Synthesis and characterization of gelatin-based quaternizable hydrogels

Abstract

Gelatin, a water-soluble natural polymer with excellent film-forming properties, exhibits high biocompatibility due to its amino acid composition, which closely resembles that of proteins. However, gelatin has poor mechanical properties and poses a risk of bacterial infection when films are composed solely of gelatin. In this study, gelatin-based crosslinked polymers with quaternary amine groups, exhibiting potential antibacterial properties, were developed. To achieve this, gelatin was first modified with methacrylate via an isocyanate-amine reaction, and the resulting material was characterized using Fourier-transform infrared spectroscopy (FTIR) and nuclear magnetic resonance spectroscopy (NMR). Hydrogels were successfully synthesized by photopolymerization of gelatin methacryloyl, with a tertiary amine-containing monomer and a four-arm crosslinker, and characterized using FTIR, scanning electron microscope (SEM), and thermogravimetric analysis (TGA). Subsequently, the hydrogel was prepared as a film on a glass surface, and quaternization of the tertiary amine groups imparted polycationic properties to the hydrogel coatings, enabling further investigation into their antibacterial potential.

Keywords

• gelatin
• photopolymerization
• hydrogels
• quaternization
• isocyanate-amine

Jelatin bazlı kuaternize edilebilir hidrojellerin sentezi ve karakterizasyonu

Abstract

Üstün film oluşturma özelliklerine sahip, suda çözünebilen doğal bir polimer olan jelatin, proteinlere benzer amino asit bileşimi sayesinde yüksek biyouyumluluk sergilemektedir. Ancak jelatinin mekanik özellikleri zayıftır ve yalnızca jelatinden oluşan filmler, bakteriyel enfeksiyon riski taşımaktadır. Bu çalışmada, potansiyel antibakteriyel özelliklere sahip kuaterner amin grupları içeren jelatin bazlı çapraz bağlı polimerlerin geliştirilmesi amaçlanmıştır. Bunun için önce jelatin izosiyanat-amin reaksiyonu kullanarak metakrilat ile modifiye edilmiş ve ardından fourier dönüşümlü kızılötesi spektroskopisi (FTIR) ve nükleer manyetik rezonans spektroskopisi HNMR ile yapısı aydınlatılmıştır. Jelatin metakriloilin tersiyer amin içeren monomer ve dört kollu bir çapraz bağlayıcı monomer ile fotopolimerleşmesi sonucu hidrojeller elde edilmiş ve FTIR, taramalı elektron mikroskobu (SEM) ve Termogravimetrik Analiz (TGA) teknikleri ile karakterize edilmiştir. Ardından, hidrojeller cam yüzeyde bir film olarak da hazırlanmış ve üçüncül amin gruplarının kuaternizasyonu ile hidrojel kaplamalara polikatyonik özellikler kazandırılarak, antibakteriyel potansiyel barındıran yeni malzemeler elde edilmiştir.

Keywords

• jelatin
• fotopolimerleşme
• hidrojeller
• kuaternizasyon
• izosiyanat-amin

References

1. Fujita, A., Kawakita, H., Saito, K., Sugita, K., Tamada, M., Sugo, T. Production of tripeptide from gelatin using collagenase-immobilized porous hollow-fiber membrane, Biotechnology Progress, 19, 1365-1367, (2003).
2. Su, K., Wang, C. Recent advances in the use of gelatin in biomedical research, Biotechnology Letters, 37, 2139–2145, (2015).
3. Klotz, B. J., Gawlitta, D., Rosenberg, A. J. W. P., Malda, J., Melchels, F. P. W. Gelatin-Methacryloyl Hydrogels: Towards Biofabrication-Based Tissue Repair, Trends in Biotechnology, 34, 394–407, (2016).
4. Dash, R., Foston, M.Ragauskas, A. J. Improving the mechanical and thermal properties of gelatin hydrogels cross-linked by cellulose nanowhiskers, Carbohydrate Polymers, 91, 638–645, (2013).
5. Xing, Q. Yates, K., Vogt, C., Qian, Z., Frost, M. C.; Zhao, F. Increasing Mechanical Strength of Gelatin Hydrogels by Divalent Metal Ion Removal, Scientific Reports, 4, 4706, (2014).
6. Costa, O. Y.A., Pijl, A., Houbraken, J., van Lith, W., Kuramae, E. E., Soil substrate source drives the microbes involved in the degradation of gelatin used as a biostimulant, Applied Soil Ecology, 189, 104906, (2023).
7. Wang, X.; Guo, J.; Zhang, Q.; Zhu, S.; Liu, L.; Jiang, X.; Wei, D.-H.; Liu, R.-S.; Li, L. Gelatin Sponge Functionalized with Gold/Silver Clusters for Antibacterial Application, Nanotechnology, 31, 134004, (2020).
8. Xiang, L., Cui, W. Biomedical application of photo-crosslinked gelatin hydrogels, Journal of Leather Science and Engineering, 3, 3, (2021).

9. Kuijpers, A. J., Engbers, G. H. M., Feijen, J., De Smedt, S. C., Meyvis, T. K. L., J. Demeester, J. Krijgsveld, S. A. J. Zaat and J.Dankert, T. K. Characterization of the Network Structure of Carbodiimide Cross-Linked Gelatin Gels, Macromolecules, 32, 3325-3333, (1999).
10. Yung, C. W., Wu, L. Q., Tullman, J. A., Payne, G. F., Bentley W. E., Barbari, T. A. Transglutaminase crosslinked gelatin as a tissue engineering scaffold, Journal of biomedical materials research. Part A, 83, 1039-104, (2007).
11. Kirchmajer, D. M., Watson, C. A., Ranson, M., Panhuis, and M. I. R., Gelapin, a degradable genipin cross-linked gelatin hydrogel, RCS Advances, 3, 1073-1081 (2013).
12. Van Den Bulcke, A. I., Bogdanov, B., De Rooze, N., Schacht, E. H., Cornelissen, M., Berghmans, H., Structural and rheological properties of methacrylamide modified gelatin hydrogels, Biomacromolecules, 1, 31-38 (2000).
13. Ghosh, R. N., Thomas, J., B. R.,V, N. G., D., Janardanan, A., Namboothiri, P. K. Mathew Peter, M. An insight into synthesis, properties and applications of gelatin methacryloyl hydrogel for 3D bioprinting, Materials Advances, 4, 5496-5529, (2023).
14. Zhu, M., Wang, Y., Ferracci, G., Zheng, J., Cho, N. -J., Lee, B.H. Gelatin methacryloyl and its hydrogels with an exceptional degree of controllability and batch-to-batch consistency, Scientific Reports, 9, 6863, (2019).
15. Velasco-Rodriguez, B., Diaz-Vidal, T., Rosales-Rivera, L.C., García-González, C.A., Alvarez-Lorenzo, C., Al-Modlej, A., Domínguez-Arca, V., Prieto, G., Barbosa, S., Soltero Martínez J.F.A, Taboada, P. Hybrid Methacrylated Gelatin and Hyaluronic Acid Hydrogel Scaffolds. Preparation and Systematic Characterization for Prospective Tissue Engineering Applications. International Journal of Molecular Sciences, 22, 6758, (2021).
16. Zhang, Y., Lv, J., Zhao, J., Ling, G., Zhang, P., A versatile GelMA composite hydrogel: Designing principles, delivery forms and biomedical applications, European Polymer Journal, 197, 112370, (2023).
17. Löwenberg, C., Julich-Gruner, K. K., Neffe, A. T., Lendlein, A., Influence of glycidylmethacrylate functional groups attached to gelatin on the formation and properties of hydrogels, MRS Online Proceedings Library (OPL), 1718, 103-108, (2015).
18. Sk, M. M., Das, P., Panwar, A., Tan, L.P. Synthesis and characterization of site selective photo-crosslinkable glycidyl methacrylate functionalized gelatin-based 3D hydrogel scaffold for liver tissue engineering, Materials Science and Engineering: C. 123, 111694, (2021).
19. Lai, T. C., Yu, J., Tsai, W. B., Gelatin Methacrylate/Carboxybetaine Methacrylate Hydrogels with Tunable Crosslinking for Controlled Drug Release, Journal of Materials Chemistry B, 4, 2304-2313, (2016).
20. Koksal, B., Onbas, R, Baskurt, M., Sahin, H., Arslan-Yildiz, A., Yildiz, U. H. Boosting up printability of biomacromolecule based bio-ink by modulation of hydrogen bonding pairs, European Polymer Journal, 141, 110070, (2020).
21. Hensarling, R. M., Rahane, S. B., LeBlanc, A. P., Sparks, B. J., White, E. M., Locklin, J. Patton, D. L. Thiol–isocyanate “click” reactions: rapid development of functional polymeric surfaces, Polymer Chemistry, 2, 88-90, (2011)..
22. Demirbilek N.; Cengiz, N.; Gevrek, T. N. Pendant isocyanate and epoxide-containing copolymers: synthesis, sequential dual-functionalization with amines, and surface modifications, Journal of Macromolecular Science, Part A, 60, 505-514 (2023).
23. Bat-Ozmatara, M.; Ünlü, A.; Gevrek, T. N., Preparation of isocyanate-containing hydrogel films as antibacterial enzyme immobilization matrices, Reactive and Functional Polymers, 192, 105695 (2024).
24. Sang L. Y., Zhou X. H., Yun F., Zhang G. L. Enzymatic synthesis of chitosan-gelatin antimicrobial copolymer and its characterisation, Journal of the Science of Food and Agriculture, 90, 58–64 (2010).
25. Butun, V.; Armes, S. P.; Billingham, N. C. Synthesis and aqueous solution properties of near-monodisperse tertiary amine methacrylate homopolymers and diblock copolymers, Polymer, 42, 5993– 6008 (2001).
26. Limer, A. J.; Rullay, A. K.; Miguel, V. S.; Peinado, C.; Keely, S.; Fitzpatrick, E.; Carrington, S. D.; Brayden, D. J.; Haddleton, D. M., Fluorescently tagged star polymers by living radical polymerisation for mucoadhesion and bioadhesion, Reactive and Functional Polymers, 66, 51–64 (2006).
27. Lenoir, S, Pagnoulle, C, Detrembleur, C, Galleni, M, Jerome, R, Antimicrobial Activity of Polystyrene Particles Coated by Photo-Crosslinked Block Copolymers Containing a Biocidal Polymethacrylate Block, e-Polymers, 074, 1–11 (2005).
28. Huang, J.; Murata, H.; Koepsel, R. R.; Russell, A. J.; Matyjaszewski, K. Antibacterial Polypropylene via Surface-Initiated Atom Transfer Radical Polymerization, Biomacromolecules, 8, 1396–1399 (2007).
29. Madrid, J. F., Barsbay, M., Abad, L., Güven, O. Grafting of N,N-dimethylaminoethyl methacrylate from PE/PP nonwoven fabric via radiation-induced RAFT polymerization and quaternization of the grafts, Radiation Physics and Chemistry, 124, 145-154, (2016).
30. Lee, S. B.; Koepsel, R. R.; Morley, S. W.; Matyjaszewski, K.; Sun, Y.; Russell, A. J. Permanent, nonleaching antibacterial surfaces. 1. Synthesis by atom transfer radical polymerization, Biomacromolecules, 5, 877–882 (2004).
31. Roy, D.; Knapp, J. S.; Guthrie, J. T.; Perrier, S. Antibacterial cellulose fiber via RAFT surface graft polymerization, Biomacromolecules, 9, 91–99 (2008).
32. Huang, J.; Koepsel, R. R.; Murata, H.; Wu, W.; Lee, S. B.; Kowalewski, T.; Russell, A. J.; Matyjaszewski, K. Antibacterial polypropylene via surface-initiated atom transfer radical polymerization, Langmuir, 24, 6785–6795, (2008).
33. Sroka, M., Zaborniak, I., Chmielarz, P., Bała, J., Wolski, K., Ciszkowicz, E., Awsiuk, K. J. Raczkowska, Grafting of Multifunctional Polymer Brushes from a Glass Surface: Surface-Initiated Atom Transfer Radical Polymerization as a Versatile Tool for Biomedical Materials Engineering, Macromolecular Chemistry and Physics, 225, 2300284, (2023).
34. Yin, J.‐J., Wahid, F., Zhang, Q., Tao, Y. -C., Zhong, C., Chu, L.-Q. Facile Incorporation of Silver Nanoparticles into Quaternized Poly (2‐(Dimethylamino) Ethyl Methacrylate) Brushes as Bifunctional Antibacterial Coatings, Macromolecular Materials and Engineering, 302, 1700069, (2017).
35. Tarakci, E. C.; Gevrek, T. N. Isocyanate Group Containing Reactive Hydrogels: Facile Synthesis and Efficient Biofunctionalization, European Polymer Journal, 175, 111338, (2022).
36. Hoch, E., Hirth, T., Tovar, G. E. M., Borchers, K. Chemical tailoring of gelatin to adjust its chemical and physical properties for functional bioprinting, Journal of Materials Chemistry B, 1, 5675-5685, (2013).
37. Resmi, R., Unnikrishnan, S., Krishnan, L. K., Krishnan, K. Synthesis and characterization of silver nanoparticle incorporated gelatin-hydroxypropyl methacrylate hydrogels for wound dressing applications, Journal of Applied Polymer Science, 134, 44529, (2016).
38. Martineau, L., Pang, N. S., Peng. H. T. Development of a Novel Biomaterial, Part II: Evaluation of a Photo Cross-linking Method, Technical Report, Defence Research & Development Canada, Toronto, (2005).
39. Eckes, K.M., Mu, X., Ruehle, M. A., Ren, P., Suggs, L.J. β sheets not required: combined experimental and computational studies of self-assembly and gelation of the ester-containing analogue of an Fmoc-dipeptide hydrogelator, Langmuir, 30, 5287-5296, (2014).
40. Martin, A. D., Wojciechowski, J. P., Warren, H., Panhuis, M., Thordarson, P. Effect of heterocyclic capping groups on the self-assembly of a dipeptide hydrogel, Soft Matter, 12, 10, 2700–2707, (2016).
41. Song, J., Jung, Y., Lee, I., Jang, J. J. Fabrication of pDMAEMA-coated silica nanoparticles and their enhanced antibacterial activity, Journal of Colloid and Interface Science, 407, 205, (2013).
42. Tu, Q., Wang, J.-C., Liu, R., He, J., Zhang, Y., Shen, S., Xu, J., Liu, J., Yuan, M. -S., Wang, J. Antifouling properties of poly(dimethylsiloxane) surfaces modified with quaternized poly(dimethylaminoethyl methacrylate), Colloids and Surfaces B: Biointerfaces, 102, 361-370, (2013).
43. Koufakis, E., Manouras, T., Anastasiadis, S. H., Vamvakaki, M., Film Properties and Antimicrobial Efficacy of Quaternized PDMAEMA Brushes: Short vs Long Alkyl Chain Length, Langmuir, 36, 3482–3493, (2020).