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Triacylglycerol accumulation and profiling in the model diatoms Thalassiosira pseudonana and Phaeodactylum tricornutum (Baccilariophyceae) during starvation

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Abstract

Although substantial economic barriers exist, marine diatoms such as Thalassiosira pseudonana and Phaeodactylum tricornutum hold promise as feedstock for biodiesel because of their ability to manufacture and store triacylglycerols (TAGs). The recent sequencing of these two marine diatom genomes by the United States Department of Energy Joint Genome Institute and the development of improved systems for genetic manipulation should allow a more systematic approach to understanding and maximizing TAG production. However, in order to best utilize these genomes and genetic tools, we must first gain a deeper understanding of the nutrient-mediated regulation of TAG anabolism. By determining both the yield and molecular species distribution of TAGs we will, in the future, be able to fully characterize the effects of genetic manipulation. Here, we lay the groundwork for understanding TAG production in T. pseudonana and P. tricornutum, as a function of nitrate and silicate depletion. Diatoms were starved of either nitrate or silicate, and TAGs were extracted with hexane from lyophilized samples taken at various time intervals following starvation. The timing of TAG production and the relative abundance of TAGs were estimated by fluorescence spectroscopy using Nile red and the total yield per biomass determined by gravimetric assay. TAGs were analyzed using thin layer chromatography, gas chromatography–mass spectrometry, and electrospray ionization mass spectrometry to identify the major TAG species produced during the growth curve. Under our conditions, the TAG yield from T. pseudonana is about 14–18% of total dry weight. The TAG yield from P. tricornutum is about 14% of total dry weight. Silicate-starved T. pseudonana accumulated an average of 24% more TAGs than those starved for nitrate; however, the chemotypes of the TAGs produced were generally similar regardless of the starvation condition employed.

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Acknowledgments

We would like to acknowledge Dr. Kenneth Sale for his careful reading of the manuscript. This work was supported by the Laboratory Directed Research and Development program at Sandia National Laboratories, which is a multi-program laboratory operated by Sandia Corp., a Lockheed Martin company, for the United States Department of Energy under contract no. DE-AC04-94AL85000.

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Author notes

  1. Sara Gaucher

    Present address: Amyris Biotechnologies, Inc, Emeryville, CA, 94608, USA

Authors and Affiliations

  1. Biosystems Research Department, Sandia National Laboratories, Livermore, CA, 94550, USA

    Eizadora T. Yu, Pamela D. Lane, Sara Gaucher & Todd W. Lane

  2. Energy Systems Department, Sandia National Laboratories, Livermore, CA, 94550, USA

    Frank J. Zendejas & Blake A. Simmons

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  1. Eizadora T. Yu
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  2. Frank J. Zendejas
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Correspondence to Todd W. Lane.

Electronic supplementary material

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Supplementary Figure 1

Tandem mass spectrum of lithiated TAG species (50:5) at m/z 831.7. Fragment ions reflect the loss of each fatty acid substituent and are observed at m/z values corresponding to [M+Li-(RnCO2H)]* and [M+Li-(RnCO2Li)] (peaks are bracketed in the figure). Product ions formed reveals two possible fatty acid compositions (14:0/16:0/20:5 and 16:0/16:1/18:4) that are consistent with the molecular composition of TAG 50:5. The relative abundances of the product ions show that the predominant FA composition for TAG 50:5 is 16:0/16:1/18:4 (GIF 108 kb)

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Supplementary Table 1

Full list of TAG molecular weight identified in hexane extracts from T. pseudonana and P. tricornutum during growth cycle (DOC 310 kb)

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Yu, E.T., Zendejas, F.J., Lane, P.D. et al. Triacylglycerol accumulation and profiling in the model diatoms Thalassiosira pseudonana and Phaeodactylum tricornutum (Baccilariophyceae) during starvation. J Appl Phycol 21, 669–681 (2009). https://doi.org/10.1007/s10811-008-9400-y

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