Multisegment Injection-Capillary Electrophoresis-Mass Spectrometry: A High-Throughput Platform for Metabolomics with High Data Fidelity

Abstract

A major constraint in large-scale mass spectrometry (MS)-based metabolomic initiatives is the low sample throughput associated with chromatographic or electrophoretic separations. Herein, we introduce multisegment injection-capillary electrophoresis-mass spectrometry (MSI-CE-MS) as a multiplexed separation platform for metabolomics that increases sample throughput up to one order of magnitude while improving overall data fidelity. We demonstrate that serial injection of seven or more discrete sample segments can be performed within a single capillary while maintaining isomeric resolution without ion suppression when using a high mass resolution time-of-flight-MS. Customized injection sequences can be devised to encode information temporally within a separation based on signal pattern recognition, which enables unambiguous identification and accurate quantification (mean bias <10%) of polar metabolites in human plasma with good reproducibility (CV ≈ 10%, n = 70). False discoveries are avoided when using a rigorous dilution trend filter to reject spurious signals and background peaks that comprise the majority (≈65%) of total detectable features. MSI-CE-MS offers an unprecedented approach to enhance sample throughput analogous to direct infusion-MS (≈3 min/sample) while delivering far greater selectivity, quantitative performance, and data quality since the same ion from different samples migrates into the ion source within a short time interval (≈2-6 min). These distinct analytical and bioinformatic merits are achieved without column switching, isotopic labeling, hardware modifications, or costly infrastructure investments.

Using direct infusion mass spectrometry for serum metabolomics in Alzheimer’s disease

“…Nevertheless, a major drawback of these conventional separation methods is the low sample throughput, so other complementary approaches are emerging in order to reduce the time of analysis. Kuehnbaum et al [11] described a multiplexed separation platform for plasma metabolomics based on multisegment injection-capillary electrophoresis-mass spectrometry, which increases sample throughput up to one order of magnitude while improving overall data fidelity. Alternatively, the use of ambient ionization techniques has extended the utility of mass spectrometry to direct analysis of complex solid materials or liquid aerosols without the need for sample pretreatment or extraction [12].…”

Personalized Metabolomics for Predicting Glucose Tolerance Changes in Sedentary Women After High-Intensity Interval Training

“…A cohort of overweight/obese women were recruited to participate in a 6-week HIIT intervention, and standardized OGTTs were performed for each subject prior to and after exercise training using a repeated measures/cross-over study design. Multi-segment injection-capillary electrophoresis-mass spectrometry (MSI-CE-MS) was used as a novel high-throughput platform in metabolomics22 for classification of plasma metabolites modulated by glucose loading and/or training status. Multiplexed separations based on MSI-CE-MS also provide an accelerated workflow for biomarker discovery since time-dependent changes in plasma metabolism are captured within a single run for an individual subject ( i.e.…”

Multisegment injections improve peptide identification rates in capillary zone electrophoresis-based bottom-up proteomics

“…A solution to this issue of a long dead time between injection and the appearance of peaks in CZE was introduced by Larmann and Jorgenson in 1993 as part of an RPLC-CZE interface (22), and redeveloped in a modified form by Kuehnbaum and colleagues in 2013 for analysis of metabolites (23). In these methods, a series of injections is performed at time periods greater than the separation window but less than the total analysis time.…”

“…In these methods, a series of injections is performed at time periods greater than the separation window but less than the total analysis time. This technology has been applied this method to the analysis of metabolites in human plasma (24–26), and demonstrated efficient generation of calibration curves, standard addition analysis, and serial dilutions (23). …”