electrophoretic deposition of calcium phosphate

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i 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.wiley-vch.de/home/muw

Electrophoretic deposition of calcium phosphatenanoparticles on a nanostructured silicon surface

Elektrophoretische Abscheidung von Calciumphosphat-Nanopartikelnauf einer nanostrukturierten Siliciumoberfl�che

M. Epple1, M. Neumeier1, D. D�rr2, R. LeHarzic2, D. Sauer2, F. Stracke2, H. Zimmermann2

Nanostructured silicon surfaces were electrophoretically coated with calcium phosphate nano-particles. Positively charged calcium phosphate nanoparticles were synthesized by precipitationand then functionalized with poly(ethyleneimine). This permits the electrophoretic depositionon a conductive surface from a dispersion in 2-propanol. The following parameters affected thedeposition of the nanoparticles: The deposition potential, the deposition time, and the deposi-tion temperature.

Keywords: Calcium phosphate / nanostructures / electrophoretic deposition /

Nanostrukturierte Silicium-Oberfl�chen wurden elektrophoretisch mit Calciumphosphat-Nano-partikeln beschichtet. Die positiv geladenen Calciumphosphat-Nanopartikel wurden durch F�l-lung und anschließende Funktionalisierung mit Poly(ethylenimin) hergestellt. Dies erlaubt dieelektrophoretische Abscheidung aus der Dispersion in 2-Propanol. Der Einfluss der ParameterSpannung, Zeit und Temperatur auf die Abscheidung wurde untersucht.

Schl�sselw�rter: Calciumphosphat / Nanostrukturen / elektrophoretische Abscheidung /

1 Introduction

For bone implants, the interaction between cells and implantsurface is very important. When the osteoblasts have a good con-tact to the implant surface, osteointegration is improved [1]. Cellscan also sense a micro- or nanostructuring of the underlying sur-face. Cell orientation was observed on titanium and hydroxyapa-tite [2], polystyrene [3], silicon [4] and even PTFE [5]. Only the sur-face texture is important for cell adhesion and orientation [6]. Itis known that micro- and nanostructured surfaces influenceimportant factors like cell adhesion, proliferation, or orientation[7–12].

We used silicon wafers structured with laser-induced periodicsurface structures (LIPSS) [13–17] as a reusable stamp to formnanostructured and bioactive surfaces. Calcium phosphate nano-particles are suitable as bioactive coating because calcium phos-phate is the inorganic component of human hard tissue [18–20].Calcium phosphate nanoparticles can be functionalized with

nucleic acids or polymers and serve as carriers for non-viral genedelivery [21–25]. We have demonstrated earlier that they can bedeposited on conducting surfaces by electrophoresis and alsostructured by laser direct writing on the lm-scale [26, 27].

The aim of this study was to electrophoretically deposit func-tionalized calcium phosphate nanoparticles inside the ripples ofa nanostructured silicon wafer to achieve a defined orientationand surface density of bioactive nanoparticles (see Ref. [28] for acomprehensive review on this topic).

2 Materials and methods

Laser induced periodic surface structuring (LIPSS) was perform-ed on silicon substrates with a Ti:sapphire tunable (kex = 690–1060 nm) laser system with an excitation wavelength of 800 nm,a pulse duration of s = 140 fs (FWHM), a high repetition rate(80 MHz), and low energy (Emax ~ 40 nJ pulse–1) [13–15]. Calciumphosphate nanoparticles were synthesized by precipitation in thepresence of poly(ethyleneimine) (PEI). An aqueous solution of18 mM calcium-L-lactate and an aqueous solution of 10.8 mMdi-ammonium hydrogenphosphate (both with pH 9, adjustedwith NH3) were pumped together in a three necked flask,equipped with an overflow, with a peristaltic pump. A PEI solu-tion (2 g L–1) was added with a second peristaltic pump. The vol-ume flow ratio of the solutions was Ca : PO4

3– : PEI = 1:1:2. Thedispersion was filtered with a Buchner funnel immediately afterpassing the overflow. The filtered calcium phosphate nanopar-

1 Inorganic Chemistry and Center for Nanointegration Duisburg-Essen(CeNIDE), University of Duisburg-Essen

2 Fraunhofer Institute for Biomedical Engineering (IBMT) and Chair forMolecular and Cellular Biotechnology, University of Saarbruecken, St.Ingbert

Corresponding author: Prof. Dr.-Ing. M. Epple, Inorganic Chemistry andCenter for Nanointegration Duisburg-Essen (CeNIDE), University ofDuisburg-Essen, Univerit�tsstraße 5–7, 45117 EssenE-mail: [email protected]

DOI 10.1002/mawe.201100730 Mat.-wiss. u. Werkstofftech. 2011, 42, No. 1

Mat.-wiss. u. Werkstofftech. 2011, 42, No. 1 Electrophoretic deposition of calcium phosphate nanoparticles

ticles were then washed with water and freeze-dried. For electro-phoretic deposition, the nanoparticles were redispersed in 2-propanol (1 mg mL–1) by ultrasonification. A nanostructured sili-con wafer (1 cm2) was used as cathode, and a stainless steelbeaker with a volume of 1.63 mL was used as anode for the elec-trophoretic deposition. Unless otherwise noted, all depositionexperiments were carried out at room temperature.

Size measurements and zeta potential measurements of theaqueous nanoparticle dispersion were performed with a MalvernZetasizer Nano ZS with a laser wavelength of 633 nm. Scanningelectron microscopy (SEM) was performed with a FEI QuantaFEG 400 instrument.

3 Results and discussion

An area of 1 cm2 was structured by LIPSS on the silicon, consist-ing of individual squares of (230 lm)2. Each field consisted ofparallel periodic nanostructures with a periodicity of approxi-mately 110 nm, Fig. 1. The PEI-functionalized calcium phos-phate nanoparticles were spherical in shape and had a diameterbetween 70 and 100 nm, Fig. 2. Dynamic light scattering gave anaverage diameter of 75 nm and a zeta potential of +38 mV, indi-cating an electrostatic stabilization and the possibility to electro-phoretically deposit these nanoparticles at the cathode. Their sizeis appropriate to fit into the ripple structure.

To avoid water electrolysis, the nanoparticles were redispersedin 2-propanol for the electrophoretic deposition. The parametersaffecting the deposition were varied: Deposition time, potential,and temperature. Figure 3 shows the effect of different depositionpotentials. The deposited amount of nanoparticles stronglydepended on the deposition voltage used. With a potential of

30 V the deposited layer was irregular, the nanoparticles formedaggregates on the surface. After deposition with 50 V there was athick layer of nanoparticles on the surface and the nanostruc-tured area was completely covered with particles. The cracks inthe deposited layer after deposition with 50 V are due to drying ofthe layer after deposition.

The influence of the deposition time on the coating structurewas small, only the amount of deposited particles increased with

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Figure 1. Scanning electron micrograph of a structured silicon surface: Left: Five adjacent squares; right: Magnification of a single field, showingthe individual ripples. Note that the whole surface was structured by parallel ripples and that the non-structured areas in the left figure serveonly as illustration of the method.

Bild 1. Rasterelektronenmikroskopische (REM) Aufnahme einer strukturierten Silicium-Oberfl�che: Links: F�nf benachbarte Quadrate, rechts:Die Vergr�ßerung eines Quadrats zeigt die individuellen ripples. Es ist zu beachten, dass die gesamte Oberfl�che mit parallelen ripples struktur-iert wurde. Die unstrukturierten Bereiche im linken Bild dienen nur der Illustration der Methode.

Figure 2. Scanning electron micrograph of PEI-functionalized calciumphosphate nanoparticles.

Bild 2. REM-Aufnahme von PEI-funktionalisierten Calciumphosphat-Nanopartikeln.

M. Epple et al. Mat.-wiss. u. Werkstofftech. 2011, 42, No. 1

increasing deposition time, Fig. 4. The silicon surface was stillvisible below the aggregated nanoparticles.

To deposit the nanoparticles only within the ripple structure,an AC potential was overlaid on the constant DC potential. Theaim was to give the deposited layer some mobility, i. e. to lower oreven reverse the adhesion of weakly bound nanoparticles (likethose sitting on the edges of the ripples) for some time to give thewhole layer more time to reach a stable state like a monolayer onthe substrate surface and within the ripples. A DC potential of5 V was overlaid with an AC potential of 10 V with 50 Hz, so thatthe effective potential changed from –5 V to +15 V.

The nanoparticle layers deposited with a combined DC/ACpotential and nanoparticles deposited with an equivalent DCpotential are shown in Fig. 5. With the combined potential thereare fewer particles deposited on the surface, but these particlesare better located along the ripples.

The temperature of the dispersion during electrophoretic dep-osition had an effect on the kinetic energy and the velocity of thenanoparticles in dispersion. The deposition from a dispersionheated up to 323 K (50 8C) resulted in more particles depositedon the surface. These particles also had a better orientation alongthe ripples. There was still an orientation of the nanoparticles

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Figure 3. Scanning electron micrographs of PEI-functionalized calcium phosphate nanoparticles deposited on nanostructured silicon waferswith 30 V for 30 s (left) and with 50 V for 30 s (right).

Bild 3. REM-Aufnahmen von nanostrukturierten Silicium-Wafern, beschichtet mit PEI-funktionalisierten Calciumphosphat-Nanopartikeln mit30 V f�r 30 s (links) und mit 50 V f�r 30 s (rechts).

Figure 4. Scanning electron micrographs of PEI-functionalized calcium phosphate nanoparticles deposited on unstructured silicon wafers with30 V for 30 s (left micrograph) and 60 s (right micrograph).

Bild 4. REM-Aufnahmen von PEI-funktionalisierten Calciumphosphat-Nanopartikeln, abgeschieden auf unstrukturierten Silicium-Wafern mit 30V f�r 30 s (linke Aufnahme) und 60 s (rechte Aufnahme).

Mat.-wiss. u. Werkstofftech. 2011, 42, No. 1 Electrophoretic deposition of calcium phosphate nanoparticles

after deposition at 70 8C, but the amount of particles depositedon the surface had decreased. Apparently, the particles had toomuch kinetic energy at higher temperatures so that the weaklybound particles desorbed from the surface.

4 Conclusions

Calcium phosphate nanoparticles functionalized with a polyelec-trolyte can be electrophoretically deposited on a LIPSS-nano-structured silicon surface. The amount of deposited nanopar-ticles strongly depended on the deposition potential, but the de-

position time had only a small effect on the structure of thedeposited layer of the nanoparticles. A combined DC/AC poten-tial led to smoother layers of nanoparticles on the structured sur-faces, possibly by giving weakly bound particles the chance tofind a more stable position within the ripples. A higher disper-sion temperature had a similar effect due to a higher kineticenergy of the nanoparticles.

Acknowledgement

We thank the Deutsche Forschungsgemeinschaft for fundingwithin the priority programme SPP1327.

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Figure 5. Scanning electron micrograph of PEI-functionalized calcium phosphate nanoparticles deposited on nanostructured silicon wafers witha combined DC/AC potential for 30 s (left) and with a comparable DC potential for 30 s (right).

Bild 5. REM-Aufnahmen von nanostrukturierten Silicium-Wafern, beschichtet mit PEI-funktionalisierten Calciumphosphat-Nanopartikeln miteiner Mischspannung f�r 30 s (links) und mit einer vergleichbaren Gleichspannung f�r 30 s (rechts).

Figure 6. Scanning electron micrograph of PEI-functionalized calcium phosphate nanoparticles deposited on nanostructured silicon wafers with10 V for 30 s from dispersions at 323 K (left) and 343 K (right).

Bild 6. REM-Aufnahmen von nanostrukturierten Silicium-Wafern, beschichtet mit PEI-funktionalisierten Calciumphosphat-Nanopartikelnaustemperierten Dispersionen bei 323 K (links) und 343 K (rechts) mit 10 V f�r 30 s.

M. Epple et al. Mat.-wiss. u. Werkstofftech. 2011, 42, No. 1

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Received in final form: November 15th 2010 T 730

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electrophoretic deposition of calcium phosphate

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