Extraction of Free Testosterone from Equine Hair on the Bead Ruptor Elite

Hair samples are commonly processed for forensic analysis. Hair trace analysis has been a major tool for forensic scientists for decades. Hair samples can be analyzed for the presence of performance enhancing drugs such as hormones and steroids. Testosterone for example, can be abused by athletes due to its ability to enhance muscle development, strength and endurance. The use of performance enhancing drugs, including testosterone, is banned in a majority of sports and scientists are investigating methods in the detection of testosterone from hair. 

A key component in creating an experimental workflow that will support the detection of testosterone from hair is the front end sample preparation method. Hair samples are a challenging sample matrix as hair is relatively resistant to disaggregation, hair tends to cling to vessel surfaces and once in powder form is easily aerosolized. An ideal mechanical method for hair disaggregation would have sufficient force to disrupt the target samples, would be compatible with a sealed tube and with the common organic solvent based extraction buffers. Historically, hair disruption methods have included mortar and pestle, cryogrinding and chemical digestion. Although these techniques are adequate, they do have their draw backs. The traditional mortar and pestle is labor and time demanding, cryogrinding is inherently low throughout and only efficient when using small sample sizes and chemical digestion is not only time consuming but uses hazardous acids and bases. Bead milling has recently been demonstrated to be an effective method for disruption of hair and fulfills all the ideal processing requirements. Bead milling is achieved through the high speed shaking of a sample in a sealed tube in the presence of small beads. The high force impacts of the beads against the sample reduces the sample size.

Herein, we evaluate the potential for the extraction of free testosterone from equine hair samples on the Omni Bead Ruptor Elite™ bead mill homogenizer coupled with sonication using the Omni Sonic Ruptor 400 ultrasonic homogenizer.

Figure 2: Testosterone Standard Curve Measured at 450 nm