Correlation between inhibition of lung surfactant function in vitro and rapid reduction in tidal volume following exposure to plant protection products in mice

Currently, testing of acute inhalation toxicity in animals is required for regulation of pesticide active ingredients and formulated plant protection products. The main outcome of the regulatory tests is "lethal concentration 50″ (LC 50), i.e. the concentration that will kill 50% of the exposed animals. However, ongoing work aims to identify New Approach Methods (NAMs) to replace animal experiments. To this end, we studied 11 plant protection products, sold in the European Union (EU), for their ability to inhibit lung surfactant function in vitro in the constrained drop surfactometer (CDS). In vivo, inhibition of lung surfactant function can lead to alveolar collapse and reduction of tidal volume. Therefore, we also assessed changes in breathing patterns of mice during exposure to the same products. Six of the eleven products inhibited lung surfactant function, and six products reduced tidal volume in mice. In vitro inhibition of lung surfactant function predicted reduction in tidal volume in exposed mice with a sensitivity of 67% and a specificity of 60%. Two products were labelled as "harmful if inhaled", both inhibited surfactant function in vitro and reduced tidal volume in mice. Lung surfactant function inhibition in vitro predicted reduction in tidal volume for plant protection products to a lesser degree than for previously tested substances. This could owe to the requirement for rigorous testing of plant protection products prior to approval that might have selected against substances that could potentially inhibit lung surfactant, e.g. due to severe adverse effects during inhalation.