LC-MS-MS Method Development and Analysis of Stimulants, Opiates, Synthetic Opiates, PCP, and Benzodiazepines in Wastewater. Preponderance of these Drugs During Football Games

  • Waseem Gul
  • Shahbaz W. Gul
  • Brandon Stamper
  • Murrell Godfrey
  • Mahmoud A. ElSohlyEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1810)


A method was developed for the analysis of stimulant drugs, opiates, synthetic opiates, PCP, and benzodiazepines in wastewater samples using liquid chromatography coupled with tandem mass spectrometry (LC-MS-MS). A total of 33 compounds (stimulant-type drugs and metabolites of opiates, synthetic opiates, PCP, and benzodiazepines) were analyzed. These drugs included amphetamine (Amp) (1), methamphetamine (Meth) (2), methylenedioxyamphetamine (MDA) (3), methylenedioxymethamphetamine (MDMA) (4), methylenedioxyethylamphetamine (MDEA) (5), benzoylecgonine (BE, the major metabolite of Coc) (6), cocaine (Coc) (7), 6-monoacetylmorphine (6-MAM, the primary urinary metabolite of heroin) (8), codeine (9), hydrocodone (10), hydromorphone (11), morphine (12), norhydrocodone (the primary urinary metabolite of hydrocodone) (13), oxycodone (14), oxymorphone (15), 2-ethylidine-1,5-dimethyl-3,3-diphenylpyrolidine (EDDP, the primary urinary metabolite of methadone) (16), fentanyl (17), meperidine (18), methadone (19), norfentanyl (the primary urinary metabolite of fentanyl) (20), normeperidine (the primary urinary metabolite of meperidine) (21), phencyclidine (PCP) (22), tramadol (23), alprazolam (24), temazepam (25), nordiazepam (26), chlordiazepoxide (27), flurazepam (28), oxazepam (29), α-OH-alprazolam (the primary urinary metabolite of alprazolam) (30), α-OH-triazolam (the primary urinary metabolite of triazolam) (31), 2-OH-ethylflurazepam (the primary urinary metabolite of flurazepam) (32), and 7-NH2-flunitrazepam (the primary urinary metabolite of flunitrazepam) (33). These drugs were chosen because of their widespread abuse. Wastewater samples were collected at both the Oxford Wastewater Treatment Plant in Oxford, Mississippi (MS), and the University Wastewater Treatment Plant in University, MS. Samples were collected on weekends on which the Ole Miss Rebel football team held home games (Vaught-Hemingway Stadium, University, MS 38677). The collected samples were analyzed using a validated method and found to contain Amp, Meth, MDMA, MDA, Coc, BE, codeine, hydrocodone, hydromorphone, morphine, norhydrocodone, oxycodone, oxymorphone, tramadol, EDDP, meperidine, normeperidine, methadone, alprazolam, α-OH-alprazolam, nordiazepam, oxazepam, and temazepam. None of the samples contained MDEA, 6-MAM, fentanyl, norfentanyl, PCP, chlordiazepoxide, flurazepam, 2-OH-ethylflurazepam, 7-NH2-flunitrazepam, and α-OH-triazolam.

Key words

Wastewater Benzodiazepines Opiates Stimulant drugs LC-MS/MS 



The authors are thankful to Mr. Randy McCluskey and employees of the Oxford Wastewater Treatment Plant, as well as Mr. David Adkisson and employees of the University Wastewater Treatment Plant for their assistance in collecting samples. The authors are also thankful to Kareem ElSohly, Candice Tolbert, Racheal Speltz, Iram Shahzadi, and Justin Reid (ElSohly Laboratories, Inc.) for their assistance.


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Waseem Gul
    • 1
    • 3
  • Shahbaz W. Gul
    • 1
  • Brandon Stamper
    • 1
    • 2
  • Murrell Godfrey
    • 2
  • Mahmoud A. ElSohly
    • 1
    • 4
    • 5
    Email author
  1. 1.ElSohly Laboratories, Inc.OxfordUSA
  2. 2.Department of Chemistry and BiochemistryUniversity of MississippiUniversityUSA
  3. 3.National Center for Natural Products ResearchUniversity of MississippiUniversityUSA
  4. 4.National Center for Natural Products ResearchUniversity of MississippiUniversityUSA
  5. 5.Department of Pharmaceutics and Drug DeliveryUniversity of MississippiUniversityUSA

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