Zeolite adsorptive membrane and preparation method thereof, combined sensor, and method for synchronously testing ammonia nitrogen and oxygen in rhizosphere of aquatic plants

Abstract

The application belongs to the technical field of in-situ analysis of water environment interface elements and environmental parameters, and particularly relates to a zeolite adsorption film, a preparation method thereof, a combined sensor, and a synchronous testing method for ammonia nitrogen and oxygen in the rhizosphere of aquatic plants. The zeolite adsorption film provided by the application comprises agar hydrogel and zeolite particles dispersed in the agar hydrogel, and the particle size of the zeolite particles is less than or equal to 1 micrometer. The agar used in the application can avoid the ammonia nitrogen analysis interference caused by using acrylamide material as the film matrix material. Meanwhile, based on the particle size of the zeolite particles used in the application being less than or equal to 1 micrometer, the micro-cutting and micro-spectrophotometric analysis method using the zeolite adsorption film can realize the sub-millimeter level high spatial resolution two-dimensional testing of ammonia nitrogen. The zeolite adsorption film provided by the application is combined with an oxygen fluorescent sensing film to realize, for the first time, the sub-millimeter level two-dimensional distribution synchronous testing of ammonia nitrogen and oxygen in the rhizosphere environment interface of aquatic plants, and obtain the sub-millimeter level two-dimensional distribution images of the two at the same position in the rhizosphere, thereby providing an important basis for revealing the response mechanism of the rhizosphere ammonia nitrogen dynamic migration-absorption process and oxygen distribution.

Description

Technical Field

[0001] This invention belongs to the field of in-situ analysis technology of interface elements and environmental parameters in aquatic environments, specifically involving zeolite adsorption membranes and their preparation methods, coupled sensors, and methods for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants. Background Technology

[0002] Aquatic plants in lakes or other water bodies can absorb nitrogen and phosphorus from sediments through their rhizosphere, thereby controlling the endogenous nitrogen load in the sediments. Aquatic plants have been widely used in lake eutrophication remediation projects. Oxygen is produced by leaves through photosynthesis and then transported to leaves, stems, and roots through aerenchyma tissues. This allows aquatic plants to secrete oxygen into rhizosphere sediments. The root oxidation zone acts as a protective layer against toxins and influences the geochemical reactions of ammonia nitrogen (nitrification and denitrification). Anaerobic microenvironments favor denitrification and the reduction of dissimilar nitrates to ammonium, while inhibiting nitrification. Therefore, the rhizosphere redox environment affects the nitrogen geochemical reactions in rhizosphere sediments and the absorption of ammonia nitrogen by the rhizosphere. Simultaneously obtaining the two-dimensional high spatial distribution of ammonia nitrogen diffusion flux and oxygen concentration in rhizosphere microzones is of great significance for revealing the relationship between ammonia nitrogen distribution and oxygen response in rhizosphere microzones, as well as the kinetics of ammonia nitrogen migration and absorption in the rhizosphere.

[0003] Currently, a relatively advanced method for simultaneous high spatial resolution testing of rhizosphere elements and oxygen in aquatic plants is the combined testing of gradient diffusion membrane (DGT) and planar optical (PO) technology. This involves combining a DGT adsorption membrane, an oxygen fluorescence sensing membrane, and a filter membrane into a single sensor. The sensor is placed in the rhizosphere for 4–8 hours, and the root box containing the aquatic plants is placed in the planar optical testing area to analyze the two-dimensional distribution of oxygen concentration in the rhizosphere. Then, the sensor is removed from the root box, and the DGT adsorption membrane is removed for two-dimensional analysis of elements immobilized by the adsorption membrane. Finally, a two-dimensional image of the diffusion flux and oxygen concentration distribution of elements in the rhizosphere sediment is plotted. Currently, the DGT adsorption membrane / oxygen fluorescence membrane combined sensor is mainly used for two-dimensional spatial analysis of inorganic phosphorus, metal elements, and oxygen in the rhizosphere; for example: ZrO adsorption membrane / oxygen fluorescence membrane (used to analyze the two-dimensional distribution of inorganic phosphorus and oxygen); SPR-IDA adsorption membrane / oxygen fluorescence membrane (used to analyze the two-dimensional distribution of metal elements and oxygen); ZrO-AT adsorption membrane / oxygen fluorescence membrane (used to analyze the one-dimensional profile of ammonia nitrogen / nitrate nitrogen / phosphorus and the two-dimensional distribution of oxygen). Only one sensor, the aforementioned ZrO-AT adsorption membrane / oxygen fluorescence membrane sensor, can simultaneously measure ammonia nitrogen and oxygen. The ZrO-AT hybrid adsorption membrane used consists of three binding agents: ZrO, A-62MP, and T-42H, with T-42H used to measure ammonia nitrogen. Because the particle diameter of T-42H cation exchange resin is >100 μm, neither T-42H nor ZrO-AT adsorption membranes can be used for sub-millimeter-level two-dimensional spatial determination of ammonia nitrogen. Currently reported DGT adsorption membrane materials for measuring ammonia nitrogen in aquatic plant rhizosphere, sediment, or water include: PrCH cation exchange resin, CMI-7000 cation exchange resin, and zeolite. The binding reagents for DGT adsorption membranes used for two-dimensional high spatial resolution ammonia nitrogen measurement must have a particle diameter ≤1 μm to enable subsequent sub-millimeter-level (~100 μm) two-dimensional ammonia nitrogen analysis. However, the particle size of the aforementioned reagents for adsorbing ammonia nitrogen (T-42H, PrCH, and CMI-7000) is all greater than 100 μm. Currently, the successfully developed zeolite adsorption membranes are prepared by polymerizing zeolite powder (particle diameter: 1 μm) with acrylamide gel. Although the zeolite particle diameter is acceptable, the amino groups in the adsorption membrane interfere with the analysis of ammonia nitrogen, introducing certain analytical errors. Furthermore, existing zeolite adsorption membranes or other adsorption membranes all use conventional tools to vertically cut the gel strips, then elute with NaCl solution and analyze using Nessler's colorimetry on a spectrophotometer, only obtaining a one-dimensional profile of rhizosphere ammonia nitrogen with a resolution ≥1 mm. Therefore, there is currently no suitable method for high spatial resolution two-dimensional distribution testing of ammonia nitrogen in the rhizosphere or sediments of aquatic plants, nor is there a novel zeolite adsorption membrane preparation method that can accurately analyze ammonia nitrogen; that is: a two-dimensional high spatial resolution analysis method for ammonia nitrogen adsorbed by zeolite adsorption membranes, a method for preparing or combining zeolite adsorption membranes and zeolite adsorption membrane / oxygen fluorescence sensing membrane sensors, and a method for testing these sensors in the rhizosphere.The aforementioned zeolite adsorption membrane / oxygen fluorescence sensing membrane sensor is the first to complete the high spatial resolution two-dimensional distribution test of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants.

[0004] In summary, existing DGT adsorption membranes for ammonia nitrogen testing in the rhizosphere of aquatic plants, or sensors for in-situ ammonia nitrogen / oxygen testing, cannot simultaneously perform sub-millimeter-level two-dimensional spatial distribution testing of ammonia nitrogen and oxygen in the rhizosphere. The key problems in this field are: (1) existing technologies cannot achieve high spatial resolution two-dimensional distribution determination of ammonia nitrogen on DGT adsorption membranes; (2) the matrix material (acrylamide) of existing zeolite adsorption membranes interferes with ammonia nitrogen analysis; other adsorption membrane fixative particles have a particle size much larger than 1 μm, making it impossible to perform sub-millimeter-level ammonia nitrogen testing; and there is a lack of zeolite adsorption membrane preparation methods that can accurately perform high spatial resolution ammonia nitrogen testing. Summary of the Invention

[0005] The purpose of this invention is to provide a zeolite adsorption membrane and its preparation method, a method for combining it with sensors, a method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants, and a subsequent method for two-dimensional micro-dissection and micro-spectrophotometric analysis of the zeolite adsorption membrane. The zeolite adsorption membrane / oxygen fluorescence sensor assembled using the zeolite adsorption membrane provided by this invention can simultaneously obtain sub-millimeter-level two-dimensional distribution images of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants, providing an important foundation for revealing the dynamic migration-absorption and oxygen response mechanism of ammonia nitrogen in the rhizosphere.

[0006] To achieve the above objectives, the present invention provides the following technical solution:

[0007] The present invention provides a zeolite adsorption membrane comprising an agar hydrogel and zeolite particles dispersed in the agar hydrogel; wherein the particle size of the zeolite particles is ≤1μm.

[0008] Preferably, the mass ratio of agar to zeolite particles in the agar hydrogel is 1:(7-8).

[0009] Preferably, the mass ratio of agar to water in the agar hydrogel is 1 g: (42-50) mL.

[0010] Preferably, the thickness of the zeolite adsorption membrane is 0.01 to 0.04 cm.

[0011] This invention provides a method for preparing the zeolite adsorption membrane described above, comprising the following steps:

[0012] Zeolite granules, agar, and pure water are heated and mixed to obtain a hot mixed solution;

[0013] The hot mixed solution is poured into a mold and cooled. After demolding, the zeolite adsorption membrane is obtained.

[0014] Preferably, the initial gel membrane obtained after demolding further includes: immersing and washing the initial gel membrane with ultrapure water four times to obtain the zeolite adsorption membrane; the immersion and washing time with ultrapure water is 2 hours each time.

[0015] This invention provides the application of the zeolite adsorption membrane described in the above technical solution or the zeolite adsorption membrane prepared by the preparation method described in the above technical solution in the detection of ammonia nitrogen in the rhizosphere of aquatic plants.

[0016] The present invention provides a combined sensor, comprising a filter membrane, a zeolite adsorption membrane and an oxygen fluorescence sensing membrane stacked together; wherein the zeolite adsorption membrane is the zeolite adsorption membrane described in the above technical solution or the zeolite adsorption membrane prepared by the preparation method described in the above technical solution.

[0017] Preferably, the filter membrane is a polyvinylidene fluoride (PVDF) filter membrane.

[0018] This invention provides a method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants, comprising the following steps:

[0019] (1) Fill a root box with sediment, aquatic plants and overlying water; the root box is provided with a removable transparent sidewall; the stems and leaves of the aquatic plants are located in the overlying water, and the roots are located in the sediment; then place it in an experimental water tank for cultivation; after the aquatic plants are successfully cultivated, a sensor is set on the inner surface of the removable transparent sidewall of the root box and in the area close to the roots of the aquatic plants. The sensor is the combined sensor described in the above technical solution, and the oxygen fluorescence sensing membrane of the combined sensor is in contact with the removable transparent sidewall of the root box.

[0020] (2) The root box equipped with the sensor was placed in the experimental water tank to conduct a rhizosphere test of aquatic plants for 4 to 8 hours.

[0021] (3) Perform planar photopolar analysis on the oxygen fluorescence sensing membrane in the coupled sensor obtained after the test to obtain a two-dimensional distribution image of the oxygen concentration in the rhizosphere of aquatic plants. The spatial resolution of the two-dimensional distribution image of the oxygen concentration in the rhizosphere of aquatic plants is 62μm×62μm.

[0022] (4) The zeolite adsorption membrane in the coupled sensor after planar optical detection is analyzed to obtain a high spatial resolution two-dimensional distribution image of ammonia nitrogen diffusion flux in the rhizosphere of aquatic plants.

[0023] Preferably, the analysis of the adsorbed ammonia nitrogen includes the following steps:

[0024] The zeolite adsorption membrane is cut with spatial resolution of 200μm×200μm;

[0025] The zeolite adsorption membrane slices obtained after cutting were immersed in sodium chloride eluent for ammonia nitrogen elution. The spatial resolution of the zeolite adsorption membrane slices was 200 μm × 200 μm, and the ammonia nitrogen eluent was obtained.

[0026] The ammonia nitrogen concentration of the eluent was determined using the Sodium colorimetric method and a micro spectrophotometer, and the diffusion flux of ammonia nitrogen in the rhizosphere sediment was calculated from the ammonia nitrogen concentration.

[0027] This invention provides a zeolite adsorption membrane, comprising agar hydrogel and zeolite particles dispersed in the agar hydrogel; the particle size of the zeolite particles is ≤1 μm. The zeolite adsorption membrane provided by this invention uses agar hydrogel as the matrix material, avoiding the interference of ammonia nitrogen analysis caused by using acrylamide as the membrane matrix material; simultaneously, the particle size of the zeolite particles used in this invention is ≤1 μm, enabling sub-millimeter-level high spatial resolution (200 μm × 200 μm) two-dimensional testing of ammonia nitrogen. The results of the embodiments show that, by using the zeolite adsorption membrane provided by this invention in conjunction with an oxygen fluorescence sensing membrane, the simultaneous testing of sub-millimeter-level two-dimensional distributions of ammonia nitrogen and oxygen at the rhizosphere interface of aquatic plants has been achieved for the first time, obtaining sub-millimeter-level two-dimensional distribution images of both, thus providing an important foundation for revealing the rhizosphere ammonia nitrogen migration-absorption and oxygen response mechanisms. Attached Figure Description

[0028] Figure 1 A schematic diagram of the membrane structure for preparing zeolite adsorption membranes or agar diffusion membranes;

[0029] Figure 2 A schematic diagram showing the structure of the zeolite adsorption membrane / oxygen fluorescence sensing membrane combined sensor and the placement of the sensor in the root box;

[0030] Figure 3 A schematic diagram of in-situ rhizosphere ammonia nitrogen / oxygen testing in root chambers and experimental water tanks using a zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor.

[0031] Figure 4 A schematic diagram of the analysis of a zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor in a box-type planar optical electrode testing equipment in a root chamber.

[0032] Figure 5 This is a schematic diagram of a circular DGT.

[0033] In the diagram: 1 is a hot agar solution or a hot agar solution mixed with zeolite; 2 is a mold; 3 is two strips of flat tempered glass (top view, dimensions: length × width × thickness = 10cm × 5cm × 0.5cm); 4 is a U-shaped plastic pad (thickness: 0.04cm or 0.01cm); 5 is the two short strips on the left and right sides of the U-shaped plastic pad (length: 5cm); 6 is the long strip below the U-shaped plastic pad (length: 10cm); 7 is three plastic clips; 8 is the narrow slit in the middle of the mold (thickness × length × width = 0.01cm or 0.04cm × 8cm × 4cm); 9 is a glass dropper with a pointed tip; 10 is a zeolite adsorption membrane; 11 is a circular DGT; 12 is an agar diffusion membrane; 13 is a zeolite... 14 is an adsorption membrane / oxygen fluorescence membrane sensor; 15 is a PVDF filter membrane; 16 is a round cap; 17 is a circular window in the center of the round cap; 18 is an oxygen fluorescence sensing membrane (length × width × thickness: 5μm × 2μm × 12μm); 19 is a root box; 20 is a removable transparent wall; 21 is a plastic screw; 22 is sediment; 23 is aquatic plants; 24 is top water; 25 is an experimental water tank; 26 is roots; 27 is stems and leaves; 28 is an inlet pipe; 29 is an outlet pipe; 30 is a fluorescent lamp; 31 is an aeration head; 32 is the liquid surface; 33 is a box-type planar photoelectric testing equipment; 34 is a dark box; 35 is a CMOS camera and lens; 36 is a filter; 37 is an LED light source; 38 is a computer control system.

[0034] Figure 6 The elution coefficients of the zeolite adsorption membrane for eluents with different NaCl concentrations are shown.

[0035] Figure 7 The mass of ammonia nitrogen accumulated on the zeolite adsorption membrane during different operating times (0–24 h);

[0036] Figure 8 The mass of ammonia nitrogen accumulated on the zeolite adsorption membrane during different operating times (0–80 h);

[0037] Figure 9 This is a two-dimensional image showing the diffusion flux of ammonia nitrogen and the oxygen concentration in a rhizosphere microregion of an aquatic plant. Detailed Implementation

[0038] The present invention provides a zeolite adsorption membrane comprising an agar hydrogel and zeolite particles dispersed in the agar hydrogel; wherein the particle size of the zeolite particles is ≤1μm.

[0039] In this invention, unless otherwise specified, all raw materials / components or test materials used in the preparation, such as zeolite particles, agar reagents or oxygen fluorescence sensing membranes, are commercially available products well known to those skilled in the art.

[0040] The zeolite adsorption membrane provided by this invention comprises agar hydrogel. In this invention, the agar hydrogel serves as the matrix material of the zeolite adsorption membrane. The preferred mass ratio of agar to water in the agar hydrogel is 1 g:(42-50) mL, more preferably 1 g:50 mL.

[0041] The zeolite adsorption membrane provided by this invention comprises zeolite particles dispersed in the agar hydrogel; the particle size of the zeolite particles is ≤1 μm. In this invention, the particle size of the zeolite particles is preferably 1 μm. The mass ratio of agar to zeolite particles in the agar hydrogel is preferably 1:(7-8), and more preferably 1:7.

[0042] In this invention, the thickness of the zeolite adsorption membrane is preferably 0.01 to 0.04 cm, and more preferably 0.01 cm or 0.04 cm.

[0043] This invention provides a method for preparing the zeolite adsorption membrane described above, comprising the following steps:

[0044] Zeolite granules, agar, and pure water are heated and mixed to obtain a hot mixed solution;

[0045] The hot mixed solution is poured into a mold and cooled. After demolding, the zeolite adsorption membrane is obtained.

[0046] This invention involves heating and mixing zeolite powder, agar, and pure water to obtain a hot mixed solution. Preferably, the heating and mixing process includes: first heating the water and then mixing it with agar to obtain a premixed solution; second heating the premixed solution to boiling to obtain a clear agar solution; and then mixing the zeolite powder and the clear agar solution while still hot. The preferred temperature for the first heating is 70–80°C.

[0047] After obtaining the hot mixed solution, the present invention injects the hot mixed solution into a mold for cooling, and after demolding, obtains the zeolite adsorption membrane. In the present invention, the structural schematic diagram of the mold is shown below. Figure 1As shown. The mold includes: two strips of tempered glass, each strip preferably measuring 10cm × 5cm × 0.5cm; a U-shaped plastic pad, preferably 0.04cm or 0.01cm thick, consisting of two short strips and a long strip connecting them, preferably 1cm wide, 5cm long, and 10cm long; and three plastic clips. In this invention, when using the mold, the two strips of tempered glass are placed face-to-face, and the U-shaped plastic pad is placed between them; the three plastic clips secure the two strips of tempered glass and the U-shaped plastic pad; a narrow slit with a thickness of 0.01cm or 0.04cm, a length of 8cm, and a width of 4cm is formed in the middle of the mold. In this invention, before pouring, the mold is preferably preheated to 80°C in an oven. The cooling process preferably involves placing the mold filled with the hot mixed solution in a room temperature environment for cooling. The cooling time is preferably 1 hour.

[0048] In this invention, after demolding, an initial gel membrane is obtained. Preferably, the invention further includes: immersing and washing the initial gel membrane four times with ultrapure water to obtain the zeolite adsorption membrane. The preferred immersion and washing time with ultrapure water is 2 hours each time. This invention preferably uses ultrapure water for immersion and washing to remove impurities and any possible color from the initial gel membrane. This invention preferably places the zeolite adsorption membrane in ultrapure water for long-term storage.

[0049] This invention provides the application of the zeolite adsorption membrane described in the above technical solution or the zeolite adsorption membrane prepared by the preparation method described in the above technical solution in the detection of ammonia nitrogen in the rhizosphere of aquatic plants.

[0050] In this invention, the preferred application is to immerse the zeolite adsorption membrane in water to detect ammonia nitrogen in the rhizosphere of aquatic plants. The pH value of the water is preferably 3.5–9, more preferably 3.5–8; the concentration of cations in the water is preferably <0.1 mol / L, and the cations include Na+. + K + Ca 2+ and Mg 2+ One or more of them.

[0051] The present invention provides a combined sensor, comprising a filter membrane, a zeolite adsorption membrane and an oxygen fluorescence sensing membrane stacked together; wherein the zeolite adsorption membrane is the zeolite adsorption membrane described in the above technical solution or the zeolite adsorption membrane prepared by the preparation method described in the above technical solution.

[0052] In this invention, the filter membrane is a polyvinylidene fluoride (PVDF) filter membrane. The thickness of the PVDF filter membrane is preferably 0.01 cm. The thickness of the oxygen fluorescence sensing membrane is preferably 12 μm. In this invention, both the PVDF filter membrane and the oxygen fluorescence sensing membrane can be commercially available products. Specifically: the oxygen fluorescence sensing membrane has a thickness of 12 μm; platinum(II)MESO-tetra(pentafluorophenyl)porphyrin (PtTFPP) is used as an oxygen indicator and is fixed on a permeable polymer matrix; the analyte interacts with the fluorescence indicator and binds, generating a fluorescence signal under the excitation of an external light source. There is a correlation between the analyte concentration and the fluorescence signal intensity; quantification can be performed by capturing the fluorescence signal using a specific camera. In a specific embodiment of this invention, the coupled sensor is a zeolite adsorption membrane / nitrogen fluorescence membrane coupled sensor (referred to as a zeolite adsorption membrane / oxygen fluorescence membrane sensor). In a specific embodiment of this invention, the length × width × thickness of the PVDF filter membrane is preferably 5 cm × 2 cm × 0.01 cm. The preferred dimensions of the zeolite adsorption membrane are 5cm × 2cm × 0.01cm (length × width × thickness). The preferred dimensions of the oxygen fluorescence sensing membrane are 5cm × 2cm × 12μm (length × width × thickness).

[0053] This invention provides a method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants, comprising the following steps:

[0054] (1) Fill a root box with sediment, aquatic plants and overlying water; the root box is provided with a removable transparent sidewall; the stems and leaves of the aquatic plants are located in the overlying water and the roots are located in the sediment; then place it in an experimental water tank for cultivation; after the aquatic plants are successfully cultivated, a sensor is set on the inner surface of the removable transparent sidewall of the root box near the roots of the aquatic plants. The sensor is the combined sensor described in the above technical solution, and the oxygen fluorescence sensing membrane of the combined sensor is in contact with the removable transparent sidewall of the root box.

[0055] (2) The root box equipped with the sensor was tested in the experimental water tank;

[0056] (3) Perform planar photopolar analysis on the oxygen fluorescence sensing membrane in the coupled sensor obtained after the test to obtain a two-dimensional distribution image of the oxygen concentration in the rhizosphere of aquatic plants. The spatial resolution of the two-dimensional distribution image of the oxygen concentration in the rhizosphere of aquatic plants is 62μm×62μm.

[0057] (4) The zeolite adsorption membrane in the coupled sensor after planar optical detection is analyzed to obtain a high spatial resolution two-dimensional distribution image of ammonia nitrogen diffusion flux in the rhizosphere of aquatic plants.

[0058] This invention involves filling a root box with sediment, aquatic plants, and overlying water. The root box has a removable transparent sidewall. The stems and leaves of the aquatic plants are located in the overlying water, while their roots are located in the sediment. The root box is then placed in an experimental water tank for cultivation. After successful cultivation, a sensor is placed on the inner surface of the removable transparent sidewall of the root box, near the roots of the aquatic plants. This sensor is a combined sensor as described in the previous technical solution, with the oxygen fluorescence sensing membrane of the combined sensor in contact with the removable transparent sidewall of the root box. The structure of the zeolite adsorption membrane / oxygen fluorescence sensing membrane combined sensor and its placement in the root box are shown in the diagram below. Figure 2 As shown. The experimental water tank ( Figure 3 Preferably, the experimental water tank is equipped with inlet and outlet pipes to achieve water reflux. During cultivation, a fluorescent lamp is installed above the experimental water tank to simulate 24-hour ambient light. At least two aeration heads are installed below the liquid surface in the experimental water tank; these aeration heads introduce air, oxygen, or argon into the water to control oxidation-reduction conditions. In the cultivation chamber, the liquid level in the experimental water tank is at least 20 cm above the top of the root box. Successful cultivation of the aquatic plant is defined as vigorous rhizosphere growth. A preferred embodiment of the present invention for setting a sensor on the inner surface of the detachable transparent sidewall of the root box near the roots of the aquatic plant is as follows: The root box is removed from the experimental water tank; the sidewall of the root box is opened, and a rectangular area of ​​the detachable transparent sidewall corresponding to a section of vigorously growing roots is selected. The length × width of the rectangular area is preferably 5cm × 2cm, and the length × width of the rectangular area is the same as the length × width of the coupled sensor. The rectangular area is marked on the detachable transparent sidewall with a marker. An oxygen fluorescence sensing membrane, a zeolite adsorption membrane, and a PVDF filter membrane are sequentially attached to the marked area on the detachable transparent sidewall. The detachable transparent sidewall is then reinstalled on the root box, ensuring that the coupled sensor is in close contact with the root section, with the root section located in the center of the coupled sensor.

[0059] This invention involves testing a root box equipped with sensors in an experimental water tank. The testing time is preferably 4–8 hours. The conditions of the experimental water tank during the testing are the same as those during the cultivation process.

[0060] This invention performs planar photopolarimetry analysis on the oxygen fluorescence sensing membrane in the coupled sensor obtained after testing to obtain a two-dimensional distribution image of the rhizosphere oxygen concentration of aquatic plants. The spatial resolution of the two-dimensional distribution image of the rhizosphere oxygen concentration of aquatic plants is 62μm×62μm. The planar photopolarimetry analysis preferably employs a box-type planar photopolarimetry testing equipment (…). Figure 4The test is conducted using a box-type planar photoluminescence testing equipment, preferably PO2100, provided by Zhongke Zhigan Environmental Technology Co., Ltd. The preferred implementation method for planar photoluminescence analysis is as follows: After removing the root chamber from the experimental water tank, immediately place the root chamber into the dark chamber of the box-type planar photoluminescence testing equipment. The detachable transparent sidewall of the root chamber, housing the coupled sensor, is aligned with a CMOS camera and lens. A filter is installed on the camera lens, preferably with a wavelength of 650nm. An LED light source is preferably positioned above the CMOS camera and lens, preferably with a wavelength of 390-400nm. A computer control system is used to set the test parameters, turn on the CMOS camera and LED light source, and test the fluorescence signal of the coupled sensor to obtain a rhizosphere fluorescence image. The spatial resolution of the fluorescence image is preferably 62μm×62μm. Simultaneously, two-point calibration is used to test the fluorescence signals of 0% saturated anhydrous sodium sulfite solution and 100% air / aerated aqueous solution, and an oxygen concentration standard curve is plotted. The rhizosphere fluorescence value is used to calculate the two-dimensional distribution value of rhizosphere oxygen concentration using the standard curve, and a computer drawing program is used to plot the two-dimensional distribution image of rhizosphere oxygen concentration.

[0061] This invention analyzes the adsorption of ammonia nitrogen by a zeolite adsorption membrane in a coupled sensor after planar photoelectric detection, obtaining a high spatial resolution two-dimensional distribution image of ammonia nitrogen diffusion flux in the rhizosphere of aquatic plants. In this invention, the analysis of adsorbed ammonia nitrogen preferably includes the following steps: spatially resolving the zeolite adsorption membrane to 200 μm × 200 μm; immersing the resulting slices in sodium chloride eluent for ammonia nitrogen elution, obtaining an ammonia nitrogen eluent; determining the ammonia nitrogen concentration in the eluent using the sodium ammonia nitrogen colorimetric method, and calculating the ammonia nitrogen diffusion flux from the ammonia nitrogen concentration.

[0062] In this invention, the spatial resolution of the zeolite adsorption membrane is preferably achieved using a fully automated paraffin microtome. Theoretically, an automated paraffin microtome can achieve a resolution of 50 μm × 50 μm; however, in practice, too high a resolution would increase the number of analyses required. Therefore, the preferred spatial resolution of this invention is 200 μm × 200 μm.

[0063] In a specific embodiment of the present invention, the preferred method for analyzing the adsorbed ammonia nitrogen is as follows: disassemble the root box, remove the coupled sensor, remove the zeolite diffusion membrane, and perform high spatial resolution two-dimensional distribution analysis of the ammonia nitrogen adsorbed by the zeolite adsorption membrane. The preferred method for analyzing the ammonia nitrogen adsorbed by the zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor is as follows: use micro-cutting + elution + spectrophotometric analysis to determine the sub-millimeter two-dimensional distribution of the ammonia nitrogen adsorbed by the zeolite adsorption membrane, wherein the spatial resolution is preferably 200 μm × 200 μm. In this invention, the preferred method for analyzing the sub-millimeter two-dimensional distribution of ammonia nitrogen adsorbed by zeolite adsorption membrane using microcutting + elution + spectrophotometry is as follows: The water adhering to the wet zeolite adsorption membrane is absorbed with absorbent paper; an adsorption membrane measuring 2cm x 5cm is placed on a flat glass plate of the same volume and 0.5mm thickness, and both are placed into the sample box of a Thermo Fisher Scientific fully automated paraffin microtome. The slice thickness is adjusted to 100μm, and the spatial resolution is achieved through manual slicing using the "handwheel rotation" mode, preferably 200μm x 200μm; the glass plate is then removed, rotated 90°, and placed back into the sample box for recutting. Then, each slice was placed into a centrifuge tube with a 1 mL specification using a needle; the label on the centrifuge tube corresponds to each position on the adsorption membrane; 100 μL of NaCl elution buffer was added to the centrifuge tube, and the elution time was 24 h; then, the ammonia nitrogen concentration of the elution buffer was determined by the sodium colorimetric method (national standard method) of ammonia nitrogen at 420 nm using an enzyme-linked immunosorbent assay (ELISA) reader: the diffusion flux of ammonia nitrogen was calculated according to formula (2): F = M / At (2); in formula (2): F is the diffusion flux of ammonia nitrogen from the sensor surface to the zeolite adsorption membrane, and the unit is ng cm. -2 s -1 M is the mass of ammonia nitrogen adsorbed on the zeolite adsorption membrane, in ng; A is the area of ​​the adsorption membrane, in cm². 2 t represents the testing time of the adsorption membrane in the sample, measured in seconds. Based on the tests conducted using a zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor, a high spatial resolution two-dimensional distribution image of ammonia nitrogen diffusion flux in the root segments of aquatic plants was plotted.

[0064] This invention provides a method for preparing a zeolite adsorption membrane and a high spatial resolution two-dimensional analysis method for adsorbing ammonia nitrogen; at the same time, this invention provides a method for assembling a novel sensor that combines a zeolite adsorption membrane with an oxygen fluorescence membrane, achieving for the first time the simultaneous testing of sub-millimeter-level two-dimensional distribution of ammonia nitrogen and oxygen at the environmental interface.

[0065] The zeolite adsorption membrane provided by this invention comprises an agar hydrogel and zeolite particles dispersed in the agar hydrogel; the particle size of the zeolite particles is ≤1 μm. This invention avoids the interference of ammonia nitrogen analysis caused by using acrylamide as the membrane matrix material; simultaneously, based on the particle size of the zeolite particles used in this invention being ≤1 μm, the micro-cutting + micro-spectrophotometric analysis method of the zeolite adsorption membrane enables sub-millimeter-level high spatial resolution two-dimensional testing of ammonia nitrogen. By using the zeolite adsorption membrane provided by this invention in conjunction with an oxygen fluorescence sensing membrane, the simultaneous testing of sub-millimeter-level two-dimensional distribution of ammonia nitrogen and oxygen at the rhizosphere interface of aquatic plants has been achieved for the first time, obtaining sub-millimeter-level two-dimensional distribution images of both at the same location in the rhizosphere, thus providing an important foundation for revealing the dynamic migration-absorption and oxygen response mechanism of rhizosphere ammonia nitrogen.

[0066] To further illustrate the present invention, the technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0067] Example 1

[0068] This embodiment provides a novel method for preparing a zeolite adsorption membrane, comprising the following steps: placing 50 mL of water in a beaker and heating it to 80°C in a glove box in a sterile room; slowly pouring 1.0 g of agar powder into the water in the beaker, stirring with a glass rod to evenly disperse it in the water until it is basically dissolved; then heating the beaker to boiling; continuing until the agar solution becomes clear; then slowly pouring 7.0 g of zeolite powder into the hot agar solution, stirring with a glass rod to evenly distribute the zeolite in the hot agar solution. The mold, heated to 80°C in an oven, is then used to pour in the agar solution 1 containing the zeolite. A schematic diagram of the structure of the mold 2 used in this embodiment is shown below. Figure 1As shown, the mold specifically includes: two flat strips of tempered glass 3, each strip having a length × width × thickness of 10cm × 5cm × 0.5cm; a U-shaped plastic pad 4, the thickness of which is 0.04cm or 0.01cm; the width of each side of the U-shaped plastic pad 4 is 1cm; the length of the two short sides 5 of the U-shaped plastic pad 4 is 5cm, and the length of the bottom long side 6 is 10cm; three plastic clips 7; the two strips of tempered glass 3 are placed face to face, with the U-shaped plastic pad 4 placed between them; the three plastic clips secure the three sides of the mold; and a narrow slit 8 in the center of the mold, with a thickness of 0.01cm or 0.04cm, a length of 8cm, and a width of 4cm. Immediately use a glass dropper 9 to drip hot agar solution 1 mixed with zeolite into the narrow slit of the mold until the slit is filled with the agar solution 1 mixed with zeolite. Cool the mold at room temperature for 1 hour; open the two strip-shaped tempered glass 3 of mold 2, carefully remove the zeolite adsorption membrane 10. The zeolite adsorption membrane 10 has dimensions of length × width × thickness = 8cm × 4cm × 0.04cm or 8cm × 4cm × 0.01cm. Place it in 1L of ultrapure water, changing the water 4 times, once every 2 hours, to remove impurities and any possible color from the zeolite adsorption membrane 10; the zeolite adsorption membrane 10 should appear uniformly white. Store the zeolite adsorption membrane 10 in ultrapure water. The 0.04cm thick zeolite adsorption membrane 10 is used to install the circular DGT11 and for ammonia nitrogen adsorption function testing; the 0.01cm thick zeolite adsorption membrane 10 is used to assemble a zeolite adsorption membrane / oxygen fluorescence membrane coupled sensor.

[0069] Preparation of agar diffusion membrane: Place 50 mL of water in a beaker and heat to 80 °C in a glove box in a sterile room; slowly pour 0.75 g of agar powder into the water in the beaker and stir with a glass rod to evenly disperse it in the water until it is basically dissolved; heat the beaker to boiling; continue heating until the agar solution becomes clear; make the mold with the following narrow slit dimensions: length × width × thickness = 8 cm × 4 cm × 0.04 cm; pour the hot agar solution into the glass mold and cool the mold at room temperature for 1 hour; open the two strips of tempered glass 3 of mold 2 and carefully remove the agar diffusion membrane 12, which has dimensions of length × width × thickness = 8 cm × 4 cm × 0.04 cm; place the agar diffusion membrane 12 in 1 L of ultrapure water for 24 hours, changing the water every 2 hours to remove impurities and any possible color from the agar diffusion membrane 12; the agar diffusion membrane 12 is transparent; store the agar diffusion membrane 12 in a 0.001 mol / L NaCl solution for long-term storage. Agar diffusion membrane 12 is used for circular DGT devices ( Figure 5The zeolite adsorption membrane 10 was also tested for functionality. The zeolite adsorption membrane 10 in the zeolite adsorption membrane / oxygen fluorescence membrane sensor 13 described below was made using the same method as described above, except that the thickness of the plastic pad 4 in the mold 2 of the zeolite adsorption membrane 10 used in the zeolite adsorption membrane / oxygen fluorescence membrane sensor 13 is 0.01 cm.

[0070] Test Example 1

[0071] The functional testing methods for zeolite adsorption membranes include: methods for determining the elution coefficient of zeolite adsorption membrane 10; kinetic testing of ammonia nitrogen absorption by zeolite adsorption membrane 10 and calculation methods for the diffusion coefficient of ammonia nitrogen in diffusion membranes; methods for testing the ammonia nitrogen adsorption capacity of adsorption membranes; and the effects of pH and cations on the ammonia nitrogen absorption of DGT by zeolite adsorption membranes. Among these:

[0072] Determination of the elution coefficient of zeolite adsorption membrane: Zeolite adsorption membrane 10 was cut into circular adsorption membranes with a diameter of 2.5 cm; three 10 mL aliquots of 2.5 mol / L precipitate were transferred. -1 NH4-N solution (pH = 7.0) was poured into three 25 mL centrifuge tubes; three circular zeolite adsorption membranes were placed in the three centrifuge tubes, and after shaking at room temperature for 24 h, the zeolite adsorption membranes were removed and treated with 0.25, 0.50, 1.0, 1.5 and 2.0 mol L⁻¹ solutions, respectively. -1 Elution with NaCl reaction solution (5 mL) for 24 h, followed by the collection of NH4-N solution and the eluent from the adsorption membrane. The ammonia nitrogen concentration in the eluent was determined using Nessler's colorimetric method. The actual mass of ammonia nitrogen accumulated on the zeolite adsorption membrane is the mass of ammonia nitrogen absorbed from the reaction solution. The elution coefficient is the ratio of the mass of ammonia nitrogen eluted from the zeolite adsorption membrane to the mass of ammonia nitrogen absorbed from the reaction solution by the adsorption membrane. The elution efficiency of the zeolite adsorption membrane with NaCl reaction solution is 100% (e.g., Figure 6 (as shown), Figure 6 The elution coefficients of the zeolite adsorption membrane are given by eluents with different NaCl concentrations.

[0073] The absorption kinetics of ammonia nitrogen fixed by zeolite adsorption membranes: The absorption kinetics of ammonia nitrogen over time by circular DGT 11 zeolite adsorption membranes can be achieved by testing five sets of circular zeolite DGTs in an ammonia nitrogen reaction solution. Circular DGTs ( Figure 5 The outer shell is made of plastic. From bottom to top, the DGT consists of a base 14, a zeolite adsorption membrane 10, an agar diffusion membrane 12, a polyvinylidene fluoride (PVDF) filter membrane 15, and a round cap 16 with a circular window 17 in the center. Ammonia nitrogen reaction solution: 3.0 mg / L - 1 NH4-N (0.002 mol L) -1NaCl; pH = 7.0 ± 0.05; Three circular DGT 11 units were placed in 7 L of reaction solution. At reaction times of 4 h, 8 h, 12 h, 16 h, and 24 h, the circular DGT 11 units were removed for elution and ammonia nitrogen analysis. Simultaneously, the ammonia nitrogen concentration in the water sample was analyzed. The RSD of the ammonia nitrogen absorption mass of each set of circular DGT 11 units was <9%; the curve showing the change in ammonia nitrogen absorption mass of the circular DGT 11 units over time is shown below. Figure 7 As shown, Figure 7 The mass of ammonia nitrogen accumulated on the zeolite adsorption membrane during different operating times (0–24 h) is the kinetic curve of ammonia nitrogen absorption by DGT, 3.0 mg / L. -1 NH4-N reaction solution; 0.002 mol / L -1 NaCl; pH = 7.0 ± 0.05, the correlation is R 2 =0.994; According to formula (1): D=aΔg / CA(1), the diffusion coefficient of ammonia nitrogen in the agar diffusion membrane can be calculated as: 1.44×10 -5 cm 2 s -1 (14℃). In formula (1): a is the slope of the linear regression equation of the mass of ammonia nitrogen absorbed by the zeolite adsorption membrane (ng) with respect to the operating time (s); Δg is the sum of the thicknesses of the diffusion membrane and the filter membrane (cm); C is the concentration of the working solution (ng / mL); A is the area of ​​the circular DGT window (cm²). 2 ).

[0074] Zeolite adsorption membrane ammonia nitrogen adsorption capacity test: Prepare 10 sets of circular DGT 11 membranes placed in a 22 mg / L solution. -1 NH4-N (0.002 mol L) -1 NaCl; pH = 7.0 ± 0.05); times for removing circular DGT: 4h, 8h, 16h, 24h, 32h, 40h, 48h, 56h, 68h, 80h; changes in the mass of ammonia nitrogen accumulated in the zeolite adsorption membrane during the operating time (0–80h) are as follows: Figure 8 As shown, Figure 8 The mass of ammonia nitrogen accumulated on the zeolite adsorption membrane during different operating times (0–80 h), i.e., the capacity curve of ammonia nitrogen absorbed by DGT, is 22.0 mg / L. -1 NH4-N reaction solution; 0.002 mol / L -1 NaCl; pH = 7.0 ± 0.05. After 56 h, the mass of ammonia nitrogen adsorbed by the zeolite adsorption membrane changed very little and remained stable at 842 ± 51 μg; indicating that the DGT installed on the zeolite adsorption membrane can complete the long-term test of ammonia nitrogen in the rhizosphere of aquatic plants.

[0075] Effect of pH on ammonia nitrogen absorption of DGT by zeolite adsorption membrane: Circular zeolite DGT ( Figure 5The test was conducted over 24 hours in ammonia nitrogen reaction solutions at a range of pH values.

[0076] Ammonia nitrogen reaction solution: 2.0 mg / L -1 NH4-N. Ammonia nitrogen DGT concentration as measured by DGT (C DGT ) and the effect of collecting liquid ammonia nitrogen concentration (C solu The ratio range is 0.90–1.10, which serves as the standard for whether it significantly affects the DGT test. Table 1 shows the C ratios of DGT from round zeolite in ammonia nitrogen standard solutions at different pH values ​​(3.5–9.0). DGT / C solu Value. The results show that the C value in the pH range of 3.5–9.0 is... DGT / C solu =0.71~0.99 (as shown in Table 1); zeolite adsorption membrane DGT in the pH range (3.5~8.0) C DGT / C solu =0.90~0.99; Zeolite adsorption membrane DGT can be used for accurate testing of ammonia nitrogen under most water pH conditions.

[0077] Table 1. C content of round zeolite DGT in ammonia nitrogen standard solutions at different pH values ​​(3.5–9.0) DGT / C solu value

[0078] pH <![CDATA[C DGT / C solu ]]> 3.51±0.04 0.90 5.02±0.03 0.94 6.02±0.02 0.99 7.14±0.07 0.95 8.01±0.05 0.92 8.54±0.08 0.85 9.01±0.08 0.71

[0079] The effect of cations on ammonia nitrogen absorption by zeolite adsorption membrane DGT: The concentration of cations in circular zeolite DGT was adjusted to a certain level (Na... + K + Ca 2+ and Mg 2+ Ammonia nitrogen reaction solution (2.0 mg / L) -1 The test was conducted in NH4-N for 24 hours.

[0080] Ammonia nitrogen concentration in liquid solution: 2.0 mg / L -1 NH4-N; containing Na + K + (0.0001~0.1mol / L), Ca 2+ or Mg 2+ The pH of the ammonia nitrogen reaction solution (0.0001~0.001mol / L) was 7.0±0.21; Ca 2+ or Mg 2+ The pH of the ammonia nitrogen reaction solution (0.01–0.1 mol / L) was 5.9 ± 0.12. The CL values ​​of the circular zeolite DGT in the ammonia nitrogen reaction solutions with different ion concentrations were measured. DGT / C soluSee Table 2. Ammonia nitrogen concentration in the reaction solution: 0.0001–0.001 mol / L -1 Na + K + Ca 2+ or Mg 2+ Measured C DGT / C solu Approaching 1; as the concentration of the above ions continues to increase, C DGT / C solu Decrease; ion concentration reaches 0.01 mol / L -1 Only Na + C DGT / C solu =0.90, meeting the test requirements; when the ion concentration is ≥0.1 mol L -1 Zeolite-DGT cannot accurately test ammonia nitrogen (Table 2). This indicates that zeolite-DGT cannot accurately test ammonia nitrogen in saline-alkali water bodies. For most water bodies with low ion concentrations, zeolite-DGT can accurately test ammonia nitrogen.

[0081] Table 2. Effect of cation concentration in the ammonia nitrogen treatment solution on the absorption of ammonia nitrogen by the zeolite adsorption membrane DGT.

[0082]

[0083] Example 2

[0084] Installation and testing method of zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13 in root box-aquatic plant rhizosphere: Prepare a zeolite adsorption membrane 10 (length × width × thickness = 5cm × 2cm × 0.01cm), a polyvinylidene fluoride (PVDF) filter membrane 15 (length × width × thickness = 5cm × 2cm × 0.01cm), and an oxygen fluorescence sensing membrane 18 (length × width × thickness = 5μm × 2μm × 12μm); construct a root box 19 (length × width × height = 14cm × 10cm × 20cm; wall thickness: 0.5cm) using PVC material. One wall is made of transparent PVC material (length × height = 14cm × 20cm, wall thickness: 0.5cm). This wall is a detachable transparent wall 20, which is fixed to the other three walls with plastic screws 21 to form a hollow cuboid; fill the root box 19 with sediment 22 and aquatic plants. (The aquatic plant used in this embodiment is Vallisneria natans) 23 and top water 24; the root box 19 was placed in the experimental water tank 25 for cultivation beforehand. After the root growth was vigorous and the cultivation was successful, the root box 19 was taken out; the detachable wall 20 of the root box 19 was opened, and a rectangular area (5cm×2cm) corresponding to a section of the wall with vigorous root 26 was selected. It was exactly the same size as the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13, and a graphic mark was made with a marker pen; the oxygen fluorescence sensing membrane 18, zeolite adsorption membrane 10 and PVDF filter membrane 16 were attached to the marked area on the detachable transparent wall 20 in sequence, and then the detachable transparent wall 20 was reinstalled in the root box 19. During the installation process, the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13 was made to be in close contact with the root section 26, and the root section was located in the center of the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13; the stem and leaves 27 were located in the top water 24. The root box 19 was placed in the experimental water tank 25 for testing for 4 hours. The experimental water tank 25 includes an inlet pipe 28 and an outlet pipe 29 to achieve water reflux within the tank. A fluorescent lamp 30 is located directly above the tank, simulating 24-hour ambient light. Two aeration heads 31 are also installed in the tank to introduce air, oxygen, or argon into the water, controlling oxidation-reduction conditions. The liquid level 32 in the tank exceeds the top of the root chamber 19 by at least 20 cm. After testing the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13 in the root chamber 19 for 4 hours in the tank, the root chamber 19 is removed.

[0085] Analysis of the oxygen fluorescence membrane: After the root box 19 of the experimental water tank 25 was removed, it was immediately placed into the dark box 34 of the box-type planar photoelectric testing equipment 33 (PO2100, Zhongke Zhigan Environmental Technology Co., Ltd.). The detachable transparent wall 20 of the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13 installed in the root box 19 was aligned with the CMOS camera and lens 35. A filter (650nm) 36 was installed on the camera lens 35. The LED light source (390~400nm) 37 was above the CMOS camera and lens 35. The test parameters were set using the computer control system 38, and the CMOS camera 35 and the LED light source (390~400nm) 37 were turned on. The fluorescence signal of the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13 was tested to obtain the fluorescence map of the rhizosphere (spatial resolution: 62μm×62μm). At the same time, the fluorescence signals of 0%-saturated anhydrous sodium sulfite solution and 100%-air / aerated aqueous solution were tested using two-point calibration, and a standard curve was plotted. Rhizosphere fluorescence values ​​were used to calculate the two-dimensional distribution of rhizosphere oxygen using a standard curve, and a computer-generated image of the rhizosphere oxygen distribution was plotted. The root box 19 was disassembled, the sensor removed, and the zeolite diffusion membrane 10 was taken out for high spatial resolution two-dimensional distribution analysis of ammonia nitrogen adsorbed by the zeolite adsorption membrane. The two-dimensional images of oxygen concentration and ammonia nitrogen diffusion flux in the rhizosphere of *Vallisneria natans* section 1, measured by the zeolite adsorption membrane / nitrogen fluorescence membrane sensor 13, are shown below. Figure 9 As shown, Figure 9 Two-dimensional images of oxygen concentration and ammonia nitrogen diffusion flux in the rhizosphere of a section of Vallisneria natans.

[0086] The analytical method for ammonia nitrogen adsorbed by the zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor 13 is as follows: The sub-millimeter two-dimensional distribution (spatial resolution: 200 μm × 200 μm) of ammonia nitrogen adsorbed by the zeolite adsorption membrane 10 is analyzed using micro-cutting + elution + spectrophotometry. The water adhering to the wet zeolite adsorption membrane is blotted dry with absorbent paper. A 2 cm × 5 cm section of the adsorption membrane is placed on a flat glass plate of the same volume and a thickness of 0.5 mm. These sections are then placed in the sample cassette of a Thermo Fisher Scientific fully automated paraffin microtome. The section thickness is adjusted to 100 μm, and manual sectioning (200 μm × 200 μm) is performed using the "handwheel rotation" mode. The glass plate is then removed, rotated 90°, and placed back into the sample cassette for re-cutting. Then, each slice was placed into a 1 mL centrifuge tube using a needle; the label on the centrifuge tube corresponds to each position on the adsorption membrane; (2) 100 μL of NaCl elution buffer (1.0 mol / L) was added to the centrifuge tube, and the elution time was 24 h; then, the concentration of ammonia nitrogen in the elution buffer was determined by the sodium colorimetric method (national standard method) of ammonia nitrogen at 420 nm using an enzyme-linked immunosorbent assay (ELISA) reader; (3) The diffusion flux of ammonia nitrogen was calculated according to formula (2): F = M / At (2); In formula (2): F is the diffusion flux of ammonia nitrogen from the sensor surface to the zeolite adsorption membrane (ng / cm). -2 s-1 M is the mass of ammonia nitrogen adsorbed on the zeolite adsorption membrane (ng); A is the area of ​​the adsorption membrane (cm²). 2 ); t is the test time (s) of the adsorption membrane in the sample.

[0087] Based on the tests conducted using the zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor 13, a high spatial resolution two-dimensional distribution image of ammonia nitrogen diffusion flux and oxygen concentration in the root segment of aquatic plants was generated. Figure 9 The dimensions of this root section of Vallisneria natans are: height × width = 12.90 mm × 7.44 mm. Theoretically, the resolution for oxygen concentration is 62 μm × 62 μm; the resolution for ammonia nitrogen diffusion flux is 200 μm × 200 μm.

[0088] As shown in the above embodiments, this invention provides a method for preparing a zeolite adsorption membrane for high spatial resolution testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants, as well as an assembly and testing method for a zeolite adsorption membrane / oxygen fluorescence membrane coupled sensor. The method is used to prepare a zeolite adsorption membrane capable of accurately measuring the diffusion flux of ammonia nitrogen in the rhizosphere of aquatic plants, to perform high spatial resolution two-dimensional analysis of ammonia nitrogen adsorbed by the membrane, and to assemble a zeolite adsorption membrane / oxygen fluorescence sensing membrane coupled sensor for high spatial resolution two-dimensional testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants. The obtained two-dimensional distribution images of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants are an important foundation for revealing the dynamic migration-absorption and oxygen response mechanisms of ammonia nitrogen in the rhizosphere.

[0089] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. Other embodiments can be obtained based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants, characterized in that, Includes the following steps: (1) Fill the root box with sediment, aquatic plants and top water; the root box is provided with a detachable transparent sidewall; the stems and leaves of the aquatic plants are in the top water and the roots are in the sediment; then place it in an experimental water tank for cultivation; after the aquatic plants are successfully cultivated, a sensor is set on the inner surface of the detachable transparent sidewall of the root box and in the area close to the roots of the aquatic plants. The sensor is a combined sensor, which includes a filter membrane, a zeolite adsorption membrane and an oxygen fluorescence sensing membrane stacked together. The zeolite adsorption membrane includes agar hydrogel and zeolite particles dispersed in the agar hydrogel. The particle size of the zeolite particles is 1 μm. The oxygen fluorescence sensing membrane of the combined sensor is in contact with the detachable transparent sidewall of the root box. (2) The root box equipped with the sensor was tested in the experimental water tank; (3) Perform planar photopolar analysis on the oxygen fluorescence sensing membrane in the coupled sensor obtained after the test to obtain a two-dimensional distribution image of the oxygen concentration in the rhizosphere of aquatic plants. The spatial resolution of the two-dimensional distribution image of the oxygen concentration in the rhizosphere of aquatic plants is 62 µm × 62 µm. (4) The zeolite adsorption membrane in the coupled sensor after planar optical detection is analyzed to obtain a high spatial resolution two-dimensional distribution image of ammonia nitrogen diffusion flux in the rhizosphere of aquatic plants; the analysis of adsorbed ammonia nitrogen includes the following steps: The zeolite adsorption membrane is cut with spatial resolution of 200 µm × 200 µm. The zeolite adsorption membrane slices obtained after cutting were immersed in sodium chloride eluent for ammonia nitrogen elution. The spatial resolution of the zeolite adsorption membrane slices was 200 µm × 200 µm, and the ammonia nitrogen eluent was obtained. The ammonia nitrogen concentration of the eluent was determined using the Sodium colorimetric method and a micro spectrophotometer, and the diffusion flux of ammonia nitrogen in the rhizosphere sediment was calculated from the ammonia nitrogen concentration.

2. The method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants according to claim 1, characterized in that, The mass ratio of agar to zeolite particles in the agar hydrogel is 1:(7~8).

3. The method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants according to claim 1 or 2, characterized in that, The mass ratio of agar to water in the agar hydrogel is 1 g: (42~50) mL.

4. The method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants according to claim 1, characterized in that, The thickness of the zeolite adsorption membrane is 0.01~0.04 cm.

5. The method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants according to claim 1, characterized in that, The method for preparing the zeolite adsorption membrane includes the following steps: Zeolite granules, agar, and pure water are heated and mixed to obtain a hot mixed solution; The hot mixed solution is poured into a mold and cooled. After demolding, the zeolite adsorption membrane is obtained.

6. The method for simultaneous testing of ammonia nitrogen and oxygen in the rhizosphere of aquatic plants according to claim 5, characterized in that, The process of obtaining the initial gel membrane after demolding also includes: washing the initial gel membrane four times with ultrapure water to obtain the zeolite adsorption membrane.

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