Determination of amphetamine derivatives (ecstasy) in urine which is commonly used among young population by GC/MS

Introduction: The illegal use of synthetic substances has become an increasing global problem resulting in significant public health and legal problems in the last decades. Amphetamines (AMPs) are a group of synthetic psychoactive drugs. The consumption of AMPs have increased significantly over the past years due to the easy availability and low cost. The majority of methods for the determination of AMPs in biological samples are based on gas chromatography–mass spectrometry (GC-MS). The aim of the present study is to develop and fully validate a simple, reliable and fast GC-MS method for quantification of the amphetamines in urine. A simple liquid-liquid extraction (LLE) was used as sample clean up procedure and the validated method was applied to authentic urine samples. Material and Methods: The study was developed in BATI institute and blank urine samples were obtained from drug-free volunteers in institute workers. Authentic urine samples were obtained from forensic and toxicology cases. Urine samples were processed by a LLE procedure as follows. An aliquot (100 μg/L) of the standard solution was added to 2.0 mL of urine. Basic hydrolysis is achieved by adding 1 mL of KOH to the solution. The urine sample is transferred to Toxitube-A (Varian, Inc) and the sample is shaken for 5 minutes. After centrifugation at 4100 rpm for 5 minutes, the organic upper layer is transferred to a clean tube and evaporated to dryness under a stream of nitrogen at 40 ◦C. The samples are reconstituted using ethyl acetate and derivitizated using Pentafluoropropionic Anhydride (PFPA) prior to GC-MS analysis. Results: The method was validated for selectivity, linearity, LOQ, LOD, imprecision and recovery parameters. The applied chromatographic method ensured the elution of AMP within a retention time of 5.59 min and produced peaks of acceptable symmetry. Quantification was performed by integration of the area under the specific chromatograms in reference to the integrated area of the deuterated analogue. Selectivity of the method was achieved by a combination of retention time, precursor and product ions (91, 118, 190). LOD and LOQ were values were found to be 5.6 and 18.8, respectively. The linearity was tested up to 1000 μg/L with an R2 > 0.9996. Conclusion: A rapid and sensitive method was validated for the determination of amphetamine in urine by GC-MS after a LLE. The method was also successfully applied to authentic samples from amphetamines users.