Surfactant-mediated matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in the analysis of small biological molecules

Grant, David Charles (2010) Surfactant-mediated matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in the analysis of small biological molecules. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Abstract

As an analytical method, matrix-assisted laser desorption/ionization (MALDI) has been used primarily for the analysis of large biomolecules. However, its applications to small molecules have been limited. To try to circumvent this limitation, a variety of surfactants have been tested as matrix ion suppressors for the analysis of small molecules by matrix-assisted laser desorption /ionization time-of flight mass spectrometry (MALDI-TOF-MS). Their addition to the common matrix α-cyano-4-hydroxycinnamic acid (CHCA) greatly reduces the presence of matrix-related ions when added at the appropriate mole ratio of CHCA/surfactant while still allowing the analyte signals to be observed. A range of cationic quaternary ammonium surfactants as well as a neutral and an anionic surfactant were tested for the analysis of phenolics, phenolic acids, peptides and caffeine. It was found that cationic surfactants, particularly cetyltrimethylammonium bromide (CTAB), were suitable for the analysis of acidic analytes. The anionic surfactant, sodium dodecylsulfate, showed promise for peptide analysis. The matrix:surfactant mole ratio was a critical parameter for suitable matrix ion suppression while allowing for an acceptable intensity of analyte signal. Of notable significance when using surfactants is that the resulting mass resolution of most analytes was improved by 25-75 %. No other study has observed this. -- Additional experiments were conducted to examine the homogeneity of the matrix:surfactant:analyte dried spots or order to explain the surfactant suppression phenomenon. Depth profiling of sample spots, by varying the number of laser shots, revealed that the surfactants tend to migrate toward the top of the droplet during dried droplet crystallization. It is likely that the analyte is also enriched in this surface region. This would lead to higher analyte/surfactant concentrations and reduced matrix-matrix interactions (known to be a major source of matrix-derived ions). -- Surfactant-mediated MALDI-TOF-MS has been successfully used for the identification and quantitation of flavonoids from three berry extracts: lowbush blueberry (Vaccinium angustifolium), lingonberry (Vaccinium vitis-idaea), and blackberry (Rubus armeniacus). The use of the matrix CHCA led to extensive fragmentation of the sugar moiety of the glycosides, whereas 2,4,6-trihydroxyacetophenone (THAP) allowed for the intact glycoside molecule to be observed. The flavonoids were also analyzed by LC-UV-ESI-MS for comparison. The intact flavonoids could be quantified with RSD values of less than 10% and are comparable to LC. However, the use of MALDI greatly reduces the analysis time compared to traditional LC-MS methods. -- In a second application, surfactant-mediated matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was successfully used in the analysis of caffeine and the vitamins riboflavin, nicotinamide and pyridoxine found in energy drinks. Of five common MALDI matrices tested, CHCA was found to be most suitable for the analysis of high sugar-containing energy drinks. Cetyltrimethylammonium bromide (CTAB) surfactant was suitable as a matrix-ion suppressor at a matrix:surfactant mole ratio of approximately 500:1. For comparative purposes, LC-ESI-MS with UV detection was used. The calibration curves showed substantial improvement when the surfactant-mediated method was used compared to traditional MALDI, where correlation coefficients of 0.989 (nicotinamide), 0.991 (pyridoxine), 0.983 (caffeine) and 0.987 (riboflavin) were attained. Reproducibility experiments gave RSD values ranging from 9.7 to 18.1% and quantitative results were comparable to LC-MS.

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