Microfluidic Picoliter-Scale Translational Spontaneous Sample Introduction And Its Application In High-Speed Capillary Electrophoresis

In conventional capillary electrophoresis (CE) systems, separation is usually performed in capillaries with a separation length of 20-100 cm, electric separation field lower than 500 V/cm, and sample volume in the range of 1-10 nL. Excellent separation efficiencies (>100 000 plates) can be obtained in less than 30 min. Since the 1990s, various high speed capillary electrophoresis (HSCE) systems featuring the use of short separation lengths (<15 cm) and high separation field (>500 V/cm) were developed, aiming to obtain both high speed (<100 s) and high efficiency (<1μm plate height) separations. As separation lengths are reduced to only a few centimeters to achieve high speed separation, narrow sample plugs less than 100μm (corresponding to picoliter scale plug volume) are usually required to ensure sufficient high separation efficiency. Various picoliter-scale sample injection approaches, including optical-gating injection, flow-gating injection, and microchip-based injection have been developed for HSCE systems.In this work, a novel microfluidic picoliter-scale sample introduction approach was developed by combining the spontaneous injection approach with a CE system based on a short capillary and slotted-vial array. A droplet splitting phenomenon at the capillary inlet end during the spontaneous sample introduction process was observed for the first time. On the basis of this phenomenon, a translational spontaneous injection approach was established to reduce sample injection volumes to the sub-100 pL range. A versatile HSCE system was built on the basis of this sample injection approach. The HSCE system was composed of a short fused-silica capillary and an automated sample introduction system with slotted sample and buffer reservoirs and a computer-programmed translational stage. The translational spontaneous sample injection was performed by linearly moving the stage, allowing the capillary inlet first to enter the sample solution and then removing it. A droplet was left at the tip end and spontaneously drawn into the capillary by surface tension effect to achieve sample injection. The stage was continuously moved to allow the capillary inlet to be immersed into the buffer solution, and CE separation was performed by applying a high voltage between the buffer and waste reservoirs. With the use of the novel system, high-speed and efficient capillary zone electrophoresis (CZE) separation of a mixture of five fluorescein isothiocyanate (FITC) labeled amino acids was achieved within 5.4 s in a short capillary with a separation length of 15 mm, reaching separation efficiencies up to 0.40μm plate height. Outstanding peak height precisions ranging from 1.2% to 3.7% RSD were achieved in 51 consecutive separations. By extension of the separation length to 50 mm, both high-speed and high-resolution CZE separation of eight FITC-labeled amino acids could be obtained in less than 21 s with theoretical plates ranging from 163 000 to 251 000 (corresponding to 0.31-0.20μm plate heights). Therefore, the separation performance of the present HSCE system is comparable to or even better than those reported in microfluidic chip-based CE systems.In the subsequent work, the picoliter-scale translational spontaneous injection approach and the HSCE system were applied in chiral separations of FITC-labeled amino acids under micellar electrokinetic chromatography (MEKC) mode. Amino acid enantiomers were resolved in 9.1 s over a separation length of 15 mm, using P-cyclodextrin (P-CD) and sodium taurocholate (STC) as chiral selectors.