Capillary electrophoresis method for the detection of nitrate and total nitrogen in high conductivity water samples

Abstract

The present invention discloses a detection system for nitrate and total nitrogen in high conductivity water samples, comprising a pH control unit in a capillary and an ultraviolet LED light reduction unit. The system has a simple structure, a simple method, a fast detection speed, a wide range of applications, can greatly reduce the use of toxic and harmful chemical reagents, and the energy consumption required for light reduction is extremely low. The present invention also discloses a capillary electrophoresis method for detecting nitrate and total nitrogen in high conductivity water samples.

Description

Technical Field

[0001] The invention relates to a capillary detection technology, in particular to a capillary electrophoresis method for detecting nitrate and total nitrogen in a water sample with a high conductivity value. Background Art

[0002] In recent years, with the rapid development of industry and agriculture, the pollution of various nutrients related to nitrogen in the environment has received more and more attention. Nitrates are easily soluble in water and have high solubility, which allows them to quickly transfer and diffuse in water and soil, causing large-scale environmental pollution. It is reported that high concentrations of nitrates can cause eutrophication of water bodies, "algae blooms", and destroy the balance of aquatic ecosystems. It is one of the important factors causing water quality deterioration. At the same time, it also has many adverse effects on human health and production and life.

[0003] Currently, many methods for determining nitrate have been reported, such as spectrophotometry, ion selective electrode method, ion chromatography, gas phase molecular absorption method, etc. When using phenol disulfonic acid spectrophotometry, when there are ions such as chlorine and nitrite in the water sample, it interferes with the accurate determination of nitrate; ion chromatography can determine multiple inorganic anions at the same time, but the instrument used is precise and requires special personnel to use and maintain; gas phase molecular absorption has a fast analysis speed and is suitable for large-scale sample testing, but its analytical instrument is large in size and requires high cost.

[0004] At present, the most common method for detecting nitrate is to reduce nitrate to nitrite and then measure it by photometry. Common methods for reducing nitrate to nitrite include copper-plated cadmium columns, zinc-cadmium reduction methods, etc. These methods have high reduction efficiency, but the chemical reagents used in the reduction process are highly toxic, posing a huge threat to human health, and are very likely to cause more serious environmental pollution. Although the ultraviolet light reduction method is not as efficient as the above methods, it is simple to operate and avoids the use of toxic and harmful reagents. Most of the ultraviolet lamps commonly used for reduction in the current market have high power, high energy consumption, large size, and require a large amount of reducing agent. The reduction process takes a long time and the operation process is too cumbersome. To this end, exploring a more portable and efficient photoreduction system to make the photoreduction process less energy-consuming and more low-carbon and environmentally friendly has gradually become a new research hotspot. Summary of the invention

[0005] The purpose of the present invention is to provide a capillary electrophoresis method for detecting nitrate and total nitrogen in high conductivity water samples, in view of the use of toxic substances in existing photoreduction reagents, high energy consumption of ultraviolet light sources used during illumination, long time consumption, etc. The system has a simple structure, easy operation, fast detection speed, wide application range, can greatly reduce the use of toxic and harmful chemical reagents, and the energy consumption required for photoreduction is extremely low.

[0006] The technical solution for achieving the purpose of the present invention is:

[0007] A detection system for nitrate and total nitrogen in high conductivity water samples, comprising a pH control unit in a capillary and an ultraviolet LED light reduction unit.

[0008] The pH control unit is composed of three sections of different pH solutions, which are formed by capillary injection, wherein the three sections of different pH solutions include an electrophoresis buffer solution of pH=3.7, a barrier solution obtained by mixing a photoreduction reagent with a pH of 8.5 and sulfate ions, and a test solution of pH=8.5, wherein a photoreduction reagent, an internal standard substance and a nitrate are added to the test solution, and the internal standard substance is sodium methanesulfonate;

[0009] The ultraviolet LED light restoration unit includes a group of LED lamps with LED lamp fixing frames forming a light source, and a driving circuit connected to the LED lamps. The LED lamp fixing frame includes a lower base with a cavity and an upper lamp frame assembled in the cavity of the lower base. The upper lamp frame can move up and down on the lower base to adjust the height of the LED lamp fixing frame. The LED lamp is assembled on the upper lamp frame; the driving circuit includes a circuit board formed by welding chips, adjustable potentiometers, capacitors, and resistors in series and parallel. The input end of the circuit board is externally connected to a power adapter, and the output end of the circuit board is connected to the LED lamp.

[0010] The capillary is provided with a detector, one end of the capillary is inserted into a first centrifuge tube filled with an electrophoresis buffer solution, the first centrifuge tube is connected to a ground end, and the other end is inserted into a second centrifuge tube filled with an electrophoresis buffer solution after injection and illumination, the second centrifuge tube is connected to a high-voltage end, the end of the capillary inserted into the first centrifuge tube is a fixed end, and the end inserted into the second centrifuge tube is an injection end.

[0011] A capillary electrophoresis method for detecting nitrate and total nitrogen in a high conductivity water sample comprises the above-mentioned detection system for nitrate and total nitrogen in a high conductivity water sample, and the method comprises the following steps:

[0012] 1) The polyimide coating is burned off at the injection end of the capillary tube, and then the capillary tube is passed through the detector and fixed in the detector, and the fixed end of the capillary tube is inserted into a first centrifuge tube filled with an electrophoresis buffer solution and a connecting end;

[0013] 2) Place the centrifuge tube containing the electrophoresis buffer solution in the headspace bottle, insert the capillary injection end into the electrophoresis buffer solution in the centrifuge tube, and use nitrogen gas with a pressure of 0.15 MPa to push the electrophoresis buffer solution in the centrifuge tube into the capillary and rinse for several minutes;

[0014] 3) Manually injecting the barrier liquid and the test liquid placed on the injection station into the capillary tube in sequence, wherein the injection time of the barrier liquid is longer than the injection time of the test liquid, and the capillary tube after injection is gently placed on the capillary tube placement rack;

[0015] 4) placing an LED light source parallel and close to both sides of the capillary injection port after the polyimide coating is removed to illuminate the capillary for photoreduction reaction;

[0016] 5) The separation voltage of the electrophoresis system was adjusted to -15 kV, and the capillary injection end after the photoreduction reaction was completed was placed in a second centrifuge tube with a high-voltage end and filled with electrophoresis buffer solution. The high-voltage power supply and the data acquisition program TraceDec Monitor were manually triggered to realize the detection.

[0017] The height of the capillary placement rack is the same as the height of the detector to ensure that the capillary is in a horizontal state during illumination to prevent sample diffusion.

[0018] The detector is a non-contact conductivity detector.

[0019] The capillary is a fused silica capillary.

[0020] During operation, a complete electrophoresis circuit is formed between the high voltage terminal electrode and the ground terminal electrode.

[0021] The electrophoresis buffer solution is prepared by mixing glacial acetic acid and TRIS and dissolving the mixture to a volume of 100 mL.

[0022] Nitrate standard stock solution: Weigh potassium nitrate and dissolve it to make up to 100 mL.

[0023] Sodium methanesulfonate: Weigh sodium methanesulfonate and dissolve it to make up to 100 mL.

[0024] The photoreduction reagent is sodium hydrogen phosphate dodecahydrate dissolved in EDTA and fixed to 100 mL.

[0025] The barrier liquid is prepared by dissolving disodium hydrogen phosphate dodecahydrate, EDTA and sodium sulfate decahydrate to a volume of 100 mL, and adjusting the pH to 8.5 with 2M sodium hydroxide solution.

[0026] Compared with the prior art, the advantages of this technical solution are:

[0027] The ultraviolet LED lamp used in the system of this technical solution has a maximum output power of 4mW. Compared with the traditional ultraviolet LED lamp with an output power of up to tens of watts, this technical solution is more energy-saving and environmentally friendly. The method of this technical solution directly performs a photoreduction reaction on the capillary column head after the injection is completed. There is no need for too many cumbersome operating steps. The operation process is extremely simple, and the toxicity of the photoreduction reagent required by the system is extremely low. The entire detection system requires a small injection volume. The detection system proposed in this technical solution only requires an injection volume of about 2nL. The detection cost of the entire system is controlled within 10,000 yuan, which is much lower than the price of other detection instruments on the market (mostly hundreds of thousands of yuan). The electrophoresis separation channel is a capillary (inner diameter 25μm, outer diameter 365μm), the typical separation voltage is -15kV, and the system injection method is manual injection.

[0028] The system structure of this method is simple, easy to operate, fast in detection, and has a wide range of applications. It can greatly reduce the use of toxic and harmful chemical reagents, and the energy consumption required for photoreduction is extremely low. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 A schematic diagram of the system structure of an embodiment;

[0030] Figure 2 : is a distribution diagram of solutions with different pH values ​​at the capillary injection port in the embodiment;

[0031] Figure 3 Schematic diagram of the driving circuit structure in the embodiment;

[0032] Figure 4 This is the electrophoresis spectrum after adding high concentration of sulfate to the barrier liquid in the embodiment;

[0033] Figure 5 Schematic diagram of the working curve of nitrate in the embodiment.

[0034] Figure 1 The middle dotted box indicates the UV reduction process at the capillary injection port. DETAILED DESCRIPTION

[0035] The content of the present invention is further described below in conjunction with the drawings and embodiments, but the present invention is not limited thereto.

[0036] Example:

[0037] Reference Figure 1A detection system for nitrate and total nitrogen in high conductivity water samples, comprising a pH control unit and an ultraviolet LED light reduction unit in a capillary, wherein the pH control unit is composed of three sections of different pH solutions, and the three sections of different pH solutions are formed by capillary injection, wherein the three sections of different pH solutions include an electrophoresis buffer solution of pH=3.7, a barrier solution obtained by mixing a photoreduction reagent with pH=8.5 and sulfate ions, and a test solution of pH=8.5, such as Figure 2 As shown, a photoreduction reagent, an internal standard substance and nitrate are added to the test solution, and the internal standard substance is sodium methanesulfonate;

[0038] The ultraviolet LED light restoration unit includes a group of LED lamps with LED lamp fixing frames forming a light source, and a driving circuit connected to the LED lamps, the LED lamp fixing frame includes a lower base with a cavity and an upper lamp frame assembled in the cavity of the lower base, the upper lamp frame can move up and down on the lower base to adjust the height of the LED lamp fixing frame, and the LED lamp is assembled on the upper lamp frame; Figure 3 As shown, the driving circuit includes a circuit board formed by welding chips, adjustable potentiometers, capacitors, and resistors in series and parallel. The input end of the circuit board is connected to an external power adapter, and the output end of the circuit board is connected to an LED lamp; in this example, the LED lamps are two ultraviolet LED lamps with a wavelength of 254nm.

[0039] In this example, the driving circuit U1's pin 1 is connected to U2's pin 2, U1's pin 2 is connected to one end of R2, and pin 3 is connected to one end of R3. The other ends of R2 and R3 are both connected to P1's pin 2. P1's pin 1 is connected to P2's pin 2. U2's pin 3 is connected to one end of R4, and the other end of R4 is connected to P2's pin 2. U2's pin 2 is connected to R1 and one end of C2. The other end of R1 is connected to U2's pin 3, and the other end of C2 is connected to P2's pin 2. U2's pin 1 is connected to P2 Pin 1 of U1 and one end of C1, and the other end of C1 is connected to pin 2 of P2. The model of U1 is LM317T, the model of U2 is LT3092, the resistor R1 is used to adjust the current, and the resistor R2 is used to adjust the voltage. The resistance value of R1 is 6Ω, the resistance value of R2 is 200Ω, the model of R3 is X103, and the model of R4 is W101. P1 is an LED lamp, P2 is a power supply, C1 is a 1μF capacitor, and C2 is a 104 capacitor.

[0040] The capillary is provided with a detector, one end of the capillary is inserted into a first centrifuge tube filled with electrophoresis buffer solution, the first centrifuge tube is connected to a ground end, and the other end is inserted into a second centrifuge tube filled with electrophoresis buffer solution after sample injection and illumination, the second centrifuge tube is connected to a high-voltage end.

[0041] A capillary electrophoresis method for detecting nitrate and total nitrogen in a high conductivity water sample comprises the above-mentioned detection system for nitrate and total nitrogen in a high conductivity water sample, and the method comprises the following steps:

[0042] 1) The polyimide coating is burned off at the injection end of the capillary tube, and then the capillary tube is passed through the detector and fixed in the detector, and the fixed end of the capillary tube is inserted into a first centrifuge tube filled with an electrophoresis buffer solution and a connecting end;

[0043] 2) Place the centrifuge tube containing the electrophoresis buffer solution in the headspace bottle, insert the capillary injection end into the electrophoresis buffer solution in the centrifuge tube, and use nitrogen gas with a pressure of 0.15 MPa to push the electrophoresis buffer solution in the centrifuge tube into the capillary and rinse for several minutes;

[0044] 3) Manually inject the barrier liquid and the test liquid placed on the injection station into the capillary tube in sequence, wherein the injection time of the barrier liquid is longer than the injection time of the test liquid. After the injection, the capillary tube is gently placed on the capillary tube placement rack. In this example, the barrier liquid is manually injected for 120 seconds and the test liquid is injected for 20 seconds;

[0045] 4) Place the LED light source parallel and close to both sides of the capillary injection port after the polyimide coating is removed to illuminate for 5 minutes to perform a photoreduction reaction;

[0046] 5) The separation voltage of the electrophoresis system was adjusted to -15 kV, and the capillary injection end after the photoreduction reaction was completed was placed in a second centrifuge tube connected to the high-voltage end and filled with electrophoresis buffer solution. The high-voltage power supply and the data acquisition program TraceDec Monitor were manually triggered to realize the detection.

[0047] The height of the capillary placement rack is the same as the height of the detector to ensure that the capillary is in a horizontal state during illumination to prevent sample diffusion.

[0048] The detector is a non-contact conductivity detector.

[0049] The capillary is a fused silica capillary.

[0050] During operation, a complete electrophoresis circuit is formed between the high voltage terminal electrode and the ground terminal electrode.

[0051] All reagents used in this example were of analytical grade or higher purity and were purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. unless otherwise specified. The resistivity of the ultrapure water used in the experiment was 18.2 MΩ·cm.

[0052] The electrophoresis buffer solution in this example is 1.2 mL of glacial acetic acid and 0.243 g of TRIS, mixed and dissolved to make up to 100 mL.

[0053] Nitrate standard stock solution (100 mM): Weigh 1.011 g potassium nitrate and dissolve it to 100 mL.

[0054] Sodium methanesulfonate (100 mM): weigh 1.181 g of sodium methanesulfonate and dissolve it to make up to 100 mL.

[0055] The photoreduction reagent is 0.895 g of disodium hydrogen phosphate dodecahydrate and 0.059 g of EDTA dissolved to a volume of 100 mL.

[0056] The barrier liquid is prepared by dissolving 0.895 g of disodium hydrogen phosphate dodecahydrate, 0.059 g of EDTA and 3.222 g of sodium sulfate decahydrate to a volume of 100 mL, and adjusting the pH to 8.5 with a 2M sodium hydroxide solution.

[0057] In this example, the sample injection table and capillary tube rack are auxiliary detection equipment, so the sample injection table, the utensils and objects on the sample injection table, and the capillary tube rack are not shown in the figure.

[0058] 1. Impact of barrier fluid:

[0059] Since the electrophoresis buffer system is acidic (pH=3.7) and the test liquid is alkaline (pH=8.5), the pH difference between the two is too large. Therefore, when the test liquid is exposed to light directly after the buffer solution, the pH of the test liquid is reduced, the photoreduction reaction is not easy to occur, and the photoreduction efficiency is extremely poor. A long "barrier liquid" is added between the buffer solution and the test liquid. The pH of the barrier liquid is the same as that of the test liquid, so that the pH of the test liquid remains stable during the photoreduction reaction, greatly improving the reduction efficiency.

[0060] Although the addition of barrier fluid improves the photoreduction efficiency, the electrodeposition effect of nitrite and methanesulfonate ion peaks is extremely poor, and severe tailing occurs. To solve this problem, high concentrations of sulfate ions are added to the barrier fluid to enhance their electrodeposition enrichment effect. The results are shown in Figure 2. Figure 4 As shown, the accumulation effect of nitrite and methanesulfonate was significantly enhanced.

[0061] 2. Working curve, detection limit and parallelism:

[0062] Using this example, a working curve is drawn for 0-500 μM nitrate standard solution. The results are as follows: Figure 5 As shown, the nitrate concentration showed a good linear relationship with the peak area of ​​nitrite in the range of 0-300 μM, and the linear correlation became worse after exceeding 300 μM. The RSD value of 11 consecutive repeated measurements of the 200 μM nitrate standard solution was 4.9%. The UV reduction efficiency of nitrate was calculated to be 66.4%, and the detection limit of the system was calculated to be 12.71 μM at a three-fold signal-to-noise ratio.

[0063] 3. Application of total nitrogen detection system:

[0064] The nitrate and total nitrogen detector described in this example was used to determine the total nitrogen content in the actual water sample. At the same time, a spike recovery experiment was performed on it. The results showed that the spike recovery rate in this example was between 97% and 109%, as shown in Table 1, indicating that the entire detection system has good stability.

[0065] Table 1 Actual water sample detection and base spike results

[0066]

[0067] This example was compared with the national standard method (alkaline potassium persulfate digestion ultraviolet spectrophotometry), and the results are shown in Table 2. The |RD| of the two methods is less than 8%, and their acceptable levels are consistent, which proves the feasibility of the low-energy column photoreduction detection system described in this example for the detection of total nitrogen and nitrate in water samples.

[0068] Table 2 Comparison results between this method and national standard method

[0069]

Claims

1. A capillary electrophoresis method for detecting nitrate and total nitrogen in high conductivity water samples, characterized in that: The invention comprises a detection system for nitrate and total nitrogen in a high conductivity water sample, wherein the detection system comprises a pH control unit in a capillary tube and an ultraviolet LED light reduction unit. The pH control unit is composed of three sections of different pH solutions, which are formed by capillary injection, wherein the three sections of different pH solutions include an electrophoresis buffer solution of pH=3.7, a barrier solution obtained by mixing a photoreduction reagent with a pH of 8.5 and sulfate ions, and a test solution of pH=8.5, wherein a photoreduction reagent, an internal standard substance and a nitrate are added to the test solution, and the internal standard substance is sodium methanesulfonate; The ultraviolet LED light restoration unit comprises a group of LED lamps with LED lamp fixing frames forming a light source, and a driving circuit connected to the LED lamps, wherein the LED lamp fixing frame comprises a lower base with a cavity and an upper lamp frame assembled in the cavity of the lower base, the upper lamp frame can move up and down on the lower base to adjust the height of the LED lamp fixing frame, and the LED lamp is assembled on the upper lamp frame; the driving circuit comprises a circuit board formed by welding electronic components of a chip, an adjustable potentiometer, a capacitor, and a resistor in series and parallel, the input end of the circuit board is externally connected to a power adapter, and the output end of the circuit board is connected to the LED lamp; The method comprises the following steps: 1) Burning off the polyimide coating at the injection end of the capillary, then passing the capillary across the detector and fixing it in the detector, inserting the fixed end of the capillary into a first centrifuge tube filled with an electrophoresis buffer solution and a connecting end; 2) Place the centrifuge tube containing the electrophoresis buffer solution in the headspace bottle, insert the capillary injection end into the electrophoresis buffer solution in the centrifuge tube, and use nitrogen gas with a pressure of 0.15 MPa to push the electrophoresis buffer solution in the centrifuge tube into the capillary and flush for several minutes; 3) Manually inject the barrier liquid and the test liquid placed on the injection table into the capillary tube in sequence, wherein the injection time of the barrier liquid is longer than the injection time of the test liquid. After the injection, the capillary tube is gently placed on the capillary tube placement rack; 4) Place the LED light source parallel and close to both sides of the capillary injection port after the polyimide coating is removed to illuminate for photoreduction reaction; 5) Adjust the separation voltage of the electrophoresis system to -15 kV, place the capillary injection end after the photoreduction reaction into the second centrifuge tube connected to the high-voltage end and filled with electrophoresis buffer solution, and manually trigger the high-voltage power supply and data acquisition program TraceDecMonitor to achieve detection.

2. The capillary electrophoresis method for detecting nitrate and total nitrogen in high conductivity water samples according to claim 1, characterized in that: The capillary is provided with a detector, one end of the capillary is inserted into a first centrifuge tube filled with an electrophoresis buffer solution, the first centrifuge tube is connected to a ground end, and the other end is inserted into a second centrifuge tube filled with an electrophoresis buffer solution after injection and illumination, the second centrifuge tube is connected to a high-voltage end, the end of the capillary inserted into the first centrifuge tube is a fixed end, and the end of the capillary inserted into the second centrifuge tube is an injection end.

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