Engineering solutions for flow control in microfluidic devices for spatial multi-dimensional liquid chromatography

Spatial multi-dimensional liquid chromatography can provide unprecedent resolving power by utilizing subsequent orthogonal separation mechanisms, while the analysis time is minimized thanks to parallel development of the analyzed fractions in each dimension. In this study, different engineering solutions have been realized aiming at achieving flow confinement and control during subsequent 1D and 2D chromatographic developments in a microfluidic device for spatial two-dimensional separations. First, the flow distributor design was optimized and physical barriers, i.e., reducing the cross section of microchannels as well as locally integrated monolithic substrates in chip segments, were assessed as a means of flow confinement. Furthermore, an on-chip activevalving, high-pressure approach was successfully developed. The flow in first-dimension separation is confined within a channel situated in a rotating axis containing through-holes. These are either closed during the 1st development or opened when aligned with the D-2 flow distributor and the D-2 channels after rotation of the axis by 90 degrees, allowing for sample transfer and executing the subsequent D-2 analysis. The on-chip active-valving concept has been successfully demonstrated in combination with stationary-phase analyte focusing prior to the D-2 development using a microfluidic spatial 2D-LC device containing polymer monolithic stationary phases locally synthesized in the parallel D-2 separation channels. Finally, this workflow was successfully employed to separate a mixture of dyes applying gradient RPLC during the D-2 development.