Effect of yeast pretreatment on the characteristics of yeast-modified electrodes as mediated amperometric biosensors for lactic acid

Institute of Chemistry, Gostauto 9, LT-01108, Vilnius, Lithuania.
Bioelectrochemistry (Amsterdam, Netherlands) (Impact Factor: 3.87). 09/2008; 74(1):188-94. DOI: 10.1016/j.bioelechem.2008.07.009
Source: PubMed

ABSTRACT Carbon paste electrode modified with baker' and wine yeast Saccharomyces cerevisiae (a source of flavocytochrome b(2)) were investigated as amperometric biosensors for L-lactic acid. Before immobilization on the electrode surface, yeast cells were pretreated with various electrolytes, alcohols and weak organic acids. Electrode responses to L-lactic acid were tested in the presence of various mediators (potassium ferricyanide, phenazine methosulfate, 2,6-dichlorophenolindophenol sodium salt hydrate, 1,2-naphthoquinone-4-sulfonic acid sodium salt). The highest (144+/-7 nA per 0.2 mM L-lactic acid) and the most stable responses were obtained after yeast pretreatment with 30% ethanol using potassium ferricyanide as a mediator. Different electrode sensitivities with mediator phenazine methosulphate probably reflected diverse changes in yeast membrane (and/or cell wall).

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Although up to now IUPAC has not accepted an official definition of the term biosensor, its electrochemical representative is defined as “a self-contained integrated device, which is capable of providing specifc quantitative or semi-quantitative analytical information using a biological recognition element (biochemical receptor) which is retained in direct spatial contact with an electrochemical transduction element’’ (Thevenot et al., 2001). Generally, the biosensor is a hybrid device containing two functional parts: a bioelement (an immobilized biologically active material) and a physical transducer. As bioelements can be used pieces of tissue, microbial cells, organelles, natural biomembranes or liposomes, receptors, enzymes, antibodies and antigens, abzymes, nucleic acids and other biomolecules and even biomimetics which imitate structural and functional features of the natural analogue. The bioelement is a recognition unit providing selective binding or biochemical/metabolic conversion of the analyte that results in changes of physical or physico-chemical characteristics of the transducer (Scheller et al., 1991; Schmidt & Karube, 1998; Gonchar et al., 2002; Nakamura & Karube, 2003; Sharma et al., 2003; Investigations on Sensor Systems and Technologies, 2006). The bioelement in such constructions is usually prepared in immobilized form and often covered with an outer membrane (or placed between two membranes in a sandwich manner), which either prevents the penetration of interfering substances into a sensitive bioselective layer and transducer surface, or creates a diffusion barrier for the analyte. Such membrane structures increase the stability of the biorecognizing element, enhance its selectivity and provide the diffusion limitations for biochemical reactions. Electrochemical, optical, piezoelectric, thermoelectric, transistor, acoustic and other elements are used as transducers in biosensor systems. Electrochemical (amperometric, potentiometric, conductometric) and optical (surface plasmon resonance) devices are the most exploited transducers in commercially available biosensors (Commercial Biosensors, 1998). Basically, biosensors can be regarded as information transducers in which the energy of biospecific interactions is transformed into information about the nature and concentration of an analyte in the sample. The most essential advantages of biosensors are excellent chemical selectivity and high sensitivity, possibility of miniaturization and compatibility with computers. Their drawbacks are limited stability and a rather complicated procedure for preparation of the biologically active material. The enzyme biosensors are the most widespread devices (Zhao & Jiang, 2010); many of them are produced commercially. Enzyme biosensors are characterized by their high selectivity. They also provide fast output due to high activity and high local enzyme concentration in a sensitive layer. The drawbacks of enzyme biosensors are insufficient stability and the high price of purified enzymes. The cell sensors, especially microbial ones, have been actively developed only in recent years (Shimomura-Shimizu & Karube, 2010a, 2010b; Su et al., 2011). Cell biosensors have a range of considerable advantages when compared to their enzyme analogues: availability of cells, low price and simple procedure of cell isolation, possibility to use long metabolic chains, avoiding purification of enzymes and coenzymes, advanced opportunity for metabolic engineering, integrity of the cell response (important in assaying total toxicity and mutagenic action of environmental pollutants), possibility to retain viability of sensoring cells and even to provide their propagation, and, in some cases, higher stability of cell elements compared to enzyme ones. The main drawbacks of microbial biosensors are a rather low signal rate due to a lower concentration of enzymes involved in cellular response, as well as low selectivity of cell output (e.g. in the case of microbial O2 electrode sensors due to a broad substrate specificity of cellular respiration). 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New genomic and proteomic approaches are being used for revealing cancer biomarkers related with genetic features, changes in gene expression, protein profiles and post-translational modifications of proteins. Recent progress in nanobiotechnology allows using nanomolecular approaches for clinical diagnostic procedures (Salata, 2004; Jain, 2007). The most important applications are foreseen in the areas of biomarker monitoring, cancer diagnosis, and detection of infectious microorganisms. Analytical nanobiotechnology uses different nanoscaled materials (gold and magnetic nanoparticles, nanoprobes, quantum dots as labels, DNA nanotags) for molecular detection (Baptista et al., 2008; Medintz et al., 2008; Sekhon & Kamboj, 2010). The use of nanomaterials in biosensors has allowed the introduction of many new signal transduction technologies into biosensorics and improvement of bioanalytical parameters of the nanosensors - selectivity, response time, miniaturization of the biorecognition unit (Jianrong et al., 2004; Murphy, 2006).
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