A fluorescent compound of purine and its preparation method and application

Abstract

The present invention discloses a fluorescent complex of copper-purine and its preparation method and application. The fluorescent complex of copper-purine is shown below #imgabs0# The present invention uses a purine ring as a matrix, and its alkaline scaffold has good conjugation properties and exhibits excellent biological and optical properties in fluorescent probes and bioimaging. The fluorescent complex of copper-purine of the present invention can selectively detect Cu 2+ , and then the pesticide residues can be detected, and can be successfully applied to the detection of pesticide residues in organisms. The fluorescent complex of the present invention has the advantages of high sensitivity, low detection cost, simple operation, and real-time detection. It has a wide range of application prospects in the fields of food and agriculture, is easy to promote, and is particularly suitable for the detection of glyphosate and glufosinate in organophosphorus pesticides.

Description

Technical Field

[0001] The present invention relates to the field of fluorescent probe and detection technology thereof, and in particular to a fluorescent compound of purine and a preparation method and application thereof. Background Art

[0002] Glyphosate is a highly effective, low-toxic organophosphorus pesticide that is widely used as a herbicide in rice, corn, wheat, and tea gardens. However, excessive use of glyphosate will produce harmful residues in soil and water environments, which not only pollutes the natural environment, but also easily accumulates in the human body through the food chain, causing serious neurological and respiratory diseases. Glyphosate is also a potential endocrine disruptor that interferes with gene expression, leading to cell cycle interruption and even cell death. The U.S. Environmental Protection Agency has set the maximum concentration of glyphosate in drinking water at 700 μg / L. Glufosinate is a non-selective herbicide that is widely used in agricultural production, forestry, and urban greening. Its main metabolites are often found in plants, soil, water bodies, and marine organisms. Residual glufosinate will acidify the soil and destroy the ecological structure and biodiversity of the soil. According to the International Food Code, the upper limit of glufosinate residues is 8 mg / L. Therefore, there is an urgent need to establish a sensitive and convenient method to accurately determine the presence of glyphosate and glufosinate in food and environmental samples. However, due to the lack of chromophores and amphoteric groups, the detection of glyphosate and glufosinate remains challenging. Traditional analytical methods for detecting organophosphorus pesticides include atomic absorption spectroscopy (AAS), inductively coupled plasma atomic emission spectroscopy (ICP-AES), inductively coupled plasma mass spectrometry (ICP-MS), solid phase extraction high performance liquid chromatography, time-of-flight resonance ionization mass spectrometry, capillary electrophoresis, and X-ray fluorescence. Although traditional detection methods provide reliable analysis results, they still have some inevitable disadvantages, such as high prices, high professional requirements for operation, cumbersome steps, and inconvenience in carrying. Therefore, it is necessary to develop various chemical fluorescence sensors with simple operation, fast response, and high sensitivity for selective detection of pesticide residues.

[0003] Purine and its derivatives are composed of large planar conjugated heterocycles containing 4 nitrogen atoms, which have good coordination ability with various metal ions. Most fluorescent sensors used for organophosphorus pesticide detection are designed and detected using the unique chemical structure of glyphosate. The phosphine group (-PO3H2) and carboxyl group (-COOH) of glyphosate are two ionizable functional groups, and reports have shown that it can effectively bind to transition metal ions. Therefore, it provides a new idea for the detection of organophosphorus pesticides. Summary of the invention

[0004] In order to solve the problems of the prior art, the present invention provides a purine fluorescent compound and its preparation method and application, wherein a purine derivative is used as a fluorescent matrix and 7-(diethylamino)-2-oxo-2H-chromene-3-carboxaldehyde is used as a specific response group. The obtained fluorescent compound has high fluorescence intensity. 2+ ions, the autofluorescence is quenched. After the addition of glyphosate or glufosinate, the fluorescence intensity of the fluorescent complex changes. 2+ The chelating ability is stronger, and then the purine skeleton fluorescent matrix is ​​released, and the fluorescence intensity of the solution is significantly restored. It is suitable for detecting the residues of organophosphorus pesticides and has good application prospects.

[0005] In order to solve the problems of the prior art, the technical solution adopted by the present invention is:

[0006] A fluorescent compound of purine, the structure of which is shown below

[0007] The method for preparing the fluorescent compound of purine comprises the following steps:

[0008] Step 1, dissolve 4-(diethylamino)-2-hydroxybenzaldehyde and diethyl malonate in ethanol, add HCl and piperidine, reflux reaction, after the reaction is completed, cool to room temperature, remove ethanol under reduced pressure, dissolve with NaOH and extract with ethyl acetate, and rotary evaporate to obtain a crude product 7-(diethylamino)-2H-chromen-2-one, which is recorded as intermediate II;

[0009] Step 2, add intermediate II to DMF, add POCl3 dropwise in an ice-water bath, and after the addition is complete, reflux at high temperature for reaction. After the reaction is complete, cool to room temperature, add a large amount of distilled water, and filter out solids to obtain 7-(diethylamino)-2-oxo-2H-chromene-3-carboxaldehyde, which is recorded as intermediate III;

[0010] Step 3, dissolving 6-chloro-9-(naphthalene-1-yl)-8-(thiophene-2-yl)-9H-purine and hydrazine hydrate in ethanol, reflux with stirring, and after the reaction is complete, removing ethanol by rotary evaporation, extracting with chloroform, and spin drying to obtain 6-hydrazine-9-(naphthalene-1-yl)-8-(thiophene-2-yl)-9H-purine, which is recorded as intermediate IV;

[0011] Step 4, dissolving intermediate IV and intermediate III in methanol, reflux with stirring, and after the reaction is complete, cooling to room temperature, filtering out the solid precipitated from the reaction solution to obtain a crude product, and drying the crude product after recrystallization to obtain (E)-7-(diethylamino)-3-((2-(9-(naphthalen-1-yl)-8-(thiophene-2-yl)-9H-purine-6-yl)hydrazono)methyl)-2H-chromen-2-one, which is recorded as fluorescent probe I based on purine hydrazone receptor.

[0012] The basic synthetic route of the reaction is as follows:

[0013]

[0014] As an improvement, in step 1, the reference amounts of 4-(diethylamino)-2-hydroxybenzaldehyde and diethyl malonate are both 5 mmol, and the reflux reaction is refluxed and stirred for 2 h under nitrogen protection.

[0015] As an improvement, the dropping speed in step 2 is 2s / drop.

[0016] As an improvement, 6-chloro-9-(naphthalen-1-yl)-8-(thiophene-2-yl)-9H-purine (0.32 g, 0.8 mmol) and hydrazine hydrate (0.25 g, 4 mmol) in step 3 are dissolved in 10 ml of methanol, refluxed with stirring at 65°C, and after the reaction is complete, the methanol is removed by rotary evaporation, and then extracted with chloroform and dried to obtain a solid compound intermediate IV.

[0017] As an improvement, in step 4, the reference amount of intermediate IV and intermediate III is 1:1.5 (reference amount is the amount of substance), the reflux temperature is 80°C, and reflux stirring is performed for 4 hours.

[0018] The application of the above-mentioned fluorescent purine compounds in the detection of organophosphorus pesticides.

[0019] As an improvement, the organophosphorus pesticide is one of glyphosate or glufosinate ammonium, or a mixture of the two.

[0020] Application of the above-mentioned purine fluorescent compounds in living cell imaging.

[0021] Beneficial effects:

[0022] Compared with the existing technology, the fluorescent compound of purine and its preparation method and application of the present invention have the following specific advantages:

[0023] 1. The present invention adopts the principle of Schiff base reaction, the reaction process is simple, and the operation is easy. The product (E)-7-(diethylamino)-3-((2-(9-(naphthalen-1-yl)-8-(thiophene-2-yl)-9H-purine-6-yl)hydrazono)methyl)-2H-chromene-2-one is a stable solid powder, which is easy to store.

[0024] 2. The preparation process uses purine ring as the matrix, and its basic scaffold has good conjugation properties, showing excellent biological and optical properties in fluorescent probes and bioimaging;

[0025] 3. In Cu 2+In terms of detection, the fluorescent probe exhibits high selectivity and sensitivity, short response time, and can quickly detect copper ions in a portable solid state. It has good portability and stability and is suitable for monitoring Cu in the environment and physiological systems. 2+ The content shows great promise;

[0026] 4. EDTP-Cu 2+ As a fluorescent complex, it can then detect glyphosate and glufosinate in organophosphorus pesticides, and can restore the fluorescence of the quenched complex itself. It can be applied to living cell imaging, which shows that it has excellent prospects in biological detection in the future. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 The nuclear magnetic resonance of the fluorescent probe synthesized in Example 1 1 HNMR spectrum;

[0028] Figure 2 The nuclear magnetic resonance of the fluorescent probe synthesized in Example 1 13 C NMR spectrum;

[0029] Figure 3 is the mass spectrum MS spectrum of the fluorescent probe synthesized in Example 1;

[0030] Figure 4 The fluorescence spectra of the fluorescent probe in a DMSO-H2O (v / v=9:1) solution are shown in Figure 1, where: (a) is the fluorescence selectivity of the fluorescent probe synthesized in Example 1 to various metal ions, and (b) is the fluorescence selectivity of the fluorescent probe synthesized in Example 1 to different concentrations of Cu 2+ The fluorescence spectrum of the response, (c) is the Job-plot spectrum of the complexation of the fluorescent probe synthesized in Example 1 with copper ions, and (d) is the color change of the test paper soaked in the fluorescent probe solution synthesized in Example 1 after being soaked in copper ion solutions of different concentrations;

[0031] Figure 5 In EtOH-H2O (v / v=9:1) solution, EDTP and Cu 2+ The fluorescence images of the standard solution of the fluorescent complex for 13 pesticides were selected specifically;

[0032] Figure 6 EDTP and Cu 2+ Fluorescence titration spectrum curves when the standard solution of the fluorescent complex is added to solutions containing (a) glyphosate or (b) glufosinate at different concentrations;

[0033] Figure 7 For various pesticides to EDTP and Cu 2+ Effect diagram of the standard solution of fluorescent complex on the detection of (a) glyphosate and (b) glufosinate.

[0034] Figure 8 EDTP and Cu 2+ The effect of restoring fluorescence of different types of pesticides in HepG-2 cells detected by standard solution of fluorescent complex. DETAILED DESCRIPTION

[0035] The following examples will enable those skilled in the art to more fully understand the present invention, but are not intended to limit the present invention in any way.

[0036] The experimental methods used in the present invention are conventional methods unless otherwise specified. The materials, reagents, etc. used in the experiments can be obtained from commercial sources unless otherwise specified. All the following reagents used in the examples are commercially available analytically pure or chemically pure.

[0037] Among them, various metal ion solutions in the embodiments are prepared by adding deionized water to chloride salt chemical reagents with a purity of more than 99%, such as anhydrous zinc chloride, anhydrous ferric chloride, etc.

[0038] Example 1

[0039] 1. Preparation of the intermediate 7-(diethylamino)-2H-chromen-2-one of formula II

[0040] In a 10mL round-bottom flask, add 4-(diethylamino)-2-hydroxybenzaldehyde (626mg, 5mmol) and diethyl malonate (606mg, 5mmol) and dissolve in ethanol, add hydrochloric acid and piperidine, and then heat and stir at 110°C for 2h. After the reaction is completed, cool to room temperature. Remove ethanol under reduced pressure, then dissolve all the crude product in 20mL of saturated sodium hydroxide aqueous solution, and extract the mixture three times with ethyl acetate, then remove water with saturated brine, and dry to obtain the crude product 7-(diethylamino)-2H-chromene-2-one It was recorded as intermediate II (927.3 mg, yield 82%). ESI-MS m / zcalcd for C 13 H 15 NO2[M+H] + 218.1, found 219.1.

[0041] 2. Preparation of the intermediate 7-(diethylamino)-2-oxo-2H-chromene-3-carbaldehyde of formula III

[0042] In a 25mL round-bottom flask, 2ml of DMF was added dropwise to 100μL of POCl3 at 0°C, stirred under nitrogen protection for 15min, then stirred at room temperature for 30min, then 7-(diethylamino)-2H-chromene-2-one (175mg, 0.806mmol) was dissolved in 2mL of DMF to form a scarlet suspension, which was slowly added dropwise to a mixed solution of phosphorus oxychloride and DMF. After complete addition, the reaction solution was stirred at 70°C for 4h. After heating, 50mL of ice water was added after cooling to room temperature, and the intermediate of formula III was obtained by suction filtration, which was a yellow solid 7-(diethylamino)-2-oxo-2H-chromene-3-carboxaldehyde. Recorded as Intermediate III (104.3 mg, yield 53%). 1 H NMR(400MHz,Chloroform-d)δ10.11(s,1H),8.24(s,1H),7.40(d,J=9.0Hz,1H),6.63(dd,J= 9.0,2.5Hz,1H),6.47(d,J=2.4Hz,1H),3.47(q,J=7.2Hz,4H),1.25(t,J=7.2Hz,6H).ESI-MS m / z:[MH] + calcd.for C 14 H 15 NO3245.10, found 246.0.

[0043] 3. Preparation of Formula IV intermediate 6-hydrazino-9-(naphthalen-1-yl)-8-(thiophene-1-yl)-9H-purine

[0044] In a 25 ml round-bottom flask, 6-chloro-9-(naphthalene-1-yl)-8-(thiophene-2-yl)-9H-purine (0.32 g, 0.80 mmol) and hydrazine hydrate (0.25 g, 4 mmol) were dissolved in 5 ml of methanol, stirred under reflux at 65 ° C, and the reaction was monitored by TLC plate until the reaction was completed. Then, the reaction solution was cooled to room temperature, the solvent was removed under reduced pressure, and extracted several times with chloroform. The organic phases were combined, dried, and spin-dried to obtain light yellow 6-hydrazine-9-(naphthalene-1-yl)-8-(thiophene-2-yl)-9H-purine,

[0045] Recorded as intermediate IV (0.23 g, yield 70%). ESI-MS m / z calcd for C 19 H 14 N6S[M+H] + 358.1, found 359.2. 1H NMR(400MHz, DMSO-d6)δ8.26(d,J=8.1Hz,1H),8.18–8.04(m,3H),7.81–7.69(m,3H),7.63–7.52(m ,3H),7.46(t,J=7.7Hz,1H),7.07(d,J=8.4Hz,1H),6.84(dt,J=7.2,4.5Hz,1H),6.58–6.53(m,1H). 13 C NMR(101MHz,DMS O-d6)δ155.0,153.0,144.1,133.9,131.6,131.1,130.5,130.1,129.2,12 8.5,128.0,127.7,127.7,127.4,127.0,126.0,121.8,117.6,79.2.ESI-MS m / z:[M+H] + calcd.for C 19 H 15 N6S 359.1, found 359.0.

[0046] 4. Preparation of fluorescent compounds based on purine hydrazone receptors

[0047] In a 50 mL round-bottom flask, intermediate IV (100 mg, 0.279 mmol) and intermediate III (103 mg, 0.419 mmol) were added and dissolved in 5 mL of ethanol. After the solid was dissolved, the mixture was refluxed and stirred at 80 ° C for 4 h. After the reaction was complete, the reactant was cooled to room temperature, filtered, and the solid was washed with ice ethanol to obtain a yellow powder (E)-7-(diethylamino)-3-((2-(9-(naphthalen-1-yl)-8-(thiophene-2-yl)-9H-purine-6-yl)hydrazono)methyl)-2H-chromen-2-one. It is recorded as fluorescent probe I based on purine hydrazone receptor (134.4 mg, yield 82%), wherein the mass spectrometry MS spectrum detected is as follows Figure 3 shown.

[0048] Nuclear magnetic resonance based on fluorescent probe compounds of purine hydrazone receptors 1 H-NMR spectrum: 1HNMR(400MHz,Chloroform-d)δ8.61(s,1H),8.50(s,1H),8.37(s,1H),8.13(d,J=7.9 Hz,1H),8.00(d,J=8.2Hz,1H),7.72–7.61(m,2H),7.55(t,J=7.6Hz,1H),7.42(t,J=9. 3Hz,2H),7.28(s,1H),7.24(s,1H),6.76(dd,J=5.0,3.8Hz,1H),6.69–6.58(m,2H),6. 50(d,J=2.5Hz,1H),3.44(q,J=7.1Hz,4H),1.23(t,J=7.1Hz,6H),0.94–0.71(m,1H), see Figure 1 .

[0049] Nuclear magnetic resonance based on fluorescent probe compounds of purine hydrazone receptors 13 C-NMR spectrum: 13 C NMR (101 MHz, Chloroform-d) δ 162.21, 157.26, 154.20, 151.73, 150.63, 147.07, 140.62, 139.63, 134.93, 131.82, 131.33, 131.21, 130.83, 130.61, 129.33, 128.93, 128.85, 128.46, 128.12, 127.54, 126.04, 122.49, 118.77, 113.68, 109.95, 109.33, 97.59, 45.31, 12.83, see Figure 2 .

[0050] Example 2

[0051] The purine hydrazone receptor-based fluorescent probe I synthesized in Example 1 was dissolved in DMSO to prepare a 1.0 mM probe stock solution. 2+ ,Al 3+ ,Cr 3+ ,Mn 2+ Mg 2+ ,Pb 2+ ,K + ,Ca 2+ ,Fe 2+ ,Na + ,Ag + ,Cd 2+ ,Pd 2+ ,Fe 3+ ,Cu 2+ ,Co + ,Ba 2+and Sn 2+ ,) Use distilled water to prepare a 3.0mM metal ion stock solution. Take 3mL of the blank stock sample DMSO-H2O (v / v=9:1) prepared in advance in a 5mL centrifuge tube, add 30μL of the probe stock solution and 50μL of the metal ion stock solution, and let it stand for 1 hour. Tested with a fluorescence spectrometer and a UV spectrophotometer, the maximum excitation wavelength was 466nm and the maximum emission wavelength was 534nm.

[0052] Figure 4 (a) is a diagram showing the selection of various metal ions by the fluorescent probe I based on the purine hydrazone receptor. 30 μL of the probe stock solution was added to 3 mL of the blank stock sample DMSO-H2O (v / v=9:1), and then 50 μL of various metal ion stock solutions were added and allowed to stand for 1 hour. The fluorescence intensity of these sample solutions was tested by fluorescence spectroscopy. The results showed that the fluorescence of the sample solution with copper ions added at 534 nm was turned off, while there was no obvious change when other ions were added.

[0053] Figure 4 (b) is a fluorescent probe based on purine hydrazone receptor I and Cu 2+ Complexation ratio diagram. Add a certain amount of probe stock solution and a certain amount of copper ion stock solution to 3mL of blank stock sample DMSO-H2O (v / v=9:1). Add the two stock solutions in a ratio of 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2, 9:1. After standing for 1 hour, test with a UV spectrophotometer. Draw a curve with the ratio of ions to the total concentration as the horizontal axis and the absorption peak intensity of the above samples at λ=534nm as the vertical axis. Figure 4 In (b), we can see that the maximum absorption intensity appears at 0.5, from which we can infer that the fluorescent probe is closely related to Cu 2+ The binding ratio of ions is 1:1.

[0054] Figure 4 (c) is a fluorescent probe based on purine hydrazone receptor I for Cu 2+ Fluorescence titration spectra of different concentrations of Cu 2 + Fluorescence intensity spectrum when it exists. Add 30μL of probe stock solution and 0-60μL (0, 1, 2, 3...15, 20, 25...60μL) of copper ion stock solution to 3mL of blank stock sample DMSO-H2O (v / v=9:1), let it stand for a period of time and then test. The final spectrum shows that: with the continuous increase of ion concentration, the intensity of fluorescence gradually weakens, and remains stable after 55μL. At the same time, the color of the probe solution under 365nm light also changes from bright yellow to dark red. The above phenomenon shows that the fluorescent probe of the present invention is effective for Cu 2+Have high sensitivity.

[0055] Figure 4 (d) shows the color change of the test paper after being soaked in the probe stock solution and different concentrations of ion stock solutions under 365nm light. First, 5 filter papers were soaked in 1mM probe stock solution. After soaking for half an hour, they were taken out and dried. Then, the 5 filter papers were soaked in 0, 0.05, 0.1, 0.5 and 1.0mM Cu 2+ The solution was taken out and dried after 30 minutes. Under the light of 365nm, the color of the filter paper was observed to change from bright yellow fluorescence to weak dark red. It can be seen that the fluorescent probe of the present invention can be made into a portable test strip for monitoring Cu 2+ The above phenomena all prove that the probe has high selectivity for copper ions.

[0056] Figure 5 This is a diagram showing the selection of various pesticides by the fluorescent probe I based on purine hydrazone receptor. 2+ 30 μL of different pesticide stock solutions (glyphosate, glufosinate, paraquat, imidacloprid, thiophanate-methyl, atrazine, avermectin, triadimefon, phoxim, diazinon, thiamethoxam, fenthion) were added to the sample stock solution EtOH-H2O (v / v=9:1) and left to stand for 1 hour. The fluorescence intensity of these sample solutions was tested by fluorescence spectroscopy. The results showed that the addition of glyphosate and glufosinate samples at 534 nm partially restored the fluorescence that was originally closed, while the addition of other pesticide stock solutions did not cause significant changes.

[0057] Figure 6 The fluorescence titration spectra of the fluorescent probe I based on the purine hydrazone receptor for glyphosate and glufosinate pesticides are shown, that is, the fluorescence intensity spectra in the presence of different concentrations of glyphosate or glufosinate. 30 μL of EDTP-Cu was added to 3 mL of blank stock sample EtOH-H2O (v / v=9:1). 2+ The sample stock solution and 0-60 μL (0, 1, 2, 3...60 μL) of pesticide stock solution were left to stand for a period of time and then tested. The resulting spectrum showed that as the concentration of the pesticide continued to increase, the intensity of the fluorescence gradually became stronger. At the same time, the color of the probe solution under 365 nm light gradually changed from dark red to yellow. The above phenomenon shows that the fluorescent probe I based on the purine hydrazone receptor of the present invention has high sensitivity to glyphosate and glufosinate in pesticides.

[0058] Figure 7 Figure 2 is the interference diagram of other different pesticide stock solutions on the detection of glyphosate and glufosinate by the purine hydrazone receptor-based fluorescent probe I. (a) is in 3 mL of EDTP-Cu 2+30 μL of glyphosate stock solution was added to the stock sample EtOH-H2O (v / v=9:1) in sequence, and then 50 μL of various interfering pesticide stock solutions were added to each blank sample. After standing for 1 hour, the fluorescence intensity was tested and compared with the fluorescence intensity of the sample solution with only glyphosate added; (b) in 3 mL of EDTP-Cu 2+ 30 μL of glufosinate stock solution was added to the stock sample EtOH-H2O (v / v=9:1) in sequence, and then 50 μL of various interfering pesticide stock solutions were added to each blank sample. The fluorescence intensity was tested after standing for 1 hour and compared with the fluorescence intensity of the sample solution with only glufosinate added. Figure 7 It can be seen that other ions can hardly cause a significant change in the original fluorescence intensity. Therefore, the selection of glyphosate and glufosinate by the fluorescent probe I based on purine hydrazone receptor of the present invention will not be interfered by other coexisting pesticides.

[0059] The practical application of monitoring copper ions and organophosphorus pesticides in living cells was studied, and HepG2 cells were used for fluorescence cell imaging experiments. First, the purine hydrazone receptor-based fluorescent probe I (2×10 -5 M) dissolved in DMSO, Cu 2+ Ions (1×10 -4 M) Prepare a solution in blank culture medium. Then, add 500 μL of cell suspension and 10 μL of Cu to well 1 and well 2, respectively. 2+ Solution. After that, the cells were incubated at 37°C for 30 minutes and then washed three times with PBS. Finally, 10 μL of the probe solution was added to well 1 and further incubated at 37°C for 50 minutes and washed three times with PBS. Finally, the cells were washed three times with PBS solution and then placed on a culture dish and fluorescence images were taken at room temperature at the excitation wavelength. After that, the cells were washed with phosphate buffered saline (PBS) and then incubated with glyphosate and glufosinate (50 μM, dissolved in 50 μM DMSO) for 1 hour. Then, EDTP-Cu 2+ (20 μM) was added to each well. After washing the cells three times with PBS, fluorescence images were acquired using a fluorescence microscope. Figure 8 shown.

[0060] from Figure 8 It can be seen that the purine hydrazone receptor fluorescent probe I has strong fluorescence in cells. 2+ After the addition of ion stock solution, the fluorescence of the probe was quenched due to the formation of fluorescent complexes. However, after adding glyphosate or glufosinate ammonium to the cells, its fluorescence was significantly restored.

[0061] The above is only a preferred specific implementation manner of the present invention, and the protection scope of the present invention is not limited thereto. Any simple change or equivalent replacement of the technical solution that can be obviously obtained by any technician familiar with the technical field within the technical scope disclosed in the present invention falls within the protection scope of the present invention.

Claims

1. A fluorescent compound of purine, characterized in that: The structure of the fluorescent compound of purine is shown below .

2. A method for preparing a fluorescent compound of purine according to claim 1, characterized in that: The following steps are involved: Step 1, dissolve 4-(diethylamino)-2-hydroxybenzaldehyde and diethyl malonate in ethanol, add HCl and piperidine, reflux reaction, after the reaction is completed, cool to room temperature, remove ethanol under reduced pressure, dissolve with NaOH and extract with ethyl acetate, and rotary evaporate to obtain a crude product 7-(diethylamino)-2H-chromen-2-one, which is recorded as intermediate II; Step 2, add intermediate II to DMF, add POCl3 dropwise in an ice-water bath, and after the addition is complete, reflux at high temperature for reaction. After the reaction is complete, cool to room temperature, add a large amount of distilled water, and filter out solids to obtain 7-(diethylamino)-2-oxo-2H-chromene-3-carboxaldehyde, which is recorded as intermediate III; Step 3: 6-chloro-9-(naphthalen-1-yl)-8-(thiophen-2-yl)-9 H -Purine and hydrazine hydrate were dissolved in methanol, refluxed and stirred, and after the reaction was complete, methanol was removed by rotary evaporation, and then extracted with chloroform and dried to obtain 6-hydrazino-9-(naphthalen-1-yl)-8-(thiophene-2-yl)-9 H -purine, denoted as intermediate IV; Step 4, dissolving intermediate IV and intermediate III in methanol, reflux with stirring, and after the reaction is complete, cooling to room temperature, filtering the reaction solution to obtain a solid precipitated to obtain a crude product, and drying the crude product after recrystallization to obtain (E)-7-(diethylamino)-3-((2-(9-(naphthalen-1-yl)-8-(thiophen-2-yl)-9H-purin-6-yl)hydrazono)methyl)-2H-chromen-2-one, which is recorded as fluorescent probe I based on purine hydrazone receptor.

3. The method for preparing a fluorescent compound of purine according to claim 2, characterized in that: In step 1, the reference amounts of 4-(diethylamino)-2-hydroxybenzaldehyde and diethyl malonate are both 5 mmol, and the reflux reaction is stirred under reflux for 2 h under nitrogen protection.

4. The method for preparing a fluorescent compound of purine according to claim 2, characterized in that: The dropping speed in step 2 is 2s / drop.

5. The method for preparing a fluorescent compound of purine according to claim 2, characterized in that: In step 3, 6-chloro-9-(naphthalen-1-yl)-8-(thiophen-2-yl)-9 H -Purine and hydrazine hydrate were dissolved in 5 ml of methanol, refluxed and stirred at 65°C. After the reaction was complete, methanol was removed by rotary evaporation, and then extracted with chloroform and dried by rotary drying to obtain solid compound intermediate IV.

6. The method for preparing a fluorescent compound of purine according to claim 2, characterized in that: In step 4, the reference amount of intermediate IV and intermediate III is 1:1.5, the reflux temperature is 80°C, and reflux stirring is performed for 4 hours.

7. Use of the fluorescent compound of purine according to claim 1 in detecting organophosphorus pesticides, wherein the application is a non-disease diagnosis method, characterized in that: The organophosphorus pesticide is one of glyphosate and glufosinate ammonium or a mixture of the two.

8. Use of the purine-based fluorescent compound according to claim 1 in living cell imaging, wherein the application is a non-disease diagnosis method.

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