Background: ethyl glucuronide (EtG) and ethyl sulfate (EtS) are phase II direct ethanol metabolites. They are specific and sensitive biomarkers in the diagnosis of an acute or chronic alcohol ab/use, depending on the biological matrix evaluated. Due to alcohol postmortem production in biological tissues, they have been recently evaluated as potential biomarkers of ethanol ingestion. Some authors observed the presence of EtG in blood also after postmortem production of ethanol while data concerning EtS are still unreliable. Aim: this in vitro study aimed to the evaluation of all developmental stages of the fly Calliphora vicina L. (Calliphoridae) as biological matrix that could be used for the determination of these two metabolites in order to discriminate an alcohol ingestion in postmortem material. Methods: different pig substrates were used during the in vitro experiments (liver, heart, muscle and tongue) to rear C. vicina. All the experiments consisted of nine different aliquots (about 30 g) of substrates. One ml of ethanol was added to three aliquots, while EtG and EtS (1µg/g) were added to other three of them. The last three samples were kept blank. An LC-MS/MS method was used to detect the presence of EtG and EtS in the substrates, in the larvae and in the pupae. Different experiments were set up in order to: a) assess the presence of EtG and EtS; b) study the variability due to the substrates characteristics; c) evaluate the possibility of false positive due to external alcohol contamination; d) estimate the potential metabolization of ethanol by the insects. Results: EtS was found in all the samples where the standard was added to the substrate, while EtG results were more scattered, mainly due to the high matrix effects observed during the instrumental analysis. A great variation in EtG and EtS analyses were observed in liver, heart and tongue, while muscle appeared to be the most stable biological matrix: the three blank aliquots as well as the three with ethanol addition didn’t lead to false positive results. EtS was found in larvae, pupae and puparia. This means that C. vicina is not able to metabolize ethanol nor EtG and EtS. Conclusion: this study showed that maggots, pupae and puparia could be useful matrix for the evaluation of antemortem alcohol ingestion. EtS is the marker of choice in this specific diagnostic field. In vivo experiments would be necessary in order to confirm the results obtained by this in vitro trial.

Ethyl glucuronide and ethyl sulfate in Calliphora vicina l. (Calliphoridae): an in vitro study on evidence of antemortem ingestion of ethanol

LAMBIASE, SIMONETTA;GROPPI, ANGELO;MORINI, LUCA
2012-01-01

Abstract

Background: ethyl glucuronide (EtG) and ethyl sulfate (EtS) are phase II direct ethanol metabolites. They are specific and sensitive biomarkers in the diagnosis of an acute or chronic alcohol ab/use, depending on the biological matrix evaluated. Due to alcohol postmortem production in biological tissues, they have been recently evaluated as potential biomarkers of ethanol ingestion. Some authors observed the presence of EtG in blood also after postmortem production of ethanol while data concerning EtS are still unreliable. Aim: this in vitro study aimed to the evaluation of all developmental stages of the fly Calliphora vicina L. (Calliphoridae) as biological matrix that could be used for the determination of these two metabolites in order to discriminate an alcohol ingestion in postmortem material. Methods: different pig substrates were used during the in vitro experiments (liver, heart, muscle and tongue) to rear C. vicina. All the experiments consisted of nine different aliquots (about 30 g) of substrates. One ml of ethanol was added to three aliquots, while EtG and EtS (1µg/g) were added to other three of them. The last three samples were kept blank. An LC-MS/MS method was used to detect the presence of EtG and EtS in the substrates, in the larvae and in the pupae. Different experiments were set up in order to: a) assess the presence of EtG and EtS; b) study the variability due to the substrates characteristics; c) evaluate the possibility of false positive due to external alcohol contamination; d) estimate the potential metabolization of ethanol by the insects. Results: EtS was found in all the samples where the standard was added to the substrate, while EtG results were more scattered, mainly due to the high matrix effects observed during the instrumental analysis. A great variation in EtG and EtS analyses were observed in liver, heart and tongue, while muscle appeared to be the most stable biological matrix: the three blank aliquots as well as the three with ethanol addition didn’t lead to false positive results. EtS was found in larvae, pupae and puparia. This means that C. vicina is not able to metabolize ethanol nor EtG and EtS. Conclusion: this study showed that maggots, pupae and puparia could be useful matrix for the evaluation of antemortem alcohol ingestion. EtS is the marker of choice in this specific diagnostic field. In vivo experiments would be necessary in order to confirm the results obtained by this in vitro trial.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11571/493846
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