A method for narrowing the ESR linewidth of photoinduced free radicals for melt dynamic nuclear polarization

Abstract

The present invention belongs to the technical field of electron paramagnetic resonance and dynamic nuclear polarization, and specifically discloses a method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization. The method of the present invention adopts an α-dicarbonyl-containing and a methylene-containing β-dicarbonyl structure as a light-induced free radical precursor as shown in the following formula. The α-dicarbonyl structure can generate light-induced free radicals at liquid nitrogen temperature, and the methylene-containing β-dicarbonyl structure is prone to keto-enol interconversion in the solution. The aforementioned light-induced free radical precursor is dissolved to contain a certain amount of active D + In a solvent, during the keto-enol interconversion process, D gradually replaces the H on the methylene in the β-dicarbonyl to obtain a methylene-deuterated keto structure, which can be illuminated at liquid nitrogen temperature to obtain a deuterated free radical with a narrower ESR line width than the undeuterated structure.

Description

Technical Field

[0001] The invention belongs to the technical field of electron paramagnetic resonance and dynamic nuclear polarization, and in particular relates to a method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization. Background Art

[0002] Melt dynamic nuclear polarization (dDNP) is one of the most successful methods to improve the sensitivity of NMR. This technique has had a positive impact on chemical and metabolic research, especially for low-γ / insensitive nuclei such as 13 Liquid NMR signals of C nuclei provide thousands of times signal enhancement.

[0003] Dynamic nuclear polarization is the process of transferring the high polarization of electrons to the nucleus through microwave irradiation, so electrons are indispensable in the polarization process. Electrons are usually provided by free radicals. In molten DNP, free radicals can be stable small molecule free radicals and photoinduced free radicals. Commonly used stable small molecule free radicals are trityl, BDPA and nitroxide free radical TEMPO.

[0004] Photoinduced free radicals are generated in situ from free radical precursors at low temperature (usually liquid nitrogen temperature) by ultraviolet or visible light irradiation. After polarizing the nucleus, they can be quenched by heating (>190K) to directly obtain a hyperpolarized sample without free radicals. The polarization can be maintained for a long time, so it is a very promising polarization reagent. Photoinduced free radical precursors generally contain α-dicarbonyl structures. The free radical precursors reported in the literature are generally α-keto acid molecules. The typical photoinduced free radical precursor is pyruvic acid. The process of pyruvic acid photoinduced free radical generation is shown in Formula 1, where Free radicals are stable at liquid nitrogen temperature and are used as melt DNP polarizers.

[0005]

[0006] Narrow ESR line free radicals (such as trityl or BDPA) can give low gamma nuclei such as 13 The C core provides good direct DNP enhancement. Although the ESR lines of photoinduced free radicals such as pyruvate free radicals are narrower than those of nitroxide free radicals such as TEMPO and TEMPOL, they are similar to 13 The Larmor frequency of C is still too wide compared to that of free radical electrons, so it is necessary to reduce the coupling of the nuclei near the free radical electrons, especially the hydrogen nuclei, to narrow its spectral line, thereby improving the DNP enhancement effect on low-γ nuclei.

[0007] A direct and effective way to reduce coupling is to deuterate the hydrogen nuclei near the free radicals. The hyperfine coupling constant of deuterated nuclei to unpaired electrons is 1 / 6.5 of that of hydrogen nuclei, so the ESR line width of the free radicals can be narrowed. In 2015, Capozzi et al. used deuterated pyruvic acid as a free radical precursor. The ESR spectrum of its photoinduced free radicals is different from the quartet of pyruvic acid photoinduced free radicals. It is a single peak with a narrow line width, which is obtained as a polarizing agent. 13 C and 129 The polarizability of Xe is twice that of the pyruvate radical. In 2019, Capozzi et al. used trimethylpyruvate and d9-trimethylpyruvate as free radical precursors. Compared with pyruvate, this molecule provides a more isolated electronic environment. The methyl proton is far away from the α carbon, the hyperfine coupling is reduced, and a clear single-line ESR spectrum can be obtained. The deuteration of the methyl group further reduces the hyperfine coupling and ESR line width. The d9-trimethylpyruvate free radical has a strong affinity to the pyruvate free radical. 13 The polarization of C obtained a polarizability close to that of the "gold standard" free radical trityl OX063.

[0008] Although a small amount of deuterated free radical precursors such as deuterated pyruvic acid are commercially available reagents and can be purchased directly, most deuterated free radical precursors usually need to be synthesized using deuterated reactants. For example, d9-trimethylpyruvic acid is synthesized from deuterated acetone in three steps (see Formula 2), and 2-oxo-2-(phenyl-d5)acetic acid is synthesized from d5-acetophenone (see Formula 3). The deuteration steps are cumbersome and uneconomical.

[0009] Summary of the invention

[0010] In order to solve the problems existing in the above-mentioned prior art, the present invention proposes a method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization. The method of the present invention utilizes the characteristic that the β-dicarbonyl structure is easy to undergo enol interconversion, and in situ deuteration of hydrogen at specific positions of the free radical precursor in a protonated deuterated solvent is performed. The method is simple and economical, does not require multi-step synthesis, and does not introduce other impurities. After irradiation, the obtained deuterated free radical precursor can obtain a light-induced free radical with a narrower ESR line width than its undeuterated structure, which can be used as a melt DNP polarizer to improve the DNP enhancement effect on low-γ nuclei.

[0011] The present invention is conceived as follows: using an α-dicarbonyl structure containing a methylene group as shown in Formula 4 as a photoinduced free radical precursor, the α-dicarbonyl structure can generate photoinduced free radicals by light irradiation at liquid nitrogen temperature (see Formula 5). The methylene-containing β-dicarbonyl structure is prone to keto-enol interconversion in solution (Formula 6), dissolving the free radical precursor shown in Formula 4 to a certain amount of active D +In a solvent, during the keto-enol interconversion process, D gradually replaces the H on the methylene in the β-dicarbonyl to obtain a methylene-deuterated keto structure. This structure can be illuminated at liquid nitrogen temperature to obtain a deuterated free radical with a narrower ESR line width than the undeuterated structure, as shown in Formula 7.

[0012] Therefore, the free radical precursor shown in Formula 4 is dissolved to contain a certain amount of active D + In the solvent, during the keto-enol interconversion process, it reacts with D + The exchange achieves deuteration of the methylene group.

[0013]

[0014]

[0015] In order to achieve the above object, the technical solution adopted by the present invention is:

[0016] A method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization comprises the following steps:

[0017] Dissolve the photoinduced free radical precursor to a certain amount of active D + The free radical precursor solution with deuterated methylene groups is obtained in a solvent; the free radical precursor solution is frozen into microbeads in liquid nitrogen, and light-induced free radicals are obtained by irradiating the microbeads. The ESR line width of the obtained photoinduced free radicals is narrowed compared to the undeuterated structure photoinduced free radicals obtained in ordinary solvents.

[0018] Specifically, the steps of obtaining light-induced free radicals by illumination are as follows: using a pipette to take a suitable volume of free radical precursor solution and drop it into liquid nitrogen to freeze it into microbeads; taking a number of microbeads and placing them into a (10 mm) finger-shaped quartz Dewar flask filled with liquid nitrogen, and then using a light source in a suitable wavelength range to irradiate the microbeads immersed in liquid nitrogen for a certain period of time to obtain light-induced free radicals that can stably exist at liquid nitrogen temperature.

[0019] Furthermore, the structural formula of the photoinduced free radical precursor is as follows:

[0020]

[0021] Wherein, R1 and R2 are each independently selected from electron-donating groups. The reasons for selection are as follows: the enol ratio is affected by the acidity of methylene hydrogen during keto-enol interconversion. The stronger the acidity, the greater the enol ratio. When R1 and R2 are electron-withdrawing groups, the acidity of methylene hydrogen is enhanced, which is more conducive to the generation of enol, and the low proportion of keto is not conducive to the generation of free radicals. Therefore, R1 and R2 select electron-donating groups to increase the proportion of keto structure, such as any one of hydroxyl, alkoxy, alkyl, dialkylamino, alkylamino, amino, amide, and acyloxy; preferably, R1 and R2 are each independently selected from any one of hydroxyl, methoxy, ethoxy, propoxy, butoxy, methyl, ethyl, propyl, butyl, dimethylamino, diethylamino, dipropylamino, dibutylamino, methylamino, ethylamino, propylamino, butylamino, formamido, acetamido, propionamido, butyramido, formyloxy, acetoxy, propionyloxy, and butyryloxy.

[0022] Preferably, R1 and R2 are each independently selected from any one of hydroxyl, methoxy, methyl and ethoxy.

[0023] Furthermore, the concentration of the free radical precursor solution is 0.1 mol / L to 2.0 mol / L, the solvent contains a protonated deuterated reagent, and the protonated deuterated reagent is selected from at least one of heavy water, deuterated ethanol, deuterated methanol, deuterated glycerol, deuterated acetic acid, etc., and the active D + The content is greater than that of active H + The content of the deuterated solvent can be a mixture of a common solvent and a protonated deuterated reagent, or a mixture of a non-protonated deuterated reagent and a protonated deuterated reagent, or a mixture of one protonated deuterated reagent or multiple protonated deuterated reagents.

[0024] Preferably, the solvent is deuterated ethanol, a mixture of deuterated ethanol and heavy water in a volume ratio of ≥1, or a mixture of heavy water and an aprotic solvent in a volume ratio of 1:(1-8).

[0025] More preferably, the photoinduced free radical precursor is 4-methoxy-2,4-oxobutyric acid, ethyl acetylacetonate or dimethyl 2-oxosuccinate; and / or

[0026] The solvent is deuterated ethanol, a mixture of deuterated ethanol and heavy water in a volume ratio of 1:1, or a mixture of heavy water and acetone or deuterated acetone in a volume ratio of 1:(1-8).

[0027] Furthermore, the volume of the free radical precursor solution was selected to be 4-5 μL, and the diameter of the microbeads was 2-3 mm.

[0028] Further, 4-6 micro beads were selected and placed in a 10 mm finger-shaped quartz Dewar flask.

[0029] Furthermore, a light source whose irradiation range includes the ultraviolet-visible absorption spectrum region of the free radical precursor is selected to irradiate the microbeads.

[0030] Furthermore, the free radical concentration is estimated through ESR testing, thereby optimizing the light source irradiation time to obtain the required free radical concentration.

[0031] The method applied to melt DNP is to dissolve the polarized target molecule and the photoinduced free radical precursor simultaneously in a solution containing a certain amount of active D + The mixed solution is then taken with a pipette and dripped into liquid nitrogen to freeze it into microbeads. Several microbeads are placed in a 10 mm finger-shaped quartz Dewar flask. A light source in a suitable wavelength range is used to irradiate the microbeads for a suitable time to obtain the required concentration of photoinduced free radicals. The microbeads are then transferred to the polarizer of the melt dynamic nuclear polarization system. The photoinduced free radicals are used as polarizers to polarize the low-γ nuclei of the target molecules.

[0032] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0033] 1. The present invention uses α-dicarbonyl and methylene-containing β-dicarbonyl structures as photoinduced free radical precursors. + The deuterated methylene group can be directly deuterated in a solvent at room temperature via keto-enol interconversion to obtain a free radical precursor with a narrow ESR linewidth.

[0034] 2. The operation is simple and convenient. The free radical precursor can be directly dissolved in a commercially available protonated deuterated solvent. There is no need to use deuterated reactants for one or more steps of synthesis, which avoids multiple operations such as separation and purification, and saves manpower and material resources. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 The D spectrum of EtOD blank solvent and the D, 1 H spectrum;

[0036] Figure 2 Hydrogen decoupling of 4-methoxy-2,4-oxobutanoic acid in EtOD and EtOH / C6D6 (450 μL / 50 μL) 13 C spectrum;

[0037] Figure 3 The ESR spectra (left) and the first integral graph (right) of free radicals generated by 4-methoxy-2,4-oxobutanoic acid in EtOH and EtOD under light.

[0038] Figure 4 The D spectrum of EtOD / D2O 1:1 blank solvent and the D spectrum of ethyl acetylacetonate in EtOD / D2O 1:1 solvent are shown in Figure 2. 1H spectrum;

[0039] Figure 5 Hydrogen decoupling of ethyl acetylacetonate in EtOH / H2O / C6D6 (230μL / 230μL / 40μL) and EtOD / D2O 1:1 13 C spectrum;

[0040] Figure 6 The ESR spectra (left) and the first integral graph (right) of free radicals generated by ethyl acetylacetonate in EtOH / H2O and EtOD / D2O under light illumination;

[0041] Figure 7 The D spectrum of EtOD / D2O 1:1 blank solvent and the D spectrum of dimethyl 2-oxosuccinate in EtOD / D2O 1:1 solvent are shown in Figure 2. 1 H spectrum;

[0042] Figure 8 Hydrogen decoupling of dimethyl 2-oxosuccinate in EtOD / D2O 1:1 and EtOH / H2O / C6D6 (230 μL / 230 μL / 40 μL) 13 C spectrum;

[0043] Fig. 9 The D spectrum and 1 H spectrum, in deuterated acetone 1 H spectrum;

[0044] Fig.10 Hydrogen decoupling of dimethyl 2-oxosuccinate in deuterated acetone and deuterated acetone / D2O (400 μL / 50 μL) 13 C spectrum;

[0045] Fig.11 The ESR spectra (left) and the first integral graph (right) of the free radicals generated by dimethyl 2-oxosuccinate in EtOH / H2O and EtOD / D2O under light.

[0046] Fig.12 The D spectrum of EtOD / D2O 1:1 blank solvent and the D spectrum of pyruvic acid in EtOD / D2O 1:1 solvent are shown in Figure 2. 1 H spectrum;

[0047] Fig.13 Decoupling of hydrogen in pyruvate in EtOD / D2O 1:1 solvent 13 C spectrum;

[0048] Fig.14The ESR spectra (left) and the first integral graph (right) of the free radicals generated by pyruvic acid in EtOH / H2O and EtOD / D2O under light.

[0049] Fig.15 The D spectrum of EtOD / D2O 1:1 blank solvent and the D spectrum of α-ketoglutaric acid in EtOD / D2O 1:1 solvent are shown in Figure 2. 1 H spectrum;

[0050] Fig.16 Decoupling of α-ketoglutarate in EtOD / D2O 1:1 solvent 13 C spectrum;

[0051] Fig.17 The ESR spectra (left) and the first integral graph (right) of the free radicals generated by α-ketoglutarate in EtOH / H2O and EtOD / D2O under light.

[0052] Fig.18 The D spectrum of EtOD / D2O 1:1 blank solvent and the D spectrum of diethyl 2-chloro-3-oxosuccinate in EtOD / D2O 1:1 solvent are shown in Table 1. 1 H spectrum;

[0053] Fig.19 Hydrogen decoupling of diethyl 2-chloro-3-oxobutanedioate in EtOH / H2O 1:1 and EtOD / D2O 1:1 13 C spectrum;

[0054] Fig. 20 The ESR spectra (left) and primary integral diagram (right) of free radicals generated by diethyl 2-chloro-3-oxosuccinate in EtOH / H2O and EtOD / D2O under light. DETAILED DESCRIPTION

[0055] The present invention is described in detail below with reference to specific embodiments and drawings, but this should not be construed as limiting the scope of protection claimed in the claims of the present invention.

[0056] Embodiment 1:

[0057] Deuterated 4-methoxy-2,4-oxobutyric acid methylene and method for generating deuterated free radicals

[0058] Weigh 36.5 mg of 4-methoxy-2,4-oxobutyric acid solid and dissolve it in 500 μL EtOD to prepare a 0.5 M deuterated solvent 4-methoxy-2,4-oxobutyric acid solution; weigh 36.5 mg of 4-methoxy-2,4-oxobutyric acid solid and dissolve it in a 450 μL:50 μL EtOH / C6D6 mixed solution to prepare a 0.5 M common solvent 4-methoxy-2,4-oxobutyric acid solution (add aprotic deuterated solvent C6D6 to uniform the field to obtain a better quality NMR spectrum. Aprotic deuterated solvents will not dissociate active D + , the solvent is equivalent to a common solvent in terms of the effect on the deuteration of methylene groups, the same below). 500 μL EtOD was taken as a blank solvent control. NMR confirmed that the H on the methylene group of the β-dicarbonyl of 4-methoxy-2,4-oxobutyric acid in the protic deuterated solvent EtOD was deuterated.

[0059] After the solution was prepared, the above two 4-methoxy-2,4-oxobutyric acid samples were tested immediately. 1 H spectrum, 13 C spectrum, HSQC spectrum and HMBC spectrum, through analysis, it was determined that the hydrogen chemical shift of -CH2- in the keto configuration was 3.83ppm, and it was buried in the -CH3 hydrogen signal of the enol structure, and the carbon chemical shift was around 44ppm; the hydrogen chemical shift of -CH= in the enol configuration was 5.96ppm, and the carbon chemical shift was around 95.5ppm. These chemical shift values ​​are in line with the theoretical / empirical value range of typical methylene and alkenyl groups. 1 The H and D spectra were compared with the D spectra of the blank solvent (see Figure 1 ), from the D spectrum, we can see that there are deuterium signal peaks at the corresponding positions of the hydrogen chemical shift in the keto form -CH2- and the enol -CH=, and there is no corresponding deuterium signal peak in the blank solvent, indicating that the deuterated signal comes from 4-methoxy-2,4-oxobutyric acid, and part of the hydrogen at these two positions is deuterated.

[0060] Comparison of hydrogen decoupling in 4-methoxy-2,4-oxobutyric acid samples with normal solvent and deuterated solvent 13 C spectrum (see Figure 2 ), it can be seen that the enol structure of 4-methoxy-2,4-oxobutyric acid -CH= in EtOH / C6D6 is decoupled to hydrogen 13 The C signal peak is a single peak, and a triplet in EtOD; the hydrogen decoupling of the keto structure of 4-methoxy-2,4-oxobutyric acid -CH2- in EtOH / C6D6 13 The C signal peak is a single peak, and it is a quintet in EtOD. This is because in common solvents or those without active D +In the solvent, 1 H pair 13 The coupling of C is removed and displayed as a single peak. In protic deuterated solvents, the hydrogen on the enol form -CH= and the keto form -CH2- is deuterated to generate the enol form -CD= and the keto form -CD2-. 1 H pair 13 The coupling of C is removed, and D 13 The coupling of C can be observed, and its split peaks are triple and quintuple, which are consistent with one D and two D for 13 C splitting. It should be noted that the enol triplet 13 The singlet on the left of the C signal is the undeuterated -CH= 13 C signal, which is equal to -CD = 13 Compared with the C signal peak, it can be seen that most of the hydrogens on -CH= are deuterated; the small peak on the left side of the keto quintet is a triplet (the rightmost signal is buried in the quintet), which is the case where only one hydrogen on -CH2- is deuterated (-CHD-). 13 C signal, undeuterated -CH2- 13 C signal was not observed. The signal intensity of the triplet (-CHD-) and the quintet (-CD2-) showed that most of the hydrogens on -CH2- were deuterated. In addition, 4-methoxy-2,4-oxobutyric acid was 13 The slight difference in C chemical shift is caused by solvent change.

[0061] From the NMR spectrum analysis, it can be clearly seen that the keto structure of 4-methoxy-2,4-oxobutyric acid -CH2- is deuterated in the protic deuterated solvent EtOD.

[0062] Weigh 36.5 mg of 4-methoxy-2,4-oxobutyric acid solid and dissolve it in 250 μL EtOH to prepare a 1 M 4-methoxy-2,4-oxobutyric acid solution in a common solvent; weigh 36.5 mg of 4-methoxy-2,4-oxobutyric acid solid and dissolve it in 250 μL EtOD to prepare a 1 M deuterated solvent 4-methoxy-2,4-oxobutyric acid solution. Use a pipette to take 5 μL of 1 M common solvent 4-methoxy-2,4-oxobutyric acid solution and drop it into a 100 mL flat-bottomed Dewar flask filled with liquid nitrogen to freeze it into microbeads with a diameter of about 2 to 3 mm. Take 4 microbeads and move them into a finger-shaped quartz Dewar flask and use a wavelength range of 280 nm-450 nm and an illumination power of 40 W / cm 2 The microbeads were irradiated with a broadband light source for 200 seconds. The irradiated microbeads were transferred to a 5mm ESR detection quartz tube at liquid nitrogen temperature and X-band ESR test was performed at -150°C. The same operation was performed on a 1M deuterated solvent 4-methoxy-2,4-oxobutyric acid solution. The test results are shown in Figure 3 As shown, the left figure is the ESR spectrum, and the right figure is the ESR first integral diagram.

[0063] Depend on Figure 3 It can be seen that in EtOH solvent, the ESR spectrum line shows splitting into two, which is attributed to the coupling of hydrogen to free electrons in the keto-CH2- of 4-methoxy-2,4-oxobutyric acid. In EtOD, most of the hydrogen in -CH2- is deuterated, and the coupling constant of deuterium is only 1 / 6.5 of that of hydrogen. At the same time, due to the widening of the anisotropic g value at low temperature, the ESR spectrum peak splitting caused by the weak hyperfine coupling of deuterium to free radical electrons becomes indistinguishable. The ESR line shape of the photoinduced free radical generated by 4-methoxy-2,4-oxobutyric acid in the deuterated solvent shows a single peak. The half-peak width (note: the half-peak width here is the width of the signal at 1 / 2 the height of the highest point of the signal, the same below) of the first integral graph of the ESR spectrum was measured. When EtOH was used as the solvent, the half-peak width was 0.92mT, and when EtOD was used as the solvent, the half-peak width was 0.45mT. It can be seen that the width of the free baseline induced by light in the deuterated solvent is significantly narrowed, which is more conducive to the polarization of the low-γ nucleus.

[0064] Embodiment 2:

[0065] Deuterated methylene group of ethyl acetylacetonate and method for generating deuterated free radicals

[0066] Take 35.11 μL of ethyl acetylacetonate, dissolve it in a mixed solution of EtOH / H2O / C6D6 with a volume of 230 μL:230 μL:40 μL (add aprotic deuterated solvent C6D6 for uniform field to obtain better quality NMR spectrum), prepare a common solvent ethyl acetylacetonate solution with a concentration of about 0.5M; take 35.11 μL of ethyl acetylacetonate, dissolve it in a mixed solution of EtOD / D2O with a volume of 250 μL:250 μL, prepare a deuterated solvent ethyl acetylacetonate solution with a concentration of about 0.5M; take 250 μL:250 μL of EtOD / D2O, prepare a blank solvent sample for control. NMR confirmed that the H on the methylene group of the β-dicarbonyl of ethyl acetylacetonate in the protic deuterated solvent EtOD / D2O was deuterated.

[0067] After the solution was prepared, the two samples of ethyl acetylacetonate were tested immediately. 1 H spectrum, 13 C spectrum, HSQC spectrum and HMBC spectrum, through analysis, it was determined that the hydrogen chemical shift of -CH2- in the keto configuration was 4.092ppm, and the carbon chemical shift was around 52.5ppm; the hydrogen chemical shift of -CH= in the enol configuration was 6.421ppm, and the carbon chemical shift was around 102.5ppm. These chemical shift values ​​are in line with the theoretical / empirical value range of typical methylene and alkenyl groups.1 The H and D spectra were compared with the D spectra of the blank solvent (see Figure 4 ), from the D spectrum, we can see that there are deuterium signal peaks at the corresponding positions of the hydrogen chemical shift in the keto form -CH2- and the enol -CH=, and there is no corresponding deuterium signal peak in the blank solvent, indicating that the deuterated signal comes from ethyl acetylacetonate, and part of the hydrogen at these two positions is deuterated.

[0068] Comparison of hydrogen decoupling in samples of normal solvent ethyl acetylacetonate and deuterated solvent ethyl acetylacetonate 13 C spectrum (see Figure 5 ), it can be seen that the enol structure of ethyl acetylacetonate -CH= in EtOH / H2O / C6D6 is decoupled to hydrogen 13 The C signal peak is a single peak, and a triplet in EtOD / D2O; the hydrogen decoupling of the keto structure of ethyl acetylacetonate -CH2- in EtOH / H2O / C6D6 13 The C signal peak is a single peak, and it is a quintet in EtOD / D2O; this is because in non-deuterated solvents, 1 H pair 13 The coupling of C is removed and displayed as a single peak. In protic deuterated solvents, the hydrogen on the enol form -CH= and the keto form -CH2- is deuterated to generate the enol form -CD= and the keto form -CD2-. 1 H pair 13 The coupling of C is removed, and D 13 The coupling of C can be observed, and its split peaks are triple and quintuple, which are consistent with one D and two D for 13 C splitting. It should be noted that the enol triplet 13 The singlet on the left of the C signal is the undeuterated -CH= 13 C signal, which is equal to -CD = 13 Compared with the C signal peak, it can be seen that most of the hydrogen on -CH= is deuterated; from the quintet (-CD2-) 13 The intensity of C signal shows that the hydrogen on -CH2- is basically deuterated; in addition, ethyl acetylacetonate is 13 The slight difference in C chemical shift is caused by solvent change.

[0069] From the NMR spectrum analysis, it can be clearly seen that the keto structure of ethyl acetylacetonate -CH2- has undergone deuteration in the protonous deuterated mixed solvent EtOD / D2O.

[0070] Take 35.11μL of ethyl acetylacetonate and dissolve it in a 125μL:125μL EtOH / H2O mixed solution to prepare a common solvent ethyl acetylacetonate solution with a concentration of about 1M; take 35.11μL of ethyl acetylacetonate and dissolve it in a 125μL:125μL EtOD / D2O mixed solution to prepare a deuterated solvent ethyl acetylacetonate solution with a concentration of about 1M. Use a pipette to take 5μL of 1M common solvent ethyl acetylacetonate solution and drop it into a 100mL flat-bottomed Dewar flask filled with liquid nitrogen to freeze it into microbeads with a diameter of about 2 to 3mm. Take 4 microbeads and move them into a finger-shaped quartz Dewar flask and use a wavelength range of 280nm-450nm and an illumination power of 40W / cm 2 The microbeads were irradiated with a broadband light source for 400 seconds. The irradiated microbeads were transferred to a 5mm ESR detection quartz tube at liquid nitrogen temperature and X-band ESR test was performed at -150°C. The same operation was performed on a 1M deuterated solvent ethyl acetylacetonate solution. The test results are shown in Figure 6 As shown, the left figure is the ESR spectrum, and the right figure is the ESR first integral diagram.

[0071] Depend on Figure 6 It can be seen that in EtOH / H2O solvent, the main peak of the ESR spectrum line shows splitting into two, which is attributed to the coupling of hydrogen to free electrons in the keto form -CH2- of ethyl acetylacetonate. In EtOD / D2O, most of the hydrogen in -CH2- is deuterated, and the coupling constant of deuterium is only 1 / 6.5 of that of hydrogen. At the same time, due to the widening of the anisotropic g value at low temperature, the ESR spectrum peak splitting caused by the weak hyperfine coupling of deuterium to free radical electrons becomes indistinguishable. The main peak of the ESR spectrum line of the photoinduced free radical generated by ethyl acetylacetonate in the deuterated solvent is a single peak. The half-peak width of the first integral graph of the ESR spectrum was measured. When EtOH / H2O was used as the solvent, the half-peak width was 2.14mT, and when EtOD / D2O was used as the solvent, the half-peak width was 1.66mT. It can be seen that the free baseline width induced by light in the deuterated solvent is significantly narrowed, which is more conducive to the polarization of low-γ nuclei.

[0072] Example 3

[0073] Deuterated methylene group of dimethyl 2-oxosuccinate and method for generating deuterated free radicals

[0074] Weigh 40 mg of solid dimethyl 2-oxosuccinate, dissolve it in a mixed solution of EtOH / H2O / C6D6 with a volume of 230 μL:230 μL:40 μL (add aprotic deuterated solvent C6D6 for uniform field to obtain a better quality NMR spectrum), and prepare a common solvent dimethyl 2-oxosuccinate solution with a concentration of about 0.5 M; weigh 40 mg of solid dimethyl 2-oxosuccinate, dissolve it in a mixed solution of EtOD / D2O with a volume of 250 μL:250 μL, and prepare a deuterated solvent 2-oxosuccinate solution with a concentration of about 0.5 M; take 250 μL:250 μL of EtOD / D2O to prepare a blank solvent sample for control. The deuterated hydrogen on the methylene group of the β-dicarbonyl of dimethyl 2-oxosuccinate in the protic deuterated solvent EtOD / D2O was confirmed by NMR.

[0075] The two samples of dimethyl 2-oxosuccinate were tested immediately after the solution was prepared. 1 H spectrum, 13 The C spectrum, HSQC spectrum and HMBC spectrum determined that the hydrogen chemical shift of -CH2- in the keto configuration was 3.959 ppm, and the carbon chemical shift was around 44.5 ppm. The hydrogen chemical shift of -CH= in the enol configuration was 6.013 ppm, and the carbon chemical shift was around 97 ppm. These chemical shift values ​​are in line with the theoretical / empirical value range of typical methylene and alkenyl groups. 1 The H and D spectra were compared with the D spectra of the blank solvent (see Figure 7 ). From the D spectrum, it can be seen that there is a deuterium signal peak at the position corresponding to the chemical shift of hydrogen in the enol -CH=, and there is no corresponding deuterium signal peak in the blank solvent, indicating that this deuterated signal comes from dimethyl 2-oxosuccinate; the hydrogen spectrum corresponding to the keto configuration shows a triplet peak, corresponding to the hydrogen signal of the structure in which one hydrogen in -CH2- is deuterated (-CHD-), and the deuterium signal can be observed at the corresponding position of the deuterium spectrum, and the peak is on the left shoulder of the solvent peak on its right side. The corresponding signal is not observed in the blank solvent deuterium spectrum, indicating that this deuterium signal comes from dimethyl 2-oxosuccinate; the NMR spectrum shows that part of the hydrogen in the enol form -CH= and keto form -CH2- of dimethyl 2-oxosuccinate is deuterated.

[0076] Comparison of hydrogen decoupling in the samples of common solvent dimethyl 2-oxosuccinate and deuterated solvent dimethyl 2-oxosuccinate 13 C spectrum (see Figure 8 ), it can be seen that the enol structure of dimethyl 2-oxosuccinate -CH= decouples hydrogen in EtOH / H2O / C6D6 13 The C signal peak is a single peak, and a triplet in EtOD / D2O; the keto structure of dimethyl 2-oxosuccinate -CH2- is decoupled from hydrogen in EtOH / H2O / C6D613 The C signal peak is a single peak. A triplet signal is observed in EtOD / D2O, which should actually be a quintet. Due to the relatively low concentration of the keto form, the split peaks on both sides could not be observed. The reasons for the change in the peak shape of dimethyl 2-oxosuccinate are as follows: + In the solvent, 1 H pair 13 The coupling of C is removed and displayed as a single peak. In the proton deuterated reagent, the hydrogen on the enol form -CH= and the keto form -CH2- is deuterated to generate the enol form -CD= and the keto form -CD2-. 1 H pair 13 The coupling of C is removed, and D 13 The coupling of C can be observed, and its split peaks are triple and quintuple, which are consistent with one D and two D for 13 C splitting. It should be noted that the enol triplet 13 The singlet on the left of the C signal is the undeuterated -CH= 13 C signal, which is equal to -CD = 13 Compared with the C signal peak, it can be seen that most of the hydrogen on -CH= is deuterated; from the quintet (-CD2-) 13 The intensity of C signal shows that the hydrogen on -CH2- is basically deuterated; dimethyl 2-oxosuccinate is in deuterated solvent and ordinary solvent 13 The slight difference in C chemical shift is caused by solvent change.

[0077] Since the ratio of enol form to keto form of dimethyl 2-oxosuccinate in the solvent of EtOD / D2O volume ratio of 1:1 is relatively low, its situation in deuterated acetone and deuterated acetone / D2O was also analyzed to further confirm its changes in protic deuterated solvents.

[0078] Weigh 16 mg of dimethyl 2-oxosuccinate solid, dissolve it in 400 μL of deuterated acetone to prepare a solution with a concentration of about 0.25 M, and test its NMR spectrum; then add 50 μL of D2O, at which point the concentration is about 0.22 M, test its NMR spectrum, and compare the changes in the spectrum before and after adding D2O.

[0079] By analyzing the concentration of dimethyl 2-oxosuccinate in deuterated acetone and deuterated acetone / D2O 1 H. 13 C, HSQC spectrum and HMBC spectrum determined that the hydrogen chemical shift of the keto configuration -CH2- is 3.911ppm, and the carbon chemical shift is around 44.5ppm; the hydrogen chemical shift of the enol configuration -CH= is 5.956ppm, and the carbon chemical shift is around 96ppm. There is a slight difference between the chemical shift values ​​in the ethanol / water solvent system. 1H spectra and their applications in deuterated acetone / D2O 1 Comparison of H and D spectra (see Fig. 9 ), it can be seen that in deuterated acetone 1 The hydrogen signal peaks of the enol and keto forms can be observed in the H spectrum. After adding D2O, 1 The H spectrum can observe that the peak intensity of the hydrogen signal of the enol -CH= is reduced, but the hydrogen signal of the keto form -CH2- is basically unobservable; the D spectrum observes the deuterium signal corresponding to the enol -CH=, and the deuterium signal corresponding to the keto form -CH2- is not observed separately, and its deuterium signal should be buried in the D signal of D2O at 3.87ppm.

[0080] Further analysis of the carbon spectra before and after the addition of D2O (see Fig.10 ), it can be seen that in deuterated acetone, the enol form -CH= and the keto form -CH2- 13 The C signals are all single peaks, indicating that they will not be deuterated in aprotic deuterated solvents. After adding D2O, the enol -CH= 13 The C signal becomes a single peak plus a triplet, and the single peak corresponds to the undeuterated structure. 13 C signal, triplet peak is -CH = hydrogen is replaced by deuterium 13 C signal; keto-CH2- 13 The C signal becomes a triplet plus a quintet. The triplet is the corresponding peak of -CH2- where one hydrogen is deuterated. 13 C signal, the quintet is -CH2-, where two hydrogens are replaced by deuterium 13 C signal. This indicates that after adding a protic deuterated solvent, the hydrogen on the enol form -CH= and the keto form -CH2- is deuterated to generate the enol form -CD= and the keto form -CD2- and -CDH-. From the NMR spectrum analysis, it can be clearly seen that the keto form structure -CH2- of dimethyl 2-oxosuccinate is deuterated in EtOD / D2O solvent and deuterated acetone / D2O solvent.

[0081] Weigh 40 mg of 2-oxosuccinic acid dimethyl ester solid, dissolve it in a 125 μL:125 μL EtOH / H2O mixed solution, and prepare a 2-oxosuccinic acid dimethyl ester solution with a concentration of about 1 M in a common solvent; weigh 40 mg of 2-oxosuccinic acid dimethyl ester solid, dissolve it in a 125 μL:125 μL EtOD / D2O mixed solution, and prepare a 2-oxosuccinic acid dimethyl ester solution with a concentration of about 1 M in a deuterated solvent. Use a pipette to take 5 μL of 1M common solvent 2-oxosuccinic acid dimethyl ester solution and drop it into a 100 mL flat-bottomed Dewar flask filled with liquid nitrogen to freeze it into microbeads with a diameter of about 2 to 3 mm. Take 4 microbeads and move them into a finger-shaped quartz Dewar flask, and use a wavelength range of 280 nm-450 nm and a light power of 40 W / cm 2The microbeads were irradiated with a broadband light source for 200 seconds. The irradiated microbeads were transferred to a 5mm ESR detection quartz tube at liquid nitrogen temperature and X-band ESR test was performed at -150°C. The same operation was performed on a 1M deuterated solvent 2-oxosuccinic acid dimethyl ester solution. The test results are shown in Fig.11 As shown, the left figure is the ESR spectrum, and the right figure is the ESR first integral diagram.

[0082] Depend on Fig.11 It can be seen that in EtOH / H2O solvent, the ESR spectrum line shows splitting into two, which is attributed to the coupling of hydrogen in the keto form -CH2- of 2-oxosuccinic acid dimethyl ester to free electrons. In EtOD / D2O, most of the hydrogen in -CH2- is deuterated, and the coupling constant of deuterium is only 1 / 6.5 of that of hydrogen. At the same time, due to the widening of the anisotropic g value at low temperature, the ESR spectrum peak splitting caused by the weak hyperfine coupling of deuterium to free radical electrons becomes indistinguishable. The ESR line shape of the photoinduced free radical generated by 2-oxosuccinic acid dimethyl ester in the deuterated solvent shows a single peak. The half-peak width of the first integral diagram of the ESR spectrum was measured. When EtOH / H2O was used as the solvent, the half-peak width was 0.77mT, and when EtOD / D2O was used as the solvent, the half-peak width was 0.37mT. It can be seen that although the ratio of enol and keto forms of dimethyl 2-oxosuccinate in the ethanol / water volume ratio of 1:1 is relatively low, the free baseline width induced by light in the deuterated solvent is still narrowed, which is more conducive to the polarization of low γ nuclei than the free radicals generated in ordinary solvents.

[0083] Comparative Example 1:

[0084] Possible methods for deuterated pyruvate and generation of deuterated free radicals

[0085] Take 16.8 μL of pyruvic acid and dissolve it in a mixed solvent of 250 μL:250 μL of EtOD / D2O to prepare a pyruvic acid solution with a concentration of about 0.5 M. Take 250 μL:250 μL of EtOD / D2O to prepare a blank solvent sample for control.

[0086] Test 0.5M pyruvic acid solution 1 H and D spectra, and compared with the blank solvent D spectrum (see Fig.12 ). From pyruvic acid 1 The H spectrum shows that a small amount of enol structure is produced, and the corresponding hydrogen chemical shift is 6.337ppm. There is no signal at the corresponding position of the deuterium spectrum, indicating that no deuterated enol structure is generated. The deuterium spectrum of pyruvate is basically similar to the deuterium spectrum of the blank solvent, and no other new peaks appear, indicating that no other deuterated structures are generated.

[0087] Testing of pyruvic acid solution 13 C spectrum (see Fig.13), no connection D is seen 13 C multiple signal peaks, further indicating that it cannot be directly deuterated in protic deuterated solvents.

[0088] Take 16.8μL of pyruvic acid and dissolve it in a mixed solvent of EtOH / H2O with a volume of 125μL:125μL to prepare a common solvent pyruvic acid solution with a concentration of about 1M; take 16.8μL of pyruvic acid and dissolve it in a mixed solvent of EtOD / D2O with a volume of 125μL:125μL to prepare a deuterated solvent pyruvic acid solution with a concentration of about 1M. Use a pipette to take 5μL of 1M common solvent pyruvic acid solution and drop it into a 100mL flat-bottomed Dewar flask filled with liquid nitrogen to freeze it into microbeads with a diameter of about 2 to 3mm. Take 4 microbeads and move them into a finger-shaped quartz Dewar flask and use a wavelength range of 280nm-450nm and an illumination power of 40W / cm 2 The microbeads were irradiated with a broadband light source for 400 seconds. The irradiated microbeads were transferred to a 5mm ESR detection quartz tube at liquid nitrogen temperature and X-band ESR test was performed at -150°C. The same operation was performed on a 1M deuterated solvent pyruvic acid solution. The test results are shown in Fig.14 As shown, the left figure is the ESR spectrum, and the right figure is the ESR first integral diagram.

[0089] Depend on Fig.14 It can be seen that the free radicals generated by pyruvic acid under light in EtOH / H2O and EtOD / D2O are all quartets, and the line width and peak splitting degree have changed slightly, but the situation where the ESR signal becomes a single peak similar to Examples 1 to 3 is not observed, indicating that it cannot be directly deuterated in protic deuterated solvents.

[0090] Comparative Example 2:

[0091] Deuteration of α-ketoglutarate (α-KG) and possible methods of generating deuterated free radicals

[0092] Weigh 36.5 mg of α-ketoglutaric acid and dissolve it in a mixed solvent of 250 μL:250 μL of EtOD / D2O to prepare an α-ketoglutaric acid solution with a concentration of about 0.5 M. Take 250 μL:250 μL of EtOD / D2O to prepare a blank solvent sample for control.

[0093] Test 0.5M α-ketoglutarate solution 1 H and D spectra, and compared with the blank solvent D spectrum (see Fig.15 ). From α-ketoglutarate 1The H spectrum shows that a small amount of enol structure is produced, and the corresponding hydrogen chemical shift is 5.826ppm. There is no signal at the corresponding position of the deuterium spectrum, indicating that no deuterated enol structure is generated. The deuterium spectrum of α-ketoglutaric acid is basically similar to the deuterium spectrum of the blank solvent, and no other new peaks appear, indicating that no other deuterated structures are generated.

[0094] Testing of α-ketoglutarate solution 13 C spectrum (see Fig.16 ), no connection D is seen 13 C multiple signal peaks, further indicating that it cannot be directly deuterated in protic deuterated solvents.

[0095] Take 36.5 mg of α-ketoglutaric acid and dissolve it in a mixed solvent of EtOH / H2O with a volume of 125 μL:125 μL to prepare a common solvent α-ketoglutaric acid solution with a concentration of about 1 M; take 36.5 mg of α-ketoglutaric acid and dissolve it in a mixed solvent of EtOD / D2O with a volume of 125 μL:125 μL to prepare a deuterated solvent α-ketoglutaric acid solution with a concentration of about 1 M. Use a pipette to take 5 μL of 1M common solvent α-ketoglutaric acid solution and drop it into a 100 mL flat-bottomed Dewar flask filled with liquid nitrogen to freeze it into microbeads with a diameter of about 2 to 3 mm. Take 4 microbeads and move them into a finger-shaped quartz Dewar, and use a wavelength range of 280nm-450nm and an illumination power of 40W / cm 2 The microbeads were irradiated with a broadband light source for 400 seconds. The irradiated microbeads were transferred to a 5mm ESR detection quartz tube at liquid nitrogen temperature and X-band ESR test was performed at -150°C. The same operation was performed on a 1M deuterated solvent α-ketoglutaric acid solution. The test results are shown in Fig.17 As shown, the left figure is the ESR spectrum, and the right figure is the ESR first integral diagram.

[0096] Depend on Fig.17 It can be seen that the main peak of the free radical generated by α-ketoglutaric acid under light in EtOH / H2O and EtOD / D2O is split into two peaks, and a small coupling splitting can be observed in each peak of the double peak, which is generally manifested as a quartet. In the first integral diagram, the small coupling is not displayed, and it appears as a double peak; the line width and the degree of splitting have changed slightly, but the situation similar to the ESR signal becoming a single peak in Examples 1 to 3 is not observed, indicating that it cannot be directly deuterated in a protic deuterated solvent.

[0097] Comparative Example 3:

[0098] Deuteration of diethyl 2-chloro-3-oxosuccinate and possible methods for the generation of deuterated free radicals

[0099] Take 45 μL of 2-chloro-3-oxosuccinic acid diethyl ester and dissolve it in a 250 μL:250 μL EtOH / H2O mixed solution to prepare a 0.5M common solvent 2-chloro-3-oxosuccinic acid diethyl ester solution; take 45 μL of 2-chloro-3-oxosuccinic acid diethyl ester and dissolve it in a 250 μL:250 μL EtOD / D2O mixed solution to prepare a 0.5M deuterated solvent 2-chloro-3-oxosuccinic acid diethyl ester solution; take 250 μL:250 μL EtOD / D2O to prepare a blank solvent sample for control. Use NMR to check whether the H on the chloromethylene group in the β-dicarbonyl of 2-chloro-3-oxosuccinic acid diethyl ester in the protic deuterated solvent EtOD / D2O is deuterated.

[0100] After the solution was prepared, the above two samples of diethyl 2-chloro-3-oxosuccinate were tested immediately. 1 H spectrum, 13 C spectrum, HSQC spectrum and HMBC spectrum, through analysis, it was determined that the hydrogen chemical shift of -CClH- in the keto configuration was 4.859ppm, and the carbon chemical shift was around 61.5ppm; the carbon chemical shift of -CCl= in the enol configuration was around 96.5ppm. Since the enol structure -CCl= does not contain hydrogen, the subsequent spectra only analyze the corresponding keto structure. 1 H and 13 C signal changes.

[0101] The deuterated solvent 2-chloro-3-oxosuccinic acid diethyl ester sample 1 The H and D spectra were compared with the D spectra of the blank solvent (see Fig.18 ), the deuterium signal corresponding to -CClH- in the keto configuration could not be observed separately from the D spectrum, which should be buried in the heavy water D signal at 4.856 ppm.

[0102] pass 13 The C spectrum further confirmed that the hydrogen decoupling of diethyl 2-chloro-3-oxosuccinate in two solvents 13 C spectrum (see Fig.19 ). In EtOH / H2O solvent, the -CClH- 13 The C signal is a single peak; in EtOD / D2O solvent, the hydrogen of -CClH- is deuterated to -CClD-. 1 H pair 13 The coupling of C is removed, and D 13 The coupling of C can be observed, and its split peak shape is a triplet, which is consistent with a D pair 13 The splitting of C.

[0103] Take 45 μL of diethyl 2-chloro-3-oxosuccinate, dissolve it in a mixed solvent of EtOH / H2O with a volume of 125 μL:125 μL to prepare a common solvent 2-chloro-3-oxosuccinate solution with a concentration of about 1 M; take 45 μL of diethyl 2-chloro-3-oxosuccinate, dissolve it in a mixed solvent of EtOD / D2O with a volume of 125 μL:125 μL to prepare a deuterated solvent 2-chloro-3-oxosuccinate solution with a concentration of about 1 M. Use a pipette to take 5 μL of 1M common solvent 2-chloro-3-oxosuccinic acid diethyl ester solution and drop it into a 100 mL flat-bottomed Dewar flask filled with liquid nitrogen to freeze it into micro beads with a diameter of about 2 to 3 mm. Take 4 micro beads and move them into a finger-shaped quartz Dewar. Use a wavelength range of 280 nm-450 nm and an illumination power of 40 W / cm 2 The microbeads were irradiated with a broadband light source for 400 seconds. The irradiated microbeads were transferred to a 5mm ESR detection quartz tube at liquid nitrogen temperature and X-band ESR test was performed at -150°C. The same operation was performed on a 1M deuterated solvent 2-chloro-3-oxosuccinic acid diethyl ester solution. The test results are shown in Fig. 20 As shown, the left figure is the ESR spectrum, and the right figure is the ESR first integral diagram.

[0104] Depend on Fig. 20 It can be seen that the free radical peak type generated by 2-chloro-3-oxosuccinic acid diethyl ester in EtOH / H2O and EtOD / D2O is basically unchanged. Although -CClH- in the middle of its β-dicarbonyl group undergoes deuteration in a protic deuterated solvent, the ESR signal is not observed to become a single peak as in similar embodiments. This may be because the coupling effect of unchlorinated hydrogen on free radicals is relatively small, so no obvious difference is observed in its coupling effect on free radicals before and after deuteration in the ESR spectrum.

Claims

1. A method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization, characterized in that: The steps include: Dissolve the photoinduced free radical precursor to a certain amount of active D + A free radical precursor solution in which the methylene groups are deuterated is obtained in a solvent; the free radical precursor solution is frozen into microbeads in liquid nitrogen, and light-induced free radicals are obtained by irradiating the microbeads; the ESR line width of the obtained light-induced free radicals is narrowed; The structural formula of the photoinduced free radical precursor is as follows: Wherein, R1 and R2 are each independently selected from electron donating groups.

2. The method for narrowing the ESR linewidth of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 1, characterized in that: The steps of obtaining light-induced free radicals by illumination are as follows: taking a suitable volume of free radical precursor solution and dropping it into liquid nitrogen to freeze it into microbeads; taking a number of microbeads and placing them into a finger-shaped quartz Dewar flask filled with liquid nitrogen, and then using a light source in a suitable wavelength range to irradiate the microbeads immersed in liquid nitrogen for a certain period of time to obtain light-induced free radicals.

3. The method for narrowing the ESR linewidth of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 1, characterized in that: R1 and R2 are each independently selected from any one of hydroxyl, alkoxy, alkyl, dialkylamino, alkylamino, amino, amide, and acyloxy; and / or The solvent contains a protonated deuterated reagent, which is active + The content is greater than that of active H + The protonated deuterated reagent is selected from at least one of heavy water, deuterated ethanol, deuterated methanol, deuterated glycerol and deuterated acetic acid.

4. The method for narrowing the ESR linewidth of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 1, characterized in that: R1 and R2 are each independently selected from any one of hydroxyl, methoxy, ethoxy, propoxy, butoxy, methyl, ethyl, propyl, butyl, dimethylamino, diethylamino, dipropylamino, dibutylamino, methylamino, ethylamino, propylamino, butylamino, formamido, acetamido, propionylamino, butyramido, formyloxy, acetoxy, propionyloxy, butyryloxy; and / or The solvent is deuterated ethanol, a mixture of deuterated ethanol and heavy water in a volume ratio of ≥1, or a mixture of heavy water and aprotic solvent in a volume ratio of 1:(1-8).

5. The method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 1, characterized in that: The photoinduced free radical precursor is 4-methoxy-2,4-oxobutyric acid, ethyl acetylacetonate or dimethyl 2-oxosuccinate; and / or The solvent is deuterated ethanol, a mixture of deuterated ethanol and heavy water in a volume ratio of 1:1, or a mixture of heavy water and acetone or deuterated acetone in a volume ratio of 1:(1-8).

6. The method for narrowing the ESR linewidth of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 1, characterized in that: The concentration of the free radical precursor solution is 0.1-2.0 mol / L.

7. The method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 2, characterized in that: The light source irradiation range includes the ultraviolet-visible absorption spectrum region of the photoinduced free radical precursor.

8. The method for narrowing the ESR linewidth of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 2, characterized in that: The diameter of the microbeads is 2 to 3 mm.

9. The method for narrowing the ESR line width of light-induced free radicals applied to melt dynamic nuclear polarization according to claim 2, characterized in that: The light source irradiation time is obtained by estimating the concentration of light-induced free radicals through ESR measurement.

10. Use of the light-induced free radicals obtained by the method of any one of claims 1 to 9 as a polarizer for melt dynamic nuclear polarization.

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