Application of a fluorogenic reagent, ammonium 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate for detection of cystine-containing peptides.
Application of a fluorogenic reagent, ammonium 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate for detection of cystine-containing peptides.
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应用荧光试剂 7-氟苯并-2-氧杂-1,3-二唑-4-磺酸铵检测含胱氨酸的肽。
DOI：
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发表时间：
1985
期刊：
影响因子：
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通讯作者：
K. Imai
中科院分区：
文献类型：
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作者：
T. Sueyoshi;T. Miyata;S. Iwanaga;T. Toyo′oka;K. Imai
建立了一种简便、灵敏、可靠的检测含胱氨酸多肽的方法。在碱性条件下，二硫键桥联的多肽被还原并用三丁基膦裂解，然后与硫醇专一性试剂7-氟苯并-2-氧杂-1，3-二唑-4-磺酸铵偶联。在60℃孵育1h后，在385 nm处激发，515 nm处发射，测量所形成的荧光导数。以氧化型谷胱甘肽为标准，标记的多肽荧光强度非常稳定，在100pmol~10nmol范围内可定量测定含二硫键的多肽。该方法可用于反相高效液相色谱柱外流出物中含胱氨酸多肽的专属性检测。
A simple, sensitive and reliable method for the detection of cystine-containing peptides has been developed. A peptide bridged with a disulfide bond was reduced and cleaved with tributylphosphine, and then coupled with a thiol specific reagent, ammonium 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate, under alkaline conditions. After incubation at 60 degrees C for 1 h, the fluorescent derivative formed was measured with excitation at 385 nm and emission at 515 nm. The intensity of fluorescence labeled to the peptide was very stable and the peptide containing disulfide was quantitatively determined in the range of 100 pmol to 10 nmol, when oxidized glutathione was used as a standard. This method was very useful for specific detection of cystine-containing peptides in the column effluents on reverse phase high performance liquid chromatography.