Fentanyl Receptor Assay II. Utilization of a Radioreceptor Assay for the Analysis of Fentanyl Analogs in Urine

A radioreceptor assay has been developed to measure fentanyl and fentanyl-like drugs in biological specimens. The assay is based on the competition of these drugs with [3H]fentanyl for opioid receptors. Rats were injected intravenously with fentanyl (15 micrograms/kg), alpha-methylfentanyl (15 micrograms/kg), (+/-)-cis-3-methylfentanyl (15 micrograms/kg), butyrylfentanyl (0.48 mg/kg), and benzylfentanyl (3.19 mg/kg). Urine samples were analyzed by radioreceptor assay (RRA), radioimmunoassay (RIA), and gas chromatography/mass spectrometry (GC/MS). The time-course of urinary analysis of fentanyl analogs showed some discrepancies. RRA measurement of urine concentrations of (+/-)-cis-3-methylfentanyl that were undetectable by RIA gave results 5-10 times higher than values obtained by GC/MS. Concentrations of alpha-methylfentanyl obtained by RRA and GC/MS were similar; however, these samples were negative by RIA. Following the administration of benzylfentanyl, urinary concentrations were not detected by RIA and only slightly detectable with RRA; however, high concentrations of benzylfentanyl were found by GC/MS in the same samples. Urine samples from animals injected with butyrylfentanyl showed high cross-reactivity with fentanyl antibody, giving values measured by radioimmunoassay about two times higher than those obtained by the other two methods. These findings suggest that this radioreceptor assay is well-suited as an initial assay for the detection of active analogs of fentanyl in urine and correlates well with other techniques in the analysis of fentanyl; however, there is substantial disagreement between techniques in the quantitation of fentanyl analogs.