American Journal of Analytical Chemistry, 2013, 4, 16-26 http://dx.doi.org/10.4236/ajac.2013.410A2003 Published Online October 2013 (http://www.scirp.org/journal/ajac) Molecular Probes in Tandem Electrospray Ionization Mass Spectrometry: Application to Tracing Chemical Changes of Specific Phospholipid Molecular Species Hiroko Tominaga 1 , Tomoe Ishihara 1 , A. K. M. Azad Shah 2 , Rumiko Shimizu 3 , Arnold N. Onyango 4 , Hideyuki Ito 5 , Toshinori Suzuki 6 , Yasuhiro Kondo 1 , Hiroshi Koaze 7 , Koretaro Takahashi 2 , Naomichi Baba 1* 1 Division of Bioscience, Graduate School of Natural Science and Technology, Okayama University, Okayama, Japan 2 Sdivision of Marine Life Science, Graduate School of Fisheries Sciences, Hokkaido University, Hakodate, Japan 3 Faculty of Pharmaceutical Sciences, Kobe Gakuin University, Kobe, Japan 4 Jomo Kenyatta University of Agriculture and Technology, Nairobi, Kenya 5 Faculty of Health and Welfare Science, Okayama Prefectural University, Okayama, Japan 6 School of Pharmacy, Shujitsu Univerisity, Okayama, Japan 7 Department of Food Science, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido, Japan Email: * babanaom-10@t.okadai.jp Received July 17, 2013; revised August 17, 2013; accepted September 15, 2013 Copyright © 2013 Hiroko Tominaga et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. ABSTRACT New ionization and detection techniques in mass spectrometry have been successfully applied for efficient analyses of complex biological systems. It is, however, still difficult to trace structural changes of a specific molecular species in such systems. In the present study, a molecular probe strategy in combination with tandem electrospray ionization mass spectrometry has been examined using synthetic deuterium-labeled phosphatidylcholine hydroperoxide (PC-OOH/D3) and ethyl-labeled phosphatidylcholine having docosahexaenoic acid side chain (DHA-PC/Et). Administration of a mix- ture of PC-OOH/D3 and DHA-PC/Et to human blood and human skin surface, followed by extraction and analysis with collision-induced tandem electrospray ionization mass spectrometry demonstrated that metabolites of both molecular probes can be detected simultaneously with strict selectivity. The present method is also found to be useful in tracing chemical changes of the unstable docosahexaenoyl group on the surface of processed fish. The activity of phospholipase A2 can also be assessed using a phospholipid molecular probe with a linoleoyl and a deuteriomethyl group via selective detection of the lyso-phospholipid product by mass spectrometry. The advantage of the present method is that no chro- matographic separation is required and analysis can be performed under strictly the same condition for different mo- lecular probes, affording multiple data by one experiment. The present strategy may be useful for tracing time-depend- ent phenomena in dynamic phospholipid biochemistry, and can be widely used for any biological and food systems. Keywords: Molecular Probe; Stable Isotope; Mass Spectrometry; Electrospray Ionization; Lipid Peroxidation; Lipid Oxidation; Skin Surface 1. Introduction Many kinds of structurally-related phospholipid species 1 (Scheme 1) occur in biological systems due to numerous combinations of different fatty acids at sn-1 and sn-2 positions and the existence of different polar head groups [1,2]. Regioisomeric relations with different fatty acyl groups at sn-1 and sn-2 positions also increase the num- ber of phospholipid species. The major fatty acids in- clude palmitic, stearic, oleic, linoleic, linolenic, arachi- donic, icosapentaenoic and docosahexaenoic acids, and the major polar head groups are choline phosphate, serine phosphate, ethanolamine phosphate, glycerol phosphate and inositol phosphate. The different phospholipid spe- cies usually have different roles in biological systems. They may localize different subcellular particles, cell membranes and tissues, or translocate according to changes in biological micro-environments. More com- plex is that phospholipid molecules change their structure moment by moment according to the changes in physio- logical conditions [3,4]. * Corresponding author. Copyright © 2013 SciRes. AJAC