Detection of non-steroidal anti-inflammatory drugs in equine plasma and urine by gas chromatography-mass spectrometry.

This research outlines a method for detecting the presence of seventeen different types of non-steroidal anti-inflammatory drugs (NSAIDs) in horse urine and blood plasma samples using gas chromatography-mass spectrometry (GC-MS).

Methodology

The researchers employed a multi-step process involving:

• The extraction of NSAIDs from biological materials, achieved through the use of diethyl ether in an acidic environment (pH 2-3).
• These ethereal extracts were then treated with certain solutions. Urine samples were washed with a saturated solution of sodium hydrogencarbonate whilst plasma samples were treated with a solid blend of sodium carbonate and sodium hydrogencarbonate.
• Following treatment, the ethereal extracts were dried and then derivatized. This was achieved through incubation at 60 degrees Celsius using methyl iodide in acetone, in the presence of solid potassium carbonate.
• This process resulted in either mono- or bismethyl derivatives of the NSAIDs.

Optimization and Screening

To ensure comprehensive methylation of all compound samples, the researchers conducted kinetic studies and determined that an incubation period of 90 minutes was optimal for screening purpose. For individual confirmation analyses, shorter incubation times sufficed.

The final stage of the process, the chromatographic analysis of the derivatives, was carried out using GC-MS and had a run time of 13 minutes.

Results

They reported that:

• In general, extraction recoveries ranged from 23.3% to 100% for plasma and from 37.5% to 83.8% for urine samples.
• Detection limits ranged from less than 5 to 25 ng/ml for both plasma and urine samples using selected-ion monitoring.

Utility and Application

The procedure was then applied successfully for the routine doping control of horse samples, allowing for screening and confirmation of NSAIDs use. This has practical implications for equine competitive sports in regards to veterinary health regulations and anti-doping rules.

Overall, this research presents a robust and effective strategy for the detection of NSAIDs in equine samples, pointing to its potential use in doping control.