## 4 Discussion
Benthic organisms may be exposed to contaminants on different routes, such as by their contact with pore water, sediment-water interface through walls of the body, and respiratory surface or by the ingestion of contaminated sediment particles [37]. Although no significant difference was found in the metal concentration in the sediment stations, the higher difference in the macro-benthic communities could be related to the effects of other environmental factors and contaminants that were not measured in this study. The Santos estuary is influenced by a number of factors, such as port and dredging activities, human sewage effluents, contribution of nutrients from the mangroves, and others [16; 25; 34]. It was reported by [16; 23] a greater bioavailability of PAHs at CPI station, such as benzo(a)anthracene, benzo(a)pyrene, and dibenzo(a,h)anthracene, which are considered very toxic for both biota and humans [38; 39]. Furthermore, these authors reported higher concentrations of Hg bioaccumulated in the oysters. Thus, all of these factors reported and the high carbon concentration (7.5%) found in the CPI station could contribute to the low biological indexes of macro-benthic community present there. It is known that the reduction in the concentration of both hydroxide and carbonate (OH\({}^{-}\) and CO\({}_{3}\)\({}^{2-}\)) could change the speciation of some metals and, consequently, raise up the fraction in free forms at lower pH [40]. One of the lines of evidence selected for this study was to analyze the mobility of metals and their toxicity in relation to acidification by enrichment of CO\({}_{2}\). The results showed a significant increase in the metals concentrations in overlying water and elutriate treatment at pH 6.5 and 6.0 using the tabular matrix and the pie charts for both sediment stations when compared with the reference case (PAI, pH 8.0). Moreover, moderate effects in the mobility of metals were observed in the gray areas (Figure 4) at pH 7.5 and 7.0 using the CPI station. A recent study showed that there are more sulfides forms in the sediments from SES than other complex forms [16]. Furthermore, these authors reported that when there is more sulfide complex than divalent metals in the environment it is supposed that these metals are complexed to the sulfides, which are less bioavailable. According to [40], metals form strong complexes with hydroxide and carbonate, which undergo significant changes in speciation as the pH of seawater decreases. Thus, for an environmental risk assessment in case of a CO\({}_{2}\) enrichment, it is important to analyze the speciation of metals to understand interactions with lower pH in seawater and how they affect the biota. The tabular matrix (Table 3) and pie charts (Figure 4) showed chronic effects associated with pH values of 7.0 and pH 6.5 for PAI and CPI sediment, respectively. However, the sediment from PAI is considerate lesser contaminated than CPI sediment, so higher bioavailability of some elements are present as carbonate forms. In addition, the results using the multivariate analysis indicated strongly correlation between certain metals (As, Zn, and Ni in seawater, and Cr, As, and Cu in elutriate) and the biological effects related to the increase in the concentration of H\({}^{+}\) protons (pH reduction). These results may confirm the statistical results reported (Table 3 and Figure 4) and provide an estimation of the metal(iod)s related biological responses (included in the 'gray and black' areas of the WoE analysis from Figure 4). Furthermore, the results of factor scores from component 2# also confirm a relationship between metal mobility, toxicity, and acidification as a positive score only at the lowest pH (6.0) in both stations. This means that the CO\({}_{2}\) enrichment in these stations could cause effects under extreme acidified conditions-.