Applied Surface Science 256 (2009) 52–55

An active nano-supported interface designed from gold nanoparticles embeddedon ionic liquid for depositing DNA

Liping Lu *, Tianfang Kang, Shuiyuan Cheng, Xiurui Guo

College of Environmental and Energy Engineering, Beijing University of Technology, Beijing 100022, PR China

A R T I C L E I N F O

Article history:

Received 1 April 2009

Received in revised form 13 July 2009

Accepted 17 July 2009

Available online 25 July 2009

Keywords:

DNA

HCHO

Ionic liquid

Electrochemical sensor

A B S T R A C T

The use of an active nano-interface designed from gold nanoparticles embedded on ionic liquid for DNA

damage resulted from formalehyde (HCHO) is reported in this article. The active nano-interface was

fabricated by depositing gold nanoparticles on the ionic liquid 1-butyl-3-methylimidazolium

tetrafluroborate ([bmim][BF4]). A glassy carbon electrode modified by this composite film was

fabricated to immobilize DNA for probing into the damage resulted from HCHO. The modifying process

was characterized by X-ray photoelectron spectroscopy, atomic force microscopy and electrochemistry

involving electrochemical impedance spectroscopy. It was found that the modified film performs

effectively in studying the DNA damage by electrocatalytic activity toward HCHO oxidation.

� 2009 Elsevier B.V. All rights reserved.

Contents lists available at ScienceDirect

Applied Surface Science

journal homepage: www.e lsev ier .com/ locate /apsusc

1. Introduction

DNA, a very important biomolecule storing the geneticinformation, plays an essential role in the determination ofhereditary characteristics. From this point of view DNA isconsidered as the major target interacting with various molecules[1]. To detect DNA damage by electrochemistry, natural DNA isusually immobilized on the surface of electrodes. Researching theDNA in vitro, it is important that DNA keeps the nativeconformation, because its image approaches the biologicalenvironments. The inherent stability of biomolecules need keepin a desired environment. A critical issue in the development of aDNA-based biosensor is the sensor material that influences directlythe sensor response [2]. Since nanoparticles (NPs) and biomole-cules are typically on the same nanometer scale, many types of NPof different sizes and compositions now available facilitate theirelectrochemical-related applications in enzyme-based sensors,immunosensors and DNA sensors [3–6].

Recently, room temperature ionic liquids (RTILs) have beenextensively used in direct electrochemistry and electroanalysisfield due to their properties of facilitating direct electron transferreaction between proteins and electrode surface [7–11]. Apartfrom its conductive property, it has many other advantages such asrobust nature, easy patterning capability and excellent adhesionproperty to substrates [12]. Ohno et al. successfully construct acontinuous ionic liquid domain along the double-strand of DNA

* Corresponding author. Fax: +86 010 67391983.

E-mail address: lipinglu@bjut.edu.cn (L. Lu).

0169-4332/$ – see front matter � 2009 Elsevier B.V. All rights reserved.

doi:10.1016/j.apsusc.2009.07.056

[13], Pang’s group verified the electrostatic interaction betweenDNA and [BMIM]BF4 [14]. RTILs open up new opportunities for thedevelopment of biosensors, biomacromolecules, and so on.

To detect DNA damage by electrochemistry, the native of DNAimmobilized on the surface of electrode should be concerned.Surface characterization and modification of electrodes andelectronic materials are important criteria in developing formsof biosensor. Facile in vitro electrochemical detection of DNAdamage by toxic metabolites could be envisioned by stable solidelectrochemical sensors coated with DNA films [15]. In this paper,gold nanoparticles and RTIL are used as modified electrodematerials for detecting the HCHO induced DNA damage.

2. Experimental

2.1. Apparatus and reagents

Calf thymus DNA (ct-DNA) was purchased from Sigma.[BMIM]BF4 (>99%) was obtained from J&K Chemical Ltd. Theywere used as received without further purification. All otherchemicals were of analytical grade. Solution of DNA was freshlyprepared in 5 mM pH 7.4 Tris–HCl buffer solution (THB). Allsolutions were prepared using doubly distilled water.

All electrochemical measurements and impedance spectro-scopy were performed on an electrochemistry workstation (CHI-660A, CHI, USA). All electrochemical experiments employed athree-electrode cell with a glassy carbon electrode (GCE) asworking electrode, and a platinum wire auxiliary electrode and asaturated calomel electrode (SCE) reference electrode. Experi-ments were carried out at 20 � 2 8C.

L. Lu et al. / Applied Surface Science 256 (2009) 52–55 53

Scanning electron microscope (SEM) image was obtained on aJEOL 6500F scanning electron microanalyser (NEC). X-ray photo-electron spectroscopy (XPS) spectra were recorded on a spectro-meter using Mg Ka X-ray radiation. Atomic force microscopy(AFM) image were recorded on a SPA 300HV with silicon tip intapping mode at room temperature.

2.2. DNA deposition

A GCE (3 mm in diameter) was polished repeatedly with 1.0,0.3, and 0.05 mm alumina slurry, followed by successive ultrasoniccleaning in acetone and doubly distilled water for 5 min. Thecleaned GCE was treated by cyclic voltammetry scanned in the0.1 M H2SO4 solution between 0 and +1.0 V at a scan rate of50 mV s�1 for 10 cycles. Then, the electrode was immersed in pure[BMIM]BF4 solution for 10 h at 4 8C. After thoroughly rinsed withwater, electrodeposition process was accomplished with cyclicvoltammetry scanning between �0.2 and +1.0 V at a scan rate of50 mV s�1 for 15 cycles from a fresh solution containing 3 mMHAuCl4 and 0.1 M KCl. The Au nanoparticles and [BMIM]BF4

modified electrode was obtained and denoted as NG/RTIL/GCE.The ct-dsDNA deposition on NG/RTIL/GCE was conducted in

0.1 mg ml�1 ct-dsDNA solution and 5% (v/v) [BMIM]BF4 undercontrolled dc potential of 0.5 V for 15 min, it is denoted as ct-DNA/NG/RTIL/GCE.

3. Results and discussion

3.1. Surface morphology of the nano-interface

Changes of surface composition can affect surface chemistry.Fig. 1 displays the SEM image observed for NG/RTIL/GCE. It clearlyshows the uniform distribution of the gold nanoparticles on the[BMIM]BF4 film. The morphology of these nanoparticles is almostidentical with cube and the size of them is about 100 nm.

XPS spectra of the NG/RTIL/GCE are presented in Fig. 2. The Au(Fig. 2A), F, N, and C (Fig. 2B) peaks were observed, showing theevidence that the Au and [BMIM]BF4 have been immobilized on thesurface. Furthermore, Fig. 2C shows the deconvolution spectra ofthe C1s spectrum, which indicate the presence of C–C (284.8 eV),C–O (286.2 eV), COOH (288.52 eV) surface functional groups. Theseoxygen containing groups are attributed to the H2SO4 oxidationprocesses [16]. Since the construction of the [BMIM]BF4, the ionicliquid film is not formed by self-assembly. But the oxygen

Fig. 1. SEM image of NG/RTIL/GCE.

Fig. 2. XPS of NG/RTIL/GCE.

containing groups on the surface of electrode may enhancecombination, such as hydrogen bond.

The ionic liquid was immobilized successfully on the surface ofthe electrode. The gold nanoparticles were of a more uniformappearance, which may be due to the presence of ionic liquidsoptimize the performance of the electrode surface. The goldnanoparticles generally possess excellent catalytic activity and offera hospitable environment for biomolecules. Fig. 3 (inset) shows theEIS of every modified procedure, which can be seen the chargetransfer resistance (Rct) was gradually reduced. IL-robed DNA kepttheir double-stranded helix structure [13]. The interaction madeDNA immobilized on the surface of the electrode stronger [14]. Thisinterface has good electrical conductivity, chemical stability andbiocompatibility. The interface has the characteristics of ionicliquids and nano-gold for the electrochemical DNA research.

Fig. 3. EIS of (a) ct-DNA/NG/RTIL/GCE, (b) ct-DNA/NG/RTIL/GCE incubated by HCHO,

(c) bare GCE, (d) GCE activated by H2SO4, (e) RTIL/GCE, (f) NG/RTIL/GCE in 5.0 mM

Fe(CN)63�/4� and 0.1 M KCl.

Fig. 4. Cyclic voltammograms of 0.02 M HCHO in 0.1 M NaHCO3–Na2CO3 solution at

GCE (a), ct-DNA/NG/RTIL/GCE (b), ct-DNA/NG/RTIL/GCE incubated in HCHO (C).

(v ¼ 100 mV=s, scan from �0.5 to 1.0 V) (inset: DPV of ct-DNA/NG/RTIL/GCE

incubated in HCHO.)

L. Lu et al. / Applied Surface Science 256 (2009) 52–5554

3.2. Eletrochemical analysis DNA damage of HCHO

The principle of electrochemical impedance sensing of DNAdamaging is based on the change of the charge transfer resistance,Rct. The negatively charged redox indicator Fe(CN)6

3�/4� waselectrostatically repelled by the resulting negatively chargedinterface. The electrochemical impedance spectroscopy (EIS)was shown in Fig. 3. It is clear that the electron transfer resistanceof Fe(CN)6

3�/4� redox couple is different. We can see from curve a,the ct-DNA is modified on the NG/RTIL/GCE, Rct = 298 V; after thect-DNA are damaged by incubated in 0.02 M HCHO at 37 8C for30 min, the Rct = 502 V. This effect was attributed to the reduceelectronic conduction via stacked bases.

Cyclic voltammograms of HCHO at bare GCE and the modifiedelectrode are comparatively shown in Fig. 4. As shown in Fig. 4,HCHO has no CV peak response at bare GCE (curve a), while the ct-DNA/NG/RTIL/GCE (curve b) leads to three current peak at about0.38, 0.61, and 0.29 V. The peak at 0.61 and 0.29 V are corresponds tothe typical oxidation of the HCHO in basic media [17]. In contrast,when ct-DNA/NG/RTIL/GCE is incubated in 0.02 M HCHO at 37 8C for30 min, the three peaks current increased greatly (Fig. 4, curve c).Additionally, the peak at 0.29 V shifts positively up to 0.35 V. The

Fig. 5. AFM of ct-DNA/NG/RTIL/GCE (left) and ct-D

peak 1 (or curve b: at 0.38 V) may ascribe to the oxidation of HCHO-guanine base, this point is supported by the result of the cyclicvoltammatry of guanine [18] originated from the degrading of DNA.The differential pulse voltammograms of the incubated ct-DNA/NG/RTIL/GCE in the 0.02 M HCHO is shown in Fig. 4 (inset), there is adistinct oxidation peak at 1.25 V, corresponding to the oxidation ofadenine [19], however, this peak was not existed when it was notHCHO. These dictate the crosslink which HCHO caused make DNAdegrade, and the stretching of the molecule may disrupt the basestacking. Meanwhile, it proved DNA on the NG/RTIL/GCE was keptthe native conformation. It is because natural DNA gives minimalvoltammetric signals, but damage exposes DNA bases to theelectrode can be observed the redox peak [20].

3.3. AFM analysis of DNA damage

Electrode surface characteristics represent an important aspecton the construction of sensitive DNA electrochemical biosensors fordetection of DNA interaction and HCHO. AFM images were obtainedon the glass carbon slices (noted GCS). The samples were prepared asthe ct-DNA/NG/RTIL/GCE. Fig. 5B is the AFM image of the ct-DNA/NG/RTIL/GCS, which were incubated in 0.02 M HCHO at 37 8C for

NA/NG/RTIL/GCE incubated by HCHO (right).

L. Lu et al. / Applied Surface Science 256 (2009) 52–55 55

30 min; Fig. 5A is the image of the ct-DNA/NG/RTIL/GCS withoutincubated. Along our consideration, the ct-DNA molecules in theirstates of compactness were different. They loosely attached to thegold nanoparticles when they were incubated in HCHO, while the ct-DNA molecules without processed were closer. The DNA moleculeswere not priority fabricated on the surface of glass carbon but thegold nanoparticles [21]. HCHO may destroy the close packing modelof the DNA molecules by interacted with the DNA.

It was known that formaldehyde can induce nucleic acidscrosslink [22]. Functional groups that are reactive are amidogroups by reactive bridges between reactive groups. The results ofthe ultraviolet experiments showed the conjugate degree increaseof DNA incubated in HCHO, which indicated may form C55N bridgesas well. Formaldehyde penetrating destroyed the 3D naturestructures of DNA, which could lead to long DNA strands relativelyloose contact. However, neither the morphology nor the con-ductivity of DNA was changed.

4. Conclusion

An active nano-supported interface was designed from goldnanoparticles embedded on ionic liquid for depositing DNA. DNAmaintained the natural structure based the extraordinary bio-compatibility of nano-gold and room temperature ionic liquids.The study of DNA damage resulted from formaldehyde proceededby AFM and electrochemistry. The results show that formaldehydecaused DNA degraded and formaldehyde penetrating combinedwith DNA forming C55N. It is the next work that the furtherdetection of the damage to DNA hybridization by this active nano-supported interface.

Acknowledgements

The authors gratefully acknowledge Beijing Municipal NaturalScience Foundation (no. 8062010), Key Project of Chinese NationalPrograms for Fundamental Research and Development (973program) (no. 2005CB724201) and BJUT Science Foundation forYouths (no. 97005013200701)

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