The human population can be exposed to copper oxides due to their use as plant protection products and feed additives. They are commercially available as powders or formulations and are broadly used. However, limited information on their particle size is available, which in turn can influence other physicochemical properties/endpoints such as dissolution and surface reactivity. For particulate materials applied in food and feed, the EFSA Guidance on Particle-Technical Requirements requires measuring the particle size distribution to assess whether a nano-specific risk assessment should be performed. In this work, sample preparation and image analysis of copper oxide particles applied as feed additive and/or as plant protection product were optimised and combined for particle size measurement using electron microscopy-based methods.

A total of 20 copper oxide materials, including five feed additives, one pesticide, and 14 benchmark materials (including four labelled as “micrometer-sized particles”), were sourced from the European market. The properties of the copper oxide particles, including size, shape, agglomeration state, and elemental composition, were determined using (scanning) transmission electron microscopy ((S)TEM) and energy-dispersive X-ray spectroscopy (EDX). Three marketed copper oxide materials were selected for further testing: two feed additives and one pesticide. To prepare dispersions for quantitative size measurement of the constituent particles of the selected copper oxide materials by (S)TEM, the conditions proposed in the generic Nanogenotox protocol were evaluated. Descriptive EM analysis was performed to assess whether notable differences in the agglomerate size and number of particles per grid surface existed between ethanol pre-treated and non pre-treated samples. In addition, the effects of increasing sonication energies on dispersions prepared in distilled water containing 0.05%BSA solution were investigated. To sample a broad size range, from nano to micro-sized particles in a representative manner, several EM-specimen preparation methods were evaluated. Normalized number-based distributions and descriptive statistics were determined for minimum and maximum Feret diameters and for the aspect ratio.

The obtained results are a first step in addressing some of the current data gaps identified in the physicochemical characterization needed for hazard assessment of copper oxides applied in the food chain.