## 4 Discussion
Microbial lipids synthesized by _Y. lipolytica_ W29 in two-stage batch cultures were mainly composed of unsaturated fatty acids (>78%), oleic acid being the predominant fatty acid. The high resemblance between the composition of _Y. lipolytica_ lipids and vegetable oils makes these bio-oils a potential substitute for vegetable oils as feedstock for biodiesel production. Moreover, the odd-chain fatty acids (heptadecenoic and margaric acids) synthesized in these conditions will improve the fuel properties of biodiesel produced from _Y. lipolytica_ lipids [23]. Other _Y. lipolytica_ strains, growing in VFAs [8; 9; 24], glycerol [25], or in hydrophobic substrates--waste cooking oils [14] and lard [26]--accumulated preferentiallyunsaturated fatty acids (oleic and linoleic acids). This behavior suggests that biogenic features of _Y. lipolytica_ prevail over the type of substrate. The potential of biodiesel to replace fossil-based fuel is evaluated by several parameters, namely kinematic viscosity, density, cetane number, iodine value, and oxidation stability, among others. These properties are affected by the fatty acid composition of oils used in the transesterification reactions, namely chain length, unsaturation degree, and number and position of double bonds. High saturated fatty acids, for example, avoid the auto-oxidation of biofuel and increase its shelf life, while the amount of unsaturated fatty acids has an effect on biodiesel's cold flow plugging properties [16]. The cetane number of diesel fuel is an index of the ignition quality (ignition delay characteristics) and defines the quality of the combustion process in a diesel engine, affecting the stability, noise, and CO emissions. A low cetane number (long ignition delay, particularly in cold environments) implies that the majority of injected biodiesel burns explosively, resulting in loud diesel combustion noise, white smoke, and hydrocarbon emissions. By contrast, a high cetane number indicates a shorter ignition delay and a more uniform and complete burning of injected fuel, leading to better exhaust air quality. However, a cetane number higher than 65 results in excessive heating of the injector and in a reduction of biofuel efficiency [16; 27]. The estimated cetane number for biodiesel produced from _Y. lipolytica_ W29 lipids (56) meets this criterion and is comparable to the cetane number of biodiesel obtained from different vegetable oils [27], and bio- oils produced by _Y. lipolytica_ NCYC 2904 [9]. The high heating value of biodiesel describes the heating energy released during the combustion, i.e., the efficiency of the biofuel. The value predicted herein (38 MJ-kg\({}^{-1}\)) is in the range of others found in the literature for biodiesel produced from pure and used vegetable oils, and bio- oils of _Y. lipolytica_ strains grown in wheat straw hydrolysate [28; 29] and VFAs [9]. The cold filter plugging point defines the lowest temperature, expressed in degrees Celsius (\({}^{\circ}\)C), at which a given volume of biodiesel easily flows through a standardized filtration device in a specific time when cooled under certain conditions [16]. Though there are no specific recommendations for the cold filter plugging point, the lower this value is, the better the cold flow properties of the biofuel. This is especially important in cold temperate countries since biodiesel with a high cold filter plugging point will clog the fuel line and filters more easily. The value of the cold filter plugging point estimated in this work (\(-0.6\)\({}^{\circ}\)C) is lower than the value obtained using vegetable oils [27] and bio- oils of _Trichosporon_ sp. [30], and higher than those obtained with lipids of other _Y. lipolytica_ strains [9; 16]. The degree of unsaturation and number of double bonds in fatty acids of oils has negative effects on the oxidative stability and biodiesel shelf life, since oxidatively unstable biodiesel leads to the formation of gums and sedimentation, decreasing the engine performance [16].