KR101336232B1 - Poly(ethylene oxide)-based block copolymer bearing boronic acid group and iron oxide nanoparticle stabilized by the same - Google Patents

Abstract

The present invention is a polyethylene oxide block copolymer having a boronic acid group that is responsive to pH and glucose and can be used as a polymer material for drug delivery, a method for preparing the same, and a tumor cell imaging and a hyperthermia. It relates to iron oxide nanoparticles stabilized by the polyethylene oxide-based block copolymer that can be used.

Description

Poly (ethylene oxide) -based block copolymer bearing boronic acid group and iron oxide nanoparticle stabilized by the same}

The present invention relates to a polyethylene oxide block copolymer having boronic acid groups and to iron oxide nanoparticles stabilized by the polyethylene oxide block copolymer. More specifically, the present invention is a polyethylene oxide block copolymer having a boronic acid group that is responsive to pH and glucose and can be used as a polymer material for drug delivery, a method for preparing the same, and cancer cell imaging and heat treatment ( It relates to iron oxide nanoparticles stabilized by the polyethylene oxide-based block copolymer that can be used for hyperthermia.

Polyethylene oxide-based polymers have been studied a lot because of the enhanced permeability and retention effect into cells.

Methods of preparing polymeric conjugated materials prepared through polyethylene oxide terminal functionalization by various conventional methods and their applications are well described in the literature [JM Harris, RB Chess, Nature Reviews Drug Discovery, 2003, 2, pages 214-221 and S. Zalipsky, Bioconjugate Chemistry, 1995, Volume 6, pages 150-165]. In this regard, the synthesis of polyethylene oxides prepared by living anionic polymerization is well described in the literature. Slomkowski, A. Duda, "Anionic ring-opening polymerization", in Ring-Opening Polymerization: Mechanism, Catalysis, Structure, Utility; Edited by DJ Brunelle, 1993, Chapter 3, pages 87-128 and RP Quirk, J. Kim, "Macromonomers and Macromonomers", in Ring-Opening Polymerization: Mechanism, Catalysis, Structure, Utility; Edited by DJ Brunelle, 1993, Chapter 9, pages 263-293].

In particular, the control of the reaction conditions is very complicated and difficult for molecular weight distribution and molecular weight control depending on the type of alkali metal of the alkoxide initiator used in the production of polyethylene oxide by living polymerization. In view of this, the living polymerization method of ethylene oxide using lithium and potassium-based alkoxides as initiators is well described in the patent literature [Korean Patent No. 122,055].

Methods for preparing block copolymers of poly (ethylene glycol) with various polymeric materials are well described in the literature [Jankova, K .; Chen, X .; Kops, J .; Batsberg, W. Macromolecules, 1998, Vol. 31, pages 538-541 and Topp, MDC; Dijkstra, PJ; Talsma, H .; Feijen, J. Macromolecules, 1997, Vol. 30, pages 8518-8520.

However, there is still a need to develop a new polyethylene oxide block copolymer that can be used as a drug delivery polymer material, is responsive to external stimuli such as pH, glucose and temperature, and can exhibit anticancer and contrast effects.

It is an object of the present invention to provide a polyethylene oxide block copolymer which is responsive to pH and glucose and can be used as a polymer material for drug delivery.

Another object of the present invention is to provide an efficient method for producing the polyethylene oxide block copolymer.

Another object of the present invention is to provide iron oxide nanoparticles stabilized by the polyethylene oxide-based block copolymers which can be used for cancer cell imaging and hyperthermia.

Still another object of the present invention is to provide a method for producing iron oxide nanoparticles stabilized by the polyethylene oxide block copolymer.

The present invention relates to a polyethylene oxide block copolymer represented by the following formula (1), (2) or (3).

[Formula 1]

(2)

(3)

Where

R is n -butyl group, sec -butyl group, tert -butyl group or tert -butoxy group,

R ₁ and R ₂ are each independently hydrogen or a methyl group,

m is an integer from 5 to 50,

n is an integer from 10 to 500,

k is an integer of 1-5.

Polyethylene oxide-based block copolymers according to the present invention

(a) obtaining polyethylene oxide by living anionic polymerization;

(b) obtaining a chain-end functionalized polyethylene oxide (compound of formula (4) wherein X is chlorine, bromine or iodine) from the polyethylene oxide;

(c) obtaining a reversible addition fragmentation transfer (RAFT) material (compound of formula 4 wherein X is xanthate) using the chain-end functionalized polyethylene oxide;

(d) 3- (acrylamido) phenylboronic acid, or 3- (meth acrylamido) phenylboronic acid or radical copolymerization, N, using the RAFT material-vinylimidazole or N-vinyl pyrrolidone and the radical Copolymerizing followed by radical copolymerization with 3- (acrylamido) phenylboronic acid or 3- (methacrylamido) phenylboronic acid to obtain a block copolymer; And

(e) denaturing the chain end of the block copolymer with azobisisobutyronitrile (AIBN).

[Chemical Formula 4]

Where

R is n -butyl group, sec -butyl group, tert -butyl group or tert -butoxy group,

R ₁ is hydrogen or a methyl group,

X is a chlorine, bromine, iodine or xanthate group,

n is an integer from 10 to 500.

When the polyethylene oxide is prepared by living anionic polymerization in the step (a), the molecular weight of the polyethylene oxide can be controlled. J .; Choi, S .; Kim, KM; Go, DH; Jeon, HJ; Lee, JY; Park, HS; Lee, CH; Park, HM Macromolecular Research, 2007, Vol. 15, pages 337-342].

The number average molecular weight of the polyethylene oxide is preferably from 500 to 100,000, more preferably from 1,000 to 10,000.

The method for preparing the chain-end functionalized polyethylene oxide of step (b), the method for preparing the RAFT material of step (c), the radical copolymerization method of step (d) and the chain terminal modification method of step (e) are described in the literature. It is described in detail in Kim, J .; Choi, S .; Kim, KM; Go, DH; Jeon, HJ; Lee, JY; Park, HS; Lee, CH; Park, HM Macromolecular Research, 2007, Vol. 15, pages 337-342].

1 is a radical copolymerization with N -vinylimidazole and 3- (methacrylamido) phenylboronic acid by sequential monomer addition using a polyethylene oxide-based RAFT material, and the chain ends are modified with AIBN. A process diagram for preparing the polyethylene oxide block copolymer represented by the formula (2) according to the present invention is shown.

Polyethylene oxide-based block copolymers according to the present invention can be usefully used as a stabilizer of iron oxide nanoparticles.

Accordingly, the present invention relates to iron oxide nanoparticles stabilized with the polyethylene oxide block copolymer according to the present invention.

Iron oxide nanoparticles stabilized with the polyethylene oxide-based block copolymer may be water-soluble and have a size of 1 to 80 nm.

Iron oxide nanoparticles stabilized with a polyethylene oxide block copolymer according to the present invention may be prepared by adding and reducing iron chloride to the polyethylene oxide block copolymer according to the present invention.

The reaction may be carried out in an aqueous solution as well as an organic solvent, the reaction temperature is preferably 5 ^o C to 70 ^o C, more preferably 10 ^o C to 50 ^o C.

The concentration of the iron chloride is preferably 0.01 to 10 g / 10 mL, NH ₄ OH, N ₂ H ₂ , NaBH ₄ , H ₂ S, Na ₂ S and the like may be used as a reducing agent, but is not limited thereto.

The polyethylene oxide-based block copolymer according to the present invention is a smart polymer material having dual response to pH and glucose, and can be used for cancer cell imaging and thermotherapy by stabilizing iron oxide nanoparticles.

In addition, since the polyethylene oxide-based block copolymer according to the present invention is water-soluble, iron oxide nanoparticles can be prepared in an aqueous solution, and easy to purify and easy to handle.

1 is a manufacturing process chart according to an embodiment of a polyethylene oxide-based block copolymer represented by the formula (2).

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are for illustrative purpose only and that the scope of the present invention is not limited to these embodiments.

≪ Example 1 >

Under high vacuum, ampoules of the various reactants were attached with a hand torch in a 1 L round flask flask and then attached to a vacuum line to remove air completely. Injecting n -butyllithium (2.4 mmol) under a argon stream with a syringe and using a dry ice / isopropanol bath instead of a water bath to reduce the reactor temperature to -78 ^o C and then evacuating the argon gas in the reactor Completely removed by pump. Again, 300 mL of purified benzene was distilled off and injected into the reactor. The temperature of the reactor was slowly raised to room temperature to completely dissolve the benzene in the reactor and change to a solution containing an initiator. Purified 30 mL (26.5 g) of ethylene oxide (EO) (30 vol%, dilute solution) was injected into the reactor by breaking the breakseal with careful attention at 0 ^o C using an ice bath. . After about 1 hour, t- BuOK (2.4 mmol in THF 20 mL) and 50 mL of purified DMSO were introduced into the reactor through a brake chamber and a stopcock. The temperature of the reactor was raised to 35 ^° C. using a water bath and stirred for 5 hours. Again using the ice water bath to 5 ^o C to proceed with the polymerization for 10 minutes, this process was repeated several times. The reaction was again performed at room temperature for 48 hours, aliquots were taken, and the solvent was removed using Rotavap ^™ . The residue was dissolved in THF again and precipitated in diethyl ether to obtain polyethylene oxide (poly (ethylene oxide): PEO). The number average molecular weight of the obtained polymer was 5,500 g / mol. The conversion of EO to a polymer was 100 mol%.

<Example 2>

To 200 mL ([POMt] = 6.3 mmol) of the alkali metal (Mt) polymer alkoxide solution obtained in Example 1, 2-bromopropionyl bromide (20 mmol in 20 mL THF) was added thereto at room temperature. The reaction was stirred for 24 hours while stirring. After the reaction was completed, the solvent was removed using Rotavap ^™ , and then dissolved in THF and precipitated in diethyl ether to obtain a pale yellow powder. The polymer thus prepared had a number average molecular weight of 5,600 g / mol by gel permeation chromatography (GPC), and a chain terminal bromination yield by ¹ H NMR spectrum analysis was 98 mol% or more.

<Example 3>

To 200 mL ([POMt] = 6.3 mmol) of the alkali metal (Mt) polymer alkoxide solution obtained in Example 1, 2-bromoisobutyryl bromide (20 mmol in 20 mL THF) was added and stirred at room temperature. The reaction was carried out for a time. After the reaction was completed, the solvent was removed using Rotavap ^™ , and then dissolved in THF and precipitated in diethyl ether to obtain a pale yellow powder. The polymer thus prepared had a number average molecular weight of 5,600 g / mol by GPC, and a chain terminal bromination yield by ¹ H NMR spectrum analysis was 98 mol% or more.

<Example 4>

5 g (0.09 mmol) of the polymer obtained in Example 2 were dissolved in 100 mL of THF purified in a 500 mL round flask under high vacuum, and 0.3 g of potassium ethyl xathantogenate (96%). Was dissolved in 30 mL THF and injected into the reactor and reacted for 24 hours under a 45 ^° C. nitrogen stream. After the reaction, the solution was precipitated in an excess of dimethyl ether to obtain a powder. Results based on PEO was analyzed by ¹ H NMR reversible addition-minute power save the (reversible addition fragmentation transfer: RAFT) number average molecular weight of the material was 5,700 g / mol, GPC results were also 5,700 g / moL.

<Example 5>

5 g (0.09 mmol) of the polymer obtained in Example 3 were dissolved in 100 mL of THF purified in a 500 mL round flask under high vacuum, and 0.3 g of potassium ethyl xathantogenate (96%). Was dissolved in 30 mL THF and injected into the reactor and reacted for 24 hours under a 45 ^° C. nitrogen stream. After the reaction, the solution was precipitated in an excess of diethyl ether to obtain a powder. Results based on PEO was analyzed by ¹ H NMR reversible addition-minute power save the (reversible addition fragmentation transfer: RAFT) number average molecular weight of the material was 5,700 g / mol, GPC results were also 5,700 g / moL.

<Example 6>

0.001 mol (5.7 g) of the PEO-RAFT material obtained in Example 4 was placed in a 250 mL round flask reactor under nitrogen stream and dissolved in 100 mL dimethylformamide (DMF), followed by 0.0002 mol of AIBN radical initiator. Injected into. The temperature is raised to 90 ^° C., and 0.01 mol (1.91 g; FW = 190.8) of 3- (acrylamido) phenylboronic acid is dissolved in 25 mL DMF and injected under a nitrogen stream with a syringe. After the injection into the reactor it was polymerized for 24 hours. The reaction was terminated by lowering the reactor temperature to room temperature, and again precipitated in excess dimethyl ether to obtain about 6.8 g of a light yellow powdery polymer. After drying in a vacuum oven, it was dissolved in 50 mL DMF again, excess AIBN was added, and the end of the chain was denatured to obtain 6.5 g of white powder. The number average molecular weight of the result was 7,300 g / moL by GPC.

&Lt; Example 7 >

0.001 mol (5.7 g) of the PEO-RAFT material obtained in Example 4 was placed in a 250 mL round flask reactor under nitrogen stream and dissolved in 100 mL dimethylformamide (DMF), followed by 0.0002 mol of AIBN radical initiator. Injected into. The temperature is raised to 90 ^° C., 0.01 mol (2.04 g; FW = 204.8) of 3- (methacrylamido) phenylboronic acid is dissolved in 25 mL DMF, and the nitrogen stream Under polymerization in the reactor for 24 hours. The reaction was terminated by lowering the reactor temperature to room temperature, and again precipitated in excess dimethyl ether to obtain about 7.6 g of a light yellow powdery polymer. After drying in a vacuum oven, it was again dissolved in 50 mL DMF, excess AIBN was added, and the end of the chain was denatured to obtain 7.5 g of a white powder. The number average molecular weight of the result was 7,500 g / moL by GPC.

&Lt; Example 8 >

0.001 mol (5.7 g) of the PEO-RAFT material obtained in Example 4 was placed in a 250 mL round flask reactor under nitrogen stream and dissolved in 100 mL dimethylformamide (DMF), followed by 0.0002 mol of AIBN radical initiator. Injected into. And its temperature is raised to 110 ^o C, N - vinylimidazole (N -vinylimidazole; FW = 94.12, d = 1.039) was mixed with 6.7 mL of 25 mL DMF is injected into the reactor under the flow of nitrogen for 24 hours with a syringe Polymerized. Again lower the temperature of the reactor to 90 ^° C. and dissolve 0.003 mole (0.61 g; FW = 204.8) of 3- (methacrylamido) phenylboronic acid in 25 mL DMF with a syringe. It was polymerized for 24 hours after being injected into the reactor under a nitrogen stream. The reaction was terminated by lowering the reactor temperature to room temperature, and again precipitated in excess dimethyl ether to obtain about 13.5 g of a light yellow powdery polymer. After drying in a vacuum oven it was again dissolved in 100 mL DMF, excess AIBN was added and the chain ends were denatured to obtain 13.4 g of white powder. The number average molecular weight of the result was 12,900 g / moL by ¹ H-NMR and GPC.

&Lt; Example 9 >

0.001 mol (5.7 g) of PEO-RAFT material obtained in Example 4 was placed in a 250 mL round flask reactor under a stream of nitrogen and dissolved in 100 mL dimethylformamide (DMF), followed by 0.0002 mol of AIBN radical initiator. Injected into. And its temperature is raised to 110 ^o C, N - vinylimidazole (N -vinylimidazole; FW = 94.12, d = 1.039) was mixed with 6.7 mL of 25 mL DMF is injected into the reactor under the flow of nitrogen for 24 hours with a syringe Polymerized. Again lower the temperature of the reactor to 90 ^o C, dissolve 0.0032 mole (0.61 g; FW = 190.8) of 3- (acrylamido) phenylboronic acid in 25 mL DMF and inject with nitrogen After the injection into the reactor under air flow it was polymerized for 24 hours. The reaction was terminated by lowering the reactor temperature to room temperature, and again precipitated in excess dimethyl ether to obtain about 13.5 g of a light yellow powdery polymer. After drying in a vacuum oven it was again dissolved in 100 mL DMF, excess AIBN was added and the chain ends were denatured to obtain 13.4 g of a white powder. The number average molecular weight of the resultant was 13,000 g / moL, respectively, by ¹ H-NMR spectrum analysis and GPC analysis.

&Lt; Example 10 >

0.001 mol (5.7 g) of the PEO-RAFT material obtained in Example 4 was placed in a 250 mL round flask reactor under nitrogen stream and dissolved in 100 mL dimethylformamide (DMF), followed by 0.0002 mol of AIBN radical initiator. Injected into. And its temperature is raised to 110 ^o C, N - vinylpyrrolidone (N -vinylpyrrolidone; FW = 111.14, d = 1.040) was mixed with 4.8 mL of 25 mL DMF is injected into the reactor under the flow of nitrogen for 24 hours with a syringe Polymerized. Again lower the temperature of the reactor to 90 ^o C, dissolve 0.0032 mole (0.61 g; FW = 190.8) of 3- (acrylamido) phenylboronic acid in 25 mL DMF and inject with nitrogen After the injection into the reactor under air flow it was polymerized for 24 hours. The reaction temperature was terminated by lowering the reactor temperature to room temperature, and again precipitated in excess dimethyl ether to obtain about 11.3 g of a white powdery polymer. After drying in a vacuum oven, it was dissolved in 100 mL DMF again, excess AIBN was added and the chain ends were denatured to obtain 11.2 g of a white powder. The number average molecular weight of the resultant was 11,000 g / moL, respectively, by ¹ H-NMR spectrum analysis and GPC analysis.

<Example 11>

0.001 mol (5.7 g) of the PEO-RAFT material obtained in Example 4 was placed in a 250 mL round flask reactor under nitrogen stream and dissolved in 100 mL dimethylformamide (DMF), followed by 0.0002 mol of AIBN radical initiator. Injected into. And its temperature is raised to 110 ^o C, N - vinylpyrrolidone (N -vinylpyrrolidone; FW = 111,14, d = 1.040) was mixed with 4.8 mL of 25 mL DMF is injected into the reactor under the flow of nitrogen into the syringe 24 Polymerized for time. Again lower the temperature of the reactor to 90 ^o C, dissolve 0.003 mol (0.61 g; FW = 204.8) of 3- (methacrylamido) phenylboronic acid in 25 mL DMF and use a syringe. It was polymerized for 24 hours after being injected into the reactor under a nitrogen stream. The reaction was terminated by lowering the reactor temperature to room temperature, and again precipitated in excess dimethyl ether to obtain about 13.5 g of a light yellow powdery polymer. After drying in a vacuum oven it was again dissolved in 100 mL DMF, excess AIBN was added and the chain ends were denatured to obtain 13.4 g of white powder. The number average molecular weight of the result was 11,100 g / moL by ¹ H-NMR and GPC.

&Lt; Example 12 >

2.0 g (0.28 mmol) of the block copolymer ((PEO b -PAPBA); MW = 7,300 g / moL) obtained in Example 6 was completely dissolved in 40 mL of distilled water in a 100 mL round flask under nitrogen stream. Next, 0.03 g of FeCl ₃ (ferric chloride; FW = 162.21) and 0.07 g of FeCl ₂ · 4H ₂ O (ferrous chloride; FW = 198.81) were dissolved in 10 mL of distilled water, and then a block copolymer solution was prepared using a syringe under a nitrogen stream. It was injected into the flask and allowed to stand for about 2 to 3 hours while stirring slowly. At this time, the color of the solution in the reactor was brown, again 1.7 mL of NH ₄ OH (25% aqueous solution) was injected using a syringe as a reducing agent and stirred for about 6 hours, and then lowered to room temperature and reacted while stirring. The brown insoluble portion was removed by filtration and the solution portion was removed using Rotavap ^™ , and then dissolved in THF and precipitated in dimethyl ether to give a dark brown powder. The size of the particle size by scanning electron microscopy ranged from 5 to 20 nanometers.

&Lt; Example 13 >

In the same manner as in Example 12, iron oxide nanoparticles as a dark brown powder using 2.0 g (0.28 mmoL) of the block copolymer ((PEO- b -PMAPBA); MW = 7,500 g / moL) obtained in Example 7 Was prepared. The size of the particle size by scanning electron microscopy ranged from 5 to 20 nanometers.

&Lt; Example 14 >

In the same manner as in Example 12, dark brown using the triple block copolymer ((PEO- b -PVIm- b -PMAPBA) obtained in Example 8; MW = 12,900 g / moL) 2.0 g (0.16 mmoL) Iron oxide nanoparticles were prepared as a powder. The size of the particle size by scanning electron microscopy ranged from 5 to 20 nanometers.

&Lt; Example 15 >

In the same manner as in Example 12, the triple block copolymer (PEO- b -PVIm- b -PAPBA) obtained in Example 9; MW = 13,000 g / moL) 2.0 g (0.15 mmoL) was used to prepare iron oxide nanoparticles, a dark brown powder. The size of the particle size by scanning electron microscopy ranged from 5 to 20 nanometers.

&Lt; Example 16 >

In the same manner as in Example 12, the triple block copolymer (PEO- b -PVP- b -PAPBA) obtained in Example 10; MW = 11,000 g / moL) 2.0 g (0.18 mmoL) was used to prepare iron oxide nanoparticles, a dark brown powder. The size of the particle size by scanning electron microscopy ranged from 5 to 20 nanometers.

&Lt; Example 17 >

In the same manner as in Example 12, the triple block copolymer (PEO- b -PVP- b -PMAPBA) obtained in Example 11; MW = 11,100 g / moL) 2.0 g (0.18 mmoL) was used to prepare iron oxide nanoparticles, a dark brown powder. The size of the particle size by scanning electron microscopy ranged from 5 to 20 nanometers.

Claims (10)

The polyethylene oxide block copolymer represented by the following general formula (1), (2) or (3):
[Chemical Formula 1]

(2)

(3)

In this formula,
R is n -butyl group, sec -butyl group, tert -butyl group or tert -butoxy group,
R ₁ and R ₂ are each independently hydrogen or a methyl group,
m is an integer from 5 to 50,
n is an integer from 10 to 500,
k is an integer of 1-5. (a) obtaining polyethylene oxide by living anionic polymerization;
(b) obtaining a chain-end functionalized polyethylene oxide represented by Formula 4 from the polyethylene oxide;
(c) obtaining a reversible addition fragmentation transfer (RAFT) material using the chain-end functionalized polyethylene oxide;
(d) 3- (acrylamido) phenylboronic acid, or 3- (meth acrylamido) phenylboronic acid or radical copolymerization, N, using the RAFT material-vinylimidazole or N-vinyl pyrrolidone and the radical Copolymerizing followed by radical copolymerization with 3- (acrylamido) phenylboronic acid or 3- (methacrylamido) phenylboronic acid to obtain a block copolymer; And
(E) a method for producing a polyethylene oxide block copolymer according to claim 1 comprising the step of modifying the chain end of the block copolymer with azobisisobutyronitrile (AIBN):
[Chemical Formula 4]

In this formula,
R is n -butyl group, sec -butyl group, tert -butyl group or tert -butoxy group,
R ₁ is hydrogen or a methyl group,
X is chlorine, bromine or iodine,
n is an integer from 10 to 500.  The method according to claim 2, wherein the number average molecular weight of the polyethylene oxide obtained in step (a) is 1000 to 10,000. delete The method of claim 2, wherein the reversible addition fragmentation transfer (RAFT) material is a compound represented by the following formula (4):
[Chemical Formula 4]

In this formula,
R is n -butyl group, sec -butyl group, tert -butyl group or tert -butoxy group,
R ₁ is hydrogen or a methyl group,
X is a xanthate group,
n is an integer from 10 to 500. Iron oxide nanoparticles stabilized with a polyethylene oxide-based block copolymer according to claim 1. The iron oxide nanoparticles of claim 6, wherein the iron oxide nanoparticles have a size of 1 to 80 nm. A method for producing iron oxide nanoparticles stabilized with a polyethylene oxide block copolymer comprising the step of adding and reducing iron chloride to the polyethylene oxide block copolymer according to claim 1. The process according to claim 8, wherein the reaction is carried out in an aqueous solution. The method of claim 8, wherein the reducing agent is NH ₄ OH, N ₂ H ₂ , NaBH ₄ , H ₂ S or Na ₂ S. 10.

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