, Volume 9, Issue 5, pp 949–959 | Cite as

Lipidomic profiling and discovery of lipid biomarkers in Stephanodiscus sp. under cold stress

  • Deying Chen
  • Xiaojun Yan
  • Jilin Xu
  • Xiaolin Su
  • Lanjuan Li
Original Article


Changes in membrane lipid composition play multiple roles in plant adaptation and survival in the face of chilling and freezing damage. The ultra-performance liquid chromatography/quadrupole-TOF-MS (UPLC/Q-TOF-MS)-based approach was developed for investigating the lipid changes during cold exposure in Stephanodiscus sp. followed by multivariate statistical analysis including principal components analysis, partial least squares discriminant analysis and orthogonal projection on latent structure discriminant analysis for data classification and potential biomarkers selection. The analysis demonstrated dramatic lipid alterations take place in both extraplastidic and plastidic membranes. Thirty-eight lipid molecules were selected and identified as putative biomarkers, including chlorophyll a degradation products, triacylglycerol, phosphatidylcholine, phosphatidylglycerol, sulfo-quinovosyldiacylglycerol, monogalactosyldiacylglyceroll, lyso-phosphatidylglycerol, lyso-phosphatidylcholine, lyso-monogalactosyldiacylglycerol and lyso-sulfoquinovosyldiacylglycerol. These metabolites have been shown previously to function in energy storage, membrane stability and photosynthesis efficiency. This study is the first one using UPLC/Q-TOF-MS-based lipidomic profiling with multivariate statistical analysis to explore the lipidomic changes of microalgae in response to stress conditions, which promotes better understanding of their physiology and ecology.


Stephanodiscus sp. Cold stress Lipidomics UPLC-Q-TOF-MS 



Ultra-performance liquid chromatography


Quadrupole time-of-flight mass spectrometry


Tandem mass spectrometry


Electrospray ionization


Principal component analysis


Projections to latent structures discriminant analysis


Orthogonal projections to latent structures discriminant analysis






















Endoplasmic reticulum



This research was supported by the National Natural Science Foundation of China (31172448), Zhejiang Natural Science Foundation, China (Y3100534 and Z3100565), Ningbo Science and Technology Research Projects, China (2011C11003), Zhejiang marine biotechnology innovation team, China (2012R10029), Ningbo Marine Algae Biotechnology Team, China (2011B81007), and partly sponsored by K. C. Wong Magna Fund in Ningbo University.

Supplementary material

11306_2013_515_MOESM1_ESM.tif (148 kb)
S-Fig. 1 PCA scores plot in negative ion scan mode for the first two components of Stephanodiscus sp. samples were harvested on time point 0, 4, 12 and 24 h, successively (TIFF 147 kb)
11306_2013_515_MOESM2_ESM.tif (158 kb)
S-Fig. 2 PLS-DA scores plot in negative ion scan mode for the first two components of Stephanodiscus sp. samples were harvested on time point 0, 4, 12 and 24 h, successively. (TIFF 157 kb)
11306_2013_515_MOESM3_ESM.tif (217 kb)
S-Fig. 3 validation plot of PLS-DA analysis on Stephanodiscus sp., (A) in positive ion scan mode; (B) in negative ion scan mode (TIFF 216 kb)
11306_2013_515_MOESM4_ESM.tif (194 kb)
S-Fig. 4 Scores scatter plot of OPLS-DA (A) and validation plot (B) of OPLS-DA analysis on Stephanodiscus sp. in positive ion scan mode, which compared group 2 versus group 3 (TIFF 194 kb)
11306_2013_515_MOESM5_ESM.tif (173 kb)
S-Fig. 5 Scores scatter plot of OPLS-DA (A) and validation plot (B) of OPLS-DA analysis on Stephanodiscus sp. in negative ion scan mode, which compared group 2 versus group 3 (TIFF 173 kb)
11306_2013_515_MOESM6_ESM.tif (188 kb)
S-Fig. 6 S-plot used in our biomarkers selection compared with group 2 versus group 1. The variables marked (red) are the metabolites selected as potential biomarkers. (A) in the positive ion mode; (B) in the negative ion mode (TIFF 188 kb)
11306_2013_515_MOESM7_ESM.tif (191 kb)
S-Fig. 7 S-plot used in our biomarkers selection compared with group 3 versus group 2. The variables marked (red) are the metabolites selected as potential biomarkers. (A) in the positive ion mode; (B) in the negative ion mode (TIFF 190 kb)
11306_2013_515_MOESM8_ESM.tif (34 kb)
S-Fig. 8 A: Growth of the culture on different days under normal condition; B: Comparison of growth of Stephanodiscus. sp. in normal condition (upper line, ∆) and cold stress condition (lower line,□) in 24 h (TIFF 34 kb)
11306_2013_515_MOESM9_ESM.pptx (157 kb)
Supplementary-PPT The variation trend plot of some representative lipid markers during the course of the cold treatment in the whole four sampling groups, which were harvested on time points 0, 4, 12 and 24 h (PPTX 157 kb)


  1. Bligh, E. G., & Dyer, W. J. (1959). A rapid method of total lipid extraction and purification. The Journal of Physiology and Biochemistry, 37, 911–917.CrossRefGoogle Scholar
  2. Dörmann, P., & Benning, C. (2002). Galactolipids rule in seed plants. Trends in Plant Science, 7, 112–118.PubMedCrossRefGoogle Scholar
  3. Dormann, P., Hoffmann-Benning, S., Balbo, I., & Benning, C. (1995). Isolation and characterization of an Arabidopsis mutant deficient in the thylakoid lipid digalactosyl diacylglycerol. Plant Cell, 7, 1801–1810.PubMedGoogle Scholar
  4. Durrett, T. P., Benning, C., & Ohlrogge, J. (2008). Plant triacylglycerols as feedstocks for the production of biofuels. The Plant Journal, 54, 593–607.PubMedCrossRefGoogle Scholar
  5. Garnier, J., Wu, B., Maroc, J., Guyon, D., & Trémolières, A. (1990). Restoration of both an oligomeric form of the light-harvesting antenna CP II and of a fluorescence state II-state I transition by Δ3-trans-hexadecenoic acid-containing phosphatidylglycerol, in cells of a mutant of Chlamydomonas reinhardtii. Biochimica et Biophysica Acta, 1020, 153–162.CrossRefGoogle Scholar
  6. Gounaris, K., & Barber, J. (1983). Monogalactosyldiacylglycerol: The most abundant polar lipid in nature. Trends in Biochemical Sciences, 8, 378–381.CrossRefGoogle Scholar
  7. Guan, X. L., He, X., Ong, W. Y., Yeo, W. K., Shui, G., & Wenk, M. R. (2006). Non-targeted profiling of lipids during kainate-induced neuronal injury. FASEB Journal, 20, 1152–1161.PubMedCrossRefGoogle Scholar
  8. Han, X., & Gross, R. W. (1994). Electrospray ionization mass spectroscopic analysis of human erythrocyte plasma membrane phospholipids. Proceedings of the National Academy of Sciences of the United States of America, 91, 10635–10639.PubMedCrossRefGoogle Scholar
  9. Han, X. L., & Gross, R. W. (2005). Shotgun lipidomics: Electrospray ionization mass spectrometric analysis and quantitation of cellular lipidomes directly from crude extracts of biological samples. Mass Spectrometry Reviews, 24, 367–412.PubMedCrossRefGoogle Scholar
  10. Han, X., Yang, K., Yang, J., Cheng, H., & Gross, R. W. (2006). Shotgun lipidomics of cardiolipin molecular species in lipid extracts of biological samples. Journal of Lipid Research, 47, 864–879.PubMedCrossRefGoogle Scholar
  11. Härtel, H., Dörmann, P., & Benning, C. (2001). Galactolipids not associated with the photosynthetic apparatus in phosphate-deprived plants. Journal of Photochemistry and Photobiology B: Biology, 61, 46–51.CrossRefGoogle Scholar
  12. Härtel, H., Lokstein, H., Dörmann, P., Trethewey, R. N., & Benning, C. (1998). Photosynthetic light utilization and xanthophylls cycle activity in the galactolipid deficient dgd1 mutant of Arabidopsis thaliana. Plant Physiology and Biochemistry, 36, 407–417.CrossRefGoogle Scholar
  13. Heaton, J. W., Yada, R. Y., & Marangoni, A. G. (1996). Discoloration of coleslaw is caused by chlorophyll degradation. Journal of Agriculture and Food Chemistry, 44, 395–398.CrossRefGoogle Scholar
  14. Heinz, E., & Roughan, G. (1983). Similarities and differences in lipid metabolism of chloroplasts isolated from 18:3 and 16:3 plants. Plant Physiology, 72, 273–279.PubMedCrossRefGoogle Scholar
  15. Hicks, G. R., Hironaka, C. M., Dauvillee, D., Funke, R. P., D’Hulst, C., Waffenschmidt, S., et al. (2001). When simpler is better. Unicellular green algae for discovering new genes and functions in carbohydrate metabolism. Plant Physiology, 127, 1334–1338.PubMedCrossRefGoogle Scholar
  16. Hobe, S., Prytulla, S., Kuhlbrandt, W., & Paulsen, H. (1994). Trimerization and crystallization of reconstituted light-harvesting chlorophyll a/b complex. EMBO Journal, 13, 3423–3429.PubMedGoogle Scholar
  17. Ivanova, P. T., Milne, S. B., Forrester, J. S., & Brown, H. A. (2004). LIPID arrays: New tools in the understanding of membrane dynamics and lipid signaling. Molecular Interventions, 4, 86–96.PubMedCrossRefGoogle Scholar
  18. Kagan, V. E., Tyurin, V. A., Jiang, J., Tyurina, Y. Y., Ritov, V. B., Amoscato, A. A., et al. (2005). Cytochrome c acts as a cardiolipin oxygenase required for release of proapoptotic factors. Nature Chemical Biology, 1, 223–232.PubMedCrossRefGoogle Scholar
  19. Köfeler, H. C., Fauland, A., Rechberger, G. N., & Trötzmüller, M. (2012). Mass spectrometry based lipidomics: An overview of technological platforms. Metabolites, 2, 19–38.CrossRefGoogle Scholar
  20. Kruse, O., Hankamer, B., Konczak, C., Gerle, C., Morris, E., Radunz, A., et al. (2000). Phosphatidylglycerol is involved in the dimerization of photosystem II. Journal of Biological Chemistry, 275, 6509–6514.PubMedCrossRefGoogle Scholar
  21. Lee, A. G. (2000). Membrane lipids: It’s only a phase. Current Biology, 10, 377–380.CrossRefGoogle Scholar
  22. Lee, S. H., Williams, M. V., DuBois, R. N., & Blair, I. A. (2003). Targeted lipidomics using electron capture atmospheric pressure chemical ionization mass spectrometry. Rapid Communications in Mass Spectrometry, 17, 2168–2176.PubMedCrossRefGoogle Scholar
  23. Li, W. Q., Li, M. Y., Zhang, W. H., Welti, R., & Wang, X. M. (2004). The plasma membrane-bound phospholipase Dδ enhances freezing tolerance in Arabidopsis thaliana. Nature Biotechnology, 22, 427–433.PubMedCrossRefGoogle Scholar
  24. Li, W., Wang, R., Li, M., Li, L., Wang, C., Welti, R., et al. (2008). Differential degradation of extraplastidic and plastidic lipids during freezing and post-freezing recovery in Arabidopsis thaliana. Journal of Biological Chemistry, 283, 461–468.PubMedCrossRefGoogle Scholar
  25. Li-Beisson, Y., Shorrosh, B., Beisson, F., Andersson, M. X., Arondel, V., Bates, P. D., et al. (2010). The arabidopsis book. Rockville: American Society of Plant Biologists.Google Scholar
  26. Liu, B., & Benning, C. (2012). Lipid metabolism in microalgae distinguishes itself. Current Opinion in Biotechnology,. doi: 1016/j.copbio.2012.08.008.Google Scholar
  27. Lu, N., Wei, D., Chen, F., & Yang, S. T. (2012). Lipidomic profiling and discovery of lipid biomarkers in snow alga Chlamydomonas nivalis under salt stress. European Journal of Lipid Science and Technology, 114, 253–265.CrossRefGoogle Scholar
  28. Lyons, J. M., & Asmundson, C. M. (1965). Solidification of unsaturated/saturated fatty acid mixtures and its relationship to chilling sensitivity in plants. Journal of the American Oil Chemists Society, 42, 1056–1058.PubMedCrossRefGoogle Scholar
  29. Lyons, J. M., Wheaton, T. A., & Pratt, H. K. (1964). Relationship between the physical nature of mitochondrial membranes and chilling sensitivity in plants. Plant Physiology, 39, 262–268.PubMedCrossRefGoogle Scholar
  30. McDonnel, A., & Staehelin, L. A. (1980). Adhesion between liposomes mediated by the chlorophyll a/b light-harvesting complex isolated from chloroplast membranes. Journal of Cell Biology, 84, 40–56.PubMedCrossRefGoogle Scholar
  31. Milne, S. B., Ivanova, P. T., DeCamp, D., Hsueh, R. C., & Brown, H. A. (2005). A targeted mass spectrometric analysis of phosphatidylinositol phosphate species. Journal of Lipid Research, 46, 1796–1802.PubMedCrossRefGoogle Scholar
  32. Milne, S., Ivanova, P., Forrester, J., & Alex Brown, H. (2006). Lipidomics: An analysis of cellular lipids by ESI-MS. Methods, 39, 92–103.PubMedCrossRefGoogle Scholar
  33. Mongrand, S., Bessoule, J. J., Cabantous, F., & Cassagne, C. (1998). The C-16:3/C-18: 3 fatty acid balance in photosynthetic tissues from 468 plant species. Phytochemistry, 49, 1049–1064.CrossRefGoogle Scholar
  34. Nussberger, S., Dörr, K., Wang, D. N., & Kühlbrandt, W. (1993). Lipid–protein interactions in crystals of plant light-harvesting complex. Journal of Molecular Biology, 234, 347–356.PubMedCrossRefGoogle Scholar
  35. Quehenberger, O., Armando, A. M., Brown, A. H., Milne, S. B., Myers, D. S., Merrill, A. H., et al. (2010). Lipidomics reveals a remarkable diversity of lipids in human plasma. Journal of Lipid Research, 51(11), 3299–3305.PubMedCrossRefGoogle Scholar
  36. Schuhmann, K., Almeida, R., Baumert, M., Herzog, R., Bornstein, S. R., & Shevchenko, A. (2012). Shotgun lipidomics on a LTQ Orbitrap mass spectrometer by successive switching between acquisition polarity modes. Journal of Mass Spectrometry, 47, 96–104.PubMedCrossRefGoogle Scholar
  37. Schwudke, D., Schuhmann, K., Herzog, R., Bornstein, S. R., & Shevchenko, A. (2011). Shotgun lipidomics on high resolution mass spectrometers. Cold Spring Harbor Perspectives in Biology, 3(9), a004614. doi: 10.1101/cshperspect.a004614.
  38. Siaut, M., Cuine, S., Cagnon, C., Fessler, B., Nguyen, M., Carrier, P., et al. (2011). Oil accumulation in the model green alga Chlamydomonas reinhardtii: Characterization, variability between common laboratory strains and relationship with starch reserves. BMC Biotechnology,. doi: 10.1186/1472-6750-11-7.PubMedGoogle Scholar
  39. Simidjiev, I., Stoylova, S., Amenitsch, H., Jávorfi, T., Mustárdy, L., Laggner, P., et al. (2000). Self-assembly of large, ordered lamellae from non-bilayer lipids and integral membrane proteins in vitro. Proceedings of the National Academy of Sciences of the United States of America, 97, 1473–1476.PubMedCrossRefGoogle Scholar
  40. Strand, A. (2004). Plastid-to-nucleus signalling. Current Opinion in Plant Biology, 7, 621–625.PubMedCrossRefGoogle Scholar
  41. Su, X. L., Xu, J. L., Yan, X. J., Zhao, P., Chen, J. J., Zhou, C. X., et al. (2012). Lipidomic changes during different growth stages of Nitzschia closterium f. minutissima. Metabolomics,. doi: 10.1007/s11306-012-0445-1.Google Scholar
  42. Taguchi, R., Houjou, T., Nakanishi, H., Yamazaki, T., Ishida, M., Imagawa, M., et al. (2005). Focused lipidomics by tandem mass spectrometry. Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, 823, 26–36.PubMedCrossRefGoogle Scholar
  43. Taylor, R., & Fletcher, R. L. (1999). Cryopreservation of eukaryotic algae—A review of methodologies. Journal of Applied Phycology, 10, 481–501.CrossRefGoogle Scholar
  44. Uemura, M., Joseph, R. A., & Steponkus, P. L. (1995). Cold acclimation of Arabidopsis thaliana (effect on plasma membrane lipid composition and freeze-induced lesions). Plant Physiology, 109, 15–30.PubMedGoogle Scholar
  45. Wang, X. M., Li, W. Q., Li, M. Y., & Welti, R. (2006). Profiling lipid changes in plant response to low temperatures. Physiologia Plantarum, 126, 90–96.CrossRefGoogle Scholar
  46. Watson, A. D. (2006). Thematic review series: Systems biology approaches to metabolic and cardiovascular disorders. Lipidomics: A global approach to lipid analysis in biological systems. Journal of Lipid Research, 47, 2101–2111.PubMedCrossRefGoogle Scholar
  47. Welti, R., Li, W., Li, M., Sang, Y., Biesiada, H., Zhou, H. E., et al. (2002). Profiling membrane lipids in plant stress responses. Role of phospholipase D alpha in freezing-induced lipid changes in Arabidopsis. Journal of Biological Chemistry, 277, 31994–32002.PubMedCrossRefGoogle Scholar
  48. Welti, R., & Wang, X. M. (2004). Lipid species profiling: A high-throughput approach to identify lipid compositional changes and determine the function of genes involved in lipid metabolism and signaling. Current Opinion in Plant Biology, 7, 337–344.PubMedCrossRefGoogle Scholar
  49. Wenk, M. R. (2005). The emerging field of lipidomics. Nature Reviews Drug Discovery, 4, 594–610.PubMedCrossRefGoogle Scholar
  50. Whitfield, P. D., Noble, P. J. M., Major, H., Beynon, R. J., et al. (2005). Metabolomics as a diagnostic tool for hepatology: Validation in a naturally occurring canine model. Metabolomics, 1, 215–225.CrossRefGoogle Scholar
  51. Wiklund, S., Johansson, E., Sjöström, L., Mellerowicz, E. J., Edlund, U., Shockcor, J. P., et al. (2008). Visualization of GC/TOF-MS-based metobolomics data for identification of biochemically interesting compounds using OPLS class models. Analytical Chemistry, 80, 115–122.PubMedCrossRefGoogle Scholar
  52. Xu, J. L., Chen, D. Y., Yan, X. J., Chen, J. J., & Zhou, C. X. (2010). Global characterization of the photosynthetic glycerolipids from a marine diatom Stephanodiscus sp. by ultra performance liquid chromatography coupled with electrospray ionization-quadrupole-time of flight mass spectrometry. Analytica Chimica Acta, 663, 60–68.PubMedCrossRefGoogle Scholar
  53. Yan, X. J., Xu, J. L., Chen, J. J., Chen, D. Y., Xu, S. L., Luo, Q. J., et al. (2012). Lipidomics focusing on serum polar lipids reveals species dependent stress resistance of fish under tropical storm. Metabolomics, 8, 299–309.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Deying Chen
    • 1
    • 2
  • Xiaojun Yan
    • 2
  • Jilin Xu
    • 2
  • Xiaolin Su
    • 1
    • 2
  • Lanjuan Li
    • 1
  1. 1.State Key Laboratory for Diagnosis and Treatment of Infectious Diseases1st Affiliated Hospital, College of Medicine, Zhejiang UniversityHangzhouChina
  2. 2.Key Laboratory of Applied Marine BiotechnologyNingbo University, Ministry of EducationNingboChina

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