Laboratory of the Government Chemist, Queens Road, Teddington, Middlesex., TW11 OLY United Kingdom

Stage 1 was designed to determine the effects of temperature, time, container type and sterility of the urine sample. The experimental design was based on a Taguchi L16 matrix mirrored to reflect the time variable requiring eight levels.

The second series of experiments examined the effects of light, radiation, agitation, storage under nitrogen and the inclusion of a preservative based on a full factorial design with five factors, each at two levels. For these variables, sterile urine samples were stored in a single container type, at 25°C for seven days prior to analysis.

The concentration of the free and conjugated testosterone and epitestosterone were determined by GC-MS using deuterium labelled internal standards after hydrolysis, C8 solid phase extraction followed by liquid-liquid clean up and enol-derivatisation.

The change in these concentrations and ratios are important as unsuitable storage conditions may result in a testosterone/epitestosterone ratio that may exceed the International Olympic Committee level of 6:1 and result in a ban or legal action.