Inability to detect free cylindrospermopsin in spiked aquatic organism extracts plausibly suggests protein binding

- Cylindrospermopsin (CYN) binding has only been previously reported in eukaryotic reticulocyte lysate.
- Lower CYN quantified after incubated with crude lysate from various organisms compared to control.
- Lower CYN quantified after incubation with various (NH4)2SO4 precipitation fractions of Egeria densa protein.
- All free CYN could be accounted for after incubation with single amino acids and glutathione.
- Free CYN % varied with exposure to animal and plant proteins.

Even though the frequency and prevalence of cylindrospermopsin producing cyanobacteria are increasing, several publications have reported the absence of free cylindrospermopsin bioaccumulation in aquatic food chains. Cylindrospermopsin modification by protein binding has been suggested, however, only one publication has investigated this with eukaryotic reticulocyte lysate and concluded that cylindrospermopsin binds non-covalently to soluble proteins larger than 100 kDa associated with eukaryotic translation. With this as the extent of knowledge regarding cylindrospermopsin binding, the present study aimed to determine whether cylindrospermopsin binding also occurred with other proteins. In the present study, proteins from various organisms were extracted, incubated with cylindrospermopsin, and the amount of free cylindrospermopsin was determined by liquid chromatography tandem mass spectroscopy. Additionally, cylindrospermopsin binding to various ammonium sulfate precipitation fractions of Egeria densa protein, as well as with selected amino acids was investigated. We find that the percentage of free cylindrospermopsin varied with exposure to various animal and plant proteins as well as with various fractions of proteins but found no binding with single amino acids.