CN114350352A - Novel carbon materials based on coffee beans and methods for detecting lead ions and PPi - Google Patents

Abstract

The invention discloses a novel carbon material based on coffee beans and a method for detecting lead ions and PPi, belonging to the nanometer carbon material technology and the application field of fluorescence detection. The method comprises the following steps: (1) dissolving coffee bean powder and sodium hydroxide in water to obtain solution A; (2) hydrothermally reacting solution A at high temperature to obtain solution B; (3) dialysis of solution B to obtain solution C; ( 4) After the solution C was filtered and dried, CQDs were obtained. In the present invention, the carbon quantum dots synthesized by one-step hydrothermal method are used as fluorescent probes. The fluorescent probes can detect Pb2+ with high sensitivity through the fluorescence quenching effect, and then detect PPi through the fluorescence recovery phenomenon. The synthesis method is green and simple, and the detection is fast and convenient. , The advantages of accurate results have high application value in the field of biomedicine.

Description

Novel carbon materials based on coffee beans and methods for detecting lead ions and PPi

technical field

The invention belongs to the nanoscale carbon material technology and the application field of fluorescence detection thereof, and particularly relates to a novel carbon material based on coffee beans and a method for detecting lead ions and PPi.

Background technique

Heavy metal pollution has seriously threatened the environment and human health. How to effectively monitor the concentration of toxic heavy metal ions in the environment has become an increasingly important issue. Lead is one of the most common toxic heavy metals, widely used in pigments, water pipes, batteries, anti-corrosion coatings, alloys and other fields, and widely exists in various fields of the environment. Meanwhile, due to the non-biodegradability of Pb ²⁺ , Pb ²⁺ easily accumulates in the human nervous and cardiovascular systems when exposed to polluted air and water sources. When the concentration of Pb ²⁺ in the blood is higher than 5μM, it can lead to anemia, reproductive dysfunction, nervous system dysfunction and developmental disorders, etc., and excessive Pb ²⁺ concentration can even lead to death. Therefore, it is necessary to establish a rapid and sensitive method for the determination of trace Pb ²⁺ .

Anions are ubiquitous throughout biological systems and play important roles in a wide range of chemical and biological processes. As an anion, pyrophosphate (PPi) is released when adenosine triphosphate is hydrolyzed to adenosine monophosphate under cellular conditions. It is part of the bioenergy cycle and DNA synthesis and is an inhibitor of several crystallization reactions. Therefore, many efforts have been devoted to developing non-biosensors for PPi in recent years, and these works have become important in medicine, biology, environment, and nutrition.

There are many traditional methods for the determination of Pb ²⁺ , such as atomic absorption spectrometry (AAS), anodic stripping voltammetry (ASV), inductively coupled plasma atomic emission spectrometry (ICP-AES) and inductively coupled plasma mass spectrometry (ICP- MS). However, these methods often require expensive and complex instrumentation, time-consuming and complicated sample pretreatment procedures. On the other hand, among the numerous PPi sensors reported so far, only a few examples are fluorescent sensors that can effectively detect PPi in 100% PPi aqueous solution. Most reports have focused on their behavior in non-aqueous solvents, and these methods are mostly based on the synthesis of organic small molecule probes, which are complex and time-consuming and require organic solvents as reaction media.

In recent years, carbon quantum dots (CQDs) have attracted great interest from researchers as a new type of fluorescent nanoparticles. Compared with organic dyes, quantum dots and noble metal nanoparticles, CQDs have excellent water solubility, good photostability, and superior biological phase. Capacitance and low toxicity, which make carbon quantum dots effective for practical applications in chemical sensors and biosensors. According to the literature survey, using different raw materials as carbon sources (such as citric acid, leaves, lemons, etc.) to prepare CQDs by hydrothermal method has the advantages of simplicity, easy operation and low cost, and has become one of the most commonly used methods for preparing CQDs. , the surface of synthesized CQDs usually has hydroxyl, carboxyl or amino groups, which can coordinate with metal ions and heavy metal ions to cause fluorescence changes.

Based on the urgent need for the preparation of novel carbon quantum dots, the present application uses coffee beans as raw materials, interacts with a certain concentration of alkali (NaOH), and synthesizes carbon quantum dots by one-step hydrothermal method, and uses them as fluorescent probes for rapid detection. Pb ²⁺ and PPi.

SUMMARY OF THE INVENTION

The object of the present invention is to provide a novel carbon material based on coffee beans and a method for detecting Pb ²⁺ and PPi, specifically a method for detecting Pb ²⁺ and PPi based on fluorescence "on-off" effect, using coffee beans as raw materials, The carbon quantum dots synthesized by one-step hydrothermal method are used as fluorescent probes. The fluorescent probes can detect Pb ²⁺ with high sensitivity through the fluorescence quenching effect, and then detect PPi through the fluorescence recovery phenomenon. The synthesis method is green and simple, and the detection is fast and convenient. , The advantages of accurate results overcome the problems of high equipment cost and complicated operation of traditional lead ion and pyrophosphate detection methods.

In order to realize the above-mentioned purpose of the invention, the technical scheme of the present invention is as follows:

In one aspect, the present invention provides a method for preparing a novel carbon material based on coffee beans, comprising the following steps:

(1) coffee bean powder and sodium hydroxide are dissolved in water to obtain solution A;

(2) solution A reacts at high temperature to obtain solution B;

(3) solution B is dialyzed to obtain solution C;

(4) After the solution C was filtered and dried, CQDs were obtained.

Preferably, in step (1), the coffee bean powder is obtained by pulverizing coffee beans, and the pulverizing methods include but are not limited to grinding, grinding, ultra-fine pulverizing, jet pulverizing, and the like.

Preferably, in step (1), the mass ratio of the coffee bean powder and sodium hydroxide is 2-4:7.2-16.8.

In this reaction, sodium hydroxide solution acts as a catalyst and a decomposer, which is crucial for the preparation of the final product. Sodium hydroxide can not only degrade the macromolecules (such as protein, cellulose, etc.) in coffee beans under alkaline conditions, thereby providing raw materials for the formation of carbon quantum dots, but also participate in a series of chemical reactions in which carbon sources generate quantum. This process is that the polymers (polysaccharides, proteins, etc.) contained in coffee can be catalytically decomposed to form amines and carboxylic acid small carbon-containing compounds under high temperature conditions containing sodium hydroxide, and the obtained small molecular carbon-containing substances are in The reaction occurs under high temperature and high pressure conditions to form carbon quantum dots.

Preferably, in step (1), the mass ratio of the coffee bean powder to water is 1:50-200.

Preferably, in step (1), the water is pure water.

Preferably, in step (1), it is fully dissolved by ultrasound.

Preferably, in step (2), the temperature of the reaction is 150-200° C., and the reaction time is 240-480 min.

Preferably, in step (2), the reaction is carried out in a polytetrafluoroethylene lining in the stainless steel reaction kettle.

Preferably, in step (3), the dialysis is performed in a dialysis bag of pure water and 1000-12,000 D, and the dialysis time is 24-72 hours.

Preferably, in step (4), the drying method is freeze-drying.

In yet another aspect, the present invention provides a novel carbon material based on coffee beans prepared according to the above preparation method.

In yet another aspect, the present invention provides the application of the above-mentioned novel carbon material based on coffee beans in the detection of Pb ²⁺ and PPi.

In another aspect, the present invention provides a method for rapidly detecting Pb ²⁺ , comprising the following steps:

S1, the CQDs obtained by the above preparation method are dissolved in ultrapure water to obtain solution D, and the fluorescence intensity of solution D and ultrapure water is mixed and tested to be F ₀ ;

S2. Mix different concentrations of Pb ²⁺ solutions with solution D respectively to obtain a mixed solution, perform a fluorescence test on the mixed solution, and obtain the fluorescence intensity value F of each mixed solution;

Taking the concentration of Pb ²⁺ as the abscissa and the fluorescence quenching rate, namely 1-F/F ₀ as the ordinate, perform linear fitting to obtain the regression equation y ₁ =k ₁ x ₁ +b ₁ , where y ₁ is the fluorescence quenching rate, x ₁ is the Pb ²⁺ concentration, k ₁ is the slope, and b ₁ is the intercept;

S3. Mix the sample to be tested with solution D to obtain a mixed solution; test the fluorescence intensity value of the obtained mixed solution to obtain the fluorescence quenching rate, and substitute it into the linear regression equation y ₁ =k ₁ x ₁ +b ₁ in step S2, The concentration of Pb ²⁺ was obtained.

Preferably, in step S1, the mass concentration of the solution D is 5-10 mg/ml, and the mixing ratio of the solution D and the ultrapure water is 1:39.

Preferably, in step S2, the concentration range of the Pb ²⁺ solution is 12-260 μM.

Preferably, in step S2, the Pb ²⁺ solution is mixed with the solution D in a ratio of 39:1, respectively.

Preferably, in step S3, the mass ratio when the sample to be tested and the solution D are mixed is 29-49:1, most preferably 39:1.

Finally, the present invention provides a method for rapidly detecting PPi, comprising the following steps:

S1. The CQDs obtained by the above preparation method are dissolved in ultrapure water to obtain solution D.

S2. Mix the Pb ²⁺ solution with the solution D to obtain a mixed solution E, carry out a fluorescence test on the mixed solution E, and obtain the fluorescence intensity value F ₀ ; add PPi of different concentrations into the mixed solution E, carry out a fluorescence test, and obtain The fluorescence intensity value F, with the concentration of PPi as the abscissa, and the fluorescence growth rate, namely F/F0-1, as the ordinate, perform linear fitting to obtain the regression equation y ₂ =k ₂ x ₂ +b ₂ , where y ₂ is the fluorescence increase rate, x ₂ is the concentration of PPi, k ₂ is the slope, and b ₂ is the intercept;

S3. Mix the sample to be tested with Pb ²⁺ solution and solution D to obtain a mixed solution, test the fluorescence intensity value of the obtained mixed solution, and substitute it into the linear regression equation y ₂ =k ₂ x ₂ +b ₂ in step S2 to obtain PPi concentration.

Preferably, in step S1, the mass concentration of the solution D is 5-10 mg/ml.

Preferably, in step S2, the concentration of the Pb ²⁺ solution is 12-260 μM.

Preferably, in step S2, the PPi concentration range of the different concentrations is 37.5-125 μM.

Preferably, in step S2, the Pb ²⁺ solution and the solution D are mixed in a ratio of 39:1.

Preferably, in step S3, the sample to be tested is mixed with the Pb ²⁺ solution and the solution D in a ratio of 38:1:1.

Preferably, in step S3, the Pb ²⁺ concentration is 10 mM.

The beneficial effects of the present invention are:

1. The present invention is used for the detection scheme of Pb ²⁺ and PPi content based on the "on-off" fluorescence effect of the carbon quantum dots, which has the advantages of high sensitivity, good selectivity, less spectral interference, and low cost, and the carbon quantum dots The point is synthesized from coffee beans, which is not only green and environmentally friendly, but also has good biocompatibility and has the potential for bioimaging.

2. The carbon quantum dots prepared by the technical solution of the present invention have the advantages of superior kinetic and thermodynamic properties, physical and chemical stability, good water solubility, good biocompatibility, little environmental harm, and high sensitivity to lead ions.

3. In the present invention, coffee beans are used as raw materials, and carbon quantum dots synthesized by one-step hydrothermal method are used as fluorescent probes, and the synthesis is green and simple.

Description of drawings

The comparison diagram of the sensitivity of the product CQDs of Fig. 1 different embodiment and comparative example to detect Pb ²⁺ ;

The FTIR diagram of the product CQDs of Fig. 2 embodiment 1;

Figure 3 is a histogram of the hemolysis rate of CQDs;

Figure 4 is a histogram of the selectivity of CQDs to detect Pb ²⁺ before pretreatment;

Figure 5 is a histogram of the selectivity of CQDs to detect Pb ²⁺ after pretreatment;

Figure 6 is a graph showing the relationship between the change of fluorescence intensity of CQDs and the concentration of Pb ²⁺ ;

Figure 7 is a graph showing the relationship between the intensity change of CQDs fluorescence recovery and the concentration of PPi;

Figure 8 is a bar graph of the selectivity of CQDs to detect PPi.

Detailed ways

The following non-limiting examples may enable those of ordinary skill in the art to more fully understand the present invention, but do not limit the present invention in any way. The following content is only an exemplary description of the scope of protection claimed in the application, and those skilled in the art can make various changes and modifications to the invention of the application according to the disclosed content, and it should also belong to the scope of protection claimed in the application. .

The present invention will be further described below by way of specific embodiments. Various chemical reagents used in the examples of the present invention are obtained through conventional commercial channels unless otherwise specified.

In the following examples, coffee bean powder is obtained by grinding coffee beans.

Example 1

1) Dissolve 3g of coffee bean powder in 30mL of pure water, then add 7.2g (6mM) of sodium hydroxide to obtain mixed solution A;

2) Transfer the solution A to the polytetrafluoroethylene lining, then load it into the stainless steel reactor and react for 480 min at 200° C. to obtain the solution B;

3) Transfer solution B to a 12000D dialysis bag, and dialyze for 48h to obtain solution C;

4) The solution C was filtered through a 0.22 μm organic filter membrane, then frozen at -60°C for 4 hours, and vacuum-dried for 12 hours to obtain brown-black powdery CQDs, which were stored at 2-8°C for later use.

Example 2

The difference from Example 1 is that 2 g of coffee bean powder was added, and the rest were the same.

Example 3

The preparation method of carbon quantum dots changes the amount of sodium hydroxide with reference to example 1, and the concrete steps are as follows:

The difference from Example 1 is that 12 g of sodium hydroxide was added, and the rest were the same.

Example 4

The difference from Example 1 is that solution A was transferred to a polytetrafluoroethylene liner, and then loaded into a stainless steel reactor to react at 150° C. for 480 min to obtain solution B, and the rest were the same.

Example 5

Different from Example 1, the specification of the dialysis bag is 1000D, and the rest are the same.

Comparative Example 1

The difference from Example 1 is that no sodium hydroxide is added, and the rest are the same.

Sensitivity test:

The product CQDs in the above examples and comparative examples were all prepared into 10 mg/mL D solution, the D solution and pure water were mixed in a ratio of 1:39, and the initial fluorescence intensity of the test was recorded as F ₀ , and then the D solution and 250 μM The Pb ²⁺ solution was mixed at a ratio of 1:39, and the fluorescence intensity after quenching was measured, denoted as F. Taking the value of F/F ₀ as the detection index of sensitivity (Fig. 1), that is, the lower the value of F/F ₀ , the higher the degree of quenching and the higher the sensitivity. It can be seen that the implementation effect of the CQDs prepared in Comparative Example 1 is poor. Comprehensive results The product CQDs of Example 1 was used as the detection material of the subsequent detection example 1 and detection example 2.

Selectivity test for detection of Pb ²⁺ by CQDs:

The final concentration of various metal ions involved in the experiment is designed to be 250 μM for selective testing. It can be seen that Cu ²⁺ , Hg ²⁺ , Fe ³⁺ interfere with the detection of Pb ²⁺ , but glutathione and Ascorbic acid was used as a chelating agent and a reducing agent to treat Cu ²⁺ , Hg ²⁺ , Fe ³⁺ and the interference disappeared. The final concentration of metal ions was 50 μM, the final concentration of glutathione was 0.08 mg/mL, and the final concentration of ascorbic acid was 0.5 mg/mL. Figures 4-5 show that the detection of Pb ²⁺ by CQDs is stable and unaffected, with high selectivity.

Test Example 1

Utilize the CQDs obtained in the above-mentioned embodiment 1 of the present invention to detect lead ion content by fluorescence spectroscopy, and the method steps are as follows:

1) Dissolve the powdered CQDs in ultrapure water, configure it into a 10 mg/mL D solution, mix the D solution and pure water in a ratio of 1:39, and test its fluorescence intensity as 10100.5, denoted as F ₀ ;

2) Mix Pb ²⁺ solutions with concentrations of 12, 35, 55, 86, 110, 130, 170, 210, and 260 μM with D solution in a ratio of 39:1, respectively, to obtain a mixed solution, and perform a fluorescence test on the mixed solution , the fluorescence intensity values obtained for each mixed solution are 9578.3, 8856.2, 8415.5, 8171.5, 6977.9, 6860.2, 5649.7, 4660.6, 3270.5, denoted as F;

Taking the concentration of Pb ²⁺ as the abscissa and the fluorescence quenching rate (1-F/F ₀ ) as the ordinate, perform linear fitting to obtain the regression equation y ₁ =0.00241x ₁ -0.0321, where y ₁ is the fluorescence quenching rate , x ₁ is the concentration of Pb ²⁺ ;

3) Mix the lake water sample containing a certain concentration of lead ions (50 μM) and the D solution according to the ratio of 39:1 to obtain a mixed solution; carry out the fluorescence test, and substitute the obtained fluorescence intensity value into the linear regression equation y ₁ =0.00241× ₁ -0.0321, the average concentration of lead ions was calculated to be 51.7 μM (n=3), and the average recovery rate was 103.4% (n=3). The excitation wavelength of fluorescence in the fluorescence assay used was 370 nm.

Test example 2

Utilize the CQDs obtained in the above-mentioned embodiment 1 of the present invention, and then use fluorescence spectroscopy to detect the PPi content, and the method steps are as follows:

1) Dissolve the powdered CQDs in ultrapure water to prepare a 10 mg/mL D solution;

2) Mix the 250 μM Pb ²⁺ solution with the D solution in a ratio of 39:1 to obtain a mixed solution E, carry out a fluorescence test on the mixed solution E, and the obtained fluorescence intensity value is 2070.2, denoted as F ₀ ; , 62.5, 75, 87.5, 100, 112.5, 125 μM of PPi solution and solution E were mixed in a ratio of 1:39, and the fluorescence intensity was measured. , 5679.2, denoted as F, take the PPi concentration as the abscissa and the fluorescence growth rate (F/F ₀ -1) as the ordinate, perform linear fitting to obtain the regression equation y ₂ =0.02051x ₂ +0.7250, where y ₂ is Fluorescence growth rate, x ₂ is the concentration of PPi;

3) Mix the PPi-containing solution (50 μM) to be tested with Pb ²⁺ solution (10 mM) and solution D in a ratio of 38:1:1 to obtain a PPi-Pb-CQDs mixed solution; perform a fluorescence test to obtain the fluorescence intensity value, substituted into the linear regression equation y ₂ =0.02051× ₂ +0.7250, the average concentration of PPi was calculated to be 50.6 μM, and the average recovery rate was 101.2%.

It can be seen that the test results of the method of the present invention are accurate and reliable, and the test steps are convenient and fast.

The above descriptions are only preferred embodiments of the present invention, and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included in the scope of the present invention. within the scope of protection.

Claims (10)

1. A preparation method of a novel carbon material based on coffee beans is characterized by comprising the following steps:

(1) dissolving coffee bean powder and sodium hydroxide in water to obtain a solution A;

(2) reacting the solution A at a high temperature to obtain a solution B;

(3) dialyzing the solution B to obtain a solution C;

(4) filtering and drying the solution C to obtain CQDs.

2. The production method according to claim 1, wherein in the step (1), the mass ratio of the coffee bean powder to the sodium hydroxide is 2-4:7.2-16.8, and the mass ratio of the coffee bean powder to the water is 1: 50-200.

3. The method as claimed in claim 1, wherein the step (2) is performed in a polytetrafluoroethylene lining in the reaction stainless steel reactor, the temperature of the reaction is 150 ℃ and 200 ℃, and the reaction time is 240 ℃ and 480 min.

4. The method as claimed in claim 1, wherein the dialysis in step (3) is carried out in a dialysis bag of pure water, 1000-12000D for a dialysis time of 24-72 hours.

5. A novel coffee bean-based carbon material produced by the production method according to any one of claims 1 to 4.

6. Use of the novel coffee bean-based carbon material according to claim 5 for detecting Pb²⁺And use in PPi.

7. Rapid detection Pb²⁺The method is characterized by comprising the following steps:

s1, dissolving CQDs prepared by the preparation method of any one of claims 1 to 4 in ultrapure water to obtain solution D, mixing the solution D and ultrapure water, and testing the fluorescence intensity of the solution D as F₀；

S2, mixing Pb with different concentrations²⁺Respectively mixing the solution with the solution D to obtain mixed solutions, and performing fluorescence test on the mixed solutions to respectively obtain fluorescence intensity values F of the mixed solutions;

with Pb²⁺Concentration is abscissa, and fluorescence quenching rate is 1-F/F₀Linear fitting is performed for the ordinate to obtain a regression equation y₁＝k₁x₁+b₁Wherein y is₁As fluorescence quenching rate, x₁Is Pb²⁺Concentration, k₁The value is the slope, b₁The value is the intercept;

s3, mixing the sample to be detected with the solution D to obtain a mixed solution; testing the fluorescence intensity value of the obtained mixed solution to further obtain the fluorescence quenching rate, and substituting the fluorescence quenching rate into the linear regression equation y in the step S2₁＝k₁x₁+b₁Obtaining Pb²⁺The concentration of (c).

8. The method according to claim 7, wherein in the step S1, the mass concentration of the solution D is 5-10 mg/ml, and the mixing ratio of the solution D and the ultrapure water is 1: 39; in step S2, Pb is²⁺The concentration range of the solution is 12-260 mu M; in step S2, Pb²⁺Mixing the solution with the solution D at a ratio of 39: 1; in step S3, the sample and the solution to be testedD, mass ratio of the mixed materials is 29-49: 1.

9. a method for rapidly detecting PPi is characterized by comprising the following steps:

s1, dissolving CQDs prepared by the preparation method of any one of claims 1 to 4 in ultrapure water to obtain solution D;

s2, adding Pb²⁺Mixing the solution with the solution D to obtain a mixed solution E, and performing fluorescence test on the mixed solution E to obtain a fluorescence intensity value F₀(ii) a Adding PPi with different concentrations into the mixed solution E, performing fluorescence test to obtain fluorescence intensity value F, and taking the concentration of PPi as abscissa, wherein the fluorescence growth rate is F/F₀-1 is a vertical coordinate, linear fitting is carried out to obtain a regression equation y₂＝k₂x₂+b₂Wherein y is₂As the rate of increase in fluorescence, x₂Is the concentration of PPi, k₂The value is the slope, b₂The value is the intercept;

s3, mixing the sample to be tested with Pb²⁺Mixing the solution and the solution D to obtain a mixed solution, testing the fluorescence intensity value of the mixed solution, and substituting the fluorescence intensity value into the linear regression equation y in the step S2₂＝k₂x₂+b₂The concentration of PPi was obtained.

10. The method according to claim 9, wherein in step S1, the mass concentration of solution D is 5-10 mg/ml; in step S2, Pb is²⁺The concentration of the solution is 12-260 mu M; in step S2, the concentration range of the PPi with different concentrations is 37.5-125 μ M; in step S2, Pb²⁺Mixing the solution with the solution D at a ratio of 39: 1; in step S3, the sample to be tested and Pb²⁺Mixing the solution and the solution D according to the ratio of 38:1: 1; in step S3, Pb is²⁺Is 10 mM.

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