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Abstract
Lipids are important mycobacterium cell wall constituents; changes are linked with drug resistance. Liquid extraction surface analysis (LESA) enables direct sampling in a highly sensitive manner. Here we describe protocols for the analysis of lipids from bacterial colonies. Lipids form various adducts, complicating spectra. Salt additives were investigated to circumvent this problem. Chloroform:methanol mixtures were studied for lipid extraction and analysis by LESA-MS. The inclusion of (ESI-compatible) acetate salts of sodium, potassium or lithium in the extraction solvent was investigated. We report the detection of bacterial cell wall lipids from mycobacterial species using LESA for the first time. Sampling protocols were optimised for the use of volatile extraction solvents. The inclusion of acetate salt additives in the sampling solvent significantly reduces spectral complexity in comparison with no additives being used. LESA offers a sensitive technique for bacterial lipid phenotyping. The inclusion of an acetate salt in the sampling solvent drives adduct formation towards a specific adduct type and thus significantly reduces spectral complexity.
Highlights
Tuberculosis (TB) continues to be the primary cause of global morbidity and mortality caused by a single infectious agent
Acyl‐phosphatidylinositol mannoside (Acyl‐Phosphatidylinositol mannosides (PIMs)) lipids, which are important cell wall constituents that have been implicated in drug‐resistance, can be detected
We present salt inclusion in Liquid extraction surface analysis (LESA) sampling solvents as an approach to promoting formation of a particular adduct type, decreasing spectral complexity
Summary
Tuberculosis (TB) continues to be the primary cause of global morbidity and mortality caused by a single infectious agent. Desorption electrospray ionisation (DESI), which desorbs analytes from a surface in a continuously sprayed jet of solvent ions, has recently been described for imaging of recombinant small molecule biocatalysts from E. coli.[4] Rapid evaporative ionisation mass spectrometry (REIMS), known as the iKnife, which continuously ablates a sample and detects gaseous ions removed from the sample, has been described for the analysis of phospholipids from bacterial colonies including P. aeruginosa, B. subtilis, and S. aureus grown on agar.[5]. We explore acetate salt addition to LESA extraction solvents for spectral simplification and structural characterisation, aiding accurate mass matching of lipid species directly to online mycobacterial databases. The extraction/ionisation solvent was 2:1 chloroform: methanol either with or without the inclusion of 10 mM acetate salt (lithium, sodium or potassium). Lithium adducts do not feature in the database; assignments made upon lithium adduct addition were searched against a calculated neutral mass after subtracting the most abundant isotopic mass of lithium from the detected m/z value
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