Determination of energy metabolites in cancer cells by porous graphitic carbon liquid chromatography electrospray ionization mass spectrometry for the assessment of energy metabolism |
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Authors: | Norbert Szoboszlai Xinghua Guo Olivér Ozohanics Júlia Oláh Ágnes Gömöry Victor G Mihucz András Jeney Károly Vékey |
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Institution: | 1. Laboratory of Environmental Chemistry and Bioanalytics, Department of Analytical Chemistry, Institute of Chemistry, Eötvös Loránd University, Pázmány Péter stny 1/A, H-1117 Budapest, Hungary;2. Institute of Analytical Chemistry and Food Chemistry, Graz University of Technology, Stremayrgasse 9, 8010 Graz, Austria;3. Research Centre for Natural Sciences of the Hungarian Academy of Sciences, Pusztaszeri u. 59-67, H-1025 Budapest, Hungary;4. 1st Institute of Pathology and Experimental Cancer Research, Semmelweis University, H-1085 Budapest, Hungary |
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Abstract: | A high performance liquid chromatography (HPLC) tandem mass spectrometric (MS/MS) method has been developed for the simultaneous determination of fifteen glucose, or acetate derived metabolites isolated from tumor cells. Glycolytic and tricarboxylic acid (TCA) cycle metabolites as well as acidic amino acids were separated on a HPLC porous graphitic carbon (PGC) column and simultaneously determined by means of triple quadrupole MS/MS using multiple reaction monitoring (MRM). Target compounds were eluted within 10 min with 8% v/v formic acid as an electronic modifier added to a 4:1 v/v methanol water mobile phase. The calibration is linear in the 1–100 μM concentration range for each analyte. The limit of detection ranges between 0.39 and 2.78 μM for the analytes concerned. To test the PGC–HPLC–MS/MS method in metabolomic studies, ZR-75.1 human mammary adenocarcinoma cells were labeled with U-13C glucose or 1-13C acetate. Applying the MRM mode, the incorporation of 13C into metabolites, isolated from the tumor cells, and derived from glucose or acetate, could be properly identified. |
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Keywords: | Targeted metabolomics Intracellular Triple quadrupole Tandem mass spectrometry Electrospray ionization |
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