A method for semi-quantitative detection of singlet oxygen using 9,10-anthracenediyl-bis(methylene)dimalonic acid

Abstract

The present invention discloses a method for semi-quantitatively detecting singlet oxygen using 9,10-anthracenediyl-di(methylene)dimalonic acid, and relates to the field of reactive oxygen species detection technology. The core technology of the present invention is: using a NaClO / H2O2 system as a standard singlet oxygen system, using a liquid chromatograph to accurately detect the characteristic product ABDAO2 produced when 9,10-anthracenediyl-di(methylene)dimalonic acid is oxidized by singlet oxygen, and establishing a standard curve for the singlet oxygen equivalent concentration, thereby semi-quantitatively detecting the singlet oxygen content in water. This detection method can accurately identify whether singlet oxygen is present in water and semi-quantitatively measure it. It has the advantages of good reproducibility, high sensitivity, no influence from the pH of the reaction system, strong specificity, and a low threshold for use. It achieves consistency in cross-research data and has broad application prospects in the field of water pollution prevention and control. It is particularly suitable for performance evaluation and reaction mechanism research of advanced oxidation processes based on singlet oxygen.

Description

Technical Field

[0001] The present invention relates to the technical field of active oxygen detection, and in particular to a method for semi-quantitatively detecting singlet oxygen using 9,10-anthracenediyl-bis(methylene)dimalonic acid. Background Art

[0002] Singlet oxygen ( 1 O2) is an excited state of oxygen molecules, in which the electrons of oxygen molecules are at a higher energy level than the ground state and have strong oxidizing properties. Compared with other ROS, 1 O2 oxygen has some unique advantages. 1 O2 has a long life span in water and has selective oxidation properties, and is less affected by water quality background interference. 1 O2-dominated reactions have been widely developed to improve the treatment efficiency of sewage systems ADDIN EN.CITE ADDIN EN.CITE.DATA, but currently 1 There are significant challenges on the O2 side that hinder the 1 The development and application of O2 advanced oxidation process 1 Understanding of the O2 decontamination mechanism. Monitoring at 1270 nm 1 Phosphorescence emission during O2 relaxation is evidenced by 1 The most direct evidence for the existence of O2. However, to detect 1 The weak emission signal of O2 requires the use of a precise phosphorescence spectrometer. 1 O2 has not been widely used in research and application. Currently, it is mainly based on the following indirect methods to indicate 1 O2 generation: (i) using sterically hindered amine (TEMP) as a probe and electron paramagnetic resonance (EPR) technology; (ii) using 1 The inhibitory effect of adding O2 quenchers (such as L-histidine, furfuryl alcohol (FFA), sodium azide (NaN3), etc.) on the degradation of pollutants; (iii) the use of 1 The lifetime of O2 in heavy water (D2O) is longer than that in light water (H2O), so the effect of D2O / H2O ratio on the degradation of pollutants was investigated; (iv) 1 Chemical probe of O2.

[0003] exist 1In the process of O2 identification, the above-mentioned indirect detection methods have been used for a long time and widely. However, recent studies have shown that the above-mentioned methods still have obvious defects, which can easily lead to false positive results. Although the traditional probe method (such as the use of 9,10-diphenylanthracene) is simple, it is limited by its specificity and stability in complex reaction systems. It is easily interfered with by other active oxygen species (such as hydroxyl radicals, superoxide anions, etc.), resulting in non-exclusive results. The application of chemical quenching methods such as NaN3 also faces the problem of poor selectivity, because it may quench other free radical substances at the same time, thereby causing misjudgment. Although electron paramagnetic resonance (EPR) provides a more direct detection path, it has harsh requirements on experimental conditions and low sensitivity. In addition, 1 The semi-quantitative detection of O2 still lacks a unified standard: the quantitative benchmarks of different methods (EPR, fluorescence, chemiluminescence) have not yet been unified, resulting in poor comparability of data across studies.

[0004] Therefore, it is urgent to develop a method that is highly selective, operable, and can 1 A new method for semi-quantitative detection of O2 is proposed to overcome the problems of high false positive rate, poor selectivity and difficulty in quantification in existing technologies. 1 The research on O2-related mechanisms and its practical application provide more reliable technical support. Based on this background, the present invention proposes a method of using 9,10-anthracenediyl-bis(methylene)dimalonic acid (ABDA) and 1 O2 reacts specifically to form a stable adduct, which is detected by liquid chromatography to achieve 1 The semi-quantitative detection method of O2 has good selectivity, reproducibility and adaptability, and effectively solves the shortcomings of existing technologies. Summary of the Invention

[0005] The present invention provides a method for semi-quantitative detection of 1 O2 method, which is to 1 The characteristic products produced by the oxidation of ABDA by O2 were semi-quantitatively analyzed to reflect the 1 O2 concentration level. The core technology of this invention is: using NaClO / H2O2 system as the standard 1 O2 system, accurate detection using liquid chromatography 1 The characteristic product ABDAO2 is produced when ABDA is oxidized by O2. 1 The standard curve of O2 equivalent concentration is used to semi-quantitatively detect the concentration of O2 in water. 1 O2 content.

[0006] The object of the present invention is to provide a method for semi-quantitatively detecting singlet oxygen using 9,10-anthracenediyl-bis(methylene)dimalonic acid, comprising the following steps:

[0007] reacting the reactant of the system to be detected with 9,10-anthracenediyl-bis(methylene)dimalonic acid at a pH of 2 to 12 to obtain a reaction solution;

[0008] The content of ABDAO2 in the reaction solution was detected by liquid chromatography to achieve semi-quantitative detection of singlet oxygen in the system to be detected.

[0009] In some embodiments of the present invention, the initial concentration of 9,10-anthracenediyl-bis(methylene)dimalonic acid is greater than 2 μM.

[0010] In some embodiments of the present invention, the reaction time is 1-60 min.

[0011] In some embodiments of the present invention, the specific method for performing semi-quantitative detection is:

[0012] A series of standard singlet oxygen generating systems with different concentrations were reacted with 9,10-anthracenediyl-bis(methylene)dimalonic acid at a pH of 8-12 to obtain a reaction solution containing ABDAO2;

[0013] The content of ABDAO2 in the reaction solution was detected by liquid chromatography, and a standard curve was drawn according to the chromatographic peak area of ​​ABDAO2.

[0014] In some embodiments of the present invention, the standard singlet oxygen generation system is H2O2 / NaClO.

[0015] In some embodiments of the present invention, the initial concentrations of H2O2 and NaClO in the standard singlet oxygen generation system are equal, and the concentrations of H2O2 or NaClO are 0.2-1.0 mM, respectively, and further, preferably 0.2, 0.4, 0.6, 0.8, 1.0 mM, etc.

[0016] In some embodiments of the present invention, the detection conditions of the liquid chromatograph are: isocratic elution mode, the eluent is an aqueous solution of 0.1% formic acid and acetonitrile, the volume ratio is 80 / 20, the flow rate is 1 mL / min, the C18 chromatographic column, and the detection wavelength is 215 nm.

[0017] In some embodiments of the present invention, if the reaction solution is alkaline, acid is added to control the pH value of the sample to 2-7 before detection by liquid chromatography. Furthermore, the acid is preferably formic acid.

[0018] The above technical solution of the present invention has the following advantages over the prior art:

[0019] (1) The present invention is realized by liquid chromatography, which has the advantages of high sensitivity, good reproducibility, simple operation, and low threshold for use.

[0020] (2) The present invention locks the 1 The chromatographic peak of the characteristic product formed by O2 can avoid the interference of other active oxygen species (hydroxyl radicals, sulfate radicals, high-valent metals, etc.). 1 Specific detection in reaction systems with coexistence of O2 1 O2.

[0021] (3) The present invention uses the NaClO / H2O2 system as a standard 1 O2 system, established 1 A universal measurement system for semi-quantitative O2 detection to achieve consistency in cross-study data. 1 O2 advanced oxidation technology in water 1 The content of O2 was semi-quantitatively determined. 1 Efficacy and mechanism of O2 advanced oxidation technology. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to make the content of the present invention more clearly understood, the present invention is further described in detail below based on specific embodiments of the present invention in conjunction with the accompanying drawings, wherein:

[0023] Figure 1 This is a liquid chromatogram of the reaction between ABDA and the NaClO / H2O2 system in Example 1 of the present invention;

[0024] Figure 2 In Example 1 of the present invention 1 Standard curve of O2 equivalent concentration;

[0025] Figure 3 ABDA and MoO4 in Example 1 of the present invention 2- Liquid chromatogram of the reaction of / H2O2 system;

[0026] Figure 4 This is a liquid chromatogram of the reaction of ABDA with the photo-RB system at different pH values ​​in Example 2 of the present invention;

[0027] Figure 5 ABDA and Fe in Example 3 of the present invention 2+ / Liquid chromatogram of PMS system reaction;

[0028] Figure 6 ABDA and Fe in Example 4 of the present invention 2+ Liquid chromatogram of the reaction of / H2O2 system. DETAILED DESCRIPTION

[0029] The present invention will be further described below with reference to the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it. However, the embodiments are not intended to limit the present invention.

[0030] Example 1

[0031] This embodiment provides a method for semi-quantitative detection of MoO4 by ABDA 2- / H2O2 system 1 The method for O2 content, the specific steps are as follows:

[0032] Step (1), 0.2, 0.4, 0.6, 0.8, and 1.0 mM H2O2 were mixed with a solution containing 20 μM ABDA, and then NaClO with the same concentration as H2O2 was added to obtain reaction solutions containing different concentrations of ABDAO2. The reaction time was 1 min, the reaction pH was 10, and the reaction temperature was 25°C.

[0033] Step (2): Use liquid chromatography to locate the chromatographic peak of ABDAO2 produced by the reaction solution in step (1); and prepare a standard curve with the chromatographic peak area of ​​ABDAO2 as the ordinate and the concentration of NaClO / H2O2 as the abscissa. The results are as follows: Figure 2 As shown, the obtained curve is: y=320616x, R 2 =0.996, further indicating that the peak area of ​​ABDAO2 and the concentration of NaClO / H2O2 are linearly related, while H2O2+NaClO→ 1 O2+H2O+NaCl, so the concentration of NaClO / H2O2 is 1 The concentration of O2 is positively correlated, so the concentration of NaClO / H2O2 is used to express 1 The equivalent concentration of O2.

[0034] Step (3), mix 5 mM H2O2 with 20 μM ABDA, and then add 0.1 mM MoO4 2- , to obtain a reaction solution, the reaction time is 20 min, the reaction pH is 10, and the reaction temperature is 25 ℃.

[0035] Step (4), detecting the chromatographic peak of ABDAO2 in the reaction solution in step (3) using a liquid chromatograph;

[0036] Step (5) Identify whether the reaction system contains ABDAO2 based on its chromatographic peak area. 1 O2, and semi-quantitative analysis was performed based on the standard curve obtained in step (2).

[0037] The results are as follows Figure 3 As shown, Figure 3 Shows ABDA and MoO4 2- Liquid chromatogram after the reaction of / H2O2 system. ABDAO2 chromatographic peak appeared after the reaction, indicating that the reaction system 1 O2, whose peak area is 209772, is substituted into the standard curve to know that the system produces 1 The O2 equivalent concentration is 0.654 mM NaClO / H2O2.

[0038] Example 2

[0039] This example is similar to Example 1, except that the detection system is replaced by the light RB system. Using a 530 nm light source to irradiate Rose Bengal (RB) can produce a wide pH range. 1 O2, so the light RB was selected to react with ABDA at pH 4, 7, and 10. The details are as follows:

[0040] Step (3): 5 μM RB and 2 μM ABDA were mixed, and the mixture was irradiated with a light source having a wavelength of 530 nm for 2 min to obtain a reaction solution.

[0041] The results are as follows Figure 4 As shown, Figure 4 It shows that at different pH, the chromatographic peak of ABDAO2 appears after ABDA reacts with the photo-RB system, that is, the reaction system exists 1 O2, whose peak areas are respectively pH 4: 160173, pH 7: 183958, pH10: 192354. Substituting into the standard curve, it can be seen that the system produces 1 The O2 equivalent concentrations are respectively 0.500 mM at pH 4, 0.574 mM at pH 7, and 0.600 mM NaClO / H2O2 at pH 10. This embodiment shows that the method of the present invention has the effect of ionizing heat under a wide range of pH conditions. 1 O2 detection capability.

[0042] Example 3

[0043] This example is similar to Example 1, except that the system to be detected is replaced by Fe 2+ / H2O2 reaction system, specifically as follows:

[0044] Step (3) Mix H2O2 and ABDA, then add Fe 2+ The reaction solution was obtained. The reaction time was 5 min and the reaction pH was 4. 2+The initial concentrations of , H2O2 and ABDA were 50 μM, 200 μM and 20 μM respectively. Figure 5 In this embodiment, Fe 2+ The / H2O2 reaction system is a typical •OH-based advanced oxidation technology. Figure 5 Shown in this non 1 In the advanced oxidation system of O2, no chromatographic peak of ABDAO2 is generated after the reaction. This shows that the detection method of the present invention can specifically detect 1 O2.

[0045] Example 4

[0046] This example is similar to Example 1, except that the system to be detected is replaced by Fe 2+ / PMS reaction system, specifically as follows:

[0047] Different from Example 1, in step (3) of this embodiment, PMS and ABDA are mixed, and Fe is added. 2+ The reaction solution was obtained. The reaction time was 5 min and the reaction pH was 4. 2+ The initial concentrations of , PMS, and ABDA were 50 μM, 200 μM, and 20 μM, respectively.

[0048] In this embodiment, Fe 2+ The / PMS reaction system is a typical Fe(IV)-based advanced oxidation technology. Figure 6 Shown in this non 1 In the advanced oxidation system of O2, no chromatographic peak of ABDAO2 is generated after the reaction. This shows that the detection method of the present invention can specifically detect 1 O2.

[0049] Obviously, the above embodiments are merely examples for clarity of explanation and are not intended to limit the implementation methods. Those skilled in the art will appreciate that other variations or modifications can be made based on the above description. It is not necessary and impossible to enumerate all implementation methods here. Obvious variations or modifications arising therefrom remain within the scope of protection of the present invention.

Claims

1. A method for semi-quantitative detection of singlet oxygen using 9,10-anthracenediyl-bis(methylene)dimalonic acid, characterized in that: The following steps are involved: reacting the reactant of the system to be detected with 9,10-anthracenediyl-bis(methylene)dimalonic acid at a pH of 2 to 12 to obtain a reaction solution; The content of ABDAO2 in the reaction solution is detected by liquid chromatography to achieve semi-quantitative detection of singlet oxygen in the system to be detected; The detection conditions of the liquid chromatograph are: isocratic elution mode, eluent: 0.1% formic acid in water and acetonitrile, volume ratio = 80 / 20, flow rate: 1 mL / min, C18 chromatographic column, detection wavelength: 215 nm; The reactant system of the system to be detected is H2O2 / NaClO.

2. The method according to claim 1, characterized in that The initial concentration of the 9,10-anthracenediyl-bis(methylene)dimalonic acid is greater than 2 μM.

3. The method according to claim 1, characterized in that The reaction time is 1-60 min.

4. The method according to claim 1, wherein Specific methods for semi-quantitative detection: A series of standard singlet oxygen generating systems with different concentrations were reacted with 9,10-anthracenediyl-di(methylene)dimalonic acid at a pH of 8 to 12 to obtain a reaction solution containing ABDAO2. The content of ABDAO2 in the reaction solution was detected by liquid chromatography, and a standard curve was drawn according to the chromatographic peak area of ​​ABDAO2; The standard singlet oxygen generation system is H2O2 / NaClO.

5. The method according to claim 4, characterized in that The initial concentrations of H2O2 and NaClO in the standard singlet oxygen generation system are equal, and the concentrations of H2O2 or NaClO are 0.2-1.0 mM, respectively.

6. The method according to claim 1, characterized in that If the reaction solution is alkaline, acid is added to control the pH value of the sample to 2-7 before detection by liquid chromatography.

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