DE102014218355B4 - Process for the preparation of surface-hydrophilized metal oxide nanoparticles - Google Patents

Abstract

A process for producing surface-hydrophilized metal oxide nanoparticles comprising the following steps: a. Activation of hydrophobic metal oxide nanoparticles containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements, and a coordinatively bound ligand shell, in an organic solvent, by adding an organophilic base comprising a strong base and a phase transfer agent, b. Addition of an active C-alkane having a functionality and a leaving group, whereby a covalent attachment of the alkane to the metal oxide nanoparticles takes place, so that a precipitate precipitates, c. Separation of the precipitate and conversion into an aqueous solution, characterized in that the covalent bonding in (b) via a carbon atom of the C-alkyl group of the active C-alkane on the nanoparticles own, occupying the surface lattice sites of the metal oxide nanoparticles, oxygen occurs.

Description

The present invention relates to a process for the hydrophilization of metal oxide nanoparticles by a specific modification of the lattice sites and the use of these.

In order to ensure high water solubility, biocompatibility and simple surface functionalization of magnetite nanoparticles, a hydrophilic surface modification is necessary. A number of different approaches are known from the prior art, which can be classified in three main categories.

In the so-called ligand exchange, an exchange of hydrophobic surface ligands (for example alkylamines or long-chain carboxylic acids) takes place by means of hydrophilic ligands having a higher affinity for the particle surface. The stabilization of metal oxide nanoparticles by the complexation of near-surface metal ions via alcohol functionalities is known and is usually carried out via multidentate ligands, such as dopamine [ Niederberger et al .: Chem. Mater. 2004, 16, 1202-1208 ]. This can be disadvantageous for detachment of the hydrophilic ligands by exchange reactions, since the underlying interactions between hydrophilic ligands and the metal ions of the particle surface are subject to chemical equilibrium.

The noncovalent coating for hydrophilic surface modification is carried out by means of amphiphilic ligands, in particular by means of block copolymers, lipids or phospholipids to form hydrophilic micelles. In the non-covalent coating by addition of additional organic (bio) polymers, however, hydrophobic components on the particle surface are not replaced, but merely enveloped, which leads disadvantageously to an increase in the hydrodynamic radius of the particles.

Moreover, those skilled in the art are also aware of possibilities for hydrophilic surface modification by means of inorganic coating, which as a rule are based on a silicate coating of the particle surface. The particle to be modified serves as a germ cell. However, the silicate coating is not sufficiently stable, so that the polysilicic acids of a silicate coating are degraded at pH values> 7 to oligosilicates and meta- or orthosilicate.

Furthermore, the international patent application describes WO 2005/095278 A1 the synthesis of metal oxide NP from a solution of benzyl alcohol and metal alcoholates via an ether elimination process [ Pinna et al .: Colloids Surf. 2004, 250, 1-3, 211-213 ].

The modification of nanoparticle surfaces by means of functionalized polymers has also been described [ Garcia, K. et al., RSC Adv. 2013. 3, 22443-22454 ]. The hydrophilization is carried out by the treatment with modified polyacrylic acid.

The preparation of monodisperse hydrophilic metal oxide nanoparticles, particularly magnetite nanoparticles <10 nm, which have high stability against aggregation (i.e., colloidal stability) and allow simple surface modifications, is not possible on the basis of methods known from the literature. Consequently, there is a great interest and a great need for a simple process for the hydrophilic surface modification of such metal oxide nanoparticles.

The present invention is therefore based on the object to provide the most selective ligand exchange reaction for the stable hydrophilization of metal oxide nanoparticles under the direct inclusion of nanoparticle-own surface atoms.

1. a. Aktivierung von hydrophoben Metalloxid-Nanopartikeln, enthaltend mindestens ein Metalloxid eines Metalls ausgewählt aus der Eisen-Platin-Gruppe des Periodensystems der Elemente, und eine koordinativ gebundene Ligandenhülle, in einem organischen Lösungsmittel, durch Zugabe einer organophilen Base,
2. b. Zugabe eines aktiven C_1-6-Alkans, aufweisend eine Funktionalität und eine Abgangsgruppe, wobei eine kovalente Anbindung des Alkans an die Metalloxid-Nanopartikel erfolgt, so dass ein Niederschlag ausfällt,
3. c. Abtrennung des Niederschlags und Überführung in eine wässrige Lösung,

wobei die kovalente Anbindung in (b) über ein Kohlenstoffatom der C_1-6-Alkylgruppe des aktiven C_1-6-Alkans an den Nanopartikel-eigenen, die Oberflächengitterplätze der Metalloxid-Nanopartikel besetzenden, Sauerstoff erfolgt.To achieve the object, a process for the preparation of surface-hydrophilized metal oxide nanoparticles is specified, comprising the steps:

1. a. Activation of hydrophobic metal oxide nanoparticles containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements, and a coordinatively bound ligand shell, in an organic solvent, by adding an organophilic base,
2. b. Addition of an active C _1-6 alkane having a functionality and a leaving group, wherein a covalent attachment of the alkane to the metal oxide nanoparticles takes place, so that a precipitate precipitates,
3. c. Separation of the precipitate and conversion into an aqueous solution,

wherein the covalent attachment in (b) via a carbon atom of the C _1-6 alkyl group of the active C _1-6 alkane takes place at the nanoparticles own, occupying the surface lattice sites of the metal oxide nanoparticles, oxygen.

For the purposes of the present invention, a "surface-hydrophilized metal oxide nanoparticle" is understood as meaning a metal oxide nanoparticle which has an oxygen-covalently bonded alkyl group having a functionality at the nanoparticle-own, the surface lattice sites of the metal oxide nanoparticles. The functionality is preferably a hydrophilic functionality (ie polar functional group) which is in particular selected from hydroxyl, amino (in particular NH ₂ groups), carboxyl, aldehyde and keto group.

The inventors have now surprisingly found that with the process according to the invention, monodisperse, surface-hydrophilized metal oxide nanoparticles, i. the standard deviation of the mean particle diameter is not more than 10%, with an average particle diameter of less than 50 nm while maintaining the desired monodisperse properties.

After the attachment of the alkane to the metal oxide nanoparticles, a covalent bond between the particle and the hydrophilic residue is advantageously present, so that no detachment of the hydrophilic constituents on the particle surface by ligand exchange reactions is possible, and thus consistent hydrophilic properties of the surface-hydrophilized metal oxide nanoparticles and at the same time high colloidal stability is ensured.

In addition, the method according to the invention allows the stable surface hydrophilization of monodisperse magnetite nanoparticles without hydrophobic constituents on the particle surface, so that particularly advantageous particle aggregation due to the interaction of hydrophobic constituents is minimized.

The complete displacement of the coordinatively bound ligand shell by the attachment of the short-chain, active C _1-6 alkane to the metal oxide nanoparticles minimizes the hydrodynamic radius of the particles.

Due to the hydrophilic functionality of the attached C _1-6 alkane radical to the surface of the metal oxide nanoparticles, the polar properties of the hydrophilic functionality are particularly advantageously transferred to the nanoparticles, so that a suspension of the hydrophilic particles in polar organic solvents (such as. Acetonitrile, acetone, methanol, ethanol) is possible.

Preferably, a negative or positive zeta potential is realized by the covalent attachment of the alkane, which leads to repulsion of the hydrophilized metal oxide nanoparticles and thus advantageously prevents the agglomeration of the particles. Preferably, the average negative zeta potential is in the range of -15 to -80 mV, more preferably in the range of to -25 to -60 mV. The mean positive zeta potential is in particular in the range of +15 to +70 mV, particularly preferably in the range of up to +25 to +50 mV.

The term metal oxide nanoparticles includes both spherical particles and so-called nanotubes or whiskers. It should be noted that the inventive method is basically independent of the particle sizes of the metal oxide nanoparticles. The dimensions of the nanoparticles are preferably less than 50 nm in at least two dimensions, preferably in the range from 1 to 20 nm. The mean particle diameter of the spherical nanoparticles is preferably less than 50 nm, more preferably in the range from 1 to 20 nm, most preferably in the range from 1 to 10 nm.

Preference is given to using metal oxide nanoparticles containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements; in particular, the metals selected from iron, ruthenium, osmium, cobalt, nickel, palladium and platinum are very particularly preferred are iron, ruthenium, osmium, cobalt and nickel. But also binary, ternary and quaternary mixtures of these metals are included.

According to a particularly preferred embodiment of the present invention, spherical metal oxide nanoparticles are used which have an average particle diameter in the range of 1 nm to 50 nm, preferably 1 to 20 nm and metal oxides of metals of the 8th group of the Periodic Table of the Elements (also iron group ), in particular iron (as hematite or magnetite) are constructed.

The organic solvent preferably has the hydrophobic metal oxide nanoparticles with a concentration of up to 10 mg / ml of solvent, more preferably in the range of 0.1 to 8 mg / ml.

The person skilled in the art is aware that the coordinatively bound ligand shell of metal oxide nanoparticles at the same time acts as a surface stabilizer and also mediates the solubility in the solvent used.

According to the invention, it is preferable to use metal oxide nanoparticles which have nonpolar side chains as coordinatively bound ligand shell and thereby mediate the solubility in the organic solvent. However, non-polar side chains are poorly or not completely soluble due to the lack of hydrogen bonds in water. Thus, the metal oxide nanoparticles used according to the invention, having a coordinatively bound ligand shell of non-polar side chains, therefore tend to coalesce in an aqueous solvent and differ from the water.

According to a preferred embodiment of the present invention, the coordinatively bound ligand shell of the metal oxide nanoparticles is composed of organic ligands. The organic ligands are preferably selected from the group of long-chain, organic C _8-20 -carboxylic acids (for example long-chain fatty acids) and / or long-chain, organic C _8-20 -alkylamines. Suitable long-chain, organic C _8-20 carboxylic acids for surface stabilization and solubilization are known in the art, with the fatty acids oleic, linoleic, stearic, palmitic, tetradecanoic, undecanoic prove to be particularly suitable. Long-chain, organic C _8-20 -alkylamines include according to the invention in particular oleylamine, dodecylamine, laurylamine, octylamine, trioctylamine, dioctylamine and hexadecylamine. In particular, the ligands are selected from oleic acid and / or oleylamine.

In the case of the hydrophobic metal oxide nanoparticles, the coordinated attachment of the ligand shell to the hydrophobic metal oxide nanoparticles is preferably carried out via amino and / or carboxyl functionalities.

The organic solvent, in which the metal oxide nanoparticles containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements, and a coordinating ligand shell are initially introduced, can be selected from the group consisting of an ether-based compound (C _n OC _n , 4 ≤ n ≤ 30), a hydrocarbon compound (C _n H _{2n + 2} , 10 ≤ n ≤ 25) and an unsaturated hydrocarbon compound (C _n H _2n , 7 ≤ n ≤ 30).

According to the invention, it is advantageous that the organic solvent has a high thermal stability or high boiling point (T _B ≥ 180 ° C.) to the high reaction temperatures in the range of 175 to 320 ° C, particularly preferably in the range of 220 to 295th To reach ° C.

The organic solvent is preferably selected on the basis of ether, specifically from the group consisting of trioctylphosphine oxide (TOPO), alkylphosphine, octyl ether, benzyl ether and phenyl ether (for example diphenyl ether), these solvents advantageously also having a certain polarity of ≥ 1.7 (according to Snyder). exhibit.

Preferably, the hydrocarbon compound is selected from the group consisting of hexadecane, heptadecane and octadecane. The unsaturated hydrocarbon compound may be selected from the group consisting of octene, heptadecene and octadecene.

According to a preferred embodiment of the invention, the organic solvent is a hydrocarbon compound, more preferably hexane.

According to the invention, the term "active C _1-6 -alkane" refers to an optionally branched C _1-6 -alkyl group which has at least one functionality and a, preferably terminal, leaving group, the leaving group undergoing a nucleophilic substitution reaction with an activated metal oxide. Nanoparticles can be replaced. The functionality is preferably a hydrophilic functionality (ie polar functional group), which is in particular selected from hydroxy, amino (in particular primary NH ₂ groups), carboxyl, aldehyde and keto group. Alternatively, the functionality is a terminal alkyne group (-C≡CR, where R is hydrogen (H) or C _1-3 alkyl group).

Most preferably, the hydrophilic functionality is an amino or carboxyl group.

It can be provided in principle that the hydrophilic functionality of the alkyl group is blocked when added in the form of active C _1-6 alkane by a protective group known in the art. The blocking of the hydrophilic functionality of the alkyl group by a protecting group is particularly well-known to those skilled in the art, when the active alkane has a C _3-6 alkyl group. The protecting group is cleaved off after the reaction so that the free (ie unprotected) hydrophilic functionality is released. The skilled worker is aware of the reaction conditions required for the removal of the protective group.

In particular, active alkanes are preferred in which the hydrophilic functionality in the bound state has a pronounced acidity, ie pKs <5, preferably <4 (eg carboxyl group) or pronounced basicity, ie pK _B <5, preferably <4 (eg amino group) and therefore acts as a solubilizer in aqueous solutions through the dipole-dipole interactions, with active alkanes having a carboxyl group being found to be particularly suitable hydrophilic functionality.

A subsequent functionalization of the hydrophilic functionality and thus a change in the zeta potential are in a simple manner, for example by coupling reactions with peptides, low molecular weight polyethers having a C ₃ to C ₁₈ carbon skeleton, especially low molecular weight polyethylene glycols having an average molecular weight of 200 to 400 g / mol , Fatty acids or C _1-20 -alkylamines possible. So is particularly advantageous when using C _1-6 alkanes having a carboxyl group as a hydrophilic functionality, subsequently a simple surface modification of the metal oxide nanoparticles, for example. By esterification, amidation with other molecules possible.

In particular, when using C _1-6 alkanes, in particular C _3-6 alkanes, ie having a propyl, butyl, pentyl or hexyl radical, it can be provided that the carboxyl group is protected in the form of a carboxylic acid ester and the release of the carboxyl group (ie conversion into the active form: -COOH) takes place only after the covalent attachment of the alkane to the metal oxide nanoparticles via a saponification reaction (for example base- or acid-catalysed hydrolysis reaction) of the carboxylic ester known to the person skilled in the art.

When using active C _1-6 alkanes, in particular C _3-6 alkanes, it may be provided that the amino group is protected in the form of a phthalimide, benzyl or alkoxycarbonyl-amine and the release of the amino group (ie conversion into the free form: -NH ₂ ) takes place only after the covalent attachment of the alkane to the metal oxide nanoparticles via a reduction or hydrogenation reaction known to those skilled in the art (for example by addition with NaBH ₄ or hydrazine or a base- or acid-catalyzed hydrolysis reaction) ,

According to an alternatively preferred embodiment of the present invention, the active alkane has as functionality a terminal alkyne group. Such surface-functionalized metal oxide nanoparticles are particularly suitable for use in a 1,3-dipolar [3 + 2] cycloaddition reaction between the alkyne group and a 1,3-dipolar organic compound. Preferably, the 1,3-dipolar compound is an azide, diazoalkane or nitrone.

An example of a 1,3-dipolar [3 + 2] cycloaddition reaction known today as the "click reaction" is the Cu (I) -catalyzed cycloaddition between terminal alkynes and azides for the synthesis of 1,4- and 1,5 disubstituted aromatic 1,2,3-triazoles [ Huisgen, R .; Szeimies, G .; Moebius, L .; Chem. Ber. 1967, 100, 2494 ]. Sharpless discovered the catalyzed type of reaction in 2002 and recognized its potential to combine any chemical "building blocks" in a highly efficient manner [ Lewis, WG; Green, LG; Grynszpan, F .; Radic, Z .; Carlier, PR; Taylor, P .; Finn, MG; Sharpless, BK; Angew. Chem., Int. Ed. 2002, 41, 2596 ]. It is now known to the person skilled in the art that this reaction can be used in the context of the principle known as "click chemistry" for the synthesis of biologically active substances and of polymers.

As a good leaving group of the active C _1-6 -alkane for a nucleophilic substitution reaction with an activated metal oxide nanoparticles halides, in particular iodide and bromide have been found to be particularly suitable.

Alternatively it can be provided that the leaving group of the active C _1-6 -alkane is an alkylsulfonic acid ester residue (eg mesyl, trifyl group) or an arylsulfonic acid residue residue (eg tosyl, nosyl group). Such radicals have a strong electron-withdrawing effect on the alkyl radical, for which reason the underlying sulfonic acid group represents a light leaving group.

The complete displacement of the coordinatively bound ligand shell on the surface of the metal oxide nanoparticles via the attachment of the short-chain, active C _1-6 alkane, in particular a C _1-3 alkane leads to a minimization of the hydrodynamic radius of the metal oxide nanoparticles over the metal oxide Nanoparticles having a coordinatively bound ligand shell. In addition, the distance between the surface of the metal oxide nanoparticles and the hydrophilic functionality by the attachment of a short-chain C _1-6 alkyl radical, in particular a C _1-3 alkyl radical to the surface of the nanoparticles is only 1 to 6 CH ₂ Group, more preferably 1 to 3 CH ₂ group.

According to a preferred embodiment of the present invention, the active C _1-6 -alkane is an α, ω-C _1-6 -carboxylic acid alkyl halide, in particular an α, ω-C _1-3 -carboxylic acid alkyl halide selected from iodoacetic acid (pKs = 3.18) , Bromoacetic acid (pK _s = 2.90), iodopropanoic acid, bromopropanoic acid. Most preferably, the active C _1-6 alkane is iodoacetic acid, bromoacetic acid or iodopropanoic acid.

According to an alternative preferred embodiment of the present invention, the active alkane is an α, ω-C _1-6 -Carbonsäurealkylsulfonsäureester, in particular an α, ω-C _1-3 -Carbonsäu-realkylsulfonsäureester.

The addition of the active alkane to the activated metal oxide nanoparticles is preferably carried out in a ratio of 10: 1 to 1: 1, calculated as mass equivalent to the nanoparticle.

According to the invention, the term "organophilic base" is understood to mean a mixture containing a strong base and a phase transfer agent, which can be mixed homogeneously or at least colloidally with an organic solvent.

Preferred strong bases are inorganic bases, in particular those selected from the group of alkali metal and alkaline earth metal hydroxides (for example NaOH, KOH, Ca (OH) ₂ ), the group of alkali metal carbonates (for example Na ₂ (CO ₃ ), K ₂ (CO ₃ ). ) and the alkali metal hydrides (eg NaH). These strong bases can be used as solids, typically at a mass concentration of at least 5 wt%, more preferably in the range of 10 to 30 wt%.

The phase transfer agent is present in a sufficient concentration to sufficiently stabilize the strong base in the organic solvent. Typically, the stabilization of the strong base is affected by the length of the hydrocarbon chain in the phase transfer agent, which can be selected by those skilled in the art. Preferably, the phase transfer agent is in a mass concentration in the range of 0.05 to 10 wt .-%, preferably 0.1 to. 5% by weight, although this may vary with the type of phase transfer agent used.

Examples of suitable phase transfer agents include quaternary onium salts, i. basic quaternary onium salts (ie hydroxides), non-basic quaternary onium salts such as quaternary onium halides (eg chlorides), hydrogensulfates, crown ethers, preferably [15] crown-5, [18] crown-6, [21] crown-7 or one of the derivatives and others known to those skilled in the art.

According to a particularly preferred embodiment of the present invention, the organophilic base comprises a mixture of a crown ether, preferably [15] crown-5, [18] crown-6, [21] crown-7 or one of the derivatives thereof, and an alkali metal carbonate, preferably Na ₂ (CO ₃ ) or K ₂ (CO ₃ ). The organophilic base preferably has the crown ether and the alkali metal carbonate in a molar ratio in the range of 10: 1 and 1:10, preferably in the range of 1: 3 and 3: 1.

When using alkali metal hydrides as a strong base it can be provided that the phase transfer agent is a saturated hydrocarbon having the general empirical formula C _n H _{2n + 2} (5 ≦ n ≦ 20), in particular paraffin oil or petroleum ether (usually a mixture of alkanes such as pentane / Hexane) is.

According to a preferred embodiment of the present invention, the strong base and the phase transfer agent are used to activate the hydrophobic metal oxide nanoparticles individually, i. added separately to the hydrophobic metal oxide nanoparticles, which forms the organophilic base in situ. It can be provided that the strong base and / or the phase transfer agent, preferably the phase transfer agent are initially charged in an organic solvent.

Particularly preferred organic solvents here are compounds based on ethers (C _n OC _n , 4 ≦ _n ≦ 30), a saturated hydrocarbon compound (C _n H _{2n + 2} , 7 ≦ n ≦ 30) and an unsaturated hydrocarbon compound (C _n H _2n , 7 ≤ n ≤ 30).

The organic solvent is particularly preferably an ether-based compound, very particularly preferably benzyl ether or phenyl ether (for example diphenyl ether).

According to an alternatively preferred embodiment of the present invention, it can be provided that the strong base and the phase transfer agent for activation of the hydrophobic metal oxide nanoparticles are initially introduced together in an organic solvent, the organophilic base being introduced ex situ.

• 1. die C_1-6-Alkylgruppen über ein Kohlenstoffatom kovalent an den Nanopartikel-eigenen, die Oberflächengitterplätze der Metalloxid-Nanopartikel besetzenden, Sauerstoff gebunden sind, wobei
• 2. die C1-6-Alkylgruppen eine hydrophile Funktionalität aufweisen.

The invention also provides surface-hydrophilized metal oxide nanoparticles of high colloidal stability, comprising at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements whose surface is formed by C _1-6 -alkyl groups, wherein:

• 1. the C _1-6 alkyl groups are covalently bound via a carbon atom to the nanoparticle-own, the surface lattice sites of the metal oxide nanoparticles, oxygen, wherein
• 2. the C1-6 alkyl groups have a hydrophilic functionality.

The surface-hydrophilized metal oxide nanoparticles of the invention are preferably prepared by the process according to the invention.

The term "colloidal stability" describes the quasi-stable state of a solution containing homogeneously distributed (i.e., unagglomerated) particles of approximately equal size, with their further growth being largely inhibited by agglomeration.

For the purposes of the present invention, nanoparticle-specific oxygen atoms mean that the oxygen atoms have been introduced into the crystal lattice during the nanoparticle growth directly in the (wet) chemical synthesis (i.e., chemical conversion of the starting materials) of the metal oxide nanoparticles. Accordingly, nanoparticle-foreign oxygen atoms would be understood to mean those which were subsequently, i. after the (wet) chemical synthesis of the metal oxide nanoparticles, in particular by the addition of further chemicals or a downstream process step (eg. Ligandaustausch), have been registered in the crystal lattice.

The person skilled in the art is fundamentally aware that the positively charged metal ions and negatively charged oxide ions of a metal oxide nanoparticle are arranged in a defined crystal lattice. Thus, the crystal lattice in metal oxide nanoparticles, according to the underlying crystal modification of the respective metal oxide, alternately formed by positively charged metal ions and negatively charged oxide ions. In a crystal lattice, one distinguishes so-called core lattice sites inside the nanoparticle, i. the ions are located inside the particle and therefore completely surrounded by nanoparticle's own oxygen and metal ions, and surface grid sites, i. The ions on the surface of the nanoparticle are covalently or coordinatively saturated in at least one position by foreign ions (for example H).

Preferably, the C _1-6 alkyl group has as functionality a hydrophilic functionality (ie polar functional group), which is in particular selected from hydroxy, amino (in particular NH ₂ groups), carboxyl, aldehyde and keto group. Alternatively, the functionality is a terminal alkyne group (-C≡CR, wherein R is hydrogen (H) or C _1-3 alkyl group) on.

Most preferably, the hydrophilic functionality is an amino or carboxyl group. It can be provided in principle that the hydrophilic functionality of the alkyl group is blocked by the direct synthesis according to the invention of the surface-hydrophilized metal oxide nanoparticles initially by a protective group known in the art. The protecting group is cleaved off after the synthesis according to the invention so that the free (i.e., unprotected) hydrophilic functionality is released. The skilled worker is aware of the reaction conditions required for the removal of the protective group.

Preference is given to C _1-6 -alkyl groups whose hydrophilic functionality has a pronounced acidity (for example carboxyl group) or basicity (for example amino group) and therefore act as solubilizers in aqueous solutions due to the dipole-dipole interactions, where C _1-6 - Alkyl groups with a carboxyl group have been found to be particularly suitable hydrophilic functionality.

According to a preferred embodiment of the present invention, the C _1-6 -alkyl group is a C _1-6 -alkylcarboxylic acid, in particular a C _2-3 -alkylcarboxylic acid selected from ethylcarboxylic acid, isopropylcarboxylic acid or n-propylcarboxylic acid. Most preferably, the C _1-6 alkyl group is ethylcarboxylic acid.

According to a preferred embodiment of the present invention, the attachment of the C _1-6 -alkyl group to the nanoparticle-own, the surface lattice sites of the metal oxide nanoparticles occupying oxygen via a terminal carbon atom of the C _1-6 alkyl group is prepared.

The percentages by weight (% by mass) denotes the relative mass-related proportion of a specific component in a mixture based on its total mass (ie 1 mg per 100 mg = 1 mass%), ie the ratio of the specific component to the sum of the masses all components contained in the mixture and the mass of the specific component itself.

The invention also provides the use of the surface-hydrophilized metal oxide nanoparticles of high colloidal stability according to the invention for subsequent functionalization of the hydrophilic functionality. The subsequent functionalization of the surface-hydrophilized metal oxide nanoparticles according to the invention is preferably carried out by coupling reactions with peptides, oligo- or polynucleotides known to the person skilled in the art, low molecular weight polyethers having a C ₃ to C ₁₈ carbon skeleton, in particular low molecular weight polyethylene glycols having a mean molecular weight of from 200 to 10 000 g / mol, activated fatty acids or C _1-20 -alkylamines.

Advantageously, the zeta potential can be modified in a simple manner by the subsequent functionalization of the hydrophilic functionality of the surface-hydrophilized metal oxide nanoparticles according to the invention.

The present invention also encompasses the use of the subsequently functionalized surface-hydrophilized metal oxide nanoparticles, in particular functionalized with peptides or oligo- or polynucleotides, as biological (active ingredient) transport probes, for example for so-called drug delivery or gene delivery.

A further subject of the present invention is the use of the surface-hydrophilized metal oxide nanoparticles according to the invention as contrast agents for magnetic resonance imaging (MRI).

The features of the invention disclosed in the specification and the claims may be of importance both individually and in any combination for the realization of the invention in its various embodiments.

embodiments

Further features and advantages of the invention will become apparent from the following embodiments, without limiting the invention to these.

The compounds of the general formula (I) according to the invention can be obtained in various ways. The manufacturing methods described below also form part of the present invention.

The above formulas and the above-described synthetic steps are in 1 summarized.

• 2: den hydrodynamischen Durchmesser von Magnetitnanopartikeln mit Essigsäure-Oberflächengruppen in Wasser (DLS-Messung).
• 3: das Zetapotential von Magnetitnanopartikeln mit Essigsäure-Oberflächengruppen in Wasser.
• 4: das Infrarot-Spektrum von eingesetzten hydrophoben Magnetitnanopartikeln sowie oberflächenhydrophilisierte Magnetitnanopartikeln (hergestellt in Beispiel 1).
• 5: das Infrarot-Spektrum der Syntheseprodukte A bis D (von unten nach oben).
• 6: das Infrarot-Spektrum der Syntheseprodukte A, E bis G (von unten nach oben).

In addition shows:

• 2 : the hydrodynamic diameter of magnetite nanoparticles with acetic acid surface groups in water (DLS measurement).
• 3 : the zeta potential of magnetite nanoparticles with surface acetic acid groups in water.
• 4 The infrared spectrum of hydrophobic magnetite nanoparticles used and surface-hydrophilized magnetite nanoparticles (prepared in Example 1).
• 5 : the infrared spectrum of the synthesis products A to D (from bottom to top).
• 6 : the infrared spectrum of the synthesis products A, E to G (from bottom to top).

Example 1: Surface Hydrophilization of Magnetite Nanoparticles

5 mg of Fe ₃ O ₄ hydrophobic nanoparticles are placed in a 50 ml flask and suspended with 1 ml of hexane. 30 mg of K ₂ CO _{3 are} added. Separately crown ether [18] crown-6 becomes equimolar to K ₂ CO ₃ dissolved in 15 ml of diphenyl ether and added to the nanoparticle suspension. With vigorous stirring ( 800 rpm), the addition of 6 mg of iodoacetic acid takes place after 15 minutes. The suspension is slowly heated to the boiling point of hexane, which is completely evaporated. Subsequently, the remaining suspension is heated to a temperature of 165 ° C. To 3 h, the reaction solution is refluxed for 90 minutes (between 220-265 ° C). Afterwards, the mixture is cooled to room temperature. The suspension is centrifuged ( 4500 rpm, 15 min), the supernatant being discarded. The remaining solid is resuspended by means of ultrasound bath in 10 ml of hexane and then centrifuged again ( 4500 rpm, 15 min). Subsequently, the solid is again resuspended by means of ultrasound bath in 10 ml of hexane and deposited with a magnet on the vessel wall. The solvent is discarded. The solid is then resuspended in methanol in an ultrasound bath and centrifuged ( 4500 rpm, 15 min). The supernatant is discarded, the solid dried and then resuspended in 1 ml of water. The rust-brown suspension is removed by dialysis membrane (Membra-Cel Dialysis Tubing: regenerated cellulose MWCO 7000 , Serva) dialyzed against deionized water. The yield of the represented hydrophilic nanoparticles is quantitative. The hydrodynamic diameter of the resulting Fe ₃ O ₄ nanoparticles is about 8 nm on average (DLS measurement, Malvern Zetasizer; 2 ).

Example 2: Measurement of the zeta potential

The zeta potential is measured by means of Zetasizer Nano ZS (Malvern Instruments Ltd., UK) with 4 mW He-Ne laser. For sample measurement, 15 scans are performed for 10 s / scan at a temperature of 25 ° C and a measurement angle of 173 ° in water (viscosity after factory setting). The binding of a multiplicity of carboxyl groups on the particle surface in Example 1 results in a negative zeta potential (mean: -41.7 mV, 3 ), which prevents the agglomeration of the particles.

Example 3: IR Spectroscopic Characterization

The IR spectroscopic characterization of the surface-hydrophilized magnetitnanoparticles prepared in Example 1 is carried out with a Nicolet iS5, Thermo Scientific at a resolution of 2 cm ^-1 . The obtained ATR-IR spectra were corrected by means of 2 point baseline and normalized to maximum.

The carboxylic acid units of the acetic acid residues can be recognized in the IR spectrum by the CO stretching vibrations at 1635 cm ^-1 (C = 0) and the deformation vibration at 1400 cm ^-1 (C-OH). The gang 3250 cm ^-1 describes the OH valence vibration of the acid. The intensive CH ₂ -Valenzschwingungen ( 2914 cm ^-1 : asymmetric; 2850 cm ^-1 : symmetric) are characteristic of the hydrophobic ligands oleylamine and (Z) -9-octadecenoic acid.

The low intensity in the range of surface-hydrophilized magnetite nanoparticles observed in this range can be attributed to the CH ₂ valence vibrations of the acid radical and indicates complete detachment of the hydrophobic ligands, which is confirmed by the results of the elemental analysis (no nitrogen detectable, see Table 1) becomes.

The determination of the composition of the different magnetite samples (ie the CHNS content) was carried out by elemental analysis (EURO EA, Elemental Analyzer, Eurovector, HEKAtech GmbH). Table 1: sample N% C% H% S% Mass [mg] Fe ₃ O ₄ ("hydrophobic") 0.17 22.29 3.76 0 1,299 0.17 22.27 3.80 0 1,407 Fe ₃ O ₄ ("hydrophilic") 0.09 3.65 0.62 0 1,182 0 3.75 0.53 0 1,131 0 3.75 0.53 0 1,649

There are about 790 nmol / mg of NP hydrophobic C18 ligands on the surface, which are composed of 8 parts of (Z) -9-octadecenoic acid and one part of oleylamine. The functionalization with iodoacetic acid leads to the attachment of 1.42 μmol acetic acid groups / mg NP.

Example 4: Synthesis product B:

5 mg of Fe ₃ O ₄ hydrophilic nanoparticles are placed in a 5 ml flask and suspended in 1 ml of water. 2 mg of EDC (10.4 .mu.mol) and 1.5 mg of mono-boc-diaminopropane (8.7 pmol) are added and reacted at 25.degree. C., 750 rpm for 8 hours. Subsequently, the particles are precipitated by the addition of 500 .mu.l of acetonitrile and deposited by means of a magnet on the vessel wall. The solvent is discarded. Subsequently, the solid is resuspended in 300 μl of water in an ultrasound bath, precipitated by the addition of 100 μl of acetonitrile and centrifuged ( 4500 rpm, 10 min). After discarding the supernatant to obtain synthesis product B.

IR-Banden[cm^-1]:

3250 (v-OH), 1660 (v_as-CO), 1305 (v-C-O (Boc-Schutzgruppe)), 1150, 1060, 850, 784 (p-CH₂)

Functionalization with Mono-Boc-Protected Diaminopropane: Synthesis Product B

IR bands [cm ^-1 ]:

3250 (v-OH), 1660 ( _as -CO), 1305 (vCO (Boc-protecting group)), 1150, 1060, 850, 784 (p-CH ₂ )

Elemental analysis: sample N% C% H% S% Mass (mg) B 0.55 5.08 0.68 0 1,222

The further functionalization with mono-Boc-diaminopropane leads to about 200 nmol Boc protected amino groups (detection amine: 196 nmol, detection carbon: 174 nmol), which corresponds to a share of 14% (carbon: 12.2%) of the possible functionalizable acid groups.

Example 5: Synthesis product C:

5 mg of synthesis product B from Example 4 are resuspended in 1 ml of dry acetonitrile by means of an ultrasound bath and admixed with 50 μl of trifluoroacetic acid. The suspension is reacted at 25 ° C, 750 rpm for 30 min. Subsequently, the particles are deposited by means of a magnet on the vessel wall, the solvent discarded and treated with 500 ul acetonitrile. In the ultrasonic bath, the particles are resuspended and separated again by means of a magnet. After repeating 3 times, synthesis product C is obtained.

IR-Banden [cm^-1]:

3250 (v-OH), 2920 (v_as-CH₂), 2850 (vs CH₂), 1660 (v_as-CO), 1423 (δ-C-OH), 1288, 1204 (TFA Rest), 1150 (TFA Rest), 1045, 850, 784 (p-CH₂) Elementaranalyse:

Cleavage of the Boc-protecting group of B: Synthesis product C

IR bands [cm ^-1 ]:

3250 (v-OH), 2920 (v _as -CH ₂ ), 2850 (vs CH ₂ ), 1660 ( _as -CO), 1423 (δ-C-OH), 1288, 1204 (TFA residue), 1150 ( TFA residue), 1045, 850, 784 (p-CH ₂ ) Elemental Analysis:

Elemental analysis: sample N% C% H% S% Mass (mg) C 0.42 5.51 0.71 0 1,085

The cleavage of the Boc protective group with trifluoroacetic acid leads according to elemental analysis to about 150 nmol free amino groups, which corresponds to a proportion of 10.5% (carbon: 41%) of the possible functionalizable acid groups.

Example 6: Synthesis product D:

5 mg of synthesis product C is resuspended in 1 ml of water and admixed with 0.5 mg of iminothiolane (3.65 μmol) (Sigma-Aldrich) and 0.1 mg of Na ₂ CO ₃ (Sigma-Aldrich). The suspension is reacted at 25 ° C, 750 rpm for 12 h. Subsequently, the particles are precipitated by the addition of 500 .mu.l of acetonitrile and deposited by means of a magnet on the vessel wall. The solvent is discarded. Subsequently, the solid is resuspended in 300 μl of water, precipitated by the addition of 100 μl of acetonitrile and centrifuged ( 4500 rpm, 10 min). After discarding the supernatant to obtain synthesis product D.

IR-Banden [cm^-1]:

3250 (v-OH), 2920 (v_as-CH₂), 2850 (v_s--CH₂), 1660 (v_as-CO), 1423 (δ-C-OH), 1288, 1150, 1095, 1045, 784 (p-CH₂)

Coupling of iminothiolane to the amino group of C: Synthesis product D

IR bands [cm ^-1 ]:

3250 (v-OH), 2920 (v _as -CH ₂ ), 2850 (v _s -CH ₂ ), 1660 ( _as -CO), 1423 (δ-C-OH), 1288, 1150, 1095, 1045 , 784 (p -CH ₂ )

Elemental analysis: sample N% C% H% S% Mass (mg) D 0.64 4.93 0.92 0.41 1,262

A conversion with iminothiolane leads according to the elemental analysis to 117 nmol / mg NP thiol groups or a reaction yield of 78% of the possible free amino groups.

Example 7: Synthesis product E:

5 mg of Fe ₃ O ₄ hydrophilic nanoparticles are placed in a 5 ml flask and suspended in 1 ml of water. 2 mg of EDC (10.4 pmol) and 3.4 mg (10.6 pmol) of Boc-Tota (IrisBiotech) are added and reacted at 25 ° C., 750 rpm for 8 h. Subsequently, the particles are precipitated by the addition of 500 .mu.l of acetonitrile and deposited by means of a magnet on the vessel wall. The solvent is discarded. Subsequently, the solid is resuspended in 300 μl of water in an ultrasound bath, precipitated by the addition of 100 μl of acetonitrile and centrifuged ( 4500 rpm, 10 min). After discarding the supernatant to obtain synthesis product E.

IR-Banden [cm^-1]:

3250 (v-OH), 1660 (v_as-CO), 1305 (v-C-O (Boc-Schutzgruppe)), 1060, 850, 784 (p-CH₂)

Functionalization of A with Boc-Tota (IRIS-Biotech): Synthesis product E

IR bands [cm ^-1 ]:

3250 (v-OH), 1660 ( _as -CO), 1305 (vCO (Boc-protecting group)), 1060, 850, 784 (p-CH ₂ )

Elemental analysis: sample N% C% H% S% Mass (mg) e 0.28 6.6 0.78 0 1,222

The further functionalization with mono-Boc-diaminopropane leads according to elemental analysis to about 177 nmol Boc-protected amino groups, which corresponds to a proportion of 12.5% (nitrogen: 7%) of the possible functionalizable acid groups.

Example 8: Synthesis product F

5 mg of synthesis product E is resuspended in 1 ml of dry acetonitrile by means of an ultrasound bath, and 50 μl of trifluoroacetic acid are added. The suspension is reacted at 25 ° C, 750 rpm for 30 min. Subsequently, the particles are deposited by means of a magnet on the vessel wall, the solvent discarded and treated with 500 ul acetonitrile. In the ultrasonic bath, the particles are resuspended and separated again by means of a magnet. After repeating 3 times, synthesis product F. is obtained.

IR-Banden [cm^-1]:

3250 (v-OH), 1660 (v_as-CO), 1405 (δ-C-OH), 1290, 1204, 1160 (δ-NH₂), 1045, 784 (p-CH₂)

Cleavage of the Boc protecting group of E: Synthesis product F

IR bands [cm ^-1 ]:

3250 (v-OH), 1660 ( _as -CO), 1405 (δ-C-OH), 1290, 1204, 1160 (δ -NH ₂ ), 1045, 784 (p-CH ₂ )

Elemental analysis: sample N% C% H% S% Mass (mg) F 0, 21 5, 74 0, 87 0 1,128

The cleavage of the Boc Scutzgruppe with trifluoroacetic leads according to the elemental analysis to about 194 nmol free amino groups, which corresponds to a share of 13.7% (nitrogen: 5.3%) of the possible functionalizable acid groups.

Example 9: Synthesis product G:

5 mg of synthesis product F is resuspended in 1 ml of water and admixed with 0.5 mg of iminothiolane (3.65 μmol) (Sigma-Aldrich) and 0.1 mg of Na ₂ CO ₃ (Sigma-Aldrich). The suspension is reacted at 25 ° C, 750 rpm for 12 h. Subsequently, the particles are precipitated by the addition of 500 .mu.l of acetonitrile and deposited by means of a magnet on the vessel wall. The solvent is discarded. Subsequently, the solid is resuspended in 300 μl of water, precipitated by the addition of 100 μl of acetonitrile and centrifuged ( 4500 rpm, 10 min). After discarding the supernatant to obtain synthesis product G.

IR-Banden [cm^-1]:

3250 (v-OH), 2960, 2920(v_as-CH₂), 2850 (v_s-CH₂), 1660 (v_as-CO), 1428 (δ-C-OH), 1045, 784 (p-CH₂)

Coupling iminothiolane to the amino group of F: Synthesis product G

IR bands [cm ^-1 ]:

3250 (v-OH), 2960, 2920 (v _as -CH ₂ ), 2850 (v _s -CH ₂ ), 1660 ( _as -CO), 1428 (δ-C-OH), 1045, 784 (p-). CH ₂ )

Elemental analysis: sample N% C% H% S% Mass (mg) G 0.35 4.74 0.93 0.04 1,284

Example 10: Synthesis product H:

10 mg of Fe ₃ O ₄ hydrophobic nanoparticles are placed in a 50 ml flask and suspended with 1.5 ml of hexane. 60 mg of K ₂ CO _{3 are} added. Separately becomes crown ether 18 -Kron-6 equimolar to K ₂ CO _{3 dissolved} in 25 ml of diphenyl ether and added to the nanoparticle suspension. With vigorous stirring ( 800 rpm), the addition of 5 .mu.l Propargylbromid takes place after 15 min. The suspension is slowly heated to the boiling point of hexane, which is completely evaporated. Subsequently, the remaining suspension is heated to 150 ° C. To 3 h, the reaction solution is maintained at reflux temperature for 90 minutes. Afterwards, the mixture is cooled to room temperature. The suspension is centrifuged ( 4500 rpm, 15 min), the supernatant being discarded. The remaining solid is resuspended by means of ultrasound bath in 10 ml of hexane and then centrifuged again ( 4500 rpm, 15 min). Subsequently, the solid is again resuspended by means of ultrasound bath in 10 ml of hexane and deposited with a magnet on the vessel wall. The solvent is discarded, the solid is resuspended in 1 ml of methanol in an ultrasound bath and centrifuged ( 4500 rpm, 15 min). The supernatant is discarded, the solid dried.

Synthesis product H:

Example 11: Synthesis product I:

12 mg of Fe ₃ O ₄ hydrophobic nanoparticles are placed in a 50 ml flask and suspended with 1.5 ml of hexane. 70 mg of K ₂ CO _{3 are} added. Separately becomes crown ether 18 -Kron-6 equimolar to K ₂ CO _{3 dissolved} in 25 ml of diphenyl ether and added to the nanoparticle suspension. With vigorous stirring ( 800 rpm), the addition of 15 mg of N- (2-bromoethyl) phthalimide is carried out after 15 minutes. The suspension is slowly heated to the boiling point of hexane, which is completely evaporated. Subsequently, the remaining suspension is heated to temperatures between 175 ° C. To 3 h, the reaction solution is held for 90 min in a temperature range between 220-265 ° C. Afterwards, the mixture is cooled to room temperature. The suspension is centrifuged ( 4500 rpm, 15 min), the supernatant being discarded. The remaining solid is resuspended by means of ultrasound bath in 10 ml of hexane and then centrifuged again ( 4500 rpm, 15 min). Subsequently, the solid is again resuspended by means of ultrasound bath in 10 ml of hexane and deposited with a magnet on the vessel wall. The solvent is discarded. The solid is then resuspended in methanol in an ultrasound bath and centrifuged ( 4500 rpm, 15 min). The supernatant is discarded, the solid dried.

Synthesis product I:

Example 12: Synthesis product K:

5 mg of synthesis product I is resuspended in 30 ml of ethanol and heated to reflux under a nitrogen atmosphere. After dropwise addition of 6 mass equivalents of hydrazine hydrate, the reaction mixture remains under reflux overnight. The mixture is then cooled to room temperature and the addition of hydrochloric acid. The suspension is centrifuged ( 4500 rpm, 15 min), the supernatant being discarded. The remaining solid is resuspended by means of ultrasound bath in 10 ml of hexane and then centrifuged again ( 4500 rpm, 15 min). Subsequently, the solid is again resuspended by means of ultrasound bath in 10 ml of hexane and deposited with a magnet on the vessel wall. The solvent is discarded. The solid is then resuspended in methanol in an ultrasound bath and centrifuged ( 4500 rpm, 15 min). The supernatant is discarded, the solid dried and then resuspended in 1 ml of water. The rust-brown suspension is removed by dialysis membrane (Membra-Cel Dialysis Tiubing: regenerated cellulose MWCO 7000 , Serva) dialyzed against deionized water. The yield of the represented hydrophilic nanoparticles is quantitative. The hydrodynamic diameter of the resulting Fe ₃ O ₄ nanoparticles is about 9 nm on average.

Coupling of iminothiolane to the amino group of I: Synthesis product K

Claims (14)

A process for producing surface-hydrophilized metal oxide nanoparticles comprising the following steps: a. Activation of hydrophobic metal oxide nanoparticles containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements, and a coordinatively bound ligand shell, in an organic solvent, by adding an organophilic base comprising a strong base and a phase transfer agent, b. Adding an active C _1-6 alkane having a functionality and a leaving group, wherein a covalent attachment of the alkane to the metal oxide nanoparticles takes place so that a precipitate precipitates, c. Separation of the precipitate and conversion into an aqueous solution, characterized in that the covalent attachment in (b) via a carbon atom of the C _1-6 alkyl group of the active C _1-6 alkane on the nanoparticles own, the surface grid sites of the metal oxide Nanoparticle occupying, oxygen takes place. Method according to Claim 1 , characterized in that spherical metal oxide nanoparticles having an average particle diameter of less than 50 nm are used. Method according to Claim 1 or 2 , characterized in that in the case of the hydrophobic metal oxide nanoparticles, the coordinate bonding of the ligand shell to the hydrophobic metal oxide nanoparticles takes place via amino and / or carboxyl functionalities. Method according to one of Claims 1 to 3 , characterized in that as the active C _1-6 alkane, a C _1-6 alkane having a hydrophilic functionality is added. Method according to one of Claims 1 to 3 , characterized in that a C _1-6 alkane having a terminal alkyne group is added as the active C _1-6 alkane, whereby such surface-functionalized nanoparticles are subsequently reacted with a 1,3-dipolar organic compound in a [3 + 2] cycloaddition be implemented. Method according to one of Claims 1 to 5 , characterized in that a quaternary onium salt, hydrogen sulfate or crown ether is used as the phase transfer agent. Method according to one of Claims 1 to 6 characterized in that the organic solvent is an ether-based compound, a hydrocarbon compound or an unsaturated hydrocarbon compound. Method according to Claim 4 , characterized in that an alkane with amino or carboxyl group is added as the active alkane with hydrophilic functionality. Method according to one of Claims 1 to 8th , characterized in that monodisperse surface-hydrophilized metal oxide nanoparticles are produced. A surface-hydrophilized metal oxide nanoparticle containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements whose surface is formed by C _1-6 -alkyl groups, wherein: a) the C _1-6 -alkyl groups via a carbon atom covalently bound to the nanoparticle-own, the surface lattice sites of the metal oxide nanoparticles, oxygen, wherein b) the C _1-6 alkyl groups have a hydrophilic functionality. Surface hydrophilized metal oxide nanoparticles according to Claim 10 , characterized in that the attachment of the C _1-6 alkyl group to the nanoparticle-own, the surface lattice sites of the metal oxide nanoparticles occupying oxygen is produced via a terminal carbon atom. Use of the surface-hydrophilized metal oxide nanoparticles according to Claim 10 or 11 for a subsequent functionalization of the hydrophilic functionality. Use of the surface-hydrophilized metal oxide nanoparticles according to Claim 10 or 11 as a contrast agent for magnetic resonance imaging. Surface-functionalized metal oxide nanoparticles containing at least one metal oxide of a metal selected from the iron-platinum group of the Periodic Table of the Elements whose surface is formed by C _1-6 -alkyl groups, wherein: c) the C _1-6 -alkyl groups via a carbon atom covalently bound to the nanoparticle-own, the surface lattice sites of the metal oxide nanoparticles, oxygen, wherein d) the C _1-6 alkyl groups have a terminal alkyne group.

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