A method for detecting Cu 2+ , BPO colorimetric fluorescent probe TZ6 and its preparation method and application

Abstract

A colorimetric fluorescent probe TZ6 for detecting Cu<supgt;2+< / supgt> and BPO, its preparation method and application, which relate to the field of fluorescent probes. The preparation method of the colorimetric fluorescent probe TZ6 includes the following steps: S1: Potassium carbonate is added to acetonitrile to obtain a mixed solution I; S2: 11-Ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazin-2(11H)-one and 2-(chloromethyl)pyridine are refluxed and stirred in the mixed solution I to obtain a mixed solution II; S3: Using a dichloromethane-methanol system as the mobile phase, the mixed solution II is separated by column chromatography to obtain the colorimetric fluorescent probe TZ6. The colorimetric fluorescent probe TZ6 provided by the present invention has good selectivity, anti-interference ability and sensitivity to both Cu<supgt;2+< / supgt> and BPO, and can detect Cu<supgt;2+< / supgt> and BPO under amphoteric acid-base conditions.

Description

Technical Field

[0001] The present invention relates to the field of fluorescent probes, and in particular to a fluorescent probe for detecting Cu 2+ , BPO colorimetric fluorescent probe TZ6 and its preparation method and application. Background Art

[0002] Benzoyl peroxide (BPO) has good bleaching properties and is therefore widely used as an additive in food processing. Studies have found that excessive BPO in food (such as wheat flour) can produce highly dangerous oxidative free radicals and transform into toxic substances such as biphenyl and benzoate, which can cause human tissue damage and disease. In order to ensure our food safety, it is particularly necessary to measure BPO in flour.

[0003] So far, a variety of analytical methods have been developed to detect BPO in food, such as high performance liquid chromatography, chemiluminescence and electrochemical analysis, but these detection methods often have complex and time-consuming pretreatment processes, and the instruments used are relatively expensive and have poor portability. Fluorescent probes have been widely used in the detection process due to their advantages such as simple operation, high sensitivity, good selectivity and low cost. It is particularly important to develop a fluorescent probe for detecting BPO in food.

[0004] Copper is the third essential trace element for the human body and plays a vital role in various cellular processes and the nervous system. Copper ions are coenzyme factors for many enzymes in the body and play an important role in various physiological processes. 2+ It is also the main source of heavy metal pollution in the natural environment. With the development of industry, more and more Cu 2+ It enters the environment through the discharge of waste gas, sewage and waste residue, harming the environment and posing a serious threat to human health. 2+ Too little or too much has a significant impact on both the natural environment and the human body. 2+ Can cause anemia, and excess Cu 2+ It can also cause neurodegenerative syndrome, liver damage in infants and young children, cirrhosis and other diseases. 2+ The fluorescent probe is of great significance for environmental protection and ensuring life and health.

[0005] There are also some methods for determining Cu in the prior art. 2+ However, most of these fluorescent probes are single-function, that is, a fluorescent probe can only recognize a single object. A multifunctional fluorescent probe refers to a fluorescent probe that can recognize different objects and produce different fluorescent signals, thereby realizing the multifunctional use of the probe. 2+There are few reports on dual-functional fluorescent probes with BPO. Summary of the invention

[0006] The present invention provides a method for detecting Cu 2+ , BPO colorimetric fluorescent probe TZ6 and its preparation method and application to Cu 2+ and BPO for testing.

[0007] In order to achieve the above object, the specific scheme adopted by the present invention is: a method for detecting Cu 2+ , a colorimetric fluorescent probe TZ6 of BPO, wherein the structural formula of the colorimetric fluorescent probe TZ6 is as follows:

[0008] .

[0009] A method for detecting Cu 2+ The method for preparing a colorimetric fluorescent probe TZ6 of BPO comprises the following steps: S1: adding potassium carbonate to acetonitrile to obtain a mixed solution I; S2: refluxing and stirring 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one and 2-(chloromethyl)pyridine in the mixed solution I to obtain a mixed solution II; S3: using a dichloromethane-methanol system as a mobile phase, separating the mixed solution II by column chromatography to obtain the colorimetric fluorescent probe TZ6.

[0010] As a further optimization of the above technical solution: the ratio of the added amounts of acetonitrile and potassium carbonate is 10 ml:1 mmol.

[0011] As a further optimization of the above technical solution: the molar ratio of 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one and 2-(chloromethyl)pyridine is 1:1.

[0012] As a further optimization of the above technical solution: the ratio of the added amounts of 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one, 2-(chloromethyl)pyridine and acetonitrile is 1mmol:1mmol:10ml.

[0013] As a further optimization of the above technical solution: in step S2, the reflux stirring time is 3-4 hours.

[0014] As a further optimization of the above technical solution: in step S2, the temperature of reflux stirring is 80°C.

[0015] A colorimetric fluorescent probe TZ6 for detecting Cu 2+ , application in BPO.

[0016] As a further optimization of the above technical solution: the colorimetric fluorescent probe TZ6 and the solution to be detected are mixed to obtain a mixed solution, and the mixed solution is subjected to fluorescence detection; if the fluorescence is enhanced, the solution to be detected contains BPO; if the fluorescence is quenched, the solution to be detected contains Cu 2+ .

[0017] As a further optimization of the above technical solution: mix the colorimetric fluorescent probe TZ6 and the solution to be tested to obtain a mixed solution, and observe the color of the mixed solution. If the color changes from yellow to colorless and transparent, the solution to be tested contains BPO; or, irradiate the mixed solution with an ultraviolet lamp with a wavelength of 365nm. If the fluorescence color changes from rose red to green, the solution to be tested contains BPO.

[0018] Compared with the prior art, the present invention has the following beneficial effects: the preparation method of the colorimetric fluorescent probe TZ6 provided by the present invention is simple, and it can be prepared by refluxing and stirring two raw materials in a mixed solution of acetonitrile and potassium carbonate, and then separating them by column chromatography; the prepared colorimetric fluorescent probe TZ6 has the advantages of long emission wavelength, visible color change, short response time, good selectivity and sensitivity, etc.

[0019] The colorimetric fluorescent probe TZ6 provided by the present invention is effective for Cu 2+ The fluorescence detection limit of BPO is only 5.29nM, and that of Cu 2+ The fluorescence detection limit is 25.68nM. It also has a fast response time and a wide pH adaptability range. When detecting BPO, the maximum fluorescence intensity can be reached within 10 minutes. It can detect Cu under both acidic and alkaline conditions. 2+ , BPO detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The colorimetric fluorescent probe TZ6 1 HNMR nuclear magnetic spectrum;

[0021] Figure 2 Fluorescence spectra of probe TZ6 (10 μM) + M (100 μM) in DMF:PBS (2:8, v:v, 0.1 μM) solvent, where M is Cu 2+ , BPO, ClO - 、Ag + 、Al 3+ , Ba 2+ , Ca 2+ 、Cd 2+ 、Co 2+ Cr 3+ , Fe 2+ , Fe 3+ , Hg 2+ Mg2+ , Mn 2+ 、Ni 2+ , Pb 2+ , Pd 2+ 、Zn 2+ 、HSO3 - 、NO2 - , S 2- 、SO3 2- 、HCO3-、H2PO4 - 、F - , Cl - 、SO4 2- 、CH3COO - , H2O2, glycine, lysine, glucose, lactose, (excitation wavelength: 398nm; slit width: 5nm);

[0022] Figure 3 Fluorescence spectra of probe TZ6 (10 μM) + M (100 μM) in DMF:PBS (2:8, v:v, 0.1 μM) solvent, where M is Cu 2+ 、Ag + 、Al 3+ , Ba 2+ , Ca 2+ 、Cd 2+ 、Co 2+ Cr 3+ , Fe 2+ , Fe 3+ , Hg 2+ Mg 2+ , Mn 2+ 、Ni 2+ , Pb 2+ , Pd 2+ 、Zn 2+ 、HSO3 - 、NO2 - , S 2- 、SO3 2- 、HCO3-、H2PO4 - 、F - , Cl - 、SO4 2- 、CH3COO - , H2O2, glycine, lysine, glucose, lactose, (excitation wavelength: 398nm; slit width: 5nm);

[0023] Figure 4 Competition experiment of probe TZ6 recognizing BPO in the presence of some common metal ion salts in DMF:H2O=2:8 solvent: 1. BPO; 2. Ca 2+ ; 3.Fe 2+ ; 4.Fe3+ ; 5.Mg 2+ ; 6.Zn 2+ ;7.Na + ; 8.KI; 9.NO3 - ; 10.CsF; 11.Cl - ; 12.HCO3 - ; 13.H2PO4 - ; 14. SO4 2- ; 15. NO2 - ; 16. H2O2; 17. Glycine; 18. Lactose; 19. Glucose; 20. Lysine, (excitation wavelength: 408nm; slit width: 2.5nm);

[0024] Figure 5 In the presence of some common metal ion salts in DMF:H2O=2:8 solvent, probe TZ6 recognizes Cu 2+ Competition experiment: 1. Probe TZ6; 2. Cu 2+ ; 3.Ag + ; 4.Al 3+ ; 5.Ba 2+ ; 6.Ca 2+ ; 7.Cd 2+ ;8.Co 2+ ; 9.Cr 3+ ; 10.Fe 2+ ; 11.Fe 3+ ; 12.Hg 2+ ; 13.Mg 2+ ; 14.Mn 2+ ; 15.Ni 2+ ; 16.Pb 2+ ; 17.Pd 2+ ; 18.Zn 2+ ; 19.HSO3 - ; 20. NO2 - ; 21.S 2- 22. CIO - ; 23.SO3 2- ; 24.HCO3 - ; 25.H2PO4 - ; 26.F - ; 27.Cl - , (excitation wavelength: 408nm; slit width: 2.5nm);

[0025] Figure 6 is the linear relationship between the concentration of BPO and the fluorescence emission intensity at 505 nm in DMF:H2O=2:8 solvent (excitation wavelength: 408 nm; slit width: 2.5 nm);

[0026] Figure 7 In DMF:H2O=2:8 solvent, Cu 2+ The linear relationship between the concentration and the fluorescence intensity at 605 nm (excitation wavelength: 408 nm; slit width: 5 nm);

[0027] Figure 8 It is the reaction time diagram of probe TZ6 and BPO;

[0028] Fig. 9 The graph shows the change of fluorescence intensity of probe TZ6 (10 μM) + BPO (50 μM) with pH in DMF:H2O=2:8 solvent;

[0029] Fig.10 In DMF:H2O=2:8 solvent, probe TZ6 (10 μM) + Cu 2+ (50 μM) as a function of pH. DETAILED DESCRIPTION

[0030] The technical scheme of the present invention is further elaborated in detail below in conjunction with specific embodiments. The parts not described and disclosed in detail in the following embodiments of the present invention should be understood as the prior art known or should be known to those skilled in the art. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art; unless otherwise specified, the reagents and materials used in the present invention are commercially available.

[0031] Example 1

[0032] A method for detecting Cu 2+ , a colorimetric fluorescent probe TZ6 of BPO, wherein the structural formula of the colorimetric fluorescent probe TZ6 is as follows:

[0033] .

[0034] Example 2

[0035] A method for detecting Cu 2+ , the preparation method of the colorimetric fluorescent probe TZ6 of BPO, the synthesis route is as follows:

[0036] .

[0037] The preparation method comprises the following steps:

[0038] S1: Add 2 mmol potassium carbonate to 20 ml acetonitrile and stir to obtain mixed solution I;

[0039] S2: 2 mmol of 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one and 2 mmol of 2-(chloromethyl)pyridine were mixed and refluxed in mixed solution I for 3-4 h at a temperature of 80°C. After the reaction was completed as monitored by TLC, mixed solution II was obtained;

[0040] S3: Using dichloromethane-methanol system as mobile phase, the mixed solution II is separated by column chromatography to obtain a light yellow product, which is the colorimetric fluorescent probe TZ6. 1 H NMR (400 MHz, CDCl3) δ in ppm: 8.54 (s, 1H), 7.76 (d, J = 1.9 Hz, 1H), 7.27 - 7.26 (m, 2H), 7.20 (t, J = 8.0 Hz, 1H), 7.15(d, J = 2.2 Hz, 1H), 7.11 (d, J = 7.6 Hz, 1H), 6.99 (t, J = 5.8 Hz, 1H), 6.93 (d, J = 7.2 Hz, 1H), 6.74 (d, J = 1.8 Hz, 1H), 3.98 - 3.95 (m, 2H), 3.76 (s,2H), 3.54 (s, 2H), 3.40 (s, 2H), 2.55 (s, 2H), 2.48 (s, 2H), 1.47 - 1.44 (m,3H).

[0041] It should be noted that in the preparation method of the colorimetric fluorescent probe TZ6, the raw material 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one was prepared according to the method disclosed in the literature “A novel benzothiazine-fused coumarin derivative for sensing hypochlorite with high performance” (Dyes And Pigments, Vol. 182 (2020) 108675). The preparation method is briefly described as follows: 2-methoxyphenothiazine is reacted with iodoethane in DMSO solvent to generate an intermediate 10-ethyl-2-methoxy-10H-phenothiazine, which is then reacted with phosphorus oxychloride in DMF solvent to generate an intermediate 10-ethyl-2-methoxy-10H-phenothiazine-3-carboxaldehyde, which is further reacted with aluminum chloride in dichloromethane and hydrolyzed to generate an intermediate 10-ethyl-2-hydroxy-10H-phenothiazine- 3-formaldehyde, and then condensed with diethyl malonate in acetonitrile solvent to generate intermediate 11-ethyl-2-oxo-2,11-dihydropyrano[2,3-b]phenothiazine-3-carboxylic acid ethyl ester, and then hydrolyzed in ethanol solvent under alkaline environment to generate intermediate 11-ethyl-2-oxo-2,11-dihydropyrano[2,3-b]phenothiazine-3-carboxylic acid, and further reacted with N-Boc-piperazine in dichloromethane solvent through N, N′-dimethylaminopyridine and 1-ethyl-(3-dimethylaminopropyl)carbodiimide are condensed to generate the intermediate 4-(11-ethyl-2-oxo-2,11-dihydropyrano[2,3-b]phenothiazine-3-carbonyl)piperazine-1-carboxylic acid tert-butyl ester, which is then de-Bocated with trifluoroformic acid in a dichloromethane solvent to generate 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one.

[0042] Example 3

[0043] A colorimetric fluorescent probe TZ6 for detecting Cu 2+ , BPO application, prepare the colorimetric fluorescent probe TZ6 solution; mix the colorimetric fluorescent probe TZ6 solution and the solution to be detected to obtain a mixed solution.

[0044] Perform fluorescence detection on the mixed solution and observe the fluorescence intensity; if the fluorescence is enhanced, the solution to be tested contains BPO; if the fluorescence is quenched, the solution to be tested contains Cu 2+ .

[0045] Fluorescence intensity and BPO, Cu in the test solution 2+The concentration is proportional to the fluorescence intensity. The BPO and Cu in the test solution can be calculated based on the fluorescence intensity. 2+ The specific calculation method of concentration is known in the art and will not be described in detail here.

[0046] Example 4

[0047] A colorimetric fluorescent probe TZ6 for detecting Cu 2+ , application in BPO, prepare colorimetric fluorescent probe TZ6 solution; mix the colorimetric fluorescent probe TZ6 solution and the solution to be tested to obtain a mixed solution; observe the color change of the mixed solution with naked eyes, if the color of the mixed solution changes from yellow to colorless and transparent, the solution to be tested contains BPO.

[0048] Example 5

[0049] A colorimetric fluorescent probe TZ6 for detecting Cu 2+ , application in BPO, prepare colorimetric fluorescent probe TZ6 solution; mix the colorimetric fluorescent probe TZ6 solution and the solution to be tested to obtain a mixed solution, irradiate the mixed solution with an ultraviolet lamp with a wavelength of 365nm, if the fluorescence color changes from rose red to green, the solution to be tested contains BPO.

[0050] Next, the performance of the colorimetric fluorescent probe TZ6 (abbreviated as: probe TZ6) described in the present invention was tested as follows.

[0051] <Optional>

[0052] 1) The probe TZ6 was dissolved in DMF:PBS (2:8, v:v, 0.1 μM) solvent to prepare a probe TZ6 molecular solution with a concentration of 10 μM; wherein, DMF:PBS (2:8, v:v, 0.1 μM) solvent refers to a mixture of DMF and PBS in a volume ratio of 2:8, and the concentration of PBS is 0.1 μM, the same below;

[0053] 2) Prepare Cu with a concentration of 100 μM 2+ , BPO, ClO - 、Ag + 、Al 3+ , Ba 2+ , Ca 2+ 、Cd 2+ 、Co 2+ Cr 3+ , Fe 2 + , Fe 3+ , Hg 2+ Mg 2+ , Mn 2+ 、Ni 2+ , Pb 2+ , Pd 2+ 、Zn2+ 、HSO3 - 、NO2 - , S 2- 、SO3 2- 、HCO3 - 、H2PO4 - 、F - , Cl - 、SO4 2- 、CH3COO - , H2O2, glycine, lysine, glucose, and lactose solutions were used as test solutions, and the solvent of the test solutions was DMF:PBS (2:8, v:v, 0.1 μM);

[0054] 3) Take several 1 ml portions of DMF:PBS (2:8, v:v, 0.1 μM) solvent, add 1 μl of the probe TZ6 molecule solution prepared in step 1) to each portion, and then add 1 μl of the different ion test solutions prepared in step 2) to each portion to obtain a mixed solution, and perform fluorescence detection on the mixed solution using a fluorescence spectrophotometer.

[0055] Test results such as Figure 2 , 3 Shown by: Figure 2 It can be seen that the fluorescence of probe TZ6 is significantly enhanced only after interacting with BPO, and the peak at 500nm increases significantly, indicating that probe TZ6 has specific selectivity for BPO. At the same time, during the detection process, the color of the solution reacting with probe TZ6 and BPO changes from yellow to colorless and transparent. When irradiated with ultraviolet light at a wavelength of 365nm, the fluorescence color changes from rose red to green, which can be used as a colorimetric fluorescent probe. Figure 3 It can be seen that the probe TZ6 only has 2+ After the interaction, there is obvious fluorescence quenching, and the peaks at 500nm and 605nm are completely quenched, showing that the probe TZ6 is effective for Cu 2+ It has specific selectivity.

[0056] <Anti-interference>

[0057] (I) Competition experiment of probe TZ6 recognizing BPO

[0058] The anti-interference ability of probe TZ6 in detecting BPO was tested by competition experiment of probe TZ6 recognizing BPO in the presence of common metal salt ions.

[0059] The specific experimental process is as follows: 1) The colorimetric fluorescent probe TZ6 was dissolved in a solvent of DMF:H2O=2:8 (volume ratio, the same below) to prepare a probe TZ6 molecular solution with a concentration of 10 μM;

[0060] 2) Take a solvent of DMF:H2O=2:8, and prepare a BPO solution and a solution in which BPO and interfering ions coexist as the test solution. The BPO concentration in the BPO solution is 20uM; the BPO concentration in the solution in which BPO and interfering ions coexist is 20uM, the interfering ion concentration is 50uM, and the interfering ion is Ca 2+ , Fe 2+ , Fe 3+ Mg 2+ 、Zn 2+ 、Na + , KI, NO3 - , CsF, Cl - 、HCO3 - 、H2PO4 - 、SO4 2- 、NO2 - , H2O2, glycine, lactose, glucose and lysine;

[0061] 3) Take several portions of 1 ml of DMF:H2O=2:8 solvent, add 1 μl of the probe TZ6 molecule solution prepared in step 1) to each portion, and then add 1 μl of the test solution prepared in step 2) to each portion to obtain a mixed solution, and perform fluorescence detection on the mixed solution using a fluorescence spectrophotometer.

[0062] Depend on Figure 4 It can be seen that in the solvent of DMF:H2O=2:8, the probe TZ6 reacts with BPO to produce a high-intensity fluorescence value (1 in the figure). The probe TZ6 still maintains a high-intensity fluorescence value when it coexists with other ions and BPO (2-20 in the figure). Figure 2 This indicates that probe TZ6 has strong selectivity and anti-interference properties for BPO.

[0063] (ii) Probe TZ6 recognizes Cu 2+ Competition experiment

[0064] Probe TZ6 recognizes Cu in the presence of common metal salt ions. 2+ Competition experiments were performed to test the detection of Cu by probe TZ6 2+ Anti-interference performance.

[0065] The specific experimental process is as follows: 1) The colorimetric fluorescent probe TZ6 was dissolved in a solvent of DMF:H2O=2:8 to prepare a probe TZ6 molecular solution with a concentration of 10 μM;

[0066] 2) Take DMF:H2O=2:8 solvent and prepare Cu 2+ Solution, Cu 2+ The solution coexisting with interfering ions is used as the test solution. 2+ Cu in solution 2+The concentration is 20uM; Cu 2+ Cu in the solution where interfering ions coexist 2+ The concentration is 20uM, the interfering ion concentration is 50uM, and the interfering ion is Ag. + 、Al 3+ , Ba 2+ , Ca 2+ 、Cd 2+ 、Co 2+ Cr 3+ , Fe 2+ , Fe 3+ , Hg 2+ Mg 2+ , Mn 2+ 、Ni 2 + , Pb 2+ , Pd 2+ 、Zn 2+ 、HSO3 - 、NO2 - , S 2- , CIO - 、SO3 2- 、HCO3 - 、H2PO4 - 、F - , Cl - ;

[0067] 3) Take several portions of 1 ml of DMF:H2O=2:8 solvent, add 1 μl of the probe TZ6 molecule solution prepared in step 1) to each portion, and then add 1 μl of the test solution prepared in step 2) to each portion to obtain a mixed solution, and perform fluorescence detection on the mixed solution using a fluorescence spectrophotometer.

[0068] Depend on Figure 5 It can be seen that in the solvent of DMF:H2O=2:8, the probe itself has a certain fluorescence intensity (1 in the figure), and the probe and Cu 2+ The reaction shows fluorescence quenching (2 in the figure), and the probe is combined with other ions and Cu 2+ The quenching still occurs under the coexistence condition (3-27 in the figure), indicating that the probe TZ6 is sensitive to Cu 2+ It has strong selectivity and anti-interference ability.

[0069] <Fluorescence detection limit>

[0070] (A) Response of probe TZ6 to different concentrations of BPO

[0071] 1) Dissolve the colorimetric fluorescent probe TZ6 in a solvent of DMF:H2O=2:8 to prepare a probe TZ6 molecular solution with a concentration of 10 μM; 2) Take a solvent of DMF:H2O=2:8, add different amounts of BPO to prepare BPO solutions of different concentrations as test solutions; the concentration range of BPO is between 0-25 μM, and the specific concentrations are 0 μM, 2 μM, 4 μM, 6 μM, 10 μM, 14 μM, 18 μM, 20 μM and 25 μM respectively; 3) Take multiple portions of DMF:H2O=2:8 solvent 1 ml, add 1 μl of the probe TZ6 molecular solution prepared in step 1) respectively, and then add 1 μl of the test solution prepared in step 2) respectively to obtain a mixed solution, perform fluorescence detection on the mixed solution with the help of a fluorescence spectrophotometer, and perform linear fitting on the obtained data.

[0072] like Figure 6 As shown, when the concentration of BPO is 0-25uM, the fluorescence intensity is linearly related to the concentration of BPO. Through the linear relationship, it can be calculated that the fluorescence detection limit of probe TZ6 for BPO is 5.29nM.

[0073] (II) Effect of probe TZ6 on different concentrations of Cu 2+ Response

[0074] According to the method of "Probe TZ6 response to different concentrations of BPO", a 10 μM probe TZ6 molecular solution and a test solution were prepared for fluorescence detection, and the obtained data were linearly fitted. The test solution was a Cu solution with a concentration range of 0-70 μM. 2+ The specific concentrations of the solutions are 0uM, 3uM, 6uM, 9uM, 12uM, 15uM, 18uM, 24uM, 27uM, 30uM, 33uM, 36uM, 39uM, 45uM, 51uM, 57uM and 63uM respectively.

[0075] like Figure 7 As shown, in Cu 2+ When the concentration is 0-70uM, the fluorescence intensity is similar to Cu 2+ The concentration of Cu is linearly related to that of TZ6. 2+ The fluorescence detection limit was 25.68 nM.

[0076] In summary, the probe TZ6 is effective for BPO and Cu 2+ The detection limits are extremely low, sufficient to detect trace amounts of BPO and Cu 2+ .

[0077] <Response Time>

[0078] Depend on Figure 8As shown, the reaction time of probe TZ6 with BPO reaches the maximum fluorescence intensity at 10 minutes. The response time is short, enabling detection to be completed within a short time.

[0079] The reaction time of probe TZ6 with Cu 2+ has the maximum quenching intensity at 30 minutes.

[0080] <pH influence>

[0081] Refer to the method of "Response of Probe TZ6 to Different Concentrations of BPO" above to prepare a 10 μM probe TZ6 molecular solution and a test solution, where the test solution is a 50 μM BPO solution; separately take multiple 1 ml portions of the solvent DMF:H2O = 2:8, add 1 μl of the probe TZ6 molecular solution and 1 μl of the test solution to each portion to obtain multiple mixed solutions, adjust the pH of the multiple mixed solutions with sodium hydroxide and hydrochloric acid respectively; test the fluorescence intensity changes at different pH values.

[0082] It can be seen from Fig. 9 that probe TZ6 can maintain a good response within the pH range of 1 - 10 for detecting BPO. Therefore, probe TZ6 can achieve the detection of BPO under both acidic and basic conditions.

[0083] Similarly, refer to the method of "Response of Probe TZ6 to Different Concentrations of BPO" above to prepare a 10 μM probe TZ6 molecular solution and a test solution, where the test solution is a 50 μM Cu 2+ solution; separately take multiple 1 ml portions of the solvent DMF:H2O = 2:8, add 1 μl of the probe TZ6 molecular solution and 1 μl of the test solution to each portion to obtain multiple mixed solutions, adjust the pH of the multiple mixed solutions with sodium hydroxide and hydrochloric acid respectively; test the fluorescence intensity changes at different pH values.

[0084] It can be seen from Fig.10 that probe TZ6 can maintain a good response within the pH range of 4 - 10 for detecting Cu 2+ . Therefore, probe TZ6 can achieve the detection of Cu 2+ under both acidic and basic conditions.

[0085] The above description of the disclosed embodiments enables those skilled in the art to implement or use the present invention. Various modifications to these embodiments will be obvious to those skilled in the art, and the general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown herein, but rather will be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for detecting Cu 2+ , BPO colorimetric fluorescent probe TZ6, characterized in that: The structural formula of the colorimetric fluorescent probe TZ6 is as follows:

2. A method for detecting Cu according to claim 1 2+ , a method for preparing a colorimetric fluorescent probe TZ6 of BPO, characterized in that: The following steps are involved: S1: Add potassium carbonate to acetonitrile to obtain mixed solution Ⅰ; S2: 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one and 2-(chloromethyl)pyridine are refluxed and stirred in the mixed solution I to obtain a mixed solution II; S3: Using a dichloromethane-methanol system as a mobile phase, the mixed solution II is separated by column chromatography to obtain the colorimetric fluorescent probe TZ6.

3. A method for detecting Cu according to claim 2 2+ , a method for preparing a colorimetric fluorescent probe TZ6 of BPO, characterized in that: The ratio of acetonitrile and potassium carbonate added is 10ml:1mmol.

4. A method for detecting Cu according to claim 2 2+ , a method for preparing a colorimetric fluorescent probe TZ6 of BPO, characterized in that: The molar ratio of 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazin-2(11H)-one to 2-(chloromethyl)pyridine is 1:

1.

5. A method for detecting Cu according to claim 2 2+ , a method for preparing a colorimetric fluorescent probe TZ6 of BPO, characterized in that: The ratio of the added amounts of 11-ethyl-3-(piperazine-1-carbonyl)pyrano[2,3-b]phenothiazine-2(11H)-one, 2-(chloromethyl)pyridine and acetonitrile is 1 mmol:1 mmol:10 ml.

6. A method for detecting Cu according to claim 2 2+ , a method for preparing a colorimetric fluorescent probe TZ6 of BPO, characterized in that: In step S2, the reflux stirring time is 3-4 hours.

7. A method for detecting Cu according to claim 2 2+ , a method for preparing a colorimetric fluorescent probe TZ6 of BPO, characterized in that: In step S2, the temperature of reflux stirring is 80°C.

8. A colorimetric fluorescent probe TZ6 as claimed in claim 1 for the preparation of a method for detecting Cu 2+ , application in BPO reagents.

9. The colorimetric fluorescent probe TZ6 according to claim 8 is used in the preparation of the detection of Cu 2+ , the use of BPO in reagents, characterized in that, Mixing the colorimetric fluorescent probe TZ6 as claimed in claim 1 and the solution to be detected to obtain a mixed solution, and performing fluorescence detection on the mixed solution; If the fluorescence is enhanced, the test solution contains BPO; if the fluorescence is quenched, the test solution contains Cu 2+ .

10. The colorimetric fluorescent probe TZ6 according to claim 8 is used in the preparation of the detection of Cu 2+ , the use of BPO in reagents, characterized in that, Mix the colorimetric fluorescent probe TZ6 as claimed in claim 1 and the solution to be detected to obtain a mixed solution, and observe the color of the mixed solution. If the color changes from yellow to colorless and transparent, the solution to be detected contains BPO; Alternatively, the mixed solution is irradiated with an ultraviolet lamp of 365 nm wavelength. If the fluorescent color changes from rose red to green, the test solution contains BPO.

Images