Metabolic and morphological changes of an oil accumulating trebouxiophycean alga in nitrogen-deficient conditions

Abstract

Oil-rich algae have promising potential for a next-generation biofuel feedstock. Pseudochoricystis ellipsoidea MBIC 11204, a novel unicellular green algal strain, accumulates a large amount of oil (lipids) in nitrogen-deficient (-N) conditions. Although the oil bodies are easily visualized by lipophilic staining in the cells, little is known about how oil bodies are metabolically synthesized. Clarifying the metabolic profiles in -N conditions is important to understand the physiological mechanisms of lipid accumulations and will be useful to optimize culture conditions efficiently produce industrial oil. Metabolome and lipidome profiles were obtained, respectively, using capillary electrophoresis- and liquid chromatography-mass spectrometry from P. ellipsoidea in both nitrogen-rich (+N; rapid growth) and -N conditions. Relative quantities of more than 300 metabolites were systematically compared between these two conditions. Amino acids in nitrogen assimilation and N-transporting metabolisms were decreased to 1/20 the amount, or less, in -N conditions. In lipid metabolism, the quantities of neutral lipids increased greatly in -N conditions; however, quantities of nearly all the other lipids either decreased or only changed slightly. The morphological changes in +N and -N conditions were also provided by microscopy, and we discuss their relationship to the metabolic changes. This is the first approach to understand the novel algal strain's metabolism using a combination of wide-scale metabolome analysis and morphological analysis.

Keywords: Capillary electrophoresis; Liquid chromatography; Mass spectrometry; Metabolic profile; Oil-rich algae.

Fig. 1

The morphological changes between nitrogen-rich and -deficient conditions. Gray color zone in m, n was cut off for cell volume calculation in Table 1. a, c Chlorophyll autofluorescence. e, g Nile red stain of oil bodies. b, d, f and h Nomarski interference images. i, j Iodine stain for starch granules. k, l Transmission electron microscopy. m, n Cell volume distributions. Bar length a–j 5.0 μm; k, l 1.0 μm. o oil body; s starch granule; n nucleus; white arrows large electron dense bodies (LEDBs); black arrow multi-membrane vesicle (MMV)

Fig. 2

The quantitative changes of detected metabolites mapped on the KEGG Atlas. The names of the metabolites are shown in Supplementary Table. ST2. Metabolites not included in the KEGG Atlas (185 metabolites) are not shown. Large red dot –N/+ N ≥ 20; Small red dot 20 > –N/+ N ≥ 2; Black dot 2 > –N/+ N ≥ 0.5; Small blue dot 0.5 > –N/+ N ≥ 0.05; Large blue dot –N/+ N < 0.05

Fig. 3

The metabolic profile of nitrogen assimilation and N-transporting metabolism in nitrogen-rich and -deficient conditions. Columns the quantity (μmol/l) in extracts normalized by genomic DNA content (μg). Bars SD; white bars +N condition; Gray bars –N condition. *p < 0.05 of the Mann–Whitney U-test

Fig. 4

The –N/+N ratio of monogalactosyldiacylglycerols (MGDGs; a) and triacylglycerols (TAGs; b). The numbers following the abbreviations of the lipid class indicates the total carbon chain length and total degree of unsaturation