A fluorescent sensor based on tripodal pillar[5]arene and its application in detecting lead ion

Abstract

The application provides a fluorescent sensor based on a tripod [5] arene, which is obtained by self-assembly complexation of a host of a tripod [5] arene TP5 and a guest of a quaternary ammonium salt in an H2O-DMSO system. A supramolecular system is constructed by host-guest interaction between the tripod [5] arene TP5 and the amphiphilic guest TPN, and then PbS fluorescent nanoparticles are generated in situ under ion induction. Fluorescence enhancement and high-sensitivity detection (LOD=7.00*10 2+ -6 M) of Pb ‑8 are realized. In addition, the fluorescent supramolecular polymer network based on the PbS fluorescent nanoparticles can be successfully applied to in-situ detection of Pb 2+ in cells.

Description

TECHNICAL FIELD

[0001] The present application relates to a fluorescent sensor based on tripodal pillar[5]arene; the present application also relates to the application of the fluorescent sensor in the fluorescence recognition of Pb 2+ in aqueous solution and cells, and belongs to the field of chemical synthesis and ion detection. BACKGROUND

[0002] Lead has good ductility, low melting point and corrosion resistance, and is widely used in industries such as battery, paint and plastic manufacturing, which has led to various environmental pollution. Lead pollution is a serious global environmental problem, which poses a significant threat to human health. It can directly enter the human body through the skin, digestive and respiratory systems. In addition, excessive lead content in water and soil will lead to lead pollution of crops, which will then be absorbed by the human body through the food chain. Since lead is non-biodegradable, the accumulation of lead ions (Pb 2+ ) in the body will cause permanent damage to the nervous system, hematopoietic system and various organs, and lead to developmental disorders in children. Even after treatment, the level of Pb 2+ in the blood is significantly reduced, but the damaged tissues and organs cannot be restored. The World Health Organization (WHO) and the International Agency for Research on Cancer have listed Pb 2+ as a carcinogen. Therefore, it is very important to develop a simple, highly selective and highly sensitive sensing probe to recognize lead ions.

[0003] Among the numerous fluorescent nanomaterials, supramolecular fluorescent nanomaterials occupy an important position. Supramolecular fluorescent nanomaterials have been widely studied in the fields of fluorescent sensing, drug delivery and optoelectronic devices due to their good external stimulus response characteristics and non-covalent assembly reversibility. Among them, pillar[n]arene is a popular macrocyclic host material in the study of supramolecular nanomaterials, which was first discovered and reported by Ogoshi in 2008. The presence of an electron-rich cavity makes pillar[n]arene an ideal candidate for constructing high-performance fluorescent nanomaterials.

[0004] The commonly used Pb 2+ detection methods include gas chromatography, high-performance liquid chromatography, excitation Raman spectroscopy and atomic absorption spectroscopy, but these methods are expensive, inconvenient to operate, time-consuming and complex to operate. However, fluorescent sensors are attracting attention due to their high sensitivity, fast analysis speed, good reproducibility, good biocompatibility, good stability and high-quality biological imaging. SUMMARY

[0005] The purpose of the present application is to provide a fluorescent sensor based on tripodal pillar[5]arene;

[0006] Another purpose of the present application is to provide the application of the fluorescent sensor in the fluorescence recognition of Pb2+ .

[0007] 1. Preparation of tripod-based [5]arene fluorescence sensors

[0008] The present invention is a fluorescent sensor based on tripod [5] aromatic hydrocarbons. The sensor is self-assembled and complexed in a H2O-DMSO system using tripod [5] aromatic hydrocarbons TP5 as a host and quaternary ammonium salt TPN as a guest, with the host and guest being self-assembled in a molar ratio of 1:3 to 1:3.2. The sensor is labeled as TN. In the H2O-DMSO system, the volume ratio of H2O to DMSO is 1:4 to 1:4.2.

[0009] Synthesis of tripodal [5] aromatic hydrocarbons: Using SNP5 as substrate, dichloromethane as solvent, and triethylamine as catalyst, 1,3,5-benzenetricarboxylic acid chloride and SNP5 were stirred at room temperature for 12 hours at a molar ratio of 1:3 to 1:3.5. The mixture was concentrated in vacuo, filtered, washed, and dried to obtain the compound tripodal [5] aromatic hydrocarbon TP5, whose molecular formula is: C 171 H204N6O 36 S3, the structural formula is as follows:

[0010] ;

[0011] The structural formula of SNP5 is: .

[0012] Figure 1 、 Figure 2 They are respectively the hydrogen spectrum and mass spectrum of the above-mentioned synthetic main body TP5, indicating that the main body TP5 was successfully synthesized.

[0013] Synthesis of guest quaternary ammonium salt (TPN): M1 is used as substrate, methanol is used as solvent, M1 and benzothiazole-2-acetonitrile are reacted at a molar ratio of 1:1 to 1:1.5 at 40 to 60°C for 4 to 6 hours, cooled, filtered, washed, and dried to obtain the guest quaternary ammonium salt, labeled TPN, with the molecular formula: C 25 H 30 N3OSBr, the structural formula is as follows:

[0014] ;

[0015] Among them, the structural formula of M1 is .

[0016] Figure 3 、 Figure 4 They are the hydrogen spectrum and mass spectrum of the above-mentioned synthesized guest TPN, respectively, indicating that the guest TPN was successfully synthesized.

[0017] Figure 5The figure is the nuclear magnetic titration spectrum of TP5 and TPN. Wherein, (a) (TP5); (b) (TP5 and 3 times equivalent TPN); (c) (TPN). It can be seen from the figure that when 3 times equivalent guest TPN is added to the host TP5, Ha and Hb on TP5 move to the low field, H2 and H3 on TPN also move to the low field, and H1 moves to the high field. These phenomena show that the quaternary ammonium salt on the guest TPN is located in the cavity of the host TP5.

[0018] II. Fluorescent recognition of Pb based on supramolecular polymer of tripodal [5] arene 2+

[0019] 1. Fluorescent properties of fluorescent sensor TN based on tripodal [5] arene

[0020] Through the study of the fluorescent properties of the fluorescent sensor TN, it is shown that the fluorescent sensor TN has good solubility and fluorescence emission properties in the DMSO-H2O system (H2O and DMSO volume ratio is 1:4 ~ 1:4.2). When the excitation wavelength is 380 nm, the fluorescent sensor TN has basically no fluorescence.

[0021] 2. Fluorescent recognition of Pb based on fluorescent sensor TN of tripodal [5] arene 2+

[0022] In the DMSO-H2O system of the fluorescent sensor (H2O and DMSO volume ratio is (1:4 ~ 1:4.2), 10 times equivalent (relative to TP5) of S 2- H2O solution is added to obtain TN-S, and then a series of cations (Zn 2+ , Pb 2+ , Cd 2+ , Ni 2+ , Fe 3+ , Co 2+ , Hg 2+ , Ag + , Ca 2+ , Cu 2+ , Mg 2+ , Cr 3+ , Ba 2+ , Tb 3+ , Eu 3+ , La 3+ , Al 3+ , Ce 3+ , Th 4+ ) are added. It is found that only the addition of Pb 2+ can turn on the fluorescence of TN-S, and the addition of other cations cannot cause significant change in the fluorescence intensity of TN-S (such as Figure 6 Therefore, TN-S can selectively recognize Pb.2+ .

[0023] At the same time, in order to avoid the interference of other cations to the experiment, we made an anti-interference experiment. The results show that the presence of other cations has no interference on the recognition of Pb 2+ by TN-S. Figure 7 Therefore, the supramolecular polymer TN-S can be used as a fluorescence sensor for recognizing Pb 2+ .

[0024] The fluorescence titration experiment shows that the minimum detection limit of the supramolecular polymer TN-S for Pb 2+ is 7.00*10 -8 M (as shown in Figure 8 and Figure 9 ), which indicates that the sensitivity of the fluorescence sensor TN-S for recognizing Pb 2+ is high.

[0025] 3. Recognition mechanism

[0026] TP5 and TPN can be complexed in a host-guest manner in a DMSO-H2O system, the quaternary ammonium salt of the guest TPN enters the cavity of the host TP5, and the fluorescence sensor TN is formed. After S 2- is added, TN-S is formed through anion-Π interaction, and then cation Pb 2+ is added, and PbS quantum dots are formed with S 2- , so that the fluorescence is enhanced (as shown in Figure 10 ).

[0027] In summary, the present application uses triptycene [5] arene as a host, uses a quaternary ammonium salt as a guest, and forms a fluorescence sensor TN in a host-guest complexation in a DMSO-H2O system. S 2- is added to the fluorescence sensor to form TN-S, and then cation Pb 2+ is added, and PbS quantum dots are formed with S 2- , so that the fluorescence is enhanced. Therefore, it can be used for single selective fluorescence recognition of Pb 2+ , and the minimum detection limit is 7.00*10 -8 M. In addition, the supramolecular system based on PbS quantum dots can be used for in-situ detection of Pb 2+ in living cells. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 is the hydrogen spectrum of the compound TP5 of the present application;

[0029] Figure 2 is the hydrogen spectrum of the compound TPN of the present application;

[0030] Figure 3 is the mass spectrum of the compound TP5 of the present application;

[0031] Figure 4 Mass spectrum of compound TPN of the present application;

[0032] Figure 5 NMR titration spectrum of TP5 and TPN;

[0033] Figure 6 Fluorescence full scan (λ = 380 nm) of DMSO-H2O solution of TN-S with different cations added;

[0034] Figure 7 Anti-interference experiment of DMSO-H2O solution of TN-S in the presence of other cations to Pb 2+ ;

[0035] Figure 8 Fluorescence titration experiment (λ = 380 nm) of TN-S to Pb 2+ ;

[0036] Figure 9 Fitting curve of fluorescence titration experiment of TN-S to Pb 2+ ;

[0037] Figure 10 TEM of PbS quantum dots formed;

[0038] Figure 11 Dark field and fluorescence confocal images of cells incubated for 4 hours with a mixture of TP5 and TPN, supramolecular polymer network containing S 2- , and supramolecular polymer network based on PbS-QD (live Hela cells (a1 bright field, a2 dark field, a3 superimposition) and TN (b1 bright field, b2 dark field, b3 superimposition), (c1 bright field, c2 dark field, c3 superimposition) TN-S, (d1 bright field, d2 dark field, d3 superimposition) TN-S + Pb 2+ (in PbS QDs) incubated for 4 h). DETAILED DESCRIPTION

[0039] The preparation of the fluorescent sensor TN of the present application and its application in the recognition of Pb 2+ will be further described below through specific implementations.

[0040] Example 1, Preparation of fluorescent sensor TN based on trisacetylenic pillar[5]arene

[0041] 1. Synthesis of host compound TP5

[0042] (1) Synthesis of compound SNP5: According to the literature Zhang, Y. M.; Yang, Q. Y.; Ma, X. Q.; Dong, H. Q.; Zhang, Y. F.; Guan, W. L.; Yao, H.; Wei, T. B.; Lin, Q. J Phys Chem A, 2020, 124 (47), 9811-9817.

[0043] (2) Synthesis of host compound TP5:

[0044] Into a 100 mL round-bottom flask was added SNP5 (1.57 g, 1.65 mmol), dichloromethane (20 mL) successively, and stirred at room temperature to make it fully dissolved, then triethylamine (0.5 mL) was added as a catalyst. Into dichloromethane (20 mL) was added trimesoyl chloride (0.13 g, 0.5 mmol), and stirred at room temperature to make it fully dissolved. The solution was slowly added into the round-bottom flask under ice-water bath with a constant pressure dropping funnel, and stirred at room temperature for 12 hours. The mixture was concentrated under vacuum, and compound TP5 was obtained by anti-solvent crystallization in dichloromethane / petroleum ether system, with a yield of 95 %. 1 H NMR (400 MHz, DMSO-d6), δ / ppm: δ 8.76 - 8.71 (m, 3H), 8.60 (s, 3H), 8.15 (d, J = 5.3 Hz, 3H), 6.76 (t, J = 5.0 Hz, 30H), 3.79 (s, 6H), 3.65 (s, 118H), 3.13 - 3.08 (m, 12H), 2.50 (s, 6H), 1.69 (d, J = 6.1 Hz, 6H), 1.52 (d, J = 6.7 Hz, 6H), 1.41 (dd, J = 16.2, 6.5 Hz, 12H). HR-MS: m / z [TP5 + 2H] 2+ Calcd for C 171 H 204 N6O 36 S33016.3663; found 3016.3696.

[0045]

[0046] 2、Guest compound quaternary ammonium salt TPN synthesis

[0047] (1) Synthesis of compound M1: According to the literature Sun, X. W.; Wang, Z. H.; Li, Y. J.; Zhang, Y. F., Zhang, Y. M.; Yao, H.; Wei, T. B.; Lin, Q. Macromolecules, 2021, 54 (1), 373-383.

[0048] (2) Synthesis of compound TPN: methanol as solvent (10 mL), benzothiazole-2-acetonitrile (0.5 mmol, 0.0871 g) and M1 (0.5 mmol, 0.1721 g) were dissolved in a 50 mL round-bottom flask, refluxed at 40 ~ 60 °C for 4 ~ 6 h, suction filtered, washed with ether for 3 times, and dried to obtain yellow solid quaternary ammonium salt with a yield of 80%; (M.P. 202-205 °C). 1 H NMR (400 MHz, DMSO- d 6) δ 8.35 (s, 1H), 8.18 (d, J = 8.4 Hz, 1H), 8.13 (d, J = 8.9 Hz, 2H), 8.06 (d, J = 8.3 Hz, 1H), 7.59 – 7.51 (m, 2H), 7.19 – 7.16 (m, 2H), 4.13 (t, J = 6.4 Hz, 2H), 3.32 (s, 2H), 3.05 (s, 9H), 1.82 – 1.71 (m, 4H), 1.53 – 1.46 (m, 2H), 1.40 – 1.33 (m, 2H).

[0049]

[0050] 3. Synthesis of fluorescent sensor TN

[0051] In a 10 mL DMSO-H2O system (H2O and DMSO in a volume ratio of 1:4), host compound TP5 (0.0030 g, 1 ×10 -6 mol) and guest compound TPN (0.0015 g, 3 ×10 -6 mol) were added to obtain fluorescent sensor TN. (TN = 1 ×10 -3 mol / L).

[0052] Example two, fluorescent recognition of Pb 2+

[0053] 2 mL of DMSO-H2O solution of fluorescent sensor TN (TN = 5 ×10 -5mol / L, H2O and DMSO volume ratio is 1:4) in a series of colorimetric tubes, first add 10eq of S 2- , then add 15 times the equivalent of cationic Zn 2+ , Pb 2+ , Cd 2+ , Ni 2+ , Fe 3+ ,Co 2+ , Hg 2+ , Ag + , Ca 2+ , Cu 2+ , Mg 2+ , Cr 3+ , Ba 2+ , Tb 3+ , Eu 3+ , La 3+ , Al 3+ , Ce 3+ and Th 4+ If the fluorescence intensity of TN increases, it means that Pb 2+ If the fluorescence of TN does not change significantly, it means that the added 2+ .

[0054] Example 3: Cell Imaging Experiment

[0055] In order to scientifically and effectively evaluate the cell activity of TN, MTT was used to conduct experiments. The activity of fluorescent probe TN was still greater than 63% after 4 hours after entering Hela cells. Therefore, it was proved that the toxicity of fluorescent probe TN to cells was relatively low and it could be used to detect Pb in cells. 2+ In addition, living HeLa cells were cultured with TN for 4 hours. The obtained HeLa cells containing supramolecular nanoparticles TN were cultured with S 2- Solution treatment, incubation for 4 hours, and then washing with buffer solution to remove the remaining TN-S. HeLa cells were treated with TN and S 2- After incubation in the solution for 4 hours, the cells showed weak fluorescence; 2+ The obtained cells were further treated with the solution, incubated for 4 hours, rinsed three times with PBS, fixed with paraformaldehyde, and then observed by laser confocal scanning microscopy (LSCM). Under LSCM, bright fluorescence emission can be observed on HeLa cells ( Figure 11 ), which proves that Pb 2+ induced in situ formation of PbS quantum dots in HeLa cells.

Claims

1. A fluorescent sensor based on tripod [5] aromatic hydrocarbons, wherein the tripod [5] aromatic hydrocarbons are used as a host and a quaternary ammonium salt is used as a guest in a DMSO-H2O binary system, wherein the host and the guest are self-assembled and complexed in a molar ratio of 1:3 to 1:3.2; The structural formula of the main tripodal columnar [5] aromatic hydrocarbon is: ； The structural formula of the quaternary ammonium salt guest is: 。 2. The tripod-column [5]arene-based fluorescence sensor according to claim 1, characterized in that: In the DMSO-H2O system, the volume ratio of H2O to DMSO is 1:4 ~ 1:4.

2.

3. The fluorescent sensor based on tripod [5] aromatic hydrocarbons as claimed in claim 1 is used for single selective fluorescence detection of Pb in aqueous system. 2+ Application in.

4. The fluorescent sensor based on tripod [5] aromatic hydrocarbons as claimed in claim 3 is used for single selective fluorescence detection of Pb in aqueous system. 2+ The application is characterized by: In the DMSO-H2O system of the fluorescence sensor, S 2- TN-S was obtained, and then cation Zn was added 2+ , Pb 2+ , Cd 2+ , Ni 2+ , Fe 3+ ,Co 2+ , Hg 2+ , Ag + , Ca 2+ , Cu 2+ , Mg 2+ , Cr 3+ , Ba 2+ , Tb 3 + , Eu 3+ , La 3+ , Al 3+ , Ce 3+ , Th 4+ H2O solution, only Pb 2+ The addition of can enhance the fluorescence intensity of TN-S, while the addition of other cations cannot cause significant changes in the fluorescence intensity of TN-S.

5. The tripod-based [5]arene fluorescent sensor as claimed in claim 1 is used for detecting Pb in cells for non-diagnostic or therapeutic purposes 2+ application.

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