Analysis of propranolol and its metabolites in postmortem human solid tissues and body fluids: LC-MS/MS approach with the standard addition method applied to a forensic case.

PURPOSE: This study aimed to develop a highly sensitive and specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of propranolol and its metabolites in human biological samples. By analyzing their presence in urine, postmortem biological fluids, and various solid tissues, the study could be of reliable forensic toxicological use for investigations in propranolol poisoning cases. In this study, the Standard Addition Method (SAM) was used for quantification, and its validation was mixed with one of the analyte's concentrations.

METHODS

A 0.1 mL aliquot of each body fluid sample or 0.1 g each of homogenized solid tissue was mixed with one of the analyte concentration standards, extracted with methanol, spiked with an internal standard (IS) using the SAM, and purified using magnesium sulfate and sodium sulfate. Following centrifugation and filtration, samples were analyzed via LC-MS/MS. Samples underwent enzymatic hydrolysis to quantify total metabolite concentrations (free plus conjugated) prior to analysis.

RESULTS

Phase I metabolites (propranolol, 4-hydroxypropranolol, propranolol glycol, N-desisopropylpropranolol, 1-naphthylenyloxyacetic acid, and 1-naphthol) and phase II metabolites (sulfate and glucuronide conjugates) were identified in urine. Among postmortem samples, propranolol was highest in the bile, followed by the lung tissue. Naphthoxylactic acid could be consistently detected in all samples except for the brain, suggesting its potential as a good biomarker for propranolol exposure.

CONCLUSION

A validated LC-MS/MS method for determining propranolol and its metabolites in forensic samples was established, and it could also be applied to the authentic human samples obtained from a propranolol poisoning case. The findings could offer substantial and reliable support for investigating propranolol-related fatalities and contribute to the comprehensive understanding of the metabolism of propranolol in the human body.