A method for preparing a gastric residence shape memory composite film

Abstract

The application relates to a preparation method of a gastric-retention shape memory composite film, which comprises the following steps: (1) mixing a polymer with dichloromethane to obtain solution 1; (2) preparing solution 2 with water as a solvent, wherein the solution 2 contains hydroxypropyl methyl cellulose and NaHCO3; (3) mixing the solution 1 and the solution 2, stirring, adding a luteolin phospholipid complex, continuing to stir, and volatilizing dichloromethane and water. The shape memory film provided by the application has good unfolding effect in the body, the deformation temperature is closer to the human body temperature, the permeation of luteolin to the biological membrane can be strengthened, the gastrointestinal absorption effect of luteolin is improved, the stomach discomfort of patients is reduced, and the bioavailability of the drug is improved.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of medicine, and particularly relates to a preparation method of a gastric-retention shape memory composite film. BACKGROUND

[0002] Luteolin is a natural flavonoid component, which is widely present in plants, especially in fruits and vegetables, such as celery, chrysanthemum, sweet pepper, carrot, onion leaves and broccoli. The chemical formula is C 15 H 10 O6. The structural formula is as shown below.

[0003]

[0004] Luteolin can form a chelate with metal ions, and is not oxidized in the chelation process. Luteolin has various biological activities, including anticancer, antioxidant, anti-inflammatory, immunomodulatory and cardioprotective effects.

[0005] Conventional oral drug therapy for treating gastric cancer has problems of not reaching the expected efficacy, poor oral effect and difficult preservation, which seriously limits the application. Luteolin has good antitumor activity in gastric cancer and is a very potential anticancer drug. However, direct application of luteolin to the human body has certain deficiencies, such as poor oral absorption, poor water solubility, short in vivo retention time and low bioavailability. Especially for the treatment of gastric diseases, there are problems of too short residence time in the stomach and inability to achieve sustained-release effect.

[0006] In order to effectively utilize luteolin, increase its oral absorption and improve its therapeutic potential, especially for the treatment of human gastric cancer, it is urgent to research a new luteolin-related dosage form. SUMMARY

[0007] In view of the problems existing in the prior art, the present application provides a preparation method of a gastric-retention shape memory composite film, which is a new gastric-retention shape memory composite film loaded with luteolin-containing phospholipid complex (LPC).

[0008] The technical scheme for solving the above technical problems is as follows:

[0009] The present application provides a preparation method of a gastric-retention shape memory composite film, comprising the following steps:

[0010] (1) mixing a polymer with dichloromethane to obtain solution 1;

[0011] (2) preparing solution 2 with water as a solvent, and solution 2 contains hydroxypropyl methyl cellulose and NaHCO3;

[0012] (3) mixing solution 1 and solution 2, stirring, adding luteolin phospholipid complex, continuing to stir, and evaporating dichloromethane and water.

[0013] The beneficial effects of adopting the above technical solutions include:

[0014] In view of the problem of high brittleness of the polymer, hydroxypropyl methyl cellulose (HPMC) is added for modification, so that it is more suitable for oral administration of the human body and reduces the discomfort of the patient's stomach. In order to increase the retention time of the gastric retention shape memory composite film, a foaming agent NaHCO3 is added to the composite film. When the composite film reaches the stomach, the acidic stomach environment causes the composite film to produce bubbles, and the floating performance is stronger.

[0015] The shape memory film prepared by the application has good unfolding effect in the body, and the deformation temperature is closer to the body temperature, which can strengthen the penetration of luteolin to the biological membrane, improve the gastrointestinal absorption effect of luteolin, and improve the bioavailability of the drug. The water solubility of luteolin is increased, the drug release behavior is good, the retention time is increased, the patient's compliance to the drug is increased, the glass transition temperature, the drug release rate, and the composite film deformation rate are suitable, the stability is good, the drug use frequency can be effectively controlled, the damage of drug toxicity to organs can be reduced, the patient's drug compliance can be increased, and the drug cost can be reduced. It has obvious inhibitory effect on gastric cancer cells.

[0016] Further, the mass ratio of the polymer to dichloromethane is 1:7.

[0017] Further, the mass of the hydroxypropyl methyl cellulose is 5% of the mass of the polymer, the mass of the NaHCO3 is 5% of the mass of the polymer, and the mass ratio of the total mass of the hydroxypropyl methyl cellulose and the NaHCO3 to the mass of the water is 1:10.

[0018] Further, the mass of the luteolin phospholipid complex is 30% of the total mass of the polymer, the hydroxypropyl methyl cellulose, and the NaHCO3.

[0019] The beneficial effects of adopting the above technical solutions include: the above method can be used to prepare a uniform composite film. The addition of hydroxypropyl methyl cellulose and NaHCO3 can effectively reduce the hardness and brittleness of the polymer, so that the composite film is easier to fold and more suitable for oral use.

[0020] Further, the preparation method of the luteolin phospholipid complex comprises the following steps: dissolving soybean lecithin and luteolin with tetrahydrofuran, stirring, and recovering under reduced pressure to obtain the luteolin phospholipid complex.

[0021] The beneficial effects of the above technical solutions include: in view of the problems of poor water solubility and low bioavailability of the main drug luteolin, the luteolin is modified in the application, coated with soy lecithin, the water solubility of luteolin is increased, the human body is more conducive to absorption, and the bioavailability of luteolin is increased.

[0022] Further, the mass ratio of luteolin to soy lecithin is 1:1.2; after being dissolved by tetrahydrofuran, the concentration of luteolin is 10 mg / mL; the stirring time is 6 h, and the stirring temperature is 50℃.

[0023] The beneficial effects of the above technical solutions include: the luteolin phospholipid complex prepared by the above method can reach a complexing rate of 91.45%, which is conducive to the preparation of the subsequent gastric retention shape memory composite film. On the other hand, the water solubility of the raw drug luteolin itself can be improved, which is more conducive to human body absorption.

[0024] Further, the polymer is selected from one or a combination of polylactic acid (PLA), polycaprolactone (PCL), and polyethylene glycol (PEG).

[0025] Preferably, the polymer includes polylactic acid (PLA). The polymer can be selected from polylactic acid (PLA) alone. Alternatively, the polymer can include one or a combination of polylactic acid (PLA), polycaprolactone (PCL), and polyethylene glycol (PEG) in addition to polylactic acid (PLA). Most preferably, the polymer includes polylactic acid (PLA) and polyethylene glycol (PEG), and the mass ratio of polylactic acid (PLA) to polyethylene glycol (PEG) is 7:3 for better effect.

[0026] The beneficial effects of the above technical solutions include: the application prepares a novel natural drug-loaded thermal-responsive gastric retention shape memory composite film for the treatment of gastric cancer, which can be used for oral chemotherapy of patients to increase the cure rate of patients with gastric cancer. The application selects luteolin with anticancer effect as the main drug, uses PLA as the main material, and modifies PLA by adding PCL and PEG to reduce the glass transition temperature of the composite film, so that the shape memory temperature is more suitable for the human body. The prepared gastric retention shape memory composite film is folded and placed in a capsule, and is orally taken by a patient into the stomach, and can be deformed back to the original shape at the temperature of the stomach, so that the area is larger than the pylorus, and the gastric retention time of the drug in the stomach is increased.

[0027] Further, in step (3), after stirring for 3.5 h, the luteolin phospholipid complex is added, and stirring is continued for 0.5 h.

[0028] Further, after the dichloromethane and water are evaporated, a molding step is further included; the molding is performed to obtain a sheet with a thickness of 1 mm, and the pressure is 10 Mpa.

[0029] The beneficial effects of the above technical scheme include: the phospholipid complex is uniformly loaded on the film agent; and the mold forming can ensure the uniform thickness of the prepared composite film and control the variable.

[0030] The application provides application of the gastric-retention shape memory composite film in preparation of a drug for treating gastric cancer.

[0031] The application provides a PLA-based gastric-retention shape memory composite film loaded with luteolin phospholipid complex (LPC). First, luteolin is complexed with soybean lecithin to prepare the LPC, so that the water solubility of the luteolin is increased. Then, the PLA-based gastric-retention shape memory composite film loaded with the LPC is prepared, PLA, PCL and PEG are used as polymer materials, and HPMC and NaHCO3 are added to modify the PLA, so that the prepared gastric-retention shape memory composite film can better rely on the temperature in the body to induce the shape memory reaction in the stomach, and the gastric retention time is increased. The system is a pioneering new drug delivery system, and can be used for future research and preparation of new gastric cancer drugs. BRIEF DESCRIPTION OF DRAWINGS

[0032] Figure 1 It is a schematic diagram of the gastric-retention drug delivery system. Figure 2 It is an ultraviolet absorption spectrum. Figure 3 It is a standard curve of luteolin. Figure 4 It is a diagram of laser irradiation of the LPC solution. Figure 5 It is a diagram of the finished product of the LPC. Figure 6 It is a morphology diagram of the LPC under an optical microscope. Figure 7 It is an ultraviolet spectrum, and in the curve corresponding to 400 nm, from bottom to top, the curves of Phospholipid, LPC and Luteolin are arranged in sequence. Figure 8 It is an infrared spectrum, and from bottom to top, the curves of LPC, Phospholipid and Luteolin are arranged in sequence. Figure 9 It is an appearance diagram of the drug-loaded gastric-retention shape memory composite film, wherein: (A) LPC-PLA; (B) LPC-PLA / PCL (9 / 1); (C) LPC-PLA / PCL (8 / 2); (D) LPC-PLA / PCL (7 / 3); (E) LPC-PLA / PEG (9 / 1); (F) LPC-PLA / PEG (8 / 2); (G) LPC-PLA / PEG (7 / 3); (H) LPC-PLA / PCL / PEG (8 / 2 / 1); (I) LPC-PLA / PCL / PEG (8 / 2 / 2); (J) LPC-PLA without HPMC and NaHCO3; (K) LPC-PLA / PCL / PEG (8 / 2 / 3). Figure 10Infrared spectrum analysis results of the gastric retention shape memory composite films, wherein: (A) from top to bottom, LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3); (B) from top to bottom, LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3); (C) from top to bottom, LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2), LPC-PLA / PCL / PEG (8 / 2 / 3). Figure 11 Differential scanning calorimetry curves of each composite film, wherein: (A) DSC curves of LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3); (B) curves corresponding to 150℃, from top to bottom, LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3); (C) curves corresponding to 175℃, from top to bottom, LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 3), LPC-PLA / PCL / PEG (8 / 2 / 2). Figure 12 Thermogravimetric curves of each composite film, wherein: (A) curves corresponding to 300℃, from top to bottom, PLA, LPC-PLA / PCL (7 / 3), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (9 / 1), LPC-PLA; (B) curves corresponding to 300℃, from top to bottom, LPC-PLA / PEG (7 / 3), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (9 / 1); (C) curves corresponding to 300℃, from top to bottom, LPC-PLA / PCL / PEG (8 / 2 / 3), LPC-PLA / PCL / PEG (8 / 2 / 2), LPC-PLA / PCL / PEG (8 / 2 / 1). Figure 13are the results of optical microscope observation of the composite films, wherein: (A) PLA; (B) LPC-PLA; (C) LPC-PLA / PCL (9 / 1); (D) LPC-PLA / PCL (8 / 2); (E) LPC-PLA / PCL (7 / 3); (F) LPC-PLA / PEG (9 / 1); (G) LPC-PLA / PEG (8 / 2); (H) LPC-PLA / PEG (7 / 3); (I) LPC-PLA / PCL / PEG (8 / 2 / 1); (J) LPC-PLA / PCL / PEG (8 / 2 / 2); (K) LPC-PLA / PCL / PEG (8 / 2 / 3). Figure 14 are the results of the floating of the gastro-retentive shape memory composite films at different times. Figure 15 are the results of the floating of the gastro-retentive shape memory composite films at different times. Figure 16 are the results of the floating of the gastro-retentive shape memory composite films at different times. Figure 17 are the results of the shape memory performance observation of the composite films at a temperature of 50 °C, wherein: (A) PLA; (B) LPC-PLA; (C) LPC-PLA / PCL (9 / 1); (D) LPC-PLA / PCL (8 / 2); (E) LPC-PLA / PCL (7 / 3); (F) LPC-PLA / PEG (9 / 1); (G) LPC-PLA / PEG (8 / 2); (H) LPC-PLA / PEG (7 / 3); (I) LPC-PLA / PCL / PEG (8 / 2 / 1); (J) LPC-PLA / PCL / PEG (8 / 2 / 2); (K) LPC-PLA / PCL / PEG (8 / 2 / 3). Figure 18 are the results of the shape memory performance observation of the composite films at a temperature of 37 °C, wherein: (A) PLA; (B) LPC-PLA; (C) LPC-PLA / PCL (9 / 1); (D) LPC-PLA / PCL (8 / 2); (E) LPC-PLA / PCL (7 / 3); (F) LPC-PLA / PEG (9 / 1); (G) LPC-PLA / PEG (8 / 2); (H) LPC-PLA / PEG (7 / 3); (I) LPC-PLA / PCL / PEG (8 / 2 / 1); (J) LPC-PLA / PCL / PEG (8 / 2 / 2); (K) LPC-PLA / PCL / PEG (8 / 2 / 3). Figure 19 are the results of the drug release curve of luteolin. Figure 20The effect of different material ratios on the in vitro release of luteolin in the composite film, the corresponding curve of 24h, from top to bottom: LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 3), LPC-PLA / PCL / PEG(8 / 2 / 2), LPC-PLA / PEG(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA, LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL(9 / 1). Figure 21 The effect of the additive amount of HPMC on the in vitro release of luteolin in LPC-PLA / PEG(7 / 3), from top to bottom: 5%, 3%, 1%. Figure 22 The effect of the additive amount of NaHCO3 on the in vitro release of luteolin in LPC-PLA / PEG(7 / 3), from top to bottom: 5%, 3%, 1%. Figure 23 The effect of the content of luteolin in the composite film on the in vitro release of luteolin in LPC-PLA / PEG(7 / 3), from top to bottom: 30%, 20%, 10%. Figure 24 The cell survival rate of SGC-7901 gastric cancer cells inoculated with non-drug composite film extract of the same concentration gradient; the results are expressed as mean ± standard deviation; (A) PLA; (B) PLA / PCL(7 / 3); (C) PLA / PEG(7 / 3); (D) LPC-PLA / PCL / PEG(8 / 2 / 3); in each group, from left to right: 24h, 48h, 72h. Figure 25 The proliferation inhibition rate of SGC-7901 gastric cancer cells to 100% drug-loaded composite film extract, from left to right in each group: LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), LPC-PLA / PCL / PEG(8 / 2 / 3). Figure 26 The morphological diagram of the blank group cells under the inverted microscope, where: A 24h; B 48h; C 72h. Figure 27 The morphological diagram of SGC-7901 gastric cancer cells cultured with 100% extract of PLA, PLA / PCL, PLA / PEG(7 / 3), PLA / PCL / PEG(8 / 2 / 3) under an inverted microscope. Figure 28Morphology of SGC-7901 gastric cancer cells cultured by 100% leaching solution of LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3), LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3), LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2), LPC-PLA / PCL / PEG (8 / 2 / 3) under an inverted microscope. Figure 29 Investigate the chromatogram for specificity, wherein: A is blank plasma; B is blank plasma + luteolin standard + internal standard; C is plasma after administration. Figure 30 Standard curve of luteolin / diosmetin (n = 6). Figure 31 Pharmacokinetic curve of luteolin, LPC and LPC-PLA / PEG (7 / 3) composite film in rats, 8h corresponding curve, from top to bottom, LPC-PLA / PEG (7 / 3), LPC, luteolin. DETAILED DESCRIPTION

[0033] The principles and features of the present application are described below in conjunction with the accompanying drawings, and the examples are only used to explain the present application and are not intended to limit the scope of the present application.

[0034] The present application provides a new gastric retention drug delivery system, which combines shape memory polymers with modified excipients to prepare drug-loaded gastric retention shape memory composite films, and explores the development and application potential of the gastric retention drug delivery system. The gastric retention drug delivery system has obvious advantages, such as sustained-release characteristics, which can increase the retention time of the drug in the stomach and maintain stable blood drug concentration, reduce the frequency of drug administration, and therefore has good clinical application prospects.

[0035] During the research, the inventors tried various preparation methods and unexpectedly found that the preparation method provided by the present application has better effect. For example, the inventors tried to first lay the non-drug-loaded composite film, and then lay the drug, but found that the method had poor effect and could not guarantee uniform distribution of the drug on the composite film. For another example, during the research, it was found that the proportion of the drug should not be too large (not more than 30%), and too large proportion of the drug makes the composite film very brittle, which is not easy to fold and use.

[0036] The application successfully designs and prepares 10 groups of luteolin phospholipid complex (LPC) polylactic acid (PLA) based gastric retention shape memory composite membranes with different polymer proportions. The polymer materials of the composite membrane include PLA, polycaprolactone (PCL), polyethylene glycol (PEG), and an appropriate amount of hydroxypropyl methyl cellulose (HPMC) and sodium bicarbonate (NaHCO3) are added to modify the composite membrane, further enhance the gastric retention effect, prolong its in vivo action time and improve the drug bioavailability. The research results of the application will lay a theoretical foundation for the development and application of new gastric retention drug delivery systems.

[0037] (1) Prepare LPC, determine the luteolin content in LPC by high performance liquid chromatography (HPLC), select the best LPC with the highest complex rate through orthogonal test, and select the prescription with the highest complex rate for the next step of preparation of gastric retention shape memory composite membrane. And the best phospholipid complex is observed by optical microscope, ultraviolet visible spectrophotometer, infrared spectroscopy and solubility analysis. According to the results of orthogonal test, the best preparation conditions of LPC are determined: luteolin and soybean lecithin are weighed according to the mass ratio of 1:1.2, an appropriate amount of tetrahydrofuran is added to make the mass concentration of the former (luteolin) 10mg / mL, constant temperature stirring at 50℃ for 6h, removal of organic solvent by rotary evaporation under reduced pressure, and drying in 40℃ vacuum drying oven overnight. By calculation, the average complex rate is 91.45%. The quality evaluation of LPC includes optical microscope observation, ultraviolet test, infrared test and solubility test. The experimental results show that luteolin can be well wrapped by soybean lecithin, and the water solubility of LPC is 2.29 times that of luteolin.

[0038] (2) 10 groups of drug-loaded gastric retention shape memory composite films were prepared, which were LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3), LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3), LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2) and LPC-PLA / PCL / PEG (8 / 2 / 3). The drug-loaded gastric retention shape memory composite films were characterized by Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA); the morphology of the drug-loaded gastric retention shape memory composite films was observed by optical microscopy; the shape memory performance, floating performance and drug release behavior of the prepared drug-loaded gastric retention shape memory composite films were tested and evaluated. The characterization results show that the drug-loaded gastric retention shape memory composite films all have good thermodynamic properties, and the materials do not denature during preparation; optical microscopy observation shows that the prepared gastric retention shape memory composite films are well mixed; drug release test shows that the drug release behavior of the composite films is good, and the release behavior is related to the ratio of polymers, the proportion of excipients and the drug content. Shape memory performance test shows that the recovery rates of the 10 kinds of gastric retention shape memory composite films prepared by the application at 50℃ are between 70-80%, and the recovery rates of the composite films at 37℃ are between 60-70%, which can realize large size deformation. The floating test results show that the composite films have good floating property, and the floating time is more than 8h, and part of the floating time is up to 48h. It is found through comprehensive characterization and evaluation that the performance of LPC-PLA / PEG (7 / 3) is better, and it is further studied for pharmacokinetics.

[0039] (3) The in vitro inhibition effect of the above-mentioned 10 groups of drug-loaded composite films on SGC-7901 gastric cancer cells was evaluated by MTT experiment. The non-drug-loaded composite film was subjected to MTT experiment in the same method to study the toxicity of the PLA, PLA / PCL (7 / 3), PLA / PEG (7 / 3) and PLA / PCL / PEG (8 / 2 / 3) composite film extracts on SGC-7901 gastric cancer cells. The inverted microscope was selected to directly observe the influence of the drug-loaded and non-drug-loaded PLA, PLA / PCL (7 / 3), PLA / PEG (7 / 3) and PLA / PCL / PEG (8 / 2 / 3) composite films on the number and morphology of SGC-7901 gastric cancer cells for 72 hours. The results show that the PLA, PLA / PCL (7 / 3), PLA / PEG (7 / 3) and PLA / PCL / PEG (8 / 2 / 3) composite film extracts do not exhibit cytotoxicity on SGC-7901 gastric cancer cells after incubation for 24 hours, 48 hours and 72 hours. The 100% drug-loaded composite film extract has obvious inhibition effect on SGC-7901 gastric cancer cells. When the composite film is LPC-PLA / PEG (7 / 3), the 100% composite film extract has stronger inhibition effect on SGC-7901 gastric cancer cells than the other drug-loaded composite film groups, which is related to the best drug release property. The inverted microscope observation results also prove this point. The non-drug-loaded composite film extract has no obvious toxicity on SGC-7901 gastric cancer cells, and the cell morphology is normal. The drug-loaded composite film extract can significantly increase the number of dead SGC-7901 gastric cancer cells and obviously change the cell morphology, and has obvious cytotoxicity on SGC-7901 gastric cancer cells.

[0040] (4) The HPLC method for determining the content of luteolin in blood plasma was established, and the pharmacokinetics of luteolin, LPC and LPC-PLA / PEG (7 / 3) in rats was studied. The pharmacokinetic analysis software DAS 2.0 was used to analyze the pharmacokinetic test results, and the pharmacokinetic parameters were analyzed by IBM SPSS Statistics 26.0. The results show that the HPLC method established in the application is simple and stable, has good specificity, and the recovery rate, precision and repeatability meet the determination requirements, and can be used for determining the content of luteolin in rat blood plasma. The pharmacokinetic test results show that the LPC-PLA / PEG (7 / 3) can effectively improve the pharmacokinetic characteristics of luteolin in rats, and has the highest bioavailability compared with luteolin and LPC.

[0041] The schematic diagram of the gastric retention drug delivery system provided by the application is shown as Figure 1 The preparation of luteolin phospholipid complex with the optimal complex ratio; the preparation of non-drug-loaded composite film; the preparation of luteolin phospholipid complex-loaded composite film; the loading of drug-loaded composite film into capsules; and the capsule is administered intragastrically in rats to expand the original shape and achieve gastric retention.

[0042] The present application uses luteolin as a model drug with anti-cancer effect and relatively low price, and the luteolin is complexed with soybean lecithin to improve the water solubility. The prepared phospholipid complex is loaded on a PLA-based thermal-responsive gastric retention shape memory composite film to prepare a multi-functional gastric retention drug delivery system with extensibility and floatability. The system can prolong the drug retention time in the stomach, increase the drug release time in the human body, improve the drug bioavailability, increase the patient's compliance to the drug, effectively control the number of drug administration, reduce the damage of drug toxicity to organs, increase the patient's medication adherence and reduce the cost of drug administration, and make more patients use the gastric retention system for anti-cancer adjuvant therapy conveniently. The system provides a novel drug delivery method for high incidence of gastric diseases and lays a foundation for the research of new drug dosage forms with high efficiency and low toxicity for the treatment of gastric diseases. The combination of the new gastric retention drug delivery system and the shape memory polymer can be used for the treatment of gastric cancer.

[0043] The following will be described by specific examples. The experimental methods used in each example are conventional experimental methods in the art unless otherwise specified. The materials, reagents and instruments used are conventional materials, reagents and instruments in the art unless otherwise specified, and can be obtained through commercial channels or prepared by conventional methods.

[0044] Experimental reagents:

[0045] Acetonitrile (chromatographically pure), tetrahydrofuran (chromatographically pure) were purchased from Merck Company in Germany; dimethyl sulfoxide (DMSO) was purchased from BIOFROX Company in Germany; MTT was purchased from Sigma Company in the United States; PBS solution was purchased from Gibco Company in the United States; trypsin was purchased from Solabio Company in the United States; sodium heparin was purchased from Alfa Aesar Company in the United States; polylactic acid, polycaprolactone, polyethylene glycol, hydroxypropyl methyl cellulose, sodium bicarbonate, soybean lecithin, and luteolin (pharmaceutical, PC>98%) were purchased from Shanghai Maikelin Biotechnology Co., Ltd.; carboxymethyl cellulose sodium (CMC-Na) was purchased from Shanghai Maikelin Biotechnology Co., Ltd.; dichloromethane (analytically pure), methanol (analytically pure), and perchloric acid (analytically pure) were purchased from National Pharmaceutical Group Chemical Reagent Co., Ltd.; luteolin (pharmaceutical, PC>98%) was purchased from Chengdu Man Sit Biological Technology Co., Ltd.; potassium dihydrogen phosphate (analytically pure) was purchased from Chengdu Kelong Chemical Reagent Factory; hydrochloric acid (analytically pure) was purchased from Tianjin Kemio Chemical Reagent Co., Ltd.; potassium bromide was purchased from Shandong Tianxin Chemical Industry Co., Ltd.; DMEM medium was purchased from Beijing Solarbio Company; fetal bovine serum was purchased from Hangzhou Sijiqing Biological Engineering Company; ethyl acetate (analytically pure) was purchased from Xilong Scientific Chemical Reagent Co., Ltd.; distilled water was prepared in the laboratory.

[0046] Experimental instruments:

[0047] High performance liquid chromatograph (Waters 1525) was purchased from Waters Corporation, USA; optical microscope (VHX-900) was purchased from Keyence Corporation, USA; Fourier infrared spectrometer (AVATAR 360) was purchased from Nicolet Instrument Corporation, USA; ultraviolet visible spectrophotometer (UV-5200PC) was purchased from Shanghai Yuanzhi Instrument Co., Ltd.; ultraviolet visible spectrophotometer (UV-7) was purchased from Mettler Toledo, Switzerland; constant temperature water bath (DK-98-IIA) was purchased from Shanghai Qunxian Instrument and Equipment Co., Ltd.; thermal gravimetric analyzer (TGA-101) was purchased from Shanghai Zhunquan Instrument and Equipment Co., Ltd.; differential scanning calorimeter (DSC-500L) was purchased from Shanghai Zhunquan Instrument and Equipment Co., Ltd.; inverted microscope (CKX41SF) was purchased from Olympus Corporation, Japan; optical microscope (CX21) was purchased from OLYMPUS, Japan; microplate reader (iMark) was purchased from Bio-Rad, USA; refrigerator (BCD-215KANDZ) was purchased from Haier.

[0048] Human gastric cancer cell SGC-7901 was provided by the School of Pharmacy, Harbin University of Commerce (Center for Pharmaceutical Engineering Technology), which can be publicly available for non-commercial purposes only to repeat the examples described in the application.

[0049] Eighteen male healthy SD rats weighing 200±10g were purchased from Changchun Baike Biological Technology Co., Ltd. During the feeding process, the temperature and humidity were appropriate, the light was good, and the bedding was replaced regularly.

[0050] Example 1 Preparation and quality evaluation of luteolin phospholipid complex

[0051] In this example, luteolin phospholipid complex (LPC) was prepared to increase the water solubility of luteolin, facilitate its oral absorption, improve its bioavailability, and make the LPC-loaded gastric retention shape memory composite film play a better pharmacological effect. The phospholipid complex with the best luteolin complexing rate was obtained through orthogonal test design, and the quality of the LPC with the best complexing rate was evaluated. The prepared phospholipid complex was irradiated with a laser pen to observe whether there was a Tyndall phenomenon; the morphological structure of the phospholipid complex was observed using an optical microscope; the spectral properties of the prepared LPC were determined using an ultraviolet visible spectrophotometer; the composition change of the phospholipid complex was analyzed using infrared spectroscopy; and the solubility of the phospholipid complex was determined to observe whether the solubility of luteolin coated with soybean lecithin was improved.

[0052] 1.1 Determination of luteolin content

[0053] The LPC was prepared by solvent evaporation method, including the following steps: weighing soybean lecithin and luteolin, and dissolving the luteolin in tetrahydrofuran to obtain a luteolin solution with a concentration of 10 mg / mL; stirring the luteolin solution at 50°C for 6 hours; removing the organic solvent by rotary evaporation under reduced pressure; drying the sample in a vacuum drying oven at 40°C overnight; and storing the dried sample.

[0054] Preparation of a solution

[0055] Preparation of a control solution: weighing 0.004 g of luteolin, dissolving the luteolin in 10 mL of 40% methanol, and sonicating for 20 minutes to obtain a control solution.

[0056] Preparation of a test solution: weighing an appropriate amount of LPC, dissolving the LPC in 10 mL of 40% methanol, and sonicating for 20 minutes to obtain a test solution.

[0057] Preparation of a negative control sample solution: weighing 0.006 g of soybean lecithin, dissolving the soybean lecithin in a 10 mL volumetric flask, and sonicating for 20 minutes to obtain a negative control sample solution.

[0058] The content of luteolin in the LPC was determined by high performance liquid chromatography (Waters 1525). 4 mg of the LPC was dissolved in 10 mL of acetonitrile, filtered through a 0.22 μm microporous filter, and the filtrate was collected to determine the content of luteolin compounded thereon (X1 is the mass of luteolin in the LPC, and X0 is the total mass of the LPC, and the calculation formula is: compounding rate = X1 / X0 x 100%).

[0059] Chromatographic conditions

[0060] Mobile phase: water containing 0.1% H3PO4:methanol (volume ratio 30:70); flow rate: 1 mL / min; injection volume: 20 uL; detection wavelength: 348 nm; chromatographic column: Agilent ZORBAX SB-C18 (4.6 x 250 mm, 5 μm); column temperature: 25°C.

[0061] (1) Selection of determination wavelength

[0062] The maximum ultraviolet absorption wavelength of luteolin was scanned by a UV-visible spectrophotometer (UV-5200PC) at 200-400 nm.

[0063] In order to determine the maximum absorption wavelength of luteolin and facilitate the establishment of the luteolin methodology, the luteolin was scanned by a UV-visible spectrophotometer at 200-400 nm. As shown in Table 1, the maximum absorption wavelength of luteolin is 348 nm, and therefore the detection wavelength of luteolin is determined to be 348 nm. Figure 2 Table 1

[0064] (2) Specificity

[0065] The control sample solution, test sample solution and negative control sample solution were determined according to the above chromatographic conditions, and the chromatographic behaviors were investigated respectively. The liquid chromatograms of the luteolin control sample solution, negative control sample solution and test sample solution were compared (the liquid chromatogram is shown), and the results showed that luteolin had a peak at 2.6 min with good peak shape, and the determination of luteolin by LPC had almost no interference, so it can be considered that the liquid phase detection method has good specificity.

[0066] (3) Standard curve

[0067] The luteolin control sample solution was precisely taken into 5 mL vials, and the concentration of luteolin was 7.81, 15.63, 31.25, 62.50, 100.00, 125.00, 200.00 μg / mL respectively. The determination was performed using the above chromatographic conditions, the abscissa was the concentration of luteolin, the ordinate was the peak area, the linear regression analysis was established, and the standard curve of luteolin was constructed.

[0068] The standard curve of luteolin is shown in Figure 3 . It can be seen from Figure 3 that the linear relationship of luteolin is good in the range of 7.81-200.00 μg / mL, and the regression equation is: y = 1.4085x + 0.7947 (R 2 = 0.9997).

[0069] (4) Precision

[0070] The precision was mainly investigated by the instrument, and the control sample mother liquor was diluted into high, medium and low concentrations, i.e. 31.25 μg / mL, 15.63 μg / mL and 7.81 μg / mL. The peak area was detected by the above chromatographic conditions, and each concentration sample was detected three times within one day to investigate the intra-day precision of the method. The same high, medium and low concentration samples were prepared, and the samples were continuously detected for three days, and each concentration sample was detected three times to monitor the inter-day precision of the instrument.

[0071] The intra-day precision analysis results (Table 1) showed that there was no significant difference in the detection of the content of the drug sample by the instrument within one day, the intra-day precision of the instrument was good, and the RSD% was between 0.16-0.26%. The inter-day precision results showed that the RSD% of the three concentrations within three days was less than 0.57%, and the inter-day precision of the instrument was good.

[0072] Table 1 Intra-day precision and inter-day precision of luteolin (n = 3)

[0073]

[0074] (5) repeatability

[0075] Take 3 parts of the same batch of LPC, use the above sample solution preparation method to prepare, use the above chromatographic conditions to determine, record the sample peak area of each sample, and monitor the repeatability.

[0076] The repeatability test results are shown in Table 2. The results show that the average content of luteolin in the same batch of LPC is 74.42%, and the RSD is 1.01%, which has good repeatability.

[0077] Table 2 repeatability of luteolin (n=3)

[0078]

[0079] (6) sample recovery rate

[0080] Precisely take 0.4 mL of LPC sample solution (luteolin concentration 48.50 μg / mL) and add it to a 2 mL EP tube, divide it into 3 groups, each group has 3 parts, a total of 9 parts. Each group is added with 0.5, 1, 1.5 mL of luteolin standard solution (15.64 μg / mL) respectively, and 1.1, 0.6, 0.1 mL of methanol is added respectively to ensure the total volume of 2 mL. Use the above chromatographic conditions, record the sample peak area of each sample, and monitor the sample recovery rate.

[0081] The sample recovery rate test results are shown in Table 3. The average sample recovery rate of luteolin is 100.42%, and the RSD is 2.03%, indicating that the recovery rate of the method is high.

[0082] Table 3 sample recovery rate of luteolin (n=9)

[0083]

[0084] (7) stability

[0085] Take appropriate amount of LPC (0.4 g), prepare the sample solution according to the above method, and place it at room temperature for 0, 12, 24 h, then determine according to the above chromatographic conditions, record the sample peak area of each sample, and monitor the stability of the sample solution within 24 h.

[0086] The stability test results are shown in Table 4. The results show that the RSD is 1.79%, indicating that the LPC solution is stable within 24 h.

[0087] Table 4 stability of luteolin

[0088]

[0089] 1.2 orthogonal test

[0090] (1) orthogonal test design

[0091] According to the results of pre-experiment, the stirring time, stirring temperature, luteolin concentration, and the ratio of luteolin to soybean lecithin (mass ratio) were taken as the factors for orthogonal design, and the factor levels are shown in Table 5.

[0092] Table 5 Orthogonal factor level table

[0093]

[0094] Orthogonal test results: Through pre-experiment, it was found that the complexation rate of phospholipid complex was mainly affected by stirring time (A), stirring temperature (B), luteolin concentration (C), and the ratio of luteolin to soybean lecithin (D), so the four factors were taken as the influencing factors for orthogonal test, and the complexation rate was taken as the evaluation index. The test results are shown in Tables 6 and 7. According to R, the order of influence of each factor on the comprehensive result was B > A > C > D. According to the analysis of variance table, the B factor had statistical significance on the comprehensive result, and the differences among the levels of A, C, and D were not statistically significant. In the A factor, K3 > K2 > K1, in the B factor, K3 > K2 > K1, in the C factor, K1 > K2 > K3, and in the D factor, K2 > K1 > K3, therefore the optimal prescription of LPC preparation process was determined as A3-B3-C1-D2, that is, luteolin and soybean lecithin were weighed according to the ratio of 1:1.2 (mass ratio), an appropriate amount of tetrahydrofuran was added to make the mass concentration of the former (i.e. luteolin) 10 mg / mL, and then stirred at 50°C for 6h.

[0095] Table 6 Test design and results

[0096]

[0097] Table 7 Analysis of variance

[0098]

[0099]

[0100] (2) Verification test

[0101] According to the optimal prescription selected by the above orthogonal experiment, three batches of LPC were prepared, each batch was determined in triplicate, and the complexation rate was fitted to evaluate the stability of the process.

[0102] The average complexation rate of the three batches of LPC prepared according to the optimal prescription selected by the orthogonal test was 91.45%, and the RSD was 1.14%, indicating that the process was stable (Table 8). The LPC used in subsequent experiments was prepared according to the optimal prescription.

[0103] Table 8 Complexation rate of three batches of LPC prepared by the optimal prescription

[0104]

[0105] 1. Quality evaluation of LPC

[0106] (1) Tyndall effect

[0107] An appropriate amount of luteolin and soy lecithin (the mass ratio of luteolin to soy lecithin is 1:1.2) was weighed, an appropriate amount of tetrahydrofuran was added to make the concentration of luteolin 10 mg / mL, and constant temperature stirring was performed under certain temperature conditions (constant temperature stirring at 50°C for 6 h). Whether the solution produced Tyndall effect was observed by laser pen irradiation, and whether the preparation of LPC was successful was preliminarily judged.

[0108] During the preparation of LPC, the LPC solution was observed by laser pen irradiation, and it was found that the solution could produce Tyndall effect Figure 4 ). It was preliminarily determined that the LPC solution had colloidal properties, rather than a mixture of liquid and particles. The LPC prepared by solvent evaporation method was dried and sieved to obtain LPC finished product, as shown in Figure 5 . It can be observed that the phospholipid complex prepared has uniform color and full particles, and the color is darker than that of the luteolin standard, which is golden yellow, so it can be preliminarily judged that the preparation of phospholipid complex is successful.

[0109] (2) Optical microscope observation

[0110] After the LPC prepared according to the above optimal prescription was dried, the prepared phospholipid complex was diluted with a small amount of distilled water, dropped on a glass slide, covered with a glass slide, and the excess water was absorbed with a water-absorbing paper. The sample was observed and photographed under an optical microscope. The objective lens was selected as 100x1.25. Microscopy is often used to observe the morphology of particles. It was found through morphological observation that the LPC particles were complete, and were in uniform spherical or near-spherical structure. There was a uniform halo around the dispersed LPC particles, which was the soy lecithin layer, indicating that the soy lecithin had good wrapping property for luteolin Figure 6 ).

[0111] (3) Ultraviolet spectrum scanning

[0112] Luteolin, phospholipid and LPC samples were dissolved in methanol to prepare sample solutions with known concentration (20 μg / mL), and scanning was performed in the wavelength range of 200-400 nm by using an ultraviolet spectrophotometer. It was analyzed that the phospholipid only had a terminal absorption near 210 nm. The absorption curves of luteolin and LPC were basically the same, and there were two obvious characteristic absorption bands at about 254 nm and 348 nm Figure 7 ). The spectrum of luteolin and phospholipid complex was similar, indicating that no new chromophore was formed during the complexing of luteolin and soy lecithin, i.e. no new substance was formed.

[0113] (4) Fourier infrared spectrum scanning

[0114] The main functional groups in the components were detected by Fourier infrared spectrometer (AVATAR 360). Fourier transform infrared spectra of luteolin, phospholipid and prepared LPC were obtained by FTIR. Potassium bromide was ground into fine powder with an appropriate amount of sample, and was compressed into a disc under a hydraulic machine of 10000 psi for 30 s. Each treated sample was scanned in the wave number region of 4000-500 cm -1 -1. The results are shown in Figure 8 Compared with luteolin and phospholipid, the infrared spectrum of LPC did not change significantly. As can be seen from Figure 8 , luteolin has a characteristic absorption peak of -OH group near 3425 cm -1 , and a characteristic absorption peak of C=C near 1692 cm -1 . Soybean lecithin has a -CH2- group representing saturated hydrocarbon at 2994 cm -1 . LPC has characteristic absorption peaks at these positions, indicating that LPC combines luteolin and soybean lecithin, and retains the properties of luteolin.

[0115] (5) Solubility determination

[0116] An excess of luteolin, a physical mixture (a mixture of luteolin and soybean lecithin), and a phospholipid complex were respectively placed in a triangular flask, double distilled water was added, and magnetic stirring was carried out at room temperature for 24 h, centrifugation was carried out at 5000 r / min for 20 min, filtration was carried out, and sample injection was determined under the above chromatographic conditions to calculate the apparent solubility.

[0117] In order to improve the delivery efficiency of oral drugs with poor water solubility in the stomach, it is necessary to improve the water solubility of the drugs in the gastrointestinal tract. The water solubility of the drugs can be improved by preparing the drugs into phospholipid complexes. The calculation results of the apparent solubility of luteolin, a physical mixture, and LPC are shown in Table 9. The solubility of LPC in water is 2.29 times higher than that of luteolin, and the solubility of LPC in n-octanol is higher than that in water.

[0118] Table 9 Determination results of apparent solubility of samples n = 3

[0119]

[0120] In this embodiment, the optimal complexing rate process of LPC was obtained by orthogonal test, and the good performance of LPC was verified by infrared, ultraviolet and other characterization experiments, which laid a foundation for the combination and application of LPC and gastric retention shape memory composite film.

[0121] Preparation and characterization of drug-loaded gastric retention shape memory composite film

[0122] The gastric retention drug delivery system provided by the present application can improve the bioavailability of the drug, locally exert the drug efficacy in the stomach, reduce the side effects of the drug, prolong the drug delivery interval, and improve the compliance of the patient. In the present embodiment, the shape memory polymer is applied to the gastric retention drug delivery system to prepare a thermally responsive gastric retention shape memory composite film. The deformability and floatability of the composite film are utilized to prolong the retention time of the drug-loaded composite film in the stomach and increase the bioavailability of the drug. First, the composite film is prepared. Polylactic acid (PLA) is used as the main material, and polycaprolactone (PCL) and polyethylene glycol (PEG) are added to modify the main material of the composite film, so as to reduce the glass transition temperature (Tg) of the composite film and make the deformation temperature of the composite film closer to the body temperature. In addition, Hypromellose (HPMC) and sodium bicarbonate (NaHCO3) are added to the composite film to increase the gastric retention time of the composite film. Subsequently, the composite film is characterized by Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), etc. The FTIR analysis is used to analyze the changes in the material composition of the prepared composite film and the stability of the composition. The DSC analysis is used to analyze the glass transition temperature and melting temperature of the prepared composite film, observe the influence of PCL and PEG on the glass transition temperature of PLA, and study whether the glass transition temperature of PLA is reduced and whether it is more suitable for responding to the deformation in the stomach temperature of the human body. The thermal stability of the prepared composite film is analyzed by thermogravimetric analysis (TGA), and the thermal decomposition temperature of the prepared composite film is observed. The optical microscope is used to observe the influence of the addition of PCL and PEG in the composite film on the apparent morphology of the PLA-based gastric retention shape memory composite film and the compatibility between the materials in the composite film. The shape memory recovery rate test is used to observe the deformation recovery of the prepared composite film at the body temperature and the glass transition temperature, respectively. The floating experiment is used to observe the floating of the prepared composite film in the simulated gastric juice, and to study whether the floating time is suitable for the gastric retention drug delivery system. The drug release from the composite film is studied using a constant temperature oscillator, and the influence of the HPMC addition ratio, the NaHCO3 addition ratio, the concentration of luteolin, and the polymer ratio of the composite film on the release of luteolin from the composite film is investigated to determine the optimal release aid ratio of the composite film. According to the comprehensive analysis of the above experimental results, the optimal LPLC-loaded gastric retention shape memory composite film prescription is screened, and the preparation for the in vivo pharmacokinetic evaluation of rats is made.

[0123] 2.1 Preparation of drug-loaded composite film and non-drug-loaded composite film

[0124] The preparation of the non-drug-loaded composite film includes the following steps:

[0125] (1) The polymer includes one or a mixture of several of PLA, PCL, PEG, and 10 formulations are prepared according to Table 10.

[0126] Table 10 Polymer ratio in thermally responsive gastric retention shape memory composite film

[0127]

[0128] (2) The polymer is mixed with dichloromethane (CH2Cl2) at a mass ratio of polymer to dichloromethane of 1:7 to obtain solution 1.

[0129] For formulation 1, the polymer is PLA at a mass ratio of PLA to dichloromethane of 1:7; for formulations 2 to 4, the polymer includes PLA and PCL at a mass ratio of (PLA+PCL) to dichloromethane of 1:7; for formulations 5 to 7, the polymer includes PLA and PEG at a mass ratio of (PLA+PEG) to dichloromethane of 1:7; and for formulations 8 to 10, the polymer includes PLA, PCL and PEG at a mass ratio of (PLA+PCL+PEG) to dichloromethane of 1:7.

[0130] (3) A fixed amount of hydroxypropyl methyl cellulose (HPMC) and NaHCO3 is dissolved in distilled water at a mass ratio of (HPMC+NaHCO3) to water of 1:10, stirred thoroughly, and left to stand to obtain solution 2.

[0131] (4) Solution 1 and solution 2 are mixed, and a mechanical stirrer is used to stir for 4 h. Subsequently, CH2Cl2 and water in the blend are evaporated at room temperature, and a sheet with a thickness of 1 mm is prepared by mold pressing at a pressure of 10 MPa to obtain a non-drug-loaded composite film.

[0132] The method for preparing a drug-loaded composite film is the same as the above method, except that in step (4), a fixed amount of LPC is added when the composite film solution is mechanically stirred for 3.5 h, and mechanical stirring is continued for 0.5 h. Subsequently, CH2Cl2 and water in the blend are evaporated, and a sheet with a thickness of 1 mm is prepared by mold pressing at a pressure of 10 MPa to obtain a drug-loaded composite film. LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3), LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3), LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2), and LPC-PLA / PCL / PEG (8 / 2 / 3) drug-loaded gastric retention shape memory composite films are prepared by the above method.

[0133] When HPMC is 5% (by mass) of the polymer, NaHCO3 is 5% (by mass) of the polymer, and LPC is 30% (by mass) of the mixture (total mass of polymer, HPMC, and NaHCO3), the appearance of the drug-loaded composite membranes obtained is shown in the figure. Figure 9 The dried composite membranes were uniformly golden in color. Small particle protrusions were present on the surface of the composite membranes, indicating LPC. This contrasts with LPC-PLA without HPMC and NaHCO3 (i.e., without added HPMC and NaHCO3). Figure 9 As shown in Figure J, it can be observed that the addition of HPMC and NaHCO3 can promote a more uniform surface of the composite membrane.

[0134] 2.2 Characterization Experiment of Gastric Retention Shape Memory Composite Membrane

[0135] (1) Fourier transform infrared spectroscopy analysis

[0136] The chemical structure of polymer materials can be studied using FTIR. The characteristic absorption peaks revealed in the results can be used to analyze the specific chemical bonds and functional groups contained in the polymer materials. Simultaneously, it can detect whether the chemical bonds and chemical structure of the material change before and after mixing. FTIR is used to evaluate the molecular properties of different samples. This invention utilizes an FTIR instrument to determine the characteristic functional groups of materials. The sample is ground into a fine powder, potassium bromide is added, and it is compressed to prepare a 5×5 mm transparent thin film. The wavelengths at 4500-300 cm⁻¹ are used. -1 The spectral range was scanned. Figure 10 The specific FTIR analysis results for LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 3) are presented.

[0137] like Figure 10 As shown in (A), the stretching vibration peak of PLA at C=O is at 1749 cm⁻¹. -1 At this point, the stretching vibration peak of COC appears at 1081 cm⁻¹. -1 1132cm -1 1184cm -1 Location; 1454cm -1 and 1381cm -1 The peaks for the stretching and bending vibrations of -CH are located at 1749 cm⁻¹. The peak for the C=O stretching vibrations of PCL appears at 1749 cm⁻¹.-1 At this point, the stretching vibration peak of COC appears at 1083 cm⁻¹. -1 1128cm -1 1181cm -1 The stretching and bending vibration peaks of -CH are at 1448 cm⁻¹. -1 and 1374cm -1 The C=O stretching vibration peak of PLA / PCL material appears at 1749 cm⁻¹. -1 At this point, the stretching vibration peak of COC appears at 1081 cm⁻¹. -1 1132cm -1 and 1184cm -1 The stretching and bending vibration peaks of -CH are at 1455 cm⁻¹. -1 and 1386cm -1 The characteristic peaks of PLA and PCL molecules did not change significantly in position, nor were any new characteristic peaks generated, indicating that no denaturation occurred between them. The C=O stretching vibration peak of PEG appeared at 1742 cm⁻¹. -1 At this point, the stretching vibration peak of COC appears at 1073 cm⁻¹. -1 1113cm -1 1204cm -1 Location; 1465cm -1 and 2876cm -1 The peaks at 1378 cm⁻¹ are the bending vibration peak and the symmetrical stretching vibration peak of -CH₂-. -1 The peak at 844 cm⁻¹ is a symmetrical deformation vibration peak of -CH₃. -1 The peak at this location is an in-plane deformation peak of -CH2CH2O-. Through analysis... Figure 10 (B) It was found that the characteristic peaks of PLA / PEG materials at these locations are basically consistent. Through analysis... Figure 10 (C) It was found that the stretching vibration peak of C=O in PLA / PCL / PEG material appears at 1750 cm⁻¹. -1 At this point, the stretching vibration peak of COC appears at 1081 cm⁻¹. -1 1130cm -1 1182cm -1 Based on the above analysis, it can be seen that in the PLA / PCL / PEG composite film, the characteristic peaks of PLA, PCL, and PEG molecules are consistent with the characteristic peaks of the individual materials.

[0138] In summary, it can be determined that LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 3) are all physical blends. Observation... Figure 10 It can also be found that these drug-loaded materials were present in 1692. -1 2994 -1 and 3425 -1 The presence of characteristic peaks nearby corresponds to the positions of LPC characteristic peaks obtained from the FTIR analysis above, indicating that LPC in the blend material has not been modified.

[0139] (2) Differential scanning calorimetry

[0140] DSC analysis is a common method for evaluating the thermal properties of materials. The powder on the prepared sample plate is filed into sample powder using a file. The DSC curve is obtained under a nitrogen flow, using a two-stage heating method. The temperature range is selected between 25-200℃, with slow heating and cooling at a rate of 10℃ / min.

[0141] The glass transition temperature (Tg) and melting point (Tm) of ten thermally responsive drug-loaded composite films were determined using a differential scanning calorimeter (DSC-500L) to analyze the changes in thermal performance parameters of the composite films during temperature changes. Figure 11 The table shows the secondary heating curves for PLA, LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 3). Table 11 provides detailed Tg and Tm values ​​for the ten composite membranes. The results show that the Tg of PLA is around 56℃, and the Tm is around 171℃. The Tg of LPC-PLA is 56.77℃. The addition of LPC, HPMC, and NaHCO3 does not affect the Tg. The Tm of LPC-PLA is 162.06℃, which is lower than that of PLA. This may be because the addition of excipients and drugs reduces the crystalline regions in the polymer matrix material, thereby lowering the Tm value. Analysis Figure 11(A) It can be seen that the Tg of the material decreases by about 10℃ after adding PCL in the composite film, indicating that there is a large modification effect, and the melting temperature of the material is maintained at about 160℃, without significant change. Analysis Figure 11 (B) It can be seen that the Tg of LPC-PLA / PEG(7 / 3) is about 10℃ lower than that of LPC-PLA, indicating that the addition of PEG has a certain modification effect on the composite material. We failed to measure the Tg of LPC-PLA / PEG(9 / 1) and LPC-PLA / PEG(8 / 2), which may be related to the overlapping of multiple peaks. We guess that the reason why the Tg value decreases due to the blending of PEG and PLA may be that PEG can enter between the polylactic acid macromolecules during melt mixing, and can establish hydrogen bonds or dipole-dipole physical interactions between atoms. Therefore, some rigid and uniform PLA-PLA interactions will be replaced by non-uniform PLA-PEG interactions. This phenomenon may provide better macromolecular movement and reduce Tg by reducing energy during the glass transition process. The addition of PEG has an effect on the melting temperature of the composite material, and within the range of our addition ratio, the Tm of the composite material decreases with the increase of the PEG ratio. The reason may be that the addition of PEG causes the PLA and PCL to appear crystal disorder and defect, and the melting point of the composite material decreases. The test results of LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 3) groups are shown in Figure 11 (C) As shown in, with the increase of the PEG ratio in LPC-PLA / PCL, the Tg and Tm values decrease in turn, and in the LPC-PLA / PCL / PEG(8 / 2 / 3) group, the Tg value and Tm value are the lowest among the ten test groups.

[0142] Table 11 Thermal performance parameters of composite film

[0143]

[0144] (3) Thermogravimetric analysis

[0145] TGA analysis is a common detection method for the thermal performance of materials. A thermogravimetric analyzer is used for testing. During testing, the material is shaved from the sample plate with a file to obtain powder as the test sample. The sample is placed in an aluminum crucible and heated from 25℃ to 800℃ at a heating rate of 10℃ / min under nitrogen atmosphere.

[0146] The mass loss curves of the 10 drug-loaded gastric retention shape memory composite films prepared by the present application under heating are shown in Figure 12As shown, the TGA curve of the gastric retention shape memory composite membrane exhibits three stages of mass loss. The first stage is attributed to the loss of water molecules (below 250℃); the second stage involves the thermal degradation of the polymer in the gastric retention shape memory composite membrane between 250-400℃; and the third stage involves the carbonization of the polymeric material (400-550℃). Figure 12 The curves show that all materials prepared in the experiment underwent thermal decomposition under heating. The temperature at which the materials were completely decomposed was around 450℃, and the residual carbon rate was 0. The degradation initiation temperature of PLA material was around 350℃, and the degradation peak temperature was around 360℃, indicating that PLA has no water content. Comparison of LPC-PLA and PLA curves reveals that ( Figure 12 (A) The addition of LPC, HPMC, and NaHCO3 significantly reduced the degradation initiation temperature of PLA, with the initial decomposition temperature of the material appearing at around 200℃. Furthermore, the addition of LPC, HPMC, and NaHCO3 significantly reduced the peak degradation temperature of PLA, with the peak degradation temperature appearing at around 250℃. This indicates that HPMC increased the water retention capacity of the composite material, enabling it to degrade significantly before 250℃. Compared to LPC-PLA, the research results for different ratios of the same material (LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), and LPC-PLA / PCL (7 / 3) showed that with increasing PCL content, the degradation initiation temperature and peak degradation temperature increased slightly. Compared to LPC-PLA, the heat resistance of the material was slightly increased. The addition of PCL to PLA increased the initial decomposition temperature of the composite material, indicating improved heat resistance. Furthermore, the initial decomposition temperature showed an upward trend with increasing PCL content. This can be attributed to the improved heat resistance of the material due to the addition of PCL. Figure 12 (B) Figure 12 (C) The results show that the initial decomposition temperature of the material decreased after the addition of PEG, indicating that the material is more easily decomposed than before modification. Furthermore, the initial decomposition temperature increased with increasing PEG content, while the complete decomposition temperature of the composite material was earlier than that of the PLA / PCL blend group. Therefore, it can be inferred that the heat resistance of the material decreased slightly after the addition of PEG. In summary, the ten groups of gastric retention shape memory composite membranes prepared exhibited good thermal stability and maintained stable properties below 200℃.

[0147] (4) Observation under an optical microscope

[0148] The microstructure of the composite material samples was observed using an optical microscope. Optical images of the composite material were taken at room temperature, focusing on areas with good light transmittance. A 20×0.5 objective lens was used. The experimental results are as follows: Figure 13As shown, all the shape memory gastric residence smart materials have typical optical microstructure. Figure 13 (A) shows that the microstructure of PLA is relatively simple, only PLA particles exist, and the PLA particles are closely arranged, which is a characterization of good compatibility between monomers. As shown in Figure 13 (B)-(K) show that the surface of the drug-loaded gastric residence shape memory composite film has a light yellow color, and the color is uniformly distributed, indicating that the LPC is well distributed in the gastric residence shape memory composite film. Compared with PLA, LPC-PLA has a slight phase separation, which is due to the addition of auxiliary materials HPMC and NaHCO3 Figure 13 (B)). By observing Figure 13 (C)-(E) found that the higher the proportion of PCL, the more uneven the surface of the composite film, and bubbles were generated, which indicated that the phase separation of PLA and PCL was more obvious. The reason is that PCL and PLA form a heterogeneous structure, and the interface adhesion is poor. By observing Figure 13 (F)-(H) found that the material compatibility of PEG and PLA is better than that of PCL and PLA, the higher the proportion of PEG, the more uniform the surface of the composite film, and the addition of PEG is beneficial to the formation of pores in the PLA material, thereby changing the pore size distribution of the material. Figure 13 (I)-(K) show the results further verify the above point, the addition of PEG can effectively improve the uniformity of the surface of the PLA-PCL composite film, and PEG has better compatibility with PLA than PCL.

[0149] (5) Floating test

[0150] The in vitro floating capacity of the gastric residence shape memory composite film was observed by floating time. 10 groups of drug-loaded composite films were placed in 50 mL beakers containing artificial gastric juice, and a glass rod was used to assist them to the bottom of the beaker, and the floating time was recorded. In addition, the floating state of the drug-loaded composite film was recorded at 0, 0.5, 1, 2, 4, 6, 8, 12, 24 and 48 h under the condition of 37.0±0.5℃.

[0151] The in vitro simulation floating test results of 10 groups of gastric residence shape memory composite films are as follows: Figure 14 and Figure 15The floating time results are shown in Table 12. The experimental results show that the floating time of the 10 groups of composite films is within 1 s. The reason is that the blending of the materials changes the dense structure of PLA into a general loose porous structure. In the process of solvent evaporation into a composite film, the porous structure forms a closed bubble small space, which promotes the drug-loaded composite film to float in the simulated gastric juice, so it can quickly float above the gastric juice when initially placed. With the extension of time, it can be observed that the gastric retention shape memory composite film releases LPC to make the solution turn yellow, and bubbles are generated on the cup wall. This is because NaHCO3 in the gastric retention shape memory composite film reacts with hydrochloric acid to generate CO2, which produces small bubbles to promote the release of LPC. Between 0-8h, the ten groups of drug-loaded composite films remain floating above the simulated gastric juice. Between 8h-12h, LPC-PLA / PCL / PEG(8 / 2 / 3) changes from floating to sinking, and the remaining drug-loaded composite films continue to show floating state. Between 12h-24h, LPC-PLA, LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(7 / 3), and LPC-PLA / PCL / PEG(8 / 2 / 2) sink. It is speculated that the reaction of NaHCO3 in the composite film is complete, no bubbles continue to be generated, and after the composite film is soaked in liquid, most of the structure pores of the composite film are immersed in water molecules, resulting in the sinking of the composite film. Between 24h-48h, LPC-PLA / PEG(9 / 1) precipitates, while LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PEG(8 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 1) do not precipitate within 48h. In summary, the 10 kinds of drug-loaded composite films prepared by the present application all maintain floating state within 8h, and part of the drug-loaded composite films have floating time exceeding 48h, indicating that the prepared drug-loaded gastric retention shape memory composite film has good floating performance.

[0152] Table 12 Floating state of composite film at different times

[0153]

[0154] (6) Shape memory performance test

[0155] The gastric-retention shape memory composite film prepared in the present application comprises a heat-sensitive shape memory polymer, and the shape memory performance of the material is characterized by a recovery rate parameter. The shape memory performance of the gastric-retention shape memory composite film is tested by a bending experiment. The sample is processed to a size of 50 mm x 4 mm x 1 mm, and a glass rod with a diameter of 5 mm is used as a mandrel. The sample and the glass rod are placed in a constant-temperature water bath at 50°C for 5 min, and then the sample is bent by the glass rod at 180°C for 10 s. Then, the sample is placed in 5°C cold water for 10 s. Finally, the sample is again placed in 50°C hot water, and a camera is used to record the shape recovery of the sample at 0 s, 3 s, 9 s and 60 s. The shape recovery time refers to the time required for the shaped polymer to recover to its original shape after being heated again. The shape recovery rate is usually indicated by a recovery angle, and the shape recovery angle is the angle change of the material at different times in the shape recovery process, i.e., the angle between the two ends of the sample after the polymer is bent and shaped. See Figure 16 , t refers to the thickness of the material, r is the bending radius, and θ is the shape recovery angle. S0 is parallel to the fixed end of the bent sample and passes through the center line end point of the circle. x is the distance of the sample from the center of the circle during the recovery process. The shape recovery rate is calculated by formula (1-1).

[0156]

[0157] The diameter of the human pylorus is 15 mm. In order to prevent the shape-recovered composite film from flowing out of the pylorus, the film preparation has certain requirements for the folding angle of the film and the length of the film. Therefore, the film preparation designed and prepared by us has a length of 50 mm. The shape recovery process of ten gastric-retention shape memory composite films under stress conditions at the Tg temperature (50°C) and the human body temperature (37°C) is recorded and calculated by a camera, and the results are shown in Figure 17 and Figure 18 . At the glass transition temperature of 50°C, the addition of the excipients HPMC and NaHCO3 has a certain effect on the shape recovery rate of the composite film. Compared with PLA alone, the addition of the excipients prolongs the shape recovery time of the composite film and presents a lower recovery rate Figure 17 A and Figure 18 B); even so, when the shape recovery time is extended to 60 s, the deformation recovery rate of the LPC-PLA group can reach 70%. From Figure 17 B and Figure 18KIt is known that the addition of PEG and PCL in PLA has little effect on the shape memory recovery performance of the composite film, and the shape recovery rate of each group of composite film within 60s is in the range of 70-90%, and the shape recovery performance is ideal. The possible reason is that PLA accounts for the largest proportion, and PLA as a fixed phase provides the corresponding driving force for shape recovery. The unmelted PLA macromolecules and microcrystals form some physical entanglement points, providing a deformation driving force. Considering that the shape memory gastric retention intelligent material will be applied to patients with gastric cancer, therefore, on the basis of investigating the deformation process of the composite film at the glass transition temperature, the deformation capacity of the composite film under the stress condition of 37℃ is further studied, so as to provide a theoretical basis for the clinical application of the composite film. The experimental results are shown in Figure 18 The decrease of stress temperature indeed affects the shape recovery ability of the ten groups of composite film (about 10% decrease), and the deformation recovery rate is about 60%, so that the diagonal edge length is 25mm, and this recovery rate also meets the size requirement of realizing gastric retention.

[0158] 2.3 Release of luteolin in the LPC-loaded gastric retention shape memory composite film

[0159] (1) Preparation of luteolin standard curve

[0160] 10mg of luteolin standard was accurately weighed and dissolved in methanol solution, and then added to a 10mL volumetric flask with simulated gastric juice to constant volume. According to the equal dilution, different concentrations of standard solution were prepared, and the absorbance was measured at 348nm by ultraviolet spectrophotometer (UV-7). The standard curve of luteolin was drawn with concentration as abscissa and absorbance as ordinate.

[0161] The luteolin standard curve is shown in Figure 19 The equation of luteolin standard curve is y=0.0067x-0.0389, R 2 =0.9969, and the linear relationship is good.

[0162] (2) Effect of polymer ratio on release of luteolin in the composite film

[0163] The release of luteolin in the gastric retention shape memory composite film was studied by using a constant temperature oscillator, and the effect of polymer ratio on the release of luteolin in the gastric retention shape memory composite film was investigated. The polymer ratio is shown in Example 2.1. On the basis of Example 2.1, the proportion of HPMC in the composite film is 1wt% (i.e. 1% of the polymer is HPMC), the proportion of NaHCO3 is 1wt% (i.e. 1% of the polymer is NaHCO3), and the proportion of LPC is 10wt% (i.e. 10% of the total mass of polymer, HPMC and NaHCO3 is LPC).

[0164] The 2.73 g of gastric retention shape memory composite film was placed in a glass beaker containing 500 mL of simulated gastric fluid, and the beaker was placed in a constant temperature shaker, which was continuously shaken at a speed of 100 rpm, and the temperature environment was set at 37℃±0.5℃. Equal amounts of test solution were taken at 1 h, 2 h, 3 h, 4 h, 6 h, 8 h, 1 d, 2 d, 3 d, 4 d, 5 d, 6 d and 7 d, and an equal volume of release medium (simulated gastric fluid) was replaced to ensure that the total volume remained unchanged throughout the release experiment. The release of luteolin was analyzed at 348 nm using a UV-visible spectrophotometer, and the concentration of luteolin in the sample was calculated by a standard curve. The in vitro release study was carried out in triplicate, and the percentage of drug release was plotted according to the average value.

[0165] This example investigates the effect of polymer ratio on the release behavior of luteolin in the composite film. Under the conditions of HPMC addition ratio of 1%, NaHCO3 addition ratio of 1%, and LPC addition ratio of 10% constant, the effect of polymer ratio on the drug release of the composite film is shown in Figure 20 The gastric retention shape memory composite film with different polymer ratios has the maximum drug release rate within the first 0-7 h, and has the characteristics of sustained drug release, and drug release can still be detected at 24 h. With the increase of PEG ratio, the cumulative release amount of the drug increases, among which the cumulative release amount of luteolin in LPC-PLA / PEG (7 / 3) is the maximum at all detection time points, and the cumulative release amount of luteolin at 24 h reaches 31.42%; the cumulative release amount of luteolin in LPC-PLA / PCL / PEG (8 / 2 / 3) is the second, and the cumulative release rate of luteolin at 24 h is 28.95%. It can be seen that PEG has a certain promoting effect on the release of luteolin in the composite film. The addition of PCL also has a certain promoting effect on the drug release of the composite film, but the addition ratio needs to reach a certain value. Figure 20The results show that the drug cumulative release amount of the LPC-PLA / PCL (7 / 3) composite film and the LPC-PLA / PCL (8 / 2) composite film is greater than that of the LPC-PLA, and the drug cumulative release amount of the LPC-PLA / PCL (8 / 2) composite film is less than that of the LPC-PLA composite film. The inventors speculate that the reason why the addition of PEG and PCL promotes drug release can be that it destroys the polymer network formed by PLA itself, and as the PCL and PEG addition ratio increases, the free space between the structure formed by PLA itself increases, thereby promoting drug release; in contrast, the effect of PEG on promoting drug release is stronger than that of PCL, which can be due to the different properties of PEG and PCL in the material. The addition of PEG can lead to the formation of large pores in PLA, which is beneficial to drug release. In addition, PCL and PLA form a heterogeneous structure in thermodynamics, and the interface adhesion is poor, and the composite film can have a phenomenon of PCL agglomeration with other materials, and the agglomeration has few pores, resulting in that the drug release of the composite film with added PCL is inferior to that of the composite film with added PEG.

[0166] (3) Effect of HPMC addition ratio on release of luteolin in composite film

[0167] The effect of HPMC addition ratio 1%, 2%, 3% (mass ratio of HPMC to polymer is 1%, 2%, 3%) on the release of luteolin in the composite film was investigated. On the basis of Example 2.1, the composite film LPC-PLA / PEG (7 / 3) was taken as the research object, and the addition ratio of NaHCO3 was 1wt% (i.e. NaHCO3 was 1% of the polymer), and the addition ratio of LPC was 10wt% (i.e. LPC was 10% of the total mass of polymer, HPMC and NaHCO3) remained unchanged.

[0168] The release amount of luteolin was determined according to the above method. With the composite film LPC-PLA / PEG (7 / 3) as the research object, under the constant conditions of NaHCO3 addition ratio 1% and LPC addition ratio 10%, the effect of the addition amount of HPMC (1%, 3% and 5%) on the release of luteolin was investigated. The experimental results are as follows Figure 21As shown, at the same drug release detection time point, the cumulative release amount of the drug increases with the increase of the HPMC addition ratio. At 24 h, the cumulative release rate of luteolin in the LPC-PLA / PEG (7 / 3) containing 5% HPMC has reached 46.85%. It is known from the above examples that the high content of HPMC will affect the deformation rate of the composite film, and in the present application, 5% of the HPMC concentration is selected as the addition ratio of the luteolin drug excipient. In addition, the LPC-PLA / PEG (7 / 3) containing different proportions of HPMC shows a biphasic release phenomenon in vitro. The gastric retention shape memory composite film shows a rapid drug release phenomenon in the first 7 h, which may be due to the fact that most of the luteolin is loaded on the surface of the composite film, which is not protected by the polymer network structure, resulting in rapid release. Another reason for the rapid release of luteolin in the first 7 h may be that the NaHCO3 on the composite film is completely reacted with the gastric hydrochloric acid in 7 h, forming pores in the composite film, thereby promoting the release of luteolin inside the composite film. The slow release phenomenon after 7 h may be due to the formation of a rigid polymer network structure between LPC, polymer and excipient, which will affect the internal void of the composite film to become smaller and reduce the entry of water molecules into the composite film.

[0169] (4) Effect of NaHCO3 addition ratio on luteolin release in the composite film

[0170] The effect of NaHCO3 addition ratio 1%, 3%, 5% (i.e. the mass percentage of NaHCO3 and polymer is 1%, 3%, 5%) on luteolin release in the gastric retention shape memory composite film was investigated. Based on Example 2.1, the composite film LPC-PLA / PEG (7 / 3) was used as the research object, and under the constant conditions of HPMC addition ratio 5wt% (HPMC is 5% of the polymer) and LPC addition ratio 10wt% (i.e. LPC is 10% of the total mass of polymer, HPMC and NaHCO3), the release amount of luteolin was determined according to the above method.

[0171] The effect of NaHCO3 addition amount (1%, 3% and 5%) on luteolin release in the composite film was investigated with the composite film LPC-PLA / PEG (7 / 3) as the research object under the constant conditions of HPMC addition ratio 5% and LPC addition ratio 10%. The experimental results are shown in Figure 22 As shown, the drug release rate of LPC-PLA / PEG (7 / 3) with different concentrations of NaHCO3 is the largest in the first 7 h, and it is found that the composite film has the characteristics of sustained drug release, and the drug release time can reach 24 h.

[0172] With the increase of NaHCO3 concentration, the cumulative release rate of the drug increased. When the concentration of NaHCO3 was 5%, the cumulative release rate of luteolin in the LPC-PLA / PCL / PEG (8 / 2 / 3) was the highest, and the cumulative release rate of luteolin in the composite film with 5% NaHCO3 was 53.76% at 24 h. According to the experimental results, the inventors inferred that the higher the concentration of NaHCO3, the more unstable the polymer network structure. When preparing the composite film, according to the factors such as the influence of NaHCO3 on the deformation rate of the composite film proved in the above examples, we selected to add 5% NaHCO3.

[0173] (5) Effect of luteolin concentration on the release of luteolin in the composite film

[0174] We further detected the drug release of LPC-PLA / PEG (7 / 3) with 10%, 20% and 30% luteolin (i.e. 10%, 20% and 30% of the total mass of the polymer, HPMC and NaHCO3). Based on Example 2.1, the concentration of HPMC was fixed at 5 wt% (5% of the polymer was HPMC), and the concentration of NaHCO3 was fixed at 5 wt% (5% of the polymer was NaHCO3). The amount of luteolin released was determined according to the above method.

[0175] We further detected the drug release of LPC-PLA / PEG (7 / 3) with 10%, 20% and 30% luteolin (i.e. 10%, 20% and 30% of the total mass of the polymer, HPMC and NaHCO3). Based on Example 2.1, the concentration of HPMC was fixed at 5 wt% (5% of the polymer was HPMC), and the concentration of NaHCO3 was fixed at 5 wt% (5% of the polymer was NaHCO3). The amount of luteolin released was determined according to the above method. Figure 23 It can be seen that with the increase of the concentration of luteolin, the cumulative release rate of the drug increased. When the concentration of luteolin was 30%, the cumulative release rate of the drug in LPC-PLA / PEG (7 / 3) was the highest, and the cumulative release rate of the drug was 75.56% at 24 h. The inventors inferred that the increase of the mass of luteolin in the drug-loaded composite film not only increased the concentration of LPC on the surface of the composite film, but also increased the free space between the structures formed by the polymer itself due to the high concentration of LPC embedded in the rigid structure of the polymer, which promoted the release of the drug. The cumulative release rate of the drug may be positively correlated with the drug loading amount, and the greater the drug loading amount, the greater the cumulative release rate. According to the above examples, the addition of luteolin to the composite film affects the deformation rate of the composite film, etc., and we selected to add 30% luteolin.

[0176] The example prepares the gastric retention shape memory composite film of LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3), LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3), LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2), and LPC-PLA / PCL / PEG (8 / 2 / 3). Comprehensive analysis of the apparent morphology, thermodynamic properties, thermal properties, floatability, deformability and drug release of the ten groups of composite films shows that the ten groups of composite films all have good performance. Among them, LPC-PLA / PEG (7 / 3) has the best performance as a gastric retention drug delivery system, and is used for later pharmacokinetic experiments.

[0177] Example 3 Evaluation of the inhibitory effect of the gastric retention shape memory composite film on gastric cancer cells

[0178] Luteolin has excellent medicinal value and is mainly used in clinic for relieving cough, eliminating phlegm, reducing inflammation, reducing uric acid, etc. Studies have found that it can inhibit the proliferation of various malignant tumors of gastric cancer, and its anti-cancer potential is huge. However, the solubility of luteolin is poor, which will directly affect the therapeutic efficacy of luteolin in treating cancer. The drug loaded in the gastric retention shape memory composite film prepared by us is LPC. The present application finds that phospholipid complexes can improve the solubility of water-insoluble drugs, allowing tumor cells to absorb more drug molecules, thereby affecting the drug to exert better efficacy. This example investigates the toxic effect of non-drug-loaded composite film on SGC-7901 gastric cancer cells and the inhibitory effect of drug-loaded composite film on SGC-7901 gastric cancer cells, and directly observes the influence of composite film on the number and morphology of SGC-7901 gastric cancer cells for 72 h through an inverted microscope.

[0179] 3.1 Experimental method

[0180] (1) Composite film combination distribution

[0181] Because some composite film groups are only different in polymer proportion when the materials are mixed, and the polymer components are the same, the same four groups of material components were selected for the study of the effect of non-drug composite films on SGC-7901 gastric cancer cells, namely PLA, PLA / PCL(7 / 3), PLA / PEG(7 / 3), and PLA / PCL / PEG(8 / 2 / 3) composite films as non-drug cell experimental groups, to observe the cytotoxicity of the four non-drug composite films on SGC-7901 gastric cancer cells. LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 3) drug-loaded composite films were subjected to SGC-7901 gastric cancer cell proliferation inhibition test.

[0182] (2) Preparation of composite film extract

[0183] The composite film was placed in a high-pressure sterilization pot and sterilized for 20 min. After sterilization, it was dried in an oven to remove surface steam moisture. After thorough drying, it was placed in a clean bench and sterilized with ultraviolet light for 30 min and was ready for use.

[0184] The pretreated 0.5 g composite film was placed in a sterilized beaker, 250 mL of cell culture solution (DMEM high-sugar medium with 10% fetal bovine serum and 1% double antibody) was added to each beaker, and it was cultured in a 37°C, 5% CO2 incubator for 24 h. The material culture solution was considered as 100% concentration of material extract. Different groups of non-drug composite films were diluted to 50%, 25%, 12.5%, and 6.25% concentrations for cell culture, and SGC-7901 gastric cancer cell toxicity test was performed. The 100% concentration of material extract of the drug-loaded composite film group was used as the experimental group for the effect of the drug-loaded composite film on SGC-7901 gastric cancer cells.

[0185] (3) SGC-7901 gastric cancer cell recovery

[0186] Firstly, the frozen SGC-7901 gastric cancer cells are recovered. The water bath is preheated, and the temperature is set to 36℃. Open the clean bench, put in the main instruments, and turn on the ultraviolet sterilization for 30 min. Take the cell freezing tube out of the liquid nitrogen and quickly melt it in the water bath. Quickly take it to the clean bench, aspirate the cells and put them into a sterile centrifuge tube, centrifuge at 1000 r / min for 5 min. Aspirate the supernatant of the centrifuge tube, add 5 mL of culture solution to the culture bottle, rinse the centrifuged cells with 1 mL of culture solution, and transfer them to the culture bottle. Observe and mark under the microscope. Wipe the cell bottle with alcohol cotton, slightly loosen the cover, and put it in the incubator.

[0187] (4) SGC-7901 gastric cancer cell passage

[0188] Observe the recovered SGC-7901 gastric cancer cells under an inverted microscope, and the growth state is good. When the cells in the cell culture bottle grow to 70-80% and the cells grow adherently, the cell morphology is irregularly long spindle-shaped, and the cell passage culture can be carried out. The reagents to be used are preheated in a constant temperature water bath at 37℃. Discard the culture solution in the cell bottle, rinse with PBS for 2-3 times, and then discard the residual PBS with a pipette gun. Add 1 mL of digestion solution (0.25% trypsin solution) to the culture bottle to digest the cells, and observe under an inverted microscope. When the cells are rounded, discard the digestion solution. Add 3 mL of new culture solution, blow the cells, and evenly distribute the new culture solution containing cells to two cell culture bottles. The passage is complete. Change the new culture solution after 24 h, and culture for 4-5 days. Observe the cell state every day.

[0189] (5) SGC-7901 gastric cancer cell inoculation

[0190] Take the SGC-7901 gastric cancer cells in good growth state, and wash the cells with PBS solution as described above. Digest the cells with trypsin, terminate the digestion with cell culture solution, and prepare a cell suspension. Count the cell suspension, dilute the suspension to a concentration of 4x10 5 Each well of the 96-well plate is inoculated with 100 μL of the cell suspension, and the culture medium containing 10% fetal bovine serum is used as a control group. The 96-well plate is placed in the cell culture incubator for 24 h. Discard the supernatant, and add 100 μL of each of the prepared four groups of 6.25%, 12.5%, 25%, and 50% non-drug composite membrane extract and 10 groups of 100% drug-loaded composite membrane extract to the 96-well plate. Mark with a marker pen, and place in the cell culture incubator for 24, 48, and 72 h, respectively, for MTT experiment.

[0191] (6) MTT experiment

[0192] After culturing SGC-7901 gastric cancer cells for 24, 48, and 72 h, 150 μL of MTT solution (5 mg / mL) was added to each well in the dark, and the plate was incubated for another 4 h. The culture solution in each well was gently aspirated without touching the edge or bottom of the well. Then, 150 μL of dimethyl sulfoxide was added to each well, and the plate was shaken for about 10 min on a micro-vibrator. The plate was placed in a microplate reader, and the OD value of each well was detected at 570 nm. The cell survival rate was calculated by the formula.

[0193]

[0194]

[0195] (7) Inverted microscope observation of the effect of gastric retention shape memory composite film on SGC-7901 gastric cancer cells

[0196] The above same method was used to culture SGC-7901 gastric cancer cells with the 4 groups of 100% non-drug-loaded composite film extracts and the 10 groups of 100% drug-loaded composite film extracts. After culturing for 24, 48, and 72 h, the number and morphology of the cells were observed under an inverted microscope, and the cells were tracked and photographed using a camera system.

[0197] 3.2 Experimental results

[0198] (1) Toxicity evaluation of non-drug-loaded gastric retention shape memory composite film on SGC-7901 gastric cancer cells

[0199] The toxicity of the 4 groups of non-drug-loaded composite films on SGC-7901 gastric cancer cells was investigated. The experimental results are shown in Table 1. Figure 24 As shown in Table 1, after culturing SGC-7901 gastric cancer cells with 6.25%, 12.5%, 25%, 50%, and 100% non-drug-loaded composite film extracts for 24 h, 48 h, and 72 h, the survival rate of the cells was greater than 78.76%, and the cell toxicity grade was grade 1 or below, indicating that PLA, PLA / PCL (7 / 3), PLA / PEG (7 / 3), and PLA / PCL / PEG (8 / 2 / 3) had no significant toxicity on SGC-7901 gastric cancer cells.

[0200] (2) Inhibition of drug-loaded gastric retention shape memory composite film on SGC-7901 gastric cancer cells

[0201] The proliferation inhibition of the 10 groups of drug-loaded composite films on SGC-7901 gastric cancer cells was investigated. The experimental results are shown in Table 2. Figure 25As shown, after incubating SGC-7901 gastric cancer cells with 10 groups of drug-loaded composite membrane extracts at 100% concentration for 24 hours, the cell proliferation inhibition rate was greater than 50%, with the maximum inhibition rate being 74.10%. The inhibition rate increased with increasing incubation time. After 72 hours, LPC-PLA / PEG (7 / 3) achieved the maximum inhibition rate against SGC-7901 gastric cancer cells, reaching 89.04%. The 100% extract of LPC-PLA / PEG (7 / 3) showed the best inhibitory effect on SGC-7901 gastric cancer cells, which is related to its superior drug release performance.

[0202] (3) Observation of the effect of the composite membrane on the proliferation of SGC-7901 cells using an inverted microscope

[0203] The effects of extracts from non-drug-loaded shape memory composite membranes and drug-loaded shape memory composite membranes on the morphology of SGC-7901 gastric cancer cells were observed using an inverted microscope. Figure 26 As shown, the live cells in the negative control group had two shapes: spherical and spindle-shaped. The effects of four groups of 100% drug-free composite membrane extracts (PLA, PLA / PCL, PLA / PEG (7 / 3), and PLA / PCL / PEG (8 / 2 / 3)) on the morphology of SGC-7901 gastric cancer cells were first observed. From a morphological perspective, the SGC-7901 gastric cancer cells cultured in the four groups of 100% drug-free composite membrane extracts showed basically normal morphology, all being spherical or spindle-shaped. Furthermore, the cell number increased significantly with time, consistent with the morphology of the negative control group. Figure 27 Subsequently, the effects of 100% drug-loaded composite membrane extracts on the morphology of cultured SGC-7901 gastric cancer cells were observed. The results showed that the morphology of SGC-7901 gastric cancer cells cultured in 10 groups of 100% drug-loaded composite membrane extracts was significantly deformed, and the number of dead cells increased. The cell count was significantly reduced compared with the negative control group. Figure 28 The above analysis further reveals that the non-drug-loaded composite membrane extract has no cytotoxicity to SGC-7901 gastric cancer cells; the drug-loaded composite membrane extract significantly increases the number of dead SGC-7901 gastric cancer cells and has a significant inhibitory effect on the proliferation of SGC-7901 gastric cancer cells.

[0204] The inhibitory effects of unloaded and drug-loaded gastric retention shape memory composite membranes on SGC-7901 gastric cancer cells were evaluated using the MTT assay. The results showed that the unloaded shape memory composite membrane had no cytotoxicity against SGC-7901 gastric cancer cells, while the drug-loaded shape memory composite membrane exhibited significant inhibitory effects, with LPC-PLA / PEG (7 / 3) showing the strongest inhibitory effect.

[0205] Example 4 In vivo pharmacokinetic evaluation of the thermally responsive gastric retention shape memory drug-loaded composite film in rats

[0206] It is found by the above examples that the ten groups of drug-loaded composite films of LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3), LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3), LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2), and LPC-PLA / PCL / PEG (8 / 2 / 3) have good gastric retention drug delivery system characteristics and obvious inhibitory effect on SGC-7901 gastric cancer cells, and the comprehensive evaluation finds that the drug-loaded composite film of LPC-PLA / PEG (7 / 3) has the best performance as a gastric retention drug delivery system. In addition, the study of the pharmacokinetics of the drug-loaded composite film in rats can systematically observe whether the drug in the drug-loaded composite film has a longer blood concentration existence time, which is of great significance for studying the absorption and excretion process of the drug-loaded composite film in the body. In this embodiment, rats are used as animal models, and the pharmacokinetic properties of the gastric retention composite film in rats are explored by gavage. The drug-loaded composite film selected is the drug-loaded composite film of LPC-PLA / PEG (7 / 3) with the best comprehensive performance in the above examples for pharmacokinetic evaluation.

[0207] Preparation of the solution

[0208] Preparation of the control solution: 25.00 mg of the control (luteolin) was accurately weighed, added to a 50 mL volumetric flask, dissolved with acetonitrile, and made up to volume to obtain a control solution with a mass concentration of 0.50 mg / mL. Then 5 mL of the control solution was added to a 50 mL volumetric flask and made up to volume to obtain a solution with a mass concentration of 50 μg / mL.

[0209] Preparation of the internal standard solution: 12.50 mg of the control (luteolin) was accurately weighed, added to a 50 mL volumetric flask, dissolved with acetonitrile, and made up to volume to obtain an internal standard solution with a mass concentration of 0.25 mg / mL. Then 2 mL of the internal standard solution was added to a 50 mL volumetric flask and made up to volume to obtain a solution with a mass concentration of 10 μg / mL. Then 1 mL was taken and added to a 10 mL volumetric flask and made up to volume to obtain a 1 μg / mL internal standard solution.

[0210] Preparation of the luteolin and luteolin phospholipid complex suspension: luteolin and LPC were diluted with 0.5% CMC-Na solution, ultrasonically dissolved, and made into a suspension with a concentration of 1.50 mg / mL (calculated as luteolin).

[0211] Preparation of the LPLC-PLA / PEG (7 / 3) loaded capsule: The LPLA / PEG (7 / 3) was heated for 10 min, and then folded into the size of the rat capsule after it became soft. The shape was fixed at a temperature of about 5°C, and then loaded into the rat capsule.

[0212] Grouping and administration: 18 male SD rats were divided into 3 groups, 6 rats in each group, and were divided into luteolin group, LPLC group and LPLC-PLA / PEG (7 / 3) loaded capsule group. The temperature in the animal room was kept at 25±0.5°C and the humidity was 50±5%. The rats were acclimated for 7 days under the condition of 12h light and dark alternation. The prepared luteolin suspension, LPLC suspension and LPLC-PLA / PEG (7 / 3) loaded capsule were administered to each rat by gavage, and the dose was 20mg / kg (based on luteolin). 12h before administration, the rats were fasted but allowed to drink water. Blood was collected at 10min, 20min, 30min, 1h, 2h, 4h, 8h and 12h after administration. After blood collection, an equal amount of normal saline was given. The blood collection process was as follows: about 600μL of blood was collected from the retro-orbital plexus of each rat into a small amount of heparin wetted 1.5mL centrifuge tube, and centrifuged at 4000r / min for 10min. The supernatant was collected to obtain rat plasma.

[0213] Pre-treatment of plasma samples: 200μL of rat plasma sample and 20μL of internal standard solution were precisely pipetted into a centrifuge tube. 100μL of 3mol / L HCl solution was added, and after vortex mixing, it was placed in a 65°C water bath for 3h. 150μL of 5% HClO4 solution was added to the centrifuge tube, and after shaking for 10min, 3mL of ethyl acetate was added for extraction, and vortexed for 15min. Then, the extraction solution was centrifuged at 10000r / min for 12min, and the upper organic phase was transferred to another empty centrifuge tube. 1.0mL of ethyl acetate was added for further extraction, and the combined extract was dried under nitrogen at 45°C. Finally, 100μL of mobile phase was added to the centrifuge tube for re-dissolution, and a 0.45μm microporous filter was used for filtration.

[0214] Establishment of luteolin HPLC analysis method

[0215] The sample concentration of luteolin was detected by internal standard method using luteolin as internal standard. The chromatographic conditions of the luteolin detection method were as follows.

[0216] Chromatographic column: Inertsil ODS-3 C18 chromatographic column (250mm×4.6mm, 5μm); mobile phase: acetonitrile-0.1% potassium dihydrogen phosphate water (55:45) elution; injection volume: 20μL; flow rate: 1mL / min; column temperature: 30°C.

[0217] Data processing and method: the pharmacokinetic program software DAS 2.1 was used to calculate the pharmacokinetic parameters of luteolin. All data were expressed as mean ± standard deviation (x ± s), and the data between two groups were analyzed by t test using SPSS 19.0 statistical software.

[0218] 4.1 Specificity of luteolin detection method

[0219] Take blank rat plasma, add a certain amount of control solution, prepare plasma samples. Another blank plasma and luteolin suspension plasma 200 μL, according to the above method of plasma and detection conditions for analysis.

[0220] The chromatographic results of rat blank plasma sample, blank plasma added with luteolin sample and plasma sample after administration are shown in Figure 29 The retention time of luteolin is 6.2 min, and the retention time of dihydroxydihydroguaiaretic acid is 10.6 min. Luteolin and dihydroxydihydroguaiaretic acid are not interfered by endogenous substances in plasma, and the method has good specificity.

[0221] 4.2 Standard curve, detection limit and quantification limit of luteolin detection method

[0222] Take appropriate amount of blank plasma, add luteolin solution with different dilution times, prepare plasma sample solution containing luteolin with concentration of 0.125 μg / mL, 0.25 μg / mL, 0.5 μg / mL, 1 μg / mL, 1.5 μg / mL and 2.5 μg / mL, repeat 6 times for each concentration. According to the above method of plasma and detection conditions for analysis.

[0223] Take drug concentration as abscissa, and take the ratio of luteolin peak area to dihydroxydihydroguaiaretic acid peak area as ordinate, and make standard curve. According to 3 times signal-to-noise ratio (S / N=3), the detection limit of luteolin is calculated, and according to 10 times signal-to-noise ratio (S / N=10), the quantification limit of luteolin is calculated.

[0224] The standard curve regression equation of luteolin detection method in rat plasma is y=0.3627x+0.2018. Luteolin has good linear relationship in the linear range of 0.125-15 μg / mL Figure 30 ). The detection limit is 0.0211 μg / mL, and the quantification limit is 0.1396 μg / mL.

[0225] 4.3 Recovery rate test and intra-day and inter-day precision test of luteolin detection method

[0226] The appropriate amount of control sample solution was precisely pipetted into the blank plasma of rats to prepare low (0.125 μg / mL), medium (1 μg / mL), and high (2.5 μg / mL) concentration plasma sample solutions, with 6 replicates in each group. 100 μL of each solution was taken, and the plasma was treated and analyzed according to the above method and detection conditions. The results were brought into the standard curve to calculate the recovery. The sample was injected 6 times within 1 day to calculate the within-day precision. Under the same conditions, the sample was injected once a day, and the injection was continuously performed for 3 days to calculate the inter-day precision.

[0227] The low, medium, and high concentration samples of luteolin were detected and analyzed, and the results are shown in Table 13. The within-day precision RSD was 3.43-7.71%, the inter-day precision was 4.70-11.93%, and the recoveries of both were between 100.00-110.00%, which indicated that the detection standard of luteolin in rat plasma was met.

[0228] Table 13 Precision evaluation of plasma (6 replicates per day, continuously measured for 3 days)

[0229]

[0230] The recovery test results are shown in Table 14. The extraction recovery of luteolin in rat plasma was 76.36-88.15%, with an RSD of 1.33-2.67%; the extraction recovery of the internal standard was 75.24-80.20%, with an RSD of 1.89-2.60%.

[0231] Table 14 Recovery test (n = 6)

[0232]

[0233]

[0234] 4.4 Stability test of luteolin in blank plasma

[0235] The appropriate amount of control sample solution was precisely pipetted into the blank plasma of rats to prepare low (0.125 μg / mL), medium (1 μg / mL), and high (2.5 μg / mL) concentration plasma sample solutions, with 6 replicates in each group. 100 μL of each solution was taken, and the plasma was treated and analyzed according to the above method and detection conditions, and the concentration of luteolin was calculated. The sample was stored in a refrigerator at -20°C for 30 days, and then the plasma treated according to the above method was analyzed according to the above detection conditions. The results were brought into the standard curve to calculate the concentration of luteolin. The stability was calculated according to the relative standard deviation.

[0236] The results of the stability experiment of luteolin in rat plasma are shown in Table 15. The RSD of the plasma sample with luteolin was less than 15% after being stored at -20°C for 30 days, indicating that luteolin is relatively stable in rat plasma.

[0237] Table 15. Results of the stability experiment in rat plasma (n = 6)

[0238]

[0239] 4.5 Blood concentration and pharmacokinetic parameter calculation

[0240] (1) Determination and analysis of the blood concentration of luteolin

[0241] The pharmacokinetic parameters of luteolin were calculated using the pharmacokinetic program software DAS 2.0. Under the above conditions, the rats in the luteolin suspension group, the LPC suspension group, and the LPC-PLA / PEG (7 / 3) complex film group were administered with luteolin at a concentration of 20 mg / kg by gavage, and the blood was collected from the orbit at fixed time points. Each experiment was repeated 6 times. The time of blood collection from the orbital venous plexus after administration was used as the horizontal coordinate, and the concentration of luteolin in the plasma was used as the vertical coordinate to obtain the plasma drug concentration-time curve (Fig. 2). Figure 31 The results showed that there were certain differences in the absorption and excretion of luteolin among the three groups. The blood concentration of luteolin reached the peak value at 2 h after gavage, while the blood concentration of LPC reached the peak value at 1.5 h after gavage, and the curve of LPC was biphasic. The sharp peak shape of the absorption peak indicated that the drug was rapidly absorbed into the systemic circulation, followed by a slower excretion phase. Compared with the luteolin group, the LPC group reached the peak value faster, which was presumably due to the fact that luteolin was wrapped in phospholipids, promoting drug release. The peak value of the blood concentration of the LPC group was 2.1 times that of the luteolin group, indicating that the preparation of luteolin into a phospholipid complex effectively improved the poor water solubility, making it easier for the human body to absorb. The maximum blood concentration peak of the LPC-PLA / PEG (7 / 3) complex film group was at 8 h, and the blood concentration was maintained before 24 h. Compared with the luteolin group (6 h) and the LPC group (18 h), the LPC-PLA / PEG (7 / 3) complex film could effectively prolong the retention time of the blood concentration, indicating that the gastric retention effect of the LPC-PLA / PEG (7 / 3) complex film was relatively ideal. However, the maximum blood concentration of the LPC-PLA / PEG (7 / 3) complex film was lower than that of the LPC group, which was presumably mainly due to the fact that the luteolin in the drug-loaded complex film was not completely released from the complex film.

[0242] (2) Calculation and analysis of the pharmacokinetic parameters

[0243] Pharmacokinetic parameters were calculated using the pharmacokinetic software DAS 2.0, conforming to an absorbable two-compartment model in rats with a weight of 1 / C. A non-compartmental model was also used to obtain the main pharmacokinetic parameters (Table 16). The results showed that after gavage administration, the AUC in the plasma of rats in the LPC group was... (0-∞) The concentration was 5697.04 μg / L·h, which was 3.54 times higher than that of luteolin, with a relative bioavailability increased by 354%, Cmax increased by 1.14 times, and mean residence time (MRT) increased. (0-t) It also significantly prolonged. The plasma AUC in rats of the LPC-PLA / PEG(7 / 3) group was significantly increased. (0-∞) The concentration was 7155.05 μg / L·h, which was 4.70 times higher than that of luteolin (AUC(0-∞)) and 470% higher relative bioavailability. max The AUC of the LPC-PLA / PEG(7 / 3) composite membrane was increased by 0.58 times compared to the LPC group. (0-∞) It increased by 0.26 times, relative bioavailability increased by 26%, and mean residence time (MRT) increased. (0-t) It also significantly prolonged.

[0244] Table 16 Pharmacokinetic parameters in rat plasma after gavage administration

[0245]

[0246]

[0247] This embodiment establishes a high-performance liquid chromatography (HPLC) method for determining luteolin content in rat plasma. Methodological investigation has demonstrated that the method is simple and stable, with good specificity, and the recovery rate, precision, and repeatability all meet the determination requirements. It can be used for the determination of luteolin content in rat plasma and the evaluation of its bioavailability in vivo.

[0248] Pharmacokinetic studies showed that the time to peak concentration (T3) of the LPC-PLA / PEG(7 / 3) group was [details missing]. max The peak concentration in the LPC group was 8 hours, which was significantly longer than that in the luteolin group and the LPC group; the peak concentration C in the LPC group was... max The peak concentration C of the LPC-PLA / PEG(7 / 3) group was 920.00 μg / L. max The effective concentration was 680.00 μg / L, which was significantly higher than that of the luteolin group; the area under the curve (AUC) of the LPC-PLA / PEG(7 / 3) group, the LPC group, and the luteolin group was also significantly higher. (0-∞)The relative bioavailability of the LPC-PLA / PEG (7 / 3) group and the LPC group was 7155.05 μg / L·h, 5697.04 μg / L·h and 1256.13 μg / L·h, respectively, which was 4.70 times and 3.54 times higher than that of the luteolin group, respectively. Among them, the bioavailability of the LPC-PLA / PEG (7 / 3) was the highest. The pharmacokinetic experiment results show that the bioavailability of luteolin in the LPC-PLA / PEG (7 / 3) in the rat body is further improved on the basis of the LPC, and the advantage of the gastric retention drug delivery system is exerted.

[0249] In summary, the shape memory polymer is combined with the gastric retention drug delivery system in the present application, luteolin phospholipid complex is used as the main drug, and 10 groups of shape memory gastric retention composite films with different materials are prepared. The performance of the 10 groups of drug-loaded composite films is screened, the best performance of the LPC-PLA / PEG (7 / 3) is subjected to the pharmacokinetic study, and it is found that the bioavailability is obviously improved compared with luteolin and LPC.

[0250] (1) Preparation and quality evaluation of luteolin phospholipid complex

[0251] LPC is prepared, which aims to improve the water solubility of luteolin and facilitate oral absorption in the human body. The drug content determination methods mainly include ultraviolet spectrophotometry, high performance liquid chromatography, liquid chromatography-mass spectrometry and the like. The present application adopts the precise high performance liquid chromatography to determine the content of luteolin. The ultraviolet wavelength scanning determines that the ultraviolet maximum absorption wavelength of luteolin is 348 nm, therefore, 348 nm is selected as the detection wavelength. At the same time, the established high performance liquid method is verified to be accurate and reliable by the specificity, precision, repeatability, sample recovery rate and stability, and can be used for detecting the content of luteolin in the LPC.

[0252] The solubility of luteolin and soybean lecithin in different solvents is detected in the pre-experiment, and it is found that both of them are soluble in ethanol and tetrahydrofuran. Further experiments show that the complexing rate of the LPC prepared in tetrahydrofuran is higher under the same conditions, which is speculated to be due to the interference of the hydrogen bond donor -OH bond in ethanol to the complexing process of phospholipid and drug, so the complexing rate is lower than that in tetrahydrofuran. Therefore, tetrahydrofuran is selected as the preparation solvent in the preparation process of the LPC.

[0253] The present application screens the best LPC prescription with the best complex rate, and the preparation method is reasonable and reliable, and the complex rate is ideal. With the complex rate as the index, the stirring time, stirring temperature, luteolin mass concentration, and luteolin to soybean lecithin ratio four evaluation indexes are investigated, and the optimal process is obtained through experimental analysis, that is, luteolin and soybean lecithin are weighed according to the mass ratio of 1:1.2, an appropriate amount of tetrahydrofuran is added to make the mass concentration of the former 10 mg / mL, constant temperature stirring is carried out at 50 DEG C for 6 h, the organic solvent is removed by rotary evaporation under reduced pressure, and the sample is dried in a vacuum drying oven at 40 DEG C overnight or in a vacuum drying oven for 48 h, and it is found through verification test that the average complex rate is 91.45%. The preparation scheme with the highest LPC complex rate is screened, and it is prepared for the drug preparation of the later prepared LCP-containing gastric retention shape memory composite film.

[0254] After the preparation of LPC, the quality evaluation and physical characterization are carried out. The optical microscope is one of the common ways for observing the particle morphology, which can use light source and glass lens to magnify the observed image. The morphological change of the phospholipid complex is observed by the optical microscope, and it is observed that the LCP particles are round and the dispersed LCP particles are surrounded by obvious light circles, which are soybean lecithin layers, indicating that the soybean lecithin has good wrapping property for luteolin. The structure absorption characteristics are analyzed by ultraviolet absorption spectrum method and infrared absorption spectrum method, and it is found that the structure characteristics of the raw material and soybean lecithin are found, and the ultraviolet absorption peak observation finds that the absorption curves of luteolin and LCP are basically the same, and there are two obvious characteristic absorption bands at about 254 nm and 350 nm, the chemical structure is not fundamentally changed, and no new substance is generated. The infrared experiment observation finds that the infrared spectrum of LCP has significant change. The improvement of water solubility is crucial for oral utilization of luteolin, the water solubility of the raw drug is enhanced by the combination of luteolin and soybean lecithin, the -OH group of luteolin can interact with the phospholipid molecule to form a complex, improve the water solubility of luteolin, strengthen the permeability of luteolin to biological membrane, improve the gastrointestinal absorption effect of luteolin, and improve the bioavailability of the drug. The water solubility of luteolin, physical mixture and LCP sample is determined, and the results show that the solubility of LCP in water is more than 2 times that of luteolin.

[0255] The phospholipid complex can be combined with various new preparation technologies, such as nanoparticles, liposomes and sub-micro emulsions, in addition to being used as an intermediate to prepare ordinary pharmaceutical dosage forms, so as to accelerate the dissolution of the drug, prolong the circulation time of the drug in the body, and improve the bioavailability of the drug. The present application combines it with a shape memory polymer to prepare a gastric retention shape memory composite film, combines the fast solubility of LCP with the gastric retention property of the composite film, and improves the oral bioavailability of luteolin.

[0256] (2) Preparation and characterization of drug-loaded gastric retention shape memory composite film

[0257] LPC is loaded in shape memory polymer to prepare a thermally responsive drug-loaded gastric retention shape memory composite film, which can prolong the retention time of the drug-loaded composite film in the stomach by using the deformable properties and floating properties of the composite film. First, LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), LPC-PLA / PCL / PEG(8 / 2 / 3) gastric retention shape memory composite films are prepared. PLA is used as the main material, and the drug loaded is the best LPC in terms of composite rate. PCL and PEG are added to the composite film to modify the brittleness of PLA and reduce the glass transition temperature of PLA, so that the deformation temperature is more suitable for the human body; the addition of HPMC, a auxiliary material, increases the water retention of the composite film as a whole; the addition of NaHCO3 in the composite film increases the generation of bubbles when the composite film contacts gastric acid in the stomach, which is beneficial to the floating of the composite film.

[0258] Subsequently, the chemical composition, thermal properties, thermal stability, apparent morphology, drug release performance, floating performance, shape memory performance of the gastric retention shape memory composite film of LPC-PLA, LPC-PLA / PCL (9 / 1), LPC-PLA / PCL (8 / 2), LPC-PLA / PCL (7 / 3), LPC-PLA / PEG (9 / 1), LPC-PLA / PEG (8 / 2), LPC-PLA / PEG (7 / 3), LPC-PLA / PCL / PEG (8 / 2 / 1), LPC-PLA / PCL / PEG (8 / 2 / 2), LPC-PLA / PCL / PEG (8 / 2 / 3) are investigated. The experimental results show that the components in the prepared drug-loaded gastric retention shape memory composite film will not affect the chemical properties of the composite film due to physical mixing, and the characteristic absorption peaks of each component can be observed by infrared spectrum analysis. By blending PLA, PBS and PEG polymers, it is found by infrared spectrum analysis that the material contains the characteristic functional groups of PLA, PEG and PBS, and the characteristic peaks of PLA, PBS and PEG do not appear obvious displacement change, and no new molecular characteristic peak appears, so it is speculated that the chemical reaction of the three molecules has not occurred. Tg is a basic parameter of thermal responsive SMPs, because it can directly determine the application situation of SMPs and the execution degree of shape memory effect (SME). If Tg is too high, the shape memory film agent in the body will have poor unfolding effect, or the high temperature will cause damage to human organs. The thermal property results show that the addition of PCL and PEG in the composite film can reduce the Tg of the composite film, so that the deformation temperature of the composite film is closer to the body temperature. And in the LPC-PLA / PCL / PEG (8 / 2 / 3) group, the Tg value reaches the lowest compared with other groups, which is 44.72℃. In the research of Dong Xue-ming et al., the Tg of the prepared PLA-based magnetic driving bone scaffold is 64.09℃. In the research of Senatov et al., a PLA-based porous scaffold containing hydroxyapatite with shape memory function is printed by FDM, and DSC detection finds that its Tg is 57.1℃. The deformation temperature of the PLA-based composite film prepared in the application is lower than the above products, and the Tg is more suitable for the human body. The thermal stability analysis results show that all materials will decompose under heating. When the temperature is about 450℃, the carbon residue rate of the material is 0%, and the material can be considered to be completely decomposed. The addition of LPC, HPMC and NaHCO3 can significantly reduce the initial degradation temperature of PLA, and the initial decomposition temperature of the material appears at about 200℃, which shows that the addition of HPMC and NaHCO3 reduces the thermal stability of the composite material compared with PLA. The addition of PCL slightly improves the heat resistance of the composite film, and the addition of PEG reduces the heat resistance of the composite film to a certain extent. Overall analysis, the thermal stability of the 10 groups of composite films is good, and the properties can remain stable below 200℃.Optical microscope observation showed that the surface of drug-loaded composite film was light yellow, and the light yellow was uniformly distributed, and the drug had been well distributed on the composite film. Compared with PLA, LPC-PLA had slight phase separation, which was the result of the addition of excipients HPMC and NaHCO3. It was found that PEG had better compatibility with PLA than PCL, and the higher the proportion of PEG, the more uniform the surface of the composite film. In the gastric retention drug delivery system, prolonging and controlling gastric emptying time can effectively improve the bioavailability of drugs in the body and reduce drug waste. Gastric retention drug delivery system helps to provide better new products, with new therapeutic possibilities and substantial benefits for patients. Increasing floating time and making the size of the dosage form larger than the pylorus are all conducive to increasing the retention time of the dosage form in the stomach. The results of the floating experiment showed that the 10 groups of drug-loaded gastric retention shape memory composite films had good floating performance, and the floating time was more than 8h. Among them, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PEG(8 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 1) can float for up to 48h, indicating that the prepared drug-loaded gastric retention shape memory composite film has good floating performance. Shape memory testing found that the addition of excipients HPMC and NaHCO3 in the composite film had a certain effect on the shape recovery rate of the composite film, which prolonged the shape recovery time of the composite film compared with PLA, and reduced the recovery rate. When the environmental temperature was 50℃, the 10 groups of drug-loaded composite films could reach a recovery rate of 70% within 60s, and each composite film combination had good shape memory recovery performance. When the environmental temperature was 37℃, the deformation recovery rate of all composite films was about 60%, which met the requirement that the gastric retention shape memory composite film should be larger than the pylorus size when it reached the stomach. Drug release is particularly important in drug dosage form design, and good drug release is related to the bioavailability of the drug. By analyzing the effect of polymer ratio, excipients and drug concentration on drug release in the composite film, the addition ratio of excipients in the composite film was optimized and the best drug release performance polymer combination was obtained. First, the best polymer combination was selected, and under the condition that the concentration of HPMC was kept constant, the drug release performance of the 10 groups of composite films was observed. The results showed that LPC-PLA / PEG(7 / 3) had the maximum cumulative drug release at all detection time points, and had the best drug release performance. Then, based on LPC-PLA / PEG(7 / 3), the best addition amount of excipients and drug concentration was investigated. The results showed that when the concentration of HPMC was 5%, the concentration of NaHCO3 was 5%, and the concentration of luteolin was 30%, the cumulative release rate of luteolin in LPC-PLA / PEG(7 / 3) was the largest. At 24h, the cumulative drug release rate of the gastric retention shape memory composite film of LPC-PLA / PEG(7 / 3) reached 75.56%.Based on drug release analysis, the optimal excipient ratio in the drug-loaded composite membrane is 5% HPMC and 5% NaHCO3. Among different polymer material combinations, the LPC-PLA / PEG(7 / 3) composite membrane exhibits the best drug release performance.

[0259] (3) Evaluation of the inhibitory effect of gastric retention shape memory composite membrane on gastric cancer cells

[0260] This invention further analyzed the inhibitory effect of drug-loaded composite membranes on the proliferation of human gastric cancer cells SGC-7901. Since the biomaterials are prepared for oral formulation, cytotoxicity must be considered first. The cytotoxicity of unloaded PLA, PLA / PCL (7 / 3), PLA / PEG (7 / 3), and PLA / PCL / PEG (8 / 2 / 3) composite membranes was detected using the MTT assay, and the morphological effects on SGC-7901 gastric cancer cells were observed using an inverted microscope, providing a preliminary basis for the application of oral formulations. The MTT results generally showed that the four groups of unloaded composite membranes exhibited no cytotoxicity. Inverted microscopy observation revealed no effect on the morphology of SGC-7901 gastric cancer cells. The inhibitory effect of 10 groups of drug-loaded composite membranes on SGC-7901 gastric cancer cells was evaluated using the MTT assay, and the morphological effects on SGC-7901 gastric cancer cells were observed using an inverted microscope. The results showed that the drug-loaded composite membranes achieved the highest inhibition rate of 89.04% against SGC-7901 gastric cancer cells. Analysis revealed that the 100% composite membrane extract of LPC-PLA / PEG(7 / 3) had the strongest inhibitory effect on SGC-7901 gastric cancer cells, which is related to its best drug release properties.

[0261] (4) Pharmacokinetic evaluation of thermoresponsive gastric retention shape memory drug-loaded composite membrane in rats

[0262] This invention establishes a method for determining the content of luteolin in rat plasma using high-performance liquid chromatography (HPLC). Method specificity validation demonstrated complete separation of the luteolin peak from impurity peaks in plasma, with a retention time of approximately 6.2 min. The method exhibited good linearity within the plasma luteolin content range of 0.125-15 μg / mL. Both intra-day and inter-day precision recoveries were between 100-110%, and precision was less than 15%, indicating good injection repeatability and meeting the assay requirements. These results indicate that the method is simple, stable, specific, has good recovery and repeatability, and high precision, making it suitable for determining the luteolin content in rat plasma and studying its pharmacokinetic properties in rats.

[0263] Pharmacokinetic parameters reflect the pharmacokinetic characteristics of drugs, and analyzing these parameters can provide a basis for developing a scientific drug regimen in clinical practice. They are important indicators for evaluating the quality of drugs. We compared and analyzed the differences in the elimination half-life (T 1 / 2 ), peak concentration (C max ), time to peak (T max ), area under the curve (AUC), and mean residence time (MRT) of luteolin, LPC, and LPC-PLA / PEG (7 / 3) complex membrane groups. The results showed that some parameters were different among the three groups. The T 1 / 2 of LPC-PLA / PEG (7 / 3) was longer than that of luteolin and LPC groups, indicating that luteolin loaded in PLA / PEG (7 / 3) had a longer elimination time in rats. Because the drug was loaded in PLA / PEG (7 / 3), the folded PLA / PEG (7 / 3) returned to its original shape at the temperature of the stomach, making the size of the drug carrier larger than the pylorus, achieving a retention effect and prolonging the duration of drug release. Therefore, it showed a longer T 1 / 2 , and could maintain a longer effective blood concentration in the animal body. The peak concentration C max is the highest concentration of drug in the plasma after administration to animals, which is used to evaluate the absorption rate and extent of the drug. The C max of the LPC group was 920.00 μg / L, which was significantly higher than the C max of the luteolin group (430.00 μg / L), indicating that the LPC group could maintain a higher blood concentration. This may be due to the effective combination of soy lecithin and luteolin in LPC, which improves the water solubility of luteolin and is beneficial to the absorption of drugs by the body. The C max of the LPC-PLA / PEG (7 / 3) complex membrane was 680.00 μg / L, which was lower than that of the LPC group. This may be due to the incomplete release of luteolin loaded in the LPC-PLA / PEG (7 / 3) complex membrane. The T max and MRT of the LPC-PLA / PEG (7 / 3) complex membrane were significantly longer than those of the luteolin and LPC groups, indicating that the LPC-PLA / PEG (7 / 3) complex membrane could achieve drug sustained release in rats, prolonging the action time of the drug. The T max of the LPC-PLA / PEG (7 / 3) group was 8 h, which was significantly delayed compared with the luteolin and LPC groups. The peak concentration of the LPC-PLA / PEG (7 / 3) group and the LPC group was also effectively improved compared with the luteolin group.

[0264] The area under the curve (AUC) (0-∞) is t0-t ∞The area under the drug-time curve within a given time frame is an important indicator for evaluating drug absorption. The AUC of the LPC group... (0-∞) The AUC of the LPC-PLA / PEG(7 / 3) composite membrane was 5697.04 μg / L·h. (0-∞) The AUC of the luteolin group was 7155.05 μg / L·h. (0-∞) The relative bioavailability (RBA) of luteolin was 1256.13 μg / L·h, indicating that preparing luteolin into LPC or LPC-PLA / PEG(7 / 3) composite membranes effectively improved the drug's absorption in rats. Bioavailability refers to the rate and extent to which a drug is absorbed into the systemic circulation and is an important parameter determining the dose-effect relationship. When drug formulations are administered via non-intravenous routes, the AUC of the oral reference standard drug is often used for comparison, i.e., relative bioavailability is evaluated. The relative bioavailability of LPC was 3.54 times higher than that of the luteolin group, and the relative bioavailability of LPC-PLA / PEG(7 / 3) was 4.70 times higher than that of the luteolin group, indicating that the LPC-PLA / PEG(7 / 3) composite membrane had the best bioavailability. LPC-PLA / PEG(7 / 3) can effectively improve the pharmacokinetic characteristics of luteolin in rats.

[0265] Luteolin exhibits extremely poor solubility and low bioavailability in water, thus limiting its clinical use. Phospholipid complexes, by encapsulating the drug with phospholipids, can improve drug solubility and bioavailability, and delay its clearance rate in vivo. Furthermore, the addition of phospholipids enhances the drug's stability, reduces interfacial tension with gastrointestinal fluids, promotes drug translocation within membranes and tissues, and effectively prolongs the drug's half-life in the blood. This invention prepares the drug into a gastric retention drug delivery system, extending the drug carrier's retention time in the stomach, which can also effectively improve the drug's gastric retention time and bioavailability. This invention uses high-performance liquid chromatography (HPLC) to study the pharmacokinetic characteristics of luteolin, LPC, and LPC-PLA / PEG(7 / 3) in rats. The results show that oral administration of LPC-loaded PLA / PEG(7 / 3) prolonged the drug's action time and improved its bioavailability. The findings of this invention will provide a basis for the clinical application of modified drug-loaded shape memory polymers.

[0266] The application prepares a new type of gastric retention shape memory composite film, uses luteolin coated by soybean lecithin as a drug model, shape memory PLA as a base material, and adds PEG and PCL in different proportions to modify the base material, and HPMC and NaHCO3 to modify the polymer material, to prepare LPC-PLA, LPC-PLA / PCL(9 / 1), LPC-PLA / PCL(8 / 2), LPC-PLA / PCL(7 / 3), LPC-PLA / PEG(9 / 1), LPC-PLA / PEG(8 / 2), LPC-PLA / PEG(7 / 3), LPC-PLA / PCL / PEG(8 / 2 / 1), LPC-PLA / PCL / PEG(8 / 2 / 2), and LPC-PLA / PCL / PEG(8 / 2 / 3) gastric retention shape memory composite films, and the best composite rate of luteolin phospholipid complex (LPC), the preparation of the drug-loaded shape memory composite film and the performance, the cytotoxicity of the shape memory composite film, and the rat in-vivo pharmacokinetics are studied. The new type of gastric retention shape memory composite film prepared by the application can effectively improve the bioavailability of luteolin, and can be better applied to the gastric retention drug delivery system as the shape memory polymer.

[0267] The above only describes the preferred embodiments of the application and is not intended to limit the application, and any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the application shall be included in the protection scope of the application.

Claims

1. A method for preparing a gastric retention shape memory composite membrane, characterized in that, Includes the following steps: The polymer is mixed with dichloromethane to obtain solution 1; the polymer is selected from any of the following mixtures; A mixture of PLA and PCL with a PLA to PCL weight ratio of 9:1; A mixture of PLA and PCL with a PLA to PCL weight ratio of 8:2; A mixture of PLA and PEG with a PLA to PEG weight ratio of 9:1; A mixture of PLA and PEG with a PLA to PEG weight ratio of 8:2; A mixture of PLA, PCL and PEG with a weight ratio of PLA, PCL and PEG of 8:2:1; (2) Solution 2 is prepared using water as a solvent. Solution 2 contains hydroxypropyl methylcellulose and NaHCO3. The mass of hydroxypropyl methylcellulose is 5% of the polymer mass, and the mass of NaHCO3 is 5% of the polymer mass. (3) Mix solution 1 and solution 2, stir, add luteolin phospholipid complex, the mass of luteolin phospholipid complex is 30% of the total mass of polymer, hydroxypropyl methylcellulose and NaHCO3; continue stirring, evaporate dichloromethane and water; the preparation method of luteolin phospholipid complex includes the following steps: dissolve soybean lecithin and luteolin in tetrahydrofuran, stir, and recover under reduced pressure to obtain luteolin phospholipid complex; (4) Molding to obtain the gastric retention shape memory composite membrane.

2. The method for preparing a gastric retention shape memory composite membrane according to claim 1, characterized in that, The mass ratio of polymer to dichloromethane is 1:

7.

3. The method for preparing a gastric retention shape memory composite membrane according to claim 1, characterized in that, The total mass ratio of hydroxypropyl methylcellulose and NaHCO3 to water is 1:

10.

4. The method for preparing a gastric retention shape memory composite membrane according to any one of claims 1-3, characterized in that, The mass ratio of luteolin to soybean lecithin was 1:1.2; after dissolving in tetrahydrofuran, the concentration of luteolin was 10 mg / mL; the stirring time was 6 h, and the stirring temperature was 50℃.

5. A method for preparing a gastric retention shape memory composite membrane according to any one of claims 1-3, characterized in that, In step (3), after stirring for 3.5 h, add the luteolin phospholipid complex and continue stirring for 0.5 h.

6. A method for preparing a gastric retention shape memory composite membrane according to any one of claims 1-3, characterized in that, A sheet with a thickness of 1 mm was produced by compression molding at a pressure of 10 MPa.

7. The application of gastric retention shape memory composite membrane in the preparation of drugs for treating gastric cancer, characterized in that, The gastric retention shape memory composite membrane is prepared using the preparation method described in any one of claims 1-6.

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