Determination of seleno-amino acids bound to proteins in extra virgin olive oils

• A two-dimensional chromatographic method coupled to ICP MS was developed.
• Seleno-methylselenocysteine and selenocysteine were determined in protein fractions of extra-virgin olive oils (EVOOs).
• Variations of selenoaminoacids and protein concentration between EVOOs varieties were observed.

An analytical method has been developed to determine seleno-amino acids in proteins extracted from extra virgin olive oils (EVOOs). Different aqueous/organic solvents were tested to isolate proteins, an acetone:n-hexane combination being the best protein precipitant. In a first dimension chromatography, extracted proteins were analysed by size exclusion chromatography (SEC) coupled to inductively coupled plasma mass spectrometry (ICP-MS) to identify S and Se associations as proteins marker. Two fractions of 66 kDa (A) and 443 kDa (B) were identified. These fractions were submitted to microwave-assisted acid hydrolysis (MAAH) to release seleno-amino acids. In a second dimension chromatography seleno-amino acids were determined by reversed-phase chromatography (RPC) coupled to ICP-MS. Seleno-methylselenocysteine was determined with values ranging from 1.03–2.03 ± 0.2 μg kg−1 and selenocysteine at a concentration of 1.47 ± 0.1 μg kg−1. Variations of protein and seleno-amino acid concentrations were observed between EVOO varieties, contributing to EVOO cultivar differentiation.