Analytical Methods Used for Identification and Determination of Synthetic Cathinones and Their Metabolites

  • Dariusz Zuba
  • Piotr Adamowicz
Part of the Current Topics in Neurotoxicity book series (Current Topics Neurotoxicity, volume 12)


Synthetic cathinones are a broad group of chemicals, which have very similar structure. Many substances seized from the drug market are isomers, homologues, analogues, etc. They follow similar metabolism processes, and often convert in the human body to the same metabolites; some being also cathinone derivatives. Moreover, active doses differ significantly among drugs, causing that the levels of cathinones and their metabolites in the body fluids and tissues are in a broad range, from low ng/mL to hundreds mg/L. A variety of analytical techniques is applied to identify synthetic cathinones in seized drugs. Unequivocal identification of an active substance is crucial, especially in countries with individual drug control system, as the legal consequences for, e.g. possession of different isomers may vary substantially. Gas chromatography–mass spectrometry (GC-MS) is the most commonly method used for preliminary identification in forensic laboratories, but it has to be supported by other techniques, e.g. by Fourier-transformed infrared spectrometry (FTIR), nuclear magnetic resonance (NMR) and liquid chromatography with different detectors, mainly tandem mass spectrometers. Synthetic cathinones are analysed in different biological matrices, including blood, serum, plasma, dried blood spots, urine, hair, oral fluid and postmortem body tissues. Samples are prepared for the analysis, e.g. by dilution, precipitation, liquid–liquid extraction (LLE) and solid phase extraction (SPE). Enzyme hydrolysis (especially for urine), washing out or digestion (for hair), and derivatization are also included in some procedures. Triple quadrupole LC-MS/MS systems are the most frequently used. Many analytical challenges cause that more sophisticated techniques, including liquid chromatography-high-resolution mass spectrometry (LC-HRMS), are increasingly applied in the analysis of biological materials for the identification and quantitation of cathinones.


Synthetic cathinones Seized material Biological material Analytical methods GC-MS LC-MS HPLC-DAD 



Attenuated total reflectance technique


Capillary electrophoresis


Chemical ionization


Direct analysis in real time-mass spectrometry


Diode array detection


Dried blood spots


Desorption electrospray ionization-mass spectrometry


Dispersive liquid–liquid microextraction


Electron impact ionization


Enzyme-linked immunosorbent assay


European Network of Forensic Science Institutes


Electrospray ionization


Fourier-transformed infrared spectrometry


Gas chromatography


Gas chromatography–electron impact–mass spectrometry


Gas chromatography with flame ionization detector


Gas chromatography–mass spectrometry


High-performance liquid chromatography with diode array detection


High-resolution mass spectrometry


Ion mobility spectrometry


Infrared spectroscopy


Liquid chromatography


Liquid chromatography-high resolution mass spectrometry


Liquid chromatography-high resolution multiple mass spectrometry


Liquid chromatography–mass spectrometry


Liquid chromatography-tandem mass spectrometry


Liquid chromatography-electrospray ionization-quadrupole time-of-flight mass spectrometry


Liquid chromatography-quadrupole time-of-flight mass spectrometry


Liquid–liquid extraction


Limit of detection


Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry


Multiple reaction monitoring


Mass spectrometry


Tandem mass spectrometry


Near infrared


Nuclear magnetic resonance


Miniaturized solid phase extraction


Protein precipitation


Triple quadrupole


Hybrid quadrupole time-of-flight analyser


Salting out liquid–liquid extraction


Selected ion monitoring


Supported liquid extraction


Solid phase extraction


Solid phase microextraction


Scientific Working Group for the Analysis of Seized Drugs


RapidFire tandem mass spectrometry


Ultra high-performance liquid chromatography with diode array detection


Ultra high-performance liquid chromatography–mass spectrometry


Ultraviolet/visible spectroscopy


Volumetric absorptive microsampling


  1. Adamowicz P, Tokarczyk B (2016) Simple and rapid screening procedure for 143 new psychoactive substances by liquid chromatography-tandem mass spectrometry. Drug Test Anal 8(7):652–667. Scholar
  2. Al-Saffar Y, Stephanson NN, Beck O (2013) Multicomponent LC-MS/MS screening method for detection of new psychoactive drugs, legal highs, in urine—experience from the Swedish population. J Chromatogr B 930:112–120. Scholar
  3. Alvarez JC, Etting I, Abe E et al (2017) Identification and quantification of 4-methylethcathinone (4-MEC) and 3,4-methylenedioxypyrovalerone (MDPV) in hair by LC-MS/MS after chronic administration. Forensic Sci Int 270:39–45. Scholar
  4. Amaratunga P, Lorenz Lemberg B, Lemberg D (2013) Quantitative measurement of synthetic cathinones in oral fluid. J Anal Toxicol 37(9):622–628. Scholar
  5. Ambach L, Hernández Redondo A, König S et al (2014) Rapid and simple LC-MS/MS screening of 64 novel psychoactive substances using dried blood samples. Drug Test Anal 6(4):367–375. Scholar
  6. Ambach L, Redondo AH, König S et al (2015) Detection and quantification of 56 new psychoactive substances in whole blood and urine by LC-MS/MS. Bioanalysis 7(9):1119–1136. Scholar
  7. Ammann D, McLaren JM, Gerostamoulos D et al (2012) Detection and quantification of new designer drugs in human blood: part 2—designer cathinones. J Anal Toxicol 36(6):381–389. Scholar
  8. Armenta S, Garrigues S, de la Guardia M et al (2015) Detection and characterization of emerging psychoactive substances by ion mobility spectrometry. Drug Test Anal 7(4):280–289. Scholar
  9. Baciu T, Borrull F, Calull M et al (2016) Enantioselective determination of cathinone derivatives in human hair by capillary electrophoresis combined in-line with solid-phase extraction. Electrophoresis 37(17-18):2352–2362. Scholar
  10. Bell C, George C, Kicman AT et al (2011) Development of a rapid LC-MS/MS method for direct urinalysis of designer drugs. Drug Test Anal 3(7–8):496–504. Scholar
  11. Bertol E, Mari F, Boscolo Berto R et al (2014) A mixed MDPV and benzodiazepine intoxication in a chronic drug abuser: determination of MDPV metabolites by LC-HRMS and discussion of the case. Forensic Sci Int 243:149–155. Scholar
  12. Concheiro M, Anizan S, Ellefsen K et al (2013) Simultaneous quantification of 28 synthetic cathinones and metabolites in urine by liquid chromatography-high resolution mass spectrometry. Anal Bioanal Chem 405:9437–9448. Scholar
  13. Concheiro M, Castaneto M, Kronstrand R et al (2015) Simultaneous determination of 40 novel psychoactive stimulants in urine by liquid chromatography-high resolution mass spectrometry and library matching. J Chromatogr A 1397:32–42. Scholar
  14. de Castro A, Lendoiro E, Fernández-Vega H et al (2014) Liquid chromatography tandem mass spectrometry determination of selected synthetic cathinones and two piperazines in oral fluid. Cross reactivity study with an on-site immunoassay device. J Chromatogr A 1374:93–101. Scholar
  15. Ellefsen KN, Anizan S, Castaneto MS et al (2014) Validation of the only commercially available immunoassay for synthetic cathinones in urine: randox drugs of abuse V biochip array technology. Drug Test Anal 6(7-8):728–738. Scholar
  16. Ellefsen KN, Concheiro M, Huestis MA (2016) Synthetic cathinone pharmacokinetics, analytical methods, and toxicological findings from human performance and postmortem cases. Drug Metab Rev 48(2):237–265. Scholar
  17. Glicksberg L, Bryand K, Kerrigan S (2016) Identification and quantification of synthetic cathinones in blood and urine using liquid chromatography-quadrupole/time of flight (LC-Q/TOF) mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 1035:91–103. Scholar
  18. Grapp M, Kaufmann C, Ebbecke M (2017) Toxicological investigation of forensic cases related to the designer drug 3,4-methylenedioxypyrovalerone (MDPV): detection, quantification and studies on human metabolism by GC-MS. Forensic Sci Int 273:1–9. Scholar
  19. Gwak S, Arroyo-Mora LE, Almirall JR (2015) Qualitative analysis of seized synthetic cannabinoids and synthetic cathinones by gas chromatography triple quadrupole tandem mass spectrometry. Drug Test Anal 7(2):121–130. Scholar
  20. Hong WY, Ko YC, Lin MC et al (2016) Determination of synthetic cathinones in urine using gas chromatography-mass spectrometry techniques. J Anal Toxicol 40(1):12–16. Scholar
  21. Johnson RD, Botch-Jones SR (2013) The stability of four designer drugs: MDPV, mephedrone, BZP and TFMPP in three biological matrices under various storage conditions. J Anal Toxicol 37(2):51–55. Scholar
  22. Joshi M, Cetroni B, Camacho A et al (2014) Analysis of synthetic cathinones and associated psychoactive substances by ion mobility spectrometry. Forensic Sci Int 244:196–206. Scholar
  23. Kerrigan S, Savage M, Cavazos C et al (2016) Thermal degradation of synthetic cathinones: implications for forensic toxicology. J Anal Toxicol 40(1):1–11. Scholar
  24. LaPointe J, Musselman B, O’Neill T et al (2015) Detection of “bath salt” synthetic cathinones and metabolites in urine via DART-MS and solid phase microextraction. J Am Soc Mass Spectrom 26(1):159–165. Scholar
  25. Lendoiro E, Jiménez-Morigosa C, Cruz A et al (2017) An LC-MS/MS methodological approach to the analysis of hair for amphetamine-type-stimulant (ATS) drugs, including selected synthetic cathinones and piperazines. Drug Test Anal 9(1):96–105. Scholar
  26. Liu C, Jia W, Li T et al (2017) Identification and analytical characterization of nine synthetic cathinone derivatives N-ethylhexedrone, 4-Cl-pentedrone, 4-Cl-α-EAPP, propylone, N-ethylnorpentylone, 6-MeO-bk-MDMA, & α-PiHP, 4-Cl-α-PHP, and 4-F-α-PHP. Drug Test Anal 9(8):1162–1171. Scholar
  27. Marinetti LJ, Antonides HM (2013) Analysis of synthetic cathinones commonly found in bath salts in human performance and postmortem toxicology: Method development, drug distribution and interpretation of results. J Anal Toxicol 37(3):135–146. Scholar
  28. Mayer M, Benko A, Huszár A et al (2013) Simultaneous determination of 4-substituted cathinones (4-MMC, 4-MEC and 4-FMC) in human urine by HPLC-DAD. J Chromatogr Sci 51(9):861–866. Scholar
  29. Mercolini L, Protti M, Catapano MC et al (2016) LC-MS/MS and volumetric absorptive microsampling for quantitative bioanalysis of cathinone analogues in dried urine, plasma and oral fluid samples. J Pharm Biomed Anal 123:186–194. Scholar
  30. Meyer MR, Maurer HH (2016) Review: LC coupled to low- and high-resolution mass spectrometry for new psychoactive substance screening in biological matrices—where do we stand today? Anal Chim Acta 927:13–20. Scholar
  31. Meyer MR, Du P, Schuster F et al (2010a) Studies on the metabolism of the α-pyrrolidinophenone designer drug methylenedioxypyrovalerone (MDPV) in rat and human urine and human liver microsomes using GC-MS and LC-high-resolution MS and its detectability in urine by GC-MS. J Mass Spectrom 45:1426–1442. Scholar
  32. Meyer MR, Wilhelm J, Peters FT et al (2010b) Beta-keto amphetamines: studies on the metabolism of the designer drug mephedrone and toxicological detection of mephedrone, butylone, and methylone in urine using gas chromatography-mass spectrometry. Anal Bioanal Chem 397(3):1225–1233. Scholar
  33. Miller B, Kim J, Concheiro M (2017) Stability of synthetic cathinones in oral fluid samples. Forensic Sci Int 274:13–21. Scholar
  34. Milman BL (2011) Chemical identification and its quality assurance. Springer, Berlin Heidelberg.
  35. Montesano C, Vannutelli G, Gregori A et al (2016) Broad screening and identification of novel psychoactive substances in plasma by high-performance liquid chromatography-high-resolution mass spectrometry and post-run library matching. J Anal Toxicol 40(7):519–528. Scholar
  36. Montesano C, Vannutelli G, Piccirilli V et al (2017) Application of a rapid μ-SPE clean-up for multiclass quantitative analysis of sixteen new psychoactive substances in whole blood by LC-MS/MS. Talanta 167:260–267. Scholar
  37. Musah RA, Cody RB, Domin MA et al (2014) DART-MS in-source collision induced dissociation and high mass accuracy for new psychoactive substance determinations. Forensic Sci Int 244:42–49. Scholar
  38. Namera A, Kawamura M, Nakamoto A et al (2015) Comprehensive review of the detection methods for synthetic cannabinoids and cathinones. Forensic Toxicol 33(2):175–194. Scholar
  39. Neifeld JR, Regester LE, Holler JM et al (2016) Ultrafast screening of synthetic cannabinoids and synthetic cathinones in urine by RapidFire-Tandem Mass Spectrometry. J Anal Toxicol 40(5):379–837. Scholar
  40. Nie H, Li X, Hua Z et al (2016) Rapid screening and determination of 11 new psychoactive substances by direct analysis in real time mass spectrometry and liquid chromatography/quadrupole time-of-flight mass spectrometry. Rapid Commun Mass Spectrom 30(S1):141–146. Scholar
  41. Odoardi S, Fisichella M, Romolo FS et al (2015) High-throughput screening for new psychoactive substances (NPS) in whole blood by DLLME extraction and UHPLC-MS/MS analysis. J Chromatogr B Analyt Technol Biomed Life Sci 1000:57–68. Scholar
  42. Ojanpera IA, Heikman PK, Rasanen IJ (2011) Urine analysis of 3,4-methylenedioxypyrovalerone in opioid-dependent patients by gas chromatography–mass spectrometry. Ther Drug Monit 33(2):257–263. Scholar
  43. Paul M, Ippisch J, Herrmann C et al (2014) Analysis of new designer drugs and common drugs of abuse in urine by a combined targeted and untargeted LC-HR-QTOFMS approach. Anal Bioanal Chem 406(18):4425–4441. Scholar
  44. Pederson CG, Friedrich DM, Hsiung C et al (2014) Pocket-size near-infrared spectrometer for narcotic materials identification. In: Proceedings of SPIE—the international society for optical engineering, 9101, Art. no. 91010O.
  45. Philp M, Shimmon R, Tahtouh M et al (2016) Development and validation of a presumptive colour spot test method for the detection of synthetic cathinones in seized illicit materials. Forensic Chem 1:39–50. Scholar
  46. Qian Z, Jia W, Li T et al (2017) Identification of five pyrrolidinyl substituted cathinones and the collision-induced dissociation of electrospray-generated pyrrolidinyl substituted cathinones. Drug Test Anal 9(5):778–787. Scholar
  47. Randox Toxicology—ELISA Accessed 4 Oct 2017
  48. Regester LE, Chmiel JD, Holler JM et al (2015) Determination of designer drug cross-reactivity on five commercial immunoassay screening kits. J Anal Toxicol 39(2):144–151. Scholar
  49. Roda E, Lonati D, Buscaglia E et al (2016) Evaluation of two different screening ELISA assays for synthetic cathinones (mephedrone/methcathinone and MDPV) with LC-MS method in intoxicated patients. J Clin Toxicol 6:3. Scholar
  50. Rosner P (2017) Mass spectra of designer drugs 2017. Wiley VCH, WeinheimGoogle Scholar
  51. Rowe WF, Marginean I, Carnes S et al (2017) The role of diode array ultraviolet detection for the identification of synthetic cathinones. Drug Test Anal. Scholar
  52. Saito T, Namera A, Osawa M et al (2013) SPME–GC–MS analysis of a-pyrrolidinovaleorophenone in blood in a fatal poisoning case. Forensic Toxicol 31(2):328–332. Scholar
  53. Salomone A, Gazzilli G, Di Corcia D et al (2016) Determination of cathinones and other stimulant, psychedelic, and dissociative designer drugs in real hair samples. Anal Bioanal Chem 408(8):2035–2042. Scholar
  54. Sørensen LK (2011) Determination of cathinones and related ephedrines in forensic whole-blood samples by liquid-chromatography-electrospray tandem mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 879(11–12):727–736. Scholar
  55. Stojanovska N, Tahtouh M, Kelly T et al (2014) Presumptive analysis of 4-methylmethcathinone (mephedrone) using Desorption electrospray ionisation—mass spectrometry (DESI-MS). Aust J Forensic Sci 46(4):411–423. Scholar
  56. Strano-Rossi S, Cadwallader AB, de la Torre X et al (2010) Toxicological determination and in vitro metabolism of the designer drug methylenedioxypyrovalerone (MDPV) by gas chromatography/mass spectrometry and liquid chromatography/quadrupole time-of-flight mass spectrometry. Rapid Commun Mass Spectrom 24:2706–2714. Scholar
  57. Swortwood MJ, Boland DM, DeCaprio AP (2013) Determination of 32 cathinone derivatives and other designer drugs in serum by comprehensive LC-QQQ-MS/MS analysis. Anal Bioanal Chem 405:1383–1397. Scholar
  58. Swortwood MJ, Hearn WL, DeCaprio AP (2014) Cross-reactivity of designer drugs, including cathinone derivatives, in commercial enzyme-linked immunosorbent assays. Drug Test Anal 6(7–8):716–727. Scholar
  59. Synthetic Cathinones (Methcathinone) ELISA Kit. Accessed 4 Oct 2017
  60. Tang MHY, Ching CK, Lee CYW et al (2014) Simultaneous detection of 93 conventional and emerging drugs of abuse and their metabolites in urine by UHPLC-MS/MS. J Chromatogr B 969:272–284. Scholar
  61. Toole KE, Fu S, Shimmon RG et al (2012) Color tests for the preliminary identification of methcathinone and analogues of methcathinone. Microgram J 9(1):27–32Google Scholar
  62. Uralets V, Rana S, Morgan S et al (2014) Testing for designer stimulants: metabolic profiles of 16 synthetic cathinones excreted free in human urine. J Anal Toxicol 38(5):233–241. Scholar
  63. Vaiano F, Busardò FP, Palumbo D et al (2016) A novel screening method for 64 new psychoactive substances and 5 amphetamines in blood by LC-MS/MS and application to real cases. J Pharm Biomed Anal 129:441–449. Scholar
  64. Westphal F, Junge T, Rosner P et al (2009) Mass and NMR spectroscopic characterization of 3,4-methylenedioxypyrovalerone: a designer drug with α-pyrrolidinophenone structure. Forensic Sci Int 190(1-3):1–8. Scholar
  65. Wiergowski M, Woźniak MK, Kata M et al (2016) Determination of MDPBP in postmortem blood samples by gas chromatography coupled with mass spectrometry. Monatsh Chem 47:1415–1421. Scholar
  66. Zuba D (2012) Identification of cathinones and other active components of ‘legal highs’ by mass spectrometric methods. TrAC Trend Anal Chem 32:15–30. Scholar

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Institute of Forensic ResearchKrakówPoland

Personalised recommendations