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Drug abuse is both an age-old and a constantly evolving problem in society. Trends in illicit drug use are highly fluid, with new formulations increasing in popularity. For this reason, methods for illicit drug detection and analysis need to be continually updated so they remain useful and relevant. A recent trend in street heroin production has seen it diluted with large amounts of tramadol in addition to the classical diluents such as acetaminophen and caffeine. This study describes a sensitive, simple and accurate high-performance liquid chromatographic method with ultraviolet detection for the simultaneous detection of heroin, 6-acetylmorphine, morphine, tramadol and O-desmethyltramadol in the blood of rats using a liquid–liquid back-extraction method. The separation was performed on LichroCART RP-18e with particle size of 5 µm (250 × 4.6 mm) with mobile phase acetonitrile–50 mM KH 2 PO 4  buffer, pH 7.1, using a gradient mode with a 1.0 mL/min flow rate. The calibration curves were linear in the concentration ranges 0.25–100 and 0.1–100 µg/mL for morphine and other analytes, respectively. Recovery values for the substances ranged between 59 and 83%. This technique was successfully used in pharmacokinetic studies measuring 6-acetylmorphine, morphine, tramadol and O-desmethyltramadol in the blood of rats intraperitoneally treated with a blend of 10 mg/kg heroin and 70 mg/kg tramadol. This technique shows promise for analysis of confiscated street heroin.

New HPLC Method to Detect Individual Opioids (Heroin and Tramadol) and their Metabolites in the Blood of Rats on Combination Treatment