Dye-sensitized rare earth up-conversion luminescent nano material as well as preparation method and application thereof
By introducing unsaturated fatty acid salt self-assembled shells and near-infrared dye modification onto UCNPs, the problems of small absorption cross-section, poor water solubility, and insufficient photostability of UCNPs were solved, enabling efficient and stable biological applications.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-10
- Publication Date
- 2026-04-28
Smart Images
Figure FT_1 
Figure FT_2 
Figure FT_3
Abstract
Description
Technical Field
[0001] This invention relates to the fields of nanomaterials and biomedical engineering technology, and in particular to a dye-sensitized rare-earth upconversion luminescent nanomaterial, its preparation method, and its application. Background Technology
[0002] Rare earth upconversion nanoparticles (UCNPs) can convert low-energy near-infrared light into high-energy visible or ultraviolet light through a multiphoton absorption process. They have advantages such as large anti-Stokes shift, strong tissue penetration depth, and low background fluorescence interference, showing broad application prospects in fields such as bioimaging, disease diagnosis and treatment.
[0003] However, traditional UCNPs and their preparation methods have the following inherent defects: (1) Small absorption cross section: The 4f-4f transition of rare earth ions is a parity-forbidden transition, which results in a low absorption cross section (usually only 1×10). -21 cm 2 -1×10 -20 cm 2 (1) Highly dependent on high-power lasers (such as 980nm or 808nm lasers) for excitation, resulting in high equipment costs and potential photothermal damage; (2) Poor water solubility: UCNPs prepared by thermal decomposition or coprecipitation are usually surface-modified with hydrophobic ligands such as oleic acid (OA), making them difficult to disperse stably in aqueous or biological media, which greatly limits their biological applications; (3) Insufficient photostability: Although near-infrared organic dyes (such as indocyanine green ICG) sensitization can significantly improve the light absorption capacity of UCNPs (the absorption cross-section is increased by about 10%), the photostability of UCNPs is still limited. 4 However, dye molecules are prone to photobleaching under light, and in aqueous phase, they are susceptible to aggregation-induced quenching (ACQ) effect, which leads to a sharp drop in luminescence efficiency.
[0004] Existing technologies attempt to improve the water solubility of UCNPs using strategies such as polymer coating, silica coating, or protein modification, but these methods still have the following shortcomings: (1) they cannot simultaneously solve the problem of dye photobleaching; (2) the coating layer may hinder energy transfer and reduce luminescence efficiency; (3) the functionalization modification steps are cumbersome and have poor reproducibility. In addition, although dye sensitization strategies (such as ICG sensitization) can combine the light-harvesting ability of dyes with the luminescence properties of rare earth ions through Förster resonance energy transfer (FRET) or Dexter energy transfer (DET) mechanisms, the stability of dye molecules remains a technical bottleneck. Summary of the Invention
[0005] The present invention aims to at least solve one of the aforementioned technical problems existing in the prior art. Therefore, one objective of the present invention is to provide a dye-sensitized rare-earth upconversion luminescent nanomaterial.
[0006] The second objective of this invention is to provide a method for preparing such dye-sensitized rare-earth upconversion luminescent nanomaterials.
[0007] The third objective of this invention is to provide a surface-functionalized dye-sensitized rare-earth upconversion nanomaterial.
[0008] The fourth objective of this invention is to provide a method for preparing such surface-functionalized dye-sensitized rare-earth upconversion nanomaterials.
[0009] The fifth objective of this invention is to provide dye-sensitized rare-earth upconversion luminescent nanomaterials, or the application of surface-functionalized dye-sensitized rare-earth upconversion nanomaterials.
[0010] To achieve the above objectives, the technical solution adopted by the present invention is as follows: A first aspect of the present invention provides a dye-sensitized rare-earth upconversion luminescent nanomaterial, comprising: a rare-earth-doped upconversion nanoparticle core, an unsaturated fatty acid salt self-assembled shell, and a near-infrared dye modified on the surface of the rare-earth-doped upconversion nanoparticle.
[0011] In some embodiments of the present invention, the diameter of the rare earth-doped upconversion nanoparticle core is 20-30 nm; the thickness of the unsaturated fatty acid salt self-assembled shell is 2.5-4.0 nm.
[0012] In some preferred embodiments of the present invention, the diameter of the rare earth-doped upconversion nanoparticle core is 24-27 nm; and the thickness of the unsaturated fatty acid salt self-assembled shell is 2.9-4.7 nm.
[0013] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles are selected from at least one of β-NaYF4:Yb,Er, β-NaYF4:Yb,Tm, and β-NaYF4:Yb,Ho.
[0014] In some preferred embodiments of the present invention, the rare earth-doped upconversion nanoparticles are β-NaYF4:Yb20%, Er2%.
[0015] In some embodiments of the present invention, the unsaturated fatty acid salt is selected from at least one of oleate, linoleate, linolenic acid, palmitate, and erucic acid.
[0016] In some preferred embodiments of the present invention, the unsaturated fatty acid salt is selected from at least one of sodium oleate (SO), sodium linoleate (SL), and sodium linolenate (SLn).
[0017] In some more preferred embodiments of the present invention, the unsaturated fatty acid salt is sodium linoleate.
[0018] In some embodiments of the present invention, the near-infrared dye is selected from cyanine dyes that have absorption in the wavelength range of 720-850 nm.
[0019] In some preferred embodiments of the present invention, the near-infrared dye is indocyanine green (ICG).
[0020] The second aspect of the present invention provides a method for preparing the dye-sensitized rare-earth upconversion luminescent nanomaterials described in the first aspect of the present invention, comprising the following steps: S1. Synthesize rare earth-doped upconversion nanoparticles with oleic acid surface-modified, remove oleic acid by ligand exchange, and obtain hydrophilic particles. S2. Disperse the hydrophilic particles and near-infrared dye in an organic solvent and react to obtain dye-sensitized nanoparticles. S3. The dye-sensitized nanoparticles are mixed and reacted with unsaturated fatty acid salts in an aqueous medium to obtain the dye-sensitized rare earth upconversion luminescent nanomaterials.
[0021] In some embodiments of the present invention, in step S1, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared by a method comprising the following steps: LnAc3·xH2O (Ln=Er, Tm, Ho), oleic acid, and 1-octadecene were mixed and heated under an inert atmosphere to remove water. After a first heating reaction, a methanol solution of sodium hydroxide and ammonium fluoride was added, and the mixture was heated to remove water and oxygen. After a second heating reaction, rare earth-doped upconversion nanoparticles with surface-modified oleic acid were obtained.
[0022] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared in a ratio of LnAc3·xH2O, oleic acid and 1-octadecene of 1 mmol: (6-9) mL: (12-18) mL.
[0023] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared, and the heating to remove water is carried out at a temperature of 80-120°C for 8-12 minutes.
[0024] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared, and the temperature of the first heating reaction is 130-190°C and the time is 20-40 min.
[0025] In some embodiments of the present invention, in the preparation of rare earth-doped upconversion nanoparticles with surface-modified oleic acid, the concentration of sodium hydroxide in the methanol solution of sodium hydroxide and ammonium fluoride is 0.008-0.012 g / mL, and the concentration of ammonium fluoride is 0.011-0.018 g / mL; the volume ratio of the methanol solution of sodium hydroxide and ammonium fluoride to oleic acid is 10:(6-9).
[0026] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared, and the temperature for heating to remove water and oxygen is 95-145°C and the time is 8-12 min.
[0027] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared at a temperature of 240-360°C for a time of 0.8-1.2 h.
[0028] In some embodiments of the present invention, the rare earth-doped upconversion nanoparticles with surface-modified oleic acid are prepared, and after the secondary heating reaction is completed, the product is collected by centrifugation and dispersed in cyclohexane.
[0029] In some embodiments of the present invention, the ligand exchange removal of oleic acid in step S1 specifically involves: A dimethyl sulfoxide solution of tetrafluoroborate nitrite was mixed with a cyclohexane solution of rare earth-doped upconversion nanoparticles with surface-modified oleic acid. The mixture was shaken and allowed to stand overnight to allow the nanoparticles to enter the lower dimethyl sulfoxide layer. The upper cyclohexane layer was removed, and the nanoparticles were collected by centrifugation to obtain the hydrophilic particles.
[0030] In some embodiments of the present invention, the concentration of the dimethyl sulfoxide solution of nitrosotetrafluoroborate is 4-6 mg / mL; and the concentration of the cyclohexane solution of rare earth-doped upconversion nanoparticles with surface-modified oleic acid is 0.08-0.12 mol / mL.
[0031] In some embodiments of the present invention, in step S2, the concentration of the near-infrared dye in the reaction system is 0.5-7.5 μg / mL.
[0032] In some preferred embodiments of the present invention, in step S2, the concentration of the near-infrared dye in the reaction system is 2.5-5.0 μg / mL.
[0033] In some embodiments of the present invention, in step S3, the concentration of the unsaturated fatty acid salt in the reaction system is 0.025-0.075 mol / L.
[0034] In some preferred embodiments of the present invention, in step S3, the concentration of the unsaturated fatty acid salt in the reaction system is 0.025-0.05 mol / L.
[0035] In some embodiments of the present invention, in step S3, the reaction temperature is 10-60°C and the time is 5-15 min.
[0036] In some preferred embodiments of the present invention, in step S3, the reaction temperature is 10-30°C and the time is 8-12 min.
[0037] A third aspect of the present invention provides a surface-functionalized dye-sensitized rare-earth upconversion nanomaterial, including the dye-sensitized rare-earth upconversion luminescent nanomaterial described in the first aspect of the present invention; wherein, the outer surface of the unsaturated fatty acid salt self-assembled shell of the dye-sensitized rare-earth upconversion luminescent nanomaterial has active functional groups exposed.
[0038] In some embodiments of the present invention, the active functional group is selected from at least one of -NH2, -COOH, -SH, and -OH.
[0039] A fourth aspect of the present invention provides a method for preparing the surface-functionalized dye-sensitized rare-earth upconversion nanomaterials described in the third aspect of the present invention, comprising the following steps: The dye-sensitized rare-earth upconversion luminescent nanomaterials were placed in an aqueous medium containing unsaturated fatty acid salts and functionalizing reagents to obtain the surface-functionalized dye-sensitized rare-earth upconversion nanomaterials.
[0040] In some embodiments of the present invention, in the aqueous medium containing unsaturated fatty acid salts and functionalizing reagents, the concentration of unsaturated fatty acid salts is 0.4-0.6 mg / mL, and the concentration of functionalizing reagents is 8 wt%-10 wt%.
[0041] In some embodiments of the present invention, the solid-liquid ratio of the dye-sensitized rare-earth upconversion luminescent nanomaterial to the aqueous medium containing unsaturated fatty acid salts and functionalizing reagents is 1 mg: (2-3) mL.
[0042] In some embodiments of the present invention, the functionalizing agent includes at least one of dodecyl alcohol, dodecylamine, dodecyl mercaptan, and lauric acid.
[0043] The fifth aspect of the present invention provides the application of the dye-sensitized rare-earth upconversion luminescent nanomaterials described in the first aspect of the present invention, or the surface-functionalized dye-sensitized rare-earth upconversion nanomaterials described in the third aspect of the present invention, in bioimaging, biodetection, or photodynamic therapy.
[0044] Compared with the prior art, the beneficial effects of the present invention are: 1) The dye-sensitized rare-earth upconversion nanoluminescent material provided by this invention overcomes the core bottlenecks of poor water solubility and weak photostability of traditional materials through the self-assembled shell coating of unsaturated fatty acid salts. This unsaturated fatty acid salt coating layer imparts excellent water dispersibility to the material through hydrophilic groups, achieving a Zeta potential of -57.9 mV and stable storage for over 30 days. Its hydrophobic segments and unsaturated double bonds effectively block water and oxygen penetration and quench singlet oxygen, resulting in a photobleaching half-life of 260 min, more than 86.7 times longer than conventional coating materials, and exhibiting excellent photostability. Benefiting from the broad-spectrum (720-850 nm) strong absorption characteristics of near-infrared dyes, this material has been successfully applied in low-power near-infrared light-emitting diodes (NIR LEDs), with power densities as low as 100 mW / cm². 2 The highly efficient upconversion luminescence excited by lasers completely eliminates the dependence on high-cost, high-risk lasers; 2) The surface-functionalized dye-sensitized rare earth upconversion nanomaterials provided by this invention introduce active functional groups such as amino and carboxyl groups during the coating process of dye-sensitized rare earth upconversion luminescent nanomaterials, which facilitates coupling with various biomolecules and expands their applications in targeted bioimaging, high-sensitivity biodetection and drug delivery. 3) The method for preparing dye-sensitized rare-earth upconversion luminescent nanomaterials provided by this invention has a simple preparation process, low cost, and significant industrialization prospects. Attached Figure Description
[0045] Figure 1 The image shows the X-ray diffraction pattern of the hydrophilic particles UCNPs in Example 1. Figure 2 This is a TEM image of the hydrophilic UCNPs in Example 1; Figure 3 Fourier transform infrared spectra of rare earth-doped upconversion nanoparticles OA-UCNPs with surface-modified oleic acid and dye-sensitized nanoparticles ICG-UCNPs in Example 1. Figure 4 This is a schematic diagram of the preparation of dye-sensitized nanoparticles ICG-UCNPs and dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn in Example 1. Figure 5 This is a TEM image of the dye-sensitized rare-earth upconversion luminescent nanomaterial ICG-UCNPs@SLn in Example 1; Figure 6 Fourier transform infrared spectra of dye-sensitized nanoparticles ICG-UCNPs and dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn in Example 1. Figure 7The Zeta potential diagram (a) and hydration particle size distribution curve (b) of dye-sensitized nanoparticles ICG-UCNPs and dye-sensitized rare earth upconversion luminescent nanomaterials ICG-UCNPs@SLn in Example 1 are shown. Figure 8 These are photographs of the solution state of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3; Figure 9 The Zeta potential diagrams are shown for the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3. Figure 10 The fluorescence emission spectra (a) and relative intensities (b) of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3 are shown. Figure 11 The image shows the decay curves (a) and photobleaching half-life (b) of the upconversion luminescence intensity of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3 under 808nm laser irradiation. Figure 12 The changes in absorbance at 410 nm after DPBF was excited by 808 nm laser for different times and then mixed with the nanomaterials in Examples 1(a), 2(b) and 3(c), and the changes in absorbance of DPBF in Examples 1-3 (d); Figure 13 The mechanism of water molecule-induced luminescence quenching of β-NaYF4:Yb20%,Er2% is shown in (a), the upconversion emission spectra of Ligand-free and ICG-UCNPs@SLn under 980nm laser excitation are shown in (b), the relative intensity is shown in (c), and the fluorescence lifetime at 540nm is shown in (d). Figure 14 The effects of the amount of near-infrared dye, the amount of unsaturated fatty acid salt, and the reaction temperature during the coating process on the luminescence properties of dye-sensitized rare-earth upconversion luminescent nanomaterials were investigated. Figure 15 The image shows a TEM image of dye-sensitized upconversion nanoparticles coated with mesoporous SiO2, as shown in Comparative Example 1. Figure 16 TEM image of dye-sensitized upconversion nanoparticles coated with Pluronic F-127 in Comparative Example 2; Figure 17 TEM image of the dye-sensitized upconversion nanoparticles coated with DSPE-PEG2000 in Comparative Example 3; Figure 18 The upconversion emission spectrum (a), luminescence photograph (b), relative fluorescence intensity diagram (c), and fluorescence lifetime (d) at 540 nm under 980 nm laser excitation are shown for the materials in Example 1 and Comparative Examples 1-3. Figure 19The upconversion luminescence intensity decay curves (a), photobleaching half-life (b), and luminescence photographs (c) of the materials in Example 1 and Comparative Examples 1-3 before and after 30 minutes of continuous irradiation are shown. Figure 20 Microscopic imaging of HepG2 cells stained with ICG-UCNPs@SLn at 0 min (a), 15 min (b), and 30 min (c) under continuous NIR LED excitation, and changes in fluorescence signal intensity (d); Figure 21 The FT-IR spectra of surface-functionalized dye-sensitized rare-earth upconversion nanomaterials @SLn-NH2 and @SLn-SH in Application Example 1 and Application Example 2 are shown. Figure 22 The diagram shows the coupling of @SLn-NH2 and RhB in Application Example 1 (a), the emission spectrum of ICG-UCNPs@SLn and the absorption spectrum of RhB (b), and the upconversion steady-state emission spectrum before and after the coupling of @SLn-NH2 and RhB (c). Figure 23 The diagram shows the coupling of @SLn-SH and AuNPs in Application Example 2 (a), the emission spectrum of ICG-UCNPs@SLn and the absorption spectrum of AuNPs (b), and the upconversion steady-state emission spectrum before and after coupling of @SLn-SH and AuNPs (c). Detailed Implementation
[0046] The present invention will be further described in detail below through specific embodiments. Unless otherwise specified, the raw materials, reagents, or apparatus used in the embodiments and comparative examples are all available from conventional commercial sources or can be obtained by existing technical methods. Unless otherwise specified, the experimental or testing methods are conventional methods in the art.
[0047] Example 1 This embodiment prepares a dye-sensitized rare-earth upconversion luminescent nanomaterial, and the steps are as follows: S11. 1 mmol ErAc3·xH2O, 7.5 mL oleic acid, and 15 mL 1-octadecene were sequentially added to a 100 mL three-necked flask. The reaction system was heated to 100 °C and held for 10 min under a nitrogen atmosphere to remove water from the system. Then, the temperature was increased to 160 °C and held for 30 min to promote uniform mixing and reaction of the precursors. After the reaction was completed, the system was cooled to room temperature. Then, 10 mL of a methanol solution containing 0.1 g sodium hydroxide and 0.148 g ammonium fluoride was added to the reaction system. The system was heated to 120 °C and held for 10 min to completely remove water and oxygen from the solvent. The temperature was then increased to 300 °C and held for 1 h under nitrogen protection to complete the crystal growth process. After the reaction was completed, the system was allowed to cool naturally to room temperature. Finally, the reaction product was collected by centrifugation at 7500 rpm for 5 min and dispersed in 10 mL cyclohexane for subsequent use, yielding rare earth-doped upconversion nanoparticles with oleic acid modified on the surface, denoted as OA-UCNPs. S12. A 5 mg / mL solution of nitrosotetrafluoroborate in dimethyl sulfoxide was mixed with a 0.1 mol / mL solution of rare earth-doped upconversion nanoparticles with surface-modified oleic acid in cyclohexane. The mixture was gently shaken for 10 min and allowed to stand overnight to allow the nanoparticles to be fully redispersed from the upper cyclohexane layer to the lower dimethyl sulfoxide layer. After removing the upper cyclohexane layer, the mixture was centrifuged at 11,000 rpm for 15 min, and the nanoparticles dispersed in dimethyl sulfoxide were collected to obtain hydrophilic particles, denoted as UCNPs. S21. Dissolve indocyanine green in dimethyl sulfoxide to prepare an indocyanine green solution with a concentration of 0.1 mg / mL; then mix 2 mL of the 2 mg / mL hydrophilic particle dimethyl sulfoxide solution with the indocyanine green solution to obtain dye-sensitized nanoparticles, denoted as ICG-UCNPs. S31. Dissolve sodium linoleate in deionized water to prepare sodium linoleate aqueous solution; then mix 1 mL of sodium linoleate aqueous solution with 200 μL of 2 mg / mL dichloromethane solution of dye-sensitized nanoparticles, stir for 10 min, and centrifuge at 11000 rpm for 15 min to obtain dye-sensitized rare earth upconversion luminescent nanomaterials, denoted as ICG-UCNPs@SLn.
[0048] Example 2 This embodiment prepares a dye-sensitized rare-earth upconversion luminescent nanomaterial, and the steps are as follows: S11. 1 mmol ErAc3·xH2O, 7.5 mL oleic acid, and 15 mL 1-octadecene were sequentially added to a 100 mL three-necked flask. The reaction system was heated to 100 °C and held for 10 min under a nitrogen atmosphere to remove water from the system. Then, the temperature was increased to 160 °C and held for 30 min to promote uniform mixing and reaction of the precursors. After the reaction was completed, the system was cooled to room temperature. Then, 10 mL of a methanol solution containing 0.1 g sodium hydroxide and 0.148 g ammonium fluoride was added to the reaction system. The system was heated to 120 °C and held for 10 min to completely remove water and oxygen from the solvent. The temperature was then increased to 300 °C and held for 1 h under nitrogen protection to complete the crystal growth process. After the reaction was completed, the system was allowed to cool naturally to room temperature. Finally, the reaction product was collected by centrifugation at 7500 rpm for 5 min and dispersed in 10 mL cyclohexane for subsequent use, yielding rare earth-doped upconversion nanoparticles with oleic acid modified on the surface, denoted as OA-UCNPs. S12. A 5 mg / mL solution of nitrosotetrafluoroborate in dimethyl sulfoxide was mixed with a 0.1 mol / mL solution of rare earth-doped upconversion nanoparticles with surface-modified oleic acid in cyclohexane. The mixture was gently shaken for 10 min and allowed to stand overnight to allow the nanoparticles to be fully redispersed from the upper cyclohexane layer to the lower dimethyl sulfoxide layer. After removing the upper cyclohexane layer, the mixture was centrifuged at 11,000 rpm for 15 min, and the nanoparticles dispersed in dimethyl sulfoxide were collected to obtain hydrophilic particles, denoted as UCNPs. S21. Dissolve indocyanine green in dimethyl sulfoxide to prepare an indocyanine green solution with a concentration of 0.1 mg / mL; then mix 2 mL of the 2 mg / mL hydrophilic particle dimethyl sulfoxide solution with the indocyanine green solution to obtain dye-sensitized nanoparticles, denoted as ICG-UCNPs. S31. Dissolve sodium oleate in deionized water to prepare an aqueous solution of sodium oleate; then mix 1 mL of the sodium oleate aqueous solution with 200 μL of a 2 mg / mL solution of dye-sensitized nanoparticles in dimethyl sulfoxide, stir for 10 min, and centrifuge at 11000 rpm for 15 min to obtain dye-sensitized rare earth upconversion luminescent nanomaterials, denoted as ICG-UCNPs@SO.
[0049] Example 3 This embodiment prepares a dye-sensitized rare-earth upconversion luminescent nanomaterial, and the steps are as follows: S11. 1 mmol ErAc3·xH2O, 7.5 mL oleic acid, and 15 mL 1-octadecene were sequentially added to a 100 mL three-necked flask. The reaction system was heated to 100 °C and held for 10 min under a nitrogen atmosphere to remove water from the system. Then, the temperature was increased to 160 °C and held for 30 min to promote uniform mixing and reaction of the precursors. After the reaction was completed, the system was cooled to room temperature. Then, 10 mL of a methanol solution containing 0.1 g sodium hydroxide and 0.148 g ammonium fluoride was added to the reaction system. The system was heated to 120 °C and held for 10 min to completely remove water and oxygen from the solvent. The temperature was then increased to 300 °C and held for 1 h under nitrogen protection to complete the crystal growth process. After the reaction was completed, the system was allowed to cool naturally to room temperature. Finally, the reaction product was collected by centrifugation at 7500 rpm for 5 min and dispersed in 10 mL cyclohexane for subsequent use, yielding rare earth-doped upconversion nanoparticles with oleic acid modified on the surface, denoted as OA-UCNPs. S12. A 5 mg / mL solution of nitrosotetrafluoroborate in dimethyl sulfoxide was mixed with a 0.1 mol / mL solution of rare earth-doped upconversion nanoparticles with surface-modified oleic acid in cyclohexane. The mixture was gently shaken for 10 min and allowed to stand overnight to allow the nanoparticles to be fully redispersed from the upper cyclohexane layer to the lower dimethyl sulfoxide layer. After removing the upper cyclohexane layer, the mixture was centrifuged at 11,000 rpm for 15 min, and the nanoparticles dispersed in dimethyl sulfoxide were collected to obtain hydrophilic particles, denoted as UCNPs. S21. Dissolve indocyanine green in dimethyl sulfoxide to prepare an indocyanine green solution with a concentration of 0.1 mg / mL; then mix 2 mL of the 2 mg / mL hydrophilic particle dimethyl sulfoxide solution with the indocyanine green solution to obtain dye-sensitized nanoparticles, denoted as ICG-UCNPs. S31. Dissolve sodium linoleate in deionized water to prepare a sodium linoleate aqueous solution; then mix 1 mL of sodium linoleate aqueous solution with 200 μL of a 2 mg / mL dimethyl sulfoxide solution of dye-sensitized nanoparticles, stir for 10 min, and centrifuge at 11000 rpm for 15 min to obtain dye-sensitized rare earth upconversion luminescent nanomaterials, denoted as ICG-UCNPs@SL.
[0050] Comparative Example 1 This comparative example prepares a mesoporous SiO2-coated dye-sensitized upconversion nanoparticle, and the steps are as follows: 1) 1 mmol of upconversion nanoparticles with oleic acid ligands on the surface (OA-UCNPs, preparation method as described in Example 1) was dissolved in 20 mL of cyclohexane and sonicated for 30 min to ensure uniform particle dispersion. After stirring at room temperature for 30 min, 2.5 mL of cyclohexane, 2.5 mL of Triton X-100, 500 μL of deionized water and 250 μL of ammonia were added to the reaction system in sequence, and the mixture was stirred at room temperature for 1 h. Then, 100 μL of tetraethoxysilane was added to the reaction system and the mixture was stirred overnight (at least 14 h). After the reaction was completed, 12.5 mL of acetone was added to terminate the reaction. The reaction product was collected by centrifugation at 7500 rpm for 5 min and washed twice with anhydrous ethanol. Finally, the product was dissolved in deionized water to obtain mesoporous SiO2-coated upconversion nanoparticles, denoted as UCNPs@mSiO2. 2) The mesoporous SiO2-coated upconversion nanoparticles were mixed with a certain amount of indocyanine green and stirred in 20 mL of deionized water for 12 h. The mixture was then centrifuged and washed to remove unreacted indocyanine green, resulting in dye-sensitized upconversion nanoparticles coated with mesoporous SiO2, denoted as ICG-UCNPs@mSiO2.
[0051] Comparative Example 2 In this comparative example, a dye-sensitized upconversion nanoparticle coated with pluronic F-127 was prepared, and the steps are as follows: 0.1 mmol of oleic acid-coated upconversion nanoparticles (OA-UCNPs, prepared according to Example 1) and a certain amount of indocyanine green were dispersed in 10 mL of dichloromethane and stirred vigorously at room temperature for 1 h to ensure thorough mixing. Subsequently, 4 mL of dichloromethane solution of Pluronic F-127 (130 mg) was added to the mixture, and the mixture was sonicated for 2 min. Then, the reaction system was stirred at room temperature overnight to allow the solvent to evaporate completely. After the solvent evaporated, 5 mL of deionized water was added to the system and centrifuged (12000 rpm, 5°C, 15 min) to remove excess indocyanine green. Finally, the obtained product was dissolved in 5 mL of deionized water to obtain dye-sensitized upconversion nanoparticles coated with Pluronic F-127, denoted as ICG-UCNPs@F-127.
[0052] Comparative Example 3 This comparative example prepares dye-sensitized upconversion nanoparticles coated with phospholipid-polyethylene glycol (DSPE-PEG2000), and the steps are as follows: Oleic acid-coated upconversion nanoparticles (OA-UCNPs, preparation method as described in Example 1) were dissolved in 5 mL of chloroform solution and the material was uniformly dispersed by ultrasound. Then, a certain amount of indocyanine green and DSPE-PEG2000 (10 mg) were added sequentially. The mixture was stirred overnight (at least 12 h) under light-protected conditions. Excess solvent was removed by rotary evaporation. The collected solid was washed with deionized water and finally redispersed in deionized water to obtain DSPE-PEG2000-coated dye-sensitized upconversion nanoparticles, denoted as ICG-UCNPs@DSPE-PEG2000.
[0053] Material characterization and performance testing Figure 1 The image shows the X-ray diffraction pattern of the hydrophilic particles UCNPs in Example 1. Figure 1 It can be seen that the hydrophilic particles UCNPs prepared in Example 1 are β-NaYF4:Yb20%,Er2% upconversion nanoparticles. Their diffraction pattern is highly consistent with the standard diffraction card (JCPDS 16-0334) of hexagonal phase β-NaYF4, indicating that the hydrophilic particles UCNPs prepared in Example 1 have a pure hexagonal phase crystal structure.
[0054] Figure 2 This is a TEM image of the hydrophilic UCNPs in Example 1, created by... Figure 2 It can be seen that the hydrophilic particles UCNPs prepared in Example 1 exhibit a uniform nanoparticle morphology, and the average particle size is calculated to be 25.6±1.2nm, indicating that the material has good size uniformity.
[0055] Figure 3 The Fourier transform infrared spectra of rare earth-doped upconversion nanoparticles OA-UCNPs and dye-sensitized nanoparticles ICG-UCNPs with surface-modified oleic acid in Example 1 are shown below. Figure 3 It can be seen that the rare earth-doped upconversion nanoparticles OA-UCNPs with surface-modified oleic acid synthesized by thermal coprecipitation in Example 1 have a high efficiency of 2400-3100 cm⁻¹. -1 A strong CH stretching vibration peak was observed within the range, which was attributed to the presence of surface oleic acid ligands. After the oleic acid ligands were stripped by ammonium fluoride and coordinated with indocyanine green, the CH stretching vibration peak was significantly weakened, and the peak at 1021 cm⁻¹ was also reduced. -1 BF4 appeared at the location - The characteristic absorption peak of the ion is at 1647 cm⁻¹. -1 The appearance of stretching vibration peaks of C=N bonds confirms the successful stripping of oleic acid ligands and the effective coordination of indocyanine green.
[0056] Figure 4This is a schematic diagram illustrating the preparation of dye-sensitized nanoparticles ICG-UCNPs and dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn in Example 1. Figure 4 It is known that when dye-sensitized nanoparticles ICG-UCNPs are dispersed in a water / dichloromethane mixture, the sulfonic acid groups of indocyanine green molecules are coordinated with the hydrophilic UCNPs, and the remaining conjugated structure exhibits strong hydrophobicity. This results in the dye-sensitized nanoparticles ICG-UCNPs being mainly distributed in the dichloromethane phase and exhibiting bright indocyanine green upconversion luminescence under 808 nm laser excitation. After being coated with sodium linoleate, the dye-sensitized rare earth upconversion luminescent nanomaterials are mainly dispersed in the aqueous phase due to the similarity and compatibility between the long carbon chain in the sodium linoleate molecule and indocyanine green, as well as the hydrophilic effect of the carboxyl group. They still maintain significant green upconversion luminescence under 808 nm laser excitation, confirming that the optical properties of the material are not significantly affected during the coating process. This result proves that the sodium linoleate coating strategy can achieve both water solubility and effectively maintain the optical properties of the material.
[0057] The dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn prepared in Example 1 were observed by transmission electron microscopy using phosphotungstic acid negative staining technique. Figure 5 The image shows a TEM image of the dye-sensitized rare-earth upconversion luminescent nanomaterial ICG-UCNPs@SLn in Example 1. Figure 5 (a) in the image is the global TEM map. Figure 5 (b) in the image is a magnified view of a portion of (a). Figure 5 It can be seen that the dye-sensitized rare earth upconversion luminescent nanomaterial ICG-UCNPs@SLn has a core-shell structure with an overall size of 32.1±1.8nm and a surface shell thickness of 3.3±0.4nm, proving that sodium linoleate was successfully coated on the surface of the dye-sensitized nanoparticles ICG-UCNPs.
[0058] Figure 6 The Fourier transform infrared spectra of dye-sensitized nanoparticles ICG-UCNPs and dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn in Example 1 are shown below. Figure 6 It can be seen that, compared with dye-sensitized nanoparticles ICG-UCNPs, dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn exhibit better performance in the 2400-3100 cm⁻¹ range. -1 The intensity of the CH stretching vibration peak is significantly enhanced within the range, and reaches 1558 cm⁻¹. -1 The presence of a characteristic absorption peak attributable to the C=C bond in the sodium linoleate molecule further confirms the successful encapsulation of sodium linoleate.
[0059] The surface charge characteristics of the material were analyzed by zeta potential testing, and the hydrated particle size was determined by dynamic light scattering (DLS) technology. Figure 7 The figures (a) and (b) are the Zeta potential diagram and hydration particle size distribution curve of dye-sensitized nanoparticles ICG-UCNPs and dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn in Example 1. Figure 7 It can be seen that, compared with dye-sensitized nanoparticles ICG-UCNPs, the surface potential of the material after coating with sodium linoleate changed from +12.1mV to -57.9mV. The reversal of positive and negative potentials not only verifies the successful anchoring of carboxylate ions in sodium linoleate molecules, but also indicates that the material surface has strong negative charge, which is conducive to maintaining a stable water-soluble system through electrostatic repulsion. After coating with sodium linoleate, the hydrated particle size of the material increased from the initial 43.8±2.1nm (hydrophilic particles UCNPs) to 164.2±8.6nm. The significant increase in hydrated particle size is directly related to the presence of the surface coating layer. This result provides kinetic evidence for the successful coating of sodium linoleate.
[0060] Figure 8 These are solution-state photographs of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3. Figure 8 It can be seen that the dye-sensitized nanoparticles treated with different unsaturated fatty acid salts (sodium linoleate, sodium oleate, sodium linoleate) in Examples 1-3 can be stably dispersed in aqueous solution and exhibit excellent light transmittance.
[0061] Figure 9 The diagram shows the Zeta potential of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3. Figure 9 It can be seen that the absolute value of the Zeta potential of the dye-sensitized rare earth upconversion luminescent nanomaterials in Examples 1-3 is relatively high, indicating that their surface charge is sufficient and the electrostatic repulsion between particles is significant, thus exhibiting good water solubility, which is consistent with the colloidal chemistry theory.
[0062] The fluorescence emission spectra of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3 were measured using a Horiba FluoroMax Plus fluorescence spectrometer, combined with 808 nm (CNI MDL-III-808, China) and 980 nm (CNI MDL-III-980, China) continuous lasers. The luminescence intensity of the sample can be obtained by calculating the integral area of the corresponding peak spectrum. Figure 10 The fluorescence emission spectra (a) and relative intensities (b) of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3 are shown below. Figure 10It can be seen that the materials coated with sodium linoleate, sodium oleate and sodium linoleate exhibit significant advantages in luminescence performance in aqueous phase. They show obvious upconversion luminescence peaks under 808nm laser excitation. Moreover, the luminescence intensity of the dye-sensitized rare earth upconversion luminescent nanomaterials in 1-3 is not much different. Based on the comprehensive analysis results of zeta potential and fluorescence intensity, sodium linoleate is preferred as the shell material.
[0063] The fluorescence intensity of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3 was recorded over time by continuous near-infrared light (808 nm) irradiation experiments. Figure 11 The figures (a) and (b) show the attenuation curves of the upconversion luminescence intensity of the dye-sensitized rare-earth upconversion luminescent nanomaterials in Examples 1-3 under 808 nm laser irradiation. Figure 11 It is known that sodium oleate, sodium linoleate, and sodium linolenate contain 1, 2, and 3 unsaturated C=C bonds, respectively. As the number of unsaturated C=C bonds in the unsaturated fatty acid salts increases, the photostability of the materials is significantly improved. The photobleaching half-lives of the materials in Examples 1-3 are 260 min, 109 min, and 205 min, respectively, indicating that the increase in the number of unsaturated C=C bonds can effectively inhibit the oxidation of indocyanine green dye by singlet oxygen, thereby significantly improving the photostability of the materials.
[0064] Using 1,3-diphenylisobenzofuran (DPBF) as the singlet oxygen ( 1 O2) detection reagent: DPBF was dissolved in dimethyl sulfoxide, and 100 μg of DPBF was added to the dye-sensitized rare earth upconversion luminescent nanomaterials in Examples 1-3 respectively. Then, the change in the intensity of the ultraviolet absorption peak at 410 nm of DPBF molecules was detected under 808 nm laser excitation from 0 to 30 minutes. Figure 12 The changes in absorbance at 410 nm after DPBF was excited by an 808 nm laser for different times and then mixed with the nanomaterials in Examples 2(a), 3(b), and 1(c), as well as the changes in absorbance of DPBF in Examples 1-3 (d), are shown. Figure 12 It can be seen that as the number of C=C double bonds in unsaturated fatty acid salts increases from 1 to 3, the absorbance decay rate of DPBF at 410 nm decreases significantly, indicating that increasing the number of C=C double bonds in unsaturated fatty acid salts can effectively inhibit the decay of absorbance. 1 The generation of O2 significantly improves the material's resistance to photobleaching.
[0065] The fluorescence intensity in the visible light region of ligand-free upconversion nanoparticles and the dye-sensitized rare-earth upconversion luminescent nanomaterial ICG-UCNPs@SLn from Example 1 under 980 nm laser excitation was measured using fluorescence spectroscopy and fluorescence lifetime spectroscopy. The fluorescence lifetime was measured using an Edinburgh FLS-1000 fluorescence spectrophotometer. Pulsed excitation light was obtained by connecting external 980 nm (CNI MDL-III-980) and 808 nm continuous-wave lasers to obtain the lifetimes in the corresponding wavelength bands. The radiative decay time τ was calculated using the formula... In the formula, I(t) refers to the luminescence intensity at time t. Figure 13 The diagram shows the mechanism of water molecule-induced luminescence quenching of β-NaYF4:Yb20%,Er2% (a), the upconversion emission spectra (b) and relative intensities (c) of Ligand-free and ICG-UCNPs@SLn under 980 nm laser excitation, and the fluorescence lifetime at 540 nm (d). Figure 13 It can be seen that water molecules can affect Yb 3+ ( 2 F 5 / 2 → 2 F 7 / 2 ) and Er 3+ ( 4 H 11 / 2 → 4 F 9 / 2 , 4 S 3 / 2 → 4 F 9 / 2 The energy level transition of Yb significantly reduces the upconversion luminescence efficiency of NaYF4:Yb,Er. The luminescence intensity of the material coated with sodium linoleate is 5.8 times higher than that of the ligand-free sample, indicating that the unsaturated fatty acid salt coating layer effectively blocks water molecules from reaching Yb. 3+ -Er 3+ The quenching effect of energy transfer significantly enhances the upconversion luminescence performance of the material. Under 980nm laser excitation, the fluorescence lifetime of the material in Example 1 is significantly longer than that of the Ligand-free sample, indicating that the unsaturated fatty acid salt coating effectively isolates water molecules and dissolved oxygen, reducing the amount of oxygen absorbed. 1 The generation of O2 and its oxidative damage to indocyanine green dye molecules significantly improve the photostability of the material.
[0066] In summary, the dye-sensitized rare-earth upconversion luminescent nanomaterials provided by this invention exhibit excellent anti-photobleaching properties for the following two reasons: (1) 1 O2 scavenging mechanism: The unsaturated C=C double bonds in unsaturated fatty acid salt molecules can efficiently capture singlet oxygen. 1 O2), forming stable epoxides, inhibiting1 O2 oxidative cleavage of indocyanine green molecules; (2) Physical isolation protection of unsaturated fatty acid salts: The unsaturated fatty acid salt coating not only blocks water molecule penetration, but also significantly reduces the diffusion rate of dissolved oxygen to the particle surface, thereby reducing 1 The generation of O2.
[0067] The dye-sensitized rare-earth upconversion luminescent nanomaterials were prepared using the method described in Example 1. The single-factor variables were controlled as follows: (1) In the step of modifying the hydrophilic particles with indocyanine green, the concentrations of indocyanine green in the reaction system were 0.5 μg / mL, 2.5 μg / mL, 5.0 μg / mL, 7.5 μg / mL, and 10.0 μg / mL, respectively; (2) In the step of coating the dye-sensitized nanoparticles with sodium linoleate, the concentrations of sodium linoleate in the reaction system were 0.005 mol / L, 0.010 mol / L, and 0.010 mol / L, respectively. 0.025 mol / L, 0.050 mol / L, 0.075 mol / L and 0.100 mol / L; (3) In the step of coating dye-sensitized nanoparticles with sodium linoleate, the reaction system temperatures were 10℃, 20℃, 30℃, 40℃, 50℃, 60℃, 70℃ and 80℃, respectively; to investigate the effects of the amount of near-infrared dye, the amount of unsaturated fatty acid salt and the reaction temperature of the coating process on the luminescence performance of dye-sensitized rare earth upconversion luminescent nanomaterials: Figure 14 The effects of the amount of near-infrared dye, the amount of unsaturated fatty acid salt, and the reaction temperature during the coating process on the luminescence properties of dye-sensitized rare-earth upconversion luminescent nanomaterials were investigated. Figure 14 (a) shows the upconversion emission spectra of dye-sensitized rare-earth upconversion luminescent nanomaterials sensitized with different amounts of indocyanine green. Figure 14 (b) shows the integral area of the upconversion emission spectrum of the dye-sensitized rare-earth upconversion luminescent nanomaterial as a function of indocyanine green concentration. Figure 14 (c) shows the upconversion emission spectra of dye-sensitized rare-earth upconversion luminescent nanomaterials coated with different amounts of sodium linoleate. Figure 14 In the figure, (d) represents the integral area of the upconversion emission spectrum of the dye-sensitized rare-earth upconversion luminescent nanomaterial as a function of sodium linoleate concentration. Figure 14 In the figure, (e) represents the upconversion emission spectrum of dye-sensitized rare-earth upconversion luminescent nanomaterials at different coating process reaction temperatures. Figure 14 In the figure, (f) represents the integral area of the upconversion emission spectrum of the dye-sensitized rare-earth upconversion luminescent nanomaterial as a function of the reaction temperature during the coating process. Figure 14It was found that the optimal dosage of indocyanine green was 2.5 μg / mL, and the luminescence intensity of the dye-sensitized rare-earth upconversion luminescent nanomaterials showed a trend of first increasing and then decreasing with the increase of indocyanine green dosage. This phenomenon is mainly due to the aggregation-induced quenching effect of the indocyanine green dye molecules. That is, at low concentrations, increasing the dosage of indocyanine green can improve the sensitization efficiency and enhance the upconversion luminescence intensity, but when the critical concentration is exceeded, the aggregation-induced quenching effect will lead to a decrease in luminescence intensity. The optimal coating amount of sodium linoleate was 0.025 mol / L, and its luminescence intensity change trend was similar to that caused by the dosage of indocyanine green. The mechanism may be that low concentrations of sodium linoleate form micelles to coat the dye-sensitized nanoparticles, and the coating effect improves with increasing concentration. However, the carboxyl groups in excessive sodium linoleate molecules will compete with indocyanine green for coordination sites, causing some indocyanine green molecules to fall off, ultimately reducing the luminescence intensity. In addition, it was verified that the reaction temperature during the coating process also affects the luminescence intensity. As the temperature increases, the luminescence intensity decreases, and the optimal reaction temperature was finally determined to be 10℃.
[0068] Figure 15 This is a TEM image of the dye-sensitized upconversion nanoparticles coated with mesoporous SiO2 in Comparative Example 1. Figure 16 This is a TEM image of the dye-sensitized upconversion nanoparticles coated with Pluronic F-127 in Comparative Example 2. Figure 17 The image shows a TEM image of the dye-sensitized upconversion nanoparticles coated with DSPE-PEG2000 in Comparative Example 3. Figures 15-17 It can be seen that coating with mesoporous SiO2, Pluronic F-127 and DSPE-PEG2000 all yielded dye-sensitized upconversion nanoparticles with uniform morphology, and the particle sizes were 44.5±2.7nm, 29±1.6nm and 27.6±1.3nm, respectively.
[0069] Figure 18 The upconversion emission spectrum (a), luminescence photograph (b), relative fluorescence intensity diagram (c), and fluorescence lifetime (d) at 540 nm under 980 nm laser excitation of the materials in Example 1 and Comparative Examples 1-3 are shown below. Figure 18 It can be seen that under 808nm laser excitation (0.5W / cm²), 2 In Example 1, the upconversion luminescence integral intensity of the dye-sensitized rare-earth upconversion luminescent nanomaterial ICG-UCNPs@SLn was 18 times, 35 times, and 5 times that of Comparative Examples 1, 2, and 3, respectively. The fluorescence lifetime was also significantly longer than the other three coating methods in Comparative Examples 1-3. This indicates that the interaction between the upconversion nanoparticles and water molecules is weakest under the unsaturated fatty acid salt coating method. This can be attributed to the water molecule isolation effect of the unsaturated fatty acid salt shell. The hydrophobic long chains of the unsaturated fatty acid salt form a dense barrier on the particle surface, blocking the interaction between water molecules and rare-earth ions (such as Yb).3+ Er 3+ The energy level coupling of the water molecules reduces the nonradiative transfer of energy to the vibrations of water molecules.
[0070] Through continuous laser irradiation experiments (808nm, 1.0W / cm²), 2 To evaluate the photobleaching resistance of materials prepared by different coating methods in Example 1 and Comparative Examples 1-3. Figure 19 The upconversion luminescence intensity decay curves (a), photobleaching half-life (b), and luminescence photographs (c) of the materials in Example 1 and Comparative Examples 1-3 before and after 30 minutes of continuous irradiation are shown below. Figure 19 It can be seen that the photobleaching half-life t of the dye-sensitized rare-earth upconversion luminescent nanomaterial ICG-UCNPs@SLn in Example 1 is... 1 / 2 =260min, significantly better than Comparative Example 1 (t 1 / 2 =3min), Comparative Example 2 (t 1 / 2 =3min) and Comparative Example 3 (t 1 / 2 =3min) of the sample.
[0071] As can be seen, the upconversion luminescence intensity and photostability of the dye-sensitized rare-earth upconversion luminescent nanomaterials coated with unsaturated fatty acid salts prepared in Example 1 are significantly better than those coated with mesoporous SiO2, Pluronic F-127, and DSPE-PEG2000 in existing technologies. This indicates that existing coating technologies lack the important unsaturated C=C chromium oxide. 1 O2 scavengers result in poor photostability, limiting their practical applications. However, the unsaturated fatty acid salt-coated dye-sensitized rare-earth upconversion luminescent nanomaterials provided by this invention overcome the trade-off between luminescence efficiency and stability in traditional coating materials through a unique synergistic mechanism of double bond protection and physical isolation, providing a new approach for the design of high-performance aqueous dye-sensitized upconversion probes.
[0072] Application Example 1 The dye-sensitized rare-earth upconversion luminescent nanomaterials ICG-UCNPs@SLn prepared in Example 1 were applied to bioimaging, and the steps are as follows: Using HepG2 cells as a model, before the experiment, HepG2 cells were cultured in a cell culture incubator at 37℃ and 5% CO2 until the cells reached a good state and 80% confluence within the microscope field of view. Then, 200 μL of ICG-UCNPs@SLn solution (1 μmol / mL) prepared in HEPES buffer was added to the cells and incubated overnight at 4℃. Finally, the cells were washed with HEPES buffer to prepare for subsequent imaging. The upconversion fluorescence signal of ICG-UCNPs@SLn in HepG2 cells was captured using a fluorescence inverted microscope under NIR LED excitation. The imaging system was a self-built NIR LED microscopy system with LED peak values of 780 / 805 nm and a power of 1.5 W / cm². 2 It is equipped with an Olympus IX73 microscope and a 60× oil immersion microscope.
[0073] Figure 20 Microscopic imaging of HepG2 cells stained with ICG-UCNPs@SLn at 0 min (a), 15 min (b), and 30 min (c) under continuous NIR LED excitation, and changes in fluorescence signal intensity (d). Figure 20 As can be seen, the dye-sensitized rare-earth upconversion luminescent nanomaterial ICG-UCNPs@SLn prepared in Example 1 is uniformly distributed in the cytoplasm and generates a strong and stable upconversion luminescence signal, indicating that it has successfully entered the cell and achieved efficient fluorescence imaging under low-power LED excitation. Furthermore, the fluorescence signal did not significantly decay under continuous irradiation from 0 to 30 minutes, indicating that the material has excellent resistance to photobleaching. Therefore, the dye-sensitized rare-earth upconversion luminescent nanomaterial provided by this invention has great application potential in bioimaging and biodetection.
[0074] Application Example 2 This application example provides a surface-functionalized dye-sensitized rare-earth upconversion nanomaterial, and the preparation steps are as follows: 1) Add 5 mg of sodium linoleate and 10 mL of deionized water to a 20 mL glass bottle and stir until a yellow transparent solution is formed. Then, add 10 wt% dodecylamine-ethanol solution to the above solution and stir for 30 min until the solution becomes clear. 2) 4 mg of the dye-sensitized rare earth upconversion luminescent nanomaterial ICG-UCNPs@SLn prepared in Example 1 was slowly added dropwise to the solution and stirred vigorously for 30 min. Subsequently, the product was collected by centrifugation (11000 rpm, 30 min), washed with deionized water and ethanol respectively, and the final product was dispersed in 2 mL of deionized water to obtain the surface-functionalized dye-sensitized rare earth upconversion nanomaterial, denoted as @SLn-NH2.
[0075] 5 mg of Rhodamine B (RhB), 7.6 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, and 11.3 mg of N-hydroxysuccinimide were dissolved in 0.5 mL of MES buffer (0.05 mol / L, pH=6.0) and stirred at room temperature for 1 h to activate RhB. Due to the limited solubility of RhB in water, the precipitate was separated by centrifugation (11000 rpm, 5 min) after activation and redispersed in 0.5 mL of deionized water. Subsequently, the activated RhB was mixed with @SLn-NH2 and stirred continuously at room temperature for 2 h to achieve covalent coupling. After the reaction was completed, the final product was collected by centrifugation (11000 rpm, 15 min) and washed repeatedly with deionized water to remove unreacted RhB. The obtained material was denoted as @SLn-RhB, and the product was resuspended in deionized water.
[0076] Application Example 3 This application example provides a surface-functionalized dye-sensitized rare-earth upconversion nanomaterial, and the preparation steps are as follows: 1) Add 5 mg of sodium linoleate and 10 mL of deionized water to a 20 mL glass bottle and stir until a yellow transparent solution is formed. Then, add 10 wt% dodecyl mercaptan-ethanol solution to the above solution and stir for 30 min until the solution becomes clear. 2) 4 mg of the dye-sensitized rare earth upconversion luminescent nanomaterial ICG-UCNPs@SLn prepared in Example 1 was slowly added dropwise to the solution and stirred vigorously for 30 min. Subsequently, the product was collected by centrifugation (11000 rpm, 30 min), washed with deionized water and ethanol respectively, and the final product was dispersed in 2 mL of deionized water to obtain the surface-functionalized dye-sensitized rare earth upconversion nanomaterial, denoted as @SLn-SH.
[0077] 2 mL of 4 mg / mL @SLn-SH solution was mixed with citric acid-stabilized AuNPs (purchased from Guangzhou Caijin Technology Co., Ltd.) at a mass ratio of 1:5. The mixture was stirred at room temperature for 12 h in Tris-HCl buffer at pH 8.0. After the reaction was completed, the product was separated by centrifugation (11000 rpm, 15 min) until the supernatant was colorless to ensure that unbound @SLn-SH was completely removed. The obtained material was denoted as @SLn-AuNPs. The product was washed several times with deionized water and finally redissolved in 2 mL of deionized water.
[0078] Figure 21 The FT-IR spectra of the surface-functionalized dye-sensitized rare-earth upconversion nanomaterials @SLn-NH2 and @SLn-SH in Application Examples 1 and 2 are obtained from... Figure 21 It can be seen that the FT-IR spectrum at 1150 cm⁻¹ -1The absorption band appearing at 2550-2600 cm⁻¹ corresponds to the stretching vibration of the CN bond, indicating the successful introduction of the amino group. -1 A weak and broad absorption peak was observed within the range, indicating the presence of thiol groups, thus confirming the successful preparation of surface-functionalized dye-sensitized rare-earth upconversion nanomaterials in Application Examples 1 and 2.
[0079] Figure 22 The diagram shows the coupling of @SLn-NH2 and RhB in Application Example 1 (a), the emission spectrum of ICG-UCNPs@SLn and the absorption spectrum of RhB (b), and the upconversion steady-state emission spectrum before and after coupling of @SLn-NH2 and RhB (c). Figure 23 The diagram shows the coupling of @SLn-SH and AuNPs in Example 2 (a), the emission spectrum of ICG-UCNPs@SLn and the absorption spectrum of AuNPs (b), and the upconversion steady-state emission spectrum before and after coupling of @SLn-SH and AuNPs (c). Figure 22 and 23 It is known that the coupling mechanism between @SLn-NH2 and RhB is as follows: 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide reacts with the carboxyl group on the surface of Rhodamine B to generate an unstable O-acylisourea intermediate. Subsequently, N-hydroxysuccinimide is added to react with this intermediate to form a stable NHS active ester. Finally, the active ester undergoes a nucleophilic substitution reaction with the amino group in @SLn-NH2 to form a stable amide bond (-CONH-). This reaction needs to be carried out in a MES buffer at pH=6.0 to avoid hydrolysis of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and ensure reaction efficiency. The coupling mechanism between @SLn-SH and AuNPs is as follows: the gold atoms on the surface of AuNPs and the thiol group form a stable Au-S bond through high-affinity covalent interaction. Furthermore, the emission spectrum of ICG-UCNPs@SLn in the 500-550 nm range overlaps with the absorption spectra of RhB and AuNPs, and the distance between the upconversion nanoparticles and RhB and AuNPs is <10 nm. According to the fluorescence resonance energy transfer mechanism (FRET), RhB and AuNPs can quench Er. 3+ The upconversion luminescence showed that the steady-state upconversion emission intensity decreased by approximately 50% and the fluorescence lifetime decreased by 43.0% and 46.1% before and after coupling, respectively. This indicates that the surface functionalization of ICG-UCNPs@SLn was successfully achieved through the coupling reaction mediated by EDC / NHS (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide / N-hydroxysuccinimide) and the covalent binding strategy of thiol-AuNPs. This study provides a reliable technical path for constructing multifunctional nanoprobes and lays an experimental foundation for their application in targeted imaging, drug delivery, and multimodal diagnosis and treatment.
Claims
1. A dye-sensitized rare-earth upconversion luminescent nanomaterial, characterized in that, include: The rare earth-doped upconversion nanoparticle core, the unsaturated fatty acid salt self-assembled shell, and the near-infrared dye modified on the surface of the rare earth-doped upconversion nanoparticle.
2. The dye-sensitized rare-earth upconversion luminescent nanomaterial according to claim 1, characterized in that, The diameter of the rare earth-doped upconversion nanoparticle core is 20-30 nm; the thickness of the unsaturated fatty acid salt self-assembled shell is 2.5-4.0 nm.
3. The dye-sensitized rare-earth upconversion luminescent nanomaterial according to claim 1, characterized in that, The rare earth-doped upconversion nanoparticles are selected from at least one of β-NaYF4:Yb,Er, β-NaYF4:Yb,Tm, and β-NaYF4:Yb,Ho. And / or, the unsaturated fatty acid salt is selected from at least one of oleate, linoleate, linolenic acid, palmitate, and erucic acid; And / or, the near-infrared dye is selected from cyanine dyes that have absorption in the wavelength range of 720-850 nm.
4. The method for preparing dye-sensitized rare-earth upconversion luminescent nanomaterials according to any one of claims 1-3, characterized in that, Includes the following steps: S1. Synthesize rare earth-doped upconversion nanoparticles with oleic acid surface-modified, remove oleic acid by ligand exchange, and obtain hydrophilic particles. S2. Disperse the hydrophilic particles and near-infrared dye in an organic solvent and react to obtain dye-sensitized nanoparticles. S3. The dye-sensitized nanoparticles are mixed and reacted with unsaturated fatty acid salts in an aqueous medium to obtain the dye-sensitized rare earth upconversion luminescent nanomaterials.
5. The method for preparing dye-sensitized rare-earth upconversion luminescent nanomaterials according to claim 4, characterized in that, In step S2, the concentration of the near-infrared dye in the reaction system is 0.5-7.5 μg / mL; And / or, in step S3, the concentration of the unsaturated fatty acid salt in the reaction system is 0.025-0.075 mol / L; And / or, in step S3, the reaction temperature is 10-60°C and the time is 5-15 min.
6. A surface-functionalized dye-sensitized rare-earth upconversion nanomaterial, characterized in that, The invention includes the dye-sensitized rare-earth upconversion luminescent nanomaterial according to any one of claims 1-3; wherein the outer surface of the unsaturated fatty acid salt self-assembled shell of the dye-sensitized rare-earth upconversion luminescent nanomaterial has active functional groups exposed.
7. The surface-functionalized dye-sensitized rare-earth upconversion nanomaterial according to claim 6, characterized in that, The active functional group is selected from at least one of -NH2, -COOH, -SH, and -OH.
8. The method for preparing surface-functionalized dye-sensitized rare-earth upconversion nanomaterials according to claim 6 or 7, characterized in that, Includes the following steps: The dye-sensitized rare-earth upconversion luminescent nanomaterials were placed in an aqueous medium containing unsaturated fatty acid salts and functionalizing reagents to obtain the surface-functionalized dye-sensitized rare-earth upconversion nanomaterials.
9. The method for preparing surface-functionalized dye-sensitized rare-earth upconversion nanomaterials according to claim 8, characterized in that, The functionalizing agent includes at least one of dodecyl alcohol, dodecylamine, dodecyl mercaptan, and lauric acid.
10. The dye-sensitized rare-earth upconversion luminescent nanomaterial according to any one of claims 1-3, or the surface-functionalized dye-sensitized rare-earth upconversion nanomaterial according to claim 6 or 7, in bioimaging, biodetection, or photodynamic therapy.