A metal-organic framework and rare earth nanocrystal composite material with thermal enhanced fluorescence temperature measurement and a preparation method and application thereof

Abstract

The application discloses a metal-organic framework and rare earth nanocrystal composite material with thermal enhancement type fluorescence temperature measurement and a preparation method and application thereof. The metal-organic framework and rare earth nanocrystal composite material prepared by the application has a nanometer level size, can enter cells, has excellent biocompatibility and can realize practical application in vivo. Under the pumping of infrared laser with strong biological penetration capacity, the composite material generates fluorescence emission through the up-conversion process of the rare earth nanocrystal, and the up-conversion fluorescence intensity of the composite material shows abnormal thermal enhancement performance with the increase of temperature. The fluorescence thermal enhancement performance of the composite material is not affected by the complex physiological environment in cells, and more accurate temperature measurement can be realized. The maximum relative sensitivity of the temperature sensing of the composite material is 1.12%·K ‑1 , and the thermal enhancement type temperature sensing can be completely applied to the physiological temperature in cells.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of nanomaterials, and particularly relates to a metal-organic framework and rare earth nanocrystal composite material with thermal enhanced fluorescence temperature measurement, a preparation method and application thereof. The composite material has abnormal thermal enhanced fluorescence performance and excellent biocompatibility, and can be applied to intracellular temperature sensing. BACKGROUND

[0002] Temperature plays a key role in many physical, chemical and biological processes. Therefore, achieving fast and accurate temperature measurement is of great importance to industrial production, medical diagnosis and treatment, and scientific research, and has been the goal pursued by researchers in various fields. Medical research shows that certain pathological cells exhibit higher temperatures than normal cells due to faster metabolism. Temperature changes related to physiological processes in the body are ~1℃, and temperature change monitoring with a thermal resolution of 0.1℃ can accurately prevent epilepsy or acute cardiovascular accidents. Photothermal therapy, which converts light energy into heat energy to kill cancer cells, induces irreversible damage to cancer cells through local temperature rise in tumor tissue, achieving optical minimally invasive treatment. Therefore, accurate measurement of temperature in physiological processes will help biomedical diagnosis and treatment, and has very important significance. However, conventional temperature measurement tools such as thermocouples, thermistors and metal thermometers are not suitable for non-contact temperature measurement at the micro-nano level in the biological field. Fluorescence temperature sensing has become one of the most promising methods in the field of physiological temperature measurement due to its non-invasiveness, real-time readout and high thermal resolution.

[0003] At present, a series of temperature-responsive materials have been developed and used for fluorescence temperature sensing, including small-molecule organic compounds, rare earth complexes, quantum dots, etc. However, most temperature probes are subject to various limitations in actual use, such as being susceptible to environmental interference and having low signal-to-noise ratio. In addition, most thermometers are excited by ultraviolet-visible light with poor tissue penetration ability, which will be disturbed by endogenous background fluorescence during detection, and will also cause photobleaching phenomenon, which requires signal correction to accurately perceive temperature. Rare earth-doped upconversion nanocrystals exhibit high anti-photobleaching, non-flashing, non-crosstalk emission and autofluorescence characteristics compared with traditional optical thermometers, and therefore have attracted widespread attention in temperature sensing, biological probes and super-resolution imaging, etc. Rare earth ions have unique 4f electron shell and "ladder-like" excitation state energy levels, and can realize upconversion emission of high-energy photons through continuous absorption of low-energy photons. In addition, the low phonon energy (~355cm -1In the matrix of fluoride nanocrystals (fluoride yttrium sodium, fluoride gadolinium sodium, etc.) of the rare earth ions, near-infrared continuous laser can excite rare earth ions to realize photon upconversion process, which shows stronger tissue penetration ability, providing a broad prospect for the application of rare earth nanocrystals in fluorescence temperature measurement. The upconversion fluorescence thermal enhancement performance shows incomparable advantages in the field of fluorescence temperature sensing compared with conventional thermal quenching type material system, because the actual application of biological nanometer thermometer requires high sensitivity, high brightness and anti-interference ability in complex environment, which can improve the imaging contrast and temperature resolution. However, the use of surface hydrophobic ligand strictly hinders the effective dispersion of nanocrystals in the physiological environment, which strictly limits the practical application of nanocrystals as optical thermometer in the biological field. SUMMARY

[0004] Based on the deficiencies in the prior art, the present application provides a metal-organic framework and rare earth nanocrystal composite material with thermal enhancement type fluorescence temperature measurement, a preparation method and application thereof, and utilizes the interaction between the composite materials to realize the thermal enhancement type fluorescence temperature measurement in the physiological temperature range.

[0005] In one aspect, the present application provides a metal-organic framework and rare earth nanocrystal composite material with thermal enhancement type fluorescence temperature measurement, which is composed of metal-organic framework and rare earth nanocrystal, and its expression is [M(L) x (G) y ]@R.

[0006] Wherein M is a metal ion in the metal-organic framework, which is at least one of Zr, Cu, Cr, Co, Cd and Fe;

[0007] L is an organic ligand in the metal-organic framework, which is at least one of terephthalic acid, 2-bromoterephthalic acid, 2-hydroxyterephthalic acid, 2-chloroterephthalic acid, 2-methylterephthalic acid, 2,5-pyridinedicarboxylic acid, 2-nitroterephthalic acid, 2-aminoterephthalic acid, 2,5-dihydroxyterephthalic acid, 2,5-dichloroterephthalic acid, 2,5-dimethylterephthalic acid and 2,5-pyrazinedicarboxylic acid, x=l~3; G represents a solvent molecule coordinated with the metal ion or in the crystal channel, which is at least one of water, acetic acid, N,N-dimethylformamide, N,N-dimethylacetamide and N,N-diethylformamide, y=0~60; R represents a rare earth nanocrystal NaLnF4, wherein Ln is a rare earth ion, which is at least one of Yb, Y, Er and Gd.

[0008] Preferably, the Ln is Y, Yb and Er, wherein the molar ratio of Y, Yb and Er is 70:29:1. In the embodiment of the present application, rare earth chloride salt LnCl3·6H2O (Ln 3+ = 70% Y 3+ / 29% Yb3+ / 1%Er 3+ The metal-organic framework material prepared and the rare earth nanocrystal (UiO-66@NaYF4:Yb / Er nanocrystal) exhibit obvious up-conversion fluorescence under excitation of infrared laser with biological penetration ability, and the survival rate of Hela cells remains at about 100% after the composite material is co-cultured with the Hela cells, which proves excellent biocompatibility of the material.

[0009] In another aspect, the application further provides a preparation method of the metal-organic framework and rare earth nanocrystal composite material, comprising the following steps:

[0010] (1) dissolving and mixing a rare earth chloride salt with oleic acid and octadecene, and heating and reacting with ammonium fluoride and sodium hydroxide under protection of inert gas, and then treating the obtained product with acid, and centrifuging to collect the surface-acidized rare earth nanocrystal;

[0011] (2) dissolving and uniformly mixing the organic ligand and the surface-acidized nanocrystal in a solvent, dissolving a metal salt in the solvent, and then adding and heating to react the two mixed solutions to obtain the metal-organic framework and rare earth nanocrystal composite material.

[0012] Optionally, in step (1), the rare earth chloride salt is ytterbium chloride, yttrium chloride, gadolinium chloride or erbium chloride; and in step (2), the metal salt is zirconium chloride, copper nitrate, chromium nitrate, cobalt nitrate, cadmium nitrate or zinc nitrate.

[0013] In the application, the carboxylic acid-containing organic ligand used for synthesizing the metal-organic framework is (a) terephthalic acid, (b) 2-bromoterephthalic acid, (c) 2-hydroxyterephthalic acid, (d) 2-chloroterephthalic acid, (e) 2-methylterephthalic acid, (f) 2,5-pyridinedicarboxylic acid, (g) 2-nitroterephthalic acid, (h) 2-aminoterephthalic acid, (i) 2,5-dihydroxyterephthalic acid, (j) 2,5-dichloroterephthalic acid, (k) 2,5-dimethylterephthalic acid, and (l) 2,5-pyrazinedicarboxylic acid. The structural formulas are as follows:

[0014]

[0015] The metal ions form coordination bonds with carboxylate in the carboxyl group of the organic ligand, and the coordination self-assembly forms the metal-organic framework material. The transition metal ions and the metal zirconium both have strong coordination with the linear organic ligand with carboxylic acid, so in the present application, the metal salt of the metal-organic framework material selects zirconium chloride as an example, but is not limited thereto, other metal salts such as copper nitrate, chromium nitrate, cobalt nitrate, cadmium nitrate or zinc nitrate can also achieve the above-mentioned effects; the organic ligand selects terephthalic acid and 2-amino terephthalic acid as an example, but is not limited thereto, the above-mentioned other linear organic ligands with carboxylic acid can also achieve the above-mentioned effects.

[0016] In the present application, the molar ratio of the metal ions in the metal salt used for synthesizing the metal-organic framework material and the organic ligand is 1:2-5.

[0017] In some embodiments of the present application, in step (3), when the metal-organic framework and the rare earth nanocrystal composite material are synthesized, the molar ratio of the metal salt and the rare earth nanocrystal reacting in the solution is 5:1-3.

[0018] In some embodiments of the present application, in step (1), when the rare earth chloride salt is dissolved and mixed with oleic acid and octadecene, the temperature is increased to 160℃ under inert gas protection, and the molten rare earth chloride salt is kept for 30 minutes;

[0019] When heated, first slowly heat to 100℃ at a heating rate of 5℃ / min, keep for 30 minutes, then heat to 290℃-300℃ at a rate of 10-15℃ / min, keep for 2-2.5 hours.

[0020] In step (2), the heating reaction condition is: heating at 100℃-120℃ for 1-3 days.

[0021] In another aspect, the present application also provides an application of the metal-organic framework and the rare earth nanocrystal composite material, or the metal-organic framework and the rare earth nanocrystal composite material prepared by the preparation method in biological temperature sensing.

[0022] Specifically, in the application, the physiological temperature of the organism is determined by the up-conversion fluorescence intensity of the metal-organic framework and the rare earth nanocrystal composite material.

[0023] Finally, the present application also provides an application of the metal-organic framework and the rare earth nanocrystal composite material, or the metal-organic framework and the rare earth nanocrystal composite material prepared by the preparation method in preparing an optical thermometer.

[0024] The metal-organic framework and rare earth nanocrystal composite material has a thermal enhancement response performance of upconversion fluorescence intensity to temperature and excellent biocompatibility, and is used for thermal enhancement type fluorescence sensing of intracellular physiological temperature.

[0025] The present application has the following specific beneficial effects:

[0026] 1. The metal-organic framework and rare earth nanocrystal composite material prepared by the present application has a nanoscale size and can enter cells, and also has excellent biocompatibility, and can realize practical application in vivo.

[0027] 2. The metal-organic framework and rare earth nanocrystal composite material prepared by the present application generates fluorescence emission through the upconversion process of the rare earth nanocrystal under the pumping of infrared laser with strong biological penetration ability, and ensures the application ability of the composite material in the biological field.

[0028] 3. The metal-organic framework and rare earth nanocrystal composite material of the present application exhibits abnormal thermal enhancement performance with the increase of temperature, and this characteristic can provide a new measurement strategy for temperature sensing.

[0029] 4. The fluorescence thermal enhancement performance of the metal-organic framework and rare earth nanocrystal composite material of the present application is not affected by the complex physiological environment in cells, and can realize more accurate temperature measurement.

[0030] 5. The metal-organic framework and rare earth nanocrystal composite material of the present application, the maximum relative sensitivity of which for temperature sensing is 1.12%·K -1 , can be completely applied to thermal enhancement type temperature sensing of intracellular physiological temperature. BRIEF DESCRIPTION OF DRAWINGS

[0031] Figure 1 is the upconversion emission intensity and relative sensitivity of the metal-organic framework and rare earth nanocrystal composite material of the present application with temperature change;

[0032] Figure 2 is the fluorescence enhancement type spectrum of the metal-organic framework and rare earth nanocrystal composite material of the present application under 980nm laser pumping with temperature change;

[0033] Figure 3 is the scanning electron microscope images of the metal-organic framework and rare earth nanocrystal composite material of the present application before (a) and after (b) standing in water environment for one year;

[0034] Figure 4 is the cell survival rate of the metal-organic framework and rare earth nanocrystal composite material of the present application after co-culturing with Hela cells.

[0035] Figure 5The metal-organic framework and rare earth nanocrystal composite material of the present application is used in the related equipment and optical path of Hela cell monitoring temperature. DETAILED DESCRIPTION

[0036] Example 1

[0037] The UiO-66-NH2@NaYF4:Yb / Er composite material is synthesized by using erbium chloride, ytterbium chloride, yttrium chloride, zirconium chloride and 2-amino terephthalic acid in a step-by-step manner, and the specific synthesis route is as follows:

[0038] 1) First, 2 mL of a methanol solution of a rare earth chloride salt LnCl3·6H2O (Ln 3+ = 70% Y 3+ / 29% Yb 3+ / 1% Er 3+ , 1 mmol) is added to a three-necked round-bottom flask together with 6 mL of oleic acid and 15 mL of octadecene. Subsequently, the mixture is heated to 160°C under a nitrogen atmosphere and kept for 30 minutes, allowing it to form a transparent light yellow solution. After cooling to room temperature, 13 mL of a methanol solution containing 2.5 mmol of sodium hydroxide and 4 mmol of ammonium fluoride is added under vigorous stirring. The solution is then slowly heated to 100°C at a temperature increase rate of 5°C / min under a nitrogen atmosphere, kept for 30 minutes, and then heated to 300°C at a rate of 15°C / min, and kept at this temperature for 1.5 hours. Finally, the product is precipitated by adding ethanol, and the nanocrystals are collected by centrifugation and then dispersed in ethanol. Next, dilute hydrochloric acid (1M) is added, and the mixture is subjected to ultrasonic treatment to remove the surface oleic acid. Finally, the NaYF4:Yb / Er nanocrystals are collected by centrifugation.

[0039] 2) 2-amino terephthalic acid (0.04 mmol) and NaYF4:Yb / Er nanocrystals (0.06 mmol) are added to N,N-dimethylformamide (9 mL), and the mixture is subjected to ultrasonic treatment for 35 minutes. Zirconium chloride (0.1 mmol) is dissolved in a mixture of N,N-dimethylformamide (1 mL) and acetic acid (1.5 mL) and subjected to ultrasonic treatment for 10 minutes. The two are mixed to obtain a reaction solution, which is transferred to a polytetrafluoroethylene-lined autoclave and placed in an oven (120°C) for 24 hours. Finally, the metal-organic framework material and rare earth nanocrystals (UiO-66-NH2@NaYF4:Yb / Er nanocrystals) are obtained by filtration and washed with N,N-dimethylformamide and anhydrous methanol.

[0040] Under the excitation of a 980 nm laser, the peak value of the upconversion fluorescence emission spectrum obtained by the metal-organic framework and rare earth nanocrystal composite material (UiO-66-NH2@NaYF4:Yb / Er) increases with the increase of temperature Figure 1), can be fitted using the following formula:

[0041]

[0042] Among them I 526 is the fluorescence intensity at 526 nm; I 537 is the fluorescence intensity at 537 nm. C is a constant, T is the absolute temperature, and ΔE is 2 H 11 / 2 and 4 S 3 / 2 The energy difference between excited states, K B is the Boltzmann constant. Then, the relative sensitivity (S r ):

[0043]

[0044] like Figure 1 As shown, the maximum relative sensitivity of the composite material is calculated to be 1.12%. -1 , which can be fully applied to biosensing.

[0045] The metal-organic framework and rare earth nanocrystal composite material (UiO-66-NH2@NaYF4: Yb / Er) exhibits obvious upconversion fluorescence under the excitation of infrared laser with biological penetration ability ( Figure 2 ), thus ensuring the application of metal-organic framework and rare earth nanocrystal composite materials in the biological field.

[0046] The overall size of the metal-organic framework and rare earth nanocrystal composite material (UiO-66-NH2 / NaYF4: Yb / Er) remains at the nanometer level, and the composite structure can remain stable in an aqueous environment ( Figure 3 ), ensuring that the material can effectively enter the cells and thus monitor the physiological temperature changes within the cells.

[0047] In order to verify the practical application ability of this material in physiological environment, we studied the biocompatibility of metal-organic framework and rare earth nanocrystal composite material (UiO-66-NH2@NaYF4: Yb / Er). -1 The cell survival rate remained above 90% ( Figure 4 ), indicating that the material has good biocompatibility and is expected to be used in the biomedical field.

[0048] In Hela cells, the metal-organic framework and rare earth nanocrystal composite material of the present invention is used in Hela cells to monitor the temperature of the related equipment and optical path. Figure 5 As shown. When 0.04mmol composite material and 150mg·mL -1After co-culturing with Hela cells, the sample was placed in a temperature detection sample stage, temperature controlled by a temperature controller, and the sample was excited by a 980 nm laser. The emitted photons were collected by a fiber-optic spectrometer and the feedback was fluorescence signal. The results showed that the upconversion fluorescence intensity of the metal-organic framework and rare earth nanocrystal composite material (UiO-66-NH2 / NaYF4:Yb / Er) was enhanced with the increase of temperature, showing abnormal thermal enhancement type fluorescence performance Figure 2 ), which provided a more accurate temperature measurement technology for intracellular physiological temperature sensing.

[0049] Example 2

[0050] UiO-66 / NaGdF4 composite material was synthesized by using erbium chloride, ytterbium chloride, gadolinium chloride, zirconium chloride and terephthalic acid in a step-by-step manner, and the specific synthesis route was as follows:

[0051] 1) First, 2 mL of methanol solution of rare earth chloride salt LnCl3·6H2O (Ln 3+ = 70% Gd 3+ / 29% Yb 3+ / 1% Er 3+ , 1 mmol) was added to a three-necked round-bottom flask together with 6 mL of oleic acid and 15 mL of octadecene. Subsequently, the mixture was heated to 160°C under a nitrogen atmosphere and maintained for 30 minutes, so as to form a transparent light yellow solution. After cooling to room temperature, 13 mL of methanol solution containing 2.5 mmol of sodium hydroxide and 4 mmol of ammonium fluoride was added under vigorous stirring. The solution was slowly heated to 100°C at a temperature increasing rate of 5°C / min under a nitrogen atmosphere, maintained for 30 minutes, and then heated to 295°C at a rate of 10°C / min, and maintained at this temperature for 1.5 hours. Finally, the product was precipitated by adding ethanol, and the nanocrystals were collected by centrifugation and dispersed in ethanol. Then, dilute hydrochloric acid (1M) was added, and the mixture was ultrasonically treated to remove the surface oleic acid. Finally, the NaGdF4:Yb / Er nanocrystals were collected by centrifugation.

[0052] 2) Terephthalic acid (0.04 mmol) and NaGdF4:Yb / Er nanocrystals (0.05 mmol) were added to N,N-dimethylformamide (9 mL), and the mixture was ultrasonically treated for 45 minutes. Zirconium chloride (0.1 mmol) was dissolved in a mixture of N,N-dimethylformamide (1 mL) and acetic acid (1 mL) and ultrasonically treated for 10 minutes. The mixture of the two was mixed to obtain a reaction solution, which was transferred to a polytetrafluoroethylene-lined autoclave and placed in an oven (120°C) for 24 hours. Finally, the metal-organic framework material and rare earth nanocrystals (UiO-66@NaGdF4:Yb / Er nanocrystals) were obtained by filtration and washed with N,N-dimethylformamide and anhydrous methanol.

[0053] The metal organic framework and rare earth nanocrystal composite material (UiO-66 / NaGdF4:Yb / Er) shows obvious upconversion fluorescence under the excitation of infrared laser with biological penetration ability, thereby ensuring the application prospect of the composite material in the biological field.

[0054] The metal organic framework and rare earth nanocrystal composite material (UiO-66 / NaGdF4:Yb / Er) has a nanoscale overall size, and the composite structure can remain stable in a water environment, thereby ensuring that the material can effectively enter cells to realize the monitoring of intracellular physiological temperature changes. After the composite material (10-150 μg·mL -1 ) is co-cultured with Hela cells, the survival rate of the Hela cells still remains above 90%, proving the excellent biocompatibility of the material.

[0055] In the Hela cells, the upconversion fluorescence intensity of the metal organic framework and rare earth nanocrystal composite material (UiO-66 / NaGdF4:Yb / Er) is enhanced with the increase of temperature, thereby showing abnormal thermal enhancement type fluorescence performance and providing a more accurate temperature measurement technology for the sensing of intracellular physiological temperature.

[0056] The above specific embodiments are used to explain and illustrate the present application, but these examples do not limit the protection scope of the present application, and various modifications or changes made by those skilled in the art on the basis of the technical solutions of the present application without creative labor are still within the protection scope of the present application.

Claims

1. A metal-organic framework and rare earth nanocrystal composite material with thermally enhanced fluorescence thermometry, characterized in that, The composite material consists of a metal-organic framework and rare earth nanocrystals, expressed as [M(L) x (G) y ]@R, M is a metal ion in the metal-organic framework, and is at least one of Zr, Cu, Cr, Co, Cd and Fe; L is an organic ligand in the metal-organic framework, and is at least one of terephthalic acid, 2-bromoterephthalic acid, 2-hydroxyterephthalic acid, 2-chloroterephthalic acid, 2-methylterephthalic acid, 2,5-pyridinedicarboxylic acid, 2-nitroterephthalic acid, 2-aminoterephthalic acid, 2,5-dihydroxyterephthalic acid, 2,5-dichloroterephthalic acid, 2,5-dimethylterephthalic acid and 2,5-pyrazinedicarboxylic acid, x = 1-3; G represents a solvent molecule coordinated with the metal ion or in the crystal pore, and is at least one of water, acetic acid, N,N-dimethylformamide, N,N-dimethylacetamide and N,N-diethylformamide, y = 0-60; R represents a rare earth nanocrystal NaLnF4, wherein Ln is a rare earth ion, and the Ln is Y, Yb and Er, and the molar ratio of Y, Yb and Er is 70:29:

1.

2. A method of preparing the metal-organic framework and rare earth nanocrystal composite material of claim 1, characterized in that, The method comprises the following steps: (1) mixing a rare earth chloride salt with oleic acid and octadecene, and heating and reacting under inert gas protection with ammonium fluoride and sodium hydroxide, treating the obtained product with acid, and centrifuging to collect a surface-acidized rare earth nanocrystal; (2) dissolving the organic ligand and the surface-acidized nanocrystal in a solvent and mixing uniformly, dissolving a metal salt in the solvent, and then adding and heating to react the two mixed solutions to obtain the metal-organic framework and rare earth nanocrystal composite material.

3. The method for preparing the metal-organic framework and rare earth nanocrystal composite material according to claim 2, characterized in that: In step (1), the rare earth chloride salt is ytterbium chloride, yttrium chloride and erbium chloride; In step (2), the metal salt is zirconium chloride, copper nitrate, chromium nitrate, cobalt nitrate, cadmium nitrate or zinc nitrate.

4. The method for preparing the metal-organic framework and rare earth nanocrystal composite material according to claim 2, characterized in that: In step (3), when the metal-organic framework and rare earth nanocrystal composite material is synthesized, the molar ratio of the metal salt and the rare earth nanocrystal in the solution is 5:1-3.

5. The method for preparing the metal-organic framework and rare earth nanocrystal composite material according to claim 2, characterized in that: In step (1), when the rare earth chloride salt is dissolved and mixed with oleic acid and octadecene, the temperature is raised to 160°C under inert gas protection, and the molten rare earth chloride salt is kept for 30 minutes; During the heating reaction, the temperature is first raised to 100°C at a rate of 5°C / min, kept for 30 minutes, and then raised to 290-300°C at a rate of 10-15°C / min, kept for 2-2.5 hours.

6. The method for preparing the metal-organic framework and rare earth nanocrystal composite material according to claim 2, characterized in that: In step (2), the heating reaction is carried out at 100-120°C for 1-3 days.

7. Use of the metal-organic framework and rare earth nanocrystal composite material of claim 1, or the metal-organic framework and rare earth nanocrystal composite material prepared by the preparation method of any one of claims 2-6, in biological temperature sensing.

8. Use according to claim 7, characterized in that, In use, the physiological temperature of the organism is determined by the upconversion fluorescence intensity of the metal-organic framework and rare earth nanocrystal composite material.

9. Use of the metal-organic framework and rare earth nanocrystal composite material of claim 1, or the metal-organic framework and rare earth nanocrystal composite material prepared by the preparation method of any one of claims 2-6, in preparing an optical thermometer.

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