Synergistic effect of crystalline phase on protein adsorption and cell behaviors on TiO2 nanotubes

  • Yanran Li
  • Yuanjun Dong
  • Yanmei Zhang
  • Yun Yang
  • Ren Hu
  • Ping Mu
  • Xiangyang Liu
  • Changjian Lin
  • Qiaoling HuangEmail author
Original Article


The objective of this study is to explore the structure–property relationships of TiO2 nanotubes (TNTs) with different crystalline phases that link to protein adsorption and cell responses. Given the formation of intact rutile nanotubular structures by furnace annealing is challenge, a combination of furnace annealing and flame annealing is employed for the preparation of rutile TNTs. TNTs with pure anatase phase and mixed anatase/rutile phases are obtained by simple furnace annealing of amorphous TNTs. Results show that BSA and FBS adsorptions are greatly enhanced on rutile TNTs, whereas no discernable difference on other crystalline phases. Rutile TNTs also present highest adsorption of fibronectin and collagen which are diminished on anatase and dual anatase–rutile phases. Interestingly, however, there is no significant difference in cell proliferation or differentiation on TNTs with different crystallites. Scrutinization of the surface properties involved in protein adsorption and cell activities, a synergistic effect of surface charge, hydroxyl groups, and roughness is found on protein adsorption which further regulates cell behaviors. Those findings provide a better understanding of the structure–property relationships of titanium-based biomaterials.


TiO2 nanotubes Crystalline phase Protein adsorption Cell behaviors 



Authors gratefully acknowledge the financial supports from the State Key Project of Research and Development (2016YFC1100300), National Natural Science Foundation of China (51571169, 21773199), Natural Science Foundation of Guangdong Province, China (2016A030310370), and 111 Project (B16029). The authors would like to thank Shengshi Guo, Rui Yu, Hao Wang, and Likun Yang for their technical supports.

Compliance with ethical standards

Conflict of interest

The authors report no conflicts of interest.


  1. An SH, Narayanan R, Matsumoto T, Lee HJ, Kwon TY, Kim KH (2011) Crystallinity of anodic TiO2 nanotubes and bioactivity. J Nanosci Nanotechnol 11:4910–4918. CrossRefGoogle Scholar
  2. Awad NK, Edwards SL, Morsi YS (2017) A review of TiO2 NTs on Ti metal: electrochemical synthesis, functionalization and potential use as bone implants. Mater Sci Eng C 76:1401–1412. CrossRefGoogle Scholar
  3. Bai Y, Park S, Park HH, Lee MH, Bae TS, Duncan W, Swain A (2011) The effect of annealing temperatures on surface properties, hydroxyapatite growth and cell behaviors of TiO2 nanotubes. Surf Interface Anal 43:998–1005. CrossRefGoogle Scholar
  4. Barthes J et al (2018) Review: the potential impact of surface crystalline states of titanium for biomedical applications. Crit Rev Biotechnol 38:423–437. CrossRefGoogle Scholar
  5. Cheng K, Hong Y, Yu M, Lin J, Weng W, Wang H (2015) Modulation of protein behavior through light responses of TiO2 nanodots films. Sci Rep 5:13354. CrossRefGoogle Scholar
  6. Firkowska-Boden I, Zhang X, Jandt KD (2018) Controlling protein adsorption through nanostructured polymeric surfaces. Adv Healthc Mater 7:1700995. CrossRefGoogle Scholar
  7. Formentín P, Catalán Ú, Pol L, Fernández-Castillejo S, Solà R, Marsal LF (2018) Collagen and fibronectin surface modification of nanoporous anodic alumina and macroporous silicon for endothelial cell cultures. J Biol Eng 12:21. CrossRefGoogle Scholar
  8. Grainger DW (2013) All charged up about implanted biomaterials. Nat Biotechnol 31:507–509. CrossRefGoogle Scholar
  9. Highberger JH (1939) The isoelectric point of collagen. J Am Chem Soc 61:2302–2303. CrossRefGoogle Scholar
  10. Hong F, Ni YH, Xu WJ, Yan YF (2012) Origin of enhanced water adsorption at 10 step edge on rutile TiO2 (110) surface. J Chem Phys 137:114707. (114701–114709) CrossRefGoogle Scholar
  11. Hong Y, Yu M, Lin J, Cheng K, Weng W, Wang H (2014) Surface hydroxyl groups direct cellular response on amorphous and anatase TiO2 nanodots. Colloids Surf B 123:68–74. CrossRefGoogle Scholar
  12. Hoshiba T, Yoshikawa C, Sakakibara K (2018) Characterization of initial cell adhesion on charged polymer substrates in serum-containing and serum-free media. Langmuir 34:4043–4051. CrossRefGoogle Scholar
  13. Huang Q et al (2017) Effect of construction of TiO2 nanotubes on platelet behaviors: structure-property relationships. Acta Biomater 51:505–512. CrossRefGoogle Scholar
  14. Jimbo R, Ivarsson M, Koskela A, Sul Y-T, Johansson CB (2010) Protein adsorption to surface chemistry and crystal structure modification of titanium surfaces. J Oral Maxillofac Res 1:1–9. CrossRefGoogle Scholar
  15. Jin GD et al (2015) Zn/Ag micro-galvanic couples formed on titanium and osseointegration effects in the presence of S. aureus. Biomaterials 65:22–31. CrossRefGoogle Scholar
  16. Kar P, Zhang Y, Farsinezhad S, Mohammadpour A, Wiltshire BD, Sharma H, Shankar K (2015) Rutile phase n- and p-type anodic titania nanotube arrays with square-shaped pore morphologies. Chem Commun 51:7816–7819. CrossRefGoogle Scholar
  17. Kulkarni M et al (2015a) Titanium nanostructures for biomedical applications. Nanotechnology 26:062002. CrossRefGoogle Scholar
  18. Kulkarni M, Patil-Sen Y, Junkar I, Kulkarni CV, Lorenzetti M, Iglic A (2015b) Wettability studies of topologically distinct titanium surfaces. Colloids Surf B 129:47–53. CrossRefGoogle Scholar
  19. Kulkarni M et al (2017) Interaction of nanostructured TiO2 biointerfaces with stern cells and biofilm-forming bacteria. Mater Sci Eng C 77:500–507. CrossRefGoogle Scholar
  20. Lai Y, Pan F, Xu C, Fuchs H, Chi L (2013) In situ surface-modification-induced superhydrophobic patterns with reversible wettability and adhesion. Adv Mater 25:1682–1686. CrossRefGoogle Scholar
  21. Lai Y et al (2018) Progress in TiO2 nanotube coatings for biomedical applications: a review. J Mater Chem B 6:1862. CrossRefGoogle Scholar
  22. Li H et al (2015a) Multifunctional wettability patterns prepared by laser processing on superhydrophobic TiO2 nanostructured surfaces. J Mater Chem B 3:342–347. CrossRefGoogle Scholar
  23. Li M, Liu Q, Jia Z, Xu X, Shi Y, Cheng Y, Zheng Y (2015b) Polydopamine-induced nanocomposite Ag/CaP coatings on the surface of titania nanotubes for antibacterial and osteointegration functions. J Mater Chem B 3:8796–8805. CrossRefGoogle Scholar
  24. Li GL et al (2016) Enhanced osseointegration of hierarchical micro/nanotopographic titanium fabricated by microarc oxidation and electrochemical treatment. ACS Appl Mater Inter 8:3840–3852. CrossRefGoogle Scholar
  25. Lv L et al (2015) The nanoscale geometry of TiO2 nanotubes influences the osteogenic differentiation of human adipose-derived stem cells by modulating H3K4 trimethylation. Biomaterials 39:193–205. CrossRefGoogle Scholar
  26. Lv L, Li K, Xie Y, Cao Y, Zheng X (2017) Enhanced osteogenic activity of anatase TiO2 film: surface hydroxyl groups induce conformational changes in fibronectin. Mater Sci Eng, C 78:96–104. CrossRefGoogle Scholar
  27. Maitz MF, Pham MT, Wieser E (2003) Blood compatibility of titanium oxides with various crystal structure and element doping. J Biomater Appl 17:303–319. CrossRefGoogle Scholar
  28. Mazare A, Paramasivam I, Schmidt-Stein F, Lee K, Demetrescu I, Schmuki P (2012) Flame annealing effects on self-organized TiO2 nanotubes. Electrochim Acta 66:12–21. CrossRefGoogle Scholar
  29. Oh S, Daraio C, Chen LH, Pisanic TR, Finones RR, Jin S (2006) Significantly accelerated osteoblast cell growth on aligned TiO2 nanotubes. J Biomed Mater Res A 78A:97–103. CrossRefGoogle Scholar
  30. Oh S, Brammer KS, Li YSJ, Teng D, Engler AJ, Chien S, Jin S (2009) Stem cell fate dictated solely by altered nanotube dimension. Proc Natl Acad Sci USA 106:2130–2135. CrossRefGoogle Scholar
  31. Othman Z, Cillero Pastor B, van Rijt S, Habibovic P (2018) Understanding interactions between biomaterials and biological systems using proteomics. Biomaterials 167:191–204. CrossRefGoogle Scholar
  32. Park J, Bauer S, Schmuki P, von der Mark K (2009) Narrow window in nanoscale dependent activation of endothelial cell growth and differentiation on TiO2 nanotube surfaces. Nano Lett 9:3157–3164. CrossRefGoogle Scholar
  33. Pittrof A, Park J, Bauer S, Schmuki P (2012) ECM spreading behaviour on micropatterned TiO2 nanotube surfaces. Acta Biomater 8:2639–2647. CrossRefGoogle Scholar
  34. Rahmati M, Mozafari M (2018) Protein adsorption on polymers. Mater Today Commun 17:527–540. CrossRefGoogle Scholar
  35. Rezaee M, Khoie SMM, Liu KH (2011) The role of brookite in mechanical activation of anatase-to-rutile transformation of nanocrystalline TiO2: an XRD and Raman spectroscopy investigation. CrystEngComm 13:5055–5061. CrossRefGoogle Scholar
  36. Spriano S, Yamaguchi S, Baino F, Ferraris S (2018) A critical review of multifunctional titanium surfaces: new frontiers for improving osseointegration and host response, avoiding bacteria contamination. Acta Biomater 79:1–22. CrossRefGoogle Scholar
  37. Su EP, Justin DF, Pratt CR, Sarin VK, Nguyen VS, Oh S, Jin S (2018) Effects of titanium nanotubes on the osseointegration, cell differentiation, mineralisation and antibacterial properties of orthopaedic implant surfaces. Bone Jt J 100-B:9–16. CrossRefGoogle Scholar
  38. Takemoto S, Yamamoto T, Tsuru K, Hayakawa S, Osaka A, Takashima S (2004) Platelet adhesion on titanium oxide gels: effect of surface oxidation. Biomaterials 25:3485–3492CrossRefGoogle Scholar
  39. Varghese OK, Gong DW, Paulose M, Grimes CA, Dickey EC (2003) Crystallization and high-temperature structural stability of titanium oxide nanotube arrays. J Mater Res 18:156–165. CrossRefGoogle Scholar
  40. Vroman L, Adams AL, Fischer GC, Munoz PC (1980) Interaction of high molecular weight kininogen, factor XII, and fibrinogen in plasma at interfaces. Blood 55:156–159Google Scholar
  41. Yu WQ, Zhang YL, Jiang XQ, Zhang FQ (2010) In vitro behavior of MC3T3-E1 preosteoblast with different annealing temperature titania nanotubes. Oral Dis 16:624–630. CrossRefGoogle Scholar
  42. Yu Y et al (2018) Osteogenesis potential of different titania nanotubes in oxidative stress microenvironment. Biomaterials 167:44–57. CrossRefGoogle Scholar
  43. Yuan Z et al (2018) Investigation of osteogenic responses of Fe-incorporated micro/nano-hierarchical structures on titanium surfaces. J Mater Chem B 6:1359–1372. CrossRefGoogle Scholar
  44. Zhang L, Liao X, Fok A, Ning C, Ng P, Wang Y (2018) Effect of crystalline phase changes in titania (TiO2) nanotube coatings on platelet adhesion and activation. Mater Sci Eng, C 82:91–101. CrossRefGoogle Scholar
  45. Zhao L, Liu L, Wu Z, Zhang Y, Chu PK (2012) Effects of micropitted/nanotubular titania topographies on bone mesenchymal stem cell osteogenic differentiation. Biomaterials 33:2629–2641. CrossRefGoogle Scholar

Copyright information

© King Abdulaziz City for Science and Technology 2019

Authors and Affiliations

  1. 1.Fujian Provincial Key Laboratory for Soft Functional Materials Research, Department of Physics and Jiujiang Research Institute, College of Physical Science and Technology, Research Institute for Biomimetics and Soft MatterXiamen UniversityXiamenChina
  2. 2.Shenzhen Research Institute of Xiamen UniversityShenzhenChina
  3. 3.State Key Laboratory for Physical Chemistry of Solid Surfaces, Department of Chemistry, College of Chemistry and Chemical EngineeringXiamen UniversityXiamenChina
  4. 4.Department of PhysicsNational University of SingaporeSingaporeSingapore

Personalised recommendations