Calix pyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nano sensor and application thereof to heparin sodium content detection

Abstract

The invention discloses a calix-pyrene tetrasulfonic acid tetrasodium salt supramolecular nano sensor and application thereof in detecting the content of heparin sodium, and is characterized in that a building unit takes calix-pyridine as a subject and pyrene tetrasulfonic acid tetrasodium salt as an object, and a supramolecular nano sensing system is built through inclusion coordination interaction of the subject and the object. The sensor can be used for monitoring the content of heparin sodium in human blood in real time, and has the advantages that the preparation method of the supramolecular nano sensor is efficient, simple and convenient, the dosage of host and guest raw materials is small, and the cost is low. Heparin sodium is the most important clinical anticoagulant drug. However, if excessive heparin sodium exists in a human body, serious complications such as cerebral hemorrhage can be induced. Therefore, the detection of the content of the heparin sodium has great significance and wide application prospect in the field of biological medicines.

Description

[0001] This patent was supported by the National Natural Science Foundation of China (21901184) and the Tianjin Natural Science Foundation (20JCQNJC00400; 20JCYBJC00330). Technical Field

[0002] The invention belongs to the technical field of supramolecular nanomaterial preparation, relates to the preparation of a supramolecular nanosensor of calixpyridine-pyrenetetrasulfonic acid tetrasodium salt, and realizes the detection of heparin sodium content by using the system. Background Art

[0003] Supramolecular assembly systems have attracted extensive attention in recent years due to their broad application prospects in many fields such as chemistry, materials, and biomedicine. See: 1) B.-P. Jiang, D.-S. Guo, Y.-C. Liu, K.-P. Wang, Y. Liu. ACS Nano .2014, 8 , 1609−1618; 2) Y.-X. Wang, Y.-M. Zhang, Y. Liu. J. Am. Chem. Soc .2015, 137 , 4543−4549. Supramolecular assembly systems have shown special advantages in the field of detection technology. See: 1) RN Dsouza, A. Hennig, WM Nau. Chem. Eur. J. 2012, 18 , 3444−3459; 2) D.-E. Wang, L. Zhao, et al. ACS Appl. Mater. Interfaces 2016, 8 , 28231−28240; 3) H. Li, H.-N. Xie, et al. Anal. Chem .2013, 85 , 1047−1052. For example, the supramolecular fluorescence sensing system constructed by negatively charged sulfonated calixarene and positively charged fluorescent dye has been successfully applied to the monitoring of the kinetics and progress of the hydrolysis reaction of many cationic enzyme reaction substrates under the catalysis of related enzymes, as well as the selective detection of related enzymes, see: 1) D.-S. Guo, VD Uzunova, X. Su, Y. Liu, WM Nau. Chem. Sci. 2011, 2 , 1722−1734; 2) M. Florea, S. Kudithipudi, A. Rei, MJ González-Álvarez, A. Jeltsch, WM Nau. Chem. Eur. J. 2012, 18 , 3521−3528; 3) D.-S. Guo, J. Yang, Y. Liu. Chem. Eur. J. 2013, 19 , 8755−8759. However, there are few reports on supramolecular nanosensing systems based on anion recognition.

[0004] Heparin sodium is a sulfated polysaccharide with many negative charges. It can bind to positively charged proteins and participate in many physiological processes. Heparin sodium is the most important anticoagulant drug in clinical practice. However, excessive heparin sodium in the human body may induce serious complications, such as cerebral hemorrhage. Therefore, it is crucial to accurately control the dosage of heparin sodium during surgery or anticoagulant treatment. See: S.-W. Li, M. Gao, etal. Chem. Commun. 2017, 53 , 4795.

[0005] Calixopyridines are a class of cyclic compounds rich in pyridinium cations, with good water solubility and the potential to bond with anionic guests. The compounds can be synthesized simply and efficiently from 3-bromomethylpyridine, see: S.Shinoda, M. Tadokoro, H. Tsukube, R. Arakawa. Chem. Commun. 1998, 181−182. However, the construction of supramolecular assemblies based on calixpyridinium anion recognition has rarely been reported. Summary of the invention

[0006] The purpose of the present invention is to provide a supramolecular nanosensing system of calixpyridine-pyrenetetrasulfonic acid tetrasodium salt in view of the above technical analysis, and to detect the content of heparin sodium by using the system.

[0007] To achieve the above purpose, the present invention discloses the following technical contents: A supramolecular nanosensor of calixpyridine-pyrene tetrasulfonic acid tetrasodium salt is characterized in that: the building unit is composed of calixpyridine as the main body and pyrene tetrasulfonic acid tetrasodium salt as the guest, and the high-resolution transmission electron microscope image of nanoparticles is assembled through the host-guest inclusion coordination interaction to construct a supramolecular nanosensor; its structural formula is as follows: .

[0008] The present invention further discloses a method for preparing a calixpyridine-pyrenetetrasulfonic acid tetrasodium salt supramolecular nanosensor, which is characterized by: dissolving calixpyridine and pyrenetetrasulfonic acid tetrasodium salt in water respectively to obtain a calixpyridine solution and a pyrenetetrasulfonic acid tetrasodium salt solution, adding the pyrenetetrasulfonic acid tetrasodium salt solution to the calixpyridine solution, and uniformly mixing to obtain a supramolecular nanosensor; wherein the concentrations of calixpyridine and pyrenetetrasulfonic acid tetrasodium salt are 50 μmol / L and 8.8 μmol / L respectively.

[0009] The present invention further discloses the application of the cuppyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nanosensor in detecting the heparin sodium content in human blood. The detection of the heparin sodium content in human blood refers to: when heparin sodium is added to the cuppyridine-pyrene tetrasulfonic acid tetrasodium salt aqueous solution, the intermolecular force between the cuppyridine and the heparin sodium is stronger, causing the pyrene tetrasulfonic acid tetrasodium salt to be gradually separated, and the heparin sodium and the cuppyridine to gradually polymerize, thereby displaying an ultraviolet signal. In this way, the cuppyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nanosensor is used to detect the heparin sodium content. The experimental results show that the cuppyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nanosensor is highly selective in detecting heparin sodium.

[0010] The advantages of the calixpyridine-pyrenetetrasulfonic acid tetrasodium salt supramolecular nanosensor disclosed in the present invention and its application in detecting heparin sodium content are: Based on the supramolecular nanosensor of cuppyridine-pyrene tetrasulfonic acid tetrasodium salt, its preparation method is efficient and simple, with small amounts of host and guest raw materials and low cost. Heparin sodium is the most important clinical anticoagulant drug. However, excessive heparin sodium in the human body may induce serious complications, such as cerebral hemorrhage. Therefore, the detection of heparin sodium content has great significance and broad application prospects in the field of biomedicine. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1 This is an experimental graph showing the change in transmittance of a calixpyridine aqueous solution at a wavelength of 350 nm as a function of its concentration; Figure 2 This is an experimental graph showing the change in transmittance of an aqueous solution of sodium heparin at a wavelength of 350 nm as a function of its concentration; Figure 3 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 350nm with the concentration of calixpyridine when calixpyridine is gradually added to an aqueous solution with a fixed concentration of heparin sodium of 19 μg / mL; Figure 4 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 350 nm with the concentration of heparin sodium when heparin sodium is gradually added to an aqueous solution with a fixed concentration of 50 μmol / L of calixpyridine; Figure 5 High-resolution transmission electron microscopy image of nanoparticles assembled from the binary supramolecular assembly of calixpyridine and heparin sodium; Figure 6 This is an experimental graph showing the change of the transmittance of the aqueous solution at a wavelength of 350 nm with time when the concentrations of calixpyridine and sodium heparin are fixed at 50 μmol / L and 36 μg / mL respectively; Figure 7 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 450 nm with the concentration of tetrasodium pyrenetetrasulfonate when tetrasodium pyrenetetrasulfonate is gradually added to an aqueous solution with a fixed concentration of 50 μmol / L of calixpyridine; Figure 8 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 450nm with the concentration of heparin sodium when the concentrations of calixpyridine and tetrasodium pyrenetetrasulfonate are fixed at 50 μmol / L and 8.8 μmol / L respectively; Fig. 9 This is a comparison chart of the changes in transmittance of three aqueous solutions of calixpyridine (100 μmol / L)−sodium hyaluronate (1.4 μmol / L), calixpyridine (50 μmol / L)−sodium chondroitin sulfate (60 μg / mL), and calixpyridine (50 μmol / L)−sodium heparin (36 μg / mL) at a wavelength of 350 nm with the addition of tetrasodium pyrenetetrasulfonic acid; Fig.10 It is a comparison chart of the changes in the transmittance of the aqueous solution of calixpyridine (50 μmol / L)-pyrenetetrasulfonic acid tetrasodium salt (8.8 μmol / L) at a wavelength of 450 nm with the addition of three polyanions: sodium hyaluronate, sodium chondroitin sulfate and sodium heparin; DETAILED DESCRIPTION

[0012] The present invention is described below with reference to the examples. The embodiments described herein do not limit the present invention. Professionals in the field may make improvements and changes according to the spirit of the present invention. These improvements and changes should be considered to be within the scope of the present invention. The scope and essence of the present invention are defined by the claims. Among them, tetrasodium pyrenetetrasulfonate is commercially available. Heparin sodium was purchased from Alfa Company; calixpyridine is synthesized simply and efficiently using 3-bromomethylpyridine as a raw material, see: S.Shinoda, M. Tadokoro, H. Tsukube, R. Arakawa. Chem. Commun. 1998, 181−182. Example

[0013] A method for preparing a calixpyridine-pyrenetetrasulfonic acid tetrasodium salt supramolecular supramolecular nanosensor comprises the following steps: dissolving calixpyridine and pyrenetetrasulfonic acid tetrasodium salt in water to obtain a calixpyridine solution and a pyrenetetrasulfonic acid tetrasodium salt solution, adding the pyrenetetrasulfonic acid tetrasodium salt solution to the calixpyridine solution, and uniformly mixing at room temperature to obtain a supramolecular supramolecular nanosensor target solution, wherein the concentrations of the calixpyridine and pyrenetetrasulfonic acid tetrasodium salt are 50 μmol / L and 8.8 μmol / L, respectively.

[0014] 1. Responsiveness of the supramolecular fluorescence sensor: (1) Figure 1 This is an experimental graph showing the change in transmittance of a calixpyridine aqueous solution at a wavelength of 350 nm as a function of its concentration. Figure 1 As shown, the transmittance of the aqueous solution of calixpyridine itself at a wavelength of 350 nm does not change with the increase of its concentration, indicating that calixpyridine itself does not have a critical aggregation concentration in the aqueous solution and calixpyridine itself cannot self-aggregate in the aqueous solution. Figure 2 This is an experimental graph showing the change in transmittance of an aqueous solution of sodium heparin at a wavelength of 350 nm as a function of its concentration. Figure 2 As shown, the transmittance of the aqueous solution of heparin sodium itself at a wavelength of 350 nm gradually decreases with the increase of its concentration, indicating that heparin sodium itself can self-aggregate in the aqueous solution. From the inflection point of the change, it can be obtained that its critical aggregation concentration is 3.02 mg / mL; Figure 3 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 350 nm with the concentration of calixpyridine when calixpyridine is gradually added to an aqueous solution with a fixed concentration of heparin sodium of 19 μg / mL (this concentration is far lower than the critical aggregation concentration of heparin sodium, so heparin sodium exists as a monomer at this concentration and does not aggregate), as shown in the figure. Figure 3 As shown in the figure, with the addition of calixpyridine, the transmittance of the system at a wavelength of 350 nm remained basically unchanged at first, and then decreased, indicating that the presence of sodium heparin can induce the aggregation of calixpyridine in aqueous solution. From the inflection point of the change, it can be obtained that when the concentration of sodium heparin is fixed at 19 μg / mL, the critical aggregation concentration of calixpyridine is 18.5 μmol / L. Figure 4 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 350 nm with the concentration of heparin sodium when heparin sodium is gradually added to an aqueous solution with a fixed concentration of 50 μmol / L of calixpyridine, as shown in Figure 4 As shown in the figure, with the addition of sodium heparin, the transmittance of the system at a wavelength of 350 nm decreased significantly at first, dropped to the lowest point, and then slowly increased. From the inflection point of the change, it can be obtained that the optimal mixing concentrations of calixpyridine and sodium heparin to form aggregates in the solution are 50 μmol / L and 36 μg / mL, respectively. Figure 5 High-resolution transmission electron microscopy images of nanoparticles assembled from the binary supramolecular assembly of calixpyridine and heparin sodium. Figure 5As shown, the morphology of the supramolecular nanoparticles is fiber-shaped. Figure 6 The experimental graph shows the change of the transmittance of the aqueous solution at a wavelength of 350 nm with time when the concentrations of calixpyridine and heparin sodium are fixed at 50 μmol / L and 36 μg / mL, respectively. Figure 6 As shown, the transmittance of the system at a wavelength of 350 nm does not change over time for up to 7 days, indicating that the cuppyridine-heparin sodium supramolecular aggregate has high stability, further proving the existence of strong intermolecular forces between cuppyridine and heparin sodium.

[0015] (2) Figure 7 This is an experimental graph showing the change of the transmittance of the system at a wavelength of 450 nm with the concentration of tetrasodium pyrenetetrasulfonate when tetrasodium pyrenetetrasulfonate is gradually added to an aqueous solution with a fixed concentration of 50 μmol / L of calixpyridine, as shown in the figure. Figure 7 As shown, with the addition of tetrasodium pyrenetetrasulfonic acid, the transmittance of the system at a wavelength of 450nm slowly decreased. The results showed that there was also an intermolecular force between cuppyridine and tetrasodium pyrenetetrasulfonic acid. Example

[0016] Application of the calixpyridine-pyrenetetrasulfonic acid tetrasodium salt supramolecular nanosensor prepared in Example 1, adding polyanion heparin sodium to the supramolecular nanosensor system, the method is as follows: Dissolve cuppyridine, tetrasodium pyrenetetrasulfonate and sodium heparin in water to obtain cuppyridine solution, tetrasodium pyrenetetrasulfonate and sodium heparin solution, add tetrasodium pyrenetetrasulfonate solution to cuppyridine solution, mix evenly to obtain supramolecular nanosensor, wherein the concentrations of cuppyridine and tetrasodium pyrenetetrasulfonate are 50 μmol / L and 8.8 μmol / L, respectively. Figure 8 As shown, after gradually adding heparin sodium solution into the target solution of the cuppyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nanosensor prepared above, the transmittance of the system at a wavelength of 450 nm gradually decreased. The results showed that the content of heparin sodium can be detected by the cuppyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nanosensor.

[0017] Comparative test: (1) When tetrasodium pyrenetetrasulfonate solution is added dropwise to the solutions of calixpyridine-sodium hyaluronate, calixpyridine-sodium chondroitin sulfate, and calixpyridine-sodium heparin, Fig. 9 As shown in the figure, with the addition of tetrasodium pyrenetetrasulfonic acid, the transmittance of the calixpyridine-sodium hyaluronate system at a wavelength of 450nm increases the fastest, followed by the calixpyridine-sodium chondroitin sulfate system, and the calixpyridine-sodium heparin system increases the slowest. This shows that the intermolecular force between calixpyridine and sodium hyaluronate is the weakest among the three, and the intermolecular force between calixpyridine and sodium heparin is the strongest among the three.

[0018] (2) When sodium hyaluronate, sodium chondroitin sulfate and sodium heparin are added to the target solution of the above-mentioned cuppyridine-pyrene tetrasulfonic acid tetrasodium salt supramolecular nanosensor, as Fig.10 As shown in the figure, after adding sodium hyaluronate, sodium chondroitin sulfate and sodium heparin respectively, the transmittance of the ternary system of calixpyridine-tetrasodium pyrenetetrasulfonate-sodium heparin at a wavelength of 450 nm decreased the fastest. The results show that the supramolecular nanosensor has a high selectivity for the determination of sodium heparin content, and the interference of sodium hyaluronate and sodium chondroitin sulfate, both of which are glycosaminoglycans, can be almost ignored during the detection.

Claims

1. A supramolecular nanosensor of calixpyridine-pyrenetetrasulfonic acid tetrasodium salt, characterized in that: The building unit is based on cuppyridine as the main body and pyrenetetrasulfonic acid tetrasodium salt as the guest. Through the host-guest inclusion coordination interaction, it is assembled into a high-resolution transmission electron microscopy image of nanoparticles to construct a supramolecular nanosensor. Its structural formula is as follows: 。 2. The method for preparing the supramolecular nanosensor of cuprylopyridine-pyrenetetrasulfonic acid tetrasodium salt according to claim 1, characterized in that: Cuppyridine and tetrasodium pyrenetetrasulfonate were dissolved in water to obtain cuppyridine solution and tetrasodium pyrenetetrasulfonate solution, respectively. The tetrasodium pyrenetetrasulfonate solution was added into the cuppyridine solution and mixed evenly to obtain a supramolecular nanosensor, wherein the concentrations of cuppyridine and tetrasodium pyrenetetrasulfonate were 50 μmol / L and 8.8 μmol / L, respectively.

3. Use of the cuppyridine-pyrenetetrasulfonic acid tetrasodium salt supramolecular nanosensor according to claim 1 in the preparation of a method for detecting the content of heparin sodium in human blood.

4. The use of claim 3, wherein the detection of the heparin sodium content in human blood refers to: when heparin sodium is added to an aqueous solution of calixpyridine-pyrenetetrasulfonic acid tetrasodium salt, the intermolecular force between calixpyridine and heparin sodium is stronger, causing pyrenetetrasulfonic acid tetrasodium salt to be separated, while heparin sodium and calixpyridine polymerize, thereby displaying an ultraviolet signal.

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