Dextromethorphan and its matabolites in rat bone tissues by gas chromatography-mass spectrometry analysis following differential microclimate decomposition

Abstract

The effect of microclimate on dextromethorphan (DXM) and dextrorphan (DXT) responses in skeletonized rat remains was examined. Animals (n=10) received dextromethorphan at 75 mg/kg by i.p. injection for comparison against drug-free controls (n=4), and across different decomposition sites. Rats were divided equally into two groups and placed at different sites for decomposition immediately following euthanasia (30 minutes post dose). Rats at Site A decomposed in a shaded forest microenvironment on a grass-covered soil substrate. Site B animals rested on rock and gravel substrate exposed to open air and direct sunlight. Site A and Site B are approximately 600 m apart. Ambient temperature and relative humidity measurements recorded by data loggers mounted 3 cm above rats at each site established microclimate differences. Bone elements (vertebrae, ribs, pelvic girdles, femora, tibiae, humeri and scapulae) were harvested, cleaned and pulverized for Microwave Assisted Extraction in methanol. Drug and metabolite extractions were isolated by solid phase extraction prior to GC/MS analyses. Mass normalized DXM and DXT levels and metabolite/drug ratios were compared across different bone elements (within and between animals) and microclimate sites. Concentrations calculated from drug responses and standard curve plots gave estimated concentrations of 399 to 10,474 ng/g for DXM and 132 to 3,668 ng/g for DXT. Max/min values across animals and bone elements examined response variation. No significant differences in DXT levels or metabolite/parent ratios were observed between sites or across different bone elements. The only significant difference for DXM levels were found in femurs compared across microclimate sites. Microclimate showed no significant influence on observed DXM or DXT values, indicating bone as a drug reservoir may behave uniformly for certain drugs across different environments of decomposition. The results of this study show limited agreement with previous work from our laboratory on drug recoveries from decomposed bone tissues.