Urine tests to detect methadone use are based on immunoassays that are designed to detect either methadone or its major urinary metabolite, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) formed by cytochrome P450 3A4 system. Immunoassays for methadone have low reactivity with EEDP and vice versa. Polymorphism of CYP 3A4 can lead to substantial variation in methadone metabolism and elimination. Those patients who rapidly metabolize the methadone dose taken can have methadone concentrations in urine below the typical cutoff of 300 ng/ml. These urine specimens, however, do contain EDDP in high concentrations and yield positive results if an EDDP-specific immunoassay is used. It has been suggested that EDDP can be a marker for monitoring methadone compliance (36). Huestis and co-workers (37) showed in their study that 2% of the specimens collected from patients who have taken methadone tested negative for methadone, whereas all specimens tested positive by the EDDP assay at a cutoff of 100 ng/ml. Theoretically, it is possible to have positive methadone and negative EEDP in a noncompliant patient who takes a dose of methadone just before urine collection, and there is insufficient time to metabolize the methadone to reach a detectable level of EDDP.

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