JP2002088075A - Aminated fullerene derivative - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a new nucleic acid-condensing reagent useful for transfection. SOLUTION: This new reagent is an aminated fullerene derivative of the formula [wherein, R1 and R2 are each a (substituted) hydrocarbon group and may form a (substituted) nitrogen-containing heterocyclic ring together with the nitrogen atom to which R1 and R2 are bound].

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to an aminated fullerene derivative which has a nucleic acid condensing ability and is useful, for example, as a nucleic acid condensing reagent, and is useful, for example, in the field of the pharmaceutical industry.

[0002]

BACKGROUND OF THE INVENTION The condensation of nucleic acids in a protein-nucleic acid complex as exemplified by the sequence of the nucleic acids on a chromosome
action is a very important biochemical problem. Transfection by condensation of nucleic acids with small organic molecules or inorganic ions is also an important research topic in connection with gene therapy (eg, Yo).
shikawa, Y. et al, FEBS Letters, 1996, vol. 396, 7
1-76; Behr, JP, Acc. Chem. Res., 1993, vol. 26, 2
74-278; etc.).

[0003] In gene therapy, it is an important point how to safely introduce a nucleic acid to a target site. At present, methods for introduction into cells include a method using a virus, a liposome method using no virus, and a lipofection method. In the method using a virus, there is a method of treating ADA (adenosine deaminase) deficiency using a retrovirus, and there is a track record, but there is a problem in the pathogenicity and immunogenicity of the virus itself. The liposome method and the lipofection method without using a virus still need to be improved in terms of gene transfer efficiency and side effects such as toxicity.

[0004] On the other hand, various methods for chemically modifying fullerenes according to the purpose have been provided so far, and various fullerene derivatives have been synthesized (for example, Friedman,
SH et al, J. Am. Chem. Soc., 1993, vol. 115, 65
06-6509; Yamago, S. et al, J. Am. Chem. Soc., 1994,
vol. 116, 1123; Taki, M. et al, J. Am. Chem. So
c., 1997, vol. 119, 926; An, YZ et al, Tetrahed
ron, 1996, vol. 52, 5179-5189; Nakamura, E. et al,
Bull. Chem. Soc. Jpn., 1996, vol. 69, 2143-2151:
Yamago, S. et al, Chemistry Letters, 1996, 395-39
6; Murata, Y. etal, The 2nd International Forum on
Chemistry of Functional Organic Chemicals (IFOC-
2), 1997, P-31, Tokyo, Japan, etc.).

Recently, an application example of a compound having an amino group as a substituent on fullerene as an optical material has been reported.
Attention has been focused on the synthesis of fullerenes having multiple amino groups (Hutchison, K .; Gao, J .; Schick, G .; Rubi
n, Y .; Wudl, FJ Am. Chem. Soc. 1999, 121, 5611-
5612). As a method for synthesizing a fullerene having an amino group as a substituent, several techniques for synthesizing an aminated fullerene from a direct addition reaction of amines have been reported (Sc
hick, G .; Kampe, K, -D .; Hirsch, AJ Chem. Soc.,
Chem. Commun. 1995, 2023-2024.), There was no procedure to obtain the product in satisfactory yield. Further, in these methods, since the obtained products differ depending on the substituents of the amines or the reaction conditions, it has been desired to establish a method for synthesizing aminofullerene. Further, the present inventors have found that a cage compound can be synthesized from the reaction of fullerene and diamine and have already reported (Isobe, H .;
Ohbayashi, A .; Sawamura, M .; Nakamura, EJ Am.
Chem. Soc. 2000, 122, 2669-2670).

[0006]

SUMMARY OF THE INVENTION An object of the present invention is to provide a novel aminated fullerene derivative having a nucleic acid condensing ability and capable of introducing a nucleic acid into cells, a method for producing the same, and a use thereof.

[0007]

Means for Solving the Problems The present inventors have found that the quadruple addition reaction of a secondary amine to fullerene proceeds well by irradiation with visible light in the presence of oxygen. Such an aminated fullerene derivative or a salt thereof is found to have an amphiphilic and extremely excellent nucleic acid condensing ability, and further found that a complex of an aminated fullerene derivative and a nucleic acid is efficiently taken into cells, The present invention has been completed.

[0008] That is, the present invention relates to a novel aminated fullerene derivative and its use.

(1) The following general formula:

Embedded image

[In the formula, R ¹ and R ² each independently represent a hydrocarbon group which may have a substituent. In addition, R ¹ and R ² may be substituted with a nitrogen atom to which they are bonded. A nitrogen-containing heterocyclic ring which may have a group (however, excluding morpholine and piperidine) may be formed. An aminated fullerene derivative represented by the formula:

(2) The following general formula:

Embedded image [Wherein, R ¹ and R ² are each independently a hydrocarbon group which may have a substituent, and in addition, R ¹ and R ² have a substituent together with the nitrogen atom to which they are bonded. And a nitrogen-containing heterocyclic ring (optionally excluding morpholine and piperidine). An aminated fullerene derivative represented by the formula:

(3) The aminated fullerene derivative according to 1, wherein R ¹ and R ² are each independently a hydrocarbon group which may have a substituent, and a salt thereof; (4) R ¹ and R ² Is an alkylated fullerene derivative and a salt thereof, each independently being an alkyl group;

(5) azetidine, wherein R ¹ and R ² may have a substituent together with the nitrogen atom to which they are attached;
2. The compound according to claim 1, which is a nitrogen-containing heterocycle selected from pyrrolidine and piperazine, (6) the substituent is a (C ₁ -C ₆ ) alkyl group, (C _1-
A C ₆ ) alkylamino group, a (C ₁ -C ₆ ) alkoxycarbonyl group, a (C ₁ -C ₆ ) alkylenedioxy group or the following general formula:

Embedded image [Wherein A is a (C ₁ -C ₆ ) alkylene group, and R ³ is (C ₁ -C ₆ )
₆ ) an alkyl group, wherein n is an integer of 1 to 5].
The compound described,

(7) The following general formula: Wherein R ¹ and R ² are piperidine having a substituent together with the nitrogen atom to which they are bonded, or a salt thereof. (8) The following general formula:

Embedded image [Wherein X is CH ₂ , N (CH ₃ ), N (tertiary butyroxycarbonyl), N—CH ₂ CH ₂ —O—CH ₂ CH ₂ —O
—CH ₂ CH ₂ —O—C ₂ H ₅ or C (OCH ₂ CH ₂ O)
It is. An aminated fullerene derivative or a salt thereof,

(9) [60] fullerene and secondary amine (II
I):

Embedded image And visible light irradiation to produce an aminated fullerene derivative

[0016]

Embedded image

[In the above formula, R ¹ and R ² are each independently a hydrocarbon group which may have a substituent. In addition, R ¹ and R ² may be substituted with a nitrogen atom to which they are bonded. A nitrogen-containing heterocyclic ring which may have a substituent may be formed. (10) Use of the aminated fullerene derivative or a salt thereof according to (9) for the purpose of condensing nucleic acids, (11) consisting of the aminated fullerene derivative or a salt thereof according to (9). (12) a nucleic acid condensate comprising a nucleic acid comprising an aminated fullerene derivative or a salt thereof according to (9) and a nucleic acid; and (13) a nucleic acid condensate to a cell using the nucleic acid condensate according to (12). How to introduce.

[0018]

That is, the gist of the present invention is to provide an aminated fullerene derivative represented by the following general formula (I) or (II), a salt thereof, a method for producing the same, and a nucleic acid condensing action thereof. is there.

[0019]

Embedded image

[In the formula, R ¹ and R ² each independently represent a hydrocarbon group which may have a substituent. In addition, R ¹ and R ² may be substituted together with the nitrogen atom to which they are bonded. A nitrogen-containing heterocyclic ring which may have a group may be formed. The aminated fullerene derivative of the present invention can be used in the form of a salt thereof. In this case, the salt is a conventional non-toxic salt, and particularly preferably a pharmaceutically acceptable salt. Specifically, inorganic acid addition salts (eg, hydrochloride, hydrobromide, sulfate, phosphate, etc.), organic carboxylic acid or sulfonic acid addition salts (eg, formate, acetate, trifluorocarbonate) Acetate, maleate, tartrate, fumarate, methanesulfonate, benzenesulfonate, toluenesulfonate, etc.), basic or acidic amino acids (eg, arginine, aspartic acid, glutamic acid, etc.) Can be mentioned.

The aminated fullerene derivative of the present invention may have various isomers due to the presence of an asymmetric carbon atom and the presence of molecular asymmetry, all of which are included in the aminated fullerene derivative of the present invention.

The "fullerene" in the aminated fullerene derivative of the present invention includes not only C60 fullerene but also
Higher fullerenes (for example, C70 fullerene, etc.) are also included.

The aminated fullerene derivative of the present invention or a salt thereof can be synthesized by a synthesis method known in the art or an appropriate modification thereof according to the structure thereof (the above and the following literatures). reference). For example, the production method of the present invention, that is, [60] fullerene (hereinafter, referred to as
The presence of oxygen excess secondary amine to be present) when referred to as "C _60", to react under visible light irradiation, to obtain aminated fullerene (I) or (II) in high yield be able to. The production method of the present invention is illustrated by the following reaction formula.

Manufacturing method

Embedded image

[In the formula, R ¹ and R ² each independently represent a hydrocarbon group which may have a substituent. In addition, R ¹ and R ² may be substituted with a nitrogen atom to which they are bonded. A nitrogen-containing heterocyclic ring which may have a group may be formed. ]

In the above and following description of the present specification, preferred examples of various definitions included in the scope of the present invention will be described in detail below.

Suitable "hydrocarbon groups" include straight-chain or branched alkyl and alkenyl groups.

As the alkyl group, C ₁ -C ₁₈ alkyl,
Preferably, C ₁ -C ₆ alkyl is used. Examples include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl, pentyl, hexyl and the like.

Examples of the alkenyl group include C ₂ -C ₁₈ alkenyl, preferably C ₂ -C ₆ alkenyl. For example, vinyl, propenyl, isopropenyl, allyl, butenyl, isobutenyl, pentenyl, hexenyl and the like can be mentioned.

"Hydrocarbon group optionally having substituent (s)"
Is a (C ₁ -C ₆ ) alkanoyl group,
Amino group, (C ₁ -C ₆ ) alkylamino group, (C _1-
C ₆ ) alkoxy group, (C ₁ -C ₆ ) alkoxycarbonyl group and the like.

Preferred “nitrogen-containing heterocycle” is a heterocycle having at least one nitrogen atom. Further, it may have one or more hetero atoms selected from the group consisting of oxygen atoms, sulfur atoms, tin, germanium and the like. Preference is given to monocyclic nitrogen-containing heterocycles, such as (1) a 3- to 7-membered (preferably 5- or 6-membered) saturated nitrogen-containing heterocyclic group containing 1 to 4 nitrogen atoms, [eg azetidine , Pyrrolidine, imidazolidine, piperidine, piperazine and the like]; (2) a 3- to 7-membered (preferably 5- or 6-membered) unsaturated nitrogen-containing heterocyclic group containing 1 to 4 nitrogen atoms, for example, pyrrole, pyrroline, Imidazoline, imidazole, pyrazole, triazole [eg, 4H-1,2,
4-triazole, 1H-1,2,3-triazole, 2
H-1,2,3-triazole and the like], tetrazole [for example 1H-tetrazole and the like] and the like; (3) 1 to 3 nitrogen atoms and oxygen atoms, sulfur atoms,
1 to 3 selected from tin atom and germanium atom
And a 3- to 7-membered (preferably 5- or 6-membered) saturated nitrogen-containing heterocyclic group containing, for example, morpholine, thiomorpholine, (4-stanapiperidine, 4-germapiperidine and the like).

The "substituent" of the "nitrogen-containing heterocyclic ring optionally having substituent (s)" includes a (C ₁ -C ₆ ) alkyl group,
(C ₁ -C ₆ ) alkanoyl group, amino group, (C ₁ -C ₆ )
Alkylamino group, (C ₁ -C ₆ ) alkoxycarbonyl group, (C ₁ -C ₆ ) alkylenedioxy group, the following general formula:

Embedded image [Wherein A is a (C ₁ -C ₆ ) alkylene group, and R ³ is (C ₁ -C ₆ )
₆ ) an alkyl group, wherein n is an integer of 1 to 5],
And so on. The substitution position of the substituent is not particularly limited, but is preferably the β-position and the γ-position of the nitrogen atom of the nitrogen-containing heterocycle bonded to fullerene.

Suitable "(C ₁ -C ₆ ) alkanoyl groups" include formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, hexanoyl, 3,3
3-dimethylbutanoyl and the like, and further, hydroxy, (C ₁ -C ₆ ) alkoxy (eg, methoxy, ethoxy, etc.), halogen (fluorine, chlorine, bromine, iodine)
May be substituted.

Preferred "(C ₁ -C ₆ ) alkylamino group"
Examples include "mono (C ₁ -C ₆ ) alkylamino group" and "di (C ₁ -C ₆ ) alkylamino group", for example, methylamino, dimethylamino, ethylamino,
Examples include diethylamino, butylamino, methylbutylamino and the like.

Suitable "(C ₁ -C ₆ ) alkoxycarbonyl group" includes methoxycarbonyl, ethoxycarbonyl and tert-butoxycarbonyl, and may be further substituted with phenyl, halogen and the like (for example, Benzyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl, etc.).

Suitable "(C ₁ -C ₆ ) alkylenedioxy" includes methylenedioxy, ethylenedioxy,
Propylenedioxy and the like. The preferred "(C ₁
Examples of the "-C ₆ ) alkylene group" include a methylene group, an ethylene group, a propylene group, a methylethylene group and the like, and preferably an ethylene group. General formula:

Embedded image [Wherein A is a (C ₁ -C ₆ ) alkylene group, and R ³ is (C ₁ -C ₆ )
₆ ) alkyl group, n is an integer of 1 to 5], and specific examples of the group represented by (1- or 2-) methoxyethyl group, (1- or 2-) ethoxyethyl group, (1- or 2-) 2-) methoxy- (1- or 2-) ethoxyethyl group, (1- or 2-) ethoxy- (1- or 2-)
An ethoxyethyl group, (1- or 2-) ethoxy- (1
-Or 2-) ethoxy- (1- or 2-) ethoxyethyl group and the like.
-Ethoxy-2-ethoxy-2-ethoxyethyl group. 2. Method for Producing Aminated Fullerene Derivative of the Present Invention The method for producing the aminated fullerene derivative of the present invention will be described more specifically.

Production Method ([60] Secondary A
Photoaddition reaction of min) [60] The target compound (I) or (II) can be produced by reacting fullerene with a secondary amine (III) in the presence of oxygen under a visible light source. As a visible light source that can be used in this reaction, a visible light source having a wavelength of 400 to 800 nanometers can be used. For example, incandescent lamps, mercury lamps and the like can be used.

The amount of the secondary amine (III) can be appropriately selected depending on the reaction rate and the like. Usually, it is used in an amount of 10 to 100 equivalents, preferably 20 to 50 equivalents, based on [60] fullerene. The reaction solvent is halogen or (C _1-
C ₆ ) Solvents capable of well dissolving fullerenes such as benzene (chlorobenzene, orthodichlorobenzene, toluene, benzene, etc.) which may be substituted with an alkyl group can be used. Preferably, chlorobenzene can be used. When the secondary amine is a solution, the secondary amine
A secondary amine can be used as a solvent.

The reaction temperature is not particularly limited, but the reaction can be carried out under cooling or under heating, preferably at 15 to 35 ° C. 3. Use Aspects of the Aminated Fullerene Derivative of the Present Invention The aminated fullerene derivative of the present invention is an amphiphilic compound and has excellent nucleic acid condensation ability. When the aminated fullerene derivative of the present invention is acted on a nucleic acid,
Based on its ability to bind to nucleic acids, the nucleic acid is bent and folded by the binding action between the duplexes in the molecule. Furthermore, a nucleic acid condensate is generated by the binding action between the molecules. This condensate formation is reversible with respect to the concentration of the aminated fullerene derivative, and the nucleic acid is regenerated by extracting and removing the aminated fullerene derivative.

The nucleic acid condensing ability of the aminated fullerene derivative depends on the synergistic effect of the size and hydrophobicity of the fullerene portion of the aminated fullerene derivative and the affinity for the phosphate group of the nitrogen-containing hydrophilic side chain. It is. The fullerene interacts with the hydrophobic portion of the nucleic acid (eg, the main groove of DNA) and the nitrogen-containing hydrophilic side chain interacts with the phosphate group of the nucleic acid, thereby folding the nucleic acid molecule. It is considered that such condensation occurs due to the aggregation of the hydrophobic portions of the nucleic acid molecules.

This means that a sample obtained by reacting the plasmid “pBR322” with the “tetraamine compound” described below at various concentrations was subjected to an agarose electrophoresis experiment (Short Pr.
otocols in Molecular Biology 3rd Ed., 1992, Wiley,
2-13) and morphological observation with an AFM microscope.

The ratio of the number of molecules of the aminated fullerene derivative to the nucleic acid may be within a range where a complete nucleic acid condensate can be formed.

Here, the formation of the nucleic acid condensate by the aminated fullerene derivative is carried out, for example, by mixing the two in a suitable buffer, but is not limited thereto, and is generally used in the art. It can be performed by the method described below. The nucleic acid condensate can also be isolated by treating the phase-transferred nucleic acid solution by a conventional method such as ethanol precipitation. Further, the original nucleic acid can be regenerated by extracting the aminated fullerene derivative from the nucleic acid solution condensed by the addition of the aminated fullerene derivative with an organic solvent such as chloroform.

As described above, the ability of the aminated fullerene derivative to condense nucleic acids is closely related to its high binding ability to nucleic acids. FIG. 3 shows the results of an experiment on the displacement of ethidium bromide into calf thymus DNA for the aminated fullerene derivative of the present invention.

Ethidium bromide on calf thymus DNA
Test test of compound aminated fullerene derivative (hereinafter referred to as “aminated fullerene”) obtained in Example 1 described below was subjected to a test. Test Method The DNA condensation ability of the aminated fullerene derivative was evaluated by a competitive binding experiment with ethidium bromide to calf thymus DNA. The method of this competitive binding experiment is described in Journal of Medicinal Chemistry, Volume 21, 6
Those described on pages 58-668 (1978) were used.

Test Results "Aminated fullerene" caused a 50% displacement of ethidium bromide at a concentration of 0.67 μM. This test result is a guideline for designing a molecule of an aminated fullerene derivative which is particularly preferable in practicing the present invention.
It is believed that an ability to bind to NA that is equal to or greater than the compounds described above is particularly desirable in practicing the present invention.

The transfection (cell transformation method) of the present invention using a nucleic acid condensate of an aminated fullerene derivative and a nucleic acid is performed by transforming cultured animal cells into a condensate of a nucleic acid and an aminated fullerene derivative in an appropriate culture solution. It is performed by adding and incubating. A method similar to the method using other chemical vectors can be used as a method for mixing the nucleic acid and the reagent, preparing a nucleic acid introduction reagent mixture, and introducing the mixture into cells. Especially the lipofection method (Cationic Lipos
ome, DNA Transfer to Cultured Cells, Ravid, K. and
Freshney, RI Eds., 1998, Chapter 11, pp 193-212
And Proceedings of National Academy of Sciences of United States
Of America, 84, 7413-7417 (19
1987)). That is, a nucleic acid-binding reagent is added to a nucleic acid solution and allowed to stand to prepare a nucleic acid-introducing reagent mixture. Next, the nucleic acid transfection reagent mixture is added to the growing cultured cells and cultured, whereby the nucleic acid complex is taken into the cells and the expression of the transgene is expressed. To calculate the transfection efficiency, a plasmid DNA having a reporter gene (generally, a reporter protein gene) that can easily measure the expression of a foreign gene is introduced into cells, and the expression level of the foreign gene in all cells is measured. Regarding the measurement of transfection efficiency,
Literature can be referred to (New Genetic Engineering Handbook, 3rd edition, Yodosha, 1999, etc.). Transfection usually has the highest expression of the foreign gene between one and several days.To measure this "transient" foreign gene expression efficiency, a GFP assay that can be easily performed without damaging the cells Is most preferred. When the exogenous gene is inserted into the chromosome in the cell and is permanently expressed for several weeks or more, it is called stable transfection. In general, in order to measure the efficiency of stable transfection, a test is particularly often performed using a drug resistance gene as a transgene.

Next, the present invention will be described specifically with reference to examples, but these examples do not limit the present invention.

Test Method for Transient Transfection Prepare 160 μL of serum-free Dulbecco's medium (4 μg) of plasmid DNA (pGreen Lantern, 5040 bp), add 4.5 μL of a 4.0 mM 0.01SHE solution of aminated fullerene, and vortex. did. After standing at room temperature for 30 minutes, the cells were suspended in 2 mL of serum-free Dulbecco's medium, and NIH 3T3 cells in logarithmic growth phase (1 × 10 ⁵ ce
lls / 35 mm dish). CO ₂ incubator (5% C
After incubating for 6 hours in O ₂ , 35 -C), remove the medium, add 2 mL of Dulbecco's medium containing 10% fetal bovine serum, and add CO ₂
Incubated for an additional 48 hours in the incubator.
The cells are observed with a fluorescence microscope and fluorescent protein (GFP)
Was observed. Test result: GFP expression was observed in 0.003% of the total cells. No expression was observed with only DNA added. This indicates that the aminated fullerene has a gene transfer ability to cultured cells.

Test Method for Stability Transfection A serum-free Dulbecco's medium solution (160 μL) containing 4 μg of plasmid DNA (pcDneo, 4500 bp) was prepared, and 4.5 μL of a 4.0 mM 0.01SHE solution of aminated fullerene was added and vortexed. After standing at room temperature for 30 minutes, 2 mL of serum-free Dulbecco's medium
And NIH 3T3 cells in logarithmic growth phase (1 × 10 ⁵ cells / 3
5 mm dish). CO ₂ incubator (5% CO ₂ , 35
After incubation in -C) for 6 hours, the medium was removed, 2 mL of Dulbecco's medium containing 10% fetal bovine serum was added, and the mixture was further incubated in a CO ₂ incubator for 48 hours. The cells were detached by trypsin treatment, and after counting, the cells were diluted with Dulbecco's medium containing 10% fetal bovine serum to obtain 1.0 × 10 ⁵ cells / dish
Was dispensed into 100 mm culture dishes so that After incubation for 24 hours, the medium was removed, and G418-containing selection medium (G418 600
8 g of microg / L, 10% fetal bovine serum) was added, and the mixture was incubated in a CO ₂ incubator for 3 weeks. The resulting colonies were counted, and the expression efficiency of the G418 resistance gene was evaluated.

Test Results Five colonies were formed, and the expression efficiency was 0.005%. No colonies were formed when only DNA was added. This indicated that aminated fullerenes could be used for permanent transformation.

The finding that the aminated fullerene derivative of the present invention not only has the ability to bind to nucleic acids but also has the ability to further condense multimolecular nucleic acids is completely new. Further, the finding that the aggregate of the generated aminated fullerene derivative and nucleic acid can be used for transfection (transformation of cells) is a completely new finding.

The nucleic acid in the present invention includes DNA, RN
A, PNA, synthetic nucleotides that hybridize with DNA or RNA, their modified ones, and mixtures or complexes of two or more thereof are shown. More specifically, genes are derived from living organisms or synthesized double-stranded DNs.
A: A single-stranded DNA, a double-stranded RNA, a single-stranded RNA, a synthetic DNA having a thioester bond, a synthetic RNA, or the like, hybridizes with a natural DNA or RNA, and in a physiological solution with the aminated fullerene derivative of the present invention. Any nucleic acid may be used as long as it forms an insoluble complex. In the embodiment of the present invention, "the use (for) the purpose of condensing nucleic acid" and the use form of the "nucleic acid condensing agent" utilize the ability of the aminated fullerene derivative of the present invention to condense nucleic acids. All forms of use are included, for example, use as a nucleic acid condensing reagent; use for introducing a vector into cells; DNA (or a derivative thereof) such as antisense DNA (or a derivative thereof) and decoy DNA (or a derivative thereof) ) Use for introduction of the fragment into cells; Use for control of gene expression based on binding to a promoter, enhancer region, etc .; Cell cycle by suppressing conversion of double-stranded DNA to single-stranded DNA Use for regulation; single-stranded DNA
For controlling the efficiency of PCR by controlling the melting point from DNA to double-stranded DNA or from double-stranded DNA to single-stranded DNA; and application to gene therapy. Is also possible.

When the aminated fullerene derivative is used in each of the above embodiments, the aminated fullerene derivative of the present invention or a salt thereof is used alone or in the form of a composition according to a conventional method in each usage mode. Achieve the intended purpose.

[0055]

EXAMPLES Example 1 Chlorobenzene (50 mg) of [60] fullerene (100 mg, 0.139 mmol) and N-methylpiperazine (488 μL, 4.44 mmol) was used.
mL) solution was stirred while irradiating with a 60 W incandescent light.
After stirring for 20 hours, the solvent and unreacted N-methylpiperazine were distilled off under reduced pressure, and the obtained brown solid was washed with hexane to obtain 155 mg (0.138 mmol, 99%) of compound (1).

[0056]

Embedded image

[0057]¹H NMR (400 MHz, CDCl_Three, δ): 2.38 (s,
6H, CH_Three), 2.42 (s, 6H, CH_Three), 2.1-2.5 (brm, 16H, NC
H_TwoCH_Two), 2.5-3.5 (br m, 16H, NCH_Two);¹³ C NMR (100 MHz, CDCl_Three, δ): 46.07 (CH_Three), 46.09
(CH_Three), 50.42 (C₆₀NCH_TwoCH_Two), 50.78 (C₆₀NCH_TwoCH_Two), 55.
65 (C₆₀NCH_Two), 55.76 (C₆₀NCH_Two), 71.53 (1C, C-O), 71.
83 (2C, C-N), 75.44 (2C, C-N), 76.31 (1C, C-O), 14
0.07 (2C, C₆₀), 141.48 (2C, C₆₀), 142.71 (2C, C₆₀),
 142.90 (2C, C₆₀), 143.30 (2C, C₆₀), 143.41 (2C, C
₆₀), 143.68 (2C, C₆₀), 143.87 (2C, C₆₀), 144.09 (2
C, C₆₀), 144.45 (2C, C₆₀), 144.93 (2C, C₆₀), 145.10
 (2C, C₆₀), 145.96 (2C, C₆₀), 146.12 (1C, C₆₀), 14
6.65 (2C, C₆₀), 146.71 (2C, C₆₀), 146.76 (2C, C₆₀),
 146.79 (2C, C₆₀), 146.9 (2C, C ₆₀), 147.22 (2C, C
₆₀), 147.50 (2C, C₆₀), 147.58 (1C, C₆₀), 148.88 (2
C, C ₆₀), 149.11 (2C, C₆₀), 149.14 (2C, C₆₀), 151.2
1 (2C, C₆₀). IR (neat, n / cm^-1) 3390 (m), 2937 (s), 2795 (s), 269
2 (w), 1651 (w), 1455 (s), 1375 (m), 1287 (s), 115
8 (w), 1111 (m), 1004 (s), 858 (w), 793 (w), 752
(s), 661 (w), 551 (w), 526 (w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 1133 (MH⁺).

Example 2 [60] Fullerene (20.0 mg, 0.0278 mmol) and piperidine
(88.0 μL, 0.890 mmol) in chlorobenzene (10 mL) is stirred while irradiating with a 60 W incandescent lamp. After stirring for 8 hours, the reaction solution is directly applied to a silica gel column (silica gel).
7 g), and developed with a mixed solvent of toluene / ethyl acetate (49/1) to obtain 16.0 mg of compound (2) (0.0147 mmol, 5
3%).

[0059]

Embedded image

Compound (2) is described in the literature (Schick, G .; Kamp
e, K, -D .; Hirsch, AJ Chem. Soc., Chem. Commun.
1995, 2023-2024).

Example 3 The following compound (3) was synthesized in the same manner as in Example 1.

[0062]

Embedded image

[0063]¹H NMR (400 MHz, CDCl_Three, d) 1.6-2.0 (br
s, 16H, OCH_Two), 3.2-3.5 (m, 16H, NCH_Two), 3.8-4.1 (m,
 16H, NCH _TwoCH_Two);¹³ C NMR (100 MHz, CDCl_{Three ,} d) 35.71 (4C, OCH_Two), 35.85
 (4C, OCH_Two), 48.80 (4C, NCH_TwoCH_Two), 49.44 (4C, NCH_TwoC
H_Two), 64.35 (4C, NCH_Two), 64.47 (4C, NCH_Two), 71.46 (1
(C, C-O), 71.80 (2C, C-N), 75.47 (2C, C-N), 76.45
(1C, C-O), 106.88 (2C, OCO), 107.04 (2C, OCO), 14
0.14 (2C, C₆₀), 141.41 (2C, C₆₀), 142.73 (2C,
C₆₀), 142.91 (2C, C₆₀), 142.94 (2C, C₆₀), 143.39
(2C, C₆₀), 143.51 (2C, C₆₀), 143.71 (2C, C₆₀), 14
3.86 (2C, C₆₀), 144.43 (1C, C₆₀), 144.58 (2C,
C₆₀), 145.03 (2C, C₆₀), 146.04 (2C, C₆₀), 146.19
(2C, C₆₀), 146.73 (2C, C₆₀), 146.76 (2C, C₆₀), 14
6.84 (2C, C₆₀), 146.98 (1C, C₆₀), 147.03 (2C,
C₆₀), 147.30 (2C, C₆₀), 147.31 (2C, C₆₀), 147.37
(2C, C₆₀), 147.57 (2C, C₆₀), 148.90 (2C, C₆₀), 14
9.16 (2C, C₆₀), 149.63 (1C, C₆₀), 149.74 (2C,
C₆₀), 151.62 (2C, C₆₀); IR (neat, n / cm^-1) 3413 (m), 2955 (s), 2883 (s), 166
8 (m), 1463 (w), 1364 (w), 1308 (w), 1231 (w), 114
6 (s), 1079 (s), 1038 (w), 946 (w), 914 (w), 849
(w), 754 (s), 666 (w), 545 (w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 1305 (MH⁺).

Example 4 The following compound (4) was synthesized in the same manner as in Example 2.

Embedded image

¹ H NMR (400 MHz, CDCl ₃ , d) 1.49 (overla
pped s, 18H, CH ₃ ), 1.50 (overlapped s, 18H, CH ₃ ),
3.1-3.2 (br m, 8H, NCH ₂ CH ₂ ), 3.2-3.3 (br m, 8H, NC
H ₂ CH ₂ ), 3.5-3.7 (br m, 16H, NCH ₂ ); ¹³ C NMR (100 MHz, CDCl ₃ , d) 28.54 (2C, CH ₃ ), 28.57
(2C, CH ₃ ), 50.41 (4C, C ₆₀ NCH ₂ CH ₂ ), 50.89 (4C, C ₆₀
NCH ₂ CH ₂ ), 52.90 (C ₆₀ NCH ₂ ), 63.08 (C ₆₀ NCH ₂ ), 71.71
(1C, CO), 71.83 (2C, CN), 75.56 (2C, CN), 76.37
(1C, CO), 137.70 ( 1C, C 60), 140.05 (2C, C 6 0), 14
1.65 (2C, C ₆₀ ), 142.89 (2C, C ₆₀ ), 143.12 (2C,
_{C 60), 143.43 (2C,} C 6 0), 143.52 (2C, C 60), 143.85
_{(2C, C 60), 143.94} (2C, C 60), 144.04 (2C, C 6 0), 14
_{4.38 (2C, C 60),} 144.86 (2C, C 60), 145.18 (1C,
_{C 60), 146.00 (2C,} C 6 0), 146.24 (2C, C 60), 146.75
_{(2C, C 60), 146.87} (4C, C 60), 146.92 (2C, C 6 0), 14
_{7.04 (2C, C 60),} 147.36 (2C, C 60), 147.64 (2C,
_{C 60), 148.95 (2C,} C 6 0), 149.07 (2C, C 60), 149.33
_{(4C, C 60), 151.12} (2C, C 60), 154.42 (2C, C 6 0), 15
4.56 (2C, C ₆₀ ), 154.62 (2C, C ₆₀ ); IR (neat, n / cm ^-1 ) 3006 (w), 2975 (m), 2929 (w), 285
6 (w), 2821 (w), 1698 (s), 1476 (w), 1453 (m), 142
1 (s), 1365 (m), 1300 (w), 1286 (m), 1252 (s), 117
1 (s), 1132 (s), 1001 (s), 862 (m), 755 (s), 699
(w), 665 (w), 581 (w), 547 (w), 536 (w), 505 (w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 1478 ( MH ⁺ ).

Example 5 The following compound (5) was synthesized in the same manner as in Example 2.

Embedded image

Compound (2) is described in the literature (Schick, G .; Kamp
e, K, -D .; Hirsch, AJ Chem. Soc., Chem. Commun.
1995, 2023-2024).

Example 6 To a solution of [60] fullerene (100 mg, 0.139 mmol) in chlorobenzene (50 mL) was added 32 equivalents of azetidine (300 μL, 4.45).
mmol) and the solution was stirred while irradiating with a 60 W incandescent light. After stirring for 1.5 hours, the reaction solution was directly loaded on a silica gel column (silica gel 35 g), and toluene,
The mixture was developed with 5%, 20%, and 50% ethyl acetate in a toluene solvent to give fullerene, compound (6) and compound (7) in 12 mg (12%), 21 mg (16%) and 21 mg (15%, respectively). )Obtained.

[0069]

Embedded image ¹ H NMR (400 MHz, CDCl ₃ , d) 2.1-2.3 (m, 8H, NCH ₂ CH ₂ ),
3.7-3.9 (m, 8H, NCH ₂ ), 3.9-4.1 (m, 8H, NCH ₂ ); ¹³ C NMR (100 MHz, CDCl ₃ , d) 17.57 (2C, NCH ₂ CH ₂ ), 17.
77 (2C, NCH ₂ CH ₂ ), 51.82 (4C, NCH ₂ ), 52.33 (4C, NCH
₂ ), 71.24 (2C, CN), 71.60 (1C, CO), 74.54 (2C, C
-N), 77.57 (1C, CO), 140.64 (2C, C ₆₀ ), 140.68 (2
C, C ₆₀ ), 143.10 (2C, C ₆₀ ), 143.51 (2C, C ₆₀ ), 143.5
_{3 (2C, C 60),} 143.59 (2C, C 60), 143.80 (2C, C 60), 1
_{44.23 (2C, C 60),} 144.37 (1C, C 60), 145.03 (2C,
C ₆₀ ), 145.05 (2C, C ₆₀ ), 145.30 (2C, C ₆₀ ), 146.02
_{(2C, C 60), 146.10} (2C, C 60), 146.55 (2C, C 60), 14
6.77 (1C, C ₆₀ ), 146.83 (2C, C ₆₀ ), 146.87 (2C,
C ₆₀ ), 147.12 (2C, C ₆₀ ), 147.15 (2C, C ₆₀ ), 147.35
_{(2C, C 60), 147.63} (4C, C 60), 148.61 (2C, C 60), 14
_{8.91 (2C, C 60),} 149.10 (2C, C 60), 149.12 (2C,
C ₆₀ ), 151.73 (2C, C ₆₀ ); IR (neat, n / cm ^-1 ) 2956 (m), 2850 (m), 1479 (w), 145
8 (w), 1162 (s), 984 (w), 852 (w), 755 (s), 555 (w),
545 (w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 961 (MH ⁺ ).

Embedded image

[0070]¹H NMR (400 MHz, CDCl_Three, d) 1.26 (s, 1H,
 OH), 2.1-2.3 (m, 8H, NCH_TwoCH_Two), 3.6-4.0 (m, 16H, N
CH_Two);¹³ C NMR (100 MHz, CDCl_Three, d) 17.21 (1C, NCH_TwoCH_Two), 1
7.75 (2C, NCH_TwoCH_Two), 17.91 (2C, NCH_TwoCH_Two), 51.85 (4
C, NCH_Two), 52.46 (4C, NCH_Two), 54.34 (2C, NCH_Two), 70.3
7 (2C, C-N), 71.77 (2C, C-N), 72.35 (1C, C-N), 82.5
5 (1C, C-O), 137.73 (2C, C₆₀), 141.27 (2C, C ₆₀), 1
42.44 (2C, C₆₀), 143.22 (2C, C₆₀), 143.32 (2C,
C₆₀), 143.50 (2C, C ₆₀), 143.58 (2C, C₆₀), 143.80
(2C, C₆₀), 143.98 (2C, C₆₀), 144.08 (2C, C ₆₀), 14
4.66 (2C, C₆₀), 145.48 (2C, C₆₀), 145.72 (2C,
C₆₀), 146.73 (1C, C ₆₀), 146.96 (2C, C₆₀), 147.02
(2C, C₆₀), 147.11 (2C, C₆₀), 147.18 (2C, C ₆₀), 14
7.69 (2C, C₆₀), 147.97 (1C, C₆₀), 148.21 (2C,
C₆₀), 148.30 (2C, C ₆₀), 148.43 (2C, C₆₀), 148.57
(2C, C₆₀), 148.65 (2C, C₆₀), 149.25 (2C, C ₆₀), 15
2.91 (2C, C₆₀), 153.23 (2C, C₆₀); IR (neat, n / cm^-1) 3410 (m), 2957 (s), 2924 (s), 285
2 (s), 1732 (m), 1636 (w), 1458 (m), 1263 (w), 123
5 (w), 1165 (m), 983 (w), 755 (s), 665 (w), 543
(w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 1018 (MH⁺).

Example 7 The following compound (8) was synthesized in the same manner as in Example 2.

Embedded image

¹ H NMR (400 MHz, CDCl ₃ , d) 1.90-1.96 (ov
erlapped t, J = 3.6 Hz, 16H, NCH ₂ CH ₂ ), 3.20 (shoul
dered s, 8H, NCH ₂ ), 3.33 (shouldered s, 8H, NCH ₂ ); ¹³ C NMR (100 MHz, CDCl ₃ , d) 23.85 (4C, NCH ₂ CH ₂ ), 2
3.95 (4C, NCH ₂ CH ₂ ), 50.12 (4C, NCH ₂ ), 50.02 (4C, N
CH ₂ ), 69.69 (2C, CN), 71.42 (1C, CO), 72.91 (2C,
CN), 77.89 (1C, CO ), 140.37 (2C, C 60), 140.93
_{(2C, C 60), 142.85} (2C, C 60), 143.23 (2C, C 60), 14
_{3.33 (2C, C 60),} 143. 67 (2C, C 60), 143.99 (2C,
C ₆₀ ), 144.85 (2C, C ₆₀ ), 144.95 (2C, C ₆₀ ), 145.06
_{(2C, C 60), 145.16} (2C, C 60), 146.07 (2C, C 60), 14
_{6.16 (2C, C 60),} 146.67 (2C, C 60), 146.77 (2C,
C ₆₀ ), 146.83 (2C, C ₆₀ ), 147.07 (2C, C ₆₀ ), 147.19
(2C, C ₆₀ ), 147.32 (2C, C ₆₀ ), 147.35 (2C, C ₆₀ ), 14
_{7.51 (2C, C 60),} 147.61 (2C, C 60), 148.84 (2C,
C ₆₀ ), 149.06 (2C, C ₆₀ ), 149.59 (2C, C ₆₀ ), 149.78
(2C, C ₆₀ ), 152.59 (2C, C ₆₀ ); IR (neat, n / cm ^-1 ) 3443 (m), 2957 (s), 2924 (s), 285
2 (s), 1739 (w), 1458 (m), 1212 (w), 1134 (m), 860
(m), 755 (s), 541 (w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 1017 (MH ⁺ ).

Example 8 A solution of [60] fullerene (100 mg, 0.139 mmol) in chlorobenzene (10 mL) was added to a 50% aqueous dimethylamine solution (467 μL,
4.44 mmol) and stirred while irradiating with a 60 W incandescent lamp. After stirring for 5 hours, the solvent, water, and unreacted dimethylamine were distilled off under reduced pressure, and the resulting brown solid was subjected to a silica gel column (silica gel 35 g, toluene / ethyl acetate (9 /
1) Develop the compound (9) 5
5.6 mg (0.0609 mmol, 44%) were obtained.

[0074]

Embedded image

¹ H NMR (400 MHz, CDCl ₃ , d) 2.87 (s, 12
H, CH ₃ ), 2.95 (s, 12H, CH ₃ ); ¹³ C NMR (100 MHz, CDCl ₃ , d) 42.78 (4C, CH ₃ ), 43.41
(4C, CH ₃ ), 71.52 (1C, CO), 72.18 (2C, CN), 75.63
(2C, CN), 76.54 ( 1C, CO) 140.38 (2C, C 60), 141.
_{36 (2C, C 60),} 142.75 (2C, C 60), 143.02 (2C, C 60), 1
_{43.35 (2C, C 60),} 143.37 (2C, C 60), 143.83 (2C,
C ₆₀ ), 143.93 (2C, C ₆₀ ), 144.27 (2C, C ₆₀ ), 144.67
_{(2C, C 60), 145.06} (2C, C 60), 145.25 (1C, C 60), 146.
_{05 (2C, C 60),} 146.24 (2C, C 60), 146.79 (2C, C 60), 1
_{46.81 (2C, C 60),} 146.89 (2C, C 60), 146.92 (2C,
C ₆₀ ), 147.09 (2C, C ₆₀ ), 147.15 (2C, C ₆₀ ), 147.35 (2
_{C, C 60), 147.63 (} 2C, C 60), 147.71 (1C, C 60), 149.00
_{(2C, C 60), 149.24} (2C, C 60), 149.92 (4C, C 60), 15
1.82 (2C, C ₆₀ ); IR (neat, n / cm ^-1 ) 3412 (s), 2922 (m), 1617 (w), 145
6 (w), 1102 (w), 616 (w); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 913 (MH ⁺ ).

Example 9 The following compound (10) was synthesized in the same manner as in Example 2.

Embedded image

¹ H NMR (400 MHz, CDCl ₃ , d) 0.8-1.0 (m,
20H, CH ₂ CH ₃ + CH ₂ CH ₃ ), 1.3-1.5 (m, 8H, NCH ₂ CH ₂ ), 1.6
-1.7 (m, 8H, NCH ₂ ), 2.80 (s, 6H, NCH ₃ ), 2.87 (s, 6
H, NCH ₃ ); ¹³ C NMR (100 MHz, CDCl ₃ , d) 14.37 (2C, CH ₃ CH ₂ ), 14.
45 (2C, CH ₃ CH ₂ ), 20.64 (2C, CH ₃ CH ₂ ), 20.77 (2C, CH
₃ CH ₂ ), 30.83 (2C, CH ₂ CH ₂ CH ₂ ), 31.00 (2C, CH ₂ CH ₂ C
H ₂ ), 38.70 (2C, CH ₂ N), 39.34 (2C, CH ₂ N), 53.93 (2C,
NCH ₃ ), 54.43 (2C, NCH ₃ ), 72.68 (1C, CO), 72.86
(2C, CN), 76.31 (2C, CN), 76.48 (1C, CO), 140.0
_{2 (2C, C 60),} 141.36 (2C, C 60), 142.57 (2C, C 60), 1
42.61 (2C, C ₆₀ ), 142.81 (2C, C ₆₀ ), 143.36 (2C,
C ₆₀ ), 143.66 (2C, C ₆₀ ), 143.74 (2C, C ₆₀ ), 144.80
_{(2C, C 60), 144.95} (1C, C 60), 145.25 (2C, C 60), 14
_{5.32 (2C, C 60),} 146.15 (2C, C 60), 146.26 (2C,
C ₆₀ ), 146.76 (2C, C ₆₀ ), 146.86 (2C, C ₆₀ ), 146.87
(2C, C ₆₀ ), 147.07 (4C, C ₆₀ ), 147.42 (2C, C ₆₀ ), 14
_{7.59 (2C, C 60),} 147.67 (1C, C 60), 147.99 (2C,
C ₆₀ ), 148.92 (2C, C ₆₀ ), 149.20 (2C, C ₆₀ ), 150.06
(2C, C ₆₀ ), 150.41 (2C, C ₆₀ ), 151.90 (2C, C ₆₀ ); IR (neat, n / cm ^-1 ) 2955 (s), 2928 (s), 2867 (m), 279
7 (w), 1725 (m), 1457 (s), 1419 (w), 1376 (w), 126
6 (w), 1243 (w), 1215 (m), 1183 (w), 1132 (w), 110
4 (w), 1064 (w), 1026 (m), 932 (w), 861 (m), 757
(s), 666 (w), 538 (m); MS m / z (LC-APCI, toluene: i-PrOH = 7: 3) 1081 (MH ⁺ ).

Example 10 To a solution of [60] fullerene (30 mg, 0.0416 mmol) in chlorobenzene (15 mL) was added 32 equivalents of N- (2-ethoxy-2-ethoxy-2-ethoxyethyl) piperazine (328 mg). , 1.33 mmol)
And add vigorous stirring while irradiating with a 60 W incandescent light. After stirring for 22 hours, the reaction solution was washed three times with water, and water and the solvent were distilled off under reduced pressure. The obtained composition was dissolved in a small amount of chloroform, and hexane was added to obtain a sticky black oil. Centrifuge at 5000 rpm for 10 minutes and mix the compound (1
64 mg (90% yield) of 1) was obtained.

¹ H NMR (400 MHz, CDCl ₃ , d) 1.1-1.3 (br
s, 12H, CH ₃ ), 2.6-2.8 (br s, 16H, C ₆₀ NCH ₂ CH ₂ ), 3.5
-3.8 (br m, 72H, C ₆₀ NCH ₂ , OCH ₂ ); ¹³ C NMR (100 MHz, CDCl ₃ , d) 15.18 (4C, CH ₃ ) 54.19
(4C, C ₆₀ NCH ₂ CH ₂ ), 54.32 (4C, C ₆₀ NCH ₂ CH ₂ ), 57.65 (4
C, C ₆₀ NCH ₂ ), 57.78 (4C, C ₆₀ NCH ₂ ), 66.52 (4C, CH ₂ CH
₃ ), 68.94 (2C, NCH ₂ ), 69.01 (2C, NCH ₂ ), 69.72 (4C,
OCH ₂ ), 70.33 (4C, OCH ₂ ), 70.51 (4C, OCH ₂ ), 70.58 (8
C, OCH ₂ ), 71.50 (1C, CO), 71.85 (2C, CN), 75.45
(2C, CN), 76.23 ( 1C, CO), 140.06 (2C, C 60), 141.
_{46 (2C, C 60),} 142.67 (2C, C 60), 142.84 (2C, C 60),
143.24 (2C, C ₆₀ ), 143.38 (2C, C ₆₀ ), 143.62 (2C, C
₆₀ ), 143.80 (2C, C ₆₀ ), 144.19 (2C, C ₆₀ ), 144.46 (2
C, C ₆₀ ), 144.91 (2C, C ₆₀ ), 145.07 (1C, C ₆₀ ), 145.9
_{2 (2C, C 60),} 146.10 (1C, C 60), 146.63 (2C, C 60), 1
_{46.68 (2C, C 60),} 146.75 (2C, C 60), 146.78 (2C,
C ₆₀ ), 146.87 (2C, C ₆₀ ), 147.20 (2C, C ₆₀ ), 147.46
_{(2C, C 60), 147.54} (1C, C 60), 148.83 (2C, C 60), 14
_{9.08 (2C, C 60),} 149.13 (2C, C 60), 149.27 (2C,
C ₆₀ ), 151.22 (2C, C ₆₀ ); IR (neat, n / cm ^-1 ) 3494 (w), 2869 (s), 1670 (m), 145
3 (m), 1373 (w), 1350 (w), 1301 (w), 1279 (w), 124
7 (w), 1114 (s), 1006 (m), 948 (w), 855 (w), 752
(m), 666 (w), 528 (w);

[0080]

The aminated fullerene derivative of the present invention is
It not only has the ability to bind to nucleic acids, but also has the ability to further condense multi-molecular nucleic acids. Furthermore,
The resulting aggregate of the aminated fullerene derivative and the nucleic acid could be used for transfection (cell transformation).

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) C07D 295/08 C07D 295/08 Z 4H006 493/10 493/10 A // C12N 15/09 C12N 15/00 A (72) Inventor Naoki Tomita 1-53-81 F-term (reference) 1-53-81 Sakuragaoka, Tama-shi, Tokyo 4B024 AA01 CA01 CA05 CA11 DA02 EA04 GA11 GA18 HA17 4C048 AA06 BB01 BC09 CC01 4C071 AA04 BB01 CC01 CC12 EE04 EE10 FF05 GG05 JJ06 LL01 4C084 AA13 MA05 NA13 ZC012 4C086 AA01 EA16 MA02 MA05 NA13 ZC01 4H006 AA01 BJ50 BN30

Claims (13)

[Claims] 1. The following general formula: [Wherein, R ¹ and R ² are each independently a hydrocarbon group which may have a substituent, and in addition, R ¹ and R ² have a substituent together with the nitrogen atom to which they are bonded. And a nitrogen-containing heterocyclic ring (optionally excluding morpholine and piperidine). Aminated fullerene derivatives represented by the formula: 2. The following general formula: [Wherein, R ¹ and R ² are each independently a hydrocarbon group which may have a substituent, and in addition, R ¹ and R ² have a substituent together with the nitrogen atom to which they are bonded. And a nitrogen-containing heterocyclic ring (optionally excluding morpholine and piperidine). ] The aminated fullerene derivative represented by these, and its salt. 3. The aminated fullerene derivative according to claim 1, wherein R ¹ and R ² are each independently a hydrocarbon group which may have a substituent. 4. The aminated fullerene derivative according to 3, wherein R ¹ and R ² are each independently an alkyl group, and a salt thereof. 5. The compound according to claim 1, wherein R ¹ and R ² are a nitrogen-containing heterocyclic ring selected from azetidine, pyrrolidine and piperazine, which may have a substituent together with the nitrogen atom to which they are bonded. 6. The substituent is a (C ₁ -C ₆ ) alkyl group,
(C ₁ -C ₆ ) alkylamino group, (C ₁ -C ₆ ) alkoxycarbonyl group, (C ₁ -C ₆ ) alkylenedioxy group or the following general formula: [Wherein A is a (C ₁ -C ₆ ) alkylene group, and R ³ is (C ₁ -C ₆ )
₆ ) an alkyl group, wherein n is an integer of 1 to 5].
A compound as described. 7. The following general formula: Wherein R ¹ and R ² are piperidine having a substituent together with the nitrogen atom to which they are bonded, or a salt thereof. 8. The following general formula:[Wherein X is N (CH_Three), N (tertiary butyroxy)
Bonyl), N-CH_TwoCH _Two-O-CH_TwoCH_Two-O-CH
_TwoCH_Two-OC_TwoH_FiveOr C (OCH_TwoCH_TwoO)
You. Or an aminated fullerene derivative represented by the formula:
salt. 9. [60] Fullerene and secondary amine (III): Are reacted under visible light irradiation to obtain an aminated fullerene derivative. [In the above formula, R ¹ and R ² are each independently a hydrocarbon group which may have a substituent. In addition, R ¹ and R ² are each a substituent together with a nitrogen atom to which they are bonded. A nitrogen-containing heterocyclic ring which may be possessed may be formed. ]. 10. Use of the aminated fullerene derivative or a salt thereof according to claim 9 for nucleic acid condensation. 11. A nucleic acid condensing agent comprising the aminated fullerene derivative according to claim 9 or a salt thereof. 12. A nucleic acid condensate comprising the aminated fullerene derivative according to claim 9 or a salt thereof and a nucleic acid. 13. A method for introducing a nucleic acid into a cell using the nucleic acid condensate according to claim 12.