Resolvin D1, protectin D1, and related docosahexaenoic acid-derived products: Analysis via electrospray/low energy tandem mass spectrometry based on spectra and fragmentation mechanisms

  • Song Hong
  • Yan Lu
  • Rong Yang
  • Katherine H. Gotlinger
  • Nicos A. Petasis
  • Charles N. Serhan


Resolvin D1 (RvD1) and protectin D1 (Neuroprotectin D1, PD1/NPD1) are newly identified anti-inflammatory lipid mediators biosynthesized from docosahexaenoic acid (DHA). In this report, the spectra-structure correlations and fragmentation mechanisms were studied using electrospray low-energy collision-induced dissociation tandem mass spectrometry (MS/MS) for biogenic RvD1 and PD1, as well as mono-hydroxy-DHA and related hydroperoxy-DHA. The loss of H2O and CO2 in the spectra indicates the number of functional group(s). Chain-cut ions are the signature of the positions and numbers of functional groups and double bonds. The observed chain-cut ion is equivalent to a hypothetical homolytic-segment (cc, cm, mc, or mm) with addition or extraction of up to 2 protons (H). The α-cleavage ions are equivalent to: [cc + H], with H from the hydroxyl through a β-ene or γ-ene rearrangement; [cm − 2H], with 2H from hydroxyls of PD1 through a γ-ene rearrangement, or 1H from the hydroxyl and the other H from the α-carbon of mono-HDHA through an α-H-β-ene rearrangement; [mc − H], with H from hydroxyl through a β-ene or γ-ene rearrangement, or from the α-carbon through an α-H-β-ene rearrangement; or [mm] through charge-direct fragmentations. The β-ene or γ-ene facilitates the H shift to γ position and α-cleavage. Deuterium labeling confirmed the assignment of MS/MS ions and the fragmentation mechanisms. Based on the MS/MS spectra and fragmentation mechanisms, we identified RvD1, PD1, and mono-hydroxy-DHA products in human neutrophils and blood, trout head-kidney, and stroke-injury murine brain-tissue.

Supplementary material

13361_2011_180100128_MOESM1_ESM.pdf (85 kb)
Supplementary material, approximately 87 KB.


  1. 1.
    Gruppo Italiano per lo Studio della Streptochinasi nell’Infarto Miocardico (GISSI)-Prevenzione Investigators. Dietary Supplementation with n-3 Polyunsaturated Fatty Acids and Vitamin E After Myocardial Infarction: Results of the GISSI-Prevenzione Trial. Lancet 1999, 354, 447–455.CrossRefGoogle Scholar
  2. 2.
    Freedman, S. D.; Blanco, P. G.; Zaman, M. M.; Shea, J. C.; Ollero, M.; Hopper, I. K.; Weed, D. A.; Gelrud, A.; Regan, M. M.; Lapostata, M.; Alvarez, J. G.; O’Sullivan, B. P. Association of Cystic Fibrosis with Abnormalities in Fatty Acid Metabolism. N. Engl. J. Med. 2004, 350, 560–569.CrossRefGoogle Scholar
  3. 3.
    Teitelbaum, J. E.; Walker, W. A. The Role of ω-3 Fatty Acids in Intestinal Inflammation. J. Nutr. Biochem. 2001, 12, 21–32.CrossRefGoogle Scholar
  4. 4.
    Bazan, N. G.. Nestle Nutrition Workshop Series. 1992, 28, 121–133.Google Scholar
  5. 5.
    Serhan, C. N.; Hong, S.; Gronert, K.; Colgan, S. P.; Devchand, P. R.; Mirick, G.; Moussignac, R. L. Resolvins: A Family of Bioactive Products of ω-3 Fatty Acid Transformation Circuits Initiated by Aspirin Treatment that Counter Proinflammation Signals. J. Exp. Med. 2002, 196, 1025–1037.CrossRefGoogle Scholar
  6. 6.
    Hong, S.; Gronert, K.; Devchand, P. R.; Moussignac, R. L.; Serhan, C. N. Novel Docosatrienes and 17S-Resolvins Generated from Docosahexaenoic Acid in Murine Brain, Human Blood, and Glial Cells: Autacoids in Anti-Inflammation. J. Biol. Chem. 2003, 278, 14677–14687.CrossRefGoogle Scholar
  7. 7.
    Marcheselli, L. V.; Hong, S.; Lukiw, W. J.; Tian, X. H.; Gronert, K.; Musto, A.; Hardy, M.; Gimenez, J. M.; Chiang, N.; Serhan, C. N.; Bazan, N. G. Novel Docosanoids Inhibit Brain Ischemia Reperfusion-Mediated Leukocyte Infiltration and Proinflammatory Gene Expression. J. Biol. Chem. 2003, 278, 43807–43817.CrossRefGoogle Scholar
  8. 8.
    Mukherjee, P. K.; Marcheselli, V. L.; Serhan, C. N.; Bazan, N. G. Neuroprotectin D1: A Docosahexaenoic Acid-Derived Docosatriene Protects Human Retinal Pigment Epithelial Cells from Oxidative Stress. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 8491–8496.CrossRefGoogle Scholar
  9. 9.
    Van Rollins, M.; Murphy, R. C. Autooxidation of Docosahexaenoic Acid: Analysis of Ten Isomers of Hydroxydocosahexanoate. J. Lipid Res. 1984, 25, 507–517.Google Scholar
  10. 10.
    Kim, H. Y.; Salem, N., Jr. Preparation and the Structural Determination of Hydroperoxy Derivatives of Docosahexaenoic Acid and Other Polyunsaturates by Thermospray LC/MS. Prostaglandins 1989, 37, 105–119.CrossRefGoogle Scholar
  11. 11.
    Kim, H. Y.; Karanian, J. W.; Shingu, T.; Salem, N., Jr. Stereochemical Analysis of Hydroxylated Docosahexanoates Produced by Human Platelets and Rat Brain Homogenate. Prostaglandins 1990, 40, 473–490.CrossRefGoogle Scholar
  12. 12.
    Cheng, C.; Gross, M. L. Applications and Mechanisms of Charge-Remote Fragmentation. Mass Spectrom. Rev. 2000, 19, 398–420.CrossRefGoogle Scholar
  13. 13.
    Murphy, R. C.; Fiedler, J.; Hevko, J. Analysis of Nonvolatile Lipids by Mass Spectrometry. Chem. Rev. 2001, 101, 479–526.CrossRefGoogle Scholar
  14. 14.
    Lee, 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, 2168–2176.CrossRefGoogle Scholar
  15. 15.
    Hsu, F. F.; Turk, J. Charge-Driven Fragmentation Processes in Diacyl Glycerophosphatidic Acids Upon Low-Energy Collisional Activation: A Mechanistic Proposal. J. Am. Soc. Mass Spectrom.. 2000, 11, 797.CrossRefGoogle Scholar
  16. 16.
    Griffiths, W. J. Tandem Mass Spectrometry in the Study of Fatty Acids, Bile Acids, and Steroids. Mass Spectrom. Rev.. 2003, 22, 81–152.CrossRefGoogle Scholar
  17. 17.
    Moe, M. K.; Strom, M. B.; Jensen, E.; Claeys, M. Negative Electrospray Ionization Low-Energy Tandem Mass Spectrometry of Hydroxylated Fatty Acids: A Mechanistic Study. Rapid Commun. Mass Spectrom.. 2004, 18, 1731–1740.CrossRefGoogle Scholar
  18. 18.
    James, P. F.; Perugini, M. A.; O’Hair, R. A. Sources of Artifacts in the Electrospray Ionization Mass Spectra of Saturated Diacylglycerophosphocholines: From Condensed Phase Hydrolysis Reactions to Gas Phase Intercluster Reactions. J. Am. Soc. Mass Spectrom. 2006, 17, 384–394.CrossRefGoogle Scholar
  19. 19.
    Ham, B. M.; Jacob, J. T.; Keese, M. M.; Cole, R. B. Identification, Quantification, and Comparison of Major Nonpolar Lipids in Normal and Dry Eye Tear Lipidomes by Electrospray Tandem Mass Spectrometry. J. Mass Spectrom. 2004, 39, 1321–1336.CrossRefGoogle Scholar
  20. 20.
    Ekroos, K.; Ejsing, C. S.; Bahr, U.; Karas, M.; Simons, K.; Shevchenko, A. Charting Molecular Composition of Phosphatidylcholines by Fatty Acid Scanning and Ion Trap MS3 Fragmentation. J. Lipid Res. 2003, 44, 2181–2192.CrossRefGoogle Scholar
  21. 21.
    Tomer, K. B.; Crow, F. W.; Gross, M. Location of Double Bond Position in Unsaturated Fatty Acids by Negative Ion MS/MS. J. Am. Chem. Soc. 1983, 105, 5487–5488.CrossRefGoogle Scholar
  22. 22.
    Murphy, R. C.; Barkely, R. M.; Berry, K. Z.; Hankin, J.; Harrison, K.; Johnson, C.; Krank, J.; McAnoy, A.; Uhlson, C.; Zarini, S. Electrospray Ionization and Tandem Mass Spectrometry of Eicosanoids. Anal. Biochem. 2005, 346, 1–42.CrossRefGoogle Scholar
  23. 23.
    Lu, Y.; Hong, S.; Tjonahen, E.; Serhan, C. N. Mediator-Lipidomics: Databases and Search Algorithms for PUFA-Derived Mediators. J. Lipid Res. 2005, 46, 790–802.CrossRefGoogle Scholar
  24. 24.
    Serhan, C. N.; Gotlinger, K.; Hong, S.; Lu, Y.; Siegelman, J.; Baer, T.; Yang, R.; Colgan, S. P.; Petasis, N. A. Anti-Inflammatory Actions of Neuroprotectin D1/Protectin D1 and Its Natural Stereoisomers: Assignments of Dihydroxy-Containing Docosatrienes. J. Immunol. 2006, 176, 1848–1859.CrossRefGoogle Scholar
  25. 25.
    Hong, S.; Tjonahen, E.; Morgan, E. L.; Yu, L.; Serhan, C. N.; Rowley, A. F. Rainbow Trout (Oncorhynchus mykiss) Brain Cells Biosynthesize Novel Docosahexaenoic Acid-Derived Resolvins and Protectins—Mediator Lipidomic Analysis. Prostaglandins Other Lipid Mediat. 2005, 78, 107–116.CrossRefGoogle Scholar
  26. 26.
    Yang, R. Synthetic Study of Lipid Mediators; Doctoral thesis, University of Southern California: Los Angeles 2005, 262 pp.Google Scholar
  27. 27.
    Wheelan, P.; Zirrolli, J. A.; Murphy, R. C. Electrospray Ionization and Low Energy Tandem Mass Spectrometry of Polyhydroxy Unsaturated Fatty Acids. J. Am. Soc. Mass Spectrom. 1996, 7, 140–149.CrossRefGoogle Scholar
  28. 28.
    Yin, H.; Musiek, E. S.; Gao, L.; Porter, N. A.; Morrow, J. D. Regiochemistry of Neuroprostanes Generated from the Peroxidation of Docosahexaenoic Acid in Vitro and in Vivo. Biol. Chem. 2005, 280, 26600–26611.CrossRefGoogle Scholar
  29. 29.
    Chiang, N.; Takano, T.; Clish, C. B.; Petasis, N. A.; Tai, H. H.; Serhan, C. N. Aspirin-Triggered 15-epi-LXA4 Generation by Costimulation of Human Peripheral Blood Cell Types and in a Mouse Acute Pperitonitis Model: Development of a Specific 15-epi-LXA4 ELISA. J. Pharmacol. Exp. Ther. 1998, 287, 779–790.Google Scholar
  30. 30. Scholar
  31. 31.
    MacMillan, D. K.; Murphy, R. C. Analysis of Lipid Hydroperoxides and Long-Chain Conjugated Keto Acids by Negative Ion Electrospray Mass Spectrometry. J. Am. Soc. Mass Spectrom. 1995, 6, 1190–1201.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2007

Authors and Affiliations

  • Song Hong
    • 1
    • 2
  • Yan Lu
    • 1
    • 2
  • Rong Yang
    • 1
    • 2
  • Katherine H. Gotlinger
    • 1
    • 2
  • Nicos A. Petasis
    • 1
    • 2
    • 3
  • Charles N. Serhan
    • 1
    • 2
  1. 1.Analytical Core, Center for Experimental Therapeutics and Reperfusion Injury, Department of Anesthesiology, Perioperative and Pain MedicineBrigham and Women’s HospitalBostonUSA
  2. 2.Department of Oral Medicine, Infection and ImmunityHarvard School of Dental Medicine, Harvard Medical SchoolBostonUSA
  3. 3.Department of Chemistry and the Loker Hydrocarbon Research InstituteUniversity of Southern CaliforniaLos AngelesUSA

Personalised recommendations