A composite fluorescent visual sensor and a preparation method and application thereof

Abstract

The application discloses a kind of composite fluorescent visualization sensor and its preparation method and application, it is related to analytical detection technical field;Composite fluorescent visualization sensor is prepared by the following method: first, synthesis of sulfhydryl succinic acid modified cadmium telluride quantum dots and N-acetyl-L-cysteine modified cadmium telluride quantum dots, then porphyrin stock solution is mixed with N-acetyl-L-cysteine modified cadmium telluride quantum dots, namely the chiral nano porphyrin solution can be obtained, finally it is mixed with the sulfhydryl succinic acid modified cadmium telluride quantum dots synthesized in advance, and composite fluorescent visualization sensor can be prepared;The composite fluorescent visualization sensor of quantum dot-nano porphyrin prepared by the application has good selectivity to L / D- theanine and has high sensitivity and anti-interference performance, and can be used for visualizing precise analysis of L / D- theanine content, with the advantages of simple operation, high sensitivity, low cost and the like.

Description

Technical Field

[0001] This invention belongs to the field of analytical detection technology, specifically relating to a composite fluorescence visualization sensor, its preparation method, and its application. Background Technology

[0002] Chiral compounds, as a special class of substances whose mirror images cannot be superimposed, occupy a central position in the fields of chemistry, biology, and materials science due to their unique stereostructure. Their enantiomers often exhibit significant differences in properties when interacting with chiral systems, a characteristic that lays an important foundation for the design and application of functional molecules. Among many natural chiral compounds, theanine, a non-protein amino acid unique to tea, is a typical example of a compound that combines biological activity and application potential, highlighting the importance of chirality. Theanine molecules possess a chiral center, and its naturally occurring form is L-theanine (also known as L-γ-glutamylethylamine), accounting for 50%-60% of the total amino acids in tea. This configuration endows it with unique physiological functions—not only participating in the regulation of nitrogen metabolism and stress response in plants, but also exhibiting calming, cognitive-enhancing, and immune-regulating effects in humans.

[0003] Due to the limited content of L-theanine in natural tea leaves and the high cost of separation and purification, the dominant product on the market is currently theanine artificially synthesized through chemical synthesis, primarily in the form of D-theanine. However, due to differences in stereochemistry, synthetically produced D-theanine exhibits significant differences in bioavailability and activity compared to its natural isomer. For example, researchers have found that the L and D enantiomers of theanine respond differently to various physiological responses in the body. L-theanine is more readily absorbed than D-theanine, and it is preferentially degraded into glutamate and ethylamine in the kidneys, while the D-form is more easily excreted through metabolism, exhibiting significantly lower bioavailability than the L-form. However, not all theanine can be added to food; only theanine obtained from tea leaves is legally permitted. Theanine prepared through chemical synthesis or microbial fermentation cannot be used as a food additive. However, some unscrupulous merchants, in order to obtain high profits, will replace L-theanine with cheap D-theanine, selling inferior products as superior ones, and using this as a competitive advantage, thereby harming consumers' health and rights. Therefore, it is necessary to test L / D-theanine.

[0004] Traditional methods for detecting chiral compounds generally employ optical rotation, gas chromatography (GC) chiral separation, and high-performance liquid chromatography (HPLC). While each method has its advantages in detecting chiral compounds, optical rotation suffers from poor specificity and low sensitivity, making it only suitable for the quantitative analysis of high-purity single chiral compounds. It is also susceptible to interference from factors such as sample concentration, temperature, and solvent, and cannot distinguish chiral isomers in complex matrices. GC chiral separation is only suitable for volatile, heat-stable chiral compounds and relies on large instruments. HPLC, on the other hand, suffers from drawbacks such as reliance on expensive chiral stationary phases, high separation difficulty, and high detection costs, making it insufficient for the on-site detection requirements of L / D-theanine in food. In recent years, quantum dots and nanoporphyrins have been increasingly applied to food detection due to their superior properties. However, currently, no analytical method can rapidly analyze and detect the two configurations of theanine and apply it to food testing. Therefore, there is a need to develop a simple, rapid, and accurate analytical method to distinguish between the two configurations of theanine. Summary of the Invention

[0005] To address the aforementioned shortcomings, this invention provides a composite fluorescence visualization sensor, its preparation method, and its application. The raw materials are readily available, the cost is low, and it can be used for the qualitative and quantitative analysis of L / D-theanine and the detection of complex matrices rich in this type of compound.

[0006] The technical solution adopted by the present invention to solve the above problems is as follows:

[0007] In a first aspect, the present invention provides a method for fabricating a composite fluorescence visualization sensor, comprising the following steps:

[0008] S1: Dissolve cadmium chloride and mercaptosuccinic acid in water and stir for the first time, then purge with nitrogen and continue stirring. Add sodium tellurite solution and sodium borohydride solution and carry out hydrothermal reaction. After filtration, dialysis and dilution, mercaptosuccinic acid modified cadmium telluride quantum dots can be obtained.

[0009] S2: Dissolve cadmium chloride and N-acetyl-L-cysteine ​​in water and stir for the second time. Then, purge with nitrogen and continue stirring. Add sodium tellurite solution and sodium borohydride solution and carry out hydrothermal reaction. After filtration, dialysis and dilution, N-acetyl-L-cysteine-modified cadmium telluride quantum dots can be obtained.

[0010] S3: N,N-dimethylformamide was mixed with 5,10,15,20-tetra(4-aminophenyl)porphyrin to obtain a porphyrin stock solution. Then, the porphyrin stock solution was mixed with N-acetyl-L-cysteine-modified cadmium telluride quantum dots and water. The pH was first adjusted to 6.4-6.6 and incubated. After incubation, the pH was adjusted to 2.4-2.6 and then stirred for the third time to obtain a chiral nanoporphyrin solution.

[0011] S4: By adding a chiral nanoporphyrin solution to cadmium telluride quantum dots modified with mercaptosuccinic acid, a quantum dot-nanoporphyrin composite fluorescent sensor can be constructed.

[0012] Further, in step S1, the mass ratio of cadmium chloride, mercaptosuccinic acid, sodium tellurite, and sodium borohydride is 0.16~0.18:0.15~0.18:0.0399~0.04:0.054~0.06, and the mass ratio of cadmium chloride to water is 0.16~0.18:40.

[0013] Further, in step S1, the first stirring adjusts the pH to 9.6~9.8; the hydrothermal reaction specifically involves a reaction temperature of 180~200℃; the filtration specifically involves using a microporous membrane for filtration; and the dialysis specifically involves dialysis in a dialysis bag for 24 hours.

[0014] Further, in step S2, the mass ratio of cadmium chloride, N-acetyl-L-cysteine, sodium tellurite, and sodium borohydride is 0.16~0.18:0.19~0.25:0.0399~0.0450:0.054~0.06; and the mass ratio of cadmium chloride to water is 0.16~0.18:40.

[0015] Further, in step S2, the second stirring adjusts the pH to 9.70±0.1.

[0016] Further, in step S3, the volume ratio of N,N-dimethylformamide to 5,10,15,20-tetra(4-aminophenyl)porphyrin is 1:9, and the volume ratio of porphyrin stock solution, N-acetyl-L-cysteine-modified cadmium telluride quantum dots, and water is 1:2 to 3:40.

[0017] Furthermore, in step S3, the third stirring specifically refers to a reaction temperature of 80-90℃.

[0018] Furthermore, in step S4, the volume ratio of the chiral nanoporphyrin solution to the cadmium telluride quantum dots modified with mercaptosuccinic acid is 1:1.

[0019] Secondly, the present invention provides a composite fluorescence visualization sensor prepared by the above-described preparation method.

[0020] Thirdly, the present invention also provides the application of a composite fluorescence visualization sensor in the detection of L / D-theanine.

[0021] Compared with the prior art, the advantages and beneficial effects of the technical solution proposed in this invention are:

[0022] Compared with traditional detection methods, this invention overcomes many of the shortcomings of traditional detection technologies. It has many advantages, such as no need for complicated pretreatment, low detection limit, strong anti-interference ability, readily available raw materials, low cost, high selectivity and simple signal reading method, and is more suitable for on-site detection. It enables the visualization and accurate analysis of chiral compounds such as L / D-theanine in actual samples (L-theanine additives, edible amino acid supplements, etc.). Attached Figure Description

[0023] Figure 1 Transmission electron microscopy images of cadmium telluride quantum dots (YQDs) modified with mercaptosuccinic acid and cadmium telluride quantum dots modified with N-acetyl-L-cysteine ​​pre-chiral induced 5,10,15,20-tetra(4-aminophenyl) nanoporphyrin (CTAPP) materials and composite fluorescence visualization sensors constructed from both.

[0024] Figure 2 Infrared spectra of YQDs, CTAPP and sensors, and theanine;

[0025] Figure 3 The circular dichroism spectrum of chiral nanoporphyrin CTAPP and its sensor is shown.

[0026] Figure 4 Particle size distribution of YQDs, CTAPP, sensors, and theanine;

[0027] Figure 5 Zeta potential diagrams of YQDs, CTAPP, sensors, and theanine;

[0028] Figure 6 The results show the fluorescence detection of L-theanine at a series of concentrations by the YQDs-CTAPP sensor and the corresponding linear fitting results; (A) Fluorescence intensity variation of L-theanine at a series of concentration gradients by the YQDs-CTAPP sensor; (B) Linear fitting results of L-theanine by the YQDs-CTAPP sensor.

[0029] Figure 7 The results show the fluorescence detection of D-theanine at a series of concentrations by the YQDs-CTAPP sensor and the corresponding linear fitting results; (A) Fluorescence intensity variation of D-theanine at a series of concentration gradients by the YQDs-CTAPP sensor; (B) Linear fitting results of D-theanine by the YQDs-CTAPP sensor.

[0030] Figure 8 UV images of L-theanine and D-theanine obtained by the YQDs-CTAPP sensor;

[0031] Figure 9Visualization, color difference, and quantitative analysis results of the YQDs-CTAPP visual sensor array for L / D-theanine at different concentrations; (A) YQDs-CTAPP sensor for 5×10 -6 -1×10 -9 Visualization of L / D-theanine at a concentration of mol / L; (B) Sensor YQDs-CTAPP at 5×10 -6 -1×10 -9 (C) Color difference diagram of L / D-theanine at a concentration of mol / L; (D) Quantitative analysis results of L-theanine by sensor YQDs-CTAPP;

[0032] Figure 10 The results of fluorescence intensity testing (A), visual sensor array visualization, and color difference map (B) of the sensor for different ratios of theanine (mixing ratio of L-theanine to D-theanine is 1:1-1:0.001).

[0033] Figure 11 The results show the quantitative analysis of different ratios of theanine (the mixing ratio of L-theanine and D-theanine is 1:1 to 1:0.001) by the sensor; where L-theanine is (A) and D-theanine is (B).

[0034] Figure 12 The images show the visualization and fluorescence intensity difference of the sensor; (A) is the visualization of the YQDs-CTAPP sensor for L / D-theanine and the other 6 common amino acids under UV light; (B) is the fluorescence intensity difference of the YQDs-CTAPP sensor for L / D-theanine and the other 6 common amino acids.

[0035] Figure 13 For concentration (1×10) -6 Common metal ions (mol / L) were used as interfering components added to the detection system. The anti-interference test diagram of the YQDs-CTAPP sensor is shown in Figure (A); the visualization diagram of the YQDs-CTAPP sensor's anti-interference against metal ions and the fluorescence channel color difference diagram are shown in Figure (B).

[0036] Figure 14 This is a pH stability test graph of the YQDs-CTAPP sensor in the pH range of 6-11. Detailed Implementation

[0037] The following examples are provided to better understand the present invention, but are not intended to limit the invention. The technical solution of the present invention will be further described in detail below using L / D-theanine, a compound found in food, in conjunction with the accompanying drawings.

[0038] This invention constructs a composite fluorescence visualization sensor based on cadmium telluride quantum dots (YQDs) modified with mercaptosuccinic acid and 5,10,15,20-tetra(4-aminophenyl) porphyrin nanoparticles (CTAPP) material pre-chirally induced by N-acetyl-L-cysteine-modified cadmium telluride quantum dots. The detection mechanism is as follows: monodisperse chiral cadmium telluride quantum dots are synthesized using an aqueous phase synthesis method, and the chiral groups on the surface of the quantum dots are completely deprotonated; the porphyrin is dispersed in a solution containing N,N-dimethylformamide (DMF), at which point the porphyrin is chiral; then the chiral quantum dots and porphyrin are mixed... Under these conditions, the repulsion between the two is too strong, and chiral induction fails. Lowering the pH enhances the electrostatic interaction between the two, promoting the formation of porphyrin aggregates, and the chirality of the quantum dots is transferred to the porphyrin aggregates. At this point, the addition of yellow quantum dots (YQDs) modified with mercaptosuccinic acid significantly reduces the fluorescence and fluorescence lifetime of the quantum dots. Further addition of L / D-theanine, due to the strong binding ability between the chiral nanoporphyrin and L / D-theanine through hydrogen bonding and π-π stacking, which is stronger than the weak electrostatic interaction between the quantum dots and the chiral nanoporphyrin, can pull the nanoporphyrin away from the quantum dot surface, restoring its fluorescence. Finally, a highly sensitive and selective strategy was developed, enabling the detection of L / D-theanine in food using a visual fluorescence sensor.

[0039] In a first aspect, the present invention provides a method for fabricating a composite fluorescence visualization sensor.

[0040] Example 1:

[0041] The method for preparing a composite fluorescence visualization sensor using mercaptosuccinic acid-modified cadmium telluride quantum dots and cadmium telluride quantum dots modified with N-acetyl-L-cysteine ​​as pre-chirally induced 5,10,15,20-tetra(4-aminophenyl) nanoporphyrin materials includes the following steps:

[0042] S1: Synthesis of cadmium telluride quantum dots (YQDs) modified with mercaptosuccinic acid:

[0043] Cadmium chloride (0.1650 g, 0.90 mmol) and mercaptosuccinic acid (0.1447 g, 1.2 mmol) were dissolved in 40 mL of ultrapure water. The solution was stirred at room temperature and pressure for 15 min, and the pH was adjusted to 9.70 ± 0.1. Then, nitrogen gas was purged and stirred for 20 min at room temperature. Sodium tellurite (0.0399 g, dissolved in 1 mL of ultrapure water) was added using a syringe, followed by sodium borohydride (0.0540 g, dissolved in 1 mL of ultrapure water). Nitrogen gas purging and stirring were continued for another 15 min. The solution was then quickly placed in a reaction vessel and subjected to hydrothermal reaction at 200 °C for 40 min to obtain a yellow fluorescent quantum dot solution (YQDs). The cadmium telluride quantum dot solution was filtered through a 0.22 μm microporous membrane, and the filtrate was transferred to a 3500 Da dialysis bag for dialysis. The water was changed every 6 h, and dialysis was continued for approximately 24 h. The obtained cadmium telluride quantum dot solution was diluted 100 times with ultrapure water as a stock solution for subsequent use. It was then stored in a refrigerator at 4°C to obtain cadmium telluride quantum dots modified with mercaptosuccinic acid.

[0044] S2: Synthesis of N-acetyl-L-cysteine-modified cadmium telluride quantum dots:

[0045] Cadmium chloride (0.1650 g, 0.90 mmol) and N-acetyl-L-cysteine ​​(0.1958 g, 1.2 mmol) were dissolved in 40 mL of ultrapure water. The solution was stirred for 15 min at room temperature and pressure, and the pH was adjusted to 9.70 ± 0.1. Nitrogen gas was then applied and the solution was stirred for 20 min at room temperature. Sodium tellurite (0.0399 g, dissolved in 1 mL of ultrapure water) was added using a syringe, followed by sodium borohydride (0.0540 g, dissolved in 1 mL of ultrapure water). Nitrogen gas was applied and the solution was stirred for another 15 min. The solution was then quickly placed in a reaction vessel and subjected to hydrothermal reaction at 200 °C for 40 min to obtain an orange fluorescent quantum dot solution. The cadmium telluride quantum dot solution was filtered through a 0.22 μm microporous membrane, and the filtrate was transferred to a 3500 Da dialysis bag for dialysis. The water was changed every 6 h, and dialysis was continued for approximately 24 h. The obtained cadmium telluride quantum dot solution was diluted 100 times with ultrapure water as a stock solution for subsequent use. It was then stored in a refrigerator at 4 ℃ to obtain N-acetyl-L-cysteine ​​modified cadmium telluride quantum dots.

[0046] S3: Synthesis of chiral nanoporphyrin solution (CTAPP):

[0047] 5,10,15,20-tetrakis(4-aminophenyl)porphyrin was dissolved in 1 mL of LDMF and prepared to a concentration of 2 × 10⁻⁶. -310 mL of 5,10,15,20-tetra(4-aminophenyl)porphyrin stock solution (mol / L) was prepared. 1 mL of the porphyrin stock solution was mixed with 3 mL of N-acetyl-L-cysteine-modified orange cadmium telluride quantum dots, and then ultrapure water was added to a final volume of 40 mL. The pH was adjusted to 6.5 ± 0.1, and the mixture was incubated for 30 min. The pH was then adjusted to 2.5 ± 0.1, and the mixture was heated in a water bath to 90 ± 3 °C with stirring for 4 h. The mixture was then stored at 4 °C to obtain the chiral nanoporphyrin solution (CTAPP).

[0048] S4: Fabrication of a quantum dot-porphyrin nanocomposite fluorescence sensor (YQDs-CTAPP):

[0049] A quantum dot-nanoporphyrin composite fluorescence sensor can be successfully constructed by adding a chiral-induced 5,10,15,20-tetra(4-aminophenyl) nanoporphyrin solution to mercaptosuccinic acid-modified yellow cadmium telluride quantum dots at a volume ratio of 1:1. The nanoporphyrin quenches the fluorescence of the quantum dots through electron transfer and fluorescence resonance energy transfer.

[0050] Example 2:

[0051] S1: Synthesis of cadmium telluride quantum dots (YQDs) modified with mercaptosuccinic acid:

[0052] Cadmium chloride (0.1833 g, 1.00 mmol) and mercaptosuccinic acid (0.1809 g, 1.5 mmol) were dissolved in 40 mL of ultrapure water. The solution was stirred for 15 min at room temperature and pressure, and the pH was adjusted to 9.70 ± 0.1. Then, nitrogen gas was purged and stirred for 20 min at room temperature. Sodium tellurite (0.0400 g, dissolved in 1 mL of ultrapure water) was added using a syringe, followed by sodium borohydride (0.0600 g, dissolved in 1 mL of ultrapure water). Nitrogen gas purging and stirring were continued for another 15 min. The solution was then quickly placed in a reaction vessel and subjected to hydrothermal reaction at 180 °C for 40 min to obtain a yellow fluorescent quantum dot solution. The cadmium telluride quantum dot solution was filtered through a 0.22 μm microporous membrane, and the filtrate was transferred to a 3500 Da dialysis bag for dialysis. The water was changed every 6 h, and dialysis was continued for approximately 24 h. The obtained cadmium telluride quantum dot solution was diluted 100 times with ultrapure water as a stock solution and stored in a refrigerator at 4 ℃ to obtain cadmium telluride quantum dots modified with mercaptosuccinic acid.

[0053] S2: Synthesis of N-acetyl-L-cysteine-modified cadmium telluride quantum dots:

[0054] Cadmium chloride (0.1833 g, 1.00 mmol) and N-acetyl-L-cysteine ​​(0.2448 g, 1.5 mmol) were dissolved in 40 mL of ultrapure water. The solution was stirred for 15 min at room temperature and pressure, and the pH was adjusted to 9.70 ± 0.1. Nitrogen gas was then applied and the solution was stirred for 20 min at room temperature. Sodium tellurite (0.0450 g, dissolved in 1 mL of ultrapure water) was added using a syringe, followed by sodium borohydride (0.0600 g, dissolved in 1 mL of ultrapure water). Nitrogen gas was applied and the solution was stirred for another 15 min. The solution was then quickly placed in a reaction vessel and subjected to hydrothermal reaction at 180 °C for 40 min to obtain an orange fluorescent quantum dot solution. The cadmium telluride quantum dot solution was filtered through a 0.22 μm microporous membrane, and the filtrate was transferred to a 3500 Da dialysis bag for dialysis. The water was changed every 6 h, and dialysis was continued for approximately 24 h. The obtained cadmium telluride quantum dot solution was diluted 100 times with ultrapure water as a stock solution and stored in a refrigerator at 4 ℃ to obtain N-acetyl-L-cysteine ​​modified cadmium telluride quantum dots.

[0055] S3: Synthesis of chiral nanoporphyrin solution (CTAPP):

[0056] 5,10,15,20-tetrakis(4-aminophenyl)porphyrin was dissolved in 1 mL of LDMF and prepared to a concentration of 2 × 10⁻⁶. -3 10 mL of 5,10,15,20-tetra(4-aminophenyl)porphyrin stock solution (mol / L) was prepared. 1 mL of the porphyrin stock solution was mixed with 2 mL of N-acetyl-L-cysteine-modified orange cadmium telluride quantum dots, and then ultrapure water was added to a final volume of 40 mL. The pH was adjusted to 6.5 ± 0.1, and the mixture was incubated for 30 min. The pH was then adjusted to 2.5 ± 0.1, and the mixture was heated in a water bath to 80 °C with stirring for 4 h. The mixture was then stored at 4 °C to obtain the chiral nanoporphyrin solution (CTAPP).

[0057] S4: Fabrication of a quantum dot-porphyrin nanocomposite fluorescence sensor (YQDs-CTAPP):

[0058] A quantum dot-nanoporphyrin composite fluorescence sensor can be successfully constructed by adding a chiral-induced 5,10,15,20-tetra(4-aminophenyl) nanoporphyrin solution to mercaptosuccinic acid-modified yellow cadmium telluride quantum dots in a 1:1 ratio. The nanoporphyrin quenches the fluorescence of the quantum dots through electron transfer and fluorescence resonance energy transfer.

[0059] Secondly, the present invention provides a composite fluorescence sensor obtained by the above preparation method.

[0060] (1) Morphology, spectrum and particle size characterization of the composite sensor:

[0061] Figure 1 Transmission electron microscopy images of cadmium telluride quantum dots (YQDs) modified with mercaptosuccinic acid and cadmium telluride quantum dots modified with N-acetyl-L-cysteine ​​prechirogeneously induced 5,10,15,20-tetra(4-aminophenyl) porphyrin nanoparticles (CTAPP), and a composite fluorescence visualization sensor constructed from both. Figure 1 It can be seen that the surface of YQDs quantum dots has lattice stripes and the particle size is very small, while the shape of nanoporphyrin CTAPP is conical and the distribution is relatively dispersed. After adding quantum dots YQDs to nanoporphyrin CTAPP, the degree of aggregation increases, the conical shape becomes more obvious, and the chiral characteristics of the material are enhanced.

[0062] Figure 2 Figure 2 shows the infrared spectra of YQDs, CTAPP, the sensor, and theanine, with the infrared curve of YQDs as the reference, at ~3500 cm⁻¹. -1 The sharp, strong peak (-OH) completely disappears in the CTAPP infrared curve, and the peak shape becomes flat with no absorption; in YQDs-CTAPP, the intensity of this peak is significantly reduced and the peak shape is broadened to ~1500 cm⁻¹. -1 The sharp, strong double peaks of YQDs (-COO) - The peak intensity also decreased significantly due to the superposition of weak CTAPP vibrations. Upon the introduction of L / D-theanine, a sharp, strong peak (-NH2) appeared at ~3500 cm⁻¹, the broad peak at ~1500 cm⁻¹ split into multiple sets of sharp, strong peaks, and the low wavenumber region changed from a gentle, weak peak to a dense, sharp, strong peak. These differences in the infrared curves reflect the influence of interactions between different components on peak intensity and shape, and also provide a feasible method for sensors to identify L / D-theanine.

[0063] Figure 3 The image shows the circular dichroism spectrum of chiral porphyrin nanoparticles CTAPP and their sensor. Figure 3 The detection results show that the nanoporphyrin CTAPP exhibits significant CD signal fluctuations in the 190-240 nm range, indicating that it possesses chirality and exhibits differences in absorption for left-handed / right-handed circularly polarized light. Similarly, the CD spectrum detection results of the YQDs-CTAPP composite system indicate that the sensor also possesses chirality.

[0064] Figure 4 Particle size distributions of YQDs, CTAPP, sensors, and theanine, based on... Figure 4The particle size distribution results (AB) show that the YQDs particles are concentrated in the range of 5-10 nm, exhibiting a narrow and sharp single-peak distribution with good particle size uniformity. After being combined with CTAPP, the particle size increases significantly to about 400 nm. Although it is still a single peak, the peak shape is broadened, indicating that aggregation occurs and the uniformity decreases slightly. After the introduction of L-theanine, the particle size drops back to 50-60 nm, the peak shape becomes sharp and symmetrical again, and the uniformity is restored. This reflects the regulatory effect of theanine on the dispersibility of the material and reflects the overall influence of different component combinations on particle size and distribution. Figure 5 The zeta potential diagrams for YQDs, CTAPP, sensors, and theanine are based on... Figure 5 The zeta potential diagram of (AC) shows that the surface of YQDs carries a strong negative charge (potential of about -50mV), resulting in strong electrostatic repulsion and good colloidal stability. When YQDs bind to CTAPP, the potential shifts to about -25mV, the negative charge weakens, the charge distribution widens, the electrostatic repulsion decreases, and the system stability slightly decreases. When L-amino acids are added to form the YQDs-CTAPP-L-amino acid system, the potential approaches neutral, the charge distribution becomes more dispersed, the electrostatic repulsion is greatly weakened, and particles are more likely to aggregate due to van der Waals forces.

[0065] (2) High selectivity of the composite sensor for L / D-theanine:

[0066] 100 μL, 1×10 -6 The mol / L L / D-theanine was replaced with 100 μL, 1×10 -5 Other common amino acids at the same mol / L concentration, such as L-phenylalanine, glycine, L-isoleucine, L-lysine, L-tryptophan, and L-alanine. In this example, the L-phenylalanine, glycine, L-isoleucine, L-lysine, L-tryptophan, and L-alanine used were all prepared at a concentration of 1×10⁻⁶ mol / L. -5 mol / L, which is ten times that of L / D-theanine. A visualization of the selectivity comparison of six common amino acids was obtained using the same method as above. Figure 12 (A) and the fluorescence intensity difference histogram ( Figure 12 (B) These common amino acids cannot restore the fluorescence of quantum dots even at high concentrations, and the fluorescence intensity is significantly lower than that of L / D-theanine, indicating that the composite fluorescence visualization sensor constructed in this invention has good selectivity for L / D-theanine.

[0067] (3) Anti-interference and stability of composite sensors

[0068] To test the anti-interference performance of the composite fluorescence visualization sensor for the detection of L / D-theanine, a concentration of 1×10⁻⁶ was used. -6 L / D-theanine at a concentration of 1 × 10 mol / L was prepared.-6 Common metal ions (Mg) at mol / L 2+ Na + K + Ca 2 + Fe 3+ These were used as interfering components and added separately to the detection system. The fluorescence detection results are as follows: Figure 13 As shown in (A), the visualization results are as follows: Figure 13 As shown in (B), even with the same concentration of interfering components added to the detection system, this composite sensor still exhibits strong anti-interference performance.

[0069] The pH of real-world environments (such as biological samples, water bodies, and industrial fluids) often fluctuates, and pH changes can affect the active components of sensors, leading to signal drift, decreased sensitivity, or even failure. Stability tests at different pH levels can clarify the sensor's applicable pH range and assess its signal stability and accuracy in complex acidic and alkaline environments, ensuring reliable detection results in practical use. Therefore, this invention tested the sensor's fluorescence response under different pH conditions, prepared buffer solutions with pH 6-11, and used them instead of water in the detection conditions. The results are as follows: Figure 14 As shown, the fluorescence intensity remains above 600FL within the pH range of 6-11, exhibiting a high level of fluorescence intensity. Therefore, this composite fluorescence visualization sensor can maintain good fluorescence response stability under different pH conditions.

[0070] Thirdly, the present invention provides an application of the composite fluorescence sensor obtained by the above preparation method in the detection of L / D-theanine.

[0071] Application Example 1:

[0072] For the detection of L / D-theanine:

[0073] It should be specifically noted that when used for L / D-theanine fluorescence detection, CTAPP (100 μL, 5×10⁻⁶ μL) is used. -6 mol / L), YQDs (60-100 μL, 3×10 -10 The concentrations of L / D-theanine were 5 × 10 mol / L. -6 3×10 -6 1×10 -6 7×10 -7 5×10 -7 3×10 -7 1×10 -7 5×10 -8 1×10 -8 1×10-9 mol / L.

[0074] Timing began after the last reactant was added, and the fluorescence emission spectrum of the system was recorded after 1 minute. The fluorescence peaks between 450 nm and 650 nm were extracted from the above fluorescence spectrum. The YQDs-CTAPP sensor was used to obtain fluorescence spectra of a series of concentrations of L / D-theanine, as shown below. Figure 6 (A) Figure 7 As shown in (A), based on this concentration gradient result, a linear fit was performed to obtain the linear fitting equation of the sensor YQDs-CTAPP for L / D-theanine, as follows: Figure 6 As shown in (B) and 7(B), it can be seen that the fluorescence intensity of the sensor increases regularly with the increase of L / D-theanine concentration, and the linear fitting results also show that the fluorescence intensity is significantly correlated with the concentration, proving the excellent detection performance of the sensor for theanine. The UV response of the sensor to L / D-theanine is as follows: Figure 8 As shown, the absorbance of the sensor gradually decreases with increasing wavelength, and the overall trend of the absorption curves of the three materials is consistent, indicating that the introduction of theanine did not change the basic absorption characteristics of the material. At the same time, the absorbance of the absorption curve peaks of the three materials changed, reflecting that the different configurations of theanine have different effects on the absorption performance of the sensor. This verifies the feasibility of the sensor for detecting L / D-theanine, and this sensor can realize the detection of L / D-theanine.

[0075] Based on the fluorescence detection results above, the sensor and a series of concentrations of L / D-theanine solution were dropped onto a 96-well plate. Under UV light irradiation, the resulting visual colorimetric images were obtained as shown below. Figure 9 As shown in (A). Then, using MATLAB software, this visualization result was subjected to qualitative and quantitative analysis, thereby obtaining the corresponding color difference image (…). Figure 9 (B) and quantitative analysis results ( Figure 9 (C), (D)). Based on the color difference diagram and quantitative analysis results, it can be seen that the sensor has good sensitivity and can achieve visual detection of L / D-theanine. Furthermore, the detection limit of this sensor for L-theanine is 1.17 × 10⁻⁶. -10 The detection limit for D-theanine was 1.69 × 10 mol / L. -10 mol / L.

[0076] Application Example 2:

[0077] For the detection of L / D-theanine in different mixing ratios:

[0078] It should be specifically noted that the concentration of L / D-theanine used in different mixing ratios is 1×10⁻⁶. -6mol / L. The ratios of L-theanine to D-theanine were 1:1, 1:0.1, 1:0.01, and 1:0.001, respectively. Timing was started after the last reactant was added, and the fluorescence emission spectrum of the system was recorded after 1 min.

[0079] By extracting the fluorescence peaks between 450 nm and 650 nm from the above fluorescence spectra, the YQDs-CTAPP sensor detected the fluorescence spectra of L / D-theanine with different mixing ratios as follows: Figure 10 As shown in (A). Similarly, based on the above fluorescence detection results, after dropping the sensor with L / D-theanine solutions of different mixing ratios onto a 96-well plate, the visualized color development results were obtained under UV light, as shown in the figure. Figure 10 As shown in (B). Then, using MATLAB software, this visualization result was subjected to qualitative and quantitative analysis, thereby obtaining the corresponding color difference map (…). Figure 10 (B) and quantitative analysis results ( Figure 11 (A), (B).) From the color difference results, it can be seen that even theanine with different mixing ratios has obvious color differences. Furthermore, according to the quantitative results, it can be seen that the sensor has high accuracy in quantifying theanine with different mixing concentrations, demonstrating the sensor's good sensitivity and enabling the visual detection of L / D-theanine mixtures with different ratios.

[0080] Application Example 3:

[0081] This invention constructs a composite fluorescence visualization sensor that, based on the identification of different concentrations of L / D-theanine, can achieve the identification of compounds in food matrices. The different concentrations of L / D-theanine were replaced with L-theanine additive 1 (purchased from Xinjiang Fufeng Biotechnology Co., Ltd.), L-theanine additive 2 (purchased from Guangdong Bozhi Food Technology Co., Ltd.), L-theanine additive 3 (purchased from Xinjiang Fufeng Biotechnology Co., Ltd.), and tea theanine tablets (purchased from Haozhou Suyu Biotechnology Co., Ltd.). These were ground and soaked to prepare concentrations of 10... -6 A solution of mol / L was prepared. Meanwhile, 0.05g of tea leaves (grade 2 West Lake Longjing Hupao and grade 2 West Lake Longjing Meijiawu teas purchased from the China Tea Research Institute) were soaked in 10ml of deionized water at 60℃ for 30min, then removed and cooled to room temperature (25℃) to obtain the corresponding tea infusion.

[0082] Similarly, timing was started after the last reactant was added, and the fluorescence emission spectrum of the system was recorded after 1 minute. All data were repeated three times. A linear regression equation was obtained using the fluorescence spectra of L-theanine at different concentrations, and the recovery rate of L-theanine was used as the evaluation index to determine the accuracy of the quantitative analysis. The results are shown in Table 1. In real food samples, the sensor can still accurately detect L-theanine, with a recovery rate between 74.0% and 110.7%.

[0083] Table 1. Recovery rate of L-theanine in real food samples by the composite fluorescence visualization sensor.

[0084]

[0085] In summary, this invention specifically relates to a composite fluorescence visualization sensor constructed using cadmium telluride quantum dots modified with mercaptosuccinic acid and 5,10,15,20-tetra(4-aminophenyl) nanoporphyrin material chirally induced by cadmium telluride quantum dots modified with N-acetyl-L-cysteine. This sensor has a detection limit of 1.17 × 10⁻⁶ for L-theanine. -10 The detection limit for D-theanine was 1.69 × 10 mol / L. -10 The sensor, with a concentration of mol / L, enables precise and visualized analysis of chiral compounds such as L-theanine in real samples (L-theanine additives, edible amino acid supplements, etc.), achieving recoveries ranging from 74.0% to 110.7%. It offers numerous advantages, including no need for complex pretreatment, low detection limit, strong anti-interference capabilities, low cost, high selectivity, and suitability for on-site detection. This provides a new approach and method for the application of sensors in the detection of complex food matrices.

[0086] Furthermore, any equivalent changes and modifications made without departing from the method and scope of the present invention should be included within the scope of the present invention.

Claims

1. The application of a composite fluorescence visualization sensor in the detection of L / D-theanine, characterized in that, The composite fluorescence visualization sensor was prepared by the following method: S1: Dissolve cadmium chloride and mercaptosuccinic acid in water and stir for the first time, then purge with nitrogen and continue stirring. Add sodium tellurite solution and sodium borohydride solution and carry out hydrothermal reaction. After filtration, dialysis and dilution, mercaptosuccinic acid modified cadmium telluride quantum dots can be obtained. S2: Dissolve cadmium chloride and N-acetyl-L-cysteine ​​in water and stir for the second time. Then, purge with nitrogen and continue stirring. Add sodium tellurite solution and sodium borohydride solution and carry out hydrothermal reaction. After filtration, dialysis and dilution, N-acetyl-L-cysteine-modified cadmium telluride quantum dots can be obtained. S3: N,N-Dimethylformamide is mixed with 5,10,15,20-tetra(4-aminophenyl)porphyrin to obtain a porphyrin stock solution. Then, the porphyrin stock solution is mixed with N-acetyl-L-cysteine-modified cadmium telluride quantum dots and water. The pH is first adjusted to 6.4-6.6, and incubation is performed. After incubation, the pH is adjusted again to 2.4-2.6, and a third stirring is performed to obtain a chiral nanoporphyrin solution. The N,N-dimethylformamide... The volume ratio of amide to 5,10,15,20-tetra(4-aminophenyl)porphyrin is 1:9, and the volume ratio of porphyrin stock solution, N-acetyl-L-cysteine-modified cadmium telluride quantum dots, and water is 1:2~3:40; the third stirring is specifically carried out at a reaction temperature of 80-90℃; S4: the chiral nanoporphyrin solution is added to the mercaptosuccinic acid-modified cadmium telluride quantum dots to construct a quantum dot-nanoporphyrin composite fluorescent sensor.

2. The application according to claim 1, characterized in that, In step S1, the mass ratio of cadmium chloride, mercaptosuccinic acid, sodium tellurite, and sodium borohydride is 0.16~0.18:0.15~0.18:0.0399~0.04:0.054~0.06, and the mass ratio of cadmium chloride to water is 0.16~0.18:

40.

3. The application according to claim 1, characterized in that, In step S1, the first stirring adjusts the pH to 9.6-9.8; the hydrothermal reaction specifically involves a reaction temperature of 180-200℃; the filtration specifically involves using a microporous membrane for filtration; and the dialysis specifically involves dialysis in a dialysis bag for 24 hours.

4. The application according to claim 1, characterized in that, In step S2, the mass ratio of cadmium chloride, N-acetyl-L-cysteine, sodium tellurite, and sodium borohydride is 0.16~0.18:0.19~0.25:0.0399~0.0450:0.054~0.06; and the mass ratio of cadmium chloride to water is 0.16~0.18:

40.

5. The application according to claim 1, characterized in that, In step S2, the second stirring adjusts the pH to 9.70±0.

1.

6. The application according to claim 1, characterized in that, In step S4, the volume ratio of the chiral nanoporphyrin solution to the cadmium telluride quantum dots modified with mercaptosuccinic acid is 1:1.

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