On practical problems in precise estimation of 5F-ADB in urine samples

Abstract

Purpose: The purpose of this work is to establish if there are any problems in precise quantitation of methyl 2-[1-(5-fluoropentyl)-1H-indazole-3-carboxamido]-3,3-dimethylbutanoate (5F-ADB) in human urine by QuEChERS extraction coupled with gas chromatography–tandem mass spectrometry (GC–MS/MS). Methods: To establish the method for quantifying 5F-ADB in human urine samples, QuEChERS extraction coupled with GC–MS/MS was applied. To elucidate 5F-ADB degradation products, liquid chromatography coupled with linear trap quadrupole-orbitrap hybrid MS (LC–MS) was used. Results: The applied QuEChERS GC–MS/MS procedure appeared to be satisfactory for 5F-ADB estimation in acidic and alkaline urine samples. Its validation parameters were the following: good linearity (R2 = 0.9988), high detection (limit of detection = 0.33 ng/mL) and quantitation (limit of quantitation = 1.1 ng/mL) sensitivities and satisfactory inter- and intraday precisions (% relative standard deviation below 5.6%). 5F-ADB recovery from acidic urine by QuEChERS procedure was slightly lower than that from urine sample with neutral pH; however, the difference in the recovery was not statistically significant. The recovery of the drug from alkaline urine is extremely low. LC–MS analysis proved the presence of 5F-ADB hydrolysis products in alkaline urine and in alkaline solution of the drug. Conclusions: The presented studies indicate that the validated QuEChERS technique can be successfully used in routine analyses of 5F-ADB in urine. Yet, due to hydrolytical instability of 5F-ADB, the medical diagnosis of the health condition of the patient suspected of 5F-ADB abuse on the basis of the drug concentration in his/her urine may be incorrect, especially when the urine is alkaline.