ELISA screening with GC-MS confirmation of the tranquilizer chlorprothixene administered in subtherapeutic doses to horses.

This research investigates the detection of the tranquilizer chlorprothixene (CPT) in horses’s urine after the drug’s administration in sub-therapeutic doses, using an enzyme-linked immunosorbent assay (ELISA) screening and a gas chromatography-mass spectrometry (GC-MS) confirmation. It was found that the presence of CPT or its metabolites in horse urine can be readily detected for up to 24 hours using the ELISA test, and chemical ionization mass spectrometry showed the presence of at least four metabolites.

Methodology

• A commercially available generic promazine ELISA kit was used, which exhibits cross-reactivity with CPT.
• Sub-therapeutic doses (4.5 mg) of chlorprothixene were administered intravenously (i.v.) and intramuscularly (i.m.) to three horses.
• The ELISA test was used to detect the presence of the tranquilizer or its metabolites in the horses’ urine for up to 24 hours after administration.
• The GC-MS method was used for the confirmation of the presence of CPT and its metabolites.

Results

• The ELISA test readily detected CPT or its metabolites in equine urine up to 24 hours after administration.
• Maximum concentrations of CPT were observed 2 hours after i.v. administration.
• No distinct peak values were observed after i.m. administration.
• Confirmation of CPT and its metabolites by electron impact mass spectrometry after sub-therapeutic administration was not successful.
• Chemical ionization mass spectrometry, however, revealed the presence of at least four metabolites including chlorprothixene sulphoxide, hydroxylated chlorprothixene and hydroxylated chlorprothixene sulphoxide.

Conclusion

• The study successfully demonstrates the use of the commercially available generic promazine ELISA kit in detecting the sub-therapeutic administration of chlorprothixene in horses.
• The ELISA test is seen to be effective in detecting CPT or its metabolites in equine urine for up to 24 h after administration.
• Despite the failure of electron impact mass spectrometry to confirm the presence of CPT and its metabolites, chemical ionization mass spectrometry was successful in this.
• This means, the method of testing proposed in this study could prove beneficial for detecting and confirming the sub-therapeutic administration of CPT in horses, important for ensuring equine welfare and fair horse racing.