CN114034692A - Formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chain and application - Google Patents
Formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chain and application Download PDFInfo
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Abstract
The invention discloses a formaldehyde colorimetric detection method based on metal nanoparticles and a triple helix chain and application thereof, wherein the formaldehyde colorimetric detection method comprises the following steps: (1) modifying an Oligo 1 probe sequence on the surface of the metal nanoparticle to obtain a metal nanoparticle probe; (2) and (3) mixing the sample to be detected with the metal nanoparticle probe, the Oligo 2 sequence, the Oligo 3 sequence and the silver ion solution prepared in the step (1), and quantifying the formaldehyde content in the sample to be detected according to the RGB value of the final color of the mixed solution. The detection method is simple, rapid, sensitive and reliable, does not need large instruments and complex operation processes, can realize field detection in extremely short time, and has strong specificity, high sensitivity and detection limit of only 0.14 mg.L‑1Has extremely high application value.
Description
Technical Field
The invention relates to the field of molecular detection, in particular to a formaldehyde colorimetric detection method based on metal nanoparticles and a triple helix chain and application thereof.
Background
The RGB color scheme is a color standard in the industry, which obtains various colors by changing three color channels of red (R), green (G) and blue (B) and superimposing them on each other, and the standard almost includes all colors that can be perceived by human vision, and is one of the most widely used color systems at present. The three color channels red, green and blue are each divided into 256 levels of brightness, darkest at level 0 and brightest at level 255. However, the application of the method in the field of biomolecule detection has not been reported.
Formaldehyde is the most common one of environmental pollutants, and has great harm to people. The main hazard of formaldehyde is its toxic effect, and excessive exposure to formaldehyde can cause respiratory, digestive, circulatory and nervous systems to be damaged to varying degrees. Furthermore, formaldehyde was identified in 2004 by the international agency for research on cancer (IARC) as a class 1 carcinogen, and long term exposure to formaldehyde carries a carcinogenic risk. In daily life, the vegetables are preserved by illegal use of formaldehyde by bad vendors, certain formaldehyde residues are brought, and once the residual formaldehyde enters a human body, the residual formaldehyde brings great harm to the human health. And other daily activities, such as house decoration, can also cause a large amount of formaldehyde residues, and are also easy to harm human health.
In the related technology, the traditional detection methods such as ultraviolet-visible absorption spectroscopy, fluorescence spectroscopy, Raman spectroscopy and the like are mainly used for detecting the formaldehyde residue, but the methods have the problems of long detection time, high detection cost, incapability of being used for on-site temporary detection and the like, so that the application range of the methods is greatly limited.
Compared with the currently reported several methods for detecting formaldehyde, such as fluorescence method, electrochemical method, chromatography method, chemiluminescence method and other instrument methods for detecting formaldehyde, the methods still have the defects of expensive equipment, complex operation steps, low detection limit and the like. Therefore, it is still necessary to explore new formaldehyde detection strategies.
Therefore, in view of the huge and ubiquitous hazard of formaldehyde, the development of a simple, rapid, sensitive and reliable formaldehyde on-site monitoring method is of great significance.
Disclosure of Invention
The present invention is directed to solving at least one of the problems of the prior art described above. Therefore, the invention provides a formaldehyde colorimetric detection method based on metal nanoparticles and a triple helix chain, which is simple to operate, short in detection time, capable of accurately determining the formaldehyde content in a sample to be detected based on a mobile phone camera, high in sensitivity and good in specificity, and capable of realizing field monitoring of formaldehyde.
In a first aspect of the present invention, there is provided an RGB colorimetric method for detecting formaldehyde, comprising the steps of:
(1) modifying an Oligo 1 probe sequence on the surface of the metal nanoparticle to obtain a metal nanoparticle probe;
(2) and (3) mixing the sample to be detected with the metal nanoparticle probe, the Oligo 2 sequence, the Oligo 3 sequence and the silver ion solution prepared in the step (1), and quantifying the formaldehyde content in the sample to be detected according to the RGB value of the final color of the mixed solution.
According to the first aspect of the present invention, in some embodiments of the present invention, the Oligo 1 probe sequence is an oligonucleotide sequence.
In some preferred embodiments of the present invention, the Oligo 1 probe sequence comprises 5-1000 homopolymeric pyrimidine sequences; the homopolypyrimidine sequence contains 5-1000 cytosines.
In some preferred embodiments of the present invention, the Oligo 1 probe sequence has a linking group attached to the 5 'or 3' end thereof for linking the Oligo 1 probe sequence to the surface of the metal nanoparticle.
In some more preferred embodiments of the invention, the linking group is a thiol group.
In some more preferred embodiments of the present invention, the Oligo 1 probe sequence is
Wherein the underlined part is the nucleic acid sequence that is complementary to and mismatched with Oligo 2 sequence.
According to the first aspect of the present invention, in some embodiments of the present invention, the Oligo 2 sequence is an oligonucleotide sequence complementary-paired only with one end of Oligo 1 probe sequence away from gold nanoparticles.
In some preferred embodiments of the present invention, the Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3).
According to the first aspect of the present invention, in some embodiments of the present invention, the Oligo 3 sequence is a homopolypurine sequence complementary-paired only with one end of Oligo 1 probe sequence near the gold nanoparticle.
In some preferred embodiments of the present invention, the Oligo 3 sequence is: 5'-AAACAAAACAAA-3' (SEQ ID NO. 4).
In some preferred embodiments of the invention, the Oligo 2 sequence is not duplicately paired with the Oligo 3 sequence for any segment of the Oligo 1 probe sequence.
According to a first aspect of the present invention, in some embodiments of the present invention, the metal nanoparticles comprise at least one of gold nanoparticles, silver nanoparticles and palladium nanoparticles.
In some preferred embodiments of the present invention, the metal nanoparticles are gold nanoparticles. The gold nanoparticles can be prepared from commercially available gold nanoparticles or by conventional methods in the field, such as a sodium citrate reduction method.
Gold nanoparticles (AuNPs) are used as an important component of the nano material, and the inventor finds that the dispersion/aggregation state of the AuNPs modified by DNA can be obviously changed, and the absorbance, the color and the particle size of the AuNPs are changed accordingly. Therefore, the AuNPs probe can display the amount of formaldehyde as an ideal colorimetric, RGB, and dynamic light scattering signal probe.
In some preferred embodiments of the present invention, the gold nanoparticles are prepared by: 2mL of freshly prepared 38.8mM sodium citrate solution was added rapidly to 20mL of boiled 1mM HAuCl chloroaurate4In solution. At this time, the reaction solution turned from light yellow to black, then to purple, and finally to wine red. And after the color of the mixture turns to wine red, continuously heating and refluxing the mixture, and stirring the mixture for 20 to 60 minutes. And cooling the reaction solution to room temperature (continuously stirring), and filtering with a nylon filter membrane of 0.1-5.0 mu m to obtain the catalyst.
According to a first aspect of the invention, in some embodiments of the invention, the silver ion solution comprises: silver nitrate and Ag (NH)3)2At least one of OH.
In some preferred embodiments of the present invention, the silver ion solution is silver nitrate.
According to a first aspect of the present invention, in some embodiments of the present invention, the RGB colorimetry comprises the specific steps of:
(1) preparing a metal nanoparticle probe:
treating the Oligo 1 probe sequence with TCEP, and then according to the Oligo 1 probe sequence: AuNPs is 180-220: 1, mixing and incubating the Oligo 1 probe sequence treated by TCEP with AuNPs for 1-100 hours, and adding NaCl with the final concentration of 0.01-1.0M for salt aging treatment to obtain the metal nanoparticle probe.
(2) Mixing was carried out according to the following system:
| components | Dosage of |
| Gold nanoparticle probe prepared in step (2) of 30nM | 28μL |
| 0.1. mu.M Oligo 2 sequence | 22μL |
| Oligo 3 sequence at 0.2. mu.M | 10μL |
| 50 μ M silver ion solution | 33μL |
| Formaldehyde sample | 33μL |
| 0.5mM spermine | 12μL |
| PBS buffer (pH 7.0) | 82μL。 |
Among them, spermine in the system is mainly used to stabilize the triple helix structure formed.
(3) The imaging device is used to take a picture of the image and the RGB values of the imaged picture are identified. And (3) realizing quantitative detection of the formaldehyde according to the linear relation between the RGB value and the formaldehyde concentration (by using a formaldehyde standard substance determination standard curve).
In some preferred embodiments of the present invention, the RGB value signal output may be an R value, a G value, a B value, an RGB total value, an RGB average value, or an RGB ratio value (R/G, R/B or G/B).
In some more preferred embodiments of the present invention, the RGB value signal output is preferably an R value.
According to the experiments of the inventor, the change range of the R value of the system is 50-180 when the concentration of the formaldehyde is from small to large.
In some preferred embodiments of the present invention, the imaging device comprises at least one of a smartphone, a camera, a camcorder, a microscope lens, and a camera.
In some preferred embodiments of the present invention, the imaging device is a smartphone.
The mobile phone is fixed by the support, the distance between the camera and the solution is about 12 cm, the light source in the shooting parameters is adjusted to an incandescent lamp mode, and the sample is shot.
The detection principle of the RGB colorimetric method in the invention is shown in the attached drawings of figures 1 and 2 in the specification. The method is mainly based on two points: 1. formation based on triple-stranded DNA: the Oligo 3 sequence designed by the invention can be hybridized with the near AuNPs end single-stranded DNA (the Oligo 1 probe sequence is near the AuNPs end) modified on the gold nanoparticle probe, so that the modified DNA (the Oligo 1 probe sequence + the Oligo 3 sequence) is more three-dimensional, and is not simply adhered to the AuNPs surface. And the designed Oligo 2 sequence can be hybridized with the modified far AuNPs end single-stranded DNA (Oligo 1 probe sequence far AuNPs end) on the gold nano-particle probe. So that the gold nanoparticle probe is respectively combined with the Oligo 2 sequence and the Oligo 3 sequence, and when Ag is added+The gold nanoparticles aggregate (Ag)+The Oligo 1 probe sequence, Oligo 2The sequences and Oligo 3 sequences form a three-stranded DNA structure, thereby aggregating the gold nanoparticles), whereas if formaldehyde is contained in the solution at this time, the aggregated gold nanoparticles are dispersed to different degrees (formaldehyde can make Ag disperse)+Resulting in the unwinding of the triplex DNA structure) and thus the development of distinct colors (from red to magenta or blue), particle size and turbidity changes, and by identifying the RGB values, the formaldehyde content can be quantitatively detected based on the fact that the RGB values are linear with the formaldehyde concentration.
In a second aspect of the invention, a colorimetric formaldehyde detection reagent is provided.
According to the second aspect of the present invention, in some embodiments of the present invention, the colorimetric formaldehyde detection reagent contains at least one of the metal nanoparticles, the Oligo 1 probe sequence, the Oligo 2 sequence and the Oligo 3 sequence in the RGB colorimetric method according to the first aspect of the present invention.
According to the second aspect of the present invention, in some embodiments of the present invention, the Oligo 1 probe sequence is an oligonucleotide sequence.
In some preferred embodiments of the present invention, the Oligo 1 probe sequence comprises 5-1000 homopolymeric pyrimidine sequences; the homopolypyrimidine sequence contains 5-1000 cytosines.
The partially complementary mismatched nucleic acid sequence is complementary to Ag+The specific recognition forms CGC tripolymer oligonucleotide (namely DNA triple helix structure) to lead the gold nanoparticles to be aggregated.
In some preferred embodiments of the present invention, the Oligo 1 probe sequence has a linking group attached to the 5 'or 3' end thereof for linking the Oligo 1 probe sequence to the surface of the metal nanoparticle.
In some more preferred embodiments of the invention, the linking group comprises a sulfhydryl group (S-C)6-)。
In some more preferred embodiments of the present invention, the Oligo 1 probe sequence is
Among them, the underlined part is a sequence partially complementary to the mismatch.
According to the second aspect of the present invention, in some embodiments of the present invention, the Oligo 2 sequence is an oligonucleotide sequence complementary-paired only with one end of Oligo 1 probe sequence away from gold nanoparticles.
In some preferred embodiments of the present invention, the Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3).
According to the second aspect of the present invention, in some embodiments of the present invention, the Oligo 3 sequence is a homopolypurine sequence complementary-paired only with one end of Oligo 1 probe sequence near the gold nanoparticle.
In some preferred embodiments of the present invention, the Oligo 3 sequence is: 5'-AAACAAAACAAA-3' (SEQ ID NO. 4).
In some preferred embodiments of the invention, the Oligo 2 sequence is not duplicately paired with the Oligo 3 sequence for any segment of the Oligo 1 probe sequence.
According to a second aspect of the present invention, in some embodiments of the present invention, the metal nanoparticles comprise at least one of gold nanoparticles, silver nanoparticles and palladium nanoparticles.
In some preferred embodiments of the present invention, the metal nanoparticles are gold nanoparticles. The gold nanoparticles can be prepared from commercially available gold nanoparticles or by conventional methods in the field, such as a sodium citrate reduction method.
In some preferred embodiments of the present invention, the gold nanoparticles are prepared by: 2mL of freshly prepared 38.8mM sodium citrate solution was added rapidly to 20mL of boiled 1mM HAuCl chloroaurate4In solution. At this time, the reaction solution turned from light yellow to black, then to purple, and finally to wine red. And after the color of the mixture turns to wine red, continuously heating and refluxing the mixture, and stirring the mixture for 20 to 60 minutes. And cooling the reaction solution to room temperature (continuously stirring), and filtering with a nylon filter membrane of 0.1-5.0 mu m to obtain the catalyst.
According to a second aspect of the invention, in some embodiments of the invention, theThe silver ion solution comprises: silver nitrate and Ag (NH)3)2At least one of OH.
In some preferred embodiments of the present invention, the silver ion solution is silver nitrate.
In a third aspect of the present invention, there is provided a colorimetric formaldehyde detection device, comprising:
a detection module, wherein the detection module contains the formaldehyde colorimetric detection reagent according to the second aspect of the invention;
an imaging module for solution color recording;
the color identification module is used for identifying the color of the solution recorded by the imaging module and obtaining an RGB value;
and the analysis module is used for calculating the formaldehyde concentration according to the RGB value obtained by the color recognition module.
According to a third aspect of the invention, in some embodiments of the invention, the color identification module is an RGB color detector.
In some preferred embodiments of the present invention, the RGB color detector is a cell phone containing a charge-coupled device image sensor CCD (or complementary metal oxide semiconductor CMOS), or other detector containing an RGB tricolor sensor.
In a fourth aspect of the present invention, there is provided a use of the colorimetric formaldehyde detection reagent according to the second aspect of the present invention or the colorimetric formaldehyde detection device according to the third aspect of the present invention in detection of formaldehyde residues in food and environment.
In some preferred embodiments of the invention, the food product comprises fresh fruit and vegetables.
In some preferred embodiments of the invention, the food product is a green vegetable.
In some preferred embodiments of the invention, the environment includes a work environment and a living environment that are easily contaminated with formaldehyde. Such as home decoration, outdoor paint brushing and the like.
The invention has the beneficial effects that:
1. the detection method is simple, rapid, sensitive and reliable, does not need large instruments and complex operation processes, can realize field detection in extremely short time, and has far higher cost and efficiency than other traditional methods.
2. The detection method of the invention needs few samples and has strong specificity, can effectively eliminate the interference of other structural analogs such as acetaldehyde, methanol, ethanol, acetone, ether, acetic acid, ammonia water, toluene and the like, can accurately detect the content of formaldehyde in a linear range, and has the detection limit of 0.14 mg.L-1And has extremely high detection sensitivity.
Drawings
Fig. 1 is a detection schematic diagram of a colorimetric formaldehyde detection method based on metal nanoparticles and a triple helix chain in an embodiment of the invention.
Fig. 2 is an enlarged view of the sequence in fig. 1 in the description of the present invention.
Fig. 3 is a specific detection result of the colorimetric formaldehyde detection method in the embodiment of the present invention, where the samples are: formaldehyde (HCHO), water (control, Blank), acetaldehyde (CH)3CHO), methanol (CH)3OH), ethanol (CH)3CH2OH), acetone (CH)3COCH3) Diethyl ether (C)2H5OC2H5) Acetic acid (CH3COOH), ammonia monohydrate (NH)3·H2O) and toluene (C)6H5CH3)。
FIG. 4 is a standard curve of a formaldehyde standard solution obtained by the colorimetric detection method for formaldehyde in the example of the present invention.
Fig. 5 is a photograph of a color identifier of the colorimetric formaldehyde detection method according to an embodiment of the present invention.
Fig. 6 is a detection result of the colorimetric formaldehyde detection method according to the embodiment of the present invention for different vegetable samples.
FIG. 7 is a UV-VIS absorption spectrum chart for verifying the accuracy of the formaldehyde colorimetric detection method in the embodiment of the present invention.
FIG. 8 is a particle size distribution diagram of a colorimetric formaldehyde detection method according to an embodiment of the present invention, which is verified by dynamic light scattering measurement, wherein A is a diagram of a formaldehyde concentration of 0.225mg/L, and B is a diagram of a formaldehyde concentration of 4.5 mg/L.
Detailed Description
In order to make the objects, technical solutions and technical effects of the present invention more clear, the present invention will be described in further detail with reference to specific embodiments. It should be understood that the detailed description and specific examples, while indicating the preferred embodiment of the invention, are intended for purposes of illustration only and are not intended to limit the scope of the invention.
The experimental materials and reagents used are, unless otherwise specified, all consumables and reagents which are conventionally available from commercial sources.
Formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chain
The formaldehyde colorimetric detection method based on the metal nanoparticles and the triple helix chain in the embodiment comprises the following steps:
(1) preparing metal nanoparticles:
in this embodiment, the metal nanoparticles used are gold nanoparticles (AuNPs) prepared by an adjusted sodium citrate reduction method, and the specific preparation method is as follows:
2mL of freshly prepared 38.8mM sodium citrate solution was added rapidly to 20mL of boiled 1mM HAuCl chloroaurate4In solution. At this time, the reaction solution turned from light yellow to black, then to purple, and finally to wine red. And after the color of the mixture turns to wine red, continuously heating and refluxing the mixture, and stirring the mixture for 20 to 60 minutes. And cooling the reaction solution to room temperature (continuously stirring), filtering with a nylon filter membrane of 0.1-5.0 mu m to obtain gold nanoparticles, and storing at 4 ℃ for later use.
Of course, commercially available gold nanoparticles, such as, for example, can also be used.
(2) Preparing a metal nanoparticle probe:
and (3) taking the gold nanoparticles prepared in the step (1), and modifying an Oligo 1 probe sequence on the surface of the gold nanoparticles to obtain the metal nanoparticle probe.
The method comprises the following specific steps:
selection of oligo 1 probe sequence:
the Oligo 1 probe sequence is an oligodeoxyribonucleotide sequence, the sequence contains 5-1000 homopolypyrimidine sequences, the homopolypyrimidine sequences contain 5-1000 cytosines (C), and the complementary and mismatched sequences of the Oligo 1 probe sequence and the Oligo 2 sequence are as follows: 5'-TCTTCTTTCCTTTCTTCT-3' (SEQ ID NO. 1). The oligodeoxyribonucleotide sequence has a thiol group attached to the 5 'or 3' end.
In this example, the Oligo 1 probe sequence is:
wherein, the underlined part is the sequence of the partial nucleic acid complementary mismatch.
b. Modification of gold nanoparticles:
(3) Sample detection:
appropriate amounts of samples were taken and mixed according to the system in table 1.
TABLE 1 detection System
| Components | Dosage of |
| Gold nanoparticle probe prepared in step (2) of 30nM | 28μL |
| 0.1. mu.M Oligo 2 | 22μL |
| Oligo | |
| 3 sequence at 0.2. mu.M | 10μL |
| 50 μ M silver ion solution | 33μL |
| Formaldehyde sample | 33μL |
| 0.5mM spermine | 12μL |
| PBS buffer (pH 7.0) | 82μL |
Wherein the total amount of the detection system is 220 mu L.
Oligo 2 sequence is an oligonucleotide sequence complementary to Oligo 1 probe, and is complementary to only one end of Oligo 1 probe sequence far away from gold nanoparticle, in this example, Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3).
The Oligo 3 sequence is a homopolypurine sequence, and is complementary paired with only one end of the Oligo 1 probe sequence close to the gold nanoparticle, in this embodiment, the Oligo 3 sequence is: 5'-AAACAAAACAAA-3' (SEQ ID NO. 4).
The Oligo 2 sequence does not repeatedly pair with Oligo 3 sequence any fragment in the Oligo 1 probe sequence.
In this example, the silver ion solution is a silver nitrate solution. Of course, other silver ion solutions that do not interfere with the results of the test may be substituted by those skilled in the art.
The system was allowed to stand at room temperature for 2min after mixing, and the color change of the solution was observed.
This is photographed using an imaging device, and the RGB values (R values in this embodiment) of the imaged photograph are recognized. And (3) realizing quantitative detection of the formaldehyde according to the linear relation between the RGB value and the formaldehyde concentration (by using a formaldehyde standard substance determination standard curve).
In this embodiment, the imaging device uses a mobile phone camera, the mobile phone is fixed by a support, the camera is spaced from the solution by about 12 cm, the light source in the shooting parameters is adjusted to the incandescent lamp mode, and the sample is photographed. And reading the RGB value of the photo by using a color recognizer, and realizing quantitative detection of the formaldehyde according to the linear relation between the RGB value and the formaldehyde concentration.
Detection effect of formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chain
(1) Quantitative detection effect (specificity):
in order to investigate the detection specificity of the above colorimetric formaldehyde detection method based on metal nanoparticles and triple helix chains, the inventors used different compounds as detection samples (each at a concentration of 4.5mg/L) for detection.
In this example, formaldehyde (HCHO), water (control, Blank), acetaldehyde (CH) were used respectively3CHO), methanol (CH)3OH), ethanol (CH)3CH2OH), acetone (CH)3COCH3) Diethyl ether (C)2H5OC2H5) Acetic acid (CH3COOH), ammonia monohydrate (NH)3·H2O) and toluene (C)6H5CH3) As a test sample.
The test method was the same as in the above example.
The results are shown in FIG. 3.
As can be seen from FIG. 3, the detection method in the embodiment of the present invention has different responses to different interferents, but only has a specific response to formaldehyde. The concrete expression is as follows: the detection method in the embodiment of the invention generates R values for the interferent with similar structure to formaldehyde, such as acetaldehyde, methanol, ethanol, acetone, diethyl ether, acetic acid, ammonia water and toluene, which are similar to the RGB values of the control sample (water), thereby indicating that the detection method in the embodiment of the invention does not generate specific response to the structural analogues, but only the RGB values of formaldehyde are increased sharply, thereby indicating that the detection method in the embodiment has specific response to formaldehyde.
(2) Detection sensitivity:
in order to investigate the detection sensitivity of the above colorimetric formaldehyde detection method based on metal nanoparticles and triple helix chains, the inventors performed tests using formaldehyde standard solutions of different concentrations (0.225mg/L, 0.45mg/L, 0.9mg/L, 1.35mg/L, 3.6mg/L, 4.5 mg/L).
The test method was the same as in the above example.
The results are shown in FIGS. 4 and 5.
FIG. 4 is a standard curve constructed based on different concentrations (0.225mg/L, 0.45mg/L, 0.9mg/L, 1.35mg/L, 3.6mg/L, 4.5mg/L) of formaldehyde standard solution and RGB values obtained by detection, wherein the constructed standard curve is as follows:
y＝17.50x+78.67；
wherein R is20.995, and y is the detected RGB value; and x is the concentration of formaldehyde. The linear range is 0.23 mg.L-1-4.5mg·L-1The detection limit is 0.14 mg.L-1。
Fig. 5 shows that the concentration of the formaldehyde solution can be accurately obtained by analyzing RGB values of formaldehyde samples with different concentrations by using a mobile phone color identifier based on the formaldehyde colorimetric detection method in the above embodiment.
From the above results, it was found that the detection method in the above examples is excellent in the detection effect on formaldehyde, high in sensitivity, and capable of achieving an effective detection limit of 0.14 mg.L-1。
Practical application effect of formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chain
In order to explore the formaldehyde based on metal nano-particles and triple helical chainsThe practical use effect of the colorimetric detection method is that the inventor purchases three parts of Chinese cabbage, asparagus lettuce, Shanghai Qing and cabbage heart respectively from Guangzhou friendship store competitive supermarket, vegetable market and seven-delicacy life near southern school district of Zhongshan university, water is used as blank control, and 4 mg.L of the colorimetric detection method is used-1The formaldehyde solution of (2) was used as a positive control, and the detection was performed by the colorimetric formaldehyde detection method in the above examples.
The test method was the same as in the above example.
The processing method of the vegetable sample comprises the following steps: the vegetable sample is cut up first, and then the juice is squeezed out by a squeezer for detection.
The results are shown in FIG. 6.
From the results, it can be seen that the colorimetric formaldehyde detection method in the above examples provides different responses to several vegetable samples. The Chinese cabbage has the most obvious formaldehyde response, which indicates that the formaldehyde residue in the Chinese cabbage is the highest. While other vegetables, such as lettuce, shanghai green, cabbage heart, etc., had almost the same RGB values as the control, indicating that almost no formaldehyde was present in these samples. On the other hand, the formaldehyde concentration and the concentration of the formaldehyde solution (4 mg. L) detected in the 3 parts of the Chinese cabbage sample can be judged according to the RGB value conditions measured by the 3 parts of the Chinese cabbage sample-1) Similarly, the formaldehyde residue may be too high. Meanwhile, the result can also well show that the formaldehyde colorimetric detection method in the embodiment can be better used for detecting the formaldehyde residue in the Chinese cabbage and other vegetables in the current season.
Comparison of Formaldehyde colorimetric detection method based on Metal nanoparticles and triple helix chain with conventional detection method
In order to verify the reliability of the detection effect of the formaldehyde colorimetric detection method based on the metal nanoparticles and the triple helix chain, the inventor also adopts the conventional formaldehyde determination method (ultraviolet visible absorption spectrum and dynamic light scattering determination method) for auxiliary verification.
The detection samples are set as formaldehyde standards with the concentrations of 0.225mg/L and 4.5 mg/L.
The detection steps of the formaldehyde colorimetric detection method based on the metal nanoparticles and the triple helix chain are the same as the above embodiment.
The detection steps of the ultraviolet visible absorption spectrum are as follows: scanning the absorbance of the system under the conditions that the width of a slit is 2.0nm and the wavelength range is 750nm-450nm, wherein the absorption peak corresponding to the system containing 0.225mg/L formaldehyde standard substance is 620nm, the peak value is lower, the absorption peak corresponding to the system containing 4.5mg/L formaldehyde standard substance is 520nm, the peak value is higher, the change is obvious, and a standard curve of A620/A520 and the concentration can be drawn.
The detection steps of the dynamic light scattering measurement method are as follows: in the solvent selected to be H2O, testing the hydrated particle size of the system under the condition that the cell is selected to be SM50, wherein the particle size corresponding to the system containing 0.225mg/L formaldehyde standard is 1204nm, the particle size corresponding to the system containing 4.5mg/L formaldehyde standard is 75nm, the change is obvious, and a standard curve of the particle size and the concentration can be drawn.
The results of the uv-vis absorption spectroscopy and the dynamic light scattering measurement are shown in fig. 7 and 8.
The result of the colorimetric formaldehyde detection method based on the metal nanoparticles and the triple helix chain in the embodiment is consistent with the given concentration of the detection sample, and after the colorimetric formaldehyde detection method based on the metal nanoparticles and the triple helix chain is verified by using the ultraviolet-visible absorption spectrum, the change of the detection solution (gold nanoparticles) is found to be as shown in fig. 7, and the change trend of the curve conforms to the change relation between the formaldehyde concentration and the absorbance of the gold nanoparticles, so that the detection result can be considered to be accurate. Meanwhile, the particle size distribution diagram of the gold nanoparticles obtained by the dynamic light scattering determination method conforms to the theoretical estimated distribution (figure 8) of the change of the formaldehyde concentration and the particle size of the gold nanoparticles, so that the result obtained by the dynamic light scattering determination method can also show that the detection result of the formaldehyde colorimetric detection method based on the metal nanoparticles and the triple helical chains is accurate.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
SEQUENCE LISTING
<110> Zhongshan university
<120> formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chain and application thereof
<130>
<160> 4
<170> PatentIn version 3.5
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<211> 18
<212> DNA
<213> Artificial sequence
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<210> 2
<211> 30
<212> DNA
<213> Artificial sequence
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<212> DNA
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Claims (10)
1. An RGB colorimetric method for detecting formaldehyde, comprising the steps of:
(1) modifying an Oligo 1 probe sequence on the surface of the metal nanoparticle to obtain a metal nanoparticle probe;
(2) mixing a sample to be detected with the metal nanoparticle probe, the Oligo 2 sequence, the Oligo 3 sequence and the silver ion solution prepared in the step (1), and quantifying the formaldehyde content in the sample to be detected according to the RGB value of the final color of the mixed solution;
the Oligo 1 probe sequence is an oligonucleotide sequence;
the Oligo 2 sequence is an oligonucleotide sequence which is complementary and matched with only one end of the Oligo 1 probe sequence, which is far away from the gold nanoparticles;
the Oligo 3 sequence is a homopolypurine sequence which is complementary and matched with one end of the Oligo 1 probe sequence close to the gold nanoparticles;
the Oligo 2 sequence is not duplicated with the Oligo 3 sequence for any fragment in the Oligo 1 probe sequence.
2. The RGB colorimetric method of claim 1, wherein the metal nanoparticles include at least one of gold nanoparticles, silver nanoparticles, and palladium nanoparticles.
3. The RGB colorimetric method of claim 1, wherein the Oligo 1 probe sequence comprises 5-1000 homopolypyrimidine sequences, and the homopolypyrimidine sequence comprises 5-1000 cytosines.
4. The RGB colorimetric method of claim 1, wherein a linker is attached to the 5 'or 3' end of the Oligo 1 probe sequence, and the linker is a thiol group.
5. The RGB colorimetric method of claim 4, wherein the Oligo 1 probe sequence is 5' -S-C6-TTTGTTTTGTTTTCTTCTTTCCTTTCTTCT-3’(SEQ ID NO.2)。
6. The RGB colorimetric method according to claim 1, wherein the Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3);
the Oligo 3 sequence is as follows: 5'-AAACAAAACAAA-3' (SEQ ID NO. 4).
7. The RGB colorimetry according to claim 1, whereinIn that the silver ion solution includes: silver nitrate and Ag (NH)3)2At least one of OH.
8. A formaldehyde colorimetric detection reagent, which is characterized in that the formaldehyde colorimetric detection reagent contains at least one of the metal nanoparticles, the Oligo 1 probe sequence, the Oligo 2 sequence and the Oligo 3 sequence in the RGB colorimetric method according to any one of claims 1 to 7.
9. Formaldehyde colorimetric detection device, its characterized in that, formaldehyde colorimetric detection device includes:
a detection module comprising the colorimetric formaldehyde detection reagent of claim 8;
an imaging module for solution color recording;
the color identification module is used for identifying the color of the solution recorded by the imaging module and obtaining an RGB value; and the analysis module is used for calculating the formaldehyde concentration according to the RGB value obtained by the color recognition module.
10. Use of the colorimetric formaldehyde detection reagent of claim 8 or the colorimetric formaldehyde detection device of claim 9 for the detection of formaldehyde residues in food and environments.
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| CN102608073A (en) * | 2012-03-09 | 2012-07-25 | 西南大学 | Quick detection method for melamine in dairy products |
| CN108753925A (en) * | 2018-05-31 | 2018-11-06 | 中国科学院宁波材料技术与工程研究所 | A kind of colorimetric detection method and kit of single nucleotide polymorphism |
| CN111235228A (en) * | 2020-01-22 | 2020-06-05 | 中山大学 | Method for detecting cancer marker CA125 in blood based on polymerase chain reaction and dynamic light scattering |
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|---|---|---|---|---|
| CN102608073A (en) * | 2012-03-09 | 2012-07-25 | 西南大学 | Quick detection method for melamine in dairy products |
| CN108753925A (en) * | 2018-05-31 | 2018-11-06 | 中国科学院宁波材料技术与工程研究所 | A kind of colorimetric detection method and kit of single nucleotide polymorphism |
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