Signal Enhancement for Peptide Analysis in Liquid Chromatography–Electrospray Ionization Mass Spectrometry with Trifluoroacetic Acid Containing Mobile Phase by Postcolumn Electrophoretic Mobility Control

Abstract

A strategy based on postcolumn electrophoretic mobility control (EMC) was developed to alleviate the adverse effect of trifluoroacetic acid (TFA) on the liquid chromatography-mass spectrometry (LC-MS) analysis of peptides. The device created to achieve this goal consisted of a poly(dimethylsiloxane) (PDMS)-based junction reservoir, a short connecting capillary, and an electrospray ionization (ESI) sprayer connected to the outlet of the high-performance liquid chromatography (HPLC) column. By apply different voltages to the junction reservoir and the ESI emitter, an electric field was created across the connecting capillary. Due to the electric field, positively charged peptides migrated toward the ESI sprayer, whereas TFA anions remained in the junction reservoir and were removed from the ionization process. Because TFA did not enter the ESI source, ion suppression from TFA was alleviated. Operation of the postcolumn device was optimized using a peptide standard mixture. Under optimized conditions, signals for the peptides were enhanced 9-35-fold without a compromise in separation efficiency. The optimized conditions were also applied to the LC-MS analysis of a tryptic digest of bovine serum albumin.

An integrated electrophoretic mobility control device with split design for signal improvement in liquid chromatography–electrospray ionization mass spectrometry analysis of aminoglycosides using a heptafluorobutyric acid containing mobile phase

“…In the original design, the length of the connecting capillary was approximately 3e5 cm and it was noticed that the migration order might have been affected by the electrophoretic effect [22]. The migration order caused by electrophoretic mobility in the connecting column can differ from that of the HPLC column.…”

“…To minimize the electrophoretic effect on the separation, the length of the connecting capillary should be as short as possible. However, the previously proposed EMC was based on the assembly of two distinct PDMS based micro devices [22]; thus, it is difficult to insert a connecting capillary of less than 3 cm between the devices. The one-piece EMC device apparently provides the following advantages over the interface proposed earlier: (i) by controlling the splitting ratio and the interval between the separation column and connecting column, this interface is capable of accommodating the flow rate from a capillary column to conventional column, (ii) the effect of electrophoretic mobility on the separation order in high-performance liquid chromatography (HPLC) is minimized by the use of a short transfer column and (iii) without the need to assemble the EMC interface before LCeMS analysis, the operation is easier.…”

“…A strategy based on post-column electrophoretic mobility control (EMC) was proposed to alleviate the adverse effect of trifluoroacetic acid (TFA) on liquid chromatography mass spectrometry (LCeMS) analysis of peptides [22]. Peptides separated through capillary LC were directed to a connecting capillary before their detection by ESI.…”

“…However, columns with a much larger inner diameter are often used in other LCeMS applications such as the analysis of aminoglycosides. Unfortunately, the EMC device used in a capillary column [22] could not be applied to a narrow-bore column because of the mismatch in flow rate between a narrow-bore column (approximately 200 mL min À1 ) and the EMC device (less than 0.2 mL min À1 ; due to electroosmotic flow (EOF)). To overcome this difficulty, the concept of splitting was adopted for the fabrication of an EMC device.…”

Membrane-Based Continuous Remover of Trifluoroacetic Acid in Mobile Phase for LC-ESI-MS Analysis of Small Molecules and Proteins

“…The suppressor was applied for the detection of fluorocinnamic acid in negative ion polarity, and sensitivity enhancement was observed. Recently, a strategy based on post-column electrophoretic mobility control was also reported to enhance the signal of peptides [6]. However, the limitation of these solutions could still be a problem, such as sample dilution caused by post-column addition or no suitability for conventional column.…”