The aim of this study was to develop and validate a method for the determination of buprenorphine (BUP), norbuprenorphine (NBUP) and naloxone (NAL) in fingernails and urine samples collected from former heroin users under suboxone substitution therapy. The analytes were extracted by solid-liquid or solid-phase extraction and were analyzed by liquid chromatography-mass spectrometry. The validation of the analytical methods developed included linearity, recovery, accuracy, precision, ion suppression, sensitivity of interfaces and limits of determination and quantification. The validated methods were applied to samples from 46 individuals. The majority of the urine samples were positive for all analytes (93.5% for BUP, 95.7% for NBUP and 84.8% for NAL). In nails, a higher detection rate was observed for NBUP and BUP (89.1%), compared with NAL (10.9%). The median values of the NBUP/BUP and the NAL/BUP ratio were 2.5 and 0.3 in urine and 0.8 and 0.3 in nails, respectively. A statistically significant correlation was found between the BUP, NBUP and total BUP (BUP and NBUP) concentrations in urine and those in nails. A weak correlation was observed between the daily dose (mg/day) and total BUP (P = 0.069), or NBUP (P = 0.072) concentrations in urine. In contrast, a strong correlation was found between the total amount of BUP administered during the last 12 months and total BUP (P = 0.038), or NBUP (P = 0.023) concentrations in urine. Moreover urine BUP, NBUP and total BUP concentrations correlated significantly. Our study demonstrated successfully the application of the developed method for the determination of the three analytes in urine and nails.
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