Reliable, sensitive, rapid and quantitative enzyme-based assay for gamma-hydroxybutyric acid (GHB)

Several assays for gamma-hydroxybutyrate (4-hydroxybutyrate, GHB) have been developed based on the enzyme gamma-hydroxybutyrate dehydrogenase (GHB-DH). Enzymatic oxidation of GHB by NAD+ is coupled to diaphorase-mediated reduction of pro-dye to yield colored product. GHB-DH from Ralstonia eutropha was cloned and expressed as a stable fusion protein easily purified by affinity chromatography. Quantitative initial velocity and endpoint versions of the assay in solution are described. Michaelis-Menten parameters for oxidation of GHB and ethanol were estimated. A semi-quantitative "dipstick" version of the assay on paper also is described. Both solution endpoint and "dipstick" assays are sensitive to about 0.05 mg GHB/mL using 10 microL of sample. Ethanol at concentrations possible in urine and agents used to stabilize physiological fluids for forensics analysis do not interfere significantly. The "dipstick" assay also allows detection of GHB in alcoholic beverages after evaporation of about one-fourth drop of beverage before testing. The enzymatic assay for GHB is reliable, sensitive, inexpensive and rapid.     

A laboratory incubation method for determining the rate of microbiological degradation of skeletal muscle tissue in soil

A controlled laboratory experiment is described, in principle and practice, which can be used for the of determination the rate of tissue decomposition in soil. By way of example, an experiment was conducted to determine the effect of temperature (12 degrees, 22 degrees C) on the aerobic decomposition of skeletal muscle tissue (Organic Texel x Suffolk lamb (Ovis aries)) in a sandy loam soil. Measurements of decomposition processes included muscle tissue mass loss, microbial CO2 respiration, and muscle tissue carbon (C) and nitrogen (N). Muscle tissue mass loss at 22 degrees C always was greater than at 12 degrees C (p < 0.001). Microbial respiration was greater in samples incubated at 22 degrees C for the initial 21 days of burial (p < 0.01). All buried muscle tissue samples demonstrated changes in C and N content at the end of the experiment. A significant correlation (p < 0.001) was demonstrated between the loss of muscle tissue-derived C (Ct) and microbially-respired C (Cm) demonstrating CO2 respiration may be used to predict mass loss and hence biodegradation. In this experiment Q10 (12 degrees C-22 degrees C) = 2.0. This method is recommended as a useful tool in determing the effect of environmental variables on the rate of decomposition of various tissues and associated materials.     

Identification of ascorbic acid and its degradation products in black powder substitutes

Low explosives such as smokeless powder, black powder, and black powder substitutes have been used in illicit pipe bombings throughout the United States. Some of the newer black powder substitutes are formulated with ascorbic acid, which gradually decomposes as the powder ages, making it difficult if not impossible for the forensic chemist to identify it by traditional bulk techniques. A sensitive method for the identification of residual levels of ascorbic acid in black powder substitutes is presented. Powder samples are extracted with a mixture of acetonitrile and bis(trimethylsilyl)acetamide (BSA), which converts carboxylic acid and alcohol functional groups to trimethylsilyl esters and ethers, respectively. Samples are then analyzed by gas chromatography-mass spectrometry (GC-MS). Results have shown that trace amounts of ascorbic acid can be identified at detection limits that are well below those for traditional bulk techniques. Degradation products for ascorbic acid (hydroxylated carboxylic acids, furanones, and lactones) can also be detected.     

Determination of group affiliation of isolated spermatozoa after separation from squamous epithelium cells

Outlined in the paper are outlooks for determining a group affiliation of sperm in traces, left in material evidence, within forensic biological expertise. A technique is suggested to separate the spermatozoids from the vaginal epithelium cells--it has been used up to now in genetics alone. The method involves the available reagents and does not need any extra training. Its elaboration and introduction would resolve one of the most often-encountered issues set by the investigating office.     

The Risk Assessment Program and the Court of Penal Execution in the Province of Buenos Aires, Argentina

This study describes the working of a new system that supervises the execution of the punishment phase in the Province of Buenos Aires, Argentina; namely, the Court of Penal Execution, as well as the risk assessment program. It also reports the results of the violent recidivism baseline risk assessment carried out in candidates for conditional release. The cohort studied was recruited during 18 months in the Judicial Department of La Plata, taking into consideration psychopathological and criminological variables. The Psychopathy Checklist-Revised (PCL-R), the Historical, Clinical, and Risk Management--20 (HCR-20), and the Violence Risk Appraisal Guide (VRAG), among others, were used as measures. During the study period, 65 candidates for conditional release were part of the baseline assessment, and they constitute the population of this study. The measures' mean scores were PCL-R, 20.57; HCR-20, 18.58; VRAG, 12.17.     

Interpretation of a 3,4-methylenedioxymethamphetamine (MDMA) blood level: discussion by means of a distribution study in two fatalities

The amphetamine derivative 3,4-methylenedioxymethamphetamine (MDMA, "Ecstasy" is a currently used or abused designer drug and fatalities are frequently encountered in forensic practice. However, the question remains open whether an MDMA blood level can be toxic or even potentially lethal. In order to provide insight in the interpretation of a detected MDMA concentration, the distribution of MDMA and its metabolite 3,4-methylenedioxyamphetamine (MDA) in various body fluids and tissues was studied and discussed in two different fatalities. Apart from peripheral blood samples (such as femoral and subclavian blood), various blood samples obtained centrally in the human body and several body fluids (such as vitreous humour) were examined. In addition, various tissues such as cardiac muscle, lungs, liver, kidneys, and brain lobes were analysed. In contrast to the peripheral blood levels, high MDMA and MDA levels were found in cardiac blood and the majority of the organs, except for the abdominal adipose tissue. The high concentrations observed in all lung lobes, the liver and stomach contents indicate that post-mortem redistribution of MDMA and MDA into cardiac blood can occur and, as a result, blood sampled centrally in the body should be avoided. Therefore, our data confirm that peripheral blood sampling remains "the golden standard". In addition, a distinct difference in peripheral blood MDMA concentrations in our two overdose cases was established (namely 0.271 and 13.508 microg/ml, respectively). Furthermore, our results suggest that, if a peripheral blood sample is not available and when putrefaction is not too pronounced, vitreous humour and iliopsoas muscle can be valuable specimens for toxicological analysis. Finally, referring to the various mechanisms of death following amphetamine intake, which can result in different survival times (e.g. cardiopulmonary complications versus hyperthermia), the anatomo-pathological findings and the toxicological results should be considered as a whole in arriving at a conclusion.     

Suicide with inert gases: addendum to Final Exit

Suicide by asphyxiation with a plastic bag placed over the head is a method that has been advocated by right-to-die groups. Recently, such groups have proposed the introduction of helium into the plastic bag as a means of hastening death. Helium is readily available at toy stores, where it is sold in tanks for balloon inflation. It produces asphyxiation by the exclusion of oxygen in enclosed spaces. We report 7 fatalities throughout an 18-month period involving plastic bag suffocation in conjunction with helium use. These fatalities coincide with publication of an update to a popular right-to-die text in which this method is described. Although right-to-die literature was absent from all scenes, this method was not previously observed in our jurisdiction, and the deaths likely reflect exposure to this information. Because of analytical difficulties in testing for helium in biologic specimens, death certification rests on scene investigation.