Heterogeneous solvent confinement enrichment SERS (Surface Enhanced Raman Scattering) detection method based on acoustic suspension

Abstract

The invention relates to the technical field of Raman spectrum detection, and discloses a heterogeneous solvent confinement enrichment SERS (Surface Enhanced Raman Scattering) detection method based on acoustic suspension. The method comprises the following steps: S1, suspending a liquid drop I containing an SERS substrate at a sound pressure node; s2, injecting a liquid drop II containing a target analyte to form a liquid drop with a core-shell structure; s3, double-side heating is performed to accelerate solvent evaporation, so that the target analyte is enriched in the SERS substrate in a confinement manner, and an enriched sample is obtained; s4, collecting the enriched sample, and carrying out SERS detection; wherein the liquid drop I and the liquid drop II are immiscible phases. According to the method provided by the invention, a three-layer sandwich structure can be formed by mediating the heterogeneous solvent and regulating and controlling the volume ratio of the oil phase and the water phase, a target analyte is confined in an interlayer or a gap of the SERS substrate, a hot-spot enrichment region is formed, a confined enrichment effect is achieved, the solute concentration is improved by three orders of magnitude, and the method is suitable for all high-Raman active molecules and has a wide application prospect. And the universality is high.

Description

Technical Field

[0001] This invention relates to the field of Raman spectroscopy detection technology, and in particular to a method for detecting SERS using heterogeneous solvent confinement based on acoustic levitation. Background Technology

[0002] Surface-enhanced Raman spectroscopy (SERS), a rapidly developing molecular vibrational analysis technique, boasts advantages such as high sensitivity, non-destructive nature, speed, and fingerprint recognition, and has been widely applied in fields such as food safety, environmental monitoring, and biomedicine. However, in practical applications, the sensitivity and selectivity of SERS detection remain limited due to low concentrations of target analytes in samples and interference from complex matrices. To overcome these issues, sample pretreatment is typically required to enrich the target analytes and lower the detection limit.

[0003] Acoustic levitation refers to the use of the interaction between sound wave radiation pressure and gravity, buoyancy, etc., to stably suspend tiny objects in the air. As a non-contact sample processing technology, it has received widespread attention in recent years. During the suspension process, droplets vibrate and deform under the action of sound waves, causing changes in the internal material distribution of the droplets and accelerating solvent evaporation, thereby achieving the concentration of the target substance.

[0004] Studies have shown that non-contact sample manipulation using acoustic levitation can achieve non-destructive enrichment of analytes in any phase. However, traditional acoustic levitation can only handle single-phase solvents (pure water or pure oil) and cannot efficiently enrich analytes whose polarity does not match that of the solvent. Furthermore, single-phase evaporation can only form randomly aggregated nanoparticles with low and uneven hot spot density. During single-phase open evaporation, analytes may diffuse with the solvent flow to the edge of the droplet and escape. Unilateral heating can also easily lead to a large temperature gradient in the droplet, causing nanoparticles to migrate to the cold zone and the assembly to break down. Summary of the Invention

[0005] The present invention aims to at least solve one of the aforementioned technical problems existing in the prior art. Therefore, the object of the present invention is to provide a method for detecting SERS using heterogeneous solvent confinement enrichment based on acoustic levitation.

[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows: This invention provides a method for detecting SERS based on acoustic levitation and confined enrichment of heterogeneous solvents, comprising the following steps: S1. Suspend droplet I containing the SERS substrate at the acoustic pressure node; S2. Inject droplet II containing the target analyte to form a core-shell structured droplet; S3. Double-sided heating accelerates solvent evaporation, confining and enriching the target analyte within the SERS substrate to obtain an enriched sample. S4. Collect the enriched sample and perform SERS detection; Wherein, droplet I and droplet II are immiscible phases.

[0007] In some embodiments of the present invention, the volume ratio of droplet I to droplet II is 1:(0.1-0.9); the volume of the core-shell structured droplet is 1-12 μL.

[0008] In some embodiments of the present invention, droplet I and droplet II are independently selected from an aqueous phase and an oil phase, respectively, and are different phases; the aqueous phase includes water; the oil phase includes alkanes with more than 5 carbon atoms.

[0009] In some preferred embodiments of the present invention, the oil phase comprises cyclohexane.

[0010] In some preferred embodiments of the present invention, the suspension of droplet I is achieved by adjusting the instrument's operating voltage and output power, with the operating voltage being 1-15V and the output power being 50-200W.

[0011] In some embodiments of the present invention, the target analyte in droplet II is a compound having a Raman-active group.

[0012] In some embodiments of the present invention, the target analyte is selected from oil-soluble or water-soluble substances.

[0013] In some embodiments of the present invention, the oil-soluble substance includes at least one of 4-mercaptophenylboronic acid (4-MBPA) and Alternaria solaniol (AOH).

[0014] In some embodiments of the present invention, the water-soluble substance includes at least one of rhodamine 6G (R6G), patulin (PAT), and methylene blue (MB).

[0015] In some embodiments of the present invention, when the target analyte is an oil-soluble substance, droplet I is an aqueous phase, droplet II is an oil phase, and the SERS substrate is selected from noble metal nanoparticles.

[0016] In some embodiments of the present invention, when the target analyte is a water-soluble substance, droplet I is an oil phase, droplet II is an aqueous phase, and the SERS substrate is selected from hydrophobically modified noble metal nanoparticles.

[0017] In some embodiments of the present invention, the noble metal nanoparticles are selected from gold nanoparticles or silver nanoparticles.

[0018] In some preferred embodiments of the present invention, the noble metal nanoparticles are selected from gold nanorods, gold nanospheres, and gold nanotriangles.

[0019] In some preferred embodiments of the present invention, the noble metal nanoparticles are gold nanorods, and the aspect ratio of the gold nanorods is (2-20):1.

[0020] In some embodiments of the present invention, the hydrophobically modified noble metal nanoparticles are prepared by hydrophobically modifying noble metal nanoparticles with fluorothiols.

[0021] In some preferred embodiments of the present invention, the fluorothiols include 1H,1H,2H,2H-perfluoro-1-decylthiols.

[0022] In some embodiments of the present invention, the hydrophobically modified noble metal nanoparticles are prepared by a method comprising the following steps: The hydrophobically modified noble metal nanoparticles were prepared by immersing them in a fluorothiol solution.

[0023] In some embodiments of the present invention, the concentration of the fluorothiol solution is 0.1-10 mmol / L.

[0024] In some embodiments of the present invention, the solvent of the fluorothiol solution includes at least one of isopropanol and n-hexane.

[0025] In some embodiments of the present invention, the soaking time is 8-24 hours.

[0026] In some embodiments of the present invention, the temperature of the double-sided heating is 30-80°C; the time is 5-60 minutes.

[0027] In some preferred embodiments of the present invention, the dual-sided heating includes the use of a ceramic heating lamp; the distance between the ceramic heating lamp and the core-shell structure droplet is 2-10 cm; the power is 25-100 W.

[0028] The basic principles of this invention are explained as follows: The core principle of this invention is to achieve three-dimensional confined enrichment of target analytes on a SERS substrate through a heterogeneous solvent system (oil / water two-phase) in an acoustic suspension environment, significantly improving detection sensitivity. Specifically: 1) Heterogeneous solvent system (confined enrichment basis): Using immiscible two-phase solvents, the structure is controlled by volume ratio. If the analyte is water-soluble (e.g., R6G), the oil phase volume is larger than the water phase volume, forming a water-in-oil (O / W) emulsion, in which the water-soluble target analyte is encapsulated in "water droplets" inside the oil phase. If the analyte is oil-soluble (e.g., 4-MBPA), the aqueous phase volume is larger than the oil phase volume, forming an oil-in-water (W / O) emulsion, in which the oil-soluble target analyte is encapsulated in "oil droplets" inside the aqueous phase. Phase separation is driven by interfacial tension between the two phases, forming a core-shell structure, which provides spatial constraints for subsequent confined enrichment. 2) Directional modification of SERS substrate (signal enhancement core): If the analyte is water-soluble, use a hydrophobic substrate that is soluble in the oil phase; if the analyte is oil-soluble, use a hydrophilic substrate that is soluble in the aqueous phase. Ensure that the substrate is compatible with the solvent and is directionally assembled at the interface between the two phases. 3) Acoustic levitation technology (dynamic enrichment platform): It suspends droplets at the acoustic pressure node and supports them with acoustic radiation force, realizing non-contact control. It uses droplet vibration and deformation to accelerate the migration of internal substances and promote the enrichment of target substances to the interface; at the same time, it eliminates the adsorption interference of the container wall and avoids SERS substrate contamination. 4) Heating-assisted solvent evaporation (concentration-driven): Double-sided heating is used to accelerate solvent evaporation, which concentrates the droplet volume from the μL level to the nL level, increasing the concentration of the target substance by 3 orders of magnitude; 5) Confined enrichment mechanism (key to improved sensitivity): During solvent evaporation, the SERS substrate self-assembles into a vertically stacked bilayer structure at the droplet interface. When heterogeneous nanoparticles are added, a three-layer "sandwich" structure is formed through volume control. The heterogeneous nanoparticles are oriented within the gaps between the two layers of the SERS substrate. During the actual enrichment of the target analyte, the hydrophilicity of the target analyte is opposed to that of the SERS substrate. The target analyte will be confined within the interlayers or gaps of the SERS substrate, forming a "hot spot" enrichment area.

[0029] Compared with the prior art, the beneficial effects of the present invention are: This invention provides a heterogeneous solvent-confined enrichment SERS detection method based on acoustic levitation. It utilizes acoustic levitation technology to promote the enrichment of target analytes at the interface, and non-contact sample processing avoids substrate contamination. Using a heterogeneous solvent as a medium, a three-layer "sandwich" structure can be formed by adjusting the oil-water phase volume ratio, confining the target analyte within the layers or gaps of the SERS substrate, creating a "hot spot" enrichment region, achieving a confined enrichment effect and increasing solute concentration by three orders of magnitude. Dual-sided heating accelerates solvent evaporation, driving concentration and forming a uniform "hot spot" region. This method is applicable to all highly Raman-active molecules. By adjusting the volume of the water and oil phases, it can achieve confined enrichment SERS detection of both water-soluble and oil-soluble targets, demonstrating strong universality. Attached Figure Description

[0030] Figure 1 This is a graph showing the enrichment test results of single-phase suspended droplets in Example 1; Figure 2 This is a graph showing the enrichment test results of immiscible two-phase suspension droplets in Example 1; Figure 3 This is a SEM image of the nanomaterial assembly structure in the aqueous phase mediated by single-phase solvent enrichment technology in Example 2. Figure 4 This is a diagram of the assembly structure of nanomaterials in the oil phase mediated by single-phase solvent enrichment technology in Example 2; Figure 5 The diagram shows the assembly of nanomaterials mediated by solvent enrichment technology in the same phase (A1) and different phase (A2) solvents in Example 2, and the SEM images of the assembled structures in the same phase (B) and different phase (C) solvents. Figure 6 SEM images of multi-nanomaterial assembly structures mediated by heterogeneous solvent enrichment technology; Figure 7 A schematic diagram of a heterogeneous solvent confined enrichment SERS detection method based on acoustic suspension; Figure 8 The diagram shows the detection of water-soluble analytes in Example 3 (A), and the SERS spectra of R6G under different detection modes (B) and at 1508 cm⁻¹. -1 The SERS signal (C) at the location; Figure 9 The diagram shows the detection of oil-soluble analytes in Example 3 (A), and the SERS spectra of 4-MBPA under different detection modes (B) and at 1071 cm⁻¹. -1 The SERS signal (C) at the location; Figure 10 The SERS spectra of R6G detected in the single-phase solvent enrichment mode (A), heterogeneous solvent confined enrichment mode (B), and heterogeneous solvent open domain enrichment mode (C) in Example 4 are shown, as well as the standard curves of R6G detected in the single-phase solvent enrichment mode (D), heterogeneous solvent confined enrichment mode (E), and heterogeneous solvent open domain enrichment mode (F). Figure 11 The SERS spectra of 4-MBPA detected in the single-phase solvent enrichment mode (A), heterogeneous solvent confined enrichment mode (B), and heterogeneous solvent open domain enrichment mode (C) in Example 5 are shown, as well as the standard curves of 4-MBPA detected in the single-phase solvent enrichment mode (D), heterogeneous solvent confined enrichment mode (E), and heterogeneous solvent open domain enrichment mode (F). Figure 12 The SERS spectra (A) and standard curve (B) of different concentrations of PAT detected in the heterogeneous solvent confinement enrichment mode in Example 6 are shown. Figure 13 The SERS spectra (A) and standard curve (B) of different concentrations of AOH detected in the heterogeneous solvent confinement enrichment mode in Example 7 are shown. Detailed Implementation

[0031] The present invention will be further described in detail below through specific embodiments. Unless otherwise specified, the raw materials, reagents, or apparatus used in the embodiments can be obtained from conventional commercial sources or by existing technical methods. Unless otherwise specified, the experimental or testing methods are conventional methods in the art.

[0032] Example 1 This embodiment provides verification of the behavior of the basic droplet in a heterogeneous solvent confined enrichment SERS detection method based on acoustic levitation: Enrichment of immiscible two-phase suspension droplets: 1) Add oil droplets to water droplets: Place 6μL of methylene blue aqueous solution in the acoustic levitation device (operating voltage 1-15V, power 50-200W) acoustic pressure node, and then inject 4μL of Sudan II cyclohexane solution into the droplet. Set the temperature of the double-sided ceramic heating lamps to 50℃ to accelerate solvent evaporation.

[0033] 2) Add water droplets to oil droplets: Place 6 μL of Sudan II cyclohexane solution in the acoustic levitation device (operating voltage 1-15V, power 50-200W) sound pressure node, and then inject 4 μL of methylene blue aqueous solution into the droplet. Set the temperature of the double-sided ceramic heating lamps to 50℃ to accelerate solvent evaporation.

[0034] Comparison using single-phase suspension droplet enrichment: 1) Water droplet enrichment: Place 10 μL of methylene blue aqueous solution in the acoustic levitation device (operating voltage 1-15V, power 50-200W) sound pressure node, and set the temperature of the double-sided ceramic heating lamps to 50℃ to accelerate solvent evaporation.

[0035] 2) Oil droplet enrichment: Place 10 μL of Sudan II cyclohexane solution in the acoustic levitation device (operating voltage 1-15V, power 50-200W) at the acoustic pressure node, and set the temperature of the double-sided ceramic heating lamps to 50℃ to accelerate solvent evaporation.

[0036] Figure 1 The image shows the enrichment test results of single-phase suspension droplets in Example 1. Figure 1 (A) in the figure represents the enrichment result of Sudan II cyclohexane solution. Figure 1 (B) in the figure represents the enrichment result of methylene blue aqueous solution, derived from... Figure 1It can be seen that 10µL of methylene blue aqueous solution and 10µL of Sudan II cyclohexane solution exhibit significant differences in evaporation characteristics in the acoustic field. The solvent evaporation rate of Sudan II cyclohexane solution is significantly higher than that of methylene blue aqueous solution. After solvent evaporation, both single-phase droplets are concentrated to a final volume of about 50nL, indicating that acoustic suspension technology can achieve sample concentration and enrichment. As the processing time increases, the droplet volume gradually decreases.

[0037] Figure 2 This is a graph showing the enrichment test results of immiscible two-phase suspension droplets in Example 1. Figure 2 (A) in the text refers to a solution of Sudan II cyclohexane added to an aqueous solution of methylene blue. Figure 2 (B) in the text refers to the addition of methylene blue aqueous solution to Sudan II cyclohexane solution. Figure 2 It was found that when 6 μL of methylene blue aqueous solution was placed at the acoustic pressure node, followed by the injection of 4 μL of Sudan II cyclohexane solution, phase separation immediately occurred due to the difference in polarity and interfacial tension between the two phases. The immiscible two-phase suspension droplets formed a clear bilayer interface, creating an oil-in-water core-shell structure. During reverse mixing, methylene blue and Sudan II also exhibited a clear bilayer interface, indicating that acoustic suspension can achieve the enrichment of immiscible two-phase suspension droplets. This verifies the phase separation characteristics of heterogeneous solvents and provides a foundation for subsequent confined enrichment.

[0038] Example 2 This embodiment provides a "sandwich" structure for nanomaterial assembly mediated by heterogeneous solvent enrichment to explain the mechanism of confined enrichment: 1) Preparation of gold nanorods: a. Transfer 5.0 mL of 100 mmol / L hexadecyltrimethylammonium bromide (CTAB) and 125 μL of 10 mmol / L HAuCl4·3H2O to a 25 mL Erlenmeyer flask, add 300 μL of 10 mmol / L NaBH4 under magnetic stirring, and incubate at 30 °C for 2 h to obtain the gold seed solution. b. Under the conditions of 30°C water bath and magnetic stirring (700 rpm), dissolve 0.7g CTAB and 0.1g sodium oleate in 25mL water and transfer to a 100mL Erlenmeyer flask. Add 960μL of 10mmol / L AgNO3 and react for 15min. Add 25mL of 1.0mmol / L HAuCl4 and react for 90min. Then add 1.8mL of 1mol / L HCl to adjust the pH to 3.0. c. 80 μL of 0.1 mol / L ascorbic acid was injected into the growth medium under magnetic stirring (1000 rpm), followed by rapid injection of 40 μL of gold seed solution. Stirring was stopped and the mixture was incubated overnight. Finally, the colloidal suspension was washed three times with a large amount of deionized water, the supernatant was discarded and dispersed in 5 mL of deionized water to obtain 5.0 mmol / L gold nanorods for later use. 2) After placing 6 μL of aqueous or oil-phase gold nanospheres at the acoustic pressure node of the acoustic levitation device, inject 4 μL of gold nanorods of a different phase from the suspended droplet into the droplet. That is, if the suspended droplet is aqueous gold nanospheres, inject oil-phase hydrophobic modified gold nanorods; if the suspended droplet is oil-phase hydrophobic modified gold nanospheres, inject aqueous gold nanorods. Set the temperature of the double-sided ceramic heating lamps to 50°C to accelerate solvent evaporation. After the water has completely evaporated, turn off the power of the levitation device and collect the sample. Hydrophobically modified gold nanorods or gold nanospheres are obtained by immersing gold nanorods or gold nanospheres in a hexane / isopropanol solution (0.1-10 mmol / L) of 1H,1H,2H,2H-perfluoro-1-decylthiol for 8-24 h to achieve hydrophobic modification.

[0039] The aforementioned nanomaterials can be substituted for each other, but it should be noted that the morphologies of the nanomaterials used at the same time must be different in order to observe the boundary structure between the nanomaterials.

[0040] As a comparison, a single-phase dissolution and enrichment technique was used to mediate the assembly of a "bilayer" structure from nanomaterials: 1) Place 10 μL of nanomaterial aqueous solution in the acoustic pressure node of the acoustic levitation device, set the temperature of the double-sided ceramic heating lamps to 50℃ to accelerate solvent evaporation, and after the water has completely evaporated, turn off the power of the levitation device and collect the sample.

[0041] 2) Place a 10 μL solution of nanomaterials in cyclohexane at the acoustic pressure node of the acoustic levitation device. Set the temperature of the double-sided ceramic heating lamps to 50°C to accelerate solvent evaporation. After the water has completely evaporated, turn off the power to the levitation device and collect the sample.

[0042] Among them, the nanomaterials are gold nanorods, gold nanospheres, gold nanotriangles, and hydrophobically modified gold nanorods, hydrophobically modified gold nanospheres, and hydrophobically modified gold nanotriangles.

[0043] Figure 3 The image shows a SEM image of the nanomaterial assembly structure in the aqueous phase mediated by single-phase solvent enrichment technology in Example 2. Figure 3 Images (a) and (b) in the image are SEM images of the assembled structure at different magnifications when gold nanospheres are used as the nanomaterial. Figure 3 Images (c) and (d) in the image show SEM images of the assembled structure at different magnifications when gold nanorods are used as the nanomaterial. Figure 3Images (e) and (f) in the figure show SEM images of the assembled structure at different magnifications when gold nanotriangles are used as nanomaterials. Figure 3 It can be seen that, using gold nanorods as a model system, after solvent evaporation, the gold nanorods aggregate around the droplets, ultimately achieving self-assembly of the gold nanorods. Unlike the traditional single-layer structure of liquid-liquid self-assembly, the acoustic suspension-mediated enrichment and assembly of nanomaterials forms a unique vertically stacked bilayer structure. When gold nanospheres and gold nanotriangles are used as nanomaterials, a vertically stacked bilayer structure is also obtained.

[0044] Figure 4 This is a diagram of the assembly structure of nanomaterials in the oil phase mediated by single-phase solvent enrichment technology in Example 2, wherein... Figure 4 Images (a) and (b) in the figure show SEM images of the assembled structure at different magnifications when hydrophobically modified gold nanospheres are used as nanomaterials. Figure 4 Images (c) and (d) in the figure show SEM images of the assembled structure at different magnifications when hydrophobically modified gold nanorods are used as nanomaterials. Figure 4 Images (e) and (f) in the figure show SEM images of the assembled structure at different magnifications when hydrophobically modified gold nanotriangles are used as nanomaterials. Figure 4 It can be seen that the nanomaterials in the oil phase still maintain similar assembly characteristics as those in the aqueous phase, indicating that the solvent type does not affect the enrichment and assembly of nanomaterials.

[0045] Figure 5 This is a schematic diagram of the assembly of nanomaterials mediated by solvent enrichment technology in homophase (A1) and heterophase (A2) solvents in Example 2, and SEM images of the assembled structures in homophase (B) and heterophase (C) solvents. Figure 5 It is evident that when gold nanorods are injected into an aqueous solution of suspended gold nanospheres, after solvent evaporation, the gold nanorods do not form a third orthogonally aligned structure within the gold nanospheres. Instead, they mix with the gold nanoparticles to form an independent bilayer structure again. When an aqueous solution of gold nanospheres is suspended at an acoustic pressure node, and then a hydrophobic gold nanorod solution is injected, the mixed droplets spontaneously form an oil-in-water emulsion structure under the regulation of interfacial tension. After solvent evaporation, the hydrophobic gold nanorods are confined within the interfacial space of the gold nanospheres, ultimately forming an oriented hydrophobic gold nanorod intermediate layer within the bilayer gap of the gold nanospheres, resulting in an independent three-layer "sandwich" structure.

[0046] Figure 6 SEM images of multi-nanomaterial assembly structures mediated by heterogeneous solvent enrichment technology, by Figure 6It can be seen that when the volume of the aqueous phase is greater than that of the oil phase, the system forms an oil-in-water emulsion, in which gold nanospheres in the aqueous phase encapsulate hydrophobic gold nanorods in the oil phase; when the volume of the oil phase is greater than that of the aqueous phase, the system transforms into a water-in-oil emulsion, in which hydrophobic gold nanorods in the oil phase encapsulate gold nanospheres in the aqueous phase; by adjusting the volumes of the water and oil phases, different target materials can be encapsulated and enriched, and this method successfully extends the two-dimensional assembly of nanomaterials to the three-dimensional dimension.

[0047] Example 3 This embodiment provides a heterogeneous solvent confined enrichment SERS detection method based on acoustic suspension. Figure 7 This is a schematic diagram of a heterogeneous solvent confined enrichment SERS detection method based on acoustic suspension. Figure 7 It can be seen that the method includes the following steps: When the target analyte is water-soluble, hydrophobically modified noble metal nanoparticles are used as the SERS substrate. The SERS substrate oil solution is placed at the acoustic pressure node, and then the target analyte aqueous solution is injected into the droplet. The volume ratio of the SERS substrate oil solution to the target analyte aqueous solution is 1:(0.1-0.9). When the target analyte is oil-soluble, noble metal nanoparticles are used as the SERS substrate. The SERS substrate aqueous solution is placed at the acoustic pressure node, and then the target analyte oil solution is injected into the droplet. The volume ratio of the SERS substrate aqueous solution to the target analyte oil solution is 1:(0.1-0.9). That is, in the heterogeneous solvent-mediated confined enrichment mode, the volume of the SERS substrate solution must be greater than the volume of the target analyte solution. The distance between the dual-sided ceramic heating lamps and the droplets is 2-10 cm, the power is 25-100 W, and the temperature is set to 30-80℃ to accelerate solvent evaporation. As the solvent evaporates, the concentration of the suspended droplets gradually increases. After the solution has completely evaporated, the power to the suspender is turned off, and the sample is collected onto the silicon wafer. The analyte can be either oil-soluble or water-soluble. For SERS analysis, a 5x objective lens, an excitation wavelength of 532 nm, a laser power of 10 mW, a grating line density of 600 gr / mm, a slit width of 200 μm, and an integration time of 5 s are selected, within a range of 200-2000 cm. -1 Data is collected within the specified range.

[0048] We set up comparative methods for SERS detection using heterogeneous solvent-mediated open-domain enrichment and single-phase system enrichment: Heterogeneous solvent-mediated open-domain enrichment mode SERS detection: The heterogeneous solvent-mediated open-domain enrichment mode is similar to the heterogeneous solvent-mediated confined enrichment mode, only requiring the volume of the SERS substrate solution to be smaller than the volume of the target analyte solution. Single-phase solvent enrichment mode SERS detection: A solution of noble metal nanoparticles in ethanol or a hydrophobically modified noble metal nanoparticle solution in ethanol is mixed with the target analyte solution at a volume ratio of 1:1. Then, 10 μL of the mixed solution is placed at the acoustic pressure node of an acoustic levitation device. The distance between the double-sided ceramic heating lamps and the droplet is 2-10 cm, the power is 25-100 W, and the temperature is set to 30-80℃ to accelerate solvent evaporation. As the solvent evaporates, the concentration of the suspended droplet gradually increases. After the solution has completely evaporated, the power to the levitation device is turned off, and the sample is collected onto a silicon wafer. The analyte can be either oil-soluble or water-soluble. For SERS analysis, a 5x objective lens, excitation wavelength of 532 nm, laser power of 10 mW, grating line density of 600 gr / mm, slit width of 200 μm, integration time of 5 s, and a wavelength of 200-2000 cm⁻¹ are used. -1 Data is collected within the specified range.

[0049] With 10 -9 Using mol / L R6G as a water-soluble probe molecule, and hydrophobically modified gold nanorods as the SERS substrate, heterogeneous solvent confined enrichment mode and heterogeneous solvent open domain enrichment mode were constructed by changing the oil phase / water phase volume ratio. The enrichment performance was studied by comparing them with the single-phase solvent enrichment mode.

[0050] Figure 8 The diagram shows the detection of water-soluble analytes in Example 3 (A), and the SERS spectra of R6G under different detection modes (B) and at 1508 cm⁻¹. -1 The SERS signal (C) at the location; by Figure 8 It can be seen that the three experimental groups are (1) single-phase solvent enrichment, (2) heterogeneous solvent confined enrichment and (3) heterogeneous solvent open domain enrichment. R6G SERS signals can be detected in all three modes. Quantitative comparison shows that the signal intensity of heterogeneous solvent confined enrichment mode (2) is 51.2% higher than that of single-phase solvent enrichment mode (1), while the signal intensity of heterogeneous solvent open domain enrichment mode (3) is 81.3% lower than that of heterogeneous solvent confined enrichment mode (2). This is because the water-in-oil structure formed by heterogeneous solvent confined enrichment mode can encapsulate the water-soluble target analyte inside. After the solvent evaporates, R6G is confined in the hydrophobic modified gold nanorod aggregates, which further achieves confined enrichment on the basis of single-phase solvent enrichment mode. In contrast, the heterogeneous solvent open domain enrichment mode forms an oil-in-water structure. After the solvent evaporates, the hydrophobic modified gold nanorods form aggregates, and R6G on the outer layer is difficult to enter the hydrophobic modified gold nanorod aggregates, resulting in a poorer enrichment effect.

[0051] With 10 -9mol / L 4-MBPA is an oil-soluble probe molecule, and gold nanorods are used as the SERS substrate. By changing the oil / water phase volume ratio, heterogeneous solvent confined enrichment mode and heterogeneous solvent open domain enrichment mode are constructed and compared with single-phase solvent enrichment mode to study their enrichment performance.

[0052] Figure 9 The diagram shows the detection of oil-soluble analytes in Example 3 (A), and the SERS spectra of 4-MBPA under different detection modes (B) and at 1071 cm⁻¹. -1 The SERS signal (C) at the location; by Figure 9 It can be seen that the three experimental groups are (1) single-phase solvent enrichment, (2) heterogeneous solvent confined enrichment, and (3) heterogeneous solvent open domain enrichment. 4-MBPA SERS signals can be detected in all three modes. Quantitative comparison shows that the signal intensity of heterogeneous solvent confined enrichment mode (2) is 98.3% higher than that of single-phase solvent enrichment mode (1), while the signal intensity of heterogeneous solvent open domain enrichment mode (3) is 68.9% lower than that of heterogeneous solvent confined enrichment mode (2). This indicates that the sample is trapped in the SERS substrate aggregate, which can achieve the confined enrichment effect and make its enrichment effect optimal. When the sample is an oil-soluble substance, the heterogeneous solvent confined enrichment mode forms an oil-in-water structure, which encapsulates the target analyte in its internal oil phase. After the solvent evaporates, 4-MBPA is confined in the gold nanorod aggregate. In contrast, the heterogeneous solvent open domain enrichment mode forms an oil-in-water structure. After the solvent evaporates, the gold nanorods form aggregates, and the 4-MBPA on the outer layer is difficult to enter the gold nanorod aggregate, resulting in a poorer enrichment effect.

[0053] Example 4 This embodiment provides a heterogeneous solvent confined enrichment SERS detection method based on acoustic suspension, applied to the detection of water-soluble substance R6G. The steps are as follows: S11. Place 6 μL of hydrophobic modified gold nanorod oil solution at the acoustic pressure node; S21. Then, 4 μL of R6G solution was injected into the droplet to form a core-shell structured droplet. S31. A ceramic heating lamp is used for double-sided heating to accelerate solvent evaporation. The distance between the ceramic heating lamp and the droplet is 2-10 cm, the power is 25-100 W, and the temperature is 30-80℃. As the solvent evaporates, the concentration of the suspended droplet gradually increases. R6G is confined and enriched in the hydrophobic modified gold nanorods to obtain the enriched sample. S41. After the solution has completely evaporated, turn off the power to the suspender and collect the sample onto the silicon wafer for SERS detection. For SERS analysis, select a 5x objective lens, excitation wavelength of 532nm, laser power of 10mW, grating line density of 600gr / mm, slit size of 200μm, integration time of 5s, and a wavelength between 200-2000cm. -1Data is collected within the specified range.

[0054] We set up SERS detection methods for open-domain enrichment in heterogeneous solvents and single-phase systems for comparison: SERS detection using open domain enrichment mode of heterogeneous solvent: The open domain enrichment mode of heterogeneous solvent is similar to the confined enrichment mode of heterogeneous solvent, only requiring the volume of the hydrophobic modified gold nanorod oil solution to be smaller than the volume of the R6G solution.

[0055] Single-phase solvent enrichment mode SERS detection: Hydrophobically modified gold nanorod ethanol solution and R6G solution were mixed at a volume ratio of 1:1. Then, 10 μL of the mixed solution was placed at the acoustic pressure node of an acoustic levitation device. The distance between the double-sided ceramic heating lamps and the droplet was 2-10 cm, the power was 25-100 W, and the temperature was set to 30-80℃ to accelerate solvent evaporation. As the solvent evaporated, the concentration of the suspended droplet gradually increased. After the solution was completely evaporated, the levitation device power was turned off, and the sample was collected onto a silicon wafer. For SERS analysis, a 5x objective lens, excitation wavelength of 532 nm, laser power of 10 mW, grating line density of 600 gr / mm, slit width of 200 μm, integration time of 5 s, and a wavelength of 200-2000 cm⁻¹ were selected. -1 Data is collected within the specified range.

[0056] Figure 10 The SERS spectra of R6G detected in Example 4 using the single-phase solvent enrichment mode (A), heterogeneous solvent confined enrichment mode (B), and heterogeneous solvent open-domain enrichment mode (C), and the standard curves of R6G detected using the single-phase solvent enrichment mode (D), heterogeneous solvent confined enrichment mode (E), and heterogeneous solvent open-domain enrichment mode (F) are provided by [the relevant source]. Figure 10 It can be seen that when R6G is detected in the three modes, its 1508cm -1 The characteristic peak SERS response values ​​are all positively correlated with concentration, and the logarithm of the characteristic peak signal intensity (log) I ) and the logarithm of concentration (log C The linear relationship is shown; the linear equations for detecting R6G in single-phase solvent enrichment mode, heterogeneous solvent confined enrichment mode, and heterogeneous solvent open domain enrichment mode are log [log(2000)], ... I =0.37log C +7.47, R=0.9994; log I =0.38log C +7.95, R=0.9969; log I =0.28log C +5.77, R=0.9960.

[0057] Example 5 This embodiment provides a heterogeneous solvent confined enrichment SERS detection method based on acoustic suspension, applied to the detection of the oil-soluble substance 4-MBPA. The steps are as follows: S11. Place 6 μL of gold nanorod aqueous solution at the acoustic pressure node; S21. Then, 4 μL of 4-MBPA cyclohexane solution was injected into the droplet to form a core-shell structured droplet. S31. A ceramic heating lamp is used for double-sided heating to accelerate solvent evaporation. The distance between the ceramic heating lamp and the droplet is 2-10 cm, the power is 25-100 W, and the temperature is 30-80 ℃. As the solvent evaporates, the concentration of the suspended droplet gradually increases. 4-MBPA is confined and enriched in the gold nanorod to obtain an enriched sample. S41. After the solution has completely evaporated, turn off the power to the suspender and collect the sample onto the silicon wafer for SERS detection. For SERS analysis, select a 5x objective lens, excitation wavelength of 532nm, laser power of 10mW, grating line density of 600gr / mm, slit size of 200μm, integration time of 5s, and a wavelength between 200-2000cm. -1 Data is collected within the specified range.

[0058] We set up SERS detection methods for open-domain enrichment in heterogeneous solvents and single-phase systems for comparison: Heterogeneous solvent-mediated open domain enrichment mode SERS detection: The heterogeneous solvent open domain enrichment mode is similar to the heterogeneous solvent confined enrichment mode, only requiring the gold nanorod oil solution volume to be adjusted to be less than the 4-MBPA solution volume.

[0059] Single-phase solvent enrichment mode SERS detection: Gold nanorod ethanol solution and 4-MBPA solution were mixed at a volume ratio of 1:1. Then, 10 μL of the mixed solution was placed at the acoustic pressure node of an acoustic levitation device. The distance between the double-sided ceramic heating lamps and the droplet was 2-10 cm, the power was 25-100 W, and the temperature was set to 30-80℃ to accelerate solvent evaporation. As the solvent evaporated, the concentration of the suspended droplet gradually increased. After the solution was completely evaporated, the power to the levitation device was turned off, and the sample was collected onto a silicon wafer. For SERS analysis, a 5x objective lens, excitation wavelength of 532 nm, laser power of 10 mW, grating line density of 600 gr / mm, slit width of 200 μm, integration time of 5 s, and a wavelength of 200-2000 cm⁻¹ were selected. -1 Data is collected within the specified range.

[0060] Figure 11 The SERS spectra of 4-MBPA detected in Example 5 using the single-phase solvent enrichment mode (A), heterogeneous solvent confined enrichment mode (B), and heterogeneous solvent open-domain enrichment mode (C), as well as the standard curves for 4-MBPA detection using the single-phase solvent enrichment mode (D), heterogeneous solvent confined enrichment mode (E), and heterogeneous solvent open-domain enrichment mode (F), are provided by... Figure 11 It can be seen that the detection of 4-MBPA under the three modes results in a value of 1071 cm⁻¹. -1The characteristic peak SERS response values ​​are all positively correlated with concentration, and the logarithm of the characteristic peak signal intensity (log) I ) and the logarithm of concentration (log C The linear relationship is shown; the linear equations for detecting R6G in single-phase solvent enrichment mode, heterogeneous solvent confined enrichment mode, and heterogeneous solvent open domain enrichment mode are log [log(2000)], ... I =0.32log C +6.87, R=0.9996; log I =0.34log C +7.37, R=0.9995; log I =0.31log C 6.26, R=0.9909.

[0061] Example 6 This embodiment provides a heterogeneous solvent confinement enrichment SERS detection method based on acoustic suspension, and its application in the detection of mycotoxins in food. The steps are as follows: S11. Place 6 μL of hydrophobic modified gold nanorod aqueous solution at the acoustic pressure node; S21. Then, 4 μL of PAT aqueous solution is injected into the droplet to form a core-shell structured droplet; S31. A ceramic heating lamp is used for double-sided heating to accelerate solvent evaporation. The distance between the ceramic heating lamp and the droplet is 2-10 cm, the power is 25-100 W, and the temperature is 30-80 ℃. As the solvent evaporates, the concentration of the suspended droplet gradually increases. 4-MBPA is confined and enriched in the gold nanorod to obtain an enriched sample. S41. After the solution has completely evaporated, turn off the power to the suspender and collect the sample onto the silicon wafer for SERS detection. For SERS analysis, select a 5x objective lens, excitation wavelength of 532nm, laser power of 10mW, grating line density of 600gr / mm, slit size of 200μm, integration time of 5s, and a wavelength between 200-2000cm. -1 Data is collected within the specified range.

[0062] Figure 12 The SERS spectra (A) and standard curve (B) for detecting different concentrations of PAT in the heterogeneous solvent confinement enrichment mode in Example 6 are provided by... Figure 12 It can be seen that PAT is at 1679cm -1 The characteristic peak intensity is positively correlated with concentration, and the logarithm of the SERS signal shows good linearity with the logarithm of concentration in the range of 0.01-90.0 μg / L. The linear equation is log... I =0.47log C PAT +3.07, correlation coefficient R=0.9996, and the method detection limit calculated based on a signal-to-noise ratio of 3 (S / N=3) is 3.6 ng / L.

[0063] Example 7 This embodiment provides a heterogeneous solvent confinement enrichment SERS detection method based on acoustic suspension, and its application in the detection of mycotoxins in food. The steps are as follows: S11. Place 6 μL of gold nanorod aqueous solution at the acoustic pressure node; S21. Then, 4 μL of AOH cyclohexane solution is injected into the droplet to form a core-shell structured droplet; S31. A ceramic heating lamp is used for double-sided heating to accelerate solvent evaporation. The distance between the ceramic heating lamp and the droplet is 2-10 cm, the power is 25-100W, and the temperature is 30-80℃. As the solvent evaporates, the concentration of the suspended droplet gradually increases, and AOH is confined and enriched in the gold nanorod to obtain an enriched sample. S41. After the solution has completely evaporated, turn off the power to the suspender and collect the sample onto the silicon wafer for SERS detection. For SERS analysis, select a 5x objective lens, excitation wavelength of 532nm, laser power of 10mW, grating line density of 600gr / mm, slit size of 200μm, integration time of 5s, and a wavelength between 200-2000cm. -1 Data is collected within the specified range.

[0064] Figure 13 The images (A) and standard curve (B) for the detection of different concentrations of AOH in the heterogeneous solvent confinement enrichment mode in Example 7 are provided by... Figure 13 It can be seen that AOH at 1000cm -1 The characteristic peak intensity is positively correlated with concentration, and the logarithm of the SERS signal shows good linearity with the logarithm of concentration in the range of 0.01-80.0 μg / L. The linear equation is log... I =0.51log C AOH +2.93, correlation coefficient R =0.9972, and the method detection limit calculated based on a signal-to-noise ratio of 3 (S / N=3) is 4.3 ng / L.

Claims

1. A method for detecting SERS based on acoustic levitation and confined enrichment of heterogeneous solvents, characterized in that, Includes the following steps: S1. Suspend droplet I containing the SERS substrate at the acoustic pressure node; S2. Inject droplet II containing the target analyte to form a core-shell structured droplet; S3. Double-sided heating accelerates solvent evaporation, confining and enriching the target analyte within the SERS substrate to obtain an enriched sample. S4. Collect the enriched sample and perform SERS detection; Wherein, droplet I and droplet II are immiscible phases.

2. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 1, characterized in that, The volume ratio of droplet I to droplet II is 1:(0.1-0.9); the volume of the core-shell structured droplet is 1-12 μL.

3. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 2, characterized in that, Droplet I and droplet II are independently selected from the aqueous phase and the oil phase, respectively, and are different phases; the aqueous phase includes water; the oil phase includes alkanes with more than 5 carbon atoms.

4. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 3, characterized in that, In droplet II, the target analyte is a compound with Raman-active groups.

5. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 4, characterized in that, When the target analyte is an oil-soluble substance, droplet I is an aqueous phase, droplet II is an oil phase, and the SERS substrate is selected from noble metal nanoparticles.

6. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 4, characterized in that, When the target analyte is a water-soluble substance, droplet I is the oil phase, droplet II is the aqueous phase, and the SERS substrate is selected from hydrophobically modified noble metal nanoparticles.

7. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 5, characterized in that, The precious metal nanoparticles are selected from gold nanoparticles or silver nanoparticles.

8. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 6, characterized in that, The hydrophobically modified noble metal nanoparticles are prepared by hydrophobically modifying noble metal nanoparticles with fluorothiols.

9. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 1, characterized in that, The temperature of the double-sided heating is 30-80℃; the time is 5-60 minutes.

10. The SERS detection method based on acoustic levitation and confined enrichment of heterogeneous solvents according to claim 9, characterized in that, The dual-sided heating includes the use of ceramic heating lamps; the distance between the ceramic heating lamps and the core-shell structure droplets is 2-10 cm; the power is 25-100 W.

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