Micromolecular manganese chelate, preparation method thereof and contrast agent

Abstract

The invention belongs to the field of medical detection, and particularly relates to a micromolecular manganese chelate (MnL), a preparation method thereof and a contrast agent. The preparation method of the micromolecular manganese chelate is simple, the micromolecular manganese chelate can be self-assembled into nanoparticles without assistance of a carrier, and the micromolecular manganese chelate has high T1 relaxation efficiency, prolonged in-vivo circulation time and excellent stability. The micromolecular manganese chelate nanoparticles have wide application prospects in the field of magnetic resonance imaging (MRI) of liver, kidney, heart and blood vessels, lymph nodes and tumors.

Description

Technical Field

[0001] The present invention belongs to the field of medical detection, and particularly relates to a small molecule manganese chelate (MnL), a preparation method thereof, and a contrast agent. Background Art

[0002] Magnetic resonance imaging (MRI) is widely used in the diagnosis of various diseases because it can non-invasively obtain anatomical details of various diseases through precise contrast between soft tissues. More than one-third of clinical MRI examinations are performed with the assistance of contrast agents. Currently, the contrast agents reported clinically are mainly small molecule gadolinium chelates such as Gd-DOTA and Gd-DTPA, etc., which have deficiencies such as low T1 relaxation efficiency and short half-life, resulting in poor imaging effects. In addition, clinical results show that a part of gadolinium-based contrast agents can cause nephrogenic systemic fibrosis in patients with renal insufficiency and organ gadolinium deposition after multiple injections, especially in the brain. Therefore, there is an urgent need to develop safer and more efficient magnetic resonance contrast agents.

[0003] In recent years, studies have found that manganese contrast agents are expected to be an ideal gadolinium-based alternative contrast agent. This is mainly due to 1) manganese ions are essential trace elements in the human body. Compared with gadolinium ions, they have lower toxicity and avoid the risk of organ deposition; 2) manganese ions have five unpaired d electrons and exhibit all the necessary physical properties of MRI contrast agents: fast water exchange rate, long longitudinal electron relaxation time, and high spin quantum number. However, the small molecule manganese contrast agents reported currently still have many deficiencies: such as low T1 relaxation efficiency, fast metabolism, and poor specificity, resulting in poor magnetic resonance imaging effects.

[0004] Therefore, there is an urgent need to develop manganese-based magnetic resonance contrast agents that are safer, have higher T1 relaxation efficiency, longer in vivo circulation time, and ideal specificity.

[0005] In view of the above reasons, the present invention is specifically proposed. Summary of the Invention

[0006] In order to solve the problems existing in the prior art, the present invention provides a small molecule manganese chelate, and the structure of the small molecule manganese chelate is shown in Formula I:

[0007]

[0008] Wherein: R is one of a hydrophobic molecule containing a benzene ring, a hydrophobic molecule containing a pyridine ring, and a hydrophobic molecule containing a linear carbon chain.

[0009] Based on the same technical concept, another solution of the present invention is to provide a preparation method of a small molecule manganese chelate, and the preparation method includes the following steps:

[0010] (1) Mix 2,6 - diacetylpyridine, a small molecule monoacylhydrazide, anhydrous manganese chloride, and anhydrous methanol, filter after reflux reaction to obtain a filtrate;

[0011] (2) Subject the filtrate to sedimentation, washing, and drying in sequence to obtain the small molecule manganese chelate.

[0012] Preferably, in step (1), the molar ratio of 2,6 - diacetylpyridine, the small molecule monoacylhydrazide, anhydrous manganese chloride, and anhydrous methanol is 1:1:0.9:1 - 100.

[0013] Preferably, in step (1), the structure of the small molecule monoacylhydrazide is as shown in formula II:

[0014]

[0015] Wherein: R is one of a hydrophobic molecule containing a benzene ring, a hydrophobic molecule containing a pyridine ring, and a hydrophobic molecule containing a linear carbon chain.

[0016] Preferably, in step (1), the reflux reaction time is 1 - 24 h.

[0017] Preferably, in step (2), subject the filtrate to sedimentation with an isopropanol / ether mixture first, then wash with ether, and finally dry under vacuum to obtain the small molecule manganese chelate.

[0018] Based on the same technical concept, another solution of the present invention is to provide a contrast agent, which is self - assembled from the small molecule manganese chelate in an aqueous medium. It should be noted that the small molecule manganese chelate MnL has amphiphilic properties and can self - assemble into nanoparticles in an aqueous medium, and the nanoparticles can be used as a contrast agent for magnetic resonance imaging.

[0019] More specifically, it can be used for magnetic resonance imaging of the brain, liver, cardiovascular system, kidneys, bladder, breast, lymph nodes, and corresponding tumors, etc.

[0020] The beneficial effects of the present invention are as follows:

[0021] 1. Compared with the preparation of traditional small molecule manganese chelates, its one - step preparation strategy is simple, efficient, and has greater potential for clinical translation.

[0022] 2. Compared with traditional small molecule manganese chelates, its amphiphilic properties enable it to self - assemble into nanoparticles in water without the assistance of other carriers, showing higher T1 relaxation efficiency, longer in - vivo circulation time, and better in - vivo magnetic resonance imaging effect.

[0023] 3. Compared with traditional small-molecule manganese chelates, its rigid structure endows it with higher stability, thus showing more ideal in vivo safety. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can also be obtained based on these drawings.

[0025] Figure 1 It is the structural diagram of the small-molecule manganese chelate MnL in Example 1.

[0026] Figure 2 It is the transmission electron microscope image of the small-molecule manganese chelate MnL nanoparticles.

[0027] Figure 3 It is the T1 relaxation efficacy result diagram of the small-molecule manganese chelate MnL nanoparticles.

[0028] Figure 4 It is the stability evaluation result diagram of the small-molecule manganese chelate MnL nanoparticles.

[0029] Figure 5 It is the liver magnetic resonance imaging result diagram of the small-molecule manganese chelate MnL nanoparticles.

[0030] Figure 6 It is the kidney magnetic resonance imaging result diagram of the small-molecule manganese chelate MnL nanoparticles. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0031] To make the objectives, technical solutions, and advantages of the present invention clearer, the following will describe the technical solutions of the present invention in detail. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of them. Based on the embodiments of the present invention, all other implementation manners obtained by those of ordinary skill in the art without creative efforts fall within the scope of protection of the present invention.

[0032] Example 1

[0033] This example provides a preparation method for a small-molecule manganese chelate. The preparation method is as follows:

[0034] Weigh 0.369 g of 2,6-diformylpyridine, 0.732 g of 4-fluorobenzoylhydrazide, and 0.27 g of anhydrous manganese chloride respectively. Dissolve them separately in 10 mL of anhydrous methanol, then add them successively to a 100 mL reaction flask. After refluxing for 24 h, filter off the precipitate while it is hot, concentrate by rotary evaporation, and finally deposit with a 10-fold volume of a mixed solution of isopropanol / ether, and dry in vacuo to obtain the target product, the small molecule manganese chelate MnL, whose structure is as shown in Figure 1 .

[0035] Example 2

[0036] This example provides a preparation method of a contrast agent, and the preparation method is as follows:

[0037] Take 20 mg of the small molecule manganese chelate MnL obtained in Example 1, dissolve it in 5 mL of methanol, then add it to 10 mL of deionized water, remove the methanol by rotary evaporation, and finally obtain nanoparticles based on the small molecule manganese chelate MnL, and use them as contrast agents.

[0038] Verification Example

[0039] (I) Transmission electron microscopy imaging of small molecule manganese chelate MnL nanoparticles

[0040] Take a small amount of the diluted dispersion of small molecule manganese chelate MnL nanoparticles and drop it onto a pure carbon film copper grid. After the water volatilizes, observe the particle size and morphology of the small molecule manganese chelate MnL nanoparticles through transmission electron microscopy. The results are as shown in Figure 2 .

[0041] It can be seen from Figure 2 that the small molecule manganese chelate MnL nanoparticles exhibit a spherical morphology, with a uniform particle size distribution and an average size of about 2.5 nm.

[0042] (II) Evaluation of the T1 relaxation efficiency of small molecule manganese chelate MnL nanoparticles

[0043] Dilute the small molecule manganese chelate MnL nanoparticles with deionized water to obtain 8 samples with different manganese concentrations (0.5, 0.4, 0.3, 0.25, 0.15, 0.1, 0.06, and 0.01 mM). Then use a 1.5 T clinical magnetic resonance scanning system (Siemens) to detect the T1 relaxation efficiency of the samples. Specifically, first fix the prepared samples in the head coil of the magnetic resonance instrument, and then obtain the T1-weighted images of each sample through an inversion recovery (IR) sequence scan (TE = 11.7 ms, TR = 30 - 3300 ms), and obtain the magnetic resonance signal values of each sample at different TR times through the threshold value. Import these signal values into the software, calculate the T1 relaxation times of each sample at the above concentration gradients through the formula, and finally use the reciprocal of the T1 relaxation time (1 / T1, s-1 ) Perform a linear fit on the sample concentration (mM) to obtain a fitting curve, and the slope of the curve is the relaxation efficiency of the sample (r1, mM -1 s -1 ) The results are as Figure 3 shown.

[0044] From Figure 3 it can be seen that: under a 1.5 T magnetic field, the T1 relaxation efficiency of the small molecule manganese chelate MnL nanoparticles is 11.6 mM -1 s -1 , which is significantly higher than the T1 relaxation efficiency of most small molecule manganese chelates reported.

[0045] (III) Evaluation of the stability of small molecule manganese chelate MnL nanoparticles

[0046] Disperse the small molecule manganese chelate MnL nanoparticles in buffers with pH values of 7.4, 5.0, and 3.0, as well as in a pH 6.0 buffer with zinc ion concentrations of 2.5 mM and 5 mM, respectively, with a final manganese concentration of 0.5 mM. Subsequently, detect the T1 relaxation time of the above solutions at different time points (0, 0.5, 1, 2, 4, 8, 24, 72 h), and evaluate the stability of the small molecule manganese chelate MnL nanoparticles by comparing the changes in the T1 relaxation time. The results are as Figure 4 shown.

[0047] From Figure 4 it can be seen that: the relaxation time of the small molecule manganese chelate MnL nanoparticles does not change significantly under different pH values and in the presence of excessive zinc ion competition, indicating that no manganese ion dissociation occurs, which proves that the small molecule manganese chelate MnL nanoparticles have good chelation stability.

[0048] (IV) Evaluation of in vivo liver and kidney magnetic resonance imaging of small molecule manganese chelate MnL nanoparticles

[0049] Evaluate the in vivo liver and kidney magnetic resonance imaging effect of the small molecule manganese chelate MnL nanoparticles by tail vein injection (dose: 0.05 mmol Mn / kg Kunming mice (30 - 32 g)). The specific steps are as follows: First, collect the magnetic resonance images of the liver and kidneys of Kunming mice before injecting the contrast agent through a 3.0 T magnetic resonance scanning system (Siemens). Subsequently, inject the small molecule manganese chelate MnL nanoparticles through the tail vein, and collect the magnetic resonance images of the liver and kidneys of Kunming mice at different times Figure 5 and Figure 6 are the magnetic resonance images of the liver and kidneys before and after drug administration, respectively.

[0050] From Figure 5It can be seen that: compared with before the injection of the small molecule manganese chelate MnL nanoparticles, the magnetic resonance signals of the heart and liver of the mice are significantly enhanced after the injection, indicating that the small molecule manganese chelate MnL nanoparticles are a potential contrast agent for cardiovascular and liver imaging.

[0051] It can be seen from Figure 6 It can be seen that: compared with before the injection of the small molecule manganese chelate MnL nanoparticles, the magnetic resonance signal of the kidney of the mice is significantly enhanced after the injection, indicating that the small molecule manganese chelate MnL nanoparticles are a potential contrast agent for kidney imaging.

[0052] As described above, it is only the specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art within the technical scope disclosed by the present invention can easily think of changes or substitutions, which should all be covered by the protection scope of the present invention. Therefore, the protection scope of the present invention should be subject to the protection scope of the claims.

Claims

1. A small molecule manganese chelate, characterized in that, The structure of the small molecule manganese chelate is shown in Formula I: Wherein: R is one of a hydrophobic molecule containing a benzene ring, a hydrophobic molecule containing a pyridine ring, and a hydrophobic molecule containing a linear carbon chain.

2. The preparation method of the small molecule manganese chelate according to claim 1, characterized in that, The preparation method comprises the following steps: (1) Mix 2,6-diacetylpyridine, a monoacylhydrazide small molecule, anhydrous manganese chloride, and anhydrous methanol, filter after reflux reaction to obtain a filtrate; (2) Subject the filtrate to sedimentation, washing, and drying in sequence to obtain the small molecule manganese chelate.

3. The preparation method of the small molecule manganese chelate according to claim 2, characterized in that, In step (1), the molar ratio of 2,6-diacetylpyridine, the monoacylhydrazide small molecule, anhydrous manganese chloride, and anhydrous methanol is 1:1:0.9:1 to 100.

4. The preparation method of the small molecule manganese chelate according to claim 2, wherein In step (1), the structure of the monoacylhydrazide small molecule is shown in Formula II: Wherein: R is one of a hydrophobic molecule containing a benzene ring, a hydrophobic molecule containing a pyridine ring, and a hydrophobic molecule containing a linear carbon chain.

5. The preparation method of the small molecule manganese chelate according to claim 2, wherein In step (1), the time of the reflux reaction is 1 to 24 h.

6. The preparation method of the small molecule manganese chelate according to claim 2, characterized in that, In step (2), subject the filtrate to sedimentation with an isopropanol / ether mixed solution first, then wash with ether, and finally dry under vacuum to obtain the small molecule manganese chelate.

7. A contrast agent, characterized in that, The contrast agent is self-assembled from the small molecule manganese chelate described in claim 1 in an aqueous medium.

8. The contrast agent according to claim 7, characterized in that, The contrast agent can be applied in magnetic resonance imaging examinations.

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