An analysis of avidin, biotin and their interaction at attomole levels by voltammetric and chromatographic techniques |
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Authors: | Rene Kizek Michal Masarik Karl J Kramer David Potesil Michele Bailey John A Howard Borivoj Klejdus Radka Mikelova Vojtech Adam Libuse Trnkova Frantisek Jelen |
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Affiliation: | (1) Department of Chemistry and Biochemistry, Mendel University of Agriculture and Forestry, Zemedelska 1, 613 00 Brno, Czech Republic;(2) Laboratory of Biophysical Chemistry and Molecular Oncology, Institute of Biophysics, Academy of Sciences of the Czech Republic, Kralovopolska 135, 612 65 Brno, Czech Republic;(3) Grain Marketing and Production Research Center, Agricultural Research Service, US Department of Agriculture, Manhattan, KS 66502, USA;(4) Department of Analytical Chemistry, Faculty of Science, Kotlarska 2, 611 37 Brno, Czech Republic;(5) ProdiGene Inc, 101 Gateway Boulevard, College Station, TX 77845, USA;(6) Department of Theoretical and Physical Chemistry, Faculty of Science, Kottarska 2, 611 37 Brno, Czech Republic |
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Abstract: | The electroanalytical determination of avidin in solution, in a carbon paste, and in a transgenic maize extract was performed in acidic medium at a carbon paste electrode (CPE). The oxidative voltammetric signal resulting from the presence of tyrosine and tryptophan in avidin was observed using square-wave voltammetry. The process could be used to determine avidin concentrations up to 3 fM (100 amol in 3 l drop) in solution, 700 fM (174 fmol in 250 l solution) in an avidin-modified electrode, and 174 nM in a maize seed extract. In the case of the avidin-modified CPE, several parameters were studied in order to optimize the measurements, such as electrode accumulation time, composition of the avidin-modified CPE, and the elution time of avidin. In addition, the avidin-modified electrode was used to detect biotin in solution (the detection limit was 7.6 pmol in a 6 l drop) and to detect biotin in a pharmaceutical drug after various solvent extraction procedures. Comparable studies for the detection of biotin were developed using HPLC with diode array detection (HPLC-DAD) and flow injection analysis with electrochemical detection, which allowed biotin to be detected at levels as low as 614 pM and 6.6 nM, respectively. The effects of applied potential, acetonitrile content, and flow rate of the mobile phase on the FIA-ED signal were also studied. |
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Keywords: | Avidin Biotin Avidin– biotin technology Square-wave voltammetry Modified electrode High-performance liquid chromatography Diode array detection Electrochemical transfer technique Carbon paste electrode Drug analysis |
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