Post Mortem Analysis of Opioids and Metabolites in Skeletal Tissue

Every year, thousands of suspicious deaths are accounted for by an overdose of opioids. Occasionally all traditional matrices are unavailable due to decomposition. Skeletal tissue may pose a valid alternative. However, reference data on post-mortem concentrations in bone tissue and bone marrow is sparse. Therefore, a liquid chromatography tandem mass spectrometry method was developed and fully validated for the analysis of 4 opioids and 2 metabolites (tramadol, O-desmethyltramadol, morphine, fentanyl, norfentanyl, codeine) in bone tissue and bone marrow. Sample preparation was performed using solid phase extraction (bone marrow), methanolic extraction (bone) and a protein precipitation (whole blood). All validation parameters were successfully fulfilled. This method was applied to analyze 22 forensic cases involving opioids. All 6 opioids were proven to be detectable and quantifiable in all specimens sampled. When tramadol blood concentrations were correlated with bone concentrations, a linear trend could be detected. The same was seen between tramadol blood and bone marrow concentration. A similar linear trend was seen when correlating codeine blood concentration with bone and bone marrow concentration. Although some variability was detected, the same linear trend was seen for morphine. For fentanyl and norfentanyl, the sample size was too small to draw conclusions, regarding correlation. As far as the authors know this is the first-time fentanyl and norfentanyl are quantified in skeletal tissue. In conclusion, due to the absence of reference data for drugs in skeletal tissue, these findings are a step forward towards a more thorough understanding of drug concentration found in post-mortem skeletal tissue.