Optimization of lipid extraction from the oleaginous yeasts Rhodotorula glutinis and Lipomyces kononenkoae

Abstract

The constant growing demand for vegetable oil for biodiesel and food is raising many environmental concerns about the sustainability of its production based on crops. Oleaginous yeasts show great potential to end with those concerns due to their high lipid productivity in small areas. To evaluate their productivity in lipids, an efficient and reproducible extraction process should be used. As no standard extraction process is available for the extraction of yeast lipids, an optimized extraction process is presented. In this work, the lipids extraction process for the yeasts Rhodotorula glutinis and Lipomyces kononenkoae is optimized using bead beating for cell rupture and introducing adaptations of the two most used extraction methods (Bligh and Dyer and Folch). For Rhodotorula g. the optimum extraction conditions are obtained by the Bligh and Dyer method applying 4.8 cycles of 47 s with 0.7 g of glass beads. For Lipomyces k. the optimum extraction conditions make use of the Folch method applying seven cycles of 42 s with 0.54 g of glass beads. These results reinforce the idea that, for each yeast, different extraction processes may be needed to correctly determine the lipid yield. The extraction procedure was further evaluated with less harmful solvents. Toluene was tested as a possible substitute of chloroform, and ethanol as a possible substitute of methanol. With the optimized extraction process, better results for Lipomyces k. were obtained using toluene and ethanol, while for Rhodotorula g. toluene proved to be a valid substitute of chloroform but ethanol is far less effective than methanol.

Evaluation of Downstream Processing, Extraction, and Quantification Strategies for Single Cell Oil Produced by the Oleaginous Yeasts Saitozyma podzolica DSM 27192 and Apiotrichum porosum DSM 27194

“…Analyzing the extraction techniques on whole lipid extraction, BD was more effective compared to F for S. podzolica probably because the ratio of the polar methanol is higher in BD leading to a more efficient extraction of polar lipids (Dong et al, 2016;Vasconcelos et al, 2018). For A. porosum both extraction methods showed no significant differences, presumably due to the insufficient cell disruption.…”

“…The miniaturized version of the methods of Folch (F) (Folch et al, 1957) and Bligh and Dyer (BD) (Bligh and Dyer, 1959), as adapted by Vasconcelos et al (2018), were slightly modified in this study. F was performed by combining 1.9 mL potassium phosphate buffer, 1 mL disrupted cells suspension, 9.66 mL chloroform and 4.83 mL methanol.…”

“…In literature ethanol-hexane extractions are commonly used for microalgae lipid recovery with likewise high extraction effectiveness (Grima et al, 1994;Silve et al, 2018). For the yeast Lipomyces kononenkoae, also the less polar solvent toluene revealed higher extraction yields over chloroform (Vasconcelos et al, 2018). Hence, hazardous chloroform methanol mixtures can be dispensed with less harmful ethanol hexane system without scarifying oil recovery for S. podzolica.…”

“…yeast strain, since the cell wall composition may vary considerably between species. In literature lots of attempts using various methods and different species can be found (Jin et al, 2012;Zhang et al, 2014;Meullemiestre et al, 2016;Vasconcelos et al, 2018). However, rating the methods is difficult, as reported results of different laboratories may not be comparable.…”

Lipidomics from sample preparation to data analysis: a primer

“…Additionally, frozen tissue pieces often contain some ice, which could distort the results when they are normalized based on frozen tissue weight [7]. Cells can be disrupted by a pebble mill with beads or a nitrogen cavitation bomb [7,9]. While the first method disrupts cell walls by applying shear force (the cells are beaten by specialized Lysing Matrix beads), the second method combines high pressure (of inert gaseous nitrogen) with subsequent fast adiabatic expansion, thus avoiding any shear stress to the biomolecules.…”