Pharmaceutical Research

, Volume 30, Issue 2, pp 523–537 | Cite as

Evaluation of Wound Healing Potential of β-Chitin Hydrogel/Nano Zinc Oxide Composite Bandage

  • Sudheesh Kumar P. T.
  • Vinoth-Kumar Lakshmanan
  • Mincy Raj
  • Raja Biswas
  • Tamura Hiroshi
  • Shantikumar V. Nair
  • Rangasamy JayakumarEmail author
Research Paper



β-chitin hydrogel/nZnO composite bandage was fabricated and evaluated in detail as an alternative to existing bandages.


β-chitin hydrogel was synthesized by dissolving β-chitin powder in Methanol/CaCl2 solvent, followed by the addition of distilled water. ZnO nanoparticles were added to the β-chitin hydrogel and stirred for homogenized distribution. The resultant slurry was frozen at 0°C for 12 h. The frozen samples were lyophilized for 24 h to obtain porous composite bandages.


The bandages showed controlled swelling and degradation. The composite bandages showed blood clotting ability as well as platelet activation, which was higher when compared to the control. The antibacterial activity of the bandages were proven against Staphylococcus aureus (S. aureus) and Escherichia coli (E.coli). Cytocompatibility of the composite bandages were assessed using human dermal fibroblast cells (HDF) and these cells on the composite bandages were viable similar to the Kaltostat control bandages and bare β-chitin hydrogel based bandages. The viability was reduced to 50–60% in bandages with higher concentration of zinc oxide nanoparticles (nZnO) and showed 80–90% viability with lower concentration of nZnO. In vivo evaluation in Sprague Dawley rats (S.D. rats) showed faster healing and higher collagen deposition ability of composite bandages when compared to the control.


The prepared bandages can be used on various types of infected wounds with large volume of exudates.


antibacterial bandage nano ZnO wound healing β-chitin hydrogel 



The authors acknowledge Department of Biotechnology (DBT), India, for the financial support under a grant (BT/PR13885/MED/32/145/2010 dated 03-01-2011). We are also grateful to Nanomission, Department of Science and Technology, India, which supported this work, under a grant of the Nanoscience and Nanotechnology Initiative program. The author “R. Jayakumar” is grateful to SERC Division, Department of Science and Technology (DST), India, for providing the fund under the scheme of “Fast Track Scheme for Young Investigators” (Ref. No. SR/FT/CS-005/2008). Raja Biswas acknowledges Ramalingaswami Fellowship, Department of Biotechnology, India, for the financial support. P T Sudheesh Kumar acknowledges the Council of Scientific and Industrial Research, India for the Senior Research Fellowship (Award No. 9/963 (0011) 2K11- EMR-1).We are also grateful to Mr. Sajin P. Ravi for his help in SEM analysis. We acknowledge K. S. Sarath for his help in confocal imaging. We are grateful to Dr. P. Reshmi, Dr. A.K.K. Unni, Sajith, Sunil and Sunitha for their help during in vivo study. We thank Amrita Centre for Nanosciences and Molecular Medicine for the infrastructure support.


  1. 1.
    Albertini B, Di Sabatino M, Calonghi N, Rodriguez L, Passerini N. Novel multifunctional platforms for potential treatment of cutaneous wounds: development and in vitro characterization. Int J Pharm. 2012. doi: 10.1016/j.ijpharm.2012.06.004.
  2. 2.
    Jayakumar R, Prabaharan M, Kumar PTS, Nair SV, Tamura H. Biomaterials based on chitin and chitosan in wound dressing applications. Biotechnol Adv. 2011;29(3):322–37.PubMedCrossRefGoogle Scholar
  3. 3.
    Kim JO. Development of clindamycin-loaded wound dressing with polyvinyl alcohol and sodium alginate. Biol Pharm Bull. 2008;31(12):2277–82.PubMedCrossRefGoogle Scholar
  4. 4.
    Ong SY, Wu J, Moochhala SM, Tan MH, Lu J. Development of a chitosan-based wound dressing with improved haemostatic and antimicrobial properties. Biomaterials. 2008;29(32):4323–32.PubMedCrossRefGoogle Scholar
  5. 5.
    Willi P, Chandra PS. Chitosan and alginate wound dressings: a short review. Trends Biomater Artif Organs. 2004;18(1):18–23.Google Scholar
  6. 6.
    Balakrishnan B, Mohanty M, Umashankar PR, Jayakrishnan A. Evaluation of an in situ forming hydrogel wound dressing based on oxidized alginate and gelatin. Biomaterials. 2005;26(32):6335–42.PubMedCrossRefGoogle Scholar
  7. 7.
    Berger J, Reist M, Mayer JM, Felt O, Gurny R. Structure and interactions in chitosan hydrogels formed by complexation or aggregation for biomedical applications. Eur J Pharm Biopharm. 2004;57(1):35–52.PubMedCrossRefGoogle Scholar
  8. 8.
    Howling GI, Dettmar PW, Goddard PA, Hampson FC, Dornish M, Wood EJ. The effect of chitin and chitosan on the proliferation of human skin fibroblasts and keratinocytes in vitro. Biomaterials. 2001;22(22):2959–66.PubMedCrossRefGoogle Scholar
  9. 9.
    Hsieh CY, Tsai SP, Wang DM, Chang YN, Hsieh HJ. Preparation of gamma-PGA/chitosan composite tissue engineering matrices. Biomaterials. 2005;26(28):5617–23.PubMedCrossRefGoogle Scholar
  10. 10.
    Murakami K, Aoki H, Nakamura S, Nakamura SI, Takikawa M, Hanzawa M, et al. Hydrogel blends of chitin/chitosan, fucoidan and alginate as healing-impaired wound dressings. Biomaterials. 2010;31(1):83–90.PubMedCrossRefGoogle Scholar
  11. 11.
    Kojima K, Okamoto Y, Kojima K, Miyatake K, Fujise H, Shigemasa Y, et al. Effects of chitin and chitosan on collagen synthesis in wound healing. J Vet Med Sci. 2010;66(12):1595–8.CrossRefGoogle Scholar
  12. 12.
    Patricia MP, Joel G, Jose V, Marjana TC, Irena P, Olivera S, et al. Keratin dressings speed epithelialization of deep partial-thickness wounds. Wound Repair Regen. 2012;20(2):236–42.CrossRefGoogle Scholar
  13. 13.
    Jayakumar R, Prabaharan M, Nair SV, Tamura H. Novel chitin and chitosan nanofibers in biomedical applications. Biotechnol Adv. 2010;28(1):142–50.PubMedCrossRefGoogle Scholar
  14. 14.
    Tamura H, Furuike T, Nair SV, Jayakumar R. Biomedical applications of chitin hydrogel membranes and scaffolds. Carbohydr Polym. 2011;84(2):820–4.CrossRefGoogle Scholar
  15. 15.
    Madhumathi K, Kumar PTS, Abhilash S, Sreeja V, Tamura H, Manzoor K, et al. Development of novel chitin/nanosilver composite scaffolds for wound dressing applications. J Mater Sci Mater Med. 2010;21(2):807–13.PubMedCrossRefGoogle Scholar
  16. 16.
    Ribeiro MP, Espiga A, Silva D, Baptista P, Henriques J, Ferreira C, et al. Development of a new chitosan hydrogel for wound dressing. Wound Repair Regen. 2009;17(6):817–24.PubMedCrossRefGoogle Scholar
  17. 17.
    Jayakumar R, Chennazhi KP, Sowmya S, Nair SV, Furuike T, Tamura H. Chitin scaffolds in tissue engineering. Int J Mol Sci. 2011;12(3):1876–87.PubMedCrossRefGoogle Scholar
  18. 18.
    Chandumpai A, Singhpibulporn N, Faroongsarng D, Sornprasit P. Preparation and physico-chemical characterization of chitin and chitosan from the pens of the squid species, loligo lessoniana and loligo formosana. Carbohydr Polym. 2004;58(4):467–74.CrossRefGoogle Scholar
  19. 19.
    Mathur NK, Narang CK. Chitin and chitosan, versatile polysaccharides from marines animals. J Chem Educ. 1990;67(11):938–42.CrossRefGoogle Scholar
  20. 20.
    Mori T, Okumura M, Matsuura M, Ueno K, Tokura S, Okamoto Y, et al. Effects of chitin and its derivatives on the proliferation and cytokine production of fibroblasts in vitro. Biomaterials. 1997;18(13):947–51.PubMedCrossRefGoogle Scholar
  21. 21.
    Mi FL, Shyu SS, Peng CK, Wu YB, Sung HW, Wang PS, et al. Fabrication of chondroitin sulfate-chitosan composite artificial extracellular matrix for stabilization of fibroblast growth factor. J Biomed Mater Res A. 2006;76(1):1–15.PubMedGoogle Scholar
  22. 22.
    Cho YW, Cho YN, Chung SH, Yoo G, Ko SW. Water-soluble chitin as a wound healing accelerator. Biomaterials. 1999;20(22):2139–45.PubMedCrossRefGoogle Scholar
  23. 23.
    Fan Y, Saito T, Isogai A. Preparation of chitin nanofibers from squid pen beta-chitin by simple mechanical treatment under acid conditions. Biomacromolecules. 2008;9(7):1919–23.PubMedCrossRefGoogle Scholar
  24. 24.
    Kumar PTS, Abhilash S, Manzoor K, Nair SV, Tamura H, Jayakumar R. Preparation and characterization of novel β-chitin/nano silver composite scaffolds for wound dressing applications. Carbohydr Polym. 2010;80(3):761–7.CrossRefGoogle Scholar
  25. 25.
    Gardner KH, Blackwell J. Refinement of structure of beta chitin. Biopolymers. 1975;14(8):1581–95.PubMedCrossRefGoogle Scholar
  26. 26.
    Ratanajiajaroen P, Ohshima M. Preparation of highly porous β-chitin structure through nonsolvent-solvent exchange-induced phase separation and supercritical CO2 drying. J Supercrit Fluids. 2012;68(1):31–8.CrossRefGoogle Scholar
  27. 27.
    Ratanajiajaroen P, Watthanaphanit A, Tamura H, Tokura S, Rujiravanit R. Release characteristic and stability of curcumin incorporated in β-chitin non-woven fibrous sheet using Tween 20 as an emulsifier. Eur Polym J. 2012;48(3):512–23.CrossRefGoogle Scholar
  28. 28.
    Ehrlich H. Chitin and collagen as universal and alternative templates in biomineralization. Int Geol Rev. 2010;52(7):661–9.CrossRefGoogle Scholar
  29. 29.
    Ehrlich H. Biological materials of marine origin: invertebrates. The Netherlands: Springer Verlag; 2010. p. 594.CrossRefGoogle Scholar
  30. 30.
    Maeda Y, Jayakumar R, Nagahama H, Furuike T, Tamura H. Synthesis, characterization and bioactivity studies of novel beta-chitin scaffold for tissue engineering applications. Int J Biol Macromol. 2008;42(5):463–7.PubMedCrossRefGoogle Scholar
  31. 31.
    Saito Y, Okano T, Gaill F, Chanzy H, Putaux JL. Structural data on the intra-crystalline swelling of beta-chitin. Int J Biol Macromol. 2000;28(1):81–8.PubMedCrossRefGoogle Scholar
  32. 32.
    Sharma V, Singh SK, Anderson D, Tobin DJ, Dhwan A. Zinc oxide nanoparticles induce genotoxicity in primary human epidermal keratinocytes. J Nanosci Nanotechnol. 2011;11(5):3782–8.PubMedCrossRefGoogle Scholar
  33. 33.
    Kocbek P, Teskac K, Kreft ME, Kristl J. Toxicological aspects of long-term treatment of keratinocytes with ZnO and TiO2 nanoparticles. Small. 2010;6(17):1908–17.PubMedCrossRefGoogle Scholar
  34. 34.
    Ng KW, Khoo SP, Heng BC, et al. The role of the tumor suppressor p53 pathway in the cellular DNA damage response to zinc oxide nanoparticles. Biomaterials. 2011;32(32):8218–25.PubMedCrossRefGoogle Scholar
  35. 35.
    Hackenberg S, Scherzed A, Kessler M, et al. Zinc oxide nanoparticles induce photocatalytic cell death in human head and neck squamous cell carcinoma cell lines in vitro. Int J Oncol. 2010;37(6):1583–90.PubMedGoogle Scholar
  36. 36.
    Nair SV, Abhilash S, Divya RVV, Deepthy M, Seema N, Manzoor K, et al. Role of size scale of ZnO nanoparticles and microparticles on toxicity toward bacteria and osteoblast cancer cells. J Mater Sci Mater Med. 2009;20(1):235–41.CrossRefGoogle Scholar
  37. 37.
    Monteiro RNA, Wiench K, Landsiedel R, Schulte S, Inman AO, Riviere JE. Safety evaluation of sunscreen formulations containing titanium dioxide and zinc oxide nanoparticles in UVB sunburned skin: an in vitro and in vivo study. Toxicol Sci. 2011;123(1):264–80.CrossRefGoogle Scholar
  38. 38.
    Stephan H, Norbert K. Dermal toxicity of ZnO nanoparticles: a worrying feature of sunscreen. Nanomedicine. 2012;7(4):461–3.CrossRefGoogle Scholar
  39. 39.
    Kumar PTS, Lakshmanan VK, Anilkumar TV, Ramya C, Reshmi P, Unnikrishnan AG, et al. Flexible and microporous chitosan hydrogel/nano ZnO composite bandages for wound dressing: in vitro and in vivo evaluation. ACS Appl Mater Interfaces. 2012;4(5):2618–29.PubMedCrossRefGoogle Scholar
  40. 40.
    Becheri A, Durr M, Nostro PL, Baglioni P. Synthesis and characterization of zinc oxide nanoparticles: application to textiles as UV-absorbers. J Nanopart Res. 2008;10(4):679–89.CrossRefGoogle Scholar
  41. 41.
    Abhilash S, Parwathy C, Deepthy M, Sreerekha PR, Nair SV, Manzoor K. Rapid dissolution of ZnO nanocrystals in acidic cancer microenvironment leading to preferential apoptosis. Nanoscale. 2011;3(9):3657–69.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2012

Authors and Affiliations

  • Sudheesh Kumar P. T.
    • 1
  • Vinoth-Kumar Lakshmanan
    • 1
  • Mincy Raj
    • 1
  • Raja Biswas
    • 1
  • Tamura Hiroshi
    • 2
  • Shantikumar V. Nair
    • 1
  • Rangasamy Jayakumar
    • 1
    Email author
  1. 1.Amrita Centre for Nanosciences and Molecular Medicine Amrita Institute of Medical Sciences and Research CentreAmrita Vishwa Vidyapeetham UniversityKochiIndia
  2. 2.Faculty of Chemistry, Materials and BioengineeringKansai UniversityOsakaJapan

Personalised recommendations