High variation in hypoxanthine determination after analytical treatment of vitreous humor samples

In the field of legal medicine the correct determination of the time of death is of great importance because an error in calculating the post-mortem interval (PMI) could be crucial in a criminal investigation. The quantification of hypoxanthine (Hx) concentration in the vitreous humor (VH) as a means to estimate PMI is useful when dealing with a recent time interval of death and has several advantages over other biological matrices. However, due to its viscous nature, VH has to be liquefied prior to its analysis, and the different procedures to liquefy it may cause alterations in Hx concentration, and hence in estimation of PMI. The aim of this study was to determine differences in Hx concentration after applying several pre-analytical treatments to different aliquots of the same VH sample. Enzymatic digestion, sonication, centrifugation and heat were the liquefying methods applied to reduce the viscosity of 105 VH samples. Analysis was performed using solid phase extraction and liquid chromatography in tandem with mass spectrometry. Statistically significant differences in the measured Hx concentrations were found between enzymatic digestion and the other three methods analyzed (sonication, centrifugation, and heat). Additionally, high data dispersion was found under heat treatment but was not observed for centrifugation or sonication, where similar and more moderate results were found. We conclude that high variations in Hx determination are observed when enzymatic digestion or heat treatment is used and we therefore recommend sonication or centrifugation to quantify Hx concentration in the VH.     

Assessing the ecotoxicological effects of long-term contaminated mine soils on plants and earthworms: relevance of soil (total and available) and body concentrations

The interactions and relevance of the soil (total and available) concentrations, accumulation, and acute toxicity of several essential and non-essential trace elements were investigated to determine their importance in environmental soil assessment. Three plant species (T. aestivum, R. sativum, and V. sativa) and E. fetida were simultaneously exposed for 21 days to long-term contaminated soils collected from the surroundings of an abandoned pyrite mine. The soils presented different levels of As and metals, mainly Zn and Cu, and were tested at different soil concentrations [12.5, 25, 50, and 100 % of contaminated soil/soil (w/w)] to increase the range of total and available soil concentrations necessary for the study. The total concentrations in the soils (of both As and metals) were better predictors of earthworm uptake than were the available concentrations. In plants, the accumulation of metals was related to the available concentrations of Zn and Cu, which could indicate that plants and earthworms accumulate elements from different pools of soil contaminants. Moreover, Zn and Cu, which are essential elements, showed controlled uptake at low concentrations. The external metal concentrations predicted earthworm mortality, whereas in plants, the effects on growth were correlated to the As and metal contents in the plants. In general, the bioaccumulation factors were lower at higher exposure levels, which implies the existence of auto-regulation in the uptake of both essential and non-essential elements by plants and earthworms.