CN114034692B - Formaldehyde colorimetric detection method based on metal nano particles and triple helix chains and application thereof - Google Patents
Formaldehyde colorimetric detection method based on metal nano particles and triple helix chains and application thereof Download PDFInfo
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Abstract
The invention discloses a formaldehyde colorimetric detection method based on metal nano particles 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) 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 detection method of the invention is simple, quick, sensitive and reliable, does not need large instruments and complex operation processes, can realize on-site detection in extremely short time, has strong specificity and high sensitivity, and has the detection limit of only 0.14 mg.L ‑1 Has 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 nano particles and triple helix chains and application thereof.
Background
RGB color mode is a color standard in industry, which is to obtain various colors by changing three color channels of red (R), green (G) and blue (B) and overlapping them with each other, and includes almost all colors perceived by human eyesight, 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, its application in the field of biomolecule detection has not been reported yet.
Formaldehyde is one of the most common environmental pollutants and is very dangerous to humans. Formaldehyde is the main hazard of its toxic effect, and excessive exposure to formaldehyde can cause various levels of damage to the respiratory, digestive, circulatory and nervous systems. Moreover, formaldehyde was identified in 2004 by the international agency for research on cancer (IARC) as a class 1 carcinogen, and there is a risk of carcinogenesis upon prolonged exposure to formaldehyde. In daily life, bad vendors illegally use formaldehyde to keep vegetables fresh and preserve quality, so that certain formaldehyde residues are brought, and once the residual formaldehyde enters a human body, the residual formaldehyde brings great harm to the health of the human body. However, other daily activities, such as house decoration, etc., also cause a large amount of formaldehyde to remain, and are also easy to harm the health of human bodies.
In the related art, the formaldehyde residue is detected by adopting the traditional detection methods such as ultraviolet-visible absorption spectroscopy, fluorescence spectroscopy, raman spectroscopy and the like, 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 method is greatly limited.
Compared with the existing formaldehyde detection methods such as fluorescence, electrochemical, chromatography and chemiluminescence, the method has the defects of expensive equipment, complex operation steps, low detection limit and the like. Thus, it is still necessary to explore new formaldehyde detection strategies.
Therefore, in view of the huge and ubiquitous harm of formaldehyde, it is of great importance to develop a simple, rapid, sensitive and reliable formaldehyde on-site monitoring method.
Disclosure of Invention
The present invention aims to solve at least one of the technical problems in the prior art described above. Therefore, the formaldehyde colorimetric detection method based on the metal nano particles and the triple helix chain is simple to operate, short in detection time, high in sensitivity and good in specificity, and formaldehyde content in a sample to be detected can be accurately detected based on a mobile phone camera, so that on-site monitoring of formaldehyde can be realized.
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) 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.
According to a first aspect of the invention, in some embodiments of the invention, the Oligo 1 probe sequence is an oligonucleotide sequence.
In some preferred embodiments of the invention, the Oligo 1 probe sequence comprises 5-1000 homopyrimidine sequences; the homopyrimidine sequence contains 5-1000 cytosine.
In some preferred embodiments of the invention, the 5 'or 3' end of the Oligo 1 probe sequence is attached with a linker group for attaching 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 sulfhydryl group.
In some more preferred embodiments of the invention, the Oligo 1 probe sequence is
Wherein the underlined bolded part is a nucleic acid sequence with a complementary mismatch to the Oligo 2 sequence.
According to a first aspect of the invention, in some embodiments of the invention, the Oligo 2 sequence is an oligonucleotide sequence that is complementary paired only with the end of the Oligo 1 probe sequence remote from the gold nanoparticle.
In some preferred embodiments of the invention, the Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3).
According to a first aspect of the invention, in some embodiments of the invention, the Oligo 3 sequence is a homopurine sequence that is complementarily paired only with the Oligo 1 probe sequence at the end of the Oligo 1 probe sequence near the gold nanoparticle.
In some preferred embodiments of the 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 paired with any fragment of the Oligo 3 sequence in the Oligo 1 probe sequence repeatedly.
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 invention, the metal nanoparticles are gold nanoparticles. The gold nanoparticles may be prepared using commercially available gold nanoparticles or by methods conventional in the art, such as sodium citrate reduction.
Gold nanoparticles (gold nanoparticles, auNPs) are used as an important component of the nanomaterial, and the inventor finds that the dispersion/aggregation state of the AuNPs after DNA modification can be changed significantly, and the absorbance, color and particle size of the AuNPs are also changed. Thus, the AuNPs probe can be used as an ideal colorimetric, RGB and dynamic light scattering signaling probe to show formaldehyde levels.
In some preferred embodiments of the invention, the gold nanoparticles are prepared by: 2mL of freshly prepared 38.8mM sodium citrate solution was rapidly added to 20mL of boiled 1mM chloroauric acid HAuCl 4 In solution. At this time, the reaction liquid changed from pale yellow to black, then purple, and finally had changed to reddish wine. Heating and refluxing are continued after the color turns to the wine red, and stirring is carried out for 20-60 minutes. Cooling the reaction solution to room temperature (continuously keeping stirring), and then filtering with a nylon filter membrane with the diameter 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 ) 2 At least one of OH.
In some preferred embodiments of the 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 specific steps of the RGB colorimetric method are:
(1) Preparation of metal nanoparticle probes:
(2) The mixing was carried out according to the following system:
| component (A) | Dosage of |
| 30nM gold nanoparticle probe prepared in step (2) | 28μL |
| 0.1. Mu.M Oligo 2 sequence | 22μL |
| 0.2. Mu.M Oligo 3 sequence | 10μL |
| 50 mu M silver ion solution | 33μL |
| Formaldehyde sample | 33μL |
| Spermine 0.5mM | 12μL |
| PBS buffer (pH=7.0) | 82μL。 |
Wherein, spermine in the system is mainly used for stabilizing the formed triple helix structure.
(3) It is photographed using an imaging device, and RGB values of the imaged photograph are identified. Quantitative detection of formaldehyde is realized according to the linear relation between RGB value and formaldehyde concentration (using formaldehyde standard substance to determine 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, a RGB total value, an RGB average value, or an RGB ratio (R/G, R/B or G/B).
In some more preferred embodiments of the invention, the RGB value signal output is preferably an R value.
According to the experiments of the inventor, the R value of the system is in the range of 50-180 when the formaldehyde concentration 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 video camera, a microscope lens, and a camera.
In some preferred embodiments of the invention, the imaging device is a smart phone.
The mobile phone is fixed by a support, the camera is separated from the solution by about 12 cm, a light source in shooting parameters is adjusted to be in an incandescent lamp mode, and a sample is shot.
The detection principle of the RGB colorimetric method is shown in figures 1 and 2 of the drawings in the specification. The method is mainly based on two points: 1. based on the formation of triple-stranded DNA: the Oligo 3 sequence designed in the invention can be hybridized with the modified near AuNPs end single-stranded DNA (Oligo 1 probe sequence near AuNPs end) on the gold nanoparticle probe, so that the modified DNA (Oligo 1 probe sequence+oligo 3 sequence) is more three-dimensional, and is not simply adhered to the surface of AuNPs. 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 nanoparticle probe. Thereby the gold nanoparticle probes are respectively combined with the Oligo 2 sequence and the Oligo 3 sequence, when Ag is added + When gold nanoparticles aggregate (Ag + The Oligo 1 probe sequence, oligo 2 sequence and Oligo 3 sequence form a triple-stranded DNA structure, thereby aggregating gold nanoparticles, but if formaldehyde is contained in the solution at this time, the gold nanoparticles in the aggregated state are dispersed to different extents (formaldehyde can cause Ag) + The resulting triplex DNA structure is unwound) and thus a distinct color (the solution changes from red to purple or blue), particle size and turbidity changes, and by recognizing the RGB value, the formaldehyde content can be quantitatively detected from the fact that the RGB value is linear with the formaldehyde concentration.
In a second aspect of the invention, a formaldehyde colorimetric detection reagent is provided.
According to a second aspect of the present invention, in some embodiments of the present invention, the formaldehyde colorimetric detection reagent comprises at least one of a metal nanoparticle, an Oligo 1 probe sequence, an Oligo 2 sequence and an Oligo 3 sequence in the RGB colorimetric method according to the first aspect of the present invention.
According to a second aspect of the invention, in some embodiments of the invention, the Oligo 1 probe sequence is an oligonucleotide sequence.
In some preferred embodiments of the invention, the Oligo 1 probe sequence comprises 5-1000 homopyrimidine sequences; the homopyrimidine sequence contains 5-1000 cytosine.
The partial complementary mismatch nucleic acid sequence is identical to Ag + Specific recognition forms CGC trimer oligonucleotides (i.e., DNA triple helix structures) to aggregate gold nanoparticles.
In some preferred embodiments of the invention, the 5 'or 3' end of the Oligo 1 probe sequence is attached with a linker group for attaching 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 invention, the Oligo 1 probe sequence is
Wherein the underlined bolded part is a sequence of partially complementary mismatches.
According to a second aspect of the invention, in some embodiments of the invention, the Oligo 2 sequence is an oligonucleotide sequence that is complementary paired only with the end of the Oligo 1 probe sequence remote from the gold nanoparticle.
In some preferred embodiments of the invention, the Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3).
According to a second aspect of the invention, in some embodiments of the invention, the Oligo 3 sequence is a homopurine sequence that is complementarily paired only with the Oligo 1 probe sequence at the end of the Oligo 1 probe sequence near the gold nanoparticle.
In some preferred embodiments of the 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 paired with any fragment of the Oligo 3 sequence in the Oligo 1 probe sequence repeatedly.
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 invention, the metal nanoparticles are gold nanoparticles. The gold nanoparticles may be prepared using commercially available gold nanoparticles or by methods conventional in the art, such as sodium citrate reduction.
In some preferred embodiments of the invention, the gold nanoparticles are prepared by: 2mL of freshly prepared 38.8mM sodium citrate solution was rapidly added to 20mL of boiled 1mM chloroauric acid HAuCl 4 In solution. At this time, the reaction liquid changed from pale yellow to black, then purple, and finally had changed to reddish wine. Heating and refluxing are continued after the color turns to the wine red, and stirring is carried out for 20-60 minutes. Cooling the reaction solution to room temperature (continuously keeping stirring), and then filtering with a nylon filter membrane with the diameter 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, the silver ion solution comprises: silver nitrate and Ag (NH) 3 ) 2 At least one of OH.
In some preferred embodiments of the invention, the silver ion solution is silver nitrate.
In a third aspect of the present invention, there is provided a formaldehyde colorimetric detection device comprising:
the detection module contains the formaldehyde colorimetric detection reagent according to the second aspect of the invention;
the imaging module is used for recording the color of the solution;
the color recognition module is used for recognizing the color of the solution recorded by the imaging module and obtaining RGB values;
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 recognition module is an RGB color detector.
In some preferred embodiments of the invention, the RGB color detector is a cell phone comprising a charge coupled device image sensor CCD (or complementary metal oxide semiconductor CMOS), or other detector comprising an RGB three primary color sensor.
In a fourth aspect, the invention provides an application of the formaldehyde colorimetric detection reagent according to the second aspect or the formaldehyde colorimetric detection device according to the third aspect in food and environmental formaldehyde residue detection.
In some preferred embodiments of the invention, the food product comprises raw fresh fruit 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 work and living environments that are susceptible to formaldehyde contamination. Such as home decoration, outdoor paint painting, etc.
The beneficial effects of the invention are as follows:
1. the detection method is simple, quick, sensitive and reliable, does not need large instruments and complex operation processes, can realize on-site detection in extremely short time, and has far higher cost and efficiency than other traditional methods.
2. The detection method of the invention has the advantages of few required samples and strong specificity, can effectively eliminate the interference of other structural analogues such as acetaldehyde, methanol, ethanol, acetone, diethyl ether, acetic acid, ammonia water, toluene and the like, can accurately detect the formaldehyde content in a linear range, and has the detection limit of 0.14 mg.L -1 Has extremely high detection sensitivity.
Drawings
FIG. 1 is a schematic diagram of a colorimetric detection method of formaldehyde based on metal nanoparticles and triple helix chains in an embodiment of the present invention.
Fig. 2 is an enlarged view of the sequence of fig. 1 of the present specification.
FIG. 3 shows the specific detection results of the colorimetric detection method for formaldehyde in the embodiment of the present invention, and the samples are respectively: formaldehyde (HCHO), water (control, blank), acetaldehyde (CH) 3 CHO), methanol (CH 3 OH), ethanol (CH) 3 CH 2 OH), acetone (CH 3 COCH 3 ) Diethyl ether (C) 2 H 5 OC 2 H 5 ) Acetic acid (CH 3 COOH), ammonia monohydrate (NH) 3 ·H 2 O) and toluene (C) 6 H 5 CH 3 )。
FIG. 4 is a standard curve of formaldehyde standard solution obtained by the colorimetric detection method of formaldehyde in the embodiment of the invention.
Fig. 5 is a photograph of a color identifier of a colorimetric detection method of formaldehyde in an embodiment of the present invention.
FIG. 6 shows the results of colorimetric detection of formaldehyde on different vegetable samples in accordance with an embodiment of the present invention.
FIG. 7 is an ultraviolet-visible absorption spectrum for verifying the accuracy of the colorimetric detection method of formaldehyde in the embodiment of the invention.
FIG. 8 is a graph of particle size distribution of formaldehyde concentration of 0.225mg/L and 4.5mg/L, showing the accuracy of the colorimetric detection method for formaldehyde in the example of the present invention by dynamic light scattering measurement.
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 the following specific embodiments. It should be understood that the detailed description is presented herein for purposes of illustration only and is not intended to limit the invention.
The experimental materials and reagents used, unless otherwise specified, are those conventionally available commercially.
Formaldehyde colorimetric detection method based on metal nano particles and triple helix chains
The formaldehyde colorimetric detection method based on the metal nano particles and the triple helix chain in the embodiment comprises the following steps of:
(1) Preparation of metal nanoparticles:
in this embodiment, the metal nanoparticles used are gold nanoparticles (AuNPs) prepared according to the adjusted sodium citrate reduction method, and the specific preparation method is as follows:
2mL of freshly prepared 38.8mM sodium citrate solution was rapidly added to 20mL of boiled 1mM chloroauric acid HAuCl 4 In solution. At this time, the reaction liquid changed from pale yellow to black, then purple, and finally had changed to reddish wine. Heating and refluxing are continued after the color turns to the wine red, and stirring is carried out for 20-60 minutes. Cooling the reaction liquid to room temperature (continuously keeping stirring), then filtering with a nylon filter membrane with the diameter of 0.1-5.0 mu m to obtain gold nano particles, and preserving at the temperature of 4 ℃ for later use.
Of course, commercially available gold nanoparticles, such as, for example, gold nanoparticles, may also be used.
(2) Preparation of metal nanoparticle probes:
and (3) modifying the Oligo 1 probe sequence on the surface of the gold nanoparticles prepared in the step (1) 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 homopyrimidine sequences, the homopyrimidine sequences contain 5-1000 cytosine (C), and the sequence of complementary mismatch between the Oligo 1 probe sequence and the Oligo 2 sequence is as follows: 5'-TCTTCTTTCCTTTCTTCT-3' (SEQ ID NO. 1). The 5 'or 3' end of the oligodeoxyribonucleotide sequence is connected with a sulfhydryl.
In this example, the Oligo 1 probe sequence is:
wherein the underlined bolded portion is the sequence of a partial nucleic acid complementary mismatch.
b. Modification of gold nanoparticles:
(3) Sample detection:
an appropriate amount of sample was taken and mixed according to the system in table 1.
TABLE 1 detection System
| Component (A) | Dosage of |
| 30nM gold nanoparticle probe prepared in step (2) | 28μL |
| 0.1. Mu.M Oligo 2 sequence | 22μL |
| 0.2. |
10μL |
| 50 mu M silver ion solution | 33μL |
| Formaldehyde sample | 33μL |
| Spermine 0.5mM | 12μL |
| PBS buffer (pH=7.0) | 82μL |
Wherein the total of the detection system is 220. Mu.L.
The Oligo 2 sequence is an oligonucleotide sequence complementary to the Oligo 1 probe, which is complementary paired only to the end of the Oligo 1 probe sequence remote from the gold nanoparticle, in this example, the Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3' (SEQ ID NO. 3).
The Oligo 3 sequence is a homopurine sequence which is complementarily paired with only one end of the Oligo 1 probe sequence near the gold nanoparticle, in this embodiment, the Oligo 3 sequence is: 5'-AAACAAAACAAA-3' (SEQ ID NO. 4).
The Oligo 2 sequence and Oligo 3 sequence do not pair repeatedly any fragment of the Oligo 1 probe sequence.
In this embodiment, the silver ion solution is a silver nitrate solution. Of course, other silver ion solutions that do not interfere with the test results may be substituted by those skilled in the art.
After the system was mixed, the mixture was allowed to stand at room temperature for 2 minutes, and the color change of the solution was observed.
It is photographed using an imaging device, and RGB values (R values in this embodiment) of the imaged photograph are identified. Quantitative detection of formaldehyde is realized according to the linear relation between RGB value and formaldehyde concentration (using formaldehyde standard substance to determine standard curve).
In this embodiment, the imaging device adopts a mobile phone camera, fixes the mobile phone with a bracket, makes the camera and the solution spaced about 12 cm apart, adjusts the light source in the shooting parameters to an incandescent lamp mode, and photographs the sample. And reading the RGB value of the photo by using a color identifier, and realizing quantitative detection of formaldehyde according to the linear relation between the RGB value and the formaldehyde concentration.
Detection effect of formaldehyde colorimetric detection method based on metal nano particles and triple helix chains
(1) Quantitative detection effect (specificity):
in order to investigate the detection specificity of the above-described formaldehyde colorimetric detection method based on metal nanoparticles and triple helix chains, the inventors used different compounds as detection samples (each at a concentration of 4.5 mg/L) for detection.
In this example, formaldehyde (HCHO), water (control, blank), acetaldehyde (CH) were used, respectively 3 CHO), methanol (CH 3 OH), ethanol (CH) 3 CH 2 OH), acetone (CH 3 COCH 3 ) Diethyl ether (C) 2 H 5 OC 2 H 5 ) Acetic acid (CH 3 COOH), ammonia monohydrate (NH) 3 ·H 2 O) and toluene (C) 6 H 5 CH 3 ) As a test sample.
The test method is the same as the above embodiment.
The results are shown in FIG. 3.
As can be seen from fig. 3, the detection method in the embodiment of the invention has different response conditions to different interferents, but has specific response to formaldehyde only. The concrete steps are as follows: the detection method in the embodiment of the invention has the advantages that R values generated by interfering substances similar to formaldehyde structures, such as acetaldehyde, methanol, ethanol, acetone, diethyl ether, acetic acid, ammonia water and toluene are similar to RGB values of a control sample (water), so that the detection method in the embodiment of the invention does not generate specific responses to the structural analogues, but only the RGB values of formaldehyde are sharply increased, and the detection method in the embodiment has specific responses to formaldehyde.
(2) Detection sensitivity:
to explore the detection sensitivity of the above-described colorimetric detection method of formaldehyde based on metal nanoparticles and triple helix chains, the inventors used formaldehyde standard solutions of different concentrations (0.225 mg/L, 0.45mg/L, 0.9mg/L, 1.35mg/L, 3.6mg/L, 4.5 mg/L) for the test.
The test method is the same as the above embodiment.
The results are shown in fig. 4 and 5.
FIG. 4 is a standard curve constructed based on formaldehyde standard solutions of different concentrations (0.225 mg/L, 0.45mg/L, 0.9mg/L, 1.35mg/L, 3.6mg/L, 4.5 mg/L) and RGB values obtained by detection, the constructed standard curve being:
y＝17.50x+78.67；
wherein R is 2 =0.995, y is the detected RGB value; x is the concentration of formaldehyde. The linear range is 0.23 mg.L -1 -4.5mg·L -1 The detection limit was 0.14 mg.L -1 。
Fig. 5 is a graph showing that the concentration of formaldehyde solution can be accurately obtained by analyzing RGB values of formaldehyde samples of different concentrations using a cell phone color identifier based on the formaldehyde colorimetric detection method in the above-described embodiment.
From the above results, it was found that the detection method in the above examples is excellent in formaldehyde detection effect, high in sensitivity, and capable of achieving an effective detection limit of 0.14 mg.L -1 。
Actual application effect of formaldehyde colorimetric detection method based on metal nano particles and triple helix chains
In order to explore the practical use effect of the formaldehyde colorimetric detection method based on metal nano particles and triple helix chains, the inventor purchased three parts of Chinese cabbage, lettuce, shanghai green and cabbage heart from Guangzhou friendship store exquisite supermarket, vegetable market and seven fresh life near the southern school district of the university of Zhongshan, and uses water as blank control, 4 mg.L -1 As a positive control, the formaldehyde solution was detected using the formaldehyde colorimetric detection method in the above example.
The test method is the same as the above embodiment.
Wherein, the processing method for the vegetable samples comprises the following steps: firstly, cutting up vegetable samples, and then extruding vegetable juice by a water extruder to obtain the vegetable juice detection device.
The results are shown in FIG. 6.
From the results, it was found that, after detection using the colorimetric detection method of formaldehyde in the above examples, the detection was performed on several vegetablesThe samples had different response profiles. Wherein, the formaldehyde response of the Chinese cabbage is most obvious, which indicates that the formaldehyde residue in the Chinese cabbage is the highest. While other vegetables, such as lettuce, shanghai green, cabbage, etc., have nearly the same RGB values as the control group, indicating that formaldehyde is almost absent in these samples. According to the RGB values measured by 3 Chinese cabbage samples, the concentration of formaldehyde and the concentration of formaldehyde solution (4mg.L) detected in 3 Chinese cabbage samples can be determined -1 ) Similarly, formaldehyde residues may be too high. Meanwhile, the result can also well illustrate that the formaldehyde colorimetric detection method in the embodiment can be well used for detecting formaldehyde residues of cabbages and other vegetables in the current season.
Comparison of formaldehyde colorimetric detection method based on metal nano particles and triple helix chains with conventional detection method
In order to verify the reliability of the detection effect of the formaldehyde colorimetric detection method based on the metal nano-particles and the triple helix chains, the inventor also adopts a conventional formaldehyde detection method (ultraviolet visible absorption spectrum and dynamic light scattering detection method) to carry out auxiliary verification.
The test samples were set to formaldehyde standards at concentrations of 0.225mg/L and 4.5 mg/L.
The detection steps of the formaldehyde colorimetric detection method based on the metal nano-particles and the triple helix chains are the same as the above embodiments.
The detection steps of the ultraviolet visible absorption spectrum are as follows: the absorbance of the system is scanned under the conditions that the slit width is 2.0nm and the wavelength ranges from 750nm to 450nm, 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 the standard curve of A620/A520 and the concentration can be also made.
The detection steps of the dynamic light scattering measurement method are as follows: in the solvent selected as H 2 O, the hydration particle size of the test system under the condition that the pool is selected as SM50, the particle size corresponding to a system containing 0.225mg/L formaldehyde standard substance is 1204nm, the particle size corresponding to a system containing 4.5mg/L formaldehyde standard substance is 75nm, the change is obvious, and a standard curve of the particle size and the concentration can be also made.
The detection results of the ultraviolet visible absorption spectrum and the dynamic light scattering assay are shown in fig. 7 and 8.
The detection result of the formaldehyde colorimetric detection method based on the metal nano particles and the triple helix chains in the embodiment is consistent with the given concentration of the detection sample, and after the formaldehyde colorimetric detection method based on the metal nano particles and the triple helix chains is verified by utilizing an ultraviolet visible absorption spectrum, the change of the detection liquid (gold nano particles) is shown in fig. 7, and the change trend of the curve accords with the relation between the formaldehyde concentration and the absorbance change of the gold nano particles, so that the detection result can be considered to be accurate. Meanwhile, the distribution diagram of the particle diameters of the gold nanoparticles obtained by the dynamic light scattering measurement method accords with the theoretical estimated distribution of the change of the concentration of formaldehyde and the particle diameter size of the gold nanoparticles (figure 8), so that the measurement result obtained by the dynamic light scattering measurement method can also indicate that the detection result of the formaldehyde colorimetric detection method based on the metal nanoparticles and the triple helix chains is accurate.
The above examples are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above examples, and any other changes, modifications, substitutions, combinations, and simplifications that do not depart from the spirit and principle of the present invention should be made in the equivalent manner, and the embodiments are included in the protection scope of the present invention.
SEQUENCE LISTING
<110> university of Zhongshan
<120> a formaldehyde colorimetric detection method based on metal nano particles and triple helix chains and application thereof
<130>
<160> 4
<170> PatentIn version 3.5
<210> 1
<211> 18
<212> DNA
<213> artificial sequence
<400> 1
tcttctttcc tttcttct 18
<210> 2
<211> 30
<212> DNA
<213> artificial sequence
<400> 2
tttgttttgt tttcttcttt cctttcttct 30
<210> 3
<211> 18
<212> DNA
<213> artificial sequence
<400> 3
agaagaaagg aaagaaga 18
<210> 4
<211> 12
<212> DNA
<213> artificial sequence
<400> 4
aaacaaaaca aa 12
Claims (8)
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, the silver ion solution and spermine 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 5' -HS-C 6 -TTTGTTTTGTTTTCTTCTTTCCTTTCTTCT-3’；
The Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3';
the Oligo 3 sequence is: 5'-AAACAAAACAAA-3'.
2. The RGB colorimetric method according to claim 1, wherein the metal nanoparticles comprise at least one of gold nanoparticles, silver nanoparticles and palladium nanoparticles.
3. The RGB colorimetric method according to claim 1, wherein the silverThe ionic solution comprises: silver nitrate and Ag (NH) 3 ) 2 At least one of OH.
4. The formaldehyde colorimetric detection reagent is characterized by comprising metal nano particles, an Oligo 1 probe sequence, oligo 2 sequences, oligo 3 sequences, silver ion solution and spermine;
the Oligo 1 probe sequence is 5' -HS-C 6 -TTTGTTTTGTTTTCTTCTTTCCTTTCTTCT-3’；
The Oligo 2 sequence is: 5'-AGAAGAAAGGAAAGAAGA-3';
the Oligo 3 sequence is: 5'-AAACAAAACAAA-3'.
5. The formaldehyde colorimetric detection reagent of claim 4, wherein the metal nanoparticles comprise at least one of gold nanoparticles, silver nanoparticles, and palladium nanoparticles.
6. The formaldehyde colorimetric detection reagent of claim 4 wherein the silver ion solution comprises: silver nitrate and Ag (NH) 3 ) 2 At least one of OH.
7. Formaldehyde colorimetric detection device, its characterized in that, formaldehyde colorimetric detection device includes:
the detection module comprises the formaldehyde colorimetric detection reagent according to any one of claims 4-6;
the imaging module is used for recording the color of the solution;
the color recognition module is used for recognizing the color of the solution recorded by the imaging module and obtaining RGB values;
the analysis module is used for calculating formaldehyde concentration according to the RGB value obtained by the color recognition module;
the solution is a mixed solution obtained by the following steps:
(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, the silver ion solution and the spermine prepared in the step (1) to obtain a mixed solution.
8. The formaldehyde colorimetric detection reagent according to any one of claims 4 to 6 or the formaldehyde colorimetric detection device according to claim 7, and the use thereof in detecting food or environmental formaldehyde residues.
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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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| Cen Xiong et al..Gold nanoparticles-based colorimetric investigation of triplex formation under weak alkalic pH environment with the aid of Ag+.Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy.2011,第79卷(第5期),956-961. * |
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