CN115920890B - Preparation method of iron single-atom doped fluorescent carbon dot nano enzyme - Google Patents
Preparation method of iron single-atom doped fluorescent carbon dot nano enzyme Download PDFInfo
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Abstract
The invention discloses a preparation method of iron single-atom doped fluorescent carbon dot nano enzyme, which takes chitosan, glutamic acid and ferrous sulfate heptahydrate as reaction precursors, synthesizes iron single-atom doped carbon dot nano enzyme base solution with fluorescence characteristic in one step through a hydrothermal method, and finally obtains the iron single-atom doped fluorescent carbon dot nano enzyme solid through dialysis and frigid drying. The method for synthesizing the iron single-atom carbon point nano-enzyme is simple to operate, can realize the iron single-atom doped fluorescent carbon point nano-enzyme without strict equipment such as ultra-high temperature pyrolysis and complex reaction steps, and can effectively avoid the problem of metal load reduction under the ultra-high temperature condition. The synthesized iron single-atom fluorescent carbon point nano-enzyme has good water solubility, photoluminescence performance and excellent peroxidase characteristics, and has higher stability, adjustable catalytic activity and feasibility of large-scale preparation compared with natural enzymes.
Description
Technical Field
The invention belongs to the technical field of synthesis of monoatomic nanoenzyme, and relates to a preparation method of iron monoatomic doped fluorescent carbon dot nanoenzyme.
Background
The monoatomic nanoenzyme is a nanoenzyme with ultrahigh catalytic activity, generally, the metal-doped carbon-point nanoenzyme needs expensive reaction equipment and special conditions, and the metal loading capacity can be reduced due to ultrahigh synthesis temperature, so that the catalytic activity of a finally obtained product is reduced. The currently reported monatomic nano-enzyme is mainly prepared by decomposing the precondition of iron coordination in a pyrolysis mode (Boosting Fenton-Like Reactions via Single Atom Fe Catalysis, environmental Science & Technology,2019,53 (19) -11391-11400). Therefore, it is of great importance to explore a method for synthesizing monoatomic nano-enzyme conveniently, green and at low cost.
Composite materials with enzymatic activity such as metals, metal compounds, metal organic frameworks and the like still have a challenge in terms of affinity and specificity compared with traditional enzymes, while carbon-based nanoezymes have good biocompatibility, but the catalytic activity of pure carbon-based nanoezymes is not high and does not necessarily have strong luminescence properties, and metal-doped carbon-based nanoezymes can sometimes improve the catalytic activity of nanoezymes (Bioadhesive injectable hydrogel with phenolic carbon quantum dot supported Pd single atom nanozymes as a localized immunomodulation niche for cancer catalytic immunotherapy, DOI:10.1016/j. Therefore, the development of the single-atom fluorescent carbon dot nano-enzyme with ultrahigh peroxidase activity, good biocompatibility and excellent fluorescence characteristic has innovative significance.
Disclosure of Invention
The invention solves the technical problem of providing a preparation method of iron single-atom doped fluorescent carbon dot nano enzyme. According to the method, chitosan, glutamic acid and ferrous sulfate heptahydrate are used as raw materials, a basic reaction liquid is obtained through one-step hydrothermal reaction, sediment is removed through centrifugation, and the iron single-atom fluorescent carbon point nano enzyme is obtained through dialysis and freeze drying.
The preparation method of the fluorescent carbon dot nano enzyme doped with the iron single atoms is characterized by comprising the following specific steps of:
step S1: dissolving chitosan in 1% (v/v) acetic acid aqueous solution, performing ultrasonic treatment to form a uniform solution, adding glutamic acid and ferrous sulfate heptahydrate powder into the dispersed chitosan solution, uniformly stirring, finally dropwise adding ethylenediamine solution, transferring the solution into a reaction kettle, and performing reaction in a high-temperature oven.
Step S2: and (3) centrifuging the reaction liquid obtained in the step (S1) to remove precipitate, dialyzing, collecting liquid, and freeze-drying the liquid to obtain a brownish-black powder solid product.
Preferably, the feeding mass ratio of the glutamic acid to the ferrous sulfate heptahydrate in the step S1 is 1:1.
Preferably, the centrifugation time in step S2 is 8-14min and the dialysis time is 10-14h.
The invention takes chitosan, glutamic acid and ferrous sulfate heptahydrate as raw materials, prepares the fluorescent carbon dot nano enzyme doped with iron single atoms through one-step hydrothermal reaction, and is a brand new preparation method of the single-atom nano enzyme with convenience, green and low cost.
Compared with the prior art, the invention has the following advantages:
1. the invention has low cost, including material cost and reaction equipment cost, and simple preparation method and post-reaction treatment;
2. compared with other synthesis methods, the temperature of the iron single-atom fluorescent carbon point nano-enzyme prepared by the method is low, and the reaction condition is mild;
3. the prepared iron single-atom fluorescent carbon dot nano-enzyme has good water solubility, and the iron single-atom fluorescent carbon dot nano-enzyme has excellent peroxidase activity under the load of an iron single atom, and besides, the excellent fluorescent property of a carbon dot is reserved, so that the iron single-atom fluorescent carbon dot nano-enzyme has higher fluorescence quantum yield, and the obtained iron single-atom fluorescent carbon dot nano-enzyme has potential application value in various fields of enzyme catalysis.
The iron single-atom fluorescent carbon point nano enzyme prepared by the invention has the performance of peroxide mimic enzyme, and can catalyze and oxidize hydrogen (H) 2 O 2 ) Hydroxyl radicals (·oh) are generated such that the chromogenic substrates 3,3', 5' -Tetramethylbenzidine (TMB) and o-phenylenediamine (OPD) are oxidized to oxidized 3,3', 5' -tetramethylbenzidine (oxTMB) and 2, 3-Diaminophenazine (DAP), respectively, and have an ultraviolet-visible response. According to enzyme steady state dynamics experiment, the iron single atom fluorescent carbon point nano enzyme has lower Mie constant (K) m ) Indicating that the enzyme has strong catalytic activity.
Drawings
FIG. 1 is a schematic diagram showing the synthesis procedure of the iron single-atom fluorescent carbon dot nano-enzyme prepared in example 2.
FIG. 2 is a TEM image of the iron single-atom fluorescent carbon dot nano-enzyme prepared in example 2.
FIG. 3 is a spherical aberration diagram of the iron single-atom fluorescent carbon nano-enzyme prepared in example 2.
FIG. 4 is a HAADF-STEM diagram of the iron single-atom fluorescent carbon dot nano-enzyme prepared in example 2.
FIG. 5 is a graph of emission spectra obtained by excitation with a fluorescence spectrometer at different excitation wavelengths.
FIG. 6 is an ultraviolet absorption diagram of the iron single-atom fluorescent carbon nanoenzyme measured with an ultraviolet spectrophotometer in the wavelength range of 200-800 nm.
FIG. 7 is an XPS diagram of the iron single-atom fluorescent carbon point nano-enzyme prepared in example 2.
Detailed Description
The above-described matters of the present invention will be described in further detail by way of examples, but it should not be construed that the scope of the above-described subject matter of the present invention is limited to the following examples, and all techniques realized based on the above-described matters of the present invention are within the scope of the present invention.
Example 1
Step S1: 0.1g of chitosan is dissolved in 10mL of 1% (v/v) acetic acid aqueous solution, ultrasonic treatment is carried out for 30min in an ultrasonic cleaner, then 0.05g of glutamic acid is added, the mixture is fully stirred to dissolve solid powder, 100 mu L of ethylenediamine is added, the mixture is transferred to a high-temperature reaction kettle, and the mixture is sealed at 180 ℃ for 6h.
Step S2: and (3) cooling the reaction kettle after the reaction in the step (S1) to room temperature, taking out black liquid obtained by the reaction, centrifuging to remove solid impurities, dialyzing for 12 hours by using a dialysis bag (MW=500-1000), and collecting liquid in the dialysis bag, and freeze-drying to obtain brown black powder which is fluorescent carbon dot powder.
Example 2
Step S1: 0.1g of chitosan is dissolved in 10mL of 1% (v/v) acetic acid aqueous solution, ultrasonic treatment is carried out for 30min in an ultrasonic cleaner, then 0.05g of ferrous sulfate heptahydrate and 0.05g of glutamic acid are added, the mixture is fully stirred to dissolve solid powder, 100 mu L of ethylenediamine is added, the mixture is transferred to a high-temperature reaction kettle, and the mixture is sealed at 180 ℃ for 6h.
Step S2: and (3) cooling the reaction kettle after the reaction in the step (S1) to room temperature, taking out black liquid obtained by the reaction, centrifuging to remove solid impurities, dialyzing for 12 hours by using a dialysis bag (MW=500-1000), and collecting liquid in the dialysis bag, and freeze-drying to obtain brown black powder, namely the iron single-atom fluorescent carbon dot nano enzyme powder.
Peroxidase activity detection: the iron single-atom fluorescent carbon dot nano-enzyme can catalyze hydrogen peroxide (H) 2 O 2 ) Generating OH to oxidize chromogenic substrate 3,3', 5' -Tetramethylbenzidine (TMB) to form blue oxidized 3,3', 5' -tetramethylbenzidine (oxTMB), the chromogenic substrate having a maximum absorption peak at 652 nm; the same principle that the monoatomic nano-enzyme catalyzesHydrogen peroxide (H) 2 O 2 ) Generating hydroxyl radicals (·oh) oxidizing o-phenylenediamine (OPD) to yellow 2, 3-Diaminophenazine (DAP) and having a maximum absorbance peak at 420 nm; the material can degrade Methylene Blue (MB) by hydrogen peroxide, the methylene blue color becomes light, and the material has maximum absorption at 665 nm.
In contrast to example 2, the fluorescent carbon dots not doped with metallic iron in example 1 have no catalase-like activity.
Compared with the prior art, the prior method (Bioadhesive injectable hydrogel with phenolic carbon quantum dot supported Pd single atom nanozymes as a localized immunomodulation niche for cancer catalytic immunotherapy, DOI: 10.1016/j.biological materials.2021.121272) comprises the steps of obtaining carbon dots by hydrothermal synthesis of raw materials at 180 ℃ for 6 hours, dialyzing, freeze-drying to obtain solid, redissolving the solid and Na 2 PdCl 4 The solution is stirred to synthesize the palladium monoatomic nano-enzyme supported by carbon points, the synthesis method needs two steps of reactions, the consumed time is long, and the Mie constant (K) of TMB m ) Is 569.4mM higher than the iron single-atom fluorescent carbon point nano-enzyme (Mie constant (K) m )＝0.139mM)。
In the embodiment 2 of the invention, the reaction conditions are mild, and the existing synthesis method (Boosting Fenton-Like Reactions via Single Atom Fe Catalysis, environmental Science&Technology,2019,53 (19) -11391-11400) was prepared by calcining the material in a tube furnace at 500℃for 5h, and adding Fe (NO) 3 ) 3 ·9H 2 The O is continuously calcined at 500 ℃ for 5 hours after being ground, the synthesis method is complicated and has high temperature, and the metal load is obviously reduced due to the high temperature.
While the above examples illustrate the synthetic methods of the present invention, and the advantages of the catalytic activity of the materials, those skilled in the art will appreciate that the present invention is not limited by the foregoing examples, which are provided by way of illustration only, and that various changes and modifications may be made without departing from the scope of the synthetic methods of the present invention, which are within the scope of the invention.
Claims (3)
1. The preparation method of the fluorescent carbon dot nano enzyme doped with the iron single atoms is characterized by comprising the following specific steps:
step S1: dissolving chitosan in acetic acid aqueous solution with volume ratio of 1%, fully dissolving by ultrasonic, adding ferrous sulfate heptahydrate and glutamic acid into uniform chitosan solution, fully stirring to dissolve solid powder to obtain mixed solution, finally dripping ethylenediamine solution into the mixed solution, uniformly stirring, transferring into a high-temperature reaction kettle, and reacting for 6h at 180 ℃;
step S2: centrifuging the reaction liquid obtained in the step S1 to remove solid impurities, dialyzing by a dialysis bag, and further freeze-drying the impurity-removed liquid to obtain black brown solid powder which is the iron single-atom fluorescent carbon dot nano enzyme;
wherein, in the step S1, the chitosan is dissolved in acetic acid aqueous solution with the volume ratio of 1% for 30min by ultrasound, and the feeding mass ratio of ferrous hexa-hydrate to glutamic acid is 1:1.
2. The method for preparing the iron single-atom doped fluorescent carbon dot nano-enzyme, which is characterized by comprising the following steps of: the instrument used for the high-temperature reaction in the step S1 is a common constant-temperature drying oven.
3. The method for preparing the iron single-atom doped fluorescent carbon dot nano-enzyme, which is characterized by comprising the following steps of: the centrifugation time in the step S2 is 8-14min, and the dialysis time is 10-14h.
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| CN116726926B (en) * | 2023-06-08 | 2024-03-22 | 合肥工业大学 | Method for detecting methyl mercaptan in Fe & Cu@CDs nano-enzyme dual mode |
| CN116735580B (en) * | 2023-08-15 | 2023-11-21 | 中国农业科学院农产品加工研究所 | Meat freshness detection sensor based on bimodal monoatomic nano enzyme and preparation method thereof |
| CN117402614B (en) * | 2023-10-17 | 2024-05-24 | 齐鲁工业大学(山东省科学院) | Carbon dot-based nano-enzyme and preparation method and application thereof |
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