DEVELOPMENT OF A LC/MS-MS METHOD FOR THE STUDY OF THE RATIOS BETWEEN MORPHINE, MORPHINE-3-beta-D-GLUCURONIDE AND MORPHINE-6-beta-D-GLUCURONIDE IN BLOOD SAMPLES FROM HEROIN FATALITIES".

In heroin fatalities the diagnosis of the cause of death may be particularly difficult because of several reasons, such as the relationship between lethal dose and current individual tolerance, the complexity of heroin metabolism, the presence of systemic dysfunction, and the contemporary use of other drugs of abuse. Thus, a wide variability is present in post-mortem blood concentration of morphine (MOR), the main metabolite of heroin, which is usually the most important analytical result for the interpretation of the cause of death. Recently, increasing interest has grown towards the role of the metabolites morphine-3-β-D-glucuronide (M3G) and morphine-6-β-D-glucuronide (M6G) in mediating heroin effects. The aim of this PhD study has been the development of a LC/MS-MS method for the determination of MOR, M3G and M6G in autopsy blood samples. An ESI-QqQ Mass Spectrometer, operating in positive ionisation and MRM mode, was used. Chromatographic separation was achieved thanks to a Reversed-Phase method, using a C18 column and a gradient elution with a binary mobile phase. SPE technique was employed to extract the analytes from biological samples. After validation, the method was applied to twenty-five blood specimens collected from cases of suspected fatal heroin overdose. The concentrations and the molar ratios of MOR, M3G and M6G were investigated. The influence of some risk factors, such as the contemporary use of alcohol, methadone or cocaine, was also studied.