Carbamazepine exposure in the sea anemones Anemonia sulcata and Actinia equina: metabolite identification and physiological responses

Abstract

There is a rising interest for both the conservation and biotechnological application of Sea anemones belonging to Anthozoa Class, (Cnidaria Phylum). Global warming has been recognised as one of the most important hazards for some species but the risk posed by pharmaceuticals and other emergent compounds present in the coastal waters they inhabited have been less studied. Anemonia sulcata and Actinia equina are widely distributed species in the Mediterranean Sea. The objective of this work has been to determine the physiological and biochemical effects of carbamazepine (CBZ) exposure as well as to characterize the degradation routes followed by this compound in water and tissues (biotransformation) of the two anemones species. Anemones provided by Oceanografic of Valencia Aquarium were acclimatized to laboratory conditions for two weeks and exposed to 1 ¿g/L and 100 ¿g/L of CBZ in artificial sea water in a semi-static system. Small portion of the tentacles were taken after two and seven days. Ion transport (measured as NKATPase activity), energetic metabolism (measured as glucose and lactate levels) and nitrogen excretion (measured as ammonia concentration in tissues) were determined. Treated samples of A. sulcata and A. equina were analysed by ultra-high-performance liquid chromatographyhigh-resolution mass spectrometry on a quadrupole-time-of-flight (QqTOF) (TripleTOF®, ABsciex). The structures of metabolites have been proposed using HRMS and HRMS/MS data. In particular, the combination of mass accuracy and the fragmentation patterns of metabolites and parent compound allowed proposing plausible structures for each metabolite. CBZ metabolites originated from aromatic and benzylic hydroxylation and epoxidation. The analysis of the water samples saved from each aquarium pointed out that a small percentage of CBZ can be degraded by hydrolysis and/or oxidation to the hydroxylated metabolites but not to the epoxidized ones. The current work constitutes the first study on the identification of Cnidaria metabolites of CBZ in species of the Anthozoa Class.