Integrating Accelerated Droplet Chemistry with LC-MS for High Throughput Quantitative Analysis

Project Summary/Abstract
There is an increasing need to improve the characterization of lipids and saccharides for clinical and
biomedical research purposes. NMR is the method of choice for obtaining detailed structural information about
saccharides. However, NMR typically requires milligram (micromole) quantities of analyte; this often exceeds
biologically relevant levels. For lipids, mass spectrometry (MS) provides an efficient avenue for rapid profiling,
but quantitative analysis is challenged by difficulty in isolating species of interest due to wide structural diversity.
A new MS approach is proposed that fundamentally addresses challenges in quantitative and qualitative MS
by utilizing online accelerated droplet chemistry. Since ion suppression effects in electrospray ionization (ESI)
MS occur during droplet formation, our method is designed to tackle this intellectual challenge exactly at the
point of droplet formation – not before by adding reagents in solution, and not after by performing gas-phase
reactions. This strategy simplifies instrumentation requirements and allows effective coupling to liquid
chromatography (LC). Selected droplet-based reactions improve signal-to-noise ratios to enable femtomole
sensitivity using <1 µL sample volume. Gas-phase ion intensities generated by our platform reflect the
corresponding analyte concentration in solution. Importantly, selected droplet-based reactions allow isomers of
lipids and saccharides to be differentiated. We propose to couple online droplet reactions with LC to enable high
throughput quantification of lipids and saccharides in complex mixtures. The specific research aims are:
Aim 1: To develop a functional contained-electrospray platform for coupling accelerated droplet
chemistry on LC-MS for saccharide analysis. A novel contained-ESI source is proposed to couple droplet
chemistry with LC-MS. Our method will enable LC mobile phase and ESI spray solvent to be independently
optimized. This orthogonal feature is expected to allow effective separation of isomeric saccharides (linkage,
anomeric, and position isomers). Selected droplet reactions will improve detectability of saccharides and provide
a second layer of identification for isomers that co-elute. The LC-contained-ESI-MS/MS platform will be validated
via high throughput combinatorial studies.
Aim 2: To develop a plasma-droplet fusing contained-electrospray source for coupling LC-MS for
lipid analysis. We propose to include etched silica capillaries on our LC-contained-ESI-MS/MS platform for
accurate quantification of all types of lipids, including triglycerides. The device is expected to enable
instantaneous determination of degree of unsaturation, C=C bond position, and bond orientation (cis/trans).
The tandem development of quantitative analytical methods for lipids and saccharides will result in
concomitant creation of versatile platforms for applications in diseases diagnosis and high throughput analysis
of rare sugars to effectively guide synthetic method development. The proposed strategy will also be valuable in
biomedical research using existing instruments without modification.

Grant Number: 5R01GM149080-03
NIH Institute/Center: NIH
Principal Investigator: Abraham Badu-Tawiah