Digest Journal of Nanomaterials and B iostructures V ol. 6, No 2, April – June 2011, p. 703 – 707 [622469]

Digest Journal of Nanomaterials and B iostructures V ol. 6, No 2, April – June 2011, p. 703 – 707

TITANIUM DIOXIDE NANOTUBES ON SILICON WAFER DESIGNATED FOR
GOX ENZYMES IMMOBILIZATION

C. RAVARIUa*
Anodization stands for a simple and low cost method to synthesize TiO 2 by
electrochemical oxidation of a metallic Ti film, deposited on a silicon wafer. In this way, a Ti -film, , E. MANEAb, C. PARVULESCUb, F. BABARADAa ,
A. POPESCUb
aFaculty of Electronics, University Politehnica , Splaiul Independentei 313,
Bucharest , Romania
bInstitute of Microtechnology, Str. Erou Iancu Nicolae 32B , Bucharest , Romania

The down -scaling of the integrated biosensors demands new methods for the enzyme
immobilization, using nanostructured materials. This work presents the preparation method for th e TiO2 nanotube by anodization and its applications to enzyme entrapping.
Two kinds of electrolytes were investigated : oily and aqueous. When the electrolyte is
glycerin the achieved TiO2 nanotubes are crown in some individual fasciculus, with
weaker inter -links. For the glucose -oxidase enzyme immobilization, nafion is used as
crosslink polymer . The surface characterization is operated by FEG SEM analysis.

(Received March 11, 2011; Accepted April 05, 2011)

Keywords : nanotube, TiO2, enzyme entrapping

1. Introduction

The synthesis of the nanostructured materials is one of the challenges among
nanotechnologies, [1]. The applications of these nanomaterials cover a large spectrum, especially in bioscience: drug delivery [2, 3, 4], nanomedicine [5], cell growth [6], or optical properties [7].
A dedicated section concerns nanostructured materials compatible with the Si -technology, in order
to make possible the integrated biosensors manufacturing, [8]. One of the immobilization methods for the biodetecting layer s uses nanoporous mater ials as adh erent substrate s. The enzymatic
membra nes are entrapped in organic compounds , [9]. On the other hand, titanium is a prevalent
material used in pro sthesis due to its bio- compatibility with the living matter , [10] . This paper
presents the electro -chemical method for the TiO
2 nanotube grown on a Si-wafer and its
application s in biosensor s technology. The influence of the electrolyte composition on the pores
nanostructure and morphology of the titanium dioxide layer, w ere investigated by SEM.

2. Experimental set- up

Among the TiO 2 layers growth methods are the sol -gel deposition [ 11] or oxidation by
anodization of a thin Ti film, [ 12]. Although the oxidation of thin folium of titanium offers in
literature many processing solutions, these methods ar e difficult to be applied to silicon integrated
micro and nano- electronic devices.
In this section are described the preparation condition for a TiO 2 layer manufactured by
anodization onto a silicon substrate. In order to ensure the compatibility with the Si-technology,
the Titanium dioxide nanotubes growth, firstly imposes a thin film deposition.

* Corresponding author: cr682003@ yahoo.com

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over 90nm, was deposited by sputtering on a p- type Si -substrate with 1 -2Ωcm resistivity. During
this process, the argon gas flow was 2.5 sccm, while the base chamber pressure was 1.82mPa.
Then, the Ti metal, was converted into TiO 2 nanostructured material, by anodization. The
electrochemical cell consists of three electrodes: a platinum sheet 7×10 cm2 with a Pt wire used as
a counter electrode, a standard calomel (Hg/HgCl 2) as a reference electrode and a work electrode
composed by a Teflon sealing holder with a direct voltage contact applied on the titanium surface
film. The work electrode (anode) and the counter electrode (cathode) were placed at 2÷ 4cm
distance in an electrolyte solution. During the anodization, the potential drop was varied from 5V
to 30V from an open circuit source, OCS, with a variable rate 0.1 ÷ 0.5 V/s. The optimal pH was
kept between 6.4÷ 7.2.
Before anodization, the samples were degreased in acetone using an ultrasonic bath for
four minutes and then rinsed in de -ionized water. After anodization, the samples were immediately
washed with a large amount of de ionized water and subsequently dried in nitrogen atmosphere.
The electrolyte was sometimes oxalic acid and phosphoric acid as aqueous solutions (type –
1), and other times salts in glycerin (type -2).
Immediately after the anodization, the TiO 2 layers are amorphous.
The morphology of the nanostructured TiO 2 was analyzed by a Field Emission Gun
Scanning Electron Microscopy (FEG SEM), one of the latest version of SEM technology that
combine the thermal emission technology using a newly developed Energy and angle selective
Backscattered detector (EsB).

3. Results

3.1. The TiO 2 growth by anodization

Because the TiO 2 layers are amorphous after anodization , the samples were treated at
different temperatures, in order to convert the oxi de in a crystalline f orm. For instance, TiO 2
anatase phase results , after annealing at 773K. The fabricated structures present vertical nanotubes,
with 100 – 400 nm width and a homogenous distribution.
Figure 1 presents the TiO 2 nanotubes developed on the Si -wafer, when the electrolyte was
sodium sul fate, glycerin, water and sodium fluoride 0.4wt%, at 30V , so type -2. The morphology is
similar with that achieved for completely aqueous electrolytes (type -1).

Fig. 1. FEG SEM images in a cross section.

The FEG SEM analysis revealed that the morphology of the TiO 2 nanotubes is depending
on the electrolyte formula. Figure 2 presents the FEG SEM image of a TiO 2 nanotubes layer made
in completely oily glycerin solution of electrolyte (ammonium fluoride in glycerin) , at 10V
anodization voltage . The nanotubes are crown in some individual fasciculus, with weaker inter –
links.
The best morphology of the TiO 2 nanotubes is achieved when the anodization occurs in
aqueous electrolytes, versus the case when the solvent is the glycerin . Th e most probable
explanation is related by the ions mobility in aqueous and oily solutions, [13 ].

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Fig. 2. Lateral view by FEG SEM of a TiO 2 nanotubes layer anodized at 10V.

Therefore, the diffusion rate of the ions in water is superior to that in gly cerin, with an
increased growth rate in this first environment.

3.2. Enzyme entrapping

The TiO 2 nanostructured material was grown on the Si -wafer, in order to offer a bio-
compatible inorganic support for enzymes entrapping, like glucose -oxidas e, GO X. The surface
characterization is continuing to be ensured by SEM.

Fig. 3 . SEM imag e of the probe with: glucozoxidase / nafion / TiO 2 / SiO 2 / Si.

Usually, the GOX entrapment on a solid surface, is possible via the cross- link method,
using a glutaraldehy de solution (GA) – 2,5% concentration, as polymerization agent and serum
albumin from bovine provenience (BSA) , [14]. In these tests, another polymer was researched –
nafion – in the same concentration as the glutaraldehyde . All the time, a buffer soluti on ensures a
constant pH=7.0. We use nafion – that is a fluoropolymer -copolymer based on sulfonated
tetrafluoroethylene because it is presenting an excellent mechanical and thermal stability, [ 15].
Fig. 3 presents the SEM image with the successive films from the Si -substrate up to GOX
enzyme. The GO X membrane adheres to the substrate, but is still scant y to the homogeneity . A
possible improvement could be the increasing of the nafion quantity.

4. Discussion

The first instance for the enzymatic membrane characterization placed on nanoporous
TiO2 appeal s to the probe spectrum.

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Fig. 4. The probe spectrum: glucose -oxidase + nafion +TiO 2/SiO 2/Si.

In fig ure 4 is presented a probe spectrum after GOX immobilization in nafion. The water
spectral bands interf ers with target bands and make difficult the interpretation. After, the water
bands extraction, two bands can be observed, with the following explanation: I -st amide (C=O),
1656 cm-1 and respectiv ely II -nd amid e (N-H), 1517 cm-1 from gluco se-oxida se. The bands from
1227 cm-1 and 1152 cm-1 can be atributed to the vibration mode for C -O or C-F, signifying the
nafion bounds. In the 750÷ 550 cm-1 domain, can be observed characteristic bands associated to
the Ti -O bound. They are deviated bands comparatively to the substrate spectrum that suggest a
TiO2 anchoring to substrate, establishing Ti -O-C bounds in 976 cm-1 spectrum.

5. Conclusion s

A nanostructured titanium di oxide on silicon was manufactured. The best TiO
2 nanotubes
were manufactured by an anodizati on process at potentials of 5 ÷10V, maintaining a pH of 6.4 ÷7.4,
preferably acid. The quantity of solvent must be enough in order to let free the anions and cations
in solution, taken into account that the upper limit concentration of the (NH 4F) salt in wat er is
40%, for saturated solution. An experimental study proofed different TiO 2 morphologies for
aqueous electrolyte or glycerin.
The TiO 2 nanotube layer has a double advantage for the glucose -oxidase immobilization:
high enough adsorbent properties for t he GOD enzyme and good electro -oxidation catalytic
properties for some o rganic substances, due to its biocompatibility with the living matter.

Acknowledgements

Project s PNII 12095, 62063, POSDRU 62557.

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