Gamma-cyclodextrin polyrotaxane with macromolecular axial chain end grafted with second-generation dendritic polylysine and preparation method of gamma-cyclodextrin polyrotaxane

Abstract

The invention provides gamma-cyclodextrin polyrotaxane. The macromolecular axial chain end of the gamma-cyclodextrin polyrotaxane is terminated by macromolecular substituted second-generation dendritic polylysine. The second-generation dendritic polylysine structure does not affect recognition and combination of a gamma-cyclodextrin cavity on a macromolecular chain axis, provided steric hindrance can effectively limit falling of gamma-cyclodextrin on the macromolecular chain axis, and the yield of polyrotaxane is increased. And multiple macromolecular active groups substituted by the second-generation dendritic polylysine can also realize the whole functionalization of the gamma-cyclodextrin polyrotaxane. The yield of the obtained gamma-cyclodextrin polyrotaxane is extremely high, the coverage rate of gamma-cyclodextrin on a macromolecular axis chain is high, the product structure is clear, the preparation process is convenient and fast, the reaction condition is mild, and the gamma-cyclodextrin polyrotaxane has huge potential application value in the field of supramolecular chemical basic synthesis.

Description

Technical Field

[0001] The invention relates to the structure and preparation method of supramolecular polymer, and in particular to a gamma-cyclodextrin polyrotaxane with a high molecular axis chain end grafted with second-generation branched polylysine and a preparation method thereof. Technical Background

[0002] Cyclodextrin is a type of cyclic oligomaltose formed by glucose units connected by α-1,4-glycosidic bonds. Common α-, β- and γ-cyclodextrins are composed of 6, 7 and 8 glucose units respectively, and their cyclic cavity radius increases successively. Cyclodextrin polyrotaxane is a structurally stable supramolecular polymer formed by introducing large-sized "capping" molecules at both ends of the chain to restrict the detachment of cyclodextrin after multiple cyclodextrin small molecules pass through the polymer chain under the action of hydrophobic force. In the synthesis process of polyrotaxane, the introduction of large-sized capping groups at both ends of the chain and the dissociation of cyclodextrin from the chain are simultaneous and competitive processes. Because the cavity radius of α-, β-, and γ-cyclodextrin increases successively, even if the polyrotaxane is prepared under the same reaction conditions, the probability of them falling off the polymer axis also increases successively, so the synthesis of α-cyclodextrin polyrotaxane is the easiest, while the synthesis of γ-cyclodextrin polyrotaxane is the most difficult.

[0003] Among the reported methods for synthesizing cyclodextrin polyrotaxanes, the methods suitable for synthesizing α-cyclodextrin polyrotaxanes are the most numerous and have the widest application range. The obtained α-cyclodextrin polyrotaxanes not only have a high yield, but also have the highest coverage of α-cyclodextrin on the polymer axis. Increasing the yield of cyclodextrin polyrotaxanes in actual production activities means improving work efficiency and saving production resources, while increasing the coverage of cyclodextrin on the polymer axis is also of practical significance. The more cyclodextrins in a single polyrotaxane macromolecule, the more effective reaction sites are provided. Therefore, improving the yield of cyclodextrin polyrotaxanes and increasing the coverage of cyclodextrin on the axis are the focus and goals of exploring new ways to prepare cyclodextrin polyrotaxanes.

[0004] There are few methods reported so far for synthesizing γ-cyclodextrin polyrotaxanes using single-chain polymer axes. Kato, K et al. (Organic-Inorganic Hybrid Slide-Ring Gels: Polyrotaxanes Consisting of Poly(dimethylsiloxane)andγ-Cyclodextrin and Subsequent Topological Cross-Linking. Macromolecules 42(18):7129-7136.) disclosed a method for synthesizing γ-cyclodextrin polyrotaxanes, but the method has a long reaction time, low yield, and low coverage of γ-cyclodextrin. Wang, Y.-C et al. (Synthesis of poly(methyl methacrylate)-based polyrotaxane via reversible addition–fragmentation chain transfer polymerization. ACS Macro Letters 9(12): 1853-1857.) and Li, J et al. (Biobased thermosensitive polyrotaxanes constructed by polymerization of cyclodextrin-triterpenoid inclusion complexes. Polymer Chemistry.) improved the yield of γ-cyclodextrin polyrotaxane by changing the components of the polymer axis during the synthesis process to enhance the recognition between γ-cyclodextrin and the chain segments, but the synthesis process is demanding and difficult. Therefore, the method for synthesizing γ-cyclodextrin polyrotaxane is in urgent need of innovation, with the goal of achieving high yield and high cyclodextrin coverage, and at the same time, it is necessary to keep the operation as simple as possible and the reaction conditions mild so as to facilitate the promotion of the synthesis method. Summary of the invention

[0005] The present invention provides a γ-cyclodextrin precise polyrotaxane and a second-generation branched polylysine-terminated γ-cyclodextrin precise polyrotaxane whose polymer axis chain end is substituted by a macromolecule, which is further prepared from the γ-cyclodextrin precise polyrotaxane. The yield of the γ-cyclodextrin polyrotaxane obtained by the present invention is extremely high, and the coverage rate of γ-cyclodextrin on the polymer axis chain is high.

[0006] The specific technical solutions of the present invention are as follows:

[0007] A γ-cyclodextrin precise polyrotaxane having a cavity of γ-cyclodextrin and a polymer axis chain, wherein one or both ends of the polymer axis chain passing through the cavity of γ-cyclodextrin have a group: Wherein R1 to R4 are the same or different and represent a reactive small molecule group selected from H, carboxyl, hydroxyl, thiol, aldehyde, succinimide, azide, alkynyl or maleimide.

[0008] The polymer axis chain is selected from polypropylene glycol, polyisobutylene, polymethyl vinyl ether, polytetrahydrofuran, polypropylene oxide, polyethylene succinate, polybutylene succinate, polycaprolactone, polyethylene glycol block polypropylene glycol or polyethylene glycol block polycaprolactone. The number of repeating units of the polymer axis chain is an integer of 20 to 400, and most preferably an integer of 33 to 165.

[0009] The structure of the above-mentioned γ-cyclodextrin precise polyrotaxane is shown as follows:

[0010] The grafting of the second-generation branched polylysine on the polymer axis chain end of the γ-cyclodextrin precision polyrotaxane does not affect the recognition and binding of the γ-cyclodextrin cavity to the polymer axis, but the resulting steric hindrance can serve as a "buffer" during the detachment of γ-cyclodextrin from the axis, which can effectively hinder the detachment of γ-cyclodextrin and improve the yield. At the same time, the second-generation branched polylysine molecules grafted on both ends of the chain have 8 amine reaction sites. After connecting the same or different active groups, these active groups can be used to further achieve the purpose of functionalization of γ-cyclodextrin polyrotaxane.

[0011] Another object of the present invention is to provide a γ-cyclodextrin polyrotaxane having a cavity of γ-cyclodextrin and a polymer axis chain, which is formed by reacting the γ-cyclodextrin precise polyrotaxane of the present invention with a capping agent, wherein the capping agent is a macromolecular compound having an active group that can react with R1 to R4.

[0012] The active group capable of reacting with R1 to R4 and R1 to R4 are the following mutually reactive group pairs: alkynyl-azide group, amino-carboxyl group, hydroxyl-carboxyl group, amino-succinimide group, hydroxyl-succinimide group, acyl halide group-amino group, acyl halide group-hydroxyl group, thiol-alkenyl group, thiol-alkynyl group, thiol-maleimide group, thiol-carboxyl group, furanyl-maleimide group, anthracenyl-maleimide group, aldehyde group-hydroxyl group, aldehyde group-amine group, isocyanate group-amine group, isocyanate group-hydroxyl group, isothiocyanate group-amine group, isothiocyanate group-hydroxyl group.

[0013] The macromolecular compound is selected from water-soluble high molecular polymers (such as polyacrylic acid, polyethylene glycol, polyethylene oxide, polyvinyl alcohol, polyacrylamide, polyvinyl pyrrolidone, polymaleic anhydride, polybetaine), fluorophores (such as fluorescein, rhodamine, coumarin, anthracene, pyrene), sugars (cyclodextrin, mannose, fructose, sucrose), polypeptides, proteins, and nucleic acids.

[0014] The reaction is a "click chemistry" reaction, including a CuAAC reaction, a thiol-ene reaction, a thiol-yne reaction, a Diels-Alder reaction, an aldol condensation reaction, an esterification reaction or an amidation reaction.

[0015] In a preferred technical solution of the present invention, R1 to R4 are selected from H, alkynyl, maleimide, and thiol, and the end-capping agent is selected from succinimidyl polyethylene glycol, azidocyclodextrin, anthracenol, carboxycoumarin, and allylcyclodextrin.

[0016] Another object of the present invention is to provide a γ-cyclodextrin polyrotaxane molecular tube, which is formed by cross-linking the γ-cyclodextrin polyrotaxane described in the present invention and can be prepared by a method disclosed in the prior art.

[0017] The γ-cyclodextrin polyrotaxane of the present invention can be prepared by the following method, comprising the following steps:

[0018] (1) Grafting primary lysine at the end of a polymer axis chain: (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid, 1-hydroxybenzotriazole and 1-ethyl-(3-dimethylaminopropyl)carbodiimide are added to an organic solution of a polymer axis chain having an amine terminal (polymer axis chain such as polypropylene glycol, polyisobutylene, polymethyl vinyl ether, polytetrahydrofuran, polypropylene oxide, polyethylene succinate, polybutylene succinate, polycaprolactone, polyethylene glycol-block-polypropylene glycol or polyethylene glycol-block-polycaprolactone. Organic solvent such as dichloromethane or DMF) to obtain a polymer chain having a primary lysine molecule grafted at the end;

[0019] (2) Grafting the second-generation branched polylysine on the polymer axis chain: Add trifluoroacetic acid to the dichloromethane solution of the polymer axis chain grafted with the first-generation lysine molecule obtained in step (1) (polymer axis chain such as polypropylene glycol, polyisobutylene, polymethyl vinyl ether, polytetrahydrofuran, polypropylene oxide, polyethylene succinate, polybutylene succinate, polycaprolactone, polyethylene glycol-block polypropylene glycol or polyethylene glycol-block polycaprolactone), stir, remove the solvent, add (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid, 1-hydroxybenzotriazole and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to obtain a polymer axis chain grafted with the second-generation branched polylysine;

[0020] (3) Introducing active small molecule groups into the polymer chain axis: adding trifluoroacetic acid to a dichloromethane solution of the product obtained in (2) (e.g., polypropylene glycol, polyisobutylene, polymethyl vinyl ether, polytetrahydrofuran, polypropylene oxide, polyethylene succinate, polybutylene succinate, polycaprolactone, polyethylene glycol-block polypropylene glycol or polyethylene glycol-block polycaprolactone), stirring, removing the solvent, and adding a compound having R1-R4 groups and capable of reacting with the amino group in lysine (e.g., 3-maleimidopropionic acid, propiolic acid or mercaptopropionic acid) to obtain a polymer axis chain of terminally grafted second-generation branched polylysine substituted with R1-R4;

[0021] (4) Synthesis of polyrotaxanes by using a polymer axis chain end-grafted with second-generation branched polylysine and γ-cyclodextrin: Add the polymer axis chain end-grafted with second-generation branched polylysine substituted with R1 to R4 obtained in step (3) to a saturated solution of γ-cyclodextrin, and stir to form a suspension of γ-cyclodextrin precise polyrotaxane; add a capping agent to the suspension and stir to obtain a γ-cyclodextrin polyrotaxane.

[0022] A preferred example of the above preparation method is as follows:

[0023] (1) Add 2-4 equivalents of (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid, 2-4 equivalents of 1-hydroxybenzotriazole and 2-4 equivalents of 1-ethyl-(3-dimethylaminopropyl)carbodiimide to a dichloromethane solution of 1 equivalent of an amine-terminated polypropylene glycol axis chain in a reactor, and stir for 12-48 hours. The solution is washed with deionized water, concentrated, and separated from impurities using a silica gel column to obtain a polymer axis chain with a grafted first-generation lysine molecule.

[0024] (2) Add 5 equivalents of trifluoroacetic acid to the dichloromethane solution of the product obtained in (1), stir for 2 hours, and use an oil pump to reduce pressure and spin dry the solvent. Add 2-4 equivalents of (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid, 2-4 equivalents of 1-hydroxybenzotriazole and 2-4 equivalents of 1-ethyl-(3-dimethylaminopropyl)carbodiimide to the dichloromethane solution of 1 equivalent of the above product, and stir for 12-48 hours. The solution is washed with deionized water, concentrated, and separated from impurities with a silica gel column to obtain a polymer axis chain of end-grafted second-generation branched polylysine.

[0025] (3) Add 5 equivalents of trifluoroacetic acid to the dichloromethane solution of the product obtained in (2), stir for 2 hours, and use an oil pump to decompress and spin-dry the solvent. Add 1-4 equivalents of 3-maleimidopropionic acid to the DMF solution of the product, and stir for 12-48 hours. After the solution is concentrated, impurities are separated using a silica gel column to obtain a polymer axis chain of terminal grafted second-generation branched polylysine substituted with maleimide.

[0026] (4) Add 1 equivalent of the product obtained in (3) to a saturated solution of γ-cyclodextrin in a reactor, and stir for 48 hours to form a γ-cyclodextrin precise polyrotaxane suspension; add 8-10 equivalents of anthracene alcohol capping agent to the suspension, and stir for 12-72 hours. After centrifugation, dissolution, precipitation, repeated washing, and drying, γ-cyclodextrin polyrotaxane is obtained.

[0027] Another object of the present invention is to provide the use of the γ-cyclodextrin precise polyrotaxane or γ-cyclodextrin polyrotaxane or γ-cyclodextrin polyrotaxane molecular tube described in the present invention in the preparation of drug delivery vectors, gene delivery vectors, diagnostic and treatment system vectors, elastomer cross-linked materials, molecular device materials, molecular transmission materials, molecular recognition materials, biomembrane transmission simulation materials or supramolecular chemical materials.

[0028] Advantages of the present invention:

[0029] (1) The present invention grafts second-generation branched polylysine on the end of the polymer axis chain, providing a large steric hindrance as a "buffer" for γ-cyclodextrin to fall off from the polymer axis chain, effectively hindering the shedding of γ-cyclodextrin, and greatly improving the yield of γ-cyclodextrin polyrotaxane and the coverage of γ-cyclodextrin on the polymer chain axis.

[0030] (2) The product structure of each step of the present invention is clearly characterized. Compared with the reported method of capping γ-cyclodextrin polyrotaxane by polymerization reaction at the chain end, the present invention has controllability and stronger operability.

[0031] (3) The present invention utilizes the multiple reaction sites exposed by the polylysine at the end of the polymer axis chain to introduce a variety of active small molecule groups, which are used to react with the capping agent to cap the γ-cyclodextrin polyrotaxane, thereby achieving functional modification of the polyrotaxane, which has flexibility and applicability.

[0032] (4) The present invention uses relatively low-priced lysine monomer as a reaction reagent, the cost of the synthesis process is low, and it is easy to promote, paving the way for further broadening the application scope of γ-cyclodextrin polyrotaxane. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 The γ-cyclodextrin polyrotaxane with the first generation of lysine terminated in the polymer axis chain was prepared in DMSO-d6 1 The H NMR graph shows the characteristic peaks of γ-cyclodextrin, polypropylene glycol grafted with primary lysine, and end-capping groups. According to NMR integral calculation, there are 4 γ-cyclodextrin molecules on each polypropylene glycol (repeating unit number is 34) axis chain, and the coverage rate of γ-cyclodextrin is 22%.

[0034] Figure 2 The γ-cyclodextrin polyrotaxane grafted with second-generation branched polylysine at the end of the polymer axis chain in DMSO-d6 1The HNMR graph shows the characteristic peaks of γ-cyclodextrin, polypropylene glycol grafted with second-generation branched polylysine, and end-capping groups. According to the NMR integral calculation, each polypropylene glycol (repeating unit number is 34) axis chain has 9 γ-cyclodextrin molecules, and the coverage of γ-cyclodextrin is 53%.

[0035] Figure 3 The 2DNOESY image of γ-cyclodextrin polyrotaxane grafted with second-generation branched polylysine at the ends of the polymer axis chain in DMSO-d6 shows strong cross signals between C(3)H (3.65ppm) and C(5)H (3.55ppm) on γ-cyclodextrin and methyl hydrogen (1.04ppm) in polypropylene glycol, indicating that γ-cyclodextrin and polypropylene glycol with second-generation branched polylysine introduced at both ends self-assembled to form an inclusion complex through hydrophobic force, and this inclusion complex was stably retained by the end-capping reaction.

[0036] Figure 4 The 2DDOSY diagram of γ-cyclodextrin polyrotaxane with second-generation branched polylysine grafted on the polymer axis chain in DMSO-d6 shows that γ-cyclodextrin, the polypropylene glycol axis chain grafted with second-generation branched polylysine and the end-capping groups at both ends show the same diffusion coefficient, indicating that γ-cyclodextrin passes through the polypropylene glycol axis chain and is fixed on the axis by the end-capping groups at both ends. The three form a stable whole, proving the successful synthesis of γ-cyclodextrin polyrotaxane.

[0037] Figure 5 The γ-cyclodextrin polyrotaxane with second-generation branched polylysine grafted to the end of the polymer chain and fluorescent functional groups introduced by end-capping agents was synthesized in DMSO-d6 1 The H NMR graph shows the characteristic peaks of γ-cyclodextrin, polypropylene glycol grafted with second-generation dendritic polylysine, and fluorescent groups at both ends. DETAILED DESCRIPTION

[0038] The present invention will be further described below in conjunction with embodiments, but the scope of the present invention is not limited to the embodiments.

[0039] Example 1: Preparation of γ-cyclodextrin polyrotaxane with primary lysine grafted to the polymer chain end

[0040] Step ①: Grafting the first generation of lysine on the polymer axis chain: At room temperature, 0.33g (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid (0.96mmol) was dissolved in 20mL of anhydrous dichloromethane, and 0.13g 1-hydroxybenzotriazole (0.96mmol) and 0.18g 1-ethyl-(3-dimethylaminopropyl)carbodiimide (0.96mmol) were added thereto, and the activation reaction was carried out for 1h. Subsequently, 0.8g of amino-terminated polypropylene glycol polymer chain (0.4mmol, molecular weight of 2000g / mol, number of repeating units of 34) dissolved in 5mL of anhydrous dichloromethane was added thereto, and the reaction was stirred for 20 hours. After the reaction was completed, it was washed with deionized water, concentrated, and separated from impurities with a silica gel column to obtain a polypropylene glycol polymer chain grafted with the first generation of lysine molecules, with a yield of 92%.

[0041] Step ②: Introducing active small molecule groups into the polymer chain axis: Dissolve the product in step ① with dichloromethane, add 1 mL of trifluoroacetic acid, stir and react for 2 hours, then use an oil pump to decompress and spin dry the solvent. Dissolve again with an organic solvent, add 0.27 g of 3-maleimidopropionic acid (1.6 mmol), 0.22 g of 1-hydroxybenzotriazole (1.6 mmol) and 0.31 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (1.6 mmol), and stir and react for 24 hours. After the reaction is completed, wash with deionized water, concentrate and separate impurities with a silica gel column to obtain a polypropylene glycol polymer chain with a first-generation lysine molecule grafted at both ends of the chain and a total of 4 maleimide groups exposed at the chain end, with a yield of 88%.

[0042] Step ③: Synthesis of polyrotaxane from polymer chain axis with grafted lysine and γ-cyclodextrin: At room temperature, 2.20g of γ-cyclodextrin (1.70mmol) was dissolved in water, and 200μL of DMF solution containing 143mg of polypropylene glycol chain with grafted lysine (0.05mmol, molecular weight of 2859g / mol, number of repeating units of 34, 0.2mmol of maleimide connected to the periphery of lysine) was added thereto. The reaction system was stirred for 48 hours to form a pseudo-polyrotaxane suspension. 333.2mg of 9-anthracene alcohol (1.6mmol) was added thereto, and the reaction was stirred at 40°C for 60 hours. The precipitate was collected by centrifugation, dissolved with DMSO, precipitated with water and methanol in turn, washed repeatedly, and dried to obtain a white powder, i.e., γ-cyclodextrin polyrotaxane, with a yield of 4%.

[0043] In this example, a polymer chain axis with terminal grafted lysine was used to synthesize a polyrotaxane with γ-cyclodextrin. The results showed that the yield of the γ-cyclodextrin polyrotaxane was low. Figure 1 The product of this example is in DMSO-d6 1H NMR graph. By calculating the characteristic peak integration, it was found that the coverage of γ-cyclodextrin on the axis in this polyrotaxane was 22%. The above results show that the steric hindrance provided by the grafted lysine at the end of the polymer chain is limited and is not enough to effectively prevent γ-cyclodextrin from falling off the axis, resulting in an unsatisfactory synthesis effect. The final yield of γ-cyclodextrin polyrotaxane is low and the coverage of γ-cyclodextrin on the axis is low.

[0044] Example 2: Preparation of γ-cyclodextrin polyrotaxane with three-generation branched lysine grafted on the polymer axis chain

[0045] Step ①: Grafting primary lysine onto the polymer axis chain end: The same reaction steps as step ① in Example 1 are adopted.

[0046] Step ②: Grafting the second-generation branched polylysine on the polymer axis chain: Dissolve the product in step ① with dichloromethane, add 1mL of trifluoroacetic acid, stir and react for 2 hours, and then use an oil pump to reduce pressure and spin dry the solvent. Dissolve it again with an organic solvent, add 0.55g (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid (1.6mmol), 0.22g 1-hydroxybenzotriazole (1.6mmol) and 0.31g 1-ethyl-(3-dimethylaminopropyl) carbodiimide (1.6mmol), and stir and react for 24 hours. After the reaction is completed, wash with deionized water, concentrate and separate impurities with a silica gel column to obtain a polymer chain grafted with a second-generation branched polylysine molecule, with a yield of 90%.

[0047] Step ③: Grafting three-generation branched polylysine on the polymer axis chain: Dissolve the product in step ② with dichloromethane, add 1mL trifluoroacetic acid, stir and react for 2 hours, and then use an oil pump to reduce pressure and spin dry the solvent. Dissolve it again with an organic solvent, add 1.11g (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid (3.2mmol), 0.43g 1-hydroxybenzotriazole (3.2mmol) and 0.61g 1-ethyl-(3-dimethylaminopropyl) carbodiimide (3.2mmol), and stir and react for 24 hours. After the reaction is completed, wash with deionized water, concentrate and separate impurities with a silica gel column to obtain a polymer chain grafted with three-generation branched polylysine molecules, with a yield of 86%.

[0048] Step ④: Introducing active small molecule groups into the polymer chain axis: Dissolve the product in step ③ with dichloromethane, add 1mL trifluoroacetic acid, stir and react for 2 hours, then use an oil pump to decompress and spin dry the solvent. Dissolve again with an organic solvent, add 1.08g 3-maleimidopropionic acid (6.4mmol), 0.86g 1-hydroxybenzotriazole (6.4mmol) and 1.22g 1-ethyl-(3-dimethylaminopropyl)carbodiimide (6.4mmol), stir and react for 24 hours. After the reaction is completed, wash with deionized water, concentrate and separate impurities with a silica gel column to obtain a polypropylene glycol polymer chain with three-generation polylysine molecules grafted at both ends of the chain and a total of 16 maleimide groups exposed at the chain end, with a yield of 80%.

[0049] Step ⑤: Synthesis of polyrotaxane from polymer chain axis of end-grafted three-generation branched polylysine and γ-cyclodextrin: At room temperature, 2.20 g of γ-cyclodextrin (1.70 mmol) was dissolved in water, and 200 μL of DMF solution containing 310 mg of polypropylene glycol chain (0.05 mmol, molecular weight 6211 g / mol, repeating unit 34, 0.8 mmol maleimide connected to the periphery of lysine) was added thereto. The reaction system was stirred for 48 hours, and the results showed that no pseudopolyrotaxane suspension could be formed.

[0050] In this embodiment, a polymer chain axis grafted with three-generation branched polylysine was used to synthesize a polyrotaxane with γ-cyclodextrin, thereby increasing the number of grafted generations of lysine. The results showed that the size of the three-generation polylysine was too large for the cavity of γ-cyclodextrin, and the three-generation branched polylysine grafted at the polymer chain end seriously hindered γ-cyclodextrin from passing through the polymer chain axis, resulting in the inability to form γ-cyclodextrin precise polyrotaxane, and thus the inability to obtain γ-cyclodextrin polyrotaxane.

[0051] Example 3: Preparation of γ-cyclodextrin polyrotaxane with second-generation branched polylysine grafted to the polymer axis chain

[0052] Step ①: Grafting primary lysine onto the polymer axis chain end: The same reaction steps as step ① in Example 1 are adopted.

[0053] Step ②: Grafting the second-generation branched polylysine onto the polymer axis chain end: The same reaction steps as step ② in Example 2 are adopted.

[0054] Step ③: Introducing active small molecule groups into the polymer chain axis: Dissolve the product in step ② with dichloromethane, add 1mL trifluoroacetic acid, stir and react for 2 hours, then use an oil pump to decompress and spin dry the solvent. Dissolve again with an organic solvent, add 0.54g 3-maleimidopropionic acid (3.2mmol), 0.43g 1-hydroxybenzotriazole (3.2mmol) and 0.61g 1-ethyl-(3-dimethylaminopropyl)carbodiimide (3.2mmol), stir and react for 24 hours. After the reaction is completed, wash with deionized water, concentrate and separate impurities with a silica gel column to obtain a polypropylene glycol polymer chain with two-generation branched polylysine molecules grafted at both ends of the chain and a total of 8 maleimide groups exposed at the chain end, with a yield of 88%.

[0055] Step ④: Synthesis of polyrotaxane from polymer chain axis of end-grafted second-generation branched polylysine and γ-cyclodextrin: At room temperature, 2.20g of γ-cyclodextrin (1.70mmol) was dissolved in water, and 200μL of DMF solution containing 199mg of polypropylene glycol chain (0.05mmol, molecular weight of 3976g / mol, number of repeating units of 34, free amino group of lysine connected with 0.4mmol maleimide group) was added thereto. The reaction system was stirred for 48 hours to form a pseudo-polyrotaxane suspension. 666.4mg of 9-anthracene alcohol (3.2mmol) was added thereto, and the reaction was stirred at 40℃ for 60 hours. The precipitate was collected by centrifugation, dissolved with DMSO, precipitated with water and methanol in turn, washed repeatedly, and dried to obtain a white powder, i.e., γ-cyclodextrin polyrotaxane, with a yield of 56%.

[0056] Figure 2 In this example, the γ-cyclodextrin polyrotaxane synthesized by grafting the second-generation branched polylysine at the end of the polymer axis chain in DMSO-d6 1 The H NMR graph shows the characteristic peaks of γ-cyclodextrin, polypropylene glycol grafted with second-generation branched polylysine, and end-capping groups. According to NMR integral calculation, each polypropylene glycol (repeating unit number is 34) axis chain has 9 γ-cyclodextrin molecules, and the coverage of γ-cyclodextrin is 53%.

[0057] Figure 3 This is the 2D NOESY image of the γ-cyclodextrin polyrotaxane synthesized by grafting the second-generation branched polylysine at the end of the polymer axis chain in this example in DMSO-d6. It can be observed that C(3)H (3.65ppm) and C(5)H (3.55ppm) on γ-cyclodextrin and methyl hydrogen (1.04ppm) in polypropylene glycol show strong cross signals, indicating that γ-cyclodextrin and polypropylene glycol with second-generation branched polylysine introduced at both ends self-assemble to form an inclusion complex through hydrophobic force, and this inclusion complex is stably retained by the end-capping reaction.

[0058] Figure 4 This is the 2D DOSY image of the γ-cyclodextrin polyrotaxane prepared by grafting the second-generation branched polylysine on the polymer axis chain in this example in DMSO-d6. It can be observed that the γ-cyclodextrin, the polypropylene glycol axis chain grafted with the second-generation branched polylysine and the end-capping groups at both ends show the same diffusion coefficient, indicating that the γ-cyclodextrin passes through the polypropylene glycol axis chain and is fixed on the axis by the end-capping groups at both ends. The three form a stable whole, proving the successful synthesis of the γ-cyclodextrin polyrotaxane.

[0059] The above structural analysis test fully proves that the target γ-cyclodextrin polyrotaxane is obtained with a yield of 56% and a coverage of γ-cyclodextrin on the axis of 53%. Compared with the existing technology, both the yield and coverage are greatly improved.

[0060] Example 4: Preparation of polymer axis chain end grafted with second-generation lysine and introduction of fluorescent functionalized γ-cyclodextrin polyrotaxane Step ①: Grafting first-generation lysine on polymer axis chain end: Take the same reaction steps as step ① in Example 1.

[0061] Step ②: Grafting the second-generation branched polylysine onto the polymer axis chain end: The same reaction steps as step ② in Example 2 are adopted.

[0062] Step ③: Introducing active small molecule groups into the polymer chain axis: adopt the same reaction steps as step ③ in Example 3.

[0063] Step ④: Synthesis of capping agent with fluorescent group:

[0064] 208 mg of 9-anthryl alcohol (1 mmol) was dissolved in 5 mL of anhydrous tetrahydrofuran, and 400 mg of sodium hydride was added thereto in an ice bath. After the reaction was smooth, the mixture was transferred to room temperature and the reaction was continued for 1 hour. Subsequently, 1.05 g (4 mmol) of 1,4-di(bromomethyl)benzene dissolved in 10 mL of anhydrous tetrahydrofuran was added thereto, and the mixture was transferred to an oil bath and refluxed at 70°C for overnight reaction. After the reaction was completed, a large amount of ice water was poured in to quench the mixture, and the mixture was extracted with dichloromethane and separated by a silica gel column to obtain the product. 313 mg of the above product (0.8 mmol) was further dissolved in 5 mL of anhydrous DMF, and 130 mg of sodium azide (2 mmol) was added, and the mixture was refluxed at 60°C for overnight reaction. After the reaction was completed, deionized water was added, the mixture was extracted with ethyl acetate, and the solvent was removed by distillation under reduced pressure to obtain the product. 524 mg of 8-hydroxy-1,3,6-pyrene trisulfonic acid trisodium salt (1 mmol) and 48 mg of sodium hydroxide (1.2 mmol) were dissolved in 20 mL of DMSO solution, 143 mg of propargyl bromide (1.2 mmol) was added, and the mixture was refluxed at 60°C overnight. After the reaction was completed, the product was precipitated with isopropanol and washed several times to obtain the product. Take 85 mg of product (0.24mmol), 112mg product (0.2mmol) was dissolved in a mixed solution of 3.2mL DMF and 0.8mL deionized water. 34.7mg pentamethyldiethylenetriamine (0.2mmol, PMDETA) and 10mg copper sulfate pentahydrate (0.04mmol) were added thereto. The reaction system was then protected by argon, 106mg sodium ascorbate (0.6mmol) was added, and the reaction was carried out at 60°C for 48 hours. After the reaction was completed, it was precipitated with isopropanol and washed several times to obtain a capping agent.

[0065] Step ⑤: Synthesis of polyrotaxane with fluorescent functionalization by end-grafting the polymer chain axis of second-generation branched polylysine with γ-cyclodextrin: At room temperature, 2.20g of γ-cyclodextrin (1.70mmol) was dissolved in water, and 200μL of DMF solution containing 199mg of polypropylene glycol chain (0.05mmol, molecular weight of 3976g / mol, number of repeating units of 34, and 0.4mmol maleimide group connected to the periphery of the free amino group of lysine) with end-grafted second-generation branched polylysine was added thereto. The reaction system was stirred for 48 hours to form a pseudo-polyrotaxane suspension. 2930mg of end-capping agent was added thereto. (3.2mmol), stirred at 40℃ for 60 hours, centrifuged to collect the precipitate, dissolved in DMSO, precipitated with water and methanol, washed repeatedly, and dried to obtain a white powder, namely γ-cyclodextrin polyrotaxane.

[0066] Figure 5 This is the 1H NMR graph of the γ-cyclodextrin polyrotaxane synthesized in this example in DMSO-d6, which shows the characteristic peaks of γ-cyclodextrin, polypropylene glycol grafted with second-generation branched polylysine, and fluorescent groups at both ends, indicating that the fluorescent function was introduced while the γ-cyclodextrin polyrotaxane molecule was successfully synthesized.

Claims

1. A γ-cyclodextrin precise polyrotaxane having a cavity of γ-cyclodextrin and a polymer axis chain, characterized in that One or both ends of the polymer axis chain passing through the cavity of γ-cyclodextrin have groups: Wherein R1 to R4 are the same or different and are selected from H, carboxyl, hydroxyl, thiol, aldehyde, succinimidyl, azido, alkynyl or maleimide.

2. The γ-cyclodextrin precise polyrotaxane according to claim 1, characterized in that The polymer axis chain is selected from polypropylene glycol, polyisobutylene, polymethyl vinyl ether, polytetrahydrofuran, polypropylene oxide, polyethylene succinate, polybutylene succinate, polycaprolactone, polyethylene glycol block polypropylene glycol or polyethylene glycol block polycaprolactone.

3. A γ-cyclodextrin polyrotaxane having a γ-cyclodextrin cavity and a polymer axis chain, characterized in that The γ-cyclodextrin precise polyrotaxane according to claim 1 or 2 is reacted with a capping agent, wherein the capping agent is a macromolecular compound having an active group capable of reacting with R1 to R4.

4. The γ-cyclodextrin polyrotaxane according to claim 3, characterized in that The active group capable of reacting with R1 to R4 and R1 to R4 are the following mutually reactive group pairs: alkynyl-azide group, amino-carboxyl group, hydroxyl-carboxyl group, amino-succinimide group, hydroxyl-succinimide group, acyl halide group-amino group, acyl halide group-hydroxyl group, thiol-alkenyl group, thiol-alkynyl group, thiol-maleimide group, thiol-carboxyl group, furanyl-maleimide group, anthracenyl-maleimide group, aldehyde group-hydroxyl group, aldehyde group-amine group, isocyanate group-amine group, isocyanate group-hydroxyl group, isothiocyanate group-amine group, isothiocyanate group-hydroxyl group, and the macromolecular compound is selected from water-soluble high molecular polymers, fluorophores, cyclodextrins, carbohydrates, polypeptides, proteins, and nucleic acids.

5. The γ-cyclodextrin polyrotaxane according to claim 4, characterized in that The water-soluble high molecular polymer is selected from polyacrylic acid, polyethylene glycol, polyethylene oxide, polyvinyl alcohol, polyacrylamide, polyvinyl pyrrolidone, polymaleic anhydride or polybetaine, the fluorophore is selected from fluorescein, rhodamine, coumarin or pyrene, and the sugar is selected from cyclodextrin, mannose, fructose or sucrose.

6. The γ-cyclodextrin polyrotaxane according to claim 3, characterized in that The reaction is a "click chemistry" reaction.

7. A γ-cyclodextrin polyrotaxane molecular tube, characterized in that The γ-cyclodextrin polyrotaxane according to any one of claims 3 to 6 is cross-linked.

8. The method for preparing a γ-cyclodextrin polyrotaxane according to any one of claims 3 to 6, characterized in that Take the following steps: (1) Grafting first-generation lysine on the polymer chain end: adding (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid, 1-hydroxybenzotriazole and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to an organic solution of a polymer chain end with an amine group, to obtain a polymer chain end-grafted with first-generation lysine molecules; (2) Grafting the second-generation branched polylysine on the polymer axis chain: adding trifluoroacetic acid to the organic solution of the polymer chain with the first-generation lysine molecules grafted on the end obtained in step (1), stirring, removing the solvent, adding (S)-2,6-di-tert-butyloxycarbonylaminocaproic acid, 1-hydroxybenzotriazole and 1-ethyl-(3-dimethylaminopropyl)carbodiimide to obtain a polymer axis chain with the second-generation branched polylysine grafted on the end; (3) Introducing active small molecular groups into the polymer chain axis: adding trifluoroacetic acid to the organic solution of the product obtained in (2), stirring, removing the solvent, and adding a compound having R1-R4 groups that can react with the amino group in lysine to obtain a polymer axis chain of terminal grafted second-generation branched polylysine substituted with R1-R4; (4) Synthesis of polyrotaxane by using a polymer axis chain end-grafted with second-generation branched polylysine and γ-cyclodextrin: Add the polymer axis chain end-grafted with second-generation branched polylysine substituted with R1 to R4 obtained in step (3) to a saturated solution of γ-cyclodextrin, and stir to form a suspension of γ-cyclodextrin precise polyrotaxane; add a capping agent to the suspension and stir to obtain a γ-cyclodextrin polyrotaxane.

9. The method according to claim 8, characterized in that In step (3), the compound having R1 to R4 groups and capable of reacting with the amino group in lysine is selected from 3-maleimidopropionic acid, propiolic acid or mercaptopropionic acid.

10. Use of the γ-cyclodextrin precise polyrotaxane described in claim 1 or 2, or the γ-cyclodextrin polyrotaxane described in any one of claims 3 to 6, or the γ-cyclodextrin polyrotaxane molecular tube described in claim 7 in the preparation of drug delivery vectors, gene delivery vectors, diagnostic and therapeutic system vectors, elastomer cross-linked materials, molecular device materials, molecular transmission materials, molecular recognition materials, biomembrane transmission simulation materials or supramolecular chemical materials.

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