Tramadol and its metabolites in rat urine were identified by LC-MS(n). Rat urine samples of 0-36 h were collected after ip 9.0 mg x kg(-1) tramadol, then the samples were enriched and purified through solid-phase extraction cartridge. Purified samples were analyzed by LC-MS(n). Possible metabolites were discovered by comparing the full scan and SIM chromatograms of the test samples with the corresponding blanks and analyzing the retention time, quasi-molecular ion and fragment ion of all chromatograms. Nine phase I metabolites and four phase II metabolites were identified in rat urine. One of the metabolites was found first time in living body. The metabolites were formed via the following metabolic pathways: O-demethylation, N-demethylation, hydroxylation, N-oxidation and conjugation. The method can be used to identify tramadol and its metabolites in other animals and human.
Analgesics, Opioid ; administration & dosage ; metabolism ; urine ; Animals ; Chromatography, High Pressure Liquid ; Injections, Intraperitoneal ; Male ; Rats ; Rats, Wistar ; Spectrometry, Mass, Electrospray Ionization ; Tandem Mass Spectrometry ; Tramadol ; administration & dosage ; metabolism ; urine

OBJECTIVE: To develop a rapid and accurate gas chromatography method and investigate the distribution of tramadol in acute poisoned rats for information of samples selection and results evaluation in forensic identification. METHODS: After an oral administration of tramadol at 1140 mg/kg (5 x LD50), concentrations of tramadol in rats' biological fluids and tissues were determined by gas chromatography. RESULTS: The limit of detection of tramadol in blood and urine was 0.1 microg/mL and the limit of detection in liver was 0.1 microg/g. The intra-day precision and inter-day precision were within 3.1% and 5.5% respectively, and the recovery of tramadol in blood was more than 98%. The average levels of tramadol displayed in descending order of heart blood, liver, peripheral blood, urine, vitreous humor, kidney, lung, spleen, heart, brain respectively. CONCLUSION The established method could meet the requirements for toxicological analysis, and the results of the study suggest that blood, urine, liver, lung and kidney are suitable samples for forensic toxicological analysis in tramadol poisoning cases.
Acute Disease ; Administration, Oral ; Analgesics, Opioid/urine* ; Animals ; Body Fluids/chemistry* ; Chromatography, Gas/methods* ; Kidney/metabolism* ; Liver/metabolism* ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reproducibility of Results ; Sensitivity and Specificity ; Substance Abuse Detection/methods* ; Tissue Distribution ; Tramadol/urine*