Development of an Enzyme-responsive Nanoemulsion-type Optode based on a Novel Response Mechanism with Independent Substrate and Fluorescent Sites

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  • WADA Nayuko
    Department of Applied Chemistry, Graduate School of Engineering, Osaka Metropolitan University
  • SUEYOSHI Kenji
    Department of Applied Chemistry, Graduate School of Engineering, Osaka Metropolitan University Precursory Reserch for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency (CREST)
  • ENDO Tatsuro
    Department of Applied Chemistry, Graduate School of Engineering, Osaka Metropolitan University
  • HISAMOTO Hideaki
    Department of Applied Chemistry, Graduate School of Engineering, Osaka Metropolitan University

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Other Title
  • 基質部位・蛍光部位を独立させる新規応答機構に基づく酵素応答性ナノエマルション型オプトードの開発
  • キシツ ブイ ・ ケイコウ ブイ オ ドクリツ サセル シンキ オウトウ キコウ ニ モトズク コウソ オウトウセイ ナノエマルションガタ オプトード ノ カイハツ

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Description

<p>Optical sensors (optodes) based on hydrophobic dyes and molecular recognition molecules have been applied to a variety of sensing applications, and are expected to be applied to medical diagnosis and life science research. In our laboratory, an enzyme-responsive dye liquid nanoemulsion (NE), which combines an NE-type optode and a novel dye liquid, was developed, and a significant reduction in response time compared to the conventional thin film type was achieved. However, because the above NE uses a fluorescent substrate that has an enzyme substrate site and a fluorescent site in one molecule, it has been difficult to examine the ratio of the substrate site to the fluorescent site, which is considered effective for further increasing sensitivity. Therefore, this study aimed to propose and demonstrate an enzyme-responsive NE-type optode in which the substrate site and the fluorescent site are introduced into NE as independent molecules. In this study, naphthol AS-TR phosphate was used as an alkaline phosphatase (ALP) substrate and acridine orange (AO) derivative, which is a cationic dye that emits strong fluorescence in a hydrophobic environment, was used as a fluorescent dye. When NE containing only ALP-responsive molecules was mixed with the ALP sample, a change in the absorption spectrum of naphthol AS-TR phosphate was observed, confirming the enzymatic reaction. Subsequently, when ALP-responsive NE containing the AO/TFPB ion pair was mixed with the ALP sample, a decrease in fluorescence intensity was observed. Naphthol AS-TR phosphate was dephosphorylated by the enzymatic reaction at the oil-water interface, and the cationic dye AO moved to NE/aq. interface to balance the electrical charge in NE. Furthermore, investigating the AO/TFPB ion pair ratio for ALP-responsive molecules, it was found that it was possible to increase the sensitivity by examining the conditions. Based on these results, the development of an enzyme-responsive NE-type optode with a new response mechanism was successfully demonstrated for the first time.</p>

Journal

  • BUNSEKI KAGAKU

    BUNSEKI KAGAKU 73 (7.8), 351-356, 2024-07-05

    The Japan Society for Analytical Chemistry

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