Lipase is one of the most commonly used biocatalysts in the food, pharmaceutical and cosmetic industries, and can be produced by Yarrowia lipolytica yeast. Despite the intensive studies of lipase from Yarrowia, there are still many open questions regarding the enzyme secretion process, especially by new isolates of this genus as well as the effect of substrates or surfactants, or both on the production of lipase. This research focused on the effect of olive oil and surfactant Tween 80 including the optimisation of the concentration of these compounds on the production of lipase by some novel Yarrowia isolates. Moreover, the optimal environmental parameters (pH, temperature) of crude enzyme synthetised by Yarrowia strains were determined. All investigated strains were able to produce lipase in both intracellular and extracellular fractions. The extracellular lipase activities were higher than the intracellular ones (Y. divulgata Y.02062 and Yarrowia lipolytica 854/4 147 U/L, 80 U/L and 474 U/L, 122 U/L, respectively). In the case of extracellular lipase, supplementing olive oil and Tween 80 enhanced significantly the synthesis and secretion of the enzyme. The lipase activity can even be enhanced by 20 times higher from 25 U/L to 474 U/L in the case of Yarrowia lipolytica 854/4 strain. In the case of intracellular, supplementation of Tween 80 generally reduces lipase activity except for the Y. lipolytica 1/4 strain, which was affected by two times the increase. The optimised concentration of olive oil and Tween 80 were determined for Y. divulgata Y.02062, Y. divulgata 5257, Y. lipolytica 1/4, and Yarrowia lipolytica 854/4 strains as 1.6% olive oil and 0.09% Tween 80, 1.6% olive oil and 0.06% Tween 80, 1.4% olive oil and 0.09% Tween 80 as well as 1.6% olive oil and 0.065% Tween 80, respectively. The optimum pH and temperature of crude lipases (intra and extracellular) synthetised by the tested Yarrowia lipolytica and Y. divulgata yeast strains were found to be pH 7.2 and 37 °C, respectively. Our results confirmed that the new isolate Y. divulgata is a very promising species for further development for industrial use as Y. lipolytica.

Lactic acid bacteria exhibit high adaptability to their environment due to their wide variety of enzymes. Despite extensive knowledge of bacterial cell walls, the subcellular localization of β-galactosidase in many probiotic lactic acid bacteria, including Limosilactobacillus fermentum, remains unclear. Determining the cellular localization of such enzymes may improve insight into bacterial metabolic mechanisms and support the development of efficient downstream processes, as well as applications. In this study, three cell disruption strategies (mechanical homogenization and chemical disruption with different agents) were applied to assess the subcellular localization of β-galactosidase from the Ll. fermentum LF08 strain. Enzyme activity was measured in a ferment broth, a supernatant and cell-associated fractions. No and very low β-galactosidase activity was detected in the ferment broth and the supernatant, respectively, when either chemical or mechanical treatment was applied, whereas the main enzyme activity was assayed in the cell suspension fraction. Combined lysozyme and CTAB treatment resulted in a 21.4-fold increase in β-galactosidase activity in the supernatant fraction (2.14 U/mL), compared with CTAB treatment alone (0.10 U/mL). Bioinformatic analyses provided additional significant information to propose the potential cell wall association (maybe the outer side of the cell wall) of the subcellular localization of β-galactosidase. This feature may support understanding of the interactions between probiotic bacteria and host tissues, as well as the development of probiotic immobilized cell systems for applications such as the elimination of lactose, designing novel functional foods.