Development of Specific and Sensitive Fluorogenic Reagents for Bioactive Amines and Their Applications

生物活性胺特异性、灵敏荧光试剂的研制及其应用

• 批准号: 01571172
• 负责人: MIURA Toshiaki
• 金额: $ 0.38万
• 依托单位: Hokkaido University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1989
• 资助国家: 日本
• 起止时间: 1989 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01571172/
• 关键词: Bioactive amines; Specific fluorogenic reagents; Orthophthlaldehyde analogs; Histamine; Glutathione; Arginin; Ornithine decarboxylase; Flow injection analysis; 蛍光定量試薬; 特異的定量法; トリプトファン; オルトフタルアルデヒドアナロ-グ; ポリアミン; オルニチン脱炭素酵素; 特異試薬

In this research project, specific, sensitive and rapid methods were established for the assay of several bioactive amines by using their specific fluorogenic reagents we had developed.1. High-performance liquid chromatographic(HPLC) method was developed for the determination of putrescine in biological samples by using 1, 2-dibenzoyl-1, 4-dihydrobenzene as a precolumn fluores-cence labelling reagent. This HPLC method was shown to be applicable to the assay of ornithine decarboxylase(ODC) in rat livers.2. By using 4-bromo-1, 2-benzenedicarbaldehyde(BBDA), a specific fluorogenic reagent for histamine, we developed a simple fluorometric method for the determination of histamine in biological samples. This reagent was applied to the flow injection analysis(FIA) method, which allows 40 samples to be assayed within 1 hour. The detection limit(S/N=5) of this FIA method for histamine was demonstrated to be 0.5 pmo1/injection.HPLC method for assay of histamine was also developed by using 2-acetylbenzaldehyde(ABA) as a postcolumn fluorescence labelling reagent. This method was highly sensitive and applicable to the assay of histamine in tissue extract only after deproteinization.3. Under the appropriate reaction conditions used ABA could be employed as a specific reagent for glutathione. Thus, this reagent was employed to the simple fluorometric method and the rapid FIA method for the assay of glutathione.4. By using 2, 3-naphthalenedicarbaldehyde, a specific fluorogenic reagent for arginine, sensitive and rapid FIA method was developed for the assay of arginine in serum. Furthermore, 2, 5-dimethoxytetrahydrofuran was found to be specific fluorogenic reagent for tryptophane and applicable to the fluorometric method for the assay of tryptophane in amino acid mixtures.

本研究利用我们研制的生物活性胺特异性荧光试剂，建立了几种生物活性胺的特异、灵敏、快速的检测方法.以1，2-二苯甲酰基-1，4-二氢苯为柱前荧光标记试剂，建立了测定生物样品中腐胺的高效液相色谱法。本方法适用于大鼠肝脏中鸟氨酸脱羧酶（ODC）的含量测定.利用组胺的特异性荧光试剂4-溴-1，2-苯二甲醛（BBDA），建立了一种测定生物样品中组胺的简便荧光法。将该试剂应用于流动注射分析（FIA）方法，该方法允许在1小时内测定40个样品。组胺的检测限（S/N=5）为0.5pmol/injection，以2-乙酰苯甲醛（阿坝）为柱后荧光标记试剂，建立了高效液相色谱法测定组胺的方法。该方法灵敏度高，适用于组织提取液脱蛋白后的组胺含量测定.在适宜的反应条件下，阿坝可作为谷胱甘肽的特异性试剂。该试剂可用于谷胱甘肽的简易荧光法和快速流动注射法测定.利用精氨酸的特异性荧光试剂2，3-萘二甲醛，建立了灵敏、快速的流动注射法测定血清中精氨酸的方法。2，5-二甲氧基四氢呋喃是测定色氨酸的特异性荧光试剂，适用于氨基酸混合物中色氨酸的测定。

项目成果

Teruki Yoshimura: "Difference between Histidine and Histamine in the Mechanistic Pathway of the Fluorescene Reaction with Orthophthalaldehyde." Analytical Biochemistry.

Teruki Yoshimura：“组氨酸和组胺在与邻苯二甲醛发生荧光反应的机制途径中的差异。”

Teruki Yoshimura: "Development of a selective fluorogenic reagent for polyamines and its application to the assay of ornithine decarboxylwse(ODC)" J. PharmacobioーDynamics. 12. S-13 (1989)

Teruki Yoshimura：“多胺选择性荧光试剂的开发及其在鸟氨酸脱羧基 (ODC) 测定中的应用”J. Pharmacobio-Dynamics 12. S-13 (1989)。

Teruki Yoshimura: "Difference between Histidine and Histamine in the Mechanistic Pathway of the Fluorescence Reaction with Orthophthalaldehyde." Analytical Biochemistry. 188. 132-135 (1990)

Teruki Yoshimura：“组氨酸和组胺在与邻苯二甲醛发生荧光反应的机制途径中的差异。”

Teruki Yoshimura: "Fluorescence Reactions of Histamine with orthophthalaldehyde(OPA)Analogs.Two Types of Analogs with the Reactivity Different from OPA." Analytical Biochemistry,.

Teruki Yoshimura：“组胺与邻苯二甲醛 (OPA) 类似物的荧光反应。两种类型的类似物的反应活性与 OPA 不同。”

三浦 敏明: "生体アミンおよび酸の特異的高感度蛍光試薬の開発研究" 薬学研究の進歩. 7. 1-11 (1991)

Toshiaki Miura：“生物胺和酸的特异性高灵敏荧光试剂的开发”《药物研究进展》7. 1-11 (1991)。