Abstract
Microalgae are proposed as feedstock organisms useful for producing biofuels and coproducts. However, several limitations must be overcome before algae-based production is economically feasible. Among these is the ability to induce lipid accumulation and storage without affecting biomass yield. To overcome this barrier, a chemical genetics approach was employed in which 43,783 compounds were screened against Chlamydomonas reinhardtii, and 243 compounds were identified that increase triacylglyceride (TAG) accumulation without terminating growth. Identified compounds were classified by structural similarity, and 15 were selected for secondary analyses addressing impacts on growth fitness, photosynthetic pigments, and total cellular protein and starch concentrations. TAG accumulation was verified using gas chromatography-mass spectrometry quantification of total fatty acids, and targeted TAG and galactolipid measurements were performed using liquid chromatography-multiple reaction monitoring/mass spectrometry. These results demonstrated that TAG accumulation does not necessarily proceed at the expense of galactolipid. Untargeted metabolite profiling provided important insights into pathway shifts due to five different compound treatments and verified the anabolic state of the cells with regard to the oxidative pentose phosphate pathway, Calvin cycle, tricarboxylic acid cycle, and amino acid biosynthetic pathways. Metabolite patterns were distinct from nitrogen starvation and other abiotic stresses commonly used to induce oil accumulation in algae. The efficacy of these compounds also was demonstrated in three other algal species. These lipid-inducing compounds offer a valuable set of tools for delving into the biochemical mechanisms of lipid accumulation in algae and a direct means to improve algal oil content independent of the severe growth limitations associated with nutrient deprivation.
Highlights
Microalgae are proposed as feedstock organisms useful for producing biofuels and coproducts
In the screening experiments described here, a control-based normalization approach was employed where the data were standardized to a negative control in terms of lipid accumulation
For quality-control analysis, a Z-factor was calculated for each analysis plate to determine the separation between the positive control for lipid accumulation and the negative control to measure the signal range (Zhang et al, 1999)
Summary
Microalgae are proposed as feedstock organisms useful for producing biofuels and coproducts. Nutrient limitation is sometimes achieved during normal growth when cultures reach saturation density when nitrogen becomes limiting and triacylglyceride (TAG)-rich lipid droplets become visible and measurable (Msanne et al, 2012; Wang et al, 2012) This is normally preceded by, or commensurate with, the cessation of protein synthesis, the degradation of chlorophylls and photosynthetic enzymes including Rubisco, and a dramatic reduction in chloroplast membrane lipids (Wase et al, 2014; Allen et al, 2015). [13C]CO2 labeling provides evidence that a large majority of the TAG side chains are derived directly from photosynthetic CO2 fixation and de novo fatty acid synthesis (Allen et al, 2015, 2017) These recent advancements indicate that, membrane glycerolipid synthesis, degradation, and acyl editing are intricately involved in TAG accumulation as a consequence of the stress-induced cessation of growth, it may be possible to extricate TAG synthesis from these reactions
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