CN104845607A - Aggregation induced luminescence nanofluorescent probe and preparation method therefor - Google Patents
Aggregation induced luminescence nanofluorescent probe and preparation method therefor Download PDFInfo
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- CN104845607A CN104845607A CN201510123300.3A CN201510123300A CN104845607A CN 104845607 A CN104845607 A CN 104845607A CN 201510123300 A CN201510123300 A CN 201510123300A CN 104845607 A CN104845607 A CN 104845607A
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Abstract
The present invention discloses an aggregation induced luminescence nanofluorescent probe which coats a chitosan fluorescent molecule with the characteristic of aggregation induced luminescence on the surface of a hydroxyl apatite nanoparticle, and the structural formula of the chitosan fluorescent molecule is shown in a formula (1). The aggregation induced luminescence nanofluorescent probe is prepared by a chemical co-precipitation process. The size of the aggregation induced luminescence nanofluorescent probe is about 110 nm, the dispersion is stable, positive changes are formed on the surface thereof, and the aggregation induced luminescence nanofluorescent probe has the feature of the aggregation induced luminescence. Compared with a traditional fluorescent probe, the aggregation induced luminescence nanofluorescent probe has the advantages that the sensitivity is high, the light stability is good, and quenching does not exist during high concentration and a fluorescence spectrum is not drifted, so that the aggregation induced luminescence nanofluorescent probe can be applied to the fields of bioimaging, fluorescent tracing detection and the like. The aggregation induced luminescence nanofluorescent probe is shown in the figure below.
Description
Technical field
The present invention relates to a kind of preparation method of fluorescent probe, especially there is the preparation method of the hydroxyapatite/chitosan namo fluorescence probe of aggregation-induced emission characteristic.
Background technology
Probe is a kind of can interaction with certain specific target molecules, realize the molecule detected target molecule, and requirement does not produce or only produces insignificant interference after interacting to being detected object.Fluorescent probe is exactly using fluorescent substance as indicator, and under the exciting of certain wavelength light, make indicator produce fluorescence, realizes the qualitative of tested substance or quantitative analysis by detecting the fluorescence produced.In the past in Two decades years, fluorescent probe causes the extensive concern of chemist and biologist for the biological procedures detecting viable cell or animal, develop multiple fluorescent bio-probes, as: small molecule fluorescent spy, inorganic nano-particle pin and fluorescent polymer etc.Traditional small-molecule fluorescent probe has aggregation inducing cancellation (ACQ) phenomenon, and namely in dilute solution, during fluorescence comparatively strong and state of aggregation, fluorescent weakening does not even have fluorescence, this greatly limits its application and development.Adopt inorganic-quantum-dot (QDs) that ACQ phenomenon can be avoided, but surface hydrophilic modification must be carried out to it, to reduce toxicity.Another method improving traditional small-molecule fluorescent probe ACQ effect is marked on macromolecular chain, forms fluorescent polymer.The segment of macromolecular chain can intercept fluorophore, alleviates the gathering of fluorophore, if but the amount of mark is comparatively large, or concentration is higher, and fluorophore has hydrophobic aromatic proton, still tends to assemble.
The ACQ effect faced for fluorescent probe technique and cytotoxicity two large problems, the discovery of aggregation-induced emission (AIE) fluorescence molecule provides splendid resolving ideas undoubtedly.AIE effect makes fluorescent probe be easy to use, and AIE system does not have toxicity to cell, can not affect stechiology and cell proliferation.AIE molecule marker can effectively be improved biocompatibility on macromolecular chain and avoid dyestuff to reveal.The macromole being used for fixing dyestuff should have good biocompatibility and comparatively high reaction activity and wetting ability.Chitosan is rich in amino natural polysaccharide as a kind of, naturally becomes excellent choosing.And hydroxyapatite has good application prospect with the biocompatibility of its excellence (forming similar with natural bone) and unique needle-like crystal structure in bio-imaging, medicine controlled releasing etc.
Summary of the invention
The object of this invention is to provide a kind of highly sensitive, good light stability, without cancellation during high density, the preparation method of the aggregation-induced emission namo fluorescence probe that fluorescence spectrum does not drift about.
Aggregation-induced emission namo fluorescence probe of the present invention, has the chitosan fluorescence molecule of aggregation-induced emission characteristic at hydroxyapatite nano particle Surface coating, the structural formula of chitosan fluorescence molecule as the formula (1),
Formula (1)
In formula: m, x, y are respectively number of repeat unit.
The preparation method of aggregation-induced emission namo fluorescence probe of the present invention, step is as follows:
1) ratio 5:5:1 Calcium Chloride Powder Anhydrous, SODIUM PHOSPHATE, MONOBASIC and citric acid monohydrate trisodium being pressed amount of substance mixes, add tri-distilled water, drip Glacial acetic acid under magnetic agitation to dissolve, the consumption of Glacial acetic acid is 2% of mixeding liquid volume, and making concentration is 5 mg/mL hydroxyapatite precursor solutions;
2) the hydroxyapatite precursor solution tri-distilled water that step 1) is obtained is diluted to 1 mg/mL, under stirring, regulates pH=10 with alkaline solution, continue stirring and spend the night, obtained hydroxyapatite nano particle solution;
3) be that to be dissolved in volumetric concentration be in 2% aqueous acetic acid for the AIE fluorescent mark chitosan of 0.1 mol%-20 mol% by mark rate, compound concentration is the AIE fluorescent mark chitosan-acetic acid solution of 0.1-5 mg/mL;
4) by step 2) obtained hydroxyapatite nano particle solution centrifugal abandoning supernatant, again with tri-distilled water cleaning, centrifugal, precipitation is joined step 3) in obtained AIE fluorescent mark chitosan-acetic acid solution, stir at least 1 h, centrifugal abandoning supernatant, adding tri-distilled water makes hydroxyapatite nano particle concentration be 1 mg/mL, stirs, obtains aggregation-induced emission namo fluorescence probe.
The present invention passes through chemical coprecipitation, with citric acid monohydrate trisodium for dispersion agent preparation table wears the hydroxyapatite nano particle of negative charge, and by the chitosan of electrostatic adsorption coated one deck aggregation-induced emission fluorescence molecule mark thereon, thus the obtained namo fluorescence probe with aggregation-induced emission characteristic.
The invention has the advantages that:
1) namo fluorescence probe is of a size of about 110nm, and shape is needle-like, and surface band positive charge, be easy to interact with cytolemma, and then enter cell interior by endocytosis, realize the object of cell imaging.
2) fluorescent probe has aggregation-induced emission characteristic, compared with conventional fluorescent probe, has highly sensitive, good light stability, and without cancellation during high density, fluorescence spectrum such as not to drift about at the advantage, is expected to be applied to the field such as bio-imaging, fluorescent tracing detection.
3) material (chitosan, hydroxyapatite) used in the present invention all has good biocompatibility.
Accompanying drawing explanation
Fig. 1 is the transmission electron microscope photo of aggregation-induced emission namo fluorescence probe.
Embodiment
The present invention is further illustrated below in conjunction with example.
Embodiment 1:
1) 0.0555 g Calcium Chloride Powder Anhydrous, 0.0468 g SODIUM PHOSPHATE, MONOBASIC and 0.0294 g citric acid monohydrate trisodium is taken in 20 mL seed bottle, add 10 mL tri-distilled waters, drip 200 μ L Glacial acetic acid under magnetic agitation to dissolve, make 5 mg/mL hydroxyapatite precursor solutions;
2) the hydroxyapatite precursor solution tri-distilled water of step 1) is diluted to 1 mg/mL, stir lower dropping 40 wt% aqueous sodium hydroxide solution regulator solution pH=10, now hydroxyapatite nano particle is separated out, and continues stirring and spends the night, obtained hydroxyapatite nano particle solution;
3) be that to be dissolved in volumetric concentration be in 2% aqueous acetic acid for the AIE fluorescent mark chitosan of 0.1 mol% by mark rate, compound concentration is the AIE fluorescent mark chitosan-acetic acid solution of 1 mg/mL;
4) 1 mL step 2 is measured) obtained hydroxyapatite nano particle solution, centrifugal rear abandoning supernatant, recentrifuge after tri-distilled water cleaning, lower sediment is joined 1 mL step 3) in obtained AIE fluorescent mark chitosan-acetic acid solution, stir 1 h, centrifugal rear abandoning supernatant, add tri-distilled water and make hydroxyapatite nano particle concentration be 1 mg/mL, stir, the obtained namo fluorescence probe with aggregation-induced emission characteristic.
The namo fluorescence probe that this example obtains is of a size of 111.9 nm, and surface potential is 11.51 mV, and its transmission electron microscope photo is shown in Fig. 1.
Embodiment 2:
1) 0.0555 g Calcium Chloride Powder Anhydrous, 0.0468 g SODIUM PHOSPHATE, MONOBASIC and 0.0557 g citric acid monohydrate trisodium is taken in 20 mL seed bottle, add 10 mL tri-distilled waters, drip 200 μ L Glacial acetic acid under magnetic agitation to dissolve, make 5 mg/mL hydroxyapatite precursor solutions;
2) the hydroxyapatite precursor solution tri-distilled water of step 1) is diluted to 1 mg/mL, stir lower dropping 40 wt% aqueous sodium hydroxide solution regulator solution pH=10, now hydroxyapatite nano particle is separated out, and continues stirring and spends the night, obtained hydroxyapatite nano particle solution;
3) be that to be dissolved in volumetric concentration be in 2% aqueous acetic acid for the AIE fluorescent mark chitosan of 20 mol% by mark rate, compound concentration is the AIE fluorescent mark chitosan-acetic acid solution of 3 mg/mL;
4) 1 mL step 2 is measured) obtained hydroxyapatite nano particle solution, centrifugal rear abandoning supernatant, recentrifuge after tri-distilled water cleaning, lower sediment is joined 1 mL step 3) in obtained AIE fluorescent mark chitosan-acetic acid solution, stir 1 h, centrifugal rear abandoning supernatant, add tri-distilled water and make hydroxyapatite nano particle concentration be 1 mg/mL, stir, the obtained namo fluorescence probe with aggregation-induced emission characteristic.
The namo fluorescence probe that this example obtains is of a size of 105.9 nm, and surface potential is 10.25 mV.
Embodiment 3:
1) 0.0555 g Calcium Chloride Powder Anhydrous, 0.0468 g SODIUM PHOSPHATE, MONOBASIC and 0.147 g citric acid monohydrate trisodium is taken in 20 mL seed bottle, add 10 mL tri-distilled waters, drip 200 μ L Glacial acetic acid under magnetic agitation to dissolve, make 5 mg/mL hydroxyapatite precursor solutions;
2) the hydroxyapatite precursor solution tri-distilled water of step 1) is diluted to 1 mg/mL, stir lower dropping 40 wt% aqueous sodium hydroxide solution regulator solution pH=10, now hydroxyapatite nano particle is separated out, and continues stirring and spends the night, obtained hydroxyapatite nano particle solution;
3) be that to be dissolved in volumetric concentration be in 2% aqueous acetic acid for the AIE fluorescent mark chitosan of 10 mol% by mark rate, compound concentration is the AIE fluorescent mark chitosan-acetic acid solution of 5 mg/mL;
4) 1 mL step 2 is measured) obtained hydroxyapatite nano particle solution, centrifugal rear abandoning supernatant, recentrifuge after tri-distilled water cleaning, lower sediment is joined 1 mL step 3) in obtained AIE fluorescent mark chitosan-acetic acid solution, stir 1 h, centrifugal rear abandoning supernatant, add tri-distilled water and make hydroxyapatite nano particle concentration be 1 mg/mL, stir, the obtained namo fluorescence probe with aggregation-induced emission characteristic.
The namo fluorescence probe that this example obtains is of a size of 111.5 nm, and surface potential is 13.28 mV.
Claims (2)
1. an aggregation-induced emission namo fluorescence probe, is characterized in that the chitosan fluorescence molecule at hydroxyapatite nano particle Surface coating with aggregation-induced emission characteristic, the structural formula of chitosan fluorescence molecule as the formula (1),
Formula (1)
In formula: m, x, y are respectively number of repeat unit.
2. prepare the method for aggregation-induced emission namo fluorescence probe according to claim 1, it is characterized in that step is as follows:
1) ratio 5:5:1 Calcium Chloride Powder Anhydrous, SODIUM PHOSPHATE, MONOBASIC and citric acid monohydrate trisodium being pressed amount of substance mixes, add tri-distilled water, drip Glacial acetic acid under magnetic agitation to dissolve, the consumption of Glacial acetic acid is 2% of mixeding liquid volume, and making concentration is 5 mg/mL hydroxyapatite precursor solutions;
2) the hydroxyapatite precursor solution tri-distilled water that step 1) is obtained is diluted to 1 mg/mL, under stirring, regulates pH=10 with alkaline solution, continue stirring and spend the night, obtained hydroxyapatite nano particle solution;
3) be that to be dissolved in volumetric concentration be in 2% aqueous acetic acid for the AIE fluorescent mark chitosan of 0.1 mol%-20 mol% by mark rate, compound concentration is the AIE fluorescent mark chitosan-acetic acid solution of 0.1-5 mg/mL;
4) by step 2) obtained hydroxyapatite nano particle solution centrifugal abandoning supernatant, again with tri-distilled water cleaning, centrifugal, precipitation is joined step 3) in obtained AIE fluorescent mark chitosan-acetic acid solution, stir at least 1 h, centrifugal abandoning supernatant, adding tri-distilled water makes hydroxyapatite nano particle concentration be 1 mg/mL, stirs, obtains aggregation-induced emission namo fluorescence probe.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106010503A (en) * | 2016-05-07 | 2016-10-12 | 浙江大学 | Chitosan-based fluorescence probe suitable for long circulation of blood and preparing method thereof |
| CN107311957A (en) * | 2017-06-21 | 2017-11-03 | 海南大学 | One kind is based on aggregation-induced emission and excited state intramolecular proton transfer compound and its preparation method and application |
| CN107502344A (en) * | 2017-09-12 | 2017-12-22 | 中南大学 | Based on dione pyrrolo-pyrrole compound and the multi-functional organic fluorescence nano particle of tetraphenyl ethylene class compound and preparation and application |
| CN107828052A (en) * | 2017-10-16 | 2018-03-23 | 华南理工大学 | Conjugated polymer with aggregation-induced emission property and its production and use |
| CN108559008A (en) * | 2018-06-25 | 2018-09-21 | 浙江大学 | A kind of n-trimethyl chitosan chloride with aggregation-induced emission characteristic/heparin composite Nano fluorescence probe and preparation method thereof |
| CN109810702A (en) * | 2019-02-20 | 2019-05-28 | 深圳大学 | A kind of namo fluorescence probe of good biocompatibility and preparation method thereof |
| CN110372685A (en) * | 2019-07-30 | 2019-10-25 | 天津大学 | The pH that new A IE molecule and its synthesis step, preparation generate active oxygen responds AIE nanoparticle approach |
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| CN104356345A (en) * | 2014-12-01 | 2015-02-18 | 四川大学 | Fluorescent graft degradable block polyurethane, bone repair material and preparation method thereof |
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| CN101643645A (en) * | 2009-07-24 | 2010-02-10 | 华中科技大学 | Calcium phosphate material marked by fluorescein isothiocyanate and preparation method thereof |
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106010503A (en) * | 2016-05-07 | 2016-10-12 | 浙江大学 | Chitosan-based fluorescence probe suitable for long circulation of blood and preparing method thereof |
| CN106010503B (en) * | 2016-05-07 | 2018-01-30 | 浙江大学 | One kind is suitable to macrocyclic chitosan-based fluorescence probe of blood and preparation method thereof |
| CN107311957A (en) * | 2017-06-21 | 2017-11-03 | 海南大学 | One kind is based on aggregation-induced emission and excited state intramolecular proton transfer compound and its preparation method and application |
| CN107502344A (en) * | 2017-09-12 | 2017-12-22 | 中南大学 | Based on dione pyrrolo-pyrrole compound and the multi-functional organic fluorescence nano particle of tetraphenyl ethylene class compound and preparation and application |
| CN107502344B (en) * | 2017-09-12 | 2019-07-19 | 中南大学 | Multi-functional organic fluorescence nano particle and preparation and application based on dione pyrrolo-pyrrole compound and tetraphenyl ethylene class compound |
| CN107828052A (en) * | 2017-10-16 | 2018-03-23 | 华南理工大学 | Conjugated polymer with aggregation-induced emission property and its production and use |
| CN108559008A (en) * | 2018-06-25 | 2018-09-21 | 浙江大学 | A kind of n-trimethyl chitosan chloride with aggregation-induced emission characteristic/heparin composite Nano fluorescence probe and preparation method thereof |
| CN108559008B (en) * | 2018-06-25 | 2020-11-13 | 浙江大学 | Quaternized chitosan/heparin composite nano fluorescent probe with aggregation-induced emission characteristic and preparation method thereof |
| CN109810702A (en) * | 2019-02-20 | 2019-05-28 | 深圳大学 | A kind of namo fluorescence probe of good biocompatibility and preparation method thereof |
| CN109810702B (en) * | 2019-02-20 | 2021-12-21 | 深圳大学 | Nano fluorescent probe with good biocompatibility and preparation method thereof |
| CN110372685A (en) * | 2019-07-30 | 2019-10-25 | 天津大学 | The pH that new A IE molecule and its synthesis step, preparation generate active oxygen responds AIE nanoparticle approach |
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