Laboratory of Analytical Chemistry, School of Chemistry, Faculty of Sciences, Aristotle University of Thessaloniki, 54124, Thessaloniki, Greece.
Laboratory of Pharmaceutical Analysis, Department of Pharmaceutical Technology, School of Pharmacy, Aristotle University of Thessaloniki, 54124, Thessaloniki, Greece.
Amino Acids. 2022 Mar;54(3):399-409. doi: 10.1007/s00726-022-03135-7. Epub 2022 Feb 19.
Histidine (His) is an essential amino acid that plays an important biological role and associated with various pathological conditions. A simple and reliable method for the determination of endogenous histidine in human saliva was optimized and validated. The analyte was separated from the saliva matrix by cation exchange chromatography and detected fluorimetrically (λ/λ = 360/440 nm) after online, specific post-column derivatization (PCD) reaction with o-phthalaldehyde. The chemical and instrumental variables of the post-column reaction were optimized using Box-Behnken experimental design to achieve maximum sensitivity. Method validation was carried out employing the total-error concept. Histidine could be analyzed reliably in the range of 0.5-5.0 μΜ, with an LOD (S/N = 3) of 50 nM. Monte Carlo simulations and capability analysis were used to investigate the ruggedness of the PCD reaction. The sampling strategy, sample preparation and stability were also investigated. Seventeen saliva samples were successfully analyzed with histidine levels being in the range of 2.7-19.5 μΜ.
组氨酸（His）是一种必需氨基酸，具有重要的生物学作用，并与各种病理状况相关。本研究优化并验证了一种用于测定人唾液内源性组氨酸的简单可靠的方法。采用阳离子交换色谱法将分析物与唾液基质分离，然后通过在线特异性柱后衍生化（PCD）反应与邻苯二甲醛衍生化，在荧光检测模式（λ/λ=360/440nm）下进行检测。采用 Box-Behnken 实验设计对 PCD 反应的化学和仪器变量进行优化，以达到最大灵敏度。采用总误差概念进行方法验证。组氨酸可在 0.5-5.0μM 的范围内可靠分析，LOD（S/N=3）为 50nM。采用蒙特卡罗模拟和能力分析研究了 PCD 反应的稳健性。还研究了采样策略、样品制备和稳定性。成功分析了 17 个唾液样本，组氨酸水平在 2.7-19.5μM 范围内。
