surface of each group of slow release films by a scanning electron microscope. After TGF-beta 1 is added into each group for dry prediction, the proliferation capacity change and toxicity of HTFs are detected by CCK 8; detecting the expression condition of the fibrin related index through immunofluorescence, q-PCR and WB.

5) In vivo experiments (animal experiments)

(1) Establishing a rabbit eye filtering surgical model: new Zealand white rabbits were anesthetized systemically with 3% pentobarbital (30mg/kg), anesthetized on the ocular surface with Dunnuoxi, fixed, and then underwent microscopic simple trabeculectomy. The eye ointment is applied to the conjunctival sac one week after the operation. And the intraocular pressure of the rabbit eyes was measured by a Tono-lab tonometer at the same time every day after the operation, and the condition of the filter bleb was recorded and morphologically graded, and side reactions were recorded.

(2) Randomly dividing 150 New Zealand white rabbits with the weight of 2.5 kg-3 kg into 7 groups with 300 eyes in total, and uniformly selecting the right eye as an operation eye; the experimental groups were as follows:

1) blank control group: the left eye of each group of experimental rabbits was not subjected to any surgery and related treatment;

2) blank surgery control group: performing GFS operation on 10 randomly grouped experimental rabbits with 10 eyes, and infiltrating the scleral flap part with physiological saline for 5min in the operation;

3) positive operation control group 1: for 10 randomly grouped experimental rabbits, 10 eyes in total, a GFS operation was performed, in which a cotton piece impregnated with 0.4mg/ml of mitomycin C (MMC, 10 mg/bottle, lot number 679AFF02, manufactured by Kyowa Co., Ltd., Japan) was placed on the scleral flap portion for 5 min;

4) positive operation control group 2: for 10 randomly grouped experimental rabbits, 10 eyes are totally treated, GFS operation is carried out, and cotton sheets soaked with 5-FU stock solution (250mg/10ml) are used in the operation and are placed on the scleral flap part for 5 min;

5) experimental groups: 120 randomly grouped experimental rabbits with 120 eyes are subjected to GFS operation, and slow release membranes (Omg/ml, 0.05mg/ml, 0.5mg/ml and 5mg/ml) with different concentrations and proper sizes are placed above and/or below a scleral flap during the operation;

after operation, monitoring the daily intraocular pressure fluctuation of rabbit eyes and side effects (such as conjunctival congestion, local inflammatory reaction, corneal epithelial injury, bleb leakage, infection and the like), and grading the blebs according to an IBAGS system (comprising four indexes: height, breadth, vascularity and Seidel test); collecting aqueous humor to carry out rosiglitazone concentration detection, and drawing a rosiglitazone in-vivo release curve; detecting the form of the filtering bleb, the opening condition of the filtering channel and the existence condition of the rosiglitazone/PHBV membrane of an experimental group by UBM every week after operation; meanwhile, filtering area tissues of the same part of each group are taken down at several time points of 1w, 2w, 4w, 6w, 8w and 12w after operation, and tissues of the same part are cut off in a blank control group for subsequent experiments.

(3) And (3) comparing the opening conditions of the filtration channels of each group at different time points by adopting HE (high intensity intrinsic) staining on the tissue of the filtration area taken off from each group, and detecting and comparing the fibrosis degree of each group by adopting a massson staining technology.

(4) And (3) adopting an immunohistochemical technology, carrying out paraffin embedding on the tissue of the filtering area removed from each group, carrying out SP1, Collagen1, alpha-SMA and CTGF antibody labeling, and detecting the expression condition of each fibrosis index.

(5) The mRNA expression of fibrosis indexes SP1, Collagen1, alpha-SMA and CTGF at different time points in the tissues is detected by using a q-PCR technology.

(6) Western Blot was used to detect the expression levels of SP1, Collagen1, alpha-SMA and CTGF proteins in the tissues of each filtration area.

The invention is further described below in connection with specific technical lines.

Preparing a rosiglitazone/PHBV sustained-release membrane: mixing PHBV powder, TCM, DMF, RSG/DMSO to form spinning solution, and preparing the rosiglitazone/PHBV sustained-release membrane by an electrostatic spinning method. As shown in fig. 3.

In vitro release and degradation experiments of rosiglitazone/PHBV sustained release membrane: and cutting each group of the slow release membranes into the slow release membranes with the same size, immersing the slow release membranes into physiological saline, and performing related detection at each time point. As shown in fig. 4.

In vitro cell experiments: the growth condition of HTFs cells on the surface of the rosiglitazone/PHBV sustained-release membrane is researched, and the fibrosis degree of HTFs under the action of the rosiglitazone/PHBV sustained-release membranes with different concentrations after the HTFs are stimulated and induced by TGF-beta 1 is researched. As shown in fig. 5.

In vivo experiments (animal experiments): establishing a rabbit eye GFS surgical model, and verifying the pharmacokinetics and safety of the rabbit eye GFS surgical model through the intervention of a rosiglitazone/PHBV sustained release membrane. As shown in fig. 6.

The present invention will be further described with reference to effects.

On the basis of deep research on the effect and mechanism of the rosiglitazone medicament in the early stage, the invention further tests the effective anti-scar effect of the rosiglitazone and simultaneously provides a new scientific problem of developing a novel rosiglitazone medicament slow release system for continuous anti-scar effect after GFS operation so as to prolong the action time of the medicament and improve the long-term success rate of anti-glaucoma operation. Therefore, the compound has bright clinical application and transformation characteristics and has the following innovations:

the rosiglitazone belongs to thiazolidinediones insulin sensitizer and is commonly used for treating type 2 diabetes, the early-stage research of the project group finds that the rosiglitazone can effectively inhibit the scar formation of glaucoma filtration surgery, innovatively applies the rosiglitazone to ophthalmology, deeply researches a rosiglitazone drug sustained release system, and lays a solid foundation for the real clinical application in the future.

As a novel spinning technology, electrostatic spinning is also commonly applied to the medical industry at present, such as cardiovascular stents, bone nails, sutures and the like. The project innovatively introduces the electrostatic spinning technology into the development of the ophthalmic drug sustained-release system, provides new possibility for the development of the electrostatic spinning technology, and also provides a new idea for the preparation of the anti-glaucoma drug sustained-release system.

PHBV has been used in the fields of packaging materials, filtration and separation and the like, and is less used in the medical field. The rosiglitazone/PHBV sustained-release membrane developed by the item is a new exploration for the application of PHBV materials, and has pioneering meaning.

The invention successfully prepares the rosiglitazone/PHBV sustained release film with small diameter, uniformity and different concentrations, and observes the surface morphology of the sustained release film by an environmental scanning electron microscope.

The rosiglitazone/PHBV sustained release membranes with different concentrations have little influence on the proliferation of HTFs and have weak toxicity; meanwhile, compared with a control group, the experimental group can effectively inhibit the fibrosis degree.

The invention successfully verifies the effectiveness, the persistence and the safety of the rosiglitazone/PHBV sustained-release membrane.

The surface morphology of the rosiglitazone/PHBV sustained-release film under a scanning electron microscope is as follows:

FIG. 7 shows the forms of rosiglitazone/PHBV sustained release films with different concentrations under a scanning electron microscope. The left side shows the surface morphology of each group under the 500-fold electron microscope, and the right side shows the surface morphology of each group under the 2,000-fold electron microscope. The final concentration of RSG in each group is as follows: control group (0mg/ml RSG), group A (0.05mg/ml RSG), group B (0.5mg/ml RSG), and group C (5mg/ml RSG).

rosiglitazone/PHBV sustained release membrane in vitro release experiment

The rosiglitazone release profile can be seen in figure 8. From the release profile, it can be seen that as the concentration of rosiglitazone increases, the initial release time of rosiglitazone advances and the concentration increases. The rosiglitazone/PHBV sustained-release membranes with different concentrations reach the peak value when in vitro release is carried out for 16 days, and the rosiglitazone can be effectively and continuously released in one month. Subsequent time rosiglitazone concentration measurements are also in progress.

In vitro degradation behavior of rosiglitazone/PHBV sustained release film

From FIG. 9, it can be seen that the in vitro degradation product of rosiglitazone/PHBV sustained release membrane is acidic, and the product PH increases with time; the higher the concentration of rosiglitazone, the more acidic the in vitro degradation product, but it has no obvious statistical significance, i.e. the concentration of rosiglitazone does not affect the in vitro degradation behavior of PHBV.

The above description is only a preferred embodiment of the present invention, and the scope of the present invention is not limited thereto, and any simple changes or equivalent substitutions of the technical solutions that can be obviously obtained by those skilled in the art within the technical scope of the present invention are within the scope of the present invention.

Claims

1. A preparation method of a PHBV/rosiglitazone sustained release membrane is characterized by comprising the following steps:

step 2, dissolving rosiglitazone in a DMSO solution, preparing an RSG/DMSO solution with the concentration range of 0.5-50mg/ml, adding the RSG/DMSO solution into the previous spinning solution, preparing final spinning solutions with different concentrations, then transferring the final spinning solutions into a glass syringe, fixing the final spinning solutions into a clamping groove of an injection pump, connecting the positive electrode of a high-voltage power supply with a dull metal needle at the front section of the syringe, connecting the negative electrode of the high-voltage power supply with an aluminum plate coated with aluminum foil paper, keeping the distance between the needle and a receiver at 10-15cm, adjusting the voltage of the high-voltage power supply to be 15KV, adjusting the propelling speed of the injection pump to be 5-8ml/h, controlling the ambient temperature to be 20-30 ℃ and the humidity to be 20-;

and 3, after spinning is finished, removing the rosiglitazone/PHBV slow release film on the aluminum foil paper, and drying for 3 days at room temperature to remove residual solvent.

2. The method for preparing the PHBV/rosiglitazone sustained release membrane of claim 1, wherein in the step 1, the PHBV powder is weighed to have a mass of 500mg to 1 g.

3. The method for preparing the PHBV/rosiglitazone sustained release membrane as claimed in claim 1, wherein in the step 1, the volume of the chloroform TCM solution is 9ml to 18 ml.

4. The method for preparing PHBV/rosiglitazone sustained release membrane according to claim 1, wherein the volume of DMF in step 1 is 1ml to 2 ml.

5. The PHBV/rosiglitazone sustained release membrane prepared by the method of claim 1 is applied to the preparation process of the medicament for resisting the scar after glaucoma operation.