Practical aspects of genetic identification of hallucinogenic and other poisonous mushrooms for clinical and forensic purposes.

AIM: To assess the usefulness of a DNA-based method for identifying mushroom species for application in forensic laboratory practice. METHODS: Two hundred twenty-one samples of clinical forensic material (dried mushrooms, food remains, stomach contents, feces, etc) were analyzed. ITS2 region of nuclear ribosomal DNA (nrDNA) was sequenced and the sequen-ces were compared with reference sequences collected from the National Center for Biotechnology Information gene bank (GenBank). Sporological identification of mushrooms was also performed for 57 samples of clinical material. RESULTS: Of 221 samples, positive sequencing results were obtained for 152 (69%). The highest percentage of positive results was obtained for samples of dried mushrooms (96%) and food remains (91%). Comparison with GenBank sequences enabled identification of all samples at least at the genus level. Most samples (90%) were identified at the level of species or a group of closely related species. Sporological and molecular identification were consistent at the level of species or genus for 30% of analyzed samples. CONCLUSION: Molecular analysis identified a larger number of species than sporological method. It proved to be suitable for analysis of evidential material (dried hallucinogenic mushrooms) in forensic genetic laboratories as well as to complement classical methods in the analysis of clinical material.

Effect of occupational lead exposure on α- and γ-tocopherol concentration in plasma.

OBJECTIVES: Changes in enzymatic antioxidant activity are frequently observed in workers occupationally exposed to lead. Few studies have investigated the influence of lead on the non-enzymatic antioxidant system. The aim of our study was to assess the influence of occupational exposure to lead on the plasma concentration of two hydrophobic forms of vitamin E: α-tocopherol and γ-tocopherol. METHODS: A sample of 401 healthy men, aged 19-62, participated in the study. In total, 340 of these subjects were employed at the Mine and Metallurgical Plant in southern Poland. The workers who were occupationally exposed to lead were divided into quartiles (groups of 85 subjects). The lead concentrations in the blood of the subjects in the control group and in the lead exposure quartiles correspond to the following ranges: 10-72 µg/l (control group); 82-206 µg/l (Q1); 209-308 µg/l (Q2); 308-394 µg/l (Q3) and 395-644 µg/l (Q4), respectively. RESULTS: Significant differences were observed only for the plasma concentration of γ-tocopherol, which differed between the control group and Q1 (by 24.1%, p=0.0368), between Q1 and Q3 (by -18.8%, p=0.0115) and between Q1 and Q4 (by -25.7%, p=0.0002). Multiple linear regression analysis showed that the statistically significant, predictive properties of the γ-tocopherol plasma concentration were as follows: triglycerides (ß=0.440)> age (ß=0.131)> whole cholesterol (ß=0.117)> blood lead concentration (ß=-0.108). For α-tocopherol, significant prognostic properties were triglycerides and total cholesterol (ß=0.485 and ß=0.399, respectively). CONCLUSIONS: Occupational exposure to lead is strongly correlated with the concentration of γ-tocopherol but not α-tocopherol.

[Medical analytical toxicology in Poland in year 2012]. / Kliniczna analityka toksykologiczna w Polsce w 2012 roku.

There are 12 centers of acute poisoning treatment and 9 round the clock toxicological laboratories. Most of the laboratories access evidence of activity run by National Clinical Toxicology Consultant. The paper presents actual status of medical toxicology laboratories in Poland and summarizes activity of the laboratories in the year 2012. In 2012 toxicological laboratories reported 113,719 assays. There were diagnosed 63.8% men and 34.8% women. The toxicological laboratories determine most substances and markers of exposition to chemical compounds important for diagnosis and treatment of acute poisonings (i.e. ethanol, methanol, ethylene glycol, acetaminophen, salicylates, anticonvulsants, carboxyhemoglobin, methemoglobin). There is not possible to determine heavy metals, all medicines and "designed" drugs of abuse in all laboratories. Limited access to reference methods, that enable to confirm results obtained by screening methods (immunological cassette and strip tests) is also a problem.

[Plasma lipid concentration in smoking and nonsmoking male adults treated from alcohol addiction]. / Zmiany profilu lipidowego w osoczu mezczyzn o róznym statusie palenia tytoniu leczonych z powodu uzaleznienia od alkoholu.

Alcohol drinking and tobacco smoking affect plasma lipid levels and are both independent risk factors of cardiovascular diseases. Alcohol and nicotine addictions are more common among man than women in Poland. The aim of the study was to evaluate changes in plasma lipid levels after cessation of heavy drinking in smoking and nonsmoking Polish male adults. Subjects were recruited from individuals who participated in an inpatient addiction program following alcohol detoxification. We recruited 119 male adults: 48 non-smokers in age between 31 and 60 years (mean 48.7 +/- 8.8) and 71 smokers in age between 30 and 60 years (mean 46.1 +/- 7.8). Each subjects provided three blood samples: at baseline, after 3 weeks, and after 6 weeks of treatment. Plasma samples were analyzed for lipids by manual precipitation and automatic enzymatic methods. Changes in plasma lipid concentrations were analyzed using two-way analysis of variances with repeated measures with smoking status as between subjects factor and time post alcohol cessation as within-subject factors. All analyses were adjusted for age, and BMI. We found that plasma levels of HDL decreased in smoking and nonsmoking subjects by 30% and 24%, respectively (p < 0.001). In smoking subjects, plasma levels of triglycerides and LDL increased significantly after 6 weeks post cessation of heavy drinking cessation by 17% and 16%, respectively (p = 0.001). We also found that total cholesterol levels remained high in smoking subjects, but decreased significantly by 7% (p = 0.022) in nonsmoking subjects after 6 weeks post cessation of heavy drinking. We concluded that cigarette smoking increased LDL and inhibited the decline in plasma cholesterol among subjects addicted to alcohol following cessation of heavy drinking. Alcohol addiction therapy should be complemented with smoking cessation to prevent increase in cardiovascular risk.

[Plasma fibrinogen concentration in smoking and nonsmoking patients treated from alcohol addiction]. / Zmiany stezenia fibrynogenu w osoczu pacjentów o róznym statusie palenia tytoniu leczonych z powodu uzaleznienia od alkoholu.

Cigarette smoking is common among persons addicted to alcohol. Both tobacco smoking and alcohol binge drinking are risk factors of many cardiovascular conditions. The risk of cardiovascular events decreases after alcohol cessation. However little is known about the effect of continues smoking on biomarkers of adverse cardiovascular events among patients treated from alcohol addiction. The aim of the study was to assess fibrinogen changes after alcohol drinking cessation among cigarette smokers and non-smokers. Total of 239 patients treated from alcohol addiction in Addiction Treatment Center (OTU) Parzymiechy, Poland were included in the study. There were total of 39 women: 11 non-smoking women, in the age range of between 31 and 59 years (mean age 47 +/- 9 years) and 28 smoking women in the age range of 31-60 years (mean age 43 +/- 8 years). Among 200 men, there were 150 smokers in the age range of between 30 and 60 years (mean age 44 +/- 8 years) and 50 non-smokers in the age range of 31 and 60 years (mean age 49 +/- 9 years). We found that among non-smoking patients fibrinogen levels remained unchanged three weeks post alcohol cessation (3.42 vs. 3.49 g/l) but after six weeks significantly decreased to the level of 3.09 g/l (p=0.00085). Among smoking patients fibrinogen levels increased after three weeks post alcohol cessation by 7.9% (z 3.41 do 3.68 g/l) and went back to a baseline level of 3.50 g/l. However those changes were not statistically significant. We found that alcohol cessation leads to decrease of fibrinogen levels only among non-smoking patients post alcohol cessation. A risk of cardiovascular diseases seemed to remain elevated among smokers treated from alcohol addiction. There is need for concomitant treatment of tobacco addiction among smoking alcoholics.

[Fatal colchicine poisoning--case report and review of literature]. / Smiertelne zatrucie kolchicyna--opis przypadku i przeglad literatury.

Colchicine is a natural pseudo-alkaloid found in plants such as the autumn crocus (Colchicum autumnale) and glory lily (Gloriosa superba), which is used to treat gout and some other rheumatological disease. Colchicine binds to tubuline and prevents its polymerization into microtubules. It is thus able to impair those cellular functions that involve microtubules, eg. it arrests mitosis in metaphase. Tissues with high mitotic activity are preferentially affected. We report suicidal colchicine poisoning leading to death after 61 hours. Clinical course was typical for colchicine action. We observed severe diarrhea, cardiovascular shock, ARDS, multiorgan system failure and DIC. Postmortem toxicological studies confirm colchicine poisoning.

[Opinionating difficulties in cases concerning the use of psychoactive agents by traffic participants]. / Trudnosci opiniodawcze w sprawach dotyczacych substancji psychoaktywnych w ruchu drogowym.

In recent years, there has been observed an increasing number of traffic participants being under the influence of drugs or other than alcohol agents that affect the central nervous system. In the period from 1997 until June 2006, 435 blood samples collected from traffic participants suspected of having ingested psychoactive agents were examined in the Forensic Medicine Department, Medical University of Silesia, Katowice. Eighty-five blood samples were positive.

Effect of L-carnitine supplementation on xenobiotic-metabolizing hepatic enzymes exposed to methanol.

The study aimed to evaluate the effect of L-carnitine on hepatic cytochrome P450-dependent monooxygenases exposed to methanol. Male Spraque-Dawley rats were given methanol (1/4 LD50 and 1/2 LD50) together with L-carnitine (1g/kg body weight). The parameters of microsome electron transport chains I and II and the levels of CYP2E1, CYP2B1/2 and CYP1A2 were measured 8, 12, 24, 48, 72 and 96 h after exposure. L-carnitine did not affect cytochrome P450 but it significantly increased at 72 and 96 h NADPH-cytochrome P450 reductase. It stimulated cytochrome b5 at 48 and 96 h and NADH-cytochrome b5 reductase activity at 12, 72 and 96 h. Methanol, especially the lower dose, inhibited cytochrome P450 after 48 h, but the higher methanol dose inhibited NADH-cytochrome b5 reductase activity in this time. L-carnitine, combined with the lower dose of methanol, stimulated NADPH-cytochrome P450 reductase after 48 h and cytochrome b5 and NADH-cytochrome b5 reductase over the whole period of observation. L-carnitine stimulated CYP2B1/2 but not CYP2E1 and CYP1A2. Methanol stimulated CYP2E1 at 24 h, but CYP1A2 at 96 h in the studied doses. CYP2B1/2 was induced by the lower dose of methanol at 24 h but by the higher one at 96 h. When given together, L-carnitine and methanol (1/2 LD50) significantly stimulated CYP2E1 up to 170% at 24 h and 145% at 96 h.

[The role of ethanol in complex poisonings with carbon monoxide and hydrogen cyanide in fire victims]. / Rola etanolu w kompleksowych zatruciach tlenkiem wegla i cyjanowodorem u ofiar pozarów.

A total of 230 cases of deaths in burning spaces dating from the years 1995-2003 were investigated in Forensic Medicine Department, Silesian University of Medicine, Katowice. HbCO and HCN found in 177 blood samples ranged from 4-95 % (mean, 31,5 %) and 0,5-40,3 microg/ml (mean, 9,98 microg/ml), respectively. Moreover, ethanol was found in 122 blood samples. Its concentration ranged from 0,89-5,0 per thousand (mean, 1,45 per thousand). A comparative analysis of HbCO and HCN levels in the groups with and without ethanol showed that the range and the mean concentration of both these xenobiotics were higher in the group with no alcohol. It was also shown that the increased ethanol caused a drop in HbCO and HCN levels. To evaluate HbCO and HCN levels, the regression and correlation analysis was used.

Interaction between blood selenium concentration and a levels of oxidative stress and antioxidative capacity in healthy children.

The study aimed at defining the relationship between blood selenium concentration (Se-B) and levels of oxidative stress and antioxidative capacity in healthy children. The studies were conducted on 337 children (mean age: 8.53±1.92 years). The groups of individuals with Se-B <1st quartile (group I, Se-B<70µg/L), with Se-B fitting the range of 1st quartile and median (group II, Se-B: 70-76.9µg/L), with Se-B between the median and 3rd quartile (group III, Se-B: 77-83.9µg/L) and those with Se-B above the 3rd quartile (group IV, Se-B≥84µg/L) were distinguished. Level of oxidative stress was defined using determination of urine malonyldialdehyde concentration (MDA) and urine 8-hydroxy-2-deoxyguanosine concentration (8-OHdg). Urine total antioxidant status (TAS) was determined. In group IV TAS was significantly higher than in groups I-III. A positive correlation was detected between Se-B and TAS. In healthy children an appropriately high Se-B seems to ensure higher total antioxidative status.

The influence of L-carnitine on methanol biotransformation in rats.

There persists a need for potent and safe inhibitors of alcohol dehydrogenase (ADH), to effectively treat methanol poisoning by slowing its rate of biotransformation to there toxic products, formaldehyde and formic acid. Only a few former papers have reported on the significant effectiveness of L-carnitine in treating ethanol poisoning as well as alcohol abuse. As are no reports on the effectiveness of L-carnitine in treating methanol poisoning till now, the current studies were conducted to investigate the influence of L-carnitine on both oxydative metabolism and elimination of methanol in rats. Male Sprague-Dawley rats, aged 3 months with the body weight of 200-230 g were divided into 6 groups at random, with two of the groups considered to be control. Rats were given drinking water (control) or methanol in two different doses of 3220 mg/kg b.m. or 6440 mg/kg b.m. intragastrically and 0.9% NaCl (control) or 6.2 mmol/kg b.m. of L-carnitine intraperitionelly. Within 96 hours after the administration of methanol and 0.9% NaCl or L-carnitine, the urine was collected and then the animals were decapitated. To determine methanol there were taken blood samples for clot, and to determine carnitine and its derivatives blood was taken into heparinized test tubes. During the autopsy liver was also secured. In all the experimental time points stated the methanol concentrations in blood, urine and liver homogenate were determined by a head-space gas chromatography.

[Diagnostic importance of formic acid detection in methyl alcohol poisoning]. / Znaczenie diagnostyczne oznaczania kwasu mrówkowego w zatruciu alkoholem metylowym.

In the paper the authors have presented a possible use of formic acid detection in biological specimens in the diagnosis of methanol poisonings. Formic acid was determined as a volatile methyl formate ester by the gas chromatographic head-space method. Based on opinions relating to methanol poisonings, formulated in the Forensic Medicine Department, Silesian School of Medicine, Katowice a potential application of the method mentioned above to forensic medicine was shown, especially in cases of late deaths after methanol intoxication and also a possibility of its use in clinical evaluation of the poisoning phase as well as monitoring the course of treatment.

[Accidents in the workplace caused by alcohol intoxication]. / Wypadkowosc w pracy w zwiazku z nietrzezwoscia.

Fatal accidents in the workplace can be caused by work conditions, aggravation of a chronic disease or alcohol intoxication. The purpose of this paper was to show the influence of ethyl alcohol on accidents in the workplace with regard to the occupation and age of the examined individuals. A group of victims whose deaths resulted from other external factors (suicide, poisoning by non alcoholic agents, etc.) was separated and not included. Statistical analysis of the autopsies carried out in the Forensic Medicine Department, Silesian University of Medicine, Katowice in the years 1992-2001 showed that accidents in the workplace amounted to 4-6% of the total number of deaths in the space of a year with alcohol (ethanol) being the causative factor in 3-15%.

Effect of methanol on endogenous and exogenous carnitine levels in rat plasma.

The effect of methanol on the levels of endogenous carnitine and its derivatives was studied in male Sprague-Dawley rats aged three months. In addition, the effect of L-carnitine supplementation on metabolic disturbances caused by methanol intoxication was studied. The rats were randomized into six groups, including two control groups. Methanol was given at 1/4 LD(50) and 1/2 LD(50)/kg b.w. (or water in control) through an intragastric tube, and L-carnitine (or 0.9% NaCl in the control) was injected intraperitoneally. The levels of plasma L-carnitine and its derivatives were measured at selected time points for four days. Following methanol administration, the rats exhibited dose-dependent increases in L-carnitine levels and altered ratios of L-carnitine and its derivatives. L-carnitine supplementation accelerated the normalization of metabolic disturbances, as indicated by the acylcarnitine to free carnitine ratio (AC/FC). The protective effect of L-carnitine is supported by the fact that 100% of the methanol-treated rats supplemented with carnitine survived, while 8/60 rats and 27/101 rats died at methanol doses of 1/4 LD(50) and 1/2 LD(50), respectively, in groups without L-carnitine supplementation.