本研究係採用賴胺酸與苄基半胱胺酸合成具有親水與疏水鏈段之高分子，並設計出不同型態之聚賴胺酸嵌段聚苄基半胱胺酸(PLL-b-PBLC)，探討此共聚高分子之成膠效應。利用聚賴胺酸與苄基半胱胺酸在水溶液中親疏水鏈段之交互作用力形成無規則線圈、β-折板等二級結構，探討此鏈段長度、比例、分枝結構對於成膠機制的影響，並利用各種儀器分析水膠高分子微結構之堆疊、排列。本實驗利用一級胺作為線性起始劑，對賴胺酸鏈段進行開環聚合，利用第一段胺基酸尾端之親核官能基進行第二段聚合，得到多種不同比例之嵌段聚胺酸，再改用分枝狀起始劑以相似步驟，合成2種分枝狀嵌段聚胺酸，來針對不同嵌段比與分枝數目來探討其成膠能力並討論其流變性質，以流變儀在不同操作條件下測試水膠特性，發現水膠具有優異回復性質，且黏彈性質與PLL-b-PBLC分枝數、鏈長有關。以CD光譜儀與FT-IR光譜確認二級結構的存在，並以軟體分析其二級結構含量，苄基半胱胺酸具有形成β-折板結構之能力以幫助凝膠化，推測水膠之強度來自於賴胺酸、芐基半胱胺酸與溶劑之間的氫鍵作用力、靜電排斥力、分子鏈間相互作用力的平衡。利用SAXS、SEM來分析水膠結構，發現PLL-b-PBLC水膠在凝膠化時，其自組裝型態接近於球型堆疊，而這些球形堆疊在SEM視野下又會再聚集成片狀結構。

In this research, we synthesized linear and star-shaped poly (L-lysine)-block-poly (L-benzyl-cysteine) (PLL-b-PBLC) block copolypeptides, and studied their hydrogelation and self- assembly in aqueous solution by varying polypeptide chain length, block ratio, and branch number. Confirming the successful synthesis of polypeptides by NMR, MALDTOF, GPC analyses, our experimental data showed that these linear and star-shaped PLL-b-PBLC block copolypeptides self-assemble to form two-dimensional or three-dimensional morphologies due to the packing of sheetlike PLBC, depending on polypeptide topology and concentration. Transparent hydrogels were formed upon further increasing the polypeptide concentration. The balance between the intermolecular hydrogen bonding interactions and charge repulsion exerted respectively by sheetlike PBLC and positively charged, coil PLL segments would dictate their critical gelation concentrations (CGCs), micro/nano structure, and mechanical properties. This study demonstrated that the sheetlike block can trigger polypeptide hydrogelation and these block copolypeptides may serve as biomimetic templates for the synthesis of nanomaterials.

(1) Boretos, J.; Eden, M. Contemporary Biomaterials: Material and Host Response, Clinical Applications, New Technology and Legal Aspects. 1984.
(2) Khademhosseini, A.; Langer, R. Microengineered Hydrogels for Tissue Engineering. Biomaterials 2007, 28, 5087–5092.
(3) Zhao, X.; Wu, H.; Guo, B.; Dong, R.; Qiu, Y.; Ma, P. X. Antibacterial Anti-Oxidant Electroactive Injectable Hydrogel as Self-Healing Wound Dressing with Hemostasis and Adhesiveness for Cutaneous Wound Healing. Biomaterials 2017, 122, 34–47.
(4) Ganta, S.; Devalapally, H.; Shahiwala, A.; Amiji, M. A Review of Stimuli-Responsive Nanocarriers for Drug and Gene Delivery. J. Control. Release 2008, 126, 187–204.
(5) Xu, C. Y.; Inai, R.; Kotaki, M.; Ramakrishna, S. Aligned Biodegradable Nanofibrous Structure: A Potential Scaffold for Blood Vessel Engineering. Biomaterials 2004, 25, 877–886.
(6) Ikeda, S.; Fasman, G. D. Optical Rotatory Dispersion of Poly-S-Carboxymethyl-l-Cysteine in Aqueous Solutions: A Beta ⇌ Random Coil Transition. J. Mol. Biol. 1967, 30 (3), 491–505.
(7) Garrett, R. H.; Grisham, C. M. Biochemistry; Cengage Learning, 2017.
(8) Schulz, G. E.; Schirmer, R. H.Patterns of Folding and Association of Polypeptide Chains; Springer, New York, NY, 1979; pp 66–107.
(9) Carl, B.; Tooze, J. Introduction to Protein Structure; Garland Science, 2012.
(10) Merrifield, R. B. Solid Phase Peptide Synthesis. I. The Synthesis of a Tetrapeptide. J. Am. Chem. Soc. 1963, 85 (14), 2149–2154.
(11) Löfblom, J. Bacterial Display in Combinatorial Protein Engineering. Biotechnol. J. 2011, 6, 1115–1129.
(12) Yazawa, K.; Numata, K. Recent Advances in Chemoenzymatic Peptide Syntheses. Molecules 2014, 19, 13755–13774.
(13) Toplak, A.; Nuijens, T.; Quaedflieg, P. J. L. M.; Wu, B.; Janssen, D. B. Peptiligase , an Enzyme for Efficient Chemoenzymatic Peptide Synthesis and Cyclization in Water. Adv. Synth. Catal. 2016, 358, 2140–2147.
(14) Hadjichristidis, N.; Iatrou, H.; Pitsikalis, M.; Sakellariou, G. Synthesis of Well-Defined Polypeptide-Based Materials via the Ring-Opening Polymerization of α-Amino Acid N-Carboxyanhydrides. Chem. Rev. 2009, 109, 5528–5578.
(15) Deming, T. J. Polypeptide and Polypeptide Hybrid Copolymer Synthesis via NCA Polymerization. Adv. Polym. Sci. 2006, 202 (1), 1–18.
(16) Deming, T. J. Facile Synthesis of Block Copolypeptides of Defined Architecture. Nature 1997, 390, 386–389.
(17) Deming, T. J. Cobalt and Iron Initiators for the Controlled Polymerization of α-Amino Acid-N-Carboxyanhydrides. Macromolecules 1999, 32, 4500–4502.
(18) Pahovnik, D.; Hadjichristidis, N.Ring-Opening Polymerization of N-Carboxyanhydrides for Preparation of Polypeptides and Polypeptide-Based Hybrid Materials with Various Molecular Architectures. In Anionic Polymerization; Springer Japan: Tokyo, 2015, 307–337.
(19) Deng, C.; Wu, J.; Cheng, R.; Meng, F.; Klok, H. A.; Zhong, Z.Functional Polypeptide and Hybrid Materials: Precision Synthesis via α-Amino Acid N-Carboxyanhydride Polymerization and Emerging Biomedical Applications. Prog. Polym. Sci. 2014, 39 (2), 330–364.
(20) Daly, W. H.; Poché, D.The Preparation of N-Carboxyanhydrides of α-Amino Acids Using Bis(Trichloromethyl)Carbonate. Tetrahedron Lett. 1988, 29, 5859–5862.
(21) J. M. van Bemmelen. Das Hydrogel Und Das Krystallinische Hydrat Des Kupferoxyds. Z, Anorg. Chem 1894, 5 (s), 466.
(22) Hoffman, A. S. Hydrogels for Biomedical Applications. Adv. Drug Deliv. Rev. 2002, 43, 3–12.
(23) Abandansari, H. S.; Aghaghafari, E.; Nabid, M. R.; Niknejad, H. Preparation of Injectable and Thermoresponsive Hydrogel Based on Penta-Block Copolymer with Improved Sol Stability and Mechanical Properties. Polymer. 2013, 54 (4), 1329–1340.
(24) Sawamura, K.; Ikeda, T.; Nagae, M.; Okamoto, S. I.; Mikami, Y.; Hase, H.; Ikoma, K.; Yamada, T.; Sakamoto, H.; Matsuda, K. I.; et al. Characterization of in Vivo Effects of Platelet-Rich Plasma and Biodegradable Gelatin Hydrogel Microspheres on Degenerated Intervertebral Discs. Tissue Eng. - Part A 2009, 15, 3719–3727.
(25) Hamedi, H.; Moradi, S.; Hudson, S. M.; Tonelli, A. E. Chitosan Based Hydrogels and Their Applications for Drug Delivery in Wound Dressings: A Review. Carbohydr. Polym. 2018, 199 (March), 445–460.
(26) Pupkaite, J.; Rosenquist, J.; Hilborn, J.; Samanta, A. Injectable Shape-Holding Collagen Hydrogel for Cell Encapsulation and Delivery Cross-Linked Using Thiol-Michael Addition Click Reaction. Biomacromolecules 2019, 20 (9), 3475–3484.
(27) Lee, J. H. Injectable Hydrogels Delivering Therapeutic Agents for Disease Treatment and Tissue Engineering. Biomater. Res. 2018, 22 (1), 1–14.
(28) Yu, F.; Cao, X.; Li, Y.; Zeng, L.; Yuan, B.; Chen, X. An Injectable Hyaluronic Acid/PEG Hydrogel for Cartilage Tissue Engineering Formed by Integrating Enzymatic Crosslinking and Diels-Alder “Click Chemistry.” Polym. Chem. 2014, 5, 1082–1090.
(29) Nowak, A. P.; Breedveld, V.; Pakstis, L.; Ozbas, B.; Pine, D. J.; Deming, T. J. Rapidly Recovering Hydrogel Scaffolds from Self-Assembling Diblock Copolypeptide Amphiphiles. Lett. to Nat. 2002, 417, 424–428.
(30) Li, Z.; Wang, F.; Roy, S.; Sen, C. K.; Guan, J. Injectable, Highly Flexible, and Thermosensitive Hydrogels Capable of Delivering Superoxide Dismutase. Biomacromolecules 2009, 10, 3306–3316.
(31) Buwalda, S. J.; Boere, K. W. M.; Dijkstra, P. J.; Feijen, J.; Vermonden, T.; Hennink, W. E. Hydrogels in a Historical Perspective: From Simple Networks to Smart Materials. J. Control. Release 2014, 190, 254–273.
(32) Hirokawa, Y.; Okamoto, T.; Kimishima, K.; Jinnai, H.; Koizumi, S.; Aizawa, K.; Hashimoto, T. Sponge-like Heterogeneous Gels: Hierarchical Structures in Poly(N-Isopropylacrylamide) Chemical Gels as Observed by Combined Scattering and Confocal Microscopy Method. Macromolecules 2008, 41, 8210–8219.
(33) Zhang, J. T.; Bhat, R.; Jandt, K. D. Temperature-Sensitive PVA/PNIPAAm Semi-IPN Hydrogels with Enhanced Responsive Properties. Acta Biomater. 2009, 5 (1), 488–497.
(34) Huang, C. J.; Chang, F. C. Polypeptide Diblock Copolymers: Syntheses and Properties of Poly(N-Isopropylacrylamide)-b-Polylysine. Macromolecules 2008, 41 (19), 7041–7052.
(35) Hua, J.; Li, Z.; Xia, W.; Yang, N.; Gong, J.; Zhang, J.; Qiao, C. Preparation and Properties of EDC/NHS Mediated Crosslinking Poly (Gamma-Glutamic Acid)/Epsilon-Polylysine Hydrogels. Mater. Sci. Eng. C 2016, 61, 879–892.
(36) Rizwan, M.; Yahya, R.; Hassan, A.; Yar, M.; Azzahari, A. D.; Selvanathan, V.; Sonsudin, F.; Abouloula, C. N. pH Sensitive Hydrogels in Drug Delivery: Brief History, Properties, Swelling, and Release Mechanism, Material Selection and Applications. Polymers (Basel). 2017, 9 (4), 137.
(37) Suzuki, M.; Hayakawa, Y.; Hanabusa, K. Thixotropic Supramolecular Gel Based on L-Lysine Derivatives. Gels 2015, 1, 81–93.
(38) Wu, D.; Xie, X.; Kadi, A. A.; Zhang, Y. Photosensitive Peptide Hydrogels as Smart Materials for Applications. Chinese Chem. Lett. 2018, 29 (7), 1098–1104.
(39) Namdeo, M.; Bajpai, S. K.; Kakkar, S. Preparation of a Magnetic- Field-Sensitive Hydrogel and Preliminary Study of Its Drug Release Behavior. J. Biomater. Sci. 2009, 20, 1747–1761.
(40) Mansky, P.; Liu, Y.; Huang, E.; Russell, T. P.; Hawker, C. Controlling Polymer-Surface Interactions with Random Copolymer Brushes. 1997, 275 (March), 1458–1461.
(41) Hou, S. S.; Fan, N. S.; Tseng, Y. C.; Jan, J. S. Self-Assembly and Hydrogelation of Coil-Sheet Poly(l -Lysine)- Block-Poly(l -Threonine) Block Copolypeptides. Macromolecules 2018, 51 (20), 8054–8063.
(42) Jarosz, T.; Gebka, K.; Stolarczyk, A. Recent Advances in Conjugated Graft Copolymers : 2019.
(43) Arandia, I.; Meabe, L.; Aranburu, N.; Sardon, H.; Mecerreyes, D.; Müller, A. J. Influence of Chemical Structures on Isodimorphic Behavior of Three Different Copolycarbonate Random Copolymer Series. Macromolecules 2020, 53 (2), 669–681.
(44) Pérez-Camargo, R. A.; Arandia, I.; Safari, M.; Cavallo, D.; Lotti, N.; Soccio, M.; Müller, A. J. Crystallization of Isodimorphic Aliphatic Random Copolyesters: Pseudo-Eutectic Behavior and Double-Crystalline Materials. Eur. Polym. J. 2018, 101 (February), 233–247.
(45) Chang, J.; Huang, S.; Section, E. Pd / Al 2 O 3 Catalysts for Selective Hydrogenation of Polystyrene- Block -Polybutadiene- Block -Polystyrene Thermoplastic. 1998, 37, 1220–1227.
(46) Shim, W. S.; Yoo, J. S.; Bae, Y. H.; Lee, D. S. Novel Injectable PH and Temperature Sensitive Block Copolymer Hydrogel. 2005, 6, 2930–2934.
(47) He, C.; Kim, S. W.; Lee, D. S. In Situ Gelling Stimuli-Sensitive Block Copolymer Hydrogels for Drug Delivery. J. Control. Release 2008, 127 (3), 189–207.
(48) Kissel, T.; Li, Y.; Unger, F. ABA-Triblock Copolymers from Biodegradable Polyester A-Blocks and Hydrophilic Poly(Ethylene Oxide) B-Blocks as a Candidate for in Situ Forming Hydrogel Delivery Systems for Proteins; 2002; Vol. 54.
(49) Ian W. Hamley. Developments in Block Copolymer Science and Technology; Hamley, I. W., Ed.; John Wiley & Sons, 2004.
(50) Murphy, R. D.; In hetPanhuis, M.; Cryan, S. A.; Heise, A. Disulphide Crosslinked Star Block Copolypeptide Hydrogels: Influence of Block Sequence Order on Hydrogel Properties. Polym. Chem. 2018, 9 (28), 3908–3916.
(51) Singh, R. P.; Nayak, B. R.; Biswal, D. R.; Tripathy, T.; Banik, K. Biobased Polymeric Flocculants for Industrial Effluent Treatment. Mater. Res. Innov. 2003, 7 (5), 331–340.
(52) Mishra, S.; Sinha, S.; Dey, K. P.; Sen, G. Synthesis, Characterization and Applications of Polymethylmethacrylate Grafted Psyllium as Flocculant. Carbohydr. Polym. 2014, 99, 462–468.
(53) Ding, J.; Zhuang, X.; Xiao, C.; Cheng, Y.; Zhao, L.; He, C.; Tang, Z.; Chen, X. Preparation of Photo-Cross-Linked PH-Responsive Polypeptide Nanogels as Potential Carriers for Controlled Drug Delivery. J. Mater. Chem. 2011, 21 (30), 11383–11391.
(54) Shen, X. Y.; Tang, C. C.; Jan, J. S. Synthesis and Hydrogelation of Star-Shaped Poly(L-Lysine) Polypeptides Modified with Different Functional Groups. Polymer. 2018, 151, 108–116.
(55) Kumar, D.; Pandey, J.; Raj, V.; Kumar, P. A Review on the Modification of Polysaccharide Through Graft Copolymerization for Various Potential Applications. Open Med. Chem. J. 2017, 11 (1), 109–126.
(56) Li, T.; Senesi, A. J.; Lee, B. Small Angle X-Ray Scattering for Nanoparticle Research. Chem. Rev. 2016, 116 (18), 11128–11180.
(57) Shibayama, M.; Isono, K.; Okabe, S.; Karino, T.; Nagao, M. SANS Study on Pressure-Induced Phase Separation of Poly(N- Isopropylacrylamide) Aqueous Solutions and Gels. Macromolecules 2004, 37 (8), 2909–2918.
(58) Percus, J. K.; Yevick, G. J. Analysis of Classical Statistical Mechanics by Means of Collective Coordinates. Phys. Rev. 1958, 110 (1), 1–13.
(59) Feigin, L. A.; Svergun, D. I.; Taylor, G. W. Structure Analysis by Small-Angle X-Ray and Neutron Scattering; Springer, 1987.
(60) Beaucage, G.; Schaefer, D. W. Structural Studies of Complex Systems Using Small-Angle Scattering : A Unified Guinier / Power-Law Approach Structural Studies of Complex Systems Using Small-Angle Scattering : A Unified Guinier / Power-Law Approach. J. Non. Cryst. Solids 1994, 172–174, 797–805.
(61) Hammouda, B. A New Guinier – Porod Model. J. Appl. Crystallogr. 2010, 43, 716–719.
(62) Tamon, H.; Ishizaka, H. SAXS Study on Gelation Process in Preparation of Resorcinol – Formaldehyde Aerogel. 1998, 206, 577–582.
(63) Luo, J.; Panzarasa, G.; Osypova, A.; Sorin, F.; Spano, F.; Rossi, R. M.; Sadeghpour, A.; Boesel, L. F. Polyphenols as Morphogenetic Agents for the Controlled Synthesis of Mesoporous Silica Nanoparticles. Chem. Mater. 2019, 31 (9), 3192–3200.
(64) Takei, T.; Ikeda, K.; Ijima, H.; Kawakami, K. Fabrication of Poly (Vinyl Alcohol) Hydrogel Beads Crosslinked Using Sodium Sulfate for Microorganism Immobilization. Process Biochem. 2011, 46 (2), 566–571.
(65) Das, A. Studies on Complex π-π and T-Stacking Features of Imidazole and Phenyl/p-Halophenyl Units in Series of 5-Amino-1-(Phenyl/p-Halophenyl)Imidazole-4-Carboxamides and Their Carbonitrile Derivatives: Role of Halogens in Tuning of Conformation. J. Mol. Struct. 2017, 1147, 520–540.