Fast and sensitive analysis of dermorphin and HYP ⁶ ‐dermorphin in equine plasma using liquid chromatography tandem mass spectrometry

Dermorphin and HYP⁶‐dermorphin are hepta‐ peptides and natural opioids originally isolated from the skin of South American frogs. They are more potent than morphine but less likely to produce drug tolerance and addiction. These properties make them ideal candidates for the doping of racehorses to enhance performance during competition. Dermorphin was recently classified as a Class I drug by Racing Commissioners International (RCI), indicating that it is a banned substance in equine athletes. To enforce this ban, a fast and sensitive method was developed for dermorphin and HYP⁶‐dermorphin analysis in equine plasma. Equine plasma (2 ml) was extracted on a mixed mode cation exchange solid‐phase column. After extraction, dermorphin and HYP⁶‐dermorphin were separated and detected using a liquid chromatography (LC) triple quadrupole linear ion trap mass spectrometry in positive multiple‐reaction‐monitoring (MRM) mode. Each analysis was 3.5 min. Four MRM transitions were used for identification of each compound. The extraction procedure was efficient and the limits of detection (LOD) were 2 pg/ml and 10 pg/ml for dermorphin and HYP⁶‐dermorphin, respectively. The method has good selectivity and precision. Results of stability studies showed that both analytes were stable at low temperature. This is the first report of dermorphin and HYP⁶‐dermorphin analysis in equine plasma, which could be adopted as a regular screening or confirmation method for controlling the abuse of these compounds in equine sports. Copyright © 2013 John Wiley & Sons, Ltd.