Disposable biosensor based on platinum nanoparticles-reduced graphene oxide-laccase biocomposite for the determination of total polyphenolic content Sandra A.V. Eremia a , Ioana Vasilescu a , Antonio Radoi b , Simona-Carmen Litescu a,n , Gabriel-Lucian Radu a a Centre of Bioanalysis, National Institute of Research and Development for Biological Sciences, 296 Splaiul Independentei, 060031 Bucharest, Romania b National Institute for Research and Development in Microtechnology (IMT-Bucharest), 126A Erou Iancu Nicolae, 077190 Bucharest, Romania article info Article history: Received 2 November 2012 Received in revised form 7 February 2013 Accepted 13 February 2013 Available online 21 February 2013 Keywords: Carbon screen-printed electrode Laccase Platinum nanoparticles Reduced graphene oxide Tea infusion Total polyphenolic content abstract A disposable amperometric biosensor was developed for the detection of total polyphenolic compounds from tea infusions. The biosensor was designed by modifying the surface of a carbon screen-printed electrode with platinum nanoparticles and reduced graphene oxide, followed by the laccase drop- casting and stabilization in neutralised 1% Nafion solution. The obtained biosensor was investigated by scanning electron microscopy and electrochemical techniques. It was observed that platinum nanoparticles-reduced graphene oxide composite had synergistic effects on the electron transfer and increased the electroactive surface area of the carbon screen-printed electrode. The constructed analytical tool showed a good linearity in the range 0.2–2 mM for caffeic acid and a limit of detection of 0.09 mM. The value of Michaelis–Menten apparent constant was calculated from the electrochemical version of Lineweaver–Burk equation to be 2.75 mM. This disposable laccase biosensor could be a valuable tool for the estimation of total polyphenolic content from tea infusions. & 2013 Elsevier B.V. All rights reserved. 1. Introduction Polyphenols are natural compounds widely found in vegeta- bles [1,2], cereals, tea leaves [3,4], fruit juices [5] and red wines [6]. Tea contains a variety of polyphenolic compounds (phenolic acids, flavonoids, tannins, catechins, etc.) having a wide range of biological activities that contribute in cancer prevention [7], anti-aging, exhibits protective effect against cardiovascular diseases [8] and brain protection against oxidative stress [9]. Fruit tea infusions have a high content of caffeic acid and therefore can show strong antioxidant properties [10]. There are many analytical methods currently involved in the determination of polyphenols from different sources: UV–vis spectrophotometry [11], chromatography [12–14], capillary elec- trophoresis [15], electrochemistry [16]. Two standard methods frequently used for the determination of total polyphenolic content from tea are Folin–Ciocalteu colorimetric method [17,18] and high performance liquid chromatography with appro- priate detection (HPLC) [19]. Spectrophotometric methods some- times lack in sensitivity and specificity (e.g. Folin–Ciocalteu), while chromatographic methods are laborious, require skilled personnel and high amounts of solvents, reagents and resources. Considering tea extracts, the major drawback of Folin–Ciocalteu method is the detection of other interfering compounds (i.e. ascorbic acid, several amino acids, carbohydrates, etc.) besides polyphenolic compounds. To overcome the drawbacks of the previously mentioned methods, biosensors represent a valuable alternative for the determination of polyphenolic content. There are several amperometric biosensors reported in the literature for the analysis of polyphenols from tea. The first enzyme-based biosensor used for the determination of polyphenols was devel- oped by Ghindilis et al. [20], the biological recognition element being laccase. Laccase [21–23] is rather used than tyrosinase [24,25] in developing biosensors for polyphenolic content deter- mination, due to its wider catalytic effect against polyphenols (it catalyses both ortho and para di- and polyphenols). To provide resolution and a better quantification of each compound from a phenolic or polyphenolic mixture, two groups of researchers developed independently two analytical tools, one based on a dual amperometric approach (laccase- and tyrosinase-based biosensors) for rapid evaluation of mixtures of phenolic com- pounds [26] and the second system was based on an array of enzymatic biosensors (tyrosinase and laccase) for the determina- tion of major polyphenols found in wine [27]. Contents lists available at SciVerse ScienceDirect journal homepage: www.elsevier.com/locate/talanta Talanta 0039-9140/$ - see front matter & 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.talanta.2013.02.029 n Corresponding author. Tel./fax: þ40 21 2200 900. E-mail address: slitescu@gmail.com (S.-C. Litescu). Talanta 110 (2013) 164–170