Preparation and application of fluorescent probe for ultra-sensitively detecting cadmium ions

Abstract

The invention discloses preparation and application of a fluorescent probe for ultra-sensitive detection of Cd < 2 + >, and aims to solve the problems that the existing fluorescent probe for identifying Cd < 2 + > ions is tedious in synthesis step, relatively long in selectivity and response time and relatively high in detection limit. The two fluorescent probes are prepared through a Schiff base reaction under mild conditions. The preparation method of the fluorescent probe 1 (HMH) comprises the following step: reacting 2, 6-diformyl-4-methylphenol with salicylhydrazine to prepare the fluorescent probe 1 (HMH). The preparation method of the fluorescent probe 2 (H3-MB) comprises the step of further reacting with 6-amino-1-hexanol on the basis of the probe HMH to obtain a target compound. The two probes disclosed by the invention are simple to synthesize and low in cost, both can realize sensing detection of Cd < 2 + > ions, the detection limit is low, the response time is short, the selectivity is good, and H3-MB can also realize visual distinguishing of Cd < 2 + > and Zn < 2 + >.

Description

Technical Field

[0001] The present invention belongs to the technical field of fluorescent probes and heavy metal detection, and relates to a method for ultra-sensitively detecting cadmium ions (Cd 2 + ) Preparation method of fluorescent probe. Background Art

[0002] Due to its strong chemical activity, mobility and irreversible toxicity, cadmium is listed as one of the heavy metal pollutants by the World Health Organization. With the development of industrialization, environmental problems of water, soil and crop pollution caused by industries such as metallurgy, electroplating, mining of lead and zinc deposits and ceramics have attracted global attention. Cadmium is a non-essential element for the human body. Once absorbed by the human body, it accumulates in the bones and kidneys through the food chain, eventually causing various diseases and endangering human health. Therefore, there is an urgent need for rapid, sensitive and selective detection technology for cadmium, which is of great significance for accurately repairing polluted water environments and reducing a series of health risks caused by heavy metal pollution.

[0003] At present, traditional methods for detecting heavy metals include atomic absorption spectroscopy (AAS), atomic fluorescence spectrophotometry (AFS), inductively coupled plasma (ICP), high performance liquid chromatography (HPLC), mass spectrometry (MS), electrochemical method, colorimetry, etc. Although these detection technologies can perform quantitative analysis more accurately, various factors such as high detection cost, complex operation, and high detection limit make these methods difficult to popularize and apply. Fluorescence detection technology shows obvious advantages due to its high selectivity, high sensitivity, and simple operation. 2+ and Zn 2+ The similar electronic configuration, coordination number and chemical properties make it difficult to distinguish. 2+ Fluorescent probes have been reported, but the currently developed fluorescent probes still have the problems of high preparation cost, high detection limit and poor selectivity. Therefore, it is of great significance to develop fluorescent probes with simple preparation, low detection limit and good selectivity. Summary of the invention

[0004] The purpose of the present invention is to solve the existing problem of identifying Cd 2+ The probe synthesis steps are complicated, the selectivity is low, the response time is long, and the detection limit is high. 2+ Fluorescent probe and preparation method thereof.

[0005] To achieve the above object, the present invention adopts the following technical solution: An ultra-sensitive fluorescent probe for detecting cadmium ions, the structural formula of which is as follows: , When R = -CHO, the fluorescent probe is named HMH; When R=-C=N-(CH2)OH, the fluorescent probe is named H3-MB.

[0006] The preparation methods of the above two fluorescent probes are as follows: 1. Fluorescent probe 1 (HMH) Preparation reaction formula: Preparation method: Dissolve 2,6-diformyl-4-methylphenol and salicylic hydrazide in ethanol, heat to reflux, the heating temperature is 60~100 ℃, react for 6~24 hours, use TCL plate to detect the reaction, after the reaction is complete, cool to room temperature, filter and precipitate. The collected supernatant is dried by rotary evaporator to obtain a crude product, and then purified by column chromatography, the eluent is PE: EA = 5: 1~100: 1, and the target compound HMH can be obtained by drying.

[0007] Wherein, the molar ratio of 2,6-diformyl-4-methylphenol to salicylic hydrazide is 1:1-1:3.

[0008] Structural Characterization of HMH: 1 H NMR (400 MHz, DMSO- d 6) δ 12.35 (s, 1H), 11.95 (s,2H), 10.37 (s,1H), 8.68 (s, 1H), 7.88 (d, J = 7.8 Hz, 1H), 7.70 (s, 1H), 7.59(s, 1H), 7.47 (t, J = 7.8 Hz, 1H), 7.00 (d, J = 8.2 Hz, 1H), 6.97 (d, J = 7.5 Hz,1H), 2.32 (s, 3H). 13 C NMR (101 MHz, DMSO-d6) δ 190.37, 164.51, 158.83, 158.54,148.22, 136.77, 134.10, 130.49, 128.77, 128.52,122.93, 119.90, 119.09,117.28, 115.67, 19.69. ESI-MS (m / z): Calcd: 298.30. Found: 299.1680 [M + H] + ;321.1533 [M+Na] + .

[0009] 2. Fluorescent probe 2 (H3-MB) Preparation reaction formula: Preparation method: A mixture of compound HMH and 6-amino-1-hexanol is dissolved in ethanol, heated to reflux at a heating temperature of 60~100°C, reacted for 6~24 hours, and detected the reaction with a TCL plate. After the reaction is complete, it is cooled to room temperature and rotary evaporated to obtain a crude product, which is then purified by silica gel chromatography with an eluent of DCM:MeOH = 2:1~100:1, and dried to obtain the target compound H3-MB.

[0010] Wherein, the molar ratio of the compound HMH to 6-amino-1-hexanol is 1:1-1:5.

[0011] Structural Characterization of H3-MB: 1 H NMR (400 MHz, DMSO-d6) δ 11.98 (s, 1H), 8.65 (s,1H), 8.56 (s, 1H), 7.93 (d, J = 8.0 Hz, 1H), 7.77 (d, J = 7.7 Hz, 1H), 7.58(d, J = 17.6 Hz, 1H), 7.23 (t, J = 7.4Hz, 1H), 6.81 (d, J = 8.1 Hz, 1H), 6.72(t, J = 7.5 Hz, 1H), 4.38 (s, 1H), 3.65 – 3.59 (m, 2H), 3.38 (s, 2H), 2.19(s,3H), 1.47 – 1.40 (m, 4H), 1.35 – 1.30 (m, 4H). 13 C NMR (101 MHz, DMSO- D 6) δ170.23, 168.19, 159.96, 150.43, 142.89,136.04, 132.75, 131.21, 128.74,123.80, 122.78, 118.52, 116.41, 114.80, 63.99, 60.39, 32.21, 32.09, 28.88,24.97, 19.29. ESI-MS (m / z): Calcd: 397.48. Found:398.1506 [M + H] + .

[0012] Another object of the present invention is to disclose the application of the above two fluorescent probes in cadmium ion detection. The HMH is a fluorescence on-type probe, and H3-MB is a fluorescence ratio probe. Among them, the detection limit of HMH is 2.78 ppb, and the response time is 1 min. The detection limit of H3-MB is 0.30 ppb, and the response time is 30 s. The sensing detection is fluorescence spectrum detection, selective detection, and time response detection. H3-MB can also visually distinguish Cd by changes in fluorescence spectrum intensity and displacement. 2+ and Zn 2 + .

[0013] The beneficial effects of the present invention are as follows: 1) The probes of the present invention are simple to synthesize, the raw materials are economical and readily available, and the post-processing is relatively simple, wherein the probe HMH only requires one step, and the probe H3-MB only requires two steps to complete; 2) The fluorescent probes prepared by the present invention can achieve low concentration Cd 2+ The detection of HMH was 2.78 ppb and the response time was 1 min. The detection limit of H3-MB was 0.30 ppb and the response time was 30 s. 3) The fluorescent probe H3-MB prepared by the present invention can distinguish Cd 2+ and Zn 2+ , which is to realize the probe in detecting Cd 2+ and Zn 2+ It is an important means to distinguish the two in the process.

[0014] Therefore, the present invention is a simple, rapid, ultrasensitive and selective Cd 2+ Fluorescent probes are expected to achieve practical applications through further functionalization. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 The probe HMH reacts with Cd in different solvents 2+ (concentration 500 ppb) fluorescence change graph.

[0016] Figure 2 is the selectivity of probe HMH to different ions in ethanol.

[0017] Figure 3 The probe HMH is used to detect Cd in ethanol. 2+ Time response plots for concentrations of 100 ppb and 50 ppb.

[0018] Figure 4 The probe HMH is used to detect different concentrations of Cd in ethanol.2+ Fluorescence spectral response diagram.

[0019] Figure 5 The probe HMH is used to detect different concentrations of Cd in ethanol. 2+ Linear fitting graph of .

[0020] Figure 6 The probe H3-MB reacts with Cd in different solvents. 2+ (concentration of 200 ppb).

[0021] Figure 7 is the selectivity of probe H3-MB to different ions in acetonitrile.

[0022] Figure 8 The probe H3-MB was used to detect Cd 2+ (concentration of 200 ppb) time response plot.

[0023] Fig. 9 The probe H3-MB was used to detect different concentrations of Cd in acetonitrile. 2+ Fluorescence spectral response diagram.

[0024] Fig.10 The probe H3-MB was used to detect different concentrations of Cd in acetonitrile. 2+ Linear fitting graph of .

[0025] Fig.11 The probe H3-MB was used to detect Cd 2+ (concentration of 200 ppb) and Zn 2+ (concentration 1 ppm) fluorescence spectrum response diagram.

[0026] Fig.12 The probe H3-MB was added with Cd 2+ and Zn 2+ Fluorescence emission image under UV light. DETAILED DESCRIPTION

[0027] The present invention is further explained below with reference to specific embodiments.

[0028] Example 1 2,6-diformyl-4-methylphenol (0.1640 g, 1.0 mmol) and salicylic hydrazide (0.1560 g, 1.0 mmol) were dissolved in ethanol, heated to reflux at 80 °C, reacted for 6 h, and the reaction was detected by TCL plate. After the reaction was complete, it was cooled to room temperature and filtered to precipitate. The collected supernatant was dried by rotary evaporator to obtain a crude product, which was then purified by column chromatography with eluent PE: EA = 10:1. The target compound HMH was obtained by drying with a yield of 94%.

[0029] Example 2 A mixture of compound HMH (0.0298 g, 0.1 mmol) and 6-amino-1-hexanol (0.0117 g, 0.1 mmol) was dissolved in ethanol and heated to reflux at 85 °C for 8 h. The reaction was monitored by TCL plate. After the reaction was complete, the product was cooled to room temperature and rotary evaporated to obtain a crude product, which was then purified by silica gel chromatography with an eluent of DCM:MeOH = 5:1. The target compound H3-MB was obtained by drying with a yield of 65%.

[0030] Fluorescent probe performance study The fluorescent probes HMH and H3-MB synthesized in Example 1 and Example 2 were dissolved in DMSO to prepare a mother solution with a concentration of 5 mM. 0.0102 g of CdCl2·2.5H2O solid powder was dissolved in 5 mL of secondary water to prepare 1000 ppm Cd 2+ The other metal ions were also dissolved in secondary water to prepare 1000 ppm stock solutions and stored in a 4°C refrigerator.

[0031] 1. Fluorescent probe HMH 1) The probe HMH synthesized in Example 1 was dissolved in methanol (MeOH), acetonitrile (ACN), dimethyl sulfoxide (DMSO), N,N-dimethylformamide (DMF), ethyl acetate (EA), tetrahydrofuran (THF), acetone (AC), dichloromethane (DCM), 1,4-dioxane (1,4-Dioxane), water (H2O), and ethanol (EtOH) to prepare a concentration of 5×10 -6 mol / L. When 500ppb Cd is added 2+ After that, the fluorescence intensity of the probe HMH in different solvents changes as shown in Figure 1 Under the excitation wavelength of 430 nm, the change in fluorescence intensity at the maximum emission wavelength of 540 nm was most obvious in the ethanol solvent. Therefore, ethanol was selected as the solvent in the subsequent tests.

[0032] 2) To determine the effect of HMH on Cd 2+ To improve the selectivity of HMH, other common ions including Na were added at a concentration of 1 ppm to a 5 μM HMH ethanol solution. + , K + 、Ag + 、Al 3+ , Mn 2+ 、Co 2+ 、Ni 2+ , ClO - .like Figure 2 As shown, when 500 ppb Cd is added2+ After addition of Cd, the fluorescence at the maximum fluorescence emission peak was significantly enhanced and the wavelength was blue-shifted to 499 nm. Although the addition of other ions would change the fluorescence, it was not significantly different from that of Cd. 2+ The changes are relatively small, so the addition of other ions can be ignored.

[0033] 3) Fluorescent probe HMH in testing Cd 2+ In the process of detection, response time is not only an important factor affecting the detection conditions, but also an important parameter for evaluating the performance of the probe. Figure 3 As shown in Figure 2, when 100 ppb of Cd is added 2+ When 50 ppbCd was added, the fluorescence intensity of the probe HMH reached equilibrium within 3 min. 2+ After that, the fluorescence intensity of HMH can reach equilibrium within 1 min. To ensure the reliability of the detection conditions, the response time in the subsequent tests was 3 min.

[0034] 4) To determine the sensitivity of the fluorescent probe HMH, 5-50 ppb Cd was added to a 5 μM HMH ethanol solution. 2+ ,like Figure 4 As shown, the fluorescence intensity increases continuously with a slight blue shift. Figure 5 The fluorescence intensity of the HMH probe was correlated with the low concentration of Cd 2+ The standard working curve of ions, linear fitting obtained y = 819.27 + 6.49 * x, correlation coefficient R 2 =0.99. According to the minimum detection limit formula LOD = 3σ / S (S is the slope of the calibration curve, σ is the standard deviation of the intercept), the detection limit LOD is calculated to be 2.78ppb, and the actual concentration can reach 5.00 ppb, thus achieving quantitative detection of Cd2+.

[0035] 2. Fluorescent probe H3-MB 1) The probe H3-MB synthesized in Example 2 was dissolved in MeOH, ACN, DMSO, DMF, EA, THF, AC, DCM, 1,4-Dioxane, H2O, and EtOH solvents to a concentration of 5×10 -6 mol / L. When 200 ppb Cd is added 2+ After that, the fluorescence intensity of the probe HMH in different solvents changes as shown in Figure 6 The fluorescence intensity change of the probe at the maximum emission wavelength of 552 nm under the excitation wavelength of 410 nm was most obvious in acetonitrile solvent. Therefore, acetonitrile was selected as the solvent in the subsequent tests.

[0036] 2) To determine the effect of H3-MB on Cd 2+To improve the selectivity of H3-MB, other common ions including Na + , K + , ClO - , Cu 2+ 、Co 2+ 、Ni 2+ 、NO3 - 、SO4 2- 、HCO3 2- .like Figure 7 As shown, when 200 ppb Cd is added 2+ After addition, the fluorescence intensity at the maximum fluorescence emission peak of 552 nm decreased, while the emission peak intensity at 498 nm increased significantly. Although the addition of other ions would cause fluorescence changes, they were not significantly different from those of Cd 2+ The changes are relatively small, so the addition of other ions can be ignored.

[0037] 3) In order to determine the fluorescence probe H3-MB in the test Cd 2+ The response time of the process, such as Figure 8 As shown in Figure 2, when 200 ppb Cd is added 2+ When the fluorescence intensity of probe H3-MB reaches equilibrium within 30 s, the probe increases the response time by 6 times compared with probe HMH. Therefore, the response time in subsequent tests is 30 s.

[0038] 4) In order to evaluate the sensitivity of the fluorescent probe H3-MB, 5-40 ppb of Cd was added to the acetonitrile solution of 5 μM HMH. 2+ ,like Fig. 9 As shown, with Cd 2+ With the continuous increase of concentration, the emission peak intensity at 552 nm gradually decreases, while the emission peak intensity at 498 nm continues to increase. Fig.10 The peak ratio of the H3-MB probe at 498 nm and 552 nm was established with the low concentration of Cd 2+ The standard working curve of the linear fitting is y=0.2314+0.3182*x, and the correlation coefficient is R 2 =0.995. According to the minimum detection limit formula LOD = 3σ / S (S is the slope of the calibration curve, σ is the standard deviation of the intercept), the detection limit LOD is calculated to be 0.30ppb, which is 9 times higher than that of the probe HMH, so the probe has high sensitivity.

[0039] 5) Due to Cd 2+ and Zn 2+ They have similar electronic configurations, coordination numbers, and chemical properties, so it is necessary to explore the selectivity of the fluorescent probe H3-MB. Fig.11 As shown, in order to detect the change of fluorescence signal, 200 ppb Cd was added 2+ and 1ppm Zn 2+ The experimental results show that Zn 2+ and Cd 2+ will enhance fluorescence, but Cd 2+ The fluorescence enhancement is more obvious, and the emission peak positions of the two differ by 5 nm. Fig.12 To add 1 ppm Cd to the 25 μM probe solution 2+ and Zn 2+ After that, the fluorescence color changes under the irradiation of ultraviolet light at 365nm. It can be seen from the figure that the fluorescence color of H3-MB is yellow. 2+ After that, the fluorescence color turned green, and the addition of Zn 2+ The fluorescence color then changes to blue-green, so this method can be used to visually distinguish Zn 2+ and Cd 2 + .

[0040] Based on the excellent performance of the above-mentioned fluorescent probes HMH and H3-MB, it is expected to quantitatively detect and screen cadmium ions in complex solvent systems with good practicality.

Claims

1. An ultrasensitive fluorescent probe for detecting cadmium ions, characterized in that: The structural formula of the fluorescent probe is as follows: ， When R = -CHO, the fluorescent probe is named HMH; When R=-C=N-(CH2)OH, the fluorescent probe is named H3-MB.

2. A method for preparing an ultrasensitive fluorescent probe for detecting cadmium ions as claimed in claim 1, characterized in that: The following steps are involved: 1) Dissolve 2,6-diformyl-4-methylphenol and salicylic hydrazide in ethanol, heat to 60-100°C, reflux for 6-24 h, and after the reaction is complete, cool to room temperature, filter the precipitate, and dry the collected supernatant by rotary evaporator to obtain a crude product, which is then purified by column chromatography with an eluent of PE:EA=5:1-100:

1. Drying can obtain the target compound HMH; 2) The compound HMH obtained in step 1) and 6-amino-1-hexanol are co-dissolved in ethanol, heated to 60-100° C., and refluxed for 6-24 h. After the reaction is complete, the mixture is cooled to room temperature and rotary evaporated to obtain a crude product, which is then purified by silica gel chromatography with an eluent of DCM:MeOH = 2:1-100:1, and dried to obtain the target compound H3-MB.

3. A method for preparing a fluorescent probe for ultrasensitive detection of cadmium ions as claimed in claim 2, characterized in that: In step 1), the molar ratio of 2,6-diformyl-4-methylphenol to salicylic hydrazide is 1:1 to 1:

3.

4. A method for preparing a fluorescent probe for ultrasensitive detection of cadmium ions as claimed in claim 2, characterized in that: In step 2), the molar ratio of HMH to 6-amino-1-hexanol is 1:1 to 1:

5.

5. Use of the ultrasensitive fluorescent probe for detecting cadmium ions as claimed in claim 1 in sensing and detecting cadmium ions.

6. The use of the ultrasensitive fluorescent probe for detecting cadmium ions in sensing and detecting cadmium ions as claimed in claim 5, characterized in that: The HMH is a fluorescence on-type probe, and H3-MB is a fluorescence ratio probe; the detection limit of HMH is 2.78 ppb, and the response time is 1 min; the detection limit of H3-MB is 0.30 ppb, and the response time is 30 s.

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