CN104844839A - Preparation method of magnetic fluorescent composite nanoparticle - Google Patents

Abstract

The invention belongs to the preparation field of nano-materials, and in particular relates to a preparation method of a magnetic fluorescent composite nanoparticle. The preparation method comprises the following steps: firstly using a magnetic nanoparticle with negative charges and modified by carboxymethyl chitosan as a core; and then under the action of poly diallyldimethylammonium choloride (PDDA), an ion cross-linking agent and a polyanion type macromolecular organic compound, linking the core with a water-soluble quantum dot by an ion crosslinking method to obtain the magnetic fluorescent composite nanoparticle. The fluorescent quantum dots provided by the present invention are uniformly distributed on the surface of the magnetic nanometer particle, wherein the particle size of the magnetic nanometer particle is 10-200 nm, and the particle size of the quantum dots is 1.5-10 nm. The preparation method according to the present invention has the advantages that the reaction conditions are mild, the operating method is simple, the prepared composite nanoparticle has favorable luminous performance and favorable magnetic performance, and the prepared composite nanoparticle can be used in the aspects of target positioning and bioluminescence imaging in organisms.

Description

A kind of preparation method of magnetic fluorescent composite nanoparticle

technical field

The invention relates to a method for preparing multifunctional nanocomposite microspheres, in particular to a method for preparing magnetic fluorescent composite nanoparticles. The composite particles have both the fluorescent properties of quantum dots and the magnetic properties of magnetic nanoparticles, and can be used as In vivo targeting and bioluminescence imaging.

Background technique

Quantum dots and magnetic nanoparticles have shown obvious advantages in many fields, especially in the field of biomedicine. However, they often only have a single function, such as fluorescent labeling of quantum dots, magnetic separation of magnetic particles, etc. If the two functions-separation and labeling functions are integrated, a new type of magnetic fluorescent nanometer can be prepared through a certain physical and chemical process. The composite material makes it a fluorescent probe and also has good magnetic response characteristics. Therefore, magnetic fluorescent nanoparticles have dual functions of magnetic separation and fluorescent tracking, and their performance and application range far exceed those of single-function nanoparticles. They can not only detect and separate biomolecules, but also realize multiple modes. Imaging, namely fluorescence imaging, magnetic resonance imaging (MRI) and confocal laser microscopy imaging.

So far, the common preparation methods of magnetic fluorescent composite materials include layer-by-layer self-assembly method, embedding method, and covalent bonding method. Encapsulation is a relatively simple and mature method to realize the assembly of magnetic nanoparticles and quantum dots. The main process is to pre-prepare magnetic nanoparticles and quantum dots, and then use surfactants, polyelectrolyte molecules, etc. to embed them together in the inner core to form magnetic fluorescent composite nano/microspheres. The use of polymers can retain the properties of the polymers while coating the two nanoparticles. Its shell material requires good biocompatibility and structural stability, and at the same time, it is easy to further surface functional modification, so as to connect with specific target molecules and finally achieve targeting.

Using biodegradable polymer materials as coating materials has the characteristics of non-toxicity, biodegradability, and good tissue compatibility, and has broad application prospects in the fields of medicine and biology. As a derivative of chitosan, carboxymethyl chitosan is a negatively charged water-soluble polysaccharide obtained by carboxymethylation of chitosan. It is non-toxic, non-irritating, non-allergenic, and non-toxic Mutagenesis, no hemolytic effect, good biocompatibility, biodegradability and adhesion, and at the same time overcome the defect that chitosan can only be dissolved in acidic solution, showing the excellent properties that ordinary chitosan does not have.

The ionic cross-linking method refers to the method in which the macromolecular chains in the raw materials are linked by chemical bonds under the action of an ionic cross-linking agent to form a network or body-shaped polymer material. The ionic cross-linking method is usually also called the physical cross-linking method. Its advantage is that no organic solvent is used in the preparation process, and it avoids the harm of most chemical cross-linking agents such as glutaraldehyde cross-linking agents to the human body, so this method is widely used .

Contents of the invention

The purpose of the present invention is to provide a method for preparing magnetic fluorescent composite nanoparticles by ion cross-linking. The composite nanoparticles prepared by the method have strong magnetic responsiveness, high photostability, simple preparation process, good dispersion and uniform size, etc. At the same time, due to the coating effect of polymer organic matter, the prepared composite nanoparticles also have the characteristics of high specific surface area, good biocompatibility, biofunctionality and biodegradability, and have a wider range of applications in the field of nanobiotechnology. Applications.

The present invention is achieved through the following measures:

A method for preparing magnetic fluorescent composite nanoparticles, using the following steps:

(a) Preparation of carboxymethyl chitosan magnetic nanocomposites (CMCH-MNPs) by blending embedding method:

① Weigh 50-500 mg of magnetic nanoparticles and dissolve them in 30 mL of PBS buffer solution with a pH = 7.4 and a concentration of 0.01mol/L, pass nitrogen gas to remove oxygen, and use an ultrasonic cleaner to disperse the magnetic particles evenly; ② Weigh 10-800 mg carboxymethyl chitosan was dissolved in 20 mL pH = 7.4 PBS buffer solution, while adding 0.1-0.5 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDAC ) to activate the groups on the surface of carboxymethyl chitosan; ③ Mix the two solutions ① and ②, shake in a constant temperature water bath shaker at 25°C for 2-6 h, and then use distilled water and PBS (pH=7.4) buffer solution to fully Washing to obtain the carboxymethyl chitosan magnetic nanocomposite;

(b) Preparation of magnetic fluorescent composite nanoparticles by ion cross-linking method:

Take the prepared concentrated aqueous phase quantum dots and dissolve them in distilled water, and add polyanionic macromolecular organic matter and ionic cross-linking agent, wherein the mass concentrations of polyanionic macromolecular organic matter and ionic cross-linking agent are 0.33-6.67 mg/ mL and 0.67-2.67 mg/mL to obtain reaction solution A; take 0.005-0.05 g of the carboxymethyl chitosan-modified magnetic nanocomposite prepared in step (a) and dissolve it in distilled water, sonicate for 20-60 min to make The magnetic nanoparticles are uniformly dispersed, and polydiallyldimethylammonium chloride is added to obtain the reaction solution B; the reaction solution A is added dropwise to the reaction solution B, and the reaction is shaken at room temperature for 2-10 h, centrifuged, distilled water After washing, the magnetic fluorescent composite nanoparticles are obtained; wherein the mass ratio of polydiallyldimethylammonium chloride: polyanionic macromolecular organic matter: ion crosslinking agent is 1:0.1-2:0.01-0.5.

The preparation of magnetic fluorescent composite nanoparticles in the present invention, the particle size of magnetic fluorescent composite nanoparticles is 10-220 nm; magnetic nanoparticles (MNPs) are metals and metal oxides with superparamagnetic, paramagnetic or ferromagnetic , selected from Fe ₃ O ₄ , Fe ₂ O ₃ , MeFe ₂ O ₄ (Me=Co, Mn, Ni), compounds NdFeB, SmCo, etc., metals Fe, Co, Ni and alloys Fe ₂ Co, Ni ₂ One of the nanoparticles of metal oxides of Fe; the preparation method of the magnetic nanoparticles includes a co-precipitation method and a hydrothermal method; the surface of the magnetic nanoparticles contains at least one of hydroxyl, amino and carboxyl groups.

The quantum dots of the present invention are water-soluble quantum dots with hydrophilic groups on the surface, and the quantum dots are II-VI group semiconductor materials, or composite materials formed by II-VI group semiconductor materials, and the particle size of the quantum dots is 1.5-10 nm; preferred quantum dots are ZnSe, CdSe, CdTe, CdS, ZnSe/ZnS, CdS/ZnS, CdSe/ZnS, CdTe/ZnS, Zn _X Cd _1-X Se, CdSe _1-X S _X , CdSe _1-X Te _X , CdSe/ZnSe, CdS/ZnSe, CdTe/ZnSe, CdSe/CdS, CdTe/CdS, CdS/Zn _X Cd _1-X S, ZnSe/Zn _X Cd _1-X S, CdSe/Zn One of _X Cd _1-X S or CdTe/Zn _X Cd _1-X S, where 0<X<1.

The ionic cross-linking agent of the present invention includes one or more of tripolyphosphate, tetrapolyphosphate, hexametaphosphate, pyrophosphate, molybdate, hyaluronic acid or γ-polyglutamic acid ;Polyanionic macromolecular organic matter sodium carboxymethyl cellulose, sodium cellulose sulfate, sodium cellulose carboxylate, cellulose carboxylate, carboxymethyl chitosan, potassium carboxymethyl cellulose or sodium carboxymethyl cellulose One or more of potassium; the carboxymethyl chitosan can be selected from O-carboxymethyl chitosan or N, O-carboxymethyl chitosan; polydiallyl dimethyl chloride The molecular weight and mass concentration of ammonium are 1×10 ⁵ ~2×10 ⁵ and 0.51~5.1 mg/mL, respectively.

The preparation of magnetic fluorescent composite nanoparticles of the present invention, the thickness of carboxymethyl chitosan on the surface of magnetic nanoparticles in step (a) is 3-10 nm; the magnetic nanocomposite of carboxymethyl chitosan in step (b) The molar ratio of matter to quantum dots is 1:1~1:20.

The quantum dot surface of the present invention contains at least one of mercapto, carboxyl, and amino groups; the hydrophilic group ligands used in the synthesis of semiconductor quantum dots include 3-mercaptopropionic acid, thioglycolic acid, L-cysteine, One or more of 2-mercaptopropionic acid, mercaptobutyric acid, mercaptovaleric acid, mercaptocaproic acid, mercaptosuccinic acid, mercaptoethanol, glutathione, mercaptopropanol or mercaptoethylamine.

Beneficial effects of the present invention :

(1) In the present invention, water-soluble carboxymethyl chitosan and other polysaccharide derivatives are selected to replace common chitosan, avoiding the use of acidic medium, eliminating the adverse effects caused by acidic medium, and expanding the scope of application in the field of biomedicine. scope of application;

(2) The present invention uses the electrostatic interaction between oppositely charged polyelectrolyte substances to assemble magnetic fluorescent composite nanoparticles through ionic cross-linking, without using organic solvents, and the reaction conditions are mild, which can protect the activity of biomolecules to the greatest extent;

(3) The present invention selects polydiallyldimethylammonium chloride (PDDA) as the cationic polyelectrolyte, which is safe, non-toxic, easily soluble in water, has good hydrolytic stability and strong cohesion, and avoids the use of Biological toxicity caused by other organic polymer electrolytes;

(4) Due to the modification of magnetic particles by carboxymethyl chitosan in the present invention, the leakage of magnetic materials and the fluorescence quenching effect on quantum dots are effectively avoided; the particle size, fluorescent properties and magnetic properties of the final product, Magnetic fluorescent composite nanoparticles with different properties can be prepared by adjusting the molar ratio of water-soluble quantum dots to magnetic nanoparticles, the concentration of polyanionic polymer organic matter, and the concentration of ion cross-linking agents, which can be used for biologically oriented drugs. Transportation and biological fluorescence imaging; at the same time, the preparation process of the invention is simple, the conditions are mild, and the active substances can be protected to the greatest extent.

Description of drawings

Fig.1 XRD patterns of magnetic fluorescent composite nanoparticles

Fig.2 Fluorescence and absorption spectra of magnetic fluorescent composite nanoparticles

Fig.3 TEM photo of magnetic fluorescent composite nanoparticles

Fig. 4 Hysteresis loops of magnetic fluorescent composite nanoparticles.

Detailed ways

The technical solution of the present invention is described below through specific examples, but the technical solution of the present invention is not limited to the specific examples.

Example 1:

1.1 Preparation of CdTe quantum dots in aqueous phase. Under nitrogen protection, dissolve 0.0945g NaBH ₄ and 0.0063g Te powder in 5 mL distilled water, heat to 40°C, and obtain NaHTe solution after completely dissolving; take 0.293 g Cd(Ac) ₂ and dissolve it in 100 mL distilled water, wait After it was completely dissolved, 2.0 mmol thioglycolic acid was added, and the pH was adjusted to 11 with 1 mol/L NaOH solution to obtain a Cd precursor solution; the Cd precursor was transferred into a three-necked flask, and NaHTe solution was quickly injected under nitrogen protection, and the oil The bath was refluxed at 100°C, magnetically stirred for 3 hours, then taken out and placed in a refrigerator to cool rapidly to room temperature to obtain a red solution, which was the prepared CdTe QDs solution.

1.2 Preparation of Fe ₃ O ₄ nanoparticles by hydrothermal method. Weigh 3 g of FeCl ₃ and dissolve it in 80 mL of ethylene glycol, stir and ultrasonically dissolve, add 2 g of polyethylene glycol with a molecular weight of 2000, and 7 g of sodium acetate, stir and ultrasonically dissolve it, and transfer the precursor solution to hydrothermal reaction In the kettle, react at 200°C for 5 h. After the reaction is completed, the obtained solution is alternately washed with water and absolute ethanol, and dried in vacuum to obtain dry Fe ₃ O ₄ magnetic nanoparticles.

1.3 The carboxymethyl chitosan magnetic nanocomposite (Fe ₃ O ₄ -CMCH) was prepared by blending embedding method. Weigh 0.08 g of Fe ₃ O ₄ magnetic nanoparticles and dissolve them in 30 mL of PBS buffer solution with a pH of 7.4 and a concentration of 0.01mol/L, pass nitrogen gas to remove oxygen, and use an ultrasonic cleaner to disperse the magnetic particles evenly to obtain a uniformly dispersed Fe ₃ O ₄ nanoparticle solution; Weigh 0.2 g carboxymethyl chitosan and dissolve it in 20 mL pH = 7.4 PBS buffer solution, add 0.1 g 1-ethyl-(3-dimethylaminopropyl) at the same time Carbodiimide hydrochloride (EDAC) activates the groups on the surface of carboxymethyl chitosan; then the carboxymethyl chitosan solution is added to the Fe ₃ O ₄ nanoparticle solution, and shaken in a constant temperature water bath shaker at 25°C for 2 h, after the reaction, the product was collected with a magnet, and then washed thoroughly with distilled water and PBS (pH=7.4) buffer solution in turn to obtain the carboxymethyl chitosan magnetic nanocomposite.

1.4 Prepare magnetic fluorescent composite nanoparticles by ionic cross-linking method. Take the concentrated water-phase CdTe quantum dots prepared in 1.1, separate and purify them with isopropanol, dissolve them in 5 mL of distilled water, and add sodium polyphosphate and carboxymethyl chitosan, wherein sodium polyphosphate and carboxymethyl chitosan The concentration of CdTe quantum dots was 1 mg/mL and 1.5 mg/mL respectively, and the concentration of CdTe quantum dots was 0.04mol/L to obtain reaction solution A; take 0.01g of carboxymethyl chitosan modified magnetic nanocomposite solution prepared in step 1.3 In 10 mL of distilled water, sonicate for 30 min to disperse the magnetic nanoparticles evenly, and add polydiallyldimethylammonium chloride with a mass concentration of 1 mg/mL to obtain a reaction solution B; add solution A to B dropwise The solution was shaken in a constant temperature water bath oscillator at 25° C. for 4 hours, the reaction product was collected by a magnet, and the reaction product was washed with distilled water to obtain a magnetic fluorescent bifunctional nanomaterial. Fig. 1 XRD patterns of magnetic fluorescent composite nanoparticles.

Example 2:

2.1 Preparation of CdTe/ZnS quantum dots by the aqueous phase method. First, the chemical co-precipitation method was used to prepare CdTe quantum dots. The preparation method was as described in Example 1 above, except that the reaction time was 4h. Take the prepared CdTe quantum dots and dissolve them in 30 mL distilled water, in which the concentration of CdTe quantum dots is 1.67×10 ^-3 mol/L, add 0.0329 g Zn(Ac) ₂ , 0.0921 g reduced glutathione, and use the configuration The best concentration is 1 mol/L NaOH solution to adjust the pH value to 8. Under magnetic stirring, the oil bath is refluxed at 100 ° C. After the magnetic stirring reaction for 2 hours, put it in the refrigerator and quickly cool it to room temperature to obtain water-soluble CdTe/ ZnS quantum dots.

2.2 Preparation of Fe ₃ O ₄ magnetic nanoparticles by hydrothermal method. Dissolve 2.78 g FeSO ₄ 7H ₂ O and 4.32 g FeCl ₃ 6H ₂ O in 30 mL of distilled water, stir to dissolve, then add 30 mL of ethylene glycol, stir evenly, and put it into a three-necked flask under nitrogen protection , adjust the pH to 9-11 with the prepared NaOH solution with a concentration of 2 mol/L, add 0.15 g of surfactant polyvinylpyrrolidone, stir well and react for 30 min to prepare the precursor of Fe ₃ O ₄ magnetic nanoparticles. The precursor of Fe ₃ O ₄ magnetic nanoparticles was transferred into a hydrothermal reactor and reacted at 160°C for 6h. After the reaction was completed, the obtained solution was alternately washed with water and absolute ethanol, and dried in vacuum to obtain dry Fe ₃ O ₄ magnetic nanoparticles.

2.3 The carboxymethyl chitosan magnetic nanocomposite (Fe ₃ O ₄ -CMCH) was prepared by blending embedding method. Weigh 0.2 g of Fe ₃ O ₄ magnetic nanoparticles and dissolve them in 30 mL of PBS buffer solution with a pH of 7.4 and a concentration of 0.01 mol/L, pass nitrogen gas to remove oxygen, and use an ultrasonic cleaner to disperse the magnetic particles evenly to obtain a uniformly dispersed Fe ₃ O ₄ nanoparticle solution; Weigh 0.5 g carboxymethyl chitosan and dissolve it in 20 mL PBS buffer solution with pH = 7.4, add 0.3 g 1-ethyl-(3-dimethylaminopropyl) at the same time Carbodiimide hydrochloride (EDAC) activated the groups on the surface of carboxymethyl chitosan; then the carboxymethyl chitosan solution was added to the Fe ₃ O ₄ nanoparticle solution and shaken in a constant temperature water bath shaker at 25°C for 4 h, after the reaction, the product was collected with a magnet, and then washed thoroughly with distilled water and PBS (pH=7.4) buffer solution in turn to obtain the carboxymethyl chitosan magnetic nanocomposite.

2.4 Preparation of magnetic fluorescent composite nanoparticles by ion cross-linking method. Take the concentrated water-phase CdTe/ZnS quantum dots prepared in 2.1, separate and purify them with isopropanol, dissolve them in 5 mL of distilled water, and add sodium tetrapolyphosphate and sodium carboxymethylcellulose, wherein sodium tetrapolyphosphate and carboxymethylcellulose The concentration of plain sodium was 1.5 mg/mL and 3 mg/mL respectively, and the concentration of CdTe/ZnS quantum dots was 0.05mol/L to obtain reaction solution A; take 0.02 g of carboxymethyl chitosan-modified magnetic The nanocomposite was dissolved in 10 mL distilled water, ultrasonicated for 30 min to disperse the magnetic nanoparticles evenly, and polydiallyldimethylammonium chloride was added with a mass concentration of 2 mg/mL to obtain reaction solution B; Add dropwise to solution B, shake in a constant temperature water bath shaker at 25°C for 6 hours, collect the reaction product with a magnet, wash the reaction product with distilled water, and obtain the magnetic fluorescent bifunctional nanomaterial. Fig. 2 is the fluorescence spectrum and absorption spectrum of the prepared magnetic fluorescent composite nanoparticles.

Example 3:

3.1 Preparation of ZnSe/ZnS quantum dot solution. First, ZnSe quantum dots were prepared by chemical co-precipitation method. Under the protection of nitrogen, 0.01g NaBH ₄ and 0.0061g Se powder were dissolved in 2 mL distilled water, heated to 40°C, and NaHSe solution was obtained after complete dissolution; 0.0439 g Zn( Ac) ₂ was dissolved in 20 mL of distilled water, and after it was completely dissolved, 0.0737 g of reduced glutathione was added, and the pH was adjusted to 11.5 with 1mol/L NaOH solution to obtain a Zn precursor solution; Into a three-necked flask, quickly inject NaHSe solution under nitrogen protection, reflux in an oil bath at 100°C, magnetically stir for 1 hour, take it out and put it in a refrigerator to cool rapidly to room temperature to obtain a colorless and transparent solution, which is the prepared ZnSe QDs solution. Take 15 mL of prepared ZnSe quantum dots, in which the concentration of ZnSe quantum dots is 2.7×10 ^-3 mol/L, add 0.0138 g Zn(Ac) ₂ , 0.0277 g reduced glutathione and 0.01 g thiourea, and use The prepared NaOH solution with a concentration of 1 mol/L adjusts the pH value to 10.5. Under magnetic stirring, the oil bath is refluxed at 100°C. After 2 hours of magnetic stirring reaction, put it in the refrigerator and quickly cool it to room temperature to obtain light yellow water-soluble high luminous efficiency ZnSe/ZnS quantum dots.

3.2 Preparation of magnetic nanoparticles. The Fe ₃ O ₄ /CoO core-shell magnetic nanoparticles are prepared by adopting the method with the patent publication number of CN101597495A.

3.3 Preparation of carboxymethyl chitosan magnetic nanocomposites by blending embedding method. Weigh 0.35 g of Fe ₃ O ₄ /CoO magnetic nanoparticles and dissolve them in 30 mL of PBS buffer solution with a pH of 7.4 and a concentration of 0.01mol/L, pass nitrogen gas to remove oxygen, and use an ultrasonic cleaner to disperse the magnetic particles evenly to obtain a dispersed Homogeneous Fe ₃ O ₄ /CoO nanoparticle solution; weigh 0.65 g carboxymethyl chitosan and dissolve it in 20 mL of PBS buffer solution with pH = 7.4, and add 0.3 g 1-ethyl-(3-dimethyl Aminopropyl) carbodiimide hydrochloride (EDAC) activates the groups on the surface of carboxymethyl chitosan; then add the carboxymethyl chitosan solution to the Fe ₃ O ₄ /CoO nanoparticle solution and shake in a constant temperature water bath Shake in a container at 25°C for 5 h. After the reaction, the product was collected with a magnet, and then thoroughly washed with distilled water and PBS (pH=7.4) buffer solution to obtain carboxymethyl chitosan magnetic nanocomposites.

3.4 Preparation of magnetic fluorescent composite nanoparticles by ionic cross-linking method. Take the concentrated aqueous phase ZnSe/ZnS quantum dots prepared in 3.1, separate and purify them with isopropanol, dissolve them in 5 mL of distilled water, and add sodium pyrophosphate and sodium cellulose sulfate, wherein the concentrations of sodium pyrophosphate and sodium cellulose sulfate are respectively 2 mg/mL and 4 mg/mL, the concentration of ZnSe/ZnS quantum dots was 0.1 mol/L, and the reaction solution A was obtained; 0.035 g of the carboxymethyl chitosan-modified magnetic nanocomposite prepared in step 3.3 was dissolved in 10 Sonicate in mL distilled water for 30 min to disperse the magnetic nanoparticles evenly, and add polydiallyldimethylammonium chloride with a mass concentration of 2 mg/mL to obtain reaction solution B; add solution A to solution B dropwise , shake in a constant temperature water bath oscillator at 25°C for 6h, collect the reaction product with a magnet, wash the reaction product with distilled water, and obtain the magnetic fluorescent bifunctional nanomaterial. Fig. 3 is a TEM photo of the prepared magnetic fluorescent composite nanoparticles.

Example 4:

4.1 Prepare CdSe quantum dots by the aqueous phase method. Under the protection of nitrogen, dissolve 0.0106 g NaBH ₄ and 0.0063 g Se powder in 2 mL distilled water, heat to 40 °C, and obtain NaHSe solution after complete dissolution; take 0.0533 g Cd(Ac ) ₂ was dissolved in 20 mL of distilled water, and after it was completely dissolved, 1.0 mmol of mercaptopropionic acid was added, and the pH was adjusted to 11.5 with 1 mol/L NaOH solution to obtain a Cd precursor solution; transfer the Cd precursor into a three-necked flask , quickly inject NaHSe solution under the protection of nitrogen, reflux the oil bath at 100 ° C, magnetically stir for 1 hour, take it out and put it in the refrigerator to cool to room temperature quickly to obtain a colorless and transparent solution, which is the prepared CdSe QDs solution.

4.2 Preparation of magnetic nanoparticles. The method of patent publication number CN 101928043 A is used to prepare α-Fe ₂ O ₃ magnetic nanoparticles.

4.3 Carboxymethyl chitosan magnetic nanocomposites (CMCH-MNPs) were prepared by blending embedding method. Weigh 0.5 g of α-Fe ₂ O ₃ magnetic nanoparticles and dissolve them in 30 mL of PBS buffer solution with a pH of 7.4 and a concentration of 0.01mol/L, pass nitrogen gas to remove oxygen, and use an ultrasonic cleaner to disperse the magnetic particles evenly to obtain a dispersed Uniform α-Fe ₂ O ₃ nanoparticle solution; weigh 0.8 g carboxymethyl chitosan and dissolve it in 20 mL of PBS buffer solution with pH = 7.4, and add 0.5 g 1-ethyl-(3-dimethyl Aminopropyl) carbodiimide hydrochloride (EDAC) activates the groups on the surface of carboxymethyl chitosan; then add the carboxymethyl chitosan solution to the α-Fe ₂ O ₃ nanoparticle solution and shake in a constant temperature water bath Shake in a container at 25°C for 6 h. After the reaction, the product was collected with a magnet, and then thoroughly washed with distilled water and PBS (pH=7.4) buffer solution to obtain carboxymethyl chitosan magnetic nanocomposites.

4.4 Preparation of magnetic fluorescent composite nanoparticles by ionic cross-linking method. Take the concentrated aqueous CdSe quantum dots prepared in 4.1, separate and purify them with isopropanol, dissolve them in 5 mL of distilled water, and add sodium molybdate and cellulose carboxylate, where the concentrations of sodium molybdate and cellulose carboxylate are 2.5 mg/mL and 6 mg/mL, and the concentration of CdSe quantum dots was 0.25 mol/L to obtain reaction solution A; take 0.05 g of the carboxymethyl chitosan-modified magnetic nanocomposite prepared in step 4.3 and dissolve it in 10 mL of distilled water , sonicate for 30 min to disperse the magnetic nanoparticles evenly, and add polydiallyldimethylammonium chloride with a mass concentration of 5 mg/mL to obtain reaction solution B; add solution A dropwise to solution B, ℃ in a constant temperature water bath oscillator for 10 h, the reaction product was collected by a magnet, and the reaction product was washed with distilled water to obtain a magnetic fluorescent bifunctional nanomaterial. Figure 4 is the hysteresis loop of the prepared magnetic fluorescent composite nanoparticles.

Claims (7)

1. the preparation of a magnetic fluorescence composite nanometer particle, it is characterized in that: be connected water-soluble quantum dot with polyanionic macromolecule organic by ionic cross-linking obtain with the sugar-modified magnetic nano-particle of electronegative carboxymethyl chitosan, diallyl dimethyl ammoniumchloride (PDDA), ion crosslinking agent, described fluorescence quantum is evenly distributed on the surface of magnetic nanoparticle with single dispersing form, wherein the size of magnetic nanoparticle is at 10-200 nm, and the particle diameter of quantum dot is at 1.5-10 nm.

2. the preparation of a kind of magnetic fluorescence composite nanometer particle according to claim 1, it is characterized in that, described preparation method comprises the following steps:

A () adopts blended entrapping method to prepare cm-chitosan magnetic nanocomposites (Fe ₃o ₄-CMCH): 1. take 50-500mg magnetic nanoparticle and be dissolved in 30 mL pH=7.4, concentration is in the PBS buffered soln of 0.01mol/L, and logical nitrogen deoxidation, utilizes Ultrasonic Cleaners to be uniformly dispersed by magnetic particle; 2. take 10-800 mg cm-chitosan to be dissolved in the PBS buffered soln of 20 mL pH=7.4, add the group that 0.1-0.5 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDAC) activates cm-chitosan surface simultaneously; 3. incite somebody to action 1. and 2. two solution mixing, 25 DEG C of vibration 2-6 h in water bath with thermostatic control vibrator, then use distilled water and PBS (pH=7.4) buffered soln fully to wash successively, namely obtain cm-chitosan magnetic nanocomposites;

B () adopts ionic cross-linking to prepare magnetic fluorescence composite nanometer particle:

The concentrated aqueous phase quantum point getting preparation is dissolved in distilled water, add polyanionic macromolecule organic and ion crosslinking agent, wherein the mass concentration of polyanionic macromolecule organic and ion crosslinking agent is respectively 0.33-6.67 mg/mL and 0.67-2.67 mg/mL, obtains reaction soln A; The magnetic nanocomposites getting the carboxymethyl chitosan of preparation in 0.005-0.05g step (a) sugar-modified is dissolved in distilled water, and ultrasonic 20-60 min, makes magnetic nanometer particles be uniformly dispersed, and adds diallyl dimethyl ammoniumchloride and obtain reaction soln B; Dropwise joined by reaction soln A in reaction soln B, oscillatory reaction 2-10 h under normal temperature, centrifugation, distilled water wash, namely obtains magnetic fluorescence composite nanometer particle; Wherein diallyl dimethyl ammoniumchloride: polyanionic macromolecule organic: the mass ratio of ion crosslinking agent is 1:0.1-2:0.01-0.5.

3. the preparation of a kind of magnetic fluorescence composite nanometer particle according to claim 1 and 2, is characterized in that, the particle diameter of described magnetic fluorescence composite nanometer particle is 10-220 nm; Described magnetic nanoparticle (MNPs) is superparamagnetic, paramagnetic or ferromagnetic metal and metal oxide, is selected from Fe ₃o ₄, Fe ₂o ₃, MeFe ₂o ₄(Me=Co, Mn, Ni), compound neodymium iron boron, SmCo etc., metal Fe, Co, Ni and alloy Fe ₂co, Ni ₂one in the nano particle of the metal oxide of Fe; The preparation method of magnetic nanoparticle comprises coprecipitation method, hydrothermal method; Magnetic nanoparticle surface is containing at least one in hydroxyl, amino, carboxyl.

4. a kind of magnetic fluorescence composite nanometer particle according to claim 1 and 2, it is characterized in that, described quantum dot is the water-soluble quantum dot of surface with hydrophilic radical, quantum dot is II-VI group semiconductor material, or be the matrix material that II-VI group semiconductor material is formed, described quantum point grain diameter is 1.5-10 nm; Preferred quantum dot is ZnSe, CdSe, CdTe, CdS, ZnSe/ZnS, CdS/ZnS, CdSe/ZnS, CdTe/ZnS, Zn _xcd _1-Xse, CdSe _1-Xs _x, CdSe _1-Xte _x, CdSe/ZnSe, CdS/ZnSe, CdTe/ZnSe, CdSe/CdS, CdTe/CdS, CdS/Zn _xcd _1-Xs, ZnSe/Zn _xcd _1-Xs, CdSe/Zn _xcd _1-Xs or CdTe/Zn _xcd _1-Xone in S, wherein 0 < X < 1.

5. a kind of magnetic fluorescence composite nanometer particle according to claim 1 and 2, it is characterized in that, described ion crosslinking agent comprises one or more in tri-polyphosphate, four poly-phosphate, hexametaphosphate, pyrophosphate salt, molybdate, hyaluronic acid or gamma-polyglutamic acid-; One or more in polyanionic macromolecule organic Xylo-Mucine, Ushercell, Mierocrystalline cellulose carboxylic acid sodium, Carboxylic Acid Fibre element, cm-chitosan, potassium carboxymethylcellulose or Xylo-Mucine potassium; Described cm-chitosan can select O-CMC or CMC; Molecular weight and the mass concentration of diallyl dimethyl ammoniumchloride are respectively 1 × 10 ⁵-2 × 10 ⁵with 0.51-5.1 mg/mL.

6. a kind of magnetic fluorescence composite nanometer particle according to claim 2, is characterized in that, in step (a), the thickness of cm-chitosan on magnetic nanoparticle surface is 3-10 nm; In step (b), the mol ratio of cm-chitosan magnetic nanocomposites and quantum dot is 1:1-1:20.

7. the preparation of water-soluble quantum dot according to claim 4, is characterized in that: described quantum dot surface contains at least one in sulfydryl, carboxyl, amino; The hydrophilic radical part used in semiconductor-quantum-point synthesis comprises one or more in 3-thiohydracrylic acid, Thiovanic acid, Cys, 2 mercaptopropionic acid, mercaptobutyric acid, mercaptopentanoic acid, mercaptohexanoic acid, dimercaptosuccinic acid, mercaptoethanol, gsh, mercaprol or mercaptoethylamine.