Mass spectrometry based detection of common vitellogenin peptides across fish species for assessing exposure to estrogenic compounds in aquatic environments

The identification of myriad of chemicals in the environment that mimic hormones and affect the endocrine functions of exposed organism is a daunting analytical challenge for environmental scientists and engineers. Many of these endocrine disrupting chemicals (EDCs) are present at very low concentrations in the aquatic systems, but yet affect the metabolic, developmental, and reproductive functions in exposed fish and wildlife. Vitellogenin (VTG) protein is a widely used biomarker in fish for assessing exposure to EDCs, and is commonly measured using species-specific immunochemical techniques. In this study, we developed a liquid chromatography tandem mass spectrometry (LC-MS/MS) method that can measure common peptides from digested VTG in multiple fish species. In the initial experiments using high resolution mass spectrometry, two peptides (ALHPELR and FIELIQLLR) were identified as common fragments in the digested VTG protein isolated from three different fish species (Pimephales promelas, Micropterus salmoides, and Fundulus heteroclitus). Then, a quantitative analysis using LC-MS/MS under selected reaction monitoring mode was developed for the detection of these two peptides in trypsin-digested plasma from female fish (positive control), estrogen-exposed male fish (test sample), and unexposed male fish (negative control) using two of the same species used for identifying the common peptides (P. promelas, and M. salmoides) and one new species (Ameiurus nebulosus) that was not included during the selection of peptides. Results from this study demonstrate the potential of LC-MS/MS as an effective cross-species method to detect VTG in fish, which can be an alternative analytical technique for assessing endocrine disruption in multiple fish species.