阅读说明：本技术 一种发光型传感器及其构筑方法与在检测植物水杨酸含量中的应用 (Light-emitting sensor, construction method thereof and application of light-emitting sensor in detection of salicylic acid content in plants ) 是由 滕旭 翟喜海 李宝英 宋伟丰 潘亚清 苏雅迪 王宇 苏保华 于 2020-12-07 设计创作，主要内容包括：一种发光型传感器及其构筑方法与在检测植物水杨酸含量中的应用,属于化学发光传感分析领域。为了建立一种高灵敏度测定植物样品中水杨酸含量的化学发光分析法,本发明提供了一种利用发光型传感器测定植物样品中水杨酸含量的方法,其检测原理是将碳量子点和鲁米诺同时负载在水滑石表面后,加入氧化剂,可以获得较强的发光信号,当水杨酸加入后,可以抑制体系发光,随着水杨酸加入量的增加,体系发光信号越弱。该发光型传感器具有灵敏度高、线性范围宽、检测速度快等优点,植物样本只需简单的提取离心处理,即可直接检测植物体内水杨酸含量。该发光型传感器可广泛应用于不同植物体内水杨酸含量的快速检测。(A luminous sensor, a construction method thereof and application thereof in detecting the content of plant salicylic acid belong to the field of chemiluminescence sensing analysis. The invention provides a method for determining the content of salicylic acid in a plant sample by using a luminous sensor, which aims to establish a chemiluminescence analysis method for determining the content of salicylic acid in the plant sample with high sensitivity. The luminous sensor has the advantages of high sensitivity, wide linear range, high detection speed and the like, and the salicylic acid content in the plant can be directly detected by simply extracting and centrifuging a plant sample. The luminous sensor can be widely applied to the rapid detection of the content of salicylic acid in different plants.)

1. The light-emitting sensor is characterized by consisting of an independently packaged hydrotalcite nano material loaded with carbon quantum dots and luminol and an independently packaged oxidant, wherein the carbon quantum dots are prepared from 5,10,15, 20-tetra (1-methyl-4-pyridine) porphyrin and sodium citrate serving as raw materials.

2. The luminescence-type sensor according to claim 1, wherein the oxidizing agent is one or more of hydrogen peroxide, urea peroxide, and tert-butyl hydroperoxide.

3. A method for producing a luminescence-type sensor according to claim 1, comprising the steps of:

step one, preparing a carbon quantum dot: dissolving 1-5g of 5,10,15, 20-tetra (1-methyl-4-pyridine) porphyrin and 1-6g of sodium citrate in 25mL of deionized water, and reacting the obtained solution at 200 ℃ for 6 hours;

step two, preparing the hydrotalcite nano material loaded with the carbon quantum dots and the luminol: adding 1-10mg of carbon quantum dots, 10-20g of hydrotalcite and 20-50mg of luminol into 1L of deionized water, stirring for 1 hour at 25 ℃, centrifuging to remove supernatant, and washing with deionized water for three times.

4. Use of the luminescent sensor according to claim 1 for rapid detection of salicylic acid content in different plants.

5. The use of claim 7, wherein the method for detecting the content of salicylic acid is as follows:

(1) mixing a plant sample to be detected with methanol according to the ratio of 1 g: 5mL of materials are mixed, the mixture is placed in an ultrasonic cell disruptor for 5min of ultrasound, after centrifugation, supernatant is taken and dried by nitrogen, 1mL of deionized water is added to prepare a solution to be detected, and then the solution to be detected is diluted to prepare salicylic acid solutions with different concentrations;

(2) mixing 100 mu L of hydrotalcite nano material loaded with carbon quantum dots and luminol simultaneously with 100 mu L of salicylic acid solution obtained in the step (1) in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier;

(3) injecting 100 mu L of oxidant into the quartz bottle to obtain a chemiluminescent signal of the salicylic acid solution sample;

(4) and (5) carrying out quantitative determination by adopting a standard addition method to obtain the content of the salicylic acid in the plant sample.

Technical Field

The invention belongs to the technical field of chemiluminescence sensing analysis, and particularly relates to a luminous sensor, a construction method of the luminous sensor and application of the luminous sensor in detection of the content of salicylic acid in plants.

Background

Salicylic acid is a plant hormone commonly existing in plants, and the content change of the salicylic acid in the plants is closely related to the disease resistance, drought resistance, salt resistance and the like of the plants. Although the research on salicylic acid for inducing plant stress resistance is increasing in recent years, many problems are still unclear, and particularly, the accurate determination of the content of salicylic acid in plants is a necessary prerequisite for completely determining various mechanisms of the salicylic acid in the plant stress resistance.

The reported methods for measuring the salicylic acid include a spectrum method, an ion chromatography method, a gas-mass combined method and the like, but most methods are used for measuring the content of the salicylic acid in wastewater, food and medicines, and the reports of the detection of the salicylic acid in a plant sample are less, mainly because the composition of the plant sample is more complex and the content of the salicylic acid in a plant body is very low. At present, the detection method for salicylic acid in plants only adopts a chromatographic method and an electrochemical method. The method has the defects of complex operation, expensive instrument and need of professional analysts, and the detection result of the electrochemical method is easily influenced by the environment such as temperature, pressure, external disturbance and the like, so that the method has the problem of poor reproducibility. In summary, it is a great challenge to establish a chemiluminescence assay for determining salicylic acid in plant samples with high sensitivity.

Disclosure of Invention

In order to establish a chemiluminescence analysis method for measuring the content of salicylic acid in a plant sample with high sensitivity, the invention provides a luminous sensor, which consists of an independently packaged hydrotalcite nano material loaded with carbon quantum dots and luminol and an independently packaged oxidant, wherein the carbon quantum dots are prepared from 5,10,15, 20-tetra (1-methyl-4-pyridine) porphyrin and sodium citrate serving as raw materials.

Further, the oxidant is one or more of hydrogen peroxide, carbamide peroxide and tert-butyl hydroperoxide which are mixed according to any proportion.

The invention also provides a construction method of the light-emitting sensor, which comprises the following steps:

step one, preparing a carbon quantum dot: dissolving 1-5g of 5,10,15, 20-tetra (1-methyl-4-pyridine) porphyrin and 1-6g of sodium citrate in 25mL of deionized water, and reacting the obtained solution at 200 ℃ for 6 hours;

step two, preparing the hydrotalcite nano material loaded with the carbon quantum dots and the luminol: adding 1-10mg of carbon quantum dots, 10-20g of hydrotalcite and 20-50mg of luminol into 1L of deionized water, stirring for 1 hour at 25 ℃, centrifuging to remove supernatant, and washing with deionized water for three times.

The invention also provides application of the light-emitting sensor in rapid detection of salicylic acid content in different plants.

Further, the application is that the luminous intensity is firstly measured, and then the salicylic acid concentration is obtained according to the relation between the luminous intensity and the salicylic acid concentration.

In one embodiment of the present invention, the method for detecting the content of salicylic acid is as follows:

(1) mixing a plant sample to be detected with methanol according to the ratio of 1 g: 5mL of materials are mixed, the mixture is placed in an ultrasonic cell disruptor for 5min of ultrasound, after centrifugation, supernatant is taken and dried by nitrogen, 1mL of deionized water is added to prepare a solution to be detected, and the solution to be detected is diluted to prepare salicylic acid solutions with different concentrations.

(2) Mixing 100 mu L of hydrotalcite nano material loaded with carbon quantum dots and luminol simultaneously with 100 mu L of salicylic acid solution obtained in the step (1) in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier;

(3) and injecting 100 mu L of oxidant into the quartz bottle by using a micro-injector to obtain a chemiluminescent signal of the salicylic acid solution sample.

(4) And (5) carrying out quantitative determination by adopting a standard addition method to obtain the content of the salicylic acid in the plant sample.

Advantageous effects

The invention provides a method for detecting the content of salicylic acid in different plants by using a light-emitting sensor, which has the principle that carbon quantum dots and luminol are simultaneously loaded on the surface of hydrotalcite, an oxidant is added, a stronger light-emitting signal can be obtained, when salicylic acid is added, the system can be inhibited from emitting light, and the system light-emitting signal is weaker along with the increase of the addition of the salicylic acid. The light-emitting sensor has the advantages of high sensitivity, wide linear range, high detection speed and the like. The salicylic acid content in the plant can be directly detected by simply extracting and centrifuging the plant sample. The concentration range of the salicylic acid detected by the luminous sensor is 10-800 mug/L, and the detection limit is 6 ng/mL. The luminous sensor can be widely applied to the rapid detection of the salicylic acid content in different plants, can be expanded to other environments to be detected of the salicylic acid, and lays a foundation for the visual research of the salicylic acid.

Drawings

FIG. 1.5, 10,15, 20-tetrakis (1-methyl-4-pyridine) porphyrin structure;

fig. 2 is a graph comparing the luminous intensity of a luminol-hydrogen peroxide system with the luminous intensity of a hydrotalcite nanomaterial-hydrogen peroxide system loaded with carbon quantum dots and luminol, where a is the luminous intensity of the luminol-hydrogen peroxide system, and b is the luminous intensity of the hydrotalcite nanomaterial-hydrogen peroxide system loaded with carbon quantum dots and luminol;

FIG. 3 is a graph of luminescence intensity of the luminescence-type sensor of the present invention at different concentrations of salicylic acid;

FIG. 4 is a stability test chart of the luminescence type sensor according to the present invention.

Detailed Description

The present invention will be further illustrated with reference to the following examples, but the present invention is not limited to the following examples.

EXAMPLE 1 construction method of light-emitting sensor

Step one, preparation of carbon quantum dots

1g of 5,10,15, 20-tetrakis (1-methyl-4-pyridine) porphyrin and 2g of sodium citrate were weighed out and dissolved in 25mL of deionized water, and reacted at 200 ℃ for 6 hours.

Step two, preparation of hydrotalcite nano material loaded with carbon quantum dots and luminol simultaneously

And (2) weighing 1mg of the carbon quantum dots prepared in the step (1), 20g of hydrotalcite and 20mg of luminol, adding into 1L of deionized water, stirring for 1 hour at 25 ℃, centrifuging, removing a supernatant, and washing with deionized water for three times for later use.

Obtaining of relationship between luminous intensity and salicylic acid concentration under different salicylic acid concentrations

Mixing 100 mu L of hydrotalcite nano material simultaneously loaded with carbon quantum dots and luminol and 100 mu L of salicylic acid solution with different concentrations in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier; then 100 mul of hydrogen peroxide is injected into the quartz glass bottle, and the luminous intensity under different salicylic acid concentrations is obtained, and the luminous intensity graph of the luminous sensor under different salicylic acid concentrations is shown in figure 3.

Investigating the chemiluminescence intensity of a hydrotalcite nano material-hydrogen peroxide system loaded with carbon quantum dots and luminol:

mixing 100 mu L of hydrotalcite nano material loaded with carbon quantum dots and luminol and 100 mu L of deionized water in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier; and injecting 100 mu L of hydrogen peroxide into the quartz bottle by using a micro-injector to obtain the luminous intensity of the hydrotalcite nano material-hydrogen peroxide system loaded with the carbon quantum dots and the luminol.

Mixing 100 mu L of luminol solution with the concentration of 40mg/L and 100 mu L of deionized water in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier; and injecting 100 mu L of hydrogen peroxide into the quartz bottle by using a micro-injector to obtain the luminous intensity of the luminol-hydrogen peroxide system.

A graph comparing the luminous intensity of the luminol-hydrogen peroxide system with the luminous intensity of the hydrotalcite nanomaterial-hydrogen peroxide system loaded with carbon quantum dots and luminol is shown in fig. 2.

The stability test of the light-emitting sensor obtained by the construction method:

mixing 100 mu L of hydrotalcite nano material simultaneously loaded with carbon quantum dots and luminol and 100 mu L of salicylic acid solution with the concentration of 20 mu g/L in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier; then 100 mul hydrogen peroxide is injected into the quartz bottle to obtain the luminous intensity under the salicylic acid concentration, the chemiluminescence signals of the salicylic acid system with the concentration are continuously and repeatedly measured, and the stability test chart of the constructed sensor is obtained and is shown in figure 4.

EXAMPLE 2 determination of salicylic acid content in Soybean seedling root samples

(1) Weighing 1g of soybean seedling root sample, adding 5mL of methanol, carrying out ultrasonic treatment for 5min by using an ultrasonic cell disruption instrument, centrifuging, taking supernatant, drying by using nitrogen, adding 1mL of deionized water to prepare a solution to be detected, diluting the solution to be detected by 5 times by using the deionized water, and preparing salicylic acid solutions with the concentrations of 10 mu g/L, 50 mu g/L, 100 mu g/L, 150 mu g/L and 200 mu g/L.

(2) Mixing 100 mu L of hydrotalcite nano material loaded with carbon quantum dots and luminol simultaneously with 100 mu L of salicylic acid solution obtained in the step (1) in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier;

(3) and (3) injecting 100 mu L of or hydrogen oxide into the quartz bottle by using a micro-injector to obtain a chemiluminescent signal of the salicylic acid solution sample.

(4) And (5) carrying out quantitative determination by adopting a standard addition method to obtain the content of the salicylic acid in the plant sample.

The salicylic acid content in the soybean seedling root sample was found to be 234. mu.g/L with a recovery rate of 99%.

Example 3 determination of salicylic acid content in Rice leaf samples

(1) Weighing 1g of rice leaf sample, adding 5mL of methanol, carrying out ultrasonic treatment for 5min by using an ultrasonic cell disruption instrument, centrifuging, taking supernatant, drying by using nitrogen, adding 1mL of deionized water to prepare a solution to be detected, diluting the solution to be detected by 5 times by using the deionized water, and preparing salicylic acid solution with the concentration of 10, 50, 100, 150 and 200 mu g/L.

(2) Mixing 100 mu L of hydrotalcite nano material loaded with carbon quantum dots and luminol simultaneously with 100 mu L of salicylic acid solution obtained in the step (1) in a quartz bottle, and placing the quartz bottle above a detection window of a photomultiplier;

(3) and (3) injecting 100 mu L of or hydrogen oxide into the quartz bottle by using a micro-injector to obtain a chemiluminescent signal of the salicylic acid solution sample.

(4) And (5) carrying out quantitative determination by adopting a standard addition method to obtain the content of the salicylic acid in the plant sample.

The salicylic acid in the rice leaf sample was found to be 365 μ g/L with a recovery of 102%.