Optimization of capillary electrophoresis coupled to negative mode electrospray ionization-mass spectrometry using polyvinyl alcohol coated capillaries. Application to a study on non-small cell lung cancer.

Despite recent developments in separation techniques, the analysis of relatively small highly polar negatively charged analytes (e.g. small organic acids, phosphorylated sugars, and underivatized amino acids) remains challenging. Capillary electrophoresis coupled to mass spectrometry (CE-MS) has been included in the untargeted metabolomics toolbox, although mostly in positive polarity. The aim of this study was to assess the use of CE-MS to analyze highly polar and negatively charged metabolites at physiological levels without the need for derivatization. After a preliminary selection, conditions regarding CE (buffers, applied potential, injection time and applied pressure), electrospray parameters (sheath liquid flow, temperature and drying gas flow, nebulizer, and capillary voltage), and fragmentor voltage were optimized using a capillary coated with polyvinyl alcohol (PVA) for the metabolic profiling of anionic compounds compared to fused silica as the reference capillary. In addition, a database of 240 metabolites with two relative migration times (RMT) obtained against methionine sulfone and 2-morpholinoethanesulfonic acid (MES) as internal standards (IS) has been compiled. Finally, the optimized method has been used to characterize the metabolic profile of blood plasma in patients with non-small cell lung cancer (NSCLC). The identified compounds are mostly amino acids and their derivatives, carboxylic acids and organic compounds from the TCA cycle, and sugars and their phosphoderivates. In addition, we performed a comparative study to find significant differences between non-small cell lung cancer (NSCLC) vs non-cancer individuals, and squamous cell carcinoma (SCC) and adenocarcinoma (ADC) vs non-cancer individuals, respectively, searching for differences between the various types of NSCLC.