Open Access Peer-reviewed Research Article

Comparative studies on enantioseparation of New Psychoactive Substances using cyclodextrin-assisted capillary electrophoresis with UV detection

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Eva-Maria Hubner
Patrick Steinkellner
Martin G. Schmid corresponding author


New psychoactive substances (NPS) count as psychoactive substances, which are slightly modified compared to illicit drugs regarding their chemical structure to circumvent law. Compared to classical drugs such as heroin, cocaine, or amphetamine, they show similar psychoactive effects, however, because of their novelty there is few knowledge about their side effects or toxicity. NPS are available as different chemical substance classes, among them chiral novel derivatives of amphetamine, cathinone, and ketamine. Since in most cases no clinical studies are available about the possibly different effects of the two enantiomers, there is a big demand for enantioseparation method development. Besides high-performance separation techniques such as gas chromatography or HPLC, capillary electrophoresis has turned out to be a powerful alternative for chiral separation development. The addition of chiral additives such as cyclodextrins to the background electrolyte often results in successful attempts. The present study compares the chiral separation power of different previously used non-charged ß-cyclodextrins, among them native ß-cyclodextrin as well as some of its derivatives such as acetyl-, and 2-hydroxypropyl-β-cyclodextrin, with the negatively charged derivatives carboxymethyl-, carboxyethyl- and succinyl-β-cyclodextrin by capillary zone electrophoresis. A total of 136 chiral NPS were investigated with these cyclodextrins, 122 of them were resolved in their enantiomers successfully by means of a simple electrolyte composition consisting of 10 mM aqueous sodium hydrogen phosphate buffer, pH 2.5 and 10 mM of the chiral selector. Furthermore, the presented method turned out to be useful to distinguish between positional isomers and examples for both enantiomer order and positional order for seized samples are given.

native β-cyclodextrin, acetyl-β-cyclodextrin, 2-hydroxypropyl-β-cyclodextrin, carboxymethyl-β-cyclodextrin, carboxyethyl-β-cyclodextrin, succinyl-β-cyclodextrin, novel psychoactive substances, chiral

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How to Cite
Hubner, E.-M., Steinkellner, P., & Schmid, M. (2022). Comparative studies on enantioseparation of New Psychoactive Substances using cyclodextrin-assisted capillary electrophoresis with UV detection. Journal of Pharmaceutical and Biopharmaceutical Research, 3(1), 187-205.


  1. Schmid MG. Optical Detection of NPS Internet Products via HPLC-DAD Systems: A Selective Review, in Light in Forensic Science: Issues and Applications. Royal Society of Chemistry, 2018, 301-332.
  2. EMCDDA. European Drug Report 2021, viewed December 22, 2021.
  3. United Nations 2013. The International Drug Control Conventions, viewed December 22, 2021.
  4. United Nations 1961. Drug Convention 1961, viewed December 22, 2021.
  5. United Nations 1972. “Psychotropic Convention 1972.” viewed December 22, 2021.
  6. Zhang YY, Li L, Ma AD, et al. Research Progress on Chiral Separation Methods and Toxic Effects of Methamphetamine Enantiomers. Fa Yi Xue Za Zhi, 2019, 35(3): 337-343.
  7. Nishimura T, Takahata K, Kosugi Y, et al. Psychomotor effect differences between l-methamphetamine and d-methamphetamine are independent of murine plasma and brain pharmacokinetics profiles. Journal of Neural Transmission, 2017, 124(5): 519-523.
  8. Jelen LA, Young AH and Stone JM. Ketamine: A tale of two enantiomers. Journal of Psychopharmacology, 2021, 35(2): 109-123.
  9. Ferris RM and Tang FL. Comparison of the effects of the isomers of amphetamine, methylphenidate and deoxypipradrol on the uptake of l-[3H]norepinephrine and [3H] dopamine by synaptic vesicles from rat whole brain, striatum and hypothalamus. The Journal of pharmacology and experimental therapeutics, 1979, 210(3): 422-428.
  10. Silva B, Palmeira A, Silva R, et al. S-(+)-Pentedrone and R-(+)-methylone as the most oxidative and cytotoxic enantiomers to dopaminergic SH-SY5Y cells: Role of MRP1 and P-gp in cathinones enantioselectivity. Toxicology and Applied Pharmacology, 2021, 416: 115442.
  11. Losacker M, Toennes SW, de Sousa Fernandes Perna EB, et al. Chiral Serum Pharmacokinetics of 4-Fluoroamphetamine after Controlled Oral Administration: Can (R)/(S)-Concentration Ratios Help in Interpreting Forensic Cases? Journal of Analytical Toxicology, 2021, 45(9): 985-992.
  12. Philogene-Khalid HL, Hicks C, Reitz AB, et al. Synthetic cathinones and stereochemistry: S enantiomer of mephedrone reduces anxiety- and depressant-like effects in cocaine- or MDPV-abstinent rats. Drug and Alcohol Dependence, 2017, 178: 119-125.
  13. Curry DW, Young MB, Tran AN, et al. Separating the agony from ecstasy: R(–)-3,4- methylenedioxymethamphetamine has prosocial and therapeutic-like effects without signs of neurotoxicity in mice. Neuropharmacology, 2018, 128: 196-206.
  14. Pitts EG, Curry DW, Hampshire KN, et al. (±)-MDMA and its enantiomers: potential therapeutic advantages of R(-)-MDMA. Psychopharmacology, 2018, 235(2): 377-392.
  15. Botanas CJ, Perez Custodio RJ, Kim HJ, et al. R-methoxetamine exerts rapid and sustained antidepressant effects and fewer behavioral side effects relative to S (+)-methoxetamine. Neuropharmacology, 2021, 193: 108619.
  16. Weiß JA, Pertl C and Schmid MG. Investigation of Two Seized Crystal Meth Labs in Austria – Parameters and Chirality Aspects of Methamphetamine Low Scale Synthesis. ARC Journal of Forensic Science, 2018, 3(1): 1-8.
  17. Losacker M, Z¨orntlein S, Schwarze B, et al. Determination of the enantiomeric composition of amphetamine, methamphetamine and 3,4-methylendioxy-N-methylamphetamine (MDMA) in seized street drug samples from southern Germany. Drug Testing and Analysis, 2021, 3118.
  18. Schmid MG and H¨agele JS. Separation of enantiomers and positional isomers of novel psychoactive substances in solid samples by chromatographic and electrophoretic techniques – A selective review. Journal of Chromatography A, 2020, 1624.
  19. Koster N, Clark CP and Kohler I. Past, present, and future developments in enantioselective analysis using capillary electromigration techniques. Electrophoresis, 2021, 42(1-2): 38-57.
  20. Hancu G, Papp LA, T´oth G, et al. The Use of Dual Cyclodextrin Chiral Selector Systems in the Enantioseparation of Pharmaceuticals by Capillary Electrophoresis: An Overview. Molecules, 2021, 26(8): 2261.
  21. Hancu G, Orlandini S, Papp LA, et al. Application of Experimental Design Methodologies in the Enantioseparation of Pharmaceuticals by Capillary Electrophoresis: A Review. Molecules, 2021, 26(15): 4681.
  22. Peluso P and Chankvetadze B. Native and substituted cyclodextrins as chiral selectors for capillary electrophoresis enantioseparations: Structures, features, application, and molecular modeling. Electrophoresis, 2021, 42(17-18): 1676-1708.
  23. Lurie IS, Klein RFX, Dal Cason TA, et al. Chiral Resolution of Cationic Drugs of Forensic Interest by Capillary Electrophoresis with Mixtures of Neutral and Anionic Cyclodextrins. Analytical Chemistry, 1994, 66(22): 4019-4026.
  24. Chinaka S, Tanaka S, Takayama N, et al. Simultaneous chiral analysis of methamphetamine and related compounds by capillary electrophoresis. Journal of Chromatography B: Biomedical Sciences and Applications, 2000, 749(1): 111-118.
  25. Lurie IS, Odeneal NG, McKibben TD, et al. Effects of various anionic chiral selectors on the capillary electrophoresis separation of chiral phenethylamines and achiral neutral impurities present in illicit methamphetamine. Electrophoresis, 1998, 19(16-17): 2918-2925.
  26. Rudaz S, Geiser L, Souverain S, et al. Rapid stereoselective separations of amphetamine derivatives with highly sulfated -cyclodextrin. Electrophoresis, 2005, 26(20): 3910-3920.
  27. Lanz M, Brenneisen R and Thormann W. Enantioselective determination of 3,4-methylenedioxymethamphetamine and two of its metabolites in human urine by cyclodextrin-modified capillary zone electrophoresis. Electrophoresis, 1997, 18(6): 1035.
  28. Huang YS, Liu JT, Lin LC, et al. Chiral separation of 3,4-methylenedioxymeth- amphetamine and related compounds in clandestine tablets and urine samples by capillary electrophoresis/fluorescence spectroscopy. Electrophoresis, 2003, 24(6): 1097-1104.
  29. Scarcella D, Tagliaro F, Turrina S, et al. Optimization of a simple method for the chiral separation of phenethylamines of forensic interest based on cyclodextrin complexation capillary electrophoresis and its preliminary application to the analysis of human urine and hair. Forensic Science International, 1997, 89(1-2): 33.
  30. Iwata YT, Inoue H, Kuwayama K, et al. Forensic application of chiral separation of amphetamine-type stimulants to impurity analysis of seized methamphetamine by capillary electrophoresis. Forensic Science International, 2006, 161(2-3): 92-96.
  31. Iio R, Chinaka S, Takayama N, et al. Simultaneous Chiral Analysis of Methamphetamine and Related Compounds by Capillary Electrophoresis/Mass Spectrometry Using Anionic Cyclodextrin. Analytical Sciences, 2005, 21(1): 15-19.
  32. Iwata YT, Garcia A, Kanamori T, et al. The use of a highly sulfated cyclodextrin for the simultaneous chiral separation of amphetamine-type stimulants by capillary electrophoresis. Electrophoresis, 2002, 23(9): 1328-1334.;2-Z
  33. Tagliaro F, Manetto G, Bellini S, et al. Simultaneous chiral separation of 3,4- methylenedioxymethamphetamine (MDMA), 3-4-methylenedioxyamphetamine (MDA), 3,4- methylenedioxyethylamphetamine (MDE), ephedrine, amphetamine and methamphetamine by capillary electrophoresis in uncoated and coated capillaries with native β-cyclodextrin as the chiral selector: Preliminary application to the analysis of urine and hair. Electrophoresis, 1998, 19(1): 42-50.
  34. Pascual-Caro S, Borrull F, Aguilar C, et al. Comparison of different chiral selectors for the enantiomeric determination of amphetamine-type substances in human urine by solid-phase extraction followed by capillary electrophoresis-tandem mass spectrometry. Electrophoresis, 2021.
  35. Burrai L, Nieddu M, Pirisi MA, et al. Enantiomeric Separation of 13 New Amphetamine-Like Designer Drugs by Capillary Electrophoresis, Using Modified Cyclodextrins. Chirality, 2013, 25(10): 617-621.
  36. Mohr S, Pilaj S and Schmid MG. Chiral separation of cathinone derivatives used as recreational drugs by cyclodextrin-modified capillary electrophoresis. Electrophoresis, 2012, 33(11): 1624-1630.
  37. Li L and Lurie IS. Regioisomeric and enantiomeric analyses of 24 designer cathinones and phenethylamines using ultra high performance liquid chromatography and capillary electrophoresis with added cyclodextrins. Forensic Science International, 2015, 254: 148-157.
  38. Merola G, Fu H, Tagliaro F, et al. Chiral separation of 12 cathinone analogs by cyclodextrin-assisted capillary electrophoresis with UV and mass spectrometry detection. Electrophoresis, 2014, 35(21-22): 3231-3241.
  39. Nowak PM, Olesek K, Wo´zniakiewicz M, et al. Simultaneous enantioseparation of methcathinone and two isomeric methylmethcathinones using capillary electrophoresis assisted by 2-hydroxyethyl- β-cyclodextrin. Electrophoresis, 2018, 39(19): 2406-2409.
  40. Taschwer M, Hofer MG and Schmid MG. Enantioseparation of benzofurys and other novel psychoactive compounds by CE and sulfobutylether β-cyclodextrin as chiral selector added to the BGE. Electrophoresis, 2014, 35(19): 2793-2799.
  41. Schutzner W and Fanali S. Enantiomers resolution in capillary zone electrophoresis by using cyclodextrins. Electrophoresis, 1992, 13(1): 687-690.
  42. H¨agele JS, Hubner EM and Schmid MG. Determination of the chiral status of different novel psychoactive substance classes by capillary electrophoresis and β-cyclodextrin derivatives. Chirality, 2020, 32(9): 1191-1207.
  43. H¨agele JS, Hubner EM and Schmid MG. Chiral separation of cathinone derivatives using β- cyclodextrin-assisted capillary electrophoresis–Comparison of four different β-cyclodextrin derivatives used as chiral selectors. Electrophoresis, 2019, 40(14): 1787-1794.