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The fast identification and quantification of analytes in the field of food safety or environmental analysis is difficult. Surface enhanced Raman scattering (SERS) is an analytical method which can be used simultaneously for the rapid identification and concentration determination of trace analytes,[1,2] usually covering a large dynamic range from nanomolar up to molar concentrations. The identification of the molecules is accomplished through the specific fingerprint of a molecule’s Raman spectrum.
For facile and straightforward SERS measurements, we present here a combination of paper-based SERS test strips with microfluidic systems on paper as a microfluidic paper-based analytical device (μPAD). The SERS μPAD is thus principally suited for cheap, fast, non-destructive, label-free and portable detection of analytes. In this system basically, the use of the microfluidic structured paper increases the sensitivity and suppresses background signals of the SERS assay.
Deposition of the SERS substrate on the test strips is simple and relies on an inkjet printer. For the optimization of the reproducibility and intensity of the SERS signal, we tested different nanoparticles, different numbers of print cycles and different paper types. The nanoparticle solutions used in the μPAD preparation were gold and silver nanoparticle solutions. The paper types were cellulose and glass fiber. SERS arrays were prepared by printing and compared to arrays prepared by spraying. The optimized μPAD was used for the identification and quantification of pure analyte solutions (e.g., adenine) and mixtures of compounds, the concentration series following Langmuir isotherms.
Relevant analytes in the field of food safety are antibiotics and pesticides. We apply the SERS microfluidic paper-based analytical devices for the detection of antibiotics (enoxacin, enrofloxacin) and pesticides.
The growing need to implement sensors such as NIR or Raman spectroscopy for the in-situ monitoring of bioprocesses which follows the standards of Quality by Design is either restricted by the impact of the huge water signal or by a disturbing fluorescence background originating from compounds in the culture media. Furthermore, the characterization of the bioprocess samples is challenging due to changing conditions in course of cultivation.
Here we evaluate two different process-suitable Raman spectroscopic approaches, namely time-gated Raman which bears the potential to extract the Raman signal from the fluorescence background, and cw- Raman with NIR excitation in combination with Surface Enhanced Raman Spectroscopy- (SERS) to investigate cell-free supernatants of Escherichia coli sampled over the course of a cultivation. A confocal Raman microscope was used as a reference for the process devices. The concentration of the analytes, glucose, acetate as well as metabolites such as cAMP, AMP and amino-acids were determined by offline by High-Performance Liquid Chromatography (HPLC) to serve as reference for the calibration of the Raman and SERS spectral data.
Multivariate evaluation of the Raman and SERS spectra by Partial Least Squares Regression (PLSR) yielded for most of the analytes robust correlations at each sampling point. Repeated investigation of the off-line samples over a larger experimental period suggested not only a high reliability of the Raman data in general but also a high repeatability of the SERS experiments. Similar spectral features in different quality and signal/noise ratios were measured with all three set-ups. Major results of the comparison of the different Raman spectroscopic approaches and their combination with SERS are summarized and conclusions are drawn on which approach provides the most accurate concentration data among the target analytes.
Acknowledgement
The authors kindly thank Mario Birkholz (IHP, Frankfurt (Oder), Germany) for the opportunity to use a confocal Raman microscope, Alex Bunker and Tapani Viitala (Division of Pharmaceutical Biosciences, Centre for Drug Research, University of Helsinki, Finland).