Department of Pharmaceutical Analytical Chemistry, Faculty of Pharmacy, Mansoura University, Mansoura, Egypt.
Department of Pharmaceutical Analytical Chemistry, Faculty of Pharmacy, Tanta University, Tanta, Egypt.
Luminescence. 2022 Jul;37(7):1162-1173. doi: 10.1002/bio.4270. Epub 2022 Jun 13.
The binding of small molecular drugs with human serum albumin (HSA) has a crucial influence on their pharmacokinetics. The binding interaction between the antihypertensive eplerenone (EPL) and HSA was investigated using multi-spectroscopic techniques for the first time. These techniques include ultraviolet-visible (UV-vis) spectroscopy, Fourier-transform infrared (FTIR), native fluorescence spectroscopy, synchronous fluorescence spectroscopy and molecular docking approach. The fluorescence spectroscopic study showed that EPL quenched HSA inherent fluorescence. The mechanism for quenching of HSA by EPL has been determined to be static in nature and confirmed by UV absorption and fluorescence spectroscopy. The modified Stern-Volmer equation was used to estimate the binding constant (K ) as well as the number of bindings (n). The results indicated that the binding occurs at a single site (K = 2.238 × 10 L mol at 298 K). The enthalpy and entropy changes (∆H and ∆S) were 58.061 and 0.258 K J mol , respectively, illustrating that the principal intermolecular interactions stabilizing the EPL-HSA system are hydrophobic forces. Synchronous fluorescence spectroscopy revealed that EPL binding to HSA occurred around the tyrosine (Tyr) residue and this agreed with the molecular docking study. The Förster resonance energy transfer (FRET) analysis confirmed the static quenching mechanism. The esterase enzyme activity of HSA was also evaluated showing its decrease in the presence of EPL. Furthermore, docking analysis and site-specific markers experiment revealed that EPL binds with HSA at subdomain IB (site III).
小分子药物与人体血清白蛋白（HSA）的结合对其药代动力学有至关重要的影响。本研究首次采用多种光谱技术研究了降压药依普利酮（EPL）与 HSA 的结合相互作用。这些技术包括紫外-可见（UV-vis）光谱、傅里叶变换红外（FTIR）光谱、天然荧光光谱、同步荧光光谱和分子对接方法。荧光光谱研究表明，EPL 猝灭了 HSA 的固有荧光。通过紫外吸收和荧光光谱证实了 EPL 猝灭 HSA 荧光的机制为静态猝灭。利用修正的 Stern-Volmer 方程估算了结合常数（K）和结合位点数（n）。结果表明，结合发生在单一结合位点（298 K 时 K = 2.238 × 10 L mol ）。焓变（∆H）和熵变（∆S）分别为 58.061 和 0.258 K J mol ，表明稳定 EPL-HSA 体系的主要分子间相互作用是疏水作用力。同步荧光光谱表明，EPL 与 HSA 结合发生在色氨酸（Tyr）残基周围，这与分子对接研究结果一致。荧光共振能量转移（FRET）分析证实了静态猝灭机制。还评估了 HSA 的酯酶酶活性，结果表明在存在 EPL 的情况下其活性降低。此外，对接分析和位点特异性标记实验表明，EPL 与 HSA 在亚结构域 IB（位点 III）结合。
