Please cite this article in press as: U.B. Trivedi, et al., Amperometric fructose biosensor based on fructose dehydrogenase enzyme, Sens. Actuators B: Chem. (2008), doi:10.1016/j.snb.2008.10.020 ARTICLE IN PRESS G Model SNB-11058; No. of Pages 7 Sensors and Actuators B xxx (2008) xxx–xxx Contents lists available at ScienceDirect Sensors and Actuators B: Chemical journal homepage: www.elsevier.com/locate/snb Amperometric fructose biosensor based on fructose dehydrogenase enzyme U.B. Trivedi a , D. Lakshminarayana a , I.L. Kothari b , P.B. Patel a , C.J. Panchal c,∗ a Department of Electronics, Sardar Patel University, Vallabh Vidyanagar-388120, Gujarat, India b Department of Bio Sciences, Sardar Patel University, Vallabh Vidyanagar-388120, Gujarat, India c Applied Physics Department, Faculty of Technology and Engineering, M. S. University of Baroda, Vadodara-390001, Gujarat, India article info Article history: Received 24 April 2008 Received in revised form 10 October 2008 Accepted 16 October 2008 Available online xxx Keywords: Amperometry Fructose Biosensor Fructose dehydrogenase (FDH) Polymer matrix abstract A low cost, portable and disposable biosensor based upon the enzyme d-fructose dehydrogenase (FDH) has been developed. A polymer matrix of polyethylenemine (PEI) and poly(carbamoylsulphonate) (PCS) hydrogel has been used for the immobilization of FDH on a platinum (Pt) tip of a screen printed graphite electrode with a ferricyanide mediator as the electron acceptor. A four channel potentiostat has been uti- lized in the electrode configuration for the amperometric measurement of fructose. The, thus, developed Pt/PCS + PEI/FDH-[Fe(CN) 6 ] 3- mediated biosensor could offer a reliable method of fructose determination in standard stock solutions. The designed fructose biosensor system has been found to show a sensitivity of 0.62 ± 0.10 nA/M. The influence of various parameters like buffer solution pH, temperature, ferricyanide concentration, etc. on the biosensor performance has been studied in the present investigation. Also, the specificity of the enzyme electrode, the reproducibility and the storage stability of the FDH based biosen- sor system have been studied and the results are presented. The sensor system could also be utilized for the determination of fructose in the real samples of fruit juice, soft drinks and honey. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Quality control is of paramount importance in food and beverage industries. In food and fermentation processes, quick and reliable analytical methods are required to analyze sugars like glucose, fructose, sucrose, etc. for better process efficiency and economy. Conventional methods for sugar analysis like spectrophotometric and titrimetric techniques [1–2] are non-specific and laborious. Other methods like HPLC and Gas Chromatography (GC) are specific but expensive and often require elaborate sample pretreatment [3]. Considering the disadvantages, there is a need to develop alternate techniques which are rapid, accurate, and reproducible and require no sample pretreatment. Also, the recent demands for high quality food products to meet the customer needs have opened up newer and improved sensor technologies that are cou- pled with production processes for quality control and consumer assurance. One of the modern techniques which overcome the disadvantages of conventional methods of sugars analysis is the biosensor. For real time testing and for online measurements in food industries, the biosensor may offer a fast, low cost, disposable and portable type of sensor technology leading to efficient production too. ∗ Corresponding author. Tel.: +91 9825094761; fax: +91 2652423898. E-mail address: cjpanchal msu@yahoo.com (C.J. Panchal). d-Fructose, one of the principal sugar components, is a widely distributed monosaccharide and an important sweetener. It is also frequently used as a sugar substitute for diabetics [4]. Several methods for fructose determination have been developed in recent years [5]. For the determination of d-fructose in food and clinical specimens, analytical technique such as titration [6], polarime- try [7], gas–liquid chromatography [8], spectrophotometric [9], enzymatic analysis [10], nuclear magnetic resonance spectropho- tometry [11], fluorimetry [12], and electrochemistry [13] have been described in the literature. However, several studies are still being carried out to obtain faster and more selective meth- ods of analysis [14]. As a sensor for d-fructose determination, a bi-enzyme electrode using hexokinase and glucose 6-phosphate dehydrogenase has been described [15]. Unfortunately, this sens- ing method was found to be too expensive, because it wasted valuable enzymes and related co-factors and it turned out to be only slightly sensitive [16]. The first description of membrane bound d-fructose dehydrogenase (FDH) was made by Yamada et al. [17], who confirmed that the enzyme catalyses the oxidation of d-fructose to 5-keto-d-fructose in the presence of a mediator. FDH is an enzyme containing pyrroloquinolinequinone (PQQ) and belongs to a group of quinoproteins that have been described as an interesting alternative for the construction of enzyme elec- trodes [18]. Later on, with the use of immobilized FDH reactors or membranes [19], different amperometric techniques for the determination of d-fructose were reported. In addition, several biosensors, based on platinum electrodes [20], glassy carbon elec- 0925-4005/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.snb.2008.10.020