Preparation method and application of a dual excitation fluorescent probe for detecting hydrazine hydrate

Abstract

The application belongs to the field of fluorescent probe and environmental analysis and detection technology, and particularly relates to a preparation method and application of a double-excitation fluorescent probe for detecting hydrazine hydrate. A double-excitation fluorescent probe is synthesized, the fluorescent probe compound has the structure of formula I, 4-chloro-7-(dimethylamino)-3-formyl coumarin and 3-ethyl-2-methyl benzothiazole iodide are used as raw materials, and the fluorescent probe is synthesized through nucleophilic substitution and aldol condensation two-step reactions. The fluorescent probe has high selectivity and high sensitivity to hydrazine in an aqueous solution. With the increase of the concentration of hydrazine, the fluorescence intensity at 446 nm gradually increases, and the fluorescence intensity at 633 nm gradually decreases. In the presence of common cations, anions and amino acids, the fluorescent probe can specifically recognize hydrazine hydrate. The probe can construct a ratio type detection system, has strong anti-interference ability, high selectivity and high sensitivity to hydrazine hydrate, and is used for quantitative detection of hydrazine hydrate in environmental samples such as water and soil. The probe is simple to operate and has wide application prospects. Formula I

Description

Technical Field

[0001] This invention belongs to the field of fluorescent probe and environmental analysis and detection technology, and particularly relates to a method for preparing and applying a dual-excitation fluorescent probe for detecting hydrazine hydrate. Background Technology

[0002] Hydrazine (N₂H₄, with hydrazine hydrate being its most common form) is a highly reactive, colorless, oily, flammable liquid with an irritating odor similar to ammonia. As an important chemical raw material and reaction intermediate, hydrazine is widely used in rocket propellants, metal corrosion inhibitors, and industrial reducing agents. It is also a key raw material in the synthesis of herbicides, insecticides, fungicides, and other pesticides.

[0003] From an environmental and health safety perspective, hydrazine is highly toxic and can enter the human body through inhalation, ingestion, and skin contact, causing acute or chronic damage. Long-term exposure can lead to organic lesions in organs such as the liver and kidneys. At the same time, as a typical environmental pollutant, hydrazine can easily enter water and soil environments through infiltration and diffusion, posing a long-term threat to ecosystems and public health. Therefore, developing rapid, sensitive, and reliable detection methods for hydrazine hydrate is of great practical significance for environmental monitoring and human health protection.

[0004] Currently, the main methods for detecting hydrazine hydrate include electrochemical methods, chromatographic analysis, and chemiluminescence methods. However, these methods generally suffer from drawbacks such as cumbersome operation procedures, reliance on large and precision instruments, and harsh detection conditions, making it difficult to meet the needs of real-time and rapid on-site detection. In contrast, fluorescence analysis has gained widespread attention in the field of rapid detection due to its advantages such as simple operation, rapid response, and intuitive signals.

[0005] Currently, various fluorescent probes based on optical signal response have been successfully designed and developed. These probes achieve targeted detection by detecting changes in fluorescence signals triggered by the specific recognition of N2H4, providing a new technical approach for the efficient and accurate detection of N2H4. However, most fluorescent probes for hydrazine hydrate are single-excitation and single-emission modes, and their detection signals are easily affected by factors such as ambient temperature, pH, instrument fluctuations, and matrix background, resulting in insufficient detection accuracy and stability. At the same time, most probes suffer from problems such as long response time, low sensitivity, and high detection limits, making them unsuitable for the detection of trace amounts of hydrazine hydrate.

[0006] Therefore, developing a fluorescent probe for hydrazine hydrate with strong anti-interference ability, high sensitivity, low detection limit, and the ability to achieve on-site visual detection has significant application value. Summary of the Invention

[0007] This invention addresses the technical problems of existing hydrazine hydrate fluorescent probes, such as poor anti-interference performance, insufficient sensitivity, and difficulty in rapid on-site detection under single excitation mode. It proposes a dual-excitation response hydrazine hydrate fluorescent probe, and provides its preparation method and detection application, achieving high selectivity, high sensitivity, and low detection limit detection of hydrazine hydrate, as well as visualized on-site detection of gaseous and liquid hydrazine hydrate.

[0008] To achieve the above objectives, the technical solution adopted by the present invention is as follows: a dual-excitation fluorescent probe for detecting hydrazine hydrate is provided, which has the chemical structure shown in Formula I:

[0009] Formula I.

[0010] A detection method for preparing a dual-excitation fluorescent probe for detecting hydrazine hydrate is also provided, specifically including the following steps:

[0011] (1) 4-Chloro-7-(dimethylamino)-3-carboxycoumarin was mixed with ethanol, and triethylamine was added as an acid-binding agent. The mixture was heated to reflux and reacted. After the reaction was completed, the intermediate 4-ethoxy-7-(dimethylamino)-3-carboxycoumarin was obtained by column chromatography. In this step, ethanol was used as both a reaction solvent and a nucleophile to replace the chlorine atom at the 4-position of coumarin, generating an ethoxylated product.

[0012] ;

[0013] (2) The intermediate obtained in step (1) was mixed with 3-ethyl-2-methylbenzothiazole iodide, and anhydrous ethanol was added as a solvent. The mixture was heated under reflux to undergo an aldol condensation reaction. After the reaction was completed, the mixture was purified by column chromatography to obtain the target dual-excitation fluorescent probe:

[0014] .

[0015] Preferably, the reaction temperature in steps (1) and (2) is 85~95℃ and the reaction time is 6~8 hours; preferably, the reaction temperature is 90℃ and the reaction time is 7 hours.

[0016] Preferably, in step (2), the molar ratio of 4-ethoxy-7-(dimethylamino)-3-carboxycoumarin to 3-ethyl-2-methylbenzothiazole iodide is 1:(0.9~1.1), and more preferably the molar ratio is 1:1.

[0017] Preferably, the column chromatography eluent in step (1) is a mixture of dichloromethane and ethyl acetate with a volume ratio of 3:1; and the column chromatography eluent in step (2) is a mixture of dichloromethane and methanol with a volume ratio of 20:1.

[0018] The application of the aforementioned dual-excitation fluorescent probe in the detection of hydrazine hydrate is also provided, and the detection targets include one or more of the following: water samples (tap water, river water, drinking water, etc.), soil samples (field soil, sand, etc.), and food samples.

[0019] Preferably, the detection adopts a dual-excitation fluorescence detection mode: the first excitation wavelength is 370~380nm (preferably 375nm), corresponding to the detection of fluorescence emission intensity at 440~450nm (preferably 446nm), and the fluorescence intensity at this wavelength gradually increases with the increase of hydrazine hydrate concentration; the second excitation wavelength is 505~515nm (preferably 510nm), corresponding to the detection of fluorescence emission intensity at 630~640nm (preferably 633nm), and the fluorescence intensity at this wavelength gradually decreases with the increase of hydrazine hydrate concentration.

[0020] By alternating the rise and fall of fluorescence signals at two wavelengths, a ratiometric detection system can be constructed, effectively offsetting interference from environmental factors and instrument fluctuations, and significantly improving detection accuracy. This probe achieves a detection limit of 20.12 nM at 446 nm and 3.10 nM at 633 nm, demonstrating excellent trace detection capabilities.

[0021] Compared with the prior art, the advantages and positive effects of the present invention are as follows:

[0022] This invention provides a dual-excitation fluorescent probe for detecting hydrazine hydrate:

[0023] (1) Dual excitation ratio response with strong anti-interference: The response mode of dual excitation wavelength and dual emission signal is adopted. The signal change of one rise and one fall can construct a ratio detection system, which can effectively offset the interference of environmental matrix, pH, instrument parameter fluctuations, etc. The detection stability and accuracy are significantly better than those of single excitation probe.

[0024] (2) High sensitivity and low detection limit: It has excellent sensitivity to hydrazine hydrate, and the lowest detection limit can reach the nanomolar level, which can meet the detection requirements of trace hydrazine hydrate in the environment.

[0025] (3) Excellent selectivity: It can still produce a specific recognition response to hydrazine hydrate in the presence of common cations, anions, amino acids and other coexisting substances, without obvious cross-interference.

[0026] (4) Wide range of applications: It can be used for the precise quantitative detection of hydrazine hydrate in solution systems, and can also be prepared into test strips to realize the visual on-site detection of liquid and gaseous hydrazine hydrate, adapting to various scenarios such as laboratory testing and on-site rapid screening.

[0027] (5) The synthesis process is simple: the target product can be obtained in two steps, the reaction conditions are mild, the purification method is simple, and it is easy to prepare on a large scale. Attached Figure Description

[0028] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the following description of the embodiments will be briefly introduced. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0029] Figure 1 The proton NMR spectrum of the fluorescent probe prepared in Example 1 of this invention ( 1 HNMR, 500MHz, DMSO-d6);

[0030] Figure 2 The carbon NMR spectrum of the fluorescent probe prepared in Example 1 of this invention ( 13 CNMR, 126MHz, DMSO-d6);

[0031] Figure 3(a) shows the fluorescence spectrum under 375 nm excitation;

[0032] Figure 3(b) is a bar chart of fluorescence intensity corresponding to different analytes under 375 nm excitation;

[0033] Figure 3(c) shows the fluorescence spectrum under 510 nm excitation;

[0034] Figure 3(d) is a bar chart of fluorescence intensity corresponding to different analytes under 510 nm excitation;

[0035] Figure 4(a) shows the results under 375nm excitation;

[0036] Figure 4(b) shows the results under 510 nm excitation;

[0037] Figure 5(a) shows the fluorescence intensity variation curves of the probe at 375 nm excitation and 446 nm emission under different pH conditions;

[0038] Figure 5(b) shows the fluorescence intensity variation curves of the probe at 510 nm excitation and 633 nm emission under different pH conditions;

[0039] Figure 6(a) shows the fluorescence spectrum under 375 nm excitation;

[0040] Figure 6(b) shows the linear fitting curve of fluorescence intensity at 446 nm under 375 nm excitation and hydrazine hydrate concentration;

[0041] Figure 6(c) shows the fluorescence spectrum under 510 nm excitation;

[0042] Figure 6(d) shows the linear fitting curve of fluorescence intensity at 633 nm under 510 nm excitation and hydrazine hydrate concentration;

[0043] Figure 7(a) shows the results under 375nm excitation;

[0044] Figure 7(b) shows the results under 510 nm excitation;

[0045] Figure 8(a) shows the fluorescence response of sandy soil matrix under low concentration of hydrazine hydrate;

[0046] Figure 8(b) shows the fluorescence response of sandy soil matrix under medium concentration of hydrazine hydrate;

[0047] Figure 8(c) shows the fluorescence response of sandy soil matrix under high concentration of hydrazine hydrate;

[0048] Figure 8(d) shows the fluorescence response of field soil matrix under low concentrations of hydrazine hydrate;

[0049] Figure 8(e) shows the fluorescence response of field soil matrix under medium concentration of hydrazine hydrate;

[0050] Figure 8(f) shows the fluorescence response of field soil matrix under high concentration of hydrazine hydrate;

[0051] Figure 9(ac) is a visualization of the response of the fluorescent probe detection strip to liquid hydrazine;

[0052] Figure 9(d) is a visualization of the results of the fumigation detection of fluorescent probe test strips and gaseous hydrazine hydrate. Detailed Implementation

[0053] To better understand the above-mentioned objectives, features, and advantages of the present invention, the present invention will be further described below in conjunction with the accompanying drawings and embodiments. It should be noted that, unless otherwise specified, the embodiments and features described in these embodiments can be combined with each other.

[0054] Numerous specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways than those described herein, and therefore the invention is not limited to the specific embodiments disclosed in the following specification.

[0055] Example 1, as Figures 1-2 As shown, a dual-excitation fluorescent probe for detecting hydrazine hydrate is prepared:

[0056] (1) Synthesis of intermediate 4-ethoxy-7-(dimethylamino)-3-carboxycoumarin

[0057] Weigh 0.2 g of 4-chloro-7-(dimethylamino)-3-carboxycoumarin and place it in a reaction flask. Add 10 mL of ethanol and then 0.52 mL of triethylamine. Stir well and heat to 90 °C. Reflux for 7 hours. After the reaction is complete, cool to room temperature and remove the solvent by rotary evaporation under reduced pressure. The crude product is purified by column chromatography (eluent volume ratio: dichloromethane: ethyl acetate = 3:1) to obtain 0.099 g of the intermediate yellow solid product, with a yield of 50.87%.

[0058] (2) Synthesis of target fluorescent probe

[0059] Weigh 0.1 g of the intermediate prepared in step (1) and 0.12 g of 3-ethyl-2-methylbenzothiazole iodide into a reaction flask, add 15 mL of anhydrous ethanol, stir well, and heat to 90 °C. Reflux for 7 hours. After the reaction is complete, cool to room temperature, remove the solvent by rotary evaporation under reduced pressure, and purify the crude product by column chromatography (eluent: dichloromethane: methanol = 20:1, volume ratio) to obtain 0.0859 g of a red solid product, with a yield of 39.59%.

[0060] The product was characterized by nuclear magnetic resonance, and the results are as follows:

[0061] 1 HNMR(500MHz,DMSO-d6)δ:8.37(d,J=8.1Hz,1H),8.28(d,J=8.5Hz,1H),8.12 (d,J=15.3Hz,1H),8.08(s,1H),7.85(d,J=7.8Hz,1H),7.82-7.73(m,2H),6. 87(dd,J=9.3,2.4Hz,1H),6.67(d,J=2.5Hz,1H),4.76(t,J=7.3Hz,2H),4.51 (q,J=7.0Hz,2H),3.13(s,6H),1.55(t,J=7.0Hz,3H),1.49(t,J=7.3Hz,3H).

[0062] 13 C{ 1 H}NMR(126MHz,DMSO-d6)δ:172.3,171.2,161.0,157.0,155.7,141.5,139.7,129.9,128.5,12 8.2,127.3,124.8,116.7,111.2,104.9,104.5,97.5,74.7,56.5,44.7,40.4,19.0,16.3,14.1.

[0063] The spectral results are consistent with the target probe structure, proving that the product synthesis was successful.

[0064] Example 2, Anti-interference performance test of dual-excitation fluorescent probe:

[0065] The fluorescent probe prepared in Example 1 was prepared into a 1 mM stock solution using DMSO.

[0066] Prepare a DMSO and HEPES buffer solution (pH=7.4) at a volume ratio of 9:1, resulting in a total volume of 240 μL. Add 30 μL of fluorescent probe stock solution, followed by 30 μL of 20 equivalents (relative to probe concentration) of different interfering standard solutions, including: Ca... 2+ Mg 2+ Zn 2+ Cr3 + Na + Mn 2+ SO4 2- H2PO4 - F - Cl - ,Br - HS - L-alanine, L-cysteine, L-histidine, L-serine, L-proline, L-valine, L-aspartic acid, and L-tryptophan were reacted. After 30 minutes of reaction, 270 μL of HEPES buffer (pH=7.4) and 2430 μL of LDMSO were added to bring the volume to a fixed level. The fluorescence emission spectra were measured at excitation wavelengths of 375 nm and 510 nm, respectively.

[0067] Another experimental group was set up: after adding the interfering substance to the same system as above, 30 μL of 20 equivalent hydrazine hydrate standard solution was added, and the fluorescence spectrum was tested after reacting for 30 minutes.

[0068] The test results are shown in Figure 3 (fluorescence response results of the fluorescent probe and hydrazine hydrate under different interfering substances under dual excitation conditions):

[0069] When various interfering substances were added, the fluorescence intensity at both wavelengths did not change significantly. After adding hydrazine hydrate, the fluorescence at 446 nm was significantly enhanced and the fluorescence at 633 nm was significantly weakened, regardless of the presence of interfering substances. The results show that the prepared fluorescent probe can still show a specific recognition response to hydrazine hydrate under the condition of coexistence of multiple common interfering substances. The probe has excellent recognition selectivity for hydrazine hydrate, and common anions, cations and amino acids do not significantly interfere with the detection.

[0070] Example 3: Testing the effect of solvent system on probe fluorescence performance

[0071] Prepare 3 mL mixtures of DMSO and HEPES buffer (pH=7.4) at volume ratios of 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2, and 9:1. Add 30 μL of 1 mM fluorescent probe stock solution to each mixture and test the fluorescence spectra at excitation wavelengths of 375 nm and 510 nm.

[0072] The test results are shown in Figure 4 (fluorescence intensity variation of the fluorescent probe in DMSO / HEPES mixtures with different volume ratios under dual excitation conditions):

[0073] The probe exhibits stable fluorescence performance in DMSO / HEPES mixtures with varying ratios. The addition of hydrazine hydrate results in significant fluorescence responses across a wide solvent ratio range, demonstrating the probe's adaptability to detection systems with varying water content.

[0074] Example 4: Test of the effect of pH on probe response performance

[0075] Prepare a series of HEPES buffer solutions with pH values ​​from 1 to 14. Prepare 240 μL mixtures of DMSO and the corresponding pH buffer at a volume ratio of 9:1. Add 30 μL of 1 mM fluorescent probe stock solution to each pH mixture, followed by 30 μL of hydrazine hydrate stock solution (concentration 2 × 10⁻⁶) for the corresponding pH. -2 After reacting for 30 minutes, 270 μL of HEPES buffer at the corresponding pH and 2430 μL of LDMSO were added to bring the volume to 3 mL. The fluorescence spectra were then measured at excitation wavelengths of 375 nm and 510 nm.

[0076] The test results are shown in Figure 5 (fluorescence response of the fluorescent probe and hydrazine hydrate under dual excitation conditions in different pH environments):

[0077] This probe exhibits a stable fluorescence response to hydrazine hydrate over a wide pH range, making it suitable for a broad pH range and meeting the detection needs of most real-world environmental samples.

[0078] Example 5: Testing the response performance of the probe to different concentrations of hydrazine hydrate

[0079] Prepare a mixed solution of DMSO and HEPES buffer (pH=7.4) at a volume ratio of 9:1, with a total volume of 240 μL. Add 30 μL of 1 mM fluorescent probe stock solution to the solution, followed by 1–30 equivalents of hydrazine hydrate standard solution. After reacting at room temperature for 30 min, add 270 μL of HEPES buffer and 2430 μL of DMSO to bring the volume to a final level. Measure the fluorescence emission spectra at excitation wavelengths of 375 nm and 510 nm.

[0080] The test results are shown in Figure 6 (fluorescence spectra and linear fitting plots of the response of the fluorescent probe to different concentrations of hydrazine hydrate under dual excitation conditions):

[0081] When excited at 375 nm, the fluorescence intensity at 446 nm gradually increased with increasing hydrazine hydrate concentration, exhibiting a good linear relationship within the concentration range of 0–90 μM. The linear equation was y = 88.09064x + 48.65904, with a correlation coefficient R0. 2 =0.9977, and the fluorescence intensity tends to stabilize when the concentration exceeds 100 μM.

[0082] When excited at 510 nm, the fluorescence intensity at 633 nm gradually decreases with increasing hydrazine hydrate concentration, showing an excellent linear relationship in the concentration range of 0~40 μM. The linear equation is y=-105.9333x+5253.6271 with a correlation coefficient R²=0.9902. After the concentration reaches 50 μM, the fluorescence intensity tends to stabilize.

[0083] The results demonstrate that the probe has ultra-high sensitivity to hydrazine hydrate, enabling accurate quantitative detection of trace amounts of hydrazine hydrate.

[0084] Example 6: Response time test of probe with hydrazine hydrate

[0085] Prepare a mixed solution of DMSO and HEPES buffer (pH=7.4) at a volume ratio of 9:1, with a total volume of 240 μL. Add 30 μL of 1 mM fluorescent probe stock solution and 30 μL of 7 equivalent hydrazine hydrate standard solution to the solution. At different reaction time points, add buffer and DMSO to make up the volume, and test the fluorescence spectra under excitation at 375 nm and 510 nm.

[0086] The test results are shown in Figure 7 (time-kinetic curves of the response of the fluorescent probe to hydrazine hydrate under dual excitation conditions):

[0087] The system reached equilibrium after about 18 minutes of reaction, and the fluorescence signal tended to stabilize, proving that the probe has a fast response speed and can meet the needs of rapid detection.

[0088] Example 7: Detection of hydrazine hydrate in actual soil samples

[0089] Weigh 1g of sand and 1g of field soil samples respectively, and add 2mL of each with a concentration of 5×10⁻⁶. -4 M, 1×10 -3 M, 2×10 -3 The soil extract was prepared by treating a standard solution of hydrazine hydrate (M) at room temperature for 10 min, followed by centrifugation at 2000 r / min to remove residue. 300 μL of the clarified extract was then added to 2670 μL of DMSO and mixed thoroughly to prepare a solution with a concentration of 5 × 10⁻⁶ m² / L. -5 M, 1×10 -4M, 2×10 -4 Add 30 μL of 1 mM fluorescent probe stock solution to M; place the system under a 365 nm UV lamp for fluorescence imaging at 1 min, 5 min, 10 min, 20 min and 30 min respectively to investigate the response time and visualization detection effect of the probe with hydrazine hydrate in actual soil matrix.

[0090] The test results are shown in Figure 8 (visualized imaging of the response of the fluorescent probe to hydrazine hydrate in different soil matrix extracts):

[0091] Figures 8(a) and (d) show low-concentration hydrazine hydrate, Figures 8(b) and (e) show medium-concentration hydrazine hydrate, and Figures 8(c) and (f) show high-concentration hydrazine hydrate. As the reaction time increases, the samples containing hydrazine hydrate gradually show obvious fluorescence changes. Different concentrations of hydrazine hydrate show distinguishable differences in fluorescence signals, proving that the probe can be used for the visual detection of hydrazine hydrate in actual soil samples.

[0092] Example 8: Preparation and application of hydrazine hydrate test strips

[0093] Cut the filter paper into 1.5cm × 2cm test paper substrates, and immerse the substrates in a solution with a concentration of 1 × 10⁻⁶. -3 The substrate is immersed in the fluorescent probe solution of M at room temperature for 2 hours. After immersion, it is removed and dried in an environment of 40~60℃ to obtain the test paper loaded with fluorescent probe.

[0094] (1) Detection of liquid hydrazine hydrate: Use a cotton swab to apply a solution of 1×10⁻⁶ hydrazine hydrate. -2 The M standard solution of hydrazine hydrate is marked on the test paper (e.g., LCU), dried, and then coated with fluorescent probe stock solution. After about 1 minute, the red color of the hydrazine hydrate contact area becomes significantly lighter. When irradiated under a 365nm ultraviolet lamp, the hydrazine hydrate area shows a characteristic fluorescent signal that can be clearly distinguished with the naked eye.

[0095] (2) Detection of gaseous hydrazine hydrate: Take a 5 mL glass vial, add 200 μL of 1% hydrazine hydrate solution, fix the test paper to the mouth of the vial and seal it; place the vial in a 60℃ oven and keep it at a constant temperature for 3 min. After taking it out, observe that the original red color of the test paper has completely faded; when placed under a 365 nm ultraviolet lamp, the test paper shows a significant fluorescence change.

[0096] The test results are shown in Figure 9 (visualization of the response of the fluorescent probe test strip to liquid and gaseous hydrazine hydrate):

[0097] Figure 9(a) shows the natural light appearance of the blank filter paper, Figure 9(b) shows the natural light effect of liquid hydrazine hydrate detection, Figure 9(c) shows the ultraviolet light effect of liquid hydrazine hydrate detection, and Figure 9(d) shows the comparative effect of gaseous hydrazine hydrate detection. This test strip can simultaneously achieve visual detection of liquid and gaseous hydrazine hydrate, is easy to operate, and has a rapid response, making it suitable for rapid on-site screening scenarios.

[0098] Example 9: Test of hydrazine hydrate in actual water samples

[0099] HEPES buffer solutions were prepared separately using river water and tap water, and hydrazine hydrate standard solution was added to prepare solutions with a concentration of 2×10⁻⁶. -3 M, 3×10 -3 M, 4×10 -3 Spiked water sample M.

[0100] The spiked water sample was mixed with 240 μL DMSO and 30 μL fluorescent probe stock solution to make the final probe concentration 10 μM and the hydrazine hydrate concentrations 20 μM, 30 μM, and 40 μM, respectively. After reacting for 30 minutes, the solution was diluted to volume with DMSO:HEPES = 9:1. The fluorescence intensity was measured at two excitation wavelengths, and the recovery rate and relative standard deviation were calculated. The results are shown in Tables 1 and 2 below.

[0101] Table 1: Results of Spiked Recycling Experiments in Tap Water and River Water (n=3) (λex=375nm)

[0102]

[0103] Table 2: Results of spiked water and river water recovery experiments (n=3) (λex=510nm)

[0104]

[0105] Experimental results show that the probe has good recovery rate in actual water samples, small relative standard deviation, and excellent detection accuracy and precision, and can be used for the quantitative detection of hydrazine hydrate in actual environmental water samples.

[0106] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any other way. Any person skilled in the art may make changes or modifications to the above-disclosed technical content to create equivalent embodiments for application in other fields. However, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A dual-excitation fluorescent probe for detecting hydrazine hydrate, characterized in that, The dual-excitation fluorescent probe has the chemical structure shown in Formula I: (Equation I).

2. A method for preparing a dual-excitation fluorescent probe for detecting hydrazine hydrate as described in claim 1, characterized in that, Includes the following steps: (1) 4-chloro-7-(dimethylamino)-3-carboxycoumarin was mixed with ethanol, triethylamine was added, and the mixture was heated under reflux. The reaction product was separated and purified to obtain the intermediate 4-ethoxy-7-(dimethylamino)-3-carboxycoumarin. (2) The intermediate obtained in step (1) is mixed with 3-ethyl-2-methylbenzothiazole iodide, anhydrous ethanol is added, and the mixture is heated under reflux. The reaction product is separated and purified to obtain the dual-excited fluorescent probe.

3. The method for preparing a dual-excitation fluorescent probe for detecting hydrazine hydrate according to claim 2, characterized in that, The reaction temperature of heating and reflux in steps (1) and (2) is 85℃~95℃, and the reaction time is 6~8 hours. The molar ratio of 4-ethoxy-7-(dimethylamino)-3-carboxycoumarin to 3-ethyl-2-methylbenzothiazole iodide in step (2) is 1:(0.9~1.1).

4. The method for preparing a dual-excitation fluorescent probe for detecting hydrazine hydrate according to claim 3, characterized in that, The reaction temperature in steps (1) and (2) is 90°C and the reaction time is 7 hours. The molar ratio of 4-ethoxy-7-(dimethylamino)-3-carboxycoumarin to 3-ethyl-2-methylbenzothiazole iodide is 1:

1.

5. The method for preparing a dual-excitation fluorescent probe for detecting hydrazine hydrate according to claim 2, characterized in that, The separation and purification in step (1) is carried out by column chromatography, and the eluent is a mixture of dichloromethane and ethyl acetate in a volume ratio of 3:1; the separation and purification in step (2) is carried out by column chromatography, and the eluent is a mixture of dichloromethane and methanol in a volume ratio of 20:

1.

6. The method for preparing a dual-excitation fluorescent probe for detecting hydrazine hydrate according to claim 2, characterized in that, In step (1), ethanol is used as both a reactant and a solvent.

7. The application of the dual-excitation fluorescent probe as described in claim 1 in the detection of hydrazine hydrate, characterized in that, The hydrazine hydrate detection targets include one or more of the following: water samples, soil samples, and food samples.

8. The application of the dual-excitation fluorescent probe according to claim 7 in the detection of hydrazine hydrate, characterized in that, The detection method employs a dual-excitation fluorescence detection approach: using 370-380nm as the first excitation wavelength to detect fluorescence intensity changes at 440-450nm, and using 505-515nm as the second excitation wavelength to detect fluorescence intensity changes at 630-640nm. As the concentration of hydrazine hydrate increases, the fluorescence intensity at 440-450nm gradually increases, while the fluorescence intensity at 630-640nm gradually decreases.

9. The application of the dual-excitation fluorescent probe according to claim 8 in the detection of hydrazine hydrate, characterized in that, The first excitation wavelength is 375nm, corresponding to a detection emission wavelength of 446nm, and the second excitation wavelength is 510nm, corresponding to a detection emission wavelength of 633nm.

Images