Combining ultrafiltration and diffusive gradients in thin films techniques for speciation/fractionation of Cu and Zn in cytosol of liver of Nile tilapia (Oreochromis niloticus)

• Published in: Metallomics 2023-04, Vol.15 (4)
• Main Authors: Gemeiner, Hendryk, Menegário, Amauri Antonio, Eismann, Carlos Eduardo, Elias, Lucas Pellegrini, Pedrobom, Jorge Henrique, Dourado, Thiago de Araujo, Chang, Hung Kiang, da Conceição, Fabiano Tomazini, Moruzzi, Rodrigo Braga
• Format: Article
• Language: English
• Subjects: Animals; Cichlids; Cytosol; Environmental Monitoring - methods; Liver - chemistry; Ultrafiltration - methods; Water Pollutants, Chemical - analysis; Zinc - analysis
• ISSN: 1756-591X; 1756-591X
• DOI: 10.1093/mtomcs/mfad018

Abstract This work aims to evaluate the size and lability of Cu and Zn bound to proteins in the cytosol of fish liver of Oreochromis niloticus by employing solid-phase extraction (SPE), diffusive gradients in thin films (DGT), and ultrafiltration (UF). SPE was carried out using Chelex-100. DGT containing Chelex-100 as binding agent was employed. Analyte concentrations were determined by ICP-MS. Total Cu and Zn concentrations in cytosol (1 g of fish liver in 5 ml of Tris–HCl) ranged from 39.6 to 44.3 ng ml−1 and 1498 to 2106 ng ml−1, respectively. Data from UF (10–30 kDa) suggested that Cu and Zn in cytosol were associated with ∼70% and 95%, respectively, with high-molecular-weight proteins. Cu-metallothionein was not selectively detected (although 28% of Cu was associated with low-molecular-weight proteins). However, information about the specific proteins in the cytosol will require coupling UF with organic mass spectrometry. Data from SPE showed the presence of labile Cu species of ∼17%, while the fraction of labile Zn species was >55%. However, data from DGT suggested a fraction of labile Cu species only of 7% and a labile Zn fraction of 5%. This data, as compared with previous data from literature, suggests that the DGT technique gave a more plausible estimation of the labile pool of Zn and Cu in cytosol. The combination of results from UF and DGT is capable of contributing to the knowledge about the labile and low-molecular pool of Cu and Zn. Graphical Abstract Graphical Abstract Combining ultrafiltration and DGT to provide information on the labile pool of Cu and Zn and its relationship with small and large-molecular weight proteins in liver cytosol of Oreochromis niloticus.