Water-soluble rare earth trinuclear supramolecular thiol-containing amino acid responsive CPL probe and preparation method thereof

Abstract

The application discloses a water-soluble rare earth trinuclear supermolecular thiol-containing amino acid response CPL probe and a preparation method thereof, relates to a thiol-containing amino acid response CPL probe and a preparation method thereof. In order to solve the problem that the existing rare earth complex is used for detecting thiol-containing amino acid and has poor water solubility, compatibility and anti-interference ability. The aldehyde group is introduced into the ligand structure, and three rare earth ions are selected as metal centers, so that the rare earth ions interact with six ligands, and a double-layer triangular sandwich structure (Et3NH)3[Eu3L6] complex is successfully constructed. In the formation process of the complex, the aldehyde group reacts with the solvent methanol to generate a hemiacetal structure, and the complex has excellent water solubility, so that the compatibility of the complex in a biological system is greatly improved. More importantly, (Et3NH)3[Eu3L6] can realize sensing of thiol-containing amino acid through significant change of a CPL spectrum, and still has excellent selectivity under the condition that multiple interferences coexist.

Description

Technical Field

[0001] This invention relates to a thiol-containing amino acid-responsive CPL probe and its preparation method. Background Technology

[0002] Amino acids play crucial roles in many biological processes, serving as core building blocks for polypeptides, proteins, and numerous other natural compounds. Among them, thiol-containing amino acids, such as penicillamine (PA), possess significant medicinal value, being the primary metabolite of penicillin antibiotics in vivo. Of the two enantiomers of penicillamine, D-penicillamine (D-PA) is particularly noteworthy. It is not only a highly effective antirheumatic agent for treating Wilson's disease but also acts as an integrator, precisely removing excess copper ions from the body and effectively alleviating symptoms. Given the vital role of D-PA in clinical medicine, developing an efficient and accurate method for its detection is essential.

[0003] Among numerous methods for detecting chiral substances, circular dichroism (CD) and circularly polarized emission (CPL) spectroscopy have attracted considerable attention. While both can detect chiral substances, CPL significantly surpasses CD in spatial resolution due to its unique advantages. This is primarily attributed to the use of luminescent probes, which are perfectly compatible with commonly used microscopy techniques, expanding the scope of detection applications. Among the many luminescent probe molecules, rare earth complexes stand out, exhibiting unparalleled advantages. Due to the unique electronic structure of rare earth elements, the optical anisotropy factor (g) generated by their ff transition processes... lum The concentration of rare earth complexes is far higher than that of ordinary organic compounds, making them excellent luminescent probes in the field of chiral detection spectroscopy.

[0004] Based on these characteristics, constructing supramolecular sensors using rare-earth complexes for the detection of thiol-containing amino acids has become a highly promising research direction. However, existing rare-earth supramolecular probes generally face the following challenges: First, their poor water solubility makes them difficult to stably exist in a completely aqueous physiological environment and effectively identify target molecules; second, their poor water solubility leads to poor compatibility with biological systems, limiting their direct application in complex biological samples; furthermore, in biological matrices containing multiple amino acids, metal ions, and other coexisting interfering substances, these probes often exhibit insufficient selectivity, making it difficult to achieve accurate identification of thiol-containing amino acids. Therefore, improving the water solubility of rare-earth probes and enhancing their compatibility and anti-interference capabilities in biological environments have become core issues that urgently need to be addressed in this field. Summary of the Invention

[0005] To address the shortcomings of existing rare-earth complexes for detecting thiol-containing amino acids, such as poor water solubility, compatibility, and interference resistance, this invention proposes a water-soluble rare-earth trinuclear supramolecular CPL probe responsive to thiol-containing amino acids and its preparation method. This invention introduces an aldehyde group into the ligand structure and selects three rare-earth ions as metal centers, enabling the rare-earth ions to interact with six ligands, successfully constructing a double-layered triangular sandwich structure (Et3NH)3[Eu3L6] complex. During the formation of the complex, the aldehyde group reacts chemically with the solvent methanol, generating a hemiacetal structure that endows the complex with excellent water solubility, greatly improving its compatibility in biological systems. More importantly, (Et3NH)3[Eu3L6] can sense thiol-containing amino acids through significant changes in the CPL spectrum, and still maintains excellent selectivity under conditions of multiple coexisting interferences.

[0006] The structural formula of the water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe of this invention is as follows:

[0007] ;

[0008] The general structural formula of the water-soluble rare-earth trinuclear supramolecular CPL probe containing thiol amino acids is (HNEt3)3[Eu3L6], where L is a ligand, and the structural formula of L is:

[0009] .

[0010] The preparation method of the water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe of the present invention is carried out according to the following steps:

[0011] Step 1: Synthesis of 4-hydroxy-3-iodoacetophenone

[0012] Add 3-5g of 4-hydroxyacetophenone, 4-6g of elemental iodine and 10-20mL of hydrogen peroxide aqueous solution to 100-200mL of water, stir at room temperature for 24h, filter to obtain solid, and recrystallize by anhydrous ethanol to obtain 4-hydroxy-3-iodoacetophenone.

[0013] Step 2: Synthesis of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone

[0014] Add 4-6 g of 4-hydroxy-3-iodoacetophenone and 8-10 g of Cs₂CO₃ to a Schlenk flask, add 100-200 mL of N,N-dimethylformamide, and stir at 110 °C for 0.5 h. Then add 5-7 g of 2-bromo-1,1-dimethoxyethane and react for 5 h. Quench the reaction solution in water and extract with dichloromethane. Combine the organic layers after extraction, and finally wash, dry, concentrate to remove solvent and purify to obtain 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)acetophenone.

[0015] Step 3: Synthesis of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl

[0016] 1-3 g of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone, 0.2-0.4 g of Pd(PPh3)4 and 1-2 g of 3-acetylphenylboronic acid were placed in a Schlenk flask; 40-60 mL of tetrahydrofuran and 20-30 mL of sodium carbonate aqueous solution were added sequentially, and the mixture was refluxed and stirred for 12 h; the reaction solution was then poured into water and extracted with ethyl acetate; the organic phases were combined after extraction, and finally washed, dried, concentrated to remove the solvent and purified to obtain 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl;

[0017] Step 4: Synthesis of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl

[0018] Dissolve 1-3 g of sodium methoxide and 3-5 g of ethyl trifluoroacetate in 10-30 mL of dimethyl ether, then add 1-3 g of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl and stir at room temperature for 24 h; pour the reaction solution into 10 mL of water, adjust the pH to 2-3, precipitate the solid, filter under reduced pressure and dry to obtain 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl;

[0019] Step 5: Synthesis of ligand L:

[0020] Dissolve 0.1-0.5 g of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl in 10-30 mL of acetonitrile, add 1-3 mL of hydrochloric acid, and stir for 30 min. After the reaction is complete, pour the solution into 10 mL of ice water, then filter under reduced pressure to obtain a solid. Wash with water until pH 7, and finally recrystallize with anhydrous ethanol to obtain ligand L.

[0021] Step 6: Synthesis of (HNEt3)3[Eu3L6]

[0022] Dissolve 0.3-0.6 g of ligand L and 0.2-0.4 g of triethylamine in 10-20 mL of methanol to obtain a reaction solution. Stir until the solution is clear. Dissolve rare earth trifluoromethanesulfonic acid salt in 3-5 mL of methanol at room temperature to obtain a rare earth salt solution. Add the rare earth salt solution to the reaction solution. After the reaction solution is completely clear, continue stirring for 12-24 hours. Then add the reaction solution to 10-20 mL of water to form a flocculent precipitate. After standing, filter to obtain (HNEt3)3[Eu3L6].

[0023] Compared with the prior art, the present invention has the following advantages:

[0024] 1. Innovative chiral recognition function: This invention forms a double-layer triangular supramolecular structure through innovative structural design, which shortens the distance between the upper and lower ligands, thereby increasing the steric hindrance and increasing the cyclization efficiency with thiol amino acids, thus successfully achieving accurate differentiation between thiol amino acids and non-thiol amino acids.

[0025] 2. Excellent anti-interference performance: Because the (HNEt3)3[Eu3L6] of this invention can only undergo cyclization reactions with thiol-containing amino acids and does not react with other amino acids, it can still accurately identify thiol amino acids even in complex sample matrices. Its selective detection performance is not affected by environmental factors. Faced with other coexisting substances such as structurally similar amino acids, the detection system exhibits significant anti-interference tolerance, effectively overcoming the technical bottleneck of traditional detection probes being susceptible to environmental interference, and providing a reliable guarantee for the accurate detection of thiol amino acids.

[0026] 3. Innovative Dynamic Response Mechanism: This invention (HNEt3)3[Eu3L6] achieves recognition through the cyclization reaction of an aldehyde probe with an amino group containing a thiol amino acid and a thiol thiazoline. The reaction forms a rigid five-membered ring. Unlike the flexible structure of the Schiff base probe with its "single-point binding," the thiol cyclization firmly fixes the chiral center to the probe through "cyclization locking," significantly amplifying the energy difference between matched and unmatched enantiomers, thereby significantly improving chiral amplification efficiency. This design brings multiple advantages, including specific cyclization triggering a strong CPL signal, improving detection sensitivity. Therefore, based on the dual recognition of bifunctional groups and chiral matching, (HNEt3)3[Eu3L6] achieves precise capture of thiol amino acids in complex matrices, solving the selectivity problem while ensuring that the signal truly reflects the concentration and configuration of the target molecule, enhancing the reliability and practicality of detection.

[0027] 4. Improved water solubility: This invention adopts a novel structural design and utilizes in-situ hemiacetalization of aldehyde groups to generate hydroxyl groups, thereby improving the overall water solubility and effectively solving the problems of poor water solubility and insufficient water stability, laying a solid foundation for the accurate identification of water-soluble amino acids. Attached Figure Description

[0028] Figure 1 This is a graph showing the non-specific detection results of (HNEt3)3[Eu3L6] obtained in Example 1;

[0029] Figure 2 The selective recognition anti-interference test diagram of (HNEt3)3[Eu3L6] prepared in Example 1 is shown.

[0030] Figure 3 The linear relationship between ΔI and D-PA concentration for (HNEt3)3[Eu3L6] prepared in Example 1. Detailed Implementation

[0031] The technical solution of the present invention is not limited to the specific embodiments listed below, but also includes any reasonable combination of the specific embodiments.

[0032] Specific Implementation Method 1: The structural formula of the water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe in this implementation method is as follows:

[0033] ;

[0034] The general structural formula of the water-soluble rare-earth trinuclear supramolecular CPL probe containing thiol amino acids is (HNEt3)3[Eu3L6], where L is a ligand, and the structural formula of L is:

[0035] .

[0036] 1. Innovative chiral recognition function: This implementation method forms a double-layer triangular supramolecular structure through innovative structural design, which shortens the distance between the upper and lower ligands, thereby increasing the steric hindrance and increasing the cyclization efficiency with thiol amino acids, thus successfully achieving accurate differentiation between thiol amino acids and non-thiol amino acids.

[0037] 2. Excellent anti-interference performance: Because the (HNEt3)3[Eu3L6] in this embodiment can only undergo cyclization reactions with thiol-containing amino acids and does not react with other amino acids, it can still accurately identify thiol amino acids even in complex sample matrices. Its selective detection performance is not affected by environmental factors. Faced with other coexisting substances such as structurally similar amino acids, the detection system exhibits significant anti-interference tolerance, effectively overcoming the technical bottleneck of traditional detection probes being susceptible to environmental interference, and providing a reliable guarantee for the accurate detection of thiol amino acids.

[0038] 3. Innovative Dynamic Response Mechanism: This implementation (HNEt3)3[Eu3L6] achieves recognition through the cyclization reaction of an aldehyde probe with the amino group containing a thiol amino acid and a thiol thiazoline. The reaction forms a rigid five-membered ring. Unlike the flexible structure of the Schiff base probe with "single-point binding," the thiol cyclization firmly fixes the chiral center to the probe through "cyclization locking," significantly amplifying the energy difference between matched and unmatched enantiomers, thereby significantly improving chiral amplification efficiency. This design brings multiple advantages, including specific cyclization triggering a strong CPL signal and improving detection sensitivity. Therefore, based on the dual recognition of bifunctional groups and chiral matching, (HNEt3)3[Eu3L6] achieves precise capture of thiol amino acids in complex matrices, solving the selectivity problem while ensuring that the signal truly reflects the concentration and configuration of the target molecule, enhancing the reliability and practicality of detection.

[0039] 4. Improved water solubility: This implementation method adopts a novel structural design, which utilizes in-situ hemiacetalization of aldehyde groups to generate hydroxyl groups, thereby improving the overall water solubility and effectively solving the problems of poor water solubility and insufficient water stability, laying a solid foundation for the accurate identification of water-soluble amino acids.

[0040] Specific Implementation Method Two: The preparation method of the water-soluble rare earth trinuclear supramolecular thiol amino acid-responsive CPL probe in this implementation method is carried out according to the following steps:

[0041] Step 1: Synthesis of 4-hydroxy-3-iodoacetophenone

[0042] Add 3-5g of 4-hydroxyacetophenone, 4-6g of elemental iodine and 10-20mL of hydrogen peroxide aqueous solution to 100-200mL of water, stir at room temperature for 24h, filter to obtain solid, and recrystallize by anhydrous ethanol to obtain 4-hydroxy-3-iodoacetophenone.

[0043] Step 2: Synthesis of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone

[0044] Add 4-6 g of 4-hydroxy-3-iodoacetophenone and 8-10 g of Cs₂CO₃ to a Schlenk flask, add 100-200 mL of N,N-dimethylformamide, and stir at 110 °C for 0.5 h. Then add 5-7 g of 2-bromo-1,1-dimethoxyethane and react for 5 h. Quench the reaction solution in water and extract with dichloromethane. Combine the organic layers after extraction, and finally wash, dry, concentrate to remove solvent and purify to obtain 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)acetophenone.

[0045] Step 3: Synthesis of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl

[0046] 1-3 g of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone, 0.2-0.4 g of Pd(PPh3)4 and 1-2 g of 3-acetylphenylboronic acid were placed in a Schlenk flask; 40-60 mL of tetrahydrofuran and 20-30 mL of sodium carbonate aqueous solution were added sequentially, and the mixture was refluxed and stirred for 12 h; the reaction solution was then poured into water and extracted with ethyl acetate; the organic phases were combined after extraction, and finally washed, dried, concentrated to remove the solvent and purified to obtain 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl;

[0047] Step 4: Synthesis of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl

[0048] Dissolve 1-3 g of sodium methoxide and 3-5 g of ethyl trifluoroacetate in 10-30 mL of dimethyl ether, then add 1-3 g of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl and stir at room temperature for 24 h; pour the reaction solution into 10 mL of water, adjust the pH to 2-3, precipitate the solid, filter under reduced pressure and dry to obtain 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl;

[0049] Step 5: Synthesis of ligand L:

[0050] Dissolve 0.1-0.5 g of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl in 10-30 mL of acetonitrile, add 1-3 mL of hydrochloric acid, and stir for 30 min. After the reaction is complete, pour the solution into 10 mL of ice water, then filter under reduced pressure to obtain a solid. Wash with water until pH 7, and finally recrystallize with anhydrous ethanol to obtain ligand L.

[0051] Step 6: Synthesis of (HNEt3)3[Eu3L6]

[0052] Dissolve 0.3-0.6 g of ligand L and 0.2-0.4 g of triethylamine in 10-20 mL of methanol to obtain a reaction solution. Stir until the solution is clear. Dissolve rare earth trifluoromethanesulfonic acid salt in 3-5 mL of methanol at room temperature to obtain a rare earth salt solution. Add the rare earth salt solution to the reaction solution. After the reaction solution is completely clear, continue stirring for 12-24 hours. Then add the reaction solution to 10-20 mL of water to form a flocculent precipitate. After standing, filter to obtain (HNEt3)3[Eu3L6].

[0053] 1. Innovative chiral recognition function: This implementation method forms a double-layer triangular supramolecular structure through innovative structural design, which shortens the distance between the upper and lower ligands, thereby increasing the steric hindrance and increasing the cyclization efficiency with thiol amino acids, thus successfully achieving accurate differentiation between thiol amino acids and non-thiol amino acids.

[0054] 2. Excellent anti-interference performance: Because the (HNEt3)3[Eu3L6] in this embodiment can only undergo cyclization reactions with thiol-containing amino acids and does not react with other amino acids, it can still accurately identify thiol amino acids even in complex sample matrices. Its selective detection performance is not affected by environmental factors. Faced with other coexisting substances such as structurally similar amino acids, the detection system exhibits significant anti-interference tolerance, effectively overcoming the technical bottleneck of traditional detection probes being susceptible to environmental interference, and providing a reliable guarantee for the accurate detection of thiol amino acids.

[0055] 3. Innovative Dynamic Response Mechanism: This implementation (HNEt3)3[Eu3L6] achieves recognition through the cyclization reaction of an aldehyde probe with the amino group containing a thiol amino acid and a thiol thiazoline. The reaction forms a rigid five-membered ring. Unlike the flexible structure of the Schiff base probe with "single-point binding," the thiol cyclization firmly fixes the chiral center to the probe through "cyclization locking," significantly amplifying the energy difference between matched and unmatched enantiomers, thereby significantly improving chiral amplification efficiency. This design brings multiple advantages, including specific cyclization triggering a strong CPL signal and improving detection sensitivity. Therefore, based on the dual recognition of bifunctional groups and chiral matching, (HNEt3)3[Eu3L6] achieves precise capture of thiol amino acids in complex matrices, solving the selectivity problem while ensuring that the signal truly reflects the concentration and configuration of the target molecule, enhancing the reliability and practicality of detection.

[0056] 4. Improved water solubility: This implementation method adopts a novel structural design, which utilizes in-situ hemiacetalization of aldehyde groups to generate hydroxyl groups, thereby improving the overall water solubility and effectively solving the problems of poor water solubility and insufficient water stability, laying a solid foundation for the accurate identification of water-soluble amino acids.

[0057] Specific Implementation Method 3: This implementation method differs from Specific Implementation Method 2 in that the mass fraction of the hydrogen peroxide aqueous solution in step 1 is 30%.

[0058] Specific Implementation Method Four: This implementation method differs from Specific Implementation Method Two in that the reagent used for washing in steps two and three is a saturated sodium chloride solution.

[0059] Specific Implementation Method 5: This implementation method differs from Specific Implementation Method 2 in that the drying reagent used in steps 2 and 3 is anhydrous sodium sulfate.

[0060] Specific Implementation Method Six: This implementation method differs from Specific Implementation Method Two in that the purification described in steps two and three uses silica gel column chromatography.

[0061] Specific Implementation Method Seven: This implementation method differs from Specific Implementation Method Two in that the concentration of the sodium carbonate aqueous solution in step three is 2 mol / L.

[0062] Specific Implementation Method Eight: This implementation method differs from Specific Implementation Method Two in that the pH adjustment in step four uses hydrochloric acid.

[0063] Specific Implementation Method Nine: This implementation method differs from Specific Implementation Method Two in that the concentration of hydrochloric acid in step five is 3 mol / L.

[0064] Specific Implementation Method 10: This implementation method differs from Specific Implementation Method 2 in that the rare earth trifluoromethanesulfonic acid salt mentioned in step 6 is Eu(OTf)3.

[0065] Example 1

[0066] The structural formula of the water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe in this embodiment is as follows:

[0067] ;

[0068] The general structural formula of the water-soluble rare-earth trinuclear supramolecular CPL probe containing thiol amino acids is (HNEt3)3[Eu3L6], where L is a ligand, and the structural formula of L is:

[0069] .

[0070] The preparation method of the water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe in this embodiment is carried out according to the following steps:

[0071] Step 1: Synthesis of 4-hydroxy-3-iodoacetophenone

[0072] 5 g of 4-hydroxyacetophenone, 4.66 g of elemental iodine and 11.5 mL of hydrogen peroxide aqueous solution were added to 150 mL of water and stirred at room temperature for 24 h. The solid was filtered and recrystallized from anhydrous ethanol to obtain 4-hydroxy-3-iodoacetophenone, which was a white crystal with a yield of 72%.

[0073] The mass fraction of the hydrogen peroxide aqueous solution is 30%.

[0074] Step 2: Synthesis of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone

[0075] Under a nitrogen atmosphere, 5 g of 4-hydroxy-3-iodoacetophenone and 9.33 g of Cs₂CO₃ were added to a Schlenk flask, followed by the addition of 150 mL of N,N-dimethylformamide and stirring at 110 °C for 0.5 h. Then, 6.45 g of 2-bromo-1,1-dimethoxyethane was added and the reaction was carried out for 5 h. The reaction solution was quenched in water and extracted with dichloromethane. After extraction, the organic layers were combined, and finally washed, dried, concentrated to remove the solvent and purified to obtain 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)acetophenone, a white crystalline solid with a yield of 75%.

[0076] The washing reagent used is a saturated sodium chloride solution;

[0077] The drying reagent used is anhydrous sodium sulfate;

[0078] The purification was performed using silica gel column chromatography.

[0079] Step 3: Synthesis of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl

[0080] Under a nitrogen atmosphere, 2 g of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone, 0.33 g of Pd(PPh3)4, and 1.12 g of 3-acetylphenylboronic acid were placed in a Schlenk flask; 50 mL of tetrahydrofuran and 25 mL of sodium carbonate aqueous solution were added sequentially, and the mixture was refluxed and stirred for 12 h; the reaction solution was poured into water and extracted with ethyl acetate; the organic phases were combined after extraction, and finally washed, dried, concentrated to remove the solvent, and purified to obtain 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl; it was a white solid; the yield was 73%.

[0081] The concentration of the sodium carbonate aqueous solution is 2 mol / L;

[0082] The washing reagent used is a saturated sodium chloride solution;

[0083] The drying reagent used is anhydrous sodium sulfate;

[0084] The purification was performed using silica gel column chromatography.

[0085] Step 4: Synthesis of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl

[0086] Dissolve 1.26 g of sodium methoxide and 3.32 g of ethyl trifluoroacetate in 20 mL of dimethyl ether, then add 1 g of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl and stir at room temperature for 24 h. Pour the reaction solution into 10 mL of water, adjust the pH to 2, precipitate the solid, filter under reduced pressure and dry to obtain 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl; a white solid; yield 80%.

[0087] The pH adjustment is performed using hydrochloric acid;

[0088] Step 5: Synthesis of ligand L:

[0089] 0.5 g of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl was dissolved in 10 mL of acetonitrile. 1.5 mL of hydrochloric acid was added, and the mixture was stirred for 30 min. After the reaction was complete, the solution was poured into 10 mL of ice water, and the mixture was filtered under reduced pressure to obtain a solid. The solid was washed with water until the pH reached 7, and finally recrystallized from anhydrous ethanol to obtain ligand L, which was a white solid.

[0090] The concentration of the hydrochloric acid is 3 mol / L;

[0091] Step 6: Synthesis of (HNEt3)3[Eu3L6]

[0092] 0.5 g of ligand L and 0.34 g of triethylamine (HNEt3) were dissolved in 20 mL of methanol to obtain a reaction solution. The solution was stirred until clear. At room temperature, rare earth trifluoromethanesulfonic acid salt was dissolved in 5 mL of methanol to obtain a rare earth salt solution. The rare earth salt solution was added to the reaction solution. After the reaction solution was completely clear, the mixture was stirred for 12 hours. Then, the reaction solution was added to 15 mL of water to form a flocculent precipitate. After standing, the precipitate was filtered to obtain (HNEt3)3[Eu3L6]; the yield was 86%.

[0093] The rare earth trifluoromethanesulfonic acid salt is Eu(OTf)3.

[0094] Figure 1 This is a non-specific detection result image of (HNEt3)3[Eu3L6] prepared in Example 1; where a is the total luminescence intensity I when 40 eq. of chiral amino acids are present. tot (I) tot =I L +I R ( ) curves, b is the total luminescence intensity change ΔI curve after adding amino acids, c is the I / I0-1 curve, and d is the ΔI / ΔI0 curve; as shown Figure 1 As shown, without the addition of amino acids, (HNEt3)3[Eu3L6] showed no CPL signal, indicating that it does not possess chiral optical activity. When alanine, valine, proline, lysine, histidine, tryptophan, and threonine were added, the CPL spectra did not change significantly, indicating that (HNEt3)3[Eu3L6] does not have a specific interaction with these amino acids. However, after adding D-PA and reacting with (HNEt3)3[Eu3L6], I... tot Both ΔI and ΔI increased significantly (2.54 × 10⁻⁶). 6 au, 2.95×10 5 au), through I tot The degree of luminescence enhancement (I) tot The degree of luminescence enhancement is defined as I / I0⁻¹, where I0 is [Eu³L⁶]. 3The initial total luminescence intensity (I / I0-1) of (HNEt3)3[Eu3L6] is significantly higher than that of the other seven chiral amino acids (I / I0-1 = 1.08) when D-PA is introduced into the system, indicating that its host-guest binding with D-PA can effectively improve luminescence efficiency. However, the other amino acids still elicit weak luminescence responses (I / I0-1 between -0.16 and +0.13), indicating the presence of non-specific interactions. Compared to I... tot The relative change in ΔI (defined as ΔI / ΔI0, where ΔI0 is the relative change in the initial total luminescence intensity of (HNEt3)3[Eu3L6]) highlights the probe's high selectivity for D-PA (ΔI / ΔI0 = 253.94), but it also exhibits a ΔI signal response to D-Cys (2.9 × 10⁻⁶). 4 Apart from au), the ΔI / ΔI0 of the remaining amino acids can be ignored. In conclusion, [Eu3L6] 3 The probe not only exhibits highly selective recognition of D-PA, but also responds to D-Cys (D-cysteine), which also contains a thiol group. Therefore, [Eu3L6] 3 ⁻ It can achieve high-precision differentiation between thiol amino acids and non-thiol amino acids.

[0095] Figure 2 The selective recognition anti-interference test diagram of (HNEt3)3[Eu3L6] prepared in Example 1 is shown. First, 40 eq. D-PA was added to (HNEt3)3[Eu3L6], and after 10 minutes, seven other amino acids (40 eq. each) were added. The luminescence asymmetry factor (g) was monitored by CPL spectroscopy. lum The changes in (HNEt3)3[Eu3L6] were observed. Test results showed that after adding 40 eq. D-PA, g lum It reached +0.30, which is an increase compared to the helical form, and its anti-interference ability is also superior. In the presence of a single interfering amino acid (such as D-His, D-Trp, etc.), the system's g... lum It can maintain 92%–98% of its initial value. However, when D-Cys is used as a disruptor, g lum The concentration decreased to 66% of the original level, a reduction compared to the helical form. This is likely due to the response of (HNEt3)3[Eu3L6] to D-Cys. To further evaluate the probe's applicability in complex systems, 40 eq. each of the six amino acids (excluding D-Cys) were mixed and added to a (HNEt3)3[Eu3L6] solution containing D-PA. lum It remains at 87% of the initial value, indicating that the probe still has excellent selectivity for D-PA under conditions of multiple interferences.

[0096] Figure 3 The figure shows the linear relationship between ΔI and D-PA concentration for (HNEt3)3[Eu3L6] prepared in Example 1. The left figure shows the response of (HNEt3)3[Eu3L6] to 0-50 eq. D-PA ΔI, and the right figure shows the relationship between ΔI and concentration of (HNEt3)3[Eu3L6] at 588.5 nm. First, we investigated the effect of D-PA concentration (0–50 equivalents) on the ΔI of the (HNEt3)3[Eu3L6] probe. A significant concentration dependence was observed at 594 nm. In the concentration range of 0–40 eq., ΔI increased with increasing D-PA concentration, indicating that the host-guest binding of the probe to D-PA was in an unsaturated state. The binding tended to plateau above 40 eq., indicating that the binding sites were close to saturation. Therefore, ΔI can be used as a quantitative detection parameter for D-PA.

Claims

1. A water-soluble rare-earth trinuclear supramolecular CPL probe containing thiol amino acids, characterized in that: The structural formula of the water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe is as follows: ； The general structural formula of the water-soluble rare-earth trinuclear supramolecular CPL probe containing thiol amino acids is (HNEt3)2[Eu3L6], where L is a ligand, and the structural formula of L is: 。 2. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe as described in claim 1, characterized in that: The preparation method of water-soluble rare-earth trinuclear supramolecular thiol amino acid-responsive CPL probe is carried out according to the following steps: Step 1: Synthesis of 4-hydroxy-3-iodoacetophenone Add 3-5g of 4-hydroxyacetophenone, 4-6g of elemental iodine and 10-20mL of hydrogen peroxide aqueous solution to 100-200mL of water, stir at room temperature for 24h, filter to obtain solid, and recrystallize by anhydrous ethanol to obtain 4-hydroxy-3-iodoacetophenone. Step 2: Synthesis of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone Add 4-6 g of 4-hydroxy-3-iodoacetophenone and 8-10 g of Cs₂CO₃ to a Schlenk flask, add 100-200 mL of N,N-dimethylformamide, and stir at 110 °C for 0.5 h. Then add 5-7 g of 2-bromo-1,1-dimethoxyethane and react for 5 h. Quench the reaction solution in water and extract with dichloromethane. Combine the organic layers after extraction, and finally wash, dry, concentrate to remove solvent and purify to obtain 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)acetophenone. Step 3: Synthesis of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl 1-3 g of 1-(3-iodo-4-(2,2-dimethoxyethoxy)phenyl)ethyl ketone, 0.2-0.4 g of Pd(PPh3)4 and 1-2 g of 3-acetylphenylboronic acid were placed in a Schlenk flask; 40-60 mL of tetrahydrofuran and 20-30 mL of sodium carbonate aqueous solution were added sequentially, and the mixture was refluxed and stirred for 12 h; the reaction solution was then poured into water and extracted with ethyl acetate; the organic phases were combined after extraction, and finally washed, dried, concentrated to remove the solvent and purified to obtain 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl; Step 4: Synthesis of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl Dissolve 1-3 g of sodium methoxide and 3-5 g of ethyl trifluoroacetate in 10-30 mL of dimethyl ether, then add 1-3 g of 3-acetyl-2-(2,2-dimethoxyethoxy)-3'-acetylbiphenyl and stir at room temperature for 24 h; pour the reaction solution into 10 mL of water, adjust the pH to 2-3, precipitate the solid, filter under reduced pressure and dry to obtain 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl; Step 5: Synthesis of ligand L: Dissolve 0.1-0.5 g of 3-(3-oxo-4,4,4-trifluorobutenyl)-2-(2,2-dimethoxyethoxy)-3'-(3-oxo-4,4,4-trifluorobutenyl)biphenyl in 10-30 mL of acetonitrile, add 1-3 mL of hydrochloric acid, and stir for 30 min. After the reaction is complete, pour the solution into 10 mL of ice water, then filter under reduced pressure to obtain a solid. Wash with water until pH 7, and finally recrystallize with anhydrous ethanol to obtain ligand L. Step 6: Synthesis of (HNEt3)2[Eu3L6] Dissolve 0.3-0.6 g of ligand L and 0.2-0.4 g of triethylamine in 10-20 mL of methanol to obtain a reaction solution. Stir until the solution is clear. Dissolve rare earth trifluoromethanesulfonic acid salt in 3-5 mL of methanol at room temperature to obtain a rare earth salt solution. Add the rare earth salt solution to the reaction solution. After the reaction solution is completely clear, continue stirring for 12-24 hours. Then add the reaction solution to 10-20 mL of water to form a flocculent precipitate. After standing, filter to obtain (HNEt3)2[Eu3L6].

3. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The mass fraction of the hydrogen peroxide aqueous solution in step one is 30%.

4. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The washing process described in steps two and three uses a saturated sodium chloride solution.

5. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The drying reagent used in steps two and three is anhydrous sodium sulfate.

6. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The purification described in steps two and three uses silica gel column chromatography.

7. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The concentration of the sodium carbonate aqueous solution in step three is 2 mol / L.

8. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The pH adjustment in step four uses hydrochloric acid.

9. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The concentration of hydrochloric acid mentioned in step five is 3 mol / L.

10. The method for preparing the water-soluble rare-earth trinuclear supramolecular thiol-containing amino acid-responsive CPL probe according to claim 2, characterized in that: The rare earth trifluoromethanesulfonic acid salt mentioned in step six is ​​Eu(OTf)3.

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