Lipid fingerprinting of Gram‐positive lactobacilli by intact cells – matrix‐assisted laser desorption/ionization mass spectrometry using a proton sponge based matrix

Abstract

A method of direct lipid analysis by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) in intact membranes, without prior extraction/separation steps, is described. Here, we demonstrate the efficacy of a strong base, 1,8-bis(dimethylamino)naphthalene (DMAN; proton sponge), as a novel matrix for MALDI-time-of-flight (TOF) MS analysis of whole cell bacteria. Initially, individual acidic low-molecular-weight analytes such as standard free fatty acids and phospholipids were analyzed using DMAN as matrix. Clear negative-mode MALDI-TOF MS spectra of all analytes show only deprotonated analyte signals at a low picomole limit of detection with the complete absence of matrix-related signals. These results indicate that DMAN represents a suitable matrix for MALDI-TOF MS analysis of mixtures of complex lipids as the intact membranes of microorganisms. DMAN was successfully applied to the analysis of Lactobacillus sanfranciscensis and L. plantarum microorganisms. Different components were sensitively detected in a single spot, including 16:0, 18:2, 18:3, and 21:0 free acids, glycolipids, phosphatidylglycerols (PGs) and cardiolipins. This method might be of general application, offering the advantage of quickly gaining information about lipid components of other gram-positive bacterial membranes.

Rapid lipid profiling of bacteria by online MALDI-TOF mass spectrometry

“…The experimental results show that the mass spectra obtained with chloroform/methanol as the extraction solvent present more mass peaks. The lipid profiles of the bacteria obtained in the experiment are basically consistent with previously reported profiles obtained using the traditional MALDI methods [15,36,38,39].…”

Modulation of the Lactobacillus acidophilus La-5 lipidome by different growth conditions

“…The methods used for L. acidophilus were thin layer chromatography and low-resolution fast atom bombardment mass spectrometry (Fernández Murga et al, 2001). The present study is one of only a few studies using mass spectrometry to identify the lipid species, and previously none of them have used state-of-the-art high-resolution FTMS-based lipidomics (Calvano et al, 2011;Drucker et al, 1995;Fernández Murga et al, 2001). Further studies using high-resolution MS techniques will show if the present findings can be generalized beyond L. acidophilus La-5.…”

“…The lipid classes included the lipids identified in another study of L. acidophilus (Fernández Murga et al, 2001), and additionally identified MLCL, PA and QGDG, but not diglycodiacylglycerol (DGDG). DGDG could be an intermediate for TGDG and QGDG, as the glycosyltransferase responsible for transferring glyco-residues is able to transfer up to four residues to the precursor, diacylglycerol (Hölzl & Dörmann, 2007 have not been identified in other lipid studies on Lactobacillus species (Calvano et al, 2011;Drucker et al, 1995;Exterkate et al, 1971;Fernández Murga et al, 2001;Machado et al, 2004;Murga et al, 2000;Tymczyszyn et al, 2005). One reason for this difference could be that the composition of the lipid classes is dependent on the bacterial species and strain and only one research group has investigated the lipid classes of the L. acidophilus species within the last 15 years (Fernández Murga et al, 2001;Murga et al, 2000), and this was not the same strain as L. acidophilus La-5.…”

“…A more recent study, using a novel method of matrix-assisted laser desorption-ionization time-of-flight mass spectrometry on whole cell bacteria, identified the lipid species of Lactobacillus sanfranciscensis and Lactobacillus plantarum (Calvano et al, 2011). We have found only one study on Lactobacillus species that gives the overall distribution for major lipid species.…”

Proteomic Analysis of Complex Protein Samples by MALDI–TOF Mass Spectrometry

“…In particular, MALDI MS has become a technique of considerable impact on many fi elds, from proteomics [ 11 , 12 ] to lipidomics [ 13 , 14 ] and, more recently, even metabolomics due to the introduction of new matrices such as ionic liquids [ 15 , 16 ], proton sponges [ 17 ] or aromatic amines [ 18 ], and metal nanoparticles [ 19 , 20 ].…”