Thermosensitive phosphazene-based polymer comprising sulfate moiety, and preparation method and use thereof

Abstract

Provided is a thermosensitive phosphazene-based polymer including an amino acid ester moiety, a polyethyleneglycol moiety, and a moiety including a sulfate group linked directly or by a linker in a predetermined ratio, a method of preparing the same, and a hydrogel-formable composition including the same. For example, a hydrogel formed from the composition may be used for tissue regeneration or drug delivery or used as a storage, a body tissue regeneration inducer, or a filler in a body.

Claims

1. A thermosensitive phosphazene-based polymer comprising: a first moiety of an amino acid ester represented by Formula 2; a second moiety of polyethyleneglycol represented by Formula 3; and a third moiety including a sulfate group; wherein the first moiety, the second moiety, and the third moiety are linked by an amino nitrogen to a phosphorous atom of a polyphosphazene backbone represented by Formula 1; and wherein the mole percent range of the first moiety, the second moiety, and the third moiety present in the phosphazene-based polymer are represented by a, b, and c, respectively, ##STR00006## wherein a is 55 mol % to 75 mol %, b is 5 mol % to 30 mol %, and c is 0.5 mol % to 20 mol %, and in Formulae 1, 2, and 3, R.sub.1 is C.sub.1-6 alkyl, C.sub.1-6 alkenyl, or C.sub.6-10 aryl-C.sub.1-6 alkyl; R.sub.2 is hydrogen, methyl, isopropyl, 1-methylpropyl, 2-methylpropyl, thiomethyl, methylthioethyl, benzyl, hydroxybenzyl, or 2-indolylmethyl; R.sub.3 is C.sub.1-6 alkyl; n is an integer of 3 to 100,000; and p is an integer of 1 to 20.

2. The thermosensitive phosphazene-based polymer according to claim 1, wherein R.sub.1 is methyl, ethyl, propyl, butyl, benzyl, or 2-prophenyl; and R.sub.3 is methyl.

3. The thermosensitive phosphazene-based polymer according to claim 1, further comprising a fourth moiety including a functional moiety for introducing a functional group into an end of the polymer.

4. The thermosensitive phosphazene-based polymer according to claim 3, further comprising a fourth moiety including at least one functional substance linked directly or by a linker to a part of or the entire functional group of the fourth moiety, wherein the functional substance; is capable of regulating a degradation rate of the polymer, a is capable of cross-linking, is capable of inducing tissue adhesion, a physiologically active substance, or a composite material formed by linear connection of two or more substances thereof.

5. The thermosensitive phosphazene-based polymer according to claim 1, wherein the phosphazene-based polymer including a sulfate group is represented by a formula of poly[(isoleucineethylester).sub.a′(aminomethoxypolyethyleneglycol 750).sub.b′(aminoethylsulfate).sub.c′phosphazene].sub.n′: wherein in the formula, a′ is 1.1 to 1.5; b′ is 0.1 to 0.6; c′ is 0.01 to 0.4, 1.6≤a′+b′+c′≤2, and n′ is an integer of 3 to 100,000.

6. A method of preparing the phosphazene-based polymer including a sulfate group according to claim 1, the method comprising: a first step of reacting polydichlorophosphazene represented by Formula 4 with an amino acid ester represented by Formula 5; a second step of further reacting the reaction mixture obtained from the first step by adding a C.sub.1-6 aminoalkanol or a hydrogen sulfate compound including an amine group at one end to the reaction mixture; a third step of further reacting the reaction mixture obtained from the second step by adding aminopolyethyleneglycol to the reaction mixture; and a fourth step of reacting a product obtained from the third step with a sulfur trioxide or a composite thereof: ##STR00007## wherein in Formulae 4 and 5, R.sub.1, R.sub.2 and n are as defined in claim 1, wherein the second step and the third step are able to be performed in a reverse order, and the fourth step is omitted when the hydrogen sulfate compound including an amine group at one end is added in the second step.

7. The method according to claim 6, wherein the sulfur trioxide is used in the form of a composite with a tertiary amine-based compound in the fourth step.

8. A hydrogel-forming composition comprising a thermosensitive phosphazene-based polymer, the phosphazene-based polymer comprising: a first moiety of an amino acid ester represented by Formula 2; a second moiety of polyethyleneglycol represented by Formula 3; and a third moiety including a sulfate group; wherein the first moiety, the second moiety, and the third moiety are linked by an amino nitrogen to a phosphorous atom of a polyphosphazene backbone represented by Formula 1; and wherein the mole percent of the first moiety, the second moiety, and the third moiety present in the phosphazene-based polymer are represented by a, b, and c, respectively, ##STR00008## wherein a is 55 mol % to 75 mol %, b is 5 mol % to 30 mol %, and c is 0.5 mol % to 20 mol %, and in Formulae 1, 2, and 3, R.sub.1 is C.sub.1-6 alkyl, C.sub.1-6 alkenyl, or C.sub.6-10 aryl-C.sub.1-6 alkyl; R.sub.2 is hydrogen, methyl, isopropyl, 1-methylpropyl, 2-methylpropyl, thiomethyl, methylthioethyl, benzyl, hydroxybenzyl, or 2-indolylmethyl; R.sub.3 is C.sub.1-6 alkyl; n is an integer of 3 to 100,000; and p is an integer of 1 to 20.

9. The hydrogel-forming composition according to claim 8, wherein the hydrogel-forming composition is converted from a sol state to a gel state by body temperature to form a hydrogel in the body.

10. The hydrogel-forming composition according to claim 9, further comprising a substance that is present in the hydrogel in the body.

11. The hydrogel-forming composition according to claim 10, wherein the hydrogel formed in the body releases the substance to the body.

12. The hydrogel-forming composition according to claim 9, wherein the hydrogel in the body absorbs water, an inorganic ion, a vitamin, a hormone, or a growth factor.

13. The hydrogel-forming composition according to claim 8, wherein the phosphazene-based polymer further comprises a carboxyl group.

14. The hydrogel-forming composition according to claim 8, wherein the hydrogel assists tissue regeneration.

15. The hydrogel-forming composition according to claim 8, wherein the hydrogel serves as a body tissue prosthesis.

16. A hydrogel formed from the hydrogel-forming composition according to claim 8.

17. The hydrogel according to claim 16, wherein the hydrogel is used as a filter to control passage of molecules or particles in a body.

18. The hydrogel according to claim 16, wherein water, an inorganic ion, a vitamin, or a hormone is stored in a gel.

19. The thermosensitive phosphazene-based polymer according to claim 1, wherein a is 60.5 mol % to 69 mol %, b is 19 mol % to 28.5 mol %, and c is 2.5 mol % to 15 mol %.

Description

BRIEF DESCRIPTION OF DRAWINGS

(1) FIG. 1 shows photographs of a solution of a sulfate group-containing phosphazene-based polymer prepared in Example 3 dissolved in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt % obtained by visual observation of a sol-gel behavior thereof.

(2) FIGS. 2 and 3 are graphs showing viscosities of solutions of sulfate group-containing phosphazene-based polymers (Examples 3 and 11) according to the present invention dissolved in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt % measured with respect to temperature, respectively.

(3) FIG. 4 is a graph illustrating surface charges of a hydroxy group-containing phosphazene-based polymer (Comparative Example 1) as a matrix polymer, a phosphazene-based polymer substituted with a carboxyl group (Comparative Example 4), and a phosphazene-based polymer substituted with a sulfate group (Example 3).

(4) FIG. 5 is a graph illustrating changes in surface charges of phosphazene-based polymers (Examples 1, 2, and 3) according to a substitution degree of the sulfate group.

(5) FIG. 6 shows photographs and a graph illustrating an in vivo degradation rate of a hydrogel of a sulfate group-containing phosphazene-based polymer prepared according to Example 3.

(6) FIG. 7 is a graph illustrating comparison results of cytotoxicity between a sulfate group-containing polyphosphazene-based polymer of Example 11 and a precursor polymer immediately before introducing the sulfate group thereinto.

(7) FIG. 8 is a graph illustrating changes in viscosity of an aqueous solution including a phosphazene-based polymer according to Example 1 and triamcinolone acetonide with respect to temperature.

(8) FIG. 9 is a graph illustrating changes in viscosity of an aqueous solution including a phosphazene-based polymer according to Example 11 and a vascular endothelial growth factor (VEGF) with respect to temperature.

(9) FIG. 10 is a graph illustrating release behaviors of a vascular endothelial growth factor (VEGF) from phosphazene-based polymer hydrogels according to Example 11 and Comparative Example 1 with time.

(10) FIG. 11 is a graph illustrating release behaviors of a stromal cell-derived factor from phosphazene-based polymer hydrogels according to Example 11 and Comparative Examples 1 and 4 with time.

(11) FIGS. 12 to 16 shows comparison results of tissue regeneration effects among hydrogels formed of phosphazene-based polymers according to Example 11 and Comparative Examples 1 and 4 alone, hydrogels formed of each of the polymers and a growth factor, hydrogels formed of each of the polymers and a stem cell, hydrogels formed of the polymer and both, natural regeneration, a growth factor alone without hydrogel, or a stem cell alone without hydrogel.

BEST MODE

(12) Hereinafter, the present invention will be described in more detail with reference to the following examples. However, these examples are for illustrative purposes only, and the present disclosure is not intended to be limited by these examples.

(13) <Identification of Compounds>

(14) In the examples below, element analysis of carbon, hydrogen, and nitrogen was carried out using Perkin-Elmer's C, H, and N analyzers in the Advanced Analysis Center of the Korea Institute of Science and Technology (KIST) in order to identify synthesized polymers. In addition, the nuclear magnetic resonance spectrum with hydrogen and phosphorus was measured by a Varian Gemini-300, and the weight average molecular weight Mw was measured by gel permeation chromatography using a Waters 1515 pump and a 2410 differentiation refractometer.

Example 1

Preparation of Poly[(isoleucineethylester).SUB.1.38.(aminomethoxypolyethyleneglycol 750).SUB.0.57.(aminoethylsulfate).SUB.0.05.phosphazene].SUB.n

(15) Dry isoleucine ethyl ester hydrochloride (IleOEtHCl, 11.65 g, 59.53 mmol) was dissolved in 200 mL of anhydrous tetrahydrofuran (THF) including 35 mL of triethylamine. A solution prepared by dissolving polydichlorophosphazene (5 g, 43.14 mmol) in 50 mL of anhydrous THF was added dropwise to the mixed solution in an acetone-dry ice bath, and then the temperature was gradually raised to 40° C. to 50° C. and reaction was performed for 24 hours. After cooling the reaction mixture to room temperature, dry aminoethanol (0.94 g, 15.53 mmol) was dissolved 100 mL of in anhydrous THF and 5 mL of triethylamine was added to the reaction mixture, and then reaction was performed at a temperature of 40° C. to 50° C. for 24 hours.

(16) After cooling the reaction mixture to room temperature, a solution prepared by dissolving dry polyethyleneglycol (6.47 g, 8.63 mmol) having a molecular weight of 750 in 200 mL of anhydrous THF and adding 5 mL of triethylamine thereto was added to the reaction mixture, and reaction was performed for 24 hours at a temperature of 40° C. to 50° C.

(17) The solution in which the reaction was completed was filtered in order to remove produced triethylamine hydrochloride, and the reaction filtrate was concentrated under reduced pressure until a small amount of the solvent remained. The concentrated solution was dissolved in 10 mL of anhydrous THF, and then an excess of hexane was added thereto to induce precipitation. After repeating this process twice or three times, the precipitates were dissolved in a small amount of methanol, placed in MWCO 12000 membrane (Spectrum Laboratories, Inc.), dialyzed against methanol at room temperature for 4 days, dialyzed against distilled water for 4 days, and dried at a low temperature to obtain a polyphosphazene polymer [NP(IleOEt).sub.1.38(AMPEG750).sub.0.57(Aminoethanol).sub.0.05].sub.n (7.21 g) including isoleucineethylester, aminomethoxypolyethyleneglycol, and aminoethanol.

(18) The polyphosphazene polymer [NP(IleOEt).sub.1.38(AMPEG750).sub.0.57(Aminoethanol).sub.0.05].sub.n obtained in the previous step was dissolved in 200 mL of anhydrous THF and 200 mL of dimethylformamide, and a solution prepared by dissolving sulfur trioxide pyridine complex (4.97 g, 31.25 mmol) in 200 mL of dimethylformamide was added to the reaction solution, and reaction was performed at a temperature of 25° C. to 40° C. for 24 hours. The reaction solution was filtered and the filtrate was concentrated under reduced pressure until a small amount of the solvent remained. The concentrated solution was placed in MWCO 1200 membrane, dialyzed against methanol for 4 days, dialyzed against distilled water for 4 days, and dried at a low temperature to obtain a final product [NP(IleOEt).sub.1.38(AMPEG750).sub.0.57(Aminoethylsulfate(AES)).sub.0.05].sub.n.

(19) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(20) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(21) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(22) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(23) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(24) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(25) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(26) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(27) Average molecular weight (M.sub.w): 2,200

Example 2

Preparation of Poly[(isoleucineethylester).SUB.1.35.(aminomethoxypolyethyleneglycol 750).SUB.0.5.(aminoethylsulfate).SUB.0.15.phosphazene].SUB.n

(28) A final product [NP(IleOEt).sub.1.35(AMPEG750).sub.0.5(AES).sub.0.15].sub.n was obtained in the same manner as in Example 1, except that the amounts of isoleucine ethyl ester hydrochloride (10.97 g, 56.08 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (12.94 g, 17.26 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (3.78 g, 23.74 mmol), dimethylformamide (total 400 mL), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(29) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(30) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(31) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(32) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(33) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(34) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(35) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(36) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(37) Average molecular weight (M.sub.w): 1900

Example 3

Preparation of Poly[(isoleucineethylester).SUB.1.32.(aminomethoxypolyethyleneglycol 750).SUB.0.38.(aminoethanol).SUB.0.1.(aminoethylsulfate).SUB.0.2.phosphazenel].SUB.n

(38) A final product [NP(IleOEt).sub.1.3(AMPEG750).sub.0.4(Aminoethanopal).sub.0.1(AES).sub.0.3].sub.n was obtained in the same manner as in Example 1, except that the amounts of isoleucine ethyl ester hydrochloride (11.15 g, 56.98 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (12.30 g, 16.4 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (3.86 g, 24.24 mmol), dimethylformamide (total 400 mL), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(39) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(40) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(41) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(42) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(43) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(44) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(45) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(46) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(47) Average molecular weight (M.sub.w): 2600

Example 4

Preparation of Poly[(isoleucineethylester).SUB.1.23.(aminomethoxypolyethyleneglycol 550).SUB.0.57.(aminoethylsulfate).SUB.0.2.phosphazene].SUB.n

(48) A final product [NP(IleOEt).sub.1.23(AMPEG550).sub.0.57(AES).sub.0.2].sub.n was obtained in the same manner as in Example 1, except that polyethyleneglycol having a molecular weight of 550 was used instead of polyethyleneglycol having a molecular weight of 750, and the amounts of isoleucine ethyl ester hydrochloride (10.38 g, 53.07 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.53 g, 8.63 mmol), polyethyleneglycol (13.53 g, 24.6 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (3.8 g, 23.9 mmol), dimethylformamide (total 400 mL), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(49) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(50) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(51) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(52) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(53) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(54) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(55) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(56) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(57) Average molecular weight (M.sub.w): 4400

Example 5

Preparation of Poly[(isoleucineethylester).SUB.1.21.(aminomethoxypolyethyleneglycol 550).SUB.0.51.(aminoethylsulfate).SUB.0.28.phosphazene].SUB.n

(58) A final product [NP(IleOEt).sub.1.21(AMPEG550).sub.0.51(AES).sub.0.28].sub.n was obtained in the same manner as in Example 4, except that the amounts of isoleucine ethyl ester hydrochloride (10.22 g, 52.22 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.74 g, 12.08 mmol), polyethyleneglycol (12.1 g, 22 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (4.03 g, 25.29 mmol), dimethylformamide (total 400 mL), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(59) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(60) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(61) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(62) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(63) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(64) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(65) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(66) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(67) Average molecular weight (M.sub.w): 2500

Example 6

Preparation of Poly[(isoleucineethylester).SUB.1.25.(aminomethoxypolyethyleneglycol 550).SUB.0.45.(aminoethanol).SUB.0.12.(aminoethylsulfate).SUB.0.18.phosphazene].SUB.n

(68) A final product [NP(IleOEt).sub.1.25(AMPEG750).sub.0.45(Aminoethanol).sub.0.12(AES).sub.0.18].sub.n was obtained in the same manner as in Example 4, except that the amounts of isoleucine ethyl ester hydrochloride (10.55 g, 53.93 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (10.68 g, 19.41 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (4.29 g, 26.94 mmol), dimethylformamide (total 400 mL), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(69) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(70) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(71) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(72) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(73) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(74) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(75) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(76) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(77) Average molecular weight (M.sub.w): 3600

Example 7

Preparation of Poly[(isoleucineethylester).SUB.1.38.(aminomethoxypolyethyleneglycol 750).SUB.0.4.(ethyl-2-(O-glycyl)lactate).SUB.0.02.(aminoethylsulfate).SUB.0.2.phosphazene].SUB.n

(78) Dry isoleucine ethyl ester hydrochloride (11.65 g, 59.54 mmol) was dissolved in 200 mL of anhydrous THF including 35 mL of triethylamine. A solution prepared by dissolving polydichlorophosphazene (5 g, 43.14 mmol) in 100 mL of anhydrous THF was added dropwise to the mixed solution in acetone-dry ice bath, and then the temperature was gradually raised to 40° C. to 50° C. and reaction was performed for 24 hours. After cooling the reaction mixture to room temperature, a solution prepared by dissolving dry ethyl-2-(O-glycyl)lactate ammonium oxalate (0.93 g, 4.3 mmol) in 100 mL of anhydrous acetonitrile to which 5 mL of triethylamine was added was gradually added dropwise to the reaction mixture, and then reaction was performed at room temperature for 8 hours.

(79) After cooling the reaction mixture to room temperature, a solution prepared by dissolving dry polyethyleneglycol (12.94 g, 17.26 mmol) having a molecular weight of 750 in 100 mL of anhydrous THF and adding 10 mL of triethylamine thereto was added to the reaction mixture and reaction was performed at a temperature of 40° C. to 50° C. for 24 hours.

(80) The solution in which the reaction was completed was filtered in order to remove produced triethylamine hydrochloride, and the reaction filtrate was concentrated under reduced pressure until a small amount of the solvent remained. The concentrated solution was dissolved in 10 mL of anhydrous THF, and then an excess of hexane was added thereto to induce precipitation. After repeating this process twice or three times, the precipitates were dissolved in a small amount of methanol, placed in MWCO 12000 membrane, dialyzed against methanol at room temperature for 4 days, dialyzed against distilled water for 4 days, and dried at a low temperature to obtain a polyphosphazene polymer [NP(IleOEt).sub.1.38(AMPEG750).sub.0.4(GlyLacOEt).sub.0.02(Aminoethanol).sub.0.2].sub.n including isoleucineethylester, aminomethoxypolyethyleneglycol, ethyl-2-(O-glycyl)lactate, and aminoethanol.

(81) The polyphosphazene polymer [NP(IleOEt).sub.1.38(AMPEG750).sub.0.4(GlyLacOEt).sub.0.02(Aminoethanol).sub.0.2].sub.n obtained in the previous step was dissolved in 200 mL of anhydrous THF and 200 mL of dimethylformamide, and a solution prepared by dissolving sulfur trioxide pyridine complex (3.73 g, 23.46 mmol) in 200 mL of dimethylformamide was added to the reaction solution, and reaction was performed at a temperature of 25° C. to 40° C. for 24 hours. The reaction solution was filtered and the filtrate was concentrated under reduced pressure until a small amount of the solvent remained. The concentrated solution was placed in MWCO 1200 membrane, dialyzed against methanol for 4 days, dialyzed against distilled water for 4 days, and dried at a low temperature to obtain a final product [NP(IleOEt).sub.1.38(AMPEG750).sub.0.4(GlyLacOEt).sub.0.02(AES).sub.0.2].sub.n.

(82) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(83) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(84) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(85) δ 1.3-1.5(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(86) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(87) δ 1.6-1.7(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(88) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(89) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(90) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(91) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(92) δ 4.0-4.4(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(93) δ 5.2-5.4(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(94) Average molecular weight (M.sub.w): 33000

Example 8

Preparation of Poly[(isoleucineethylester).SUB.1.23.(aminomethoxypolyethyleneglycol 550).SUB.0.43.(ethyl-2-(O-glycyl)lactate).SUB.0.04.(aminoethylsulfate).SUB.0.3.phosphazene].SUB.n

(95) A final product [NP(IleOEt).sub.1.23(AMPEG550).sub.0.43(GlyLacOEt).sub.0.04(AES).sub.0.3].sub.n was obtained in the same manner as in Example 7, except that polyethyleneglycol having a molecular weight of 550 was used instead of polyethyleneglycol having a molecular weight of 750, and the amounts of isoleucine ethyl ester hydrochloride (10.38 g, 53.07 mmol), polydichlorophosphazene (5 g, 43.14 mmol), ethyl-2-(O-glycyl)lactate ammonium oxalate 1.86 g, 12.6 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (10.2 g, 18.55 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (4.5 g, 28.27 mmol), dimethylformamide (total 400 mL), anhydrous acetonitrile (100 mL), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(96) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(97) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(98) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(99) δ 1.3-1.5(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(100) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(101) δ 1.6-1.7(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(102) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(103) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(104) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(105) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(106) δ 4.0-4.4(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(107) δ 5.2-5.4(b, —NHCH.sub.2COOCH(CH.sub.3)COOCH.sub.2CH.sub.3),

(108) Average molecular weight (M.sub.w): 34000

Example 9

Preparation of Poly[(isoleucineethylester).SUB.1.38.(aminomethoxypolyethyleneglycol 750).SUB.0.32.(aminoethylsulfate).SUB.0.15.(aminoethylsuccinate).SUB.0.15.phosphazene].SUB.n

(109) Dry isoleucine ethyl ester hydrochloride (11.65 g, 59.53 mmol) was dissolved in 200 mL of anhydrous THF including 35 mL of triethylamine. A solution prepared by dissolving polydichlorophosphazene (5 g, 43.14 mmol) in 100 mL of anhydrous THF was added dropwise to the mixed solution in acetone-dry ice bath, and then the temperature was gradually raised to 40° C. to 50° C. and reaction was performed for 24 hours. After cooling the reaction mixture to room temperature, a solution prepared by dissolving dry aminoethanol (0.94 g, 15.53 mmol) in 100 mL of anhydrous THF and adding 5 mL of triethylamine thereto was added to the reaction mixture, and then reaction was performed at a temperature of 40° C. to 50° C. for 24 hours.

(110) After cooling the reaction mixture to room temperature, a solution prepared by dissolving dry polyethyleneglycol (8.41 g, 11.21 mmol) having a molecular weight of 750 in 200 mL of anhydrous THF and adding 5 mL of triethylamine was added to the reaction mixture and reaction was performed at a temperature of 40° C. to 50° C. for 24 hours.

(111) The solution in which the reaction was completed was filtered in order to remove produced triethylamine hydrochloride, and the reaction filtrate was concentrated under reduced pressure until a small amount of the solvent remained. The concentrated solution was dissolved in 10 mL of anhydrous THF, and then an excess of hexane was added thereto to induce precipitation. After repeating this process twice or three times, the precipitates were dissolved in a small amount of methanol, placed in MWCO 12000 membrane, dialyzed against methanol at room temperature for 4 days, dialyzed against distilled water for 4 days, and dried at a low temperature to obtain a polyphosphazene polymer [NP(IleOEt).sub.1.38(AMPEG750).sub.0.32(Aminoethanol).sub.0.3].sub.n including isoleucineethylester, aminomethoxypolyethyleneglycol, and aminoethanol.

(112) The polyphosphazene polymer [NP(IleOEt).sub.1.38(AMPEG750).sub.0.32(Aminoethanol).sub.0.3].sub.n obtained in the previous step was dissolved in 200 mL of anhydrous THF and 200 mL of dimethylformamide, and a solution prepared by dissolving 1 equivalent of sulfur trioxide pyridine complex (1.4 g, 8.78 mmol) in 200 mL of dimethylformamide was added to the reaction solution, and reaction was performed at a temperature of 25° C. to 40° C. for 24 hours. After cooling the reaction solution to room temperature, 2 equivalents of anhydrous succinic acid (1.76 g, 17.57 mmol) and 2 equivalents of dimethylaminopyridine (2.15 g, 17.57 mmol) were added to the reaction mixture, and then reaction was further performed at a temperature of 25° C. to 40° C. for 24 hours. The reaction solution was filtered and the filtrate was concentrated under reduced pressure until a small amount of the solvent remained, and the concentrated solution was placed in MWCO 1200 membrane, dialyzed against methanol for 4 days, dialyzed against distilled water for 4 days, and dried at a low temperature to obtain a final product [NP(IleOEt).sub.1.38(AMPEG750).sub.0.32(AES).sub.0.15(Aminoethylsuccinate).sub.0.15].sub.n.

(113) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(114) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(115) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(116) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(117) δ 2.5-2.7(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(118) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(119) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(120) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(121) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(122) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(123) Average molecular weight (M.sub.w): 2200

Example 10

Preparation of Poly[(isoleucineethylester).SUB.1.22.(aminomethoxypolyethyleneglycol 550).SUB.0.5.(aminoethylsulfate).SUB.0.1.(aminoethylsuccinate).SUB.0.18.phosphazene].SUB.n

(124) A final product [NP(IleOEt).sub.1.22(AMPEG750).sub.0.5(AES).sub.0.1(Aminoethylsuccinate).sub.0.18].sub.n was obtained in the same manner as in Example 9, except that polyethyleneglycol having a molecular weight of 550 was used instead of polyethyleneglycol having a molecular weight of 750, and the amounts of isoleucine ethyl ester hydrochloride (10.30 g, 52.64 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (11.86 g, 21.58 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (1.38 g, 8.67 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous succinic acid (1.72 g, 17.18 mmol), 2 equivalents of dimethylaminopyridine (2.1 g, 17.18 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(125) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(126) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(127) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(128) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(129) δ 2.5-2.7(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(130) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(131) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(132) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(133) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(134) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(135) Average molecular weight (M.sub.w): 3500

Example 11

Preparation of Poly[(isoleucineethylester).SUB.1.39.(aminomethoxypolyethyleneglycol 750).SUB.0.31.(aminoethylsulfate).SUB.0.13.(aminoethylglutarate).SUB.0.17.phosphazene].SUB.n

(136) A final product [NP(IleOEt).sub.1.39(AMPEG750).sub.0.31(AES).sub.0.13(AminoethylGlutarate).sub.0.17].sub.n was obtained in the same manner as in Example 9, except that anhydrous glutaric acid was used instead of anhydrous succinic acid, and the amounts of isoleucine ethyl ester hydrochloride (11.74 g, 59.98 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (10.03 g, 13.37 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (1.42 g, 8.91 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous glutaric acid (3.65 g, 32.01 mmol), 2 equivalents of dimethylaminopyridine (3.91 g, 32.01 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(137) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(138) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(139) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(140) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(141) δ 2.1-2.32(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(142) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(143) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(144) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(145) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(146) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(147) Average molecular weight (M.sub.w): 4500

Example 12

Preparation of Poly[(isoleucineethylester).SUB.1.21.(aminomethoxypolyethyleneglycol 550).SUB.0.49.(aminoethylsulfate).SUB.0.2.(aminoethylglutarate).SUB.0.1.phosphazene].SUB.n

(148) A final product [NP(IleOEt).sub.1.21(AMPEG550).sub.0.49(AES).sub.0.2(AminoethylGlutarate).sub.0.1].sub.n was obtained in the same manner as in Example 11, except that polyethyleneglycol having a molecular weight of 550 was used instead of polyethyleneglycol having a molecular weight of 750, and the amounts of isoleucine ethyl ester hydrochloride (10.22 g, 52.2 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (11.63 g, 21.14 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (1.12 g, 11.14 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous glutaric acid (3.59 g, 31.5 mmol), 2 equivalents of dimethylaminopyridine (3.85 g, 31.49 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(149) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(150) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(151) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(152) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(153) δ 2.1-2.32(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(154) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(155) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(156) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(157) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(158) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(159) Average molecular weight (M.sub.w): 4600

Example 13

Preparation of Poly[(isoleucineethylester).SUB.1.36.(aminomethoxypolyethyleneglycol 750).SUB.0.28.(aminoethylsulfate).SUB.0.21.(aminoethyladipate).SUB.0.15.phosphazene].SUB.n

(160) A final product [NP(IleOEt).sub.1.36(AMPEG750).sub.0.28(AES).sub.0.21(AminoethylAdipate).sub.0.15].sub.n was obtained in the same manner as in Example 9, except that anhydrous adipic acid was used instead of anhydrous succinic acid, and the amounts of isoleucine ethyl ester hydrochloride (11.48 g, 59.67 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (9.06 g, 12.08 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (1.48 g, 9.32 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous adipic acid (2.63 g, 20.54 mmol), 2 equivalents of dimethylaminopyridine (2.51 g, 20.54 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(161) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(162) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(163) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(164) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(165) δ 1.52-1.64(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2CH.sub.2COOH),

(166) δ 2.3-2.32(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2CH.sub.2COOH),

(167) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(168) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(169) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(170) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(171) Average molecular weight (M.sub.w): 3700

Example 14

Preparation of Poly[(isoleucineethylester).SUB.1.21.(aminomethoxypolyethyleneglycol 550).SUB.0.49.(aminoethylsulfate).SUB.0.12.(aminoethyladipate).SUB.0.18.phosphazene].SUB.n

(172) A final product [NP(IleOEt).sub.1.21(AMPEG550).sub.0.49(AES).sub.0.12(AminoethylAdipate).sub.0.18].sub.n was obtained in the same manner as in Example 13, except that polyethyleneglycol having a molecular weight of 550 was used instead of polyethyleneglycol having a molecular weight of 750, and the amounts of isoleucine ethyl ester hydrochloride (10.22 g, 52.2 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (11.63 g, 21.14 mmol) having a molecular weight of 550, sulfur trioxide pyridine complex (1.35 g, 11.02 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous adipic acid (3.14 g, 24.52 mmol), 2 equivalents of dimethylaminopyridine (3 g, 24.52 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were adjusted.

(173) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(174) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(175) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(176) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(177) δ 1.52-1.64(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2CH.sub.2COOH),

(178) δ 2.3-2.32(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2CH.sub.2COOH),

(179) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(180) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(181) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(182) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(183) Average molecular weight (M.sub.w): 2900

Example 15

Preparation of Poly[(isoleucineethylester).SUB.1.4.(aminomethoxypolyethyleneglycol 750).SUB.0.3.(aminoethylsulfate).SUB.0.16.(aminoethylsuccinatepolyethyleneimine).SUB.0.14.phosphazene].SUB.n

(184) [NP(IleOEt).sub.1.4(AMPEG750).sub.0.3(AES).sub.0.16(Aminoethylsuccinate).sub.0.14].sub.n was obtained in the same manner as in Example 9 using isoleucine ethyl ester hydrochloride (11.82 g, 60.4 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (9.71 g, 12.94 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (1.43 g, 8.98 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous succinic acid (1.8 g, 18 mmol), 2 equivalents of dimethylaminopyridine (2.2 g, 18 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL).

(185) The obtained [NP(IleOEt).sub.1.4(AMPEG750).sub.0.3(AES).sub.0.16(Aminoethylsuccinate).sub.0.14].sub.n was dissolved in 200 mL of chloroform, and isobutylchloroformate (0.25 g) and triethylamine (5 mL) were added thereto, followed by activation for 40 minutes. Then, polyethyleneimine (PEI, 16.2 g, 9 mmol) having a molecular weight of 1800 dissolved in chloroform was added to the solution and reaction was performed. After 18 hours, the reaction mixture was concentrated under reduced pressure, precipitated using a KF solution, dialyzed against distilled water at 4° C. for 3 days, and dried at a low temperature to obtain a final product [NP(IleOEt).sub.1.4(AMPEG750).sub.0.3(AES).sub.0.16(AminoethylsuccinatePEI).sub.0.14].sub.n.

(186) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(187) δ 0.7-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(188) δ 1.1-1.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(189) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-PEI),

(190) δ 2.5-2.7(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-PEI),

(191) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4,

(192) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(193) δ 3.4-3.8(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3), —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-PEI),

(194) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(195) Average molecular weight (M.sub.w): 3400

Example 16

Preparation of Poly[(isoleucineethylester).SUB.1.41.(aminomethoxypolyethyleneglycol 750).SUB.0.4.(aminoethylsulfate).SUB.0.12.(aminoethylsuccinateimidazole).SUB.0.07.phosphazene].SUB.n

(196) [NP(IleOEt).sub.1.41(AMPEG750).sub.0.4(AES).sub.0.12(Aminoethylsuccinate).sub.0.07].sub.n was obtained in the same manner as in Example 9 using isoleucine ethyl ester hydrochloride (11.9 g, 60.83 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.5 g, 8.2 mmol), polyethyleneglycol (12.94 g, 17.26 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (1.24 g, 7.82 mmol), dimethylformamide (total 400 mL), 2 equivalents of anhydrous succinic acid (1.57 g, 15.73 mmol), 2 equivalents of dimethylaminopyridine (1.92 g, 15.73 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL).

(197) The obtained [NP(IleOEt).sub.1.41(AMPEG750).sub.0.4(AES).sub.0.12(Aminoethylsuccinate).sub.0.07].sub.n was dissolved in 400 mL of THF. Each of the solutions respectively prepared by dissolving 10 equivalents of diisopropylcarbodiimide (4.38 g) and 10 equivalents of n-hydroxysuccinimide (4 g) in 50 mL of THF was added to the polymer solution, followed by activation for 40 minutes. Then, a solution prepared by dissolving 5 equivalents of 1-(3-aminopropylimidazole) (2.18 g) in THF was added thereto, reaction was performed in an ice bath for 5 hours and then at room temperature for hours to obtain a final product [NP(IleOEt).sub.1.41(AMPEG750).sub.0.4(AES).sub.0.12(Aminoethylsuccinatelmidazole).sub.0.7].sub.n.

(198) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(199) δ 0.7-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(200) δ 1.1-1.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(201) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(202) δ 2.5-2.7(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-Imi),

(203) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4),

(204) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(205) δ 3.4-3.8(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3), —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-Imi)

(206) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3)

(207) δ 6.8-7.8(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-Imi)

(208) Average molecular weight (M.sub.w): 4500

Example 17

Preparation of Poly[(isoleucineethylester).SUB.1.49.(aminomethoxypolyethyleneglycol 750).SUB.0.34.(aminoethylsulfate).SUB.0.10.(aminoethylsuccinatepolypeptide).SUB.0.07.phosphazene].SUB.n

(209) [NP(IleOEt).sub.1.49(AMPEG750).sub.0.34(AES).sub.0.1(Aminoethylsuccinate).sub.0.07].sub.n was obtained in the same manner as in Example 9 using isoleucine ethyl ester hydrochloride (12.58 g, 64.28 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.26 g, 4.31 mmol), polyethyleneglycol (11 g, 14.67 mmol) having a molecular weight of 750, sulfur trioxide pyridine complex (1.32 g, 8.3 mmol), dimethylformamide (total 400 mL), anhydrous succinic acid (0.83 g, 8.3 mmol), dimethylaminopyridine (1.02 g, 8.3 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL).

(210) A solution of 5 equivalents of hexylamine (0.29 g, 2.84 mmol) and 1 equivalent of polypeptide (amino acid sequence: CRRRRHHHHHHGGGGGRGDS, 1.29 g, 0.57 mmol) was added to a flask in which 5 g of the polymer obtained as described above was dissolved in 400 mL of dimethyl sulfoxide, and reaction was performed at room temperature for 24 hours. Then, the resultant was dialyzed against distilled water at 4° C. for 3 days, and dried at a low temperature to obtain a final product [NP(IleOEt).sub.1.49(AMPEG750).sub.0.34(AES).sub.0.1(AminoethylsuccinateCRRRRHHHHHHGGG GGRGDS).sub.0.07].sub.n.

(211) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl3, ppm):

(212) δ 0.7-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(213) δ 1.1-1.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(214) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-PEI),

(215) δ 2.5-2.7(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-PEI),

(216) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2SO.sub.4),

(217) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(218) δ 3.4-3.8(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3), —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CONH-PEI),

(219) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(220) δ 8.73(b, CHNCHNHCH, imidazole),

(221) Average molecular weight (Mw): 1840

Example 18

Preparation of Poly[(isoleucineethylester).SUB.1.40.(aminomethoxypolyethyleneglycol 750).SUB.0.30.(aminoethylmethacrylateethylsulfate).SUB.0.16.(aminoethylmethacrylate).SUB.0.14.phosphazene].SUB.n

(222) Dry isoleucine ethyl ester hydrochloride (9.79 g, 50.04 mmol) was dissolved in 500 mL of anhydrous tetrahydrofuran including 30 mL of triethylamine. A solution prepared by dissolving poly(dichlorophosphazene) (5.00 g, 43.14 mmol) in 100 mL of tetrahydrofuran was added dropwise to the mixed solution in an acetone-dry ice bath, and then the temperature was gradually raised to 40° C. to 50° C. and reaction was performed for 24 hours. After cooling the reaction mixture to room temperature, a solution prepared by dissolving dry aminomethacrylate hydrochloride (3.58 g, 21.57 mmol) in 100 mL of dimethylformamide was added to a reaction mixture, and reaction was performed at 40° C. to 50° C. for 24 hours. After cooling the reaction mixture to room temperature again, a solution prepared by dissolving dry polyethyleneglycol (8.07 g, 14.67 mmol) having a molecular weight of 750 in 200 mL of anhydrous tetrahydrofuran and adding 5 mL of triethylamine thereto was added to the reaction mixture, and reaction was performed at a temperature of 40° C. to 50° C. for 24 hours.

(223) The reaction solution was filtered to remove produced triethylamine hydrochloride and the filtrate was concentrated under reduced pressure until a small amount of the solvent remained. The concentrated solution was dissolved in 10 mL of tetrahydrofuran, and an excess of hexane was added thereto to induce precipitation. After repeating this process twice or three times, the precipitates were dissolved in a small amount of methanol, placed in MWCO 1200 membrane (Spectrum Laboratories, Inc.), dialyzed against methanol at room temperature for 5 days, dialyzed against distilled water for 5 days, and dried at a low temperature to obtain a poly(dichlorophosphazene) polymer [NP(IleOEt).sub.1.40(AMPEG750).sub.0.30(AEMA).sub.0.30].sub.n.

(224) Then, a solution prepared by dissolving sulfur trioxide pyridine complex (1.32 g, 8.3 mmol) in dimethylformamide (total 400 mL) was added dropwise to the reaction mixture, and reaction was performed at a temperature of 40° C. to 50° C. for 24 hours to obtain 9.5 g of [NP(IleOEt).sub.1.40(AMPEG750).sub.0.30(AEMAES).sub.0.16(AEMA).sub.0.14].sub.n (yield: 82%).

(225) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(226) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(227) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(228) δ 1.4 to 1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(229) δ 1.9(s, —NHCH.sub.2CH.sub.2O.sub.2C(CH.sub.3)C═CH.sub.2),

(230) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2O.sub.2C(CH.sub.3)CCHCH2SO4,)

(231) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3),

(232) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.11CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(233) δ 3.9 to 4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(234) δ 5.5(s, —NHCH.sub.2CH.sub.2O.sub.2C(CH.sub.3)C═CH.sub.2),

(235) δ 6.1(s, —NHCH.sub.2CH.sub.2O.sub.2C(CH.sub.3)C═CH.sub.2),

(236) Average molecular weight (M.sub.w): 41000

Comparative Example 1

Preparation of Poly[(isoleucineethylester).SUB.1.26.(aminomethoxypolyethyleneglycol 750).SUB.0.44.(aminoethanol).SUB.0.36.phosphazene].SUB.n

(237) A final product [NP(IleOEt).sub.1.26(AMPEG750).sub.0.44(Aminoethanol).sub.0.36].sub.n was obtained in the same manner as in Example 1, except that isoleucine ethyl ester hydrochloride (10.64 g, 54.36 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.79 g, 12.94 mmol), polyethyleneglycol (14.24 g, 18.98 mmol) having a molecular weight of 750, anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were used without perform the reaction with sulfur trioxide pyridine complex.

(238) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(239) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(240) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(241) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(242) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2OH, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(243) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(244) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(245) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(246) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(247) Average molecular weight (M.sub.w): 6800

Comparative Example 2

Preparation of Poly[(isoleucineethylester).SUB.1.38.(aminomethoxypolyethyleneglycol 750).SUB.0.32.(aminoethylsuccinate).SUB.0.15.phosphazene].SUB.n

(248) A final product [NP(IleOEt).sub.1.32(AMPEG750).sub.0.31(Aminoethylsuccinate).sub.0.36].sub.n was obtained in the same manner as in Example 9, except that isoleucine ethyl ester hydrochloride (11.14 g, 56.95 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.95 g, 15.53 mmol), polyethyleneglycol (10.35 g, 13.8 mmol) having a molecular weight of 750, anhydrous succinic acid (1.16 g, 11.64 mmol), dimethylaminopyridine (1.42 g, 11.64 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were used.

(249) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(250) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(251) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(252) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(253) δ 2.5-2.7(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(254) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2OH, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(255) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(256) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(257) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(258) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2COOH),

(259) Average molecular weight (M.sub.w): 4400

Comparative Example 3

Preparation of Poly[(isoleucineethylester).SUB.1.48.(aminomethoxypolyethyleneglycol 750).SUB.0.41.(aminoethylglutarate).SUB.0.11.phosphazene].SUB.n

(260) A final product [NP(IleOEt).sub.1.48(AMPEG750).sub.0.41(AminoethylGlutarate).sub.0.11].sub.n was obtained in the same manner as in Example 9, except that isoleucine ethyl ester hydrochloride (12.5 g, 63.85 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.29 g, 4.7 mmol), polyethyleneglycol (13.27 g, 17.68 mmol) having a molecular weight of 750, anhydrous glutaric acid (3.17 g, 27.81 mmol), dimethylaminopyridine (3.4 g, 27.81 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were used.

(261) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(262) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(263) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(264) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(265) δ 2.1-2.32(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(266) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2OH, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(267) δ 2.9-3.2(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(268) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(269) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(270) δ 3.9-4.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(271) Average molecular weight (M.sub.w): 5600

Comparative Example 4

Preparation of Poly[(isoleucineethylester).SUB.1.44.(aminomethoxypolyethyleneglycol 750).SUB.0.34.(aminoethyladipate).SUB.0.23.phosphazene].SUB.n

(272) A final product [NP(IleOEt).sub.1.44(AMPEG750).sub.0.34(AminoethylAdipate).sub.0.23].sub.n was obtained in the same manner as in Example 9, except that isoleucine ethyl ester hydrochloride (12.16 g, 62.13 mmol), polydichlorophosphazene (5 g, 43.14 mmol), aminoethanol (0.61 g, 9.92 mmol), polyethyleneglycol (11 g, 14.67 mmol) having a molecular weight of 750, anhydrous adipic acid (2.91 g, 22.71 mmol), dimethylaminopyridine (2.77 g, 22.71 mmol), anhydrous THF (total 1000 mL), and triethylamine (total 20 mL) were used.

(273) Nuclear Magnetic Resonance Spectrum with Hydrogen (CDCl.sub.3, ppm):

(274) δ 0.8-1.1(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(275) δ 1.1-1.4(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(276) δ 1.4-1.8(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(277) δ 1.52-1.64(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2CH.sub.2COOH),

(278) δ 2.3-2.32(b, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2CH.sub.2COOH),

(279) δ 2.67-3.2(b, —NHCH.sub.2CH.sub.2OH, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(280) δ 3.4(s, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3),

(281) δ 3.4-3.9(b, —NH(CH.sub.2CH.sub.2O).sub.16CH.sub.3, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3),

(282) δ 3.94.3(b, —NHCH(CH(CH.sub.3)CH.sub.2CH.sub.3)COOCH.sub.2CH.sub.3, —NHCH.sub.2CH.sub.2OCOCH.sub.2CH.sub.2CH.sub.2COOH),

(283) Average molecular weight (M.sub.w): 4430

Experimental Example 1

Sol-Gel Change of Sulfate Group-Containing Phosphazene-Based Polymer According to Temperature Change

(284) Each of the phosphazene-based polymers including or not including a sulfate group prepared according to Examples 1 to 18 and Comparative Examples 1 to 4 was dissolved in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt %. Then, the resultant was placed in a chamber of a viscometer (Brookfield DV-III+ Rheometer) equipped with an automatic water distiller TC-501, and a sol-bel transition behavior thereof was observed at a shear rate of 0.1 to 1.7 per second while raising a temperature by 0.33° C. per minute.

(285) FIG. 1 shows photographs of the solution of the sulfate group-containing phosphazene-based polymer prepared in Example 3 dissolved in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt % obtained by visual observation of the sol-gel behavior thereof. It was confirmed that the solution state was maintained at a temperature below the initial gelation temperature and turned to a gel state at a temperature higher than the body temperature of 37° C.

(286) In addition, gel properties of the polymers according to Examples 1 to 18 and Comparative Examples 1 to 4 with respect to temperature are shown in Table 1 below.

(287) TABLE-US-00001 TABLE 1 Initial Gel strength gelation at body temperature temperature Polymer Structure (° C.) (Pa   s) Example 1 [NP(IleOEt).sub.1.38(AMPEG750).sub.0.57(AEs).sub.0.05].sub.n 16 325 Example 2 [NP(IleOEt).sub.1.35(AMPEG750).sub.0.50(AEs).sub.0.15].sub.n 20 368.7 Example 3 [NP(IleOEt).sub.1.32(AMPEG750).sub.0.38(Aminoethanol).sub.0.10(AES).sub.0.20].sub.n 28 145 Example 4 [NP(IleOEt).sub.1.23(AMPEG550).sub.0.57(AEs).sub.0.20].sub.n 13 300 Example 5 [NP(IleOEt).sub.1.21(AMPEG550).sub.0.51(AEs).sub.0.28].sub.n 20 232.5 Example 6 [NP(IleOEt).sub.1.25(AMPEG550).sub.0.45(Aminoethanol).sub.0.12(AEs).sub.0.18].sub.n 20 292.5 Example 7 [NP(IleOEt).sub.1.38(AMPEG750).sub.0.40(GlyLacOEt).sub.0.02(AEs).sub.0.20].sub.n 15 580 Example 8 [NP(IleOEt).sub.1.23(AMPEG550).sub.0.43(GlyLacOEt).sub.0.04(AEs).sub.0.3].sub.n 23 255 Example 9 [NP(IleOEt).sub.1.38(AMPEG750).sub.0.32(AEs).sub.0.15(Aminoethylsuccinate).sub.0.15].sub.n 28 245 Example 10 [NP(IleOEt).sub.1.22(AMPEG550).sub.0.50(AEs).sub.0.10(Aminoethylsuccinate).sub.0.18].sub.n 22 175 Example 11 [NP(IleOEt).sub.1.39(AMPEG750).sub.0.31(AEs).sub.0.13(AminoethylGlutarate).sub.0.17].sub.n 32 181 Example 12 [NP(IleOEt).sub.1.21(AMPEG550).sub.0.49(AEs).sub.0.20(AminoethylGlutarate).sub.0.1].sub.n 25 125 Example 13 [NP(IleOEt).sub.1.36(AMPEG750).sub.0.28(AEs).sub.0.21(AminoethylAdipate).sub.0.15].sub.n 29 400 Example 14 [NP(IleOEt).sub.1.21(AMPEG550).sub.0.49(AEs).sub.0.12(AminoethylAdipate).sub.0.18].sub.n 19 456.5 Example 15 [NP(IleOEt).sub.1.40(AMPEG750).sub.0.3(AEs).sub.0.16(AminoethylAdipate).sub.0.14].sub.n 31 251 Example 16 [NP(IleOEt).sub.1.41(AMPEG750).sub.0.40(AEs).sub.0.12(AminoethylsuccinateImidazole).sub.0.07].sub.n 30 230 Example 17 [NP(IleOEt).sub.1.49(AMPEG750).sub.0.34(AEs).sub.0.10(Ami- 28 270 noethylsuccinateCRRRRHHHHHHGGGGGRGDS).sub.0.07].sub.n Example 18 [NP(IleOEt).sub.1.4(AMPEG750).sub.0.30(AEMAEs).sub.0.16(AminoethylMethacylate).sub.0.14].sub.n 18 325 Comparative [NP(IleOEt).sub.1.26(AMPEG750).sub.0.44(Aminoethanol).sub.0.36].sub.n 10 206 Example 1 Comparative [NP(IleOEt).sub.1.32(AMPEG750).sub.0.32(Aminoethylsuccinate).sub.0.36].sub.n 16 662 Example 2 Comparative [NP(IleOEt).sub.1.48(AMPEG750).sub.0.41(AminoethylGlutarate).sub.0.11].sub.n 19 893.7 Example 3 Comparative [NP(IleOEt).sub.1.33(AMPEG750).sub.0.44(AminoethylAdipate).sub.0.23] 23 225 Example 4

(288) In Table 1 above, the term ‘initial gelation temperature’ refers to a temperature where a viscosity of an aqueous solution of the polymer (a viscosity of 2 Pas or lower when measured by a viscometer) starts to gradually increase, specifically, a temperature where a viscosity measured by the viscometer is 10 Pas or higher, and the term ‘gel strength at body temperature’ refers to a strength of a polymer gel measured at 37° C.

(289) As shown in Table 1, it was confirmed that the polymers consisting of the same substituents (Examples 1 and 2) may have different initial gelation temperatures and strengths according to the ratio of respective substituents.

(290) Furthermore, viscosities of the solutions of the sulfate group-containing phosphazene-based polymers (Examples 3 and 11) according to the present invention dissolved in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt % were measured with respect to temperature and the results are shown in FIGS. 2 and 3. A state flowing fast like water was observed from a low temperature to room temperature since the viscosity thereof was 0, and a state flowing slow unlike water was observed at around the body temperature due to an increase in the viscosity. In general, a gel state that does not flow in the direction of gravity was observed at a viscosity of 100 Pa.Math.s or greater, and thus it was confirmed that the state was changed to a gel state.

Experimental Example 2

Surface Charge Change of Sulfate Group-Containing Phosphazene-Based Polymer

(291) In order to measure surface charges of the sulfate group-containing phosphazene-based polymers according to the present invention, each of the polymers was dissolved in phosphate-buffered saline (pH 7.4) at a concentration of 1 wt % and a Zeta-potential thereof was measured (Zetasizer Nano ZS, Malvern instrunets Ltd., Malvernm UK). Various biologically effective factors may be retained in a hydrogel of the polymer and sustained-release thereof may be achieved due to ionic interactions according to types and degrees of surface charges of the polymers.

(292) In order to observe changes in surface charges caused by introduction of the sulfate group, surface charges of a hydroxy group-containing phosphazene-based polymer (Comparative Example 1) as a matrix polymer of the sulfate group-containing phosphazene polymer, the phosphazene-based polymer substituted with a carboxyl group (Comparative Example 4), and a sulfate group-containing phosphazene polymer (Example 3) were respectively measured and the results are shown in FIG. 4. As shown in Table 1, when the total amount of substituents is considered as 2, amounts of anionic group-containing substituents of the phosphazene-based polymers according to Comparative Example 1, Comparative Example 4, and Example 3 were 0, 0.26, and 0.2, respectively. Referring to FIG. 4, the polymers prepared according to Comparative Example 1, Comparative Example 4, and Example 3 exhibited zeta potentials of −0.21 mV, −3.93 mV, and −6.23 mV, respectively, indicating that the polymer including the sulfate group has a stronger negative charge although it has fewer substituents.

(293) Furthermore, in order to measure changes in surface charges of the polymers according to a substitution degree of the sulfate group, surface charges of the group-containing phosphazene-based polymers prepared as described above were measured and shown in FIG. 5. When the total amount of substituents is considered as 2, amounts of the sulfate groups of the phosphazene-based polymers according to Examples 1, 2, and 3 were 0.05, 0.15, and 0.2 respectively. Surface charges of the polymers according to Examples 1, 2, and 3 were measured as −3.38 mV, −4.83 mV, and −6.23 mV, respectively, indicating that surface charge varies according to the amount of the sulfate group.

Experimental Example 3

Observation of In Vivo Gelation and Biodegradability by Injection of Aqueous Solution of Sulfate Group-Containing Phosphazene Polymer

(294) In order to identify an in vivo degradation rate of the sulfate group-containing phosphazene-based polymer hydrogel, changes in amounts of the polymer hydrogel over time were measured after injection into mice. Specifically, the sulfate group-containing phosphazene-based polymer according to Example 3 was dissolved in phosphate-buffered saline (pH 7.4) at a concentration of 10 wt % and loaded into a 31G syringe and then subcutaneously injected into the mice. A subcutaneous area was excised over time, and changes in residual amounts of the sulfate group-containing phosphazene-based polymer were measured. The results are shown in FIG. 6.

Experimental Example 4

Evaluation of Cytotoxicity of Sulfate Group-Containing Phosphazene Polymer

(295) In order to evaluate cytotoxicity of the sulfate group-containing polyphosphazene-based polymer, the polymer was dissolved in a cell culture at a concentration of up to 30 mg/ml and added to a 96 well plate in which 10,000 Fibroblast (NIH3T3) cells were distributed at different concentrations. The cells were cultured overnight or more and cell viability was measured to evaluate cytotoxicity of the sulfate group-containing polyphosphazene-based polymer, and the results are shown in FIG. 7. By simultaneously evaluating the sulfate group-containing polyphosphazene-based polymer according to Example 11 and a precursor polymer immediately before introducing the sulfate group thereinto, it was proved that cytotoxicity was prevented by introducing the sulfate group. Interestingly, it was found that the sulfate group contained in the hydrogel of the phosphazene-based polymer alleviated cytotoxicity although the polyphosphazene-based polymer was used at a high concentration.

Experimental Example 5

Observation of Sol-Gel Transition of Aqueous Solution of Sulfate Group-Containing Phosphazene-Based Polymer Including Synthetic Drug According to Temperature Change

(296) In order to identify the possibility of the phosphazene-based polymer according to the present invention as a drug carrier, an aqueous solution of the polymer loaded with a synthetic drug was prepared, gel properties thereof according to temperature change were observed, and the results are shown in FIG. 8. In FIG. 8, viscosity of a solution prepared by dissolving the sulfate group-containing phosphazene-based polymer according to Example 1 in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt % and viscosity of a solution prepared by further dissolving Triamcinolone acetonide (synthetic corticosteroiod for intra-articular use) in the same solution at a concentration of 1 wt % according to temperature were shown.

(297) ##STR00005##

Experimental Example 6

Observation of Sol-Gel Transition of Sulfate Group-Containing Phosphazene-Based Polymer Including Protein Drug According to Temperature Change

(298) In order to identify the possibility of the phosphazene-based polymer according to the present invention as a drug carrier, an aqueous solution of the polymer loaded with a protein drug was prepared, gel properties thereof according to temperature change were observed, and the results are shown in FIG. 9. In FIG. 9, viscosity of a solution prepared by dissolving the sulfate group-containing phosphazene-based polymer according to Example 11 in phosphate-buffered saline (pH 7.4) at 4° C. at a concentration of 10 wt % and viscosity of a solution prepared by further dissolving vascular endothelial growth factor (VEGF) in the same solution at a concentration of 50 μg/ml according to temperature were shown.

Experimental Example 7

Observation of In Vitro Releasing Behavior of Vascular Endothelial Growth Factor (VEGF) from Sulfate Group-Containing Phosphazene-Based Polymer Hydrogel

(299) 50 μg of a vascular endothelial growth factor (VEGF) was added to 1 mL of a solution prepared by dissolving the phosphazene-based polymer according to Example 11 in phosphate-buffered saline at a concentration of 10 wt %, and then 300 μl of the mixture was loaded on a milli cell, followed by formation of a hydrogel at 37° C. The phosphazene-based polymer hydrogel loaded with the vascular endothelial growth factor (VEGF) was added to 6 mL of a release solution (phosphate-buffered saline, pH 7.4) and stirred at 37° C. in a water bath at 50 rpm, and the release solution was replaced with 6 mL of a new release solution at a predetermined time. The released vascular endothelial growth factor (VEGF) was quantified by analyzing the release solution collected at a predetermined time using an ELIZA KIT.

(300) A release behavior of the vascular endothelial growth factor (VEGF) from the phosphazene-based polymer hydrogel over time is shown in FIG. 10. As shown in FIG. 10, while the phosphazene-based polymer hydrogel not including the sulfate group did not inhibit initial release of the vascular endothelial growth factor (VEGF), it was confirmed that the sulfate group-containing phosphazene-based polymer hydrogel efficieintly inhibited an excessive release of the vascular endothelial growth factor (VEGF) loaded thereon at the initial stage and the release was controlled for a long time. This may be understood because the vascular endothelial growth factor may be trapped in the hydrogel for a long time due to ionic interactions between the vascular endothelial growth factor (VEGF) cationic at pH 7.4 and the sulfate group-containing phosphazene-based polymer anionic in the same environment.

Experimental Example 8

Observation of In Vitro Release Behavior of Stromal Cell-Derived Factor-1 (SDF-1) from Sulfate Group-Containing Phosphazene-Based Polymer Hydrogel

(301) 50 μg of a stromal cell-derived factor-1 (SDF-1) was added to 1 mL of a solution prepared by dissolving the phosphazene-based polymer according to Example 11 in phosphate-buffered saline at a concentration of 10 wt %, and then 300 μl of the mixture was loaded on a milli cell, followed by formation of a hydrogel at 37° C. The phosphazene-based polymer hydrogel loaded with the stromal cell-derived factor-1 (SDF-1) was added to 6 mL of a release solution (phosphate-buffered saline, pH 7.4) and stirred at 37° C. in a water bath at 50 rpm, and the release solution was replaced with 6 mL of a new release solution at a predetermined time. The released stromal cell-derived factor-1 (SDF-1) was quantified by analyzing the release solution collected at a predetermined time using an ELIZA KIT.

(302) A release behavior of the stromal cell-derived factor-1 (SDF-1) from the phosphazene-based polymer hydrogel over time is shown in FIG. 11. As shown in FIG. 11, while the phosphazene-based polymer hydrogel not including the sulfate group did not inhibit initial release of the stromal cell-derived factor-1 (SDF-1), the carboxyl group-containing phosphazene-based polymer hydrogel controlled the release behavior of the stromal cell-derived factor-1 (SDF-1) relatively well. In this case, the sulfate group-containing phosphazene-based polymer hydrogel showed a far less initial release amount than the previous release patterns observed in Comparative Examples 1 and 4. This may be understood because the stromal cell-derived factor-1 (SDF-1) may be trapped in the hydrogel for a long time due to ionic interactions between the stromal cell-derived factor-1 (SDF-1) cationic in the pH 7.4 environment and the sulfate group-containing phosphazene-based polymer anionic in the same environment.

Experimental Example 9

Evaluation of Skin Regeneration Effect of Sulfate Group-Containing Polyphosphazene-Based Polymer in Skin-Damaged Animal Model of Mice

(303) The sulfate group-containing phosphazene-based polymer prepared according to Example 11 was dissolved in phosphate-buffered saline at a concentration of 10 wt % and mice having severe skin damage on the back were treated with the solution to evaluate self-skin regeneration efficacy. The sulfate group-containing polyphosphazene-based polymer hydrogel alone may promote the skin regeneration. Even when a biologically effective factor or a stem cell was added thereto, skin regeneration may also be promoted due to improved abilities to store the biologically effective factor or to deliver the biologically effective factor to the damaged region. Efficacy of the sulfate group-containing phosphazene-based polymer hydrogel is shown in more detail in FIGS. 12, 13, 14, 15, and 16. When skin regeneration of damaged areas was induced using the hydrogel according to Example 11 as shown in FIGS. 12, 13, 14, 15, and 16, statistically significant (*p<0.05) regeneration effects were observed when compared with the comparative examples without using the hydrogel and the polymer of Comparative Example 1.

(304) In short, the phosphazene-based polymer according to the present invention may efficiently introduce a biologically effective factor into a hydrogel via a sulfate group to promote migration, growth, and differentiation of cells, thereby exhibiting superior regeneration efficacy when compared with natural regeneration or the polymer hydrogels according to Comparative Examples 1 and 4.