Two-dimensional (2D) chromatographic techniques have become very popular in the past decade for analyzing simple, yet challenging problems to complex samples. Compared to one-dimensional (1D) chromatography, 2D chromatographic techniques have higher selectivity and resolving power assuming the retention mechanisms are complementary. If the retention mechanisms are complementary, the theoretical peak capacity of the system will be the product of the individual peak capacities. Such systems will have immense applications in pharmaceutical and other industries.
One of the major challenges encountered in multi-dimensional separations is the incompatibility of solvents used in the two dimensions that can result in severe band dispersion or broadening and peak deterioration limiting most two-dimensional separations to reverse phase in both dimensions. Since the retention mechanisms in a reversed phase – reversed phase (RP-RP) separation are driven primarily by the hydrophobic nature of the analytes in both dimensions, the separation is not fully orthogonal and the effective peak capacity is much smaller than the theoretical peak capacity, resulting from an inability to access the full chromatographic space. Alternatively, coupling “reversed phase” to “normal phase” will offer the most complementary, useful separation due to differences in the retention mechanisms. As addressed earlier, designing such a system would be challenging if not impractical. Supercritical fluid chromatography (SFC), a form of “normal phase” liquid chromatography (NPLC), can address the incompatibility issues between dimensions. SFC is superior to NPLC due to its versatility, improved efficiency, higher throughput, and faster analysis times. Supercritical fluids have low viscosity and high diffusivity (similar to gases) to allow higher flow rates, faster re-equilibration times and have a high density (similar to liquids) to provide a high solvating power. This technique would have wider applications in pharmaceutical and other industries ranging from peak purity assessment to simultaneous achiral-achiral/chiral separations to high-throughput analysis. Also, this technique would be quite powerful compared to current RP-RP separations. Additionally, the resolved fractions can be isolated from non-polar supercritical CO2 mobile phase if desired.
The proof of concept (design) and applications of a fully automated 2D-LC-SFC system in achiral-chiral analysis of pharmaceutical compounds with multiple chiral centers, including the analysis of residual chiral metabolite in complex plasma sample, has been demonstrated. The key component of the system is the low-volume high-capacity interface between the dimensions enabling selective retention of primary column eluent and its reinjection to the secondary SFC for further separation.
The objective of this Request for Information is towards the development and demonstration of a commercially-feasible 2D-LC-SFC system that can meet the current and future analytical needs.