A solvent-free method for preparing red fluorescent carbon dots and its application

Abstract

The present invention discloses a solvent-free preparation method and application of red fluorescent carbon dots, belonging to the technical field of functional polymer materials. The disclosed method involves reacting o-phenylenediamine, a polyphenol compound, and an inorganic metal salt via a solvent-free process, followed by purification of the reaction product to obtain the red fluorescent carbon dots. The preparation process for these red fluorescent carbon dots is simple, low-cost, pollution-free, and uniform, making them easy to produce on a large scale. The red fluorescent carbon dots have broad application prospects in enhancing photosynthetic energy conversion in microalgae. Experimental studies have shown that these red fluorescent carbon dots strongly absorb yellow-green light (480-600 nm), which is poorly utilized by microalgae. They are excited by light in this range, emitting red fluorescence with a wavelength of 600-700 nm, which can be efficiently utilized by microalgae. This converts light into energy and improves light energy utilization.

Description

Technical Field

[0001] The present invention belongs to the technical field of polymer functional materials, and in particular relates to a red fluorescent carbon dot prepared by a solvent-free method and a method and application thereof. Background Art

[0002] Photosynthesis is the most important chemical reaction on Earth. Photosynthetic organisms use it to provide energy for all life on Earth. Microalgae utilize sunlight as the energy source for photosynthesis, but the visible light range they utilize is limited. Blue-violet light with wavelengths between 400 and 480 nm is strongly absorbed by chlorophyll and carotenoids, while red-orange light with wavelengths between 600 and 700 nm is absorbed by chlorophyll and promotes carbon fixation in microalgae. Yellow-green light with wavelengths between 500 and 600 nm contributes little to photosynthesis. Therefore, fully utilizing the solar radiation reaching the Earth's surface for photosynthesis is key to enhancing microalgae's photosynthetic energy conversion.

[0003] As a type of 0D carbon-based nanomaterial, carbon dots (Cdots) possess the characteristics of nanometer size, non-toxicity, good biocompatibility, rich functional groups, and the ability to generate photoinduced electrons. They have great potential for enhancing photosynthesis. Cdots can combine with chloroplasts to form a photoconversion system. Through photoconversion or energy transfer, they promote electron transfer in the photosystem, enhance photosynthetic activity and efficiency, and thus promote carbon reactions and accumulate more biomass. Cdots also have excellent luminescence properties, and the emission wavelength can be adjusted; to date, the reported luminescence range covers almost the entire visible light region. Theoretically, if the fluorescence emission spectrum of Cdots is consistent with the absorption spectrum of microalgae, the excitation spectrum of Cdots can be absorbed by chloroplasts and used for photosynthesis, thereby improving photosynthetic efficiency.

[0004] In addition, the current synthesis methods of carbon dots are mostly solvothermal synthesis. Since the reaction process is uncontrollable, the synthesized carbon dots are uneven and usually require further separation and purification, which has many limitations. Summary of the Invention

[0005] The purpose of the present invention is to provide a solvent-free preparation method and application of red fluorescent carbon dots, so as to solve the technical problem that the existing solvent thermal synthesis method is difficult to meet large-scale production due to the limitation of the preparation process.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions:

[0007] The present invention discloses a method for preparing red fluorescent carbon dots by a solvent-free method, comprising the following steps:

[0008] Add o-phenylenediamine, polyphenol compound and inorganic metal salt into a mortar and grind thoroughly to obtain a mixed powder;

[0009] The mixed powder is transferred to a reactor and heated for reaction, and after the reaction is completed, the carbonized solid is obtained after cooling;

[0010] The carbonized solid is dissolved and then purified to obtain red fluorescent carbon dot powder.

[0011] Furthermore, the molar ratio of the o-phenylenediamine, the polyphenol compound and the inorganic metal salt is 1:(0.08-1):(0.05-0.1).

[0012] Furthermore, the polyphenol compound is one of dopamine hydrochloride, tea polyphenols, tannic acid, gallic acid and 3,4-dihydroxybenzoic acid.

[0013] Furthermore, the inorganic metal salt is AlCl3 or AlCl3·6 H2O;

[0014] The mixed powder is transferred to a reactor and heated for reaction at a temperature of 180-200° C. for 10-12 h.

[0015] Furthermore, the carbonized solid is dissolved, and the mass ratio of the carbonized solid to the solvent is 1:(1-100); the solvent is methanol or ethanol.

[0016] Furthermore, the purification process includes sequentially performing membrane filtration, dialysis, centrifugation and freeze-drying;

[0017] The pore size of the filter membrane is 0.22 μm; the dialysis is performed using a dialysis bag with a molecular weight cutoff of 1000 Da; the centrifugal speed is 8000-10000 rpm, and the centrifugation time is 5-10 min.

[0018] The present invention also discloses red fluorescent carbon dots prepared by the above preparation method. The ultraviolet absorption wavelength and excitation wavelength of the red fluorescent carbon dots are 480-600 nm, and the emission peak wavelength range is 600-700 nm.

[0019] The present invention also discloses the application of the red fluorescent carbon dots in enhancing the photosynthetic energy conversion of microalgae.

[0020] Further, the following steps are included:

[0021] First, microalgae in the logarithmic growth phase are inoculated into a liquid culture medium, and then a red fluorescent carbon dot solution is added to the liquid culture medium, which is then placed in a constant temperature shaker for culture.

[0022] Furthermore, the red fluorescent carbon dot solution is obtained by mixing red fluorescent carbon dot powder and dimethyl sulfoxide; the concentration of the red fluorescent carbon dot solution is (0-10) μg / mL;

[0023] The culture temperature in the constant temperature shaker is 25±1°C, the photoperiod is 12 / 12 h, and the light source is LED;

[0024] The liquid culture medium is BG-11 culture medium, which is sterilized at 121° C. for 20 min; and the microalgae is Chlorella pyrenoidosa.

[0025] Compared with the prior art, the present invention has the following beneficial effects:

[0026] The present invention discloses a method for preparing red fluorescent carbon dots by a solvent-free method. The method adopts o-phenylenediamine, a polyphenol compound and an inorganic metal salt aluminum chloride to synthesize the red fluorescent carbon dots by a solvent-free method. During the preparation process, the reaction pressure of the reaction process is reduced, the safety of the experiment is greatly improved, and the material synthesis cost is reduced. In addition, the preparation process is simple and easy to operate, the reaction process is controllable, and industrial production can be realized.

[0027] Furthermore, the raw materials and preparation process used in this method follow the concept of green environmental protection, and the entire process has almost no pollution to the environment.

[0028] The present invention also discloses red fluorescent carbon dots prepared by the above method. The red fluorescent carbon dots in the present invention are synthesized by a solvent-free method, have small particle size, stable properties, a simple preparation process, and good biocompatibility. According to relevant experimental results, the red fluorescent carbon dots in the present invention have strong absorption of yellow-green light (480-600 nm) with low utilization rate by microalgae, and can be excited by light in this region to emit red fluorescence with a wavelength range of 600-700 nm that can be efficiently utilized by microalgae, thereby achieving light conversion and improving light energy utilization. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 This is the emission spectrum of the red fluorescent carbon dots prepared by the present invention;

[0030] Figure 2 The UV-visible absorption spectrum of the red fluorescent carbon dots prepared by the present invention is shown in FIG.

[0031] Figure 3 This is a transmission electron microscope image of the red fluorescent carbon dots prepared by the present invention;

[0032] Figure 4 This is a confocal image of Chlorella pyrenoidosa / red fluorescent carbon dots of the present invention;

[0033] Where: a-bright field image; b-Chlorella pyrenoidosa (λ ex =488 nm); c-red fluorescent carbon dots (λ ex =561 nm); confocal image of d-Chlorella pyrenoidosa / red fluorescent carbon dots;

[0034] Figure 5 : The growth curves of Chlorella pyrenoidosa cultured without red fluorescent carbon dots or with different concentrations of red fluorescent carbon dots in the present invention;

[0035] Figure 6 In the present invention, the light intensity is 600 W / m 2 Under the same conditions, the dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots;

[0036] Figure 7 The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots within 30 min under different light intensities in the present invention;

[0037] Figure 8 It is the ATP content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots in the present invention. DETAILED DESCRIPTION

[0038] To facilitate understanding of the features and effects of the present invention by those skilled in the art, the following provides a general description and definition of the terms and expressions used in the specification and claims. Unless otherwise indicated, all technical and scientific terms used herein have the ordinary meanings as understood by those skilled in the art regarding the present invention. In the event of conflict, the definitions in this specification shall prevail.

[0039] The theories or mechanisms described and disclosed herein, whether correct or incorrect, should not limit the scope of the present invention in any way, that is, the present invention can be implemented without being limited by any specific theory or mechanism.

[0040] All features, such as values, amounts, contents, and concentrations, described herein as numerical ranges or percentage ranges are provided for simplicity and convenience only. Accordingly, the description of numerical ranges or percentage ranges should be considered to include and specifically disclose all possible subranges and individual values ​​within the range (including integers and fractions).

[0041] In this document, unless otherwise specified, “include,” “including,” “contains,” “has” or similar terms cover the meanings of “consisting of” and “mainly consisting of,” for example, “A includes a” covers the meanings of “A includes a and other” and “A only includes a.”

[0042] In this document, for the sake of brevity, not all possible combinations of the various technical features in each embodiment or example are described. Therefore, as long as there are no contradictions in the combination of these technical features, the various technical features in each embodiment or example can be combined in any way, and all possible combinations should be considered to be within the scope of this specification.

[0043] The present invention provides a method for preparing red fluorescent carbon dots by a solvent-free method, comprising the following steps:

[0044] O-phenylenediamine, polyphenol compounds and inorganic metal salts were mixed evenly in proportion and ground into powder in an agate mortar. The mixed powder was then transferred to a stainless steel reactor, heated at 200°C for 12 hours. After the reaction was completed, it was naturally cooled to room temperature. The obtained carbonized solid was dissolved in anhydrous ethanol and filtered with a filter membrane. Red fluorescent carbon dot powder was obtained through dialysis, centrifugation and freeze-drying.

[0045] Preferably, the polyphenol compound is one of dopamine hydrochloride, tea polyphenols, tannic acid, gallic acid, and 3,4-dihydroxybenzoic acid.

[0046] Preferably, the inorganic metal salt is AlCl3 or AlCl3·6H2O.

[0047] Preferably, the molar ratio of o-phenylenediamine, polyphenol compound and inorganic metal salt is 1: (0.08-1): (0.05-0.1).

[0048] Preferably, the pore size of the filter membrane is 0.22 μm; the dialysis is performed using a dialysis bag with a molecular weight cut-off of 1000 Da; the centrifugal speed is 8000-10000 rpm, and the centrifugation time is 5-10 min.

[0049] Preferably, the ultraviolet absorption wavelength and excitation wavelength of the red fluorescent carbon dots are 480-600 nm, and the emission peak wavelength range is 600-700 nm.

[0050] The invention also discloses red fluorescent carbon dots prepared by the method.

[0051] The present invention also discloses the application of the red fluorescent carbon dots in enhancing the photosynthetic energy conversion of microalgae.

[0052] Preferably, the specific steps are: first, inoculating microalgae in the logarithmic growth phase into a liquid culture medium, then adding different concentrations of red fluorescent carbon dots (0, 1, 2.5, 5, 10 μg / mL) to the culture medium, and then placing it in a digital constant temperature shaker for cultivation, the temperature is 25±1°C, the photoperiod is 12 / 12 h (bright / dark), the light source is LED, all culture media and conical flasks are sterilized at 121°C for 20 min, the device for culturing Chlorella is a 250 mL conical flask, and it is sealed with a bio-permeable sealing film to prevent contamination by other microorganisms.

[0053] Preferably, the microalgae is Chlorella pyrenoidosa, and the liquid culture medium is a sterile BG-11 culture medium suitable for the growth of blue-green algae.

[0054] Preferably, the red fluorescent carbon dots of different concentrations are prepared by dissolving red fluorescent carbon dot powder in dimethyl sulfoxide solvent to form a red fluorescent carbon dot solution.

[0055] Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiments are only used to illustrate the present invention and are not used in limiting the scope of the present invention.In addition, should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms fall equally within the scope limited by the appended claims of the application.

[0056] The following examples were prepared using conventional instruments and equipment in the art. Experimental methods in the following examples, where specific conditions are not specified, were generally performed under conventional conditions or according to the conditions recommended by the manufacturer. The various raw materials used in the following examples, unless otherwise specified, were conventional commercially available products, with specifications conventional in the art. In the present specification and the following examples, unless otherwise specified, "%" indicates percentage by weight, "part" indicates parts by weight, and "ratio" indicates weight ratio.

[0057] Example 1

[0058] A method for preparing red fluorescent carbon dots by a solvent-free method comprises the following steps:

[0059] 5 mmol of o-phenylenediamine, 5 mmol of dopamine hydrochloride, and 0.33 mmol of AlCl₃·6H₂O were mixed and ground uniformly in an agate mortar for 10 min to a uniform powder. The mixed powder was then transferred to the inner liner of a 30 mL autoclave, which was placed in an oven at 200°C for 12 h. After the reaction, the autoclave was removed and naturally cooled to room temperature. 1.57 g of the resulting carbonized solid was dissolved in anhydrous ethanol and filtered through a 0.22 μm polyethersulfone membrane to remove large particles. The mass ratio of carbonized solid to solvent was 1:10. The filtered solution was placed in a dialysis bag with a molecular weight cutoff of 1000 Da and dialyzed against deionized water for 3 days, with the deionized water replaced every 12 h. The dialyzed solution was centrifuged at 10,000 rpm for 5 min, the supernatant removed, and the resulting solid was freeze-dried to obtain a dark purple powder, which is the red fluorescent carbon dot powder.

[0060] The red fluorescent carbon dot powder is dissolved in an organic solvent (methanol, ethanol, etc., an organic solvent that can dissolve carbon dots and has high volatility) to produce a purple-red red fluorescent carbon dot solution with red fluorescence.

[0061] Example 2

[0062] A method for preparing red fluorescent carbon dots by a solvent-free method comprises the following steps:

[0063] 5 mmol o-phenylenediamine, 1.25 mmol tea polyphenols, and 0.33 mmol AlCl₃·6H₂O were mixed and ground uniformly in an agate mortar for 10 min to a uniform powder. The mixed powder was then transferred to the inner liner of a 30 mL autoclave, which was placed in an oven at 200°C for 12 h. After the reaction, the autoclave was removed and naturally cooled to room temperature. 0.97 g of the resulting carbonized solid was dissolved in anhydrous ethanol and filtered through a 0.22 μm polyethersulfone membrane to remove large particles. The mass ratio of carbonized solid to solvent was 1:10. The filtered solution was placed in a dialysis bag with a molecular weight cutoff of 1000 Da and dialyzed against deionized water for 3 days, with the deionized water replaced every 12 h. The dialyzed solution was centrifuged at 10,000 rpm for 5 min, the supernatant removed, and the resulting solid was freeze-dried to obtain a dark red powder, which is the red fluorescent carbon dots.

[0064] The red fluorescent carbon dot powder is dissolved in an organic solvent (methanol, ethanol, etc., an organic solvent that can dissolve carbon dots and has high volatility) to produce a purple-red red fluorescent carbon dot solution with red fluorescence.

[0065] Example 3

[0066] A method for preparing red fluorescent carbon dots by a solvent-free method comprises the following steps:

[0067] 5 mmol o-phenylenediamine, 0.4 mmol tannic acid, and 0.33 mmol AlCl₃·6H₂O were mixed and ground uniformly in an agate mortar for 10 min to a uniform powder. The mixed powder was then transferred to the inner liner of a 30 mL autoclave, which was placed in an oven at 200°C for 12 h. After the reaction was complete, the autoclave was removed and naturally cooled to room temperature. 1.30 g of the resulting carbonized solid was dissolved in anhydrous ethanol and filtered through a 0.22 μm polyethersulfone membrane to remove large particles. The mass ratio of carbonized solid to solvent was 1:10. The filtered solution was placed in a dialysis bag with a molecular weight cutoff of 1000 Da and dialyzed against deionized water for 3 days, with the deionized water replaced every 12 h. The dialyzed solution was centrifuged at 10,000 rpm for 5 min, the supernatant removed, and the resulting solid was freeze-dried to obtain a dark gray powder, which is the red fluorescent carbon dots.

[0068] The red fluorescent carbon dots are dissolved in an organic solvent (methanol, ethanol, etc., an organic solvent that can dissolve carbon dots and has high volatility) to produce a purple-red red fluorescent carbon dot solution with red fluorescence.

[0069] Example 4

[0070] A method for preparing red fluorescent carbon dots by a solvent-free method comprises the following steps:

[0071] 5 mmol o-phenylenediamine, 3.3 mmol gallic acid, and 0.33 mmol AlCl₃·6H₂O were mixed and ground uniformly in an agate mortar for 10 min to a uniform powder. The mixed powder was then transferred to the inner liner of a 30 mL autoclave, which was placed in an oven at 200°C for 12 h. After the reaction, the autoclave was removed and naturally cooled to room temperature. 1.18 g of the resulting carbonized solid was dissolved in anhydrous ethanol and filtered through a 0.22 μm polyethersulfone membrane to remove large particles. The mass ratio of carbonized solid to solvent was 1:10. The filtered solution was placed in a dialysis bag with a molecular weight cutoff of 1000 Da and dialyzed against deionized water for 3 days, with the deionized water replaced every 12 h. The dialyzed solution was centrifuged at 10,000 rpm for 5 min. The supernatant was removed, and the resulting solid was freeze-dried to obtain a dark brown powder, which is the red fluorescent carbon dots.

[0072] The red fluorescent carbon dots are dissolved in an organic solvent (methanol, ethanol, etc., an organic solvent that can dissolve carbon dots and has high volatility) to produce a purple-red red fluorescent carbon dot solution with red fluorescence.

[0073] Example 5

[0074] A method for preparing red fluorescent carbon dots by a solvent-free method comprises the following steps:

[0075] 5 mmol of o-phenylenediamine, 5 mmol of 3,4-dihydroxybenzoic acid, and 0.33 mmol of AlCl₃·6H₂O were mixed and ground uniformly in an agate mortar for 10 min to a uniform powder. The mixed powder was then transferred to the inner liner of a 30 mL autoclave, which was placed in an oven at 200°C for 12 h. After the reaction, the autoclave was removed and cooled to room temperature. 1.39 g of the resulting carbonized solid was dissolved in anhydrous ethanol and filtered through a 0.22 μm polyethersulfone membrane to remove large particles. The mass ratio of carbonized solid to solvent was 1:10. The filtered solution was placed in a dialysis bag with a molecular weight cutoff of 1000 Da and dialyzed against deionized water for 3 days, with the deionized water replaced every 12 h. The dialyzed solution was centrifuged at 10,000 rpm for 5 min, the supernatant removed, and the resulting solid was freeze-dried to obtain a yellow-brown powder, which is the red fluorescent carbon dots.

[0076] The red fluorescent carbon dots are dissolved in an organic solvent (methanol, ethanol, etc., an organic solvent that can dissolve carbon dots and has high volatility) to produce a purple-red red fluorescent carbon dot solution with red fluorescence.

[0077] Application Example 1

[0078] Chlorella vulgaris in the logarithmic growth phase was inoculated into a sterile BG-11 liquid culture medium suitable for the growth of blue-green algae, and different concentrations of red fluorescent carbon dots obtained in Example 1 (0, 1, 2.5, 5, 10 μg / mL) were added thereto. The cells were then placed in a digital thermostat shaker for culture at a temperature of 25±1°C, a photoperiod of 12 / 12 h (bright / dark), and an LED light source. All culture media and conical flasks were sterilized at 121°C for 20 min. The Chlorella vulgaris culture apparatus was a 250 mL conical flask sealed with a bio-permeable sealing film to prevent contamination by other microorganisms. Confocal laser scanning microscopy (CLSM) was used to confirm the interaction between Chlorella vulgaris and the red fluorescent carbon dots; the absorbance of the culture solution at a wavelength of 680 nm (OD 680 ) was used to monitor the growth activity of Chlorella pyrenoidosa. Samples were taken every day and the absorbance at 680 nm was measured using UV-Vis-NIR to characterize its biomass and draw a growth curve. A portable dissolved oxygen meter was used to detect the dissolved oxygen content. 2Under different light intensities, the dissolved oxygen content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured every 5 minutes. The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured within 30 minutes. The ATP content in Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was determined using an ATP detection kit.

[0079] Application Example 2

[0080] Chlorella vulgaris in the logarithmic growth phase was inoculated into a sterile BG-11 liquid culture medium suitable for the growth of blue-green algae, and different concentrations of red fluorescent carbon dots obtained in Example 2 (0, 1, 2.5, 5, 10 μg / mL) were added thereto. The cells were then placed in a digital thermostat shaker for culture at a temperature of 25±1°C, a photoperiod of 12 / 12 h (bright / dark), and an LED light source. All culture media and conical flasks were sterilized at 121°C for 20 min. The Chlorella vulgaris culture apparatus was a 250 mL conical flask sealed with a bio-permeable sealing film to prevent contamination by other microorganisms. Confocal laser scanning microscopy (CLSM) was used to confirm the interaction between Chlorella vulgaris and the red fluorescent carbon dots; the absorbance of the culture solution at a wavelength of 680 nm (OD 680 ) was used to monitor the growth activity of Chlorella pyrenoidosa. Samples were taken every day and the absorbance at 680 nm was measured using UV-Vis-NIR to characterize its biomass and draw a growth curve. A portable dissolved oxygen meter was used to detect the dissolved oxygen content. 2 Under different light intensities, the dissolved oxygen content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured every 5 minutes. The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured within 30 minutes. The ATP content in Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was determined using an ATP detection kit.

[0081] Application Example 3

[0082] Chlorella vulgaris in the logarithmic growth phase was inoculated into a sterile BG-11 liquid culture medium suitable for the growth of blue-green algae, and different concentrations of red fluorescent carbon dots obtained in Example 3 (0, 1, 2.5, 5, 10 μg / mL) were added thereto. The cells were then placed in a digital thermostat shaker for culture at a temperature of 25±1°C, a photoperiod of 12 / 12 h (bright / dark), and an LED light source. All culture media and conical flasks were sterilized at 121°C for 20 min. The Chlorella vulgaris culture apparatus was a 250 mL conical flask sealed with a bio-permeable sealing film to prevent contamination by other microorganisms. Confocal laser scanning microscopy (CLSM) was used to confirm the interaction between Chlorella vulgaris and the red fluorescent carbon dots; the absorbance of the culture solution at a wavelength of 680 nm (OD 680 ) was used to monitor the growth activity of Chlorella pyrenoidosa. Samples were taken every day and the absorbance at 680 nm was measured using UV-Vis-NIR to characterize its biomass and draw a growth curve. A portable dissolved oxygen meter was used to detect the dissolved oxygen content. 2 Under different light intensities, the dissolved oxygen content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured every 5 minutes. The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured within 30 minutes. The ATP content in Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was determined using an ATP detection kit.

[0083] Application Example 4

[0084] Chlorella vulgaris in the logarithmic growth phase was inoculated into a sterile BG-11 liquid culture medium suitable for the growth of blue-green algae, and different concentrations of red fluorescent carbon dots obtained in Example 4 (0, 1, 2.5, 5, 10 μg / mL) were added thereto. The cells were then placed in a digital thermostat shaker for culture at a temperature of 25±1°C, a photoperiod of 12 / 12 h (bright / dark), and an LED light source. All culture media and conical flasks were sterilized at 121°C for 20 min. The Chlorella vulgaris culture apparatus was a 250 mL conical flask sealed with a bio-permeable sealing film to prevent contamination by other microorganisms. The interaction between Chlorella vulgaris and the red fluorescent carbon dots was confirmed using confocal laser scanning microscopy (CLSM); the absorbance of the culture solution at a wavelength of 680 nm (OD 680 ) was used to monitor the growth activity of Chlorella pyrenoidosa. Samples were taken every day and the absorbance at 680 nm was measured using UV-Vis-NIR to characterize its biomass and draw a growth curve. A portable dissolved oxygen meter was used to detect the dissolved oxygen content. 2Under different light intensities, the dissolved oxygen content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured every 5 minutes. The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured within 30 minutes. The ATP content in Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was determined using an ATP detection kit.

[0085] Application Example 5

[0086] Chlorella vulgaris in the logarithmic growth phase was inoculated into a sterile BG-11 liquid culture medium suitable for the growth of blue-green algae, and different concentrations of red fluorescent carbon dots obtained in Example 5 (0, 1, 2.5, 5, 10 μg / mL) were added thereto. The cells were then placed in a digital thermostat shaker for culture at a temperature of 25±1°C, a photoperiod of 12 / 12 h (bright / dark), and an LED light source. All culture media and conical flasks were sterilized at 121°C for 20 min. The Chlorella vulgaris culture apparatus was a 250 mL conical flask sealed with a bio-permeable sealing film to prevent contamination by other microorganisms. Confocal laser scanning microscopy (CLSM) was used to confirm the interaction between Chlorella vulgaris and the red fluorescent carbon dots; the absorbance of the culture solution at a wavelength of 680 nm (OD 680 ) was used to monitor the growth activity of Chlorella pyrenoidosa. Samples were taken every day and the absorbance at 680 nm was measured using UV-Vis-NIR to characterize its biomass and draw a growth curve. A portable dissolved oxygen meter was used to detect the dissolved oxygen content. 2 Under different light intensities, the dissolved oxygen content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured every 5 minutes. The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured within 30 minutes. The ATP content in Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was determined using an ATP detection kit.

[0087] Performance testing:

[0088] The red fluorescent carbon dot powder prepared by the present invention was tested, and the results are shown in the attached figure. Figure 1 To the attached Figure 3 As shown. Figure 1 This is the emission spectrum of red fluorescent carbon dot powder. The emission wavelength range is 600-700 nm, indicating that the red fluorescent carbon dots of the present invention can convert yellow-green light, which has a low utilization rate of microalgae, into red light that can be efficiently utilized by microalgae, and have a high match with the photosynthesis spectrum of microalgae.

[0089] Figure 2This is the UV-visible absorption spectrum of the red fluorescent carbon dot powder. From the figure, we can see that the red fluorescent carbon dots have strong absorption in the 480~600 nm region, which matches the excitation spectrum.

[0090] Figure 3 This is a transmission electron microscope image of red fluorescent carbon dot powder. It can be seen that the average size of the red fluorescent carbon dots is less than 20 nm and they are well dispersed.

[0091] The red fluorescent carbon dots obtained in Example 1 at different concentrations (0, 1, 2.5, 5, and 10 μg / mL) were co-cultured with Chlorella pyrenoidosa. The interaction between Chlorella pyrenoidosa and the red fluorescent carbon dots was confirmed using confocal laser scanning microscopy (CLSM). The absorbance of the culture solution at a wavelength of 680 nm (OD 680 ) was used to monitor the growth activity of Chlorella pyrenoidosa, and the absorbance at 680 nm was measured by UV-Vis-NIR to characterize its biomass and draw a growth curve; a portable dissolved oxygen meter was used to detect the dissolved oxygen content. 2 Under different light intensities, the dissolved oxygen content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured every 5 minutes. The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was measured within 30 minutes. The ATP content in Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots was determined using an ATP detection kit. The results are shown in the attached figure. Figure 4 To the attached Figure 8 shown.

[0092] Figure 4 This is a confocal image of Chlorella pyrenoidosa / red fluorescent carbon dots. Under excitation at a wavelength of 488 nm, the spontaneous green fluorescence of the chloroplasts can be clearly observed. Under excitation at a wavelength of 561 nm, the red fluorescent carbon dots emit red fluorescence, and the red fluorescence of the red fluorescent carbon dots highly overlaps with the green fluorescence of Chlorella pyrenoidosa, indicating that the red fluorescent carbon dots are bound to Chlorella pyrenoidosa.

[0093] Figure 5 Figure 2 shows the growth curves of Chlorella pyrenoidosa cultured without or with different concentrations of red fluorescent carbon dots. The growth of Chlorella pyrenoidosa reflects an increase in overall biomass and cell number. As photoautotrophs, their biomass is derived from photosynthetic carbon fixation. Furthermore, photosynthesis also provides the energy and materials required for cell division. Therefore, the accelerated growth of Chlorella pyrenoidosa directly indicates that the red fluorescent carbon dots promote photosynthesis in Chlorella pyrenoidosa.

[0094] Figure 6 For a light intensity of 600 W / m 2Under the same conditions, the dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots showed that the oxygen production of the group with the addition of red fluorescent carbon dots increased significantly, indicating that the photosynthetic efficiency was significantly improved after the addition of red fluorescent carbon dots.

[0095] Figure 7 The dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots within 30 minutes under different light intensities. Higher light intensity increased the dissolved oxygen production of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots; however, with the increase of light intensity, the amount of oxygen evolved decreased. It may be that with the increase of light intensity, the light of Chlorella gradually became saturated, and its own photosynthetic rate gradually reached saturation.

[0096] Figure 8 Figure 2 is the ATP content of Chlorella pyrenoidosa and Chlorella pyrenoidosa / red fluorescent carbon dots. It can be seen that the ATP production of Chlorella pyrenoidosa with the addition of red fluorescent carbon dots was significantly increased, successfully improving the photosynthetic efficiency of Chlorella pyrenoidosa.

[0097] The above content is only for explaining the technical idea of ​​the present invention and cannot be used to limit the protection scope of the present invention. Any changes made on the basis of the technical solution in accordance with the technical idea proposed by the present invention shall fall within the protection scope of the claims of the present invention.

Claims

1. A method for preparing red fluorescent carbon dots by a solvent-free method, characterized in that: The following steps are involved: Grinding o-phenylenediamine, polyphenol compound and inorganic metal salt to obtain mixed powder; The mixed powder is transferred to a reactor for heating reaction, and after the reaction is completed, the carbonized solid is obtained after cooling; The carbonized solid is dissolved and then purified to obtain red fluorescent carbon dot powder; The molar ratio of o-phenylenediamine, polyphenol compound and inorganic metal salt is 1: (0.08-1): (0.05-0.1); The polyphenol compound is one of dopamine hydrochloride, tea polyphenols, tannic acid, gallic acid and 3,4-dihydroxybenzoic acid; The inorganic metal salt is AlCl3 or AlCl3·6 H2O; The mixed powder is transferred to a reactor for heating reaction at a temperature of 180-200°C for 10-12 hours; The purification process includes sequentially performing membrane filtration, dialysis, centrifugation and freeze-drying; The pore size of the filter membrane is 0.22 μm; the dialysis is performed using a dialysis bag with a molecular weight cutoff of 1000 Da; the centrifugal speed is 8000-10000 rpm, and the centrifugation time is 5-10 min.

2. The method for preparing red fluorescent carbon dots by a solvent-free method according to claim 1, wherein: Dissolving the carbonized solid is to mix the carbonized solid with a solvent, wherein the mass ratio of the carbonized solid to the solvent is 1:(1-100); the solvent is methanol or ethanol.

3. A red fluorescent carbon dot, characterized in that: The red fluorescent carbon dots are prepared by the preparation method according to claim 1 or 2, and the ultraviolet absorption wavelength and excitation wavelength of the red fluorescent carbon dots are 480-600 nm, and the emission peak wavelength range is 600-700 nm.

4. The use of the red fluorescent carbon dots according to claim 3 in enhancing photosynthetic energy conversion of microalgae, characterized in that: The following steps are involved: First, microalgae in the logarithmic growth phase are inoculated into a liquid culture medium, and then a red fluorescent carbon dot solution is added to the liquid culture medium, which is then placed in a constant temperature shaker for culture.

5. The use of red fluorescent carbon dots in enhancing photosynthetic energy conversion of microalgae according to claim 4, characterized in that: The red fluorescent carbon dot solution is obtained by mixing red fluorescent carbon dot powder and dimethyl sulfoxide; the concentration of the red fluorescent carbon dot solution is (0-10) μg / mL; The culture temperature in the constant temperature shaker is 25±1°C, the photoperiod is 12 / 12 h, and the light source is LED; The liquid culture medium is BG-11 culture medium, which is sterilized at 121° C. for 20 min; and the microalgae is Chlorella pyrenoidosa.

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