**5. Conclusions**

Based on the theoretical discussion presented, regarding the right selection of a single extraction protocol, it results clear that for functional speciation studies, it is of utmost relevance to evaluate and correlate the phytoavailable/phytotoxic HM levels determined, as affected by the ameliorating agronomic materials added, with plant growth, through the application of HM-sensitive plant bioassays.

Total concentration cannot be used as a guide to determine the area impacted by the acid spill, because no difference in total metal(loid) concentrations was noted between polluted and baseline sediments after control and remedial measures were taken. On the other hand, As fractionation is an excellent option. Two other reported strategies for the identification of areas of the river impacted by the acid solution spill are based on the detection of gypsum formed after the addition of calcite, and of jarosite formed only under the acidic conditions prevailing at the dam. However, these two methods require sophisticated equipment, as amorphous particles are practically invisible to most analytical techniques. In their stead, sequential extraction conforms to a relatively simple and inexpensive method. Monitoring metal(loid) behavior is recommended in this area to evaluate changes in sediments. The eventual attainment of an equilibrium that would form more insoluble compounds, mainly from Cu, is expected. This F1-fraction recovered metal can be desorbed, which would likely impact macro-invertebrate populations of the river.
