Detection of the Brominating Activity of Myeloperoxidase Using Fluorescein

IF 0.8 4区 化学 Q4 SPECTROSCOPY Journal of Applied Spectroscopy Pub Date : 2024-05-10 DOI:10.1007/s10812-024-01723-x
D. V. Grigorieva, I. V. Gorudko, V. E. Reut, A. V. Simakin, V. A. Kostevich, N. P. Gorbunov, O. M. Panasenko, A. V. Sokolovc
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Abstract

A study was carried out on the spectralluminescent properties of fl uorescein after its reaction with various reactive oxygen and halogen species (\({O}_{2}^{\bullet-},\) H2O2, HOCl, HOBr, HOSCN, N-chloramine, taurine N-chloramine, and taurine N-bromamine) as well as in the myeloperoxidase (MPO)–H2O2–Cl/Br/SCN system. Reaction with only HOBr or with the MPO–H2O2–Br system turns fluorescein into a compound with an absorption maximum at 518 nm. The fluorescence maximum is recorded at 540 nm when excited at 520 nm, corresponding to eosin Y (brominated fluorescein). Conditions with phosphatebuffered saline (PBS) at pH 7.4 containing 137 mM NaCl, 5 mM fluorescein, 15–30 mM NaBr, and 25–50 mM H2O2 were found to be optimal for detecting HOBr in solution. A qualitative method for determining the brominating activity of MPO in vitro has been proposed. This method was used to study the effect of physiological and synthetic inhibitors as well as reactive oxygen and halogen species scavengers on the brominating activity of MPO. Our results indicate that fluorescein holds promise for use in a fluorescent method for detecting the brominating activity of mammalian hemecontaining peroxidases.

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使用荧光素检测髓过氧化物酶的溴化活性
研究了荧光素与各种活性氧和卤素(\({O}_{2}^{/bullet-},\) H2O2、HOCl、HOBr、HOSCN、N-氯胺、牛磺酸 N-氯胺和牛磺酸 N-溴胺)以及髓过氧化物酶(MPO)-H2O2-Cl-/Br-/SCN-体系反应后的光谱发光特性。仅与 HOBr 反应或与 MPO-H2O2-Br 系统反应,荧光素会变成一种在 518 纳米波长处有吸收最大值的化合物。在 520 纳米波长处激发时,荧光最大值在 540 纳米波长处记录,与曙红 Y(溴化荧光素)相对应。检测溶液中 HOBr 的最佳条件是 pH 值为 7.4 的磷酸盐缓冲液(PBS),其中含有 137 mM NaCl、5 mM 荧光素、15-30 mM NaBr 和 25-50 mM H2O2。有人提出了一种测定 MPO 体外溴化活性的定性方法。该方法用于研究生理抑制剂、合成抑制剂以及活性氧和卤素清除剂对 MPO 溴化活性的影响。我们的研究结果表明,荧光素有望用于检测哺乳动物含血过氧化物酶溴化活性的荧光方法。
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来源期刊
CiteScore
1.30
自引率
14.30%
发文量
145
审稿时长
2.5 months
期刊介绍: Journal of Applied Spectroscopy reports on many key applications of spectroscopy in chemistry, physics, metallurgy, and biology. An increasing number of papers focus on the theory of lasers, as well as the tremendous potential for the practical applications of lasers in numerous fields and industries.
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