Forensic Science Laboratory, Kyoto Prefectural Police H.Q., 85-3, Yabunouchi-cho, Kamigyo-ku, Kyoto-shi, Kyoto, 602-8550, Japan.
Graduate School of Biomedical Sciences, Course of Pharmaceutical Sciences, Nagasaki University, 1-14 Bunkyo-machi, Nagasaki, 852-8521, Japan.
J Chromatogr A. 2022 Aug 16;1677:463275. doi: 10.1016/j.chroma.2022.463275. Epub 2022 Jun 30.
A new fluorescence derivatization technique for the determination of the thyroid hormones, 3,3',5-triiodo-L-thyronine (T, triiodothyronine) and 3,3',5,5'-tetraiodo-L-thyronine (T, L-thyroxine), in human serum was developed based on the Sonogashira coupling reaction. This derivatization reaction was recently utilized by our research group as a promising solution for the derivatization of ortho-substituted aryl halides that suffer from steric hindrance. T and T possess amino groups that could be derivatized by many reagents; however, these reagents are not useful in the case of biological analysis as they could non-selectively react with many biogenic amines and amino acids. Thus, herein we aimed at labeling the iodo-phenyl group of T and T as a selective fluorescence labeling approach suitable for biological analysis. The fluorescent alkyne, 2-(4-ethynylphenyl)-4,5-diphenyl-1H-imidazole (DIB-ET), can label the ortho-substituted aryl halides T and T in the presence of palladium and copper as catalysts, overcoming the steric hindrance of ortho-substitution. Furthermore, the application of the proposed method for the selective analysis of T and T in biological samples was successfully performed even in the presence of numerous biological components. The formed fluorescent derivatives produced from the reaction of DIB-ET and T and T could be determined by an HPLC system with fluorescence detection. The proposed method was successfully applied for the selective and sensitive determination of T and T in human serum with detection limits (S/N = 3) of 4.0 and 6.1 ng/mL and the recovery rate in the ranges of 84.3-92.1% and 81.3-84.9%, respectively. Therefore, the proposed method could be used as a new simple tool for the simultaneous determination of T and T in biological samples.
一种基于 Sonogashira 偶联反应的新荧光衍生化技术，用于测定人血清中的甲状腺激素 3,3',5-三碘-L-甲状腺原氨酸 (T，三碘甲状腺原氨酸) 和 3,3',5,5'-四碘-L-甲状腺原氨酸 (T，L-甲状腺素)。该衍生化反应最近被我们的研究小组用作一种有前途的方法，用于衍生具有空间位阻的邻取代芳基卤化物。T 和 T 都具有氨基，可以被许多试剂衍生化；然而，这些试剂在生物分析中并不有用，因为它们可能会与许多生物胺和氨基酸非选择性反应。因此，我们旨在将 T 和 T 的碘苯基基团标记为一种适合生物分析的选择性荧光标记方法。荧光炔烃 2-(4-乙炔基苯基)-4,5-二苯基-1H-咪唑 (DIB-ET) 可以在钯和铜作为催化剂的存在下标记邻取代芳基卤化物 T 和 T，克服了邻取代的空间位阻。此外，即使在存在大量生物成分的情况下，该方法也成功地应用于生物样品中 T 和 T 的选择性分析。通过 HPLC 系统与荧光检测，可以检测到由 DIB-ET 与 T 和 T 反应生成的荧光衍生物。该方法成功地应用于人血清中 T 和 T 的选择性和灵敏测定，检测限 (S/N = 3) 分别为 4.0 和 6.1 ng/mL，回收率分别在 84.3-92.1%和 81.3-84.9%范围内。因此，该方法可用于生物样品中 T 和 T 的同时测定的新简单工具。
