2, and 29.8 min (Fig. 1), which correspond to palmitic acid (C16:0), a mixed peak of linoleic (C18:2) and oleic (C18:1) acids, MG-132 mw and ergosterol, respectively. The ethanol extract obtained from W. sebi mycelia showed concentration-dependent lysis of bovine erythrocytes (Fig. 2a). The hemolysis rate (1/t50) of 0.1 min−1 was produced by 25 μg mL−1 of the extract TS obtained after the cultivation at 20% NaCl. If W. sebi was cultivated at the lower 5% NaCl, the same rate of hemolysis was observed only after the addition of approximately 200 μg mL−1 of the extract TS, making this eightfold less active. To further explore the nature of hemolytically active compounds, the most abundant fatty acids in the
extract (C18:1, C18:2, and C16:0) were also tested for their hemolytic potential (Fig. 2b), both separately or in an equimolar mixture. Their hemolytic activity was comparable to that of the W. sebi extract and was associated with the unsaturated forms (C18:1 and C18:2). Ergosterol, which was detected in considerable amounts in the extract (Fig. 1), was also tested for hemolysis and found inactive. buy Buparlisib Exposure to 100 °C significantly affected this ethanolic extract activity, as there was almost total loss of hemolytic activity in comparison with the control (Fig. 3a). The same loss of the activity after heating
to 100 °C could be observed with the equimolar mixture of three tested fatty acids (Fig. 2b). A significant increase in hemolytic activity of the W. sebi extract was observed
at pH above 8.5 (Fig. 3b), and the higher ionic strengths also induced significant increases, although small, in the hemolytic activity (Fig. 3c). As shown on Fig. 4a, the SUVs containing phosphocholine (i.e. those formed with DPPC, DOPC, and POPC) and/or sphingomyelin completely prevented lysis of the erythrocytes that otherwise occurred Celecoxib in first few minutes of assay. This suggests that the phospholipids with a choline headgroup in their structures can bind the hemolytically active compound(s) in the extract and thus diminished their activity toward the erythrocytes. Additionally, the fluorescence of the calcein released from the SUVs was measured after the addition of the extract. Here, the percentage of released calcein was highest in cholesterol-containing vesicles (Fig. 4b), indicating that membranes with a higher degree of fluidity are more susceptible to lysis induced by this W. sebi ethanolic extract. Wallemia sebi is an important pan-global contaminant of foods and feeds preserved with low aw. It can contaminate food not only as an airborne or soil-borne contaminant, but it can also be inoculated with the preservative itself (Butinar et al., 2011). Wallemia sebi can grow over a wide range of aw (0.997–0.690) in glucose/fructose media (Pitt & Hocking, 1997), but in media with NaCl as the major solute, the lowest aw for its growth was reported as 0.80 (Zalar et al., 2005; Plemenitaš et al., 2008), which corresponds to 4.5 M NaCl.