Production of BioSeNPs in fungus Phycomyces blakesleanus is accompanied by decrease in intracellular thiols as detected by in vivo EPR

Abstract

Selenium represents an essential microelement for many organisms, and exists in several oxidation states. Selenite (Se4+) is often toxic due to its water solubility and bioavailability, but can be reduced to insoluble Se0 by many microorganisms including fungi. Fungus Phycomyces blakesleeanus reduces Se4+ to Se0 and forms selenium nanoparticles (BioSeNPs) as witnessed by red coloration of the mycelium after 24h of treatment with 2, 5 and 10 mM Se4+. SEM micrographics and EDS spectra confirmed presence of BioSeNPs with characteristic peaks at 1.4 (SeLα), 11.2 (SeKα) and 12.5 (SeKβ) keV. One of the mechanisms of selenite toxicity is considered to be production of reactive oxygen species that then triggers response of cellular antioxidative systems, among which important part is played by thiols such as glutathione. Glutathione is also believed to play a more direct role in reduction of Se4+ to Se0 as proposed by the Painter reaction. Thiol specific biradical EPR probe (RSSR) was used for EPR in vivo detection of intracellular thiolgroup modifications in mycelium of fungus P. blakesleanus treated by 0.1, 0.5, 1, 2, 5 and 10 mM selenite (Na2SeO3) for 24 h. Decrease of available –SH groups is detected as the decrease in ratio of mono- and biradical peak in the spectrum of the given sample. This ratio was 43.05 for control, and didn’t change with 0.1 mM treatment (42.23), but halved to 23.55 with 0.5 mM Se4+. It further decreased 10 × to minimal values of 4.31, 4.21, 4.1 and 3.5 with 1, 2, 5 and 10 mM Se4+, respectively. This cannot be solely attributed to biomass decrease as it never decreased more than 45%. P. blakesleeanus forms BioSeNPs from Se4+ and intracellular thiols are involved in this process either through neutralization of ROS or directly in the reduction of Se4+, but most probably both.