Journal of Chromatography B, 985 (2015) 75–84 Contents lists available at ScienceDirect Journal of Chromatography B jou rn al hom ep age: www.elsevier.com/locate/chromb Combination of direct infusion mass spectrometry and gas chromatography mass spectrometry for toxicometabolomic study of red blood cells and serum of mice Mus musculus after mercury exposure M.A. García-Sevillano a,b,c , T. García-Barrera a,b,c,∗∗ , F. Navarro c,d , N. Abril c,e , C. Pueyo c,e , J. López-Barea c,e , J.L. Gómez-Ariza a,b,c,∗ a Department of Chemistry and Materials Science, Faculty of Experimental Sciences, University of Huelva, Campus de El Carmen, Av. Fuerzas Armadas s/n, 21007 Huelva, Spain b Research Center on Health and Environment (CYSMA), University of Huelva, Huelva, Spain c International Campus of Excellence on Agrofood (ceiA3), University of Huelva, Huelva, Spain d Department of Environmental Biology and Public Health, Faculty of Experimental Sciences, University of Huelva, Campus El Carmen, 21007 Huelva, Spain e Department of Biochemistry and Molecular Biology, S. Ochoa Building, University of Córdoba, Rabanales Campus, 14071 Córdoba, Spain a r t i c l e i n f o Article history: Received 1 November 2014 Received in revised form 27 December 2014 Accepted 17 January 2015 Available online 25 January 2015 Keywords: Toxicometabolomics Red blood cells Serum Mercury Mus musculus Mass spectrometry a b s t r a c t Although mercury (Hg) is an important environmental and occupational pollutant, its toxicological effects, especially in serum and red blood cells (RBCs), have been scarcely studied. A toxicometabolomics workflow based on high resolution mass spectrometry approaches has been applied to investigate the toxicological effects of Hg in Mus musculus mice after subcutaneous injection for 10 days, which pro- duced inflammation and vacuolization, steatosis and karyolysis in the hepatic tissue. To this end, direct infusion mass spectrometry (DIMS) of polar and lipophilic extracts from serum and RBCs, using positive and negative mode of acquisition (ESI+/ESI-), and gas chromatography–mass spectrometry were used. A quantitative analysis of reversible oxidized thiols in serum proteins demonstrated a strong oxidative stress induction in the liver of Hg-exposed mice. Endogenous metabolites alterations were identified by partial least squares-discriminant analysis (PLS-DA). Mercury-exposed mice show perturbations in energy metabolism, amino acid metabolism, membrane phospholipid breakdown and oxidative stress- related metabolites in serum along the exposure. This work reports for the first time the effects of Hg-exposure on RBCs metabolic pathways, and reveals disturbances in glycolysis, membrane turnover, glutathione and ascorbate metabolisms. © 2015 Elsevier B.V. All rights reserved. Abbreviations: CA, arbonic anhydrase; CHAPS, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate; CK, creatine kinase; DIMS, direct infusion mass spec- trometry; EDTA, ethylenediaminetetraacetic acid; FFA, free fatty acids; GC–MS, gas chromatography–mass spectrometry; HCA, homocysteic acid; HEPES, hydroxyethyl piperazineethanesulfonic acid; IAM, iodoacetamide; IAF, iodoacetamide fluorescein; DMSO, Dimethyl sulfoxide; LC–MS, liquid chromatography–mass spectrometry; MSTFA, N-methyl-N-(trimethylsilyl) trifluoroacetamide; NMR, nuclear magnetic resonance; GSSG, oxidized glutathione; PLS-DA, partial least squares-discriminant analysis; PMSF, phenylmethanesulfonyl fluoride; PCs, phosphatidyl-cholines; RBCs, red blood cells; GSH, reduced glutathione; RNS, reactive nitrogen species; ROS, reactive oxygen species; TIC, total ion current; TAG, triacylglycerols; TMCS, trimethylchlorosilane; QqQ-TOF, triple quadrupole-time-of-flight; VIP, variable influence on the projection. ∗ Corresponding author at: Department of Chemistry and Materials Science, Faculty of Experimental Sciences, University of Huelva, Campus de El Carmen, Av. Fuerzas Armadas s/n, 21007 Huelva, Spain. Tel.: +34 959 219968; fax: +34 959 219942. ∗∗ Corresponding author at: Department of Chemistry and Materials Science, Faculty of Experimental Sciences, University of Huelva, Campus de El Carmen, Av. Fuerzas Armadas s/n, 21007 Huelva, Spain. Tel.: +34 959 219962; fax: +34 959 219942. E-mail addresses: tamara@dqcm.uhu.es (T. García-Barrera), ariza@uhu.es (J.L. Gómez-Ariza). http://dx.doi.org/10.1016/j.jchromb.2015.01.029 1570-0232/© 2015 Elsevier B.V. All rights reserved.