ADVANCED ENGINEERING MATERIALS Dol:10.1002/adem.200800026 Nanostructured Titanium for Biomedical Applications By Ruslan Z.Valliev,Irina P.Semenova,Vladimir V.Latysh,Henry Rack,Terry C.Lowe,Jiri Petruzelka, Ludek Dluhos,Daniel Hrusak and Jarmila Sochova Metallic materials,for example,stainless steel,titanium avenues and concepts for medical implants,providing bene- and its alloys,and tantalum,are widely used for medical im- fits in all areas of medical device technology plants in trauma surgery,orthopedic and oral medicine.-31 Numerous clinical studies of medical devices fabricated Successful incorporation of these materials for design,fabri- from commercial purity (CP)titanium for trauma,orthopae- cation and application of medical devices require that they dic and oral medicine has proven its excellent biocompatibil- meet several critical criteria.Paramount is their biocompat- ity.However the mechanical strength of CP titanium is rela- ibility as expressed by their relative reactivity with human tis- tively low compared to other metals used in biomedical sues.Another is their ability to provide sufficient mechanical devices.Whereas the strength of this material can be in- strength,especially under cyclic loading conditions to ensure creased by either alloying or secondary processing,for exam- the durability of the medical devices made therefrom.Finally ple rolling,drawing,etc.,these enhancements normally come the material should be machinable and formable thereby en- with some degradation in biometric response and fatigue be- abling device fabrication at an affordable cost.In this paper haviour.Recently it has been shown that nanostructuring of we show that nanostructured commercial purity titanium CP titanium by SPD processing can provide a new and prom- produced by severe plastic deformation (SPD)opens new ising alternative method for improving the mechanical prop- erties of this material This approach also has the benefit of enhancing the biological response of the CP titanium sur- face,I91 This paper reports the results of the first developments [*Prof.R.Z.Valiev,Dr.I.P.Semenova and studies of nanostructured titanium (n-Ti),produced as Institute of Physics of Advanced Materials long-sized rods with superior mechanical and biomedical Ufa State Aviation Technical University properties and demonstrates its applicability for dental im- 12 K.Marx str.,Ufa 450000 Russia plants.The effort was conducted using commercially pure E-mail:RZValiev@mail.rb.ru Grade 4 titanium [C-0.052%,O2-0.34%,Fe -0.3%,N- Dr.V.V.Latush 0.015%,Ti-bal.(wt.pct.)].Nanostructuring involved SPD Innovation Scientific and Technical Center skra> processing by equal-channel angular pressinglol followed by 81 Pushkin str. thermo-mechanical treatment(TMT)using forging and draw- Ufa 450077 Russia ing to produce 7 mm diameter bars with a 3 m length.This Prof.H.Rack processing resulted in a large reduction in grain size,from School of Materials Science and Engineering the 25 um equiaxed grain structure of the initial titanium rods Clemson University to 150 nm after combined SPD and TMT processing,as Clemson,SC 29634 LISA shown in Figure 1.The selected area electron diffraction pat- Dr.T.C.Lowe tern,Figure 1(c),further suggests that the ultra fine grains Los Alamos National Laboratory contained predominantly high-angle non-equilibrium grain Los Alamos boundaries with increased grain-to-grain internal stresses.1 NM 87545 USA A similar structure for CP Ti can be produced in small Prof.J.Petruzelka discs using other SPD methods,for example-high pressure FS,VSB-Technicki univerzita Ostrava torsion(HPT)as studied in detail.s In the present work it tr.17 listopadu 15 was essential to produce homogeneous ultrafine-grained Ostrava-Poruba,CZ 708 33 structure throughout a three-meter length rod to enable the E-mail:jiri.petruzelka@vsb.cz pilot production of implants and provide sufficient material J.Sochova,Dr.L.Dluhos for thorough testing of the mechanical and bio-medical prop- Timplant,Sjednoceni77 erties of the nanostructured titanium. Ostrava-Polanka,CZ 725 25 Table 1 illustrates mechanical property benefits attainable Dr.D.Hrusak by nanostructuring of CP titanium,for example,the strength FN Plzen of the nanostructured titanium is nearly twice that of conven- Alej Svobody 80 tional CP titanium.Notably this improvement has been Plzen CZ 323 00-Czech Republic achieved without the drastic ductility reductions (to below WY InterScience ADVANCED ENGINEERING MATERIALS 2008,10,No.8 2008 WILEY-VCH Verlag GmbH Co.KGaA,Weinheim
DOI: 10.1002/adem.200800026 Nanostructured Titanium for Biomedical Applications By Ruslan Z. Valiev,* Irina P. Semenova, Vladimir V. Latysh, Henry Rack, Terry C. Lowe, Jiri Petruzelka, Ludek Dluhos, Daniel Hrusak and Jarmila Sochova Metallic materials, for example, stainless steel, titanium and its alloys, and tantalum, are widely used for medical implants in trauma surgery, orthopedic and oral medicine.[1–3] Successful incorporation of these materials for design, fabrication and application of medical devices require that they meet several critical criteria. Paramount is their biocompatibility as expressed by their relative reactivity with human tissues. Another is their ability to provide sufficient mechanical strength, especially under cyclic loading conditions to ensure the durability of the medical devices made therefrom. Finally the material should be machinable and formable thereby enabling device fabrication at an affordable cost. In this paper we show that nanostructured commercial purity titanium produced by severe plastic deformation (SPD) opens new avenues and concepts for medical implants, providing benefits in all areas of medical device technology. Numerous clinical studies of medical devices fabricated from commercial purity (CP) titanium for trauma, orthopaedic and oral medicine has proven its excellent biocompatibility.[3] However the mechanical strength of CP titanium is relatively low compared to other metals used in biomedical devices. Whereas the strength of this material can be increased by either alloying or secondary processing, for example rolling, drawing, etc., these enhancements normally come with some degradation in biometric response and fatigue behaviour. Recently it has been shown that nanostructuring of CP titanium by SPD processing can provide a new and promising alternative method for improving the mechanical properties of this material.[4–8] This approach also has the benefit of enhancing the biological response of the CP titanium surface.[9] This paper reports the results of the first developments and studies of nanostructured titanium (n-Ti), produced as long-sized rods with superior mechanical and biomedical properties and demonstrates its applicability for dental implants. The effort was conducted using commercially pure Grade 4 titanium [C – 0.052 %, O2 – 0.34 %, Fe – 0.3 %, N – 0.015 %, Ti-bal. (wt. pct.)]. Nanostructuring involved SPD processing by equal-channel angular pressing[10] followed by thermo-mechanical treatment (TMT) using forging and drawing to produce 7 mm diameter bars with a 3 m length. This processing resulted in a large reduction in grain size, from the 25 lm equiaxed grain structure of the initial titanium rods to 150 nm after combined SPD and TMT processing, as shown in Figure 1. The selected area electron diffraction pattern, Figure 1(c), further suggests that the ultra fine grains contained predominantly high-angle non-equilibrium grain boundaries with increased grain-to-grain internal stresses.[11] A similar structure for CP Ti can be produced in small discs using other SPD methods, for example – high pressure torsion (HPT) as studied in detail.[8] In the present work it was essential to produce homogeneous ultrafine-grained structure throughout a three-meter length rod to enable the pilot production of implants and provide sufficient material for thorough testing of the mechanical and bio-medical properties of the nanostructured titanium. Table 1 illustrates mechanical property benefits attainable by nanostructuring of CP titanium, for example, the strength of the nanostructured titanium is nearly twice that of conventional CP titanium. Notably this improvement has been achieved without the drastic ductility reductions (to below COMMUNICATIONS ADVANCED ENGINEERING MATERIALS 2008, 10, No. 8 © 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 1 – [*] Prof. R. Z. Valiev, Dr. I. P. Semenova Institute of Physics of Advanced Materials Ufa State Aviation Technical University 12 K. Marx str., Ufa 450000 Russia E-mail: RZValiev@mail.rb.ru Dr. V. V. Latysh Innovation Scientific and Technical Center «Iskra» 81 Pushkin str. Ufa 450077 Russia Prof. H. Rack School of Materials Science and Engineering Clemson University Clemson, SC 29634 USA Dr. T. C. Lowe Los Alamos National Laboratory Los Alamos NM 87545 USA Prof. J. Petruzelka FS, VŠB – Technická univerzita Ostrava tr. 17 listopadu 15 Ostrava-Poruba, CZ 708 33 E-mail: jiri.petruzelka@vsb.cz J. Sochova, Dr. L. Dluhos Timplant®, Sjednocení 77 Ostrava – Polanka, CZ 725 25 Dr. D. Hrusak FN Plzen Alej Svobody 80 Plzen CZ 323 00 – Czech Republic
ADVANCED ENGINEERING MATERIALS Valiev et al./Nanostructured Titanium for Biomedical Applications One objective of this effort was to design fabricate and implant a nanostructured CP Grade 4 titanium dental post to clinically demonstrate the benefits associated with na- nostructuring outlined previously.Toward this end,a reduced diameter implant post Nanoimplant was designed and fabricated. a b) c This implant sustains the same load as a con- Fig.1.Microstructure of Grade 4 CP Ti:a)the initinl coarse grained rod;b,c)after ECAP TMT (Optical and electron photomicrographs). ventional 3.5 mm-diameter titanium im plant,the former having the added capabili- ty of being used as a pillar in cases of Table 1.Mechanical properties of conventionally processed and nanostructured CP Grade 4 titanium. insufficient thickness of the alveolar bone. The certified system of Timplant manu- State Processing/treatment UTS,MPa YS,MPa Elongation,Reduction Fatigue factured to standard EN ISO 13485:2003 was conditions % Area,% strength at 10cycles used during development of the Nanoim- plant implant.The implants are shown in Conventional Ti 700 530 25 分 340 As received Figure 4,the nanoimplant intraosseal diame- ter 2.4 mm,having a strength equivalent to nTi 1240 1200 12 42 620 ECAP+TMT the conventional of 3.5 mm diameter im- plant. annealed 940 840 16 530 Ti-6AL-4V ELI To date over 250 Nanoimplants have been implanted,most of them as immediate load implants,with all results indicating the excellent primary stability of Nanoimplants 10%elongation to failure)normally seen after rolling or when compared to other implant types [http://www.tim- drawing. plant.cz/e_stomatolog.asp].For example,a 55-year-old male Further room temperature,laboratory air fatigue studies of with edentulous mandible and maxilla was treated by inser- nanostructured and conventional CP titanium were per- tion of conical implants laterally and NanoimplantsR in the formed per ASTME 466-96 at a load ratio R (min/omax)=0.1 narrow anterior part.Primary retention of all implants was and loading frequency of 20 Hz.Table 1 also shows that the very good;on the day of surgery the patient received a com- fatigue strength of nanostructured CP titanium at 105 cycles plete provisional bridge.Post-operation healing at the sur- is almost two times higher than conventional CP titanium gery site occurred without complications,with subsequent at- and exceeds that of the Ti-6Al-4V alloy.12 tachment of a definitive metalloceramic bridge completing Cytocompatibility tests utilizing fibroblast mice cells L929 the treatment. were undertaken to verify the previously reported benefits of Thus,nanostructuring of titanium by SPD processing has nanostructured CP titanium vis a vis conventional coarse made material with significantly superior mechanical perfor- grained CP Ti.This study was performed as described else- mance when compared to conventional CP Grade 4 titanium. where2 with hydrofluoric acid surface etching being per- formed prior to cell exposure.Figure 2 shows the etched con- ventional and nanostructured titanium surfaces,respectively. The differences in surface roughness of these materials are T五 CPTi easily seen,a homogeneous and nanometer-sized roughness being apparent for nanostructured titanium compared with the much coarser structure for etched CP Grade 4 titanium. The cell attachment investigation shows that fibroblast co- lonization of the CP Grade 4 titanium surface dramatically in- creases after nanostructuring,Figure 3.For example,the sur- face cell occupation for conventional CP Ti was 53.0%after 72 hrs in contrast to 87.2%for nanostructured CP Grade 4 (Tab.2).The latter observations also confirm the previous studies,4 showing that cell-adhesion on nanostructured titanium is greater than on conventional CP Grade 4 tita- nium.This result suggests that a high osteointegration rate 300x should be expected with nanostructured CP Grade 4 titanium Fig.2.Surface relief after hydrofluoric acid treatment of nanostructured (left)and Cp when compared to conventional titanium. Grade 4 titanium(right)surfaces. http://www.aem-journal.com 2008 WILEY-VCH Verlag GmbH Co.KGaA,Weinheim ADVANCED ENGINEERING MATERIALS 2008,10,No.8
10 % elongation to failure) normally seen after rolling or drawing. Further room temperature, laboratory air fatigue studies of nanostructured and conventional CP titanium were performed per ASTM E 466-96 at a load ratio R (rmin/rmax) = 0.1 and loading frequency of 20 Hz. Table 1 also shows that the fatigue strength of nanostructured CP titanium at 106 cycles is almost two times higher than conventional CP titanium and exceeds that of the Ti-6Al-4V alloy.[1,2] Cytocompatibility tests utilizing fibroblast mice cells L929 were undertaken to verify the previously reported benefits of nanostructured CP titanium vis à vis conventional coarse grained CP Ti. This study was performed as described elsewhere,[12] with hydrofluoric acid surface etching being performed prior to cell exposure. Figure 2 shows the etched conventional and nanostructured titanium surfaces, respectively. The differences in surface roughness of these materials are easily seen, a homogeneous and nanometer-sized roughness being apparent for nanostructured titanium compared with the much coarser structure for etched CP Grade 4 titanium. The cell attachment investigation shows that fibroblast colonization of the CP Grade 4 titanium surface dramatically increases after nanostructuring, Figure 3. For example, the surface cell occupation for conventional CP Ti was 53.0 % after 72 hrs in contrast to 87.2 % for nanostructured CP Grade 4 (Tab. 2). The latter observations also confirm the previous studies,[9,13,14] showing that cell-adhesion on nanostructured titanium is greater than on conventional CP Grade 4 titanium. This result suggests that a high osteointegration rate should be expected with nanostructured CP Grade 4 titanium when compared to conventional titanium. One objective of this effort was to design, fabricate and implant a nanostructured CP Grade 4 titanium dental post to clinically demonstrate the benefits associated with nanostructuring outlined previously. Toward this end, a reduced diameter implant post Nanoimplant® was designed and fabricated. This implant sustains the same load as a conventional 3.5 mm-diameter titanium implant, the former having the added capability of being used as a pillar in cases of insufficient thickness of the alveolar bone. The certified system of Timplant® manufactured to standard EN ISO 13485:2003 was used during development of the Nanoimplant® implant. The implants are shown in Figure 4, the nanoimplant intraosseal diameter 2.4 mm, having a strength equivalent to the conventional of 3.5 mm diameter implant. To date over 250 Nanoimplants® have been implanted, most of them as immediate load implants, with all results indicating the excellent primary stability of Nanoimplants® when compared to other implant types [http://www.timplant.cz/e_stomatolog.asp]. For example, a 55-year-old male with edentulous mandible and maxilla was treated by insertion of conical implants laterally and Nanoimplants® in the narrow anterior part. Primary retention of all implants was very good; on the day of surgery the patient received a complete provisional bridge. Post-operation healing at the surgery site occurred without complications, with subsequent attachment of a definitive metalloceramic bridge completing the treatment. Thus, nanostructuring of titanium by SPD processing has made material with significantly superior mechanical performance when compared to conventional CP Grade 4 titanium. Valiev et al./Nanostructured Titanium for Biomedical Applications COMMUNICATIONS 2 http://www.aem-journal.com © 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim ADVANCED ENGINEERING MATERIALS 2008, 10, No. 8 Fig. 1. Microstructure of Grade 4 CP Ti: a) the initial coarse grained rod; b, c) after ECAP + TMT (Optical and electron photomicrographs). Table 1. Mechanical properties of conventionally processed and nanostructured CP Grade 4 titanium. State Processing/treatment conditions UTS, MPa YS, MPa Elongation, % Reduction Area, % Fatigue strength at 106 cycles 1 Conventional Ti As received 700 530 25 52 340 2 nTi ECAP + TMT 1240 1200 12 42 620 3 annealed Ti-6Al-4V ELI 940 840 16 45 530 Fig. 2. Surface relief after hydrofluoric acid treatment of nanostructured (left) and CP Grade 4 titanium (right) surfaces
ADVANCED ENGINEERING Valiev et al./Nanostructured Titanium for Biomedical Applications MATERIALS preliminary results being extremely encouraging. Further clinical studies are presently underway with an enlarged,1000 patient,population. M Received:January 30,2008 Final version:May 26,2008 ATIO Fig.3.Occupation of the mice fibroblast cells 1929 after 24 hours;Nanostructured (left)and con- ventional (right)CP Grade 4 titanium. [ M.Niinomi,Metal Mater.Trans A 2002,33 A, 云 477. Table 2.Surface cell occupation for conventional and nanostructured CP Grade 4 titanium [21 R.Boyer,G.Welsch,E.Collings,Mater.Proper- ties Handbook:Titanium Alloys ASM Interna- tional,1998. Material Surface treatment Occupied surface Ipct.] after 72 hours [31 D.M.Brunette,P.Tengvall,M.Textor,P.Thom- CP Gr.4Ti Machining,followed by 53.0 sen,Titanium in Med.Springer-Verlag Berlin hydrofluoric acid etching Heidelberg,2003. Nanostructured Gr.4 Ti 872 [4]R.Z.Valiev,Nature Mater.2004,3,511. [5]V.V.Stolyarov,V.V.Latysh,R.Z.Valiev, Y.T.Zhu,T.C.Lowe,Investigations and Appl. of Severe Plastic Deformation,Kluwer Academic Publishers 2000. [6]Y.T.Zhu,T.C.Lowe,R.Z.Valiev,V.V.Stolyarov, V.V.Latysh,G.I.Raab,U.S Patent 6399 215,2002. [7]V.V.Latysh,I.P.Semenova,G.H.Salimgareeva,I.V. Kandarov,Y.T.Zhu,T.C.Lowe,R.Z.Valiev,Mater. Sci.F0rum2006,503-504,763. [8]R.Z.Valiev,A.V.Sergueeva,A.K.Mukherjee,Scr.Ma- ter200349,669. [9]C.Yao,E.B.Slamovich,J.Qazi,H.J.Rack,T.J.Web- ster,Ceram.Trans.2005,159,239. [10]R.Z.Valiev,T.G.Langdon,Progr.Mater.Sci 2006,51, Fig.4.3.5 mm diameter Timplan(above)and 2.4 mm diameter Nanoimplant (be 881. loe以. [11]R.Z.Valiev,R.K.Islamgaliev,I.V.Alexandrov,Progr. Mater..Sci.2000,45,103. Furthermore,cytocompatibility studies with fibroblast mice [12]J.Petruzelka,L.Dluhos,D.Hrusak,J.Sochova,Sb.Ved. cells L929 have indicated that the nanostructured Ti surface Pr.Vys.Sk.banske-Tech.Univ.Ostrava.2006,roc.LII.c. has significantly higher cell colonization,suggesting more 1.cd.1517.ISSN1210-0471,177. rapid osseointegration.Nanostructured(Nanoimplants)im- [13]S.Faghihi,A.P.Zhilyaev,J.A.Szpunar,F.Azari, plants have been successfully designed and fabricated.Clini- H.Vali,M.Tabrizian,Ado.Mater.2007,19,1069. cal trials with over 250 patients,most of them receiving im- [14]S.Faghihi,F.Azari,A.P.Zhilyaev,J.A.Szpunar, mediate load implants,have shown no adverse effects, H.Vali,M.Tabrizian,Biomater.2007,28,3887 ADVANCED ENGINEERING MATERIALS 2008,10,No.8 @2008 WILEY-VCH Verlag GmbH Co.KGaA,Weinheim http://www.aem-journal.com
Furthermore, cytocompatibility studies with fibroblast mice cells L929 have indicated that the nanostructured Ti surface has significantly higher cell colonization, suggesting more rapid osseointegration. Nanostructured (Nanoimplants®) implants have been successfully designed and fabricated. Clinical trials with over 250 patients, most of them receiving immediate load implants, have shown no adverse effects, preliminary results being extremely encouraging. Further clinical studies are presently underway with an enlarged, 1000 patient, population. Received: January 30, 2008 Final version: May 26, 2008 – [1] M. Niinomi, Metal Mater. Trans A 2002, 33 A, 477. [2] R. Boyer, G. Welsch, E. Collings, Mater. Properties Handbook: Titanium Alloys ASM International, 1998. [3] D. M. Brunette, P. Tengvall, M. Textor, P. Thomsen, Titanium in Med. Springer-Verlag Berlin Heidelberg, 2003. [4] R. Z. Valiev, Nature Mater. 2004, 3, 511. [5] V. V. Stolyarov, V. V. Latysh, R. Z. Valiev, Y. T. Zhu, T. C. Lowe, Investigations and Appl. of Severe Plastic Deformation, Kluwer Academic Publishers 2000. [6] Y. T. Zhu, T. C. Lowe, R. Z. Valiev, V. V. Stolyarov, V. V. Latysh, G. I. Raab, U.S Patent 6399 215, 2002. [7] V. V. Latysh, I. P. Semenova, G. H. Salimgareeva, I. V. Kandarov, Y. T. Zhu, T. C. Lowe, R. Z. Valiev, Mater. Sci. Forum 2006, 503–504, 763. [8] R. Z. Valiev, A. V. Sergueeva, A. K. Mukherjee, Scr. Mater 2003, 49, 669. [9] C. Yao, E. B. Slamovich, J. Qazi, H. J. Rack, T. J. Webster, Ceram. Trans. 2005, 159, 239. [10] R. Z. Valiev, T. G. Langdon, Progr. Mater. Sci 2006, 51, 881. [11] R. Z. Valiev, R. K. Islamgaliev, I. V. Alexandrov, Progr. Mater. Sci. 2000, 45, 103. [12] J. Petruzˇelka, L. Dluhoš, D. Hrušák, J. Sochová, Sb. Ved. Pr. Vys. Šk. bánské – Tech. Univ. Ostrava. 2006, roc. LII. c. 1. cl. 1517. ISSN 1210-0471, 177. [13] S. Faghihi, A. P. Zhilyaev, J. A. Szpunar, F. Azari, H. Vali, M. Tabrizian, Adv. Mater. 2007, 19, 1069. [14] S. Faghihi, F. Azari, A. P. Zhilyaev, J. A. Szpunar, H. Vali, M. Tabrizian, Biomater. 2007, 28, 3887. ______________________ Valiev et al./Nanostructured Titanium for Biomedical Applications COMMUNICATIONS ADVANCED ENGINEERING MATERIALS 2008, 10, No. 8 © 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim http://www.aem-journal.com 3 Table 2. Surface cell occupation for conventional and nanostructured CP Grade 4 titanium. Material Surface treatment Occupied surface [pct.] after 72 hours CP Gr. 4 Ti Machining, followed by hydrofluoric acid etching 53.0 Nanostructured Gr. 4 Ti 87.2 Fig. 3. Occupation of the mice fibroblast cells L929 after 24 hours; Nanostructured (left) and conventional (right) CP Grade 4 titanium. Fig. 4. 3.5 mm diameter Timplant® (above) and 2.4 mm diameter Nanoimplant® (below)
ADVANCED ENGINEERING MATERIALS Valiev et al./Nanostructured Titanium for Biomedical Applications Nanostructured Titanium for Biomedical Applications R.Z.Valiev,*I.P.Semenova,V.V.Latysh,H.Rack,T.C.Lowe,J.Petruzelka,L.Dluhos, D.Hrusak,I.Sochova In this paper the authors show that nanostructured commercial purity titanium produced by severe plastic deformation(SPD)opens new avenues and concepts for medical im- plants,providing benefits in all areas of medical device technology. ADV.ENG.MATER.2008,10............................... ■.…■ http://www.aem-journal.com 2008 WILEY-VCH Verlag GmbH Co.KGaA,Weinheim ADVANCED ENGINEERING MATERIALS 2008,10,No.8
Valiev et al./Nanostructured Titanium for Biomedical Applications COMMUNICATIONS 4 http://www.aem-journal.com © 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim ADVANCED ENGINEERING MATERIALS 2008, 10, No. 8 Nanostructured Titanium for Biomedical Applications R. Z. Valiev,* I. P. Semenova, V. V. Latysh, H. Rack, T. C. Lowe, J. Petruzelka, L. Dluhos, D. Hrusak, J. Sochova In this paper the authors show that nanostructured commercial purity titanium produced by severe plastic deformation (SPD) opens new avenues and concepts for medical implants, providing benefits in all areas of medical device technology. ADV. ENG. MATER. 2008, 10 ............................................... ...