Determination of butorphanol in horse race urine by immunoassay and gas chromatography-mass spectrometry.

This research was aimed at developing a method to detect the presence of butorphanol, a pain-relieving drug, in the urine samples from race horses. The study used ELISA kits for screening and gas chromatography-mass spectrometry (GC-MS) for absolute confirmation.

Methodology

• The research began by conducting an enzymatic hydrolysis on urine samples that were 5 ml in size.
• The samples were then extracted via solid-phase extraction.
• The residues were evaporated, derivative and injected into a gas chromatography-mass spectrometry (GC-MS) system for further analysis.

Findings

• The ELISA tests carried out on 20 microliters of each sample were able to detect the presence of butorphanol up to 104 hours after intramuscular administration of 8 mg of Torbugesic (a branded form of butorphanol).
• The GC-MS method could detect the drug up to 24 hours in a full scan mode or up to 31 hours in Selected Ion Monitoring (SIM) mode.

Validation of GC-MS method in SIM mode

• Nalbuphine was used as an internal standard for the validation of the GC-MS method in SIM mode.
• This validation method recorded linearity studies spanning from 10 to 250 ng per milliliter.
• The recovery rate for the method was consistently close to 100%, demonstrating a high level of accuracy.
• In intra-assay precision was between 4.1% and 14.9% and inter-assay precision varied between 9.3% and 45.1%.
• The method showed stability for 10 days.
• Both the limit of detection and limit of quantitation for the method were recorded at 10 ng/ml and 20 ng/ml respectively.

In conclusion, the research has provided an effective method to test for butorphanol in urine samples from racehorses. This could be useful to enforce strict drug rules in sporting events.