Metabolomics-Based Investigation on the Metabolic Changes in Crassostrea gigas Experimentally Exposed to Galvanic Anodes

Abstract

Cathodic protection is widely used to protect metal structures from corrosion in marine environments using sacrificial galvanic anodes. These anodes, either in Zinc, or preferentially nowadays in Al-Zn-In alloys, are expected to corrode instead of the metal structures. This leads to the release of dissolved species, Zn²⁺, Al³⁺, and In³⁺, and solid phases such as Al(OH)₃. Few studies have been conducted on their effects on marine organisms, and they concluded that further investigations are needed. We therefore evaluated the effects of Zn and Al-Zn-In anodes on oysters stabulated in tanks, under controlled conditions defined through a comparison with those prevailing in a given commercial seaport used as reference. We analyzed the entire metabolome of gills with a non-targeted metabolomic approach HRMS. A modelling study of the chemical species, corresponding to the degradation products of the anodes, likely to be present near the exposed oysters, was also included. We identified 16 and two metabolites modulated by Zn- and Al-Zn-In-anodes, respectively, that were involved in energy metabolism, osmoregulation, oxidative stress, lipid, nucleotide nucleoside and amino acid metabolisms, defense and signaling pathways. The combination of chemical modelling and metabolomic approach, used here for the first time, enlightened the influence of Zn present in the Al-Zn-In anodes.

Keywords: aluminum; bioaccumulation; biological effects; corrosion products modelling; metabolomics; oysters; sacrificial galvanic anode; zinc.

Figure 1

Schematic representation of the experimental setup consisting of potentiostat, galvanic anode, passive sampler, and oysters exposed to the product of anode dissolution in an open-circuit tank. The LSNM-NP passive samplers were set among the oysters, inside the area demarcated by the dashed blue line (C.-E.: Counter-electrode; Ref.: reference electrode; WE: working electrode).

Figure 2

The PCA scores of the oyster samples in each group with the ESI positive (LC-Positive) and negative (LC-Negative) ion modes. T1 represents principal component 1, t2 represents principal component 2, and t3 represents principal component 2. The dotted ellipse represented the confidence limit (95%) of Hotelling’s T2 statistic. The reference samples (“Reference”), the Zn-anode exposed (“Zn-exposed”), and the Al-Zn-In anode exposed (“Al-exposed”) samples are visually grouped in orange, blue, and grey ellipses, respectively.

Figure 3

PLS-DA scores plotted for Reference and Zn-exposed samples for ESI positive (LC-Positive) and negative (LC-Negative) ion modes (top) and for Reference and Al-exposed samples for ESI negative (LC-Negative) ion mode (bottom). t1 represents principal component 1, t2 represents principal component 2, and t3 represents principal component 2. The dotted ellipse represented the confidence limit (95%) of Hotelling’s T2 statistic. The reference samples (“Reference”), the Zn-anode exposed (“Zn-exposed”), and the Al-Zn-In anode exposed (“Al-exposed”) samples are visually grouped in orange, blue, and grey ellipses, respectively.

Figure 4

Box plots of the relative abundance of significant compounds (16 identified by the PLS-DA model (Figure 3)) in M. gigas gills. Grey plots represent the reference samples, and red plots represent data obtained on oysters exposed to the dissolution of Zn-anode (the mean value is given by the dark diamond-shaped symbol). Metabolites were classified by family: Group 1: Amino acids and derivatives; Group 2: Nucleotides and nucleosides; Group 3: Carnitine acids and derivatives; Group 4: Others.

Figure 5

Box plots of the relative abundance of significant compounds (two identified by the PLS-DA model (Figure 3)) in M. gigas gills. Grey plots represent the reference samples, and red plots represent data obtained on oysters exposed to the dissolution of Al-Zn-In-anode (the mean value is given by the dark diamond-shaped symbol). Metabolites were classified by family: Group 1: Amino acids and derivatives; Group 4: Others.