STR genotyping of DNA extracted from used Triage kits

This study examined whether short tandem repeat (STR) genotyping can be performed using DNA remaining in Triage kits used to screen for drugs of abuse in urine. STR genotyping was successful for 15 loci using 12 kits stored for 1–6 months at room temperature. These results suggest that STR genotyping for human identification can be performed using DNA extracted from used Triage kits.     

Detection of methamphetamine and amphetamine in a skeletonized body buried for 5 years

A 28-year-old male methamphetamine abuser, who had been buried for 5 years after being killed by strangulation, was found skeletonized. Methamphetamine and amphetamine in the significantly denatured fatty material of the bone marrow were analyzed by gas chromatography and gas chromatography-mass spectrometry. The confirmation of the chemicals was carried out by chemical ionization (CI) mass chromatography, CI mass spectrometry and CI mass fragmentography. The concentrations of methamphetamine and amphetamine determined by CI mass fragmentography were 1.0 mumol/100 g and 0.1 mumol/100 g, respectively. The method used would seem to be very useful for determination of methamphetamine and amphetamine in marked putrefied biological materials.     

Determination of dimethoate in blood and hemoperfusion cartridge following ingestion of formothion: a case study

A 57-year-old male who had ingested not more than 22 g of formothion was semicomatose on admission to hospital, approximately 1.5 h after ingestion. Dimethoate, a hydrolyzed formothion, was found in blood samples collected from the patient and in the charcoal column in the direct hemoperfusion cartridge which was used 6 to 7.5 h after ingestion. It was extracted and purified by Extrelut column extraction. A gas chromatograph, equipped with a flame photometric detector and a gas chromatograph-mass spectrometer, were used to detect and confirm the presence of dimethoate. The blood dimethoate concentrations which were taken approximately 1.5 and 6 h after ingestion were 21.4 and 12.7 micrograms/g, respectively. A blood dimethoate concentration of 21.4 micrograms/g would appear to indicate a high level of formothion intoxication. The total amount of dimethoate found in the charcoal column used was 15 mg.     

Determination of disulfoton and its metabolites in the body fluids of a Di-Syston intoxication case

Disulfoton and its metabolites, two sulfoxides and two sulfones, in the body fluids of a patient who had ingested Di-Syston were analyzed by FPD-GC and GC/MS. After the chemicals in the extract (Fraction 1) obtained by Extrelut column extraction were analyzed, disulfoton and sulfoxides in Fraction 1 were oxidized into sulfones. The sulfones in the extract (Fraction 2) obtained by Extrelut column extraction were analyzed and the estimated concentrations of metabolite were calculated. The concentrations of disulfoton and the sum of the metabolites in the blood collected on admission were 0.093 nmol/g (25.4 ng/g) and 4.92 nmol/g (corresponding to 1.35 micrograms/g of disulfoton), respectively. These concentrations appear to indicate a severe level of disulfoton intoxication.     

Correlation between 'on admission' blood toluene concentrations and the presence or absence of signs and symptoms in solvent abusers

For 1 year, from June 1987 to June 1988, toluene concentrations in blood samples of patients admitted to a psychiatric hospital after inhaling solvent vapor, collected on admission and 4 h later, were analyzed by gas chromatograph. Toluene levels in the first urine samples collected after admission were also analyzed and case histories were kept listing age, sex and physical and psychiatric effects. In all, 51 cases were studied--34 males and 17 females. (1) The average age of the males was 21.4 years and of the female 16.2 years. (2) The toluene concentrations in the blood collected on admission ranged from 0.3 to 22.8 micrograms/g. (3) Physical signs were observed in 9 patients with an "on admission" blood toluene concentrations of more than 3.0 micrograms/g; twice as many subjects (18), however, with blood toluene concentration greater than 3.0 micrograms/g were without physical signs. (4) The blood toluene concentrations of three cases in the condition known as twilight state were more than 10.0 micrograms/g. (5) In 24 cases with blood toluene concentrations below 3.0 micrograms/g, there were no physical signs. (6) Five subjects with blood toluene concentrations in the 0.8-5.2 micrograms/g range showed neuropsychiatric effects; however, 23 subjects in the same blood toluene concentration range did not exhibit psychiatric effects, and none of the subjects with blood toluene concentrations greater than 5.2 micrograms/g, 15 in all, had such effects.     

Postmortem molecular screening for mutations in ryanodine receptor type 1 (RYR1) gene in psychiatric patients suspected of having died of neuroleptic malignant syndrome

Postmortem diagnosis of neuroleptic malignant syndrome (NMS) is difficult to perform, because the clinical symptoms just before death are not usually available. Malignant hyperthermia (MH) is a catastrophic, life-threatening hypermetabolic syndrome triggered by certain anesthetics. Ryanodine receptor type 1 (RYR1) gene mutations are known to be involved in susceptibility to MH. Similarities in clinical features, such as elevated body temperature, between NMS and MH have led to the suggestion that NMS is a neurogenic form of MH. In this study, we analyzed possible mutations of the RYR1 gene in 11 psychiatric patients suspected at autopsy to have died of NMS. All cases were suspected of having elevated body temperature at death, and their causes of death could not be determined by autopsy examinations. Two mutations (R4645Q and A612T) in the RYR1 gene were identified. The R4645Q mutation has previously been reported in MH patients, but five heterozygous mutations were also found in 400 Japanese control alleles. The other mutation was novel, and was not found in the same control alleles. The results of this study provide the first successful identification of RYR1 mutations in psychiatric patients suspected at autopsy of having died of NMS. However, the association between RYR1 gene mutations and cause of death in psychiatric patients suspected of dying of NMS remains unclear.     

Mitochondrial DNA and STR typing of matter adhering to an earphone

STR typing and mitochondrial DNA (mtDNA) sequencing were performed on the matter adhering to an earphone found at a crime scene. Experimental studies were carried out using the earphones provided by volunteers. By means of immunohistochemistry, keratinocytes and a portion of nucleated epithelial cells were proven to exist in the contents from the earphones. DNA was extracted by means of the phenol/chloroform method, and the low quantity of extracted DNA was found to be highly degraded. Six STR loci, CSFIPO, TPOX, TH01, F13A01, FESFPS and vWA, were PCR amplified and typed by using two triplex systems (CTT and FFv Multiplexes, Promega, WI), and an amelogenin locus was determined as well. Although partial profiles were observed in some experimental samples, all STR loci could be typed when a considerable amount of high molecular weight DNA was obtained (>0.5 ng/microL). Amplification and sequencing of mtDNA hypervariable region I(15997-16401) and hypervariable region 11(29-408) were all successful. The mitochondrial DNA sequence of the actual case sample, comprising two hypervariable regions and a total of 785 base pairs, showed eight mutations and two insertions with respect to the standard published reference sequence. The genotype was unique in the three published Japanese databases. These results suggest that it is possible to analyze mtDNA from minute amounts of materials and from degraded materials more effectively and routinely in forensic practice.