A near-infrared emitting europium-doped carbon dot and its preparation method and application

Abstract

The present invention provides near-infrared emitting europium-doped carbon dots, their preparation method, and applications. The preparation method comprises dissolving p-phenylenediamine, ethylenediaminetetraacetic acid, and europium chloride in water and performing a hydrothermal reaction; after the reaction, cooling the mixture to room temperature to obtain a reaction solution; and filtering, purifying the reaction solution through dialysis, and drying the mixture to obtain the near-infrared emitting europium-doped carbon dots. The near-infrared emitting europium-doped carbon dots, their preparation method, and applications are simple, and the resulting nanoprobes can achieve near-infrared emission without further modification.

Description

Technical Field

[0001] The present invention belongs to the technical field of nanomaterials, and in particular relates to near-infrared emitting europium-doped carbon dots and a preparation method and application thereof. Background Art

[0002] Carbon dots, due to their unique photostability, low toxicity, and good biocompatibility, have demonstrated great potential in medical treatment, imaging, antibacterial, and antimicrobial applications. Among them, near-infrared fluorescent carbon dots have low scattering and absorption in biological tissues, are less phototoxic to cells and tissues, and have higher tissue penetration, facilitating in situ imaging, labeling, and phototherapy of organisms. However, carbon dots primarily emit blue and green light (with wavelengths of 450-495nm and 520-570nm, respectively). The preparation of long-wavelength (yellow and red) luminescent carbon dots is difficult, resulting in poor imaging effects in cells and making them unsuitable for imaging.

[0003] In view of this, the present invention proposes a new carbon dot and its preparation method and application, which is a near-infrared emitting europium-doped carbon dot with a higher near-infrared fluorescence range. Summary of the Invention

[0004] The object of the present invention is to provide a method for preparing europium-doped carbon dots that emit near-infrared light. The preparation method is simple and the nanoprobes can achieve near-infrared emission without further modification.

[0005] In order to achieve the above objectives, the technical solutions adopted are:

[0006] A method for preparing near-infrared emitting europium-doped carbon dots comprises the following steps:

[0007] After dissolving p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride in water, a hydrothermal reaction is carried out; after the reaction is completed, the mixture is cooled to room temperature to obtain a reaction solution;

[0008] The reaction solution is filtered, dialyzed for purification, and dried to obtain the near-infrared emitting europium-doped carbon dots.

[0009] Furthermore, the mass ratio of p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride is 1-3:1-3:0.25-3.

[0010] Furthermore, the mass ratio of p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride is 3:3:1.

[0011] Furthermore, the temperature of the hydrothermal reaction is 180-210° C., and the time is 8-12 hours.

[0012] Furthermore, the filtration is performed using a 0.22 μm organic filter membrane.

[0013] Furthermore, the dialysis purification is performed by dialysis using a dialysis bag with a molecular weight cut-off of 500-1000Da.

[0014] Another object of the present invention is to provide near-infrared emitting europium-doped carbon dots prepared by the above-mentioned preparation method. Compared with imaging in the visible light band (400-650nm), near-infrared light (NIR-I, 700-900nm) can be used for cell imaging due to its deep penetration ability, high resolution, and low interference background.

[0015] A near-infrared emitting europium-doped carbon dot is prepared by the above-mentioned preparation method.

[0016] Furthermore, the fluorescence excitation spectrum of the europium-doped carbon dots is in the range of 300-700 nm, and the fluorescence emission spectrum is in the range of 600-800 nm.

[0017] Another object of the present invention is to provide an application of the above-mentioned europium-doped carbon dots, which can be used for cell imaging based on their high near-infrared fluorescence range, deep penetration ability, high resolution, and low interference background.

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] 1. In the technical solution of the present invention, p-phenylenediamine and ethylenediaminetetraacetic acid are used as precursors, and europium chloride is doped as a metal dopant. A deep purple europium-doped carbon dot solid is successfully obtained through a one-step hydrothermal reaction.

[0020] 2. In the technical solution of the present invention, the synthesis process of this method is simple, and the precursor can be directly purchased and is cheap.

[0021] 3. In the technical solution of the present invention, the synthesized near-infrared emitting europium-doped carbon dots have good water dispersibility, stable structure, uniform particle size, rich surface groups, and excellent optical properties. They have good biocompatibility and can be used as fluorescent imaging materials with good imaging sensitivity and penetration. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 1. Transmission electron microscope image (a) and particle size distribution diagram (b) of near-infrared emitting europium-doped carbon dots prepared by the present invention.

[0023] Figure 2 This is the X-ray photoelectron spectrum of near-infrared emitting europium-doped carbon dots.

[0024] Figure 3 The ultraviolet absorption, fluorescence excitation and fluorescence emission spectra of the prepared near-infrared europium-doped fluorescent carbon dots.

[0025] Figure 4 This is a comparison chart of the red shift of the prepared near-infrared europium-doped carbon dots.

[0026] Figure 5 It is a bar graph of cell survival rate after the prepared near-infrared europium-doped carbon dots were incubated with HeLa cells for 24 hours.

[0027] Figure 6 This is the fluorescence imaging of the prepared near-infrared europium-doped carbon dots after incubation with HeLa cells for 4 hours. DETAILED DESCRIPTION

[0028] To further illustrate the near-infrared-emitting europium-doped carbon dots, their preparation methods, and applications, and to achieve the intended purpose of the present invention, the following describes in detail the near-infrared-emitting europium-doped carbon dots, their preparation methods, and applications, along with their specific implementation methods, structures, features, and efficacy, in conjunction with preferred embodiments. In the following description, different references to "one embodiment" or "embodiment" do not necessarily refer to the same embodiment. Furthermore, specific features, structures, or characteristics of one or more embodiments may be combined in any suitable manner.

[0029] The following will further introduce in detail the near-infrared emitting europium-doped carbon dots and their preparation method and application in conjunction with specific embodiments of the present invention:

[0030] In order to prepare optical materials with better optical properties, the present invention proposes a near-infrared emitting europium-doped carbon dot and its preparation method and application. The technical solution adopted is:

[0031] A method for preparing near-infrared emitting europium-doped carbon dots comprises the following steps:

[0032] After dissolving p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride in water, a hydrothermal reaction is carried out; after the reaction is completed, the mixture is cooled to room temperature to obtain a reaction solution;

[0033] The reaction solution is filtered, dialyzed for purification, and dried to obtain the near-infrared emitting europium-doped carbon dots.

[0034] Preferably, the mass ratio of p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride is 1-3:1-3:0.25-3.

[0035] Further preferably, the mass ratio of p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride is 3:3:1.

[0036] Preferably, the temperature of the hydrothermal reaction is 180-210° C., and the time is 8-12 hours.

[0037] Preferably, the filtration is performed using a 0.22 μm organic filter membrane.

[0038] Preferably, the dialysis purification is performed by dialysis using a dialysis bag with a molecular weight cut-off of 500-1000Da.

[0039] A near-infrared emitting europium-doped carbon dot is prepared by the above-mentioned preparation method.

[0040] Preferably, the fluorescence excitation spectrum of the europium-doped carbon dots is in the range of 300-700 nm, and the fluorescence emission spectrum is in the range of 600-850 nm.

[0041] All reagents and solvents used in this example were of analytical grade and purchased from Nanjing Huada Chemical Instrument Co., Ltd. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was purchased from Energy Chemical Co., Ltd.

[0042] Example 1.

[0043] The specific steps are as follows:

[0044] (1) Using p-phenylenediamine and ethylenediaminetetraacetic acid as precursors and europium chloride as a metal dopant, 0.3 g of p-phenylenediamine, 0.3 g of ethylenediaminetetraacetic acid, and 0.1 g of europium chloride were dissolved in 20 ml of water. The mixture was placed in a sealed reactor and subjected to hydrothermal reaction at 210°C for 12 hours. After the reaction was completed, the mixture was cooled to room temperature to obtain a reaction solution.

[0045] (2) The reaction liquid in the kettle was collected, filtered through a 0.22 μm filter membrane to remove large particle impurities, placed in a 500-1000 Da dialysis bag, dialyzed and purified in deionized water for 24 h to remove residual organic molecules, and then dried to obtain a dark purple europium-doped carbon dot solid powder (PEu-CDs).

[0046] Example 2.

[0047] The europium-doped carbon dots prepared in Example 1 were tested:

[0048] (1) Transmission electron microscopy test and particle size distribution

[0049] The europium-doped carbon dots prepared in Example 1 were examined by transmission electron microscopy and particle size distribution. Figure 1 As shown. Figure 1 It can be seen that the carbon dots prepared by near-infrared europium doping are spherical ( Figure 1 (a)), the particle size is small, the particle size distribution is uniform, and it is narrow and symmetrical. The average particle size is about 1.70±0.06nm after statistical calculation.

[0050] (2) X-ray photoelectron spectroscopy

[0051] The europium-doped carbon dots prepared in Example 1 were subjected to X-ray photoelectron spectroscopy to determine whether europium was doped into the carbon dots and the elements contained therein. Figure 2 As shown. Figure 2 It can be seen that PEu-CDs are mainly composed of four elements: carbon, oxygen, nitrogen and europium, indicating that europium has been successfully doped into carbon dots.

[0052] (2) Optical performance

[0053] Method: The europium-doped carbon dots prepared in Example 1 were dispersed in ethanol (concentration: 4 mg / mL) to prepare a stock solution, and then 100 μL was added to 1.9 mL of PBS buffer (pH = 7.4) (final concentration: 0.2 mg / mL) to test its ultraviolet absorption and fluorescence excitation and emission spectra.

[0054] Results: The UV absorption, fluorescence excitation and fluorescence emission spectra of the prepared near-infrared europium-doped fluorescent carbon dots are as follows: Figure 3 As shown. Figure 3 The prepared near-infrared europium-doped carbon dots exhibit a broad UV-visible absorption range, with the most pronounced absorption peak at 531 nm. Their excitation range is broad, covering the 300-700 nm range. In particular, at the optimal excitation wavelength of 588 nm, they exhibit optimal emission in the first near-infrared region at 752 nm.

[0055] The emission wavelengths of europium-doped carbon dots PEu-CDs and undoped europium P-CDs were tested under the same test conditions.

[0056] Results: Using p-phenylenediamine and ethylenediaminetetraacetic acid as precursors and europium chloride as a metal dopant, a synergistic effect was produced, which could make its emission wavelength reach the near-infrared region. Compared with the emission wavelength of P-CDs without europium, the europium-doped carbon dots PEu-CDs were red-shifted by 140nm, and the effect was very significant.

[0057] Example 3: Fluorescence Imaging

[0058] Methods: Fluorescence imaging was performed using an Olympus FV-1000 confocal microscope (Japan). HeLa cells were cultured in DMEM supplemented with 10% FBS and antibiotics (100 units / mL penicillin and 100 μg / mL streptomycin) in an incubator with 5% carbon dioxide at 37°C. The cytotoxicity of PEu-CDs in HeLa cells was determined using the MTT assay.

[0059] First, cells were plated in 96-well culture plates before treatment. After 24 hours of culture, the cells were incubated with different concentrations of carbon dots (europium-doped carbon dots prepared in Example 1) (0, 10, 50, 100, 150, and 200 μg / mL, respectively) for 24 hours. 10 μL of MTT was then added to each well to fully react with the cells. Control experiments were performed under the same conditions, but without the addition of PEu-CDs. Each experiment was performed seven times to ensure the credibility of the experimental data. The activities obtained from the MTT assay (measured by seven independent biological replicates) are expressed as mean ± SD.

[0060] Results: From Figure 5 It can be seen that when the concentration of europium-doped near-infrared carbon dots is 200 μg / mL, the cell survival rate is still over 80%, indicating that europium-doped near-infrared carbon dots have no obvious toxicity and are cell compatible.

[0061] The results of laser confocal imaging are shown in Figure 6 , the excitation wavelength is 650nm, and bright red light is obtained respectively. It can be seen from the figure that near-infrared europium-doped carbon dots can enter cells and mark living cells. The bright red fluorescence is mainly concentrated in the cytoplasm area. These preliminary results show that near-infrared carbon dots can be used for cell imaging.

[0062] Example 4.

[0063] The specific steps are as follows:

[0064] (1) Using p-phenylenediamine and ethylenediaminetetraacetic acid as precursors and europium chloride as a metal dopant, 0.3 g of p-phenylenediamine, 0.3 g of ethylenediaminetetraacetic acid, and 0.3 g of europium chloride were dissolved in 30 ml of water. The mixture was placed in a sealed reactor and subjected to a hydrothermal reaction at 180°C for 12 hours. After the reaction was completed, the mixture was cooled to room temperature to obtain a reaction solution.

[0065] (2) The reaction liquid in the kettle was collected, filtered through a 0.22 μm filter membrane to remove large particle impurities, placed in a 500-1000 Da dialysis bag, dialyzed and purified in deionized water for 24 h to remove residual organic molecules, and then dried to obtain a dark purple europium-doped carbon dot solid powder.

[0066] Example 5.

[0067] The specific steps are as follows:

[0068] (1) Using p-phenylenediamine and ethylenediaminetetraacetic acid as precursors and europium chloride as a metal dopant, 0.2 g of p-phenylenediamine, 0.2 g of ethylenediaminetetraacetic acid, and 0.2 g of europium chloride were dissolved in 30 ml of water. The mixture was placed in a sealed reactor and subjected to a hydrothermal reaction at 200°C for 10 hours. After the reaction was completed, the mixture was cooled to room temperature to obtain a reaction solution.

[0069] (2) The reaction liquid in the kettle was collected, filtered through a 0.22 μm filter membrane to remove large particle impurities, placed in a 500-1000 Da dialysis bag, dialyzed and purified in deionized water for 24 h to remove residual organic molecules, and then dried to obtain a dark purple europium-doped carbon dot solid powder.

[0070] Example 6.

[0071] The specific steps are as follows:

[0072] (1) Using p-phenylenediamine and ethylenediaminetetraacetic acid as precursors and europium chloride as a metal dopant, 0.1 g of p-phenylenediamine, 0.1 g of ethylenediaminetetraacetic acid, and 0.1 g of europium chloride were dissolved in 20 ml of water. The mixture was placed in a sealed reactor and subjected to a hydrothermal reaction at 210°C for 8 hours. After the reaction was completed, the mixture was cooled to room temperature to obtain a reaction solution.

[0073] (2) The reaction liquid in the kettle was collected, filtered through a 0.22 μm filter membrane to remove large particle impurities, placed in a 500-1000 Da dialysis bag, dialyzed and purified in deionized water for 24 h to remove residual organic molecules, and then dried to obtain a dark purple europium-doped carbon dot solid powder.

[0074] Comparative Example 1: Effect of Precursor Selection on Europium-Doped Carbon Dots

[0075] The specific operation steps are the same as those in Example 1, except for the selection of precursors. The selection of precursors and their effects on europium-doped carbon dots are shown in Table 1.

[0076] Table 1

[0077] serial number Precursor result 1 cellulose Unsuccessful preparation of europium-doped carbon dots 2 urea Unsuccessful preparation of europium-doped carbon dots 3 Melamine Unsuccessful preparation of europium-doped carbon dots 4 o-phenylenediamine Unsuccessful preparation of europium-doped carbon dots 5 m-phenylenediamine Unsuccessful preparation of europium-doped carbon dots 6 o-phenylenediamine, citric acid (1:1) Unsuccessful preparation of europium-doped carbon dots 7 m-phenylenediamine, citric acid (1:1) Unsuccessful preparation of europium-doped carbon dots

[0078] Europium-doped carbon dots are difficult to prepare. As shown in Table 1, using europium chloride as a metal dopant has a limited range of precursor options, and conventional precursors cannot produce europium-doped carbon dots.

[0079] Comparative Example 2.

[0080] The specific operation steps are the same as those in Example 1, except for the selection of precursors and metal dopants. The selection of precursors and metal dopants and their effects on europium-doped carbon dots are shown in Table 2.

[0081] Table 2

[0082]

[0083]

[0084] From the combination of Table 1 and Table 2, it can be seen that only when p-phenylenediamine and ethylenediaminetetraacetic acid are used as precursors and europium chloride is used as a metal dopant, can europium-doped carbon dots be successfully prepared, and a synergistic effect is produced, with a red shift of 140 nm (e.g. Figure 4 ), the effect is very significant.

[0085] Comparative Example 3.

[0086] The specific operation steps are the same as those in Example 1, except for the ratio of the precursor to the metal dopant. The ratio of the precursor to the metal dopant and its effect on the europium-doped carbon dots are shown in Table 3.

[0087] Table 3

[0088] serial number p-phenylenediamine / g Ethylenediaminetetraacetic acid / g Europium chloride / g result 1 0.3 0.3 0.025 Optimal emission wavelength 620nm 2 0.3 0.3 0.05 Optimal emission wavelength 620nm 3 0.3 0.3 0.1 Optimal emission wavelength 750nm 4 0.3 0.3 0.2 Optimal emission wavelength 670nm 5 0.3 0.3 0.3 Optimal emission wavelength 630nm

[0089] The above is merely a preferred embodiment of the present invention and does not constitute any form of limitation to the embodiments of the present invention. Any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the embodiments of the present invention are still within the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing near-infrared emitting europium-doped carbon dots, characterized in that: The following steps are involved: After dissolving p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride in water, a hydrothermal reaction is carried out; after the reaction is completed, the mixture is cooled to room temperature to obtain a reaction solution; The reaction solution is filtered, dialyzed for purification, and dried to obtain the near-infrared emitting europium-doped carbon dots.

2. The preparation method according to claim 1, characterized in that The mass ratio of p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride is 1-3:1-3:0.25-3.

3. The preparation method according to claim 2, characterized in that The mass ratio of p-phenylenediamine, ethylenediaminetetraacetic acid and europium chloride is 3:3:

1.

4. The preparation method according to claim 1, characterized in that The temperature of the hydrothermal reaction is 180-210° C., and the time is 8-12 hours.

5. The preparation method according to claim 1, characterized in that The filtration is carried out using a 0.22 μm organic filter membrane.

6. The preparation method according to claim 1, characterized in that The dialysis purification is performed by dialysis using a dialysis bag with a molecular weight cut-off of 500-1000Da.

7. A near-infrared emitting europium-doped carbon dot, characterized in that: The invention is prepared by the preparation method according to any one of claims 1 to 6.

8. The europium-doped carbon dots according to claim 7, characterized in that The fluorescence excitation spectrum of the europium-doped carbon dots is in the range of 300-700 nm, and the fluorescence emission spectrum is in the range of 600-800 nm.

9. Use of the europium-doped carbon dots according to claim 7 in cell imaging.

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