Development of a D-amino acids electrochemical sensor based on immobilization of thermostable D-proline dehydrogenase within agar gel membrane |
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Authors: | Tani Yuji Tanaka Katsuhito Yabutani Tomoki Mishima Yuji Sakuraba Haruhiko Ohshima Toshihisa Motonaka Junko |
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Institution: | a Department of Chemical Science and Technology, Institute of Technology and Science, The University of Tokushima, Minami-josanjima 2-1, Tokushima 770-8506, Japan b Tsurui Chemical Co. Ltd., 2-1-1 Niihama Arai-cho, Takasago, Hyogo 676-8686, Japan c Institute of Genetic Resources, Faculty of Agriculture, Kyushu University, Hakozaki, Higashi-ku 6-10-1, Fukuoka 812-8581, Japan |
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Abstract: | A novel biosensor for determination of d-amino acids (DAAs) in biological samples by using an electrode based on immobilization of a thermostable d-Proline dehydrogenase (d-Pro DH) within an agar gel membrane was developed. The electrode was simply prepared by spin-coating the agar solution with the d-Pro DH on a glassy carbon (GC) electrode.An electrocatalytic oxidation current of 2,6-dichloroindophenol (DCIP) was observed at −100 mV vs. Ag/AgCl with the addition of 5 and 20 mmol L−1d-proline. The current response and its relative standard deviation were 0.15 μA and 7.6% (n = 3), respectively, when it was measured in a pH 8.0 phosphate buffer solution containing 10 mmol L−1d-proline and 0.5 mmol L−1 DCIP at 50 °C. The current response of d-proline increased with increase of the temperature of the sample solution up to 70 °C. The electrocatalytic response at the d-Pro DH/agar immobilized electrode subsequently maintained for 80 days. Finally, the d-Pro DH/agar immobilized electrode was applied to determination of DAAs in a human urine sample. The determined value of DAAs in the human urine and its R.S.D. were 1.39 ± 0.12 mmol L−1 and 8.9% (n = 3), respectively. |
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Keywords: | Electrochemical biosensor Thermostable enzyme Agar Amino acids d-Proline" target="_blank">d-Proline d-Amino acid sensor" target="_blank">d-Amino acid sensor |
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