## 3 Results
### Yeast Lipid Analysis  
All screened strains were analyzed through GC analysis, as regards the fatty acid (FA) composition of the total recovered lipids (\"Folch\" viz. C/M 2/1 extract) at the stationary growth phase of the trials (FA composition analysis performed at the end of fermentation when lactose was totally assimilated), during cultivation on either lactose-based media or on second CW (the obtained results during growth on second CW are depicted in Table 5a). SCOs of _C. curvatus_ ATCC 20509 are mainly composed of unsaturated fatty acids (UFA) (59.9% \(w/w\) of total lipids) and were particularly rich in the \({}^{\Delta 9}\)C18:1 FA (52.8% \(w/w\) of total lipids), while the saturated FAs present a value of 40.1%, being mostly composed of C16:0 (29.3% \(w/w\) of total lipids). FA content of _C. curvatus_ NRRL Y-1511 was composed of UFAs (58% \(w/w\)) and saturated FAs (SFA) (42% \(w/w\)). In contrast to ATCC 20509, NRRL Y-1511 lipids included a slightly higher value of poly-unsaturated FAs (PUFA; 9.3% vs. 5.7% \(w/w\)). A difference was observed in the FA composition between the 2 strains of _P. laurentii_. Specifically, FAs of _P. laurentii_ NRRL YB-3594 were more unsaturated (65.5% \(w/w\)) of total lipids) than them of _P. laurentii_ NRRL Y-2536 (53.5% \(w/w\)). This difference is due to the high content of \({}^{\Delta 9}\)C18:1 (56.4% vs. 46.1%) and low content of C16:0 (19.1% vs. 28.2%) FAs. In addition, _P. laurentii_ NRRL YB-3594 lipids included higher values of PUFAs (8.3% vs. 4.3% \(w/w\)) than those of _P. laurentii_ NRRL Y-2536. As far as the FA content of _D. lunsenii_ ACA-DC 5079 was concerned, \({}^{\Delta 9}\)C18:1 was the predominant FA (44.8% \(w/w\) of total lipids), followed by C16:0 (26.3% \(w/w\)). The intracellular lipids of this strain were, also, composed of \({}^{\Delta 9}\)C16:1 (12% \(w/w\)), the PUFA content was low (1.9% \(w/w\) of total lipids), but the mono-unsaturated fatty acids (MUFA) content was high (60.4% \(w/w\)). Moreover, the FA analysis concerning the fed-batch culture of _C. curvatus_ ATCC 20509, is presented in Table 5b. As the cultivation progressed, intra-cellular lipids of _C. curvatus_ ATCC 20509 were principally composed of unsaturated fatty acids (60.1% at 321 h vs. 52.4% at 70 h), rich in \({}^{\Delta 9}\)C18:1 (53.7% of total lipids), while SFAs presented some reduction (47.6% \(w/w\) at 70 h vs. 39.9% \(w/w\) at 321 h), because of C16:0 content decrease (24.3% \(w/w\) at 321 h vs. 36.5% \(w/w\) at 70 h). C18:0 content was slightly increased (14.3% \(w/w\) at 321 h vs. 10.4% \(w/w\) at 70 h). Small quantities of C14:0 (0.5% \(w/w\) of total lipids), 17:0 (0.7% \(w/w\) of total lipids) and C22:0 (0.1% \(w/w\) of total lipids) were observed (not presented). As previously, therefore, FA composition analysis demonstrated predominance of oleic and palmitic acid, constituting thus the cellular lipids of this strain as perfect materials for synthesis of \"second generation\" biodiesel, in accordance with the literature reports [33]. Total cellular lipids (\"Folch\" viz. C/M 2/1 extract) were analyzed through TLC analysis (Figure 5a). The strains _C. curvatus_ ATCC 20509, _C. curvatus_ NRRL Y-1511, _P. laurentii_ NRRL Y-2536, and _P. laurentii_ NRRL YB-3594 accumulated lipids, mainly in the form of triacylglycerglycerols, although the quantities of storage lipids were not high (in all cases K\({}_{\rm L/X}\) values were \(\leq\)0.16 g/g; see Table 4). Other lipid compounds non-identified (i.e., monoacylglycerolglycerols, diacylglycerols, phospholipids, ergosterol, and steryl-esters, the latter one presumably being revealed at the top of the TLC plate; see: [49]) were also detected in non-negligible quantities. Another interesting result was associated with the strain  
Figure 4: Representation of lipid accumulation progress of _Cryptococcus curvatus_ ATCC 20509 fed-batch cultivation in treated second CW supplemented with added CW-derived lactose.