A method for identifying chiral flavor substances based on covalent organic framework nanochannels

Abstract

The present application relates to a method for identifying chiral flavor substances based on covalent organic framework nanochannels, which belongs to the technical field of chiral substance identification and nanochannel analysis. The identification method comprises the following steps: synthesizing a covalent organic framework (COF) membrane by an interfacial polymerization method; preparing Chiral-AuNPs with good chiral signals; using the COF membrane as a nanochannel and the Chiral-AuNPs as a recognition unit to convert weak chiral interactions into nanoscale aggregation behavior, thereby hindering the transmission of ions in the COF nanochannel and achieving the purpose of distinguishing chiral substances. A new type of label-free biomimetic chiral nanochannel sensing platform is constructed for the identification of flavor substance enantiomers. The present application utilizes pure COF membrane as a nanochannel and can be applied to medicine, food testing and environmental science. Compared with other chiral recognition methods, it has the advantages of high sensitivity, simple operation, low cost and good economy.

Description

Technical Field

[0001] The present application relates to a method for identifying chiral flavor substances based on covalent organic framework nanochannels, belonging to the technical field of chiral substance identification and nanochannel analysis. Background Art

[0002] Enantiomers are a pair of stereochemically mirror-image molecules with non-superimposable spatial structures. They are fundamental phenomena and laws of nature. Enantiomers are closely related to human life, possessing identical physical properties. However, they exhibit significant differences in chiral contexts such as biomedicine, food safety, and environmental effects. For example, the S-isomer of naproxen has higher pharmacological activity than the R-isomer, amino acids are present only in natural fruit juices as L-isomers, and the herbicidal activity of phenoxyalkanoic acid herbicides is limited to the R-isomer. Notably, in the vast majority of chiral enantiomers, only one enantiomer is active, while the inactive other enantiomer poses a serious threat to human health, such as residues of chiral pesticides and the emission of various enantiomeric pollutants. Therefore, chiral recognition is crucial in medicine, food, and environmental science.

[0003] Currently, methods for enantiomer separation and determination primarily include chromatography, spectroscopy, and electrochemistry. While chromatography offers excellent selectivity, disadvantages such as expensive instrumentation, complex chiral column preparation, and specialized operation limit its application for high-throughput analysis of chiral molecules at low concentrations. In comparison, both spectroscopy and electrochemistry offer advantages such as high sensitivity and low cost. However, in practical applications, these methods still face limitations such as complex sample pretreatment and low recognition efficiency, hindering the growing demand for advanced bioanalysis. Therefore, there is an urgent need to develop a highly sensitive, high-throughput, real-time identification method to ensure the safety of the relevant enantiomers.

[0004] Nanochannel analysis, a sensing technology that effectively integrates Coulter counting with channel ion current measurement, has attracted considerable attention due to its high sensitivity, rapid response, and ease of miniaturization. In recent years, researchers both domestically and internationally have reported a variety of chiral recognition sensors based on various chiral recognition methods (host-guest interactions, chiral functional group recognition, etc.) and nanochannel materials (polyethylene terephthalate, polyimide membranes, anodized aluminum oxide, etc.), drawing on the transport mechanisms of drugs / metabolites and transmembrane proteins in nature. Covalent organic frameworks (COFs), with their unique properties of high porosity, well-defined pore structure, and controllable pore size, have potential applications in isomer recognition. Currently, most nanochannel membranes are prepared by modifying the membrane surface to incorporate a monolayer of chiral receptors, enabling enantiomer recognition. However, this approach suffers from the limited applicability of chiral recognition units to the membrane, necessitating a more broadly applicable method for identifying chiral flavor substances. Summary of the Invention

[0005] In order to solve the above problems, this application provides a method for identifying chiral flavor substances based on covalent organic framework nanochannels. First, a covalent organic framework (COF) membrane is synthesized as a nanochannel by interfacial polymerization; chiral gold nanoparticles (Chiral-AuNPs) are synthesized by classical redox reactions as recognition units to construct a new label-free biomimetic chiral nanochannel sensing platform, which is applied to the recognition of flavor substance enantiomers.

[0006] Technical Solution

[0007] The first aspect of the present application proposes a method for identifying chiral flavor substances based on covalent organic framework nanochannels, comprising the following steps:

[0008] Step S1: dissolving tris(4-aminophenyl)amine and trialdehyde phloroglucinol in an organic solvent to obtain an organic phase, and adding water dropwise on the surface of the organic phase to form an organic phase / inorganic phase interface;

[0009] Step S2: adding the catalyst dropwise to the inorganic phase, and after the interfacial reaction, a thin film is obtained at the interface, which is then washed to obtain a COF membrane; after the interfacial reaction is completed, a red thin film is obtained, which is then washed to obtain a COF membrane.

[0010] Step S3: dissolving chloroauric acid and chiral amino acid in water to obtain solution A, and then adding sodium borohydride solution dropwise to obtain Chiral-AuNPs solution after reaction;

[0011] Step S4: installing the COF membrane in a flow cell, wherein the COF membrane divides the flow cell into a first cell body and a second cell body; fixing the first electrode in the first cell body, and fixing the second electrode in the second cell body; adding a Chiral-AuNPs solution and an electrolyte into the flow cell;

[0012] Step S5: adding the chiral flavor substance to be tested into the flow cell to perform a current-voltage curve test to identify the chiral flavor substance.

[0013] In one embodiment, the first electrode is an Ag / AgCl electrode.

[0014] In one embodiment, the second electrode is an Ag / AgCl electrode.

[0015] In one embodiment, the organic solvent in step S1 is selected from one or a combination of dichloromethane and dichloroethane;

[0016] In one embodiment, ultrasonic treatment is used to better dissolve tris(4-aminophenyl)amine and trialdehyde phloroglucinol; the treatment time is 15 minutes.

[0017] In one embodiment, in step S2, the catalyst is an acetic acid solution;

[0018] Preferably, the concentration of the catalyst acetic acid solution is 3.0 mol / L;

[0019] In one embodiment, the volume ratio of the catalyst to the organic solvent is 9-12;

[0020] In one embodiment, the washing step uses acetone and ultrapure water for washing; by using acetone and ultrapure water for washing, unreacted monomers can be effectively removed.

[0021] In one embodiment, the reaction container in step S1 is a culture dish;

[0022] In one embodiment, in step S5, during the test, the transmembrane potential between the first electrode and the second electrode is -0.2V-0.2V, and the scanning rate is 10mV / s.

[0023] In one embodiment, the molar ratio of the tris(4-aminophenyl)amine to the trialdehyde phloroglucinol is 0.9-1.2;

[0024] Preferably, the molar ratio of the tris(4-aminophenyl)amine to the trialdehyde phloroglucinol is 1:1;

[0025] In one embodiment, the concentration of tris(4-aminophenyl)amine is 0.4-0.8 mmol / L;

[0026] In one embodiment, the concentration of trialdehyde phloroglucinol is 0.4-0.8 mmol / L.

[0027] Preferably, step S2 is carried out under sealed conditions;

[0028] In one embodiment, the sealing is performed using plastic wrap;

[0029] In one embodiment, the interfacial reaction time in step S2 is 24-72 hours, preferably 48 hours.

[0030] In one embodiment, the concentration of the chloroauric acid solution in solution A is 0.07-0.09 mg / mL.

[0031] In one embodiment, the molar ratio of sodium borohydride to chloroauric acid is 0.5-2.0.

[0032] In one embodiment, the chiral amino acid is selected from L-cysteine, D-cysteine, L(+)-penicillamine, D-penicillamine;

[0033] In one embodiment, the concentration of the chiral amino acid in solution A is 1-3 mmol / L.

[0034] In one embodiment, the concentration of Chiral-AuNPs in step S4 is 1-3 mmol / L;

[0035] Using chiral-AuNPs as recognition units for chiral substances can transform weak chiral interactions into nanoscale aggregation, thereby hindering ion transport within the COF nanochannels and achieving the purpose of distinguishing chiral substances. The current change rate (I-I0) / I0 after the chiral-AuNPs interact with the enantiomers is different due to the different electrostatic and hydrogen bonding forces between the molecules, resulting in different (I-I0) / I0, thus achieving the purpose of chiral recognition.

[0036] In one embodiment, the electrolyte in the electrolyte is selected from at least one of potassium chloride, sodium chloride, sodium dihydrogen phosphate, and disodium hydrogen phosphate.

[0037] Preferably, the electrolyte in the electrolyte is potassium chloride.

[0038] In one embodiment, the concentration of the electrolyte is 1 μmol / L

[0039] In one embodiment, the pH in the flow cell is between 5 and 9.

[0040] In one embodiment, the material of the flow cell is tetrafluoroethylene or quartz glass.

[0041] In a preferred embodiment, the molar concentration of trialdehyde phloroglucinol is 0.6 mmol.

[0042] In a preferred embodiment, the molar concentration of tris(4-aminophenyl)amine is 0.6 mmol.

[0043] In a preferred embodiment, the tris(4-aminophenyl)amine and trialdehyde phloroglucinol are dissolved in dichloromethane at a molar ratio of 1:1.

[0044] In a preferred embodiment, the concentration of the chloroauric acid solution is 0.089 mg / mL.

[0045] In a preferred embodiment, the concentration of sodium borohydride is 0.1 mol / L.

[0046] In a preferred embodiment, the concentration of potassium chloride is 1 μmol / L and the pH value in the flow cell is 7.4.

[0047] The second aspect of the present application is to provide an application of a method for identifying chiral flavor substances based on covalent organic framework nanochannels, which can be applied to the fields of medicine, food testing and environmental science for chiral recognition.

[0048] The third aspect of the present application is to provide a chiral flavor substance identification device, which uses the above method to identify chiral flavor substances;

[0049] The chiral flavor substance identification device includes a circulation cell, a first electrode, a second electrode, a COF membrane, a Chiral-AuNPs identification unit, an electrolyte, and an ammeter; the COF membrane is installed in the circulation cell, and the COF membrane divides the circulation cell into a first cell body and a second cell body; the first electrode is fixed in the first cell body, and the second electrode is fixed in the second cell body; the first electrode is connected to a power supply, the second electrode is connected to a power supply, and the ammeter is set in a loop formed by the first electrode and the second electrode.

[0050] Beneficial effects

[0051] The present application discloses a method for identifying chiral flavor substances based on covalent organic framework nanochannels. The method synthesizes a COF membrane by interfacial polymerization. The COF membrane can be easily and controllably obtained by this method. The COF membrane has good ion current performance as a nanochannel. By preparing Chiral-AuNPs with good chiral signals as recognition units, weak chiral interactions are converted into nanoscale aggregation behavior, thereby hindering the transmission of ions in the COF nanochannel, achieving the purpose of distinguishing chiral substances. Compared with other nanochannel materials, the method of the present application has the advantages of high throughput and good ion current performance, which is conducive to the development of new nanochannel sensing platforms. Compared with other chiral recognition methods, it has the advantages of high sensitivity, simple operation, low cost and good economy. It realizes high-throughput and high-selectivity recognition of chiral flavor substances. BRIEF DESCRIPTION OF THE DRAWINGS

[0052] Figure 1 This is a schematic diagram of the method for identifying chiral flavor substances based on covalent organic framework nanochannels in this application;

[0053] Figure 2 is the N2 adsorption-desorption isotherm (a) and pore size distribution diagram (b) of the COF membrane prepared in Example 1;

[0054] Figure 3 is a SEM cross-sectional image of the COF film prepared in Example 1;

[0055] Figure 4 is a TEM image of chiral gold nanoparticles prepared in Example 2;

[0056] Figure 5is a schematic diagram of the current-voltage curve for identifying limonene isomers at different concentrations;

[0057] Figure 6 is the current change rate of Chiral-AuNPs prepared with different chiral amino acids in recognizing limonene isomers. DETAILED DESCRIPTION

[0058] Examples of what is included in the claims

[0059] Example 1: Preparation of Covalent Organic Framework (COF) Film

[0060] Tris(4-aminophenyl)amine and trialdehyde phloroglucinol were dissolved in dichloromethane (5 mL) at a molar ratio of 1:1 (molar concentration of 0.6 mmol);

[0061] After 15 minutes of ultrasonic treatment, the mixture was poured into a glass petri dish as the organic phase. Ultrapure water was then slowly added dropwise to the surface of the organic phase until a stable organic / inorganic phase interface was formed.

[0062] An acetic acid solution (3.0 mol / L, 60 μL) was slowly added dropwise to the inorganic phase as a catalyst, and the mixture was sealed with plastic wrap. Interfacial polymerization was carried out at room temperature for 48 hours to obtain a red film. The red film was removed and washed with ultrapure water and acetone to remove unreacted monomers, thereby obtaining the target product COF film.

[0063] Figure 2 The N2 adsorption-desorption isotherm and pore size distribution of COF membrane are shown in Figure 2. Figure 2 As shown, it was demonstrated that a covalent organic framework (COF) membrane with nanopores was successfully prepared, and the pore size of the COF membrane was ~1.667nm.

[0064] Figure 3 is the SEM cross-sectional image of the COF film, as shown in Figure 3 As shown, the thickness of the COF film is about 641 nm, which has sufficient mechanical strength to be used for nanochannel analysis technology.

[0065] Example 2: Preparation of Chiral Gold Nanoparticles (Chiral-AuNPs)

[0066] Dissolve chloroauric acid and L-cysteine (L-Cys) in 100 ml of ultrapure water to a chloroauric acid concentration of 0.089 mg / mL and an L-cysteine concentration of 2 mmol / L. Then, add sodium borohydride solution (0.1 mol / L, 200 μL) dropwise and stir vigorously for 2 hours to obtain the target product, Chiral-AuNPs (L-Cys-AuNPs).

[0067] Dissolve chloroauric acid and D-cysteine (D-Cys) in 100 ml of ultrapure water to a concentration of 0.089 mg / mL chloroauric acid and 1 mmol / L D-cysteine. Then, add sodium borohydride solution (0.1 mol / L, 200 μL) dropwise and stir vigorously for 2 hours to obtain the target product, Chiral-AuNPs (D-Cys-AuNPs).

[0068] Chloroauric acid and L(+)-penicillamine (L-Pen) were dissolved in 100 ml of ultrapure water to a chloroauric acid concentration of 0.089 mg / mL and a L(+)-penicillamine concentration of 3 mmol / L. Sodium borohydride solution (0.1 mol / L, 200 μL) was then added dropwise and stirred vigorously for 2 hours to obtain the target product, Chiral-AuNPs (L-Pen-AuNPs).

[0069] Dissolve chloroauric acid and D-penicillamine (D-Pen) in 100 ml of ultrapure water to a chloroauric acid concentration of 0.089 mg / mL and a D-penicillamine concentration of 2 mmol / L. Then, add sodium borohydride solution (0.1 mol / L, 200 μL) dropwise and stir vigorously for 2 hours to obtain the target product, Chiral-AuNPs (D-Pen-AuNPs).

[0070] Figure 4 This is the TEM image of L-Cys-AuNPs prepared in Example 2. Figure 4 As shown, the prepared L-Cys-AuNPs are monodisperse nanoparticles with a size of approximately 3.5 ± 0.8 nm.

[0071] Example 3: Different chiral recognition units recognize limonene isomers: (S)-(-)-Limonene and (R)-(+)-Limonene

[0072] The COF membrane obtained in Example 1 was installed in the middle of a polytetrafluoroethylene flow cell, and the chiral gold nanoparticles obtained in Example 2 were used as recognition units to perform current-voltage (IV) curve tests. The transmembrane potential was set to -0.2 V to 0.2 V, the scan rate was 10 mV / s, and potassium chloride was used as the electrolyte.

[0073] Figure 5 Schematic diagram of the current-voltage curve for identifying different concentrations of limonene isomers. The lighter the color in the figure, the greater the transmembrane ion current (S)-(-)-Limonene concentration. Figure 4It can be clearly seen that the transmembrane ion current changes significantly with the increase in (S)-(-)-Limonene concentration. For example, at 0.2V, the transmembrane ion current decreases as the (S)-(-)-Limonene concentration increases. However, for (R)-(+)-Limonene, its transmembrane ion current value remains almost unchanged. This is because (S)-(-)-Limonene preferentially binds and aggregates with L-Cys-AuNPs. The electrostatic and hydrogen bonding interactions between its molecules hinder ion transport within the nanochannel, resulting in a decrease in current. On the other hand, (R)-(+)-Limonene passes smoothly through the COF nanochannel, with a negligible effect on the current, thus achieving chiral recognition.

[0074] Figure 6 For different chiral gold nanoparticles as recognition unit identification limonene isomer current change rate, it can be seen that the recognition unit prepared by adding the chiral amino acid of different configurations has different current change rate for limonene isomer.And when not adding chiral recognition unit (Bare), the current change rate is basically the same, it is impossible to effectively identify limonene isomer.After adding four kinds of chiral recognition units prepared in Example 2, there is a larger difference in the current change rate of (S)-(-)-Limonene and (R)-(+)-Limonen, which proves that different chiral recognition units can effectively identify limonene isomer.Wherein when L-Cys-AuNPs is as recognition unit, the current change rate difference between limonene isomers is the largest, and efficient identification can be achieved.

[0075] Example 4: Identification of flavor substances: (S)-1,2-propylene glycol and (R)-1,2-propylene glycol

[0076] The COF membrane obtained in Example 1 was installed in the center of a polytetrafluoroethylene flow cell. The L-Cys-AuNPs prepared in Example 2 served as the recognition element. Current-voltage (IV) curve testing was performed with a transmembrane potential ranging from -0.2 V to 0.2 V, a scan rate of 10 mV / s, and potassium chloride as the electrolyte. The concentration of the substance to be recognized was 1.0 mmol / L.

[0077] Example 5: Identification of flavor substances: (S)-(+)-2-methylbutanoic acid and (R)-2-methylbutanoic acid

[0078] The COF membrane obtained in Example 1 was installed in the center of a polytetrafluoroethylene flow cell. The L-Cys-AuNPs prepared in Example 2 served as the recognition element. Current-voltage (IV) curve testing was performed with a transmembrane potential ranging from -0.2 V to 0.2 V, a scan rate of 10 mV / s, and potassium chloride as the electrolyte. The concentration of the substance to be recognized was 1.5 mmol / L.

[0079] Example 6: Identification of flavor substances: (S)-(+)-sec-butanol and (R)-(-)-sec-butanol

[0080] The COF membrane obtained in Example 1 was installed in the center of a polytetrafluoroethylene flow cell. The L-Cys-AuNPs prepared in Example 2 served as the recognition element. Current-voltage (IV) curve testing was performed with a transmembrane potential ranging from -0.2 V to 0.2 V, a scan rate of 10 mV / s, and potassium chloride as the electrolyte. The concentration of the substance to be recognized was 2.0 mmol / L.

[0081] Table 1 Current change rate for identifying different flavor substances

[0082]

[0083] The results of identifying different flavor substances in Examples 4-6 are shown in Table 1. From the data in the table, it can be seen that there are obvious differences in the current change rates between the isomers of propylene glycol, methylbutyric acid, and sec-butanol, that is, effective identification can be achieved, and effective identification can be achieved within a certain concentration range.

[0084] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for identifying chiral flavor substances based on covalent organic framework nanochannels, characterized in that: The following steps are involved: Step S1: dissolving tris(4-aminophenyl)amine and trialdehyde phloroglucinol in an organic solvent to obtain an organic phase, and adding water dropwise on the surface of the organic phase to form an organic phase / inorganic phase interface; Step S2: adding the catalyst dropwise to the inorganic phase, and after the interface reaction, a thin film is obtained at the interface, which is then washed to obtain a COF membrane; Step S3: dissolving chloroauric acid and chiral amino acid in water to obtain solution A, and then adding sodium borohydride solution dropwise to obtain Chiral-AuNPs solution after reaction; Step S4: installing the COF membrane in a flow cell, wherein the COF membrane divides the flow cell into a first cell body and a second cell body; fixing the first electrode in the first cell body, and fixing the second electrode in the second cell body; adding the Chiral-AuNPs solution and the electrolyte into the flow cell; Step S5: adding the chiral flavor substance to be tested into the flow cell to perform a current-voltage curve test to identify the chiral flavor substance; The chiral amino acid is selected from L-cysteine, D-cysteine, L(+)-penicillamine and D-penicillamine.

2. The method for identifying chiral flavor substances based on covalent organic framework nanochannels according to claim 1, characterized in that: The organic solvent is selected from one or a combination of dichloromethane and dichloroethane; In step S2, the catalyst is a 3.0 mol / L acetic acid solution, and the volume ratio of the catalyst to the organic solvent is 9-12.

3. The method for identifying chiral flavor substances based on covalent organic framework nanochannels according to claim 1, characterized in that: In step S5, the transmembrane potential between the first electrode and the second electrode is -0.2 V to -0.2 V, and the scanning rate is 10 mV / s.

4. The method for identifying chiral flavor substances based on covalent organic framework nanochannels according to claim 1, characterized in that: The molar ratio of the tris(4-aminophenyl)amine to the trialdehyde phloroglucinol is 0.9-1.2; The concentration of the tris(4-aminophenyl)amine is 0.4-0.8 mmol / L; the concentration of the trialdehyde phloroglucinol is 0.4-0.8 mmol / L.

5. The method for identifying chiral flavor substances based on covalent organic framework nanochannels according to claim 1, characterized in that: In the solution A, the concentration of the chiral amino acid is 1-3 mmol / L; the concentration of the chloroauric acid is 0.07-0.09 mg / mL; The concentration of the Chiral-AuNPs solution in step S4 is 1-3 mmol / L.

6. The method for identifying chiral flavor substances based on covalent organic framework nanochannels according to claim 1, characterized in that: The molar ratio of the sodium borohydride to the chloroauric acid is 0.5-2.

0.

7. The method for identifying chiral flavor substances based on covalent organic framework nanochannels according to claim 1, characterized in that: The electrolyte in the electrolyte is selected from at least one of potassium chloride, sodium chloride, sodium dihydrogen phosphate, and disodium hydrogen phosphate.

8. A chiral flavor substance identification device, characterized in that: A chiral flavor substance is identified using the method according to any one of claims 1 to 7; the chiral flavor substance identification device comprises a flow cell, a first electrode, a second electrode, a COF membrane, a Chiral-AuNPs identification unit, an electrolyte, and an ammeter; the COF membrane is installed in the flow cell, and the COF membrane divides the flow cell into a first cell body and a second cell body; the first electrode is fixed in the first cell body, and the second electrode is fixed in the second cell body; The first electrode is connected to a power source, the second electrode is connected to a power source, and the ammeter is arranged in a loop formed by the first electrode and the second electrode.

9. Use of the method for identifying chiral flavor substances based on covalent organic framework nanochannels according to any one of claims 1 to 7 or the chiral flavor substance identification device according to claim 8, characterized in that: It is used in medicine, food testing and environmental science for chiral recognition.

Images