Optical enzyme sensor for determining L-lysine content using L-lysine oxidase from the rockfish Sebastes schlegeli.
L-lysine (L-Lys) in living bodies is critical for metabolism; therefore, determination of its levels in food is important. Most enzymatic methods for L-Lys analysis are performed using L-lysine oxidase (LyOx), but commercially manufactured LyOx is generally not highly selective for L-Lys among amino acids. We previously isolated LyOx as an antibacterial protein secreted from the skin of the rockfish Sebastes schlegeli. In the present study, we developed an optical enzyme sensor system for rapid and continuous determination of L-Lys using this LyOx. The system comprised an immobilized LyOx membrane, an optical oxygen probe, a flow system, and a personal computer. The amount of L-Lys was detected as a decrease in the oxygen concentration due to the LyOx reaction. The specificity of the sensor was examined against various amino acids. The sensor response was specific for L-Lys. Good reproducibility was obtained in 58 assays. The response of the sensor using commercially prepared LyOx was unstable compared with the response using LyOx isolated in our laboratory. Our sensor system could be used for 5 weeks without our having to change the enzyme membrane. The calibration curve for a standard L-Lys solution was linear from 0.1 to 3.0 mmol L(-1). One assay could be completed within 2 min. The sensor was applied to determine the L-Lys content in food samples such as bonito cooking water and scallop hepatopancreas. The values obtained using the sensor and conventional high-performance liquid chromatography methods were well correlated.
Endo
Hideaki
H
Department of Ocean Sciences, Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato-ku, Tokyo, 108-8477, Japan. endo@kaiyodai.ac.jp
Hayashi
Yoshitaka
Y
Kitani
Yoichiro
Y
Ren
Huifeng
H
Hayashi
Tetsuhito
T
Nagashima
Yuji
Y
D000594:Amino Acid Oxidoreductases / Q000378:metabolism*; D000818:Animals; D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods*; D002138:Calibration; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism*; D005399:Fishes / Q000378:metabolism*; D008239:Lysine / Q000032:analysis* / Q000378:metabolism*; D010084:Oxidation-Reduction; D015203:Reproducibility of Results; D013379:Substrate Specificity
D004800:Enzymes, Immobilized; D000594:Amino Acid Oxidoreductases; C041123:L-lysine oxidase; D008239:Lysine