Toxicity in vitro and in Zebrafish Embryonic Development of Gold Nanoparticles Biosynthesized Using Cystoseira Macroalgae Extracts

Abstract

Sofia Machado,1,* Noelia González-Ballesteros,2,* Anabela Gonçalves,1,3 Luana Magalhães,1 Marisa Sárria Pereira de Passos,4,5 Maria Carmen Rodríguez-Argüelles,2 Andreia Castro Gomes1,3 1Centre of Molecular and Environmental Biology (CBMA), Universidade do Minho, Campus de Gualtar, Braga, 4710-057, Portugal; 2CINBIO, Departamento de Química Inorgánica, Universidade de Vigo, Vigo, 36310, Spain; 3Institute of Science and Innovation for Bio-Sustainability (IB-S), Universidade do Minho, Campus de Gualtar, Braga, 4710-057, Portugal; 4International Iberian Nanotechnology Laboratory (INL), Braga, 4715-330, Portugal; 5European Commission, Joint Research Centre (JRC), Ispra, 21027, Italy*These authors contributed equally to this workCorrespondence: Andreia Castro GomesInstitute of Science and Innovation for Bio-Sustainability (IB-S), Universidade do Minho, Departamento de Biologia, Campus de Gualtar, Braga, 4710-057, PortugalTel [phone]Email [email protected] Carmen Rodríguez-ArgüellesCINBIO, Departamento de Química Inorgánica. Universidade de Vigo, Vigo, 36310, SpainTel [phone]Email [email protected]: Research on gold nanoparticles (AuNPs) occupies a prominent place in the field of biomedicine nowadays, being their putative toxicity and bioactivity areas of major concern. The green synthesis of metallic nanoparticles using extracts from marine organisms allows the avoidance of hazardous production steps while maintaining features of interest, thus enabling the exploitation of their promising bioactivity.Objective: To synthesize and characterize AuNPs using, for the first time, macroalga Cystoseira tamariscifolia aqueous extract (Au@CT).Methods: Algal aqueous extracts were used for the synthesis of AuNPs, which were characterized using a wide panel of physicochemical techniques and biological assays.Results: The characterization by UV-Vis spectroscopy, transmission electron microscopy, Z-potential and infrared spectroscopy confirmed that Au@CT were stable, spherical and polycrystalline, with a mean diameter of 7.6 ± 2.2 nm. The antioxidant capacity of the extract, prior to and after synthesis, was analyzed in vitro, showing that the high antioxidant potential was not lost during the synthesis. Subsequently, in vitro and in vivo toxicity was screened, by comparing two species of the genus Cystoseira (C. tamariscifolia and C. baccata) and the corresponding biosynthesized gold nanoparticles (Au@CT and Au@CB). Cytotoxicity was tested in mouse (L929) and human (BJ5ta) fibroblast cell lines. In both cases, only the highest (nominal) test concentration of both extracts (31.25 mg/mL) or Au@CB (12.5 mM) significantly affected cell viability, as measured by the MTT assay. These results were corroborated by a Fish Embryo Acute Toxicity (FET) test. Briefly, it was shown that, at the highest (nominal) tested concentration (31.25 mg/mL), CT extract induced significantly higher cytotoxicity and embryotoxicity than CB extract. However, it was demonstrated that Au@CT, but not Au@CB, were generally non-toxic. At sub-lethal (nominal) test concentrations (1.25 and 2.5 mM), Au@CT affected zebrafish embryonic development to a much lesser extent than Au@CB. In vitro wound healing assays also revealed that, while other experimental conditions did not impact cell migration, CT and Au@CT displayed a moderate positive effect.Conclusion: Au@CT and Au@CB display promising features, desirable for biomedical applications, as wound healing.Keywords: gold nanoparticles, green synthesis, Cystoseira sp., zebrafish embryotoxicity test, bioactivity, toxicity

Keywords

• gold nanoparticles
• green synthesis
• cystoseira sp.
• zebrafish embryotoxicity test
• bioactivity
• toxicity