JP2001081082A - New fluorescent labeling reagent 4-acylamino-7- mercapto-2,1,3-benzoxadiazole - Google Patents

New fluorescent labeling reagent 4-acylamino-7- mercapto-2,1,3-benzoxadiazole

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Publication number
JP2001081082A
JP2001081082A JP29718899A JP29718899A JP2001081082A JP 2001081082 A JP2001081082 A JP 2001081082A JP 29718899 A JP29718899 A JP 29718899A JP 29718899 A JP29718899 A JP 29718899A JP 2001081082 A JP2001081082 A JP 2001081082A
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JP
Japan
Prior art keywords
compound
benzoxadiazole
reacting
solution
aabd
Prior art date
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Japanese (ja)
Inventor
Kazuhiro Imai
一洋 今井
Tomofumi Mita
智文 三田
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Tokyo Chemical Industries Co Ltd
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Tokyo Kasei Kogyo Co Ltd
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Priority to JP29718899A priority Critical patent/JP2001081082A/en
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  • Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
  • Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
  • Heterocyclic Carbon Compounds Containing A Hetero Ring Having Nitrogen And Oxygen As The Only Ring Hetero Atoms (AREA)

Abstract

PROBLEM TO BE SOLVED: To obtain a new compound which is a compound quantitatively reactive with carboxyl group and producing a strong fluorescent substance and useful for high-sensitivity determination of carboxylic acids required by a field to which the medicine, pharmacy, agriculture and clinical chemistry belong and other fields. SOLUTION: This compound is represented by the formula [R is a (substituted) alkyl], e.g. 4-acetylamino-7-mercapto-2,1,3-benzoxadiazole. The compound is obtained by reacting 4-chloro-7-nitro-2,1,3-benzoxadiazole as a starting substance with potassium O-ethyl dithiocarbonate, dissolving the resultant yellow crystal in a mixed solution of methanol with dichloromethane and concentrated hydrochloric acid, adding and reacting an iron powder therewith, adding and reacting the resultant red powder with a mixed liquid of pyridine with acetic anhydride then distilling off the pyridine and acetic anhydride, dissolving the formed residue in ethanol, adding an aqueous solution of sodium hydroxide, adding a dilute hydrochloric acid to the reactional mixture, concentrating and purifying the resultant mixture.

Description

【発明の詳細な説明】DETAILED DESCRIPTION OF THE INVENTION

【０００１】[0001]

【産業上の利用分野】本発明はカルボキシル基と定量的
に反応し，強い蛍光性物質を生成する４−アシルアミノ
−７−メルカプト−２，１，３−ベンゾオキサジアゾー
ルに関するものであって，医薬，薬学，農学，臨床化学
の属する分野，および他の分野で要求されているカルボ
ン酸類の高感度定量に供するものである。BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a 4-acylamino-7-mercapto-2,1,3-benzoxadiazole which reacts quantitatively with a carboxyl group to produce a strong fluorescent substance. It is used for highly sensitive quantification of carboxylic acids required in the fields of medicine, pharmacy, agriculture, clinical chemistry, and other fields.

【０００２】[0002]

【従来の技術】脂肪酸，胆汁酸，プロスタグランジンな
どのカルボキシル基を有する生理活性物質やカルボキシ
ル基を有する医薬品などは生体内で重要な役割を演じて
おり，その存在量を正確に把握することが，医薬，薬
学，農学，臨床化学の属する分野，および他の分野で要
求されている。プロスタグランジン，医薬品などは極少
量で強い生理作用を示すものが多く，生体内での存在量
は極わずかである。したがって，高感度な定量法が望ま
れている。2. Description of the Related Art Physiologically active substances having a carboxyl group such as fatty acids, bile acids, and prostaglandins and pharmaceuticals having a carboxyl group play important roles in living organisms, and it is necessary to accurately determine the abundance thereof. Are required in the fields of medicine, pharmacy, agriculture, clinical chemistry, and other fields. Many prostaglandins and pharmaceuticals show strong physiological effects even in extremely small amounts, and their abundance in living bodies is extremely small. Therefore, a highly sensitive quantitative method is desired.

【０００３】近年，微量成分を検出，定量する手段とし
て高い感度と選択性を有する蛍光−液体クロマトグラフ
ィー（以下，ＨＰＬＣ）が多用されている。そして，よ
り高い感度と選択性を得るため，分析対象物を蛍光標識
することが行われており，いくつかの優れた蛍光標識試
薬が報告されている。例えば，カルボン酸類の蛍光標識
試薬として４−ブロモメチル−７−メトキシクマリン
［Ａｎａｌ．Ｃｈｅｍ．，４９，４４２（１９７
７）］，４−ブロモメチル−６，７−ジメトキシクマリ
ン［Ｊ．Ｃｈｒｏｍａｔｏｇｒ．，２１７，４９１（１
９８３）］，３−ブロモアセチル−６，７−メチレンジ
オキシクマリン［Ａｎａｌ．Ｓｃｉ．，８，６６３（１
９９２）］，３−ブロモアセチル−７−メトキシクマリ
ン［Ａｎａｌ．Ｓｃｉ．，８，６９５（１９９２）］，
４−ブロモメチル−７−アセトキシクマリン［Ｊ．Ｃｈ
ｒｏｍａｔｏｇｒ．，２３４，１２１（１９８２）］，
３−ブロモメチル−６，７−ジメトキシ−１−メチル−
２（１Ｈ）−キノキサリノン［Ａｎａｌ．Ｓｃｉ．，
１，２９５（１９８５）］，９−アントラニルジアゾメ
タン［Ａｎａｌ．Ｌｅｔｔ．，１３，１９１（１９８
０）］，１−ピレニルジアゾメタン［Ａｎａｌ．Ｓｃ
ｉ．，５，６８１（１９８９）］，ダンシルカタベリン
［Ａｎａｌｙｓｔ，１１５，１３６３（１９９０）］，
４−（５，６−ジメトキシ−２−ベンゾイミダゾイル）
ベンゾヒドラジド［Ａｎａｌ．Ｃｈｅｍ．，６０，２０
６７（１９８８）］，２−（４−ヒドラジノカルボニル
フェニル）−４，５−ジフェニルイミダゾール［Ｊ．Ｃ
ｈｒｏｍａｔｏｇｒ．，６１９，１（１９９３）］，２
−（２，３−ナフタルイミド）エチルトリフラート
［Ｊ．Ｃｈｒｏｍａｔｏｇｒ．，５０８，１３３（１９
９０）］などがある。In recent years, fluorescence-liquid chromatography (hereinafter, HPLC) having high sensitivity and selectivity has been widely used as a means for detecting and quantifying trace components. Then, in order to obtain higher sensitivity and selectivity, an analyte is fluorescently labeled, and some excellent fluorescent labeling reagents have been reported. For example, as a fluorescent labeling reagent for carboxylic acids, 4-bromomethyl-7-methoxycoumarin [Anal. Chem. , 49 , 442 (197
7)], 4-bromomethyl-6,7-dimethoxycoumarin [J. Chromatogr. , 217 , 491 (1
983)], 3-bromoacetyl-6,7-methylenedioxycoumarin [Anal. Sci. , 8 , 663 (1
992)], 3-bromoacetyl-7-methoxycoumarin [Anal. Sci. , 8 , 695 (1992)],
4-bromomethyl-7-acetoxycoumarin [J. Ch
romatogr. , 234 , 121 (1982)],
3-bromomethyl-6,7-dimethoxy-1-methyl-
2 (1H) -quinoxalinone [Anal. Sci. ,
1 , 295 (1985)], 9-anthranyldiazomethane [Anal. Lett. , 13 , 191 (198
0)], 1-pyrenyldiazomethane [Anal. Sc
i. , 5 , 681 (1989)], dansyl cataberine [Analyst, 115 , 1363 (1990)],
4- (5,6-dimethoxy-2-benzimidazoyl)
Benzohydrazide [Anal. Chem. , 60 , 20
67 (1988)], 2- (4-hydrazinocarbonylphenyl) -4,5-diphenylimidazole [J. C
Chromatogr. , 619 , 1 (1993)], 2
-(2,3-naphthalimide) ethyl triflate [J. Chromatogr. , 508 , 133 (19
90)].

【０００４】[0004]

【発明が解決しようとする課題】しかしながら，これら
蛍光標識試薬は試薬それ自体が強い蛍光を有しており，
標識反応の際に生じる副生成物も強い蛍光を有してい
る。そのため，微量のカルボン酸類定量の際に影響を与
え，満足の行く定量結果が得られない場合がある。ま
た，これら蛍光標識試薬による蛍光標識体の蛍光，励起
波長は比較的短波長であるため，複雑な組成である生体
試料中のカルボン酸類定量に当たり，夾雑物の影響を受
けやすいという問題点も有している。However, these fluorescent labeling reagents themselves have strong fluorescence,
By-products generated during the labeling reaction also have strong fluorescence. For this reason, it may affect the determination of a small amount of carboxylic acids, and a satisfactory quantitative result may not be obtained. In addition, since the fluorescence and excitation wavelength of the fluorescent label by these fluorescent labeling reagents are relatively short, there is also a problem that quantification of carboxylic acids in a biological sample having a complicated composition is easily affected by impurities. are doing.

【０００５】[0005]

【課題を解決するための手段】そこで，発明者は上記問
題点を解決すべく鋭意研究を重ねた結果，本発明化合物
がＨＰＬＣによるカルボン酸類を分離，定量するための
優れた蛍光標識試薬であることを見出し，本発明を完成
するに至った。すなわち，本発明は下記構造式（１）The inventors of the present invention have conducted intensive studies to solve the above problems, and as a result, the compound of the present invention is an excellent fluorescent labeling reagent for separating and quantifying carboxylic acids by HPLC. This led to the completion of the present invention. That is, the present invention provides the following structural formula (1)

【０００６】[0006]

【化２】 Embedded image

【０００７】で表される４−アシルアミノ−７−メルカ
プト−２，１，３−ベンゾオキサジアゾールに関するも
のである。本発明に係る上記構造式（１）で表される化
合物は文献未載の新規化合物であり，その製造法として
は，例えば，４−クロロ−７−ニトロ−２，１，３−ベ
ンゾオキサジアゾール（以下，ＮＢＤ−Ｃｌ）を出発物
質として下記反応式に従って製造することができる。The present invention relates to a 4-acylamino-7-mercapto-2,1,3-benzoxadiazole represented by the formula: The compound represented by the above structural formula (1) according to the present invention is a novel compound not described in any literature, and its production method includes, for example, 4-chloro-7-nitro-2,1,3-benzoxadiazine. It can be produced according to the following reaction formula using azole (hereinafter, NBD-Cl) as a starting material.

【０００８】[0008]

【化３】 Embedded image

【０００９】本発明化合物の１つである下記構造式
（２）の４−アセチルアミノ−７−メルカプト−２，
１，３−ベンゾオキサジアゾール（以下，ＡＡＢＤ−Ｓ
Ｈ）を代表例として，参考例を用いて本発明化合物の有
用性を明らかにする。One of the compounds of the present invention, 4-acetylamino-7-mercapto-2, of the following structural formula (2):
1,3-benzoxadiazole (hereinafter, AABD-S
Using H) as a representative example, the usefulness of the compound of the present invention will be clarified using a reference example.

【００１０】[0010]

【化４】 Embedded image

【００１１】参考例１ ＡＡＢＤ−ＳＨによる酢酸の標識体とＡＡＢＤ−ＳＨと
の蛍光強度の比較 酢酸のＡＡＢＤ−ＳＨ標識体である４−アセチルアミノ
−７−チオアセトキシ−２，１，３−ベンゾオキサジア
ゾール（以下，ＡＡＢＤ−ＳＣＯＣＨ３）とＡＡＢＤ−
ＳＨとをそれぞれアセトニトリルに溶解，１．０μＭと
し，蛍光スペクトルを測定した。その結果を表１に示
す。Reference Example 1 Comparison of Fluorescence Intensity between AABD-SH and AABD-SH Labeled Acetic Acid 4-Acetylamino-7-thioacetoxy-2,1,3-benzoate, which is an AABD-SH labeled acetic acid oxadiazole (hereinafter, AABD-SCOCH 3) and AABD-
SH and each were dissolved in acetonitrile to make 1.0 μM, and the fluorescence spectrum was measured. Table 1 shows the results.

【００１２】[0012]

【表１】 [Table 1]

【００１３】上記スペクトルにおいて，（ａ）はＡＡＢ
Ｄ−ＳＨ，（ｂ）はＡＡＢＤ−ＳＣＯＣＨ３を示す。表
から明らかなようにＡＡＢＤ−ＳＨはＡＡＢＤ−ＳＣＯ
ＣＨ３と比較し，その蛍光は無視できる程小さい。した
がって，過剰のＡＡＢＤ−ＳＨがカルボン酸の定量に際
し，影響を与える可能性は少ない。In the above spectrum, (a) is AAB
D-SH, shows a (b) is AABD-SCOCH 3. As is clear from the table, AABD-SH is AABD-SCO
Compared to CH 3 , its fluorescence is negligibly small. Therefore, it is unlikely that an excessive amount of AABD-SH will affect the determination of carboxylic acid.

【００１４】参考例２ ＡＡＢＤ−ＳＨによるカルボン酸の標識 カルボキシル基を有する化合物としてカプリル酸，カプ
リン酸，ラウリン酸，ミリスチン酸，パルミチン酸を取
り上げ，蛍光標識，ＨＰＬＣ分析を行った。 １．蛍光標識試液 ＡＡＢＤ−ＳＨをジクロロメタン溶解，２０ｍＭの溶液
を調製した。 ２．被検サンプル溶液 カプリル酸，カプリン酸，ラウリン酸，ミリスチン酸，
パルミチン酸をアセトニトリルに溶解，各１００μＭの
混合溶液を調製した。 ３．トリフェニルホスフィン（以下，ＴＰＰ）溶液 ＴＰＰをアセトニトリルに溶解，２０ｍＭの溶液を調製
した。 ４．２，２’−ジピリジルジスルヒド（以下，ＤＰＤ
Ｓ）溶液 ＤＰＤＳをアセトニトリルに溶解，２０ｍＭの溶液を調
製した。 ５．蛍光標識試液，被検サンプル溶液，ＴＰＰ溶液，Ｄ
ＰＤＳ溶液それぞれ２０μｌをバイアルビンに取り，室
温下で１５分間放置して蛍光標識反応を終了した。次い
で，アセトニトリル２０μｌを加え，この混合溶液１．
０μｌを下記条件のＨＰＬＣに注入し，蛍光検出を行っ
た。得られたクロマトグラムを表２に示す。 ホンプ ：日立Ｌ−６３００インテリジェントポンプ カラム ：ＯＤＳ−８０ＴｓＱＡ（２５０Ｘ４．６ｍｍｉ．ｄ．， ５μｍ） 溶出液 ：Ａ液；８０％（Ｖ／Ｖ）アセトニトリル−水 Ｂ液；アセトニトリル ：Ａ１００％（０−１５分），Ａ→Ｂ１００％（１５−４ ５分）Ｂ１００％（４５分以降） 流速 ：毎分１．０ｍｌ 注入量 ：１．０μｌ 蛍光検出器 ：日立Ｌ−１０８０ 検出波長 ：励起波長３６８ｎｍ，蛍光波長５２４ｎｍReference Example 2 Labeling of Carboxylic Acid with AABD-SH Caprylic acid, capric acid, lauric acid, myristic acid, and palmitic acid were taken as compounds having a carboxyl group, and fluorescent labeling and HPLC analysis were performed. 1. AABD-SH was dissolved in dichloromethane to prepare a 20 mM solution. 2. Test sample solution Caprylic acid, capric acid, lauric acid, myristic acid,
Palmitic acid was dissolved in acetonitrile to prepare mixed solutions of 100 μM each. 3. Triphenylphosphine (hereinafter, TPP) solution TPP was dissolved in acetonitrile to prepare a 20 mM solution. 4.2, 2'-dipyridyl disulfide (hereinafter referred to as DPD
S) Solution DPDS was dissolved in acetonitrile to prepare a 20 mM solution. 5. Fluorescent labeling reagent, test sample solution, TPP solution, D
20 μl of each PDS solution was placed in a vial and left at room temperature for 15 minutes to complete the fluorescent labeling reaction. Then, 20 μl of acetonitrile was added, and the mixed solution was added as follows.
0 μl was injected into the HPLC under the following conditions, and fluorescence detection was performed. Table 2 shows the obtained chromatograms. Pump: Hitachi L-6300 Intelligent Pump Column: ODS-80TsQA (250 × 4.6 mmid, 5 μm) Eluent: Solution A; 80% (V / V) acetonitrile-water B solution; Acetonitrile: A 100% (0-15) Min), A → B 100% (15-45 min) B100% (45 min or later) Flow rate: 1.0 ml / min Injection volume: 1.0 μl Fluorescence detector: Hitachi L-1080 Detection wavelength: excitation wavelength 368 nm, fluorescence Wavelength 524nm

【００１５】[0015]

【表２】 [Table 2]

【００１６】上記クロマトグラムにおいて，（１）はカ
プリル酸のＡＡＢＤ−ＳＨ標識体，（２）はカプリン酸
のＡＡＢＤ−ＳＨ標識体，（３）はラウリン酸のＡＡＢ
Ｄ−ＳＨ標識体，（４）はミリスチン酸のＡＡＢＤ−Ｓ
Ｈ標識体，（５）はパルミチン酸のＡＡＢＤ−ＳＨ標識
体である。ＡＡＢＤ−ＳＨのメルカプト基は脱水縮合剤
の存在下，室温で短時間の内にカルボキシル基と反応
し，蛍光標識反応を完結した。そして，表２から明らか
なようにカプリル酸，カプリン酸，ラウリン酸，ミリス
チン酸，パルミチン酸のそれぞれのＡＡＢＤ−ＳＨ標識
体はクロマトグラフ上，十分に分離し，精度良くそれぞ
れを定量することができる。しかもその検出限界（Ｓ／
Ｎ＝３）は１０ｆｍｏｌと高感度であった。In the above chromatogram, (1) is an AABD-SH label of caprylic acid, (2) is an AABBD-SH label of capric acid, and (3) is an AAB of lauric acid.
D-SH label, (4) is AABD-S of myristic acid
The H label, (5) is an AABD-SH label of palmitic acid. The mercapto group of AABD-SH reacted with the carboxyl group within a short period of time at room temperature in the presence of a dehydrating condensing agent to complete the fluorescent labeling reaction. And, as is clear from Table 2, each AABD-SH label of caprylic acid, capric acid, lauric acid, myristic acid, and palmitic acid can be sufficiently separated on a chromatograph, and each of them can be quantified accurately. . Moreover, its detection limit (S /
N = 3) was as high as 10 fmol.

【００１７】[0017]

【作用】本発明化合物の一つであるＡＡＢＤ−ＳＨはほ
とんど無蛍光で，カルボン酸と反応してチオエステルを
生成した場合に強い蛍光を示す。また，ベンゾオキサジ
アゾール骨格に直結したメルカプト基は，脱水縮合剤の
存在下，カルボキシル基と速やかに反応完結する。得ら
れる蛍光標識体は安定な化合物で，ＨＰＬＣでの分離分
析が可能である。AABD-SH, one of the compounds of the present invention, is almost non-fluorescent and shows strong fluorescence when it reacts with a carboxylic acid to form a thioester. Further, the mercapto group directly bonded to the benzoxadiazole skeleton quickly completes the reaction with the carboxyl group in the presence of a dehydrating condensing agent. The obtained fluorescent label is a stable compound and can be separated and analyzed by HPLC.

【００１８】[0018]

【実施例】以下に本発明の好ましい実施例を記載する
が，これは例示の目的であり，本発明を制限するもので
はない。本発明の範囲内では変形が可能なことは当業者
には明らかであろう。DESCRIPTION OF THE PREFERRED EMBODIMENTS Preferred embodiments of the present invention will be described below, but these are for the purpose of illustration and do not limit the present invention. It will be apparent to those skilled in the art that variations are possible within the scope of the invention.

【００１９】実施例１ ＡＡＢＤ−ＳＨの合成 アセトニトリル３０ｍｌにＮＢＤ−Ｃｌ５００ｍｇを溶
解させる。この溶液にジチオ炭酸カリウムＯ−エチル４
００ｍｇを加え，室温で３０分攪拌する。その後，アセ
トニトリルを留去し，得られた残渣のカラム精製を行
い，黄色結晶６６０ｍｇを得た。この黄色結晶２００ｍ
ｇをメタノール１５ｍｌ，ジクロロメタン２ｍｌ，濃塩
酸５ｍｌとの混合溶液に溶解させ，次いで鉄粉２００ｍ
ｇを加え，３０分間攪拌する。反応混合物を減圧下濃縮
し，蒸留水５０ｍｌを加える。次いで酢酸エチル１００
ｍｌで抽出する。抽出液を留去し，得られた残渣のカラ
ム精製を行い，赤色粉末７０ｍｇを得た。この赤色粉末
５０ｍｇをピリジン１ｍｌと酢酸無水物１ｍｌの混合試
液に加え，６０℃で３０分間攪拌した後，ピリジン，酢
酸無水物を留去する。残渣をエタノールに溶解し，１０
％水酸化ナトリウム水溶液１ｍｌを加え，６０℃で１０
分間攪拌する。反応混合物に希塩酸を加え酸性とし，減
圧下濃縮する。濃縮液を酢酸エチル５０ｍｌで抽出す
る。抽出液を留去し，残渣のカラム精製を行いＡＡＢＤ
−ＳＨの黄色結晶２５ｍｇを得た。Example 1 Synthesis of AABD-SH 500 mg of NBD-Cl is dissolved in 30 ml of acetonitrile. To this solution is added potassium dithiocarbonate O-ethyl 4
Add 00 mg and stir at room temperature for 30 minutes. Thereafter, acetonitrile was distilled off, and the obtained residue was subjected to column purification to obtain 660 mg of yellow crystals. 200m of this yellow crystal
g in a mixed solution of 15 ml of methanol, 2 ml of dichloromethane and 5 ml of concentrated hydrochloric acid.
g and stir for 30 minutes. The reaction mixture is concentrated under reduced pressure, and 50 ml of distilled water is added. Then ethyl acetate 100
Extract with ml. The extract was distilled off, and the obtained residue was subjected to column purification to obtain 70 mg of a red powder. 50 mg of this red powder is added to a mixture of 1 ml of pyridine and 1 ml of acetic anhydride, and the mixture is stirred at 60 ° C. for 30 minutes, and then pyridine and acetic anhydride are distilled off. Dissolve the residue in ethanol and add 10
1% aqueous sodium hydroxide solution at 60 ° C.
Stir for a minute. The reaction mixture is acidified by adding dilute hydrochloric acid, and concentrated under reduced pressure. The concentrate is extracted with 50 ml of ethyl acetate. The extract is distilled off, the residue is purified by column, and AABD
25 mg of -SH yellow crystals were obtained.

【００２０】ＡＡＢＳ−ＳＨの物性値は次の通りであ
る。 融点：２１４℃，１Ｈ ＮＭＲ（重クロロホルム）：σ
８．１７（１Ｈ，ｄ，ｊ＝８．０Ｈｚ），７．９１（１
Ｈ，ｓ），７．２３（１Ｈ，ｄ，ｊ＝８．０），４．０
６（１Ｈ，ｓ），２．２７（３Ｈ，ｓ），ＡＰＣｌ−Ｍ
Ｓ：ｍ／ｚ＝２０９（Ｍ＋）。The physical properties of AABS-SH are as follows. Melting point: 214 ° C., 1 H NMR (deuterated chloroform): σ
8.17 (1H, d, j = 8.0 Hz), 7.91 (1
H, s), 7.23 (1H, d, j = 8.0), 4.0
6 (1H, s), 2.27 (3H, s), APCl-M
S: m / z = 209 (M + ).

【００２１】[0021]

【効果】上記のように本発明化合物の代表例であるＡＡ
ＢＤ−ＳＨは，縮合剤の存在下，カルボキシル基と速や
かに反応し，チオエステルを生成する。ＡＡＢＤ−ＳＨ
それ自身は殆ど無蛍光であるが，チオエステルとなった
時に強い蛍光を有する。この時の蛍光，励起波長はとも
に長波長である。また，ＡＡＢＤ−ＳＨによるカルボン
酸の標識体であるチオエステルは，安定な化合物で，Ｈ
ＰＬＣで分離分析が可能である。以上のようにＡＡＢＤ
−ＳＨは蛍光−ＨＰＬＣ分析に当たり，夾雑物の影響を
受け難く，高感度でカルボン酸を定量することができ
る。また，ＡＡＢＤ−ＳＨそれ自身はほとんど無蛍光で
あること，および縮合剤の存在下，速やかに反応完結す
ることから，ポストラベル化への応用が可能である。こ
のようにＡＡＢＤ−ＳＨは，極微量のカルボン酸の検
出，定量に応用でき，医学，薬学，農学，臨床化学の属
する分野，および他の分野において広く利用されうる優
れた蛍光標識試薬と言える。As described above, AA which is a typical example of the compound of the present invention
BD-SH reacts quickly with a carboxyl group in the presence of a condensing agent to form a thioester. AABD-SH
It is almost non-fluorescent by itself, but has strong fluorescence when converted to a thioester. The fluorescence and excitation wavelengths at this time are both long wavelengths. A thioester, which is a labeled carboxylic acid with AABD-SH, is a stable compound,
Separation analysis is possible by PLC. AABD as described above
-SH is less affected by contaminants in fluorescence-HPLC analysis and can quantify carboxylic acid with high sensitivity. In addition, since AABD-SH itself is almost non-fluorescent and completes the reaction quickly in the presence of a condensing agent, it can be applied to post-labeling. Thus, AABD-SH is an excellent fluorescent labeling reagent that can be applied to the detection and quantification of trace amounts of carboxylic acids and can be widely used in the fields of medicine, pharmacy, agriculture, clinical chemistry, and other fields.

───────────────────────────────────────────────────── フロントページの続き Ｆターム(参考） 2G043 AA01 BA14 CA03 DA02 EA01 GA07 GB21 JA01 KA02 LA01 NA13 2G054 AA02 BA04 BB08 CD01 CE02 EA03 FA12 FA34 GA04 GB02 4C056 AA01 AB02 AC04 AD03 AE03 FA01 FB01 FB07 FB10 FC01 ──────────────────────────────────────────────────続 き Continued on the front page F term (reference) 2G043 AA01 BA14 CA03 DA02 EA01 GA07 GB21 JA01 KA02 LA01 NA13 2G054 AA02 BA04 BB08 CD01 CE02 EA03 FA12 FA34 GA04 GB02 4C056 AA01 AB02 AC04 AD03 AE03 FA01 FB01 FB07 FB10 FC01

Claims (1)

【特許請求の範囲】[Claims] 【請求項１】下記構造式 【化１】 （Ｒは置換されていてもよいアルキル基を表す）で示さ
れる新規蛍光標識試薬４−アシルアミノ−７−メルカプ
ト−２，１，３−ベンゾオキサジアゾール。(1) The following structural formula: (R represents an alkyl group which may be substituted), a novel fluorescent labeling reagent 4-acylamino-7-mercapto-2,1,3-benzoxadiazole.
JP29718899A 1999-09-13 1999-09-13 New fluorescent labeling reagent 4-acylamino-7- mercapto-2,1,3-benzoxadiazole Withdrawn JP2001081082A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
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Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
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Country Link
JP (1) JP2001081082A (en)

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2008051790A (en) * 2006-03-15 2008-03-06 Noguchi Inst Method for trace mass analysis
JP2008261707A (en) * 2007-04-11 2008-10-30 Toyota Motor Corp Deterioration degree measuring method of synthetic resin
EP2492680A1 (en) 2011-02-28 2012-08-29 Tanita Corporation Method of purifying 8-isoprostane
JP2015038114A (en) * 2007-07-26 2015-02-26 ピエール、ファーブル、メディカマン Novel fluorescent derivatives of polyamines, method for preparing the same and applications thereof as diagnosis tools in treatment of cancerous tumors
JP2016136880A (en) * 2015-01-27 2016-08-04 公益財団法人東洋食品研究所 Detection agent and detecting method of spore-forming bacteria

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2008051790A (en) * 2006-03-15 2008-03-06 Noguchi Inst Method for trace mass analysis
JP2008261707A (en) * 2007-04-11 2008-10-30 Toyota Motor Corp Deterioration degree measuring method of synthetic resin
JP2015038114A (en) * 2007-07-26 2015-02-26 ピエール、ファーブル、メディカマン Novel fluorescent derivatives of polyamines, method for preparing the same and applications thereof as diagnosis tools in treatment of cancerous tumors
EP2492680A1 (en) 2011-02-28 2012-08-29 Tanita Corporation Method of purifying 8-isoprostane
JP2016136880A (en) * 2015-01-27 2016-08-04 公益財団法人東洋食品研究所 Detection agent and detecting method of spore-forming bacteria

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