F₂-isoprostanes in Fish mucus: A new, non-invasive method for analyzing a biomarker of oxidative stress

F₂-isoprostanes (F₂-isoPs) are a reliable biomarker class for oxidative stress in vivo in animals. These compounds are traditionally measured in matrices like liver and plasma, however social and environmental pressures warrant the development of non-lethal and non-invasive methods to assess animal health. Therefore, this study aimed to develop a high-performance liquid chromatography tandem mass spectrometry (HPLC-ESI-MS/MS) method to separate and detect F₂-isoPs in fish mucus. The method was developed and validated for four native F₂-isoP isomers using Northern pike mucus (Esox lucius). Linearity was observed between 5 and 1000 pg/μL. The limits of detection of the four F₂-IsoP isomers ranged from 0.63 to 2.0 ng/g. Recoveries ranged from 78 to 95%, and matrix effects were small (<10%). The between-day and within-day repeatability for all target analytes was lower than 20% RSD. Endogenous F₂-isoPs were measured in the pike mucus (5.3-28.8 ng/g). A preliminary study of baseline F₂-isoP concentrations in lake trout (Salvelinus namaycush) captured from five lakes at the IISD-Experimental Lakes Area in Northwestern Ontario, Canada, was also conducted to test the interspecies applicability of the method. Endogenous F₂-isoPs were quantified in lake trout (6.3-132 ng/g). Lake trout samples displayed large variability within and between the different lakes, which suggests sampling methods may require adjustment for this species. This work developed a sensitive analytical method for measuring F₂-isoPs in fish mucus, however several further studies are required to determine its ability to accurately measure oxidative stress in fish species.