Journal of Materials Science

, Volume 53, Issue 8, pp 5891–5908 | Cite as

Antibiotic peptide-modified nanostructured titanium surface for enhancing bactericidal property

  • Chen Zhu
  • Wei-wei Zhang
  • Shi-yuan Fang
  • Rong Kong
  • Gang Zou
  • Ni-Rong Bao
  • Jian-Ning Zhao
  • Xi-Fu Shang


The infections associated with titanium-based biomaterials have been one of the most serious postoperative complications in the orthopedic surgery. Great efforts have been made to improve the antimicrobial property of titanium-based biomaterials by virtue of the surface modification strategy. From the biomimetic perspective of vegetation roots anchoring soil, alkali treatment was conducted on metallic titanium to produce a nanoroot-structured surface in the present study; then, antimicrobial peptide was anchored within the nanoroot surface by vacuum extraction and lyophilization. As a result, the obtained antibacterial peptide-leashed titanium surface showed a hierarchical structure combining the designed nanoroot topography and the anchored antibiotic peptide. Furthermore, this modified surface could steadily release for more than 10 h in a time-dependent manner. As a consequence, the elaborate antimicrobial peptide-loaded surface demonstrated a powerful antibacterial and biofilm-resistant capability against two types of Staphylococcus, without significant cytotoxicity. Specifically, Peptide-2 can kill the most planktonic and sessile bacteria for two gram-positive bacteria. Therefore, the integration of antibacterial peptide onto titanium-based implant surface may be a hopeful tool to prevent implant-associated infections in the orthopedic surgery.



This work was supported by the National Natural Science Foundation of China (Grant No. 81401815), the China Postdoctoral Science Foundation (Grant No. 2015M582900) and the Jiangsu Postdoctoral Science Foundation (Grant No. 1501146C).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Huo K, Gao B, Fu J, Zhao L, Chu PK (2014) Fabrication, modification, and biomedical applications of anodized TiO2 nanotube arrays. RSC Adv 4:17300–17324CrossRefGoogle Scholar
  2. 2.
    Yang W-E, Hsu M-L, Lin M-C, Chen Z-H, Chen L-K, Huang H-H (2009) Nano/submicron-scale TiO2 network on titanium surface for dental implant application. J Alloys Compd 479:642–647CrossRefGoogle Scholar
  3. 3.
    Krzakala A, Kazek-Kesik A, Simka W (2013) Application of plasma electrolytic oxidation to bioactive surface formation on titanium and its alloys. RSC Adv 3:19725–19743CrossRefGoogle Scholar
  4. 4.
    Geetha M, Singh AK, Asokamani R, Gogia AK (2009) Ti based biomaterials, the ultimate choice for orthopaedic implants—a review. Prog Mater Sci 54:397–425CrossRefGoogle Scholar
  5. 5.
    Darouiche RO (2004) Treatment of infections associated with surgical implants. N Engl J Med 350:1422–1429CrossRefGoogle Scholar
  6. 6.
    Hetrick EM, Schoenfisch MH (2006) Reducing implant-related infections: active release strategies. Chem Soc Rev 35:780–789CrossRefGoogle Scholar
  7. 7.
    Huang Y, Zha G, Luo Q et al (2014) The construction of hierarchical structure on Ti substrate with superior osteogenic activity and intrinsic antibacterial capability. Sci Rep 4:6172CrossRefGoogle Scholar
  8. 8.
    Costerton JW, Stewart PS, Greenberg EP (1999) Bacterial biofilms: a common cause of persistent infections. Science 284:1318–1322CrossRefGoogle Scholar
  9. 9.
    Montanaro L, Speziale P, Campoccia D et al (2011) Scenery of Staphylococcus implant infections in orthopedics. Future Microbiol 6:1329–1349CrossRefGoogle Scholar
  10. 10.
    Garrett TR, Bhakoo M, Zhang Z (2008) Bacterial adhesion and biofilms on surfaces. Prog Nat Sci 18:1049–1056CrossRefGoogle Scholar
  11. 11.
    Subbiahdoss G, Kuijer R, Grijpma DW, van der Mei HC, Busscher HJ (2009) Microbial biofilm growth vs. tissue integration: “the race for the surface” experimentally studied. Acta Biomater 5:1399–1404CrossRefGoogle Scholar
  12. 12.
    Cunha A, Elie A-M, Plawinski L et al (2016) Femtosecond laser surface texturing of titanium as a method to reduce the adhesion of Staphylococcus aureus and biofilm formation. Appl Surf Sci 360:485–493CrossRefGoogle Scholar
  13. 13.
    Simchi A, Tamjid E, Pishbin F, Boccaccini AR (2011) Recent progress in inorganic and composite coatings with bactericidal capability for orthopaedic applications. Nanomed Nanotechnol Biol Med 7:22–39CrossRefGoogle Scholar
  14. 14.
    Campoccia D, Montanaro L, Arciola CR (2013) A review of the biomaterials technologies for infection-resistant surfaces. Biomaterials 34:8533–8554CrossRefGoogle Scholar
  15. 15.
    Antoci V Jr, King SB, Jose B et al (2007) Vancomycin covalently bonded to titanium alloy prevents bacterial colonization. J Orthop Res 25:858–866CrossRefGoogle Scholar
  16. 16.
    Romano A, Mayorga C, Torres MJ et al (2000) Immediate allergic reactions to cephalosporins: cross-reactivity and selective responses. J Allergy Clin Immunol 106:1177–1183CrossRefGoogle Scholar
  17. 17.
    Sharma R, Sharma CL, Kapoor B (2005) Antibacterial resistance: current problems and possible solutions. Indian J Med Sci 59:120–129CrossRefGoogle Scholar
  18. 18.
    Necula BS, Fratila-Apachitei LE, Zaat SA, Apachitei I, Duszczyk J (2009) In vitro antibacterial activity of porous TiO2–Ag composite layers against methicillin-resistant Staphylococcus aureus. Acta Biomater 5:3573–3580CrossRefGoogle Scholar
  19. 19.
    Hang R, Gao A, Huang X et al (2014) Antibacterial activity and cytocompatibility of Cu–Ti–O nanotubes. J Biomed Mater Res Part A 102:1850–1858CrossRefGoogle Scholar
  20. 20.
    Foldbjerg R, Dang DA, Autrup H (2011) Cytotoxicity and genotoxicity of silver nanoparticles in the human lung cancer cell line, A549. Arch Toxicol 85:743–750CrossRefGoogle Scholar
  21. 21.
    Hancock RE (2005) Mechanisms of action of newer antibiotics for Gram-positive pathogens. Lancet Infect Dis 5:209–218CrossRefGoogle Scholar
  22. 22.
    Li B, Jiang B, Boyce BM, Lindsey BA (2009) Multilayer polypeptide nanoscale coatings incorporating IL-12 for the prevention of biomedical device-associated infections. Biomaterials 30:2552–2558CrossRefGoogle Scholar
  23. 23.
    Liu X, Chu PK, Ding C (2010) Surface nano-functionalization of biomaterials. Mater Sci Eng R Rep 70:275–302CrossRefGoogle Scholar
  24. 24.
    Boyce BM, Lindsey BA, Clovis NB et al (2012) Additive effects of exogenous IL-12 supplementation and antibiotic treatment in infection prophylaxis. J Orthop Res 30:196–202CrossRefGoogle Scholar
  25. 25.
    Noore J, Noore A, Li B (2013) Cationic antimicrobial peptide LL-37 is effective against both extra- and intracellular Staphylococcus aureus. Antimicrob Agents Chemother 57:1283–1290CrossRefGoogle Scholar
  26. 26.
    Brogden KA (2005) Antimicrobial peptides: pore formers or metabolic inhibitors in bacteria? Nat Rev Microbiol 3:238–250CrossRefGoogle Scholar
  27. 27.
    Hancock RE, Rozek A (2002) Role of membranes in the activities of antimicrobial cationic peptides. FEMS Microbiol Lett 206:143–149CrossRefGoogle Scholar
  28. 28.
    Matsuzaki K (2009) Control of cell selectivity of antimicrobial peptides. Biochem Biophys Acta 1788:1687–1692CrossRefGoogle Scholar
  29. 29.
    Costa F, Carvalho IF, Montelaro RC, Gomes P, Martins MC (2011) Covalent immobilization of antimicrobial peptides (AMPs) onto biomaterial surfaces. Acta Biomater 7:1431–1440CrossRefGoogle Scholar
  30. 30.
    Sass V, Schneider T, Wilmes M et al (2010) Human β-defensin 3 inhibits cell wall biosynthesis in Staphylococci. Infect Immun 78:2793–2800CrossRefGoogle Scholar
  31. 31.
    Harder J, Bartels J, Christophers E, Schröder J-M (2001) Isolation and characterization of human β-defensin-3, a novel human inducible peptide antibiotic. J Biol Chem 276:5707–5713CrossRefGoogle Scholar
  32. 32.
    Zhu C, Tan H, Cheng T et al (2013) Human β-defensin 3 inhibits antibiotic-resistant Staphylococcus biofilm formation. J Surg Res 183:204–213CrossRefGoogle Scholar
  33. 33.
    Maisetta G, Batoni G, Esin S et al (2006) In vitro bactericidal activity of human β-defensin 3 against multidrug-resistant nosocomial strains. Antimicrob Agents Chemother 50:806–809CrossRefGoogle Scholar
  34. 34.
    Wu Z, Hoover DM, Yang D et al (2003) Engineering disulfide bridges to dissect antimicrobial and chemotactic activities of human β-defensin 3. Proc Natl Acad Sci 100:8880–8885CrossRefGoogle Scholar
  35. 35.
    Funderburg N, Lederman MM, Feng Z et al (2007) Human β-defensin-3 activates professional antigen-presenting cells via Toll-like receptors 1 and 2. Proc Natl Acad Sci 104:18631–18635CrossRefGoogle Scholar
  36. 36.
    Warnke PH, Springer IN, Russo PA et al (2006) Innate immunity in human bone. Bone 38:400–408CrossRefGoogle Scholar
  37. 37.
    Reubens B, Poesen J, Danjon F, Geudens G, Muys B (2007) The role of fine and coarse roots in shallow slope stability and soil erosion control with a focus on root system architecture: a review. Trees 21:385–402CrossRefGoogle Scholar
  38. 38.
    Huang H-H, Wu C-P, Sun Y-S, Yang W-E, Lee T-H (2014) Surface nanotopography of an anodized Ti–6Al–7Nb alloy enhances cell growth. J Alloys Compd 615(Supplement 1):S648–S654CrossRefGoogle Scholar
  39. 39.
    Li J, Wang G, Wang D, Wu Q, Jiang X, Liu X (2014) Alkali-treated titanium selectively regulating biological behaviors of bacteria, cancer cells and mesenchymal stem cells. J Colloid Interface Sci 436:160–170CrossRefGoogle Scholar
  40. 40.
    Zhu C, Tan H, Cheng T et al (2013) Human beta-defensin 3 inhibits antibiotic-resistant Staphylococcus biofilm formation. J Surg Res 183:204–213CrossRefGoogle Scholar
  41. 41.
    Ma M, Kazemzadeh-Narbat M, Hui Y et al (2012) Local delivery of antimicrobial peptides using self-organized TiO2 nanotube arrays for peri-implant infections. J Biomed Mater Res Part A 100:278–285CrossRefGoogle Scholar
  42. 42.
    Wang J, Li J, Qian S et al (2016) Antibacterial surface design of titanium-based biomaterials for enhanced bacteria-killing and cell-assisting functions against periprosthetic joint infection. ACS Appl Mater Interfaces 8:11162–11178CrossRefGoogle Scholar
  43. 43.
    Cheng H, Xiong W, Fang Z et al (2016) Strontium (Sr) and silver (Ag) loaded nanotubular structures with combined osteoinductive and antimicrobial activities. Acta Biomater 31:388–400CrossRefGoogle Scholar
  44. 44.
    Li J, Liu X, Wen C (eds) (2015) Surface coating and modification of metallic biomaterials. Woodhead Publishing, SawstonGoogle Scholar
  45. 45.
    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 123:68–74CrossRefGoogle Scholar
  46. 46.
    Ciampi S, Böcking T, Kilian KA, James M, Harper JB, Gooding JJ (2007) Functionalization of acetylene-terminated monolayers on Si(100) surfaces: a click chemistry approach. Langmuir 23:9320–9329CrossRefGoogle Scholar
  47. 47.
    Jia Z, Xiu P, Li M et al (2016) Bioinspired anchoring AgNPs onto micro-nanoporous TiO2 orthopedic coatings: trap-killing of bacteria, surface-regulated osteoblast functions and host responses. Biomaterials 75:203–222CrossRefGoogle Scholar
  48. 48.
    Huang H-H, Wu C-P, Sun Y-S, Lee T-H (2013) Improvements in the corrosion resistance and biocompatibility of biomedical Ti–6Al–7Nb alloy using an electrochemical anodization treatment. Thin Solid Films 528:157–162CrossRefGoogle Scholar
  49. 49.
    Huang H-H, Wu C-P, Sun Y-S, Yang W-E, Lin M-C, Lee T-H (2014) Surface nanoporosity of β-type Ti–25Nb–25Zr alloy for the enhancement of protein adsorption and cell response. Surfa Coat Technol Part B 259:206–212CrossRefGoogle Scholar
  50. 50.
    Yang W-E, Lan M-Y, Lee S-W, Chang J-K, Huang H-H (2015) Primary human nasal epithelial cell response to titanium surface with a nanonetwork structure in nasal implant applications. Nanoscale Res Lett 10:1–10CrossRefGoogle Scholar
  51. 51.
    Sun Y-S, Liu J-F, Wu C-P, Huang H-H (2015) Nanoporous surface topography enhances bone cell differentiation on Ti–6Al–7Nb alloy in bone implant applications. J Alloys Compd 643(Supplement 1):S124–S132CrossRefGoogle Scholar
  52. 52.
    Divya Rani VV, Manzoor K, Menon D, Selvamurugan N, Nair SV (2009) The design of novel nanostructures on titanium by solution chemistry for an improved osteoblast response. Nanotechnology 20:195101CrossRefGoogle Scholar
  53. 53.
    Sun Y-S, Chang J-H, Huang H-H (2016) Enhancing the biological response of titanium surface through the immobilization of bone morphogenetic protein-2 using the natural cross-linker genipin. Surf Coat Technol 303:289–297CrossRefGoogle Scholar
  54. 54.
    An YH, Friedman RJ (1998) Concise review of mechanisms of bacterial adhesion to biomaterial surfaces. J Biomed Mater Res 43:338–348CrossRefGoogle Scholar
  55. 55.
    Gottenbos B, van der Mei HC, Busscher HJ (2000) Initial adhesion and surface growth of Staphylococcus epidermidis and Pseudomonas aeruginosa on biomedical polymers. J Biomed Mater Res 50:208–214CrossRefGoogle Scholar
  56. 56.
    Hancock RE, Sahl HG (2006) Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies. Nat Biotechnol 24:1551–1557CrossRefGoogle Scholar
  57. 57.
    Westerhoff HV, Juretic D, Hendler RW, Zasloff M (1989) Magainins and the disruption of membrane-linked free-energy transduction. Proc Natl Acad Sci USA 86:6597–6601CrossRefGoogle Scholar
  58. 58.
    Sahl HG, Pag U, Bonness S, Wagner S, Antcheva N, Tossi A (2005) Mammalian defensins: structures and mechanism of antibiotic activity. J Leukoc Biol 77:466–475CrossRefGoogle Scholar
  59. 59.
    Henzler Wildman KA, Lee DK, Ramamoorthy A (2003) Mechanism of lipid bilayer disruption by the human antimicrobial peptide, LL-37. Biochemistry 42:6545–6558CrossRefGoogle Scholar
  60. 60.
    Dhople V, Krukemeyer A, Ramamoorthy A (2006) The human beta-defensin-3, an antibacterial peptide with multiple biological functions. Biochem Biophys Acta 1758:1499–1512CrossRefGoogle Scholar
  61. 61.
    Bechinger B (2005) Detergent-like properties of magainin antibiotic peptides: a 31P solid-state NMR spectroscopy study. Biochem Biophys Acta 1712:101–108CrossRefGoogle Scholar
  62. 62.
    Shai Y (2002) Mode of action of membrane active antimicrobial peptides. Biopolymers 66:236–248CrossRefGoogle Scholar
  63. 63.
    Strandberg E, Ulrich AS (2004) NMR methods for studying membrane-active antimicrobial peptides. Concepts Magn Reson Part A 23:89–120CrossRefGoogle Scholar
  64. 64.
    Powers J-PS, Tan A, Ramamoorthy A, Hancock RE (2005) Solution structure and interaction of the antimicrobial polyphemusins with lipid membranes. Biochemistry 44:15504–15513CrossRefGoogle Scholar
  65. 65.
    Peschel A, Jack RW, Otto M et al (2001) Staphylococcus aureus resistance to human defensins and evasion of neutrophil killing via the novel virulence factor MprF is based on modification of membrane lipids with l-lysine. J Exp Med 193:1067–1076CrossRefGoogle Scholar
  66. 66.
    Matsuzaki K (1999) Why and how are peptide–lipid interactions utilized for self-defense? Magainins and tachyplesins as archetypes. Biochim Biophys Acta BBA Biomembr 1462:1–10CrossRefGoogle Scholar
  67. 67.
    Bierbaum G, Sahl H-G (1985) Induction of autolysis of staphylococci by the basic peptide antibiotics Pep 5 and nisin and their influence on the activity of autolytic enzymes. Arch Microbiol 141:249–254CrossRefGoogle Scholar
  68. 68.
    Li Q, Zhou Y, Dong K, Guo X (2010) Potential therapeutic efficacy of a bactericidal-immunomodulatory fusion peptide against methicillin-resistant Staphylococcus aureus skin infection. Appl Microbiol Biotechnol 86:305–309CrossRefGoogle Scholar
  69. 69.
    Yang D, Chertov O, Bykovskaia SN et al (1999) Beta-defensins: linking innate and adaptive immunity through dendritic and T cell CCR6. Science 286:525–528CrossRefGoogle Scholar
  70. 70.
    Garcia JR, Jaumann F, Schulz S et al (2001) Identification of a novel, multifunctional beta-defensin (human beta-defensin 3) with specific antimicrobial activity. Its interaction with plasma membranes of Xenopus oocytes and the induction of macrophage chemoattraction. Cell Tissue Res 306:257–264CrossRefGoogle Scholar
  71. 71.
    Chaly YV, Paleolog EM, Kolesnikova TS, Tikhonov II, Petratchenko EV, Voitenok NN (2000) Neutrophil alpha-defensin human neutrophil peptide modulates cytokine production in human monocytes and adhesion molecule expression in endothelial cells. Eur Cytokine Netw 11:257–266Google Scholar
  72. 72.
    Niyonsaba F, Ushio H, Hara M et al (2010) Antimicrobial peptides human beta-defensins and cathelicidin LL-37 induce the secretion of a pruritogenic cytokine IL-31 by human mast cells. J Immunol 184:3526–3534CrossRefGoogle Scholar
  73. 73.
    Batoni G, Maisetta G, Esin S, Campa M (2006) Human beta-defensin-3: a promising antimicrobial peptide. Mini Rev Med Chem 6:1063–1073CrossRefGoogle Scholar
  74. 74.
    Grynpas MD, Marie PJ (1990) Effects of low doses of strontium on bone quality and quantity in rats. Bone 11:313–319CrossRefGoogle Scholar
  75. 75.
    Fukuda A, Takemoto M, Saito T et al (2011) Bone bonding bioactivity of Ti metal and Ti–Zr–Nb–Ta alloys with Ca ions incorporated on their surfaces by simple chemical and heat treatments. Acta Biomater 7:1379–1386CrossRefGoogle Scholar
  76. 76.
    Li J, Zhang W, Qiao Y et al (2014) Chemically regulated bioactive ion delivery platform on a titanium surface for sustained controlled release. J Mater Chem B 2:283–294CrossRefGoogle Scholar
  77. 77.
    Jiang X, Zhao J, Wang S et al (2009) Mandibular repair in rats with premineralized silk scaffolds and BMP-2-modified bMSCs. Biomaterials 30:4522–4532CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  1. 1.Department of Orthopaedic SurgeryAnhui Provincial Hospital, The First Affiliated Hospital of University of Science and Technology of ChinaHefeiPeople’s Republic of China
  2. 2.Department of Orthopaedic Surgery, Jinling HospitalNanjing University School of MedicineNanjingPeople’s Republic of China
  3. 3.Department of GeriatricsAnhui Provincial Hospital, The First Affiliated Hospital of University of Science and Technology of ChinaHefeiChina
  4. 4.Department of Polymer Science and EngineeringUniversity of Science and Technology of ChinaHefeiChina

Personalised recommendations