A novel biosensor for the detection of zearalenone family mycotoxins in milk.
ABSTRACT In this study, a method for detecting estrogenic mycotoxin residues in milk was developed utilizing bioluminescent whole-cell biosensors. Milk products of various compositions were spiked with the estrogenic mycotoxins zearalenone and its metabolites zearalanone, alpha-zearalanol, beta-zearalanol, alpha-zearalenol and beta-zearalenol. The estrogenic response was detected by a whole-cell biosensor based on a genetically modified Saccharomyces cerevisiae strain that in the presence of an estrogenic compound produces firefly luciferase-enzyme and further light emission within a system provided with D-luciferin substrate. The results show that the yeast sensor reacts to mycotoxins with typical sigmoidal response at nanomolar concentrations. The response differs in different milk products with regard to the fat content of the milk. Due to short assay time of less than 3h and automation the approach can be used as a bioavailability and activity screening method prior to more detailed chemical analysis.
- SourceAvailable from: Matti Karp[Show abstract] [Hide abstract]
ABSTRACT: A bacterial biosensor method for the selective determination of a bioavailable organomercurial compound, methylmercury, is presented. A recombinant luminescent whole-cell bacterial strain responding to total mercury content in samples was used. The bacterial cells were freeze-dried and used as robust, reagent-like compounds, without batch-to-batch variations. In this bacteria-based sensing method, luciferase is used as a reporter, which requires no substrate additions, therefore allowing homogenous, real-time monitoring of the reporter gene expression. A noninducible, constitutively light-producing control bacterial strain was included in parallel for determining the overall cytotoxicity of the samples. The specificity of the total mercury sensor Escherichia coli MC1061 (pmerRBlux) bacterial resistance system toward methylmercury is due to a coexpressed specific enzyme, organomercurial lyase. This enzyme mediates the cleavage of the carbon-mercury bond of methylmercury to yield mercury ions, which induce the reporter genes and produce a self-luminescent cell. The selective analysis of methylmercury with the total mercury strain is achieved by specifically chelating the inorganic mercury species from the sample using an optimized concentration of EDTA as a chelating agent. After the treatment with the chelating agent, a cross-reactivity of 0.2% with ionic mercury was observed at nonphysiological ionic mercury concentrations (100 nM). The assay was optimized to be performed in 3 h but results can already be read after 1 h incubation. Total mercury strain E. coli MC1061 (pmerRBlux) has been shown to be highly sensitive and capable of determining methylmercury at a subnanomolar level in optimized assay conditions with a very high dynamic range of two decades. The limit of detection of 75 ng/l (300 pM) allows measurement of methylmercury even from natural samples.Analytical and Bioanalytical Chemistry 04/2011; 400(4):1041-9. · 3.66 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Increases in food production and the ever-present threat of food contamination from microbiological and chemical sources have led the food industry and regulators to pursue rapid, inexpensive methods of analysis to safeguard the health and safety of the consumer. Although sophisticated techniques such as chromatography and spectrometry provide more accurate and conclusive results, screening tests allow a much higher throughput of samples at a lower cost and with less operator training, so larger numbers of samples can be analysed. Biosensors combine a biological recognition element (enzyme, antibody, receptor) with a transducer to produce a measurable signal proportional to the extent of interaction between the recognition element and the analyte. The different uses of the biosensing instrumentation available today are extremely varied, with food analysis as an emerging and growing application. The advantages offered by biosensors over other screening methods such as radioimmunoassay, enzyme-linked immunosorbent assay, fluorescence immunoassay and luminescence immunoassay, with respect to food analysis, include automation, improved reproducibility, speed of analysis and real-time analysis. This article will provide a brief footing in history before reviewing the latest developments in biosensor applications for analysis of food contaminants (January 2007 to December 2010), focusing on the detection of pathogens, toxins, pesticides and veterinary drug residues by biosensors, with emphasis on articles showing data in food matrices. The main areas of development common to these groups of contaminants include multiplexing, the ability to simultaneously analyse a sample for more than one contaminant and portability. Biosensors currently have an important role in food safety; further advances in the technology, reagents and sample handling will surely reinforce this position.Analytical and Bioanalytical Chemistry 01/2012; 403(1):75-92. · 3.66 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Mycotoxins are secondary metabolites produced by several filamentous fungi that grow on various food and feed. These compounds elicit a wide spectrum of toxicological effects, representing a health risk for both, humans and animals. Mycotoxin analysis is an important tool in controlling fungal contamination of food and feed. This article reviews the techniques used for their determination, and the advantages and limitations of each method are critically discussed.ROM. J. BIOCHEM. 01/2010; 47(1):79.