Detection of testosterone propionate administration in horse hair samples

A sensitive and specific method has been developed to detect semi-quantitatively testosterone in horse hair samples. The method involved a washing step with sodium dodecylsulfate aqueous solution. The mane and tail hair samples (100 mg) were dissolved in 1 mL of sodium hydroxide for 15 min at 95 °C in the presence of d3-boldenone used as internal standard. The next three steps involved diethyl ether extraction and a solid phase extraction on Isolute C18 (EC) cartridges eluted with methanol. The residue was derivatized by adding 100 μL of acetonitrile and 30 μL of PFPA then incubating for 15 min at 60 °C. After evaporation, 30 μL of hexane was added and 2.5 μL was injected into the column (a bonded phase fused silica capillary column DB5MS, 30 m × 0.25 mm i.d. × 0.25 μm film thickness) of a Trace GC chromatograph. In order to improve the sensitivity of the method, damping gas flow has been optimized. Testosterone was identified in MS2 full scan mode on the Polaris Q instrument. The assay was capable of detecting less than 1 pg mg−1. The recovery was close to 90%. The analysis of tail and mane samples collected from a gelding horse having received a single dose of testosterone propionate (1 mg kg−1) showed the presence of testosterone in the range of 1–6 pg mg−1 in hair collected during 5 months after administration.