Fast and sensitive method for the diagnosis and follow-up of anticoagulant rodenticides poisoning in animal whole blood.

Rodent control strategies are primarily based on the use of anticoagulant rodenticides (ARs), making them widely used worldwide. However, due to their high toxicity and availability, ARs are among the leading causes of animal poisoning in Europe. They are the primary agents involved in intoxication in cats and the second in dogs. Additionally, their long persistence in the body can lead to secondary exposure, particularly in wild predators. The laboratory findings and clinical signs of intoxication can range from increased clotting time (prolonged prothrombin time and activated partial thromboplastin time) to severe bleeding and death. Despite the prevalence and severity of this intoxication, only a few methods are available for the identification and quantification of ARs in animals, and most of them are suitable only for post-mortem diagnosis. In this study, we present the validation of a rapid and sensitive method for the identification and quantification of ARs in animal whole blood, using a small sample volume. The developed LC-MS/MS method demonstrated high accuracy and precision at the limit of quantification (LOQ), as well as at low, medium, and high concentrations. It exhibited higher sensitivity (LOQ 0.1 - 0.3 ng/mL) compared to previously published methods. After validation, the method was successfully applied to real cases of suspected poisoning events, resulting in the identification of several positive samples. The examples presented in this study highlight the utility of this method for diagnosis and follow-up, emphasizing the importance of method sensitivity in order to avoid misclassifying truly positive samples as negative.

Analysis of Metaldehyde in Animal Whole Blood and Serum by Gas Chromatography-Mass Spectrometry.

Metaldehyde, a widely used molluscicide, is the third cause of intoxication by pesticides in domestic animals in Europe. Most mammalian species are susceptible, and its exposure may lead to death within a few hours. While metaldehyde intoxication diagnosis is in most cases presumptive, based on the symptomatology or from "postmortem" analysis, few analytical methods are currently available for live animals. The aim of this work was to describe a fast analytical method for the specific and quantitative determination of metaldehyde in animal whole blood and serum at concentrations of toxicological significance. A liquid-liquid extraction with chloroform and gas chromatography-mass spectrometry quantification are proposed. The method limit of quantification (LOQ) was 0.04 µg/mL in serum and whole blood. The method was linear in the range from 0.04 to 200 µg/mL. The recovery was between 93% and 102% for LOQ, low, medium and high spike concentrations. Intra- and inter-assay relative standard deviation was <12% in all spike concentrations in both serum and whole blood, apart from one of the experiments at LOQ in whole blood, which accounted for 17.7%. The method was applied to real intoxication cases, and the concentration found in positive samples was between 29 and 69 µg/mL. The proposed method provides high sensitivity, accuracy and precision and can be used to assist in the diagnosis of metaldehyde poisoning.