Elsevier

Electrochimica Acta

Volume 114, 30 December 2013, Pages 285-295
Electrochimica Acta

Simultaneous detection of guanine, adenine, thymine, and cytosine at polyaniline/MnO2 modified electrode

https://doi.org/10.1016/j.electacta.2013.10.064Get rights and content

Highlights

  • PANI/MnO2 modified electrode was fabricated.

  • PANI/MnO2 was investigated in the simultaneous detection of all four DNA bases.

  • PANI/MnO2 exhibited low detection limit, good sensitivity, and wide linear range.

  • Concentrations of all four DNA bases in a calf thymus DNA sample were determined.

Abstract

In this study, PANI/MnO2 nanocomposite was synthesized by the reaction of polyaniline (PANI) and KMnO4 in the aqueous medium. Material was characterized by X-ray diffraction, nitrogen sorption, scanning electron microscopy, transmission electron microscopy, thermogravimetric analysis, and FT-IR. PANI/MnO2 modified electrode was investigated in the simultaneous detection of all four DNA bases (guanine, adenine, thymine, and cytosine) using cyclic and differential pulse voltammetries. pH was optimized to obtain the best peak potential separation and response current. This enabled us for the simultaneous voltammetric determination of all four DNA bases at physiological condition (pH 7). PANI/MnO2 nanocomposite exhibited high-current sensitivity for these analytes compared to MnO2 and PANI modified electrodes. Under the optimum conditions, PANI/MnO2 exhibited low detection limit, good sensitivity, and wide linear range for the simultaneous detection of G, A, T, and C. Moreover, the proposed method was successfully applied in the determination of G, A, T, and C contents in a calf thymus DNA sample with satisfactory results. The reliability and stability of the modified electrode provide a good possibility for applying the technique in the routine analysis for a selected class of electroactive organic/bio-molecules.

Introduction

Deoxyribonucleic acid (DNA) is an important biological macromolecule that plays crucial roles in the storage of genetic information and protein biosynthesis [1], [2]. This genetic material is present in the cells of all living organisms. DNA is a block copolymer with four types of deoxyribonucleotide monomers differentiated by their bases [1], [2]. Purine bases (guanine (G), and adenine (A)) and pyrimidine bases (thymine (T) and cytosine (C)) are found in DNA molecular structure and they are involved in cellular energy transduction and signaling mediated by enzymatic oxidation reactions [3]. Mutation occurs due to change in the sequence of DNA. One basic mutation in mitochondrial DNA can cause heart and muscle disease [4]. Therefore, abnormal changes of DNA bases in organism lead to the deficiency of the immunity system and may indicate the presence of various diseases, including cancer, Alzheimer's, epilepsy, and HIV infection [4], [5]. The detection of DNA bases is of great significance for clinical diagnosis as well as insight into fundamental mechanism of genetic information [6], [7]. Detecting DNA mutations is important and then to find appropriate ways to prevent or treat disease. Several analytical methods have been developed to detect DNA bases [8], [9] such as, microchip capillary electrophoresis [10], [11], flow injection chemiluminescence [12], [13], ion pairing liquid chromatography [14], laser-induced fluorescence detection [15], and micellar electrokinetic chromatography [16]. Although these methods exhibit some merits, expensive instruments, complicated operations, or time-consuming sample pretreatments are usually involved [17]. Electrochemical techniques are promising for the analysis of DNA bases due to their advantages of rapidity, convenience, low-cost and ease of miniaturization for small-volume sample [18]. Among these techniques, DNA biosensor has emerged as a promising alternative for microbial detection due to the specificity of hybridization between the probe and the complementary target sequence [18], [19]. However, the non-specific adsorption in the hybridization system increases the strong demand for the development of a sensitive and specific DNA biosensor.

The electrocatalytic oxidation of purine bases has been investigated [20]. However, the electrochemical detection of pyrimidine bases is rarely reported. It is difficult to obtain accurate oxidation signals of pyrimidine bases because of their high positive oxidation potentials and slow electron transfer kinetics [21]. In order to overcome these limitations, electrode materials with a wide potential window and high electrocatalytic activities are required.

Inorganic materials, such as metals, semiconductors, and metal oxides have been the subject of intensive research because of their potential applications in electronics, photonics, and catalysis [22]. Manganese dioxide is one of the most attractive inorganic materials because of its wide range of applications in catalysts, molecular-sieves, and electromagnetic materials [23], [24], [25]. MnO2 has become the most widely used electrode material available since Leclanché first discovered its use as an additive in batteries [26]. Birnessite structure (δ-MnO2) has attracted recent interest from the point of view of its structural, morphological modification, delamination [27], and inherent lithium insertion/deinsertion properties with respect to its application as a lithium battery electrode [28], [29]. Conducting polymer containing metal oxides based nanocomposites have attracted a great deal of interest because of the unexpected properties, synergistically derived from both components [30], [31], [32]. In this area, polyaniline (PANI) is considered as the most promising conducting polymers that has many advantages such as high doping/de-doping rate during charge/discharge process, easy polymerization and high environmental stability [33], [34]. Polyaniline (PANI) is an intrinsically conductive polymer (ICP), which can conduct electric currents without the addition of any conductive (inorganic) substances [35]. However, the electroactivity (i.e., redox behavior) of polyaniline is strongly dependent on the pH value of electrolyte solution, and is greatly weakened at pH > 4. Low electrocatalytic activity at physiological pH (neutral pH environment) condition greatly limits the applications of PANI in bioelectrochemistry [36]. It is reported that doping PANI with metals/metal oxides yields a redox active polymer, which exhibits good electrocatalytic activity in neutral or basic aqueous solution [37]. Furthermore, MnO2 based nanocomposites are known to be used in neutral aqueous electrolytes that can meet the requirements of “green electrolyte” [27], [28], [29]. The oxidation activity of δ-MnO2 is mainly associated to the presence of Mn4+. Nanocomposites based on conducting polymer and MnO2 have been intensively investigated during recent years due to their potential applications as electrode material [38], [39], [40], [41].

Our research is focused on the developments of nanoporous materials such as polyanilines, metal oxides, mesoporous zeolites and find their catalytic and electrocatalytic applications [42], [43], [44], [45], [46], [47], [48], [49], [50], [51], [52]. In this study, we report the synthesis and application of PANI/MnO2 nanofibers. MnO2 was coated onto PANI nanofibers by a direct coating method via the reduction of KMnO4. PANI/MnO2 modified electrode is used in the simultaneous detection of guanine, adenine, thymine, and cytosine. In the present study based on PANI/MnO2 nanocomposite, MnO2 along with polyaniline synergistically contributes in increasing the electrochemical performance even at physiological pH condition.

Section snippets

Materials

All chemicals used in this study were of A.R. grade. Aniline, ammonium peroxydisulphate (APS), and potassium permanganate (KMnO4) were obtained from Spectrochem India Pvt. Ltd. Guanine, adenine, thymine, cytosine, valine, and Nafion (5%, v/v) were purchased from Aldrich. Aniline was vacuum distilled and stored in the dark prior to its use. Deionized water from Millipore Milli-Q system (Resistivity 18.2  cm) was used in the electrochemical studies. Deoxyribonucleic acid sodium salt from the calf

Characterization

XRD investigation confirms that MnO2 obtained under our synthesis condition belong to monoclinic potassium birnessite (δ-MnO2) (JCPDS no. 80-1098) (Fig. 1a). Two broad diffractions at of 20.5° and 25.3° were obtained in the XRD pattern of PANI, which represent the periodicities parallel (1 0 0) and perpendicular (1 1 0) to the PANI chain, respectively (Fig. 1b) [54], [55]. XRD pattern of the PANI/MnO2 shows diffraction corresponding to the monoclinic K-birnessite MnO2 (JCPDS no. 80-1098) (Fig. 1

Conclusion

Combining the advantageous features of MnO2 and PANI, organic–inorganic hybrid PANI/MnO2 was prepared. PANI/MnO2 modified electrode shows promising activity in the electrochemical oxidation of guanine, adenine, thymine, and cytosine with high sensitivity, selectivity, and antifouling properties. Influence of various electrochemical reaction parameters such as scan rate and pH were evaluated to find the optimum condition for the analysis. PANI/MnO2 modified electrode (prepared with 0.06 M KMnO4

Acknowledgments

We are pleased to acknowledge Council of Scientific and Industrial Research (CSIR), New Delhi, for financial support under CSIR (01(2423)/10/EMR-II). AP is grateful to CSIR, New Delhi for SRF fellowship. We acknowledge Director, IIT Ropar for constant encouragements.

References (68)

  • H. Yu et al.

    Development of a high performance liquid chromatography method and a liquid chromatography–tandem mass spectrometry method with the pressurized liquid extraction for the quantification and confirmation of sulfonamides in the foods of animal origin

    J. Chromatogr. B

    (2011)
  • H.C. Chen et al.

    Determination of the heterogeneity of DNA methylation by combined bisulfite restriction analysis and capillary electrophoresis with laser-induced fluorescence

    J. Chromatogr. A

    (2012)
  • X.W. Zhang et al.

    Sweeping under controlled electroosmotic flow and micellar electrokinetic chromatography for on-line concentration and determination of trace phlorizin and quercitrin in urine samples

    J. Pharm. Biomed. Anal.

    (2011)
  • W. Sun et al.

    Direct electrochemistry of guanosine on multi-walled carbon nanotubes modified carbon ionic liquid electrode

    Electrochim. Acta

    (2009)
  • H. Shiraishi et al.

    Electrochemical detection of E. coli 16S rDNA sequence using air-plasma-activated fullerene-impregnated screen printed electrodes

    Bioelectrochemistry

    (2007)
  • Y.K. Ye et al.

    Rapid detection of ssDNA and RNA using multi-walled carbon nanotubes modified screen-printed carbon electrode

    Biosens. Bioelectron.

    (2005)
  • C.A. Frysz et al.

    Carbon filaments and carbon black as a conductive additive to the manganese dioxide cathode of a lithium electrolytic cell

    J. Power Sources

    (1996)
  • P. Le Goff et al.

    Synthesis, ion exchange and electrochemical properties of lamellar phyllomanganates of the birnessite group

    Mater. Res. Bull.

    (1996)
  • H.Y. Lee et al.

    Supercapacitor behavior with KCl electrolyte

    J. Solid State Chem.

    (1999)
  • Y. Xian et al.

    Glucose biosensor based on Au nanoparticles–conductive polyaniline nanocomposite

    Biosens. Bioelectron.

    (2006)
  • M.U. Anu Prathap et al.

    Synthesis of nanoporous metal oxides through the self-assembly of phloroglucinol–formaldehyde resol and tri-block copolymer

    J. Colloid Interface Sci.

    (2011)
  • M.U. Anu Prathap et al.

    Direct synthesis of metal oxide incorporated mesoporous SBA-15, and their applications in non-enzymatic sensing of glucose

    J. Colloid Interface Sci.

    (2012)
  • M.U. Anu Prathap et al.

    Hydrothermal synthesis of CuO micro-/nanostructures and their applications in the oxidative degradation of methylene blue and non-enzymatic sensing of glucose/H2O2

    J. Colloid Interface Sci.

    (2012)
  • M.U. Anu Prathap et al.

    Synthesis of NiCo2O4 and its application in the electrocatalytic oxidation of methanol

    Nano Energy

    (2013)
  • M.U. Anu Prathap et al.

    Tailoring properties of polyaniline for simultaneous determination of a quaternary mixture of ascorbic acid, dopamine, uric acid, and tryptophan

    Sens. Actuators, B

    (2013)
  • M.U. Anu Prathap et al.

    Synthesis of mesostructured polyaniline using mixed surfactants, anionic sodium dodecylsulfate and non-ionic polymers and their applications in H2O2 and glucose sensing

    Colloids Surf., B

    (2012)
  • M.U. Anu Prathap et al.

    Facile preparation of polyaniline/MnO2 nanofibers and its electrochemical application in the simultaneous determination of catechol, hydroquinone, and resorcinol

    Sens. Actuators, B

    (2013)
  • R. Srivastava et al.

    Morphologically controlled synthesis of copper oxides and their catalytic applications in the synthesis of propargylamine and oxidative degradation of methylene blue

    Colloids Surf., A

    (2011)
  • Q.M. Pham et al.

    Preparation of polyaniline–titanium dioxide hybrid materials in supercritical CO2

    Synth. Met.

    (2009)
  • A.H. Kamel et al.

    Electrochemical determination of antioxidant capacities in flavored waters by guanine and adenine biosensors

    Biosens. Bioelectron.

    (2008)
  • V. Brabec

    433—Nucleic acid analysis by voltammetry at carbon electrodes

    Bioelectrochem. Bioenerg.

    (1981)
  • W. Sun et al.

    Direct electrocatalytic oxidation of adenine and guanine on carbon ionic liquid electrode and the simultaneous determination

    Biosens. Bioelectron.

    (2008)
  • Z.H. Wang et al.

    β-Cyclodextrin incorporated carbon nanotubes-modified electrodes for simultaneous determination of adenine and guanine

    J. Electroanal. Chem.

    (2006)
  • V. Dharuman et al.

    DNA hybridization detection on electrical microarrays using coulostatic pulse technique

    Biosens. Bioelectron.

    (2006)
  • Cited by (122)

    • Investigation of thymine as a potential cancer biomarker employing tryptophan with nanomaterials as a biosensor

      2023, Spectrochimica Acta - Part A: Molecular and Biomolecular Spectroscopy
    View all citing articles on Scopus
    View full text