Optimized extraction, quantification and characterization of seven environmentally relevant metallic nanoparticles in sewage sludge: Occurrence, composition and environmental risk assessment.

The quantitative analysis of metallic nanoparticles (MNPs) in environmental matrices is crucial for understanding their occurrence, fate, and ecological risks. This study presents the protocol for extracting and quantifying seven environmentally relevant MNPs (Ag, Au, Ce, Cu, Pb, Ti and Zn NPs) from sewage sludge using single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS). Several extraction parameters and solutions, including milli-Q water, tetrasodium pyrophosphate (TSPP), and tetramethylammonium hydroxide (TMAH), were systematically evaluated. TSPP at 2.5 mM in a 1:100 sludge-to-reagent ratio was identified as optimal, providing high particle number recovery (>75 %) and mass recovery (>84.5 %) while minimizing particle transformation. Application to Croatian wastewater treatment plant sludge samples resulted in particle concentrations of 1 × 10-3 × 10 particles/g, with extracted MNPs representing < 2 % of total metal content for most elements. Scanning electron microscopy coupled with energy dispersive X-ray spectroscopy revealed that Ag, Cu, Pb, Ti and Zn-based NPs are possibly of anthropogenic origin, which was corroborated by enrichment factor analysis. Risk assessment indicated negligible to low ecological risks under current agricultural scenarios, though highly soluble species (Ag and Zn NPs) may pose greater long-term environmental risks. This methodology addresses critical gaps in environmental monitoring capabilities for engineered NPs.