Application of magneto-optical bifunctional material based on beta-diketone rare earth gadolinium complex in fluorescence imaging

Abstract

The invention provides application of a magneto-optical bifunctional material based on a beta-diketone rare earth gadolinium complex in fluorescence imaging, and belongs to the technical field of application of rare earth complexes. According to the application of the magneto-optical bifunctional material based on the beta-diketone rare earth gadolinium complex in fluorescence imaging, the expression of the magneto-optical bifunctional material is [Gd (dppz) (BTA) 3], BTA is benzoyl trifluoroacetone, dppz is dipyridino [3, 2-a: 2 ', 3'-c] phenazine, and Gd is gadolinium. The material has remarkable magnetic and fluorescent properties, can be used for carrying out broad-spectrum imaging on six common cancer cells, is remarkably superior to the existing magneto-optical bifunctional material which only has a specific imaging function on one kind of cancer cells, and shows wide biomedical application potential.

Description

Technical Field

[0001] The present invention belongs to the technical field of rare earth complex application, and particularly relates to β -Magneto-optical dual-functional materials of diketonate rare earth gadolinium complexes for application in fluorescence imaging. Background Art

[0002] Magneto-optical dual-functional materials can fully combine magnetism and luminescence, and the rational design of such materials has attracted more and more people's interest. Among them, rare earth organic complexes play an important role in the research of magnetic refrigeration materials and bioimaging materials, because of their rich ligand design, 4f single electrons that can be used for magnetic resonance imaging, and excellent fluorescence properties (good monochromaticity, long fluorescence lifetime, high fluorescence quantum yield, large Stokes shift), etc. Reasonable ligand design can make rare earth complex magneto-optical dual-functional composite materials widely used in the field of medical diagnosis and treatment.

[0003] because β -Diketones have strong coordination ability and high absorption coefficient, which can not only realize energy exchange with rare earth ions, but also control the coordination environment of rare earth ions, construct high-performance magnetic refrigeration materials, and form β -Diketone rare earth organic complexes have stable structures and strong binding ability. In addition, since rare earth ions themselves have excellent optical and electromagnetic properties, through the combination of rare earth ions and suitable β -Diketone ligand binding, the obtained β -Diketone rare earth organic complexes have unique electrical, optical, magnetic, thermal and other properties, especially fluorescence properties.

[0004] However, different β- The structure of diketone rare earth complexes has certain differences in their functions. Therefore, it is necessary to study the broad-spectrum applicability of β -The magneto-optical dual-functional materials of diketone rare earth complexes are of great significance. Summary of the invention

[0005] The present invention proposes a method based on β- The magneto-optical dual-functional materials of diketonate rare earth gadolinium complexes are applicable to a variety of cancer cells within the concentration range used in fluorescence imaging studies.

[0006] The present invention proposes a method based on β- Application of magneto-optical dual-function material of diketonate rare earth gadolinium complex in fluorescence imaging, characterized in that the expression of the magneto-optical dual-function material is [Gd(dppz)(BTA) 3 ], wherein BTA is benzoyltrifluoroacetone, dppz is dipyrido[3,2-a:2',3'-c]phenazine, and Gd is gadolinium.

[0007] Furthermore, the magneto-optical dual-functional material performs specific imaging on six types of cancer cells, including human cervical cancer cells, human breast cancer cells, human non-small cell lung cancer cells, human gastric cancer cells, liver cancer cell line G2 type and human prostate cancer cells.

[0008] Furthermore, the concentration of the magneto-optical dual-functional material in fluorescence imaging is 10-20 μmol mL -1 .

[0009] Furthermore, the fluorescence emission wavelength of the magneto-optical dual-function material in fluorescence imaging is 618-621 nm.

[0010] Furthermore, the magneto-optical dual-function material is prepared by the following steps: (1) adding benzoyltrifluoroacetone and triethylamine to a methanol solvent, stirring until dissolved, adding gadolinium trichloride hexahydrate, stirring to obtain a mixed solution; (2) adding dipyrido[3,2-a:2',3'-c]phenazine to the mixed solution, stirring in a semi-closed beaker at room temperature, and filtering to obtain a filtrate; (3) The filtrate is allowed to stand at room temperature for evaporation to obtain the target product.

[0011] Furthermore, in step (1), the usage ratio of benzoyltrifluoroacetone, triethylamine, methanol solvent and gadolinium trichloride hexahydrate is 0.3 mmol: 0.72 mmol: 20 mL: 0.1 mmol.

[0012] Furthermore, in step (2), the molar ratio of dipyrido[3,2-a:2',3'-c]phenazine to gadolinium trichloride hexahydrate is 1:1. Furthermore, in step (1), the stirring time is not less than 24 hours; In step (2), the stirring time is not less than 24 hours.

[0013] Furthermore, in step (3), the room temperature is 15-30°C.

[0014] The present invention has the following advantages: The magneto-optical dual-functional material [Gd(dppz)(BTA) 3 ], with significant magnetic and fluorescent properties, can perform broad-spectrum imaging of six common cancer cells, significantly better than existing magneto-optical dual-functional materials that can only perform specific imaging of one type of cancer cell, showing a wide range of biomedical application potential. This is consistent with the ligand [Gd(dppz)(BTA) 3] have a certain relationship with the structure of dipyrido[3,2-a:2',3'-c]phenazine (dppz) ligand. The complex formed by the dipyrido[3,2-a:2',3'-c]phenazine (dppz) ligand has high coordination stability and complex structure. In addition, dppz has good cell membrane affinity and can efficiently enter cells through direct penetration or endocytosis. At the same time, it exhibits low toxicity, good biocompatibility and certain targeting. It is suitable for drug delivery and cell imaging and can be widely used in cancer diagnosis, targeted therapy and magnetic materials. It has important scientific research value and commercial prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0016] Figure 1 Prepared by Comparative Examples 1-2 and Example 1 of the present invention β- The crystal structure of the smallest asymmetric unit and coordination diagram of the magneto-optical dual-functional material of the diketonate rare earth gadolinium complex; Figure 2 Prepared by Comparative Examples 1-2 and Example 1 of the present invention β- The X-ray powder diffraction pattern (PXRD) of the magneto-optical dual-functional material of the diketonate rare earth gadolinium complex and the theoretical X-ray powder diffraction pattern of the material; Figure 3 Prepared by Comparative Examples 1-2 and Example 1 of the present invention β- Magneto-optical dual-functional materials based on rare earth gadolinium diketone complexes χ M T value dependence curve on temperature; Figure 4 Prepared by Example Comparative Examples 1-2 and Example 1 β- Magneto-optical dual-functional materials based on rare earth gadolinium diketone complexes χM -1 and T Relationship diagram and Curie-Weiss fitting diagram; Figure 5 Prepared by Comparative Examples 1-2 and Example 1 of the present invention β- The relationship between the magnetization intensity of the diketonate rare earth gadolinium complex magneto-optical dual-functional material and the applied magnetic field; Figure 6 For comparative examples 1-2 and embodiment 1 of the present invention β- Fluorescence emission spectrum of diketonate rare earth gadolinium complex magneto-optical dual-functional material; Figure 7 The present invention is prepared by comparative examples 1-2 and embodiment 1β- Cancer cell imaging of diketonate rare earth gadolinium complex magneto-optical dual-functional material. DETAILED DESCRIPTION

[0017] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. In the absence of conflict, the embodiments of the present invention and the features in the embodiments can be combined with each other.

[0018] The embodiment of the present invention proposes a β- Application of magneto-optical dual-function material of diketonate rare earth gadolinium complex in fluorescence imaging, wherein the expression of the magneto-optical dual-function material is [Gd(dppz)(BTA) 3 ], wherein BTA is benzoyltrifluoroacetone, dppz is dipyrido[3,2-a:2',3'-c]phenazine, and Gd is gadolinium.

[0019] In one embodiment of the present invention, the magneto-optical bifunctional material performs specific imaging on six types of cancer cells: human cervical cancer cells (HeLa), human breast cancer cells (MCF-7), human non-small cell lung cancer cells (A549), human gastric cancer cells (Scg7901), liver cancer cell line G2 (Hep-G2) and human prostate cancer cells (DU-145).

[0020] In one embodiment of the present invention, the concentration of the magneto-optical dual-function material used in fluorescence imaging is 10-20 μmol mL -1 Within this range, based on β- The magneto-optical dual-functional material of diketonate rare earth gadolinium complex has no obvious inhibitory effect on cells. [Gd(dppz)(BTA) 3 ] enters the cytoplasm, and very little enters the cell nucleus. In one embodiment of the present invention, the fluorescence emission wavelength of the magneto-optical dual-function material in fluorescence imaging is 618-621 nm. In one embodiment of the present invention, the β- The preparation method of diketone rare earth gadolinium complex magneto-optical dual-function material comprises the following steps: (1) adding benzoyltrifluoroacetone (BTA) and triethylamine to a methanol solvent, stirring until dissolved, adding a rare earth chloride, stirring to obtain a mixed solution; (2) adding an auxiliary ligand dipyrido[3,2-a:2',3'-c]phenazine (dppz) to the mixed solution, stirring in a semi-closed beaker at room temperature, and filtering to obtain a filtrate; (3) The filtrate is allowed to stand at room temperature for evaporation to obtain the target product.

[0021] The preparation method proposed in the embodiment of the present invention comprises the following steps: β- Diketone ligand benzoyltrifluoroacetone (BTA) and rare earth chloride GdCl 3 6H 2 O. After adding triethylamine to the methanol solvent and stirring continuously, add the auxiliary ligand [3,2-a:2',3'-c]phenazine (dppz), continue stirring and filter, seal the obtained mixed system, let it stand, and evaporate the solvent to obtain a pure crystalline sample. The method uses methanol as a solvent and can prepare a magneto-optical dual-functional material by a solution method under stirring conditions. The process is simple, the raw material cost is low, the safety is good, and the yield and purity of the obtained target product are high. The magneto-optical dual-functional material [Gd(dppz)(BTA) 3 ], has excellent magnetic cooling properties and cancer cell fluorescence imaging effects, can image a variety of cancer cells, and a variety of cancer cells survive significantly within the concentration range used in fluorescence imaging studies.

[0022] Preferably, in step (1), the ratio of benzoyltrifluoroacetone (BTA), triethylamine, methanol solvent, and rare earth chloride is 0.3 mmol: 0.72 mmol: 20 mL: 0.1 mmol. The rare earth chloride is added in the form of a methanol solution of the rare earth chloride.

[0023] Preferably, in step (1), the rare earth chloride salt is gadolinium trichloride hexahydrate (GdCl 3 6H 2 O).

[0024] Preferably, in step (2), the molar ratio of dipyrido[3,2-a:2',3'-c]phenazine (dppz) to the rare earth chloride is 1:1.

[0025] Preferably, in step (1), the stirring time is not less than 24 hours. Preferably, in step (2), the stirring time is not less than 24 hours.

[0026] Preferably, in step (3), the room temperature is 15-30° C. More preferably, the room temperature is 25° C. Hereinafter, the present invention will be described in detail with reference to the accompanying drawings.

[0027] Example 1 based on β The method for preparing a magneto-optical dual-functional material of a diketonate rare earth gadolinium complex comprises the following steps: BTA (0.0649 g, 0.3 mmol) and triethylamine (Et 3 N) (0.1 mL, 0.72 mmol) was added to 10 ml of CH 3After the solution was stirred for 10 minutes, 10 ml of CH 3 OH GdCl after dissolution 3 6H 2 O (0.0372 g, 0.1 mmol) salt solution, seal the beaker with plastic wrap and stir for 24 h; then add dppz (0.0282 g, 0.1 mmol) into the beaker, seal and continue stirring for 24 h, filter the obtained mixed system through filter paper into a new beaker; seal the beaker with plastic wrap and let it stand for two weeks to obtain a pure sample [Gd(dppz)(BTA) 3 ] and recorded as complex 3.

[0028] Comparative Example 1 based on β The method for preparing a magneto-optical dual-functional material of a diketonate rare earth gadolinium complex comprises the following steps: BTA (0.0649 g, 0.3 mmol) and triethylamine (Et 3 N) (0.1 mL, 0.72 mmol) was added to 10 ml of CH 3 After the solution was stirred for 10 minutes, 10 ml of CH 3 OH GdCl after dissolution 3 6H 2 O (0.0372 g, 0.1 mmol) salt solution, seal the beaker with plastic wrap and stir for 24 h, then add dpq (0.0232 g, 0.1 mmol) into the beaker, seal and continue stirring for 24 h, filter the obtained mixed system through filter paper into a new beaker; seal the beaker with plastic wrap and let it stand for two weeks to obtain a pure sample [Gd(dpq)(BTA) 3 ] 2 , recorded as complex 1.

[0029] Comparative Example 2 based on β The method for preparing a magneto-optical dual-functional material of a diketonate rare earth gadolinium complex comprises the following steps: DBM (0.0672 g, 0.3 mmol) and triethylamine (Et 3 N) (0.1 mL, 0.72 mmol) was added to 10 ml of CH 3 After the solution was stirred for 10 minutes, 10 ml of CH 3 OH GdCl after dissolution 3 6H 2O (0.0372 g, 0.1 mmol) salt solution, seal the beaker with plastic wrap and stir for 24 h, then add dpq (0.0232 g, 0.1 mmol) into the beaker, seal and continue stirring for 24 h, filter the obtained mixed system through filter paper into a new beaker; and seal the beaker with plastic wrap, and after standing for two weeks, obtain the pure sample [Gd (dpq) (DBM) 3 ] and recorded as complex 2.

[0030] Test Example 1 Example 1 and Comparative Examples 1 to 2 are based on β- Performance test of magneto-optical dual-function materials of diketonate rare earth gadolinium complex (1) Crystal structure characterization and purity testing The reflection data of the complexes obtained in Comparative Examples 1 to 2 and Example 1 were collected by Smart-APEXII CCD X-ray single crystal diffractometer, and the data were analyzed by SHELXS / L-2014 program to obtain the central metal elements. The geometric configuration of Comparative Example 1 is a triangular dodecahedron ( D 2d ), Comparative Example 2 and Example 1 are both tetragonal inverse prism configurations ( D 4d ), the results are shown in Figure 1 .

[0031] The collected images of the complexes of Comparative Examples 1 to 2 and Example 1 were compared with the theoretical X-ray diffraction images by using a D8 X-ray powder diffractometer (PXRD). The obtained diffraction peaks (experimental) and the simulated crystal data (simulated) had good coincidence, indicating that the three examples based on β- The magneto-optical dual-functional material of diketonate rare earth gadolinium complex has high crystallinity and high purity. Figure 2 .

[0032] (2) Magnetic susceptibility temperature dependence test The temperature dependence of magnetic susceptibility of Comparative Examples 1-2 and Example 1 of the present invention was tested using MPMS-XL7 superconducting quantum interference magnetic measuring instrument. The results are shown in Figure 3 . Figure 3 Demonstrated based on β- The magneto-optical dual-function materials of diketonate rare earth gadolinium complexes are compared in Example 1 (a), Example 2 (b) and Example 1 (c). χ M T The value changes with temperature. In the 1000 Oe DC field and 2-300 K temperature range, the χ M TThe values ​​are 17.14, 7.88 and 8.00 cm³ K mol, respectively. -1 , which is consistent with the expected values ​​(15.76, 7.88, and 7.88 cm³ K mol -1 ) are consistent. (3) Relationship between magnetization intensity and applied magnetic field The magnetization measurement of the complexes obtained in Comparative Examples 1 and 2 and Example 1 of the present invention was studied at a temperature of 2 to 10 K in the range of 0 to 7 T. The results are shown in FIG. Figure 4 The MH curves of Comparative Examples 1-2 and Example 1 of the present invention show that the magnetization intensity of complexes 1-3 increases rapidly with the increase of magnetic field under 0-2 T low magnetic field, while under 2-7 T high magnetic field, the increase rate of magnetization intensity gradually slows down and tends to be stable. Under 2 K and 7 T conditions, the magnetization intensity M of Comparative Example 1 (a) is 15.17 Nβ, which is similar to the two Gd³⁺( g =2, S = 7 / 2) is close to the theoretical value of 14 Nβ; the M of Comparative Example 2 (b) and Example 1 (c) are 6.99 and 7.04 Nβ, respectively, which is close to the theoretical value of Gd³⁺( g = 1, S = 7 / 2) is consistent with the theoretical value 7 Nβ. (4) χM -1 and T Relationship and Curie-Weiss fitting Between 2-300k, the preparations obtained in Comparative Examples 1-2 and Example 1 of the present invention were subjected to χ M -1 right T The linear fitting results are shown in Figure 5 The obtained complex has magneto-optical dual-functional materials χ M -1 and T The relationship diagram and Curie-Weiss fit are good, indicating that the three complexes obey the Curie-Weiss law, among which, C 1 = 16.83 cm 3 K mol -1 , θ 1 = 0.41 k; C 2 = 7.79 cm 3 K mol -1 , θ 2 = 0.33 k; C 3 = 7.89 cm 3K mol -1 , θ 3 = 0.34 k, indicating that ferromagnetic coupling exists in these three Gd 3+ By applying Maxwell's relations to the experimentally measured magnetization data, the magnetic entropy change ( ). Under the conditions of temperature of 2 K and magnetic field of 7 T, the magnetic entropy change reaches the maximum value, which is 16.63 J·kg -1 ·K -1 , 15.19 J·kg -1 ·K -1 and 15.24 J·kg -1 ·K -1 Magnetic entropy change of Gd-based complexes ( ) increases with decreasing temperature, showing low-temperature magnetic refrigeration characteristics and has practical application potential. From the above test, it can be seen that the entropy changes of Comparative Examples 1-2 and Example 1 under various magnetic fields and temperatures reach the maximum when T = 2 K and H = 7 T. 16.63 J Kg respectively -1 K -1 , 15.19 J Kg -1 K -1 , 15.24J Kg -1 K -1 , exhibits low-temperature magnetic refrigeration properties, and is expected to become a magnetic refrigeration material used in practice. Application Example 1 based on β- Cancer cell imaging experiment of diketonate rare earth gadolinium complex magneto-optical dual-functional material The results of the comparative examples 1 and 2 and the embodiment 1 of the present invention were tested by using an F-7000 fluorescence spectrophotometer. β- The fluorescence emission spectrum of the diketonate rare earth gadolinium complex is shown in the figure below. Figure 6 , Comparative Examples 1 to 2 ( Figure 6 a) has a fluorescence emission wavelength of 612-614 nm; Example 1 ( Figure 6 The fluorescence emission wavelength of complex 1, complex 2 and complex 3 is red.

[0033] Comparative Examples 1-2 and Example 1 of the present invention were prepared into turbid suspensions (concentration 20 μmol mL -1 ), and several cancer cells were co-cultured with the suspension of complex 1, complex 2, and complex 3 for 4 h, and then the fluorescence color development of the complexes was studied using a Zeiss LSM900 laser confocal microscope. The results are shown in Figure 7, wherein Comparative Examples 1 to 2 ( Figure 7 a, 7b) can be used for fluorescence imaging in HeLa cells, Example 1 ( Figure 7 In c), fluorescence imaging can be performed on six common cancer cells (HeLa, MCF-7, A549, Scg7901, Hep-G2, and DU-145). The three cells were observed to be internalized, and most of them entered the cytoplasm, and only a small part entered the nucleus. From the above application examples, it can be obtained that comparative example 1 [Gd (dpq) (BTA) 3 ] 2 、Comparative Example 2 [Gd (dpq) (DBM) 3 ] can only show highly selective imaging ability for HeLa cells, while the magneto-optical dual-functional material [Gd(dppz)(BTA) 3 ], which is capable of broad-spectrum imaging of six common cancer cells (HeLa, MCF-7, A549, Scg7901, Hep-G2, and DU-145), demonstrates potential for broad biomedical applications and has achieved unexpected technical results. Compared with the ligand 2,3-bis(2-pyridyl)quinoxaline (dpq), the complex formed by the dipyrido[3,2-a:2',3'-c]phenazine (dppz) ligand has higher coordination stability and more complex structure. In addition, dppz has good cell membrane affinity and can efficiently enter cells through direct penetration or endocytosis. It also exhibits low toxicity, good biocompatibility and certain targeting. It is suitable for drug delivery and imaging of various cancer cells. It can be widely used in cancer diagnosis, targeted therapy and magnetic materials, and has important scientific research value and commercial prospects.

[0034] The above are only preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. Based on β- Application of magneto-optical dual-functional materials of diketonate rare earth gadolinium complexes in fluorescence imaging, characterized in that: The expression of the magneto-optical dual-functional material is [Gd(dppz)(BTA)3], wherein BTA is benzoyltrifluoroacetone, dppz is dipyrido[3,2-a:2',3'-c]phenazine, and Gd is gadolinium.

2. The use according to claim 1, characterized in that: The magneto-optical dual-function material performs specific imaging on six types of cancer cells: human cervical cancer cells, human breast cancer cells, human non-small cell lung cancer cells, human gastric cancer cells, liver cancer cell line G2 type and human prostate cancer cells.

3. The use according to claim 1, characterized in that: The concentration of the magneto-optical dual-functional material in fluorescence imaging is 10-20 μmol mL -1 .

4. The use according to claim 1, characterized in that: The fluorescence emission wavelength of the magneto-optical dual-function material in fluorescence imaging is 618-621 nm.

5. The use according to claim 1, characterized in that: The magneto-optical dual-function material is prepared by the following steps: (1) adding benzoyltrifluoroacetone and triethylamine to a methanol solvent, stirring until dissolved, adding gadolinium trichloride hexahydrate, stirring to obtain a mixed solution; (2) adding dipyrido[3,2-a:2',3'-c]phenazine to the mixed solution, stirring in a semi-closed beaker at room temperature, and filtering to obtain a filtrate; (3) The filtrate is allowed to stand at room temperature for evaporation to obtain the target product.

6. The use according to claim 5, characterized in that: In step (1), the usage ratio of benzoyltrifluoroacetone, triethylamine, methanol solvent and gadolinium trichloride hexahydrate is 0.3 mmol:0.72 mmol:20 mL:0.1 mmol.

7. The use according to claim 5, characterized in that: In step (2), the molar ratio of dipyrido[3,2-a:2',3'-c]phenazine to gadolinium trichloride hexahydrate is 1:

1.

8. The use according to claim 5, characterized in that: In step (1), the stirring time is not less than 24 hours; In step (2), the stirring time is not less than 24 hours.

9. The use according to claim 5, characterized in that: In step (3), the room temperature is 15-30°C.

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