Method for constructing photo-cidnp dye molecule pairs and applications thereof

Abstract

The application discloses a method for constructing a Photo-CIDNP dye molecule pair, which is composed of dye molecules A and B. After the dye molecule A absorbs a photon, the inter-system crossing effect is used to reach the triplet state. The dye molecule B receives the energy transfer of the triplet state dye molecule A to realize the triplet state, and then a Photo-CIDNP hyperpolarization process occurs with a target molecule to realize the significant enhancement of the nuclear magnetic resonance signal of the target molecule. The dye molecule A is a triplet state sensitizer, which can be selected from a ruthenium metal complex. The dye molecule B can be selected from a naphthalimide derivative, a pyrene derivative or a rhodamine derivative. The Photo-CIDNP dye molecule pair constructed by the method can significantly enhance the nuclear magnetic resonance signal intensity of the target molecule, and effectively solves the low sensitivity problem inherent in nuclear magnetic resonance.

Description

Technical Field

[0001] This invention belongs to the field of nuclear magnetic resonance hyperpolarization technology, specifically relating to a method for constructing photochemically induced dynamic nuclear polarization (Photo-CIDNP) dye molecule pairs and their applications. Background Technology

[0002] Nuclear magnetic resonance (NMR) has been widely used in various fields, such as material structure analysis and sample composition identification. The sensitivity of NMR is related to the magnetic nuclei (such as...) 1 H, 19 F, 13 C and 15 The nuclear spin polarizability of N is directly proportional to that of magnetic atoms. At room temperature and commercial magnetic field strengths, the nuclear spin polarizability of magnetic atoms is particularly low; for example, at room temperature and a magnetic field of 9.4 T, the nuclear spin polarizability of hydrogen atoms is... 1 The nuclear spin polarizability of H is only three parts per ten thousand, resulting in low sensitivity of NMR. The low sensitivity of NMR severely limits its in-depth application in various fields. Improving nuclear spin polarizability is one of the most direct and effective means to enhance NMR sensitivity. The means of improving nuclear spin polarizability is called hyperpolarization technology. At present, hyperpolarization technology includes dynamic nuclear polarization (DNP), fused dynamic nuclear polarization (d-DNP), spin polarization exchange light pumping (SEOP), secondary hydrogen-induced hyperpolarization (PHIP), and photochemical-induced dynamic nuclear polarization (Photo-CIDNP). Among them, Photo-CIDNP is currently the only hyperpolarization technology that can repeatedly hyperpolarize and sample in situ in room temperature solution, and has advantages that other hyperpolarization technologies cannot match (Eills James, et al. A1, Chemical Reviews 2023, 123(4), 1417-1551.). Photo-CIDNP can theoretically improve the sensitivity of NMR by several orders of magnitude. The hyperpolarization mechanism of Photo-CIDNP can be explained by the radical pair theory (RP). Under illumination, dye molecules undergo electron transfer reactions with the target molecule to generate radical pairs. These radical pairs recombine to achieve hyperpolarization of the target molecule. The performance of Photo-CIDNP in enhancing NMR signals is related to the selected target molecule, dye molecule, and magnetic field strength. The magnetic field strength is determined by the NMR spectrometer used; therefore, developing suitable dye molecules is crucial for achieving Photo-CIDNP hyperpolarization of the target molecule. Currently, no experience or rules for Photo-CIDNP dye molecule design have been reported. It is worth noting that when Photo-CIDNP enhances the target molecule, it simultaneously enhances all magnetic nuclei in the radical-forming region of the target molecule, including… 1 H, 19 F, 13 C and 15N, etc., are usually detected in target molecules. 1 An enhanced H NMR signal indicates an increase in the corresponding magnetic nuclei of the target molecule, such as... 13 The C signal will also be enhanced, but the enhancement magnitude will vary. This is determined by the characteristics of Photo-CIDNP hyperpolarization technology (GLCloss, Chemically Induced Dynamic Nuclear Polarization, 1974, Academic Press).

[0003] Currently reported target molecules for Photo-CIDNP hyperpolarization are mainly several amino acids, including tryptophan, tyrosine, histidine, cysteine, and methionine. For these amino acids, only a few Photo-CIDNP dye molecules have been reported so far, including riboflavin, fluorescein, ATTO-12, ruthenium metal complexes, bipyridine, and benzophenone (Okuno, Yusuke et al., Photochemically Induced Dynamic Nuclear Polarization: Basic Principles and Applications. In eMagRes, 2017, 283-314. Sobol, Aet. al., AttoThio 12 as a promising dye for photo-CIDNP, J Chem Phys 2019, 151(23), 234201.). All reports describe Photo-CIDNP using a single dye molecule. This requires the dye molecule to complete the entire process from photon absorption to target molecule hyperpolarization, necessitating a range of physicochemical properties such as visible light extinction coefficient, ISC efficiency, suitable redox potential, and free radical stability. The stringent screening conditions of the single-dye molecule model not only limit the development range of Photo-CIDNP dye molecules but also restrict the enhancement of the target molecule's NMR signal. Developing new methods for Photo-CIDNP dye molecule development can expand the range of Photo-CIDNP dye molecules, achieve significant enhancement of the target molecule's NMR signal, and advance the application of Photo-CIDNP hyperpolarization technology. Summary of the Invention

[0004] To address the aforementioned problems, the first objective of this invention is to provide a method for constructing Photo-CIDNP dye molecule pairs. This method significantly differs from previously reported single-molecule Photo-CIDNP methods; this invention uses a pair of dye molecules to form a dye molecule pair to achieve Photo-CIDNP.

[0005] Photo-CIDNP dye molecule pair design principles, such as Figure 1 As shown, the Photo-CIDNP dye molecule pair includes dye molecule A and dye molecule B. Under illumination, dye molecule A absorbs a photon and transitions to a singlet excited state, then transitions to a triplet state through intersystem crossing (ISC). Dye molecule B accepts energy from triplet dye molecule A through triplet energy transfer to form a triplet state. Triplet dye molecule B then undergoes an electron transfer reaction with the target molecule to form a radical pair. During recombination, the radical pair achieves hyperpolarization of the target molecule, significantly enhancing the nuclear self-resonance signal intensity of the target molecule. In the hyperpolarization process of the Photo-CIDNP dye molecule pair, dye molecule A is responsible for photon absorption and triplet state generation. Dye molecule A can be considered a triplet sensitizer, used to sensitize the generation of triplet dye molecule B. Dye molecule B directly participates in the hyperpolarization process of the target molecule, playing a decisive role.

[0006] A second objective of this invention is to enhance the NMR signal of target molecules using the aforementioned Photo-CIDNP dye pair. The aforementioned Photo-CIDNP dye pair has been used to significantly enhance the NMR signals of tyrosine, indole, tryptophan, and their derivatives.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] Photochemically induced dynamic nuclear polarization (Photo-CIDNP) dye molecule pairs, consisting of dye molecules A and B, wherein dye molecule A is a ruthenium metal complex selected from one or more of the structures shown below:

[0009]

[0010] The dye molecule B is one or more of a naphthalimide derivative, a pyrene derivative, or a rhodamine derivative;

[0011] Dye molecule A has high triplet quantum efficiency and can rapidly transition to the triplet state after absorbing light energy; dye molecule B accepts the triplet energy of dye molecule A and transitions to the triplet state. A reversible electron transfer reaction occurs between triplet dye molecule B and the target molecule, resulting in a significant enhancement of the NMR signal of the target molecule.

[0012] Preferably, the dye molecule A is ruthenium triphenanthroline or ruthenium triphenanthroline, with the following structure:

[0013]

[0014] Preferably, the dye molecule B is naphthalimide, with the following structure:

[0015]

[0016] The group R1 is one of phosphate, sulfonate or carboxylate.

[0017] Preferably, the dye molecule B is a pyrene derivative with the following structure:

[0018]

[0019] Groups R2, R3, R4, or R5 are one of hydrogen atoms, carboxylate groups, sulfonate groups, or phosphate groups.

[0020] Preferably, the dye molecule B is a rhodamine derivative with the following structure:

[0021]

[0022] The group R6 or R7 is one of hydrogen atom, methyl, ethyl, propyl, isopropyl, butyl or isobutyl.

[0023] In the above-mentioned method for preparing Photo-CIDNP dye molecule pairs, the concentration ratio of dye molecule A to dye molecule B is 1000:1 to 1:1000, preferably 50:200.

[0024] The above Photo-CIDNP dye pairs are used to enhance NMR signals of tyrosine and its derivatives, indole and its derivatives, or tryptophan and its derivatives.

[0025] Furthermore, the structures of the above-mentioned indole and tryptophan derivatives are shown in the figure below:

[0026]

[0027] Among them, groups R8, R9, and R 10 R 11 It is either a hydrogen atom or a fluorine atom, preferably R. 10 It is a fluorine atom, R8, R9 and R 11 It is a hydrogen atom, namely 6-F tryptophan or 6-F indole.

[0028] Furthermore, the aforementioned NMR signal is 1 H NMR, 19 F NMR, 13 C NMR or 15 One or more of N NMR, preferably 1 H NMR and 19 F NMR.

[0029] The beneficial effects of this invention are as follows:

[0030] This invention provides a method for constructing Photo-CIDNP dye molecule pairs. Using this method, dye molecules lacking Photo-CIDNP properties can be combined in an appropriate manner to construct Photo-CIDNP dye molecule pairs, which exhibit excellent NMR signal enhancement performance. This method not only expands the selection range of Photo-CIDNP dye molecules but also has the potential to significantly enhance the NMR signals of specific target molecules. This invention provides various Photo-CIDNP dye molecule pairs that significantly enhance the NMR signals of corresponding target molecules, demonstrating the method's good universality. Attached Figure Description

[0031] Figure 1 This is the design principle of the Photo-CIDNP dye molecule pairs described in this invention.

[0032] Figure 2 Showing a single ruthenium triphenanthroline ( Figure 2 A) or pyrene tetrasulfonate ( Figure 2 B) Tyrosine under light / no light conditions 1 H NMR spectral enhancement performance test; dye molecule pair composed of triphenanthroline ruthenium and tetrasulfonate pyrene ( Figure 2 C) Tyrosine under light / no light conditions 1 H NMR spectral enhancement performance test. Red line corresponds to illumination conditions, blue line corresponds to no illumination conditions.

[0033] Figure 3 This shows the effects of different concentration ratios of ruthenium triphenanthroline and pyrene tetrasulfonate on tyrosine. 1 H NMR enhancement amplitude.

[0034] Figure 4 This shows the effect of different concentrations of ruthenium triphenanthroline and pyrene tetrasulfonate on tyrosine under a fixed ratio of ruthenium triphenanthroline and pyrene tetrasulfonate. 1 H NMR enhancement amplitude.

[0035] Figure 5 Showing a single ruthenium triphenanthroline ( Figure 5 A) or pyrene tetrasulfonate ( Figure 5 B) 6-F tryptophan under both light and dark conditions 19 F NMR spectral enhancement performance test; dye molecule pair composed of triphenanthroline ruthenium and tetrasulfonate pyrene ( Figure 5 C) 6-F tryptophan under both light and dark conditions 19 F NMR spectral enhancement performance test. Red line corresponds to illumination conditions, blue line corresponds to no illumination conditions.

[0036] Figure 6This demonstrates the effect of a dye molecule pair composed of phenanthrene, ruthenium triphenate, and pyrene tetrasulfonate on tryptophan under both light- and dark conditions. 1 HNMR spectral enhancement performance test. The red line corresponds to the illumination condition, and the blue line corresponds to the no-illumination condition.

[0037] Figure 7 The dye pair consisting of ruthenium tripyridine and pyrene tetrasulfonate is shown to inhibit the activity of 6-F tryptophan under both light- and dark conditions. 19 F NMR spectral enhancement performance test. Red line corresponds to illumination conditions, blue line corresponds to no illumination conditions.

[0038] Figure 8 This demonstrates the effect of the dye pair composed of triphenanthroline ruthenium and naphthalimide on 6-F indole under both presence and absence of light / light conditions. 19 FNMR spectral enhancement performance test. The red line corresponds to the illumination condition, and the blue line corresponds to the no-illumination condition.

[0039] Figure 9 This demonstrates the effect of the triphenanthroline ruthenium and sulfonylrhodamine B dye pair on 6-F indole under both light and dark conditions. 19 F NMR spectral enhancement performance test. Red line corresponds to illumination conditions, blue line corresponds to no illumination conditions. Detailed Implementation

[0040] To more clearly illustrate the present invention, the following description, in conjunction with preferred embodiments and accompanying drawings, further clarifies the invention. Those skilled in the art should understand that the specific description below is illustrative rather than restrictive and should not be construed as limiting the scope of protection of the present invention.

[0041] The following formula is used to calculate the enhancement magnitude (or enhancement factor) of the nuclear magnetic resonance signal:

[0042] ε = Peak area after illumination / Peak area before illumination

[0043] Example 1

[0044] The Photo-CIDNP dye molecule pairs and substrate structures used in Example 1 are shown below:

[0045]

[0046] Photo-CIDNP dye molecules are composed of ruthenium triphenanthroline and pyrene tetrasulfonate, with tyrosine as the target molecule, i.e., the polarization substrate.

[0047] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthroline, pyrene tetrasulfonate, and tyrosine, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 50 μM ruthenium triphenanthroline, 50 μM pyrene tetrasulfonate, 50 μM tyrosine, 3 mM glucose, 0.2 μM glucose oxidase, and 0.125 μM catalase. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared samples were stored at room temperature in the dark for 1 hour before testing.

[0048] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 1 H NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 1 HNMR spectrum.

[0049] Analysis of experimental results: Figure 2 A demonstrates the effect of ruthenium triphenanthroline alone on tyrosine under light irradiation. 1 The H NMR spectrum did not exhibit enhancement properties. Figure 2 B demonstrates the effect of pyrene tetrasulfonate alone on tyrosine under light irradiation. 1 The 1H NMR spectrum did not show any enhancement. This result indicates that neither ruthenium triphenanthroline nor pyrene tetrasulfonate alone can enhance the NMR signal of tyrosine. Figure 2 C demonstrates that the dye pair composed of ruthenium triphenanthroline and pyrene tetrasulfonate significantly enhances tyrosine under light conditions. 1 The H NMR spectrum signal showed an 8.1-fold increase in the NMR signal of the hydrogen atom at the 2,6 position of tyrosine.

[0050] Example 2

[0051] Example 2 used the same Photo-CIDNP dye pairs and substrates as in Example 1. The purpose of Example 2 was to investigate the effect of different concentration ratios of ruthenium triphenanthroline and pyrene tetrasulfonate on the enhancement amplitude of tyrosine NMR signals.

[0052] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthroline, pyrene tetrasulfonate, and tyrosine, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 50 μM ruthenium triphenanthroline, 50 μM tyrosine, 3 mM glucose, 0.2 μM glucose oxidase, 0.125 μM catalase, and 0, 12.5, 25, 50, 100, 150, 200, 250, and 300 μM pyrene tetrasulfonate, respectively. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared samples were stored at room temperature in the dark for 1 hour before testing.

[0053] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 1 H NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 1 HNMR spectrum.

[0054] Analysis of experimental results: Figure 3 The curves showing the enhancement amplitude of tyrosine NMR signal are presented as the concentration ratio of ruthenium triphenanthroline and pyrene tetrasulfonate increases from 1:0 to 1:6. The results indicate that the enhancement amplitude of tyrosine NMR signal gradually increases to a plateau with the increase of the concentration ratio of the two dye molecules. When the ratio of ruthenium triphenanthroline and pyrene tetrasulfonate is 1:4, the enhancement amplitude of tyrosine NMR signal has stabilized.

[0055] Example 3

[0056] Example 3 used the same Photo-CIDNP dye molecule pairs and substrates as in Example 1. The purpose of Example 2 was to investigate the effect of increasing the concentrations of ruthenium triphenanthroline and pyrene tetrasulfonate on the enhancement amplitude of tyrosine NMR while maintaining a fixed concentration ratio of ruthenium triphenanthroline and pyrene tetrasulfonate.

[0057] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthroline, pyrene tetrasulfonate, and tyrosine, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 50 μM tyrosine, 3 mM glucose, 0.2 μM glucose oxidase, and 0.125 μM catalase. Maintain a fixed ruthenium triphenanthroline to pyrene tetrasulfonate concentration ratio of 1:4. The concentrations of ruthenium triphenanthroline are 0, 5, 10, 20, 30, 40, 50, 60, 70, 80, and 100 μM, with corresponding increases in pyrene tetrasulfonate concentration. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared sample was stored at room temperature in the dark for 1 hour before testing.

[0058] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 1 H NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~1.2 W). 1 HNMR spectrum.

[0059] Analysis of experimental results: Figure 4 When the concentration ratio of ruthenium triphenanthroline to pyrene tetrasulfonate was fixed at 1:4, gradually increasing the concentration of ruthenium triphenanthroline and pyrene tetrasulfonate resulted in a gradual increase in the enhancement amplitude of the NMR signal of tyrosine, which eventually reached a plateau.

[0060] Example 4

[0061] The Photo-CIDNP dye molecule pairs and substrate structures used in Example 4 are shown below:

[0062]

[0063] Photo-CIDNP dye molecules are composed of ruthenium triphenanthroline and pyrene tetrasulfonate, with 6-F tryptophan as the target molecule, i.e., the polarization substrate.

[0064] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthroline, pyrene tetrasulfonate, and 6-F tryptophan, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 30 μM ruthenium triphenanthroline, 30 μM pyrene tetrasulfonate, 50 μM 6-F tryptophan, 3 mM glucose, 0.2 μM glucose oxidase, and 0.125 μM catalase. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared samples were stored at room temperature in the dark for 1 hour before testing.

[0065] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 19 F NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 19 F NMR spectrum.

[0066] Analysis of experimental results: Figure 5 A demonstrates the effect of ruthenium triphenanthroline alone on 6-F tryptophan under light irradiation. 19 The FNMR spectrum did not exhibit enhanced performance. Figure 5 B demonstrates the effect of pyrene tetrasulfonate alone on 6-F tryptophan under light irradiation. 19 The FNMR spectrum did not show any enhancement. This result indicates that neither ruthenium triphenanthroline nor pyrene tetrasulfonate alone can enhance the NMR signal of 6-F tryptophan. Figure 5 C demonstrates that the dye pair composed of ruthenium triphenanthroline and pyrene tetrasulfonate significantly enhances 6-F tryptophan expression under light conditions. 19 The FNMR spectrum signal showed that the NMR signal of fluorine atoms in 6-F tryptophan was enhanced by 119.4 times.

[0067] Example 5

[0068] The Photo-CIDNP dye molecule pairs and substrate structures used in Example 5 are shown below:

[0069]

[0070] Photo-CIDNP dye molecules are composed of ruthenium triphenanthroline and pyrene tetrasulfonate, with tryptophan as the target molecule, i.e., the polarizing substrate.

[0071] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthroline, pyrene tetrasulfonate, and tryptophan, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 50 μM ruthenium triphenanthroline, 50 μM pyrene tetrasulfonate, 50 μM tryptophan, 3 mM glucose, 0.2 μM glucose oxidase, and 0.125 μM catalase. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared samples were stored at room temperature in the dark for 1 hour before testing.

[0072] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 1 H NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 1 HNMR spectrum.

[0073] Analysis of experimental results: Figure 6 The dye pair composed of ruthenium triphenanthroline and pyrene tetrasulfonate was shown to significantly enhance tryptophan expression under light irradiation. 1 The H NMR spectrum signal showed a 5.8-fold increase in the signal at the 6-hydrogen atom of tryptophan.

[0074] Example 6

[0075] The Photo-CIDNP dye molecule pairs and substrate structures used in Example 6 are shown below:

[0076]

[0077] Photo-CIDNP dye molecules are composed of ruthenium tripyridine and pyrene tetrasulfonate, with 6-F tryptophan as the target molecule, i.e., the polarization substrate.

[0078] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenylpyridine, pyrene tetrasulfonate, and 6-F tryptophan, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 30 μM ruthenium triphenylpyridine, 30 μM pyrene tetrasulfonate, 30 μM 6-F tryptophan, 3 mM glucose, 0.2 μM glucose oxidase, and 0.125 μM catalase. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared samples were stored at room temperature in the dark for 1 hour before testing.

[0079] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 19 F NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 19 F NMR spectrum.

[0080] Analysis of experimental results: Figure 7 The dye pair composed of ruthenium tripyridine and pyrene tetrasulfonate was shown to significantly enhance 6-F tryptophan under light conditions. 19 The NMR spectrum signal of 6-F tryptophan showed a 40.5-fold increase in the NMR signal of fluorine atoms.

[0081] Example 7

[0082] The Photo-CIDNP dye molecule pairs and substrate structures used in Example 7 are shown below:

[0083]

[0084] Photo-CIDNP dye pairs consist of ruthenium triphenanthroline and naphthalene imide, with 6-F indole as the target molecule, i.e., the polarization substrate. Naphthalene imide was synthesized according to the literature method (Zhang, L et al., Rationally Designed Surfactants for Few-Layered Graphene Exfoliation: Ionic Groups Attached to Electron-Deficient π-Conjugated Unit through Alkyl Spacers. ACS Nano 2014, 8(7), 6663-6670.).

[0085] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthroline and naphthaleneimide, and dissolve them separately in 200 mM phosphate buffer (pH 7.4) to prepare 2 mM stock solutions. Weigh out a certain amount of 6-F indole and dissolve it in dimethyl sulfoxide to prepare a 2 mM stock solution. Add the samples sequentially to a 5 mm NMR sample tube, with a total sample volume of 400 μL. The final sample concentrations added are as follows: 10% v / v heavy water, 50 μM ruthenium triphenanthroline, 200 μM naphthaleneimide, 50 μM 6-F indole, 3 mM glucose, 0.2 μM glucose oxidase, and 0.125 μM catalase. Finally, add a certain amount of 200 mM phosphate buffer (pH 7.4) to bring the final sample volume to 400 μL. The prepared samples were stored at room temperature in the dark for 1 hour before testing.

[0086] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 19 F NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 19 F NMR spectrum.

[0087] Analysis of experimental results: Figure 8 The dye pair composed of triphenanthroline ruthenium and naphthalimide was shown to significantly enhance the 6-F indole production under light conditions. 19 The NMR spectrum signal of 6-F indole showed a 22-fold increase in the NMR signal of fluorine atoms.

[0088] Example 8

[0089] The Photo-CIDNP dye molecule pairs and substrate structures used in Example 8 are shown below:

[0090]

[0091] Photo-CIDNP dye molecules are composed of triphenanthrene-ruthenium triphenate and sulfonylrhodamine B, with 6-F indole being the target molecule, i.e., the polarizing substrate.

[0092] Experimental preparation procedure: Weigh out a certain amount of ruthenium triphenanthrene-1,4- ...

[0093] Sample testing: Insert the optical fiber into the sample NMR tube, then place it in the NMR spectrometer. After field locking and shimming are completed, sampling begins. Control data are collected in the absence of light. 19 F NMR spectra were acquired and enhanced under 488 nm laser excitation (power ~0.7 W). 19 F NMR spectrum.

[0094] Analysis of experimental results: Figure 9 The dye pair composed of triphenanthroline ruthenium and sulfonylrhodamine B significantly enhanced 6-F indole under light conditions. 19The NMR spectrum signal of 6-F indole showed a 5-fold increase in the NMR signal of fluorine atoms.

[0095] In this invention, we replaced two dye molecules A, namely ruthenium triphenanthroline and ruthenium terpyridine, and three dye molecules B, namely pyrene tetrasulfonate, naphthimide, and rhodamine sulfonate B. When dye molecule A was chosen as ruthenium triphenanthroline or ruthenium terpyridine, the resulting Photo-CIDNP dye pairs all exhibited excellent NMR signal enhancement performance, indicating that dye molecule A can be selected from ruthenium metal complexes with similar structural properties. Similarly, when dye molecule B was chosen as pyrene tetrasulfonate, naphthimide, and rhodamine sulfonate B, the resulting Photo-CIDNP dye pairs also exhibited excellent NMR signal enhancement performance. Dye molecule B can also be a derivative of naphthimide, pyrene, or rhodamine. We also investigated the effect of changes in the concentration ratio between the two dye molecules on the amplitude of NMR signal enhancement. Although we have not exhaustively listed all Photo-CIDNP dye pair combinations, the examples we present include different Photo-CIDNP dye pair combinations. All Photo-CIDNP dye pair combinations exhibit excellent NMR signal enhancement effects. These results fully demonstrate that the principle of the Photo-CIDNP dye pair method proposed in this invention is correct and feasible, and also show that the method of this invention has good universality. Furthermore, in our examples, neither ruthenium triphenanthroline nor pyrene tetrasulfonate alone exhibits Photo-CIDNP hyperpolarization effects; however, the Photo-CIDNP dye pair formed by their combination shows excellent Photo-CIDNP hyperpolarization effects, indicating that the method of this invention can expand the development scope of Photo-CIDNP dye molecules. Using the method of this invention, not only can Photo-CIDNP dye pairs be developed to significantly enhance NMR signals, but the development scope of Photo-CIDNP dye molecules can also be expanded, which is of great significance for accelerating the in-depth and widespread application of NMR hyperpolarization technology.

[0096] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not intended to limit the implementation of the present invention. For those skilled in the art, other variations or modifications can be made based on the above description. It is impossible to exhaustively list all the implementation methods here. All obvious variations or modifications derived from the technical solutions of the present invention are still within the protection scope of the present invention.

Claims

1. A photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair, characterized in that, Composed of dye molecules A and B, wherein dye molecule A is a ruthenium metal complex selected from one or more of the following structures: The dye molecule B is one or more of a naphthalimide derivative, a pyrene derivative, or a rhodamine derivative.

2. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 1, characterized in that, The dye molecule A is ruthenium triphenanthroline or ruthenium triphenanthroline, with the following structure:

3. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 1 or 2, characterized in that, The dye molecule B is naphthalimide, with the following structure: The group R1 is one of phosphate, sulfonate or carboxylate.

4. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 3, characterized in that, The group R1 is a sulfonate group.

5. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 1 or 2, characterized in that, The dye molecule B is a pyrene derivative with the following structure: Groups R2, R3, R4, or R5 are one of hydrogen atoms, carboxylate groups, sulfonate groups, or phosphate groups.

6. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 5, characterized in that, The groups R2, R3, R4 and R5 are all sulfonates.

7. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 1 or 2, characterized in that, The dye molecule B is a rhodamine derivative with the following structure: The group R6 or R7 is one of hydrogen atom, methyl, ethyl, propyl, isopropyl, butyl or isobutyl.

8. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 7, characterized in that, Both R6 and R7 are ethyl groups.

9. The photochemically induced dynamic nuclear polarization Photo-CIDNP dye molecule pair according to claim 1, characterized in that, The concentration ratio between dye molecule A and dye molecule B is in the range of 1:1000 to 1000:

1.

10. The use of any one of the photochemically induced dynamic nuclear polarization Photo-CIDNP dye pairs according to claims 1 to 9 in enhancing the nuclear magnetic resonance signals of tyrosine and its derivatives, indole and its derivatives, or tryptophan and its derivatives.

11. The application according to claim 10, characterized in that, The structures of the indole and its derivatives are as follows: Groups R8, R9, R 10 or R 11 It is either a hydrogen atom or a fluorine atom.

12. The application according to claim 10, characterized in that, The structures of the tryptophan and its derivatives are as follows: Groups R8, R9, R 10 or R 11 It is either a hydrogen atom or a fluorine atom.

13. The application according to any one of claims 10 to 12, characterized in that, The nuclear magnetic resonance signal mentioned is 1 HNMR, 19 F NMR or 13 One or more of C NMR.

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