CN114441503B - Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis - Google Patents

Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis Download PDF

Info

Publication number
CN114441503B
CN114441503B CN202210072658.8A CN202210072658A CN114441503B CN 114441503 B CN114441503 B CN 114441503B CN 202210072658 A CN202210072658 A CN 202210072658A CN 114441503 B CN114441503 B CN 114441503B
Authority
CN
China
Prior art keywords
melamine
solution
sers
concentration
internal standard
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active
Application number
CN202210072658.8A
Other languages
Chinese (zh)
Other versions
CN114441503A (en
Inventor
杨龙坤
张彤彤
李志鹏
张慧娟
张�浩
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Capital Normal University
Original Assignee
Capital Normal University
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Capital Normal University filed Critical Capital Normal University
Priority to CN202210072658.8A priority Critical patent/CN114441503B/en
Publication of CN114441503A publication Critical patent/CN114441503A/en
Application granted granted Critical
Publication of CN114441503B publication Critical patent/CN114441503B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/65—Raman scattering
    • G01N21/658—Raman scattering enhancement Raman, e.g. surface plasmons
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y30/00—Nanotechnology for materials or surface science, e.g. nanocomposites
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y40/00—Manufacture or treatment of nanostructures
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W30/00—Technologies for solid waste management
    • Y02W30/50—Reuse, recycling or recovery technologies
    • Y02W30/62—Plastics recycling; Rubber recycling

Abstract

The embodiment of the invention discloses a melamine quantitative detection method based on Surface Enhanced Raman Spectroscopy (SERS) internal standard analysis, which comprises the following steps: preparing silver nanoparticle aggregate with internal standard molecules (citrate) by a laser-induced reduction method as a SERS substrate; collecting SERS signals, calculating the ratio of characteristic peaks of melamine molecules to characteristic peaks of the internal standard molecules as the relative intensity of the characteristic peaks of the melamine molecules, and fitting the concentration of melamine solution and the relative intensity (I R ) A functional relationship between; and dropwise adding a melamine solution with unknown concentration onto the SERS substrate, repeatedly collecting SERS signals at different positions, calculating the average value of the relative intensities, and reversely estimating the concentration of melamine in the solution according to the functional relation. According to the invention, quantitative detection of melamine molecules is realized by adopting a SERS internal standard analysis method, and the method has the advantages of simple sample preparation, rapid detection and high sensitivity and accuracy.

Description

Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis
Technical Field
The invention relates to the technical field of Surface Enhanced Raman Spectroscopy (SERS) detection and quantitative analysis, in particular to a melamine quantitative detection method based on surface enhanced Raman spectroscopy internal standard analysis.
Background
Melamine is a triazine nitrogen-containing heterocyclic organic compound, and the nitrogen content is up to 66%. In the food industry, the Kjeldahl method is generally adopted, and the protein content is estimated by detecting the nitrogen content. Therefore, some lawless persons mix melamine into water or bad fresh milk, and the protein content reaches the standard during detection.
The efficient and accurate quantitative detection of melamine molecules becomes a key to solve the problem. At present, common methods for melamine detection comprise high performance liquid chromatography, liquid chromatography-mass spectrometry/mass spectrometry, gas chromatography-mass spectrometry combined method, infrared spectrometry, electrochemical method, enzyme-linked immunosorbent assay, colorimetry and the like. However, the above methods have the disadvantages of complicated process, long detection period, expensive required instruments, high detection cost and the like.
The raman scattering spectrum has a characteristic peak of a high recognition degree 'molecular fingerprint', can reflect chemical structure information of molecules, and is attracting a great deal of attention. However, the scattering cross section of conventional raman scattering tends to be very small (-10) -30 cm 2 ) Limiting the application of raman spectroscopy techniques. And Surface Enhanced Raman (SERS) is a phenomenon in which the raman scattering intensity is greatly enhanced after a molecule to be measured is adsorbed on a rough metal surface or a metal nanostructure surface having a sub-nano space. It is attributed to the sharp increase in electric field strength in the sub-nanometer space caused after laser irradiation, forming hot spots. However, the non-uniformity of the enhancing effect of the SERS substrate and the strong fluctuation of the peak intensity of the object to be detected caused by experimental conditions become a great obstacle for realizing the quantitative monitoring of melamine molecules.
Thus, there is a need to provide a SERS method that can be used for quantitative detection of melamine.
Disclosure of Invention
The invention aims to provide a melamine quantitative detection method based on surface-enhanced Raman spectrum internal standard analysis, which aims to solve at least part of the technical problems of the existing detection method.
In order to achieve the above object, an embodiment of the present invention provides a method for quantitatively detecting melamine based on surface-enhanced raman spectroscopy internal standard analysis, including:
step 1, preparing silver nanoparticle aggregate with internal standard molecules by a laser-induced reduction method as a surface-enhanced Raman scattering (SERS) substrate; comprising the following steps:
uniformly mixing a silver nitrate solution and a sodium citrate solution to obtain a reaction solution, placing the reaction solution on conductive glass, and covering a cover glass above the reaction solution; converging laser at the interface of the reaction solution and the conductive glass, and forming silver nanoparticle aggregates at the interface through laser-induced reduction after irradiation for preset time; washing the formed silver nanoparticle aggregate with deionized water, and removing unreacted solution to obtain a silver nanoparticle aggregate with citrate serving as a SERS substrate, wherein the citrate is the internal standard molecule;
step 2, obtaining the ratio of the characteristic peak of the melamine molecule in the SERS signal to the characteristic peak of the sodium citrate molecule as the relative intensity of the characteristic peak of the melamine molecule, and fitting the concentration of the melamine solution and the relative intensity (I R ) A functional relationship between; comprising the following steps:
detecting the surface enhanced Raman scattering spectrum of melamine molecules in the aqueous solution on the SERS substrate to obtain a spectrum of 703cm -1 Characteristic peak of melamine molecule and 1380cm -1 Characteristic peaks of citrate ions;
1380cm of the citrate ion -1 The characteristic peak is used as an intensity internal standard to obtain the relative intensity of the characteristic peak of the melamine moleculeThe relative standard deviation of the relative intensities is less than the relative standard deviation of the melamine molecular characteristic peaks;
detecting SERS signals of melamine water solutions at different concentrations, and fitting the functional relation according to the obtained relative intensities;
and step 3, dropwise adding a melamine solution with unknown concentration onto the SERS substrate, repeatedly collecting SERS signals at different positions, calculating the relative intensity average value, and reversely estimating the concentration of melamine in the solution according to the functional relation.
In one embodiment, the relative standard deviation of the relative intensities of the characteristic peaks of the melamine molecules is less than 10%.
In one embodiment, when the concentration of the aqueous melamine solution is 10 -6 M-10 -8 Within M, the relative intensity is linearly related to the concentration of the aqueous melamine solutionIs tied up.
In one embodiment, the SERS signals of the melamine water solution at different concentrations are detected, and the functional relation is fitted according to the obtained relative intensities, including: and obtaining an average value of a plurality of obtained values of the relative intensities, and determining the concentration of melamine molecules in the solution according to the average value and the linear function relation.
Compared with the prior art, the invention has at least the following advantages:
compared with the prior art, the method has the advantages of simple sample preparation, rapid detection, high sensitivity and accuracy and the like, and specifically comprises the following steps:
(1) Silver nano-aggregates were prepared by simple laser-induced reduction as SERS substrates, which naturally carry internal standard molecules (citrate).
(2) Intensity calibration by internal standard moleculesCan effectively eliminate melamine peak intensity fluctuation caused by factors such as non-uniformity of SERS substrate or experimental conditions, and I R The relative standard deviation of (2) can be controlled within 10 percent, and reliable assurance is provided for quantitative analysis.
(3) At 10 -6 M-10 -8 Within the M concentration range, I R Has good linear relation with the melamine concentration, which takes the melamine concentration as the abscissa, I R The linear function relation is obtained by fitting the ordinate: y= 1.99568 ×x+0.09674.
(4) The linear relation is taken as a quantitative curve, and melamine (7.5 multiplied by 10) in the aqueous solution and the milk solution is obtained through SERS intensity internal standard analysis -7 The detection recovery rate of M) can reach 103% and 112% respectively.
Drawings
Fig. 1 is a flow chart of a melamine quantitative detection method based on surface enhanced raman spectroscopy internal table analysis provided by the embodiment of the invention.
Fig. 2 is a schematic diagram of a method for preparing a SERS substrate according to an embodiment of the present invention.
Fig. 3 is a scanning electron micrograph of a silver nanoparticle aggregate prepared according to the method provided by the embodiment of the present invention.
FIG. 4 shows the detection of melamine molecules (2.5X10) in aqueous solution on the substrate according to the method provided by the embodiment of the invention -7 M) and a schematic diagram of the raman characteristic peak of the melamine powder obtained by detection. .
FIG. 5 shows a concentration of 2.5X10 measured by the method according to the embodiment of the invention -7 Schematic representation of raman scattering spectra of M melamine solution at 703cm measured at different positions of the substrate -1 Absolute intensity variation at melamine molecular characterization peak and I R Is a schematic representation of the intensity variation of (a).
Fig. 6 is a schematic diagram of raman scattering spectra of different concentrations detected by a method according to an embodiment of the present invention.
FIG. 7 is a graph showing the ratio of melamine molecular concentration to the two peak intensities (I R ) Schematic representation of the functional relationship between them.
Fig. 8 is a schematic diagram of raman scattering spectra of melamine aqueous solutions, milk solutions, and blank milk solutions of unknown concentrations detected according to the method provided by the embodiment of the present invention.
Fig. 9 is a schematic diagram of reverse estimating melamine concentration in aqueous and milk solutions according to the method provided by the embodiment of the invention.
Fig. 10 is an analytical chart showing the recovery rate obtained during the estimation of melamine aqueous solution and milk solution of unknown concentration according to the method provided by the example of the present invention.
Detailed Description
In the drawings, the same or similar reference numerals are used to denote the same or similar elements or elements having the same or similar functions. Embodiments of the present invention will be described in detail below with reference to the accompanying drawings.
In the description of the present invention, the terms "center", "longitudinal", "lateral", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", and the like indicate an orientation or a positional relationship based on that shown in the drawings, only for convenience of description and simplification of the description, and do not indicate or imply that the apparatus or element to be referred to must have a specific orientation, be configured and operated in a specific orientation, and therefore should not be construed as limiting the scope of protection of the present invention.
In the case of no conflict, the technical features in the embodiments and the implementation modes of the present invention may be combined with each other, and are not limited to the embodiments or implementation modes where the technical features are located.
Surface Enhanced Raman (SERS) refers to a phenomenon in which the raman scattering intensity is greatly enhanced after a molecule to be measured is adsorbed on a rough metal surface or a metal nanostructure surface having a sub-nano space. SERS has a characteristic peak of a molecular fingerprint with high recognition degree, can reflect chemical structure information of molecules, and is widely applied to biomolecule detection. It is essentially a near field phenomenon, where only those molecules (so-called "hot spots") that are within a highly enhanced electromagnetic field (typically 2 nm) make a significant contribution to the overall SERS signal. However, field enhancement of hot spots is sensitive to coupling between nanostructures, and signal fluctuations often occur due to non-uniformity of hot spots in the metal nanostructures; meanwhile, due to the problems of uncertainty of molecular adsorption behaviors and the like, high-precision quantitative analysis and detection of molecules under low concentration are difficult, and practical application of SERS quantification is hindered.
To solve this problem, attempts have been made to introduce an internal standard method to calibrate SERS signals to achieve quantitative detection of molecules. The internal standard method is to introduce an internal standard substance with the same enhancement function as the analyte molecules, and simultaneously detect the Raman signals of the analyte molecules and the internal standard substance under the same condition, so that the instability and the nonuniformity of the SERS substrate are calibrated, a more accurate qualitative and quantitative analysis method is realized, and the practical application problem of SERS quantification can be effectively solved. However, the introduction of additional internal standard substances may increase the difficulty of the experiment or cause competitive adsorption to affect the experimental signal. The present invention seeks to find a substrate with an internal standard substance.
The embodiment of the invention provides a melamine quantitative detection method based on surface-enhanced Raman spectrum internal standard analysis, which utilizes a silver nanoparticle aggregate synthesized by laser-induced reduction and provided with internal standard molecule citrate as a SERS substrate, and realizes quantitative detection of melamine molecules by an internal standard method. As shown in fig. 1, the method includes:
step 1, preparing a silver nanoparticle aggregate with internal standard molecules by a laser-induced reduction method as an SERS substrate; comprising the following steps:
uniformly mixing a silver nitrate solution and a sodium citrate solution to obtain a reaction solution, placing the reaction solution on conductive glass, and covering a cover glass above the reaction solution; converging laser at the interface of the reaction solution and the conductive glass, and forming silver nanoparticle aggregates at the interface through laser-induced reduction after irradiation for preset time; washing the formed silver nanoparticle aggregate with deionized water to remove unreacted solution; the sodium citrate participates in the reduction process, so that citrate ions are adsorbed on the surface of the silver nanoparticle aggregate, the silver nanoparticle aggregate with internal standard molecule citrate is obtained, and the silver nanoparticle aggregate is used as an SERS substrate.
Step 2, obtaining the ratio of the characteristic peak of the melamine molecule to the characteristic peak of the sodium citrate molecule in the SERS signal as the relative intensity of the characteristic peak of the melamine molecule, and fitting the concentration of the melamine solution and the relative intensity (I R ) A functional relationship between, comprising:
detecting the surface enhanced Raman scattering spectrum of melamine molecules in the aqueous solution on the SERS substrate to obtain a spectrum of 703cm -1 Characteristic peak of melamine molecule and 1380cm -1 Characteristic peaks of citrate ions;
1380cm of the citrate ion -1 The characteristic peak is used as an intensity internal standard to obtain the relative intensity of the characteristic peak of the melamine moleculeThe relative standard deviation of the relative intensities is less than the relative standard deviation of the characteristic peaks of the melamine molecules;
and detecting SERS signals of melamine water solutions at different concentrations, and fitting the functional relation according to the obtained relative intensity values.
In one example, melamine molecules (2.5X10) in aqueous solution are detected on the substrate -7 M) can be observed at the same time at 703cm -1 Characteristic peak of melamine molecule and 1380cm -1 Is characterized by the characteristic peak of citrate ion. It was found that, at this time, if only melamine molecules were analyzed to be located at 703cm -1 Characteristic peak intensity, relative Standard Deviation (RSD) of the experiment was 20%; if citrate ions are 1380cm -1 Characteristic peaks at the points as intensity internal standardsCalculation I R To 8.9%; comprising the following steps:
the substrate was placed on a stage and the melamine solution (2.5X10 -7 M) dropwise adding the mixture onto a substrate, regulating laser, collecting surface-enhanced Raman scattering spectrum signals of melamine molecules, and observing the signals at 703cm -1 Characteristic peaks of melamine molecules at 1380cm -1 The characteristic peak of the citrate ion is detected, and the characteristic peak of the citrate ion can exist stably all the time through time stability detection. Further detection of the raman spectra of melamine molecules at different positions on the substrate, we found that if only melamine molecules were analyzed at 703cm -1 The characteristic peak at the position has larger absolute peak intensity fluctuation and about 20 percent of Relative Standard Deviation (RSD) due to the influence of factors such as the heterogeneity of the SERS substrate or experimental conditions; at this time, if the citrate ion is 1380cm -1 Characteristic peak at the position as an intensity internal standardReanalytical melamine molecular characterizationThe relative intensity of the characteristic peaks can then reduce RSD to 8.9%.
Wherein, through the intensity calibration of internal standard molecules, namely using the relative intensity of melamine molecular characteristic peaks, the fluctuation of melamine peak intensity caused by factors such as the heterogeneity of SERS substrates or experimental conditions can be effectively eliminated, and I R The relative standard deviation of the (C) can be controlled within 10 percent, and reliable assurance is provided for quantitative analysis.
In one example, detect 10 -5 M-10 -8 Raman scattering spectra of melamine aqueous solutions with 11 different concentrations in the M concentration range were calculatedRSD and I of (a) R Can find melamine molecule I at different concentrations R RSD of (a) is less than 10%, and at 10 -6 M-10 -8 Within the M concentration range, I R Has good linear relation with the melamine concentration, which takes the melamine concentration as the abscissa, I R Obtaining the linear function relation for ordinate fitting: y= 1.99568 ×x+0.09674; comprising the following steps:
the concentrations were 1.0X10 respectively -5 mol/L、7.5×10 -6 mol/L、5.0×10 -6 mol/L、2.5×10 -6 mol/L、1.0×10 -6 mol/L、7.5×10 -7 mol/L、5.0×10 -7 mol/L、2.5×10 -7 mol/L、1.0×10 -7 mol/L、5.0×10 -8 mol/L、1.0×10 -8 The 11 melamine aqueous solutions of mol/L are respectively dripped on the substrate, and the Raman spectrum of the substrate is detected. For each concentration, the melamine molecules were read 703cm by multiple measurements -1 The absolute peak intensity of the characteristic peak is calculated and the RSD is found to be more than 20%. Accordingly, citrate ions 1380cm -1 Characteristic peaks at the points as intensity internal standards Discovery I R RSD of (a) is less than 10%, and at 10 -6 M-10 -8 Within the M concentration range, I R Has good linear relation with the melamine concentration. On the abscissa of melamine concentration, I R And obtaining the linear function relation for ordinate fitting.
And step 3, dropwise adding a melamine solution with unknown concentration onto the SERS substrate, repeatedly collecting SERS signals at different positions, calculating the average value of the relative intensities, and reversely estimating the concentration of melamine in the solution according to the functional relation.
In one example, an aqueous melamine solution or a milk solution of unknown concentration is dripped onto the substrate, raman spectrum signals are acquired for a plurality of times at different positions, and I is calculated according to the spectrum R And reversely estimating the concentration of melamine in the solution according to the linear function relation, comparing the concentration with the actual concentration, and calculating to obtain the detection recovery rates of the melamine water solution and the milk solution which can reach 103% and 112% respectively. Comprising the following steps:
collecting Raman spectrum signals at different positions on the substrate, and reading 703cm of melamine molecular characteristic peaks -1 The absolute peak intensity at the position and the characteristic peak of citrate ion 1380cm -1 Absolute peak intensities at different positions are calculatedAnd find I R And inversely estimating the concentration of melamine molecules in said solution from the average and said linear functional relationship, and calculating the recovery rate thereof.
And (2) taking the linear relation in the step (2) as a quantitative curve, and obtaining detection recovery rates of melamine in the aqueous solution or the milk solution respectively reaching 103% and 112% through SERS intensity internal standard analysis.
In order to make the melamine quantitative detection method based on the surface enhanced Raman spectrum more clear, the following detailed description is provided by a specific embodiment. It should be understood that the specific parameters in this embodiment are all preferred, but not exclusive, and those skilled in the art may adjust each parameter in the following examples to obtain other embodiments without departing from the technical spirit of the present invention, which all fall within the protection scope of the present invention.
This embodiment is described by the following aspects: 1. weighing a sample; 2. an instrument device; preparation of sers substrates (silver nanoparticle aggregates); 4. detection and analysis of Melamine (Melamine).
1. Weighing a sample:
(1) 18.5mg of silver nitrate and 22.5mg of sodium citrate were weighed out by a balance and dissolved in 10mL of deionized water (resistivity of 18 M.OMEGA.cm), respectively.
(2) 12.61mg of melamine powder was weighed by a balance and dissolved in 10mL of deionized water to give a concentration of 1.0X10 -2 1mL of melamine aqueous solution with mol/L is added into 9mL of deionized water for dilution to obtain the melamine aqueous solution with the concentration of 1.0X10 -3 melamine aqueous solution in mol/L. This step was repeated until a concentration of 1.0X10 was obtained -8 melamine aqueous solution in mol/L.
(3) 7.5mL of the extract was taken at a concentration of 1.0X10 -5 Adding 2.5mL deionized water into mol/L aqueous solution of melamine to dilute to obtain a solution with a concentration of 7.5X10 -6 melamine aqueous solution in mol/L. According to the above method, a concentration of 5.0X10 was obtained, respectively - 6 mol/L、2.5×10 -6 mol/L、7.5×10 -7 mol/L、5.0×10 -7 mol/L、2.5×10 -7 mol/L、5.0×10 -8 Aqueous melamine solutions in mol/mol.
(4) 12.61mg of melamine are weighed by a balance and dissolved in 10mL of milk (by way of example only, water or other desired solution) to give a concentration of 1.0X10 -2 The melamine milk solution with mol/L is taken, 1mL is added into 9mL of milk for dilution, and the concentration is 1.0X10 -3 melamine milk solution in mol/L. This step was repeated until a concentration of 1.0X10 was obtained -5 melamine milk solution in mol/L.
(5) 1mL of the solution was taken to have a concentration of 1.0X10 -5 The melamine milk solution with mol/L is diluted by deionized water to obtain the concentration of 1.0 respectively×10 -7 mol/L and 7.5X10 -7 melamine milk solution in mol/L.
2. Instrument apparatus:
clean graduated conductive glass, clean coverslips, semi-microbalances (e.g., a Metrele-Toril multi METTER TOLEDOO semi-microbalance), raney's micro Raman spectrometer RH13325 (R-2000), lasers at 532 nm.
Preparation of sers substrate:
fig. 2 shows a schematic diagram of a system for preparing SERS substrates, including clean graduated conductive glass (ITO glass), reactant mixtures, clean coverslips, lasers at 532nm wavelength, nanoparticles on the ITO glass prepared, and an objective lens for observing the nanoparticles.
The preparation process comprises the following steps:
(1) Clean conductive glass (ITO glass) is placed on an objective table under a Raman spectrometer microscope, and a clear image of the conductive glass scale is found by adjusting the focal length.
(2) And uniformly mixing 0.01mol/L silver nitrate and 0.009mol/L sodium citrate according to a volume ratio of 1:1 to obtain a reaction solution. 200 mu L of reaction solution is dripped on the conductive glass, all graduated places are covered with the mixed solution, and a clean cover slip is covered above the reaction solution.
(3) And adjusting the focal length to enable light to be converged at the interface of the reaction solution and the conductive glass. A position was determined at the interface, and the silver nanoparticle aggregate was obtained by irradiating with a laser having a wavelength of 532nm and a power of 1mW for 90 s. Fig. 3 shows a scanning electron micrograph of a silver nanoparticle aggregate.
(4) And taking down the sample, gently flushing the substrate with deionized water, washing off unreacted solution, and drying to obtain the substrate.
Wherein, the steps (2) - (4) are carried out in a dark place.
4. Detection and analysis of melamine:
(1) Silver nanoparticle aggregates with internal standard molecules (citrate) were prepared by laser induced reduction and used as SERS substrates: uniformly mixing a silver nitrate solution and a sodium citrate solution to obtain a reaction solution, placing the reaction solution on conductive glass, and covering a cover glass above the reaction solution; converging laser at the interface of the reaction solution and the conductive glass, and irradiating for a preset time to prepare silver nanoparticle aggregates at the interface through laser-induced reduction;
(2) Rinsing the substrate with deionized water, removing unreacted solution, and drying at room temperature; wherein, because sodium citrate is used in the reduction process, the surface of the silver nanoparticle aggregate is adsorbed with rich citrate ions
(3) The substrate was placed on a stage and 200 μl of aqueous melamine solution was added dropwise to the substrate, and a clean coverslip was covered over the solution. And after the clear image of the substrate is found by adjusting the focal length, selecting a measuring point on the substrate, selecting laser with the wavelength of 532nm, wherein the laser power is 0.16mW, the integration time is 10s, and collecting a molecular Raman scattering spectrum signal. By this Raman spectrum, it was clearly observed that the spectrum was located at 703cm -1 Characteristic peaks of melamine molecules at 1380cm -1 The characteristic peaks of citrate ions at the two positions of the characteristic peaks are not overlapped. FIG. 4 shows the detection of melamine molecules (2.5X10 in aqueous solution on the substrate -7 M) raman characteristic peaks of melamine powder.
(4) And (3) selecting laser with the wavelength of 532nm, wherein the laser power is 0.16mW, and the Raman scattering spectrum signals of the substrate in (3) are collected every 1 minute, and the integration time of each time is 1s. As can be seen from analysis of the collected signals, under the same conditions, citrate ions of 1380cm -1 The absolute peak intensity of the characteristic peak has stronger time stability.
(5) At 2.5X10 -7 For example, the concentration is 2.5X10 -7 Dripping a mol/L melamine aqueous solution onto the substrate, detecting Raman scattering spectrum signals of melamine molecules at different positions on the substrate, and finding that if only the melamine molecules are analyzed to be located at 703cm -1 Characteristic peaks at the position due to factors such as non-uniformity of SERS substrate or experimental conditionsThe absolute peak intensity fluctuation is large, and the Relative Standard Deviation (RSD) of the experiment is 20%; at this time, if the citrate ion is 1380cm -1 Characteristic peaks at the sites were used as intensity internal standard (I 703 cm -1 /I 1380 cm -1 ，I R ) After analysis of the relative intensity of melamine molecular characteristic peaks, RSD can be reduced to 8.9%; FIG. 5 shows a concentration of 2.5X10 -7 Schematic representation of raman scattering spectra of aqueous melamine solutions at M, 703cm measured at different positions of the substrate -1 Absolute intensity variation and relative intensity I of melamine molecular characteristic peak R Is a variation of the schematic diagram.
(6) Repeating the step (5), and detecting the concentration to be 1.0X10 respectively -5 mol/L、7.5×10 -6 mol/L、5.0×10 - 6 mol/L、2.5×10 -6 mol/L、1.0×10 -6 mol/L、7.5×10 -7 mol/L、5.0×10 -7 mol/L、2.5×10 -7 mol/L、1.0×10 -7 mol/L、5.0×10 -8 mol/L、1.0×10 -8 Raman scattering spectra of 11 melamine aqueous solutions in mol/L were calculated for melamine molecules 703cm -1 RSD and I with absolute peak intensity of characteristic peak R Is a RSD of (c). It can be found that reference I R The RSD of the melamine molecules at different concentrations is reduced to within 10% and at 10% -6 M-10 -8 Within the M concentration range, I R Has good linear relation with the melamine concentration, takes the melamine concentration as the abscissa, I R Fitting the ordinate to obtain the linear function relation; FIG. 6 shows a schematic diagram of Raman scattering spectra of melamine molecules at different concentrations, FIG. 7 shows the difference between the concentration of melamine molecules and I R And fitting to obtain a linear function relation diagram.
(7) Respectively dripping melamine water solution and milk solution with unknown concentration onto the substrate, finding the substrate under a microscope, selecting laser with wavelength of 532nm, laser power of 0.16mW and integration time of 10s, repeatedly collecting Raman scattering spectrum signals of melamine molecules at different positions on the substrate, and reading 703cm of melamine molecules -1 The absolute peak intensity of the characteristic peak and 1380cm of citrate ion -1 Absolute peak intensity of the characteristic peak at the position, and calculating I R The average value was obtained. And reversely estimating the concentration of melamine in the solution in the linear function relation according to the value, comparing the concentration with the actual concentration, and calculating to obtain the detection recovery rates of the melamine water solution and the milk solution which can reach 103% and 112% respectively. Fig. 8 shows a schematic diagram of raman scattering spectra obtained by detecting melamine aqueous solutions, milk solutions and blank milk solutions of unknown concentrations. Fig. 9 shows a schematic diagram of reverse-speculated melamine concentration in an unknown aqueous melamine solution and in a milk solution. Fig. 10 shows an analytical chart for estimating the recovery rate obtained during the course of the melamine aqueous solution and the milk solution of unknown concentration.
The melamine molecule 703cm in the substrate is detected by adopting the melamine quantitative detection method based on the surface enhanced Raman spectrum internal standard analysis provided by the embodiment of the invention -1 The absolute peak intensity of the characteristic peak and 1380cm of citrate ion -1 The absolute peak intensity of the characteristic peak is used for calculating the relative intensity I by using citrate ions adsorbed on the silver nanoparticle aggregate as an internal standard R And fitting the melamine concentration to I R Thereby eliminating peak intensity fluctuation of the object to be detected caused by non-uniformity of the SERS substrate and experimental conditionsWe found that at various concentrations (10 -5 M-10 -8 M)，I R Is less than 10% and at 10 -6 M-10 -8 Within the M concentration range, I R Has good linear relation with the concentration of melamine. Furthermore, we examined the melamine content (7.5X10) in the aqueous solution and the milk sample, respectively -7 M), the detection recovery rate can reach 103% and 112% respectively. The internal standard analysis method provides a feasible scheme for quantitative detection of melamine in liquid.
Finally, it should be pointed out that: the above embodiments are only for illustrating the technical solution of the present invention, and are not limiting. Those of ordinary skill in the art will appreciate that: the technical schemes described in the foregoing embodiments may be modified or some of the technical features may be replaced equivalently; such modifications and substitutions do not depart from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims (4)

1. The melamine quantitative detection method based on the surface enhanced Raman spectrum internal standard analysis is characterized by comprising the following steps of:
step 1, preparing a silver nanoparticle aggregate with internal standard molecules by a laser-induced reduction method as an SERS substrate; comprising the following steps:
uniformly mixing a silver nitrate solution and a sodium citrate solution to obtain a reaction solution, placing the reaction solution on conductive glass, and covering a cover glass above the reaction solution; converging laser at the interface of the reaction solution and the conductive glass, and forming silver nanoparticle aggregates at the interface through laser-induced reduction after irradiation for preset time; washing the formed silver nanoparticle aggregate with deionized water, and removing unreacted solution to obtain silver nanoparticle aggregate adsorbed with citrate as SERS substrate, wherein the citrate is the internal standard molecule;
step 2, obtaining the ratio of the characteristic peak of the melamine molecule to the characteristic peak of the sodium citrate molecule in the SERS signal as the relative intensity of the characteristic peak of the melamine molecule, and fitting the concentration of the melamine solution and the relative intensity I R A functional relationship between; comprising the following steps:
detecting the surface enhanced Raman scattering spectrum of melamine molecules in the aqueous solution on the SERS substrate to obtain a spectrum of 703cm -1 Characteristic peak of melamine molecule and 1380cm -1 Characteristic peaks of citrate ions;
1380cm of the citrate ion -1 The characteristic peak is used as an intensity internal standard to obtain the relative intensity I of the characteristic peak of the melamine molecule R The method comprises the steps of carrying out a first treatment on the surface of the The relative intensity I R Is smaller than the melamine molecular characteristic peak I 703cm -1 Is a relative standard deviation of (2);
detecting SERS signals of melamine water solutions at different concentrations, and fitting the functional relation according to the obtained relative intensities;
and step 3, dropwise adding a melamine solution with unknown concentration onto the SERS substrate, repeatedly collecting SERS signals at different positions, calculating the relative intensity average value, and reversely estimating the concentration of melamine in the solution according to the functional relation.
2. The method according to claim 1, characterized in that the relative standard deviation of the relative intensities of the characteristic peaks of the melamine molecules is less than 10%.
3. The method according to claim 1, wherein when the concentration of the aqueous melamine solution is 10 -6 M-10 - 8 In the M range, the relative intensity is linearly related to the concentration of the aqueous melamine solution.
4. A method according to claim 3, characterized in that if the melamine solution is an aqueous solution, the detected recovery of melamine comprises 103%; if the melamine solution is a milk solution, the detection recovery rate of melamine comprises 112%.
CN202210072658.8A 2022-01-21 2022-01-21 Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis Active CN114441503B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202210072658.8A CN114441503B (en) 2022-01-21 2022-01-21 Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202210072658.8A CN114441503B (en) 2022-01-21 2022-01-21 Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis

Publications (2)

Publication Number Publication Date
CN114441503A CN114441503A (en) 2022-05-06
CN114441503B true CN114441503B (en) 2023-07-28

Family

ID=81367656

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202210072658.8A Active CN114441503B (en) 2022-01-21 2022-01-21 Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis

Country Status (1)

Country Link
CN (1) CN114441503B (en)

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115746360B (en) * 2022-11-24 2023-12-12 无锡学院 Flexible surface-enhanced Raman scattering substrate with adjustable gap, and preparation method and application thereof

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103411950A (en) * 2013-06-24 2013-11-27 吉林大学 Method for detecting tripolycyanamide in milk based on surface-enhanced Raman activity chip
CN104122246A (en) * 2013-04-28 2014-10-29 同方威视技术股份有限公司 Raman-spectrum measuring method for detecting content of melamine in milk products with different matrixes
CN104568909A (en) * 2015-02-10 2015-04-29 吕志伟 Method for detecting content of dicyandiamide in milk based on surface enhanced Raman scattering technology
WO2021057513A1 (en) * 2019-09-25 2021-04-01 暨南大学 Phenol recognition sers probe, preparation thereof, use thereof, and sers-based universal ultrasensitive immunoassay method

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2010057212A1 (en) * 2008-11-17 2010-05-20 Oxonica Materials, Inc. Melamine assay methods and systems

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104122246A (en) * 2013-04-28 2014-10-29 同方威视技术股份有限公司 Raman-spectrum measuring method for detecting content of melamine in milk products with different matrixes
CN103411950A (en) * 2013-06-24 2013-11-27 吉林大学 Method for detecting tripolycyanamide in milk based on surface-enhanced Raman activity chip
CN104568909A (en) * 2015-02-10 2015-04-29 吕志伟 Method for detecting content of dicyandiamide in milk based on surface enhanced Raman scattering technology
WO2021057513A1 (en) * 2019-09-25 2021-04-01 暨南大学 Phenol recognition sers probe, preparation thereof, use thereof, and sers-based universal ultrasensitive immunoassay method

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
采用增强拉曼检测技术对牛奶中三聚氰胺的检测;陈安宇;焦义;刘春伟;崔子健;郭浔;;中国卫生检验杂志(第08期);全文 *

Also Published As

Publication number Publication date
CN114441503A (en) 2022-05-06

Similar Documents

Publication Publication Date Title
Drolc et al. Nitrate and nitrite nitrogen determination in waste water using on-line UV spectrometric method
Fontana et al. Raman spectroscopic sensors for inorganic salts
US20170199123A1 (en) Detection method of heavy metal ions and sensor using the same
CN114441503B (en) Melamine quantitative detection method based on surface enhanced Raman spectrum internal standard analysis
TWI592651B (en) Metal ion detection equipment and metal ion detection method
EP2939263B1 (en) Determination of free hydrogen fluoride levels in a buffered oxide etchant
EP2982967A1 (en) Method and system for real time in-situ monitoring of a solution during a solution based process
CN108303388B (en) Method for in-situ quantitative characterization of complex organic matter and metal ion complexing process
Al Bratty Visible spectrophotometric determination of chlorpheniramine maleate and diphenhydramine hydrochloride in raw and dosage form using potassium permanganate
US8932874B2 (en) Control over ammonium fluoride levels in oxide etchant
CN114486852B (en) Method for detecting target molecule
Bilba et al. Determination of trace amounts of palladium (II) by solid-phase spectrophotometry
Yang et al. Development of an iodine sensor based on fluorescence energy transfer
Kenney et al. Applications of fourier-transform infrared spectroscopy in geomicrobiology
RU2391659C1 (en) Method of detecting silver using polymethacrylate matrix
Ou-Yang et al. Resonance light scattering of 1-hydroxypyrene-ethyl violet-anionic surfactant system and its analytical application
Wójcik et al. A new approach in voltammetric profiling of wines and whiskies based on a useful faradaic signal component
Zarei et al. Simultaneous Spectrophotometric Determination of Aluminum and Iron in Micellar Media by Using the H‐Point Standard Addition Method
Periolatto et al. Routine Monitoring of Trace Arsenic in Water by Lab-on-a-chip Technology: a Preliminary Study
Sung et al. Direct and simultaneous determination of Co and Cu on a silicon wafer using a chemiluminescence system
US9677998B2 (en) Control over etching molecule levels in etching compositions
CN114636686A (en) SERS quantitative working curve method based on intensity ratio
Isoe et al. Determination of trace silica in highly purified water based on delayed quenching of Rhodamine B through nanoparticle formation with molybdosilicate
Rajan Spectrophotometric Estimation of Oxolamine citrate in Bulk and Pharmaceutical Dosage Form by Second Order Derivative Method
CN104132923B (en) The detection method of light chrome yellow concentration in one heavy metal species concentrated acid liquid

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant