A three-dimensional SERS substrate for detecting thiram and its preparation method

Abstract

The present invention discloses a preparation method for a three-dimensional SERS substrate for detecting thiram, comprising: growing zinc oxide nanorods on a polyvinylidene fluoride membrane to obtain a ZnO / P substrate; coating a gold film on the surface of the zinc oxide nanorods to obtain an Au / ZnO / P substrate; modifying the Au / ZnO / P substrate and adsorbing silver nanoparticles on the surface of the modified Au / ZnO / P substrate to obtain an Ag / Au / ZnO / P substrate, which is the three-dimensional SERS substrate. The present invention also provides the substrate obtained by the above preparation method. The present invention introduces bimetallic nanoparticles as enhancement materials, synergistically combines the respective advantages of Ag and Au, and utilizes the three-dimensional morphology of ZnO nanorods to generate rich SERS "hot spots" and enhancement effects. At the same time, due to the special heterointerface structure of the bimetallic material, a stronger coupling effect can be generated with thiram, realizing rapid in-situ high-sensitivity detection of thiram.

Description

Technical Field

[0001] The present invention relates to the technical field of surface enhanced Raman spectroscopy detection, and particularly relates to a three-dimensional SERS substrate for detecting thiram and a preparation method thereof. Background Art

[0002] With the wide application of pesticides such as thiram in modern agriculture and the discovery of their potential toxicity, their exposure in the environment has attracted increasing attention. It is reported that the adverse reactions of thiram include skin diseases, organ failure and skeletal dysplasia. In addition, the concentration of thiram in organisms can be further increased due to food chain enrichment, and even a small amount of residue can cause significant damage after long-term environmental exposure. In view of this, a variety of analytical techniques including high performance liquid chromatography, gas chromatography-mass spectrometry, chemiluminescence and fluorescence methods have been used for the monitoring of thiram in the environment. Although the results are relatively accurate, the above methods are still limited by defects such as complex operation, long processing time and high cost, which prompts researchers to develop more convenient and effective detection techniques.

[0003] Due to its non-invasive, ultrasensitive and fast advantages, surface enhanced Raman spectroscopy (SERS) technology has been developed for multiple fields such as environmental monitoring, biosensing, medical diagnosis, food safety, etc. The substrate material is the core of the development of SERS technology, as it determines the specificity and sensitivity of SERS detection. Based on zinc oxide (ZnO), due to its good biocompatibility and high resistance to changes in environmental pH or temperature, it has been used to prepare semiconductor / noble metal composite substrates, and due to its strong electromagnetic and chemical enhancement effects, the detection sensitivity of SERS has been greatly improved. At present, some studies have demonstrated that SERS substrates coupled with ZnO nanorods and noble metals have excellent performance in the quantitative analysis of thiram. For example, Quan et al. synthesized three-dimensional inclined ZMRs / Ag arrays on ITO substrates and found that they had excellent SERS sensitivity when detecting thiram. However, the SERS substrates involved in these studies are all rigid materials and are not suitable for non-planar in-situ detection in curved structures, which greatly limits their development in practical applications.

[0004] The pore size of polyvinylidene fluoride (PVDF) membrane is adjustable, and it has the characteristics of strong permeability, high chemical stability and easy modification. Therefore, it is developed for the research and development of SERS flexible substrates. Chen et al. in situ grew dense and uniform gold nanoparticles on the high-curvature surface of PVDF. The PVDF@Au nanofiber porous membrane can be used for the efficient detection of thiram (the detection limit is 0.1 nM). The results show that the PVDF membrane can be used as a good supporting material for manufacturing SERS substrates. Its inherent roughness is conducive to generating "hot spots", and the large surface area provides more adsorption sites for target molecules. Its excellent performance makes it a promising alternative matrix for rapid in-situ detection of environmental pollutants.

[0005] Chinese patent document CN114354572A discloses a three-dimensional flexible SERS substrate and a method for detecting putrescine and cadaverine based on this substrate. The substrate in situ chemically grows AuNPs on the surface of ZnO nanorods to prepare a three-dimensional rose-branch-like nanostructure. Among them, the three-dimensional rose-branch-like SERS substrate modified with p-MBA can achieve high detection sensitivity and rapid detection of putrescine and cadaverine in samples. However, the Raman enhancement effect of the single noble metal gold nanoparticles used in this patent is limited and still needs to be further improved. In addition, this type of substrate is suitable for putrescine and cadaverine, and has a low binding efficiency with the pollutant thiram, and cannot achieve high-sensitivity detection of thiram. Summary of the Invention

[0006] Therefore, the technical problem to be solved by the present invention is to provide a three-dimensional SERS substrate for detecting thiram and its preparation method to overcome the defects of complex operation, long processing time and high cost in the existing thiram detection.

[0007] For this purpose, the present invention adopts the following technical solutions:

[0008] The present invention provides a preparation method for a three-dimensional SERS substrate for detecting thiram, comprising the following steps:

[0009] S1: Grow zinc oxide nanorods on a polyvinylidene fluoride (PVDF) membrane to obtain a ZnO / P substrate;

[0010] S2: Coat a gold film on the surface of the zinc oxide nanorods to obtain an Au / ZnO / P substrate;

[0011] S3: Modify the Au / ZnO / P substrate, and adsorb silver nanoparticles on the surface of the modified Au / ZnO / P substrate to obtain an Ag / Au / ZnO / P substrate, which is the three-dimensional SERS substrate.

[0012] Further, the preparation method of the Ag nanoparticles is as follows: add 7-10 mg of silver nitrate into 50 mL of boiling glycerol-water mixture, then add 1-5 mL of trisodium citrate, continuously stir for 1-1.5 h under the boiling state to obtain a silver nanoparticle colloid with a particle size of 30±8 nm;

[0013] The volume percentage of glycerol in the glycerol-water mixture is 40%-50%;

[0014] The mass fraction of trisodium citrate in the trisodium citrate solution is 3%.

[0015] In step S3, the modification method is to immerse the Au / ZnO / P substrate into an ethanol solution of 10-20 mM modifier for 0.5-1 h, and then rinse it thoroughly with pure water;

[0016] The modifier is any one of mercaptoethylamine, 3-aminopropyltriethoxysilane or polyethyleneimine.

[0017] In step S3, the method of adsorbing Ag nanoparticles on the surface of the Au / ZnO / P substrate is to immerse the modified substrate into a 0.5-2 mM Ag nanoparticle colloid solution for 6-12 h and then take it out.

[0018] The growth method in step S1 is to immerse the PVDF membrane into a mixed solution of zinc nitrate hexahydrate and hexamethylenetetramine or a mixed solution of zinc acetate dihydrate, oleylamine and dodecanol, heat at 90-100 °C for 4-5 h to grow a grass-like zinc oxide nanorod array, the top diameter of the zinc oxide nanorods is 40±5 nm, the bottom diameter is 60±5 nm, and the length is 500±50 nm.

[0019] In the mixed solution of zinc nitrate hexahydrate and hexamethylenetetramine, the concentrations of zinc nitrate hexahydrate and hexamethylenetetramine are 25-30 mM respectively, and the solvent is water;

[0020] In the mixed solution of zinc acetate dihydrate, oleylamine and dodecanol, the concentrations of zinc acetate dihydrate, oleylamine and dodecanol are 25-30 mM respectively, and the solvent is water.

[0021] In step S2, the method of coating the gold film is to deposit a gold film with a thickness of 35-55 nm on the ZnO / P array at a speed of 1 nm / s using a magnetron sputtering machine under argon protection to obtain the Au / ZnO / P substrate.

[0022] Before step S1, it also includes the pretreatment of the PVDF membrane. The pretreatment is to soak the PVDF membrane in ethanol for 5-10 min, after cleaning, immerse it in a zinc acetate dihydrate ethanol solution or a mixed solution of zinc nitrate hexahydrate and sodium nitrate for 10-20 min, dry it with nitrogen, and finally heat it at 140-150 °C for 4-5 h.

[0023] In the zinc acetate dihydrate ethanol solution, the concentration of zinc acetate dihydrate is 25 - 30 mM, and the solvent is ethanol;

[0024] In the mixed solution of zinc nitrate hexahydrate and sodium nitrate, the concentrations of zinc nitrate hexahydrate and sodium nitrate are 25 - 30 mM respectively, and the solvent is water.

[0025] The present invention also provides a three - dimensional SERS substrate for detecting thiram, and the substrate is prepared by the above - mentioned preparation method.

[0026] The technical solution of the present invention has the following advantages:

[0027] (1) The three - dimensional SERS substrate obtained by the present invention further introduces bimetallic nanoparticles as enhancement materials on the basis of the existing technology. This can synergistically combine the plasma activity of Ag and the stability of Au. At the same time, the three - dimensional morphology of the clustered ZnO nanorods combines with the bimetallic nanoparticles to generate richer SERS "hot spots" and enhancement effects, while retaining the advantages of the PVDF membrane substrate such as high permeability, high chemical stability, and easy modification. Therefore, it has good development potential.

[0028] (2) The three - dimensional sea - cucumber - like Ag / Au / ZnO / P substrate in the present invention has good specificity for thiram. Due to its unique heterojunction interface structure, the coupling between the bimetallic material and thiram is stronger. At the same time, the lowest concentration of thiram that can be detected by this substrate can reach 0.48 ng / cm 2 , which is much lower than the concentration allowed by the national food safety standard (GB 2763 - 2021), and has good application prospects in the in - situ detection of environmental pollutants.

[0029] (3) The present invention limits the preparation method and particle size of Ag nanoparticles. The three - dimensional SERS substrate finally obtained by using the specific preparation method of the present application has better Raman intensity. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] In order to more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the following will briefly introduce the drawings required for the description of the specific embodiments or the prior art. Obviously, the drawings in the following description are some embodiments of the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0031] Figure 1 It is a schematic diagram of the preparation process of the three - dimensional SERS substrate of the present invention;

[0032] Figure 2Scanning electron microscope image of the three-dimensional SERS substrate obtained in Example 1 of the present invention;

[0033] Figure 3 SERS spectra of 4-MBA at different concentrations on the three-dimensional SERS substrate obtained in Example 1 of the present invention;

[0034] Figure 4 SERS intensity of 4-MBA at 1075 cm -1 at 20 randomly selected detection points;

[0035] Figure 5 SERS spectra of thiram at different concentrations on the three-dimensional SERS substrate obtained in Example 1 of the present invention;

[0036] Figure 6 Typical SERS spectra of thiram and interfering substances (both at 1×10 -5 M);

[0037] Figure 7 Detection of different concentrations of thiram on apple peels using the three-dimensional SERS substrate obtained in Example 1 of the present invention;

[0038] Figure 8 Raman signal intensities of thiram on the substrate of Example 1 and the substrate of Comparative Example 1 at different time points in Test Example 6;

[0039] Figure 9 Raman signal intensities of the substrate of Example 1 and the substrate of Comparative Example 1 with thiram in the saturated state in Test Example 6.

[0040] Figure 10 Raman signal intensities of thiram on the substrate of Example 1 and the substrate of Comparative Example 2 in Test Example 7.

[0041] Figure 11 Raman signal intensities of thiram on the substrate of Example 1 and the substrate of Comparative Example 3 in Test Example 8. Detailed implementation method

[0042] The following examples are provided to better understand the present invention further. They are not limited to the best implementation mode, and do not constitute a limitation to the content and protection scope of the present invention. Any product identical or similar to the present invention obtained by anyone under the inspiration of the present invention or by combining the features of the present invention with those of other prior arts falls within the protection scope of the present invention.

[0043] For those without specific experimental steps or conditions indicated, the operations or conditions of the conventional experimental steps described in the literature in this field can be followed. For reagents or instruments without the manufacturer indicated, they are all conventional reagent products that can be obtained through commercial purchase.

[0044] Example 1

[0045] This example provides a method for preparing a three-dimensional SERS substrate, and its preparation process is as Figure 1 shown, and the specific steps are as follows:

[0046] (1) Add 50 mL of glycerol-water mixture (40% glycerol, v / v) to a 250 mL round-bottom flask and heat it to boiling; add 9 mg of silver nitrate to the boiling solution, and then add 1 mL of trisodium citrate (3%, w / v). Stir the reaction mixture continuously at the boiling state for 1 h, and the color of the solution will quickly change from colorless to yellow and finally to dark red. After the reaction is completed, wait for the colloid to cool to room temperature and store the colloid at 4 °C.

[0047] (2) Immerse a PVDF membrane with a size of 2×2 cm 2 in ethanol for 5 min, clean it, then immerse the PVDF membrane in a 30 mM zinc acetate dihydrate ethanol solution for 10 min, dry it with nitrogen, and then heat it at 146 °C for 4 h. The obtained PVDF membrane is then immersed in a mixed solution of 25 mM zinc nitrate hexahydrate and hexamethylenetetramine and heated at 90 °C for 4 h to obtain a grass-like zinc oxide nanorod array (ZnO / P).

[0048] (3) Put the obtained ZnO / P substrate into a magnetron sputtering machine and evacuate it; under the protection of Ar 2 , the magnetron sputtering machine deposits a 45 nm thick gold film on the ZnO / P array at a speed of 1 nm / s to obtain an Au / ZnO / P substrate.

[0049] (4) Immerse the obtained Au / ZnO / P substrate in a 10 mM ethanethiolamine ethanol solution for 1 h and then rinse it with water. Immerse the ethanethiolamine-modified substrate in the 1 mM Ag nanoparticle colloid solution obtained in step (1) for 10 h and then take it out. After thoroughly cleaning it with pure water, an Ag / Au / ZnO / P substrate, that is, the three-dimensional SERS substrate, is obtained. The electron microscopy image results of the obtained sample are as Figure 2 shown, and the Ag nanoparticles are very closely coupled to the surface of the Au film, and the substrate generates uniformly dispersed high-density SERS hot spots.

[0050] Example 2

[0051] This example provides a method for preparing a three-dimensional SERS substrate, and the specific steps are as follows:

[0052] (1) Add 50 mL of glycerol - water mixture (50% glycerol, v / v) to a 250 - mL round - bottom flask and heat it to boiling; add 8 mg of silver nitrate to the boiling solution, and then add 3 mL of sodium citrate (3%, w / v). Continuously stir the reaction mixture at boiling for 1 h. The color of the solution will quickly change from colorless to yellow and finally to dark red. After the reaction is completed, let the colloid cool to room temperature and store the colloid at 4 °C.

[0053] (2) Immerse a PVDF membrane with a size of 2×2 cm 2 in ethanol for 6 min, clean it, and then immerse the PVDF membrane in a mixed solution of 30 mM zinc nitrate hexahydrate and sodium nitrate for 10 min, dry it with nitrogen, and then heat it at 140 °C for 5 h. The obtained PVDF membrane is then immersed in a mixed solution of 30 mM zinc acetate dihydrate, oleylamine, and dodecanol and heated at 100 °C for 5 h to obtain a grass - like zinc oxide nanorod array (ZnO / P).

[0054] (3) Place the obtained ZnO / P substrate into a magnetron sputtering machine and evacuate it; under Ar 2 protection, the magnetron sputtering machine deposits a 55 - nm - thick gold film on the ZnO / P array at a speed of 1 nm / s to obtain an Au / ZnO / P substrate.

[0055] (4) Immerse the obtained Au / ZnO / P substrate in a 10 - mM ethanol solution of 3 - aminopropyltriethoxysilane for 0.5 h and then rinse it with water. Immerse the substrate modified with mercaptoethylamine in the 2 - mM Ag nanoparticle colloid solution obtained in step (1) for 12 h, then take it out and thoroughly wash it with pure water to obtain an Ag / Au / ZnO / P substrate, which is the three - dimensional SERS substrate.

[0056] Example 3

[0057] This example provides a method for preparing a three - dimensional SERS substrate, and the specific steps are as follows:

[0058] (1) Add 50 mL of glycerol - water mixture (50% glycerol, v / v) to a 250 - mL round - bottom flask and heat it to boiling; add 10 mg of silver nitrate to the boiling solution, and then add 5 mL of sodium citrate (3%, w / v). Continuously stir the reaction mixture at boiling for 1 h. The color of the solution will quickly change from colorless to yellow and finally to dark red. After the reaction is completed, let the colloid cool to room temperature and store the colloid at 4 °C.

[0059] (2) Immerse a PVDF membrane with a size of 2×2 cm 2The PVDF membrane was soaked in ethanol for 5 min, cleaned, and then immersed in a 25 mM zinc acetate dihydrate ethanol solution for 10 min, dried with nitrogen, and then heated at 150 °C for 5 h. The obtained PVDF membrane was then immersed in a mixture of 30 mM zinc nitrate hexahydrate and hexamethylenetetramine, and heated at 95 °C for 5 h to obtain a grass-like zinc oxide nanorod array (ZnO / P).

[0060] (3) The obtained ZnO / P substrate was placed in a magnetron sputtering machine and evacuated; under Ar 2 protection, the magnetron sputtering machine deposited a 35-nm-thick gold film on the ZnO / P array at a speed of 1 nm / s to obtain an Au / ZnO / P substrate.

[0061] (4) The obtained Au / ZnO / P substrate was immersed in a 20 mM mercaptoethylamine ethanol solution for 0.5 h, rinsed with water, and the mercaptoethylamine-modified substrate was immersed in the 0.5 mM Ag nanoparticle colloidal solution obtained in step (1) for 6 h, then taken out and thoroughly cleaned with pure water to obtain an Ag / Au / ZnO / P substrate, namely the three-dimensional SERS substrate.

[0062] Comparative Example 1

[0063] The substrate BigAu / Au / ZnO / P obtained in Example 1 of Chinese Patent Document CN114354572A was used.

[0064] Comparative Example 2

[0065] This comparative example provides a three-dimensional SERS substrate, which is different from Example 1 in that the preparation method of Ag nanoparticles in step (1) is changed to obtain Ag nanoparticles by ultraviolet light irradiation reduction method. The preparation process is to photodeposit Ag from a 1 mM silver nitrate aqueous solution using a low-intensity ultraviolet fluorescent lamp (λmax = 365 nm, power density 5 mW / cm 2 ) for 40 min. The obtained Ag nanoparticles have a particle size controlled to be 30 ± 8 nm.

[0066] Comparative Example 3

[0067] This comparative example provides a three-dimensional SERS substrate, which is different from Example 1 in that trisodium citrate in step (1) is changed to ascorbic acid to obtain Ag nanoparticles with a diameter of 60 ± 5 nm. The method is as follows: 50 mL of a glycerol-water mixture (40% glycerol, v / v) was added to a 250 mL round-bottom flask and heated to boiling; 9 mg of silver nitrate was added to the boiling solution, and then 1 mL of ascorbic acid (3%, w / v, dissolved in water) was added. The reaction mixture was continuously stirred at the boiling state for 1 h. After the reaction was completed, the colloid was cooled to room temperature and stored at 4 °C.

[0068] Experimental Example 1

[0069] 4-Mercaptobenzoic acid (4-MBA) was used as a Raman probe molecule to test the uniformity, sensitivity and stability of the three-dimensional SERS substrate obtained in Example 1 of the present invention; the three-dimensional SERS substrate obtained in Example 1 was immersed in a 4-MBA ethanol solution, and then dried at room temperature. Raman signal acquisition was performed using a Renishaw Raman spectrometer. The laser wavelength used was 785 nm, the integration time was 10 s, and the acquisition power was 3.5×10 -6 W~1.4×10 -4 W. The specific implementation steps are as follows:

[0070] (1) Take a prepared SERS substrate, cut it into small substrates of the same size, and immerse them in 4-MBA ethanol solutions with different concentrations (10 -6 -10 -11 M) for 30 min. After drying, collect their Raman signals and compare their signal intensities. Measure the SERS spectra of 4-MBA with different concentrations on the substrate. As Figure 3 shown, it was observed that as the concentration of 4-MBA increased, the Raman peak intensities at 1075 cm -1 and 1587 cm -1 also increased. When the concentration decreased to 10 -11 M, the characteristic peaks of 4-MBA could still be distinguished. It shows that the three-dimensional SERS substrate obtained in Example 1 of the present invention has a wide detection range and high sensitivity.

[0071] (2) Take 20 random points on the same substrate to detect the SERS response of 10 -6 M 4-MBA and compare their signal intensities. As Figure 4 shown, the Raman peak intensity at 1075 cm -1 was basically stable. The calculated relative standard deviation was 4.29%, which met the allowable range of SERS detection error. It shows that the Ag / Au / ZnO / P substrate has good uniformity, which is closely related to the three-dimensional ordered Ag / Au / ZnO / P heterostructure and the inherent uniformity of the PVDF membrane.

[0072] Experimental Example 2

[0073] The three-dimensional SERS substrate obtained in Example 1 of the present invention was used to detect thiram molecules; the SERS substrate was immersed in a thiram ethanol solution, and then dried at room temperature. Raman signal acquisition was performed using a Renishaw Raman spectrometer. The laser wavelength used was 785 nm, the integration time was 10 s, and the acquisition power was 3.5×10 -6 W~1.4×10 -4W. The specific implementation steps are as follows:

[0074] Take a prepared SERS substrate, cut it into small substrates of the same size, and immerse them in thiram ethanol solutions with different concentrations (10 -5 -10 -11 M) for 30 min. After drying, collect their Raman signals and compare their signal intensities. The detection mechanism of thiram involved is that when thiram molecules interact with the metal surface, a resonance radical structure is formed, resulting in the cleavage of the disulfide bond (S-S) of thiram. This phenomenon generates two dimethyl residues through strong chemisorption of chemical ligands on the surface of AgNPs, causing changes in the SERS spectrum. Therefore, the intensity of the Raman peak increases with the increase in the concentration of thiram. The characteristic peak at 1380 cm -1 (attributed to C-N stretching and symmetric CH 3 deformation), having the strongest intensity, is selected for subsequent detection. Measure the SERS spectra of thiram molecules with different concentrations on the substrate. As Figure 5 shown, it is observed that as the concentration of thiram molecules increases, the intensity of the Raman peak at 1380 cm -1 also increases. When the concentration decreases to 10 -10 M, the characteristic peak of thiram molecules can still be detected.

[0075] Test Example 3

[0076] Test the selectivity of the three-dimensional SERS substrate obtained in Example 1 of the present invention for thiram molecules. The specific implementation steps are as follows:

[0077] (1) Take a prepared SERS substrate and cut it into small substrates of the same size. Select dinotefuran, atrazine, acetamiprid, sulfamethoxazole, and ciprofloxacin as interfering substances and use them together with thiram for detection. The concentration of all pollutants is 10 -5 M.

[0078] (2) As Figure 6 shown, only thiram molecules have recognizable Raman spectral peaks, and no Raman peaks are found for other pollutants. And compared with the signal intensity of thiram molecules, the signal intensity of the mixture does not change significantly. In other words, the addition of other pollutants has little effect on the detection of thiram. The results prove that this Ag / Au / ZnO / P flexible substrate has excellent selectivity and is suitable for the selective detection of thiram in complex systems.

[0079] Test Example 4

[0080] Use the three-dimensional SERS substrate obtained in Example 1 of the present invention to actually detect thiram in actual samples. The specific implementation steps are as follows:

[0081] (1) Apples purchased from a local supermarket were cleaned separately with deionized water and ethanol. Solutions of thiram in ethanol with different concentrations were sprayed onto the apple skins, and then they were dried in air.

[0082] (2) Subsequently, the SERS substrate pre-wetted with ethanol was pasted onto the apple skin surface and kept for 10 s to complete the capture of the analyte before SERS measurement, and then it was peeled off for subsequent SERS detection.

[0083] (3) The results are as Figure 7 shown. Even when the concentration was as low as 0.48 ng / cm 2 , the characteristic peak of thiram at 1380 cm -1 could still be observed, and the detection limit was much lower than the allowable limit of apple skin (2×10 -6 g / cm 2 ). The results indicate that the three-dimensional sea cucumber-like Ag / Au / ZnO / P flexible substrate can be successfully applied to the extraction and rapid detection of pollutants in real environments.

[0084] Test Example 5

[0085] The detection limits of thiram by different methods are shown in Table 1:

[0086] Table 1 Detection limits (LODs) of thiram by different methods

[0087]

[0088] As shown in the above table, the SERS technology of the Ag / Au / ZnO / P substrate in this application shows the most prominent performance, superior to the vast majority of traditional substrates. These results indicate that the Ag / Au / ZnO / P SERS substrate provides a powerful tool for the trace detection of thiram in real samples.

[0089] Test Example 6

[0090] For thiram with the same concentration (10 -5 M), the substrates obtained in Example 1 of this application and the substrate of Comparative Example 1 were used for detection. As Figure 8 and Figure 9 shown, the substrate Ag / Au / ZnO / P in Example 1 of this application has a better enhancement effect. Moreover, using substrates of the same size, the reaction of the Ag / Au / ZnO / P substrate with thiram in solution reached saturation within 30 min, and increasing the reaction time could no longer improve the Raman intensity; however, the reaction of the BigAu / Au / ZnO / P substrate with thiram took 1 h or more to reach saturation ( Figure 8 ), and the signal was relatively weak ( Figure 9)。It shows that due to the unique heterogeneous interface structure of the bimetallic material, the reaction with thiram will be more rapid and effective, and the three-dimensional sea cucumber-like Ag / Au / ZnO / P substrate in the present invention has better specificity for thiram.

[0091] Test Example 7

[0092] Test the Raman intensities of the substrates obtained in Test Example 1 and Comparative Example 2.

[0093] The results are as Figure 10 shown. It can be seen that the Raman intensity of the substrate prepared with Ag particles synthesized on the substrate by the ultraviolet reduction method in Comparative Example 2 is lower than that of the substrate synthesized in Example 1 of the present application.

[0094] Test Example 8

[0095] Test the Raman intensities of the substrates obtained in Test Example 1 and Comparative Example 3.

[0096] The results are as Figure 11 shown. It can be seen that the Raman intensity of the substrate prepared with larger-sized Ag particles in Comparative Example 3 is lower than that of the substrate synthesized in Example 1 of the present application.

[0097] Obviously, the above embodiments are merely examples given for clear illustration and are not limitations on the implementation manners. For those of ordinary skill in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation manners here. And the obvious changes or modifications derived therefrom still fall within the protection scope of the present invention.

Claims

1. A preparation method for a three-dimensional SERS substrate for detecting thiram, characterized in that, it comprises the following steps: S1: Grow zinc oxide nanorods on a polyvinylidene fluoride membrane to obtain a ZnO / P substrate; S2: Coating a gold film on the surface of the zinc oxide nanorods to obtain an Au / ZnO / P substrate; S3: Modify the Au / ZnO / P substrate, and adsorb silver nanoparticles on the surface of the modified Au / ZnO / P substrate to obtain an Ag / Au / ZnO / P substrate, which is the three-dimensional SERS substrate; The growth method in step S1 is to immerse the polyvinylidene fluoride membrane in a mixed solution of zinc nitrate hexahydrate and hexamethylenetetramine or a mixed solution of zinc acetate dihydrate, oleylamine and dodecanol, and heat at 90 - 100 °C for 4 - 5 h to grow a grass-like zinc oxide nanorod array. The top diameter of the zinc oxide nanorods is 40 ± 5 nm, the bottom diameter is 60 ± 5 nm, and the length is 500 ± 50 nm.

2. The preparation method according to claim 1, characterized in that, the preparation method of the silver nanoparticles is to add 7 - 10 mg of silver nitrate to 50 mL of a boiling glycerol-water mixture, then add 1 - 5 mL of trisodium citrate solution, and continuously stir for 1 - 1.5 h under boiling conditions to obtain a silver nanoparticle colloid with a particle size of 30 ± 8 nm; the volume percentage of glycerol in the glycerol-water mixture is 40% - 50%; the mass fraction of trisodium citrate in the trisodium citrate solution is 3%.

3. The preparation method according to claim 1 or 2, characterized in that, in step S3, the modification method is to immerse the Au / ZnO / P substrate in an ethanol solution of a modifier with a concentration of 10 - 20 mM for 0.5 - 1 h, and then rinse it clean with pure water; the modifier is any one of mercaptoethylamine, 3-aminopropyltriethoxysilane or polyethyleneimine.

4. The preparation method according to claim 1, characterized in that, in step S3, the method of adsorbing Ag nanoparticles on the surface of the Au / ZnO / P substrate is to immerse the modified substrate in a 0.5 - 2 mM Ag nanoparticle colloid solution for 6 - 12 h and then take it out.

5. The preparation method according to claim 1, characterized in that, in the mixed solution of zinc nitrate hexahydrate and hexamethylenetetramine, the concentrations of zinc nitrate hexahydrate and hexamethylenetetramine are 25 - 30 mM respectively, and the solvent is water; in the mixed solution of zinc acetate dihydrate, oleylamine and dodecanol, the concentrations of zinc acetate dihydrate, oleylamine and dodecanol are 25 - 30 mM respectively, and the solvent is water.

6. The preparation method according to claim 1, characterized in that, in step S2, the coating method of the gold film is to coat a gold film with a thickness of 35 - 55 nm on the ZnO / P array at a speed of 1 nm / s using a magnetron sputtering machine under argon protection to obtain an Au / ZnO / P substrate.

7. The preparation method according to claim 1, characterized in that, Before step S1, it also includes the pretreatment of the polyvinylidene fluoride membrane. The pretreatment is to soak the polyvinylidene fluoride membrane in ethanol for 5 - 10 min, clean it, then immerse it in a zinc acetate dihydrate ethanol solution or a mixed solution of zinc nitrate hexahydrate and sodium nitrate for 10 - 20 min, dry it with nitrogen, and finally heat it at 140 - 150 °C for 4 - 5 h.

8. According to the preparation method described in claim 7, characterized in that, in the zinc acetate dihydrate ethanol solution, the concentration of zinc acetate dihydrate is 25 - 30 mM, and the solvent is ethanol; in the mixed solution of zinc nitrate hexahydrate and sodium nitrate, the concentrations of zinc nitrate hexahydrate and sodium nitrate are 25 - 30 mM respectively, and the solvent is water.

9. A three-dimensional SERS substrate for detecting thiram, characterized in that, the substrate is prepared by the preparation method described in any one of claims 1 - 8.

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