Process for the Production of Storable Implants with an Ultrahydrophilic Surface

Abstract

The present invention concerns a process for the production of implants with an ultrahydrophilic surface as well as the implants produced in that way and also processes for the production of loaded, so-called bioactive implant surfaces of metallic or ceramic materials, which are used for implants such as artificial bones, joints, dental implants or also very small implants, for example what are referred to as stents, as well as implants which are further produced in accordance with the processes and which as so-called “delivery devices” allow controlled liberation, for example by way of dissociation, of the bioactive molecules from the implant materials.

Claims

1. A process for the production of a storable implant with an ultrahydrophilic surface, the process comprising the steps of: providing an implant with an ultrahydrophilic surface with dynamic contact angles from 0° to 10° upon wetting of the surface of the implant with water, placing the implant into a salt-containing aqueous solution, which is inert in relation to the surface and which encloses the implant on all sides, and evaporating the salt-containing aqueous solution to dryness under formation of an exsiccation layer that stabilizes and protects the surface of the implant.

2. The process of claim 1, wherein the salt-containing aqueous solution has a total ion concentration of more than 0.5 mol/l.

3. The process of claim 1, wherein the salt-containing aqueous solution has a total ion concentration of more than 1 mol/l.

4. The process of claim 1, wherein the step of evaporation affords a salt layer covering at least the ultrahydrophilic surface of the implant.

5. The process of claim 4, wherein the salt layer has a layer thickness of 1 to 500 μm.

6. The process of claim 1, wherein the step of evaporation gives a salt layer enclosing the implant on all sides.

7. The process of claim 1, comprising the additional step of sterilization of the implant.

8. The process of claim 7, wherein sterilization of the implant includes sterilization with ionizing radiation.

9. The process of claim 8, wherein the ionizing radiation is electromagnetic radiation.

10. The process of claim 1, further comprising the step of immobilising peptides on the ultrahydrophilic surface.

11. The process of claim 10, wherein the peptides immobilized on the ultrahydrophilic surface are present in a concentration of more than 200 ng/cm.sup.2.

12. The process of claim 10, wherein said peptides immobilized on the ultrahydrophilic surface are selected from the group consisting of bone growth factors, TGF proteins, BMP proteins including BMP-2 and BMP-7, vessel growth factors, VEGF, angiotropin, ubiquitin, antibiotics, and mixtures thereof.

13. A storable implant with an ultrahydrophilic surface produced by the process according to claim 1.

14. A process for the production of a storable implant with an ultrahydrophilic surface, the process comprising the steps of: providing an implant with an ultrahydrophilic surface with dynamic contact angles from 0° to 10° upon wetting of the surface of the implant with water, placing the implant into a salt-containing aqueous solution, which is inert in relation to the surface and which encloses the implant on all sides, and introducing the implant and the salt-containing aqueous solution into a transport packaging.

15. The process of claim 14 additionally comprising the step of closing the transport packaging in a gas-tight and liquid-tightfashion.

16. The process of claim 14, wherein the salt-containing aqueous solution has a total ion concentration of more than 0.5 mol/l.

17. The process of claim 14, wherein the salt-containing aqueous solution has a total ion concentration of more than 1 mol/l.

18. The process of claim 14, comprising the additional step of sterilization of the implant.

19. The process of claim 18, wherein sterilization of the implant includes sterilization with ionizing radiation.

20. The process of claim 19, wherein the ionizing radiation is electromagnetic radiation.

21. The process of claim 14, further comprising the step of immobilising peptides on the ultrahydrophilic surface.

22. The process of claim 21, wherein the peptides immobilized on the ultrahydrophilic surface are present in a concentration of more than 200 ng/cm.sup.2.

23. The process of claim 21, wherein said peptides immobilized on the ultrahydrophilic surface are selected from the group consisting of bone growth factors, TGF proteins, BMP proteins including BMP-2 and BMP-7, vessel growth factors, VEGF, angiotropin, ubiquitin, antibiotics, and mixtures thereof.

24. A storable implant with an ultrahydrophilic surface produced by the process according to claim 14.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0045] The invention is further described with reference to the accompanying Figures in which:

[0046] FIG. 1 shows a diagram relating to the magnitude of the advancing and receding angles respectively in relation to the treatment time of a metal surface with chromosulphuric acid.

[0047] FIGS. 2A-2C show electron-microscope images of an ultrahydrophilic, nanostructured surface on cp-titanium by the chromosulphuric acid processes at various levels of magnification. FIG. 2A shows a cleaned Industrial Standard SLA Surface (Sand-Blasted, Large-Grit, Acid-Etched) in a 25,000 Times Magnification. FIG. 2B shows a SLA Surface after Treatment in Chromosulphuric Acid at 240° C. for 60 Minutes in a 25,000 Times Magnification. FIG. 2C shows a representation of the Globular Nanostructure at 150,000 Times Magnification on an Electropolished Titanium Surface.

[0048] FIG. 3 shows a diagram relating to the liberation rate of BMP of metal surfaces subjected to various treatments.

[0049] FIGS. 4A-4C show scanning electron-microscope recordings of chromosulphuric acid-treated SLA titanium plates (14×14×1.5 mm) after gamma sterilisation in an exsiccation buffer (60 min CSS with HNO.sub.3, with quenching, gamma sterilised in PBS, θ=0°) at magnification of 1:100 (FIG. 4A), a magnification of 1:10,000 (FIG. 4B), and a magnification of 1:100,000 (FIG. 4C).

[0050] FIGS. 5A-5B show the EDX analysis of an ultrahydrophilic plate with exsiccation layer after gamma sterilisation (FIG. 5A) and after removal of the exsiccation layer with water (FIG. 5B).

DETAILED DESCRIPTION OF THE INVENTION

[0051] As FIG. 1 shows treatment of the metal surface with chromosulphuric acid for the production of ultrahydrophilic metal surfaces leads to surprising results. For that purpose titanium plates were incubated at 240° C. in concentrated chromosulphuric acid. As was surprisingly found, ultrahydrophilic surfaces (contact angle: <10°; contact angle hysteresis: ˜0°; standard deviations (n=5) are specified), are obtained in a time window of 30-60 minutes.

[0052] The inventors found similar minimum curves with 316L steel, titanium alloys and cobalt-chromium alloys. Table 1 shows that four to five times more BMP-2 can be bonded (adsorbed) on the ultrahydrophilic surface, than on the control. The bonded amount of BMP-2 on the untreated titanium surface is still below the surface treated with HNO.sub.3.

[0053] Those ultrahydrophilic surfaces have a very high surface energy which in the ultrahydrophilic range (contact angle <11°′ contact angle hysteresis ˜0° has a critical surface tension γc=71-72 dynes/cm. The high surface energy leads to the adsorption of suitable proteins.

[0054] In the production of the ultrahydrophilic surface a novel nanostructure is produced by the new chromosulphuric acid treatment (FIG. 2). Globular, interconnected structures of a diameter of 50-100 nm are produced on the titanium surface which is smooth prior to the treatment, with nanopores of a diameter of 50-100 nm being present between those globular structures. The nanostructures are presumably involved in the provision of ultrahydrophilicity. The conditions in that respect are in detail as follows for illustrations A, B and C:

A. Cleaned Industrial Standard SLA Surface (Sand-Blasted, Large-Grit, Acid-Etched) in a 25,000 Times Magnification

[0055] The surface was sand-blasted with corundum and then etched in an acid bath (HCl/H.sub.2SO.sub.4). The surface exhibits a smooth microstructure without any sign of a nanostructure.

B. SLA Surface after Treatment in Chromosulphuric Acid at 240° C. for 60 Minutes in a 25,000 Times Magnification

[0056] The chromosulphuric acid functionally produces a hydrophilic surface and structurally a “globular” nanostructure, besides the SLA microstructure. The diameter of the interconnected nano-balls is about 50-100 nm, with nanopores being formed in the same order of magnitude.

C. Representation of the Globular Nanostructure at 150,000 Times Magnification on an Electropolished Titanium Surface

[0057] The nanospheres are of a diameter of about 50 nm and are connected together. Pores of a diameter of 10-100 nm are formed between the spheres.

[0058] In a further step, peptides like bone growth factors can be immobilised on those nanostructures by means of physisorptive or chemisorptive bonding, presumably by virtue of hydrophilic interactions, on the implant material. That makes it possible to produce a chemotactically acting and/or biologically active, so-called juxtacrine implant surface which leads to colonisation, proliferation and differentiation of bone cells. It is thus possible to produce so-called active implants which, in relation to molecules liberated from the surface, exhibit a chemotactic action on cells, in the case of BMPs on osteoblasts, even at a distance of 500 to 1000 μm.

[0059] Preferably adequate loading of the oxidised metal surface is achieved by the peptides being applied in a physiological buffer solution in a concentration which is sufficient to achieve a loading of more than 200 ng/cm.sup.2, preferably more than 500 ng/cm.sup.2 and more preferably more than 1000 ng/cm.sup.2 of the peptide on the oxide surface of the metal implant.

[0060] In general that loading is achieved with a physiological buffer solution of peptides in a concentration of more than 1 μg/ml, preferably more than 200 μg/ml of buffer solution.

[0061] According to the invention the peptides are biomolecules which are advantageous in terms of biocompatibility of the implant insofar as they counteract possible rejection of the implant and/or promote the implant growing into place.

[0062] As mentioned hereinbefore preferably proteins from the class of TGF proteins, in particular the bone growth promoting proteins from the class of bone growth factors bone morphogenic proteins, or the class of vessel growth factors such as VEGF or angiotropin or also ubiquitin can be used as peptides. The term transforming growth factor (TGF) is used to denote in particular the group (subgroup) of (i) transforming growth factors beta (TGF-β) and the group (subgroup) of (ii) bone morphogenetic proteins (BMP). The latter are osteoinductive proteins which stimulate bone regeneration and bone healing insofar as they cause proliferation and differentiation of precursor cells to osteoblasts. In addition they promote the formation of alkaline phosphatases, hormone receptors, bone-specific substances such as collagen type 1, osteocalcin, osteopontin, osteonectin, bone sialoprotein (BSP) and finally mineralisation.

[0063] Advantageously for immobilisation purposes it is possible to use a protein of that class alone, in combination with further members of that class or also together with biomolecules such as proteins of other classes or low-molecular hormones or also antibiotics to improve immune rejection. In that case those further molecules can also be immobilised on the surface by way of bonds cleavable in the physiological medium.

[0064] It was already earlier found on the part of the inventors that the number of oxide groups can surprisingly be increased by the surface of the metal being treated with hot, preferably bottom sediment-free chromosulphuric acid. In contrast to the expectation that the metal dissolves under those conditions, a substantially uniform 5-50 nm thick hydrophilic oxide layer is produced on the surface of the metal when using that acid. The process is so careful that even coronary vessel supports, referred to as stents (which for example can be made from high-quality steel or titanium) can be coated without destroying the thin delicate mesh (50-150 μm diameter). In particular transition metal surfaces cleaned with dilute acid such as titanium, steel, steel alloys such as Cr-Mo-steel or steel or pure titanium surfaces or titanium alloys, treated with chromosulphuric acid, are suitable as materials for the oxide treatment.

[0065] Both in the case of polished implants and also in the case of sand-blasted (SLA-surfaces) implants or implants coated with metal plasmas (for example titanium plasma spray or TPS), the ultrahydrophilic oxide layer, after treatment of the metal surface, under defined conditions, can be of a thickness of 10 nm up to 300 nm and can be constructed in the form of nanostructures, as shown in FIG. 2, of different geometries (for example round or polygonal). In that respect pure titanium or titanium alloys (for example (TiAlV4, TiAlFe2,5), aluminium or stainless steel (for example V2A, V4A, chromium-nickel 316L, Cr-Mo-steel) can be used as metal for the implant. A commercially available chromosulphuric acid with 92% by weight H.sub.2SO.sub.4, 1.3% by weight of CrO.sub.3 and of a density of 1.8 g/cm.sup.3, as is available for example from Merck, is preferably used for producing a thin smooth layer of metal oxide.

[0066] The novel process according to the invention allows ultrahydrophilic surfaces to be produced in all cases, in contrast to earlier processes. The three preferred process steps: (i) the novel CSS treatment (shock heating), (ii) quenching in concentrated sulphuric acid, and (iii) the novel EDTA washing method reduce the chromium content (EDX method) on the surface below the detection limit. The EDX detection limit is at 0.2-0.5 atom %. The novel chromium-free ultrahydrophilic surfaces exhibit the particular novel properties set forth hereinafter in respect of BMP-2 bonding and stabilisation by the salt layer.

[0067] If a thicker metal oxide layer (>1000 nm) is to be provided at the metal surface and/or preferably an oxide layer with small micro- and nanopores the above-described chromosulphuric acid is diluted with water to a density of 1.5 to 1.6 g/cm.sup.3. In a treatment, which then follows as described hereinbefore, of the metal implant surface with the acid diluted in that way, a “rough” surface layer with depressions and pores is formed, so that the surface available for loading with peptides is increased in size. By adjusting different densities in respect of the chromosulphuric acid and different treatment times and temperatures it is therefore possible to apply a multiplicity of different oxide layers with different properties to metal surfaces, with a high level of adhesive strength.

[0068] The ultrahydrophilic surface produced by the chromosulphuric acid can impart the hydrophilic properties in respect of prolonged storage in air and in pure water. Under those conditions the contact angle can rise after 1-2 hours to values of 20-40°. The ultrahydrophilic surface can be stabilised in accordance with the invention by means of a salt solution as the stabilisation agent. In accordance with the invention it is also alternatively possible to use as such stabilisation agents, alcohols in the homologous alkane, alkene and alkine series which can be straight-chain or branched and can have up to 20 carbon atoms, in particular up to 6 carbon atoms, in particular water-free methanol and ethanol, as well as phenolic compounds, the latter also in aqueous solution. Preferably, as mentioned hereinbefore, stabilisation is possible by various aqueous salt solutions which can be ordered in accordance with their salting-out effect in relation to proteins (Table 2). These involve for example the anions SO.sub.4−, HPO.sub.4.sup.−−, CH.sub.3COO.sup.−, Cl.sup.−, Br.sup.−, NO.sub.4.sup.−, ClO.sub.4.sup.−, CNS.sup.−, ClCH.sub.2COO.sup.−, F.sub.3CCOO.sup.−, Cl.sub.2CHCOO.sup.−, Cl.sub.3CCOO.sup.−, Br.sub.3CCOO.sup.− or the cations NH.sub.4.sup.+, Rb.sup.+, K.sup.+, Na.sup.+, Cs.sup.+, Li.sup.+, Mg.sup.++, Ca.sup.++, Ba.sup.++ as well as tetraalkylammonium cations like (CH.sub.3).sub.4N.sup.+, (C.sub.2H5).sub.4N.sup.+, (C.sub.3H7).sub.4N.sup.+, (C.sub.4H9).sub.4N.sup.+. NaCl salt solutions above 0.15 mol/l are preferred, particularly preferred above 0.5 mol/l, quite particularly preferred in the region of 1 mol/l. The ultrahydrophilic surfaces are almost unlimitedly stable in such solutions. Such high levels of salt concentration occur for short times during evaporation even from dilute buffer mixtures as specified hereinbefore. A preferred salt concentration in the buffer solution is 135 to 140 mM NaCl, 8 to 8.2 mM Na.sub.2HPO.sub.4, 2.6 to 2.8 mM KCl, 1.4 to 1.6 mM KH.sub.2PO.sub.4 at a pH in the range of 7.3 to 7.5. Evaporation to dryness leads to high local levels of salt concentration. In that respect the HPO.sub.4.sup.−−, which has substantially stronger salting-out properties than Cl.sup.− can exert a particular stabilising influence on the oxide layer.

[0069] The invention is therefore also directed to a process of making the implants provided with an oxide layer with nanostructures storage-stable by means of such “stabilisation agents”.

[0070] In the most general form the present invention therefore also concerns a process for stabilisation of the ultrahydrophilic surfaces by shielding the surfaces from influences which detrimentally influence ultrahydrophilicity. Thus an embodiment of the process according to the invention is also directed to the implant with a hydrophilic surface being put into a solvent which contains dissolved therein a coating agent which detrimentally influences the ultrahydrophilic surface neither in solution nor in the coating. The solvent is evaporated and the coating agent remains behind on the implant with the ultrahydrophilic surface and encloses the implant. In that way the implant can reliably preserved for long-term storage. An embodiment of the solution with coating agent can be the above-described aqueous salt solution which easily acquires salting-out properties upon evaporation. A further embodiment can be a solution of a zwitterionic organic substance, for example an amino acid, for example glycine, which can have a similar salting-out effect as SO.sub.4.sup.−−, HPO.sub.4.sup.−−. Further non-volatile organic substances can be polyhydric alcohols like glycerine or monosaccharides like glucose and also disaccharides like sucrose as well as inositols which also have a strong influence on the water structure of a surface and which afford a coating after evaporation of the solvent.

[0071] The implants coated in accordance with the invention have long-term storage stability and can be used after washing off the coating of salts or organic coating agent for loading with the peptides acting as mediators.

[0072] The invention is thus also directed to a process for loading the surface of an implant with peptides, in which peptides are applied to the surface of the implant, which are immobilised on the surface of the implant as a result of physisorptive or chemisorptive interactions between the peptides and the ultrahydrophilic surface of the implant.

[0073] In that respect the peptides are used in a physiological buffer solution at a concentration which is sufficient to achieve a loading of more than 200 ng/cm.sup.2, preferably more than 500 ng/cm.sup.2 and more preferably more than 1,000 ng/cm.sup.2 of the peptide on the oxide surface of the metal implant.

[0074] In general the peptides are used in a physiological buffer solution in a concentration of more than 1 μg/ml, preferably more than 200 μg/ml of buffer solution.

[0075] In accordance with the invention growth factors from the class of TGF proteins, in particular the BMP proteins, preferably BMP-2 or BMP-7, the vessel growth factors such as VEGF or angiotropin, ubiquitin, antibiotics or mixtures thereof are used as peptides.

[0076] If the mediators used are difficult to dissolve in the medium under the coupling conditions, solubility can be increased by the addition of surfactants and/or detergents and the reaction can be performed. Thus, bone growth factors and other mediators which are difficult to dissolve, at pH-values >6, can be kept in solution by ionic or non-ionic detergents in the concentration range of 0.05-10%, preferably 1-5% by weight, in particular at 0.066% SDS at pH-values >6, in particular at pH 8-12, quite particularly pH 9-11, especially pH 10.0, for bonding processes in the alkaline pH range without a loss in biological activity. Thus the invention is also directed to a process for the production of implants coated with peptides, in which the implant with an ultrahydrophilic surface is treated with a preferably alkaline buffer solution containing one or more detergents. Such a process can include in particular a treatment with an NBS buffer comprising 125 mM Na-borate buffer, 0.066% sodium dodecylsulphate (pH 10.0).

[0077] Accordingly the invention is also directed to a process of loading implants with bone growth factors, in particular BMP-2, in which the ultrahydrophilic surface of the implant is treated with a solution of the bone growth factor at a pH-value of 9 to 11, preferably 10. For that purpose for example an implant, for example the implant removed from the dry packaging and still covered with the exsiccation layer, is preferably directly treated with a buffered solution of the bone growth factor at a pH-value of 9 to 11, preferably 10, without the exsiccation layer having to be previously washed off.

[0078] Surprisingly therefore the inventors succeeded in producing a coating on the ultrahydrophilic surface of an implant which is selected from metallic materials such as pure titanium, metallic titanium alloys, chromium/nickel/ aluminium/vanadium/cobalt alloys (for example TiAlV4, TiAlFe2,5), high-quality steels (for example V2A, V4A, chromium-nickel 316L), ceramic materials, in particular hydroxyapatite, aluminium oxide or a combination of the metallic materials with ceramic materials thereof, in which the metallic material is in the form of a composite material with ceramic material, with bone growth factors, in particular BMP-2, wherein the coating on the ultrahydrophilic surface is performed in aqueous buffered solution either in the acid range in the range between pH 4 and 5, in particular at pH 4.5, or in the weakly alkaline range between pH 9 and 11, preferably pH 10. The coating operation in the alkaline range can advantageously be effected in the presence of detergents such as SDS.

[0079] A particularly preferred embodiment of the process according to the invention provides for applying to the ultrahydrophilic surface of the implant BMP-2 or BMP-7 in a physiological buffer solution in a concentration of more than 1 μg BMP-2 or BMP-7/ml buffer solution, preferably more than 200 μg BMP-2 or BMP-7/ml buffer solution. Those aforementioned concentrations are generally sufficient to achieve a loading of more than 200 ng BMP-2 or BMP-7/cm.sup.2, preferably more than 500 ng BMP-2 or BMP-7/cm.sup.2 and more preferably more than 1000 ng BMP-2 or BMP-7/cm.sup.2 of the peptide on the oxidised surface of the metal implant.

[0080] The implants with an ultrahydrophilic surface, produced by means of the process according to the invention, are also subject-matter of the invention. Thus the invention also concerns implants in which the implant material comprises titanium, titanium alloys, aluminium, stainless steel, steel alloys, chromium-bearing alloys, ceramic materials such as hydroxyapatite or combinations thereof. In that case the implant can be a joint or bone prosthesis, a dental implant or in particular a coronary vessel support coated with a peptide, for example BMP-2 (a so-called coronary stent, length about 10 mm) to therapeutically prevent or alleviate the later complication of restenosis caused by proliferation of smooth vessel muscle cells, in order thereby to promote healing and compatibility.

[0081] The influence of the materials modified in accordance with the process of the invention on bone cells was investigated in animal experiments, the modified materials being produced for that purpose in plate or dumbbell form. It was observed in that case that, 4 weeks after being introduced into the animals, accelerated bone formation occurred with contact in relation to the implant surface by BMP-2 on the materials.

[0082] The present invention is set forth in further detail by reference to the following examples.

Modification of Metals (Titanium, 316L Stainless Steel):

[0083] The experiments described hereinafter involved the use of either mechanically polished/electropolished, anodically oxidised titanium plates, titanium alloy plates pre-etched with other acids, sand-blasted or plasma-sprayed with porous titanium alloy, with or without chromosulphuric acid treatment. Equally stainless, mechanically polished/electropolished steels are used, with or without chromosulphuric acid treatment.

Cleaning Process

[0084] Prior to each use the materials were cleaned by heating at 80° C. in 5% HNO.sub.3 for 2 hours. After renewed washing in water the plates were dried by washing in 30 ml in dry methanol. Thereafter they were either directly further used or treated with chromosulphuric acid.

Chromosulphuric Acid Treatment

[0085] In the chromosulphuric acid treatment the titanium plates were shock-heated at 210-240° C. in chromosulphuric acid (92% H.sub.2SO.sub.4, 1.35 CrO.sub.3), incubated for 30-90 min at that temperature and then quenched with concentrated sulphuric acid at room temperature. Thereupon the metal samples were washed with 10×50 ml water, treated with 2×30 min 10% EDTA ((pH 7) ultrasound) and then 1-3×30 min in boiling 10% EDTA (pH 7) and thereafter washed for 30 min with boiling water and rinsed with water. That resulted in an ultrahydrophilic surface which is substantially chromate-free, that is to say no free chromium ions can now be detected on the surface.

EXAMPLE 1

Immobilisation of rhBMP-2 on Ultrahydrophilic Titanium Plates

[0086] The pretreated titanium plates were washed with 125 mM Na-borate buffer, 0.066% sodium dodecylsulphate, pH 10.0, and equilibriumed. BMP-2 which was initially present in 50 mM tris, pH 8.0, 1000 mM NaCl, 5 mM EDTA, 33 mM 3-[(3-cholamido-propyl)dimethylammonio]-propane sulfonic acid buffer (=CPDP buffer) was dialysed in relation to 125 mM Na-borate buffer, 0.066% sodium dodecylsulphate, pH 10.0 (=NBS buffer) and incubated in a concentration of 0.2-0.3 mg/ml for 12-14 hours at room temperature with shaking with the titanium plates. Thereupon they were washed 4× with borate buffer and then with water.

TABLE-US-00001 TABLE 1 immobilisation of rhBMP-2 on HNO3-treated or chromosulphuric acid-treated titanium plates (5 × 10 × 1 mm) Immobilised 125I-rhBMP-2 Modification of electropolished titanium [ng/cm2] HNO3-treated (θ.sub.A/θ.sub.R = 40°/20°)  273 ± 107 (4) ultrahydrophilic (θ.sub.A/θ.sub.R = 1°/1°) 1272 ± 636 (4) Immobilisation buffer: 125 mM borate/0.066% SDS, pH 10.0, C.sub.rhBMP-2 = 0.25 mg/ml, n = 4. θ.sub.A: dynamic advancing angle, θ.sub.R: dynamic receding angle

TABLE-US-00002 TABLE 2 ordering of the salts according to their salting- out and salting-in effect in relation to proteins Salting-out effect               Salting-in effect Anionic: SO.sub.4.sup.−− > HPO.sub.4.sup.−− > CH.sub.3COO.sup.− > Cl.sup.− > Br.sup.− > NO.sub.4.sup.− > ClO.sub.4.sup.− > I.sup.− > CNS.sup.− CH.sub.3COO.sup.− > ClCH.sub.2COO.sup.− > F.sub.3CCOO.sup.− > Cl.sub.2CHCOO.sup.− > Cl.sub.3CCOO.sup.− > Br.sub.3CCOO.sup.− Cationic: (CH.sub.3).sub.4N.sup.+ > NH.sub.4.sup.+ > Rb.sup.+, K.sup.+, Na.sup.+, Cs.sup.+ > Li.sup.+ > Mg.sup.++ > Ca.sup.++ > Ba.sup.++ Cationic/ hydrophobic (CH.sub.3).sub.4N.sup.+ > (C.sub.2H5).sub.4N.sup.+ >> (C.sub.3H7).sub.4N.sup.+, (C.sub.4H9).sub.4N.sup.+

[0087] The action of the salts occurs inter alia by way of the water structure. The salts stabilise or destabilise the ultrahydrophilic surface by way of the water molecules bonded to the surface and ionic groups. NaCl salt solutions involving 0.15 mol/l are preferred, particularly preferably 0.5 mol/l, quite particularly preferably in the region of 1 mol/l.

EXAMPLE 2

Liberation of rhBMP-2 from Ultrahydrophilic Titanium Plates

[0088] As shown in FIG. 3 the adsorption of rhBMP-2 on an ultrahydrophilic titanium surface and liberation from the surface is markedly improved in comparison with a titanium surface only treated with dilute HNO3 for cleaning purposes, as can be seen by reference to the liberation kinetics, shown in FIG. 3, of rhBMP-2 from an ultrahydrophilic titanium surface. The great capacity differences will also be clear. The illustrated liberation curves can be adapted with a 3-phase exponential function. In the case of the control there are actually only 2 phases. Liberation has been measured over 62 days. The half-value times of liberation and the amounts of rhBMP-2 liberated are set out in Table 3.

TABLE-US-00003 TABLE 3 Liberation of rhBMP-2 from Ultrahydrophilic Titanium Plates Ultrahydrophilic titanium HNO3-treatment surface (θ.sub.A/θ.sub.R = 40°/20°) (θ.sub.A/θ.sub.R = 1°/1°) Γo = 181 ng/cm.sup.2 Γo = 1551 ng/cm.sup.2 Liberated Liberated T½ amount/day T½ amount/day Liberation phase days ng/day days ng/day 1st phase (2 0.28 36 0.37 193 days) 2nd phase (42 39 0.5 33 14 days) 3rd phase (18 39 0.5 231 7 days) Total amount 102 1100 liberated in 62 days: Γo: Immobilised rhBMP-2 amount/cm.sup.2 at the time t = 0.

[0089] As can be seen from FIG. 4 the scanning electron-microscope recordings of chromosulphuric acid-treated SLA titanium plates (14×14×1.5 mm) after gamma sterilisation in an exsiccation buffer (60 min CSS with HNO.sub.3, with quenching, gamma sterilised in PBS, θ=0°) show a sterilised ultrahydrophilic titanium oxide surface, provided with microcaverns, with a protective layer of dried-in exsiccation buffer which after washing off of the “exsiccation protective layer” gives a purely ultrahydrophilic titanium oxide surface as, as shown in FIG. 5, the EDX analyses of an ultrahydrophilic plate with exsiccation layer after gamma sterilisation (A) and after removal of the exsiccation layer with water (B) show.

[0090] The storage stability of the ultrahydrophilic surfaces is shown by means of the dependency of the dynamic contact angles of gamma-sterilised, ultrahydrophilic SLA titanium plates (14×14×1.5 mm) provided with an exsiccation layer according to the invention, on the storage time as set forth in Table 4, wherein the reference to SLA titanium plates is used to denote titanium plates which have sand-blasted and acid-etched surfaces. As shown, an “unprotected” hydrophilic surface is already less hydrophilic after a few hours in air while the contact angles of the gamma-sterilised, ultrahydrophilic SLA titanium plates provided with the exsiccation layer according to the invention are almost constant without change at 0° after up to 24 weeks storage.

TABLE-US-00004 TABLE 4 Sample prior to chromosulphuric Storage time after chromosulphuric acid treatment acid treatment (exsiccation-coated and gamma-sterilised) 18 hours control 6 15 4 8 18 24 stored in air (0-days) days days weeks weeks weeks weeks Dynamic contact angles according to Wilhelmy Sample θ.sub.A θ.sub.R θ.sub.A θ.sub.R θ.sub.A θ.sub.R θ.sub.A θ.sub.R θ.sub.A θ.sub.R θ.sub.A θ.sub.R θ.sub.A θ.sub.R θ.sub.A θ.sub.R SLA-1 59.2 0.0 0.0 0.0 0.0 0.0 6.1 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 SLA-2 53.3 0.0 0.0 0.0 5.7 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 SLA-3 100.8 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 SLA-4 94.3 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 SLA-5 86.5 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 Average 78.8 0.0 0.0 0.0 1.1 0.0 1.2 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 value (□) Standard 21.3 0.0 0.0 0.0 2.5 0.0 2.7 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 deviation (s)