阅读说明：本技术 一种可视化快速、灵敏检测炭疽杆菌的试剂及其检测方法 (Reagent for visual, rapid and sensitive detection of bacillus anthracis and detection method thereof ) 是由 何炜 卢先林 兰婷 田秦秦 李明华 张邦乐 于 2021-01-11 设计创作，主要内容包括：本发明涉及一种实现炭疽杆菌生物标志物二吡啶甲酸可视化检测的Eu～(3+)-小分子配合物传感器及其制备方法。将邻苯二酚类化合物与Eu～(3+)进行螯合,得到具有比色和荧光双响应的配合物,通过二次吡啶甲酸与配合物反应发生的光物理学性质改变,实现对二吡啶甲酸的快速可视化检测。本发明配合物具有合成简单、检测快速的特点,对目标检测物有很高的灵敏度及选择性,进而实现裸眼监测二吡啶甲酸动态水平变化,并可负载于试纸上,成为快速可视化检测炭疽杆菌的优异工具,具有广泛的应用前景。(The invention relates to Eu for realizing visual detection of anthrax bacillus biomarker dipyridyl formic acid 3+ -small molecule complex sensors and methods of making the same. Reacting catechol compound with Eu 3+ Chelating to obtain a complex with colorimetric and fluorescent dual responses, and realizing rapid visual detection of the dipicolinic acid through photophysical property change generated by the reaction of the secondary picolinic acid and the complex. The complex has the characteristics of simple synthesis and quick detection, has high sensitivity and selectivity on a target detection object, further realizes the monitoring of the dynamic level change of the dipicolinic acid by naked eyes, can be loaded on test paper, and becomes the quick visual detection of the anthraxExcellent tools of bacillus, and wide application prospect.)

1. A detection reagent, characterized in that: the reagent is a complex formed by a compound shown in a structural formula (I) and trivalent europium ions,

and R is selected from hydrogen, hydroxyl, amino, nitro, halogen, C1-C4 alkyl and C1-C4 alkoxy.

2. The detection reagent according to claim 1, wherein: r is selected from hydrogen, hydroxyl and amino.

3. The detection reagent according to claim 1, wherein the complex is prepared by a method comprising: dissolving soluble trivalent europium salt and a compound shown in a structural formula (I) in water and drying.

4. The detection reagent according to claim 3, wherein: the molar ratio of the trivalent europium salt to the compound shown in the structural formula (I) is as follows: 1:100 to 100: 1.

5. The detection reagent according to claim 4, wherein: the molar ratio of the trivalent europium salt to the compound represented by the structural formula (I) is preferably 1:10 to 10: 1.

6. The method for preparing a detection reagent according to claim 1, wherein: dissolving soluble trivalent europium salt and a compound shown in a structural formula (I) in water and drying.

7. A sensor comprising the complex of claim 1.

8. The sensor of claim 7, wherein: the sensor is characterized in that the complex is loaded on a carrier, and the carrier comprises paper, a glass sheet, a plastic sheet and a metal sheet.

9. The sensor of claim 7, wherein: the sensor can detect the concentration range of pyridine-2, 6-dicarboxylic acid from 0.1 mu M to 100 mu M.

10. The use of the complex of claim 1 for detecting Bacillus anthracis.

Technical Field

The invention relates to a reagent for visually, rapidly and sensitively detecting bacillus anthracis and a detection method thereof, belonging to the technical field of luminescence.

Background

Bacillus anthracis is a gram-positive spore bacterium and belongs to aerobic bacteria. Anthrax is a major disease of herbivores and can cause infection through spores in the soil and on the coat. Anthrax spores can indirectly infect humans through the respiratory, digestive and skin passages of humans, with cutaneous cellulitis being the most common. As the anthrax spores have extremely strong resistance in the environment, the anthrax spores are always listed as first biological warfare agents in various countries in the military, and pyridine-2, 6-dicarboxylic acid (DPA) is used as a main component of the bacillus anthracis shell layer and approximately occupies 5-15% of the mass of the bacillus anthracis, so that the dynamic level of the bacillus anthracis can be indirectly detected by visually and quantitatively detecting the content of the DPA.

Currently, methods for detecting bacillus anthracis spores mainly include biological methods and chemical methods. The biological analysis method mainly includes polymerase chain reaction and immunoassay, and the chemical method includes high performance liquid chromatography, capillary electrophoresis, electrochemical method, surface enhanced Raman spectroscopy, etc. Although these methods have their own advantages, these methods still have the disadvantages of long analysis time, complicated operation, high cost, etc., and cannot achieve the purpose of rapid detection. In recent years, the unique luminescence characteristics of trivalent lanthanide ions, such as large stokes shift, narrow emission band, excellent fluorescence lifetime, and the like, are widely applied to the preparation of photosensitive materials. At present, researches on detection of pyridine-2, 6-dicarboxylic acid by a fluorescence method are increasing, however, fine nanoparticles or other nano means are often needed, and complicated synthesis and purification steps are often involved, so that the reaction period is long, and the synthesis cost is high. Therefore, the search for a novel detection method which is simple to synthesize, green, efficient, low in cost and capable of being popularized in a large scale is still a key field of research.

Disclosure of Invention

The invention aims to provide a reagent for efficiently, visually, specifically and selectively detecting a bacillus anthracis marker pyridine-2, 6-dicarboxylic acid and a detection method thereof aiming at the defects in the current technology for detecting pyridine-2, 6-dicarboxylic acid.

The invention is realized as follows:

a reagent for visual, rapid and sensitive detection of Bacillus anthracis is a complex formed by a compound shown in a structural formula (I) and trivalent europium ions,

the R is selected from hydrogen, hydroxyl, amino, nitro, halogen, C1-C4 alkyl and C1-C4 alkoxy, and is preferably hydrogen and hydroxyl.

The preparation method of the complex comprises the following steps: dissolving soluble trivalent europium salt and a compound shown in a structural formula (I) in water and drying. The molar ratio of the trivalent europium salt to the compound shown in the structural formula (I) is as follows: 1:100 to 100:1, preferably 1:10 to 10: 1.

The sensor for detecting the bacillus anthracis is obtained by loading a complex formed by a compound shown in a structural formula (I) and trivalent europium ions on a carrier, wherein the complex is loaded on the carrier, the carrier comprises paper, glass sheets, plastic sheets, metal sheets and the like, and the sensor can detect the concentration range of pyridine-2, 6-dicarboxylic acid to be 0.1-100 mu M.

Application of a complex formed by a compound shown in a structural formula (I) and trivalent europium ions in detecting bacillus anthracis.

The inventor researches and discovers that Eu is in the presence of o-phenylenediphenol Schiff base compound shown in the structural formula (I)³⁺Fluorescence quenching of complex o-phenylenediphenol Schiff base ligand complexes, since numerous hydroxyl groups of the o-phenylenediphenol Schiff base compounds can preferentially react with Eu³⁺Coordination, causing the rare earth complex to dissociate. The invention utilizes a plurality of hydroxyl groups of o-catechol Schiff base compounds to preferentially coordinate with rare earth ions, but can not sensitize rare earth ions Eu³⁺Luminescence to cause fluorescence quenching, followed by addition of rare earth ion Eu³⁺Pyridine-2, 6-dicarboxylic acid with stronger coordination, sensitized rare earth ions emit light and have strong fluorescenceThe detection limit is respectively 8.3 nM and 10.4 nM, and the fluorescence intensity at 615nM can be obviously enhanced along with the gradual increase of the concentration of the pyridine-2, 6-dicarboxylic acid; the interference of other amino acids and aromatic compounds can be eliminated, and in an anti-interference experiment, the sensor only recognizes pyridine-2, 6-dicarboxylic acid (DPA) under the interference of nicotinamide adenine binuclear glyconic acid, phthalic acid, isophthalic acid, L-glutamic acid, L-aspartic acid, 2-picolinic acid, nicotinic acid, isonicotinic acid, benzoic acid, terephthalic acid and the like, and simultaneously has good anti-interference capability under the coexistence of 5 times of interferents, so that the purpose of specific detection is achieved. The rare earth ion composite organic small molecular complex realizes the detection of pyridine-2, 6-dicarboxylic acid with simple operation and low cost, and achieves the purposes of visualization, high sensitivity, good selectivity and strong anti-interference capability.

The invention has the beneficial effects that: (1) the reported method for detecting pyridine-2, 6-dicarboxylic acid by a fluorescence method needs to prepare accurate nanoparticles or other nanometer means, and complicated synthesis and purification steps lead to long ligand preparation period and high synthesis cost. The invention can achieve the purpose of high-sensitivity detection only by simple mixing in the aqueous solution, has simple and convenient operation, greenness, high efficiency and low cost, and can realize the rapid and accurate analysis of the level change of the pyridine-2, 6-dicarboxylic acid by using visual observation and simple fluorescence intensity comparison. (2) The selectivity experiment shows that: phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, L-glutamic acid, L-aspartic acid, nicotinamide adenine binuclear glycine and the like exist in the system, and the complex only enables the fluorescence intensity to generate obvious change and color change visible to naked eyes under the action of pyridine-2, 6-dicarboxylic acid, so that the method has good selectivity. Meanwhile, the pyridine-2, 6-dicarboxylic acid solution is added into the interfering substance solution by 5 times, and the fluorescence intensity is basically consistent with that of the pyridine-2, 6-dicarboxylic acid solution with the same concentration when the pyridine-2, 6-dicarboxylic acid solution is independently added, so that the detection of the complex on the pyridine-2, 6-dicarboxylic acid is not interfered even under the coexistence of the interfering substance amino acid and the aromatic ligand, and the selectivity is excellent. (3) The invention can realize the rapid detection of trace pyridine-2, 6-dicarboxylic acid, the detection limit can respectively reach 8.3 nM and 10.4 nM, and the infection concentration is four orders of magnitude lower than that of anthracnose.

Drawings

FIG. 1 is a spectrum of light emission of the host solution of example 2 without pyridine-2, 6-dicarboxylic acid and with 100. mu.M pyridine-2, 6-dicarboxylic acid;

FIG. 2 is a linear fit graph of the bulk solution of example 2 with the concentration of pyridine-2, 6-dicarboxylic acid as the abscissa and the fluorescence intensity at 615nm as the ordinate during the fluorescence titration;

FIG. 3 is a bar graph of the fluorescence intensity of the host solution at 615nm for each acid type and for the pyridine-2, 6-dicarboxylic acid containing host solution at 5 times the interfering substance in example 2. (1-11 are sequentially phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, L-glutamic acid, L-aspartic acid, niacinamide adenine binuclear glycine, rebuke pyridine-2, 6-diacid);

FIG. 4 is a spectrum of light emission of the host solution of example 3 without pyridine-2, 6-dicarboxylic acid and with 100. mu.M pyridine-2, 6-dicarboxylic acid;

FIG. 5 is a graph of a linear fit of the bulk solution of example 3 during fluorescence titration with pyridine-2, 6-dicarboxylic acid concentration as the abscissa and fluorescence intensity at 615nm as the ordinate;

FIG. 6 is a bar graph of the fluorescence intensity of the host solution at 615nm for each acid and the bar graph of the host solution containing pyridine-2, 6-dicarboxylic acid in the 5-fold interferent solution of example 3. (1-11 are sequentially phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, L-glutamic acid, L-aspartic acid, nicotinamide adenine binuclear glycine, rebuke pyridine-2, 6-diacid).

Detailed Description

In order to illustrate the present invention more clearly, the following examples are given without any limitation to the scope of the invention.

The technical scheme of the invention is as follows:

an aqueous solution for visual, rapid and sensitive detection of a bacillus anthracis marker pyridine-2, 6-dicarboxylic acid comprises the following components:

the application method of the aqueous solution for visually detecting the anthrax bacillus marker pyridine-2, 6-dicarboxylic acid mainly comprises the following steps of cutting filter paper into the size of a pH test strip, soaking the pH test strip in the solution for 30 min, and drying the pH test strip at 50 ℃ to obtain the test strip for detecting the anthrax bacillus marker pyridine-2, 6-dicarboxylic acid;

immersing the filter paper strip into a solution to be measured, standing for 0.5-1 second, then drawing out, and observing the color under sunlight; if the DC-1 shows blue-purple color, the solution contains pyridine-2, 6-dicarboxylic acid, and if the DC-2 shows blue color, the solution contains pyridine-2, 6-dicarboxylic acid; the absence of a red or blue color means that the solution does not contain pyridine-2, 6-dicarboxylic acid.

Example 1

(1) The preparation routes of catechol Schiff base DC-1 (R = H) and DC-2 (R = OH) are shown as the following formulas,

the preparation method of the DC-1 comprises the following steps:

in a dry 50 mL two-necked round bottom flask, 276 mg (2 mmol) of 3, 4-dihydroxybenzaldehyde, 152 mg (1 mmol) of 3, 5-diaminobenzoic acid and 15 mL of methanol (chromatographic grade) were added in this order, and the solid was dissolved with stirring. And transferring the reaction bottle into an oil bath kettle, heating to 85 ℃, and carrying out reflux reaction for 3 hours to change the reaction liquid from light yellow to red solid turbidity. After the reaction, the reaction mixture was filtered, and the filter cake was washed with ethyl acetate and DCM for several purifications and dried to obtain 391 mg of a red solid. Yield: 98 percent. Melting point:> 200℃；IR (KBr): ν3410, 3074, 2939, 2831, 1662, 1562, 1450, 1385, 1288, 1196, 1165, 1119, 1003, 868, 813, 748, 621 cm-1；¹H NMR (400 MHz, DMSO-d6) δ: 6.86 (d, J = 7.6 Hz, 2 H), 7.24 (d, J = 7.6 Hz, 3 H), 7.43 (s, 2 H), 7.54 (s, 2 H), 8.50 (s, 2 H)；¹³C NMR (100 MHz, DMSO-d6 ) δ: 167.5, 162.0, 152.6, 150.1, 146.4, 132.6, 129.3, 124.9, 123.4, 116.0, 114.8 ppm；MS [m/z, ESI+] = 415 [M+Na]⁺。

the preparation method of the DC-2 comprises the following steps:

in a dry 50 mL two-necked round bottom flask, 308 mg (2 mmol) of 2,3, 4-trihydroxybenzaldehyde, 152 mg (1 mmol) of 3, 5-diaminobenzoic acid and 15 mL of methanol (chromatographic grade) are added in this order, and the solid is dissolved with stirring. And transferring the reaction bottle into an oil bath kettle, heating to 85 ℃, and carrying out reflux reaction for 3 hours to change the reaction liquid from a light yellow solution into a red solid to be turbid. After the reaction, the reaction mixture was filtered, and the filter cake was washed with methanol and DCM for several purifications and dried to give 410 mg of a red solid. Yield: 96 percent. Melting point:> 200℃；IR (KBr): ν3394, 3340, 3298, 3070, 3004, 2924, 2847, 1701, 1627, 1593, 1524, 1458, 1377, 1277, 1231, 1153, 1142, 1080, 991, 872, 802, 768, 667 cm-1；¹H NMR (400 MHz, DMSO-d6) δ: 6.46 (d, J = 8.4, 2 H), 7.02 (d, J = 8.8 Hz, 2 H), 7.66 (s, 1 H), 7.74 (s, 2 H), 8.94 (s, 2 H)。

according to a similar synthesis method, a ligand with R being amino, nitro, halogen, ethyl and methoxy can be synthesized.

Example 2

(1) 19.0 mg of DC-1 was weighed into a 100mL beaker, 19.6 mL of water was added, stirred at room temperature for 50 minutes until completely dissolved and uniformly dispersed, and then 400. mu.L of 0.1M EuCl was added thereto₃·H₂The aqueous solution was stirred for 30 minutes and the resulting solution was recorded as a host solution.

(2) 3mL of the bulk solution was taken in a quartz cell, and the fluorescence emission spectrum of the blank was measured 11 times at an excitation wavelength of 280 nm. Solutions of pyridine-2, 6-dicarboxylic acid (DPA) at different concentrations were added with a 1. mu.L sample injector (fluorescence titration). The fluorescence test adopts a OmniFluo900 series fluorescence spectrometer of Beijing Zhuoli Han light, uses a 75W xenon lamp as an excitation light source, and is provided with an excitation monochromator, an emission monochromator, a single photon counter and a refrigeration type photomultiplier. The fluorescence emission spectrum was measured with 280nm as the maximum excitation wavelength and 615nm as the maximum emission wavelength.

Subsequently, we carried out a series of examples, and measured a series of fluorescence data by gradually increasing the concentration of the pyridine-2, 6-dicarboxylic acid aqueous solution in step (2), showing the corresponding change in fluorescence. FIG. 1 is a graph showing the emission spectra of the host solution without pyridine-2, 6-dicarboxylic acid (black) and after addition of 100. mu.M pyridine-2, 6-dicarboxylic acid (red).

FIG. 2 is a linear fit graph of the pyridine-2, 6-diacid concentration as abscissa and the fluorescence intensity at 615nm as ordinate of the host solution during the fluorescence titration. Linear fitting was performed with pyridine-2, 6-dicarboxylic acid concentration as abscissa (concentration range 0-100. mu.M) and fluorescence intensity at 615nm as ordinate. As the concentration of pyridine-2, 6-dicarboxylic acid increases,I ₆₁₅the fluorescence intensity also gradually increases and shows a good linear relationship in the concentration range of 5-35 mu M, R²=0.996, detection limit of 8.3 nM. In addition, the color of the bulk solution changed from colorless to red before and after the addition of 90. mu.M pyridine-2, 6-dicarboxylic acid solution under irradiation of an ultraviolet lamp at 254 nm.

(3) Taking 3mL of main solution into a quartz cell, additionally sucking 10 parts of the same solution (11 groups of the same solution in total), selecting 1 part, adding 15 mu L of 18 mM pyridine-2, 6-dicarboxylic acid solution, and respectively adding 10 different-component interference component solutions (respectively phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, L-glutamic acid, L-aspartic acid and nicotinamide adenine binuclear glycine) with the same concentration and volume into the other 10 parts of solution (at the moment, the concentrations of rebuke pyridine-2, 6-dicarboxylic acid and all interference solutions in the quartz cell are 90 mu M, stirring uniformly, and measuring the fluorescence emission spectrum at the maximum excitation wavelength of 280nm and the maximum emission wavelength of 615 nm.

(4) 3mL of the main body solution was put in a quartz cell, 50. mu.L of 18 mM interfering component solution (phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, glutamic acid, aspartic acid, nicotinamide adenine dinuclear glycine, respectively) (at this time, the interfering solution concentration in the quartz cell was 450. mu.M) was added thereto, the mixture was stirred uniformly, and the fluorescence emission spectrum was measured at a maximum excitation wavelength of 280nm and a maximum emission wavelength of 615 nm. Then, 15. mu.L of 18 mM pyridine-2, 6-dicarboxylic acid solution (in this case, the concentration of the interfering solution in the quartz cell was 450. mu.M, and the concentration of the pyridine-2, 6-dicarboxylic acid solution was 90. mu.M) was added thereto, and the mixture was stirred uniformly, and the fluorescence emission spectrum was measured at a maximum excitation wavelength of 280nm and a maximum emission wavelength of 615 nm.

FIG. 3 is a bar graph (green) showing fluorescence intensity at 615nm after addition of various types of acidic solutions (orange) to the host solution and pyridine-2, 6-dicarboxylic acid to the host solution containing the interfering solution, respectively. As can be seen from the figure, the host solution has very obvious fluorescence response to the pyridine-2, 6-dicarboxylic acid solution, and can still realize the fluorescence response to the pyridine-2, 6-dicarboxylic acid under the condition of 5 times of coexistence of amino acid and aromatic ligand, which proves that the host solution still has excellent selectivity to the pyridine-2, 6-dicarboxylic acid.

After the pyridine-2, 6-dicarboxylic acid solution and the interference component solution are respectively added into the main solution, the fluorescence intensity at 615nm is obviously changed. Wherein, the main solution has very obvious fluorescence response to the pyridine-2, 6-dicarboxylic acid solution, so the complex can selectively identify the pyridine-2, 6-dicarboxylic acid. In addition, under the irradiation of sunlight and an ultraviolet lamp of 254nm, the naked eye can observe that the color of the pyridine-2, 6-dicarboxylic acid can be changed into bluish purple and red respectively.

(5) And (3) cutting the common filter paper into filter paper strips with the size of the pH test paper, soaking the filter paper strips in the solution obtained in the step (3), and drying the filter paper strips at 50 ℃ to obtain the test paper for detecting the pyridine-2, 6-dicarboxylic acid. And (3) immersing the filter paper in 100 mu M pyridine-2, 6-dimethyl acid solution and the interference solution respectively, standing for 0.5-1 second, then drawing out, and observing the color under sunlight and 254nm ultraviolet lamp.

Under an ultraviolet lamp, only the test strip soaked in the pyridine-2, 6-dimethyl acid solution is red by naked eyes, and the others are colorless; under the sunlight, only the test paper strip soaked in the pyridine-2, 6-dimethyl acid solution is blue-purple and the others are colorless.

Example 3

(1) Accurately weighing 29.0 mg DC-2 in a 100mL beaker, adding 19.6 mL water, stirring at room temperature for 50 minutes until completely dissolved and uniformly dispersed, and then adding 400. mu.L of 0.1M EuCl₃·H₂The aqueous solution was stirred for 30 minutes and the resulting solution was recorded as a host solution.

(2) 3mL of the bulk solution was taken in a quartz cell, and the fluorescence emission spectrum of the blank was measured 11 times at an excitation wavelength of 280 nm. Solutions of pyridine-2, 6-dicarboxylic acid (DPA) at different concentrations were added with a 1. mu.L sample injector (fluorescence titration). The fluorescence test adopts a OmniFluo900 series fluorescence spectrometer of Beijing Zhuoli Han light, uses a 75W xenon lamp as an excitation light source, and is provided with an excitation monochromator, an emission monochromator, a single photon counter and a refrigeration type photomultiplier. The fluorescence emission spectrum was measured with 280nm as the maximum excitation wavelength and 615nm as the maximum emission wavelength.

FIG. 4 is a graph showing the emission spectra of the host solution of example 3 without pyridine-2, 6-dicarboxylic acid (black) and with 100. mu.M pyridine-2, 6-dicarboxylic acid (red). Subsequently, we carried out a series of examples, and measured a series of fluorescence data by gradually increasing the concentration of the pyridine-2, 6-dicarboxylic acid aqueous solution in step (2), showing the corresponding change in fluorescence.

FIG. 5 is a linear fit graph of the main solution during the fluorescence titration process with pyridine-2, 6-dicarboxylic acid concentration as the abscissa and fluorescence intensity at 615nm as the ordinate. Linear fitting was performed with pyridine-2, 6-dicarboxylic acid concentration as abscissa (concentration range 0-100. mu.M) and fluorescence intensity at 615nm as ordinate. As the concentration of pyridine-2, 6-dicarboxylic acid increases,I ₆₁₅the fluorescence intensity also gradually increases and shows a good linear relationship in the concentration range of 5-30 mu M, R²=0.997, detection limit 10.4 nM. In addition, the color of the bulk solution changed from colorless to red before and after the addition of 90. mu.M pyridine-2, 6-dicarboxylic acid solution under irradiation of an ultraviolet lamp at 254 nm.

(3) Taking 3mL of main solution into a quartz cell, additionally sucking 10 parts of the same solution (11 groups of the same solution in total), selecting 1 part, adding 15 mu L of 18 mM pyridine-2, 6-dicarboxylic acid solution, and respectively adding 10 different-component interference component solutions (respectively phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, L-glutamic acid, L-aspartic acid and nicotinamide adenine binuclear glycine) with the same concentration and volume into the other 10 parts of solution (at the moment, the concentrations of rebuke pyridine-2, 6-dicarboxylic acid and all interference solutions in the quartz cell are 90 mu M, stirring uniformly, and measuring the fluorescence emission spectrum at the maximum excitation wavelength of 280nm and the maximum emission wavelength of 615 nm.

(4) 3mL of the main body solution was put in a quartz cell, 50. mu.L of 18 mM interfering component solution (phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, 2-picolinic acid, benzoic acid, glutamic acid, aspartic acid, nicotinamide adenine dinuclear glycine, respectively) (at this time, the interfering solution concentration in the quartz cell was 450. mu.M) was added thereto, the mixture was stirred uniformly, and the fluorescence emission spectrum was measured at a maximum excitation wavelength of 280nm and a maximum emission wavelength of 615 nm. Then, 15. mu.L of 18 mM pyridine-2, 6-dicarboxylic acid solution (in this case, the concentration of the interfering solution in the quartz cell was 450. mu.M, and the concentration of the pyridine-2, 6-dicarboxylic acid solution was 90. mu.M) was added thereto, and the mixture was stirred uniformly, and the fluorescence emission spectrum was measured at a maximum excitation wavelength of 280nm and a maximum emission wavelength of 615 nm.

FIG. 6 is a bar graph (green) showing fluorescence intensity at 615nm after addition of various types of acidic solutions (orange) to the host solution and pyridine-2, 6-dicarboxylic acid to the host solution containing the interfering solution, respectively. As can be seen from the figure, the host solution has very obvious fluorescence response to the pyridine-2, 6-dicarboxylic acid solution, and can still realize the fluorescence response to the pyridine-2, 6-dicarboxylic acid under the condition of 5 times of coexistence of amino acid and aromatic ligand, and the host solution still has excellent selectivity to the pyridine-2, 6-dicarboxylic acid.

(5) And (3) cutting the common filter paper into filter paper strips with the size of the pH test paper, soaking the filter paper strips in the solution obtained in the step (3), and drying the filter paper strips at 50 ℃ to obtain the test paper for detecting the pyridine-2, 6-dicarboxylic acid. And (3) immersing the filter paper in 100 mu M pyridine-2, 6-dimethyl acid solution and the interference solution respectively, standing for 0.5-1 second, then drawing out, and observing the color under sunlight and 254nm ultraviolet lamp.

Under an ultraviolet lamp, only the test strip soaked in the pyridine-2, 6-dimethyl acid solution is red by naked eyes, and the others are colorless; under the sunlight, only the test paper strip soaked in the pyridine-2, 6-dimethyl acid solution is observed to be blue by naked eyes, and the others are colorless.

In conclusion, the invention realizes naked eye identification and differential detection of pyridine-2, 6-dicarboxylic acid by using the complex formed by compounding the catechol Schiff bases (DC-1 and DC-2) and europium ions, and the detection limits are 8.3 nM and 10.4 nM respectively. And can eliminate the interference of other amino acids and aromatic substances such as phthalic acid, isophthalic acid, terephthalic acid, nicotinic acid, isonicotinic acid, benzoic acid, L-glutamic acid and the like; meanwhile, the method has excellent selectivity under the coexistence of 5 times of interferents, realizes simple and convenient low-cost detection of the pyridine-2, 6-dicarboxylic acid, and simultaneously achieves the purposes of high sensitivity, good selectivity and strong anti-interference capability.

Preliminary tests show that ligands in which R is amino, nitro, halogen, ethyl, methoxy also have the properties described in examples 2 and 3.