Quantitative / semi-quantitative detection method and application of paraquat or diquat

Abstract

The invention provides a quantitative / semi-quantitative detection method of paraquat or diquat and application, and relates to the technical field of analysis and detection. The quantitative detection method comprises the following steps: adding a reducing sugar solution and an alkaline buffer solution into a to-be-detected sample, uniformly mixing, determining an absorbance value by using a microplate reader, and calculating according to a standard curve to obtain the concentration of paraquat or diquat so as to realize quantitative detection of paraquat or diquat, in addition, semi-quantitative detection can be realized by a colorimetric method after a reducing sugar solution and an alkaline solution are added. Compared with a quantitative or semi-quantitative method in the prior art, the technical scheme provided by the invention is safer and more economical, has the characteristics of simplicity in operation, high analysis speed, stable solution color development and high clinical accessibility, and is beneficial to batch comparison of samples collected at different time points; environment detection of paraquat or diquat and rapid and real-time detection of blood and urine samples of poisoned patients are realized.

Description

Technical Field

[0001] The present invention relates to the technical field of analysis and detection, and in particular to a quantitative / semi-quantitative detection method and application of paraquat or diquat. Background Art

[0002] While the use of pesticides improves agricultural efficiency, it also poses a threat to human safety. Among them, paraquat and diquat are widely used as pesticide herbicides due to their excellent performance. However, during the use, improper operation may cause paraquat (PQ) or diquat (DQ) poisoning. Diquat is a conductive contact herbicide that can be quickly absorbed by plant tissues and loses its activity quickly after contact, so it is widely used; and in the removal of grass weeds, it is usually used together with paraquat to achieve better weed control effects. The symptoms, treatment plans and prognosis of paraquat and diquat poisoning are closely related to the intake. After paraquat is absorbed, it is distributed in the blood and organs throughout the body through the blood circulation. It is mostly excreted from the kidneys in its original form without metabolism. Therefore, plasma and urine are often used as test samples for paraquat in poisoned patients. Therefore, the test results of paraquat or diquat concentrations in blood and urine are of guiding significance for clinical treatment.

[0003] At present, the detection methods for blood and / or urine samples containing paraquat and diquat mainly include gas chromatography, high performance liquid chromatography, liquid chromatography-mass spectrometry, capillary electrophoresis, surface enhanced Raman spectroscopy, enzyme-linked immunosorbent assay, electrochemical method, etc. However, the above methods may have complicated sample pretreatment process, long detection time, or expensive equipment, resulting in long detection time or high cost. Spectrophotometry is also a rapid detection method for paraquat or diquat concentration detection, which has the advantages of high sensitivity and simple operation. In the prior art, the commonly used spectrophotometric method for detecting the content of paraquat and diquat is the sodium bicarbonate-sodium dithionite colorimetric method, but the colorimetric method based on sodium bicarbonate-sodium dithionite is unstable, resulting in poor detection accuracy. For example, Chinese invention patent CN110220892A discloses a diquat semi-quantitative detection spectrum formulation and urine detection method, which adopts the sodium bicarbonate-sodium dithionite colorimetric method and removes protein by double reagents. Since the solubility of protein in a mixed solvent of acetonitrile and 20% trichloroacetic acid is very low, the invention directly tests the absorbance of PQ and DQ for quantitative analysis after removing protein from serum by centrifugation; however, the colorimetric development of this method is extremely unstable, so that accurate quantitative analysis cannot be achieved, so only semi-quantitative detection can be performed. Moreover, in the actual application process, the above method has safety hazards due to the fact that the reagent sodium dithionite powder required to be used has the properties of spontaneous combustion, explosiveness, toxicity, and flammability when in contact with water, which further limits the promotion and application of this detection method.

[0004] The present invention is based on the prior art and aims to improve the color development of the sodium bicarbonate-sodium dithionite colorimetric method, which has the problem of unstable color development and potential safety hazards of the color developer. The invention proposes an optical detection method for rapidly measuring the concentration of paraquat and diquat. The method can safely, stably and rapidly quantify the content of paraquat or diquat in the sample to be tested. Summary of the invention

[0005] In view of the problems existing in the prior art, the present invention provides a quantitative / semi-quantitative detection method and application of paraquat or diquat, which can safely, stably and quickly quantitatively or semi-quantitatively detect the content of paraquat or diquat in samples such as serum and urine.

[0006] In order to achieve the above-mentioned purpose, the present invention provides a method for quantitatively detecting the content of paraquat or diquat, comprising adding a color stabilizer to a sample to be tested, measuring the absorbance value after mixing evenly, and calculating the concentration of paraquat or diquat according to a standard curve to achieve quantitative detection of paraquat or diquat; the color stabilizer is a reducing sugar solution and an alkaline buffer solution.

[0007] Preferably, the absorbance value detection equipment includes an ultraviolet spectrophotometer, a fully automatic / semi-automatic biochemical analyzer, an ELISA reader, etc., but is not limited thereto.

[0008] Preferably, in the reducing sugar solution, the reducing sugar includes any one of fructose, glucose, lactose, galactose and maltose, or a combination of several of them.

[0009] Preferably, the alkaline buffer solution is any one of a dipotassium hydrogen phosphate buffer solution, a disodium hydrogen phosphate buffer solution, a sodium hydroxide-sodium carbonate buffer solution, and a potassium hydroxide-sodium carbonate buffer solution.

[0010] Preferably, the quantitative detection method of paraquat or diquat comprises the following steps: S1. Prepare paraquat standard solution and diquat standard solution respectively; S2. Establish a regression equation for paraquat or diquat; The reducing sugar solution and the alkaline buffer solution are added to the paraquat standard solution and the diquat standard solution respectively, and the absorbance is detected after being evenly mixed, and a standard curve of absorbance and paraquat or diquat concentration is drawn to establish a regression equation; S3. Pre-treatment of samples to be tested Take a sample containing paraquat or diquat to prepare a sample solution to be tested; S4. Determination of Paraquat or Diquat Content in Test Samples The reducing sugar solution and the alkaline buffer solution are added to the sample solution to be tested, and absorbance is detected after being evenly mixed. The content of paraquat or diquat is calculated according to the regression equation of the concentration of paraquat or diquat.

[0011] Preferably, in S1, the concentrations of the paraquat standard solution and the diquat standard solution are 0-320 μg / mL, excluding 0.

[0012] Preferably, the detection wavelength in S2 is its characteristic absorption wavelength, the characteristic absorption wavelength of paraquat is 500-700 nm, and the characteristic absorption wavelength of diquat is 350-400 nm.

[0013] More preferably, the detection wavelength of paraquat is 600 nm and the detection wavelength of chloramphenicol is 380 nm.

[0014] In some specific embodiments, the regression equation and the degree of fit (R 2 ) are: Regression equation of paraquat standard solution: b=0.0218a-0.0005, R 2 =0.999; The regression equation of the standard solution of diquat: y=0.0433x-0.0042, R 2 =0.999.

[0015] Preferably, the concentration of the reducing sugar solution is 5-960 mM.

[0016] Preferably, the alkaline buffer solution has a pH value of ≥11 and a concentration of 0.1-2M.

[0017] Preferably, in S3, the content of paraquat or diquat in the sample solution to be tested should be within the linear range of the regression equation. If the concentration of the prepared sample solution to be tested exceeds the linear range, the sample to be tested should be diluted or concentrated again, and then the operation of S4 should be continued to detect the content of paraquat or diquat.

[0018] Preferably, in S4, the adding ratio of the reducing sugar solution and the alkaline buffer solution is the same as that in S2.

[0019] Preferably, the sample to be tested includes a blood sample or a urine sample.

[0020] As one of the purposes of the invention, the present invention also provides a semi-quantitative detection method for paraquat or diquat, which uses a colorimetric control method to semi-quantitatively detect the concentration of paraquat and diquat in water.

[0021] Specifically, the semi-quantitative detection method includes adding a color stabilizer to the sample to be tested, and performing a color comparison with a standard color scale card to achieve semi-quantitative detection of paraquat or diquat.

[0022] Preferably, the semi-quantitative detection method of paraquat or diquat comprises: respectively adding the reducing sugar solution and the alkaline buffer solution to a paraquat standard solution and / or a diquat standard solution, mixing them evenly, and arranging them in a color development order of the concentration gradient of the standard solution to prepare a standard color scale card; preparing a sample solution to be tested, adding the reducing sugar solution and the alkaline buffer solution, allowing the sample solution to be tested to develop color, and comparing the color with that of the standard color scale card, thereby achieving semi-quantitative detection of paraquat or diquat.

[0023] Preferably, the color of the standard color scale card gradually deepens with the increase of concentration; the color of the standard color scale card of paraquat gradually changes from colorless to blue, and the color of the standard color scale card of diquat gradually changes from colorless to green.

[0024] The quantitative / semi-quantitative detection of paraquat or diquat based on the above technical solution can be applied to the environment, including the detection of paraquat or diquat content in samples such as water, soil, and food, or the detection of paraquat or diquat content in samples such as serum or urine of poisoned patients.

[0025] Different from the prior art method of semi-quantitative analysis by directly measuring the absorbance of paraquat or diquat, the inventive principle of the present invention includes: in the presence of an alkaline buffer solution, a reducing agent can reduce paraquat or diquat to form a cationic free radical that absorbs visible light, thereby developing color; under aerobic conditions, the free radical will be oxidized to the original paraquat or diquat, so that the solution does not develop color. The concentration and stability of the reducing agent determine the stability of the cationic free radical. The present invention uses reducing sugar as a reducing agent instead of sodium dithionite to enhance the color development efficiency and color development stability.

[0026] Beneficial technical effects obtained by the present invention: (1) The color development method using reducing sugars according to the technical solution of the present invention is safer and more economical than the color development methods of the prior art, such as the sodium bicarbonate-sodium dithionite color development method. It also has the characteristics of simple operation and fast analysis speed. It can realize environmental detection of paraquat or diquat, and in particular, it can also realize rapid and real-time detection of blood and urine samples of poisoned patients, which is beneficial to the clinical diagnosis of paraquat and diquat and has guiding significance for the rescue of patients.

[0027] (2) Compared with the existing sodium bicarbonate-sodium dithionite colorimetric method, the semi-quantitative detection method of the present invention uses reducing sugar instead of sodium dithionite as a colorimetric agent, and the color development is more stable. The absorbance remains basically unchanged within 0.5 hours, and the color is still clearly discernible after 24 hours, which is conducive to large-scale high-throughput testing of samples and comparison of samples collected at different time points.

[0028] (3) The present invention uses an optical detection method to quantitatively detect paraquat or diquat. Compared with commonly used quantitative methods, the present invention has the advantages of being simple to operate, convenient and quick, and not requiring the use of large instruments. It can reduce detection costs and shorten detection time. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 It is the UV-visible absorption spectrum of the paraquat standard solution after adding the color stabilizer in Example 1 of the present invention.

[0030] Figure 2 It is a UV-visible absorption spectrum of the diquat standard solution after adding a color stabilizer in Example 1 of the present invention.

[0031] Figure 3 This is a standard curve diagram established according to absorbance and paraquat concentration in Example 1 of the present invention.

[0032] Figure 4 This is a standard curve diagram established according to absorbance and diquat concentration in Example 1 of the present invention.

[0033] Figure 5 This is the standard color scale card after the paraquat in Example 2 of the present invention is colored.

[0034] Figure 6 It is the standard color scale card after the diquat of Example 2 of the present invention is developed.

[0035] Figure 7 It is a comparison chart of the absorbance change over time of paraquat after color development in Example 1 of the present invention and Comparative Example 1.

[0036] Figure 8 It is a comparison chart of the absorbance change over time of diquat after color development in Example 1 of the present invention and Comparative Example 1.

[0037] Figure 9a-9e They are respectively graphs showing the change in absorbance of paraquat over time under different pH conditions in Example 3 of the present invention.

[0038] Figure 10a-Figure 10e They are respectively graphs showing the change in absorbance of diquat over time under different pH conditions in Example 3 of the present invention.

[0039] Figure 11a-Figure 11bThey are respectively graphs showing the change in absorbance over time of paraquat and diquat concentrations detected at different fructose concentrations in Example 4 of the present invention.

[0040] Fig.12a and Figure 12b The following are comparison diagrams of the absorbance changes over time (800 min) of paraquat and diquat concentrations detected in Examples 5 to 7 of the present invention and Comparative Example 2. DETAILED DESCRIPTION

[0041] In view of the defects of the prior art, the inventors of this case have proposed the technical solution of the present invention after long-term research and extensive practice. The technical solution mainly utilizes reducing sugar instead of sodium dithionite as a color developer, which makes the color development more stable, and the absorbance remains basically unchanged within 0.5 hours. The color is still clearly discernible after 24 hours, which is conducive to large-scale high-throughput testing of samples and comparison of samples collected at different time points, and realizes semi-quantitative detection of paraquat or diquat; moreover, in combination with the optical detection method, quantitative detection of paraquat or diquat is realized by establishing a standard curve and a regression equation.

[0042] The technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0043] Unless otherwise specified, the experimental materials used in the following examples can be purchased from conventional biochemical reagent companies.

[0044] Example 1 The method of the present invention for rapid quantitative detection of paraquat and diquat content based on optical detection method specifically comprises the following steps: (1) Preparation of paraquat and diquat standard solutions Prepare paraquat standard solution and diquat standard solution with gradient concentrations of 4 μg / mL, 8 μg / mL, 20 μg / mL, 40 μg / mL, 80 μg / mL, 160 μg / mL, 240 μg / mL, and 320 μg / mL respectively; (2) Establishment of standard curve Take 50 μL of the above-mentioned gradient concentration paraquat standard solution and diquat standard solution, add 50 μL of 960 mM fructose solution and 100 μL of 1 M potassium dihydrogen phosphate buffer solution with a pH of 12, mix well, and use an enzyme reader to measure the absorbance at wavelengths of 600 nm and 380 nm.

[0045] See also Figure 1 and Figure 2 They are respectively the UV-visible absorption spectra of paraquat standard solution and diquat standard solution after adding color stabilizer. The measurement results are shown in Table 1 and Table 2, and standard curves are established based on absorbance and paraquat and diquat concentrations.

[0046] See also Figure 3 and Figure 4 , the regression equation and fitting degree (R 2 )as follows: Paraquat: b = 0.0218a-0.0005, R 2 =0.999; Diquat: y=0.0433x-0.0042, R 2 =0.999.

[0047] Table 1 Absorbance value record of paraquat standard solution

[0048] Table 2 Absorbance value record of diquat standard solution

[0049] (3) Pretreatment of the sample to be tested The sample containing paraquat and diquat was diluted at different times and the method of step (4) of this example was used to determine whether it was within the linear range of the regression equation (0-320 μg / mL, excluding 0). If not, the sample was further diluted.

[0050] (4) Measurement of paraquat and diquat concentrations Take 50 μL of the paraquat and diquat-containing sample treated in step (3), add 50 μL of 960 mM fructose solution and 100 μL of 1 M potassium dihydrogen phosphate buffer solution with a pH of 12, mix well, and use an ELISA reader to measure the absorbance at wavelengths of 600 nm and 380 nm. The concentrations of paraquat and diquat are calculated using the measured absorbance and the standard curve obtained in step (2).

[0051] (5) Recovery rate Take 1 mL of blank solution and add working solution to determine the recovery of the method. The results are shown in Tables 3 and 4.

[0052] It can be seen from the results in Table 3 and Table 4 that the quantitative detection of paraquat and diquat using the technical solution of this embodiment has good accuracy and reliability.

[0053] Table 3 Recovery of paraquat in water, artificial urine and fetal bovine serum

[0054] Table 4 Recovery of diquat in water, artificial urine and fetal bovine serum

[0055] Example 2 This embodiment provides a method for semi-quantitative detection of paraquat and diquat content based on reducing sugar colorimetric method, which specifically comprises the following steps: (1) Preparation of paraquat and diquat standard solutions Prepare paraquat and diquat standard solutions with concentrations of 0 μg / mL, 3 μg / mL, 10 μg / mL, 30 μg / mL, and 100 μg / mL, respectively; (2) Preparation of standard color scale cards Take 2 mL of the above paraquat and diquat standard solutions respectively, add 86.4 mg of fructose and 1 mL of lyophilized powder made of 1 M potassium hydrogen phosphate buffer solution at pH = 12, mix well and prepare a standard color scale card according to the color of the solution.

[0056] Standard color scale card Figure 5 and Figure 6 As shown, Figure 5 This is the standard color scale card for paraquat. Paraquat appears blue after color development. Figure 6 This is the standard color scale card of diquat. Diquat appears green after color development. Both of them become darker in color with increasing concentration.

[0057] (3) Pretreatment of the sample to be tested The test samples containing paraquat and diquat were diluted at different times, and the method of step (2) of this embodiment was used to determine whether their colors were within the range of the standard color scale card. If not, the samples were further diluted. 2 mL of paraquat and diquat were taken, respectively, and 1 mL of freeze-dried powder prepared from 1 M potassium hydrogen phosphate buffer solution at pH = 12 was added. The mixture was mixed evenly and centrifuged to obtain the supernatant.

[0058] (4) Measurement of concentrations of paraquat and diquat Take 1 mL of the sample containing paraquat and diquat treated in step (3), add 43 mg of fructose, mix well, and compare with the standard color scale card to obtain the concentrations of paraquat and diquat in the sample.

[0059] Example 3

[0060] This example adopts the same technical scheme as Example 1, and simultaneously examines the pH values ​​of the dipotassium hydrogen phosphate buffer solution (dipotassium hydrogen phosphate-sodium hydroxide solution) adjusted to 11, 11.5, 12, 12.5, and 13 using different concentrations of sodium hydroxide.

[0061] The concentrations of potassium hydrogen phosphate were 0.1 M, 0.25 M, 0.5 M, 0.75 M, and 1 M, respectively.

[0062] In the measurement of the concentration of paraquat and diquat in step (4) of Example 1, different buffer solutions are selected, specifically comprising: taking 50 μL of the paraquat and diquat sample to be tested after treatment in step (3), adding 50 μL of 960 mM fructose solution, mixing evenly, using the above-mentioned gradient concentration and pH gradient potassium hydrogen phosphate-sodium hydroxide solution as the alkaline buffer solution, and then using an ELISA instrument to measure the absorbance changes at wavelengths of 600 nm and 380 nm over time.

[0063] See also Figure 9a-9e , which are the absorbance variation of paraquat over time under different potassium dihydrogen phosphate concentrations at pH 11, 11.5, 12, 12.5 and 13 respectively. It can be seen from the figure that the ultraviolet absorption stability at pH 11 and 11.5 is relatively poor and the color development stability is relatively slow compared with that at pH=12; the stability is better at pH=12, and then the stability decreases slightly with the increase of pH; considering that higher pH has certain corrosiveness, pH=12 is preferred in clinical applications.

[0064] See also Figure 10a-Figure 10e , which are the absorbance variation of diquat over time under the conditions of pH 11, 11.5, 12, 12.5 and 13 respectively. The variation pattern and effect are the same as the results of paraquat detection.

[0065] Example 4

[0066] This example adopts the same technical scheme as that of Example 1, except that the final concentrations of the fructose solutions selected in this example after mixing are 5 mM, 10 mM, 20 mM, 40 mM, 60 mM, 120 mM, 240 mM, 360 mM, 480 mM and 960 mM, respectively, and the other conditions and steps are the same.

[0067] See also Fig.11a and Fig.11b The graphs are respectively the ultraviolet absorption variation over time of paraquat and diquat concentrations detected in this example. It can be seen from the figure that within the concentration range of 5-960 mM, the detection effect is equivalent to that of Example 1, and has good stability.

[0068] Example 5

[0069] This example adopts the same technical scheme as that of Example 1, except that, in this example, after adding fructose solution and alkaline buffer solution to the paraquat or diquat test sample, the absorbance change at 600 nm and 380 nm over time is measured by an ELISA instrument, respectively, once every 10 minutes.

[0070] The other steps are the same as those in Example 1.

[0071] Example 6

[0072] This example adopts the same technical solution as Example 1, and simultaneously examines the effect of using sodium carbonate-sodium hydroxide with pH=12 as a buffer solution for paraquat and diquat concentration detection.

[0073] The concentration of sodium carbonate was 1M.

[0074] In the measurement of paraquat and diquat concentrations in step (4), different buffer solutions are selected, specifically including: taking 50 μL of the paraquat and diquat-containing sample treated in step (3), adding 50 μL of 960 mM fructose solution and 100 μL of sodium carbonate-sodium hydroxide solution with a pH of 12 as an alkaline buffer solution, and then using an ELISA reader to measure the absorbance changes at wavelengths of 600 nm and 380 nm over time, once every 10 minutes.

[0075] The other steps are the same as those in Example 1.

[0076] Example 7

[0077] This example adopts the same technical solution as Example 1, and simultaneously examines the effect of using sodium hydroxide at pH=12 as an alkaline solution to detect the concentrations of paraquat and diquat.

[0078] In the measurement of paraquat and diquat concentrations in step (4), different alkaline solutions are selected, specifically including: taking 50 μL of the paraquat and diquat-containing sample treated in step (3), adding 50 μL of 960 mM fructose solution and 100 μL of pH=12 sodium hydroxide solution as alkaline buffer solution, and then using an ELISA reader to measure the absorbance changes at wavelengths of 600 nm and 380 nm over time, once every 10 minutes.

[0079] The other steps are the same as those in Example 1.

[0080] Comparative Example 1

[0081] In this comparative example, the sodium dithionite colorimetric method was used to perform semi-quantitative detection of paraquat and diquat, and the method was compared with the optical detection method and colorimetric method of Example 2.

[0082] Paraquat solution and diquat solution were taken separately, and reducing sugar and dipotassium hydrogen phosphate buffer solution were added respectively according to the method of Example 2. The final concentrations of the paraquat solution and diquat solution were 30 μg / mL.

[0083] Separately prepare a paraquat solution and a diquat solution to which a freshly prepared mixture of sodium bicarbonate (100 mg / 2 mL) and sodium dithionite powder (50 mg) is added to obtain the same final concentration.

[0084] First, a color contrast was performed, and the changes in their absorbance over time were measured using an enzyme-labeled instrument every 10 minutes.

[0085] See also Figure 7 This is a comparison chart of the absorbance change over time of paraquat after color development in Example 2 of the present invention and Comparative Example 1, Figure 8 The following is a comparison of the absorbance changes over time after the color development of diquat in Example 2 of the present invention and Comparative Example 1. As can be seen from the figure, after the color development is stable, the absorbance values ​​of the two methods are equivalent, and the color development speed is equivalent. However, reducing sugar is used as a color developer, and the color development is more stable. The absorbance remains basically unchanged within 30 minutes, and the color is still clearly discernible after 800 minutes, so it is more suitable for large-scale high-throughput testing of samples and comparison of samples collected at different time points, so as to achieve semi-quantitative detection of paraquat or diquat. From a quantitative analysis, within about 100 minutes, the absorbance value of the sodium dithionite color development method continues to increase, and then the absorbance value quickly drops to 0, and the sample absorbs light failure; while the absorbance value of the sample with fructose added remains stable within 24 hours, with almost no change. Obviously, the optical detection stability of the method of adding reducing sugar is significantly higher than that of the sodium dithionite color development method.

[0086] Comparative Example 2

[0087] This comparative example adopts the same technical solution as Example 1, and simultaneously examines the effect of using a 16.7 mg / mL sodium bicarbonate solution as a buffer solution for paraquat and diquat concentration detection.

[0088] The pH of sodium bicarbonate solution is the natural pH value (pH ~ 8).

[0089] In the measurement of paraquat and diquat concentrations in step (4), different buffer solutions are selected, specifically including: taking 50 μL of the paraquat and diquat-containing sample treated in step (3), adding 50 μL of 960 mM fructose solution and 100 μL of sodium bicarbonate solution as alkaline buffer solution, and then using an ELISA reader to measure the absorbance changes at wavelengths of 600 nm and 380 nm over time, once every 10 minutes.

[0090] The other steps are the same as those in Example 1.

[0091] See also Fig.12a and Figure 12b The absorbance changes over time (0-800 min) of paraquat and diquat concentrations detected in Examples 5-7 and Comparative Example 2 are respectively shown. As can be seen from the figure, the response speed of potassium dihydrogen phosphate-potassium hydroxide (Example 5) as an alkaline buffer solution is the fastest, and the absorbance remains stable, followed by sodium carbonate-sodium hydroxide solution (Example 6). Sodium hydroxide solution (Example 7) can also detect absorbance, but compared with other alkaline buffer solutions, the response speed is relatively slow, and the color development time is relatively short. The reason is that after sodium hydroxide is mixed with reducing sugar as an alkaline buffer solution, the pH changes greatly, resulting in poor system stability and a short color development time, and the buffer solution cannot play the role of balancing pH. When sodium bicarbonate solution (Comparative Example 2) is used as an alkaline buffer solution, absorbance cannot be detected.

[0092] In summary, compared with the traditional sodium dithionite color development method, the optical detection method for detecting the concentration of paraquat and diquat used in the present invention can achieve a more stable color development effect. The technical solution of the present invention can provide sufficient time for intuitive comparison of the two test results of clinical symptoms in clinical practice, and has more practical clinical detection value.

[0093] In addition, the inventors of this case also referred to the aforementioned embodiments and conducted experiments with other raw materials, process operations, and process conditions described in this specification, and obtained relatively ideal results.

[0094] The various aspects, embodiments, features and examples of the present invention should be considered as illustrative in all aspects and are not intended to limit the present invention, the scope of the present invention is defined only by the claims. Other embodiments, modifications and uses will be apparent to those skilled in the art without departing from the spirit and scope of the claimed invention.

[0095] The use of headings in the present invention is not meant to limit the present invention; both the Summary and Detailed Description sections may apply to any aspect, embodiment, or feature of the present invention.

[0096] Throughout the present disclosure, where a composition is described as having, comprising, or including particular components, or where a process is described as having, comprising, or including particular process steps, it is contemplated that the compositions taught by the present invention also consist essentially of, or consist of, the recited components, and that the processes taught by the present invention also consist essentially of, or consist of, the recited process steps.

[0097] Unless specifically stated otherwise, use of the terms "include," "includes," "including," "have," "has," or "having" should generally be construed as open ended and non-limiting.

[0098] It should be understood that the order of the various steps or the order in which a particular action is performed is not very important, as long as the teachings of the present invention remain operable. In addition, two or more steps or actions may be performed simultaneously.

[0099] The above are only preferred embodiments of the present invention, which do not limit the protection scope of the present invention. For those skilled in the art, the present invention may have various modifications and changes. Within the spirit and principle of the present invention, changes, modifications, replacements, integrations and parameter changes to these embodiments by conventional substitutions or capable of achieving the same functions without departing from the principles and spirit of the present invention all fall within the protection scope of the present invention.

Claims

1. A method for quantitative detection of paraquat or diquat, characterized in that: A color stabilizer is added to the sample to be tested, and after being mixed evenly, the ultraviolet light absorbance value is measured, and the concentration of paraquat or diquat is calculated according to the standard curve to achieve quantitative detection of paraquat or diquat; The color development stabilizer is a reducing sugar solution and an alkaline buffer solution.

2. The quantitative detection method according to claim 1, characterized in that: In the reducing sugar solution, the reducing sugar includes any one or a combination of fructose, glucose, lactose, galactose and maltose; The alkaline buffer solution is any one of a dipotassium hydrogen phosphate buffer solution, a disodium hydrogen phosphate buffer solution, a sodium hydroxide-sodium carbonate buffer solution, and a potassium hydroxide-sodium carbonate buffer solution.

3. The quantitative detection method according to claim 1, characterized in that The quantitative detection method of paraquat or diquat comprises the following steps: S1. Prepare paraquat standard solution and diquat standard solution with gradient concentration respectively; S2. Establish a regression equation for paraquat or diquat; The reducing sugar solution and the alkaline buffer solution are added to the paraquat standard solution and the diquat standard solution respectively, and the absorbance is detected after being evenly mixed, and a standard curve of absorbance and paraquat concentration or a standard curve of diquat concentration is drawn to establish a regression equation; S3. Pre-treatment of samples to be tested Take a sample containing paraquat or diquat to prepare a sample solution to be tested; S4. Determination of Paraquat or Diquat Content in Test Samples The reducing sugar solution and the alkaline buffer solution are added to the sample solution to be tested, and absorbance is detected after being evenly mixed. The content of paraquat or diquat is calculated according to the regression equation of the concentration of paraquat or diquat.

4. The quantitative detection method according to claim 3, characterized in that: In S1, the concentration of paraquat standard solution or diquat standard solution was 0-320 μg / mL; In S2, the absorbance value of the paraquat standard solution is detected at a wavelength of Its characteristic absorption wavelength; The detection wavelength of the absorbance value of the diquat standard solution is its characteristic absorption wavelength; The concentration of the reducing sugar solution after mixing is 5-960 mM; the pH of the alkaline buffer solution before mixing is ≥11, and the concentration is 0.1-2 M.

5. The quantitative detection method according to claim 3, characterized in that: In S3, the content of paraquat or diquat in the sample solution to be tested should be within the linear range of the regression equation. If the concentration of the prepared sample solution to be tested exceeds the linear range, the sample to be tested should be diluted or concentrated again, and then the operation of S4 should be continued to perform quantitative detection of the content of paraquat or diquat.

6. The quantitative detection method according to claim 3, characterized in that: In S4, the adding ratio of the reducing sugar solution and the alkaline buffer solution is the same as that in S2.

7. A semi-quantitative detection method for paraquat or diquat, characterized in that: Add a color stabilizer to the sample to be tested and compare the color with the standard color scale card to achieve semi-quantitative detection of paraquat or diquat.

8. The semi-quantitative detection method according to claim 7, characterized in that: The reducing sugar solution and the alkaline buffer solution are added to the paraquat standard solution and / or the diquat standard solution respectively, mixed evenly, and arranged in the color development order of the concentration gradient of the standard solution to prepare a standard color scale card; The sample solution to be tested is prepared, and after adding the reducing sugar solution and the alkaline buffer solution, the sample solution to be tested develops color, and is compared with the color of the standard color scale card to achieve semi-quantitative detection of paraquat or diquat.

9. The semi-quantitative detection method according to claim 7, characterized in that: The color of the standard color scale card gradually deepens with the increase of concentration; the color of the standard color scale card of paraquat gradually changes from colorless to blue, and the color of the standard color scale card of diquat gradually changes from colorless to green.

10. Use of the quantitative detection method according to any one of claims 1 to 6, or the semi-quantitative detection method according to any one of claims 7 to 9 in the detection of paraquat or diquat content in water, soil, serum or urine samples.

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