Ultra-centrifugation force in adaptive evolution changes the cell structure of oleaginous yeast Trichosporon cutaneum into a favorable space for lipid accumulation.

Microbial lipid production from lignocellulose biomass provides an essential option for sustainable and carbon-neutral supply of future aviation fuels, biodiesel, as well as various food and nutrition products. Oleaginous yeast is the major microbial cell factory but its lipid-producing performance is far below the requirements of industrial application. Here we show an ultra-centrifugation fractionation in adaptive evolution (UCF) of Trichosporon cutaneum based on the minor cell density difference. The lightest cells with the maximum intracellular lipid content were isolated by ultra-centrifugation fractionation in the long-term adaptive evolution. Significant changes occurred in the cell morphology with a fragile cell wall wrapping and enlarged intracellular space (two orders of magnitude increase in cell size). Complete and coordinate assimilations of all nonglucose sugars derived from lignocellulose were triggered and fluxed into lipid synthesis. Genome mutations and significant transcriptional regulations of the genes responsible for cell structure were identified and experimentally confirmed. The obtained T. cutaneum MP11 cells achieved a high lipid production of wheat straw, approximately five-fold greater than that of the parental cells. The study provided an effective method for screening the high lipid-containing oleaginous yeast cells as well as the intracellular products accumulating cells in general.

[Simultaneous qualitative and quantitative determination of seven anticoagulant rodenticides in whole blood and urine samples using on-line solid phase extraction with liquid chromatography-linear ion trap mass spectrometry].

A new and sensitive analytical method has been developed for the simultaneous determination of seven anticoagulant rodenticides in whole blood and urine samples by liquid chromatography-linear ion trap mass spectrometry (LC-LIT/MS) with on-line solid phase extraction (on-line SPE). The samples were treated with acetonitrile, followed by dilution, centrifugation, and filtration. The resulting solution was injected into the LC system directly and processed by on-line SPE column for enrichment and purification. Separation was performed on a C18 column with mixed mobile phases of methanol and 0.02 mol/L ammonium acetate aqueous solution for gradient elution. The analytes were detected by the mass spectrometer with electrospray ionization (ESI) in negative mode. MS2 full scan signals of the target parent ions within the locked retention time window were recorded. Self-built database searching was performed for qualitative confirmation, and MS2 fragment ions with high sensitivity and specificity were selected for quantification. Simultaneous qualitative and quantitative analyses of the seven rodenticides were achieved in this way. Good linearities were obtained within the investigated mass concentration ranges of the seven rodenticides, with r2 ≥ 0.9958 in blood and r2 ≥ 0.9946 in urine. The LODs varied from 0.02 ng/mL to 1.00 ng/mL, and the LOQs varied from 0.10 ng/mL to 4.00 ng/mL. The recoveries at three spiked levels in blood and urine samples ranged from 81.0% to 113.9%, with RSDs of 0.1%-6.2% (n = 6). The developed method is simple, sensitive, and can be used for the rapid detection and accurate quantification of the seven anticoagulant rodenticides in whole blood and urine samples.