A rare earth supramolecular fluorescent aggregate and its preparation method and application

Abstract

The present invention discloses a rare earth supramolecular fluorescent aggregate, a preparation method, and an application thereof, belonging to the field of analytical chemistry technology. The preparation method of the rare earth supramolecular fluorescent aggregate comprises the following steps: mixing a rare earth salt solution and a 2,6-pyridinedicarboxylic acid solution to conduct a coordination reaction to obtain a mixed solution A; mixing a polyoxyethylene polyoxypropylene ether triblock polymer solution and a carboxymethyl β-cyclodextrin solution, and then conducting a host-guest inclusion complex reaction to obtain a mixed solution B; and mixing mixed solutions A and B to cause coordination and supramolecular aggregation reactions to obtain a rare earth supramolecular fluorescent aggregate. The preparation method of the present invention is simple and easy, and the prepared supramolecular fluorescent aggregate has excellent fluorescence properties and stimulus response characteristics.

Description

Technical Field

[0001] The present invention relates to the technical field of analytical chemistry, and more particularly to a rare earth supramolecular fluorescent aggregate, a preparation method and an application thereof. Background Art

[0002] Since Lehn proposed the concept of supramolecular chemistry in 1995, supramolecular aggregates have had a profound impact on various branches of chemistry, especially in materials science. These supramolecular aggregates, self-assembled through non-covalent interactions, have endowed target materials with numerous remarkable properties, such as shape memory, adaptability, self-healing, encryption, and recyclability.

[0003] Cyclodextrins, or CDs, are cyclic oligosaccharides composed of multiple glucose molecules with a unique cavity structure. Their inner cavity is hydrophobic, while their outer shell is hydrophilic. This property allows CDs to serve as host molecules, forming supramolecular aggregates with various guest molecules through host-guest interactions. CD-based supramolecular aggregates not only possess diverse structural morphologies but also exhibit remarkable stimuli-responsive properties in response to external stimuli. For example, in response to changes in temperature, light, pH, or the presence of specific chemicals, CD-based supramolecular aggregates can undergo structural changes, thereby altering their physical or chemical properties. This stimuli-responsive nature holds great promise for CD supramolecular aggregates in areas such as smart materials, targeted drug delivery, and sensors. For example, in biomedicine, CD-based supramolecular aggregates can serve as smart drug carriers, responding to internal or external stimuli. By changing their structure under specific conditions of pH, temperature, or light, they can control the rate and location of drug release, thereby achieving targeted and controlled drug delivery. In the field of materials, cyclodextrin-based supramolecular aggregates can be used to construct smart materials with adaptive properties that can self-adjust their physical or chemical properties according to environmental conditions. In environmental science, cyclodextrin-based molecular aggregates can be used as sensors that respond to environmental stimuli, used to detect and report various changes in the environment, such as pollutant concentrations, temperature fluctuations, or radiation intensity. In the field of energy science, stimulus-responsive supramolecular aggregates also have great potential and can be used to manufacture smart batteries and solar cells that change their conductivity or energy storage properties in response to light, temperature, or chemical stimuli. In short, the application prospects of stimulus-responsive cyclodextrin supramolecular aggregates are diverse and full of possibilities.

[0004] In recent years, the construction of cyclodextrin supramolecular aggregates has evolved from single-stimulus-responsive to multi-stimulus-responsive, further enhancing their application in various fields. Multi-stimulus responsiveness of supramolecular aggregates is typically achieved by covalently modifying cyclodextrin building blocks or by introducing additional functional groups into guest molecules. However, this inevitably increases the complexity of molecular design and synthesis. Currently, most multi-stimulus-responsive supramolecular aggregates developed and prepared are cumbersome and require laborious chemical synthesis steps. This contradicts the inherent advantages of non-covalent interactions in supramolecular chemistry and hinders large-scale industrial production, thus limiting their practical application. Furthermore, covalently modified cyclodextrins can pose numerous unforeseen risks and issues. For example, the introduction of functional groups during the modification process often reduces the water solubility of cyclodextrin molecules, thereby limiting their in vivo application. Furthermore, covalent modification can alter the spatial conformation of cyclodextrins, thereby affecting their binding ability to guest molecules and weakening the stability of the inclusion complex. To complicate matters further, covalently modifying cyclodextrins can also raise biocompatibility issues. For example, certain modification groups may be toxic or immunogenic, potentially causing adverse effects on organisms. Summary of the Invention

[0005] In response to the above problems, the present invention provides a rare earth supramolecular fluorescent aggregate and its preparation method and application. The preparation method of the present invention is simple, and the prepared supramolecular fluorescent aggregate has excellent fluorescence properties and multiple stimulus response characteristics.

[0006] In a first aspect, the present invention provides a method for preparing a rare earth supramolecular fluorescent aggregate, comprising the following steps:

[0007] Mixing a rare earth salt solution and a 2,6-pyridinedicarboxylic acid solution to carry out a coordination reaction to obtain a mixed solution A;

[0008] The polyoxyethylene polyoxypropylene ether triblock polymer solution and the carboxymethyl β-cyclodextrin solution are mixed and subjected to a host-guest inclusion complex reaction to obtain a mixed solution B;

[0009] After the mixed solution A and the mixed solution B are mixed, coordination and supramolecular aggregation reactions occur to obtain rare earth supramolecular fluorescent aggregates.

[0010] In a preferred embodiment of the present invention, in the mixed solution A, the molar ratio of the rare earth salt to 2,6-pyridinedicarboxylic acid is 1:1-2;

[0011] The rare earth element in the rare earth salt is one of europium, dysprosium, terbium, and samarium, and the rare earth salt is one of chloride, nitrate, and sulfate.

[0012] In a preferred embodiment of the present invention, the coordination reaction is carried out at 50-60° C. for 1-2 hours.

[0013] It can be understood that the reaction temperature can be, but is not limited to, 50°C, 51°C, 52°C, 53°C, 54°C, 55°C, 56°C, 57°C, 58°C, 59°C, 60°C;

[0014] Furthermore, the reaction time can be, but is not limited to, 1 h, 1.1 h, 1.2 h, 1.3 h, 1.4 h, 1.5 h, 1.6 h, 1.7 h, 1.8 h, 1.9 h, or 2 h.

[0015] In a preferred embodiment of the present invention, the host-guest inclusion complex reaction is carried out at 40-50° C. for 10-12 hours.

[0016] It can be understood that the reaction temperature can be, but is not limited to, 40°C, 41°C, 42°C, 43°C, 44°C, 45°C, 46°C, 47°C, 48°C, 49°C, or 50°C;

[0017] Furthermore, the reaction time can be, but is not limited to, 10 h, 10.5 h, 11 h, 11.5 h, or 12 h.

[0018] In a preferred embodiment of the present invention, the molar ratio of the rare earth salt in the mixed solution A to the carboxymethyl β-cyclodextrin in the mixed solution B is 1:1-7.

[0019] It can be understood that the molar ratio of the rare earth salt to the carboxymethyl β-cyclodextrin can be, but is not limited to, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, or 1:7.

[0020] The molar ratio of the rare earth salt to the carboxymethyl β-cyclodextrin is 1:3.

[0021] In a preferred embodiment of the present invention, in the mixed solution obtained by mixing the mixed solution A and the mixed solution B, the concentration of the polyoxyethylene polyoxypropylene ether triblock polymer is 0.150-0.600 mM.

[0022] It can be understood that the concentration of the polyoxyethylene polyoxypropylene ether triblock polymer can be, but is not limited to, 0.150 mM, 0.225 mM, 0.375 mM, 0.450 mM, 0.525 mM, or 0.600 mM.

[0023] More preferably, the concentration of the polyoxyethylene polyoxypropylene ether triblock polymer is 0.375 mM.

[0024] In a preferred embodiment of the present invention, the coordination and supramolecular aggregation reaction is carried out at 40-50° C. for 20-24 hours.

[0025] It can be understood that the reaction temperature can be, but is not limited to, 40°C, 41°C, 42°C, 43°C, 44°C, 45°C, 46°C, 47°C, 48°C, 49°C, or 50°C;

[0026] Furthermore, the reaction time can be, but is not limited to, 10 h, 10.5 h, 11 h, 11.5 h, or 12 h.

[0027] In a second aspect, the present invention provides rare earth supramolecular fluorescent aggregates prepared by the above preparation method.

[0028] In a third aspect, the present invention provides the use of the above-mentioned rare earth supramolecular fluorescent aggregates in the preparation of drug targeted release carriers, such as those used as targeted release carriers for related therapeutic drugs in diseases related to abnormal enzyme expression (such as pancreatitis) and cancer cell tissues.

[0029] At the same time, rare earth supramolecular fluorescent aggregates can also be used for the detection of nitro compounds and heavy metal ions, the nitro compound is nitrophenol, and the heavy metal ion is copper ion.

[0030] Compared with the prior art, the present invention has the following beneficial effects:

[0031] (1) The preparation method of the present invention is simple and easy, does not require complicated synthesis steps, and the raw materials can be directly purchased commercially. In the preparation process, both the host and guest molecules are non-toxic and can be used as drug additives. The carboxyl substituent on the carboxymethyl-β-cyclodextrin (CMCD) used is hydrophilic, which can further improve the water solubility of cyclodextrin. The stability of the inclusion compound can be enhanced by rare earth ion coordination.

[0032] This invention utilizes the host-guest inclusion complex between a carboxyl-containing cyclodextrin derivative, CMCD, and a triblock polymer (F127) to construct a supramolecular fluorescent ordered aggregate that can coordinate and aggregate with rare earth europium ions. Because rare earth europium ions have weak luminescence properties, an organic ligand, 2,6-pyridinedicarboxylic acid (DPA), is introduced into the supramolecular aggregate to enhance the fluorescence properties of the rare earth europium ions.

[0033] (2) The rare earth supramolecular fluorescent aggregates prepared by the present invention contain cyclodextrin molecules that can be hydrolyzed by α-amylase, so when α-amylase is added, the aggregates disintegrate and produce fluorescence quenching. When the pH is lower than 7, the DPA and CMCD ligands will dissociate from the central metal europium ion, so the aggregates will also disintegrate and produce fluorescence quenching. According to the principle of energy competition quenching, since the nitro compound absorbs the excitation light energy and competes with the ligands of the rare earth complex, the ligand cannot transfer energy to the central ion, resulting in fluorescence quenching. In addition, when metal copper ions that can coordinate with DPA and CMCD ligands are added, the rare earth ions and ligands are no longer coordinated due to the strong complexation of the copper ions, and the rare earth fluorescent aggregates that lose the sensitization effect of the organic ligand will also undergo fluorescence quenching. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 The fluorescence emission spectra of Eu[III] at an excitation wavelength of 286 nm in the complexes prepared with Eu[III] and CMCD at different molar ratios (a) and (b) Figure 1 (b) Changes in fluorescence emission intensity at 615 nm recorded in Figure a;

[0035] Figure 2 The relationship between the intensity of the optimal fluorescence emission peak of Eu[III]@PPRs at 615nm and the concentration of F127 is shown in the figure.

[0036] Figure 3 Fluorescence emission spectra of Eu[III], Eu[III]@CMCD, and Eu[III]@PPRs;

[0037] Figure 4 Fluorescence decay curves of Eu[III]@CMCD and Eu[III]@PPRs in H2O (A) and D2O (B) at 615 nm.

[0038] Figure 5 DLS (a), TEM (b), SEM (c) and EDX element distribution maps (df) of Eu[III]@PPRs. The scale bars in bf are all 500 nm.

[0039] Figure 6 Fluorescence emission spectra of Eu[III]@PPRs in the presence of 100 U / mL α-amylase at 37.0°C over time (A) (the inset shows the relationship between fluorescence intensity recorded at 615 nm and enzymatic hydrolysis time) and DLS data collected from Eu[III]@PPRs after 8 h of enzymatic hydrolysis (B) (the inset shows spherical aggregates of released Eu[III] after enzymatic hydrolysis);

[0040] Figure 7Figure 2 shows the fluorescence emission spectra of Eu[III]@PPRs at different pH values as the pH changes (A) (the inset shows the curve of fluorescence intensity recorded at 615 nm as a function of pH) and the DLS data collected from Eu[III]@PPRs at pH 1.2 (B) (the inset shows the small-sized solid spherical particles aggregated by PPRs at pH 1.2).

[0041] Figure 8 High-resolution image (a) of Eu[III]@PPRs at pH 1.2 and EDX element distribution maps of Eu (b), N (c), and O (d);

[0042] Figure 9 Fluorescence quenching emission spectrum of Eu[III]@PPRs by p-nitrophenol (PNP) (a) and the quenching linear relationship curve recorded at 615 nm (b);

[0043] Figure 10 (a) The fluorescence quenching emission spectrum of Eu[III]@PPRs by copper ions and the quenching linear relationship curve recorded at 615 nm (b) are shown. DETAILED DESCRIPTION

[0044] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0045] The existing technology for preparing supramolecular polymers is relatively complex, and there are several problems in the preparation of cyclodextrin supramolecular polymers: (1) the introduction of functional groups into cyclodextrins increases their hydrophobicity and reduces their water solubility; (2) covalent modification also affects the binding ability of cyclodextrins with guest molecules, weakening the stability of the inclusion complex; (3) some modified groups may also be toxic or immunogenic. Therefore, researchers still need to work hard to simplify the chemical composition of cyclodextrins and guest molecules and propose more optimized design schemes to meet the challenges of preparing smart supramolecular materials with multi-stimulus responsiveness.

[0046] Based on this, the present invention prepares a rare earth supramolecular fluorescent aggregate using rare earth salts, 2,6-pyridinedicarboxylic acid, a polyoxyethylene polyoxypropylene ether triblock polymer, and carboxymethyl β-cyclodextrin as raw materials. Both the host and guest molecules used in the present invention are non-toxic reagents. The carboxyl substituent in carboxymethyl β-cyclodextrin can increase the hydrophilicity of cyclodextrin. During the preparation process, the host-guest inclusion complex of CMCD and F127 is utilized to construct a supramolecular fluorescent ordered aggregate that can coordinate and aggregate with rare earth europium ions. The presence of rare earth ions can enhance the stability of the inclusion complex. In addition, because rare earth ions have weak luminescence properties, an organic ligand of pyridinedicarboxylic acid is introduced into the supramolecular aggregate to enhance the fluorescence properties of the rare earth ions and aggregates.

[0047] The supramolecular fluorescent aggregates prepared by this invention can produce fluorescence quenching in response to α-amylase and pH changes. In diseases associated with abnormal enzyme expression (such as pancreatitis) and cancer cell tissues, the prepared rare earth supramolecular fluorescent aggregates with unique stimulus-responsive properties are expected to be used as targeted delivery vehicles for therapeutic drugs, enabling optical diagnosis and treatment of related tumors and diseases. Furthermore, the aggregates can quantitatively detect the levels of copper ions and p-nitrophenol in aqueous solutions.

[0048] The specific preparation method is as follows:

[0049] The present invention provides a method for preparing a rare earth supramolecular fluorescent aggregate, comprising the following steps:

[0050] Step 1: Mixing a rare earth salt solution and a 2,6-pyridinedicarboxylic acid solution to carry out a coordination reaction to obtain a mixed solution A;

[0051] Step 2: mixing the polyoxyethylene polyoxypropylene ether triblock solution and the carboxymethyl β-cyclodextrin solution and performing a host-guest inclusion reaction to obtain a mixed solution B;

[0052] Step 3: Mixing the mixed solution A and the mixed solution B causes coordination and supramolecular aggregation reactions, and obtaining rare earth supramolecular fluorescent aggregates after stabilization.

[0053] Specifically, during the preparation, the rare earth salt solution, 2,6-pyridinedicarboxylic acid solution, polyoxyethylene polyoxypropylene ether triblock polymer solution and carboxymethyl β-cyclodextrin solution were all prepared using HEPSE buffer as a solvent. The concentration of the HEPSE buffer was 10 mM and the pH was 7.4.

[0054] The N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid (HEPES) used in the preparation process is 99.5%, with the molecular formula C8H 18N2O4S, molecular weight 238.30 g / mol, purchased from Shanghai MacLean Biochemical Technology Co., Ltd.; 2,6-pyridinedicarboxylic acid (abbreviated as DPA), 99%, molecular formula C7H5NO4, molecular weight 167.12 g / mol, purchased from Shanghai Titan Technology Co., Ltd.; carboxymethyl-β-cyclodextrin sodium salt (abbreviated as CMCD), 99%, molecular formula C 56 H 77 Na7O 49 , molecular weight 1695.11g / mol, and polyoxyethylene polyoxypropylene ether triblock polymer PluronicF127 (F127), 99%, PEO 100 PPO 64 PEO 100 , molecular weight 12600.00 g / mol, were purchased from Sigma;

[0055] In a preferred embodiment of the present invention, in the mixed solution A of step 1, the molar ratio of the rare earth salt to 2,6-pyridinedicarboxylic acid is 1:1-2;

[0056] The rare earth element in the rare earth salt is one of europium, dysprosium, terbium, and samarium, and the rare earth salt is one of a chloride, a nitrate, or a sulfate. Examples include europium chloride, dysprosium chloride, terbium chloride, samarium chloride, samarium nitrate, europium nitrate, and europium sulfate. The rare earth salt used in the present invention is europium chloride, specifically europium chloride hexahydrate, 99.99% with a molecular formula of EuCl3·6H20 and a molecular weight of 366.41 g / mol, purchased from Shanghai MacLean Biochemical Technology Co., Ltd.

[0057] In a preferred embodiment of the present invention, in step 1, the coordination reaction is carried out at 50-60° C. for 1-2 hours.

[0058] In a preferred embodiment of the present invention, in step 2, the host-guest inclusion complex reaction is carried out at 40-50°C for 10-12 hours; after the host-guest inclusion complex reaction is completed, it is stabilized at 25-28°C for 24-30 hours to make the aggregate morphology uniform and set aside.

[0059] In a preferred embodiment of the present invention, in step 3, the molar ratio of the rare earth salt in the mixed solution A to the carboxymethyl β-cyclodextrin in the mixed solution B is 1:1-7.

[0060] In a preferred embodiment of the present invention, in the mixed solution obtained by mixing the mixed solution A and the mixed solution B, the concentration of the polyoxyethylene polyoxypropylene ether triblock polymer is 0.150-0.600 mM.

[0061] In a preferred embodiment of the present invention, in step 3, the coordination and supramolecular aggregation reaction is carried out at 40-50° C. for 20-24 hours, and then stabilized at 25-28° C. for 24-30 hours to obtain rare earth fluorescent supramolecular aggregates.

[0062] The present invention produces a rare earth supramolecular fluorescent aggregate. It can be used to prepare a targeted drug release carrier, such as a carrier for the targeted release of therapeutic drugs in diseases associated with abnormal enzyme expression (such as pancreatitis) and cancer cell tissues. It can also be used in the detection of nitro compounds and heavy metal ions. The fluorescent response of the rare earth supramolecular aggregate to nitro compounds and heavy metal ions will facilitate the practical application of rare earth supramolecular aggregates in the detection field. Nitro compounds such as nitrophenols and heavy metal ions such as copper ions are suitable.

[0063] During the test, the nitrophenol used in the present invention is p-nitrophenol (abbreviated as PNP), 99%, molecular formula C6H5NO3, molecular weight 139.12g / mol, purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.; copper ions are provided by copper sulfate, copper sulfate is analytical grade, molecular formula CuSO4·5H2O, molecular weight 249.68g / mol, purchased from Shanghai Huaxing Chemical Plant; sodium hydroxide, analytical grade, molecular formula NaOH, molecular weight 40.00g / mol, purchased from Tianjin Tianda Chemical Experimental Plant;

[0064] The present invention utilizes the coordination aggregation of rare earth europium complexes and pseudopolyrotaxanes (PPRs) to construct a rare earth supramolecular fluorescent aggregate with multiple stimulus responses. Among them, the guest molecule Pluronic F127 in PPRs is an ideal nanocarrier for loading and controlled release of anticancer drugs. In order to connect the pseudopolyrotaxane with the rare earth europium complex, the present invention selects commercially available carboxyl-substituted cyclodextrin (carboxymethyl-β-cyclodextrin, abbreviated as CMCD) as the host molecule of PPRs. When a rare earth europium complex sensitized by 2,6-pyridinedicarboxylic acid (abbreviated as DPA) is added, the PPRs can further coordinate and aggregate with the rare earth complex through the carboxyl groups on the CMCD to form fluorescence-enhanced supramolecular vesicles, thereby preparing rare earth supramolecular fluorescent aggregates. In an environment where α-amylase and hydrochloric acid are introduced, the fluorescence intensity of the rare earth supramolecular fluorescent aggregates is quenched due to changes in the aggregation structure. This rare earth supramolecular fluorescent aggregate also produces fluorescent stimulation in response to copper ions and p-nitrophenol. The simplified assembly strategy proposed in this paper, which does not require complex molecular design and synthesis steps, provides new ideas for the design and construction of multi-stimulus-responsive supramolecular aggregates, thereby facilitating their further application in chemical analysis, diagnosis, and biomedicine.

[0065] In the following specific examples, unless otherwise specified, all raw materials can be obtained from commercial sources. The instruments used in the present invention are as follows: Shimadzu fluorescence spectrophotometer, model RF5301PC, wavelength range 220-800 nm; steady-state / transient fluorescence spectrometer, FLS920, Edinburgh, UK; transmission electron microscope TEM JEM-2100plus, voltage 200 kV; nanoparticle size and zeta potential analyzer (DLS), Malvern Zetasizer Nano ZS90, UK; scanning electron microscope SEM Hitachi Regulus 8100; atomic force microscope AFM Bruker Dimension ICON; pH meter, model Leici PHS-3C; analytical balance, model AR1140, range 0.0001g-110g; Zhicheng constant temperature incubation shaker, model ZWY-200D, power supply 220V±10%, 50-60Hz / 3.6A, maximum power 700W, Shanghai Zhicheng Analytical Instrument Manufacturing Co., Ltd.; Chinese LCD desktop ultrasonic cleaner, model KM-300DE, ultrasonic power 300W, ultrasonic frequency 40kHz, power supply 220V / 50Hz, Kunshan Meimei Ultrasonic Instrument Co., Ltd.; electromagnetic stirrer, model HS-12, power supply 220V 50Hz / 60Hz, power 180W. The water used in the following examples was high-purity water.

[0066] The invention is further described below with reference to specific examples. Firstly, the fluorescence intensity variation of different molar ratios of CMCD to Eu[III] is studied, and the appropriate CMCD addition ratio for constructing supramolecular aggregates is determined.

[0067] It should be noted that the concentration of the HEPSE buffer used in the present invention is 10 mM and the pH of the solution is 7.4.

[0068] The preparation method of EuCl3 solution is as follows: dissolve EuCl3·6H20 in HEPSE buffer to obtain 50mM EuCl3 solution;

[0069] The DPA solution was prepared as follows: DPA was dissolved in HEPSE buffer to obtain a 25 mM DPA solution;

[0070] The preparation method of CMCD solution is as follows: dissolve CMCD in HEPSE buffer to obtain a 10 mM CMCD solution;

[0071] The preparation method of F127 solution is as follows: dissolve F127 in HEPSE buffer to obtain 7.5 mM F127 solution;

[0072] The specific technical solutions are as follows:

[0073] Example 1

[0074] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0075] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0076] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0077] A 0.5 mL 10 mM CMCD solution was diluted to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 hours and stabilized at 25°C for 24 hours. Then, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. After mixing, sonication was continued for another 24 hours. The complex, Eu[III]@CMCD, was obtained after stabilization at 25°C for another 24 hours.

[0078] Example 2

[0079] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0080] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0081] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0082] 1.0 mL of a 10 mM CMCD solution was diluted to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 hours and stabilized at 25°C for 24 hours. Then, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. After mixing, sonication was continued for another 24 hours, and the complex, Eu[III]@CMCD, was obtained after stabilization at 25°C for another 24 hours.

[0083] Example 3

[0084] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0085] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0086] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0087] 1.5 mL of 10 mM CMCD solution was diluted to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h. Then, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. After mixing, sonication was continued for 24 h and the sample was stabilized at 25°C for 24 h to obtain the complex Eu[III]@CMCD.

[0088] Example 4

[0089] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0090] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0091] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0092] Take 2.0 mL of a 10 mM CMCD solution and dilute to 4.5 mL with 10 mM HEPES buffer. The sample is then sonicated at 40°C for 12 hours and stabilized at 25°C for 24 hours. Then, 0.5 mL of the Eu[III] solution prepared in step 1 is added dropwise in a 40°C water bath. After mixing, sonication is continued for 24 hours. The complex, Eu[III]@CMCD, is obtained after stabilization at 25°C for 24 hours.

[0093] Example 5

[0094] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0095] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0096] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0097] 2.5 mL of a 10 mM CMCD solution was diluted to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 hours and stabilized at 25°C for 24 hours. Then, 0.5 mL of the previously prepared 10 mM Eu[III] solution was added dropwise in a 40°C water bath. After mixing, sonication was continued for another 24 hours, and the sample was stabilized at 25°C for another 24 hours to obtain the Eu[III]@CMCD complex.

[0098] Example 6

[0099] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0100] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0101] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0102] Take 3.0 mL of a 10 mM CMCD solution and dilute to 4.5 mL with 10 mM HEPES buffer. The sample is then sonicated at 40°C for 12 hours and stabilized at 25°C for 24 hours. Then, 0.5 mL of the Eu[III] solution prepared in step 1 is added dropwise in a 40°C water bath. After mixing, sonication is continued for 24 hours and the solution is stabilized at 25°C for 24 hours to obtain the complex Eu[III]@CMCD.

[0103] Example 7

[0104] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0105] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0106] Step 2: Preparation of rare earth fluorescent supramolecular aggregate Eu[III]@CMCD:

[0107] 3.5 mL of a 10 mM CMCD solution was diluted to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 hours and stabilized at 25°C for 24 hours. Then, 0.5 mL of the previously prepared 10 mM Eu[III] solution was added dropwise in a 40°C water bath. After mixing, sonication was continued for another 24 hours, and the sample was stabilized at 25°C for another 24 hours to obtain the complex Eu[III]@CMCD.

[0108] The fluorescence intensity of the complexes Eu[III]@CMCD prepared in Examples 1 to 7 was tested. The molar ratios of Eu[III]:CMCD in Examples 1 to 7 were 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, and 1:7, respectively. Figure 1 It can be seen that with the increase of CMCD addition, the fluorescence intensity of Eu[III] first increases steadily, and reaches the maximum value when the molar ratio of Eu[III] / CMCD is 1:3. If the CMCD is increased beyond this ratio, the fluorescence intensity of Eu[III] will decrease. This is because the CMCD and Eu[III] 3+ There is no antenna effect between them, and its ability to sensitize rare earth ions to luminescence is weaker than DPA, so Eu 3+ The fluorescence intensity of the CMCD decreases with the competitive coordination of the CMCD. Furthermore, the optimal molar ratio of Eu[III] / CMCD was determined to be 1:3, at which the supramolecular aggregates were constructed and their fluorescence properties were studied.

[0109] Example 8

[0110] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0111] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0112] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0113] 0.10 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0114] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Eu[III]@PPRs:

[0115] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. Mixing was followed by continuous sonication for 24 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Eu[III]@PPRs was finally obtained.

[0116] Example 9

[0117] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0118] Mix 2.0 mL of 10 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0119] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0120] 0.15 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0121] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Eu[III]@PPRs:

[0122] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. Mixing was followed by continuous sonication for 24 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Eu[III]@PPRs was finally obtained.

[0123] Example 10

[0124] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0125] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0126] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0127] 0.25 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0128] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Eu[III]@PPRs:

[0129] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. Mixing was followed by continuous sonication for 24 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Eu[III]@PPRs was finally obtained.

[0130] Example 11

[0131] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0132] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0133] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0134] 0.30 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0135] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Eu[III]@PPRs:

[0136] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Eu[III] (10 mM) solution prepared in step 1 was added dropwise in a 40°C water bath. Mixing was followed by continuous sonication for 24 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Eu[III]@PPRs was obtained.

[0137] Example 12

[0138] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0139] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0140] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0141] 0.35 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0142] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Eu[III]@PPRs:

[0143] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. Mixing was followed by continuous sonication for 24 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Eu[III]@PPRs was finally obtained.

[0144] Example 13

[0145] Step 1, 10 mM europium complex [Eu(DPA)2] - (Abbreviated as Eu[III]) solution configuration:

[0146] Mix 2.0 mL of 50 mM EuCl 3 solution and 8.0 mL of 25 mM DPA solution, and stir at 50° C. for 2 h to prepare a 10 mM Eu[III] solution for later use.

[0147] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0148] 0.40 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 40°C for 12 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0149] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Eu[III]@PPRs:

[0150] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Eu[III] solution prepared in step 1 was added dropwise in a 40°C water bath. Mixing was followed by continuous sonication for 24 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Eu[III]@PPRs was finally obtained.

[0151] Example 14

[0152] Step 1, Dysprosium complex [Dy(DPA)2] - (abbreviated as Dy[III]) solution configuration:

[0153] 2.0 mL of 50 mM DyCl 3 solution and 4.0 mL of 25 mM DPA solution were mixed, and the mixture was stirred at 60° C. for 1 h to prepare a Dy[III] solution for later use.

[0154] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0155] 0.25 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 50°C for 10 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0156] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Dy[III]@PPRs:

[0157] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Dy[III] solution prepared in step 1 was added dropwise in a 50°C water bath. Mixing was followed by ultrasonication for 20 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Dy[III]@PPRs was finally obtained.

[0158] Example 15

[0159] Step 1, terbium complex [Tb(DPA)2] - (abbreviated as Tb[III]) solution configuration:

[0160] 2.0 mL of 50 mM TbCl 3 solution and 6.0 mL of 25 mM DPA solution were mixed, and the mixture was stirred at 55° C. for 1.5 h to prepare a Tb[III] solution for later use.

[0161] Step 2: Preparation of pseudopolyrotaxane (PPRs) solution of CMCD and F127:

[0162] 0.25 mL of 7.5 mM F127 and 1.5 mL of 10 mM CMCD solution were mixed at room temperature and brought to 4.5 mL with 10 mM HEPES buffer. The sample was then sonicated at 45°C for 11 h and stabilized at 25°C for 24 h to obtain the PPRs solution, which was then used.

[0163] Step 3: Preparation of rare earth fluorescent supramolecular aggregates Tb[III]@PPRs:

[0164] To the 4.5 mL PPRs solution prepared in step 2, 0.5 mL of the Tb[III] solution prepared in step 1 was added dropwise in a 45°C water bath. Mixing was followed by continuous sonication for 22 h. After stabilization at 25°C for 24 h, the rare earth fluorescent supramolecular aggregate Tb[III]@PPRs was finally obtained.

[0165] When the Eu[III] / CMCD ratio was 1:3, Examples 8-13 adjusted the host-guest inclusion ratio of CMCD and F127 by changing the concentration of added F127, and constructed a series of Eu[III]@PPRs supramolecular fluorescent aggregates. The optimal addition concentration of F127 was explored using the fluorescence intensity of the aggregates at 615 nm as the evaluation index. Figure 2 The fluorescence intensity change curves of Eu[III]@PPRs at 615 nm at different F127 concentrations (the concentration of F127 is 0-0.600 mM) in Examples 3 and 8-13.

[0166] from Figure 2 As can be seen, the fluorescence intensity of the system peaks at an F127 concentration of 0.375 mM and then remains stable as the F127 concentration increases. This is because F127 begins to form micelles when the F127 concentration exceeds 0.375 mM. At 25°C, the critical micelle concentration of F127 in a pH 7.4 buffer is 0.40 mM, indicating that micellar F127 cannot be included by CMCD. Therefore, using Example 10 as an example, the fluorescence properties and aggregation behavior of rare earth supramolecular aggregates will be explored below.

[0167] Figure 3 The fluorescence emission spectra of the blank sample of the rare earth complex (added with an equal volume of HEPSE buffer solution, Eu[III]), added with CMCD (Eu[III]@CMCD prepared in Example 3), and added with PPRs (Eu[III]@PPRs prepared in Example 10) are compared at a maximum excitation wavelength of 286 nm. Figure 3As can be seen in the figure, the fluorescence intensity of Eu[III] increased significantly after the addition of CMCD and PPRs. Under the same Eu[III] concentration conditions, the fluorescence intensity of Eu[III] coordinated with PPRs increased by 5.2 times compared to when CMCD coordinated without PPRs (Eu[III]@CMCD), indicating that CMCD is more effective in coordinating with rare earth complexes by forming PPRs.

[0168] In the fluorescence emission spectrum, the two emission bands at 596 and 615 nm are attributed to Eu 3+ of 5 D0→ 7 The magnetic dipole of F1 and 5 D0→ 7 The electric dipole transition of F2. The ratio of the peak intensity of the electric dipole transition to the magnetic dipole transition is K=I( 5 D0→ 7 F2) / I( 5 D0→ 7 F1), can be used as a measure of the degree of symmetry of the coordination environment of europium ions or to illustrate changes in the microchemical environment around rare earth ions. A higher K value indicates a higher asymmetry of the coordination structure of the rare earth ions and a stronger interaction between the rare earth complex and its host matrix. It can be calculated from the fluorescence emission spectrum that the K value (4.48) of Eu[III] coordinated with PPRs is higher than the K value (3.81) when it is directly coordinated with CMCD. This shows that after forming PPRs, CMCD has a stronger coordination effect with Eu[III], making the coordination result of the central rare earth europium ion more asymmetric and the fluorescence more monochromatic. The data of fluorescence lifetime measurement also show ( Figure 4 (Figure A) There are two lifetimes in the Eu[III]@CMCD sample: a short lifetime of 0.311ms (38%) and a long lifetime of 1.440ms (62%). This is due to the presence of some Eu[III] in the system that is not coordinated with CMCD, which leads to the existence of some short lifetimes and also illustrates the instability of the coordination environment of the central europium ion in the Eu[III]@CMCD sample. However, in the Eu[III]@PPRs sample, only a long lifetime of 1.512ms was fitted, and the fluorescence lifetime was longer than that of Eu[III]@CMCD. This shows that under the action of PPRs, the coordination environment of Eu[III] is single and more stable.

[0169] By measuring the fluorescence lifetime of Eu[III]@CMCD and Eu[III]@PPRs in heavy water as solvent ( Figure 4(Figure B) The number of coordinated water molecules present in the first coordination sphere of the rare earth europium complexes in both samples can be calculated. The number of water molecules in the first coordination sphere is 0.43 for Eu[III]@CMCD and 0.03 for Eu[III]@PPRs, respectively. This indicates that the presence of PPRs effectively reduces the number of coordinated water molecules, thereby reducing fluorescence quenching caused by the non-radiative vibrational deactivation of the OH groups in the water molecules. This enhances the fluorescence intensity of the rare earth complex Eu[III] in the Eu[III]@PPRs sample.

[0170] Figure 5 Dynamic light scattering (DLS), transmission electron microscopy (TEM), scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDX) data of Eu[III]@PPRs rare earth fluorescent supramolecular aggregates prepared in Example 10. DLS data ( Figure 5 Figure a) shows that the average particle size of the aggregates formed by Eu[III]@PPRs is 690nm. At the same time, the TEM image ( Figure 5 b) and SEM images ( Figure 5 Hollow vesicle spherical aggregates with diameters ranging from 454 nm to 659 nm were observed in Figure c). Figure 5 The arrow in Figure c marks the collapsed hollow structure of the vesicle. Figure 5 EDX data analysis of Figures e and f of the vesicles showed that the vesicles were enriched with a large amount of europium and nitrogen, indicating that the rare earth complex Eu[III] was indeed bound into the spherical aggregates of the vesicles.

[0171] Next, the performance of the rare earth supramolecular fluorescent aggregate prepared in Example 10 was tested.

[0172] (1) Preparation of enzymatic fluorescence response experimental solution

[0173] To the prepared rare earth supramolecular fluorescent aggregate solution, 12.5 μL of an 8000 U / mL aqueous solution of α-amylase (derived from Aspergillus oryzae) was added, bringing the α-amylase concentration to 100 U / mL. The sample was then placed in a 37°C water bath, and the change in fluorescence intensity was measured at regular intervals.

[0174] Since the 1,4-glycosidic bond of β-CD can be hydrolyzed by α-amylase, the supramolecular aggregates of Eu[III]@PPRs will produce fluorescence response to α-amylase due to the degradation of CMCD. Figure 6 Figure A shows the curve of the change of Eu[III]@PPRs fluorescence intensity over time when the temperature is 37°C and the α-amylase concentration in the system is 100 U / mL. Figure 6The arrows in Figure A indicate the time sequence from 0h to 8.0h. Figure 6 As can be seen in A, after adding α-amylase, the fluorescence intensity of the supramolecular aggregates gradually decreased with time. After 7 hours, the fluorescence intensity no longer changed, indicating that the supramolecular aggregates were completely enzymatically degraded at about 7 hours, and the fluorescence of the system no longer changed. DLS data showed that ( Figure 6 (Figure B) shows that after enzymatic hydrolysis, a large number of aggregates with an average particle size of 9.7 nm are present in the system. These are F127 monomer molecules released after CMCD hydrolysis. Furthermore, less than 1% of the system exhibits signals with a particle size larger than 600 nm, indicating that a small amount of undigested supramolecular aggregates may remain. TEM analysis revealed that the small number of 600 nm aggregates are hollow spherical aggregates that have released rare earth europium complexes.

[0175] (2) Preparation of pH fluorescence response experimental solution

[0176] In the prepared rare earth supramolecular fluorescent aggregate solution, a 3M HCl aqueous solution was used as the acidity regulating liquid and added to the aggregate system via a microliter syringe. After stirring at room temperature for several minutes, the adjusted pH value of the solution was determined using a pH acidity meter, and then fluorescence measurement was performed.

[0177] Since rare earth europium complexes with carboxylic acid ligands are stable in neutral environments but undergo ligand dissociation under acidic conditions, this acid-base dependent property between rare earth europium ions and organic ligands leads to the fluorescence sensitivity of supramolecular vesicles to pH. Figure 7 Figure 2 Fluorescence emission spectrum and pH response curve of Eu[III]@PPRs fluorescent supramolecular aggregates after adjusting the solution pH with 3M HCl solution. Figure 7 The direction of the arrow in Figure a is the order from pH 7.4 to 1.2. Figure 7 It can be seen that as the pH of the system gradually decreases, the fluorescence intensity of Eu[III]@PPRs also gradually decreases. At a pH of 1.2, the fluorescence decreases to 6% of the initial fluorescence intensity, and at a pH of 1.2, the fluorescence is basically quenched completely. This shows that at lower pH values, both CMCD and DPA ligands completely dissociate from the rare earth europium ions, resulting in complete quenching of the aggregate fluorescence. In addition, when the pH value of the system drops to 1.2, a peak with a particle size of 125 nm appears on the DLS curve ( Figure 7 b). At the same time, low-contrast aggregated particles with an average diameter of 110 nm were also found in the TEM image ( Figure 7 b inset). Since the size of these particles is much larger than the size of the micelles formed by F127, and the EDX elemental analysis of TEM also shows that there is no europium ion aggregation in the spherical particles ( Figure 8). Therefore, these small aggregates are formed by the aggregation of dissociated PPRs. This may be because at lower pH values, the carboxyl substituents on CMCD are protonated, which reduces the electrostatic repulsion between PPRs and increases their hydrophobicity. Therefore, after the vesicles disintegrate, the PPRs can aggregate together to form small solid spherical particles.

[0178] (3) Preparation of fluorescence quenching titration experimental solution

[0179] In the experiments on the fluorescence quenching titration of supramolecular fluorescent aggregates by nitro compounds and metallic copper ions, 0.8 μL of 250 mM p-nitrophenol (PNP) or copper sulfate solution was added to the prepared rare earth supramolecular fluorescent aggregate solution each time, so that the concentration of the detection substance in the solution changed to 0, 40, 80, 120, 160, and 200 μM, respectively. After oscillation and ultrasonic mixing, the change in its fluorescence intensity was measured.

[0180] According to the principle of energy competition quenching, nitro compounds absorb excitation light energy, competing with the ligands of the rare earth complexes. This prevents the ligands from transferring energy to the central ion, leading to fluorescence quenching. Furthermore, when copper ions, which can coordinate with DPA and CMCD ligands, are added, the strong complexation of the copper ions prevents the coordination between the rare earth ions and the ligands. Consequently, the rare earth fluorescent aggregates, deprived of the sensitizing effect of the organic ligands, also experience fluorescence quenching. Based on this, the fluorescence quenching properties of the aggregates for nitro compounds and copper ions were investigated.

[0181] Figure 9 Figure 3 is the fluorescence emission spectrum of Eu[III]@PPRs after adding different concentrations of p-nitrophenol (PNP) and its fluorescence quenching linear relationship curve. Figure 9 The arrows in Figure a indicate the order of PNP concentration from 0 to 200 μM. The fluorescence emission spectra show that the fluorescence intensity of the rare earth fluorescent aggregates gradually decreases as the PNP concentration increases. Using the Stern–Volmer equation, I0 / I=K sv [C]+1 can describe the quenching efficiency of PNP on supramolecular fluorescent aggregates, where I0 is the fluorescence intensity without PNP, I is the fluorescence intensity after adding PNP, and [C] is the added concentration of PNP. At an emission wavelength of 616 nm, I0 / I is plotted against the added concentration of PNP [C] ( Figure 9 b) The linear equation obtained by fitting is I0 / I=0.00357[C]+0.97889, and the correlation of the equation is R 2 =0.99535, the fluorescence quenching constant K can be known from the slope sv =3.57L / mol. According to the calculation formula of detection limit LOD=3σ / K sv, the minimum detection limit of rare earth fluorescent aggregates for PNP can be obtained, where σ is the standard deviation of the y-axis intercept of the Stern–Volmer line equation. By calculation, the minimum detection limit of rare earth fluorescent aggregates for PNP is LOD=8.226×10 -6 mol / L.

[0182] Figure 10 Figure 3 is the fluorescence emission spectrum of Eu[III]@PPRs after adding copper sulfate solution of different concentrations and its fluorescence quenching linear relationship curve. Figure 10 The arrows in Figure a indicate the order in which the concentration of copper sulfate solution increases from 0 to 200 μM. The fluorescence emission spectra show that the fluorescence intensity of the supramolecular fluorescent aggregates gradually decreases as the concentration of copper ions increases. Using the modified Stern–Volmer equation, ln(I0 / I)=K sv [C]+1 can describe the quenching efficiency of copper ions on rare earth fluorescent aggregates, where I0 is the quenching efficiency of copper ions without adding Cu. 2+ The fluorescence intensity of I is the fluorescence intensity of Cu 2+ The fluorescence intensity after [C] is Cu 2+ The concentration of Cu is calculated by ln(I0 / I) at the emission wavelength of 615nm. 2+ The added concentration [C] is plotted ( Figure 10 Figure b), the linear equation obtained by fitting is ln(I0 / I)=0.00772[C]-0.06229, and the correlation of the equation is R 2 =0.99157, the fluorescence quenching constant K can be known from the slope sv =7.72L / mol. According to the calculation formula of detection limit LOD=3σ / K sv , we can get the rare earth fluorescent aggregates to Cu 2+ The lowest detection limit of Cu is σ, where σ is the standard deviation of the y-intercept of the Stern–Volmer line equation. 2+ The lowest detection limit is LOD = 2.421 × 10 -7 mol / L.

[0183] Although the preferred embodiments of the present invention have been described, those skilled in the art may make additional changes and modifications to these embodiments once they have learned the basic creative concept. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.

[0184] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if such changes and modifications fall within the scope of the claims and their equivalents, the present invention is intended to include such changes and modifications.

Claims

1. A method for preparing rare earth supramolecular fluorescent aggregates, characterized in that: The following steps are involved: A rare earth salt solution and a 2,6-pyridinedicarboxylic acid solution are mixed to carry out a coordination reaction to obtain a mixed solution A; the rare earth element in the rare earth salt is europium; The polyoxyethylene polyoxypropylene ether triblock polymer solution and the carboxymethyl β-cyclodextrin solution are mixed and subjected to a host-guest inclusion complex reaction to obtain a mixed solution B; After the mixed solution A and the mixed solution B are mixed, coordination and supramolecular aggregation reactions occur to obtain rare earth supramolecular fluorescent aggregates.

2. The method for preparing a rare earth supramolecular fluorescent aggregate according to claim 1, characterized in that: In the mixed solution A, the molar ratio of the rare earth salt to 2,6-pyridinedicarboxylic acid is 1:1-2; Rare earth salts are a type of chloride, nitrate, or sulfate.

3. The method for preparing a rare earth supramolecular fluorescent aggregate according to claim 1, characterized in that: The coordination reaction is carried out at 50-60 °C for 1-2 h.

4. The method for preparing a rare earth supramolecular fluorescent aggregate according to claim 1, wherein: The host-guest inclusion complex reaction is carried out at 40-50 °C for 10-12 h.

5. The method for preparing a rare earth supramolecular fluorescent aggregate according to claim 1, characterized in that: The molar ratio of the rare earth salt in the mixed solution A to the carboxymethyl beta-cyclodextrin in the mixed solution B is 1:1-7.

6. The method for preparing a rare earth supramolecular fluorescent aggregate according to claim 1, characterized in that: In the mixed solution obtained by mixing the mixed solution A and the mixed solution B, the concentration of the polyoxyethylene polyoxypropylene ether triblock polymer is 0.15-0.60 mM.

7. The method for preparing a rare earth supramolecular fluorescent aggregate according to claim 1, characterized in that: The coordination and supramolecular aggregation reactions were carried out at 40-50 °C for 20-24 h.

8. A rare earth supramolecular fluorescent aggregate prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the rare earth supramolecular fluorescent aggregate according to claim 8 in preparing a drug targeted release carrier.

10. Use of the rare earth supramolecular fluorescent aggregate according to claim 8 in detecting nitro compounds and heavy metal ions, characterized in that: The nitro compound is p-nitrophenol, and the heavy metal ion is copper ion.

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