CN114031593B - Nano fluorescent material, nano fluorescent probe, and preparation method and application thereof - Google Patents
Nano fluorescent material, nano fluorescent probe, and preparation method and application thereof Download PDFInfo
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- CN114031593B CN114031593B CN202111007160.5A CN202111007160A CN114031593B CN 114031593 B CN114031593 B CN 114031593B CN 202111007160 A CN202111007160 A CN 202111007160A CN 114031593 B CN114031593 B CN 114031593B
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- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
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- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6432—Quenching
Abstract
The invention relates to the technical field of nano biological materials, and provides a nano fluorescent material, a nano fluorescent probe, a preparation method and application thereof. The nano fluorescent material disclosed by the invention can be used for synthesizing a nano fluorescent probe through water phase self-assembly, and the nano fluorescent probe has good water phase stability and light stability. Because the molecular structure has a group of a specific targeting bacterial structure, the fluorescence of the nano fluorescent probe before bacteria are not identified is very weak, but the fluorescence signal is enhanced after the nano fluorescent probe is specifically combined with the bacteria, so that the naked eye bacteria detection and the bacterial imaging of a fluorescence means can be realized; after the bacteria detection is completed, the controllable and in-situ sterilization function can be realized, and the nano probe integrates diagnosis and treatment functions, and can be applied to the fields of biological imaging, bacteria detection and monitoring, sanitation and epidemic prevention and the like.
Description
Technical Field
The invention relates to the technical field of nano biological materials, in particular to a nano fluorescent material, a nano fluorescent probe, a preparation method and application thereof.
Background
Infections caused by pathogenic bacteria result in about one third of the mortality rate worldwide, which constitutes a serious threat to global public health. Various classes of antibacterial drugs currently have become effective weapons for safeguarding human health and saving human life, however, due to their irrational use, they are gradually triggering a global bacterial resistance crisis. In clinical application, the early and rapid diagnosis and the timely treatment of pathogenic bacteria infection are of great significance for improving the infection treatment effect and relieving the bacterial drug resistance situation. For pathogen detection, the current clinical diagnosis mainly depends on the traditional detection technology (such as tissue section culture, histological analysis and peripheral blood sampling) assisted by biochemical detection, and although a reliable bacteria identification result can be obtained, the method has the defects of time and labor waste and low efficiency, and often causes serious treatment delay. In the treatment of bacterial infections, a number of effective therapeutic strategies have been developed, including cationic polymers, antimicrobial peptides, photothermal/photodynamic therapeutic agents and gaseous drugs (e.g. CO, NO) and the like, are widely used in research against bacterial and drug-resistant bacterial infections. However, bacterial diagnosis and treatment are currently independent procedures in clinical practice, which inevitably lead to delays in optimal treatment time, reduced treatment efficacy and increased economic and psychological burden for patients. Thus, there has been a great interest in developing integrated antimicrobial strategies for effective bacterial detection and targeted therapy.
The fluorescence detection technology has the characteristics of simplicity, rapidness and sensitivity, and becomes an indispensable technology in bacteria detection and imaging application. Among various fluorescent materials (molecular fluorescent probes, fluorescent proteins, nanoprobes, etc.) that have been developed, nanoprobes exhibit small-sized unique physicochemical properties due to their size and structural particularities, and show great potential in bacterial diagnosis and treatment. The integration of fluorescent probe molecules with bactericidal functional components is a common approach to the construction of an antibacterial system with integrated bacterial detection and function. However, most of the current constructions are prepared by chemically attaching or physically encapsulating the fluorescent dye and the bactericidal component into the nanoparticle, and have disadvantages of complicated preparation process, low drug loading efficiency, poor stability, unexpected release or leakage of the fluorescent dye and the bactericidal component, and the like, which severely limit the clinical application thereof. In addition, some nanocarriers may cause toxic and inflammatory reactions during degradation or metabolism in the human body.
Disclosure of Invention
In view of the above, the present invention is directed to a nano fluorescent probe. Compared with the traditional diagnosis and treatment integrated antibacterial system, the nano fluorescent probe has the advantages of excellent bacteria detection specificity, better stability and biocompatibility, and extremely simple preparation process.
The invention provides a nano fluorescent material, which has a structure shown in a formula I:
wherein R is 1 Aryl selected from-H or C6-C30; r is R 2 Selected from amide groups or ester groups, m is an integer from 0 to 15, and n is an integer from 0 to 10.
In the nano fluorescent material provided by the invention, R is as follows 1 is-H, R 2 Is an amide group.
The nano fluorescent material provided by the invention has a structure shown in a formula VI:
the nano fluorescent material provided by the invention is prepared from compounds shown in the formulas II and III:
wherein R is 1 Aryl selected from-H or C6-C30;
R 3 selected from-COOH, -NH 2 、-CHO、-COCl、(CH 2 ) m COOH、(CH 2 ) m NH 2 、(CH 2 ) m CHO or (CH) 2 ) m COCl, m is an integer from 0 to 15;
r4 is selected from-OH, -COOH or-NH 2 -n is an integer from 0 to 10.
In some embodiments, R 1 is-H, R 3 is-COOH, R4 is-NH 2 。
In some specific embodiments, the nano fluorescent material provided by the invention is prepared from a compound shown in a formula II-a and a compound shown in a formula III-a:
the invention also provides a preparation method of the nano fluorescent material, which comprises the following steps:
reacting a compound of formula II with a compound of formula III at 0 ℃ in a protective atmosphere to obtain an intermediate;
removing Boc protecting groups from the intermediate to obtain a compound shown in a formula I;
in the present invention, the protective atmosphere is a protective atmosphere known to those skilled in the art, and is not particularly limited, but nitrogen is preferable in the present invention.
In the invention, the reaction of the compound of formula II and the compound of formula III is a condensation reaction. The specific type of condensing agent used in the condensation reaction of the present invention is not particularly limited, and may be any condensing agent commonly used in the art. The condensation reaction is carried out with stirring at room temperature.
In the invention, the removing agent used for removing the Boc protecting group is trifluoroacetic acid, HCl/tetrahydrofuran or HCl/dioxane, and the trifluoroacetic acid is preferred. In some embodiments of the invention, the step of removing the Boc protecting group is: the intermediate compound was slowly added to the solvent containing trifluoroacetic acid at 0 ℃.
In the preparation method of the fluorescent nanomaterial, after the Boc protecting group is removed to obtain the crude product of the compound shown in the formula I, the preparation method further comprises the step of purifying the crude product.
The invention also provides a nano fluorescent probe which is prepared by self-assembling the nano fluorescent material in an aqueous solvent.
In some embodiments, the nano-fluorescent probe of the invention is prepared by self-assembling a nano-fluorescent material shown in formula II in an aqueous solvent.
Wherein the solvent is aqueous solution, and can be H 2 O, PBS buffer, naCl solution, etc.
In some embodiments, the nano-fluorescent probe of the present invention is prepared by self-assembling a compound represented by formula VI in a mixed solution of DMSO and water.
The fluorescent nano probe is prepared by one-step simple water phase small molecule self-assembly mainly by utilizing the combined actions of pi-pi interaction, hydrophilic/hydrophobic action, electrostatic action and the like among molecular structures.
Experiments show that the nano probe has the capabilities of rapid, sensitive and specific bacterial fluorescence detection and structural imaging, and has the sterilization function after detection, namely the diagnosis and treatment integrated antibacterial function.
Therefore, the invention also provides application of the fluorescent nano probe in bacteria detection, analysis and/or sterilization.
The invention provides a method for detecting, monitoring and analyzing bacteria, which comprises the steps of incubating a nano probe and a bacterial suspension liquid for 30min-12h at 37 ℃, carrying out UV irradiation, carrying out naked eye visualization detection, and carrying out structural observation on fluorescence imaging by utilizing a laser confocal or fluorescence microscope.
Wherein the bacterial suspension is preferably a bacterial LB suspension. The co-incubation time is preferably 1h to 6h.
The invention also provides a sterilization method, which comprises the steps of incubating the nano probe solution and bacterial LB suspension for 30min-12h at 37 ℃, exposing the nano probe solution to visible light, and continuously irradiating for 30min-12h.
Further, the co-incubation time is preferably 1h to 6h.
The nano fluorescent probe constructed by the invention can be disassembled into small molecules by being induced by bacteria, and then is metabolized and utilized by the bacteria to enter a cell wall structure to activate the fluorescence of the small molecules, so that the nano fluorescent probe has advantageous bacterial responsiveness and specificity, can realize the rapid, sensitive, accurate and broad-spectrum detection of the bacteria, can perform fluorescence imaging on the bacterial structure, and is beneficial to further assisting in bacterial identification.
After bacteria detection is realized, the nano fluorescent probe constructed by the invention can release CO under the irradiation of visible light, so that the nonspecific broad-spectrum killing of bacteria is realized, the nano fluorescent probe is effective to drug-resistant bacteria, and the bacteria are not easy to induce drug resistance.
The nano fluorescent probe has good dispersibility and stability in a water phase environment, can realize the functions of bacteria detection and sterilization under multiple scenes, and can be applied to the fields of biological imaging, bacteria detection and monitoring, epidemic prevention and the like.
The invention relates to a nano probe and an application method thereof in sterilization, wherein the application method comprises the following steps: exposing the bacterial solution metabolized with the molecular fluorescent probe to visible light, and continuously irradiating for 30min-12h; more preferably, the irradiation time is 1h to 5h.
Compared with the prior art, the invention has the beneficial effects that:
the nano probe provided by the invention has hydrophilic and hydrophobic elements in the molecular structure, and in the water phase self-assembly process, the common driving actions such as intermolecular pi-pi interaction, hydrophilic/hydrophobic action, electrostatic action and the like can induce the self-assembly of molecules to form nano particles with uniform size and better water-solubility stability, so that the nano probe has the advantages of low-cost and easily obtained raw materials, simple preparation method, easiness in realizing industrial production and the like, and solves the problems of limited application and the like caused by poor water solubility, easiness in photo-bleaching and the like of the traditional molecular probe.
After self-assembling the nano fluorescent probe molecules into nano particles, the fluorescent quenching effect can occur, and the light stability is good; in addition, the CO release capability of the light-shielding type solar energy storage battery is obviously inhibited, so that the light-shielding type solar energy storage battery does not need to be protected from light during storage, and has the advantage of easy storage.
The nano fluorescent probe provided by the invention is dissociated into small molecules under the induction of bacterial metabolism, can be metabolized into a cell wall structure by bacteria, activates molecular fluorescence, realizes the 'turn-on' fluorescence detection and structural imaging of bacteria, and has the advantages of specificity, accuracy, rapidness and the like.
The nano fluorescent probe provided by the invention has specific bacterial identification and sterilization capability, can only detect and kill bacteria in various complex and practical scenes (such as a scene where bacteria coexist with mammalian cells, a scene where bacteria exist in phagocytes and a scene in bacterial biofilms), and can be applied to the fields of biological imaging, bacteria detection and monitoring, epidemic prevention and the like.
Drawings
FIG. 1 is an intermediate compound 2 of example 1 1 H NMR spectrum;
FIG. 2 is an ESI-MS spectrum of intermediate compound 2 of example 1;
FIG. 3 is a diagram of Compound 3 of example 1 1 H NMR spectrum;
FIG. 4 is an ESI-MS spectrum of compound 3 of example 1;
FIG. 5 is a graph showing absorption and emission spectra of Compound 3 in DMSO in example 1;
FIG. 6 is a fluorescence emission spectrum of the molecule of Compound 3 and nanoparticle formation in example 1;
FIG. 7 is a TEM image of nanoparticles aggregated by compound 3 of example 1 a) and an aggregate TEM image formed by compound 1;
FIG. 8 is a graph showing the bacterial detection performance of the nanoprobe in example 1;
FIG. 9 is a graph of CLSM fluorescence (9 a) and bright field (9 b) after incubation of aggregates formed by Compound 1 with bacteria in comparative example 1.
Detailed Description
The invention is further illustrated below with reference to the drawings and specific examples, but the invention is not limited to these specific embodiments. The instruments, materials, reagents and the like used in the examples are conventional instruments and reagents unless otherwise specified, and are commercially available.
Example 1: the embodiment provides a bacterial response type diagnosis and treatment integrated nano fluorescent probe, the specific synthetic route is shown in the following chart, and the specific process is as follows:
1) Synthesis of Compound 2:
compound 1 (0.332 g) was dissolved in THF at 0deg.C, N 2 N-hydroxysuccinimide (NHS, 0.105 g) and 1-ethyl-3 (3-dimethylpropylamine) carbodiimide (EDCI, 0.221 g) were added to compound 3 in an atmosphere and the reaction was stirred at room temperature for 6h. Triethylamine (2.3 mmol) was added dropwise to the reaction system, and after stirring for 30 minutes, boc-D-Dap-OH (0.204 g) was added thereto and stirred at room temperature overnight. After the reaction is finished, the obtained crude product is purified by centrifugation, washing and drying in sequence, and a yellow product compound is obtained. FIG. 1 is a diagram of Compound 2 1 HNMR spectra, as can be seen from the figure: 1 h NMR (DMSO-d 6, 400 MHz) delta 8.86 (s, 1H), 8.76 (s, 1H), 8.39 (d, j=8.3 hz, 2H), 8.36 (s, 1H), 8.27 (d, j=8.3 hz, 1H), 8.10 (d, j=8.4 hz, 1H), 8.03 (d, j=8.4 hz, 2H), 7.70 (t, j=7.6 hz, 1H), 7.59 (t, j=7.5 hz, 1H), 6.43-6.38 (m, 1H), 4.18 (q, j=6.7 hz, 1H), 3.76 (s, 1H), 3.74 (s, 1H), 3.63-3.58 (m, 1H), 1.39 (s, 9H). FIG. 2 is an ESI-MS spectrum of Compound 2, from which it can be seen: [ MH ]]Theoretical molecular weight 517.5, actual molecular weight 517.2.
2) Synthesis of Compound 3:
compound 2 (0.202 g) was added to DCM (4 mL), TFA (3 mL) was slowly added at 0deg.C, and the reaction was stirred at room temperature for 4h. After the reaction, the crude product was obtained by concentration by distillation under reduced pressure. The crude product is purified by centrifugation, washing and drying to finally obtain a yellow product HF-D-Ala. FIG. 3 is a diagram of Compound 3 1 H NMR spectra, as can be seen from the figure: 1 h NMR (DMSO-d 6, 400 MHz) delta 8.95 (s, 1H), 8.87 (s, 1H), 8.42 (d, j=8.1 hz, 2H), 8.36 (s, 1H), 8.28 (d, j=8.3 hz, 1H), 8.10 (d, j=9.2 hz, 1H), 8.08 (d, j=8.7 hz, 2H), 7.71 (t, j=7.5 hz, 1H), 7.60 (t, j=7.5 hz, 1H), 3.70 (t, j=6.7 hz, 1H), 3.61 (dt, j=7.2 hz, j=6.8 hz, 1H), 3.46 (t, j=6.4 hz, 1H). FIG. 4 is an ESI-MS spectrum of Compound 3, from which it can be seen: [ MH ]] - Is 417.4 and the actual molecular weight is 417.3.
3) Preparation of a nanoprobe based on compound 3:
and adding the compound 3 into DMSO, stirring to dissolve the compound quickly, taking a proper amount of solution which is uniformly mixed, adding deionized water into the solution, and preparing the compound by a precipitation method.
Example 2: molecular fluorescent probe 3 and nanoparticle spectrum information formed by same
UV-Vis spectral characterization of compound 3: compound 3 was dissolved in DMSO, 2mL of the solution was removed, added to a cuvette, and its absorbance spectrum was collected in the visible wavelength range.
The fluorescence spectrum characterization of the nanoparticle comprises the following specific steps: 2mL of the aqueous nanoparticle solution was added to a cuvette, and its fluorescence emission spectrum was collected at a wavelength of 445-700nm, with an excitation light wavelength of 415nm.
As shown in fig. 5, compound 3 has a maximum absorption wavelength of 415nm in DMSO, a maximum emission wavelength of 619nm, and a Stokes shift of 204nm. Maximum absorption wavelength of nanoparticles in water
FIG. 6 is a graph of fluorescence emission of Compound 3 and the nanoprobe formed therefrom, from which it can be seen that Compound 3 exists in molecular form in DMSO, which has a strong emission peak around 620 nm; after it aggregates in the aqueous phase to form nanoparticles, its emission peak decays greatly, hardly emitting fluorescence, due to aggregation-induced quenching.
Example 3: stability characterization of the nano-fluorescent probe:
the step of characterizing the nanoparticle by Dynamic Light Scattering (DLS) is: the nanoparticles were first prepared and added to a cuvette for direct use in the test, and the results are shown in table 1.
Table 1 is a particle size and stability characterization in water of the nanoprobe of example 1
The result shows that the particle size of the nano fluorescent probe particle prepared by the invention is about 58nm, and the particle size of the nano fluorescent probe particle can be always stabilized at about 60nm after the nano fluorescent probe particle is dispersed in water for 5 days, and the main reason is that the nano fluorescent probe particle can be uniformly and stably dispersed in a water phase environment due to the amino acid hydrophilic group in a molecular structure, so that the stability of the nano probe in the water phase environment is improved.
Example 4: and (3) TEM structural characterization of the nano fluorescent probe:
the morphology difference between the aggregate formed by the compound 1 and the nano probe of the compound 3 is explored by TEM, as shown in fig. 7, the compound 3 can be self-assembled into a spherical nano particle with the size of about 50-60nm in an aqueous phase solvent, and the aggregate formed by the compound 1 is in a disordered and entangled state, which indicates that the hydrophilic element of the amino acid in the molecular structure is important to the morphology and stability of the nano particle.
Example 5: characterization of CO release of molecular fluorescent probe:
5mL of HF-D-Ala in DMSO was added to a sealed vial, and after sealing, the vial was irradiated with a visible light (410 nm,10 mW/cm) 2 ) The vial was irradiated for 30min and the CO content of the vial was detected by a CO detector.
The water-soluble stability of the nanoparticle is further confirmed by detecting the release amount of CO, and the specific steps are as follows: the aqueous solution containing the nanoparticles was selected and transferred to a sealable vial, sealed with visible light (410 nm,10 mW/cm) 2 ) And (3) irradiating, and detecting the content of CO in the bottle after irradiation by using a CO detector. The results are shown in Table 2.
Table 2 shows the amounts of CO released by Compound 3 and nanoprobe in example 1 under dark and light conditions
The properties of the nanoparticles and their molecular forms to release CO are shown in table 2, and under dark conditions, neither the nanoparticles nor their molecular forms release CO; whereas in pure DMSO, visible light irradiation of the molecular fluorescent probe solution may release about 187ppm CO, whereas nanoparticles release only about 6ppm CO. The energy absorbed by the molecules is dissipated in a non-radiative transition mode mainly due to the inhibition of the aggregation state of the molecules, so that CO release cannot be activated, and the nano probe has better photostability than the molecular form of the nano probe, and light-induced CO leakage is avoided.
Example 6: characterization of bacterial detection performance of the nano fluorescent probe:
nanoparticles (30. Mu.M) were incubated with LB suspension liquid medium containing bacteria on a shaker at 37 ℃. Subsequently, bacterial pellets were collected by centrifugation and the luminescence of the bacteria was observed under a portable ultraviolet lamp.
As shown in fig. 8, the treatment of both bacterial precipitates (staphylococcus aureus and escherichia coli) with nanoparticles emitted intense red fluorescence under the uv lamp, while the bacterial precipitates without treatment did not emit any fluorescence, demonstrating that the bacteria were able to induce the nanoprobe to disaggregate, illuminating the bacteria by eliminating the aggregation-induced quenching effect, allowing detection of the bacteria.
Example 7: characterization of bactericidal performance of the nano fluorescent probe:
the sterilization performance is verified by a plate colony counting method, and the specific steps are as follows: the bacteria metabolized with fluorescent probe molecules were irradiated with visible light (410 nm,10 mW/cm) 2 ) After 1h of irradiation, the bacterial liquid is coated on an LB solid agar plate, and is placed in a 37 ℃ incubator for 24h of cultivation, and the sterilization efficiency is calculated by recording the number of bacterial colonies. The results are shown in Table 3.
Table 3 shows the sterilization rates of the nanoprobe of example 1 under visible light irradiation at different times;
the result shows that the bacterial death rate after molecular marking is related to the irradiation time of visible light, and the longer the irradiation time is, the higher the sterilization rate of CO is; after 2h of irradiation, complete inactivation of staphylococcus aureus (s.aureus), escherichia coli (e.coli) and methicillin-resistant staphylococcus aureus (MRSA) was achieved.
Example 8: cytotoxicity test of nano fluorescent probe:
the cytotoxicity of the nano fluorescent probe on the mouse fibroblast (L929 cell) is evaluated by adopting a CCK-8 method, and the specific steps are as follows: l929 cells were first resuscitated and transferred to high sugar DMEM at 37℃with 5% CO 2 Is cultured in an incubator of (a). A DMEM dispersion of nano-fluorescent probes (100. Mu.M, 2% DMSO/DMEM, v/v) was prepared. L929 cells (1X 10) 4 cells/well) were incubated in 96-well plates for 24h, and 100. Mu.L of DMEM dispersion containing the nanofluorescent probe was added. Then using visible light (410 nm,10 mW/cm) 2 ) Irradiating for 1h, and incubating in a dark condition in an incubator for 24h. After co-incubation, cells were gently washed with PBS buffer, DMEM was removed, 10% CCK-8 solution (v/v) was added to each well, and incubated for 2h at 37℃in an incubator. Finally, the absorbance at 450nm is measured by a multifunctional enzyme-labeled instrument.
The cytotoxicity results are shown in Table 4.
Table 4 shows cytotoxicity of the nanoprobe of example 1
The result shows that the cell survival rate of the nano fluorescent probe and the cells after co-culture is higher than 90% under illumination, and the nano fluorescent probe has no obvious adverse effect on the activity of the L929 cells.
Comparative example 1: aggregate formed by Compound 1 and related Performance test
(1) Preparation of aggregates formed by Compound 1
And adding the compound 1 into DMSO, stirring to quickly dissolve, taking a proper amount of mixed and dissolved solution, and adding deionized water into the solution to prepare the aggregate of the compound 1.
(2) Stability test of aggregates formed by compound 1 the procedure for characterizing the aggregates of compound 1 by Dynamic Light Scattering (DLS) was: first, an aggregate of the compound 1 was prepared, and it was put into a cuvette for direct testing, and the analysis of the test results is shown in table 5.
Table 5 shows particle size and stability in water of the nanoprobe of example 1
The results show that the aggregates obtained in comparative example 1 (aggregates formed by compound 1) are unstable under the same preparation operation as the nanoprobe, mainly because no hydrophilic group in the molecular structure provides sufficient hydrophilic/hydrophobic action and electrostatic interaction.
(3) TEM test of aggregates formed by Compound 1
The morphology of the aggregates formed by compound 1 was examined by TEM, as shown in fig. 7, and the aggregates formed by compound 3 were in a disordered, entangled state, which also demonstrates that the hydrophilic motifs in the molecular structure are important for both morphology and stability of the nanoparticle.
(4) Bacterial detection Performance test of aggregates formed by Compound 1
The aggregates formed by compound 1 were incubated with LB suspension liquid medium containing the bacteria on a shaker at 37 ℃. Subsequently, bacterial pellet was collected by centrifugation and fluorescence of the bacteria (staphylococcus aureus) was observed by laser confocal microscopy. From the analysis of the results, it was confirmed that the aggregate formed by compound 1 did not realize the ability of bacteria detection imaging.
(5) Test of the sterilizing Performance of aggregates formed by Compound 1
The sterilization performance is verified by a plate colony counting method, and the specific steps are as follows: bacteria incubated with the aggregates formed by Compound 1 were incubated with visible light (410 nm,10mW/cm 2 ) After 1h of irradiation, the bacterial liquid is diluted and coated on an LB solid agar plate, and is placed in a 37 ℃ incubator for 24h of cultivation, and the sterilization efficiency is calculated by recording the number of bacterial colonies. The results are shown in Table 6.
Table 6 shows the sterilization rates of the nanoprobes of the aggregates formed by the compound 1 under visible light irradiation at different times;
from the results, it was found that the bacterial death rate of the incubation with the aggregate formed by compound 1 was independent of the irradiation time of visible light, and the bactericidal effect was long, which was related to the inhibition of CO release by compound 1 due to aggregation.
Conclusion:
from the results, the nano probe has the advantages of simple synthesis and preparation, good water-soluble stability, good light stability and the like. When bacteria are infected, the nano probe can specifically identify the bacteria, and the bacteria are lightened through fluorescence, so that the detection of bacterial infection is realized; after illumination, bacteria metabolizing the molecular probes can release CO for in-situ sterilization, namely, the nanometer provided by the invention can realize specific detection and treatment of bacteria at the same time.
It will be apparent to those skilled in the art that various changes and modifications can be made to the present invention without departing from the principles of the invention, and such changes and modifications fall within the scope of the appended claims.
Claims (7)
1. A nano fluorescent material having the structure shown below:
2. the method for preparing the nano fluorescent material according to claim 1, wherein the process is as follows:
3. a nano fluorescent probe prepared by self-assembling the nano fluorescent material of claim 1 in an aqueous solvent.
4. The nano-fluorescent probe of claim 3, wherein the solvent is H 2 O, PBS buffer or NaCl solution.
5. Use of the fluorescent nanomaterial of claim 1 or the nano-fluorescent probe of claim 3 or 4 for the preparation of a reagent for bacterial detection, monitoring, analysis and/or sterilization.
6. A method for detecting, monitoring and analyzing bacteria of non-diagnostic purpose, characterized in that after incubating the nano fluorescent probe according to claim 3 or 4 with bacterial suspension at 37 ℃ for 30min-12h, UV irradiation and naked eye visualization detection are carried out, and structural observation is carried out on fluorescence imaging by using a laser confocal or fluorescence microscope.
7. A method for sterilizing for non-therapeutic purposes, which is characterized in that the nano fluorescent probe according to claim 3 or 4 is incubated with a bacterial suspension at 37 ℃ for 30min-12h, and then is exposed to visible light for continuous irradiation for 30min-12h.
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| JP2014141465A (en) * | 2012-12-27 | 2014-08-07 | Tosoh Corp | Derivatives of 4h-chromone, process for producing the same, and method for detecting nucleic acid using the same |
| WO2016123141A1 (en) * | 2015-01-26 | 2016-08-04 | Berreau Lisa M | Carbon monoxide releasing molecules and associated methods |
| CN110437150A (en) * | 2019-06-06 | 2019-11-12 | 中国科学技术大学 | Carbon monoxide donor molecule with photoluminescent property and its preparation method and application |
| CN110981841A (en) * | 2019-11-15 | 2020-04-10 | 郑州大学 | Coumarin-based wash-free fluorescent probe for detecting gram-positive bacteria and application |
| CN112661740A (en) * | 2020-12-29 | 2021-04-16 | 中国科学技术大学 | Donor molecule for photoresponse cooperative release of carbon monoxide and nitric oxide, derivative thereof, preparation method and application |
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| WO2018214932A1 (en) * | 2017-05-26 | 2018-11-29 | The Hong Kong University Of Science And Technology | Organic luminogens |
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| JP2014141465A (en) * | 2012-12-27 | 2014-08-07 | Tosoh Corp | Derivatives of 4h-chromone, process for producing the same, and method for detecting nucleic acid using the same |
| WO2016123141A1 (en) * | 2015-01-26 | 2016-08-04 | Berreau Lisa M | Carbon monoxide releasing molecules and associated methods |
| CN110437150A (en) * | 2019-06-06 | 2019-11-12 | 中国科学技术大学 | Carbon monoxide donor molecule with photoluminescent property and its preparation method and application |
| CN110981841A (en) * | 2019-11-15 | 2020-04-10 | 郑州大学 | Coumarin-based wash-free fluorescent probe for detecting gram-positive bacteria and application |
| CN112661740A (en) * | 2020-12-29 | 2021-04-16 | 中国科学技术大学 | Donor molecule for photoresponse cooperative release of carbon monoxide and nitric oxide, derivative thereof, preparation method and application |
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