An analytical strategy for the identification of carbamates, toxic alkaloids, phenobarbital and warfarin in stomach contents from suspected poisoned animals by thin-layer chromatography/ultraviolet detection

Abstract

In this study, an analytical strategy to identify brucine, strychnine, methomyl, carbofuran (alkaline compounds), phenobarbital, and warfarin (acid compounds) using thin-layer chromatography (TLC) screening with ultraviolet (UV) detection at 254 nm in stomach content is shown. The optimum mobile phase was found to be a chloroform: ethyl acetate: diethylamine (0.5:8.5:1) mixture for alkaline substances while a mixture of chloroform: acetone (9:1) has given better results for acidic substances. As for extraction, an equal proportion between distillated water and crude material (1:1) is required. For alkaline compounds, a filtration system was created in order to avoid any interferences from the biological matrix while for acidic compounds only centrifugation (4000 rpm/10 minutes) was required to obtain an appropriate sample. After the respective pretreatments, a one-step liquid–liquid extraction (LLE) has been employed for alkaline substances using a 3 mL of chloroform: ethyl ether (2:1) mixture for 2 min while acidic analytes used 3 mL of chloroform only during 5 min. For both methodologies described, the respective organic layers were dried down and re-suspended with 50 µL of methanol for further TLC plate application. The methodologies have been developed, successfully validated and applied to gastric contents from real case samples of suspected animal poisoning. Positive results from TLC/UV screening were confronted with HPLC-UV and confirmed by GC-MS.

Keywords:

• Pesticides
• stomach content
• thin-layer chromatography
• drugs
• animal intoxication

Acknowledgments

The authors would like to thank Fundo de Incentivo à Extensão (FIEX) for the scholarship provided to Diulia Oliveira Dias and also to the Veterinary Hospital of Santa Maria for the animal stomach contents provided for this study. Many thanks are also due to Dr. Thiago Belarmino de Souza for his valuable considerations and also to Dra. Tatiana Emanuelli for granting the HPLC-UV use.

Disclosure statement

No potential conflict of interest was reported by the authors.