一些研究探索了各种蛋白质在固液界面上的吸附,揭示缓冲溶液在生物过程中的关键作用。然而,对该缓冲液对熔融石英上蛋白质吸收的影响的综合评价尚缺乏。这项研究采用了消逝波腔衰荡光谱(EW-CRDS)来评估缓冲溶液和pH值对三种球状蛋白的吸附动力学的影响:血红蛋白(Hb),肌红蛋白(Mb),和细胞色素c(Cyt-C)在熔融二氧化硅上。EW-CRDS工具,衰荡时间为1.4μs,最小可检测吸光度为1×10-6,能够在固液界面进行精确的光学测量。研究了三种血红素蛋白的吸附行为在pH7在三种不同的溶剂:去离子(DI)水,三(羟甲基)-氨基甲烷盐酸盐(Tris-HCl),和磷酸盐缓冲盐水(PBS)。对于每种蛋白质,表面覆盖,吸附和解吸常数,表面平衡常数通过我们的EW-CRDS工具进行光学测量。根据每种溶剂的性质,蛋白质在二氧化硅表面表现出完全不同的吸附趋势。与未缓冲的(DI水)溶液相比,在Tris-HCl和PBS缓冲液的存在下,Mb在二氧化硅表面的吸附均得到抑制。相比之下,Cyt-C吸附似乎相对不受缓冲液选择的影响,因为它涉及与表面的强静电相互作用。值得注意的是,Hb表现出相反的趋势,在Tris-HCl和PBS缓冲液存在下具有增强的蛋白质吸附。pH研究表明,蛋白质与表面之间的静电相互作用对蛋白质在二氧化硅表面的吸附有主要影响。当pH值在蛋白质的等电点附近时,吸附作用最大。这项研究证明了高灵敏度的EW-CRDS工具能够在低表面覆盖率下以快速分辨率实时研究瞬逝场限制的蛋白质种类的吸附事件。使其成为研究固液界面生物分子动力学的有价值的工具。
Several studies have explored the adsorption of various proteins onto solid-liquid interfaces, revealing the crucial role of buffer solutions in biological processes. However, a comprehensive evaluation of the buffer\'s influence on protein absorption onto fused silica is still lacking. This study employs evanescent-wave cavity ring-down spectroscopy (EW-CRDS) to assess the influence of buffer solutions and pH on the adsorption kinetics of three globular proteins: hemoglobin (Hb), myoglobin (Mb), and cytochrome c (Cyt-C) onto fused silica. The EW-CRDS tool, with a ring-down time of 1.4 μ s and a minimum detectable absorbance of 1 × 10 - 6 , enabled precise optical measurements at solid-liquid interfaces. The three heme proteins\' adsorption behavior was investigated at pH 7 in three different solvents: deionized (DI) water, tris(hydroxymethyl)-aminomethane hydrochloride (Tris-HCl), and phosphate buffered saline (PBS). For each protein, the surface coverage, the adsorption and desorption constants, and the surface equilibrium constant were optically measured by our EW-CRDS tool. Depending on the nature of each solvent, the proteins showed a completely different adsorption trend on the silica surface. The adsorption of Mb on the silica surface was depressed in the presence of both Tris-HCl and PBS buffers compared with unbuffered (DI water) solutions. In contrast, Cyt-C adsorption appears to be relatively unaffected by the choice of buffer, as it involves strong electrostatic interactions with the surface. Notably, Hb exhibits an opposite trend, with enhanced protein adsorption in the presence of Tris-HCl and PBS buffer. The pH investigations demonstrated that the electrostatic interactions between the proteins and the surface had a major influence on protein adsorption on the silica surface, with adsorption being greatest when the pH values were around the protein\'s isoelectric point. This study demonstrated the ability of the highly sensitive EW-CRDS tool to study the adsorption events of the evanescent-field-confined protein species in real-time at low surface coverages with fast resolution, making it a valuable tool for studying biomolecule kinetics at solid-liquid interfaces.