C/N ratio and carbon source-dependent lipid production profiling in Rhodotorula toruloides

Abstract

Microbial oils are lipids produced by oleaginous microorganisms, which can be used as a potential feedstock for oleochemical production. The oleaginous yeast Rhodotorula toruloides can co-produce microbial oils and high-value compounds from lowcost substrates, such as xylose and acetic acid (from hemicellulosic hydrolysates) and raw glycerol (a byproduct of biodiesel production). One step towards economic viability is identifying the best conditions for lipid production, primarily the most suitable carbon-to-nitrogen ratio (C/N). Here, we aimed to identify the best conditions and cultivation mode for lipid production by R. toruloides using various low-cost substrates and a range of C/N ratios (60, 80, 100, and 120). Turbidostat mode was used to achieve a steady state at the maximal specific growth rate and to avoid continuously changing environmental conditions (i.e., C/N ratio) that inherently occur in batch mode. Regardless of the carbon source, higher C/N ratios increased lipid yields (up to 60% on xylose at a C/N of 120) but decreased the specific growth rate. Growth on glycerol resulted in the highest specific growth and lipid production (0.085 g lipids/gDW*h) rates at C/Ns between 60 and 100. We went on to study lipid production using glycerol in both batch and fed-batch modes, which resulted in lower specific lipid production rates compared with turbisdostat, however, fed batch is superior in terms of biomass production and lipid titers. By combining the data we obtained in these experiments with a genome-scale metabolic model of R. toruloides, we identified targets for improvements in lipid production that could be carried out either by metabolic engineering or process optimization. Keywords Rhodotorula toruloides. Microbial oil. Biorefineries. Genome-scale metabolic model. Flux balance analysis. Carbon to nitrogen ratio. Alternative substrates Helberth Júnnior Santos Lopes and Nemailla Bonturi contributed equally to this work.

Biomass, lipid accumulation kinetics, and the transcriptome of heterotrophic oleaginous microalga Tetradesmus bernardii under different carbon and nitrogen sources

“…Lipid accumulation in oleaginous microalgae depends on diverse factors, such as the strain in use, nutritional imbalances of medium components, the available carbon (C) and nitrogen (N) source, trophic mode (autotrophy, mixotrophy, or heterotrophy), cultivation conditions, and culture time [ 1 , 11 ]. Multiple strategies have been proposed for lipid production improvement in various microorganisms.…”

Physiological Traits of Dihomo-γ-Linolenic Acid Production of the Engineered Aspergillus oryzae by Comparing Mathematical Models

“…In our work, the carbon-limited condition was elaborated at the lipid-accumulating phase of the engineered strain, although such condition could not boost the lipid content over 20% of dry biomass, as known for the phenotype of oleaginous strains (Ochsenreither et al, 2016;Gientka et al, 2017). Generally, the cultivation at high C/N ratio has been supposed to be a strategy for enhancing the lipid content in biomass (Lopes et al, 2020), but the PUFAs proportionally decreased in total lipid, pertaining a poor quality of PUFA-containing oils. For the high C/N ratio condition, the residual glucose usually remains in the biowaste derived from the production process in addition to the drawback in high medium cost that might be attributed to the overall operation cost for PUFA production.…”

Biomass, lipid accumulation kinetics and the transcriptome of heterotrophic oleaginous microalga Tetradesmus bernardii under different carbon and nitrogen regime

“…Fatty acid biosynthesis is often limited by the supply of NADPH, as the elongation of fatty acids during their biosynthesis by two carbon units requires two NADPH molecules [11]. Oxidative pentose phosphate pathway provides NADPH to the cell through the oxidation of glucose.…”