First evidence of protein profile alteration due to the main cocaine metabolite (benzoylecgonine) in a freshwater biological model

Illicit drugs represent not only a great social problem but are also considered an environmental problem because their use and, often, abuse release large amounts of parent compounds, and especially their metabolites, into freshwaters. One of the most commonly used drugs is cocaine, which is the second most prevalent drug in Europe (accounting for almost 30% of all cocaine users worldwide). Cocaine is rapidly metabolised in humans to benzoylecgonine (35-54%), ecgonine methyl ester (32-49%) and norcocaine (5%), which are eliminated in the urine and are only partially removed by wastewater treatment plants (WWTPs). Because no studies have previously been carried out to evaluate the possible risks due to cocaine and its metabolites in non-target organisms, we applied a multi-disciplinary approach to investigate the possible environmental risk related to benzoylecgonine (BE), the main metabolite of cocaine. Previous studies carried out by means of a biomarker suite and the redox-proteomic approach showed an imbalance of anti-oxidant enzyme activities and several genotoxic effects to be caused by environmental BE concentrations in the freshwater bivalve Zebra mussel (Dreissena polymorpha). This report presents the results obtained in the last step of this study, based on a proteomics analysis. We analysed the protein expression profile in the gills of Zebra mussels exposed to two different concentrations (0.5 and 1 μg/L) of BE for 14 days through 2-DE and mass spectrometry analysis (RP-UPLC ESI-LTQ-Orbitrap). Our results highlight significant changes in some proteins in gill cells whose functions are crucial for overall metabolism. In particular, we detected a probable effect of BE on calcium homeostasis and a consequent imbalance of oxidative stress, as verified for vertebrates.