Signal Enhancement by Electrophoretic Mobility Difference in LC/ESI-MS and CE/ESI-MS

On line coupling of separation technique with mass spectrometry is a common analytical approach for the analysis of trace or complicate samples. However, the common solution compositions used in liquid chromatography or capillary electrophoresis generally contain nonvolatile salts or additives, which produce interferences and ion suppressions to ESI-MS. In this study, the concept of electrophoretic mobility difference among analytes and salts was utilized. The analytes were selectively transported to MS and signals of analytes in LC-ESI-MS and CE-ESI-MS were elevated. A strategy of postcolumn electrophoretic mobility control (EMC) was proposed to alleviate ion suppression caused by ion pairing reagents in LC-ESI-MS. Peptides analysis using LC-ESI-MS with trifluoroacetic acid (TFA) containing mobile phase, as well as aminoglycans analysis with heptafluorobutyric acid (HFBA) containing mobile phase were discussed. With regard to studies of signal suppression in CE-ESI-MS, the pressure assisted double junction interface was developed to overcome the adverse effect of nonvolatile cation salt of cetyltrimethylammonium bromide (CTAB) on triazine analysis using CTAB dynamic coating CE-ESI-MS..
In analysis of charged analytes using reversed phase liquid chromatography, a small amount of ion pairing reagent was usually added in the mobile phase to neutralize the analytes , increase the hydrophobicity of analytes and thus improve chomatographic performance . However, the strong ion pairs between ion pairing reagents and protonated analytes can not be dissociated during the electrospray ionization process and thus result in serious signal suppression in ESI-MS. The EMC was used to alliveate the problem. The EMC device was a simple electric field setup comprising a low sheath flow interface, a liquid junction and a connecting capillary (3 cm), which was located at the terminus of LC column. An electric filed was created across the connecting capillary. Due to the electric field, the positive charged peptides migrated toward the ESI sprayer, whereas the ion pairing reagent TFA anions remained in the junction reservoir and did not enter the ESI source. Ion pairs were successively dissociated.and ion suppression from TFA was alleviated. The signals for the peptides were enhanced 9-35 times. The optimized conditions were also applied to the LC-MS analysis of a tryptic digests of bovine serum albumin. Using the EMC, good chromatographic performance from TFA was maintained.
Aminoglycans are highly polar compounds, and thus are usually analyzed using reversed phase liquid chromatography with the addition of HFBA ion pairing reagents. In this study, the EMC device was utilized to dissociate ion pairs between HFBA anions and positive charged aminoglycoside cations in capLC-ESI-MS. Based on the comparison of signal intensities with and without EMC, a lower detection limit in full scan in ESI-MS as well as a better signal sensitivity in MS/MS can be achieved with EMC under optimized conditions. In addition, we also attempted to apply the EMC concept on a higher flow rate analysis (e.g., 250 μL/min). The flow injection analysis (FIA) was used to demonstrate the possibility of a high flow rate separation system coupling with EMC. The coupling of EMC with a higher flow rate system was successful after splitting flow rate to the several hundred nanoliters per minute.The signal of analytes increased due to the removement of HFBA.
A new interface for CE-ESI-MS, pressure assisted double junction interface, was proposed by the combination of pressure and electric field to alleviate the MS signal suppressions resulted by non-volatile CTAB when analyzing positive charged triazines by CTAB dynamic coating CE-ESI-MS. The pressure assisted double junction interface was constructed by the polymerization of poly(dimethylsilane)(PDMS), including a short connecting capillary(1 cm) to connect two liquid junction reservoirs. One reservoir was used as an ESI-MS interface while the other was used to connect CE separation column. The liquid reservoir connected to the separation column was sealed by a poly(ethylene ketone)(PEEK) tubing with an inserted Pt wire, and a pressure was introduced using by a syringe pump. Due to (1) the pressure flow introduced from a syringe pump toward ESI sprayer, (2) the larger electrophoretic mobility difference among analytes and CTA+ by the addition of organic phase, and (3) a reversed electric field applied across the connecting capillary, only the analytes of lower electrophoretic mobilities can be carried toward MS following the direction of pressure flow, whereas of the higher electrophoretic mobility CTA+ was controlled by the electric field to remain in the junction reservoir and did not enter the ESI source. In this way, the problems of signal suppressions caused by positive charged salt CTA+ and contamination of ion source can be successfully solved and the signals of analytes were elevated for 20-60 fold.