KR20220097180A - Method for manufacturing magnetic nanoparticles conjugated with chelating ligand and magnetic nanoparticles conjugated with chelating ligand manufactured thereby - Google Patents

Abstract

The present invention provides (a) magnetite (Fe ₃ O ₄ ) by modifying the particle surface to introduce a functional group capable of forming a peptide bond with nitrilotriacetic acid (NTA), and (b) the functional group on the surface A method for preparing magnetic nanoparticles conjugated with a chelate ligand comprising the step of conjugating nitrilotriacetic acid to the surface of the magnetite particles by reacting the introduced magnetite particles with nitrilotriacetic acid, and magnetic nanoparticles conjugated with a chelate ligand prepared thereby As for, according to the present invention, magnetite (magnetite, Fe ₃ O ₄ ) The surface of the nanoparticles, (3-aminopropyl) triethoxysilane (APTES) or polyethylenimine (PEI) using an amino group functionalized or using polyacrylic acid (PAA) Unlike the prior art, mass synthesis is possible by preparing magnetic nanoparticles by conjugating a chelating ligand (NTA) after functionalizing the carboxyl group.

Description

Method for manufacturing magnetic nanoparticles conjugated with chelate ligands and magnetic nanoparticles prepared by conjugated chelate ligands

The present invention relates to a method for preparing magnetic nanoparticles having a chelate ligand attached to the surface and magnetic nanoparticles to which a chelate ligand is conjugated.

Chelate-based sensors play an important role in environmental and biological fields, such as metal and chemical detectors, humidity sensors, and biosensors.

In a chelate-based sensor, metal ions are attached only to the chelating ligand on the surface of the sensing particle and serve as a stimulator, and the sensing particle can be stimulated by a chemical substance that is secondary to a chelate and metal ion pair. .

This mechanism is generally determined by the type of chelate with different affinity for functional groups and other substances, such as nitrilotriacetic acid (NTA), ethylenediaminediacetate (EDDA), and ethylenediaminetetraacetic acid (EDTA).

A study on synthesizing magnetic nanoparticles surface-modified with chelates such as Fe ₃ O ₄ /PAM(polyacrylamide)/NTA-Ni ²⁺ , etc. is reported for improved performance of chelate-based sensors and easy recovery of sensor particles However, according to the prior art such as the corresponding synthesis method, complex reaction conditions such as reflux for a long time and injection of inert gas during synthesis are required, which makes it unsuitable for mass synthesis.

M.T. Gabr and F.C. Pigge, Dalton T. 45 (2016) 14039-14043. H. Guo, M. Li, S. Tu, and H. Sun, IOP Conf. Ser.: Mater. Sci. Eng. 322 (2018) 022017.

An object of the present invention is to provide a novel manufacturing method capable of mass-synthesizing magnetic nanoparticles surface-modified with a chelate, unlike the prior art, and magnetic nanoparticles conjugated with a chelate ligand prepared thereby.

In order to achieve the above technical object, the present invention provides a functional group capable of forming a peptide bond with nitrilotriacetic acid (NTA) by modifying the surface of the magnetite (Fe ₃ O ₄ ) particle (a) introducing a functional group, and (b) reacting the magnetite particles having the functional group introduced to the surface with nitrilotriacetic acid to bond nitrilotriacetic acid to the surface of the magnetite particles.

In addition, in step (a), the magnetite (Fe ₃ O ₄ ) particle surface is modified with (3-aminopropyl) triethoxysilane (APTES) to introduce an amino group and react with glutaraldehyde to form a terminal A method for preparing magnetic nanoparticles conjugated with a chelate ligand, characterized in that it forms an aldehyde group, is proposed.

In addition, in step (a), the magnetite (Fe ₃ O ₄ ) particle surface is modified with polyethyleneimine (PEI) to introduce an amino group and react with glutaraldehyde to form an aldehyde group at the end, characterized in that A method for preparing magnetic nanoparticles conjugated with a chelating ligand is proposed.

In addition, in step (a), magnetite (Fe ₃ O ₄ ) We propose a method for producing magnetic nanoparticles conjugated with a chelate ligand, characterized in that the particle surface is modified with polyacrylic acid (PAA) to introduce a carboxyl group.

And, in another aspect of the present invention, a magnetic nanoparticle to which a chelating ligand prepared according to the above preparation method is conjugated is proposed.

According to the present invention, magnetite (magnetite, Fe ₃ O ₄ ) The surface of the nanoparticles, (3-aminopropyl) triethoxysilane (APTES) or polyethylenimine (PEI) to functionalize an amino group, or polyacrylic acid (PAA) to functionalize a carboxyl group Unlike the prior art, mass synthesis is possible by preparing magnetic nanoparticles by conjugating a chelating ligand (NTA) after fusion.

1 is a TEM image of as-synthesized Fe ₃ O ₄ and intermediate functionalized Fe ₃ O ₄ nanoparticles.
2 is an FT-IR spectrum of as-synthesized Fe ₃ O ₄ and intermediate functionalized Fe ₃ O ₄ nanoparticles.
3 is a schematic diagram showing the chemical structure of NTA-conjugated Fe ₃ O ₄ nanoparticles synthesized through various types of intermediate functionalization.
4 is an FT-IR image of NTA-conjugated Fe ₃ O ₄ nanoparticles synthesized through various types of intermediate functionalization.
5 is a TEM spectrum of NTA-conjugated Fe ₃ O ₄ nanoparticles synthesized through various types of intermediate functionalization.
6 is a result of measuring the surface charge of as-synthesized Fe ₃ O ₄ nanoparticles, intermediate functionalized Fe ₃ O ₄ nanoparticles, and NTA-conjugated Fe ₃ O ₄ nanoparticles. .
7 is a hysteresis curve of as-synthesized Fe ₃ O ₄ nanoparticles and NTA-conjugated Fe ₃ O ₄ nanoparticles.

In describing the present invention, if it is determined that a detailed description of a related well-known function or configuration may unnecessarily obscure the gist of the present invention, the detailed description thereof will be omitted.

Since the embodiment according to the concept of the present invention may have various changes and may have various forms, specific embodiments will be illustrated in the drawings and described in detail in the present specification or application. However, this is not intended to limit the embodiment according to the concept of the present invention to a specific disclosed form, and should be understood to include all changes, equivalents, or substitutes included in the spirit and scope of the present invention.

The terms used herein are used only to describe specific embodiments, and are not intended to limit the present invention. The singular expression includes the plural expression unless the context clearly dictates otherwise. In the present specification, terms such as "comprise" or "have" are intended to designate that the described feature, number, step, operation, component, part, or a combination thereof exists, but one or more other features or numbers , it should be understood that it does not preclude the possibility of the existence or addition of steps, operations, components, parts, or combinations thereof.

Hereinafter, the present invention will be described in more detail by way of examples.

Embodiments according to the present specification may be modified in various other forms, and the scope of the present specification is not to be construed as being limited to the embodiments described below. The embodiments of the present specification are provided to more completely explain the present specification to those of ordinary skill in the art.

<Example>

1. intermediate functionalized Fe ₃ O ₄ Nanoparticles and NTA-conjugated Fe ₃ O ₄ Synthesis of nanoparticles

(1-1) Core-shell Fe functionalized with amino groups using APTES ₃ O ₄ @SiO ₂ Synthesis of nanoparticles

First, Fe ₃ O ₄ nanoparticles were synthesized according to a known in-situ SiO ₂ coating process [GS An, SW Choi, DH Chae, HS Lee, H.-J. Kim, Y. Kim, Y.-G. Jung, and S.-C. Choi, Ceram. Int. 43 (2017) 12888-12892.; GS An, JS Han, JR Shin, DH Chae, JU Hur, H.-Y. Park, Y.-G. Jung, and S.-C. Choi, Ceram. Int. 44 (2018) 12233-12237]

After adjusting the solution of Fe ₃ O ₄ nanoparticles (2g) dispersed in a mixed solvent containing 450mL ethanol and 50mL distilled water to basic pH, (3-aminopropyl)triethoxysilane (APTES) 4 wt% of an ethanol-based solution was added dropwise. . After vigorous stirring for 8 hours, the amino-functionalized nanoparticles were collected by magnetic field and washed several times with ethanol and distilled water.

(1-2) Fe with amino groups functionalized using PEI ₃ O ₄ Synthesis of nanoparticles

Fe ₃ O ₄ nanoparticles (2g) were dissolved in 100 mL of distilled water and 4 wt% of polyethylenimine (PEI) solution. Next, the solution was mixed at room temperature for 30 minutes, heated to 80° C., reacted for 10 hours with vigorous stirring, and cooled until it reached room temperature. The obtained nanoparticles were collected by inducing a magnetic field, washed several times with distilled water, and then stored as a solution in distilled water.

(1-3) Fe functionalized with amino groups ₃ O ₄ Formation of NTA chelate ligands on the surface of nanoparticles

A total of 0.1 g of amino group-functionalized Fe ₃ O ₄ nanoparticles containing APTES and PEI were prepared as a solution with 100 mL of distilled water and mechanically stirred before reaction. Then, a glutaraldehyde solution was injected and mechanically stirred for 8 hours. The particles were collected by applying a magnetic field, washed with distilled water to remove unreacted reagents, and the acidity was adjusted to neutral. Then, it was redispersed in 100 mL of distilled water and mixed with a solution of Nα,Nα-Bis(carboxymethyl)-Llysine hydrate (ABNTA, C ₁₀ H ₁₈ N ₂ O _6· xH ₂ O). The reaction was carried out for 24 hours at room temperature under vigorous stirring conditions. Finally, Fe ₃ O ₄ nanoparticles to which a chelating ligand is attached were collected through a magnet. It was then washed and stored as a solution in distilled water.

(2) Fe functionalized with a carboxyl group using PAA ₃ O ₄ Chelate Functionalization of Nanoparticles

A dispersion solution (300 mL) containing 10 g of Fe ₃ O ₄ nanoparticles washed several times with distilled water was prepared, and polyacrylic acid (PAA) was prepared as a 100 mL solution of 4 wt%. These solutions were mixed in a round bottom flask and heated to 75° C. for 4 hours, then the mixed solution was mechanically stirred at 300 rpm until cooled to room temperature. Next, 0.25 g of PAA-attached Fe ₃ O ₄ was redispersed in 100 mL of distilled water and mixed with 100 mL of ABNTA to obtain a solution having a concentration of 0.1 wt%, followed by reaction for 8 hours through mechanical stirring. Thereafter, the reacted Fe ₃ O ₄ nanoparticles were washed and stored as a solution in distilled water.

2. Fe ₃ O ₄ and intermediate functionalized Fe ₃ O ₄ change in shape

Transmission electrons to investigate the surface and microstructure of as-synthesized Fe ₃ O ₄ nanoparticles, Fe ₃ O ₄ amino-functionalized with APTES and PEI, and Fe ₃ O ₄ functionalized with carboxyl groups with PAA. Microscopic (TEM) analysis was performed ( FIG. 1 ). The synthesized Fe ₃ O ₄ nanoparticles were observed to have a size of about 300 nm. The size and shape of the APTES modified particles were almost identical to the synthesized particles. However, the PEI-functionalized Fe ₃ O ₄ particles were found to have a hemispherical shape with a size of approximately 300 nm. A coating layer with a thickness of 3 to 4 nm was formed on the surface of the PEI-treated Fe ₃ O ₄ particles. In contrast, Fe ₃ O ₄ nanoparticles treated with PAA for functionalization with carboxyl groups had a smooth and opaque layer with a thickness of 5-6 nm formed on the surface. The core size of the PAA-treated Fe ₃ O ₄ was about 320 nm, which was slightly larger than the synthesized Fe ₃ O ₄ , and had a nearly perfect spherical shape compared to the PEI-treated Fe ₃ O ₄ particles.

3. Fe ₃ O ₄ and intermediate functionalized Fe ₃ O ₄ functional group of

FT-IR spectra of as-synthesized Fe ₃ O ₄ , amino-functionalized Fe ₃ O ₄ using APTES and PEI, and carboxyl-functionalized Fe ₃ O ₄ using PAA are shown in FIG. 2 . For each particle, the Fe-O peak was observed at about 600 cm ^-1 . After surface modification, a prominent amine peak was observed at 1,500 ~ 1,700 cm ^-1 due to bending of the NH bending bond. Since the condensation of SiO ₂ is accompanied by the functionalization of APTES, the OH stretching peak and the Si-O-Si peak were observed at about 3,000 and 950-1,250 cm ^-1 , respectively. In the case of another amino functionalization using PEI, a CH bending peak was observed at 1,300 cm ^-1 as a polymer carbon chain was attached to the particle surface. In addition, CN bending and NH stretching bonds were observed at 1,000 ~ 1,200 and 3,300 ~ 3,500 cm ^-1 , respectively, in the spectra of the particles in which PEI was used, and these ranges were, in the spectrum of the particles in which APTES was used, OH stretching bonding and Si- It was a range in which a strong peak due to O-Si bonding appeared. Although some differences were observed, it was confirmed that both surface-treated particles were successfully functionalized with amine groups. In contrast, in the case of the PAA-treated particles, CH bending bonds derived from polymer carbon chains were observed at 1,300 ~ 1,400 cm ^-1 , which was similar to that of the PEI-treated particles. Significant carboxyl group bonds were found at 1,600-1,800 and 3,000 cm ^-1 for C=O and OH extension bonds, respectively. OH extension bonding was observed at 2,700-3,300 cm ^-1 , which was slightly lower than the range of APTES-treated particles (3,000-3,700 cm ^-1 ). Considering the difference in OH elongation bonding, it was confirmed that a carboxyl group was successfully attached to the Fe ₃ O ₄ surface, and most of the OH bonding was derived from a carboxylic acid rather than a hydroxyl group.

4. Fe ₃ O ₄ Discussion of the intermediate functionalization process of

From the above analysis results, it was observed that there was a difference in the amino-functionalized particles using APTES and PEI. The functionalization mechanism of APTES is based on hydrolysis and condensation similar to the Stober method. During APTES treatment, the existing SiO ₂ layer was etched with ammonia solution, and unreacted hydroxyl groups were exposed through hydrolysis. These hydroxyl groups were used as condensation sites with APTES, and this functional group was observed at 3,000 to 3,700 cm ^-1 in the FT-IR spectrum of Fe ₃ O ₄ treated with APTES. However, as evident in Fe ₃ O ₄ functionalized with PEI and PAA, no interlayer was involved in the functionalization process. Therefore, these precursors were directly attached to the surface of Fe ₃ O ₄ nanoparticles. However, due to the molecular structure of APTES, Si-O-Si bonds could not be formed continuously. Therefore, no coating layer was observed on the surface of the APTES-treated particles. For the PEI and PAA treated particles, the directly bound carbon chains produced an opaque layer on the Fe ₃ O ₄ nanoparticle surface.

5. Fe ₃ O ₄ Overview of NTA Conjugation and Intermediate Functionalization Processes

3 shows the mechanism of NTA attachment to the surface of Fe ₃ O ₄ nanoparticles and the chemical structure of Fe ₃ O ₄ functionalized in various ways. Since the NTA attachment mechanism used a condensation reaction including electrostatic attraction, in the case of amines, Fe ₃ O ₄ particles were attached with glutaraldehyde before conjugation with NTA. As can be seen from the FT-IR spectrum, the middle SiO ₂ layer or carbon layer showed the greatest difference between APTES-treated Fe ₃ O ₄ and PEI-treated Fe ₃ O ₄ . Such differences may lead to different conjugation properties with glutaraldehyde due to different concentrations of activation sites, electrostatic forces and chemical affinity with glutaraldehyde. Glutaraldehyde can form another peptide bond with the amine group of NTA to bind with various metal ions such as Ni and W. However, the PAA-treated Fe ₃ O ₄ particles contain a carboxylic acid capable of forming a peptide bond with an amine group. Therefore, PAA-treated Fe ₃ O ₄ can be directly conjugated with NTA and does not require glutaraldehyde as a cross-linker.

6. NTA-conjugated Fe ₃ O ₄ change in shape

FT-IR spectra of NTA-attached Fe ₃ O ₄ particles obtained through various types of intermediate functionalization are shown in FIG. 4 . In contrast to the APTES-attached particles, an accentuated peak was observed in the spectrum of the NTA-attached particles. The unique peaks observed in the FT-IR spectrum of Fe ₃ O ₄ treated with APTES shown in FIG. 2 were also observed at 600 cm ^-1 (Fe-O) and 1,300 cm ^-1 (CH). The C = O peak, which is a representative bond of the carboxyl group, coexisted with the NH peak at 1,500 ~ 1,700 cm ^-1 . This unique peak of NTA was also observed in the FT-IR spectrum of particles with NTA attached to PEI. This peak contrasts with the spectrum of PEI-attached Fe ₃ O ₄ nanoparticles. In general, it was found that most of the difference in the FTIR spectrum of each particle to which NTA was attached was almost the same as the difference in the spectrum of the amino-functionalized particle. In contrast, NTA-attached Fe ₃ O ₄ nanoparticles obtained through carboxylation with PAA were 1,300 cm ^-1 (CH), 1,395 to 1,440 cm ^-1 (OH) similar to APTES and PEI-treated particles. , showed a prominent NTA peak at 1,500 ~ 1,700 cm ^-1 (N-H and C=O coexistence), but its intensity was lower than that of APTES and PEI-treated particles. In addition, the OH extension bond shifted from 2,700 to 3,300 cm ^-1 (PAA-treated particles) to 3,000 to 3,700 cm ^-1 . This difference between the amino or carboxyl functionalized particles and the NTA-binding particles suggests that NTA was successfully attached to the surface of the Fe ₃ O ₄ particles.

7. NTA-conjugated Fe ₃ O ₄ functional group of

In the TEM image of NTA attached to the Fe ₃ O ₄ particles treated with APTES in FIG. 5 , the core-shell structure was observed with a spherical Fe ₃ O ₄ core and a fluffy shell. . The size of the core was about 300 nm, and the surface of the coating layer was observed to be wavy. However, each of the particles was not individually coated, but covered with a coating layer in the agglomerate state. The thickness of the coating layer was measured to be 15 to 30 nm, which was found to be thicker than the APTES-treated particles. In contrast, in the images of PEI-treated Fe ₃ O ₄ particles, round particles with a size of 300 nm were observed along with other small particles (size 150-250 nm).

These particles were agglomerated and connected to a thin coating layer (8-15 nm). In the case of the PAA-treated particles, the size of the core Fe ₃ O ₄ particles was observed to be about 300 nm, similar to the APTES and PEI-treated particles. However, unlike other samples, the PAA treated particles remained separated. The coated layer covered the Fe ₃ O ₄ nanoparticles individually and was observed to be about 10 nm thick.

8. NTA-conjugated Fe with various kinds of intermediate functional groups ₃ O ₄ surface charge of

In order to analyze the repulsive force on the particles, the surface charge at each functionalization step of the Fe ₃ O ₄ sample is shown in FIG. 6 . As-synthesized Fe ₃ O ₄ had a slight negative charge insufficient to generate sufficient repulsive force for stable dispersion. However, after APTES treatment, the surface charge increased to 35.4 eV due to the positive (+) amine group. However, after NTA conjugation, it was changed to a negative charge of -23.1 eV. A similar trend was observed for the PEI-treated particles whose surface charge was changed from 33.1 eV to -28.2 eV. However, after PAA treatment, a strong negative charge (-58.7 eV) was formed on the particle surface and changed to -46.2 eV when NTA was attached.

9. NTA-conjugated Fe with various kinds of intermediate functional groups ₃ O ₄ magnetic properties of

A hysteresis curve of the sample to which Fe ₃ O ₄ and NTA synthesized through APTES, PEI, and PAA treatment is attached is shown in FIG. 7 . No coercive force or residual magnetization was found in any of the hysteresis curves, and each particle exhibited a paramagnetic curve. However, the saturation magnetization of Fe ₃ O ₄ as it was synthesized was the highest at 120 emu/g. This was followed by the APTES and PEI treated samples, which exhibited values of 106 and 98 emu/g, respectively, which were 88.3% and 81.7% of the saturation magnetization maximum, respectively. The PAA-treated particles showed the lowest value at 94 emu/g, which was 78.3% of the as-synthesized Fe ₃ O ₄ .

In short, it was observed that NTA was properly attached with a negative (-) surface charge through the NTA attachment process. However, an inversion of the surface charge occurred in the samples amino-functionalized by APTES and PEI. In this reversal process, the electrostatic force was weakened due to the pH reversal and the change of functional groups, and it was difficult to maintain dispersion stability. Accordingly, aggregation that can be observed in the TEM image shown in FIG. 5 occurred. However, although the surface charge of the particles treated with PAA was slightly reduced during the NTA bonding process, they were sufficiently strong in terms of maintaining dispersibility.

On the other hand, the magnetization of NTA-attached specimens is directly related to the reversible separation and redistribution considering the high saturation magnetization and low residual magnetization of these specimens. The magnetic hysteresis curves of each specimen came from the Fe ₃ O ₄ core particles and show the differences due to the phase change caused by the coating layer or the external environment. In the process of NTA conjugation through amino or carboxyl functionalization, Fe ₃ O ₄ nanoparticles may have been damaged through basic or acidic reaction conditions, but their paramagnetic properties were maintained after NTA attachment and sufficiently high saturation for magnetic separation showed magnetism.

The present invention is not limited to the above embodiments, but can be manufactured in various different forms, and those of ordinary skill in the art to which the present invention pertains can take other specific forms without changing the technical spirit or essential features of the present invention. It will be understood that it can be implemented as Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive.

Claims (5)

(a) introducing a functional group capable of forming a peptide bond with nitrilotriacetic acid (NTA) by modifying the magnetite (Fe ₃ O ₄ ) particle surface; and
(b) reacting the magnetite particles having a functional group introduced thereon with nitrilotriacetic acid to bond nitrilotriacetic acid to the surface of the magnetite particles;
A method for producing magnetic nanoparticles conjugated with a chelate ligand comprising a. The method of claim 1,
In step (a),
A chelate characterized in that the surface of the magnetite (Fe ₃ O ₄ ) particle is modified with (3-aminopropyl) triethoxysilane (APTES) to introduce an amino group and react with glutaraldehyde to form an aldehyde group at the end. A method for manufacturing a magnetic nanoparticle conjugated with a ligand. The method of claim 1,
In step (a),
Magnetite (Fe ₃ O ₄ ) The surface of the particle is modified with polyethyleneimine (PEI) to introduce an amino group and react with glutaraldehyde to form an aldehyde group at the end of the magnetic nanoparticles conjugated with a chelate ligand. manufacturing method. In step (a),
Magnetite (Fe ₃ O ₄ ) A method for producing magnetic nanoparticles conjugated with a chelate ligand, characterized in that the surface of the particle is modified with polyacrylic acid (PAA) to introduce a carboxyl group. A magnetic nanoparticle conjugated with a chelating ligand prepared according to any one of claims 1 to 4.

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