A fluorine-containing amphiphilic block polymer, a preparation method thereof, and application thereof
By synthesizing fluorinated amphiphilic block polymers, the problems of low loading rate and complex synthesis of block polymers in the prior art have been solved, achieving efficient drug loading and targeted therapy with good biocompatibility and drug release effect.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- HAINAN NORMAL UNIV
- Filing Date
- 2023-05-09
- Publication Date
- 2026-08-04
AI Technical Summary
Existing block polymers have low loading rates when loading poorly soluble antitumor drugs, and the synthesis steps are cumbersome and produce many byproducts, making it difficult to achieve efficient targeted drug release and good biocompatibility.
Fluorinated amphiphilic block polymers were synthesized by controlled polymerization. Polyethylene glycol was used as the hydrophilic block and fluorinated acrylate as the hydrophobic block. The monomer ratio was controlled, and the polymers were self-assembled into spherical particles, loaded with poorly soluble drugs, and activated by infrared light to generate reactive oxygen species for targeted therapy.
It achieves high drug loading rate, simplifies synthesis steps, improves drug solubility and bioavailability, enables targeted release of polymer at tumor sites, and exhibits cytotoxicity under light irradiation, with good biocompatibility.
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Figure CN116751347B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to a fluorinated amphiphilic block polymer, its preparation method and its application, belonging to the technical fields of biomaterials, polymer materials and pharmaceutical carrier materials. Background Technology
[0002] Malignant tumors are one of the leading causes of death worldwide. To date, governments around the world invest significant human and material resources annually in the diagnosis and treatment of cancer to improve its effectiveness. Despite some progress, the results remain unsatisfactory.
[0003] Block polymers are a special type of polymer prepared by linking two or more polymer segments with different properties together. These polymers have controllable molecular weights, narrow molecular weight distributions, and designable molecular structures and compositions, making them one of the most significant and challenging research areas in polymer science. Block polymers with specific structures exhibit properties different from simple linear polymers, many random copolymers, and even mixtures of homopolymers, and have wide applications in biomedicine, construction, chemical engineering, and other fields. Amphiphilic block polymers can encapsulate poorly soluble, highly toxic, and bioavailable drugs into nanoparticles, which can then be transported to the tumor site via the EPR effect within the tumor. Therefore, developing new drugs in this field is a hot topic, although current loading rates are low.
[0004] Chinese invention patent CN201710336880.3 discloses a method for preparing and applying a drug-loaded amphiphilic polycarbonate-polyethylene glycol polymer using Schiff bases. The method utilizes polyethylene glycol as a hydrophilic block and connects a poorly soluble antitumor drug to the carbonate structure via a Schiff base as a hydrophobic block, thus preparing multifunctional biodegradable polymer carrier materials of different molecular weights. This polymer carrier material exhibits amphiphilic properties and can be used to prepare polymeric nanomicelles for the drug, improving the solubility and bioavailability of the poorly soluble drug. It also possesses pH sensitivity for targeted drug release, and the released drug matrix exhibits good hydrophilicity, allowing for rapid tissue metabolism and excellent biocompatibility. However, due to the characteristics of the reaction, a specific ratio of hydrophilic to hydrophobic monomers is required, preventing optimal loading of the largest possible drug volume. Furthermore, the synthesis steps are cumbersome, and numerous byproducts are generated. Summary of the Invention
[0005] To address the aforementioned technical problems, this invention provides a fluorinated amphiphilic block polymer, its preparation method, and its applications. The fluorinated amphiphilic block polymer of this invention is synthesized by controlled polymerization of a hydrophilic material, polyethylene glycol, and a hydrophobic material, fluorinated acrylate. The ratio of hydrophilic to hydrophobic materials can be freely changed to maximize the loading of the hydrophobic perfluorocrown ether. The synthesis process of this fluorinated amphiphilic block polymer is simple; it can self-assemble into spherical particles in aqueous solution, exists stably in blood, and generates reactive oxygen species upon reaching the tumor region under infrared light irradiation. It has no toxic effect on cells in the absence of light irradiation.
[0006] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is as follows:
[0007] The structure of a fluorinated amphiphilic block polymer is as follows:
[0008]
[0009] Wherein, R1 is a hydrophilic group, R2 is a fluorinated hydrophobic group, and n and m are the degrees of polymerization.
[0010] R1 can be any acrylate hydrophilic group, preferably polyethylene glycol methyl ether acrylate. The monomer ratio can be controlled through ATRP polymerization; n and m can be arbitrary values, preferably 10 ≤ n ≤ 15, 5 ≤ m ≤ 10, and n ≥ m. More preferably, both n and m are 10. At this point, the polymer's drug loading rate can reach its maximum.
[0011] The polymers mentioned above contain fluorine, and their nm values can be changed arbitrarily without affecting each other, in order to achieve the optimal ratio for loading drugs.
[0012] To further improve encapsulation performance, R2 is perfluorodecyl, perfluoromethyl, perfluoroethyl, or perfluorohexylethylmethyl.
[0013] The hydrophilic segment (the part in brackets with a degree of polymerization of n) of the above-mentioned fluorinated amphiphilic block polymer has a molecular weight of 500 to 10,000, accounting for 5% to 95% of the total molecular weight of the polymer; the hydrophobic segment (the part in brackets with a degree of polymerization of m of the fluorinated polymer) has a molecular weight of 500 to 10,000, accounting for 5% to 95% of the total molecular weight of the polymer.
[0014] The aforementioned hydrophilic polymer blocks utilize polyethylene glycol (PEGA), the most commonly used and FDA-approved excipient. The molecular weight of PEGA in hydrophilic blocks is typically selected from 500 to 20,000. PEGA has excellent water solubility and a relatively large exclusion volume. PEGA can also inhibit the interaction between micelles and protein or cell surfaces, prolonging the half-life of drugs encapsulated by polymer micelles in vivo.
[0015] The monomers used to prepare the hydrophobic segments are at least one of perfluorodecyl acrylate, perfluoromethacrylate, perfluoroethyl acrylate, or perfluorohexylethyl methacrylate. These hydrophobic polymers exhibit good physicochemical properties and biocompatibility.
[0016] The preferred structure of the fluorinated amphiphilic block polymer is as follows:
[0017]
[0018] The preparation method of the above-mentioned fluorinated amphiphilic polymer includes the following steps:
[0019] 1) and The polymerization reaction is catalyzed by CuBr2 to obtain structure;
[0020] 2) and A polymerization reaction occurs, yielding the final product.
[0021]
[0022] In step 1), Preferred At this point, the product structure obtained in step 1) is: In step 2), Preferred The optimal product structure obtained is as follows:
[0023] In other words, a further preferred method for preparing the above-mentioned fluorinated amphiphilic polymer includes the following steps:
[0024] 1) and Polymerization reaction occurs via CuBr2 catalyst to obtain structure;
[0025] 2) Further addition A polymerization reaction occurs, yielding the final product.
[0026] The polyethylene glycol methyl ether acrylate and perfluorodecyl acrylate used in this application are both biocompatible materials.
[0027] Both steps 1) and 2) are carried out at room temperature, and the reaction is simple and controllable.
[0028] The solvent used in step 1) above is a mixture of isopropanol and trifluoroethanol in a volume ratio of 1:(0.9 to 1.1), and more preferably a volume ratio of 1:1.
[0029] The aforementioned fluorinated amphiphilic polymers are used to encapsulate hydrophobic substances and improve their water solubility.
[0030] The fluorinated amphiphilic block polymer of this application can self-assemble into spheres in aqueous solution to load drugs in the core.
[0031] The aforementioned fluorinated amphiphilic polymers are used as prodrugs for loaded antitumor drugs.
[0032] Poorly soluble antitumor drugs include doxorubicin, dihydroporphyrin, or perfluorocrown ether.
[0033] During the self-assembly process, the aforementioned fluorinated amphiphilic block polymers can encapsulate perfluorocrown ethers (PFCs) within the hydrophobic core. Furthermore, the hydrophobic monomers in the blocks of this application contain fluorine, enabling the dissolution of a larger volume of PFCs. PFCs possess advantages such as good physicochemical stability, high biocompatibility, good oxygen solubility, and low cost, and are widely used as a substitute for artificial blood. The oxygen loading in PFCs depends only on the van der Waals forces between the PFCs and oxygen molecules, and is unaffected by temperature, pH, storage, and transportation.
[0034] The method for loading antitumor drugs is as follows: a fluorinated amphiphilic polymer is dissolved in water, a poorly soluble antitumor drug is added, and the mixture is sonicated for 10-15 minutes. Undissolved substances are removed by centrifugation, and the poorly soluble antitumor drug is encapsulated in the core of the hydrophobic part of the fluorinated amphiphilic polymer.
[0035] Any techniques not mentioned in this invention are based on existing technologies.
[0036] This invention relates to a fluorinated amphiphilic polymer, prepared by controlled polymerization of hydrophilic block polyethylene glycol and hydrophobic block fluorinated acrylate. The polymer can be prepared in any molar ratio of monomers and applied to drug delivery systems. This polymer carrier exhibits amphiphilic properties, which can improve the solubility and bioavailability of poorly soluble drugs. Furthermore, the polymer has a small molecular weight, allowing self-assembled spherical molecules to easily penetrate the complex microenvironment within tumors, achieving drug release. In the later stages of drug release, the small molecules are rapidly metabolized by tissues, exhibiting good biocompatibility. The polymer preparation process is simple and efficient, with mild reaction conditions, high yield, no toxic byproducts, and uniform molecular weight distribution, making it suitable for multi-directional expansion and large-scale production. The polymer itself is non-toxic to cells; the cytotoxicity of tumor cells only becomes apparent after the loaded drug is exposed to light, facilitating targeted drug therapy. Attached Figure Description
[0037] Figure 1 NMR spectrum of the polymer intermediate of this invention;
[0038] Figure 2NMR spectra of the polymer products of this invention;
[0039] Figure 3 Molecular weight distribution diagram of the polymer of this invention;
[0040] Figure 4 Transmission electron microscope image of the polymer of this invention;
[0041] Figure 5 Particle size distribution diagram of the polymer of this invention;
[0042] Figure 6 UV concentration diagram of the polymer of this invention;
[0043] Figure 7 UV drug loading rate diagram of the polymer of this invention;
[0044] Figure 8 PFC loading diagram of the polymer of this invention;
[0045] Figure 9 Oxygen-carrying control diagram of the polymer of this invention;
[0046] Figure 10 A diagram showing the generation of reactive oxygen species in the polymer of this invention;
[0047] Figure 11 Polymer cell viability graph of this invention;
[0048] Figure 12 The cell viability diagram of polymer-loaded cells without light exposure in this invention;
[0049] Figure 13 Cell viability diagram under light irradiation with polymer-loaded drugs in this invention;
[0050] Figure 14 Diagram of the polymer acid-release drug of this invention; Detailed Implementation
[0051] To better understand the present invention, the following embodiments further illustrate the content of the present invention, but the content of the present invention is not limited to the following embodiments.
[0052] The experimental materials and instruments used in this application are well known to those skilled in the art and related fields. The implementation examples are merely exemplary experiments and do not cover or exhaust all of the inventor's experiments. Their purpose is simply to illustrate the preparation method of the polymer structure designed in the invention using practical examples.
[0053] Example 1: The preparation method of fluorinated amphiphilic block polymer is as follows:
[0054] (one) Preparation
[0055] Take a 20ml sample vial, add copper bromide (23mg, 0.1mmol), dissolve it in a mixture of 4ml isopropanol (2ml) and trifluoroethanol (2ml), then add the catalyst tris[2-(dimethylamino)ethyl]amine (20mg, 0.01mmol). Dissolve the copper bromide solid by sonication. Then add polyethylene glycol monomethyl ether acrylate (molecular weight 480, 2g, 3.8mmol) and the initiator ethyl 2-bromoisobutyrate (150mg, 0.76mmol). React at room temperature with air purged for 3 hours to obtain a gel-like liquid as a polymer intermediate. Check the reaction rate using NMR; the reaction rate is 98%. Figure 1 This is the NMR spectrum of the polymer intermediate.
[0056] (two) Preparation of .
[0057] The product from the first step was added to the hydrophobic monomer perfluorodecyl acrylate (2 g, 15 mmol) without contact with air. The reaction was continued for 20 h, then stopped. The product was dissolved in 5 L of distilled water and dialyzed (using a MW3500 dialysis bag) for 48 h, changing the water every 8 h. Dialysis was complete when the solution became colorless and transparent, yielding the target polymer PEGA. 10 -PFOEA 10 The NMR spectroscopy showed a reaction rate of 95%. Figure 2 The image shows the NMR spectrum of the product.
[0058] In step (II) above, the amount of the hydrophobic monomer perfluorodecyl acrylate was adjusted to 1g, and the rest of the steps were performed as described above to obtain PEGA. 15 -PFOEA5;
[0059] (III) Molecular weight distribution of polymers
[0060] 10 mg of the dialyzed product was dissolved in 1 ml of tetrahydrofuran, passed through a 0.22 μm molecular sieve, and then the molecular weight distribution of the polymer was examined using gel permeation chromatography. Figure 3 This is a molecular weight distribution diagram of the polymer, from... Figure 3 It can be seen that the molecular weight distribution of this amphiphilic block polymer is very uniform.
[0061] (iv) Morphology of the polymer (transmission electron microscopy image)
[0062] 1 mg of the dialysis product was dissolved in 1 ml of deionized water and sonicated. A drop was placed on the organic copper grid of a transmission electron microscope (TEM). After the water evaporated, its morphology was observed using the TEM. Figure 4 The image shows a transmission electron microscope (TEM) image of the polymer. Figure 4 It can be seen that the size of this amphiphilic block polymer is 50-100 nm, and it is spherical under microscopic conditions.
[0063] (v) Particle size distribution of polymer
[0064] 100 mg of the dialysis product was dissolved in 10 ml of deionized water and sonicated. 1 ml of the supernatant was placed in a four-sided transparent cuvette, and its particle size distribution was measured using a Malvern laser particle size analyzer. Figure 5 The particle size distribution of the polymer is determined by... Figure 5 The particle size distribution of the amphiphilic block polymer is consistent with that observed by transmission electron microscopy.
[0065] Example 2: Fluorinated amphiphilic block polymer loaded with PFCs (perfluoro-15-crown ether-5, abbreviated as PFCs)
[0066] Dissolve 30 mg of the polymer obtained in Example 1 in 3 ml of water, add 30 μL of PFCs, and sonicate (45 kW 99%) for 15 min to allow it to be loaded into the hydrophobic core of the polymer (PFCs are insoluble in water, and whether they are loaded can be directly observed macroscopically). Figure 8 As shown, when no small white droplets appear at the bottom of the solution, it indicates that the PFCs have been fully loaded. Nitrogen gas is then introduced for 15 minutes to remove pre-dissolved gases from the water, followed by oxygen for 15 minutes to ensure the material is fully loaded with oxygen. The oxygen concentration was measured using a portable dissolved oxygen meter, and the dissolved oxygen level was significantly higher than that in the control group water. Figure 9 The oxygen loading of the polymer is the same as that of other drugs loaded with it. PEGA-PFOEA is the PEGA obtained in Example 1. 10 -PFOEA 10 . Figure 10 The power is 0.5W / cm. 2 The reactive oxygen species content after 10 minutes of infrared light irradiation was compared with that of water in the control group. The drug-loaded material released under acidic tumor conditions, as shown in [reference needed]. Figure 14 .
[0067] Example 3: Fluorinated amphiphilic block polymer loaded with DOX (doxorubicin)
[0068] Take the polymer PEGA obtained in Example 1 10 -PFOEA 10 30 mg of DOX was dissolved in 3 ml of water, and 3 mg of DOX was added. The mixture was sonicated (45 kW, 99%) for 15 min to allow the drug to be loaded into the hydrophobic core of the polymer. The sonicated material was then centrifuged at 8000 rpm to remove any drug that failed to be loaded into the polymer. The drug loading rate of the material was measured using a UV spectrophotometer, and the result was 22.5%. The drug loading rate is defined as the ratio of the mass of the loaded drug to the total mass of the loaded material and the drug.
[0069] Example 4: Fluorine-containing amphiphilic block polymer loaded with Ce6 (dihydroporphyrin)
[0070] Take the polymer PEGA obtained in Example 1 10 -PFOEA 10 30 mg was dissolved in 3 ml of water, and 3 mg of Ce6 was added. The mixture was sonicated (45 kW, 99%) for 15 min to allow the drug to be loaded into the hydrophobic core of the polymer. The sonicated material was then centrifuged at 8000 rpm to remove any drug that failed to be loaded into the polymer. The drug loading rate of the material was measured using a UV spectrophotometer, and the result was 23%.
[0071] Example 5: Fluoroamophilic block polymer loaded with DOX and Ce6
[0072] Take the polymer PEGA obtained in Example 1 10 -PFOEA 10 30 mg was dissolved in 3 ml of water, and 3 mg of DOX and 3 mg of Ce6 were added. The mixture was sonicated (45 kW, 99%) for 15 min to allow the drug to be loaded into the hydrophobic core of the polymer. The sonicated material was then centrifuged at 8000 rpm to remove any drug that failed to be loaded into the polymer. The drug loading rate of the material was measured using a UV spectrophotometer. Figure 6-7 As shown, the test result was 28%; Figure 6 In the diagram, 400 is the absorption peak of Ce6, 500 is the absorption peak of DOX, ad is the standard curve, e is the curve after drug loading, and the concentrations are the concentrations of photosensitizer and doxorubicin in the standard curve. PEGA-PFOEA is the PEGA obtained in Example 1. 10 -PFOEA 10 .
[0073] In vitro experiments
[0074] The synthesized products obtained in the examples and the drug-loaded synthesized products were prepared into 10 mg / mL stock solutions using PBS (phosphate buffer), and diluted to 1000 μg / mL and 100 μg / mL immediately before use.
[0075] Experimental procedure: Primary cultured HeLa cells were taken and inoculated at 1×10⁻⁶. 6Cells were seeded at 90 μL / mL in 96-well plates and incubated in a CO2 incubator for 24 h. After cell attachment, 10 μL of PBS solution containing the extract was added to bring the final extract concentration to 100 μg / mL. After 48 h of incubation, 10 μL of MTT (3-(4,5-dimethylthiazol-2)-2,5-diphenyltetrazolium bromide) solution (5 mg / mL) was added to each well, and the plates were incubated for another 4 h. After discarding the supernatant, 100 μL of DMSO (dimethyl sulfoxide) was added to each well, and the plates were shaken. The absorbance of each well was measured at 570 nm using a microplate reader. A control group (using the same volume of PBS instead of the extract) was also set up. Cell viability was calculated as (OD value of extract group / OD value of control group) × 100%. The results are shown below. Figure 11 , 12 13. Figure 11 The material exhibits cytotoxicity alone, remaining non-toxic to cells even at very high concentrations. Figure 12 The material is non-toxic to cells after drug loading without light exposure, and is non-toxic compared to the concentration of light exposure. Figure 13 The material exhibits cytotoxicity after light exposure, showing toxicity even at very low concentrations. (From...) Figure 11 , 12 As can be seen from 13, the polymer itself is non-toxic to cells, but it becomes cytotoxic after being exposed to light and can be used for targeted drug therapy.
Claims
1. A method for preparing a fluorinated amphiphilic block polymer, characterized in that: Includes the following steps: 1) Take a 20ml sample vial, add 0.1mmol of copper bromide, dissolve it in a mixed solution of 2ml isopropanol and 2ml trifluoroethanol, then add 0.01mmol of tris[2-(dimethylamino)ethyl]amine catalyst, and sonicate to dissolve the copper bromide solid; then add 3.8mmol of polyethylene glycol monomethyl ether acrylate with a molecular weight of 480 and 0.76mmol of ethyl 2-bromoisobutyrate initiator; react at room temperature with air removed for 3h to obtain a gel-like liquid as a polymer intermediate; 2) Add 15 mmol of the hydrophobic monomer perfluorodecyl acrylate to the product from step 1) without contact with air, continue the reaction for 20 h and then stop the reaction. Dissolve the product in 5 L of distilled water and dialyze for 48 h, changing the water every 8 h. When the solution becomes colorless and transparent, the dialysis is complete and the fluorinated amphiphilic block polymer is obtained.
2. The use of a fluorinated amphiphilic polymer prepared by the method of claim 1, characterized in that, Used to encapsulate hydrophobic substances and improve their water solubility.
3. The use of a fluorinated amphiphilic polymer prepared by the method of claim 1, characterized in that, Used to load anti-tumor drugs.
4. The use as described in claim 3, characterized in that, The fluorinated amphiphilic polymer is dissolved in water, a poorly soluble antitumor drug is added, and the mixture is sonicated for 10-15 minutes. Undissolved substances are removed by centrifugation. The poorly soluble antitumor drug is encapsulated in the core of the hydrophobic part of the fluorinated amphiphilic polymer. The poorly soluble antitumor drug is doxorubicin, dihydroporphyrin, or perfluorocrown ether, or a mixture of doxorubicin and dihydroporphyrin.