Raman Microspectroscopy of Individual Algal Cells: Sensing Unsaturation of Storage Lipids in vivo

Ota Samek,Petr Kotas,Jan Tříska,Zdeněk Pilát,Pavel Zemánek,Martin Trtílek,Alexandr Jonáš,Ladislav Nedbal

doi:10.3390/s100908635

Abstract

Algae are becoming a strategic source of fuels, food, feedstocks, and biologically active compounds. This potential has stimulated the development of innovative analytical methods focused on these microorganisms. Algal lipids are among the most promising potential products for fuels as well as for nutrition. The crucial parameter characterizing the algal lipids is the degree of unsaturation of the constituent fatty acids quantified by the iodine value. Here we demonstrate the capacity of the spatially resolved Raman microspectroscopy to determine the effective iodine value in lipid storage bodies of individual living algal cells. The Raman spectra were collected from three selected algal species immobilized in an agarose gel. Prior to immobilization, the algae were cultivated in the stationary phase inducing an overproduction of lipids. We employed the characteristic peaks in the Raman scattering spectra at 1,656 cm−1 (cis C═C stretching mode) and 1,445 cm−1 (CH2 scissoring mode) as the markers defining the ratio of unsaturated-to-saturated carbon-carbon bonds of the fatty acids in the algal lipids. These spectral features were first quantified for pure fatty acids of known iodine value. The resultant calibration curve was then used to calculate the effective iodine value of storage lipids in the living algal cells from their Raman spectra. We demonstrated that the iodine value differs significantly for the three studied algal species. Our spectroscopic estimations of the iodine value were validated using GC-MS measurements and an excellent agreement was found for the Trachydiscus minutus species. A good agreement was also found with the earlier published data on Botryococcus braunii. Thus, we propose that Raman microspectroscopy can become technique of choice in the rapidly expanding field of algal biotechnology.

Highlights

Photosynthetic organisms transform the energy of solar photons into the free energy of chemical bonds that provide for nearly the entire energy supply of Earth’s biosphere
We present Raman spectra of storage lipid bodies measured with Raman microspectroscopy in individual cells of three algal species: Botryococcus sudeticus, Chlamydomonas sp., and Trachydiscus minutus
In this paper we have demonstrated the potential of Raman microspectroscopy for the fast and spatially resolved characterization of the composition of selected intracellular regions in individual living algal cells

Summary

Introduction

Photosynthetic organisms transform the energy of solar photons into the free energy of chemical bonds that provide for nearly the entire energy supply of Earth’s biosphere. Most of the ocean photosynthesis occurs in planktonic algae. This enormous capacity of algae to transform the solar radiation into energy-rich compounds and to remove CO2 from the atmosphere justifies the current interest of science and industry. Algae are considered as a potent source of biofuels of higher generation that will not compete for land with food production and that will contribute to biological capture of atmospheric CO2 to mitigate the global climate change. In parallel to hydrogen and alcohols, the most often considered products from algae for the fuel industry are algal lipids [2]. Typical storage lipids in algae are triacylglycerols: tri-esters of glycerol with saturated or unsaturated fatty acids. We focus on the degree of fatty acid unsaturation which is the key parameter that determines the application potential for fuels or as dietary supplements or for pharmaceutical raw materials

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Results

Conclusion