| Summary: | Research on nanomaterial exposure-related health risks is still quite limited; this includes standardizing methods for measuring metals in living organisms. Thus, this study validated an atomic absorption spectrophotometry method to determine fertility and bioaccumulated iron content in <i>Drosophila melanogaster</i> flies after feeding them magnetite nanoparticles (Fe<sub>3</sub>O<sub>4</sub>NPs) dosed in a culture medium (100, 250, 500, and 1000 mg kg<sup>−1</sup>). Some NPs were also coated with chitosan to compare iron assimilation. Considering both accuracy and precision, results showed the method was optimal for concentrations greater than 20 mg L<sup>−1</sup>. Recovery values were considered optimum within the 95–105% range. Regarding fertility, offspring for each coated and non-coated NPs concentration decreased in relation to the control group. Flies exposed to 100 mg L<sup>−1</sup> of coated NPs presented the lowest fertility level and highest bioaccumulation factor. Despite an association between iron bioaccumulation and NPs concentration, the 500 mg L<sup>−1</sup> dose of coated and non-coated NPs showed similar iron concentrations to those of the control group. Thus, <i>Drosophila</i> flies’ fertility decreased after NPs exposure, while iron bioaccumulation was related to NPs concentration and coating. We determined this method can overcome sample limitations and biological matrix-associated heterogeneity, thus allowing for bioaccumulated iron detection regardless of exposure to coated or non-coated magnetite NPs, meaning this protocol could be applicable with any type of iron NPs.
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