چكيده لاتين :
Introduction Single cell oils (SCO) are neutral lipids (triacyglycerides = TAGs) synthesized by microorganisms such as microalgae and oleaginous yeasts. Although many yeast species are able to produce SCO, only a few strains that have the capacity to store more than 20% cell mass as lipids are considered as “oleaginous”. The increasing cost of vegetable oils is turning the use of microbial lipids into a competitive alternative for the production of biodiesel fuel. Oleaginous yeast is able to produce lipid up to 60% of the dry cell weight, therefore, it can be used as the biodiesel feedstock. This study focused on the ability to synthesize lipids by the poorly characterized oleaginous yeast and Rhodosporidium diobovatum, when growing on different carbon sources under batch conditions. To the best of my knowledge, this study is the first detailed report of lipid synthesis and produced biodiesel fuel characterization by R. diobovatum grown on glucose or glycerol.
Materials and Methods The growth, lipid synthesis, and carbon use efficiency of oleaginous yeast and Rhodosporidium diobovatum, were investigated under nitrogen-limiting conditions with glucose (217 and 434 mM) and glycerol (434mM) as the carbon sources. The yeast, R. diobovatum 08-225, was obtained from the Phaff Yeast Culture Collection, at the University of California at Davis (UCDFST). To compare growth and cell mass production of R. diobovatum in GMY medium with glucose or glycerol, OD600, biomass production (dried cell weight), pH, ammonium ion (NH4+), glucose and glycerol concentrations were monitored at different time-points. Samples (4 mL) for cell growth, dry cell weight (dcw), glucose and glycerol consumption, and ammonium ion (NH4+) concentration, were taken regularly every 6 h for the initial 30 h, and then at the time-points of 48, 72, 96 and 120 hours post-inoculation (h pi). Five (5) mL samples for lipid extraction and characterization were taken every 24 h pi. In addition, the physical properties of produced biodiesel fuel were compared with international standard criteria, diesel and other biodiesel (produced from vegetable oils, animal fat and microalgae) fuel properties.
Results and Discussion In all the experiments conducted, cultures displayed a very short lag phase (less than 6 h pi) followed by an exponential growth phase from 6 to 18 h pi, and then reached stationary phase between 18 to 24 h, pi. Although NH4+ concentrations were depleted between 24 and 30 h pi, dry cell weight continued to increase, putatively due to the accumulation of neutral lipids in the cells. The glucose and glycerol cultures produced approximately equal amounts of cell mass, but the glucose cultures (434 mM) produced greater amounts of lipid (49.49 ± 0.89 % dcw) than the glucose (217 mM) and glycerol cultures (434 mM) at 120 h pi. Fatty acid profiles were similar, with minor variations, in R. diobovatum grown on glucose or glycerol. The main fatty acid species were palmitic acid (C16:0), oleic acid (C18:1), and linoleic acid (C18:2) in all experiments conducted, and these species accounted for about 90% of total fatty acid composition in the cells grown either substrate. For glucose and glycerol substrates, both were found to have good estimated cetane number values (57.8 for 217 mM glucose, 57.5 for 434 mM glucose and 58.6 for 434 mM glycerol concentrations), which would place them above the accepted value for cetane number values. However, the estimated cloud point for all the cultures and was higher than those found in many vegetable oils. This could be the result of higher levels of palmitic acid synthesized by different oleaginous yeast. Likewise, the predicted value for the cloud point of R. diobovatum cultures with glycerol were higher than R. diobovatum cultures with glucose and other oleaginous yeast species, because of the different FAMEs composition produced by each species. In general, a mathematical estimate from the FAMEs profile derived from R. diobovatum lipids expressed that, the physical properties of produced biodiesel including kinematic viscosity, cloud point, cetane number (CN), Iodine value (IV) and higher heating value (HHV) were acceptable by U.S. biodiesel and EU biodiesel standards.
Conclusion Lipid synthesis by the red oleaginous yeast, Rhodospiridium diobovatum, was investigated under nitrogen-limiting conditions using glucose or glycerol as substrates. Our results indicate that R. diobovatum is a robust strain for oil production. The study also demonstrates extensive accumulation of neutral lipids containing high percentages of oleic acid, palmitic acid, and linoleic acid by R. diobovatum, and suggests that this species could be an effective strain to biodiesel and nutritional oils. Further investigations are needed to achieve the greatest biomass with optimized lipid accumulation for commercial purposes.
