Aluminum ion fluorescent probe as well as preparation method and application thereof
By grafting benzoylhydrazide on carboxymethyl chitosan, an aluminum ion fluorescent probe was prepared, which solved the problems of poor water solubility and poor specificity in aluminum ion detection by existing small molecule fluorescent probes, and achieved efficient and specific detection of aluminum ions.
Patent Information
- Application Number
- CN202411294547.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-09-14
- Publication Date
- 2025-03-28
AI Technical Summary
The existing small molecule fluorescent probes have problems of poor water solubility and poor specificity in aluminum ion detection, which limits their application effect.
Carboxymethyl chitosan is used as a support, and benzohydrazide is grafted on carboxymethyl chitosan through Schiff base condensation reaction to prepare an aluminum ion fluorescent probe, which improves its water solubility and selectivity to aluminum ions.
Highly efficient and specific detection of aluminum ions is achieved, with the minimum detection limit of aluminum ions being 0.33 μmol/L, which can maintain anti-interference under the conditions of the presence of a variety of cations and anions.
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Abstract
Description
Technical Field
[0001] The invention relates to the technical field of aluminum ion detection, in particular to an aluminum ion fluorescent probe and a preparation method and application thereof. Background Art
[0002] Al 3+ It is one of the non-essential trace elements in the human body and organisms. Excessive intake of aluminum ions may cause nervous system and bone diseases. As an industrial pollutant, monitoring its environmental concentration helps to assess the impact of industrial activities on the ecology. In addition, by detecting the aluminum content in water and food, it can also ensure water quality and food safety and protect public health. Therefore, it is necessary to develop a fast, accurate and environmentally friendly Al 3+ The detection method is of vital importance.
[0003] The prior art discloses a variety of small molecule fluorescent probes of various types and functions. Although small molecule fluorescent probes play an important role in many fields, they still have some limitations, such as poor water solubility and poor specificity, which limit the application of small molecule fluorescent probes in aluminum ion detection. Summary of the invention
[0004] In view of the deficiencies in the above-mentioned prior art, the present invention provides an aluminum ion fluorescent probe and a preparation method and application thereof. The aluminum ion fluorescent probe of the present invention uses carboxymethyl chitosan as a carrier, uses benzoyl hydrazide as a fluorescent material, and grafts the fluorescent material onto the carboxymethyl chitosan. Carboxymethyl chitosan improves the water solubility and selectivity of the fluorescent material to aluminum ions. The aluminum ion fluorescent probe is used as a fluorescent probe to overcome the problems of poor water solubility and poor specificity of the small molecule fluorescent probe in the prior art, so that the minimum detection limit of aluminum ions is 0.33 μmol / L, and anti-interference specific detection of aluminum ions can be achieved under the conditions of the presence of a variety of cations and anions.
[0005] To achieve the above object, the technical solution of the present invention is as follows:
[0006] The present invention protects an aluminum ion fluorescent probe, which is a carboxymethyl chitosan derivative, and has the chemical structural formula: Wherein, n represents the degree of polymerization of carboxymethyl chitosan, and its value range is 100-1000.
[0007] The present invention also protects a method for preparing the aluminum ion fluorescent probe, comprising the following steps:
[0008] Carboxymethyl chitosan and glyoxal are used as raw materials. The two are mixed in water and subjected to Schiff base condensation reaction. Then, according to the principle of like dissolves like, ethanol is used for precipitation to obtain an intermediate compound. The process of this step is That is, an aldehyde group of glyoxal is grafted onto carboxymethyl chitosan.
[0009] Intermediate compounds and benzoyl hydrazide as raw materials, the two are mixed in water and subjected to Schiff base condensation reaction, the solvent is removed, and the aluminum ion fluorescent probe is obtained by Soxhlet extraction; the process of this step is: That is, the grafting of benzoyl hydrazide onto carboxymethyl chitosan was achieved through glyoxal.
[0010] Preferably, the mass ratio of carboxymethyl chitosan to glyoxal is 1.0-1.2:1.2-1.44, and the amount of glyoxal is higher than that of carboxymethyl chitosan, so as to ensure that more glyoxal is connected to carboxymethyl chitosan.
[0011] Preferably, the synthetic intermediate compound The conditions for the Schiff base condensation reaction are: reflux reaction for 4-6 hours. If the reaction time is too short, the reaction will be incomplete, and if the reaction time is too long, it will waste time and energy.
[0012] Preferably, the intermediate compound The mass ratio of benzoyl hydrazide to benzoyl hydrazide is 0.5-0.6:0.16-0.19. The purpose of this ratio range is to ensure that more benzoyl hydrazide is connected to the intermediate compound as much as possible. superior.
[0013] Preferably, when synthesizing the aluminum ion fluorescent probe, the conditions for the Schiff base condensation reaction are: reflux reaction for 6-8 hours. If the reaction time is too short, the reaction is incomplete, and if the reaction time is too long, time is wasted and energy is consumed.
[0014] Preferably, the extractant for Soxhlet extraction is ethanol.
[0015] The present invention also protects the application of the above aluminum ion fluorescent probe in the preparation of a detection kit or a detection test paper for aluminum ions in the detection and identification environment or in biological samples.
[0016] Preferably, the minimum detection limit of aluminum ions is 0.33 μmol / L.
[0017] Preferably, the aluminum ion fluorescent probe is used in the preparation of a kit or a test strip for detecting aluminum ions using fluorescence.
[0018] Preferably, the aluminum ion fluorescent probe is used in the preparation of a kit or a test paper for identifying the fluorescence intensity of aluminum ions in the environment or in a biological sample at a maximum emission wavelength of 464 nm through fluorescence spectroscopy.
[0019] Compared with the prior art, the present invention has the following beneficial effects:
[0020] 1. The present invention prepares an aluminum ion fluorescent probe with excellent fluorescence properties, which is denoted as a carboxymethyl chitosan derivative CMCS-P, i.e., a carboxymethyl chitosan-based benzohydrazide derivative. The aluminum ion fluorescent probe is used as an efficient probe in pure water to demonstrate the ability to detect aluminum ions (Al3+). In the aluminum ion fluorescent probe, carboxymethyl chitosan is used as an excellent carrier of the functional fluorescent probe, and benzohydrazide is used as the fluorescent material. Carboxymethyl chitosan improves the water solubility and selectivity of the fluorescent material to aluminum ions. The reason why carboxymethyl chitosan improves the selectivity of the fluorescent material is that carboxymethyl chitosan contains a large number of rich hydroxyl groups, which provide a strong affinity with Al3+. 3+ binding site.
[0021] The principle of using aluminum ion fluorescent probe for aluminum ion detection is that aluminum ion fluorescent probe can show very weak fluorescence response, which may be due to the transfer of lone pair electrons of –C=N– group to benzoyl hydrazide unit and the radiationless decay of C=N bond in excited state. 3+ The fluorescence response at 464 nm was significantly switched on and off, suggesting that the nitrogen and oxygen atoms in the Schiff base may be involved in the formation of the CMCS-P-Al complex. 3+ , which limits the isomerization of the C=N bond and the photoinduced electron transfer process, resulting in a significant enhancement of the fluorescence intensity at a wavelength of 464 nm.
[0022] 2. The present invention uses the prepared aluminum ion fluorescent probe as a high-efficiency enhanced fluorescent probe to achieve specific recognition and detection of aluminum ions, specifically: among the 16 common metal cations (Al 3+ , Fe 3+ , Fe 2+ , Cu 2+ , Pb 2+ 、Zn 2+ Mg 2+ , Ca 2+ 、Na + , K + Cr 3+ 、Cd 2+ 、Co 2+ , Hg 2+ 、Ni 2+ 、Ag + , Ba 2+ ) and 7 anions (NO3 - Br - 、ClO4 - ,I - 、CO3 2- 、H2PO4 - 、SO4 2- ) 3+After addition, the probe CMCS-P showed an obvious fluorescence peak at 464nm, accompanied by signal enhancement.
[0023] 3. In order to verify the importance of carboxymethyl chitosan in aluminum ion fluorescent probe, the present invention also conducted a comparative study to compare the ungrafted carboxymethyl chitosan with It is a fluorescent material, denoted as small molecule compound P. Small molecule compound P is prepared from benzoyl hydrazide and glyoxal. When conducting the same specific detection experiment as the aluminum ion fluorescent probe, it was found that small molecule compound P had no obvious spectral changes for all metal cations and anions, indicating that small molecule compound P has no specific detection function. This further reflects the necessity of carboxymethyl chitosan grafting, and also explains that the fluorescent probe CMCS-P specifically recognizes Al3+ and realizes the detection of 0.33μmol / L Al3+ in a pH 6.0 aqueous solution.
[0024] 4. The present invention uses carboxymethyl chitosan as a carrier and benzoyl hydrazide as a fluorescent material. Under the action of glyoxal, the fluorescent material is grafted onto the carrier through a two-step Schiff base condensation reaction to obtain an aluminum ion fluorescent probe. The preparation method of the present invention is simple and the raw materials are cheap and easily available. BRIEF DESCRIPTION OF THE DRAWINGS
[0025] Figure 1 The synthetic route of the carboxymethyl chitosan derivative CMCS-P of Example 1 and the small molecule compound P of Comparative Example 1 is shown.
[0026] Figure 2 The infrared spectra of the carboxymethyl chitosan derivative CMCS-P, carboxymethyl chitosan CMCS and the intermediate compound CMCS-1 of Example 1 are shown.
[0027] Figure 3 In the figure, a) is the fluorescence selectivity diagram of the carboxymethyl chitosan derivative CMCS-P of Example 1 for aluminum ions in different metal cations and anions; b) is the fluorescence selectivity diagram of the small molecule compound P of Comparative Example 1 for different metal cations and anions.
[0028] Figure 4 This is a graph showing the analytical performance of the carboxymethyl chitosan derivative CMCS-P in Example 1 for different aluminum ion concentrations, wherein the small graph is a graph showing the relationship between the fluorescence intensity of the carboxymethyl chitosan derivative CMCS-P and the aluminum ion concentration. DETAILED DESCRIPTION
[0029] The specific embodiments of the present invention are described in detail below, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the protection scope of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified.
[0030] The present invention uses carboxymethyl chitosan as a raw material to prepare a carboxymethyl chitosan derivative CMCS-P. The reason for using carboxymethyl chitosan as a raw material is that chitosan has the characteristics of safety and non-toxicity, large biomass, natural degradation, and easy modification. More importantly, it can be used as a carrier of small molecule fluorescent probes. Therefore, the present invention utilizes the advantages of both organic small molecule dyes and chitosan, and is designed in a targeted manner, and an environmentally friendly chitosan-based fluorescent probe that can specifically bind to a test object is prepared, thereby solving the bottleneck encountered in the development of small molecule fluorescent probes.
[0031] In addition, in the present invention, carboxymethyl chitosan is grafted with a fluorescent material. Carboxymethyl chitosan is not only used as a carrier, but also studies have found that carboxymethyl chitosan can promote the specific selection of the fluorescent material for aluminum ions, that is, through the joint action of carboxymethyl chitosan and the fluorescent material, specific detection of aluminum ions is achieved, and a high detection sensitivity is achieved at a wavelength of 464nm.
[0032] The technical solution of the present invention is studied using embodiments and comparative examples below, and the specific research methods and results are as follows:
[0033] Example 1
[0034] The preparation method of carboxymethyl chitosan derivative CMCS-P comprises the following steps:
[0035] S1. Carboxymethyl chitosan and glyoxal are mixed in water, and then refluxed for 6 hours to obtain a reaction solution. The reaction solution is precipitated with ethanol, and filtered and dried to obtain a gray intermediate compound CMCS-1.
[0036] Wherein, the mass ratio of carboxymethyl chitosan to glyoxal is 1.0g:1.2g.
[0037] The intermediate compound CMCS-1 was mixed with benzoyl hydrazide in water, and then refluxed for 8 hours. The solvent was then removed to precipitate a black-brown solid. The black-brown solid was washed with anhydrous ethanol and then subjected to Soxhlet extraction for 4 hours. Ethanol was used as the eluent for the Soxhlet extraction. After drying, a black-brown pure carboxymethyl chitosan derivative CMCS-P was obtained.
[0038] Among them, the mass ratio of the intermediate compound CMCS-1 to benzoyl hydrazide is 0.5g:0.1632g.
[0039] Example 2
[0040] The preparation method of carboxymethyl chitosan derivative CMCS-P comprises the following steps:
[0041] S1. Carboxymethyl chitosan and glyoxal are mixed in water, and then refluxed for reaction for 5 hours to obtain a reaction solution. The reaction solution is precipitated with ethanol, and filtered and dried to obtain a gray intermediate compound CMCS-1.
[0042] The mass ratio of carboxymethyl chitosan to glyoxal is 1.1 g:1.3 g.
[0043] The intermediate compound CMCS-1 was mixed with benzoyl hydrazide in water, and then refluxed for 7 hours. The solvent was then removed to precipitate a black-brown solid. The black-brown solid was washed with anhydrous ethanol and then subjected to Soxhlet extraction for 4 hours. Ethanol was used as the eluent for the Soxhlet extraction. After drying, a black-brown pure carboxymethyl chitosan derivative CMCS-P was obtained.
[0044] Among them, the mass ratio of the intermediate compound CMCS-1 to benzoyl hydrazide is 0.55g:0.18g.
[0045] Example 3
[0046] The preparation method of carboxymethyl chitosan derivative CMCS-P comprises the following steps:
[0047] S1. Carboxymethyl chitosan and glyoxal are mixed in water, and then refluxed for 4 hours to obtain a reaction solution. The reaction solution is precipitated with ethanol, and filtered and dried to obtain a gray intermediate compound CMCS-1.
[0048] The mass ratio of carboxymethyl chitosan to glyoxal is 1.2 g:1.44 g.
[0049] The intermediate compound CMCS-1 was mixed with benzoyl hydrazide in water, and then refluxed for 6 hours. The solvent was then removed to precipitate a black-brown solid. The black-brown solid was washed with anhydrous ethanol and then subjected to Soxhlet extraction for 4 hours. Ethanol was used as the eluent for the Soxhlet extraction. After drying, a black-brown pure carboxymethyl chitosan derivative CMCS-P was obtained.
[0050] Among them, the mass ratio of the intermediate compound CMCS-1 to benzoyl hydrazide is 0.6g:0.19g.
[0051] Example 4
[0052] The preparation method of carboxymethyl chitosan derivative CMCS-P comprises the following steps:
[0053] S1. Carboxymethyl chitosan and glyoxal are mixed in water, and then refluxed for reaction for 5 hours to obtain a reaction solution. The reaction solution is precipitated with ethanol, and filtered and dried to obtain a gray intermediate compound CMCS-1.
[0054] The mass ratio of carboxymethyl chitosan to glyoxal is 1.1 g:1.4 g.
[0055] The intermediate compound CMCS-1 was mixed with benzoyl hydrazide in water, and then refluxed for 8 hours. The solvent was then removed to precipitate a black-brown solid. The black-brown solid was washed with anhydrous ethanol and then subjected to Soxhlet extraction for 4 hours. Ethanol was used as the eluent for the Soxhlet extraction. After drying, a black-brown pure carboxymethyl chitosan derivative CMCS-P was obtained.
[0056] The mass ratio of the intermediate compound CMCS-1 to benzoyl hydrazide is 0.6 g:0.17 g.
[0057] Comparative Example 1
[0058] The preparation method of the small molecule compound P, wherein the small molecule compound P is prepared from benzoyl hydrazide and glyoxal, is different from the embodiment in that benzoyl hydrazide is not grafted onto carboxymethyl chitosan, and comprises the following steps:
[0059] Benzoyl hydrazide and glyoxal are used as raw materials, the two are mixed in anhydrous ethanol and subjected to Schiff base condensation reaction, and then filtered to obtain a small molecule compound P.
[0060] The mass ratio of benzoyl hydrazide to glyoxal is 1.0-1.2:3-3.6, and the conditions for the Schiff base condensation reaction are: reflux for 4-6 hours.
[0061] In this embodiment, the reflux reaction was performed for 4 h, and the mass ratio of benzoyl hydrazide to glyoxal was 450 mg:1500 mg.
[0062] Examples 1-4 of the present invention all prepared aluminum ion fluorescent probes with water solubility and excellent specific detection performance for aluminum ions. The following is a study using the carboxymethyl chitosan derivative CMCS-P obtained in Example 1 and the small molecule compound P obtained in Comparative Example 1 as examples. The specific research methods and results are as follows:
[0063] The structural formula of carboxymethyl chitosan derivative CMCS-P is: The structural formula of the small molecule compound P is: The synthetic routes of the two are Figure 1 shown.
[0064] Figure 2 This is the infrared spectrum of the carboxymethyl chitosan derivative CMCS-P. Figure 2From the infrared absorption spectrum, we can see that the intermediate compound CMCS-1 and the carboxymethyl chitosan derivative CMCS-P were obtained. The infrared spectrum data of the two are as follows:
[0065] CMCS-1: IR (KBr, cm -1 ): 3461.42,2930.23,2355.45,1645.44,1400.59,1326.59,1261.14,1250.91,1113.86,1072.01,906.67,708.98,651.25,613.10,587,552.94.
[0066] CMCS-P:IR(KBr,cm -1 ): 3402.58, 3224.77, 5055.00, 2928.18, 2355.45, 1652.05, 1604.77, 1576.14, 1535.86, 1490.23, 1402.01, 1314.08, 1278.64, 1148.64, 1109.77, 1071.20, 906.82, 798.86, 693.92. The infrared spectrum shows that the target structure product is obtained in the present invention.
[0067] The following is a test of the optical recognition of aluminum ions by the carboxymethyl chitosan derivative CMCS-P:
[0068] 1. Specific optical property determination:
[0069] S1. Add deionized water to a 20 mmol / L HEPES buffer solution and adjust the pH of the system to 6.0 to obtain a test solution.
[0070] S2, take some 50 μL of the aqueous solution of the carboxymethyl chitosan derivative CMCS-P (2000ppm), and then add them to 5mL colorimetric tubes, respectively, and then add the same concentration of metal cations or anions (the concentration is 0.01mol / L) thereto, and dilute to the scale of 5mL with the detection solution, shake well, and the carboxymethyl chitosan derivative CMCS-P after dilution is 20ppm. Test different colorimetric tubes, the test conditions are: excitation / emission slit = 10 / 10nm, excitation wavelength is 350nm.
[0071] S3, Figure 3 (a) is the fluorescence selectivity spectrum of the probe carboxymethyl chitosan derivative CMCS-P. After adding the same concentration of metal cations or anions to the water (20 mmol / L HEPES buffer solution, pH 6.0) solution of the probe carboxymethyl chitosan derivative CMCS-P (20 ppm), the metal cations are selected from (Al 3+, Fe 3+ , Fe 2+ , Cu 2+ , Pb 2+ 、Zn 2+ Mg 2+ , Ca 2+ 、Na + , K + Cr 3+ 、Cd 2+ 、Co 2+ , Hg 2+ 、Ni 2+ 、Ag + Or Ba 2+ ), the anion is selected from (NO3 - Br - 、ClO4 - ,I - 、CO3 2- 、H2PO4 - or SO4 2- ), only Al 3+ The addition of CMCS-P resulted in an increase in the fluorescence peak intensity at 464 nm, indicating that the carboxymethyl chitosan derivative CMCS-P can achieve the effect of Al 3+ Excellent anti-interference and specific detection.
[0072] Figure 3 (b) is the fluorescence selectivity spectrum of the small molecule compound P. After adding the same concentration of metal cations or anions to the ethanol solution of the probe small molecule compound P (10 μmol / L), the results show that there is no obvious signal change for both metal cations and anions, indicating that carboxymethyl chitosan not only improves the selectivity and water solubility of the probe carboxymethyl chitosan derivative CMCS-P, but more importantly, carboxymethyl chitosan is indispensable.
[0073] 2. Detection limit determination:
[0074] S1. Add deionized water to a 20 mmol / L HEPES buffer solution and adjust the pH of the system to 6.0 to obtain a test solution.
[0075] S2. Pipette some 50 μL of the aqueous solution of the carboxymethyl chitosan derivative CMCS-P (2000 ppm), and then add them into 5 mL colorimetric tubes respectively, and then add different concentrations of aluminum ion standard solutions thereto respectively, dilute to the scale of 5 mL with the detection solution, shake well, and after dilution, the carboxymethyl chitosan derivative CMCS-P is 20 ppm. Test different colorimetric tubes, and the test conditions are: excitation / emission slit = 10 / 10 nm, and the excitation wavelength is 350 nm.
[0076] S3, Figure 4The fluorescence titration spectrum of aluminum ions is shown in Figure 1. After adding 1-20 μmol / L aluminum ions to the water (20 mmol / L HEPES buffer, pH 6.0) solution of the probe carboxymethyl chitosan derivative CMCS-P (20 ppm), the fluorescence peak intensity at 464 nm gradually increases with the increase of aluminum ion concentration. This may be due to the coordination effect between the aluminum ions and the probe carboxymethyl chitosan derivative CMCS-P after the carboxymethyl chitosan derivative CMCS-P is complexed with aluminum ions, which induces signal enhancement. The linear range of the probe carboxymethyl chitosan derivative CMCS-P's response to aluminum ions is 1-20 μmol / L, and the detection limit is 0.33 μmol / L.
[0077] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.
Claims
1. Aluminum ion fluorescent probe, characterized in that: The aluminum ion fluorescent probe is a carboxymethyl chitosan derivative, and its chemical structure is: Wherein, n represents the degree of polymerization of carboxymethyl chitosan, and its value range is 100-1000.
2. A method for preparing the aluminum ion fluorescent probe according to claim 1, characterized in that: The steps include: Carboxymethyl chitosan and glyoxal are used as raw materials. The two are mixed in water and subjected to Schiff base condensation reaction. Then, ethanol is used for precipitation to obtain an intermediate compound. Intermediate compounds and benzoyl hydrazide as raw materials, which are mixed in water and subjected to Schiff base condensation reaction, and then the solvent is removed and the aluminum ion fluorescent probe is obtained through Soxhlet extraction.
3. The method for preparing the aluminum ion fluorescent probe according to claim 2, characterized in that: The mass ratio of carboxymethyl chitosan to glyoxal is 1.0-1.2:1.2-1.
44.
4. The method for preparing the aluminum ion fluorescent probe according to claim 2, characterized in that: Synthetic intermediate compounds The conditions for the Schiff base condensation reaction are: reflux reaction for 4-6 hours.
5. The method for preparing the aluminum ion fluorescent probe according to claim 2, characterized in that: Intermediate Compounds The mass ratio of benzoyl hydrazide to benzoyl hydrazide is 0.5-0.6:0.16-0.
19.
6. The method for preparing the aluminum ion fluorescent probe according to claim 2, characterized in that: When synthesizing aluminum ion fluorescent probe, the conditions of Schiff base condensation reaction are: reflux reaction for 6-8h.
7. The method for preparing the aluminum ion fluorescent probe according to claim 2, characterized in that: The extraction solvent for Soxhlet extraction is ethanol.
8. An application of the aluminum ion fluorescent probe according to claim 1, characterized in that: Application in the preparation of detection and identification of aluminum ion detection kits or test strips in the environment or biological samples.
9. The use of the aluminum ion fluorescent probe according to claim 8, characterized in that: Application of aluminum ion fluorescent probe in preparing a kit or test paper for detecting aluminum ions using fluorescence.
10. The use of the aluminum ion fluorescent probe according to claim 8, characterized in that: The aluminum ion fluorescent probe is used in preparing a kit or a test paper for identifying the fluorescence intensity of aluminum ions in the environment or in biological samples at a maximum emission wavelength of 464 nm through fluorescence spectroscopy.