Europium-doped carbon dots capable of near-infrared emission and preparation method and application of europium-doped carbon dots

Abstract

The invention relates to near-infrared emission europium-doped carbon dots as well as a preparation method and application thereof. The invention relates to a preparation method of near-infrared emission europium-doped carbon dots, which comprises the following steps: dissolving p-phenylenediamine, ethylenediamine tetraacetic acid and europium chloride in water, and carrying out hydrothermal reaction; after the reaction is finished, cooling to room temperature to obtain reaction liquid; and filtering the reaction liquid, dialyzing and purifying, and drying to obtain the near-infrared emission europium-doped carbon dots. According to the near-infrared emission europium-doped carbon dots as well as the preparation method and the application thereof, the preparation method is simple, and the prepared nanoprobe can achieve near-infrared emission without further modification.

Description

Technical Field

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

[0002] Carbon dots have shown great potential in medical treatment, imaging, antibacterial, and antimicrobial applications due to their unique photostability, low toxicity, and good biocompatibility. Among them, near-infrared fluorescent carbon dots have low scattering and absorption in biological tissues, low phototoxicity to cells and tissues, and higher tissue penetration, which is beneficial for in situ imaging, labeling, and phototherapy of organisms. However, the fluorescence emission of carbon dots is mainly blue and green light (wavelength ranges are 450-495nm and 520-570nm, respectively), and the preparation of long-wavelength (yellow and red) luminescent carbon dots is relatively difficult, resulting in poor imaging effects in cells, and thus not suitable for imaging.

[0003] In view of this, the present invention proposes a new carbon dot and a preparation method and application thereof, 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 with near-infrared emission. The preparation method is simple and the nanoprobes can be prepared without further modification to achieve near-infrared emission.

[0005] In order to achieve the above purpose, the technical solution adopted is:

[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 reaction is cooled to room temperature to obtain a reaction solution;

[0008] The reaction solution is filtered, purified by dialysis, 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 a near-infrared emitting europium-doped carbon dots prepared by the above-mentioned preparation method. Compared with imaging in the visible light (400-650nm) band, 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 the method is simple, and the precursor can be purchased directly 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, good biocompatibility, and can be used as fluorescent imaging materials with good imaging sensitivity and penetration. BRIEF DESCRIPTION OF THE DRAWINGS

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

[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] In order to further explain the near-infrared emitting europium-doped carbon dots and their preparation methods and applications of the present invention and achieve the intended purpose of the invention, the following is a detailed description of the near-infrared emitting europium-doped carbon dots and their preparation methods and applications, their specific implementation methods, structures, features and their effects proposed in accordance with the present invention in combination with the preferred embodiments. In the following description, different "one embodiment" or "embodiment" does not necessarily refer to the same embodiment. In addition, specific features, structures or characteristics in one or more embodiments may be combined in any suitable form.

[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 a preparation method and application thereof, and 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 reaction is cooled to room temperature to obtain a reaction solution;

[0033] The reaction solution is filtered, purified by dialysis, 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 comprises 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) p-phenylenediamine and ethylenediaminetetraacetic acid are used as precursors, and europium chloride is used as a metal dopant. Take 0.3 g p-phenylenediamine, 0.3 g ethylenediaminetetraacetic acid and 0.1 g europium chloride, dissolve them in 20 ml water, place them in a reaction kettle, and perform a hydrothermal reaction at 210°C in a sealed container for 12 hours. After the reaction is completed, cool to room temperature to obtain a reaction solution.

[0045] (2) Collect the reaction liquid in the kettle, filter the reaction liquid with a 0.22 μm filter membrane to remove large particle impurities, place it in a 500-1000 Da dialysis bag, dialyze and purify it in deionized water for 24 hours to remove residual organic molecules, and then dry it 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 tested 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. After statistical calculation, the average particle size is about 1.70±0.06nm.

[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 mother solution, and then 100 μL was added to 1.9 mL of PBS buffer with pH = 7.4 (final concentration was 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 shown in Figure 3 As shown. Figure 3 It can be seen that the prepared near-infrared europium-doped carbon dots have a wide UV-visible absorption range, among which the most significant absorption peak is located at 531nm. Its excitation range is wide, covering the range of 300-700 nanometers. In particular, at the optimal excitation wavelength of 588nm, it exhibits the best emission in the near-infrared region with a wavelength of 752nm.

[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 and 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 medium containing 10% FBS and antibiotics (100 units / mL penicillin and 100 μg / mL streptomycin) in an incubator with 5% carbon dioxide and a temperature of 37°C. The cytotoxicity of PEu-CDs in HeLa cells was determined using the MTT assay.

[0059] First, cells were loaded into 96-well culture plates before treatment. After 24 hours of culture, cells were cultured with carbon dots (Europium-doped carbon dots prepared in Example 1) at different concentrations (0, 10, 50, 100, 150, 200 μg / mL, respectively) for 24 hours. Then 10 μL MTT was added to each well to fully react with the cells. The control experiment was performed under the same conditions, but PEu-CDs were not added. Each experiment was performed seven times to ensure the credibility of the experimental data. The activity obtained by the MTT test (measured by seven independent biological replicates) is 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 region. 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) p-phenylenediamine and ethylenediaminetetraacetic acid are used as precursors, and europium chloride is used as a metal dopant. Take 0.3 g p-phenylenediamine, 0.3 g ethylenediaminetetraacetic acid and 0.3 g europium chloride, dissolve them in 30 ml water, place them in a reaction kettle, and perform a hydrothermal reaction at 180°C in a sealed container for 12 hours. After the reaction is completed, cool to room temperature to obtain a reaction solution.

[0065] (2) Collect the reaction liquid in the kettle, filter the reaction liquid with a 0.22 μm filter membrane to remove large particle impurities, place it in a 500-1000 Da dialysis bag, dialyze and purify it in deionized water for 24 hours to remove residual organic molecules, and then dry it to obtain a dark purple europium-doped carbon dot solid powder.

[0066] Example 5.

[0067] The specific steps are as follows:

[0068] (1) p-phenylenediamine and ethylenediaminetetraacetic acid are used as precursors, and europium chloride is used as a metal dopant. Take 0.2 g p-phenylenediamine, 0.2 g ethylenediaminetetraacetic acid and 0.2 g europium chloride, dissolve them in 30 ml water, place them in a reaction kettle, and perform a hydrothermal reaction at 200°C in a sealed container for 10 hours. After the reaction is completed, cool to room temperature to obtain a reaction solution.

[0069] (2) Collect the reaction liquid in the kettle, filter the reaction liquid with a 0.22 μm filter membrane to remove large particle impurities, place it in a 500-1000 Da dialysis bag, dialyze and purify it in deionized water for 24 hours to remove residual organic molecules, and then dry it to obtain a dark purple europium-doped carbon dot solid powder.

[0070] Example 6.

[0071] The specific steps are as follows:

[0072] (1) p-phenylenediamine and ethylenediaminetetraacetic acid are used as precursors, and europium chloride is used as a metal dopant. Take 0.1 g p-phenylenediamine, 0.1 g ethylenediaminetetraacetic acid and 0.1 g europium chloride, dissolve them in 20 ml water, place them in a reaction kettle, and perform a hydrothermal reaction at 210°C in a sealed container for 8 hours. After the reaction is completed, cool to room temperature to obtain a reaction solution.

[0073] (2) Collect the reaction liquid in the kettle, filter the reaction liquid with a 0.22 μm filter membrane to remove large particle impurities, place it in a 500-1000 Da dialysis bag, dialyze and purify it in deionized water for 24 hours to remove residual organic molecules, and then dry it 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 the precursor. The selection of the precursor and its effect on the 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 Metaphenylenediamine 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, when europium chloride is used as a metal dopant, the range of precursors that can be selected is small, and conventional precursors cannot prepare 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] It can be seen from Table 1 and Table 2 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 140nm (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 EDTA / g Europium chloride / g result 1 0.3 0.3 0.025 Optimum emission wavelength 620nm 2 0.3 0.3 0.05 Optimum emission wavelength 620nm 3 0.3 0.3 0.1 Optimum emission wavelength 750nm 4 0.3 0.3 0.2 Optimum emission wavelength 670nm 5 0.3 0.3 0.3 Optimum emission wavelength 630nm

[0089] The above is only a preferred embodiment of the embodiment of the present invention, and does not impose any form of limitation on the embodiment of the present invention. Any simple modification, equivalent changes and modifications made to the above embodiment based on the technical essence of the embodiment of the present invention are still within the scope of the technical solution of the embodiment 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 reaction is cooled to room temperature to obtain a reaction solution; The reaction solution is filtered, purified by dialysis, 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 preparation method is described in 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 300-700nm, and the fluorescence emission spectrum is in the range of 600-800nm.

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

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