Selective Separation of Highly Similar Proteins on Ionic Liquid-Loaded Mesoporous TiO₂

Abstract

Separating proteins from their mixtures is an important process in a great variety of applications, but it faces difficult challenges as soon as the proteins are simultaneously of similar sizes and carry comparable net charges. To develop both efficient and sustainable strategies for the selective separation of similar proteins and to understand the underlying molecular mechanisms to enable the separation are crucial. In this work, we propose a novel strategy where the cholinium-based amino acid [Cho][Pro] ionic liquid (IL) is used as the trace additive and loaded physically on a mesoporous TiO 2 surface for separating two similar proteins (lysozyme and cytochrome c). The observed selective adsorption behavior is explained by the hydration properties of the [Cho][Pro] loaded on the TiO 2 surface and their partially dissociated ions under different pH conditions. As the pH is increased from 5.0 to 9.8, the degree of hydration of IL ions also increases, gradually weakening the interaction strength of the proteins with the substrates, more for lysozymes, leading to their effective separation. These findings were further used to guide the detection of the retention behavior of a binary mixture of proteins in high-performance liquid chromatography, where the introduction of ILs did effectively separate the two similar proteins. Our results should further stimulate the use of ILs in the separation of proteins with a high degree of mutual similarity.

Cation Chain Length of Nonhalogenated Ionic Liquids Matters in Enhancing SERS of Cytochrome c on Zr–Al–Co–O Nanotube Arrays

“…Recently, trace amounts of biocompatible ILs were added into protein−nanostructured TiO 2 SERS substrate systems to adjust their interactions through the dissociation− hydration of ILs and to further promote protein adsorption, allowing an exceptionally low limit of protein detection down to 10 −9 M with high SERS EFs. 18,19 We have reported that a phosphonium-based IL as an additive can enhance the interaction of a protein with TiO 2 nanotube array substrates and also facilitate protein adsorption and thus boost the SERS performance. 15 Nevertheless, to the best of our knowledge, the effect of the composition and structure of ILs, in particular, the chain length of alkyl chains in ILs on the interaction of the probe molecule with the nanostructured SERS substrate and thus the SERS performance, has not been investigated.…”

“…Unlike organic solvents, ionic liquids (ILs) are good alternative additives to change the microenvironment and to regulate the protein–SERS substrate interactions. − ILs are organic salts composed of organic cations and either organic or inorganic anions with a melting point of <100 °C; depending on the structure and size of the cation and anion, many ILs are liquid at ambient temperature and can potentially replace organic solvents. Recently, trace amounts of biocompatible ILs were added into protein–nanostructured TiO 2 SERS substrate systems to adjust their interactions through the dissociation–hydration of ILs and to further promote protein adsorption, allowing an exceptionally low limit of protein detection down to 10 –9 M with high SERS EFs. , We have reported that a phosphonium-based IL as an additive can enhance the interaction of a protein with TiO 2 nanotube array substrates and also facilitate protein adsorption and thus boost the SERS performance . Nevertheless, to the best of our knowledge, the effect of the composition and structure of ILs, in particular, the chain length of alkyl chains in ILs on the interaction of the probe molecule with the nanostructured SERS substrate and thus the SERS performance, has not been investigated.…”