Casey R. McAlpin

Analysis of lipids: metal oxide laser ionization mass spectrometry.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been used for lipid analysis; however, one of the drawbacks of this technique is matrix interference peaks at low masses. Metal oxide surfaces are described here for direct, matrix-free analysis of small (MW < 1000 Da) lipid compounds, without interferences in the resulting spectra from traditional matrix background peaks. Spectra from lipid standards produced protonated and sodiated molecular ions. More complex mixtures including vegetable oil shortening and lipid extracts from bacterial and algal sources provided similar results. Mechanistic insight into the mode of ionization from surface spectroscopy, negative ion mass spectrometry, and stable isotope studies is also presented. The metal oxide system is compared to other reported matrix-free systems.

Modified MALDI MS fatty acid profiling for bacterial identification.

Bacterial fatty acid profiling is a well-established technique for bacterial identification. Ten bacteria were analyzed using both positive- and negative-ion modes with a modified matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) approach using CaO as a matrix replacement (metal oxide laser ionization MS (MOLI MS)). The results show that reproducible lipid cleavage similar to thermal in situ tetramethyl ammonium hydroxide saponification/derivatization had occurred. Principal component analysis showed that replicates from each organism grouped in a unique space. Cross validation (CV) of spectra from both ionization modes resulted in greater than 94% validation of the data. When CV results were compared for the two ionization modes, negative-ion data produced a superior outcome. MOLI MS provides clinicians a rapid, reproducible and cost-effective bacterial diagnostic tool.

Enhanced matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometric analysis of bacteriophage major capsid proteins with β‐mercaptoethanol pretreatment

Bacteriophage (phage) proteins have been analyzed previously with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). However, analysis of phage major capsid proteins (MCPs) has been limited by the ability to reproducibly generate ions from MCP monomers. While the acidic conditions of MALDI-TOF MS sample preparation have been shown to aid in disassembly of some phage capsids, many require further treatment to successfully liberate MCP monomers. The findings presented here suggest that beta-mercaptoethanol reduction of the disulfide bonds linking phage MCPs prior to mass spectrometric analysis results in significantly increased MALDI-TOF MS sensitivity and reproducibility of Yersinia pestis-specific phage protein profiles.

Identification of bacteria by fatty acid profiling with direct analysis in real time mass spectrometry.

RATIONALE Bacterial fatty acid profiling is a well-established technique for bacterial identification. Current methods involving esterification and gas chromatography/mass spectrometry (GC/MS) or matrix-assisted laser desorption/ionization (MALDI) analysis are effective, but there are potential benefits to be gained by investigating ambient ionization methods that can provide rapid analysis without derivatization or additional sample handling. METHODS Lipid extracts from colonies of five Gram-positive and five Gram-negative pathogenic bacteria were analyzed by Direct Analysis in Real Time (DART) ionization coupled with a time-of-flight mass spectrometer. Fatty acid profiles were obtained from the negative-ion DART mass spectra without additional derivatization or sample preparation. RESULTS Fatty acid profiles obtained from the deprotonated molecules [M - H](-) were found to be highly species-specific and reproducible. Leave-one-out cross validation (LOOCV) for principal component analysis (PCA) showed 100% correct classification accuracy. CONCLUSIONS The results of this preliminary feasibility study show good precision and accuracy, and the fatty acid patterns are clearly distinctive for each of the ten species examined. The speed and ease of analysis and the high classification accuracy for this initial study indicate that DART is an effective method for bacterial fatty acid profiling.

Extension of metal oxide laser ionization mass spectrometry to analytes with varied chemical functionalities

RATIONALE Applications of metal oxide laser ionization mass spectrometry (MOLI MS) have been limited to compounds that provide protons for ionization upon adsorption to a metal oxide surface. The addition of a small molecule, which can act as a proton source and extend MOLI MS applications to a wider variety of analytes, was investigated. METHODS Mass spectrometric measurements were made with a PerSeptive Biosystems Voyager DE-STR MALDI mass spectrometer, using NiO-based MOLI to generate ions. NiO was pretreated in this study with a number of small molecules to act as proton sources during ionization. Peptides, monosaccharides, and a crude oil sample were analyzed with pretreated and untreated NiO. RESULTS Candidate small molecule proton donors were evaluated based on their effect on signal-to-noise ratio and peak intensity for tristearin. The best performing proton source was used for subsequent analyses. The analysis of monosaccharides, peptides and compounds in heavy crude oil was enhanced by the use of proton source treated NiO for MOLI MS. Proton/cation exchange following ionization was observed. CONCLUSIONS Addition of a proton source to NiO enhanced the MOLI MS response for peptides, carbohydrates, and compounds in a crude oil sample. Efficient ionization of these samples effectively extended applications of MOLI MS beyond lipid analytes containing esters.