Fluidity of cadaveric blood after sudden death: Part II. Mechanism of release of plasminogen activator from blood vessels

Experiments by the vascular perfusion technique in isolated hind leg of the dog showed that after addition of various vasoactive drugs to the perfusate, fibrinolytic activity (plasminogen activator level) of the effluent was elevated. The effects of the drugs were completely inhibited only by the specific blocker of their receptor on the vascular wall. High K+ level of the perfusate also induced abundant release of plasminogen activator from the vascular wall.     

Fluidity of cadaveric blood after sudden death: Part I. Postmortem fibrinolysis and plasma catecholamine level

In rats, fibrinolytic activity and plasma catecholamine levels increased rapidly after death. Postmortem fibrinolysis was affected by the method of sacrifice, but catecholamine level was not always affected. Immediately after death, the plasma adrenalin level was higher than the noradrenalin level, but then adrenalin remained unchanged or tended to decrease while noradrenalin tended to increase gradually with time. In human subjects after rapid death, the plasma catecholamine and fibrinolytic activity levels were high. However, in cases after slow death they were low.     

Fluorometric determination of pseudocholinesterase activity in postmortem blood samples

A fluorometric assay using 3-(p-hydroxyphenyl) propionic acid (HPPA) was conducted to determine the activity of pseudocholinesterase (ChE) [Enzyme Commission (EC) No. 3.1.1.8] in postmortem blood samples so as to test for organophosphate poisoning. By the enzymatic reaction of ChE, its substrate, benzoylcholine, produces choline, which is oxidized by choline oxidase to generate hydrogen peroxide. HPPA is oxidized by hydrogen peroxide and peroxidase to become the fluorogenic dimer whose concentration is measured fluorometrically at an excitation emission wavelength of 320 nm and an elimination emission wavelength of 404 nm. The selectivity and sensitivity of the present method were found to be superior to those of conventional pH and spectrophotometric methods.     

Detection and quantification of low levels of benzoylecgonine in equine urine

Cocaine (COC) is a highly addictive plant alkaloid expressing strong psychostimulatory effect. It has no medical use in equine veterinary practice. The contamination of the environment with cocaine such as its presence on the US paper currency has been reported few times. There are anecdotal reports of low benzoylecgonine (BE) concentrations (usually much less than 100 ng/mL) being found in urine of race horses. In order to protect horsemen against harsh penalties associated with the presence of trace amounts of BE in horse urine as a result of environmental contamination, in February 2005 the Illinois Racing Board issued new medication rules that established the threshold level of 150 ng/mL for BE in equine urine. The penalties associated with this rule provide for increasing fines ($250, $500, $1000) with successive positive reports against a trainer for levels of BE below 150 ng/mL. A total of 19,315 urine samples were collected over the 2-year period of time from winning horses (both harness and thoroughbred) at race tracks in Illinois for routine drug screening (ELISA). The presence of BE was confirmed by GC/MS in 28 urine samples (0.14%). The concentration range for BE in harness horses (21 detections) was < 5-91 ng/mL, and for thoroughbred (seven detections) was 7-52 ng/mL. To date, the laboratory has not reported concentrations of BE that exceed the established threshold concentration of 150 ng/mL.     

Levels of cocaine and its metabolites in washed hair of demonstrated cocaine users and workplace subjects

In a study of subjects in drug rehabilitation programs, cocaine and cocaine metabolite levels were determined in the hair of 75 subjects who had produced cocaine-positive urine results. The hair was analyzed after being washed with the 3.75 h wash procedure developed by this laboratory. In addition, results of testing 73 non-users are presented, as well as levels of cocaine, benzoylecgonine (BE), cocaethylene, and norcocaine from workplace population samples. The data support a recommendation of reporting as positive a sample with cocaine of 500 pg/mg hair and either a 5% ratio of benzoylecgonine (BE) to cocaine in samples, or the presence of cocaethylene at 50 pg/mg hair, or norcocaine at 50 pg/mg hair for samples < or =2000 pg cocaine/mg hair. For samples with cocaine present at >2000 pg/mg hair, the data indicate that a ratio of 5% BE may be an overly conservative approach. In appropriately washed hair samples, cocaine users can produce hair levels of <5% BE and thus a minimum BE cutoff in lieu of a ratio could be considered.     

Urinary excretion of benzoylecgonine following ingestion of Health Inca Tea

Four males ingested one cup of Health Inca Tea which contained 1.87 mg of cocaine. Urine specimens collected for 36 h post-ingestion were analyzed for benzoylecgonine (BE) by EMIT-d.a.u., TDx and gas chromatography/mass spectrometry (GC/MS). Positive immunoassay results were obtained for 21-26 h post tea ingestion. Discrepant immunoassay results occurred with only one specimen: EMIT positive; TDx negative, 0.25 mg/l; GC/MS, 0.273 mg/l. Quantitative TDx results were well correlated with GC/MS results, r2 = 0.963, n = 45. Maximum urinary BE concentrations ranged from 1.4-2.8 mg/l, occurring from 4-11 h, post ingestion. Total BE excretion in 36 h ranged from 1.05 to 1.45 mg, 59-90% of the ingested cocaine dose. Urinary excretion rate constant (Km) ranged from -0.073 to 0.111/h. Health Inca Tea ingestion should be considered when interpreting urinary BE concentrations.     

High-performance liquid chromatography with column switching for the determination of cocaine and benzoylecgonine concentrations in vitreous humor

A sample concentration technique was adapted for the determination of cocaine and benzoylecgonine (BE) concentrations in vitreous humor. Vitreous humor (0.5 mL) was diluted 1:1 with water and applied through a filter onto a 3-cm preconcentration column. Following a simple wash step, the analytes were flushed directly onto a reversed-phase analytical high-performance liquid chromatography (HPLC) system. Absolute recoveries were high (above 90%) and the chromatograms were free from interference. Analysis for the drug and its breakdown product was performed using ultraviolet (UV) visible photodiode array detection, which allowed confirmation of peak identity. Recognizable UV spectra could be measured with as little as 20 ng on column. Comparison of the drug levels in 27 blood and vitreous humor samples showed that, while there was only a low correlation between the blood and vitreous concentrations (R = 0.70), vitreous cocaine and BE determinations were good indicators of antemortem cocaine use. In almost all cases, the vitreous BE concentrations were higher than the cocaine concentrations. The technique was easy to perform and the vitreous samples were especially compatible with this low-labor analytical procedure.     

Analysis of nitrite in adulterated urine samples by capillary electrophoresis

A simple method for analyzing nitrite in urine has been developed to confirm and quantify the amount of nitrite in potentially adulterated urine samples. The method involved separation of nitrite by capillary electrophoresis and direct UV detection at 214 nm. Separation was performed using a bare fused silica capillary and a 25 mM phosphate run buffer at a pH of 7.5. Sample preparation consisted of diluting the urine samples 1:20 with run buffer and internal standard, and centrifuging for 5 min at 2500 rpm. The sample was hydrodynamically injected, then separated using -25 kV with the column maintained at 35 degrees C. The method had upper and lower limits of linearity of 1500 and 80 microg/mL nitrite, respectively, and a limit of detection of 20 microg/mL. The method was evaluated using the National Committee for Clinical Laboratory Standards (NCCLS) protocol (Document EP10-A2), and validated using controls, standards, and authentic urine samples. Ten anions, ClO-, CrO4(-2), NO3-, HCO3-, I-, CH3COO-, F-, SO4-, S2O8(-2), and Cl-, were tested for potential interference with the assay. Interferences with quantitation were noted for only CrO4(-2) and S2O8(-2). High concentrations of Cl- interfered with the chromatography. The method had acceptable accuracy, precision, and specificity.     

On the stability of the irritant dibenz-[b,f]-[1,4]-oxazepine (substance CR)

The objective of the present study was to determine the duration and conditions of persistence of the irritant dibenz-[b,f]-[1,4]-oxazepine (substance CR) on the environmental objects. The quantitative analysis of the substance on cotton fabric specimens was carried out using the approved method of high performance liquid chromatography (HPLC) with UV detection at different time periods and envirobmental conditions. It was shown that the main factor determining the lifetime of dibenz-[b,f]-[1,4]-oxazepine on the cotton fabric is the ambient conditions. By way of example, the "open" and "closed", storage of such specimens during 300 days resulted in the decrease of the amount of substance CR to 22.5 and 79% of the initial level respectively. By the end experiment (day 600) these values lowered to 3 and 52.5% respectively. Taken together, the results of the study indicate that dibenz-[b,f]-[1,4]-oxazepine can be described as a substance resistant to environmental impacts. It is comparable in terms of stability with such known irritants as capsicum oleoresin and pelargonic acid morpholide.     

钾中毒死亡家兔的尸体化学变化

目的研究家兔钾中毒死亡后的尸体化学变化特征,为钾中毒的法医学鉴定提供参考。方法采用浓度为0.3%和1%的KCl葡萄糖溶液分别以全速和100滴/min的速度输给家兔,至其死亡,测定家兔输液前后血、尿电解质浓度,比较两种不同输液方式所致钾中毒死亡后尸体化学变化特征。结果输钾前后,家兔血清K (SK)浓度升高,血清Na 、Ca2 、Cl-及HCO3-浓度均降低,全血K (TK)及血清Mg2 浓度变化无显著性差异。0.3%KCl组致死输液时间长于1%组(P=0.006),致死输钾量无显著性差异(P=0.062);TK、血清Na 、Mg2 及Cl-浓度变化值具有显著差异,SK、Ca2 、HCO3-浓度变化值无显著差异;尿量和尿液各电解质浓度指标未见显著性差异。结论尸体SK、TK及血清Mg2 浓度升高,有助于钾中毒的法医学死后诊断。     

挤压伤大鼠血清对血管内皮细胞凋亡的作用及机制探讨

目的观察挤压伤大鼠血清对血管内皮细胞凋亡的影响及可能机制。方法培养小牛主动脉内皮细胞,观察挤压伤大鼠血清对培养的血管内皮细胞凋亡及胞内钙浓度的影响,并检测血浆内皮素-1(endothelin-1,ET-1)及心钠素(atrial natriuretic peptide,ANP)的含量。结果与正常大鼠血清相比,挤压伤大鼠血清致细胞的凋亡百分数由(8.26±1.75)%降为(2.75±0.90)%,胞内钙浓度由(96.98±3.95)nmol/L增加到(118.79±3.22)nmol/L,挤压伤大鼠血浆内皮素-1和心钠素含量显著增高。结论肢体挤压伤大鼠血清可抑制血管内皮细胞的凋亡,此效应可能与内皮素-1和心钠素诱发胞内钙浓度增加有关。     

Positive and negative ion mass spectrometry of tricyclic antidepressants

Positive electron impact (EI), positive chemical ionization (CI), and negative CI mass spectra of eight tricyclic antidepressants are presented. In the positive EI mode, peak(s) at m/z 193 and/or 195, which corresponded to the tricyclic nucleus, appeared for five compounds; a peak at m/z 58 was common to compounds having a gamma-dimethylaminopropyl group as their side chain. Molecular ions appeared for all compounds though they were very small in some compounds in the positive EI mode. In the positive CI mode, [M + H]+ quasi-molecular peaks appeared together with [M + C2H5]+ peaks in five compounds; the ion at m/z 196, which corresponded to the tricyclic nucleus also appeared in five compounds. In the negative CI mode with 1 Torr chamber pressure, [M - 1]- quasi-molecular ions were observed for all compounds except for lofepramine; [M + 43]- anions, which probably corresponded to [M + C3H4]-, appeared in five compounds; strong [M + Cl]- ions appeared in carpipramine and clocapramine. The anions due to the tricyclic nucleus also appeared in this mode. In the negative CI mode at low pressure (0.01 Torr), the spectra were generally similar to those in the 1 Torr negative CI mode. However, the cluster anions never appeared at the low pressure. Some data on extraction for some antidepressants from human urine and plasma, and their separation by gas chromatography, are also presented.     

Validation of an ion-trap gas chromatographic-mass spectrometric method for the determination of cocaine and metabolites and cocaethylene in post mortem whole blood

The coingestion of cocaine (COC) and ethanol is a very frequent occurrence and is known to increase the risk of morbidity and mortality. The formation occurs of a transesterification product, the cocaethylene (CE), which is even more toxic than cocaine. In order to study the role of ethanol as an agent of interaction in lethal cocaine intoxication, and to establish its influence in post mortem cocaine concentrations, an ion-trap gas chromatographic-mass spectrometric method (GC-MS) was validated to quantify simultaneously the agent and its biotransformation products, benzoylecgonine (BE), ecgoninemethylester (EME) and the 'biomarker' of the interaction, the CE present in whole blood. Deuterated internal standards were added to 2 ml of post mortem whole blood and extracted in Bond Elut Certify columns. The residues were evaporated and derivatized with N-methyl-N-t-butyldimethylsilyltrifluoroacetamide (MTBSTFA). Detection was performed by electron impact ionization. The monitored ions were m/z 82/85 for EME-tert-butyldimethylsilyl (TBDMS)/EME-d3-TBDMS; m/z 182/185 for COC/COC-d3; m/z 196/199 for CE/CE-d3 and m/z 282/285 for BE-TBDMS/BE-d3-TBDMS. The limits of detection and quantification were found to be 25 ng and 50 ng ml(-1), respectively, for COC and CE, and 50 and 100 ng ml(-1) for BE and EME. Accuracy was different for each of the compounds, varying from 65 to 98%. The dynamic range of the assay was 50-2000 ng ml(-1).     

Changed compounds of hydroxyzine in a human urine

During a forensic toxicological GC/MS screening, changed compounds of hydroxyzine were found in a man's urine taken about 20 h after ingesting sake (Japanese liquor) and tranquilizers. The structures of those compounds were assigned to 4-chlorodiphenylmethane(I), p-chlorbenzophenone(III), norchlorcyclizine(IV), 7-(p-chloro-alpha-phenylbenzyl)-2-hydroxy-2H,3H-oxazolo-[3,2 -alpha] piperazine(V), 1-(p-chloro-alpha-phenylbenzyl)-4-methylpiperazine-3-one(VI), N-(p-chloro-alpha-phenylbenzyl)-N'-formylmethyl-N'- hydroxyethyl-1,2-ethenediamine(VII) and 1-(p-chloro-alpha-phenylbenzyl)-4-[2-ethoxy(2-ethoxy)-ethyl] piperazine(VIII) on the basis of the fragmentation patterns and relative retention times except VIII which was identified using the synthesized compound. I, V, VI, VII and VIII have not been reported previously, but the possibility of metabolites of hydroxyzine is still uncertain.     

National Institute for Occupational Safety and Health; occupational exposure to inorganic lead: request for comments and information; republication--NIOSH. Request for comments and information relevant to occupational exposure to inorganic lead

NIOSH is reviewing its recommendations contained in the document Criteria for a Recommended Standard.... Occupational Exposure to Inorganic Lead, Revised Criteria--1978 [NIOSH 1978]. The evaluation of recent literature indicates that the NIOSH recommended exposure limit [REL] of 100 micrograms/m3 as an 8-hour time-weighted average [TWA] in that document does not sufficiently protect workers from the adverse effects of exposure to inorganic lead. NIOSH is requesting comments and information relevant to the evaluation of the potential health risks associated with occupational exposure to inorganic lead, as well as case reports or other data that demonstrate adverse health effects in workers exposed to inorganic lead at or below the OSHA permissible exposure limit [PEL] of 50 micrograms/m3 as an 8-hour TWA and any information pertinent to evaluating the technical feasibility of establishing a more protective REL for inorganic lead. NIOSH is also soliciting information on worker blood lead levels [BLLs] including data on methodologies used in measuring BLLs in the workplace and information that can be used for comparing airborne inorganic lead concentrations to observed BLLs. NIOSH intends to analyze the feasibility of developing preventive measures including an REL that would provide better protection for workers. In the interim, NIOSH plans to adopt the more protective current OSHA PEL as its REL.     

Accuracy of age estimation of radiographic methods using developing teeth

Developing teeth are used to assess maturity and estimate age in a number of disciplines, however the accuracy of different methods has not been systematically investigated. The aim of this study was to determine the accuracy of several methods. Tooth formation was assessed from radiographs of healthy children attending a dental teaching hospital. The sample was 946 children (491 boys, 455 girls, aged 3-16.99 years) with similar number of children from Bangladeshi and British Caucasian ethnic origin. Panoramic radiographs were examined and seven mandibular teeth staged according to Demirjian's dental maturity scale [A. Demirjian, Dental development, CD-ROM, Silver Platter Education, University of Montreal, Montreal, 1993-1994; A. Demirjian, H. Goldstein, J.M. Tanner, A new system of dental age assessment, Hum. Biol. 45 (1973) 211-227; A. Demirjian, H. Goldstein, New systems for dental maturity based on seven and four teeth, Ann. Hum. Biol. 3 (1976) 411-421], Nolla [C.M. Nolla, The development of the permanent teeth, J. Dent. Child. 27 (1960) 254-266] and Haavikko [K. Haavikko, The formation and the alveolar and clinical eruption of the permanent teeth. An orthopantomographic study. Proc. Finn. Dent. Soc. 66 (1970) 103-170]. Dental age was calculated for each method, including an adaptation of Demirjian's method with updated scoring [G. Willems, A. Van Olmen, B. Spiessens, C. Carels, Dental age estimation in Belgian children: Demirjian's technique revisited, J. Forensic Sci. 46 (2001) 893-895]. The mean difference (+/-S.D. in years) between dental and real age was calculated for each method and in the case of Haavikko, each tooth type; and tested using t-test. Mean difference was also calculated for the age group 3-13.99 years for Haavikko (mean and individual teeth). Results show that the most accurate method was by Willems [G. Willems, A. Van Olmen, B. Spiessens, C. Carels, Dental age estimation in Belgian children: Demirjian's technique revisited, J. Forensic Sci. 46 (2001) 893-895] (boys -0.05+/-0.81, girls -0.20+/-0.89, both -0.12 y+/-0.85), Demirjian [A. Demirjian, Dental development, CD-ROM, Silver Platter Education, University of Montreal, Montreal, 1993-1994] overestimated age (boys 0.25+/-0.84, girls 0.23+/-0.84, both 0.24 y+/-0.86), while Nolla [C.M. Nolla, The development of the permanent teeth, J. Dent. Child. 27 (1960) 254-266] and Haavikko's [K. Haavikko, The formation and the alveolar and clinical eruption of the permanent teeth. An orthopantomographic study, Proc. Finn. Dent. Soc. 66 (1970) 103-170] methods under-estimated age (boys -0.87+/-0.87, girls -1.18+/-0.96, both -1.02 y+/-0.93; boys -0.56+/-0.91, girls -0.79+/-1.11, both -0.67 y+/-1.01, respectively). For individual teeth using Haavikko's method, first premolar and second molar were most accurate; and more accurate than the mean value of all developing teeth. The 95% confidence interval of the mean was least for mean of all developing teeth using Haavikko (age 3-13.99 years), followed by identical values for Demirjian and Willems (sexes combined).     

The development and evaluation of a 125I radioimmunoassay for the measurement of LSD in body fluids

A radioimmunoassay (RIA) has been developed for the direct detection of LSD in biological fluids. The radiotracer, (+)-2-[125I]iodo-LSD, allows the use of gamma-counting rather than the liquid scintillation counting currently used for existing 3H radioimmunoassays. The assay is specific for LSD and very closely related compounds. It is inexpensive, sensitive, simple to use and small volumes of samples (50 microliter) can be assayed directly without the need for any time-consuming extraction procedures. The cut-off levels are 1.2 ng/ml in blood and 3.0 ng/ml in urine. The results obtained using the 125I assay described in this work compare very favourably with those obtained using the 3H assay currently used by Home Office Forensic Science Laboratories. The advantages of the assay make it a most appropriate method for the routine screening of LSD in biological samples of forensic interest.     

Classification of two steroids, prostanozol and methasterone, as Schedule III anabolic steroids under the Controlled Substance Act. Final rule

With the issuance of this Final Rule, the Administrator of the DEA classifies the following two steroids as "anabolic steroids' under the Controlled Substances Act (CSA): prostanozol (17[beta]-hydroxy-5[alpha]-androstano[3,2-c]pyrazole) and methasterone (2[alpha],17[alpha]-dimethyl-5[alpha]-androstan-17[beta]-ol-3-one). These steroids and their salts, esters, and ethers are Schedule III controlled substances subject to the regulatory control provisions of the CSA.     

Comparison of daily urine,sweat, and skin swabs among cocaine users

This study (1) compares urine, skin swabs, and PharmChek sweat patches for monitoring drug use; (2) measures possible environmental contamination in recent cocaine (COC) users; and (3) evaluates various immunoassays (IA) for screening COC in diverse matrices. Unique aspects include daily urine monitoring of 10 participants for 4 weeks, multiple monitoring methods, analysis for all specimens by IA and gas chromatography (GC)/mass spectrometry (MS), and the potential for continued illicit drug use by participants. Urine served as the "gold standard" specimen for determining drug use. Only cocaine and related substances were detected.Trace amounts of drugs were found on the skin (<50 ng per swab) of urine-negative participants' hands or forehead. In contrast, larger quantities of COC were found on the skin of individuals with BE-positive urines or individuals living with drug users (up to 20 microg per swab). Patch COC amounts among the three regular users (250-9000, 0-240, 160-22,000 ng per patch) exceeded BE (50-950, none, 30-2200 ng per patch). Pre-swabs, valuable for interpreting the source or time frame of positive patch results, contained substantial COC (38-1160, 0-152, 34-762 ng per swab) prior to patch application; therefore, patch results may represent current use, prior use, contamination, or a combination. In three individuals with no indication of cocaine use, false positives (defined as sweat patch positive when urine specimens were <300ng BE/ml) occurred at a 7% rate. Proposed cut-off concentrations of 75 ng cocaine per patch and 300 ng BE/ml urine curtail the incidence of false positives in this limited population. Three immunoassays were compared to screen specimens for cocaine: a modified, manual Microgenics CEDIA; a Cozart ELISA; and an OraSure ELISA. CEDIA's limit of detection (LOD) was 81ng/ml, compared with LODs of 4 ng/ml for the Cozart ELISA and 1.5 ng/ml for the OraSure ELISA. Cozart correlated with OraSure results for COC concentrations <2000 ng per swab (n=117), r(2)=0.79.     

Evaluation of some oxygen, sulfur, and selenium substituted ninhydrin analogues, nitrophenylninhydrin and benzo[f]furoninhydrin

Six ninhydrin analogues containing oxygen, sulfur, and selenium substituents at the C-5 position, 5-(4-nitrophenyl)ninhydrin, and benzo[f]furoninhydrin were evaluated as fingerprint development reagents. The analogues all showed good fingerprint color development but were not superior to ninhydrin in this respect. The benzo[f]furoninhydrin complex was strongly luminescent at room temperature following zinc complexation, while the remaining analogues required cooling to -196 degrees C to produce optimum luminescence. The benzo[f]furo, nitrophenyl, and methyl selenide analogues showed the best potential as fingerprint reagents with the benzo[f]furo analogue comparing favorably with DFO.