A Kratom Metabolite Causes False Positive Urine Drug Screening Results for Methadone
Authors: Pierre Christina , Gineste Catherine , Bazydlo Lindsay 
Published: October 2020

Abstract

Background
The synthetic opioid methadone is utilized in pain management and opioid addiction therapy. Patients with methadone prescriptions are monitored for compliance using immunoassay-based urine drug screens (UDS) for methadone and its primary metabolite, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP). It is well documented that immunoassays are subject to false positive results due to cross-reactivity of antibody reagents with structurally related and unrelated compounds. A 30-year-old male with a history of polysubstance abuse presented to the emergency department with altered mental status and was noted to have a positive UDS for EDDP. Reflex mass spectrometry testing was negative for both methadone and EDDP. Upon improvement, the patient endorsed use of powder derived from the tropical plant Kratom, whose main components, mitragynine and 7-hydroxymitragynine, function as opioid receptor agonists. Kratom use has become popularized in the United states in spite of warnings by the United States Food and Drug Administration about its potential to cause addiction, abuse, and dependence. Here we describe our investigation of Kratom as a positive interferent in the Thermo Scientific CEDIA Methadone Metabolite (EDDP) immunoassay.

Methods
Patient specimens that underwent urine drug screens as part of routine clinical care at the University of Virginia Health System over 3 non-contiguous months meeting the following criteria were included in the study: EDDP screen ≥10 arbitrary absorbance units, negative methadone screen, and negative methadone or EDDP mass spectrometry confirmation (if available). Urine drug screens were performed on the Olympus AU480 Chemistry Analyzer using the Thermo Scientific CEDIA Methadone Metabolite (EDDP) immunoassay and the Siemens Syva Emit II Plus Methadone assay. Specimens were tested for mitragynine, 7-hydroxymitragynine, methadone and EDDP using the Sciex X500R QTOF with Sciex Exion Liquid Chromatography.

Results
A total of 48 specimens met the inclusion criteria, where 50% (24/48) confirmed positive for mitragynine and/or 7-hydroxymitragynine and negative for methadone/ EDDP. Methadone and/or EDDP was detected in 33% (16/48), none of which confirmed positive for mitragynine or 7-hydroxymitragynine and 17% were negative for all four analytes. Urine spiked with a methanol extract prepared from Kratom powder produced no signal in the EDDP screen. Spiking drug-free urine with 10,000 ng/mL mitragynine and 10,000 ng/mL 7-hydroxymitragynine both resulted in indeterminate EDDP screens. Since several drugs are glucuronidated during phase II metabolism we performed enzymatic hydrolysis on urine specimens and rescreened the specimens using the EDDP immunoassay. Upon hydrolysis, mitragynine and/or 7-hydroxymitragynine positive specimens showed a significantly higher percent increase in absorbance on the EDDP screen compared to those in which methadone and/or EDDP was detected (p=0.0031).

Conclusion
Our studies suggest that a Kratom metabolite can cause false positive results in the Thermo Scientific CEDIA Methadone Metabolite (EDDP) immunoassay. Kratom use should be investigated in cases of false positive urine drug screens for EDDP.