WO2010076946A1 - Nanoparticulates, complex nanoparticulates, and manufacturing method thereof - Google Patents

Abstract

The present invention relates to nanoparticulates, complex nanoparticulates, and a manufacturing method thereof. The nanoparticulates of the present invention which are gadolinium oxide nanoparticulates coated with the biocompatibility ligand can be used as a contrast media for diagnosing diseases such as brain illness, because of the capability to easily pass through the blood-brain barrier. Also, as the manufacturing method of the present invention is simpler than other manufacturing methods for contrast media, the manufacturing method of the present invention may reduce production costs and be utilized in the medical diagnosis field. Also, the contrast media including the complex nanoparticulates of the present invention contains complex particulates manufactured by coating gadolinium oxide particulates with manganese oxide, wherein the gadolinium oxide and the manganese oxide are conventionally used as main components of the contrast media. The complex nanoparticulates manufactured by combining the complex particulates and the biocompatibility ligand can maximize contrast effect and reduce ecotoxicity.

Description

Nanoparticles themselves, complex nanoparticles themselves and their preparation

The present invention relates to a nano-particle itself, a composite nano-particle itself, and a method for producing the same.

Magnetic Resonance Imaging (MRI) is a method of acquiring anatomical, physiological, and biochemical information images of a body using spin relaxation of hydrogen atoms in a magnetic field. It is a noninvasive method of obtaining human or animal body organs It is an excellent imaging device that can be imaged in real time.

In life sciences and medicine, we use a method to increase the image contrast by injecting materials from outside in order to make various and precise use of MRI. Such materials are called contrast agents. The contrast between tissues on the MRI image is due to the different tissues' different relaxation of the water molecule nuclear spin in the tissue to equilibrium, which affects the relaxation It causes the difference in the degree of relaxation between the tissues and induces a change in the MRI signal, thereby making the contrast between the tissues clearer.

Contrast agents differ in their characteristics and functions depending on the constituents of the constituent materials, and there is a difference in utilization and precision depending on the target to be injected. Enhanced contrast using contrast agents is achieved by increasing or decreasing the perimeter and imaging signal of a particular organ or tissue to produce more vivid imaging. Generally, the contrast agent that makes the image signal of the body part which wants to obtain the MRI image relatively bright is called a positive contrast agent, and the contrast agent which makes it relatively darker than the surrounding is called a negative contrast agent.

Examples of the paramagnetic nanoparticles used for the contrast agent include manganese (Mn) and gadolinium (Gd). These paramagnetic nanoparticles have problems such as insolubility or toxicity, and in order to solve this problem, There is an attempt to coat.

There is a kind of barrier in the brain nerve tissue that prevents certain substances from entering the blood vessels. These functional barriers are called the blood-brain barrier (BBB). For example, when trypan blue is injected intravenously and observed, this dye appears in all other intercellular spaces, but the central nervous system does not appear. This is because many kinds of drugs and metabolites are blocked by the blood-brain barrier and can not penetrate the brain nerve tissue. Because of these features of the brain, it is difficult to identify the disease present in the brain as a general contrast agent, and an MRI contrast agent that can pass through the cerebral blood vessel barrier has been actively studied as a contrast agent for diagnosing brain diseases.

Conventionally, U.S. Patent No. 6,638,494 relates to a superpowder nanoparticle containing a metal such as iron oxide, and attaches a specific carboxylic acid to the surface of the nanoparticle to prevent aggregation and precipitation of the nanoparticle due to gravity or magnetic field Is disclosed. The specific carboxylic acid is an aliphatic dicarboxylic acid such as maleic acid, tartaric acid or glutaric acid or an aliphatic polydicarboxylic acid such as citric acid, cyclohexane or tricarboxylic acid .

However, the above-mentioned contrast agent is a method for attaching a carboxylic acid to the surface of a metal particle to prevent agglomeration of the nanoparticle. Although the method of bonding the carboxylic acid to the nanoparticle is simple, It is not verified that the nanoparticles to which the carboxylic acid is bound are bio-toxic, and it can not be used as a contrast agent for diagnosing brain diseases.

Therefore, in the art, a contrast agent having low bio-toxicity is required, while the size of the contrast medium is small so that it is not collected in the liver and exists at a high concentration in the blood and can sufficiently pass through the broken cerebral blood vessel barrier.

It is an object of the present invention to solve the problems of the prior art described above, and it is an object of the present invention to provide a nano-particle itself coated with a biocompatible ligand so as not to cause coagulation and sedimentation of the paramagnetic nano- To provide a simple manufacturing method that can be used.

In order to achieve the above object, the present invention provides a nano-particle itself coated with a biocompatible ligand on gadolinium oxide nanoparticles.

The biocompatible ligand may have an antibody or protein bound thereto.

The biocompatible ligand may be polyethylene glycol, lactobionic acid or D-glucuronic acid.

The diameter of the gadolinium oxide nanoparticles is preferably 0.5 to 9 nm, more preferably 0.5 to 2.5 nm.

The diameter of the biocompatible ligand-coated gadolinium oxide nanoparticle itself is preferably 0.5 to 10 nm, more preferably 0.5 to 3 nm.

According to another aspect of the present invention, there is provided a method for preparing a nanoparticle comprising: 1) adding a metal precursor and a biocompatible ligand to a polar organic solvent to obtain a mixture; 2) stirring the mixture of step 1) at a high temperature while supplying air to obtain a reactant; And 3) adding an organic solvent to the reactant to obtain a final product;

.

The polar organic solvent in the step 1) may be triethylene glycol, tripropylene glycol, or the like.

The metal precursor of the step 1) may be GdCl 3 .6H 2 O or Gd (NO 3) 3.

The diameter of the metal precursor is preferably 0.5 to 9 nm, more preferably 0.5 to 2.5 nm.

The biocompatible ligand of step 1) may be bound to an antibody or protein.

The biocompatible ligand of step 1) may be polyethylene glycol, lactobionic acid or D-glucuronic acid.

The metal precursor and the biocompatible ligand of step 1) may be added in a molar ratio of 1: 0.5 to 0.5: 1.

The air in the step 2) preferably contains 5 to 50% or more of oxygen.

The high temperature of step 2) is preferably 200 to 300 ° C.

In the step 2), stirring is preferably performed for 20 to 30 hours.

The organic solvent in step 3) may be acetone, methyl ethyl ketone or the like.

The diameter of the nanoparticle itself coated with the biocompatible ligand is preferably 0.5 to 10 nm, more preferably 0.5 to 3 nm.

Also provided is an MRI contrast agent comprising a nano-particle itself coated with a biocompatible ligand on gadolinium oxide nanoparticles.

The present invention also provides a composite nano-particle itself in which a biocompatible ligand is bound to a composite particle obtained by coating gadolinium oxide particles with manganese oxide.

The biocompatible ligand may be D-glucuronic acid, polyethylene glycol or lactobionic acid.

The diameter of the composite nano-particles itself is preferably 0.5 to 3 nm.

According to another aspect of the present invention, there is provided a method of preparing a composite nano-particle itself, comprising the steps of: 1) preparing gadolinium oxide particles using a gadolinium precursor; 2) coating the gadolinium oxide particles with a manganese precursor to prepare composite particles; And 3) binding the biocompatible ligand to the composite particle; .

The method for producing the gadolinium oxide particles of the step 1) is characterized in that the gadolinium precursor is mixed with distilled water and reacted at 200 to 300 ° C.

The method for preparing the composite particles of the step 2) comprises: adding a manganese precursor to a solution containing the gadolinium oxide particles of the step 1) and reacting the mixture at a temperature of 200 to 300 ° C to form a gadolinium oxide particle It is characterized by being coated.

The step 3) is characterized in that a biocompatible ligand such as D-glucuronic acid, polyethylene glycol or lactobionic acid is added to the solution containing the composite particles of step 2), and the reaction is carried out at 130 to 170 ° C.

The diameter of the composite nano-particles itself is preferably 0.5 to 3 nm.

According to another aspect of the present invention, there is provided a composite nano-particle itself, wherein a biocompatible ligand is bound to a composite particle wherein gadolinium oxide particles are coated with manganese oxide.

The diameter of the composite nano-particles itself is preferably 0.5 to 3 nm.

The nanoparticles coated with the biocompatible ligand on the oxidized gadolinium nanoparticles according to the present invention are coated with a biocompatible ligand in order to prepare gadolinium oxide in nanoscale and to neutralize the toxicity of gadolinium oxide, , Can be transmitted to the brain through the blood-brain barrier, and thus can be used as a contrast agent for diagnosing brain diseases.

In addition, the method for preparing nanoparticles according to the present invention is a method for adding a biocompatible ligand to a gadolinium precursor solution and then reacting the biocompatible ligand to bind the biocompatible ligand to the gadolinium oxide nanoparticle. And it is easy to manufacture the nano-particles themselves by a simple method.

In addition, the contrast agent containing the composite nano-particles according to the present invention can be produced by preparing composite particles in which gadolinium oxide particles are coated with manganese oxide using gadolinium oxide and manganese oxide, which are conventionally used as main components of the contrast agent, By combining the biocompatible ligand with the complex nano-particle itself, the biomedical toxicity can be reduced while maximizing the contrast effect.

Therefore, the nano-particles themselves and the composite nano-particles themselves have superior imaging effects and can be used in medical diagnosis fields.

1 is an electron micrograph of a nano-particle itself coated with a biocompatible ligand, polyethylene glycol, on gadolinium oxide nanoparticles prepared according to an embodiment of the present invention.

FIG. 2 is an electron micrograph of a nano-particle itself coated with lactobionic acid, which is a biocompatible ligand, on gadolinium oxide nanoparticles prepared according to an embodiment of the present invention.

FIG. 3 is an electron micrograph of a nano-particle itself coated with a biocompatible ligand, D-glucuronic acid, on the gadolinium oxide nanoparticles prepared according to an embodiment of the present invention.

4 is an electron micrograph of a composite nano-particle itself prepared according to an embodiment of the present invention.

FIG. 5 is a graph showing characteristics of nano-particles manufactured according to the present invention in accordance with changes in absolute temperature and magnetic field.

FIG. 6 shows the FT-IR spectrum of the nano-particles themselves prepared according to the present invention.

FIG. 7 is a view showing the binding state of a biocompatible ligand to a composite nano-particle itself prepared according to an embodiment of the present invention.

FIG. 8 shows a map image according to the concentration of the nano-particles themselves prepared according to the present invention.

Figure 9 relates to the relaxation according to the concentration of the nano-particles themselves prepared according to the present invention.

10 is a measurement result of the relaxation property of the composite nano-particles produced according to an embodiment of the present invention.

11 is a brain MRI photograph of a mouse for experimental administration of a contrast agent containing nano-particles prepared according to the present invention.

FIG. 12 is a MRI photograph of a contrast agent prepared in accordance with an embodiment of the present invention and administered to an animal.

The present applicant has been studying a contrast agent for the diagnosis of brain diseases using gadolinium oxide. The inventors of the present invention have found that when a biocompatible polymer such as polyethylene glycol, lactobionic acid or D-glucuronic acid is coated with gadolinium oxide, Coated with a biocompatible polymer, and that the composite nano-particles coated with the biocompatible polymer can pass through the cerebral blood vessel barrier, and that the contrast effect is excellent. Thus, the present invention has been completed.

The present invention provides a nanoparticle itself coated with a biocompatible ligand on gadolinium oxide nanoparticles.

The gadolinium oxide nanoparticle is a portion constituting the core of the nano-particles according to the present invention, and the diameter of the gadolinium oxide nanoparticles is preferably 0.5 to 9 nm. When the diameter of the nanoparticles of the present invention exceeds 9 nm, the size of the nanoparticle according to the present invention is not easily controlled, and the nanoparticle itself may not easily pass through the blood vessel barrier.

The biocompatible ligand according to the present invention may be bound to an antibody or protein. Although the antibody is not particularly limited, it is also possible to use a mutated variable region or constant region in order to increase the affinity for a specific region in a living body. In addition, although the protein is not particularly limited, a therapeutic protein or a protein hormone can be used. The protein may be mutated to increase the affinity for a specific site in a living body.

The biocompatible ligand is not particularly limited, but it is not limited as long as it is a biocompatible polymer. Preferably, polyethylene glycol, lactobionic acid, D-glucuronic acid and the like can be used alone or in combination.

The diameter of the nano-particles having the biocompatible ligand coated on the gadolinium oxide nanoparticles is preferably 0.5 to 10 nm. If the gadolinium oxide nano-particle itself is more than 10 nm, it is difficult to pass through the cerebral blood vessel and it is difficult to use it as a contrast agent for diagnosis of brain diseases.

Further, in the method for producing a nano-particle itself of the present invention,

1) adding a metal precursor and a biocompatible ligand to a polar organic solvent to obtain a mixture;

2) stirring the mixture of step 1) at a high temperature while supplying air to obtain a reactant; And

3) adding an organic solvent to the reaction product to obtain a final product;

.

First, the step 1) is a step of adding a metal precursor and a biocompatible ligand to a polar organic solvent to obtain a mixture.

The polar organic solvent in the step 1) is not particularly limited and is not limited as long as it is a polar organic solvent capable of inducing the reaction of the metal precursor and the biocompatible ligand. Preferably, triethylene glycol, tripropylene glycol, have.

The metal precursor is not particularly limited, and preferably a gadolinium precursor can be used. More preferably, GdCl 3 .6H 2 O and Gd (NO 3) 3 can be used. The diameter of the metal precursor is preferably 0.5 to 9 nm. When the diameter of the nanoparticles used in the present invention exceeds 9 nm, it is difficult to control the size of the nanoparticles according to the present invention, and the nanoparticles themselves may not easily pass through the blood vessel barrier have.

The biocompatible ligand according to the present invention may be conjugated with an antibody or protein. Although the antibody is not particularly limited, it is also possible to use a mutated variable region or constant region in order to increase the affinity for a specific region in a living body. In addition, although the protein is not particularly limited, a therapeutic protein or a protein hormone can be used. The protein may be mutated to increase the affinity for a specific site in a living body.

The biocompatible ligand is not particularly limited, and is not limited as long as it is a biocompatible polymer. Preferably, polyethylene glycol, lactobionic acid, D-glucuronic acid and the like can be used singly or in combination.

The metal precursor and the biocompatible ligand may be mixed at a molar ratio of 1: 0.5: 0.5 to 1 to coat the metal precursor with the biocompatible ligand.

Next, the step 2) is a step of obtaining a reactant by stirring at a high temperature while supplying air to the mixture of the step 1).

The air is not particularly limited, and air in the air may be used. Preferably 5 to 50% of oxygen may be used. In the reaction, oxygen contained in the air reacts with the gadolinium metal precursor to produce gadolinium oxide. The method of supplying the air is not particularly limited, but it is preferable to bubble the reaction solution to supply air.

In addition, the step of supplying air may be carried out simultaneously with the preparation of the mixture, but may be performed after the mixture has an appropriate reaction time. It is preferable that the high temperature is 200-300 占 폚 if the metal precursor has a temperature capable of coating the biocompatible ligand. The reaction may be carried out by stirring for 20 to 30 hours to facilitate the reaction. When the temperature is less than 200 ° C, it is difficult to induce a reaction to coat the biocompatible ligand with the metal precursor. When the temperature exceeds 300 ° C, the reaction rate does not increase with increasing temperature. The stirring is not limited as long as the reactant can be easily produced. The reaction product produced in this step may be a state where the biocompatible ligand is bound or coated on the surface of the gadolinium oxide nanoparticles and the biocompatible ligand is somewhat unstable due to the high temperature. Therefore, there is a need for additional steps as follows to obtain it.

Finally, step 3) is a step of adding an organic solvent to the reactant to obtain a final product.

The organic solvent may be precipitated using ketones such as acetone and methyl ethyl ketone, while the stability of the reactant in step 2) is enhanced, and the organic solvent is not limited as long as it is an organic solvent capable of precipitating. The temperature of the added organic solvent is not particularly limited, and an organic solvent at room temperature may be added to lower the temperature of the reactant, or may be added after the reactant is cooled. The reaction product may be cooled to obtain an appropriate final product. The method of cooling the reaction product is not limited as long as it is a method for obtaining an appropriate final product. However, the reaction product can be slowly cooled at room temperature or quenched. The final product produced in this step is a precipitate of the nano-particles themselves, to which the biocompatible ligand, which is a reactant in step 2), is added. When the precipitate is added to a solution suitable for use as distilled water or other contrast agent, It is a form that can be easily used as a contrast agent.

The preparation of the nanoparticle itself coated with the biocompatible ligand on the gadolinium oxide nanoparticle is described in detail. A gadolinium precursor and an organic solvent such as polyethylene glycol, lactobionic acid or D-glucurononic acid are added to an organic solvent such as triethylene glycol. And the mixture was reacted by stirring at 260 캜 for 24 hours while bubbling air in the air.

The reaction product after the above synthesis step may include a nano-particle itself coated with a biocompatible ligand on the gadolinium oxide nanoparticles, and a polar organic solvent and an unreacted material depending on the amount of the reactant used. From this reaction product, a separation step is carried out to separate and obtain the nanoparticle itself coated with the biocompatible ligand on the oxidized gadolinium nanoparticle.

The separation step may be performed by using a membrane filtration method or the like to separate and obtain the nano-particles themselves coated with the biocompatible ligand from the reaction product, preferably the precipitation separation method. Specifically, a precipitant is added to the reaction product to precipitate the nanoparticle itself coated with the biocompatible ligand on the gadolinium oxide nanoparticles. Then, the supernatant is removed from the reactor to separate and obtain a precipitate (nano-particle itself coated with biocompatible ligand on gadolinium oxide nanoparticles). As the precipitant, ketones such as acetone or methyl ethyl ketone can be used. At this time, the biocompatible ligand-coated gadolinium oxide precipitate can be dried to produce a powdery product. In addition, the dried powder can be made into a liquid product by dissolving in distilled water or the like, and the dried powder has excellent dispersibility in distilled water.

According to the manufacturing method of the present invention described above, the nanoparticle itself coated with the biocompatible ligand on the gadolinium oxide nanoparticles by a single synthesis step is simply produced. That is, as noted above, the synthesis step involving the reaction oxidizes the metal precursor to the metal oxide of the nanoscale particles, while at the same time coating the biocompatible ligand on the surface of the metal oxide. In addition, the final product according to the present invention is advantageous in that it is manufactured in an ultrafine size, has quick absorption of human body, and has a low manufacturing cost. In addition, since the synthesis is carried out in a single step in the reactor, mass production can be achieved by designing the reactor scale to be large.

Meanwhile, the gadolinium oxide nanoparticle according to the present invention has a core-shell structure in which a biocompatible ligand is coated on the surface of a metal oxide.

The diameter of the gadolinium oxide nano-particles themselves according to the present invention is preferably 0.5 to 10 nm.

The gadolinium oxide nanoparticles according to the present invention are not particularly limited but can be used for various purposes such as MRI contrast agent and drug delivery system. Since the metal oxide constituting the core is gadolinium oxide, it is useful as a T1 contrast agent in MRI Can be used.

Gadolinium-based contrast agents do not pass through the cerebral blood vessels and some are toxic and are not suitable as contrast agents for brain diseases. However, the contrast agent of the present invention is a biocompatible ligand used to reduce toxicity, such as polyethylene glycol, coated with nano-sized gadolinium oxide, resulting in a slow metabolism of the liver, through biocompatible ligand coating Resulting in reduced toxicity of the contrast agent. Particularly, gadolinium is the most suitable element to be used as a T1 contrast agent, but the use of the human body due to toxicity was limited. However, the present invention overcomes these shortcomings of gadolinium and completed the contrast agent for brain diseases.

An auxiliary component may be added to enhance the stability of the active ingredient of the MRI contrast agent containing the gadolinium oxide nanoparticle itself, which is produced by the production method according to the present invention, as an active ingredient. Examples of suitable auxiliary components include sodium citrate The stability of the gadolinium oxide nano-particles themselves according to the present invention can be enhanced.

The present invention also provides a composite nano-particle itself in which a biocompatible ligand is bound to a composite particle obtained by coating gadolinium oxide particles with manganese oxide.

The gadolinium oxide particles are nano-sized particles formed by using a gadolinium precursor such as GdCl 3 .6H 2 O or Gd (NO 3) 3. The gadolinium oxide is a paramagnetic substance having a weak magnetic property. When used as a contrast agent, the gadolinium oxide has an advantage that the lesion is clearly visible and the boundaries of the mass are clarified in contrast to low signals of muscles, bone, and blood vessels. However, the above-mentioned gadolinium oxide has a disadvantage of high bio-toxicity, and in order to solve the disadvantage of the gadolinium oxide, the nanoparticulate gadolinium oxide particles are coated with manganese oxide using a manganese precursor to form nano- . The manganese oxide is also used as a contrast agent. The manganese oxide is stable enough to show no toxicity even when the manganese oxide is tested in an animal without coating the biocompatible ligand. When coated with manganese oxide, the imaging effect of gadolinium oxide particles is almost maintained and toxicity can be reduced.

Therefore, the composite particles coated with manganese oxide on the gadolinium oxide particles have a high contrast effect, and the stability of the particles can be enhanced.

When the composite particle is used in a living body, a complex nano-particle itself may be formed by binding a biocompatible ligand such as D-glucuronic acid, polyethylene glycol or lactobionic acid in order to enhance bio-toxicity and stability. The biocompatible ligand is a long-chain material, bound to the surface of the composite particle, and can be wrapped around the composite particle or formed into a branched shape. In this way, the composite particle coated with manganese oxide on the gadolinium oxide particle forms a core, and the biocompatible ligand surrounding the composite particle forms a complex nano-particle having a shell shape.

The diameter of the composite nano-particles itself is preferably 0.5 to 3 nm. If the diameter of the composite nano-particles itself is less than 0.5 nm, the particles tend to aggregate and the dispersion may not be achieved. As a result, the composite nano-particles themselves may not be uniformly dispersed into the brain tissue, If it exceeds 3 nm, the dispersibility of the liposomes is improved, but it is not suitable for use as a contrast agent for diagnosis of brain diseases to be used in the present invention.

Further, the method for producing the composite nano-particle itself according to the present invention comprises:

1) preparing gadolinium oxide particles using a gadolinium precursor;

2) coating the gadolinium oxide particles with a manganese precursor to prepare composite particles; And

3) binding the biocompatible ligand to the composite particle;

.

Hereinafter, each step of the present invention will be described in detail.

The method of preparing the gadolinium oxide particles of the step 1) may be performed by mixing the gadolinium precursor with distilled water and reacting the mixture at 200 to 300 ° C. It can be carried out by bubbling a gas containing oxygen into the solution so that the gadolinium precursor can be produced as gadolinium oxide particles. At this time, the temperature range is a temperature range allowing the gadolinium precursor to become gadolinium oxide particles.

Next, a method for preparing the composite particles of the step 2) comprises: adding a manganese precursor to a solution containing the gadolinium oxide particles of the step 1), and reacting the mixture at a temperature of 200 to 300 ° C to remove the gadolinium oxide particles from the manganese precursor May be coated with manganese oxide. The manganese precursor may be bubbled with oxygen-containing gas so as to be manganese oxide. At this time, the temperature range is a reaction temperature at which manganese oxide is coated on gadolinium oxide. Under such conditions, the composite particles can be produced by coating the gadolinium oxide particles with manganese oxide.

Finally, in the step 3), a biocompatible ligand is bound to the composite particle by adding a biocompatible ligand such as D-glucuronic acid, polyethylene glycol or lactobionic acid to the solution containing the complex particle, RTI ID = 0.0 > C, < / RTI > In this case, the temperature range is a reaction temperature allowing the biocompatible ligand to be easily bonded to the composite particle. Under the above conditions, hydrogen is removed from the carboxylic acid of the biocompatible ligand, and the remaining oxygen and the manganese oxide of the composite particles are bonded. This can be confirmed by 1600 C = O oscillations of FT-IR (Fourier transform spectroscopy), which was not found in the ligand alone after the binding of manganese and ligand.

Through the reaction of step 3), the solution contains gadolinium oxide particles, manganese oxide particles, composite particles coated with gadolinium oxide and manganese oxide, and composite nano particles in which the biocompatible ligand is combined with the composite particles. Here, in the method of separating the complex nano-particles themselves according to the present invention, when the distilled water is added to the mixed solution, only the ligand-bound complex nano-particles themselves are precipitated, and in addition, the substance becomes dissolved in water. The precipitated material may be separated to separate the complex nano-particles themselves.

The separated complex nano-particles may be washed once or more with a solution that does not affect the properties of the distilled water or other materials of the complex nano-particles of the present invention. The composite nano-particles obtained through the above steps may have a diameter of 0.5 to 3 nm.

The present invention also provides a contrast agent comprising a complex nano-particle itself in which a biocompatible ligand is bound to a composite particle wherein gadolinium oxide particles are coated with manganese oxide.

The complex nano-particles themselves are paramagnetic nanoparticles of 3 nm or less and have water-soluble biocompatible ligands coated therein. 10, the complex nano-particle self-relieving rate (10 to 15 s-1 / M) of the present invention is more than twice as high as the relaxation rate (4 s-1 / M) Quot; < / RTI > value.

The higher the relaxation rate, the more relaxation of the hydrogen nucleus, which shows that the contrast effect of the contrast agent is increased.

The diameter of the composite nano-particles itself is preferably 0.5 to 3 nm.

An auxiliary component may be added to enhance the stability of the contrast agent comprising the complex nano-particle itself prepared by the method of the present invention as an active ingredient. As a preferable auxiliary component, sodium citrate or the like may be mixed The stability of the gadolinium oxide nanoparticle itself according to the invention can be enhanced.

Hereinafter, the present invention will be described in detail with reference to the preferred embodiments, which will be readily apparent to those skilled in the art to which the present invention pertains. The present invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

Example 1: Preparation of polyethylene glycol-coated gadolinium oxide nanoparticles

5 mmol of GdCl3 占 H2O, 5 占 퐉 ol of polyethylene glycol and 40 ml of triethylene glycol were mixed. The mixture was allowed to react at 260 占 폚 for 24 hours while stirring with magnetic air while bubbling with air in the atmosphere. After the mixture is cooled to room temperature, acetone is added to the reaction product to precipitate polyethylene glycol-coated gadolinium oxide nano-particles themselves. The supernatant was removed, and the precipitate was dried in air to obtain a powder.

Example 2: Preparation of lactobionic acid-coated gadolinium oxide nanoparticles

Lactobionic acid-coated gadolinium oxide nanoparticles were prepared in the same manner as in Example 1, except that lactobionic acid was used instead of polyethylene glycol.

Example 3: Preparation of gadolinium oxide nanoparticles self-coated with D-glucuronic acid

Gadolinium oxide nanoparticles coated with D-glucuronic acid were prepared in the same manner as in Example 1, except that D-glucuronic acid was used instead of polyethylene glycol.

Example 4: Composite nanoparticle itself prepared by bonding tripropylene glycol to a gadolinium oxide-manganese oxide composite particle

GdCl 3 .6H 2 O (5 mmol) was added to 40 ml of distilled water and reacted with stirring at 260 ° C for 24 hours while bubbling air to prepare gadolinium oxide particles. Again, MnCl 2 .4H 2 O (5 mmol) was added thereto and reacted at 260 ° C for 24 hours while bubbling air. Tripropylene glycol (5 mmol) was added thereto and reacted at 150 ° C for 24 hours. After completion of the reaction, the reaction mixture was cooled at room temperature, and water was added to precipitate the pore. The separated mouths were washed three times with distilled water to prepare composite nano-particles themselves.

Example 5: Composite nanoparticle itself prepared by bonding D-glucuronic acid to gadolinium oxide-manganese oxide composite particle

The complex nano-particles themselves were prepared in the same manner as in Example 4, except that D-glucuronic acid was used instead of tripropylene glycol.

Example 6: Composite nanoparticle itself prepared by binding lactobionic acid to a gadolinium oxide-manganese oxide composite particle

The complex nano-particles were prepared in the same manner as in Example 4 except that lactobionic acid was used instead of tripropylene glycol.

EXPERIMENTAL EXAMPLE 1 Measurement of Gadolinium Oxide Nanoparticle Size

In order to measure the nano-particles themselves and the composite nano-particles prepared in Examples 1 to 4 by an electron microscope, the nano-particles themselves and the composite nano-particles themselves were dispersed in methanol, and specimens were prepared. This was measured with a high resolution electron microscope (manufactured by JEOL, model: JEM 2100F). The results are shown in Figs. 1 to 4.

As shown in FIGS. 1 to 4, the biocompatible ligand-coated gadolinium oxide nanoparticles according to the present invention were homogeneous and showed a particle size of 0.5 to 3 nm. This indirectly indicates that the gadolinium oxide according to the present invention can be easily absorbed by the human body.

Experimental Example 2: Parametric measurement of gadolinium oxide nano-particle itself

Example 1 produced by the manufacturing method according to the present invention was put into a SQUID magnetometer (manufacturer: Quantum Design, model: MPMS 7) and measured. The results are shown in Figs. 5 (a) and 5 (b).

Fig. 5 (a) shows the specimen of Example 1 while keeping the magnetic field constant. It can be seen that the magnetic moment of the gadolinium oxide nanoparticles of Example 1 decreases as the temperature increases.

5 (b) shows the change of the specimen of the first embodiment according to the change of the magnetic field in a state where the temperature is kept constant. The gadolinium oxide of Example 1 had a moment value of 0 emu / g when the magnetic field was 0 Oe.

As a result, it was confirmed that the nanoparticles coated with the biocompatible ligand on the oxidized gadolinium nanoparticles according to the present invention were paramagnetic.

Experimental Example 3: Measurement of biopolymer coating of gadolinium oxide nano-particles themselves

Example 1 produced by the manufacturing method according to the present invention was put into an FT-IR infrared spectrophotometer (manufactured by Mattson Instruments Ins., Model: Galaxy 7020A) and measured. The results are shown in Figs. 6 (a) and 6 (b) and Fig.

FIG. 6 (a) is an FT-IR spectrum of polyethylene glycol, and FIG. 6 (b) is an FT-IR spectrum of polyethylene glycol-coated gadolinium oxide nanoparticles. First, the functional group peak of polyethylene glycol is observed in FIG. 6 (b), indicating that the gadolinium oxide nanoparticles are coated with polyethylene glycol. Also in Fig. 6 (b), it can be seen that the carbonyl group of polyethylene glycol is chemically bonded to the gadolinium oxide nanoparticles at a C = O peak at 1600 cm < -1 >.

Further, in FIG. 7, the functional group peak of tripropylene glycol, which is a biocompatible ligand, is observed in the complex nano-particle itself, and it can be seen that the composite particle is coated with a biocompatible ligand, tripropylene glycol.

Experimental Example 4: Determination of relaxation and paramagnetism of gadolinium oxide nano-particles by concentration

The gadolinium oxide nanoparticles prepared in Example 1 were suspended in distilled water at concentrations of 0.0625, 0.125, 0.25 and 0.5 mM to prepare samples. This was measured with an MRI apparatus (manufactured by Mattson Instruments Ins., Model: Galaxy 7020A), and the results are shown in Figs. 8 (a) and (b) and Fig.

As shown in FIGS. 8 (a) and 8 (b), the relaxation rate time becomes shorter as the concentration of gadolinium oxide nanoparticle itself increases. The gadolinium oxide nanoparticles according to the present invention exhibit concentration-dependent T1 and T2 map image. The T1 and T2 map images were measured and the reciprocal values were taken to obtain r1 and r2 values, and the results are shown in Fig.

As shown in FIG. 9, the slopes of the equations obtained by substituting the values of r1 and r2 are 11.6 and 13.4, respectively. If we calculate this as r2 / r1, we can see that the value is close to 1. Theoretically, the r2 slope value always appears to be higher than the r1 slope value, but it is similar, indicating that the contrast agent according to the present invention is paramagnetic. In addition, the slope value of r1, which is the relaxation rate of the gadolinium oxide nanoparticles according to the present invention, was 11.6. This indicates that the relaxation rate of the gadolinium oxide nano-particles of the present invention is higher when the relaxation rate of the gadolinium complex used in the prior art is 4, which is the average value of r1. This indicates that the gadolinium oxide nanoparticles according to the present invention can exhibit the same contrasting effect even when a low concentration of contrast agent is used. Therefore, it has been found that the gadolinium oxide nano-particles themselves according to the present invention can be used as an effective contrast agent even at a low concentration.

Experimental Example 5: Relaxation and parametric measurement of complex nano-particles according to concentration

The complex nano-particles prepared in Example 1 were suspended in distilled water at a concentration of 2, 1, 0.5, 0.25, 0.125 and 0.0625 mM to prepare samples. This was measured with an MRI apparatus (manufactured by Mattson Instruments Ins., Model: Galaxy 7020A), and the results are shown in Fig.

As shown in Fig. 10, the reciprocal values of the R1 and R2 values of the complex nano-particles themselves according to the present invention were taken and r1 and r2 values were obtained.

The reason for the difference between r1 and r2 is that the complex nano-particle itself has different relaxation rates in the z-axis and the x-y axis when the hydrogen nuclear spin relaxes. The r1 value is related to the x-y axis relaxation rate and the r2 value is related to the relaxation rate in the z-axis. The two relaxations can be independently measured, and the composite nano-particles themselves have a high T1 contrast, which indicates that the r1 value is higher than 10.

Experimental Example 6: In vivo contrast effect of contrast agent containing nano-particles according to the present invention

An anesthetic is maintained by inhalation of 350 g of a brain tumor-induced experimental rat (SD-rat, Hyochang Science) mixed with isoflurane (Choongwae Pharmaceutical Co., Ltd.) with N2O and O2 and continuously inhaled. 520 μl (0.007 mM) of the contrast agent according to Example 3 of the present invention was injected into anesthetized experimental rats through a rat tail vein, and the experimental animals were placed in an MRI apparatus (manufactured by GE, model: Excite) . The results are shown in Figs. 11 (a) and 11 (b).

Fig. 11 (a) is a brain image of a laboratory mouse before administration of the contrast agent according to the present invention, and Fig. 11 (b) is a brain image of administration of the contrast agent of Example 3. Fig. As shown in FIG. 11 (b), when the contrast agent according to the present invention is administered, tumors induced in a laboratory rat as shown in FIG. 11 (b) can be seen.

Therefore, the gadolinium oxide nanoparticles prepared according to the present invention can be usefully used as a contrast agent.

Experimental Example 7: In vivo contrast effect of contrast agent containing complex nano-particles according to the present invention

An anesthetic is maintained by inhalation of 350 g of a brain tumor-induced experimental rat (SD-rat, Hyochang Science) mixed with isoflurane (Choongwae Pharmaceutical Co., Ltd.) with N2O and O2 and continuously inhaled. 520 μl (0.007 mM) of the contrast agent according to Example 3 of the present invention was injected into anesthetized experimental rats through a rat tail vein, and the experimental animals were placed in an MRI apparatus (manufactured by GE, model: Excite) . The results are shown in Fig.

As shown in FIG. 12, when the contrast agent containing the complex nano-particles according to the present invention was administered, it was found that the contrast enhancement effect on the internal organs of the mice was shown. This suggests that the complex nano particles of the present invention are small in size and easy to access to all organs and have a high contrast effect. This indirectly indicates that the brain tumor can be used to diagnose a brain tumor more easily when the brain tumor is induced.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, is intended to cover various modifications and equivalent arrangements included within the spirit and scope of the appended claims. Of course.

Claims (28)

1. Nanoparticles themselves coated with biocompatible ligands on gadolinium oxide nanoparticles.
2. The method according to claim 1,

   Wherein the biocompatible ligand is bound to an antibody or a protein.
3. The method according to claim 1,

   Wherein the biocompatible ligand is at least one selected from the group consisting of polyethylene glycol, lactobionic acid, and D-glucuronic acid.
4. The method according to claim 1,

   Wherein the diameter of the gadolinium oxide nanoparticles is 0.5 to 9 nm.
5. The method according to claim 1,

   Wherein the biocompatible ligand-coated gadolinium oxide nanoparticle itself has a diameter of 0.5 to 10 nm.
6. 1) adding a metal precursor and a biocompatible ligand to a polar organic solvent to obtain a mixture;

   2) stirring the mixture of step 1) at a high temperature while supplying air to obtain a reactant; And

   3) adding an organic solvent to the reaction product to obtain a final product;

   Wherein the biocompatible ligand is coated on the gadolinium oxide nanoparticles.
7. The method according to claim 6,

   Wherein the polar organic solvent in step 1) is at least one selected from the group consisting of triethylene glycol and tripropylene glycol.
8. The method according to claim 6,

   Wherein the metal precursor of step 1) is at least one selected from the group consisting of GdCl3 6H2O and Gd (NO3) 3.
9. The method according to claim 6,

   Wherein the metal precursor has a diameter of 0.5 to 9 nm.
10. The method according to claim 6,

    Wherein the biocompatible ligand of step 1) is bound to an antibody or a protein.
11. The method according to claim 6,

    Wherein the biocompatible ligand of step 1) is at least one selected from the group consisting of polyethylene glycol, lactobionic acid, and D-glucuronic acid.
12. The method according to claim 6,

    Wherein the metal precursor and the biocompatible ligand of step 1) are added in a molar ratio of 1: 0.5 to 0.5: 1.
13. The method according to claim 6,

    Wherein the air of step 2) contains 5 to 50% or more of oxygen.
14. The method according to claim 6,

    Wherein the high temperature of step 2) is 200 to 300 占 폚.
15. The method according to claim 6,

    And stirring in the step 2) is performed for 20 to 30 hours.
16. The method according to claim 6,

    Wherein the organic solvent in step 3) is at least one selected from the group consisting of acetone and methyl ethyl ketone.
17. The method according to claim 6,

    Wherein the biocompatible ligand-coated nanoparticle has a diameter of 0.5 to 10 nm.
18. An MRI contrast agent comprising a nanoparticle itself coated with a biocompatible ligand of the gadolinium oxide nanoparticle of claim 1 or 6.
19. Composite nanoparticle itself in which a biocompatible ligand is bound to a composite particle coated with gadolinium oxide particles with manganese oxide.
20. 20. The method of claim 19,

    Wherein the biocompatible ligand is at least one selected from the group consisting of D-glucuronic acid, polyethylene glycol, and lactobionic acid.
21. 20. The method of claim 19,

    Wherein the complex nano-particle itself has a diameter of 0.5 to 3 nm.
22. 1) preparing gadolinium oxide particles using a gadolinium precursor;

    2) coating the gadolinium oxide particles with a manganese precursor to prepare composite particles; And

    3) binding the biocompatible ligand to the composite particle;

    Wherein the composite nano-particles are produced by a method comprising the steps of:
23. 23. The method of claim 22,

    Wherein the gadolinium precursor is mixed with distilled water and reacted at a temperature of 200 to 300 ° C.
24. 23. The method of claim 22,

    The method for preparing the composite particles of the step 2) comprises: adding a manganese precursor to a solution containing the gadolinium oxide particles of the step 1) and reacting the mixture at a temperature of 200 to 300 ° C to form a gadolinium oxide particle Wherein the coating layer is coated on the surface of the composite nano-particles.
25. 23. The method of claim 22,

    In the step 3), a biocompatible ligand, which is at least one selected from the group consisting of D-glucuronic acid, polyethylene glycol and lactobionic acid, is added to the solution containing the composite particles of the step 2) And reacting the resultant nanoparticles.
26. 23. The method of claim 22,

    Wherein the diameter of the composite nano-particle itself is 0.5 to 3 nm.
27. Contrast agent containing a complex nano-particle itself in which a biocompatible ligand is bound to a composite particle coated with gadolinium oxide particles with manganese oxide.
28. 28. The method of claim 27,

    Wherein the complex nano-particle itself has a diameter of 0.5 to 3 nm.

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