A simple and isocratic protein-based high performance liquid chromatography method for the enantioseparation of amphetamine derivatives

Abstract

New amphetamine derivatives, a substance class belonging to the New Psychoactive Substances (NPS), have fundamentally changed the global drug market during the last years. Also known as “legal highs” they are sold under various names like "bath salts", "air fresheners" and so forth. Structures of well-known illicit drugs such as amphetamine, methamphetamine and 3,4-methylenedioxyamphetamine are slightly modified to circumvent law and samples are traded worldwide via various internet shops. Many of these new derivatives possess a chiral centre and the pharmacological differences of the two enantiomers are not yet investigated for many of them. This makes the continuous development of analytical methods for their chiral separation crucial.The aim of this study was to develop a simple, isocratic HPLC-UV method for enantioseparation and to test this method on a set of 26 amphetamine derivatives, including latest products on the drug market. A Daicel Chiralpak® CBH 150×3.0 mm column (5 µm particle size) served for chiral separation. The influence of the organic modifier and type, molarity and pH of the buffer on separation results was investigated. UV detection was performed at 210 and 254 nm. As final mobile phase, a mixture of 5 mM sodium dihydrogen phosphate buffer (pH = 6.9) / methanol (85 : 15 v/v) was chosen. Under these conditions, all 26 tested amphetamines were separated successfully into their enantiomers. Thus, the results showed that all analytes were traded as racemic mixtures.