# Unconventional Extraction Methods of Oleaginous Yeast Cell Pretreatment and Disruption
## 4 Discussion
[37], PEF was used as a method of extracting proteins from waste brewing yeast. By applying 2% yeast biomass solution and an electric voltage of 10 kV, the authors observed 2.788% protein extraction efficiency. The results obtained within the current work allowed the extraction of almost 0.5% of protein from yeast cells using PEF but the physiology and cell wall structure of the yeast itself may be a cause of the differentiated results. US also cause cell wall disintegration, contributing to the release of proteins and lipids from cells. The mechanisms of ultrasound-assisted extraction are based on fragmentation, erosion, sonoporation, and detexturation [38]. The mass transfer to the liquid phase is improved due to the collapse of cavitation bubbles, and moreover, this process is characterized primarily by a much shorter time and lower energy consumption compared to conventional extraction methods [39]. Some authors [26] stated that the use of US causes a high degree of loosening of the yeast cell wall and membrane, which is also very useful during the extraction of microbial oil. In the current study, the US did not have an impact on the improvement of lipid extraction yield but proved its use in protein release from yeast cells. On the other hand, application of US for _Trichosporon_ sp. Biomass proved to be the most efficient method for lipid extraction with the obtained lipid yield of 43% (Table 1). The achieved results were higher than those obtained with Soxhlet (30%) and Folch (36%) methods [40]. [PERSON] and [PERSON] [41] compared lipid recovery with the use of different biomass concentrations of oleaginous yeast _Naganishia liquefaciens_ NITTS2, the chloroform: methanol ratio, and ultrasound power density. The recovery of 99.3 \(\pm\) 0.1% was achieved after 30 min at 30 \({}^{\circ}\)C when 60 g/L of biomass, a solvent ratio of 1:1, and 0.6 W/mL of US power were used. Sonication is a valuable method for permeabilizing the yeast cell wall and releasing intracellular components like proteins. The biomass of _Saccharomyces cerevisiae_ 2200 was subjected to the US treatment and the content of protein released from the cells reached 320 mg/g of the dry yeasts [26]. The third unconventional method used in the current study was HPP. According to the obtained results, it can be seen that the use of HPP as a permeabilization method did not improve the lipid extraction yield, hence it can be stated that such pretreatment is not suitable for _Y. lipolytica_ cells. The use of high-pressure processing is rather scarce for yeast biomass treatment and more often high-pressure homogenization was applied for metabolite extraction. HPH was used in the previously mentioned preliminary studies [27], as well as in the papers of [PERSON] et al. [42] and [PERSON] et al. [36]. As a result of using homogenization at a high pressure of 2000 bar for 5 min with 15 passes of the sample through the homogenizer, 95% disruption efficiency was observed for _S. podzolica_ and only 53% for _Apidrichum porosum_, which in the case of the selected extraction methods, i.e., Folch, Bligh-Dyer, or ethanol-hexane mixture, translated into 37.8-46.9% and 14.1-15.0% of whole lipid/CDW, respectively [42]. In [PERSON] et al. 's [36] research, the use of HPH was the most efficient method for _Y. lipolytica_ cell disruption, which resulted in 83.8% oil extraction yield with only 19.8% for the control method (Table 1). In the case of their study, a cell suspension of 15% DM was passed through the homogenizer 20 times at a pressure of 1500 bar. The obtained results were 2- to 4-fold higher in comparison with the application of US or PEF. Selecting the appropriate permeabilization method is a crucial step before extraction, as described in the above discussion. On the other hand, selecting the extraction method itself is another key factor when obtaining valuable substances from the inside of yeast cells. [PERSON] et al.