Metabolomics assessment of vitamin D impact in Pam3CSK4 stimulation.

Bacteria are amongst the leading causes of mortality worldwide. Although several studies have proposed the possible therapeutic role of vitamin D in bacterial infection, the exact mechanism through which vitamin D functions in antibacterial immunity remains elusive. The metabolic reconfigurations induced by vitamin D in bacterial infection can therefore be explored through metabolomics, a multidisciplinary 'omics' science that evaluates the metabolic changes of a biological system by identifying and quantifying metabolites under specific conditions. In the present study, cultured U937 macrophages were treated with Pam3CSK4/mL, 1,25(OH)2D3 and a combination of Pam3CSK4/mL and 1,25(OH)2D3. These treatment/stimulated U937 cells were compared to untreated cells in order to measure the metabolic effect of vitamin D (1,25(OH)2D3). Intracellular metabolomics was performed using nuclear magnetic resonance. The obtained data were subjected to chemometric modelling and statistical analyses, which revealed a clear distinction between the metabolic profiles of Pam3CSK4 stimulated cells, 1.25(OH)2D3 supplemented cells and cells supplemented with a combination of Pam3CSK4/1.25(OH)2D3 as compared to the untreated cells. Significant differences (p < 0.05) were identified in 32 metabolites linked to bioenergy production, redox reaction regulation, inflammation and protein synthesis. The generated results illustrate that 1.25(OH)2D3 reprogrammes the metabolic profile of U937 cells stimulated with Pam3CSK4.

Untargeted metabolomics analysis reveals dynamic changes in azelaic acid- and salicylic acid derivatives in LPS-treated Nicotiana tabacum cells.

To counteract biotic stress factors, plants employ multilayered defense mechanisms responsive to pathogen-derived elicitor molecules, and regulated by different phytohormones and signaling molecules. Here, lipopolysaccharide (LPS), a microbe-associated molecular pattern (MAMP) molecule, was used to induce defense responses in Nicotiana tabacum cell suspensions. Intracellular metabolites were extracted with methanol and analyzed using a liquid chromatography-mass spectrometry (UHPLC-qTOF-MS/MS) platform. The generated data were processed and examined with multivariate and univariate statistical tools. The results show time-dependent dynamic changes and accumulation of glycosylated signaling molecules, specifically those of azelaic acid, salicylic acid and methyl-salicylate as contributors to the altered metabolomic state in LPS-treated cells.

Development and Optimization of an UPLC-QTOF-MS/MS Method Based on an In-Source Collision Induced Dissociation Approach for Comprehensive Discrimination of Chlorogenic Acids Isomers from Momordica Plant Species.

Chlorogenic acids (CGA) have been profiled in the leaves of Momordica balsamina, Momordica charantia, and Momordica foetida. All three species were found to contain the trans and cis isomers of 4-acyl para-coumaroylquinic acid (pCoQA), caffeoylquinic acid (CQA), and feruloylquinic acid (FQA). To the best of our knowledge, this is the first report of pCoQA and FQA and their cis isomers in these Momordica species. These profiles were obtained by a newly developed UPLC-qTOF-MS method based on the in-source collision induced dissociation (ISCID) method optimized to mimic the MS(2) and MS(3) fragmentation of an ion trap-based MS. The presence of the cis isomers is believed to be due to high UV exposure of these plants. Furthermore, the absence of the 3-acyl and 5-acyl CGA molecules points to a metabolic mark that is unusual and represents a very interesting biochemical phenotype of these species. Our optimized ISCID method was also shown to be able to distinguish between the geometrical isomers of all three forms of CGA, a phenomenon previously deemed impossible with other common mass spectrometry systems used for CGA analyses.