Nano material as well as preparation method and application thereof

Abstract

The invention relates to the technical field of luminescent materials, in particular to a nano material and a preparation method and application thereof. The invention provides a nanometer material, the nanometer material is NaYbF4: Er (at) NaYF4 (at) Cy7.5 (at) DPPC, the nanometer material comprises a core-shell structure with the component of NaYbF4: Er (at) NaYF4, and the core-shell structure is modified with DPPC and compounded with Cy7.5. When the nano material provided by the invention is used for an optical nano indicator or sensor for detecting the membrane potential under an NIR-IIb window, the sensor generates emitted light (1500-1700nm) under the NIR-IIb window under the excitation of 808nm near-infrared incident light. By utilizing the nano sensor, the membrane potential change of cells marked by the sensor can be visually detected by means of optical imaging.

Description

Technical Field

[0001] The present invention relates to the technical field of luminescent materials, and in particular to a nanomaterial, a preparation method and an application thereof. Background Art

[0002] Electrophysiological activity is a key indicator for studying neural processes and neurological diseases (such as temporal lobe epilepsy). Its essence is the generation and propagation of action potentials driven by ion flow across cell membranes. Although implantable electrodes, the "gold standard," can directly record electrophysiological signals from brain tissue, this approach is inevitably invasive, and the gradual formation of scar tissue around the electrodes eventually isolates them from the neural cells, rendering them ineffective. Despite the recent emergence of more biocompatible flexible electrodes, the wired connection between the electrodes and the detection equipment still interferes with the normal behavior of experimental animals. In contrast, optical imaging technology offers unique advantages in monitoring membrane potential signals due to its high spatiotemporal resolution and low invasiveness. In recent years, a variety of molecular or nanosensors for membrane potential detection have emerged. Traditional voltage-sensitive dyes (VSDs), including di-2-ANEPPS, di-4-ANEPPS, and RH237, respond to fluctuations in the membrane potential of excitable cells through the molecular Stark effect. However, when anchored to the cell membrane, the fluorescence response of VSDs is very limited due to spectral shifts. In order to amplify the fluorescence changes induced by transmembrane voltage fluctuations, related studies have begun to use electron transfer (ET) and Resonance Energy Transfer (FRET) processes. These mechanisms enable voltage-sensitive signals to be generated not only at the molecular scale but also at the nanoscale, allowing the resulting fluorescence signal to reflect changes in membrane potential. However, in excitable cells, there is a correlation between intracellular calcium influx and action potentials. The energy transfer mechanisms involved in both ET and FRET are affected by fluctuations in intracellular calcium concentration, further complicating the relationship between fluorescence signals and voltage fluctuations. Furthermore, the fluorescence response of currently reported indicators / sensors is primarily confined to the visible spectrum. The scattering and absorption of the visible spectrum by biological tissues severely impacts the accuracy of optical imaging techniques for membrane potential detection. In contrast, luminescence in the near-infrared II (NIR-II) spectral range (1000-1700 nm), particularly in the NIR-IIb window (1500-1700 nm), offers excellent tissue penetration, low scattering, and minimal in vivo side effects. Therefore, the development of optical nanoindicators / sensors for membrane potential detection in the NIR-IIb window will be an important future development direction in the field of optical membrane potential detection. Summary of the Invention

[0003] In view of the technical problem mentioned above that when the indicator in the prior art is anchored on the cell membrane, the fluorescence response of the voltage-sensitive dye is very limited due to spectral shift and is easily affected by the fluctuation of ion flow on both sides of the cell membrane, the present invention provides a nanomaterial and its preparation method and application.

[0004] To achieve the above-mentioned and other related purposes, the first aspect of the present invention provides a nanomaterial, which is NaYbF4:Er@NaYF4@Cy7.5@DPPC. The nanomaterial includes a core-shell structure composed of NaYbF4:Er@NaYF4, and the core-shell structure is modified with DPPC and compounded with Cy7.5.

[0005] Preferably, the NaYbF4:Er@NaYF4 in A1) contains Er in a molar fraction of 1 to 20%.

[0006] Preferably, the thickness of the shell layer close to the core in A2) is 1.4-4.1 nm, for example, 1.4-2 nm, 2-2.5 nm, 2.5-3 nm, 3-3.5 nm, 3.5-4.1 nm, etc.

[0007] Preferably, the diameter of the core in A3) is 7-12 nm, for example, 7-8 nm, 8-9 nm, 9-10 nm, 10-11 nm, 11-12 nm, etc.

[0008] Preferably, the diameter of the nanoparticles in A4) is 15-30 nm, for example, 15-17 nm, 17-19 nm, 19-21 nm, 21-23 nm, 23-25 nm, 25-27 nm, 27-29 nm, 29-30 nm, etc.

[0009] Preferably, A5) the mass ratio of NaYbF4:Er@NaYF4 to the nanomaterial is (1:1.5)-(1:5).

[0010] Preferably, A6) the mass molar ratio of the NaYbF4:Er@NaYF4 to the Cy7.5 is 20 mg:(2.5-20 nmol);

[0011] Preferably, A7) the mass ratio of DPPC to the nanoparticles is (0.5:1.5)-(4:5).

[0012] A second aspect of the present invention provides a method for preparing a nanomaterial, comprising the following steps:

[0013] 1) a rare earth trifluoroacetate, sodium trifluoroacetate, oleic acid, oleylamine, and 1-octadecene are first mixed, heated, and washed, and a product of the first wash is second mixed with 1-octadecene, oleic acid, sodium trifluoroacetate, and yttrium trifluoroacetate, heated, and washed to obtain NaYbF4:Er@NaYF4;

[0014] 2) The NaYbF4:Er@NaYF4 obtained in step 1) is mixed with phosphatidylcholine and dichloromethane for a third time to obtain NaYbF4:Er@NaYF4 modified with phosphatidylcholine;

[0015] 3) The phosphatidylcholine-modified NaYbF4:Er@NaYF4 obtained in step 2) is mixed with the Cy7.5 solution for the fourth time to obtain NaYbF4:Er@NaYF4@Cy7.5@DPPC (Cy7.5-ErNP).

[0016] Preferably, in step 11), the rare earth trifluoroacetate is selected from one or more of ytterbium trifluoroacetate and erbium trifluoroacetate.

[0017] Preferably, in step 12) of step 1), Yb 3+ The molar fraction of Er is 0.99~0.80, 3+ The mole fraction is 0.01 to 0.20.

[0018] Preferably, in step 13) before the first washing, the molar ratio of the rare earth trifluoroacetate to sodium trifluoroacetate is 1:1.

[0019] Preferably, in step 14) before the first washing, the molar volume ratio of the rare earth trifluoroacetate to oleic acid is 1 mmol:3.2 mL.

[0020] Preferably, in step 15) before the first washing, the molar volume of the rare earth trifluoroacetate and oleylamine is 1 mmol:3.3 mL.

[0021] Preferably, in step 16) before the first washing, the molar ratio of the rare earth trifluoroacetate to 1-octadecene is 1 mmol:6.4 mL.

[0022] Preferably, in step 17), the first heating is performed to a temperature of 90 to 120°C.

[0023] Preferably, in step 18) of step 1), the first heating is followed by maintaining the mixture under vacuum for 25 to 60 minutes.

[0024] Preferably, in 19) step 1), the first heating includes a third heating.

[0025] Preferably, in step 110), the first washing comprises washing with ethanol and washing with a mixed solution of ethanol and cyclohexane in sequence.

[0026] Preferably, in step 111) after the first washing, the product of the first washing is dispersed in a cyclohexane solution or a n-hexane solution.

[0027] Preferably, in step 112) of step 1), the mass volume ratio of the first washed product to the 1-octadecene is (1:1) to (1:1.5).

[0028] Preferably, in step 113) after the first washing, the volume ratio of the 1-octadecene to the oleic acid is 1:1.

[0029] Preferably, in step 114) of step 1), the volume mass ratio of the 1-octadecene to the sodium trifluoroacetate after the first washing is 6.4 ml:0.4 mmol.

[0030] Preferably, in step 115) after the first washing, the volume mass ratio of the 1-octadecene to the yttrium trifluoroacetate is 6.4 ml:0.4 mmol.

[0031] Preferably, in step 1) of 116), the second heating temperature is 90-120°C, for example, 90-100°C, 100-110°C, 110-120°C, etc.

[0032] Preferably, in step 117), the second heating is maintained under vacuum for 25 to 60 minutes, for example, 25 to 30 minutes, 30 to 35 minutes, 35 to 40 minutes, 40 to 45 minutes, 45 to 50 minutes, 50 to 55 minutes, 55 to 60 minutes, etc.

[0033] Preferably, in step 1) of 118), the fifth heating is performed after the second heating.

[0034] Preferably, in 119) step 1), cooling is also included before the second washing.

[0035] Preferably, in step 1) of 120), the second washing comprises washing with ethanol and washing with a mixed solution of ethanol and cyclohexane in sequence.

[0036] Preferably, in step 121), after the second washing, the product of the second washing is dispersed in a cyclohexane solution or a n-hexane solution.

[0037] Preferably, in step 21) of step 2), the mass ratio of the NaYbF4:Er@NaYF4 to the phosphatidylcholine is (1:0.5) to (1:4).

[0038] Preferably, in 22) step 2), the mass volume ratio of the phosphatidylcholine to the dichloromethane is (40 mg: 2 ml) to (40 mg: 10 ml).

[0039] Preferably, in 23) step 2), stirring is performed during the third mixing.

[0040] Preferably, in step 24) of step 2), after the third mixing, rotary evaporation is performed to disperse in a solvent.

[0041] Preferably, in step 31) of step 3), the mass molar ratio of the modified phosphatidylcholine NaYbF4:Er@NaYF4 to the Cy7.5 is (30-100 mg): (2.5-20 nmol), for example, it can be (30-40 mg): (2.5-20 nmol), (40-50 mg): (2.5-20 nmol), (50-60 mg): (2.5-20 nmol), (60-70 mg): (2.5-20 nmol), (70-80 mg): (2.5-20 nmol), (80-90 mg): (2.5-20 nmol), (90-100 mg): (2.5-20 nmol), etc.

[0042] Preferably, in step 3) of 32), the solvent of the Cy7.5 solution is dimethyl sulfoxide.

[0043] Preferably, in step 3), the fourth mixing further comprises oscillation.

[0044] Preferably, in 191) feature 19), the third heating is carried out under nitrogen protection.

[0045] Preferably, in 192) feature 19), the third heating temperature is 245-260°C; for example, it can be 245-250°C, 250-255°C, 255-260°C, etc.

[0046] Preferably, in 193) feature 19), the third heating is maintained for 30 to 60 minutes, for example, it can be 30 to 35 minutes, 35 to 40 minutes, 40 to 45 minutes, 45 to 50 minutes, 50 to 55 minutes, 55 to 60 minutes, etc.

[0047] Preferably, in 194) feature 19), the third heating includes a fourth heating.

[0048] Preferably, in 11011) feature 110), in the mixed solution of ethanol and cyclohexane, the volume ratio of ethanol to cyclohexane is 1:1.

[0049] Preferably, in 1102) feature 110), the ethanol and cyclohexane mixed solution is washed 1 to 3 times.

[0050] Preferably, in feature 1111), the mass volume ratio of the product of the first washing to the cyclohexane solution or the n-hexane solution is (1:1) to (1:1.5), for example, it can be (1:1) to (1:1.1), (1:1) to (1:1.2), (1:1) to (1:1.3), (1:1) to (1:1.4), (1:1) to (1:1.5), etc.

[0051] Preferably, in 1181) feature 118), the fifth heating is carried out under nitrogen protection.

[0052] Preferably, in 1182) feature 118), the fifth heating temperature is 290-310°C, for example, it can be 290-295°C, 295-300°C, 300-305°C, 305-310°C, etc.

[0053] Preferably, in 1183) feature 118), the fifth heating time is 60 to 90 min, for example, it can be 60 to 65 min, 65 to 70 min, 70 to 75 min, 75 to 80 min, 80 to 85 min, 85 to 90 min, etc.

[0054] Preferably, in 1184) feature 118), the fifth heating further comprises a sixth heating;

[0055] Preferably, in 1201) feature 120), in the mixed solution of ethanol and cyclohexane, the volume ratio of ethanol to cyclohexane is 1:1.

[0056] Preferably, in 1202) feature 120), the ethanol and cyclohexane mixed solution is washed 1 to 3 times.

[0057] Preferably, in feature 1211), the mass volume ratio of the product of the second washing to the cyclohexane solution or the n-hexane solution is (1:1) to (1:1.5); for example, it can be (1:1) to (1:1.1), (1:1) to (1:1.2), (1:1) to (1:1.3), (1:1) to (1:1.4), (1:1) to (1:1.5), etc.

[0058] Preferably, in 231) feature 23), the stirring time is 10 to 40 min, for example, it can be 10 to 15 min, 10 to 40 min, 15 to 20 min, 20 to 25 min, 25 to 30 min, 30 to 35 min, 35 to 40 min, etc.

[0059] Preferably, in 241) feature 24), the solvent is water.

[0060] Preferably, in 242) feature 24), the mass volume ratio of the solid obtained after the rotary evaporation to the solvent is (30-100) mg:1 mL, for example, it can be (30-40) mg:1 mL, (40-50) mg:1 mL, (50-60) mg:1 mL, (60-70) mg:1 mL, (70-80) mg:1 mL, (80-90) mg:1 mL, (90-100) mg:1 mL, etc.

[0061] Preferably, in feature 321), the molar volume ratio of Cy7.5 to dimethyl sulfoxide is 5 nmol:10 ul.

[0062] Preferably, in 331) feature 33), the oscillation time is 0.5 to 5 min, for example, 0.5 to 51 min, 1 to 1.5 min, 1.5 to 2 min, 2 to 2.5 min, 2.5 to 3 min, 3 to 3.5 min, 3.5 to 4 min, 4 to 4.5 min, 4.5 to 5 min, etc.;

[0063] Preferably, in 332) feature 33), the oscillation includes centrifugation and dispersion in a solvent.

[0064] Preferably, in 1941) feature 194), the fourth heating temperature is 250-270°C, for example, it can be 250-255°C, 255-260°C, 260-265°C, 265-270, etc.

[0065] Preferably, in 1942) feature 194), the fourth heating time is 10 to 20 minutes, for example, it can be 10 to 15 minutes, 15 to 20 minutes, etc.

[0066] Preferably, in 11841) feature 1184), the temperature of the sixth heating is 290-310°C, for example, it can be 290-295°C, 295-300°C, 300-305°C, 305-310°C, etc.

[0067] Preferably, in 11842) and feature 1184), the sixth heating time is 10 to 20 minutes, for example, it can be 10 to 15 minutes, 15 to 20 minutes, etc.

[0068] Preferably, in 2421) feature 242), the solvent is water.

[0069] Preferably, in 3321) feature 332), the solvent is water.

[0070] A third aspect of the present invention provides an application of a nanomaterial, wherein the nanomaterial is used as an optical nanoindicator or sensor for detecting membrane potential in the NIR-IIb window.

[0071] A fourth aspect of the present invention provides an optical indicator comprising the aforementioned nanoparticles, wherein the optical indicator is used for detecting membrane potential in the NIR-IIb window.

[0072] The fifth aspect of the present invention provides an optical nanosensor comprising the above-mentioned nanoparticles. The optical indicator in the present invention is used for detecting membrane potential in the NIR-IIb window.

[0073] This study successfully achieved membrane potential detection in the NIR-IIb window by constructing an organic-inorganic composite system composed of a voltage-sensitive dye molecule (Cy7.5) and a lanthanide luminescent nanomaterial (NaYbF4: Er@NaYF4). This system detects membrane potential without being affected by fluctuations in ion currents across the cell membrane. Furthermore, the present invention also enabled the spatiotemporal dynamics of abnormal discharges in epileptic lesions in the mouse hippocampus, with optical imaging results demonstrating significantly higher spatial resolution than those achieved with dye molecules with detection wavelengths less than 1000 nm.

[0074] The above technical solution has at least one of the following beneficial effects:

[0075] 1) This nanomaterial enables visual detection of membrane potential changes in cells labeled by the sensor through optical imaging. A notable feature of this sensor design is that both the incident and emitted light have good tissue penetration and low scattering, enabling further applications in visually detecting membrane potential in tissue samples and living organisms.

[0076] 2) The nanomaterials in the present invention have good uniformity.

[0077] 3) When the nanomaterials of the present invention are used as optical nanoindicators or sensors for membrane potential detection in the NIR-IIb window, the sensor generates emission light in the NIR-IIb window (1500-1700 nm) under the excitation of 808 nm near-infrared incident light. BRIEF DESCRIPTION OF THE DRAWINGS

[0078] Figure 1 These are the transmission electron microscope images and high-resolution transmission electron microscope images of NaYbF4:Er@NaYF4 of Example 1.

[0079] Figure 2 X-ray diffraction (XRD) patterns of NaYbF4:Er@NaYF4, NaYF4 and NaYbF4 in Example 1.

[0080] Figure 3 Fourier transform infrared spectra (FTIR) of NaYbF4:Er@NaYF4@DPPC and DPPC of Example 1. Figure 4 : This is the down-conversion emission spectrum of NaYbF4:Er@NaYF4@DPPC under 808nm near-infrared excitation before and after Cy7.5 dye sensitization in aqueous solution in Example 1.

[0081] Figure 5 This is the emission spectrum of NaYbF4:Er@NaYF4@DPPC in Example 2 labeled with Cy7.5 at different concentrations in aqueous solution.

[0082] Figure 6 This is the sensitization enhancement effect of Cy7.5 in Example 3 on NaYbF4:Er@NaYF4@DPPC with different thicknesses of the first shell layer.

[0083] Figure 7 This is a comparison chart of the luminescence intensity of NaYbF4:Er@NaYF4@DPPC with different first shell thicknesses in Example 3 under 808nm laser and optimal Cy7.5 concentration.

[0084] Figure 8 This is the recording of the changes in cell membrane potential under the control of square wave voltage by the optical nanosensor Cy7.5-ErNP in Example 4.

[0085] Figure 9 This is a verification of the sensitivity of the optical nanosensor Cy7.5-ErNP of Example 4 to common cations in the physiological environment.

[0086] Figure 10 The optical nanosensor Cy7.5-ErNP of Example 5 can achieve stable labeling of the cell membrane of HL-1 cardiomyocytes.

[0087] Figure 11 This is the visualization of the action potential generated by the optical nanosensor Cy7.5-ErNP in Example 5 on HL-1 cardiomyocytes in the NIR-IIb window.

[0088] Figure 12 The injection coordinates of the optical nanosensor Cy7.5-ErNP in Example 6 into the mouse brain area.

[0089] Figure 13 These are the H&E and TUNEL section staining results of the kainic acid-induced mouse epilepsy model in Example 6.

[0090] Figure 14These are the NIR-IIb window optical recording results after the optical nanosensor Cy7.5-ErNP of Example 6 was injected into the hippocampus and motor cortex of the kainic acid-induced mouse epilepsy model.

[0091] Figure 15 This is the EEG recording of the hippocampus and motor cortex in the kainic acid-induced mouse epilepsy model of Example 6. DETAILED DESCRIPTION

[0092] The technical solution of the present invention is described below through specific examples. It should be understood that the one or more method steps mentioned in the present invention do not exclude the presence of other method steps before and after the combination step or the insertion of other method steps between these explicitly mentioned steps; it should also be understood that these embodiments are only used to illustrate the present invention and are not used to limit the scope of the present invention. Moreover, unless otherwise specified, the numbering of each method step is only a convenient tool for identifying each method step, and is not intended to limit the order of arrangement of each method step or to define the scope of the present invention. Changes or adjustments in their relative relationships, without substantially changing the technical content, should also be regarded as the scope of the present invention.

[0093] Example 1

[0094] Preparation of NaYbF4:5% Er@NaYF4@Cy7.5@DPPC (abbreviated as Cy7.5-ErNP):

[0095] 1 mmol of rare earth trifluoroacetate (Yb=0.95, Er=0.05), 1 mmol of sodium trifluoroacetate, 3.2 mL of oleic acid (OA), 3.3 mL of oleylamine (OM) and 6.4 mL of 1-octadecene were mixed in a 100 mL three-necked flask to obtain a mixed solution;

[0096] The mixed solution was heated to 100°C and maintained under vacuum for 30 minutes. Subsequently, under nitrogen protection, the mixed solution was heated to 250°C for 45 minutes and then to 260°C for 15 minutes. After the mixed solution was cooled to room temperature, an equal volume of ethanol was added and centrifuged to obtain a solid. The solid was then washed three times with a 1:1 v / v ethanol:cyclohexane solution and finally dispersed in 4 mL of cyclohexane.

[0097] In a separate 100 mL three-necked flask, 2 mL of the cyclohexane solution was mixed with 6.4 mL of 1-octadecene, 6.4 mL of oleic acid (OA), 0.4 mmol of sodium trifluoroacetate, and 0.4 mmol of yttrium trifluoroacetate. The solution was heated to 100°C under vacuum for 30 minutes, then heated to 295°C under nitrogen for 75 minutes, and then to 300°C for 15 minutes.

[0098] After the system cooled naturally, an equal volume of ethanol was added and centrifuged to obtain a solid. The solid was then washed three times with a 1:1 v / v ethanol:cyclohexane solution and redispersed in 4 mL of cyclohexane. The resulting core-shell NaYbF4:5% Er@NaYF4 lanthanide luminescent nanoparticles were obtained.

[0099] The X-ray diffraction patterns and transmission electron microscopy results of the obtained nanoparticles are shown in Figure 1 , Figure 2 As shown. The X-ray diffraction pattern of NaYbF4:5%Er@NaYF4 was compared with the standard cards of cubic phase NaYF4 and NaYbF4 crystals, and the results were highly consistent. This proves that the nanocrystals prepared by this method are all cubic phase. In the transmission electron microscopy characterization results, the statistical result of the diameter of NaYbF4:5%Er@NaYF4 is about 10.18nm, and the 0.34nm interplanar spacing on the surface of the nanocrystal is almost consistent with the (11 1) interplanar spacing of NaYF4 crystals, which once again proves that the nanocrystals prepared by this method are NaYbF4:5%Er@NaYF4 with a core-shell structure.

[0100] The cyclohexane solution containing 20 mg of NaYbF4:5% Er@NaYF4 was added to 8 mL of dichloromethane solution containing 40 mg of phosphatidylcholine and stirred for 20 minutes to mix thoroughly. The organic solvent in the solution was quickly evaporated by rotary evaporation, and the resulting solid was redispersed in 5 mL of water; after further centrifugation, it was dispersed in 2 mL of water. The transformation of the surface ligand of NaYbF4:5% Er@NaYF4 is shown in Figure 2. Figure 3 As shown. NaYbF4:5%Er@NaYF4@DPPC (NaYbF4:Er@NaYF4 modified with phosphatidylcholine) at 1740 cm -1 It has obvious carbonyl stretching vibration characteristic peak at 2850cm -1 and 2920cm -1 There are obvious CH stretching vibration characteristic peaks, which are consistent with the infrared characteristic peaks of DPPC. This shows that DPPC is successfully coated on the outer layer of NaYbF4:5%Er@NaYF4.

[0101] Then, 1 mL of NaYbF4:5% Er@NaYF4 aqueous dispersion modified with phosphatidylcholine was taken, and 10 μL of dimethyl sulfoxide solution containing 5 nmol Cy7.5 was added to the dispersion. After vigorous shaking for 1 minute, centrifugation was performed again and the resulting solid was redispersed in 1 mL of water, which is NaYbF4:5% Er@NaYF4@Cy7.5@DPPC (abbreviated as Cy7.5-ErNP).

[0102] The luminescence intensity of the NIR-IIb window of NaYbF4:5%Er@NaYF4 and NaYbF4:5%Er@NaYF4@Cy7.5@DPPC with the same concentration under 808nm near-infrared light excitation was measured by spectroscopy. Figure 4 Compared with NaYbF4:5%Er@NaYF4, the luminescence intensity of NaYbF4:5%Er@NaYF4@Cy7.5@DPPC in the NIR-IIb window is significantly enhanced under 808nm near-infrared light excitation.

[0103] Example 2

[0104] Optimization of the Luminescence Enhancement Effect of Cy7.5 in Cy7.5-ErNP on the NIR-IIb Window of Lanthanide Luminescent Nanoparticles

[0105] As shown in Example 1, a cyclohexane solution containing 20 mg of NaYbF4:5% Er@NaYF4 was added to an 8 mL dichloromethane solution containing 40 mg of phosphatidylcholine and stirred for 20 minutes to mix evenly. After quickly evaporating the organic solvent of the solution by rotary evaporation, the obtained solid was redispersed in 5 mL of water; after centrifugation again, it was dispersed in 2 mL of water. The above operation was repeated 7 times to prepare a total of 7 samples. Then, 10 ul of dimethyl sulfoxide solution containing 0 / 2.5 / 5.0 / 7.5 / 10 / 15 / 20 nmol Cy7.5 was added to the dispersion respectively. After vigorous shaking for 1 minute, it was centrifuged again and the obtained solid was redispersed in 1 mL of water. The NIR-IIb window emission spectrum of Cy7.5-ErNP under 808 nm near-infrared light excitation was determined by spectrometry under different concentrations of Cy7.5, as shown in FIG. Figure 5 As shown in the figure, at a Cy7.5 concentration of 5 nmol / mL, Cy7.5-ErNP can achieve the optimal luminescence enhancement effect.

[0106] Example 3

[0107] Optimization of the first shell thickness in Cy7.5-ErNPs for the luminescence enhancement effect of lanthanide luminescent nanoparticles in the NIR-IIb window

[0108] By adjusting the dosage of sodium trifluoroacetate and yttrium trifluoroacetate during the NaYF4 shell synthesis process, NaYbF4:5%Er@NaYF4 nanoparticles with four different shell thicknesses (2.0±0.6, 2.4±0.6, 3.0±0.7, and 3.5±0.6 nm) were synthesized. 0.4 nmol, 0.8 nmol, 1.2 nmol, and 1.6 nmol of sodium trifluoroacetate and yttrium trifluoroacetate were added, respectively (using 0.4 nmol as an example, 0.4 nmol of sodium trifluoroacetate and 0.4 nmol of yttrium trifluoroacetate were added). Then, as in Example 1, 20 mg of NaYbF4:5%Er@NaYF4 with different first shell thicknesses was added to 8 mL of dichloromethane solution containing 40 mg of phosphatidylcholine and stirred for 20 minutes to mix thoroughly. After the organic solvent of the solution was quickly evaporated by rotary evaporation, the obtained solid was redispersed in 5 mL of water; after centrifugation again, it was dispersed in 2 mL of water.

[0109] Then, 10 μl of dimethyl sulfoxide solution containing the same amount of Cy7.5 was added to the dispersion, shaken vigorously for 1 minute, centrifuged again, and the resulting solid was redispersed in 1 mL of water. The NIR-IIb window emission spectrum of Cy7.5-ErNP under 808 nm near-infrared light excitation was measured using spectrometry under different first shell thicknesses. Figure 6 and 7 As shown, Cy7.5-ErNP with a NaYF4 shell thickness of 2.0 nm can achieve the highest multiple enhancement effect and the strongest luminescence of the material in the NIR-IIb window under 808 nm near-infrared light excitation.

[0110] Example 4

[0111] The Cy7.5-ErNP synthesized in Example 2 with a Cy7.5 concentration of 5nmol / mL was used to detect changes in membrane potential under square wave voltage stimulation: two silver chloride electrodes with a distance of about 3cm were fixed in parallel on a culture dish where HEK293T adherent cells were pre-cultured, and the electrodes were connected to a periodic square wave voltage signal; and the square wave voltage was applied to the cells. Under the generation of the square wave voltage, the cell membrane potential was induced to a fixed value; when the square wave voltage disappeared, the membrane potential returned to the resting level. Cy7.5-ErNP with a potential change of 100mV generated by the cell membrane was added to the culture dish, incubated with the cells for 3 hours, and then the cells were imaged by a near-infrared camera under 808nm near-infrared light excitation. The results are shown in the figure. Figure 8 As shown in the figure, the luminous intensity of the second near-infrared region of the sensor is significantly improved when the square wave voltage is generated, and changes periodically with the voltage signal.

[0112] To verify the insensitivity of Cy7.5-ErNP to the ion flow on both sides of the membrane accompanying the action potential, Cy7.5-ErNP was exposed to different concentrations of Na + or K + The NIR-IIb luminescence intensities of Cy7.5-ErNP under the conditions of ion concentration close to the physiological environment were also compared. Figure 9 The results showed that Cy7.5-ErNPs were not disturbed by changes in ion concentration in the environment.

[0113] Example 5

[0114] HL-1 cells grown adherently in Claycomb medium are a cell line that retains cardiac electrophysiological properties and can spontaneously generate action potentials. Figure 10 As shown, co-localization imaging captured the upconversion luminescence of Cy7.5-ErNP anchored on the membrane surface under 808nm excitation, as well as calcein AM (green) in HL-1 cells. Cy7.5-ErNP synthesized in Example 2 with a concentration of 5nmol / mL and a dosage of 500ug / mL was co-incubated with HL-1 cells for 6 hours. With the assistance of a customized microscopic imaging system, after Cy7.5-ErNP was labeled to HL-1 cells using a near-infrared second-zone CMOS camera, the optical nanosensor continuously recorded the luminescence intensity in the NIR-IIb window. Figure 11 As shown, Cy7.5-ErNPs enabled visualization of cell action potentials, and the optical signal of Cy7.5-ErNPs was detected by a 1450 nm long-pass filter.

[0115] Example 6

[0116] Optical imaging in the NIR-IIb window has an overwhelming advantage in tissue penetration depth compared to other short-wavelength imaging methods, enabling high-spatial-resolution in vivo imaging. Therefore, Cy7.5-ErNPs have been further used for low-invasive optical detection and diagnosis of abnormal discharges in epileptic lesions in the mouse hippocampus.

[0117] First, 8-week-old healthy male C57BL / 6 mice were selected and their skulls fixed under anesthesia. After cleaning the skull hair, the scalp was cut along the midline of the skull, and the skull surface was cleaned with a cotton swab dipped in 10% H2O2. An electric drill was used to drill a hole in the left hippocampus (AP: -2mm, ML: -1.5mm). A microsyringe containing alkaloid acid was inserted into the hole and lowered to a depth of 2mm under motor control. 0.2μL (5mM) alkaloid acid solution was injected to complete the induction of hippocampal epileptic lesions. Subsequently, holes were drilled in another area of the hippocampus (AP: -2.8mm, ML: -3.0mm) and the motor cortex (AP: -0.8mm, ML: 0.8mm). 5uL of Cy7.5-ErNP solution (20mg / mL) synthesized in Example 2 with a concentration of 5nmol / mL was injected into both locations using a microsyringe. Finally, the cut skin was sealed with biological glue, and the mouse was allowed to wake up naturally. The injection position of Cy7.5-ErNP is as follows Figure 12 As shown. H&E and TUNEL staining results of model mouse brain tissue, as shown Figure 13 As shown in Figure 2, significant neuronal apoptosis and fragmented DNA were observed in the hippocampus ipsilateral to kainic acid injection. Comparison of hematoxylin and eosin (H&E) staining results of hippocampal sections from control mice and kainic acid-treated mice. Significant nuclear loss was observed in the CA2 / CA3 regions of the hippocampus ipsilateral to kainic acid injection.

[0118] The optical signal of the NIR-IIb window of Cy7.5-ErNP from the mouse brain was recorded by an InGaAs camera with a 1500nm long-pass filter. Figure 14 As shown in the figure, the light signal showed a platform-like oscillation accompanied by the stiffening of the tail, which is consistent with the EEG characteristics of mouse epileptic seizures. In addition, the implanted electrodes were also used as the "gold standard" to detect the electroencephalogram (EEG) during the abnormal discharge stage of the epileptic lesions in the hippocampus of kainic acid-induced mice. Silver electrodes were implanted in the skulls of kainic acid-induced mice for electroencephalogram (EEG) recording. The distribution of EEG signals in the hippocampus and motor cortex during the mouse epileptic seizure was recorded, as shown in the figure. Figure 15 The EEG results were consistent with the optical detection results of Cy7.5-ErNP in the NIR-IIb window, both of which recorded abnormal epileptic discharges lasting about 20 seconds.

[0119] The above embodiments are merely illustrative of the principles and effects of the present invention and are not intended to limit the present invention. Anyone skilled in the art may modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by one of ordinary skill in the art without departing from the spirit and technical principles disclosed herein are intended to be covered by the claims of the present invention.

Claims

1. A nanomaterial, characterized in that: The nanomaterial is NaYbF4:Er@NaYF4@Cy7.5@DPPC, and the nanomaterial comprises a core-shell structure of components NaYbF4:Er@NaYF4, wherein the core-shell structure is modified with DPPC and compounded with Cy7.

5.

2. The nanomaterial according to claim 1, characterized in that Include at least one of the following technical features: A1) The NaYbF4:Er@NaYF4 contains 1 to 20% by mole of Er; A2) The thickness of the shell layer close to the core is 1.4-4.1 nm.

3. The nanomaterial according to claim 1, characterized in that Include at least one of the following technical features: A3) the diameter of the core is 7 to 12 nm; A4) the diameter of the nanoparticles is 15 to 30 nm; A5) The mass ratio of NaYbF4:Er@NaYF4 to the nanomaterial is (1:1.5)-(1:5); A6) the mass molar ratio of the NaYbF4:Er@NaYF4 to the Cy7.5 is 20 mg:(2.5-20 nmol); A7) The mass ratio of DPPC to the nanoparticles is (0.5:1.5)-(4:5).

4. The method for preparing a nanomaterial according to any one of claims 1 or 2, characterized in that: The following steps are involved: 1) a rare earth trifluoroacetate, sodium trifluoroacetate, oleic acid, oleylamine, and 1-octadecene are first mixed, heated, and washed, and a product of the first wash is second mixed with 1-octadecene, oleic acid, sodium trifluoroacetate, and yttrium trifluoroacetate, heated, and washed to obtain NaYbF4:Er@NaYF4; 2) The NaYbF4:Er@NaYF4 obtained in step 1) is mixed with phosphatidylcholine and dichloromethane to obtain NaYbF4:Er@NaYF4 modified with phosphatidylcholine; 3) The phosphatidylcholine-modified NaYbF4:Er@NaYF4 obtained in step 2) is mixed with the Cy7.5 solution for the fourth time to obtain NaYbF4:Er@NaYF4@Cy7.5@DPPC (Cy7.5-ErNP).

5. The nanomaterial according to claim 3, characterized in that Include at least one of the following technical features: 11) In step 1), the rare earth trifluoroacetate is selected from one or more of ytterbium trifluoroacetate and erbium trifluoroacetate; 12) In step 1), Yb 3+ The molar fraction of Er is 0.99~0.80, 3+ The mole fraction is 0.01 to 0.20; 13) In step 1), before the first washing, the molar ratio of the rare earth trifluoroacetate to sodium trifluoroacetate is 1:1; 14) In step 1), before the first washing, the molar volume ratio of the rare earth trifluoroacetate to oleic acid is 1 mmol:3.2 mL; 15) In step 1), before the first washing, the molar volume of the rare earth trifluoroacetate and oleylamine is 1 mmol:3.3 mL; 16) In step 1), before the first washing, the molar ratio of the rare earth trifluoroacetate to 1-octadecene is 1 mmol:6.4 mL; 17) In step 1), the first heating is to a temperature of 90 to 120°C; 18) In step 1), the first heating is followed by maintaining the mixture under vacuum for 25 to 60 minutes; 19) In step 1), the first heating includes a third heating; 110) In step 1), the first washing sequentially comprises washing with ethanol and washing with a mixed solution of ethanol and cyclohexane; 111) In step 1), after the first washing, the product of the first washing is dispersed in a cyclohexane solution or a n-hexane solution; 112) In step 1), the mass volume ratio of the first washed product to the 1-octadecene is (1:1) to (1:1.5); 113) In step 1), after the first washing, the volume ratio of the 1-octadecene to the oleic acid is 1:1; 114) In step 1), after the first washing, the volume mass ratio of the 1-octadecene to the sodium trifluoroacetate is 6.4 ml:0.4 mmol; 115) In step 1), after the first washing, the volume mass ratio of the 1-octadecene to the yttrium trifluoroacetate is 6.4 ml:0.4 mmol; 116) In step 1), the second heating is to a temperature of 90 to 120°C; 117) In step 1), the second heating is followed by maintaining the mixture under vacuum for 25 to 60 minutes; 118) In step 1), the second heating is followed by a fifth heating; 119) In step 1), cooling is further included before the second washing; 120) In step 1), the second washing comprises washing with ethanol and washing with a mixed solution of ethanol and cyclohexane in sequence; 121) In step 1), after the second washing, the product of the second washing is dispersed in a cyclohexane solution or a n-hexane solution; 21) In step 2), the mass ratio of the NaYbF4:Er@NaYF4 to the phosphatidylcholine is (1:0.5) to (1:4); 22) In step 2), the mass volume ratio of the phosphatidylcholine to the dichloromethane is (40 mg:2 ml) to (40 mg:10 ml); 23) In step 2), stirring is performed during the third mixing; 24) In step 2), after the third mixing, the mixture is subjected to rotary evaporation and dispersed in a solvent; 31) In step 3), the mass molar ratio of the modified phosphatidylcholine NaYbF4:Er@NaYF4 to the Cy7.5 is (30-100 mg): (2.5-20 nmol); 32) In step 3), the solvent of the Cy7.5 solution is dimethyl sulfoxide; 33) In step 3), the fourth mixing further includes oscillation.

6. The method for preparing the nanomaterial according to claim 4, wherein: Include at least one of the following technical features: 191) In feature 19), the third heating is performed under nitrogen protection; 192) In feature 19), the third heating step is performed to a temperature of 245-260°C; 193) In feature 19), the third heating is maintained for 30 to 60 minutes; 194) In feature 19), the third heating includes a fourth heating; 11011) In feature 110), the volume ratio of ethanol to cyclohexane in the mixed solution of ethanol and cyclohexane is 1:1; 1102) In feature 110), the number of washing times of the mixed solution of ethanol and cyclohexane is 1 to 3 times; 1111) In feature 111), the mass volume ratio of the product of the first washing to the cyclohexane solution or the n-hexane solution is (1:1) to (1:1.5); 1181) In feature 118), the fifth heating is performed under nitrogen protection; 1182) In feature 118), the fifth heating is to a temperature of 290-310°C; 1183) In feature 118), the fifth heating time is 60 to 90 minutes; 1184) In feature 118), the fifth heating further comprises a sixth heating; 1201) In feature 120), in the mixed solution of ethanol and cyclohexane, the volume ratio of ethanol to cyclohexane is 1:1; 1202) In feature 120), the washing number of the ethanol and cyclohexane mixed solution is 1 to 3 times; 1211) In feature 121), the mass volume ratio of the second washed product to the cyclohexane solution or the n-hexane solution is (20 mg:1 mL) or (20 mg:1.5 mL); 231) In feature 23), the stirring time is 10 to 40 minutes; 241) In feature 24), the solvent is water; 242) In feature 24), the mass volume ratio of the solid obtained after rotary evaporation to the solvent is (30-100) mg:1 mL. 321) In feature 321), the molar volume ratio of Cy7.5 to dimethyl sulfoxide is 5 nmol:10 ul; 331) In feature 33), the oscillation time is 0.5 to 5 minutes; 332) In feature 33), the shaking includes centrifugation and dispersion in a solvent.

7. The nanomaterial according to claim 5, characterized in that Include at least one of the following technical features: 1941) In feature 194), the fourth heating is to a temperature of 250-270°C; 1942) In feature 194), the fourth heating time is 10 to 20 minutes; 11841) In feature 1184), the sixth heating temperature is 290-310°C; 11842) In feature 1184), the sixth heating time is 10 to 20 minutes; 2421) Feature 242), wherein the solvent is water; 3321) Feature 332), wherein the solvent is water.

8. An application of nanomaterials, characterized in that: The nanomaterial according to any one of claims 1 to 3 is used as an optical nanoindicator or sensor for detecting membrane potential in the NIR-IIb window.

9. An optical indicator, characterized in that The nanomaterial comprises the nanomaterial according to any one of claims 1 to 3.

10. An optical nanosensor, characterized in that: The nanomaterial comprises the nanomaterial according to any one of claims 1 to 3.

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