Forensic Toxicology

Forensic Toxicology dr Paweł Szpot Chair and Department of ForensicMedicine WroclawMedical University

Introduction: Forensic toxicology encompasses the analysis of drugs and chemicals including themost common drugs of abuse and also focuses on the interpretation, that is, the understandingand appreciation of the results of this testing in a medical–legal context.The results of toxicology testing are often presented to courtsfor the adjudication of an issue but are very often misunderstood.We need to remember that a test is not a test. A test result is only as good as thequestion it is asked to answer. Toxicology test results must, therefore, be introducedby qualified toxicologists. The traditional specimens used in testing include blood or its component parts, that is, plasma or serum, and urine. This is in part because these are the easiest to collect. In addition, in the case of blood, or its components, it represents the dynamic state of drug distribution in the body with the best relation to the state of the individual’s pharmacologic condition (therapeutic, impairment, and death). In the case of urine, we have a static fluid that generally does not correlate with the pharmacological effects in an individual, rather it represents high concentrations of drugs and metabolites and demonstrates prior use. Thus, the ready accessibility and knowledge of the pharmacokinetics and distribution of drugs caused toxicologists to focus on these specimens.

Topics: • Biologicalevidencepossible to preserve and use in toxicologicalanalyses. • Toxicsubstancesclassification for the forensicpurposes. • Toxicmetals and theircharacteristics. • Toxicgases and easilyevaporatingsubstances, toxicorganicsolvents. • Toxic non-narcoticsubstancesderived from plants. • Narcoticssubstancesintoxications – characteristics, ways od administering, mechanisms of action, possibilities of detection in organic materials. • Carbon monoxideintoxication – sources, characteristics, mechanism of action, possibilities of detection in organic materials. • Forensicinterpretation of stateafterconsumingethanol, drunkstate and stateafterconsumingnarcoticsubstances. • Possibilities of calculatingethanollevelat a giventime in the past usingresults of toxicologicaltestsperformed.

Biologicalevidencepossible to preserve and use in toxicologicalanalyses. Material should include: • Place of xenobiotic penetration into the body • Blood which distributes xenobiotic to the organs • Place of xenobioticelimination • Place of a possible xenobiotic accumulation To the untargeted study we take: stomachgastricbuffers the small intestinethe content of the small intestinebloodliverbilekidneyurine It’srecommended by the Polish Society of Forensic Medicine and Criminology totake50 gof tissueand 30 ml of tissue fluids

Packing for storing the taken material fortoxicological studies has to be: Chemically pure and inert Hermetically sealed The material should be properly described. The description should include information such as: • Name of deceased • Age (if known)or date of birth • Type of material • Date of Section • Name of person performing theautopsy • The registration number of books autopsy

From: A Simplified Guide to ForensicToxicology

Material type and quantity - specialcases -

Signs of injection Cut about 1-2 cm2 of skin, subcutaneous tissue and the superficial layer of muscle from place which shows signs of injection. This section is inserted into the plastic tube. In comparison, to the other tube you have to cut out the same size piece of skin, but from place which does not contain a trace ofinjection. The fragment with a trace of injection will have higher concentration of xenobiotic that has entered the body by injection

Suspectedcarbonmonoxidepoisoning • In the case of carbon monoxide poisoning collect blood from the right and left ventricle to compare the concentration of carboxyhemoglobin in the blood. If someone died in an atmosphere containing carbon monoxide,therewill be higher concentration of carboxyhemoglobin in the left ventricle (coming fromlungs).

Hair and nails • Hair and nails are a new trend in forensictoxicology.They may be taken for toxicological studies from bodies in whichany of soft tissues havesurvived (e. g. inthe exhumation of the old corpses).

Toxicsubstancesclassification for the forensicpurposes. Classification by Prof. S. Raszeja „Medycyna sądowa”

Anotherclassification • due to changes in the body: The poisons that cause distinct changes: • 1. locally acting - acids, alkalis, bromine, iodine - if they come into contact with body tissue theycausenecrosis and denaturation of proteins • 2. parenchymal - metals, salts of heavy metals, fungi –damagingthe kidneys, liver, heart The poisons that may not result in significant changes (but not necessarily): • 1. The volatile - act mainly on the central nervous system - the solvents, alcohols, hydrocarbons • 2. The blood - carbon monoxide, cyanides, phenols, benzene, medicines, drugs, pesticides • 3. Acting on a functional way - without causing any apparent damage visible at autopsy – drugsand pesticides.

Basic effects of possible multiple interactions concerning the main toxic and/or essential metals and metalloids in mammals Arsenic toxicity is a global health problem affecting millions of people. Contamination is caused by arsenic from natural geological sources leaking into aquifers, contaminating drinking water and may also occur from mining and other industrial processes. Arsenic is present as a contaminant in many traditional remedies. Arsenic trioxide is now used to treat acute promyelocyticleukaemia. Absorption occurs predominantly from ingestion from the small intestine, though minimal absorption occurs from skin contact and inhalation. Arsenic exerts its toxicity by inactivating up to 200 enzymes, especially those involved in cellular energy pathways and DNA synthesis and repair. Acute arsenic poisoning is associated initially with nausea, vomiting, abdominal pain, and severe diarrhoea. Encephalopathy and peripheral neuropathy are reported. Chronic arsenic toxicity results in multisystem disease. Arsenic is a well documented human carcinogen affecting numerous organs.

Basic effects of possible multiple interactions concerning the main toxic and/or essential metals and metalloids in mammals

Toxicgases and easilyevaporatingsubstances, toxicorganicsolvents. • Toxicmethanol, ethyleneglycol, isopropylalcohol • Methyl alcohol - colorless liquid with a slightly sweet, strongaroma,properties very similar to ethyl alcoholtherefore, in industries where it is used, thereare cases of mistaken consumption of alcoholinstead of ethyl. Absorbedthrough the skin, lungs (vapors),the poisoning occurs mostly via the oralroute. In the body breaks down into formaldehydeand then to formic acid, which are verytoxic and lead to metabolic acidosis • Lethal dose is 30 to 150 ml

Methanol • loss of consciousness, impaired breathing,impaired liver function and damage to theoptic nerve and retina causing blurredvision, blindness, may causeswelling of the brain,kidney failure

Isopropyl alcohol • Isopropyl alcohol (2-propanol)colorless, volatile liquid with a bitter characteristic smell. Application: component ofwindow cleaning fluids component of antifreeze fluid for cooling hospitals (liquids to decontaminatethe surface) For a healthy human adult (weighing approximately 80 kg) lethal dose by ingestion of 2-propanol is about 240 ml

Isopropanol • Isopropanolvaporactlocally, irritating the eyes, mucousmembranes of the nose and throatAfteringestioncausesnausea, abdominalpain, vomitingJust as ethanolaffects the central nervous system,causingagitation, blurredvision High concentrations of isopropanol: convulsions, coma, respiratory distress respiratory paralysis Abnormalheartrate, bloodpressure, blood clottingdisorders, liverdamage and kidney

Ethyleneglicol colorless, odorless, oily and sweet liquid,soluble in water and ethanol Toxic dose of 30-50 ml, Lethal dose of 100-150 ml Application:solvent for inkscomponent of antifreezefluid for coolingautomotiveengines, a component of brake fluidpoisoningoccurs by mistakenorsuicidaldrinkingliquidcontainingethyleneglycol

Ethyleneglicol morphological changes: The organs and tissues states the following:swelling,bleedings,Degenerative changes of the heart muscle, liver and kidney,The precipitated calcium oxalate crystals in renal tubular light-histological marker

Carbon monoxidepoisoning Determination of carboxyhemoglobin (CoHb) in the blood • About75% of all poisonings are caused by a suicide from carbon monoxide • Carbon monoxide is a toxic gas, colorless, odorless, it is very difficult for detection • Carbon monoxide is a product of incomplete combustion of organic matter due to insufficient oxygen supply to enable complete oxidation to carbon dioxide (CO2)

Toxicitymechanism of carbonmonoxide • Carbon monoxide connectswith hemoglobin to carboxyhemoglobin (HbCO) in thehuman blood. This prevents oxygen binding to hemoglobin, reducing the oxygen-carrying capacity of the blood andleading to hypoxia

Carbon monoxide is absorbed into the body from the respiratory tract (very rapidly absorbed by the alveoli), the skin Carbon monoxide binds to hemoglobin (Hb), ferritin, and myoglobin 200 - 300 times faster than oxygen Prolonged exposure can lead tobrain damage and even death. • medical opinion is based on the results ofan autopsy or examination and the determination of carboxyhemoglobin in the blood of the deceased • on the basis of determination of blood carboxyhemoglobin obtain information about the cause of the poisoning or death

Clinical Signs & Symptoms associated with COPoisoning and correlated COHblevels

Toxic non-narcotic substances derived from plants • The EuropeanYew (Taxusbaccata, family: Taxaceae) is a coniferous, evergreen poisonousshrubortreecommonlyused for ornamentallandscaping (e.g., parks, gardens, and churchyards) or for pharmaceutical purposes. The toxicnature of the yew plant hasbeenknown for morethan 2000 years. The lethaldose for anadultisreported to be 50 g of yewneedlesequal to 250 mg taxinealkaloidsorapproximately 3 mg taxine per kilogram body weight. • Taxusbaccatacontains a complexmixturecharacterized by the taxane ring system skeleton, including non-alkaloidalditerpenoids (e.g., 10-deactetylbaccatin III), alkaloidalditerpenoids [e.g., paclitaxel (taxol A), taxineB], orphenolicconstituents (e.g., 3,5-dimethoxyphenol), flavonoids (e.g., myricetin), and biflavonoids (e.g., bilobetin), amongothers.

Toxic non-narcoticsubstancesderived from plants • Amanita phalloides, commonly known as the death cap, is a deadly poisonous basidiomycete fungus, one of many in the genus Amanita. Widely distributed across Europe, A. phalloides forms ectomycorrhizas with various broadleaved trees. In some cases, the death cap has been introduced to new regions with the cultivation of non-native species of oak, chestnut, and pine. The large fruiting bodies (mushrooms) appear in summer and autumn; the caps are generally greenish in color, with a white stipe and gills. • These toxic mushrooms resemble several edible species (most notably caesar's mushroom and the straw mushroom) commonly consumed by humans, increasing the risk of accidental poisoning. A. phalloides is one of the most poisonous of all known toadstools. It has been involved in the majority of human deaths from mushroom poisoning, possibly including the deaths of Roman Emperor Claudius in AD 54 and Holy Roman Emperor Charles VI in 1740. It has been the subject of much research, and many of its biologically active agents have been isolated. The principal toxic constituent is α-amanitin, which damages the liver and kidneys, often fatally. • As the common name suggests, the fungus is highly toxic, and is responsible for the majority of fatal mushroom poisonings worldwide. Itsbiochemistry has been researched intensively for decades, and 30 grams (1 oz), or half a cap, of this mushroom is estimated to be enough to kill a human. In 2006, a family of three in Poland was poisoned, resulting in one death and the two survivors requiring liver transplants. Some authorities strongly advise against putting suspected death caps in the same basket with fungi collected for the table and to avoid touching them.Furthermore, the toxicity is not reduced by cooking, freezing, or drying.

Digitalis • A group of medicines extracted from foxglove plants are called digitalin. The use of D. purpurea extract containing cardiac glycosides for the treatment of heart conditions was first described in the English-speaking medical literature by William Withering, in 1785,which is considered the beginning of modern therapeutics. • Digitalis toxicity (Digitalis intoxication) results from an overdose of digitalis and causes nausea, vomiting and diarrhoea, as well as sometimes resulting in xanthopsia (jaundiced or yellow vision) and the appearance of blurred outlines (halos), drooling, abnormal heart rate, cardiac arrhythmias, weakness, collapse, dilated pupils, tremors, seizures, and even death. Bradycardia also occurs. Because a frequent side effect of digitalis is reduction of appetite, some individuals have used the drug as a weight-loss aid.

Artemisiaabsinthium • Artemisia absinthium (absinthium, absinthe wormwood, wormwood, common wormwood, green ginger or grand wormwood) is a species of Artemisia, native to temperate regions of Eurasia and Northern Africa and widely naturalized in Canada and the northern United States. It is grown as an ornamental plant and is used as an ingredient in the spirit absinthe as well as some other alcoholic drinks. • Artemisiaabsinthiumcontainsthujone, a GABAA receptor antagonistthatcancauseepileptic-likeconvulsions and kidneyfailurewheningested in largeamounts. Edgar Degas Dans un café”l'Absinthe” 1873

Tansy • Tansy (Tanacetum vulgare) is a perennial, herbaceous flowering plant of the aster family, native to temperate Europe and Asia. • Tansy was used to treat intestinal worms, rheumatism, digestive problems, fevers, sores, and to “bring out” measles.During the Middle Ages and later, high doses were used to induce abortions.

Mycotoxin • The term 'mycotoxin' is usually reserved for the toxic chemical products produced by fungi that readily colonize crops. One mold species may produce many different mycotoxins, and the same mycotoxin may be produced by several species. • Aflatoxins.The aflatoxins were isolated and characterized after the death of more than 100,000 turkey poults (turkey X disease) was traced to the consumption of a mold-contaminated peanut meal. The four major aflatoxins are called B1, B2, G1, and G2 based on their fluorescence under UV light (blue or green) and relative chromatographic mobility during thin-layer chromatography. Aflatoxin B1 is the most potent natural carcinogen known and is usually the major aflatoxin produced by toxigenic strains.

Forensic ToxicologyofEthyl Alcohol • According to statistics of the National Highway Traffic Safety Administration,17126 persons were killed in alcohol-related accidents in 1996.This constitutes 40.9% of all people killed in traffic accidents that year. Ifone counts only those deaths in which one or more persons had a bloodalcohol content above the legal limit of 0.1% at the time of the accident, thefigure of alcohol-related deaths is reduced to 13,395, or approximately 32%of all fatal crashes. In comparison, approximately 15% of totalroad deaths in Great Britain involve accidents in which one or more peoplehave a blood alcohol content (BAC) over the legal limit of 0.08.

The ethanol concentration exceeds 1‰ in 50% of drivers responsible for accidents; in about 30-40% of them the ethanol level exceeds 1.5 ‰

The insobriety condition at the moment of accident may be demonstrated using blood or respiratory air tests Evidencemethodsto determinate of the ethanol in blood: • head-spacegaschromatogrphy • Stationary analyzers with spectrophotometricdetector IR Alkometr A2.0

dr hab. G. Buszewicz (Lublin)

Alcohol has been a global companion for many hundred of years One important thing to remember about alcohol is that it is a drug, and people get addicted to it. It's known as alcoholism and people who suffer from it are called alcoholics. The truth is, alcoholism is a disease, and it can be very, very hard for alcoholics to stop drinking. So why do people start drinking to begin with?

1)"Drinking helps me deal with my problems.„ 2)"Drinking helps me have a good time.„ 3)"Drinking makes me high and happy.„ 4) "Drinkingiscool.„ 5)"Drinking keeps me warm when it's cold out.„ 6)”I can't have adrinking problem, because I only drink beer andwine, not hard liquor.„ 7)"I only drink because all my friends drink too."

Stateafterconsumingethanoloccurs when the alcohol content in the body is or leads to: 1) blood alcoholcontentfrom 0.2 to 0.5 ‰ or 2) from 0.1 mg to0.25 mg of alcohol in 1 dm3 of breath Drunkstate occurs when the alcohol concentrationin the body is or leads to: • blood alcoholcontentabove 0.5 ‰ of alcohol or • the presence of exhaled above 0.25 mg of alcohol in 1 dm3

Erik M. P. Widmark (1889-1945) was among the first researchers to study in a systematic way the absorption, distribution, and elimination of ethanol in the body; in addition, he formulated his results in mathematical terms. Widmark's research during the first decades of lastcentury paved the way for innovative traffic safety legislation that stipulated punishable limits of alcohol in the blood of a person driving a car.

‰ ~ 0,55 (0,47÷0,64) distributioncoefficient [‰] ~ 0,68 (0,51÷0,85) c0= Ct+β60 x t Cmax 0 tC-max (ok. 30 – 60 min) t ex-post Widmarkequation A = c0 · W · r [g] [kg] β60 = 0,07 – 0,28 (0,1-0,2) ‰/h coefficient of alcohol elimination from the body

Retrospective calculations can be performed in cases which meet the the following conditions: • 1) the concentration of alcohol in the blood (or breath) during the first study must behigher than 0.4 ‰, • 2) the duration of the event to the first study can not be longer than 5-6 hours • 3) At the time the event suspect must be in the elimination phase of alcohol, • 4) the suspect should not consume alcohol between the event and studies ofconcentration of alcohol.

Forensic Toxicology