Mediator-lipidomics: databases and search algorithms for PUFA-derived mediators
2005; Elsevier BV; Volume: 46; Issue: 4 Linguagem: Inglês
10.1194/jlr.d400020-jlr200
ISSN1539-7262
AutoresYan Lu, Song Hong, Eric Tjonahen, Charles N. Serhan,
Tópico(s)Eicosanoids and Hypertension Pharmacology
ResumoLipid mediators (LMs) derived from PUFAs play important roles in health and disease. Databases and search algorithms are crucial, but currently unavailable, for accurate and prompt analysis of LMs via liquid chromatography-ultraviolet-tandem mass spectrometry (LC-UV-MS/MS). A novel algorithm and databases, cognoscitive-contrast-angle algorithm and databases (COCAD), were developed for the identification of LMs based on the integration of standard MS/MS spectra with chromatograms and UV spectra. Segment naming and empirical fragmentation rules were introduced to determine MS/MS ion identities, along with ion intensities used by COCAD in matching the unknown to those of authentic standards. The structures of potential LMs without synthetic and/or authentic products as standards were identified by developing theoretical databases and algorithms based on virtual LC-UV-MS/MS spectra and chromatograms. The performance of these databases and algorithms was tested by identifying LMs in murine tissues.These results indicate that COCAD has many advantages for profiling and identification of LMs compared with the conventional dot-product algorithm. Lipid mediators (LMs) derived from PUFAs play important roles in health and disease. Databases and search algorithms are crucial, but currently unavailable, for accurate and prompt analysis of LMs via liquid chromatography-ultraviolet-tandem mass spectrometry (LC-UV-MS/MS). A novel algorithm and databases, cognoscitive-contrast-angle algorithm and databases (COCAD), were developed for the identification of LMs based on the integration of standard MS/MS spectra with chromatograms and UV spectra. Segment naming and empirical fragmentation rules were introduced to determine MS/MS ion identities, along with ion intensities used by COCAD in matching the unknown to those of authentic standards. The structures of potential LMs without synthetic and/or authentic products as standards were identified by developing theoretical databases and algorithms based on virtual LC-UV-MS/MS spectra and chromatograms. The performance of these databases and algorithms was tested by identifying LMs in murine tissues. These results indicate that COCAD has many advantages for profiling and identification of LMs compared with the conventional dot-product algorithm. Lipid mediators (LMs) biosynthesized from precursor PUFAs such as eicosanoids, resolvins, neuroprotectins, and docosatrienes are local autacoids that play critical roles in human physiology and many prevalent diseases. These include cardiovascular disease, Alzheimer's disease, psoriasis, asthma, and arthritis as well as the physiologic events of inflammation and resolution (1Serhan C.N. Endogenous chemical mediators in anti-inflammation and pro-resolution.Curr. Med. Chem. 2002; 1: 177-192Google Scholar, 2Serhan C.N. Clish C.B. Brannon J. Colgan S.P. Chiang N. Gronert K.K. Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2-nonsteroidal antiinflammatory drugs and transcellular processing.J. Exp. Med. 2000; 192: 1197-1204Crossref PubMed Scopus (954) Google Scholar, 3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar). Lipidomics of LMs, namely the study of the identity, function, and profiling of potent bioactive LMs as well as their biosynthesis and metabolic inactivation, can provide a useful approach for qualifying LMs as diagnostic markers in metabolomics of disease and health and possibly in the development and assessment of new therapeutics (5Balazy M. Eicosanomics: targeted lipidomics of eicosanoids in biological systems.Prostaglandins Other Lipid Mediat. 2004; 73: 173-180Crossref PubMed Scopus (44) Google Scholar, 6Serhan C.N. Mediator lipidomics.Prostaglandins Other Lipid Mediat. 2005; In pressGoogle Scholar). As a subdivision of lipidomics, mediator-lipidomics is further subdivided into eicosanomics, which focuses on the classes of bioactive mediators derived from arachidonic acid, including, for example, prostaglandins (PGs), leukotrienes (LTs), lipoxins (LXs), epoxyeicosatrienoic acids, and isoeicosanoids, as well as LMs that are derived from other lipid structures, such as sphingolipids, diacylglycerides, lysophospholipids, and platelet-activating factors (for recent references, see 5, 6). Although the physicochemical properties of individual LMs can give different chromatographic behaviors and mass spectra that are instrument- and chromatographic system-dependent, to facilitate profiling analyses between laboratories and assembling of system metabolomic maps, it is essential to first construct databases and search algorithms that permit accurate, prompt identification of LMs in complex biological matrices such as human blood, urine, and inflammatory exudates. To this end, one specific and sensitive tool for chemical identification in mediator-lipidomics is liquid chromatography-ultraviolet-tandem mass spectrometry (LC-UV-MS/MS) with atmospheric pressure ionization (7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar, 8Griffiths W.J. Yang Y. Sjoevall J. Lindgren J. Electrospray/collision-induced dissociation mass spectrometry of mono-, di- and tri-hydroxylated lipoxygenase products, including leukotrienes of the B-series and lipoxins.Rapid Commun. Mass Spectrom. 1996; 10: 183-196Crossref PubMed Scopus (27) Google Scholar, 9Wenk M.R. Lucast L. Paolo G.D. Romanelli A.J. Suchy S.F. Nussbaum R.L. Cline G.W. Shulman G.I. McMurray W. Camilli P.D.D. Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry.Nat. Biotechnol. 2003; 21: 813-817Crossref PubMed Scopus (207) Google Scholar, 10Lee S.H. Williams M.V. DuBois R.N. Blair L.A. Targeted lipidomics using electron capture atmospheric pressure chemical ionization mass spectrometry.Rapid Commun. Mass Spectrom. 2003; 17: 2168-2176Crossref PubMed Scopus (157) Google Scholar), especially electrospray ionization (ESI). Low-energy ionization of LMs with ESI generates primarily molecular (or pseudomolecular) ions for low/intermediate energy collision-induced dissociation (CID) MS/MS analysis and can avoid unwanted degradation products (7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar). MS/MS spectra are used extensively to identify and elucidate the structures of LMs derived from PUFAs (2Serhan C.N. Clish C.B. Brannon J. Colgan S.P. Chiang N. Gronert K.K. Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2-nonsteroidal antiinflammatory drugs and transcellular processing.J. Exp. Med. 2000; 192: 1197-1204Crossref PubMed Scopus (954) Google Scholar, 3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar, 7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar, 8Griffiths W.J. Yang Y. Sjoevall J. Lindgren J. Electrospray/collision-induced dissociation mass spectrometry of mono-, di- and tri-hydroxylated lipoxygenase products, including leukotrienes of the B-series and lipoxins.Rapid Commun. Mass Spectrom. 1996; 10: 183-196Crossref PubMed Scopus (27) Google Scholar, 9Wenk M.R. Lucast L. Paolo G.D. Romanelli A.J. Suchy S.F. Nussbaum R.L. Cline G.W. Shulman G.I. McMurray W. Camilli P.D.D. Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry.Nat. Biotechnol. 2003; 21: 813-817Crossref PubMed Scopus (207) Google Scholar, 11Chiang N. Takano T. Clish C.B. Petasis N.A. Tai H. Serhan C.N.N. Aspirin-triggered 15-epi-lipoxin A4 (ATL) generation by human leukocytes and murine peritonitis exudates: development of a specific 15-epi-LXA4 ELISA.J. Pharmacol. Exp. Ther. 1998; 287: 779-790PubMed Google Scholar). LC-UV-MS/MS can also provide more spectral information and criteria for those compounds with specific UV chromophores and assist in the elucidation of both known and novel LM structures. In this context, many LMs derived from PUFAs possess conjugated double bond systems that are critical components for their bioactions; each gives a characteristic UV spectrum (2Serhan C.N. Clish C.B. Brannon J. Colgan S.P. Chiang N. Gronert K.K. Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2-nonsteroidal antiinflammatory drugs and transcellular processing.J. Exp. Med. 2000; 192: 1197-1204Crossref PubMed Scopus (954) Google Scholar, 3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar, 7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar, 8Griffiths W.J. Yang Y. Sjoevall J. Lindgren J. Electrospray/collision-induced dissociation mass spectrometry of mono-, di- and tri-hydroxylated lipoxygenase products, including leukotrienes of the B-series and lipoxins.Rapid Commun. Mass Spectrom. 1996; 10: 183-196Crossref PubMed Scopus (27) Google Scholar, 9Wenk M.R. Lucast L. Paolo G.D. Romanelli A.J. Suchy S.F. Nussbaum R.L. Cline G.W. Shulman G.I. McMurray W. Camilli P.D.D. Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry.Nat. Biotechnol. 2003; 21: 813-817Crossref PubMed Scopus (207) Google Scholar). Unesterified PUFAs frequently coexist in samples, and these compounds contain 1,4-cis-pentadiene subunits that do not give a specific UV chromophore signature. For example, leukotrienes contain a conjugated triene chromophore and lipoxins possess a conjugated tetraene chromophore. Mediator-lipidomic databases containing MS/MS as well as UV spectra and chromatographic profiles with appropriate search algorithms are imperative for accurate and timely analysis of LMs (12Lu, Y., S. Hong, E. Tjonahen, and C. N. Serhan. 2003. Lipid mediator lipidomics: database and search algorithm of electrospray ionization/tandem mass and ultraviolet spectra for structural elucidation. (5th Winter Eicosanoid Conference. Baltimore, MD, March 9–12, 2003).Google Scholar). Mediator-lipidomic analysis is summarized in Scheme 1. Although GC-MS can be used in many cases for structure identification, the required high column temperatures limit its use because many of the PUFA-derived LMs are thermolabile (7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar). Search algorithms for GC-MS electron impact ionization spectra have been well studied (13Stein S.E. Scott D.R. Optimization and testing of mass spectral library search algorithms for compound identification.J. Am. Soc. Mass Spectrom. 1994; 5: 859-866Crossref PubMed Scopus (582) Google Scholar, 14Stein S.E. Chemical substructure identification by mass spectral library searching.J. Am. Soc. Mass Spectrom. 1995; 6: 644-655Crossref PubMed Scopus (106) Google Scholar, 15Ausloos P. Clifton C.L. Lias S.G. Mikaya A.I. Stein S.E. Tchekhovskoi D.V. Sparkman O.D. Zaikin V. Zhu D.D. The critical evaluation of a comprehensive mass spectral library.J. Am. Soc. Mass Spectrom. 1999; 10: 287-299Crossref PubMed Scopus (174) Google Scholar), as have those for both GC-MS chromatograms and mass spectra (16Mallard, G. W., and J. Reed. 1997. Automated mass spectral deconvolution and identification system. U. S. Department of Commerce, Technology Administration, National Institute of Standards and Technology, Gaithersburg, MD.Google Scholar). The contrast-angle (or dot-product, = cosine of the contrast-angle) algorithm is widely used. The contrast-angle is the angle between two spectra represented as vectors composed of ordered peak intensities (13Stein S.E. Scott D.R. Optimization and testing of mass spectral library search algorithms for compound identification.J. Am. Soc. Mass Spectrom. 1994; 5: 859-866Crossref PubMed Scopus (582) Google Scholar). Because databases and search algorithms for LMs on comprehensive LC-UV-MS/MS are not yet available, we initially used MassFrontier™ (ThermoFinnigan) GC-MS mass spectral commercial software to construct a LM database and search algorithm. The search algorithm for MassFrontier is dot-product and was developed by Stein and Scott (13Stein S.E. Scott D.R. Optimization and testing of mass spectral library search algorithms for compound identification.J. Am. Soc. Mass Spectrom. 1994; 5: 859-866Crossref PubMed Scopus (582) Google Scholar, 14Stein S.E. Chemical substructure identification by mass spectral library searching.J. Am. Soc. Mass Spectrom. 1995; 6: 644-655Crossref PubMed Scopus (106) Google Scholar). The dot-product algorithm in MassFrontier uses the intensities of all peaks in MS/MS as a vector for computation and is not concerned with the identities of ions, whether they are molecular ions, ions derived from molecular ions, or ions from interfering substances in the samples (13Stein S.E. Scott D.R. Optimization and testing of mass spectral library search algorithms for compound identification.J. Am. Soc. Mass Spectrom. 1994; 5: 859-866Crossref PubMed Scopus (582) Google Scholar, 14Stein S.E. Chemical substructure identification by mass spectral library searching.J. Am. Soc. Mass Spectrom. 1995; 6: 644-655Crossref PubMed Scopus (106) Google Scholar, 15Ausloos P. Clifton C.L. Lias S.G. Mikaya A.I. Stein S.E. Tchekhovskoi D.V. Sparkman O.D. Zaikin V. Zhu D.D. The critical evaluation of a comprehensive mass spectral library.J. Am. Soc. Mass Spectrom. 1999; 10: 287-299Crossref PubMed Scopus (174) Google Scholar, 16Mallard, G. W., and J. Reed. 1997. Automated mass spectral deconvolution and identification system. U. S. Department of Commerce, Technology Administration, National Institute of Standards and Technology, Gaithersburg, MD.Google Scholar). Using the ions generated from interfering background can decrease the percentage correct for best match of the search. Even the ions generated from unknown LMs contribute differently to the identification. For LMs, ions generated by cleavage of the carbon-carbon bond inside the carbon chain have greater diagnostic yield for determining the structures than the ions formed by the loss of water or CO2, which are common features of most PUFA-derived LMs (3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar, 7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar, 8Griffiths W.J. Yang Y. Sjoevall J. Lindgren J. Electrospray/collision-induced dissociation mass spectrometry of mono-, di- and tri-hydroxylated lipoxygenase products, including leukotrienes of the B-series and lipoxins.Rapid Commun. Mass Spectrom. 1996; 10: 183-196Crossref PubMed Scopus (27) Google Scholar, 11Chiang N. Takano T. Clish C.B. Petasis N.A. Tai H. Serhan C.N.N. Aspirin-triggered 15-epi-lipoxin A4 (ATL) generation by human leukocytes and murine peritonitis exudates: development of a specific 15-epi-LXA4 ELISA.J. Pharmacol. Exp. Ther. 1998; 287: 779-790PubMed Google Scholar). The relationships between ESI-MS/MS spectra and LM structures have been studied extensively (2Serhan C.N. Clish C.B. Brannon J. Colgan S.P. Chiang N. Gronert K.K. Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2-nonsteroidal antiinflammatory drugs and transcellular processing.J. Exp. Med. 2000; 192: 1197-1204Crossref PubMed Scopus (954) Google Scholar, 3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar, 7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar, 8Griffiths W.J. Yang Y. Sjoevall J. Lindgren J. Electrospray/collision-induced dissociation mass spectrometry of mono-, di- and tri-hydroxylated lipoxygenase products, including leukotrienes of the B-series and lipoxins.Rapid Commun. Mass Spectrom. 1996; 10: 183-196Crossref PubMed Scopus (27) Google Scholar, 9Wenk M.R. Lucast L. Paolo G.D. Romanelli A.J. Suchy S.F. Nussbaum R.L. Cline G.W. Shulman G.I. McMurray W. Camilli P.D.D. Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry.Nat. Biotechnol. 2003; 21: 813-817Crossref PubMed Scopus (207) Google Scholar). The identification and elucidation of the unknown structures of novel LMs by LC-UV-MS/MS are based on the relationships between structural features (such as functional groups and double bonds) and characteristics of the spectra and chromatograms (MS/MS ions, UV spectra, and LC retention times). On C18 reversed-phase LC, chromatographic retention times (CRTs) generally increase in the following order: trihydroxy LMs, dihydroxy LMs, monohydroxy LMs, and then the nonoxidized PUFA precursors (such as arachidonic acid) are eluted, depending on the mobile phase used. For positional isomers of hydroxy-containing LMs, the CRTs decrease as the hydroxy groups locate closer to the methyl group at the ω-end of the long-chain PUFA. For instance, the CRT of 5S,14R,15S-trihydroxy-6E,8Z,10E,12E-eicosatetraenoic acid (LXB4) is usually shorter than that of 5S,6R,15S-trihydroxy-7E,9E,11Z,13E-trans-11-cis-eicosatetraenoic acid (LXA4). The CRT of native LXA4 (containing a 15S-hydroxy at carbon 15) is shorter than that of the aspirin-triggered 15-epi-LXA4 but greater than that of 11-trans-LXA4, which is the natural all-trans-containing isomer of LXA4 (17Serhan C.N. On the relationship between leukotriene and lipoxin production by human neutrophils: evidence for differential metabolism of 15-HETE and 5-HETE.Biochim. Biophys. Acta. 1989; 1004: 158-168Crossref PubMed Scopus (64) Google Scholar). The band multiplicity and maximum absorbance wavelength (λmax) of UV spectra are additional class- or series-specific signatures of LMs, such as the presence of an asymmetric singlet band with λmax ∼ 235 nm for the conjugated diene present within the lipoxygenase pathways. These include monohydroxy-eicosatetraenoic acids (mono-HETEs); a triplet with λmax ∼ 270 nm for the leukotrienes, such as the conjugated triene within LTB4 (5S,12R-dihydroxy-6Z,8E,10E,14Z-eicosatetraenoic acid); a triplet with λmax ∼ 300 nm for conjugated tetraene, as present within LXA4 and LXB4; and, for example, an asymmetric singlet with λmax ∼ 242 nm from the two conjugated dienes interrupted by a methylene group, as present in the double lipoxygenation product 5S,15S-di-HETE (17Serhan C.N. On the relationship between leukotriene and lipoxin production by human neutrophils: evidence for differential metabolism of 15-HETE and 5-HETE.Biochim. Biophys. Acta. 1989; 1004: 158-168Crossref PubMed Scopus (64) Google Scholar). For compounds without a class and/or series specific chromophore or with λmax in vacuum UV range (actually undetectable on our LC-UV-MS/MS instrument) such as 1,4-cis-pentadiene-containing PUFAs as well as some prostaglandins, namely PGE2 (11α,15S-dihydroxy-9-oxo-prosta-5Z,13E-dien-1-oic acid) and PGF2α (9α,11α,15S-trihydroxy-prosta-5Z,13E-dien-1-oic acid), which do not possess conjugated double bond systems (17Serhan C.N. On the relationship between leukotriene and lipoxin production by human neutrophils: evidence for differential metabolism of 15-HETE and 5-HETE.Biochim. Biophys. Acta. 1989; 1004: 158-168Crossref PubMed Scopus (64) Google Scholar, 18Kiss L. Bieniek E. Weissmann N. Schütte H. Sibelius U. Günther A. Bier J. Mayer K. Henneking K. Padberg W. Grimm H. Seeger W. Grimminger F.F. Simultaneous analysis of 4- and 5-series lipoxygenase and cytochrome P450 products from different biological sources by reversed-phase high-performance liquid chromatographic technique.Anal. Biochem. 1998; 261: 16-28Crossref PubMed Scopus (31) Google Scholar), the UV chromophore component is not used for identification. When the conjugated tetraene of LXA4 isomers is in the all-trans geometry, the λmax is shifted to 302 nm instead of 300 nm (17Serhan C.N. On the relationship between leukotriene and lipoxin production by human neutrophils: evidence for differential metabolism of 15-HETE and 5-HETE.Biochim. Biophys. Acta. 1989; 1004: 158-168Crossref PubMed Scopus (64) Google Scholar). The relationship between MS/MS spectra and LM structures is of interest and presented vide infra. With current chemical analytical technologies, most LMs are identified manually by comparing the spectra and chromatographic behaviors acquired from sample tissues with those of authentic standards of known LMs; when authentic standards are not available, as in the case of novel LMs and their metabolites, basic chemical structures can be obtained on the basis of the relationship between structures and features of their spectra and chromatographic behaviors compared with those of synthetic and biogenic products prepared to assist in the assignment. We routinely identify LMs by comparing the unknown spectra (MS/MS, GC-MS, and UV spectra) and CRTs with those of authentic and synthetic standards if available or with a theoretical database that consists of virtual UV and MS/MS spectra and CRTs for discovering potentially novel LMs (3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar, 12Lu, Y., S. Hong, E. Tjonahen, and C. N. Serhan. 2003. Lipid mediator lipidomics: database and search algorithm of electrospray ionization/tandem mass and ultraviolet spectra for structural elucidation. (5th Winter Eicosanoid Conference. Baltimore, MD, March 9–12, 2003).Google Scholar) if standards are not available. We initially developed a theoretical database and algorithm according to the relationships between LM structures and their spectral and chromatographic characteristics (12Lu, Y., S. Hong, E. Tjonahen, and C. N. Serhan. 2003. Lipid mediator lipidomics: database and search algorithm of electrospray ionization/tandem mass and ultraviolet spectra for structural elucidation. (5th Winter Eicosanoid Conference. Baltimore, MD, March 9–12, 2003).Google Scholar). The proposed structures of novel potential LMs in the theoretical databases were based on PUFA precursors and established biosynthetic pathways. In this report, we focus on constructing mediator-lipidomic databases and search algorithms useful for the identification of LM structures using LC-UV-MS/MS with the following objectives: 1) to assemble a database using currently available mass spectral software; 2) to construct a cognoscitive-contrast-angle algorithm and databases (COCAD) to improve the identification of LMs using MS/MS ion identities that currently cannot be determined with available software; and 3) to develop a theoretical database and algorithm for assessing potentially novel and/or unknown structures of LMs and their metabolites in biologic matrices. It is quite meaningful to develop mediator-lipidomic databases and algorithms using ion trap mass spectrometers that are relatively cheaper and popular. Moreover, the fragmentation rules and patterns for CID spectra from triple-quadruple mass spectrometers, another popular MS instrument, are similar to those we encounter using the ion trap (7Murphy R.C. Fiedler J. Hevko J. Analysis of nonvolatile lipids by mass spectrometry.Chem. Rev. 2001; 101: 479-526Crossref PubMed Scopus (236) Google Scholar, 19Serhan C.N. Jain A. Marleau S. Clish C. Kantarci A. Behbehani B. Colgan S.P. Stahl G.L. Merched A. Petasis N.A. Chan L. Van Dyke T.E.E. Reduced inflammation and tissue damage in transgenic rabbits overexpressing 15-lipoxygenase and endogenous anti-inflammatory lipid mediators.J. Immunol. 2003; 171: 6856-6865Crossref PubMed Scopus (332) Google Scholar, 20Aliberti J. Hieny S. Sousa C.R. Serhan C.N. Sher A. Lipoxin-mediated inhibition of IL-12 production by DCs: a mechanism for regulation of microbial immunity.Nat. Immunol. 2002; 3: 76-82Crossref PubMed Scopus (240) Google Scholar). The databases and algorithms described in this report should be a useful initiation that can be extended to other types of MS instruments. Chromatograms and spectra of samples and standards were acquired in negative-ion mode using a Finnigan LC-UV-ESI-LCQ ion trap tandem mass spectrometer (ThermoFinnigan, San Jose, CA). The entire LC-UV-MS system control and data analysis were performed using Xcalibur® software 1.3 (ThermoFinnigan) (2Serhan C.N. Clish C.B. Brannon J. Colgan S.P. Chiang N. Gronert K.K. Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2-nonsteroidal antiinflammatory drugs and transcellular processing.J. Exp. Med. 2000; 192: 1197-1204Crossref PubMed Scopus (954) Google Scholar, 3Serhan C.N. Hong S. Gronert K. Colgan S.P. Devchand P.R. Mirick G. Moussignac R.L.L. Resolvins: a family of bioactive products of omega-3 fatty acid transformation circuits initiated by aspirin treatment that counter proinflammation signals.J. Exp. Med. 2002; 196: 1025-1037Crossref PubMed Scopus (1362) Google Scholar, 4Hong S. Gronert K. Devchand P.R. Moussignac R.L. Serhan C.N.N. Novel docosatrienes and 17S-resolvins generated from docosahexaenoic acid in murine brain, human blood and glial cells: autocoids in anti-inflammation.J. Biol. Chem. 2003; 278: 14677-14687Abstract Full Text Full Text PDF PubMed Scopus (844) Google Scholar, 11Chiang N. Takano T. Clish C.B. Petasis N.A. Tai H. Serhan C.N.N. Aspirin-triggered 15-epi-lipoxin A4 (ATL) generation by human leukocytes and murine peritonitis exudates: development of a specific 15-epi-LXA4 ELISA.J. Pharmacol. Exp. Ther. 1998; 287: 779-790PubMed Google Scholar, 12Lu, Y., S. Hong, E. Tjonahen, and C. N. Serhan. 2003. Lipid mediator lipidomics: database and search algorithm of electrospray ionization/tandem mass and ultraviolet spectra for structural elucidation. (5th Winter Eicosanoid Conference. Baltimore, MD, March 9–12, 2003).Google Scholar). We used a LUNA C18-2 (Phenomenex, Torrance, CA) column packed with 5 μm spherical C-18 particles. The dimension was 100 mm (or 150 mm) long and 2 mm inner diameter. The column was kept in a column heater (30°C). The LC system used a P-4000 quaternary LC pump (ThermoFinnigan). When the column was 100 mm long, the mobile phase (methanol-water-acetic acid, 65:35:0.01) flo
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