Analysis of Buprenorphine, Norbuprenorphine, and Their Glucuronides in Urine by Liquid Chromatography-Mass Spectrometry

Buprenorphine is used for the treatment of chronic pain and also in treatment of heroin addiction as an alternative to methadone. As the availability of buprenorphine increases, so does the risk for abuse and the pressure on forensic and clinical laboratories to analyze for it. Buprenorphine and its dealkylated metabolite are excreted in urine, almost exclusively as glucuronides. The aim of the present study was to evaluate electrospray liquid chromatography tandem mass spectrometry (LC-MS-MS) for the rapid screening and quantitation of buprenorphine and its metabolites in urine. Three approaches were evaluated: (1) direct injection of diluted urine for measurement of glucuronides, (2) direct injection of diluted urine after enzymatic hydrolysis for the quantitation of buprenorphine and norbuprenorphine, and (3) quantitation of buprenorphine and norbuprenorphine after enzymatic hydrolysis and solid-phase extraction (SPE). One hundred six samples were subjected to procedure 1 and, when positive, further quantitated using procedure 2. Only samples with low analyle concentrations (< 20 µg/L) were subject to SPE. Concentrations of buprenorphine and norbuprenorphine in patients (N = 16) ranged between 31 and 1080 µg/L and 48–2050 µg/L, respectively. In suspected abusers (N = 33), the ranges were 2.3–796 µg/L and 5.0–2580 µg/L. In four of the authentic samples, both the buprenorphine and norbuprenorphine concentrations were below the 20-µg/L cutoff. We concluded that LC-MS-MS analysis of the glucuronides provided an adequate screening method, but that the direct method for quantitation sometimes had to be complemented with a concentration by SPE, providing increased sensitivity, thus lowering the cutoff from 20 to 1 µg/L urine.