Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK

Purposes: To study the effect of hydrothermal treatment on cytotoxicity of Titanium Nitride - Hydroxyapatite (TiN-HA) coated on Polyetheretherketone (PEEK)

Materials and Methods: Twelve pieces of PEEK, TiN-HA coated PEEK and TiN-HA coated PEEK with hydrothermal treatment were prepared. The toxicity tests on L929 cell were done by indirect contact method at 24 hours, 7 days, 14 days and 35 days. MTT assay was performed to evaluate cell viability. Cellular morphology was evaluated using a phase-contrast microscope. The surface morphology of coated films was also assessed by scanning electron microscopy and atomic force microscopy.

Results: PEEK, and TiN-HA coated PEEK did not have cytotoxicity effect on fibroblast cells both prior- and after- hydrothermal treatment groups. The cell viability in the same material condition was increased when duration increased and fibroblasts showed an elongated morphology and a good congruence. Additionally, coated hydroxyapatite increased material surface roughness.

Conclusions: Hydrothermal treatment of TiN-HA coated films were not cytotoxic to fibroblasts and surface roughness was performed by hydroxyapatite in the coated layer.

1. Skirbutis G, Dzingutė A, Masiliūnaitė V, Šulcaitė G, Žilinskas JJS. A review of PEEK polymer's properties and its use in prosthodontics. Stomatologija 2017; 19(1): 19-23.

2. Kurtz SM, Devine JN. PEEK biomaterials in trauma, orthopedic, and spinal implants. Biomaterials 2007; 28(32): 4845-4869.

3. Toth JM, Wang M, Estes BT, Scifert JL, Seim HB, Turner AS.Polyetheretherketone as a biomaterial for spinal applications. Biomaterials 2006; 27(3): 324-334.

4. Rabiei A, Sandukas S. Processing and evaluation of bioactive coatings on polymeric implants.J Biomed Mater Res A 2013; 101(9): 2621-2629.

5. Murphy W, Black J, Hastings G. Handbook of biomaterial properties. 2nd ed. New York: Springer; 2016.

6. Ma R, Tang T. Current strategies to improve the bioactivity of PEEK. Int J Mol Sci 2014; 15(4): 5426-5445.

7. Wang H, Xu M, Zhang W, Kwok DTK, Jiang J, Wu Z, et al. Mechanical and biological characteristics of diamond-like carbon coated poly aryl-ether-ether-ketone. Biomaterials 2010; 31(32): 8181-8187.

8. Almasi D, Iqbal N, Sadeghi M, Sudin I, Abdul Kadir MR, Kamarul T. Preparation Methods for Improving PEEK's Bioactivity for Orthopedic and Dental Application: A Review. Int J Biomater 2016; article ID 8202653, 12 pages.

9. Hench. LL. An introduction to bioceramics. 2nd ed. London: Imperial College Press; 2013.

10. Tang SM, Cheang P, AbuBakar MS, Khor KA, Liao K. Tension–tension fatigue behavior of hydroxyapatite reinforced polyetheretherketone composites. Int J Fatigue 2004; 26(1): 49-57.

11.Sawase T, Yoshida K, Taira Y, Kamada K, Atsuta M, Baba K. Abrasion resistance of titanium nitride coatings formed on titanium by ion‐beam‐assisted deposition. J Oral Rehabil 2005; 32(2): 151-157.

12. Boonyawan D, Waruriya P, Suttiat K. Characterization of titanium nitride–hydroxyapatite on PEEK for dental implants by co-axis target magnetron sputtering. Surf Coat Technol 2016; 306: 164-170.

13. Shi X, Xu L, Le TB, Zhou G, Zheng C, Tsuru K, et al. Partial oxidation of TiN coating by hydrothermal treatment and ozone treatment to improve its osteoconductivity. Mater Sci Eng C 2016; 59: 542-548.


14. Piscanec S, Ciacchi LC, Vesselli E, et al. Bioactivity of TiN-coated titanium implants. Acta Mater 2004; 52(5): 1237-1245.

15. Ha S-W, Mayer J, Koch B, Wintermantel E. Plasma-sprayed hydroxylapatite coating on carbon fibre reinforced thermoplastic composite materials. J Mater Sci Mater Med 1994; 5(6): 481-484.

16. Faghihi-Sani M-A, Arbabi A, Mehdinezhad-Roshan A. Crystallization of hydroxyapatite during hydrothermal treatment on amorphous calcium phosphate layer coated by PEO technique. Ceram Int 2013; 39(2): 1793-1798.

17. Wang BC, Chang E, Lee TM, Yang CY. Changes in phases and crystallinity of plasma-sprayed hydroxyapatite coatings under heat treatment: A quantitative study. J Biomed Mater Res 1995; 29(12): 1483-1492.

18. Yang CW, Lui TS, Chang E. Low temperature crystallization and structural modification of plasma-sprayed hydroxyapatite coating with hydrothermal treatment. Adv Mat Res 2007; 15: 147-152.

19. Yang Y, Kim KH, Ong JL. A review on calcium phosphate coatings produced using a sputtering process: an alternative to plasma spraying. Biomaterials 2005; 26(3): 327-337.

20. Huang Y, Song L, Liu X, et al. Hydroxyapatite coatings deposited by liquid precursor plasma spraying: controlled dense and porous microstructures and osteoblastic cell responses. Biofabrication 2010; 2(4): 045003.

21. Wang H, Zhi W, Lu X, et al. Comparative studies on ectopic bone formation in porous hydroxyapatite scaffolds with complementary pore structures. Acta Biomaterialia 2013; 9(9): 8413-8421.

22. Chang BS, Lee iCKfC-K, Hong KS, et al. Osteoconduction at porous hydroxyapatite with various pore configurations. Biomaterials 2000; 21(12): 1291-1298.23. Hing KA. Bioceramic Bone Graft Substitutes: Influence of Porosity and Chemistry. Int J Appl Ceram Technol 2005; 2(3): 184-199.

24. Han CM, Lee EJ, Kim HE, et al. The electron beam deposition of titanium on polyetheretherketone (PEEK) and the resulting enhanced biological properties. Biomaterials 2010; 31(13): 3465-3470.

25. Hunter A, Archer CW, Walker PS, Blunn GW. Attachment and proliferation of osteoblasts and fibroblasts on biomaterials for orthopaedic use. Biomaterials 1995; 16(4): 287-295.


26. Nupangtha W, Boonyawan D. Fabrication and Physical Properties of Titanium Nitride/Hydroxyapatite Composites on Polyether Ether Ketone by RF Magnetron Sputtering Technique. Journal of Physics: Conference Series 2017; 901(1): 012131.

27. Thai industrial standard: TISI. Biologic evaluation of medical device (Part 5: Test for in vitro cytotoxicity ISO 10993-5). 3rd ed. Rajathevee Bangkok; 2009: 5.

28. Thai industrial standard: TISI. Biologic evaluation of medical device (Part 12: Sample preparation and reference materials ISO 10993-12). 3rd ed. Rajathevee Bangkok; 2007: 7.

29. Geurtsen W, Leyhausen G. Biological aspects of root canal filling materials–histocompatibility, cytotoxicity, and mutagenicity. Clin Oral Investig 1997; 1(1): 5-11.

30. Cenni E, Ciapetti G, Granchi D, Arciola CR, et al. Established Cell Lines and Primary Cultures in Testing Medical Devices In Vitro. Toxicol In Vitro 1999; 13(4): 801-810.

31. Shi X, Xu L, Munar ML, Ishikawa K. Hydrothermal treatment for TiN as abrasion resistant dental implant coating and its fibroblast response. Mater Sci Eng C 2015; 49: 1-6.

32. Chou L, Marek B, Wagner WR. Effects of hydroxylapatite coating crystallinity on biosolubility, cell attachment efficiency and proliferation in vitro. Biomaterials 1999; 20(10): 977-985.

33. Jarcho M, Kay JF, Gumaer KI, Doremus RH, Drobeck HP. Tissue, cellular and subcellular events at a bone-ceramic hydroxylapatite interface. J Bioeng 1977; 1(2): 79-92.

34. Yuan H, Li Y, de Bruijn JD, de Groot K, Zhang X. Tissue responses of calcium phosphate cement: a study in dogs. Biomaterials 2000; 21(12): 1283-1290.

35. Weng J, Liu Q, Wolke JGC, Zhang X, de Groot K. Formation and characteristics of the apatite layer on plasma-sprayed hydroxyapatite coatings in simulated body fluid. Biomaterials 1997; 18(15): 1027-1035.

36. Porter AE, Patel N, Skepper JN, Best SM, Bonfield W. Comparison of in vivo dissolution processes in hydroxyapatite and silicon-substituted hydroxyapatite bioceramics. Biomaterials 2003; 24(25): 4609-4620.

37. Sun L, Berndt CC, Gross KA, Kucuk A. Material fundamentals and clinical performance of plasma-sprayed hydroxyapatite coatings: A review. J Biomed Mater Res B Appl Biomater 2001; 58(5): 570-592.

38. Nakasato Y, Takebe J. Analysis of Thin Hydroxyapatite Layers Formed on Anodic Oxide Titanium after Hydrothermal Treatment in Rat Bone Marrow Cell Culture. Prosthodontic Research & Practice 2005; 4(1): 32-41.

39. Massaro C, Baker MA, Cosentino F, Ramires PA, Klose S, Milella E. Surface and biological evaluation of hydroxyapatite-based coatings on titanium deposited by different techniques. J Biomed Mater Res 2001; 58(6): 651-657.

40. van Dijk K, Schaeken HG, Wolke JGC, Jansen JA. Influence of annealing temperature on RF magnetron sputtered calcium phosphate coatings. Biomaterials 1996; 17(4): 405-410.

41. Paital SR, Dahotre NB. Calcium phosphate coatings for bio-implant applications: Materials, performance factors, and methodologies. Mater Sci Eng R Rep 2009; 66(1): 1-70.

42. Gittens RA, Olivares-Navarrete R, Schwartz Z, Boyan BD. Implant osseointegration and the role of microroughness and nanostructures: Lessons for spine implants. Acta Biomater 2014; 10(8): 3363-3371.

43. Ponche A, Bigerelle M, Anselme K. Relative influence of surface topography and surface chemistry on cell response to bone implant materials. Part 1: physico-chemical effects. Proc Inst Mech Eng H 2010; 224(12): 1471-1486.

44.Zhu X, Son DW, Ong JL, Kim KH. Characterization of hydrothermally treated anodic oxides containing Ca and P on titanium. J Mater Sci Mater Med 2003; 14(7): 629-634.


45. Albrektsson T, Wennerberg A. Oral implant surfaces: Part 1-review focusing on topographic and chemical properties of different surfaces and in vivo responses to them. Int J Prosthodont 2004; 17(5): 536-543.

46. Karageorgiou V, Kaplan D. Porosity of 3D biomaterial scaffolds and osteogenesis. Biomaterials 2005 Sep; 26(27): 5474-5491.

47. Ducheyne P, Radin S, King L. The effect of calcium phosphate ceramic composition and structure on in vitro behavior. I. Dissolution. J Biomed Mater Res 1993; 27(1): 25-34.

48. Radin SR, Ducheyne P. The effect of calcium phosphate ceramic composition and structure on in vitro behavior. II. Precipitation. J Biomed Mater Res 1993; 27(1): 35-45.

Buranapanich V, Suttiat K, Wanachantararak P, Yavirach P. Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK: Original articles. CM Dent J [Internet]. 2020 Sep 01 [cited 2024 May 09];41(3):135-178. Available from: https://www.dent.cmu.ac.th/cmdj/frontend/web/?r=site/viewarticle&id=73

Buranapanich, V., Suttiat, K., Wanachantararak, P. & Yavirach, P. (2020). Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK. CM Dent J, 41(3), 135-178. Retrieved from: https://www.dent.cmu.ac.th/cmdj/frontend/web/?r=site/viewarticle&id=73

Buranapanich, V., Suttiat Kullapop,Wanachantararak Phenphichar and Yavirach Piriya . 2020. "Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK." CM Dent J, 41(3), 135-178. https://www.dent.cmu.ac.th/cmdj/frontend/web/?r=site/viewarticle&id=73

Buranapanich, V. et al. 2020. 'Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK', CM Dent J, 41(3), 135-178. Retrieved from https://www.dent.cmu.ac.th/cmdj/frontend/web/?r=site/viewarticle&id=73

Buranapanich, V., Suttiat, K., Wanachantararak, P. and Yavirach, P. "Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK", CM Dent J, vol.41, no. 3, pp. 135-178, Sep. 2020.

Buranapanich, V., Suttiat, K., Wanachantararak, P., et al. "Effect of Hydrothermal Treatment on Cytotoxicity of Titanium Nitride-hydroxyapatite Coated PEEK." CM Dent J, vol.41, no. 3, Sep. 2020, pp. 135-178, https://www.dent.cmu.ac.th/cmdj/frontend/web/?r=site/viewarticle&id=73