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ID 54058
フルテキストURL
著者
Harada, Airi Department of Chemistry, Graduate School of Natural Science and Technology, Okayama University
Sasaki, Keiko Department of Earth Resources Engineering, Graduate School of Engineering, Kyushu University
Kaneta, Takashi Department of Chemistry, Graduate School of Natural Science and Technology, Okayama University
抄録
Here we describe the application of an in-capillary enzyme assay using micellar electrokinetic chromatography (MEKC) in the determination of enzyme activity in a crude culture medium containing lignin peroxidase released from Phanerochaete chrysosporium (P. chrysosporium). The method consists of a plug–plug reaction between lignin peroxidase and its substrate, veratryl alcohol, the separation of the product, veratraldehyde, from the other components including the enzyme and the culture medium, and the determination of the enzyme activity from the peak area of veratraldehyde produced by the plug–plug reaction. This method is more sensitive than conventional spectrophotometry since the background originates from the enzyme and the culture medium can be removed via MEKC separation. Veratraldehyde was separated at −10 kV in a background electrolyte containing 50 mM tartrate buffer (pH 2.5) and 50 mM sodium dodecyl sulfate (SDS) after a plug–plug reaction in the capillary for 5 min. The calibration curve of veratraldehyde was linear up to 4 pmol (500 μM) with a limit to quantification of 0.026 pmol (3.2 μM) (SN = 10). The activity of lignin peroxidase was directly measured from the peak area of veratraldehyde. The activity of lignin peroxidase released from P. chrysosporium into the medium for 7 days was successfully determined to be 3.40 UL−1.
キーワード
Lignin Peroxidase
Phanerochaete chrysosporium
Micellar electrokinetic chromatography
Capillary electrophoresis
Enzyme assay
備考
© 2016. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/
発行日
2016-04-01
出版物タイトル
Journal of Chromatography A
1440巻
出版者
Elsevier Science B.V.
開始ページ
145
終了ページ
149
ISSN
0021-9673
NCID
AA10985833
資料タイプ
学術雑誌論文
言語
English
OAI-PMH Set
岡山大学
著作権者
http://creativecommons.org/licenses/by-nc-nd/4.0/
論文のバージョン
author
DOI
関連URL
isVersionOf https://doi.org/10.1016/j.chroma.2016.02.062