Final Report Summary - MARAGNANO (The behaviour, fate and ecotoxicological effects of silver nanoparticles in estuarine and coastal waters) AgNPs are the most widely used NPs in consumer products (textiles, deodorants, gels, cosmetics, household appliances, etc.). The increasing production of manufactured goods containing AgNPs is potentially leading to increased emissions to the environment, with possible detrimental effects on humans and ecosystems. In this project (www.maragnanoproject.com) we have obtained data that will help to fill the gap already present regarding the behaviour, fate and toxic effects of AgNPs in complex natural waters, particularly in estuarine and coastal waters. Moreover, we have also obtained important data (aggregation/oxidation kinetics, surface plasmon resonance band characterization, silver speciation, etc.) under realistic environmental concentrations conditions (nM range). The research was strongly interdisciplinary, linking chemistry (AgNP manufacturing and chemical NP measurements), oceanography (AgNP behaviour and fate) and biology (AgNP toxicity), and deals with a societal pollution issue from consumer products for which no legislation is currently available. We are confident that those findings will assist us in developing new approaches to oversee and regulate AgNPs, detect if they are present in European Union marine waters in the near future and provide policy makers with important tools to prepare and enforce environmental legislation.Summary overview of the main results obtained in this project:- The manufacture of AgNPs stable in aqueous solution (around 6 months) with different sizes and coatings.- The coating layer has been revealed as a key factor in the aggregation/oxidation behaviour of AgNP in solution. Electrostatically stabilized AgNPs readily aggregate in waters with salinities higher than 4.5-9 and their likely fate is sedimentation and kinetic deposition if no additional stabilisation from organic matter is considered. Sterically stabilised AgNPs remain as individual NPs for a longer duration in waters column, even at high salinities of seawater (around 35), and consequently they are available for uptake by marine organisms. The ionic strength is not the only factor that influenced the aggregation rate of the polymer-coated AgNPs. The discharge of AgNPs into natural marine waters resulted in additional stabilisation from organic matter independently of the steric/electrostatic behaviour provided by the polymer coating, except when high concentrations of Ag+ species are present.- We have developed a novel method that allows detection of AgNPs in aqueous samples at the expected environmental concentrations (pM-nM range). To our best knowledge, this is a new achievement and will allow us to observe AgNP in natural waters and study their behaviour at realistic concentrations. The work undertaken until now, allowed us to detect the surface plamon resonance band, characteristic of AgNPs, in solutions doped with less than 50 nmol/L of silver. Nevertheless, lower detections limits should be achieved in order to be able to detect AgNPs already present in marine waters. Improvements in the spectrophotometer sensibility, the light source intensity and stability, together with the development of materials with lower light absorption in the long path cells (>2 m), will help to reduce the AgNPs detection limits already achieved on this work.- We have observed a significant (49% decrease in chlorophyll content and 10% decrease in quantum yield values, compared to a control in absence of silver) toxicity effect of NM300K AgNPs at μM concentrations to the species Chaetoceros curvisetus in absence of Ag+ ions in solution. No oxidation of NM300K AgNPs was detected after 24 h of deposition in culture media containing phytoplankton. Silver ion speciation in the culture media is dominated by chloride ions; while the presence of Ag+ ions represents around 0.01% of the total species at silver concentrations ranging between 0 to 3 μM.- The obtained dynamic light scattering and UV-Visible data from samples with different coatings at varying ionic strengths were described with accuracy by the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory providing useful parameters to characterize the NPs stability, such as critical coagulation concentration (CCC), aggregation efficiency, Hamaker constants and fractal dimensions, in artificial NaCl solutions and natural marine waters. Electrostatically stabilized AgNPs (coated with alginate or tannic acid) presented lower CCCs values than the sterically stabilised AgNPs (coated for example with BPEI, GA, or Tween 20). The application of second order kinetic rate equations allowed us to accurately describe the aggregation kinetics of AgNPs and distinguish between different aggregation steps. The application of numerical simulations showed that AgNPs oxidation is increased with the AgNPs concentration reduction.Conclusions- We have evidenced the importance of the NP purification step during manufacturing and also the need of good NP characterization in terms of silver ions measurement in the stock solution. An inefficient purification step when AgNPs are manufactured will lead to the presence of unreacted silver ions in the NP stock solution. Moreover, a high affinity of the non-reduced silver ions present in solution for the NP coating and/or NPs surfaces will also produce the same toxicity effect even if intense purification is carried out. Therefore, if unreacted silver ions are present in the NPs stock solution they will have an influence on AgNPs behaviour in terms of aggregation/oxidation effects. This phenomenon is poorly understood and was studied in this project using gum arabic-coated (GA) AgNPs. When the AgNPs coated with GA were subjected to high concentrations of Ag+ in the stock solution (AgNP-GAH with >40% of the total silver content bound to the NPs and/or coating surfaces), the steric stabilisation of gum arabic coating was not noticeable. This was associated to changes in the coating layer structure and/or adsorption due to interactions with different silver-chloride species present in excess and interacting with the AgNPs.- We have set up a spectrophotometric method that allows detection of AgNPs in aqueous samples at the expected environmental concentrations (pM-nM range). Nevertheless, further improvements to analytical techniques are required to reduce the detection limits for AgNPs in order to allow detection in marine waters (probably at concentrations - We were able to study the behaviour and fate of AgNPs coated with very different polymers in complex environments, such as estuarine and marine waters. Moreover, the toxicity effects of AgNP/Ag species over a marine diatom were also determined. As a first approximation, we used synthetic NaCl solutions of increasing concentrations in order to understand aggregation mechanisms and extrapolate the results to more complex matrixes. Chloride ions are the main components in marine waters and also a key factor in silver speciation, so their use in this work provided us with a good approximation for the real behaviour of AgNPs in natural waters. The organic matter content, the specific coating bound to the NPs surface and the ionic strength of the medium, were found to be key factors in the AgNPs behaviour and fate.- The DLVO theory provided realistic descriptions of the aggregation of NPs in solution. Moreover, the application of second order kinetic rate equations allowed us to accurately describe the aggregation kinetics of AgNPs and distinguish between different aggregation steps. final1-tem-image-agnp.tif final1-agnp-solution.jpg final1-sem-image-agnps-in-nacl-solution.tif Related documents final1-project-logo.pdf
