Preparation of surface plasmon coupled heterogeneous composite membrane structure and application of surface plasmon coupled heterogeneous composite membrane structure in amikacin detection

Abstract

The invention discloses preparation of a surface plasmon coupled heterogeneous composite film structure and application of the surface plasmon coupled heterogeneous composite film structure in amikacin detection, and belongs to the field of analysis and detection. AgNPs are adsorbed through electrostatic interaction by utilizing surface amination of a PDMS film to form an AgNPs-PDMS-AgNPs film, the AgNPs-PDMS-AgNPs film covers and is overlaid on an Au film to construct a heterostructure, the thickness of the PDMS film is regulated and controlled, and the surface plasmon coupled heterogeneous composite film structure is obtained. The AgNPs thin film on the surface and the surface plasmon of the heterostructure AgNPs / Au thin film can generate a coupling effect, so that the SERS detection sensitivity of amikacin on the AgNPs thin film on the surface is improved, and the detection limit of amikacin is reduced. The amikacin detection limit is 3.79 * 10 <-11 > M, the linear detection range is 10 <-5 >-10 <-10 > M, and the relative standard deviation of SERS intensity is less than 10%. The method disclosed by the invention has the characteristics of high sensitivity and convenience in operation in SERS (Surface Enhanced Raman Scattering) detection.

Description

Technical Field

[0001] The present invention relates to the field of analysis and detection, and in particular to the preparation of a surface plasmon-coupled heterogeneous composite film structure and its application in amikacin detection. Background Art

[0002] Amikacin is a commonly used aminoglycoside antibiotic, and its mechanism of action is to inhibit the synthesis of bacterial proteins. It has good antibacterial activity against most Enterobacteriaceae, Pseudomonas aeruginosa, some other Pseudomonas, Alcaligenes, etc. It is clinically used for anti-infection treatment. Amikacin has a broad antibacterial spectrum and strong antibacterial activity, and has significant bactericidal effects on a variety of bacteria. However, the toxic side effects of amikacin are manifested as damage to the kidneys, hearing, nerve and muscle paralysis, and damage to the absorption function of the intestine. Long-term consumption of such livestock products with excessive residues will directly cause harm to humans. Therefore, in order to ensure food safety, GB 31650-2019 "National Food Safety Standard Maximum Residue Limits of Veterinary Drugs in Food" stipulates that the maximum residue limits (MRLs) of amikacin drugs are 100 μg / kg. Establishing rapid screening and detection of amikacin residues in livestock products is of great significance to maintaining human dietary health.

[0003] Currently reported detection methods for similar antibiotics such as amikacin mainly include microbiological method, high performance liquid chromatography, fluorescence method, chemiluminescence method, capillary electrophoresis method, etc. Among them, the microbiological method is complicated to operate, has low sensitivity and is time-consuming. Due to the lack of chromophores in the structure of aminoglycoside antibiotics, the fluorescence method often requires the use of fluorescent reagents or other reagents for derivatization before determination, which is often subject to photobleaching and interference. The operation method is cumbersome and there are many experimental interference factors. The high performance liquid chromatography operation process is complicated and the detection time is long. It is not suitable for rapid screening of large quantities of samples. Summary of the invention

[0004] The purpose of the present invention is to solve the above-mentioned problems in the prior art and provide the preparation of a surface plasmon coupled heterogeneous composite film structure and its application in the detection of amikacin. By preparing a heterogeneous composite film structure and adjusting the thickness of the PDMS film, the surface plasmons of the AgNPs film and the heterogeneous structure AgNPs / Au film will be coupled, thereby increasing the degree of mutual coupling of the surface plasmons and enhancing the Raman spectrum of amikacin in the AgNPs film. The PDMS film thickness and the concentration of silver nanoparticles of the AgNPs film are optimized. Under the conditions of the optimal PDMS film thickness and silver nanoparticle concentration, the quantitative analysis of amikacin is achieved by establishing a standard curve of the intensity and concentration of the SERS characteristic peak. The linear detection range of amikacin is 10 -5 -10 -10 M, the detection limit is 3.79×10 -11The relative standard deviation of the SERS intensity is less than 10%. The heterogeneous composite membrane structure prepared by the present invention has high sensitivity, good universality and practical prospects.

[0005] In order to achieve the above object, the present invention adopts the following technical scheme:

[0006] The preparation of a surface plasmon coupled heterogeneous composite film structure comprises the following steps:

[0007] 1) AgNPs are adsorbed on the PDMS membrane by electrostatic action to form an AgNPs film, thereby obtaining an AgNPs-PDMS-AgNPs film;

[0008] 2) Au thin film is deposited on the glass sheet by vacuum evaporation;

[0009] 3) The AgNPs-PDMS-AgNPs film is covered and superimposed on the Au film to obtain a heterogeneous composite film structure;

[0010] In step 1), the PDMS membrane is subjected to hydrophilization and amination treatments.

[0011] In step 1), the thickness of the PDMS membrane is 100-500 μm.

[0012] The thickness of the Au film is 1-2 μm.

[0013] The application of the surface plasmon coupled heterogeneous composite film structure in SERS detection of amikacin.

[0014] The amikacin sample solution was dripped onto the AgNPs film on the heterogeneous composite membrane structure for SERS detection. The SERS characteristic peak position of amikacin was different from that of other aminoglycoside antibiotics of the same type, so qualitative analysis was performed. A standard curve was established between the intensity value of the SERS characteristic peak of amikacin and the concentration of amikacin for quantitative analysis.

[0015] The characteristic SERS peak of amikacin is at 930 cm -1 The SERS characteristic peaks are shown at .

[0016] The linear detection range of amikacin is 10 -5 ~10 -10 M, the detection limit is 3.79×10 -11 M.

[0017] The relative standard deviation of the SERS intensity detected at the same concentration of the present invention is less than 10%.

[0018] Compared with the prior art, the technical solution of the present invention has the following beneficial effects:

[0019] The present invention is based on the surface plasmon coupling phenomenon of the heterogeneous composite film structure, which greatly improves the SERS intensity of amikacin adsorbed on the AgNPs film. The heterogeneous composite film structure prepared by the present invention has the advantages of simple and convenient operation, low cost and good reproducibility in detection. The experimental results show that the linear detection range of amikacin is 10 -5 ~10 -10 M, the detection limit is 3.79×10 -11 M, the relative standard deviation of SERS intensity is less than 10%. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 A in the figure is PDMS membrane. Figure 1 B is the AgNPs film. Figure 1 C in the middle is an optical photograph of AgNPs-PDMS-AgNPs film covered with superimposed Au film.

[0021] Figure 2 This is the SERS spectrum of the detection of amikacin solution dropped on the AgNPs-PDMS-AgNPs film covered with the superimposed Au film.

[0022] Figure 3 Optical photographs of PDMS films with different thicknesses.

[0023] Figure 4 A is the SEM image of AgNPs film. Figure 4 B in the middle is the particle size histogram of silver nanoparticles on the film.

[0024] Figure 5 SERS spectra of amikacin on AgNPs-PDMS-AgNPs film covered with Au film under different PDMS film thickness conditions; from top to bottom a→c, a is 100 μm PDMS film, b is 300 μm PDMS film, and c is 500 μm PDMS film.

[0025] Figure 6 The SERS spectra of amikacin on the AgNPs-PDMS-AgNPs film covered with the Au film under different silver nanoparticle concentration conditions; from top to bottom a→c, a is the original solution, b is the concentrated solution, and c is the diluted solution.

[0026] Figure 7 A in the middle is the SERS spectrum of amikacin detected on the AgNPs-PDMS-AgNPs film covered with the Au film, which is paralleled ten times. Figure 7 B is the detection of amikacin on the AgNPs-PDMS-AgNPs film covered with the Au film. The SERS spectrum at 930 cm-1 is paralleled ten times. -1Relative standard deviation of SERS peak intensity at .

[0027] Figure 8 A in the middle is the SERS spectra of different concentrations of amikacin detected on the AgNPs-PDMS-AgNPs film covered with the Au film. Figure 8 B is amikacin at 930 cm -1 Linear relationship between SERS intensity at and negative logarithm of concentration.

[0028] Fig. 9 SERS spectra of amikacin and other aminoglycoside antibiotics (tobramycin, sisomicin, gentamicin, streptomycin) solutions detected on AgNPs-PDMS-AgNPs film covered with Au film. DETAILED DESCRIPTION

[0029] In order to make the technical problems, technical solutions and beneficial effects to be solved by the present invention clearer and more understandable, the present invention is further described in detail below in conjunction with the accompanying drawings and embodiments.

[0030] The present invention comprises the following steps:

[0031] 1) Preparation and modification of polydimethylsiloxane (PDMS) membrane.

[0032] 2) AgNPs are adsorbed on the PDMS membrane by electrostatic action to form an AgNPs film.

[0033] The first step is to synthesize the silver nanoparticle solution by using trisodium citrate and ascorbic acid method, and to obtain a silver nanoparticle solution with uniform particle size by controlling the concentration of reactants, reaction temperature and reaction time. The second step is to immerse the amino PDMS membrane in the silver nanoparticle solution, and to adsorb the silver nanoparticles on the PDMS membrane through electrostatic adsorption to form an AgNPs-PDMS-AgNPs film.

[0034] 3) AgNPs-PDMS-AgNPs film is covered with Au film to construct a heterogeneous composite film structure. The Au film is a 1 μm Au film coated on a glass plate by vacuum evaporation.

[0035] The AgNPs film is superimposed on the Au film, and since PDMS is flexible, the two can fit closely together. The heterogeneous composite film structure is a composite of the surface AgNPs film and the heterogeneous structure AgNPs / Au film.

[0036] 4) SERS quantitative analysis and detection of amikacin on the AgNPs film on the surface of the composite membrane.

[0037] Ultrapure water was used as solvent to prepare different concentrations of 10 -5 ~10 -10 M of amikacin solution; at an excitation wavelength of 785 nm, different concentrations of amikacin solution were added and SERS detection was performed. The SERS characteristic peak 930 cm in the spectrum was recorded. -1 The intensity values ​​were compared with the corresponding concentrations of amikacin to establish a standard curve for quantitative analysis. According to the different SERS characteristic peaks of amikacin, tobramycin, sisomicin, gentamicin, and streptomycin, the strongest SERS peak of amikacin was 930 cm -1 The strongest SERS peaks of tobramycin, sisomicin, gentamicin, and streptomycin are at 1550 cm -1 In addition, the I of amikacin 1020 / I 1120 The intensity ratio of aminoglycosides and other antibiotics is greater than 1. 1020 / I 1120 The intensity ratio was less than 1, achieving specific detection of amikacin.

[0038] The standard curve of the quantitative analysis detection is to establish a standard curve with the logarithmic value of the series concentration of the standard amikacin as the horizontal coordinate and the peak intensity at the SERS characteristic peak as the vertical coordinate; the linear detection range of the standard is 10 -5 ~10 -10 M, the detection limit is 3.79×10 -11 The relative standard deviation of the SERS intensity detected at the same concentration was less than 10%.

[0039] By adjusting the thickness of the PDMS film, the surface AgNPs film and the surface plasmon of the heterostructured AgNPs / Au film are coupled to improve the sensitivity of SERS detection of amikacin on the surface AgNPs film. The SERS intensity is increased by 2 times, and the detection limit of amikacin is reduced to 3.79×10 -11 M.

[0040] Specific examples are given below.

[0041] Example 1

[0042] 1) Preparation and modification of polydimethylsiloxane (PDMS) membrane.

[0043] In the first step, the Sylgard 184 prepolymer and the catalyst are fully mixed in a suitable mass ratio of 10:1, cured in an oven, and cut into the required thickness. In the second step, the PDMS is immersed in a piranha solution for hydrophilization; then immersed in an APTES ethanol solution for amino treatment. Specifically, the concentration ratio of the piranha solution is (H2SO4:H2O2 = 3:1), and the PDMS membrane is immersed in a 30°C piranha solution for 30 s to obtain a hydroxylated PDMS membrane; the APTES ethanol solution is a 5% APTES ethanol solution, and the hydroxylated PDMS membrane is placed in a 5% APTES ethanol solution and heated in an oven at 70°C for 3h to prepare a positively charged polydimethylsiloxane film.

[0044] Figure 1 A in the figure is an optical photograph of the PDMS membrane.

[0045] Figure 3 Optical photographs of PDMS films with different thicknesses. From left to right, the thicknesses are 100, 300, and 500 μm.

[0046] 2) AgNPs are adsorbed on the PDMS membrane by electrostatic action to form an AgNPs film.

[0047] The first step is to synthesize the silver nanoparticle solution by using trisodium citrate and ascorbic acid method, and by controlling the concentration of reactants, reaction temperature and reaction time, a silver nanoparticle solution with uniform particle size is obtained.

[0048] In the second step, the amino-modified PDMS membrane is immersed in a silver nanoparticle solution, and the silver nanoparticles are adsorbed on the PDMS membrane through electrostatic adsorption to form an AgNPs-PDMS-AgNPs film.

[0049] Figure 1 B is the optical photograph of AgNPs-PDMS-AgNPs film.

[0050] Specifically, in this embodiment, the mass of trisodium citrate is 50 mg, the mass of ascorbic acid is 100 mg, and the two are dissolved in 400 mL of ultrapure water at the same time; the concentration of the silver nitrate solution is 0.01 mM, and the volume is 1 mL; the reaction temperature is 80° C. in a water bath; and the reaction time is 40 min.

[0051] The AgNPs film is formed by electrostatic adsorption on the PDMS film. 10 mL of silver nanoparticle solution is taken and the amino PDMS film is immersed in the silver nanoparticle solution. The silver nanoparticle solution is negatively charged and the PDMS film is positively charged, and a silver nanoparticle film is formed by electrostatic adsorption. The formed silver nanoparticle film will reflect silver-gray light. The formed silver nanoparticle film is rinsed with ultrapure water and dried with nitrogen for later use.

[0052] 3) AgNPs-PDMS-AgNPs film is covered with Au film to construct a heterostructure. The Au film is made by vacuum evaporation to deposit 1 μm Au on a glass plate.

[0053] Figure 1 C in the middle is an optical photograph of AgNPs-PDMS-AgNPs film covered with superimposed Au film.

[0054] 4) Detection of amikacin on AgNPs-PDMS-AgNPs film covered with Au film.

[0055] Ultrapure water was used as solvent to prepare 10 -5 M amikacin solution, AgNPs-PDMS-AgNPs film was covered on the Au film, and 10 -5 M of amikacin solution for SERS detection.

[0056] Figure 2 This is the SERS spectrum of the detection of amikacin solution dropped on the AgNPs-PDMS-AgNPs film covered with the superimposed Au film. It can be seen that the SERS signal is enhanced compared to when there is no heterostructure.

[0057] 5) SEM characterization of silver nanoparticle film.

[0058] Figure 4 A in the middle is a SEM image of a silver nanoparticle film, and the inset is a SEM image of a silver nanoparticle film at 500 nm. Figure 4 B in the middle is the particle size histogram of the silver nanoparticles on the film. It can be seen that the silver nanoparticles are spherical with a particle size of about 46.07±11.34nm.

[0059] Example 2

[0060] SERS detection of amikacin on AgNPs-PDMS-AgNPs film covered with Au film under different PDMS film thickness conditions.

[0061] The concentration of amikacin solution was fixed at 10 -5M, PDMS films with thicknesses of 100, 300 and 500 μm were modified to prepare silver nanoparticle films; the AgNPs-PDMS-AgNPs film was covered on the Au film, and the amikacin solution was dropped to collect the SERS spectra. Figure 5 It can be seen that when the thickness of the PDMS film is 100 μm, the SERS intensity of amikacin reaches the strongest. Therefore, in subsequent experiments, a 100 μm thick AgNPs-PDMS-AgNPs film was used to cover the Au film to detect amikacin.

[0062] Example 3

[0063] SERS detection of amikacin on AgNPs-PDMS-AgNPs film covered with Au film under different silver nanoparticle concentration conditions.

[0064] The concentration of amikacin solution was fixed at 10 -5 M, the thickness of PDMS was adjusted to 100 μm, and the amino-modified PDMS membrane was immersed in the synthesized silver nanoparticle solution stock solution, stock solution concentrate, and stock solution dilution, respectively, taken out, rinsed with ultrapure water, dried with nitrogen gas for later use, and SERS spectra were collected. Figure 6 It can be seen that when the stock solution is used, the SERS intensity of amikacin reaches the highest, so the optimal silver nanoparticle concentration of the amikacin detection system in subsequent experiments is the stock solution concentration synthesized in the above steps.

[0065] Example 4

[0066] The uniformity of amikacin was detected on AgNPs-PDMS-AgNPs film covered with Au film.

[0067] The AgNPs-PDMS-AgNPs film was covered on the stacked Au film, and amikacin solution was added. The SERS spectra were randomly collected ten times on the substrate with an excitation wavelength of 785 nm. The SERS peak intensities of amikacin collected ten times in parallel were compared to determine the uniformity of the SERS substrate.

[0068] Figure 7 A in the middle is the SERS spectrum of amikacin measured ten times in parallel; Figure 7 B is amikacin at 930 cm -1 The SERS peak intensity bar graph was obtained by measuring the SERS peak intensity ten times in parallel. It can be seen that the relative standard deviation (RSD) calculated for the characteristic SERS peak intensity of 10 sets of parallel samples is 5.33%, which is less than 10%, indicating that the uniformity of the SERS substrate is good and can be applied to the SERS quantitative analysis of amikacin.

[0069] Example 5

[0070] SERS quantitative analysis and detection of amikacin on AgNPs-PDMS-AgNPs film covered with superimposed Au film.

[0071] Ultrapure water was used as solvent to prepare different concentrations of 10 -5 ~10 -10 M of amikacin solution, AgNPs-PDMS-AgNPs film was covered on the Au film, and different concentrations of amikacin solution were added for SERS detection. The SERS characteristic peak 930 cm in the spectrum was recorded. -1 The intensity values ​​were used to establish a standard curve with the corresponding amikacin concentration for quantitative analysis.

[0072] Figure 8 As can be seen in Figure A, the SERS intensity of amikacin decreases with decreasing concentration.

[0073] Figure 8 B is the standard curve established after testing the standard solutions of different concentrations of amikacin. -5 ~10 -10 M, amikacin 930 cm -1 The negative logarithmic relationship between the intensity of the characteristic peak and the concentration of amikacin, (y=2813.49x+29141.05), R 2 =0.9951. In addition, the detection limit is 3.79×10 -11 M.

[0074] Example 6

[0075] SERS-specific detection of amikacin and other homologues on AgNPs-PDMS-AgNPs film covered with superimposed Au film.

[0076] Ultrapure water was used as the solvent to prepare a 10 -5 M of amikacin (AMK), tobramycin (TOB), sisomicin (SISO), gentamicin (GEN), and streptomycin (STR) were dropped on the AgNPs-PDMS-AgNPs film covered with the stacked Au film for SERS detection.

[0077] Fig. 9 The experimental results of Example 6 show that the strongest SERS peak of amikacin is 930 cm -1 The strongest SERS peak of other aminoglycoside antibiotics is 1550 cm -1 Therefore, the composite PDMS silver nanoparticle film can specifically detect amikacin in amikacin and other aminoglycoside antibiotic solutions.

Claims

1. Preparation of a surface plasmon coupled heterogeneous composite film structure, characterized in that: The following steps are involved: 1) AgNPs are adsorbed on the PDMS membrane by electrostatic action to form an AgNPs film, thereby obtaining an AgNPs-PDMS-AgNPs film; 2) Au thin film is deposited on the glass sheet by vacuum evaporation; 3) The AgNPs-PDMS-AgNPs film is covered and stacked on the Au film to obtain a heterogeneous composite film structure.

2. The preparation of the surface plasmon coupled heterogeneous composite film structure according to claim 1, characterized in that: In step 1), the PDMS membrane is subjected to hydrophilization and amination treatments.

3. The preparation of the surface plasmon coupled heterogeneous composite film structure according to claim 1, characterized in that: In step 1), the thickness of the PDMS membrane is 100-500 μm.

4. The preparation of the surface plasmon coupled heterogeneous composite film structure according to claim 1, characterized in that: The thickness of the Au film is 1-2 μm.

5. A surface plasmon coupled heterogeneous composite film structure, characterized in that: Prepared as described in any one of claims 1 to 4.

6. Use of the surface plasmon coupled heterogeneous composite film structure according to claim 5 in SERS detection of amikacin.

7. The use according to claim 6, characterized in that: The amikacin sample solution was dripped onto the AgNPs film on the heterogeneous composite membrane structure for SERS detection. The SERS characteristic peak position of amikacin was different from that of other aminoglycoside antibiotics of the same type, so qualitative analysis was performed. A standard curve was established between the intensity value of the SERS characteristic peak of amikacin and the concentration of amikacin for quantitative analysis.

8. The use according to claim 7, characterized in that: The characteristic SERS peak of amikacin is at 930 cm -1 The SERS characteristic peaks are shown at .

9. The use according to claim 7, characterized in that: The linear detection range of amikacin is 10 -5 ~10 -10 M, the detection limit is 3.79×10 -11 M.

10. The use according to claim 7, characterized in that: The relative standard deviation of the detected SERS intensity at the same concentration is less than 10%.

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