Sensitive LC-MS/MS method for Palytoxin using cationization

Palytoxins are produced by marine dinoflagellates, cyanobacteria and certain corals and is a potent toxin in test animals. Palytoxins are within the marine biotoxins the group with the largest molecular structure. Legislation... [ view full abstract ]

Palytoxins are produced by marine dinoflagellates, cyanobacteria and certain corals and is a potent toxin in test animals. Palytoxins are within the marine biotoxins the group with the largest molecular structure. Legislation for palytoxins has not been established yet but there is an EFSA opinion where it is stated that in order to avoid exceeding the acute reference dose a shellfish portion of 400g should not contain more than 12ug PlTX (30ug/kg shellfish meat). When ionized with electrospray ionization (ESI) palytoxin shows a complex MS spectrum with multiple salt adducts (Ca, Na and K), multiple charge states and in-source fragmentation. The most abundant ion present is less than 30% of the sum of all ions formed therefore sensitivity is relatively low.

To gain sensitivity there was attempted to simplify the ionization by applying cationization. As cation lithium was used. The presence of lithium directed the ionization mainly to the formation of [M+H+2Li]³⁺ and [M+H+Li]²⁺, where the abundance of [M+H+2Li]³⁺ was around 60% of the total abundance. Fragments are mainly losses of H₂O. When measured with MS/MS the most sensitive transition is [M+H+2Li]³⁺ m/z 897.8>1215.7, where m/z 1215.7 is not yet elucidated. In the final procedure 0.25mM LiI and 0.31mM formic acid were added to the mobile phase. The developed method had a limit of quantitation of 1ng/ml, which is low for the measurement of intact palytoxin. During the presentation the development as well as the application to palytoxins will be presented.