Sustainable separation using high-temperature liquid chromatography (HTLC): mechanism, packing materials, challenges,...

A review of high-temperature liquid chromatography (HTLC) highlights its sustainability advantages over the traditional HPLC approach that uses hazardous organic solvents. This approach has supported the green analytical chromatography concept, most notably the replacement of toxic solvents with pure water in the mobile phase. While the existing literature contains many reviews on HTLC theory, detection, and practical applications, none have provided a comprehensive analysis covering in-depth details of all these aspects collectively. Therefore, our review addresses these gaps and highlights the greenness and sustainability of HTLC in addition to the field's modern applications and advanced practices. Implementing this environmentally benign, single-component system necessitates overcoming several technical challenges, including operating at elevated temperatures to improve water's elution strength, engineering stationary phases compatible with aqueous environments, and harnessing the distinct interaction patterns that arise between polar eluents and specialized stationary phases. Some of these improvements include new stationary phases made from a mix of organic and inorganic materials, as well as metal oxide-based stationary phases, which have shown high stability at moderate temperatures (60-90 °C). While optimized derivatives of silica-based phases (e.g., bridged ethylsiloxane/silica hybrids) show utility for biomolecule separations in the 60-90 °C range-offering improved efficiency and selectivity-their instability at elevated temperatures (>120 °C) limits applicability in HTLC. The retention behavior on such phases can be described by both linear and nonlinear van't Hoff relationships, depending on the temperature-dependent interactions between the analyte and the mobile phase, and between the analyte and the stationary phase at elevated temperature. Furthermore, challenges such as stationary phase collapse, often observed in reversed-phase systems under purely aqueous conditions, can be effectively mitigated by flushing the column in the reverse flow direction. This combination of advanced materials and method optimization highlights the potential of HTLC to align high analytical performance with environmentally conscious practices.