CN108794510B - A double-radical compound composed of triarylmethyl radical and nitroxide radical and salt thereof, preparation method and application thereof - Google Patents

Abstract

制备方法包括：化合物A1与化合物A2发生酸胺缩合反应，得到化合物A3；化合物A3与化合物A4发生酸胺缩合反应，得到双自由基化合物及其盐。该双自由基极化剂同时具有氮氧自由基和三芳基甲基自由基部分具有如下优势：频率匹配性更优，CE效率更高；在魔角旋转的条件下不存在去极化效应，高场DNP增强效果更优；具有的较强的交换相互作用；具有良好的水溶性，因此DNP增强倍数高；对R₁、R₂、R₃、R₄、R₅、R₆基团的修饰可适度提高交换相互作用、提高双自由基的弛豫时间和双自由基的水溶性，从而提高极化剂的DNP性质。The present invention provides a biradical compound composed of triarylmethyl radical and nitroxide radical and salt thereof, a preparation method and application thereof, and the structural formula of the biradical compound is represented by the following formula 1:

The preparation method includes: compound A1 and compound A2 undergo acid-amine condensation reaction to obtain compound A3; compound A3 undergoes acid-amine condensation reaction with compound A4 to obtain diradical compounds and salts thereof. The biradical polarizer has both nitroxide radicals and triarylmethyl radical moieties, and has the following advantages: better frequency matching, higher CE efficiency; no depolarization effect under the condition of magic angle rotation, High-field DNP enhancement effect is better; has strong exchange interaction; has good water solubility, so DNP enhancement ratio is high; R ₁ , R ₂ , R ₃ , R ₄ , R ₅ , R ₆ groups Modification can moderately improve the exchange interaction, the relaxation time of diradicals and the water solubility of diradicals, thereby improving the DNP properties of polarizers.

Description

A two-radical compound composed of triarylmethyl radical and nitroxide radical and salt thereof, preparation method and application thereof

technical field

The invention relates to the technical field of nuclear magnetic resonance, in particular to a diradical compound composed of a triarylmethyl radical and a nitroxide radical and a salt thereof, a preparation method and application thereof.

Background technique

Nuclear magnetic resonance technology is an important means for the characterization of chemical structures of substances, and is widely used in chemistry, materials science, biochemistry and other fields, especially solid-state nuclear magnetic resonance technology makes it possible to analyze the structure of biological macromolecules such as protein molecules, but its application is limited. Limited to inherently low sensitivity. Aiming at this defect, there are currently two efforts. On the one hand, it is to improve the magnetic field strength of the instrument, thereby increasing the polarizability of the nucleus under thermal equilibrium, but this method is expensive and requires high technical requirements; The polarization efficiency of the nucleus is enhanced by the DNP (DNP) technology, thereby enhancing the signal strength and improving the sensitivity. The choice of polarizer is the key to the dynamic nuclear polarization experiment, which directly determines the signal enhancement factor of DNP. The DNP enhancement factor of the existing polarizing agents is much lower than the theoretical value ( ¹ H, 658-fold; ¹³ C, 2600-fold).

Since the DNP technique was first proposed 60 years ago, all experiments have relied on single spin centers, such as nitroxide radicals TEMPO and metal atoms as polarization sources. Currently, for DNP technology, the most effective mechanism is CE (crosseffect) mechanism, and CE mechanism is closely related to the dipole interaction and spin interaction between electrons. In 2003, Kan-Nian Hu and Robert G. Griffin first proposed the concept of nitroxide radicals, linking two TEMPOs with PEG chains, which enhanced the dipole interaction and increased the signal intensity by 4 times compared to single nitroxide radicals. . Since then, nitroxide radicals have been attracting attention as DNP polarizers. However, nitroxide still has problems of poor stability, poor water solubility, wide single peak linewidth, short relaxation time and weak exchange interaction, and the nitroxide rotates at the magic angle. There is a nuclear depolarization effect under the condition of , which weakens the efficiency of nitroxide radical DNP, and the enhancement effect of high-field DNP is not good, which limits the development of dynamic nuclear polarization technology and the improvement of solid-state NMR sensitivity.

According to the relevant research on the use of nitroxide as a polarizer for DNP, it is confirmed that factors such as electron relaxation time, exchange interaction, and water solubility can affect the DNP properties of polarizers. For nitroxide radicals, the enhancement factor of DNP can be effectively increased by prolonging the relaxation time. Relevant studies have shown that for nitroxide radicals, the relaxation time has reached the optimal conditions, and there is no room for improvement. Studies have confirmed that the exchange interaction is closely related to the high-field DNP properties of polarizers. The depolarization effect of oxygen biradical is obvious), which is also an important factor for its high-field DNP properties to be superior to nitroxide biradicals. Therefore, moderately enhancing the exchange interaction is also an important consideration in the design of TN biradical polarizers. At low temperature, the polarizer has a self-aggregation phenomenon, which makes the polarization transfer loss and the transfer efficiency decrease. Therefore, improving the water solubility and enhancing the dispersion of the polarizer in the solution is also an effective method to optimize the polarizer.

SUMMARY OF THE INVENTION

The problem to be solved by the present invention is to provide a di-radical compound composed of triarylmethyl radical and nitroxide radical and its salt, its preparation method and application. The enhancement multiple is high, which can effectively enhance the sensitivity of solid-state NMR.

In order to solve the above-mentioned technical problems, an object of the present invention is to provide a kind of two-radical compound and salt thereof composed of triarylmethyl radical and nitroxide radical, and its structural formula is represented by following formula 1:

in,

R ₁ is selected from one of H or C1-C18 saturated alkyl group, C1-C18 saturated alkyl group represents a straight or branched chain alkyl group containing 1-18 carbon atoms, such as methyl, ethyl, propyl, Butyl, isopropyl, etc., preferably, R ₁ is H or CH ₃ ;

中，不同的氨基酸侧链R取代基不同，式1中R₂与R对应，R₂选自天然或非天然氨基酸中的侧链R取代基，优选的，R₂为H、CH₃或天然氨基酸或非天然氨基酸的相应侧链取代基中的一种；R ₂ is selected from one of H, C1-C6 saturated alkyl and corresponding side chain substituents of natural amino acids or unnatural amino acids, C1-C6 saturated alkyl represents a straight or branched chain containing 1-6 carbon atoms Alkyl, such as methyl, ethyl, propyl, butyl, isopropyl, etc., the corresponding side chain substituents of natural amino acids or unnatural amino acids refer to: amino acid general formula

In, different amino acid side chain R substituents are different, in formula 1 R ₂ corresponds to R, R ₂ is selected from the side chain R substituents in natural or unnatural amino acids, preferably, R ₂ is H, CH ₃ or natural one of the corresponding side chain substituents of an amino acid or an unnatural amino acid;

R ₃ is selected from one of H, C1-C18 saturated alkyl or (CH ₂ CH ₂ O) _n H, 1≤n≤20, C1-C18 saturated alkyl represents a straight chain containing 1-18 carbon atoms Or branched alkyl, such as methyl, ethyl, propyl, butyl, isopropyl, etc., preferably, R ₃ is one of H or (CH ₂ CH ₂ O) _n H, 1≤n≤ 20, more preferably, R ₃ is H or (CH ₂ CH ₂ O) ₂ H;

X为O、S、CHOH、NCH₃或NH，m＝0、1或2，优选的，R₄、R₅为CH₃或R₄与R₅相连形成

The same or different R ₄ and R ₅ are selected from alkyl or R ₄ is connected with R ₅ to form

X is O, S, CHOH, NCH ₃ or NH, m=0, 1 or 2, preferably, R ₄ and R ₅ are CH ₃ or R ₄ is connected with R ₅ to form

R ₆ is CH ₃ , CD ₃ , CH ₂ CH ₂ OH or connected with two R ₆ on carbon to form a ring structure, preferably, R ₆ is CH ₃ , CD ₃ or connected with two R ₆ on carbon is a ring structure.

The compound connects nitroxide radicals with trityl radicals through a linking chain, forming a biradical polarizer with both nitroxide radicals and triarylmethyl radicals. Nitroxyl radical-triarylmethyl radical type diradical has the following advantages over dinitroxide radical: First, for high-field DNP, the CE (cross effect) mechanism is the most effective mechanism , when the difference in electron spin frequency matches the Raman frequency of the nucleus, the CE mechanism can achieve maximum efficiency, while the frequency matching of nitroxide-triarylmethyl-type bis-radical is better than that of bis-nitrogen Oxygen free radicals, so its CE efficiency is higher. Secondly, under the condition of magic angle rotation, the bis-nitroxide radical has the depolarization effect of the nucleus, which weakens the DNP efficiency, but the nitroxyl radical-triarylmethyl radical type double radical does not depolarize the nitroxide radical. polarization effect, which makes the high-field DNP enhancement effect better than that of nitroxide radicals. In addition, the experimental results suggest that the nitroxide radical-triarylmethyl radical type diradical has a stronger exchange interaction than nitroxide radicals, which is significantly different from that of nitroxide radicals. It has been reported in the literature that for high-field DNP, the effect of dipole interaction is very small, and the dominant role is exchange interaction. Therefore, the stronger exchange interaction also makes this compound more efficient than nitroxide biradical compounds. High high-field DNP enhancement factor. But the exchange interaction is not as strong as possible. Only when the exchange interaction is just right can the DNP efficiency be maximized. Finally, the triarylmethyl radical has good water solubility and abundant modifiable sites, which can be used for functional derivatization for different purposes.

时可延长氮氧自由基部分的弛豫时间，R₆为CD₃时可提高三芳基甲基自由基部分的弛豫时间；3)R₂、R₃、R₄、R₅、R₆的修饰可提高双自由基的水溶性。R₂为谷氨酸、天冬氨酸、丝氨酸、苏氨酸相应的中的取代基；R₃为(CH₂CH₂O)_nH(1≤n≤20)；R₄与R₅相连形成

其中X为O、S、CHOH、NCH₃或NH；或者R₆为CH₂CH₂OH时可提高双自由基的水溶性。In the technical solution, preferably, R ₁ , R ₂ and R ₃ are not simultaneously hydrogen or R ₄ , R ₅ and R ₆ are not simultaneously methyl, preferably, R ₁ , R ₂ and R ₃ are not simultaneously hydrogen. According to the related research on the use of nitroxide biradicals as DNP polarizers, it is confirmed that electron relaxation time, exchange interaction, water solubility and other factors will affect the DNP properties of polarizers. Interaction and water solubility, 1) the modification of R ₁ , R ₂ and R ₃ groups can moderately improve the exchange interaction; 2) the modification of R ₄ , R ₅ , R ₆ groups can improve the relaxation time of diradicals , in which R ₄ is connected with R ₅ to form

When R ₆ is CD ₃ , the relaxation time of the triarylmethyl radical part can be prolonged; 3) R ₂ , R ₃ , R ₄ , R ₅ , R ₆ Modification can improve the water solubility of diradicals. R ₂ is the corresponding substituent in glutamic acid, aspartic acid, serine and threonine; R ₃ is (CH ₂ CH ₂ O) _n H (1≤n≤20); R ₄ is connected to R ₅ form

Wherein X is O, S, CHOH, NCH ₃ or NH; or when R ₆ is CH ₂ CH ₂ OH, the water solubility of the diradical can be improved.

In the technical scheme, preferably, the natural amino acids are glycine, alanine, valine, leucine, isoleucine, methionine, proline, phenylalanine, tyrosine, tryptophan , serine, threonine, asparagine, glutamine, cysteine, lysine, arginine, histidine, aspartic acid or glutamic acid, unnatural amino acids are sarcosine or tertiary Leucine.

In the technical scheme, preferably, the ring structure is a substituted or unsubstituted four-membered ring, five-membered ring or six-membered ring, preferably, the ring structure is cyclobutane, cyclopentane, cyclohexane, tetrahydrofuran , tetrahydropyran, thiocyclopentane, cyclobutanol, cyclopentanol or cyclohexanol, more preferably, the cyclic structure is tetrahydropyran or cyclohexanol.

In the technical scheme, preferably, it is represented by one of the following formulas 2-10:

In the technical scheme, preferably, it is represented by one of the following formulas 3-7:

Another object of the present invention is to provide a method for preparing the above-mentioned diradical compound and salt thereof, comprising:

The first step is to make the compound with the structure of formula A1 and the compound with the structure of formula A2 to undergo acid amine condensation reaction to obtain the compound with the structure of formula A3;

The second step is to make the compound with the structure of formula A3 and the compound with the structure of formula A4 to undergo acid-amine condensation reaction to obtain the biradical compound and its salt

In the technical scheme, preferably, the first step includes dissolving the compounds A1 and A2 in a first solvent, adding a first condensing agent and a first catalyst for acid-amine condensation reaction, and the first solvent is dichloromethane , a mixture of one or more of ethyl acetate, benzene, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, tetrahydrofuran and ether, the first A condensing agent is DCC (dicyclohexylcarbodiimide), EDCI (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride), BOP (benzotriazole- 1-yloxytris(dimethylamino)phosphonium hexafluorophosphate), TBTU (O-benzotriazole-N,N,N',N'-tetramethylurea tetrafluoroborate) , HBTU (benzotriazole-N,N,N',N'-tetramethylurea hexafluorophosphate), DIC (N,N-diisopropylcarbodiimide) and HATU (2-( One or more combinations in 7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate), the first catalyst is HOBt (1- One or more combinations of hydroxybenzotriazole), DMAP (4-dimethylaminopyridine) and DIPEA (N,N-diisopropylethylamine);

Preferably, the first solvent is a mixture of one or more of N,N-dimethylacetamide, tetrahydrofuran and dichloromethane;

Preferably, the first condensing agent is one or more combinations of EDCI, HATU and BOP;

Preferably, the first catalyst is HOBt.

In the technical scheme, preferably, the second step includes dissolving compounds A3 and A4 in a second solvent, adding a second condensing agent and a second catalyst for acid-amine condensation reaction, and the second solvent is dichloromethane, acetic acid A mixture of one or more of ethyl ester, benzene, toluene, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, tetrahydrofuran and diethyl ether, the second condensation The mixture is a combination of one or more of DCC, EDCI, BOP, TBTU, HBTU, DIC and HATU, and the second catalyst is a combination of one or more of HOBt, DMAP and DIPEA;

Preferably, the second solvent is a mixture of one or more of dichloromethane, N,N-dimethylformamide and ethyl acetate;

Preferably, the second condensing agent is one or a combination of HATU, HBTU and BOP;

Preferably, the second catalyst is HOBt.

One object of the present invention is to provide the application of the above-mentioned diradical compounds and their salts in dynamic nuclear polarization.

The invention has the advantages and positive effects that: the compound connects the nitroxide radical with the triarylmethyl radical through the linking chain, forming a double radical polarization with both nitroxide radical and triarylmethyl radical agent. The advantages of the double radical polarizer are: the frequency matching of nitroxide radical-triarylmethyl radical type double radical is better than that of nitroxide radical, and the CE efficiency is higher; Under the condition of magic angle rotation, the nitroxyl radical type diradical has no depolarization effect, which makes its high-field DNP enhancement effect better than that of dinitroxide radical; nitroxyl radical-triarylmethyl radical type The strong exchange interaction of diradicals is obviously different from that of nitroxide diradicals, and because the influence of high-field DNP dipole interaction is very small, the dominant role is the exchange interaction. Therefore, the strong exchange interaction The effect also makes the compound have a higher high-field DNP enhancement multiple than the nitroxide biradical compound; the triarylmethyl radical has good water solubility, and is rich in modifiable sites, which can be used for functional derivation for different purposes. The modification of R ₁ , R ₂ , R ₃ , R ₄ , R ₅ , and R ₆ groups in the present invention can moderately improve the exchange interaction, improve the relaxation time of diradicals and the water solubility of diradicals, Thereby enhancing the DNP properties of the polarizer.

Description of drawings

Fig. 1 is a high-resolution mass spectrum of compound 1 in Example 1 of the present invention.

Fig. 2 is a high-resolution mass spectrum of compound 2 in Example 2 of the present invention.

3 is a high-resolution mass spectrum of compound 3 in Example 3 of the present invention.

4 is a high-resolution mass spectrum of compound 4 in Example 4 of the present invention.

Fig. 5 is a high-resolution mass spectrum of compound 5 in Example 5 of the present invention.

6 is a high-resolution mass spectrum of compound 6 in Example 6 of the present invention.

7 is a high-resolution mass spectrum of compound 7 in Example 7 of the present invention.

FIG. 8 is a high-resolution mass spectrum of compound 8 in Example 8 of the present invention.

9 is a high-resolution mass spectrum of compound 9 in Example 9 of the present invention.

Detailed ways

The specific embodiments of the present invention are further introduced below in conjunction with the examples:

Example 1

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

The preparation steps of compound 1 are as follows in formula 11:

1. Preparation of compound A4:

Second, the preparation of compound 1:

1. Dissolve compound A1 (1-1.5eq) and HOBt (2-5eq) in redistilled DCM, stir, add DIPEA (3-10eq), BOP (0.9-1.5eq) first dissolve in redistilled DCM, then The reaction solution was added, activated for 0.5-2h, then compound A2 (0.9-2eq) was dissolved in redistilled DCM, added to the reaction solution, and reacted at room temperature for 0.5-18h. Treatment: extracted with DCM, water, washed with citric acid, saturated sodium carbonate solution, saturated brine in turn, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated by normal phase silica gel column, mobile phase: ethyl acetate: petroleum Ether=1:4-2:1 to give product A3 in 65-95% yield.

2. Use trifluoroacetic acid or formic acid to excise the Boc group of compound A3, condition: A3 is stirred at room temperature for 0.5-8h in pure acid or acid in dichloromethane solution (acid ratio is 1:5 to 2:1); then After the solvent and acid were removed in vacuo, A3 with Boc excised was obtained by alkalization and extraction.

3. Dissolve A4 (0.9-1.5eq) in DMF and add DIPEA (3-20eq), dissolve HBTU (0.9-2eq) in DMF, add dropwise to the reaction solution, activate for 0.5-2h, and remove Boc The A3 was dissolved in an appropriate amount of DMF, slowly added dropwise (5-60min) to the reaction solution, and the reaction was complete after 4-24h. Dilute hydrochloric acid was added, extracted with ethyl acetate and water, the organic layers were combined, concentrated under reduced pressure, neutralized with NaOH solution to form a salt, and separated by reverse-phase silica gel column chromatography. Yields are around 15-50%.

市售可购买。化合物A2

市售可购买。In this embodiment, compound A1 is

Commercially available. Compound A2

Commercially available.

Figure 1 shows the high-resolution mass spectrometry data of compound 1 in this example.

3. DNP-enhancing effect experiment of compound 1

The DNP experiment is to dissolve the double radical polarizer into DNP juice (glycerol/water/heavy water 60%/30%/10%) to prepare a solution with a concentration of 10mM, and the substrate is ¹³ C- ¹⁵ N proline. DNP experiments were carried out on an 800MHz Bruker BioSpin527GHz solid-state NMR spectrometer at a temperature of 100K.

The DNP enhancement fold of compound 1 was 57.

Embodiment 2

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

1. The preparation steps of compound 2 are the same as the method in Example 1.

市售可购买。In this embodiment, compound A1 is

Commercially available.

Figure 2 shows the high-resolution mass spectrometry data of compound 2 in this example

2. DNP-enhancing effect experiment of compound 2

The experimental method of DNP enhancement performance test is the same as that of compound 1 test.

The DNP enhancement fold of compound 2 was 76. Compared with compound 1, the DNP enhancement factor has increased by 33%, and the improvement effect is remarkable.

Embodiment 3

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

The preparation step of compound 3 is as follows formula 13:

1. Preparation of compound 3:

Diethylene glycolamine (0.5-2eq), imidazole (2-5eq), DMAP (0.05-0.5eq), tert-butyldimethylsilyl chloride (0.5-2eq) were placed in a reaction flask, argon protection, ice bath, adding anhydrous DCM, stirring at zero temperature for half an hour, then transferring to room temperature and stirring for 2-18 hours to complete the reaction. Post-reaction treatment: washing with saturated brine, and evaporating to dryness to obtain pure product 6, which is a colorless and transparent oil with a yield of 75-95%.

The product obtained in the previous step (0.9-2eq) and nitroxide radical (1eq) were dissolved in anhydrous THF, under argon protection, stirred at room temperature for 2 hours, and then added in batches of sodium triacetoxyborohydride (1.5eq), After stirring at room temperature for 16 hours, the reaction was complete. It was concentrated under reduced pressure, extracted with DCM, washed with saturated sodium bicarbonate and saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and separated by normal phase silica gel column chromatography. The mobile phase was 0-5% methanol/dichloromethane. Yield 55-75%.

The remaining steps are the same as above. In addition, when removing Boc, TBS will also be completely removed.

The raw materials are commercially available.

Figure 3 shows the high-resolution mass spectrometry data of compound 3 in this example.

2. Experiment of DNP-enhancing effect of compound 3

The experimental method of DNP enhancement performance test is the same as compound 1.

The DNP enhancement factor of compound 3 is 44. Although the enhancement factor is not as good as that of compound 1, it still maintains a considerable DNP enhancement factor while weakening the intermolecular hydrogen bond interaction. The performance is significantly better than that of compound 1. For example, in the test of some lipid-soluble protein or membrane protein samples, the water solubility and dispersibility of compound 3 are better than that of compound 1, which can reflect better performance.

Embodiment 4

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

1. Preparation of compound 4:

The preparation method of compound 4 is the same as that of compound 3.

Figure 4 shows the high-resolution mass spectrometry data of compound 4 in this example.

2. Experiment of DNP-enhancing effect of compound 4

The experimental method of DNP enhancement performance test is the same as compound 1.

The DNP enhancement factor of compound 4 is 20. Although the enhancement factor is lower than that of compound 1, it still maintains a considerable DNP enhancement factor while weakening the intermolecular hydrogen bond interaction. The performance is significantly better than that of compound 1. For example, in the test of some lipid-soluble protein or membrane protein samples, the water solubility and dispersibility of compound 4 are better than that of compound 1, which can reflect better performance.

Embodiment 5

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

The preparation procedure of compound 5 is as follows formula 14:

1. Preparation of compound 5:

2,2,6,6-Tetramethyl-4-piperidinone (0.5-2eq), 4-hydroxycyclohexanone (3-6eq) and _NH4Cl (3-6eq) were dissolved in DMSO, And heated and stirred at 40-60 ° C for 5-12 hours. Add dilute hydrochloric acid to acidify, extract with petroleum ether, adjust pH=9 with saturated sodium carbonate solution, concentrate under reduced pressure to obtain the crude product, which is purified by normal phase silica gel column chromatography, the mobile phase is 1-5% MeOH/DCM, to obtain the product As a white solid, the yield is 10-30%.

30% H ₂ O ₂ (5-10 eq) was added to an aqueous solution of the product from the previous step (0.5-2 eq) and Na ₂ WO ₄ ·2H ₂ O (0.3-1 eq) in an ice bath, and stirred at room temperature overnight. Concentration under reduced pressure gave the crude product, which was purified by silica gel column chromatography, the mobile phase was 1-3% MeOH/CH ₂ Cl ₂ , and the product was a red oil with a yield of 70-85%.

The subsequent reaction is the same as compound 3.

Raw materials are commercially available.

Figure 5 shows the high-resolution mass spectrometry data of compound 5 in this example.

2. Experiment of DNP-enhancing effect of compound 5

The DNP enhancement multiple of compound 5 is 72 times, which is 26% higher than that of compound 1, and the improvement effect is obvious.

Embodiment 6

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

The preparation procedure of compound 6 is as follows formula 15:

1. Preparation of compound 6:

To a solution of 2,2,6,6-tetramethyl-4-piperidinone (0.5-2eq) in toluene (5-15mL) was added paraformaldehyde (1-2eq), the resulting solution was heated at 100-120°C Heating and stirring and refluxing for 0.5-2 hours, then dropwise addition of formic acid (0.5-2eq), after refluxing for 3-8 hours, the reaction solution was cooled to room temperature. It was extracted with ethyl acetate, washed with 1N NaOH and saturated brine in this order. The organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product, which was purified by normal phase silica gel column chromatography, and the mobile phase was ethyl acetate: petroleum ether=1:3 to obtain the product in a yield of 40 -60%.

Dissolve 1,2,2,6,6-pentamethyl-4-piperidinone (0.5-2eq), 4-hydroxycyclohexanone (3-6eq) and _NH4Cl (3-6eq) in DMSO , and heated and stirred at 50-70 ° C for 6-12 hours. Acidify with dilute hydrochloric acid, extract with petroleum ether, adjust pH=9 with saturated sodium carbonate solution, concentrate under reduced pressure to obtain crude product, which is purified by normal phase silica gel column chromatography, mobile phase is 5% MeOH/DCM, and the product is white Solid, 12% yield.

30% H ₂ O ₂ (5-12 eq) was added to an aqueous solution of the product from the previous step (0.5-2 eq) and Na ₂ WO ₄ ·2H ₂ O (0.3-1 eq) in an ice bath, and stirred at room temperature overnight. Concentration under reduced pressure gave the crude product, which was purified by silica gel column chromatography, the mobile phase was 1-3% MeOH/CH ₂ Cl ₂ , and the product was a red oil with a yield of 75-90%.

The subsequent reaction is the same as compound 3.

The raw materials are all commercially available.

FIG. 6 is the high-resolution mass spectrometry data of compound 6 in this example.

2. Experiment of DNP-enhancing effect of compound 6

The experimental method of DNP enhancement performance test is the same as compound 1.

The DNP enhancement fold of compound 6 was 95.

Embodiment 7

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

1. Preparation of compound 7:

The preparation method of compound 7 is the same as that of compound 1.

市售可购买。In this embodiment, compound A1 is

Commercially available.

Figure 7 shows the high-resolution mass spectrometry data of compound 7 in this example.

2. Experiment of DNP-enhancing effect of compound 7

The experimental method of DNP enhancement performance test is the same as compound 1.

The DNP enhancement factor of compound 4 was 40.

Embodiment 8

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

1. Preparation of compound 8:

The preparation method of compound 8 is the same as that of compound 1.

市售可购买。In this embodiment, compound A1 is

Commercially available.

FIG. 8 is the high-resolution mass spectrometry data of compound 8 in this example.

2. Experiment of DNP-enhancing effect of compound 8

The experimental method of DNP enhancement performance test is the same as compound 1.

The DNP enhancement factor of compound 8 was 30.

Embodiment 9

The biradical compound and its salt that a kind of triarylmethyl radical and nitroxide radical described in this embodiment are formed, the structural formula is as follows:

1. Preparation of compound 9:

The preparation method of compound 9 is the same as that of compound 1.

Figure 9 shows the high-resolution mass spectrometry data of compound 9 in this example.

2. Experiment of DNP-enhancing effect of compound 9

The experimental method of DNP enhancement performance test is the same as compound 1.

The DNP enhancement fold of compound 8 was 43.

Several embodiments of the present invention have been described in detail above, but the above contents are only preferred embodiments of the present invention and cannot be considered to limit the scope of implementation of the present invention. All equivalent changes and improvements made according to the scope of the application of the present invention should still belong to the scope of the patent of the present invention.

Claims (13)

1. A diradical compound consisting of a triarylmethyl radical and a nitroxide radical and a salt thereof, the structural formula of which is represented by the following formula 1:

wherein,

R₁is H or CH₃；

R₂Is H or CH₃；

R₃Is H;

R₄and R₅Are connected to form

R₆Is CH₃。

2. The diradical compound and the salt thereof according to claim 1, wherein: r₁、R₂And R₃Not hydrogen at the same time.

3. The diradical compound and the salt thereof according to claim 1, wherein: represented by the following formula 6 or 7:

4. a method for producing the diradical compound and the salt thereof according to any one of claims 1 to 3, wherein: the method comprises the following steps:

firstly, carrying out acid amine condensation reaction on a compound A1 and a compound A2 to obtain a compound A3;

secondly, carrying out acid-amine condensation reaction on a compound A3 and a compound A4 to obtain the diradical compound and salt thereof;

5. the method for producing a diradical compound and a salt thereof according to claim 4, wherein: the first step comprises the steps of dissolving a compound A1 and A2 in a first solvent, adding a first condensing agent and a first catalyst to perform acid-amine condensation reaction, wherein the first solvent is one or a mixture of more of dichloromethane, ethyl acetate, benzene, toluene, N-dimethylformamide, N-dimethylacetamide, dimethyl sulfoxide, tetrahydrofuran and diethyl ether, the first condensing agent is one or a combination of more of DCC, EDCI, BOP, TBTU, HBTU, DIC and HATU, and the first catalyst is one or a combination of more of HOBt, DMAP and DIPEA.

6. The method for producing a diradical compound and a salt thereof according to claim 5, wherein: the first solvent is one or a mixture of N, N-dimethylacetamide, tetrahydrofuran and dichloromethane.

7. The method for producing a diradical compound and a salt thereof according to claim 5, wherein: the first condensing agent is one or a combination of more of EDCI, HATU and BOP.

8. The method for producing a diradical compound and a salt thereof according to claim 5, wherein: the first catalyst is HOBt.

9. The method for producing a diradical compound and a salt thereof according to any one of claims 4 to 8, wherein: the second step comprises the steps of dissolving a compound A3 and A4 in a second solvent, adding a second condensing agent and a second catalyst for carrying out acid-amine condensation reaction, wherein the second solvent is one or a mixture of more of dichloromethane, ethyl acetate, benzene, toluene, N-dimethylformamide, N-dimethylacetamide, dimethyl sulfoxide, tetrahydrofuran and diethyl ether, the second condensing agent is one or a combination of more of DCC, EDCI, BOP, TBTU, HBTU, DIC and HATU, and the second catalyst is one or a combination of more of HOBt, DMAP and DIPEA.

10. The method for producing a diradical compound and a salt thereof according to claim 9, wherein: the second solvent is one or a mixture of more of dichloromethane, N-dimethylformamide and ethyl acetate.

11. The method for producing a diradical compound and a salt thereof according to claim 9, wherein: the second condensing agent is one or a combination of HATU, HBTU and BOP.

12. The method for producing a diradical compound and a salt thereof according to claim 9, wherein: the second catalyst is HOBt.

13. Use of the diradical compound and the salt thereof according to any one of claims 1-3 for dynamic nuclear polarisation.

Images