Screening of chlorpropamide in horse plasma by high-performance liquid chromatography with ultraviolet absorbance detection, and confirmation by gas chromatography-mass spectrometry.

The research article describes a method to detect and confirm the presence of a drug called chlorpropamide in horse plasma, using high-performance liquid chromatography, and gas chromatography-mass spectrometry. This forms a crucial part of antidoping control in the equine sports industry.

Methodology and Chromatographic Techniques

• The researchers developed a method specifically to screen chlorpropamide in plasma samples collected from horses.
• The plasma sample of 1 milliliter was extracted with a chemical solvent named dichloromethane for analysis.
• High-Performance Liquid Chromatography (HPLC), a technique used to separate, identify and quantitate each component in a mixture, was used to screen the samples.
• The presence of chlorpropamide was then confirmed using another technique called Gas Chromatography-Mass Spectrometry (GC-MS). This technique is particularly effective in identifying individual substances within a test sample.

Detection Limit

• The research found that the detection limit for chlorpropamide was 3.5 nanograms per milliliter, at a signal-to-noise ratio of three. The signal-to-noise ratio is a measure that compares the level of a desired signal to the level of background noise.

Derivatization and Analysis

• Derivatization, a technique used in chemistry to transform a chemical compound into a product of similar chemical structure, of Chlorpropamide was carried out using N,O-bis-(trimethylsilyl)trifluoroacetamide with 1% trimethylchlorosilane. This process allowed greater stability, accuracy, and sensitivity in GC-MS analysis.
• Subsequently, plasma samples collected after the administration of Diabinese (a trade-name for Chlorpropamide) returned positive results for the presence of chlorpropamide, up to five days after administration, in all samples analysed.

Significance

• The research is significant as the detection of drugs like chlorpropamide in equine sports is crucial for ensuring fair competition and the welfare of the horses. The development of this reliable detection method adds to the set of available methods to ensure compliance with anti-doping regulations.