There is a current debate on whether the toxicity of engineered ZnO nanoparticles (NPs) can be traced back to their nanoscale properties or rather to the simple fact of their relatively high solubility and consequent release of Zn2+ ions. In this work, the emerging electroanalytical technique AGNES (Absence of Gradients and Nernstian Equilibrium Stripping), which is specially designed to determine free metal ion concentration, is shown to be able to measure the Zn2+ concentration resulting from dissolution of ZnO nanoparticles dispersed in aqueous salt solutions. Three NP samples from different sources (having average primary particle diameters of 6, 20, and 71 nm) were tested and compared with bulk ZnO material. The enhanced solubility of the nanoparticles with decreasing primary radius allows for an estimation of the surface energy of 0.32 J/m2. AGNES also allows the study of the kinetics of Zn2+ release as a response to a change in the solution parameters (e.g., pH, ZnO concentration). A physicochemical model has been developed to account for the observed kinetic behavior. With this model, only one kinetic parameter is required to describe the time dependence of the free Zn2+ concentration in solution. Good agreement with this prediction is obtained when, starting from an equilibrated NP dispersion, the pH of the medium is lowered. Also, the independence of this parameter from pH, as expected from the model, is obtained at least in the pH range 7–9. When dissolution is studied by dispersing ZnO nanoparticles in the medium, the kinetic parameter initially decreases with time. This decrease can be interpreted as resulting from the increase of the radius of the clusters due to the agglomeration/aggregation phenomena (independently confirmed). For the larger assayed NPs (i.e., 20 and 71 nm), a sufficiently large pH increase leads to a metastable solubility state, suggesting formation of a hydroxide interfacial layer.

AGNES (absence of gradients and Nernstian equilibrium stripping) is a stripping technique consisting of two conceptual steps: (i) application of a potential program (e.g. a step at a fixed potential) generating a known concentration gain between the outer and inner concentrations of the metal at the mercury electrode surface together with null gradients of the concentration profiles (inside and outside the mercury electrode) and (ii) determination of the concentration of reduced metal inside the amalgam in a stripping step. In the present implementation, the stripping step under diffusion limited conditions leads to a measured current just proportional to the free metal ion concentration. In this paper we present the basic principles of the technique, analytical expressions for a simplified model of its voltammetric implementation and a numerical study for a more refined model together with preliminary experimental results in the Cd(II)/nitrilotriacetic acid system showing how this technique can be used as an alternative to other techniques (such as ion selective electrodes) in order to determine free metal activities or concentrations in the presence of complex mixtures avoiding complications such as electrodic adsorption or complexation kinetics.

Absence of gradient and Nernstian equilibrium stripping (AGNES) senses the free ion concentration of Zn(II) in solutions containing different ligands, being unaffected by the lack of reversibility of the Zn 2+ /Zn 0 couple under the conditions assayed. In the presence of oxalate, the determination of [Zn 2+ ] agrees with the stability and solubility constants of this sparingly soluble salt, once the precipitation kinetics are taken into account. Different strategies have been analysed and implemented in order to reduce the preconcentration time with the standard electrode of the polarographic stand (smallest drop radius around 0.141 mm): (i) using a lower preconcentration factor when there is no need of enhanced limit of detection; (ii) splitting the deposition stage into two, with a first potential step under diffusion limited conditions; (iii) the analysis of the chronoamperometric response in the deposition stage allows its duration to be adjusted, especially if non-inert complexes contribute to the arriving flux of metal to the mercury electrode. The two-potential-steps strategy is assessed as the most suitable in a general case.

Absence of Gradients and Nernstian Equilibrium Stripping (AGNES) is a recently suggested electroanalytical technique designed for the determination of the free concentration of heavy metals (such as Zn, Cd or Pb) which is here developed and applied to seawater samples. A key improvement for the implementation of AGNES with complex matrices is the development of a new blank, called the shifted blank (presented in this work for the first time), which can be applied to the same sample where the measurement is intended. The careful selection of the required parameters for the determination of the free Zn concentration (or activity) at the nanomolar level is described in detail. The methodology has been validated with a synthetic solution containing Zn and nitrilotriacetic acid (NTA) and then applied, as a first case, to two coastal seawater samples taken close to Barcelona and Tarragona (Catalonia, North-Eastern Spain) finding values in the range of 1-3nM, representing around 25% of total Zn. This technique can, in the near future, be crucial in helping to elucidate the role of the free zinc(II) concentration in natural waters.

In this study, the effect of ZnO nanoparticles and ZnCl2 on growth, reproduction and accumulation of zinc in Daphnia magna was determined in a 21-day chronic toxicity test. A variety of techniques were used to distinguish the free zinc ion, dissolved, nanoparticle and aggregated zinc fraction in the Daphnia test medium. The results showed similar chronic effects on growth, reproduction and accumulation for the ZnO nanoparticles (EC10, 20, 50 reproduction: 0.030, 0.049, 0.112 mg Zn/l) and the ZnCl2 (EC10, 20, 50 reproduction: 0.014, 0.027, 0.082 mg Zn/l). A large fraction of the nanoparticles rapidly dissolved after introduction in the exposure medium. Aggregation of nanoparticles was also observed but within 48 h of exposure most of these ZnO aggregates were dissolved. Based on the combined dissolution kinetics and toxicity results, it can be concluded that the toxicological effects of ZnO nanoparticles at the chronic level can be largely attributed to the dissolved fraction rather than the nanoparticles or initially formed aggregates.