Preparation of host-guest doped tadf nanoparticles and application thereof in sted imaging
By designing a β-dicarbonyl fluorine-boron framework curcumin compound and using host-guest doping methods, TADF nanoparticles were prepared, solving the problems of low excited-state oscillator strength and weak aggregated-state luminescence in STED probe materials, and achieving low saturation power and high resolution STED imaging.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- CHINA AGRI UNIV
- Filing Date
- 2025-01-23
- Publication Date
- 2026-07-21
AI Technical Summary
Existing STED imaging probe materials suffer from problems such as low excited-state oscillator strength, weak aggregated-state luminescence, and easy quenching of long-lived triplet states, resulting in high power loss and insufficient imaging resolution.
Curcumin compounds with a β-dicarbonyl fluorine-boron framework were designed, and TADF nanoparticles were prepared by host-guest doping. By utilizing their charge transfer characteristics and long fluorescence lifetime, the luminescence intensity and stability were improved, achieving low saturation power and high resolution.
High-resolution STED imaging at low power was achieved. The nanoparticles exhibited bright deep red light emission, excellent optical stability, and high luminescence brightness, with a resolution of 122.5 nm.
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Figure CN119978006B_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of stimulated radiation loss (STED) super-resolution imaging probe preparation technology, specifically to a method for designing and synthesizing curcumin compound molecules based on a β-dicarbonyl fluorine boron framework and preparing low-power STED nanoparticle probes. Background Technology
[0002] With the development of modern biology and micro / nanotechnology, traditional optical microscopes can no longer meet the imaging resolution requirements of researchers. Stimulated emission depletion (STED) technology, as a super-resolution imaging method, has advantages such as fast imaging speed, no need for complex image post-processing, and relatively simple sample preparation.
[0003] The principle of STED super-resolution imaging technology is to selectively consume excited-state atoms in the edge region of the excited spot through stimulated emission (SE), causing these atoms to return to the ground state through SE, thereby reducing the emission range and achieving a resolution less than the diffraction limit. According to the STED imaging resolution formula: R = λ / [2NA(1+I STED / I sat ) 1 / 2 ], where NA is the numerical aperture of the objective lens, I STED To reduce light intensity, I sat This is the saturation power. Therefore, high-intensity I is typically required. STED To improve imaging resolution, this can lead to photobleaching of fluorophores and damage to biological samples. Furthermore, low saturation power ensures high resolution with low power loss. From Equation I... sat = hc / (τλ STED σ STED It can be seen that σ STED For stimulated radiation cross section, λ STED Let τ be the wavelength of the loss light and τ be the fluorescence lifetime. Therefore, developing novel STED probe materials with low saturation power and suitable fluorescence lifetime has significant practical application value and real-world significance.
[0004] In recent years, thermally activated delayed fluorescence (TADF) materials have attracted widespread attention in the fields of organic light-emitting diodes (OLEDs) and time-resolved imaging due to their advantages such as high exciton utilization efficiency and long emission time. Therefore, TADF molecules usually exhibit a large stimulated emission cross section and a long fluorescence lifetime, which is very beneficial for achieving low saturation power. However, there are few reports on the use of TADF compounds as STED imaging probes. The main reasons are as follows: (i) Most TADF molecules rely on the spatial separation between the highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) to reduce singlet-triple state splitting, thereby realizing the RISC process. However, space charge separation usually leads to low excited-state oscillator intensity and weak emission, which is not conducive to obtaining a large stimulated emission cross section; (ii) In the aggregated state, DA molecular systems with conjugated planar structures are prone to strong π-π interactions, leading to fluorescence quenching, which affects the stimulated emission loss efficiency and reduces imaging resolution; (iii) The long-lifetime triplet excited state of TADF usually relaxes due to intramolecular vibrations or is quenched by oxygen in the environment. Therefore, designing TADF material systems with large stimulated emission cross sections and long fluorescence lifetimes is of great significance, as it can provide new insights and perspectives for exploring novel STED probes. Summary of the Invention
[0005] Developing novel STED probe materials has significant practical application value and significance. TADF molecules typically exhibit large stimulated emission cross-sections and long fluorescence lifetimes, which are highly advantageous for achieving low saturation power. However, charge separation in most TADF molecules currently leads to low excited-state oscillator strength and weak aggregated-state luminescence, hindering the acquisition of large stimulated emission cross-sections. Furthermore, in the aggregated state, strong π-π interactions cause fluorescence quenching, affecting stimulated emission loss efficiency and reducing imaging resolution. Finally, the long-lived triplet excited states of TADFs are often relaxed by intramolecular vibrations or quenched by oxygen in aqueous environments. Therefore, research on the use of TADF materials as STED imaging probes is currently scarce.
[0006] This patent mainly designs and synthesizes curcumin derivatives with a β-dicarbonyl fluorine-boron framework, and constructs host-guest type TADF nanoparticles to achieve low-power STED imaging:
[0007] A curcumin-like compound containing a β-dicarbonyl fluorine-boron skeleton, wherein the structure of Formula I is as follows:
[0008]
[0009] Wherein, R is selected from thiophene groups, preferably thiophene alkyl groups, more preferably 2-thiophene-methyl, 5-bromothiophene-2-methyl or dibenzothiophene-2-methyl.
[0010] In some preferred embodiments, the compound represented by Formula I has the following specific structure:
[0011]
[0012] In some preferred embodiments, the compound represented by Formula I is obtained via the following reaction route:
[0013]
[0014] Step 1: Add ethyl acetate (EtOAc) to a mixture of compound 1, ethyl diacetate, and boron trifluoride diethyl ether (BF3·Et2O), and heat to carry out the first contact reaction;
[0015] Step 2: Compound 2 and tri-tert-butylboronic acid ester B(n-OBu)3 are added to the mixture from step 1, followed by the addition of n-butylamine BuNH2 to carry out the second contact reaction;
[0016] Step 3: Cool and purify the crude product to obtain curcumin compound with a β-dicarbonyl fluoroboron skeleton.
[0017] In some preferred embodiments, in step 2, compound 2 is selected from any one of the following groups: 2-thiophene-2-methyl, 5-bromothiophene-2-methyl, and dibenzothiophene-2-carboxaldehyde.
[0018] In some preferred embodiments, the conditions for the first contact reaction in step 1 include: a reaction time of 0.5 to 1 h; a reaction temperature of 50 to 60 °C; preferably, a reaction time of 0.5 h and a reaction temperature of 55 °C; more preferably, compound 1 and boron trifluoride diethyl ether (BF3•Et2O) are weighed into a branch-tube reactor, ethyl acetate is added as a solvent, and the reaction is carried out for 0.5 hours under oil bath heating at 55 °C.
[0019] and / or
[0020] The conditions for the second contact reaction include: a reaction time of 8-12 h; a reaction temperature of 50-60 °C; preferably, a reaction time of 12 h and a reaction temperature of 55 °C; more preferably, compound 2 and tri-tert-butylboronic acid ester B(n-OBu)3 are added to the reaction solution, n-butylamine BuNH2 is added to the reaction system after 0.5-1 hour, and n-butylamine BuNH2 is added to the reaction system again after 5-7 hours.
[0021] In some preferred embodiments, the molar equivalent ratio of compound 1 (ethyl diacetoacetate), boron trifluoride ether (BF3·Et2O), compound 2, tritert-butylboronate (B(n-OBu)3), and n-butylamine (BuNH2) in the reaction system is 1~1.2:1.1~1.3:2.5~3:2.5~3:0.6~0.8.
[0022] In some preferred embodiments, in step 3, purification is performed using a silica gel column; preferably, the elution reagent for purification is petroleum ether / V: dichloromethane / V = 1~1.2:1; more preferably, the product is dark red and the yield is 50~60%.
[0023] Secondly, the present invention provides a method for preparing host-guest doped TADF nanoparticles using the curcumin derivative containing the β-dicarbonyl fluorine-boron framework, comprising the following steps:
[0024] Step a: The curcumin derivative containing the β-dicarbonyl fluorine boron skeleton is used as the host molecule, and the guest molecule, poloxamer F127 and tetrahydrofuran solvent are dissolved to obtain a mixed solution.
[0025] Step b: The mixed solution is rapidly injected into deionized water, sonicated, and then allowed to stand at room temperature; the above system is repeatedly centrifuged and washed with deionized water to obtain a host-guest doped TADF nanoparticle solution.
[0026] In step a, the main molecule is selected from any one of the following groups: BDSF, BRSF, and BBSF;
[0027] The guest molecule is selected from any of the following groups: 4,4'-bis(9-carbazole)biphenyl (CBP), 1,3-biscarbazole-9-ylbenzene (mCP) and 1,3,5-tris(1-phenyl-1H-benzimidazol-2-yl)benzene (TPBi).
[0028] The molar equivalent ratio of the host molecule to the guest molecule is 2~5:1, and 10~11 mg of poloxamer F127 is added to each 1 mL of tetrahydrofuran solvent.
[0029] In step b, the volume ratio of the mixed solution to deionized water is 1:9~10, the ultrasonic time is 1~2 minutes, the frequency is 20~25 MHz, and after standing for 12~24 hours, it is centrifuged at a speed of 9000~10000 rpm.
[0030] Thirdly, the present invention provides a low-power host-guest doped TADF nanoparticle as described above.
[0031] Fourthly, the present invention also provides an application of the aforementioned low-power host-guest doped TADF nanoparticles in STED imaging.
[0032] The beneficial effects of this invention are as follows:
[0033] 1. In this invention, β-dicarbonyl fluorine boron is selected as the electron acceptor and groups containing different thiophene structures are used as electron donors. The designed and synthesized compounds have obvious charge transfer characteristics and exhibit bright deep red light emission in their molecular aggregate state.
[0034] 2. This invention utilizes host-guest doping to improve the luminescence intensity, optical stability, and fluorescence lifetime of nanoparticles in aqueous solutions.
[0035] 3. The host-guest doped TADF nanoparticles of this invention achieve low saturation power (150.46 mW) and high-resolution STED imaging (122.5 nm).
[0036] 4. The synthetic route for the curcumin derivative with the β-dicarbonyl fluorine-boron framework described in this invention is highly efficient and yields high reaction rates. The preparation method for host-guest doped nanoparticles is simple and economical, exhibiting high luminescence brightness and TADF characteristics. As STED nanoparticle probes, they demonstrate strong optical stability and low saturation power, thus enabling ultra-high resolution imaging. Attached Figure Description
[0037] Figure 1 A schematic diagram of the synthesis process of curcumin compounds with a β-dicarbonyl fluorine boron skeleton.
[0038] Figure 2 The chemical structure of curcumin compounds with a β-dicarbonyl fluorine boron skeleton.
[0039] Figure 3 TADF properties characterization of curcumin compounds with a β-dicarbonyl fluoroboron skeleton.
[0040] Figure 4 This is a schematic diagram of the preparation process of host-guest doped nanoparticles.
[0041] Figure 5 Transmission electron microscopy images of host-guest doped nanoparticles, scale bar 500 nm.
[0042] Figure 6 (a) Absorption / emission spectrum of BRSF@CBP nanoparticles.
[0043] Figure 7 The fluorescence intensity of BRSF@CBP nanoparticles at (a) different excitation powers, and (b) without...
[0044] At the same time, the efficiency of loss.
[0045] Figure 8(a) Confocal and (b) STED images of BRSF@CBP nanoparticles, (c, d)
[0046] (e) Magnified image at the yellow dashed line, scale bar 3000 nm, resolution in confocal and super-resolution modes, scale bar 500 nm. Detailed Implementation
[0047] This patent primarily provides a method for synthesizing curcumin compounds containing a β-dicarbonyl fluorine-boron framework. A TADF-type nanoparticle probe was prepared through host-guest doping, ultimately achieving high-resolution STED imaging at low power. The experimental scheme mainly consists of the following six parts:
[0048] Example 1: Synthesis of curcumin compounds containing a β-dicarbonyl fluorine boron skeleton
[0049] This embodiment 1 provides a method for synthesizing curcumin compounds containing a β-dicarbonyl fluorine boron skeleton. Figure 1 and Figure 2 The β-dicarbonyl fluorine-boron skeleton exhibits a strong electron-withdrawing effect, and its photophysical properties vary significantly depending on the molecular packing state. Compounds based on this skeleton are generally reported to possess excellent stimulated emission properties, demonstrating superior photophysical properties and assembly performance. Based on this, β-dicarbonyl fluorine-boron was selected as the electron acceptor, and curcumin compounds containing the β-dicarbonyl fluorine-boron skeleton were designed and synthesized. Curcumin derivatives containing the β-dicarbonyl fluorine-boron skeleton have advantages such as simple synthetic steps, readily available and inexpensive raw materials, simple purification methods, and high yields. The compounds shown were obtained according to the reaction route:
[0050] Step 1: Compound 1 undergoes a first contact reaction with BF3·Et2O in the presence of ethyl acetate (EtOAc).
[0051] In step 1, compound 1 and boron trifluoride diethyl ether (BF3•Et2O) are weighed into a sidewall reactor, and ethyl acetate (EtOAc) is added as a solvent. The reaction is carried out under oil bath heating at 50-60 °C for 0.5-1 hours.
[0052] Step 2: Compound 2 and tri-tert-butylboronic acid ester B(n-OBu)3 are added to the above reaction system to carry out a second contact reaction in the presence of n-butylamine BuNH2.
[0053] In step 2, compound 2 includes, but is not limited to, 2-thiophenecarboxaldehyde, 5-bromothiophene-2-carboxaldehyde, and dibenzothiophene-2-carboxaldehyde. Compound 2 and tri-tert-butylboronic acid ester are added to the reaction solution. After 0.5–1 hour, n-butylamine is added to the reaction system, and after 5–7 hours, n-butylamine is added again. The reaction time is 8–12 hours, and the oil bath heating temperature is 50–60°C. The molar equivalent ratio of compound 1, boron trifluoride diethyl ether, compound 2, tri-tert-butylboronic acid ester, and n-butylamine is 1–1.2 : 1.1–1.3 : 2.5–3 : 2.5–3 : 0.6–0.8.
[0054] Step 3: After cooling, the crude product is purified by silica gel column chromatography.
[0055] In step 3, the elution reagent is a mixed solution of petroleum ether and dichloromethane with a volume ratio of 1 to 1.2:1, which finally yields a dark red product with a yield of 50 to 60%.
[0056] The 1H NMR spectral data are as follows:
[0057] BDSF: 1H NMR (600 MHz, Chloroform-d) δ 8.33 (s, 2H), 7.60 (s, 2H), 7.48 (s, 2H), 7.21 (s, 2H), 7.16 (s, 2H), 4.46 (s, 2H), 1.49 (s, 3H).
[0058] BRSF: 1H NMR (600 MHz, Chloroform-d) δ 8.17 (d, J = 14.9 Hz, 2H), 7.22 (d, J = 4.0 Hz, 2H), 7.13 (d, J = 4.0 Hz, 2H), 7.10 (d, J = 15.0 Hz, 2H), 4.45 (q, J = 7.1 Hz, 2H), 1.48 (t, J = 7.1 Hz, 3H).
[0059] BBSF: 1H NMR (600 MHz, Chloroform-d) δ 8.39 - 8.32 (m, 4H), 8.19 (dd,J = 6.7, 2.0 Hz, 2H), 7.89 (d, J = 8.3 Hz, 2H), 7.84 (dd,J = 6.7, 1.8 Hz,2H), 7.73 (dd, J = 8.4, 1.4 Hz, 2H), 7.56 - 7.49 (m, 6H), 4.58 (q, J = 7.1Hz, 2H), 1.27 (s, 3H).
[0060] Example 2: Characterization of properties of curcumin compounds containing a β-dicarbonyl fluorine boron skeleton
[0061] A CBP:BRSF solution with a mass ratio of 20:1 was prepared using CBP as the host molecule, BRSF as the guest molecule, and ultradry chlorobenzene as the solvent. The solution was stirred to ensure complete homogeneity. BRSF / CBP films were then prepared using spin coating. 200 μL of this solution was spin-coated onto a clean glass substrate at 3500 rpm / min for 30 s. The substrate was then annealed at 80°C for 30 min to remove residual trace solvent. Figure 3 The photoluminescence decay curve of the BRSF / CBP film at 700 nm was depicted, exhibiting double exponential decay, with instantaneous and delayed component lifetimes of 3.92 ns and 21.8 μs, respectively. Furthermore, the temperature sensitivity of fluorescence decay is an important property of TADF molecules; the delayed component lifetime significantly decreases to 9.86 μs at 78 K. Therefore, the emission peak at 700 nm originates from TADF emission. Figure 3 As shown in b, the instantaneous and delayed spectra of the BRSF / CBP film are similar, indicating that they originate from the same singlet excited state. Therefore, BRSF exhibits TADF properties.
[0062] Example 3: Preparation of host-guest doped nanoparticles
[0063] like Figure 4 As shown in Example 3, this invention provides a method for preparing host-guest doped nanoparticles. A curcumin compound with a β-dicarbonyl fluorine-boron backbone as the host molecule and a guest molecule are embedded in poloxamer F127, a polymer with good biocompatibility, to form host-guest doped nanoparticles.
[0064] Step 1: Weigh the host molecule, guest molecule, poloxamer F127 and tetrahydrofuran solvent and place them in a beaker and stir until dissolved.
[0065] Step 2: Rapidly inject the mixed solution into deionized water, sonicate for a period of time, and then allow it to stand at room temperature. Repeatedly centrifuge the above system and wash with deionized water to obtain a host-guest doped nanoparticle solution.
[0066] In step 1, the host molecules include, but are not limited to, BDFS, BRSF, and BBSF, and the guest molecules include, but are not limited to, 4,4'-bis(9-carbazole)biphenyl (CBP), 1,3-dicarbazole-9-ylbenzene (mCP), and 1,3,5-tris(1-phenyl-1H-benzimidazol-2-yl)benzene (TPBi). The molar equivalent ratio of the host and guest molecules is 2 to 5:1, and 10 to 11 mg of poloxamer F127 is added to each 1 mL of tetrahydrofuran solvent.
[0067] In step 2, the volume ratio of the mixed solution to deionized water is 1:9~10, the sonication time is 1~2 minutes, the frequency is 20~25 MHz, and after standing for 12~24 hours, it is centrifuged at a speed of 9000~10000 rpm.
[0068] Example 4 Morphology characterization of host-guest doped nanoparticles
[0069] Taking BRSF@CBP nanoparticles as an example, 10 mg of poloxamer F127 was added to THF solutions of BDSF, BBSF, and BRSF (0.00005 mmol) and CBP (0.0001 mmol), respectively. Then, 1 mL of the above mixed solution was rapidly injected into 9 mL of deionized water under ultrasonication. After standing at room temperature for 12 h, the solution was washed several times with deionized water and centrifuged to obtain three nanoparticle solutions. Figure 5 As shown, transmission electron microscopy images reveal that the nanoparticles of BDSF@CBP, BRSF@CBP, and BBSF@CBP have uniform spherical morphologies with sizes of 50-70 nm, 30-40 nm, and 40-50 nm, respectively.
[0070] Example 5 Optical characterization of host-guest doped TADF nanoparticles
[0071] like Figure 6 As shown in Figure a, the absorption peak of BRSF@CBP nanoparticles is located at 509 nm, exhibiting deep red emission at a peak of 701 nm, and possessing a large Stokes shift of 192 nm. Simultaneously, the lifetime of BRSF@CBP nanoparticles is 8.28 ns, demonstrating a relatively long TADF lifetime in solution. Figure 6 b).
[0072] Example 6: STED properties of host-guest doped TADF nanoparticles
[0073] Based on the peak positions of the absorption and emission spectra of BRSF@CBP nanoparticles, 470 nm continuous wavelength light was selected as the excitation beam, and 775 nm continuous wavelength laser light was selected as the STED beam. With increasing STED power, the fluorescence intensity of BRSF@CBP nanoparticles significantly decreased. When the STED power reached 435.75 mW, the stimulated emission loss efficiency of BRSF@CBP nanoparticles reached 67%. Figure 7 a). The saturation power of BRSF@CBP nanoparticles is 150.46 mW, lower than most reported STED nanoparticle probes, enabling STED imaging at low power. Furthermore, the optical stability of the probe is also a crucial factor in super-resolution imaging. Since STED imaging requires very high loss power, it easily leads to photobleaching of molecules, thus requiring the probe to have strong resistance to photobleaching. To verify that the weakening of light intensity in the experiment was due to photoloss rather than photobleaching, the "on and off" of the loss wavelength was controlled and repeated for 600 s. It was ultimately found that BRSF@CBP experienced only about 7% light loss in each cycle (…). Figure 7 (b) This eliminates the reduction in fluorescence intensity caused by photobleaching, and also indicates that BRSF@CBP has excellent optical stability.
[0074] Example 7: STED imaging of host-guest doped TADF nanoparticles
[0075] In the confocal and STED images of BRSF@CBP nanoparticles ( Figure 8 a) In confocal mode, the sample can only be displayed as a bright spot with a large luminous area, and the precise internal information of the spot cannot be resolved, thus the achievable resolution is approximately 189.44 nm. In STED mode, the large spot is actually a small luminous point, and the highest achievable resolution is approximately 122.50 nm. Figure 8 b).
[0076] While the specific embodiments of the present invention have been described above in conjunction with the accompanying drawings, this is not intended to limit the scope of protection of the present invention. Those skilled in the art should understand that, based on the technical solutions disclosed in the present invention, various modifications or variations that can be made by those skilled in the art without creative effort should be included within the scope of protection of the present invention.
Claims
1. A curcuminoid compound containing a β-dicarbonyl fluorine-boron skeleton, the structure of which is as follows: 。 2. A method for preparing a curcumin-like compound containing a β-dicarbonyl fluorine-boron skeleton as described in claim 1, characterized in that, This includes obtaining the compound shown in Formula I via the following reaction route: ， Step 1: Add ethyl acetate (EtOAc) to the mixture of compound 1, namely ethyl diacetate and boron trifluoride diethyl ether (BF3·Et2O), heat to carry out the first contact reaction, and react for 0.5 hours in an oil bath at 55 °C. Step 2: Add compound 2, namely 5-bromothiophene-2-carboxaldehyde and tri-n-butyl borate B(n-OBu)3, to the reaction system after 0.5 to 1 hour. Then add n-butylamine BuNH2 to the reaction system after 5 to 7 hours. Step 3: Cool and purify the crude product to obtain curcumin compound with a β-dicarbonyl fluoroboron skeleton.
3. The method for preparing a curcumin-like compound containing a β-dicarbonyl fluorine-boron skeleton according to claim 2, wherein, The molar equivalent ratio of compound 1, boron trifluoride ether BF3·Et2O, compound 2, tri-n-butyl borate B(n-OBu)3, and n-butylamine BuNH2 in the reaction system is 1~1.2:1.1~1.3:2.5~3:2.5~3:0.6~0.
8.
4. The method for preparing a curcuminoid compound containing a β-dicarbonyl fluorine-boron skeleton according to claim 2, wherein, In step 3, purification is performed using a silica gel column; the elution reagent used for purification is petroleum ether / V: dichloromethane / V = 1~1.2:
1.
5. A method for preparing a curcuminoid compound containing a β-dicarbonyl fluorine-boron skeleton according to claim 2 or 4, wherein, In step 3, the product is dark red, and the yield is 50-60%.
6. A low-power host-guest doped TADF nanoparticle for STED imaging, the nanoparticle comprising BRSF as the host molecule of claim 1 and 4,4'-bis(9-carbazole)biphenyl CBP as the guest molecule, the nanoparticle having a size of 30-40 nm, an absorption peak at 509 nm, exhibiting deep red light emission at a peak of 701 nm, having a large Stokes shift of 192 nm, and a lifetime of 8.28 ns.
7. A method for preparing low-power host-guest doped TADF nanoparticles for STED imaging as described in claim 6, comprising the following steps: Step a: Dissolve the host molecule, the guest molecule, poloxamer F127, and tetrahydrofuran solvent to obtain a mixed solution; Step b: The mixed solution is rapidly injected into deionized water, sonicated, and then allowed to stand at room temperature; the above system is repeatedly centrifuged and washed with deionized water to obtain a host-guest doped TADF nanoparticle solution. In step a, the main molecule is BRSF; The guest molecule is: 4,4'-bis(9-carbazole)biphenyl CBP; The molar equivalent ratio of the host molecule to the guest molecule is 2~5:1, and 10~11 mg of poloxamer F127 is added to each 1 mL of tetrahydrofuran solvent; In step b, the volume ratio of the mixed solution to deionized water is 1:9~10, the ultrasonic time is 1~2 minutes, the frequency is 20~25 MHz, and after standing for 12~24 hours, it is centrifuged at a speed of 9000~10000 rpm.
8. The application of the low-power host-guest doped TADF nanoparticles for STED imaging as described in claim 6 in the preparation of STED imaging probes.