Blood Compatibility of Amphiphilic Poly(N-α-acrylamide-L-lysine-b-dimethylsiloxane) Block Copolymers
ABSTRACT
Amphiphilic block copolymers poly(LysAA-b-DMS) consisting of a hydrophilic poly(N-α-acrylamide-L-lysine) [poly(LysAA)] segment with different molecular weights and a hydrophobic polydimethylsiloxane (PDMS) segment were prepared as follows. The precursor copolymer poly(Boc-LysAA-OtBu-b-PDMS) was obtained from radical polymerization of N-α-acrylamide-N-ε-tert-butoxycarbonyl-L-lysine-tert-butylester (Boc-LysAA-OtBu) initiated with 4,4’-azobis(polydimethylsiloxane 4-cyanopentanoate) (azo-PDMS) with the molecular weight of PDMS Mw = 4.3 × 103 in the presence of 2-mercaptoethanol (2-ME) as a chain-transfer agent. Removal of the protecting groups of the precursor copolymer was carried out in 80% trifluoroacetic acid aqueous solution to give poly(LysAA-b-DMS)-1-3. The weight average molecular weight of poly(LysAA-b-DMS)-1-3 was Mw = 1.02 × 104 – 2.52 × 104. From the 1H-NMR and fluorescence spectra measurements, poly(LysAA-b-DMS)-1-3 was determined to self-organize and form core-shell micelles in water. The critical micelle concentration (CMC) increased to 1000 - 4000 mg·L–1 with increasing molar ratio of the poly(LysAA) segment from 0.42 to 0.65. From morphological analysis with a scanning probe microscope (SPM), poly(LysAA-b-DMS) has microphase-separated structures made up of hydrophilic and hydrophobic regions with the domain size ranging from several tens to several hundreds of nanometers. Inhibition of thrombin activity of poly(LysAA-b-DMS) was evaluated from the Michaelis constant (KM) and catalytic activity (kcat) for the enzymatic reaction of thrombin and synthetic substrate S-2238 in the presence of poly(LysAA-b-DMS). The KM and kcat were 0.10 - 0.11 mM and 4.04 × 105 – 4.26 × 105 min–1, respectively. Fibrinolytic activity was also verified from the transformation of plasminogen to plasmin by tissue plasminogen activator (t-PA) using synthetic substrate S-2251 in the presence of poly(LysAA-b-DMS). The KM and kcat were 0.07 mM and 5.73 × 106 –5.95 × 106 min–1, respectively.
Amphiphilic block copolymers poly(LysAA-b-DMS) consisting of a hydrophilic poly(N-α-acrylamide-L-lysine) [poly(LysAA)] segment with different molecular weights and a hydrophobic polydimethylsiloxane (PDMS) segment were prepared as follows. The precursor copolymer poly(Boc-LysAA-OtBu-b-PDMS) was obtained from radical polymerization of N-α-acrylamide-N-ε-tert-butoxycarbonyl-L-lysine-tert-butylester (Boc-LysAA-OtBu) initiated with 4,4’-azobis(polydimethylsiloxane 4-cyanopentanoate) (azo-PDMS) with the molecular weight of PDMS Mw = 4.3 × 103 in the presence of 2-mercaptoethanol (2-ME) as a chain-transfer agent. Removal of the protecting groups of the precursor copolymer was carried out in 80% trifluoroacetic acid aqueous solution to give poly(LysAA-b-DMS)-1-3. The weight average molecular weight of poly(LysAA-b-DMS)-1-3 was Mw = 1.02 × 104 – 2.52 × 104. From the 1H-NMR and fluorescence spectra measurements, poly(LysAA-b-DMS)-1-3 was determined to self-organize and form core-shell micelles in water. The critical micelle concentration (CMC) increased to 1000 - 4000 mg·L–1 with increasing molar ratio of the poly(LysAA) segment from 0.42 to 0.65. From morphological analysis with a scanning probe microscope (SPM), poly(LysAA-b-DMS) has microphase-separated structures made up of hydrophilic and hydrophobic regions with the domain size ranging from several tens to several hundreds of nanometers. Inhibition of thrombin activity of poly(LysAA-b-DMS) was evaluated from the Michaelis constant (KM) and catalytic activity (kcat) for the enzymatic reaction of thrombin and synthetic substrate S-2238 in the presence of poly(LysAA-b-DMS). The KM and kcat were 0.10 - 0.11 mM and 4.04 × 105 – 4.26 × 105 min–1, respectively. Fibrinolytic activity was also verified from the transformation of plasminogen to plasmin by tissue plasminogen activator (t-PA) using synthetic substrate S-2251 in the presence of poly(LysAA-b-DMS). The KM and kcat were 0.07 mM and 5.73 × 106 –5.95 × 106 min–1, respectively.
KEYWORDS
Poly(N-α-acrylamide-L-lysine), Polydimethylsiloxane, Block Copolymer, Molecular Assembly, Blood Compatibility, S-2238/S-2251, Biomedical Polymer Material
Poly(N-α-acrylamide-L-lysine), Polydimethylsiloxane, Block Copolymer, Molecular Assembly, Blood Compatibility, S-2238/S-2251, Biomedical Polymer Material
Cite this paper
nullK. Sugiyama, N. Tanigawa and K. Shiraishi, "Blood Compatibility of Amphiphilic Poly(N-α-acrylamide-L-lysine-b-dimethylsiloxane) Block Copolymers," Journal of Biomaterials and Nanobiotechnology, Vol. 2 No. 4, 2011, pp. 337-346. doi: 10.4236/jbnb.2011.24042.
nullK. Sugiyama, N. Tanigawa and K. Shiraishi, "Blood Compatibility of Amphiphilic Poly(N-α-acrylamide-L-lysine-b-dimethylsiloxane) Block Copolymers," Journal of Biomaterials and Nanobiotechnology, Vol. 2 No. 4, 2011, pp. 337-346. doi: 10.4236/jbnb.2011.24042.
References
[1] A. Kishida, T. Furuzono, T. Oshige, I. Maruyama, T. Matsumoto, H. Itoh, M. Murakami and M. Akashi, “Study of the surface properties of ultrathin films of poly (dimethylsiloxane)-polyamide multiblock copolymers,” Angewante Makromolekulare Chemie, Vol. 220, No. 7, 1994, pp. 89-97. [2] T. Furuzono, E. Yashima, A. Kishida, I. Maruyama, T. Matsumoto and M. Akashi, “A Novel Biomaterial Poly (dimethylsiloxane)-Polyamide Multiblock Copolymer 1. Synthesis and Evaluation of Blood Compatibility,” Journal of Biomaterals Science-Polymer Edition, Vol. 5, No. 1-2, 1993, pp. 89-98. [3] E. Nyilas, “Polysiloxane-polyurethane block copolymers” US Patent No. 3562352, 1971. [4] K. Sugiyama, K. Isobe and K. Shiraishi, “Introduction of tetramethyldisiloxane moiety into polyetherurethaneurea”, Nippon KagakuKaishi, Vol. 1997, No. 11, pp. 816-820. [5] K. Sugiyama, M. Tanigawa and K. Shiraishi, “Characterization of polysimethylsiloxane block copolymers containing hydrophobic polymethacrylate segments”, Nippon KagakuKaishi, Vol. 1998, No. 8, pp. 551-557. [6] K. Sugiyama and H. Aoki, “Surface modified polymer microspheres obtained by the emulsion copolymerization of 2-methacryloyloxyethyl phosphorylcholine with various vinyl monomers”, Polymer Journal, Vol. 26, No. 5, 1994, pp. 561-569. [7] K. Sugiyama, K. Shiraishi, K. Okada and O. Matsuo, “Biocompatible Block Copoymers Composed of Polydimethylsiloxane and Poly[2-(methacryloyloxy)ethyl phosporylcholine] Segments”, Polymer Journal, Vol. 31, No. 10, 1999, pp. 883-886. [8] K. Sugiyama, K. Shiraishi and T. Matsumoto , “Assembly of Amphiphilic Poly[2-(methacryloyloxy)ethyl phosphorylcholine] with Cholesteryl Moieties as Terminal Groups,” Journal of Polymer Science: Part A: Polymer Chemistry, Vol. 41, 2003, pp. 1992-2000. [9] K. Shiraishi, M. Sugiyama, Y. Okamura and K. Sugiyama, “Cholesteryl moiety terminated amphiphilic polymethacrylates containing nucleic acid base for drug delivery”, Journal of Applied Polymer Science, Vol. 103, 2007, pp. 3064-3075. [10] K. Shiraishi, K. Miura, G. Asami, M. Kohta and K. Sugiyama, “Analysis of pH Response for the Amphoteric Poly(O-methacrloyl-L-serine and Interaction with Serum Protein by Fluorescence Spectroscopy,” Japanese Journal Polymer Science and Technology, Vol. 60, No. 1, 2003, pp. 30-37. [11] N. Tanigawa, K. Shiraishi and K. Sugiyama, “Blood Compatibility of Self-Assembled Poly(N-α-methacrylamide-L-lysine-b-dimethylsiloxane) Copolymers,” Japanese Journal of Polymer Science and Technology, Vol. 65, No. 2, 2008, pp. 150-156. [12] K. Shiraishi, M. Kohta and K. Sugiyama,”Preparation of Zwitterionic Polyacrylamide Modified with L-Lysine and Its Effect on Fibrinolytic Activity,” Chemistry Letters, Vol. 33, No. 6, 2004, pp. 646-647. [13] J. Brandrup and E. H. Immergut, “Polymer Handbook 2nd Ed, “ pIII-139, John Willy & Sons, N.Y., 1975. [14] G. S. Kwon, M. Naito, M. Yokoyama, T. Okano, Y. Sakurai and K. Kataoka, “Micelles Based on AB Block Copolymers of Poly(ethylene oxide) and Poly(β-Benzyl- L-Aspartate),” Langmuir, Vol. 9, No. 4, 1993, pp. 945- 949. [15] G. S. Kwon, M. Naito, M. Yokoyama, T. Okano, Y. Sakurai and K. Kataoka, ”Physical Entrapment of Adriamycin in AB Block-Copolymer Micelles,” Pharmaceutical Research, Vol. 12, No. 2, 1995, pp. 192-195. [16] N. Tanigawa, K. Shiraishi and K. Sugiyama, “Characterization of Thermo-Responsive Poly[N-(2-Hydroxypropyl)Methacrylamide-Dimethylsiloxane] Block Copolymers,” Journal of the Society of Material Science Japan, Vol. 55, No. 4, 2006, pp. 391-396. doi: 10.2472/jsms.55.391 [17] N. Tanigawa, K. Shiraishi and K. Sugiyama, “Molecular Assembly and Blood Compatibility of Poly[2-(methacryloyloxy)ethyl phosporylcholine-b-dimethylsiloxane] Block Copolymers,” Japanese Journal of Polymer Science and Technology, Vol. 64, No. 6, 2007, pp. 373- 379. doi:10.1295/koron.64.373 [18] D. A. Smith, “The Thermal Decomposition of Azonitrile Polymers,” Die Makromolekulare Chemie, Vol. 103, 1967, pp. 301-303. [19] S. B. Cho, K. Nakanishi, T. Kokubo, N. Soga, C. Ohtsuki, T. Nakamura, K. Kitsugi and T. Yamamuro, “Dependence of Apatite Formation on Silica-Gel. On Its Structure - Effect of Heat-Treatment,” Journal of the American Chemical Society, Vol. 78, No. 7, 1995, pp. 1769- 1774. [20] J. E. Chung, M. Yokoyama, K. Suzuki, T. Aoyagi, Y. Sakurai and T. Okano, ”Reversible Thermo-responsive Alkyl-terminated poly(N-isopropylacrylamide) Core- Shell Micellar structures,” Colloids and Surfaces B-Biointerfaces, Vol. 9, No. 1-2, 1997, pp. 37-48. [21] K. Akiyoshi, S. Deguchi, N. Moriguchi, S. Yamaguchi and J. Sunamoto, “Self-Aggregates of Hydrophobized Polysaccharides in Water-Formation and Characteristics of Nanoparticles,” Macromolecules, Vol. 26, No. 12, 1993, pp. 3062-3068. [22] A. Girolami, L.Saggin and G.Boeri, “Factor X Assays Using Chromogenic Substrate S-2238”, American Journal of Clinical Pathology, Vol. 73, No. 3, 1980, pp. 400- 402. [23] H. Fukao, S. Ueshima, T. Takaishi, K. Okada and O. Matsuo, “Enhancement of Tissue-Type Plasminogen Activator (t-PA) Activity by Purified t-PA Receptor Expressed in Human Endothelial Cells”, Biochimica et Biophysica Acta-Molecular Cell Research, Vol. 1356, No. 1, 1997, pp. 111-120. [24] K. D. Fowers and J. Kope?ek, “Development of a Fibrinolytic surface : Specific and Non-specific Binding of Plasminogen,” Colloids & Surfaces B-Biointerfaces, Vol. 9, No. 6, 1997, pp. 315-330. [25] P. H. Warkentin, K. Johansen, J. L. Brash and J. Lundstrom, “The Kinetics and Affinity of Binding of GLU- Plasminogen Specific to the ε-Amino Group of L-Lysine : Its potential Application to Modified Biomaterials, ” Journal of Colloid & Interface Science, Vol. 199, No. 2, 1998, pp. 131-139. [26] P. H. Warkentin, “Selective Plasminogen Binding : Cystein-Lysine-Dextran Protein Interactions,” Biomaterials, Vol. 19, No. 19, 1998, pp. 1753-1761. [27] K. Shiraishi, S. Wakisaka, J. Satozaki and K. Sugiyama, “Preparation and Fibrinolytic Activity of Poly(acrylamide) Containing-L-Lysine Moiety” will be published somewhere else.
[1] A. Kishida, T. Furuzono, T. Oshige, I. Maruyama, T. Matsumoto, H. Itoh, M. Murakami and M. Akashi, “Study of the surface properties of ultrathin films of poly (dimethylsiloxane)-polyamide multiblock copolymers,” Angewante Makromolekulare Chemie, Vol. 220, No. 7, 1994, pp. 89-97. [2] T. Furuzono, E. Yashima, A. Kishida, I. Maruyama, T. Matsumoto and M. Akashi, “A Novel Biomaterial Poly (dimethylsiloxane)-Polyamide Multiblock Copolymer 1. Synthesis and Evaluation of Blood Compatibility,” Journal of Biomaterals Science-Polymer Edition, Vol. 5, No. 1-2, 1993, pp. 89-98. [3] E. Nyilas, “Polysiloxane-polyurethane block copolymers” US Patent No. 3562352, 1971. [4] K. Sugiyama, K. Isobe and K. Shiraishi, “Introduction of tetramethyldisiloxane moiety into polyetherurethaneurea”, Nippon KagakuKaishi, Vol. 1997, No. 11, pp. 816-820. [5] K. Sugiyama, M. Tanigawa and K. Shiraishi, “Characterization of polysimethylsiloxane block copolymers containing hydrophobic polymethacrylate segments”, Nippon KagakuKaishi, Vol. 1998, No. 8, pp. 551-557. [6] K. Sugiyama and H. Aoki, “Surface modified polymer microspheres obtained by the emulsion copolymerization of 2-methacryloyloxyethyl phosphorylcholine with various vinyl monomers”, Polymer Journal, Vol. 26, No. 5, 1994, pp. 561-569. [7] K. Sugiyama, K. Shiraishi, K. Okada and O. Matsuo, “Biocompatible Block Copoymers Composed of Polydimethylsiloxane and Poly[2-(methacryloyloxy)ethyl phosporylcholine] Segments”, Polymer Journal, Vol. 31, No. 10, 1999, pp. 883-886. [8] K. Sugiyama, K. Shiraishi and T. Matsumoto , “Assembly of Amphiphilic Poly[2-(methacryloyloxy)ethyl phosphorylcholine] with Cholesteryl Moieties as Terminal Groups,” Journal of Polymer Science: Part A: Polymer Chemistry, Vol. 41, 2003, pp. 1992-2000. [9] K. Shiraishi, M. Sugiyama, Y. Okamura and K. Sugiyama, “Cholesteryl moiety terminated amphiphilic polymethacrylates containing nucleic acid base for drug delivery”, Journal of Applied Polymer Science, Vol. 103, 2007, pp. 3064-3075. [10] K. Shiraishi, K. Miura, G. Asami, M. Kohta and K. Sugiyama, “Analysis of pH Response for the Amphoteric Poly(O-methacrloyl-L-serine and Interaction with Serum Protein by Fluorescence Spectroscopy,” Japanese Journal Polymer Science and Technology, Vol. 60, No. 1, 2003, pp. 30-37. [11] N. Tanigawa, K. Shiraishi and K. Sugiyama, “Blood Compatibility of Self-Assembled Poly(N-α-methacrylamide-L-lysine-b-dimethylsiloxane) Copolymers,” Japanese Journal of Polymer Science and Technology, Vol. 65, No. 2, 2008, pp. 150-156. [12] K. Shiraishi, M. Kohta and K. Sugiyama,”Preparation of Zwitterionic Polyacrylamide Modified with L-Lysine and Its Effect on Fibrinolytic Activity,” Chemistry Letters, Vol. 33, No. 6, 2004, pp. 646-647. [13] J. Brandrup and E. H. Immergut, “Polymer Handbook 2nd Ed, “ pIII-139, John Willy & Sons, N.Y., 1975. [14] G. S. Kwon, M. Naito, M. Yokoyama, T. Okano, Y. Sakurai and K. Kataoka, “Micelles Based on AB Block Copolymers of Poly(ethylene oxide) and Poly(β-Benzyl- L-Aspartate),” Langmuir, Vol. 9, No. 4, 1993, pp. 945- 949. [15] G. S. Kwon, M. Naito, M. Yokoyama, T. Okano, Y. Sakurai and K. Kataoka, ”Physical Entrapment of Adriamycin in AB Block-Copolymer Micelles,” Pharmaceutical Research, Vol. 12, No. 2, 1995, pp. 192-195. [16] N. Tanigawa, K. Shiraishi and K. Sugiyama, “Characterization of Thermo-Responsive Poly[N-(2-Hydroxypropyl)Methacrylamide-Dimethylsiloxane] Block Copolymers,” Journal of the Society of Material Science Japan, Vol. 55, No. 4, 2006, pp. 391-396. doi: 10.2472/jsms.55.391 [17] N. Tanigawa, K. Shiraishi and K. Sugiyama, “Molecular Assembly and Blood Compatibility of Poly[2-(methacryloyloxy)ethyl phosporylcholine-b-dimethylsiloxane] Block Copolymers,” Japanese Journal of Polymer Science and Technology, Vol. 64, No. 6, 2007, pp. 373- 379. doi:10.1295/koron.64.373 [18] D. A. Smith, “The Thermal Decomposition of Azonitrile Polymers,” Die Makromolekulare Chemie, Vol. 103, 1967, pp. 301-303. [19] S. B. Cho, K. Nakanishi, T. Kokubo, N. Soga, C. Ohtsuki, T. Nakamura, K. Kitsugi and T. Yamamuro, “Dependence of Apatite Formation on Silica-Gel. On Its Structure - Effect of Heat-Treatment,” Journal of the American Chemical Society, Vol. 78, No. 7, 1995, pp. 1769- 1774. [20] J. E. Chung, M. Yokoyama, K. Suzuki, T. Aoyagi, Y. Sakurai and T. Okano, ”Reversible Thermo-responsive Alkyl-terminated poly(N-isopropylacrylamide) Core- Shell Micellar structures,” Colloids and Surfaces B-Biointerfaces, Vol. 9, No. 1-2, 1997, pp. 37-48. [21] K. Akiyoshi, S. Deguchi, N. Moriguchi, S. Yamaguchi and J. Sunamoto, “Self-Aggregates of Hydrophobized Polysaccharides in Water-Formation and Characteristics of Nanoparticles,” Macromolecules, Vol. 26, No. 12, 1993, pp. 3062-3068. [22] A. Girolami, L.Saggin and G.Boeri, “Factor X Assays Using Chromogenic Substrate S-2238”, American Journal of Clinical Pathology, Vol. 73, No. 3, 1980, pp. 400- 402. [23] H. Fukao, S. Ueshima, T. Takaishi, K. Okada and O. Matsuo, “Enhancement of Tissue-Type Plasminogen Activator (t-PA) Activity by Purified t-PA Receptor Expressed in Human Endothelial Cells”, Biochimica et Biophysica Acta-Molecular Cell Research, Vol. 1356, No. 1, 1997, pp. 111-120. [24] K. D. Fowers and J. Kope?ek, “Development of a Fibrinolytic surface : Specific and Non-specific Binding of Plasminogen,” Colloids & Surfaces B-Biointerfaces, Vol. 9, No. 6, 1997, pp. 315-330. [25] P. H. Warkentin, K. Johansen, J. L. Brash and J. Lundstrom, “The Kinetics and Affinity of Binding of GLU- Plasminogen Specific to the ε-Amino Group of L-Lysine : Its potential Application to Modified Biomaterials, ” Journal of Colloid & Interface Science, Vol. 199, No. 2, 1998, pp. 131-139. [26] P. H. Warkentin, “Selective Plasminogen Binding : Cystein-Lysine-Dextran Protein Interactions,” Biomaterials, Vol. 19, No. 19, 1998, pp. 1753-1761. [27] K. Shiraishi, S. Wakisaka, J. Satozaki and K. Sugiyama, “Preparation and Fibrinolytic Activity of Poly(acrylamide) Containing-L-Lysine Moiety” will be published somewhere else.