Active Solvent Modulation: A Valve-Based Approach To Improve Separation Compatibility in Two-Dimensional Liquid Chromatography

Abstract

Two-dimensional liquid chromatography (2D-LC) is increasingly being viewed as a viable tool for solving difficult separation problems, ranging from targeted separations of structurally similar molecules to untargeted separations of highly complex mixtures. In spite of this performance potential, though, many users find method development challenging and most frequently cite the "incompatibility" between the solvent systems used in the first and second dimensions as a major obstacle. This solvent strength related incompatibility can lead to severe peak distortion and loss of resolution and sensitivity in the second dimension. In this paper, we describe a novel approach to address the incompatibility problem, which we refer to as Active Solvent Modulation (ASM). This valve-based approach enables dilution of D effluent with weak solvent prior to transfer to theD column but without the need for additional instrument hardware. ASM is related to the concept we refer to as Fixed Solvent Modulation (FSM), with the important difference being that ASM allows toggling of the diluent stream during each D separation cycle. In this work, we show that ASM eliminates the major drawbacks of FSM including complex elution solvent profiles, baseline disturbances, and slowD re-equilibration and demonstrate improvements in D separation quality using both simple small molecule probes and degradants of heat-treated bovine insulin as case studies. We believe that ASM will significantly ease method development for 2D-LC, providing a path to practical methods that involve both highly complementaryD and D separations and sensitive detection.

Optimizing separations in online comprehensive two‐dimensional liquid chromatography

“…In contrast, all approaches in which the 1 D effluent is modified before 2 D injection are referred to as "active modulation". To reduce the deterioration of RPLC × RPLC performance due to injectionsolvent effects (e.g., solvent strength or pH), Stoll et al have demonstrated the feasibility of adding a postcolumn dilution solvent before [53,54] and recently within the modulation interface ("active solvent modulation" [55]). Weakening the 1 D effluent increases the retention on the 2 D column head (focusing effect) and this improves peak-shape symmetry, and enhances peak height and peak capacity, despite the larger volume injected.…”

Merging metabolomics and lipidomics into one analytical run

“…Several studies proposed on-line combination of orthogonal separations either using parallel separations, 15 single heart-cut 16 or multiple heart cut LC separations. [17][18][19] 2D-LC has proved to be a powerful approach in lipidomics. [20][21][22][23] Rampler et al recently introduced an on-line combination of HILIC and reversed phase chromatography for increased coverage of hydrophilic and hydrophobic lipids.…”

Recent Advances in Chromatography for Pharmaceutical Analysis

“…In this context, several 2D-LCxSFC applications have recently been proposed for simultaneous achiral-chiral analysis, 27 25 It is clear that the greatest challenge in 2D-LC method development is to overcome incompatibility between the solvent systems, namely, the influence that the effluent of the first dimension can have on the performance of the second dimension. However, Stoll et al 29 paved the way to new possibilities and applications by proposing a valve-based approach, referred to as Active Solvent Modulation (ASM), specifically designed to address this incompatibility issue. Without the need for additional instrument hardware, ASM enables the dilution of the effluent of the first dimension with a weak solvent prior to transferring to the second dimension by using an 8 -port / 4 -position valve equipped with a bypass path.…”

“…The benefits of the approach were demonstrated using small molecule probes and the degradants of heat-treated bovine insulin as case studies. 29 It is also worth mentioning that several possible implementations of 2D-LC rely on how the effluent of the first dimension is transferred to the second dimension ( Figure 5).…”