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Background Although the post-mortem increase of ammonium in biological fluids is well known, ammonium analysis in vitreous humour has never been used in recent times for the determination of the post-mortem interval. The present work represents a new application of capillary electrophoresis with indirect UV detection in the field of forensic analysis. Methods The electrophoretic separation was carried out in a running buffer made of 5 mM imidazole, 5 mM 18-crown-6 ether and 6 mM d,l-α-hydroxybutyric acid (HIBA). To overcome the lack of optical absorption of ammonium, indirect UV detection was applied. The used wavelength was 214 nm. Results The method showed good linearity in the concentration range from 0.16 to 5.0 mM. The limit of detection, 0.039 mmol/L, was established on the basis of the linearity curve. Precision and bias studies carried out on the pure ammonium solutions and in real biological samples, revealed %RSDs well below 20%. A preliminary application to real cases where the death time was precisely known (14 bodies) was carried out plotting vitreous humour ammonium vs. post-mortem interval with a resulting good linear correlation until 100 h post-mortem. Conclusions After validation in real cases, the present method can become a powerful tool to unravel one of the most challenging issues of forensic investigation: determination of the time of death.

clinical chemistry and laboratory medicine (cclm)

A new method for the determination of ammonium in the vitreous humour based on capillary electrophoresis and its preliminary application in thanatochemistry