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Forensic Toxicology
Toxicologists
Detect and identify the presence of drugs
and poisons in body fluids, tissues, and
organs
Work in health facilities such as hospitals
and clinics as well as crime labs
Most frequently asked to determine the
presence of alcohol in the body
Alcohol is the most
widely abused drug in
western countries.
Due to prevalence of
abuse, laws have
been instituted to
regulate driving while
under the influence –
intoxication.
The degree of intoxication can be
influenced by:
Body weight
Rate of absorption through stomach and small
intestines directly into the blood stream
Influenced by presence/absence of food
Total time taken to consume the alcohol
Type of alcohol consumed – diluted/undiluted
The longer total time required for complete
absorption, the lower will be the peak alcohol
concentration
Methods had to be
developed to assess
levels of intoxication
at the time of arrest
These had to be
reliable to withstand
court presentation
What does Alcohol do to the Body?
It depresses the CNS – Brain
Extent of depression is directly proportional to
concentration of alcohol within the nerve cells
Specifically the forebrain
Eventually alcohol will affect the central and
rear portions of the brain
Most resistant and last to fail are in the
medulla which regulates respiration and heart
activity
Most tests rely on blood – the medium for
circulating alcohol throughout the body
Research confirms the direct proportional
relationship of:
Blood alcohol concentration to brain alcohol
concentration
Maximum Blood Alcohol Content (BAC) may
not be reached until 2-3 hours after time of
consumption – more likely within 30-90 min.
from final drink until absorption process is
complete
What does Alcohol do to the Body?
Alcohol will become uniformly distributed
throughout the watery portion of the body
at complete absorption – 2/3 of body
volume
Liver begins the elimination process of 95-
98% of the alcohol by oxidation
In the presence of the enzyme alcohol
dehydrogenase, alcohol is converted into
acetaldehyde and then to acetic acid which is
oxidized to carbon dioxide and water
What does Alcohol do to the Body?
Remaining 2-5% is excreted unchanged in
the breath, urine, and perspiration.
Amount exhaled is in direct proportion to
concentration of alcohol in the blood
Using Henry’s Law, Alveoli air is approximately
2,100:1 – alcohol content in 1 ml of blood =
2,100 ml of alcohol content in alveoli air
Elimination or “burn off” rate can vary by
30% between individuals, but average is
0.015% w/v (weight/vol.) per hour
What does Alcohol do to the Body?
Alcohol’s movement through the circulatory
system
After alcohol is ingested it moves down the
esophagus into the stomach – where about 20% of
the alcohol is absorbed through the stomach lining
into the portal vein.
Remaining 80% is absorbed into the blood through
the lining of the small intestines
Once in the blood it is transported to the liver, heart,
lungs, back to heart to be distributed to all parts of
the body
What does Alcohol do to the Body?
1. Blood rich in carbon dioxide is pumped
from the heart into the lungs through the
pulmonary arteries. (Arteries are blood
vessels carrying blood away from the heart;
veins are blood vessels carrying blood to the
heart.)
2. In the lungs, CO2 in the blood is
exchanged for O2.
3. The oxygen-rich blood is carried back to
the heart through the pulmonary veins.
4. This oxygen-rich blood is then pumped
from the heart to the many tissues and
organs of the body, through the systemic
arteries.
5. In the tissues, the arteries narrow to tiny
capillaries. Here, O2 in the blood is
exchanged for CO2.
6. The capillaries widen into the systemic
veins, which carry the carbon-dioxide-rich
blood back to the heart.
Circulatory/Respiratory System
Circulatory/Respiratory System
During period of alcohol absorption,
alcohol concentration within arterial
(oxygenated) blood will be considerably
higher than in venous (deoxygenated)
blood.
Blood drawn for testing comes from the
veins in the arm. The true level of alcohol
may not be accurate during the early
phases of absorption. Alcohol levels will
equalize as absorption continues into later
and final stages.
Breath-Test Instruments
Breathalyzer Test
Widely used until
1990
A device for collecting
and measuring the
alcohol content of
alveolar breath
A subject blows into a mouthpiece that
leads to a metal cylinder which causes a
piston to rise to a height that exposes two
vent holes. When collection of alveolar
breath is complete the piston drops back
into place trapping the collected ~52.5 ml
or 1/40 of 2.100 ml of alveolar breath.
Breathalyzer Test
This trapped alveolar breath can then be
analyzed as it passes into a glass ampoule
containing 3 ml of 0.025% potassium
dichromate and 0.025% silver nitrate in
sulfuric acid and water.
Any alcohol present in the sample
dissolves in the dichromate solution and is
oxidized to acetic acid.
Breathalyzer Test
The oxidation process destroys the potassium
dichromate – the extent of which is measured by
the Breathalyzer and relates to the quantity of
alcohol passed into the ampoule.
This is a spectrophotometer specially designed
to measure the absorption of light passing
through the potassium dichromate solution at a
single wavelength.
Breathalyzer Test
REACTION:
2K2Cr2O7 + 3C2H5OH + 8H3SO4
Potassium Ethyl Sulfuric
dichromate alcohol acid
2Cr2(SO4)3 + 2K2SO4 + 3CH3COOH + 11H2O
Chromium Potassium Acetic Water
sulfate sulfate Acid
Breathalyzer Test
Potassium dichromate
Yellow in color and absorbs visible light
wavelengths of 420 nm
The Breathalyzer indirectly determines the
quantity of alcohol consumed by
measuring the absorption of light by
potassium dichromate before and after its
reaction to alcohol
Breathalyzer Test
Newer Technology - Infrared
Designed to minimize operator
error and free of chemical
reagents
Operates on infrared light
absorption and/or fuel cells
Infrared light absorption uses
filters selected to a wavelength of
infrared light at which alcohol will
absorb.
Greater alcohol concentration
decreases intensity of light measured
by a photoelectric detector
Infrared light signal is proportional to
concentration of alcohol present in captured
breath sample
Processed by an electronic microprocessor and
% BAC is displayed in a digital readout on a card
for permanent record
A second simultaneous infrared wavelength is
also used to check for acetone or other chemical
interferences and warns operator of the
presence of an “interferant”
Newer Technology - Infrared
Fuel cell converts a fuel and an oxidant
into an electrical current.
Breath alcohol = the fuel
Atmospheric oxygen (O2) = oxidant
Alcohol is converted into acidic acid within the
fuel cell which generates a current that is
proportional to the quantity of alcohol present
in the breath.
Device is microprocessor controlled which reduces
operator error and provides a printout for
permanent record
Newer Technology – Fuel Cell
Newer Technology – Fuel Cell
Newer Technology
Both techniques utilize a slope detector
which insures that the sample
administered is from deep long breaths
which relate to the true BAC of the subject
being tested
Field Sobriety Tests
Normally performed to ascertain the
degree of the suspect’s physical
impairment and whether or not an
evidential test is justified.
Roadside breath tests
Preliminary and non-evidential
Probable cause for later tests
Psychophysical tests – reliable and effective
Horizontal gaze nystagmus,
Involuntary jerking of eye as it moves to side as
intoxication increases the ability of the eye to follow an
object as it moves to the side without jerking decreases.
If BAC is at .10%, eye jerking will begin at 45° angle as
BAC increases jerking will begin at increasingly small
angles
Walk and turn – a divided attention task
Tests subject’s ability to comprehend and simultaneously
carry out more than two instructions. This ability
decreases with increasing consumption of alcohol
One leg stand – a divided attention task
Field Sobriety Tests
Analysis of Blood for Alcohol
Gas Chromatography
Alcohol is separated from other volatiles in blood.
Compares resultant alcohol peak area to ones
obtained with known blood-alcohol standards
Can be calculated with a high degree of accuracy
Oxidation of alcohol to acetaldehyde in the presence
of the enzyme alcohol dehydrogenase ad a coenzyme
nicotin-amide-adenine dinucleotide (NAD). As
oxidation proceeds, NAD NADH. The extent of
conversion is related to alcohol concentration.
Collection and Preservation of
Blood – Live Suspect
Must be drawn under medically accepted
conditions by a qualified person
Must use a sterile needle or lancet
Cleanse with a non-alcoholic disinfectant
such as:
Aqueous benzalkonium chloride (Zepiran)
Aqueous mercurie chloride
Providone-iodine (Betadine)
Seal blood sample in an airtight container
with an anticoagulant (EDTA or Potassium
oxalate) and preservative (sodium
fluoride)
Place in refrigerated storage for delivery to
toxicology ASAP
If procedures are not strictly adhered to,
then BAC decreases and never increases –
so sloppy police and lab work benefits the
suspect.
Collection and Preservation of
Blood – Live Suspect
Requires added precautions
Must account for bacterial action so collect
from a number of different body sites.
Each sample is kept in separate airtight containers
with an anticoagulant, sodium fluoride, and
refrigerated.
Collect urine and vitreous humor fluids – both
usually do not suffer from postmortem ethyl
alcohol production to any significant extent.
Collection and Preservation of
Blood – Postmortem Samples
Alcohol and the Law
A BAC of .08% or
.10% in most states
in equivalent to
intoxication for most
drivers
Federal standard for
bus and truck
operators is .04%
Implied consent comes with obtaining a license
to operate a motor vehicle
Driver has a choice – submit to test or be
subject to loss of license for a given period of
time.
Schmerber v. California
Court ruled against defendant – the Fifth Amendment
only prohibits self-incrimination. Being compelled to
furnish “physical” evidence such as fingerprints,
photographs, measurements, and blood samples are
not protected by the Fifth Amendment
Alcohol and the Law
Role of the Toxicologist
First attempt to narrow down type of toxic substance
based on victim’s symptoms, postmortem pathological
exam, exam of victim’s personal effects, nearby
presence of empty drug containers or household
chemicals
Next, use general screening procedures
Test for small quantities that may be remaining
Must be aware that the body always processes
substances – few substances enter and completely leave
the body in the same chemical state – most are
metabolized – so test for the metabolite
Must also assess for substance’s toxicity
Toxicological Techniques
90% of drugs analyzed is for alcohol and
cocaine
The toxicologist must devise an analytical
scheme that will successfully detect, isolate, and
specifically identify a toxin
Large number of drugs are either acidic (H+) or
alkaline (OH-)
By controlling the pH of a water solution into which
blood, urine, or tissues are dissolved, the toxicologist
can control the type of drug that will be recovered.
Acidic Drugs
Acid drugs are extracted from an acidified
water solution with organic solvents such
as chloroform.
Barbiturates
Acetylsalicylic acid (aspirin)
Alkaline (Basic) - Drugs
Basic drugs are extracted from an alkaline
water solution with organic solvents
Phencyclidine
Methadone
Amphetamines
Cocaine
Sample
Acidic Drugs Basic Drugs
Extraction at appropriate pH
SCREENING TEST
Immunoassay
Gas Chromatography
Thin-Layer Chromatography
CONFIRMATION TEST
Gas Chromatography-Mass Spectrometry
Heavy Metal Poisons
Rarely encountered
Arsenic, bismuth, antimony, mercury, and
thallium
Extraction into HCL solution – insert a copper
strip (Reinsch Test) – positive presence of a
heavy metal will develop a silvery or dark
coating on the copper.
Confirm with atomic absorption
spectrophotometry, emission spectroscopy, or
X-Ray differaction
Carbon Monoxide – A Commonly
Encountered Poison
CO binds with hemoglobin carboxyhemoglobin
Methods to determine concentration of CO in the
blood:
Spectrophotometric methods examine bloods visible
spectrum to determine the relative amount of
carboxyhemoglobin to oxyhemoglobin or total
hemoglobin
Treat a volume of blood with a reagent to liberate the
CO then measure with gas chromatography.
Measured as percent saturation – 50-60% saturation
is considered fatal when combined with .2% BAC 35-
45% saturation maybe fatal
Smokers usually have an 8-10% saturation
In fire evaluation – determination of victim’s
condition prior to fire.
If victim was alive at time of fire they would
have a high % CO saturation
If victim was dead at time of fire they would
have low % CO saturation since they could not
breath in any of the fumes
Carbon Monoxide – A Commonly
Encountered Poison

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cha_6_-_toxicologia y visión forense.ppt

  • 2. Toxicologists Detect and identify the presence of drugs and poisons in body fluids, tissues, and organs Work in health facilities such as hospitals and clinics as well as crime labs Most frequently asked to determine the presence of alcohol in the body
  • 3. Alcohol is the most widely abused drug in western countries. Due to prevalence of abuse, laws have been instituted to regulate driving while under the influence – intoxication.
  • 4. The degree of intoxication can be influenced by: Body weight Rate of absorption through stomach and small intestines directly into the blood stream Influenced by presence/absence of food Total time taken to consume the alcohol Type of alcohol consumed – diluted/undiluted The longer total time required for complete absorption, the lower will be the peak alcohol concentration
  • 5. Methods had to be developed to assess levels of intoxication at the time of arrest These had to be reliable to withstand court presentation
  • 6. What does Alcohol do to the Body? It depresses the CNS – Brain Extent of depression is directly proportional to concentration of alcohol within the nerve cells Specifically the forebrain Eventually alcohol will affect the central and rear portions of the brain Most resistant and last to fail are in the medulla which regulates respiration and heart activity
  • 7. Most tests rely on blood – the medium for circulating alcohol throughout the body Research confirms the direct proportional relationship of: Blood alcohol concentration to brain alcohol concentration Maximum Blood Alcohol Content (BAC) may not be reached until 2-3 hours after time of consumption – more likely within 30-90 min. from final drink until absorption process is complete What does Alcohol do to the Body?
  • 8. Alcohol will become uniformly distributed throughout the watery portion of the body at complete absorption – 2/3 of body volume Liver begins the elimination process of 95- 98% of the alcohol by oxidation In the presence of the enzyme alcohol dehydrogenase, alcohol is converted into acetaldehyde and then to acetic acid which is oxidized to carbon dioxide and water What does Alcohol do to the Body?
  • 9. Remaining 2-5% is excreted unchanged in the breath, urine, and perspiration. Amount exhaled is in direct proportion to concentration of alcohol in the blood Using Henry’s Law, Alveoli air is approximately 2,100:1 – alcohol content in 1 ml of blood = 2,100 ml of alcohol content in alveoli air Elimination or “burn off” rate can vary by 30% between individuals, but average is 0.015% w/v (weight/vol.) per hour What does Alcohol do to the Body?
  • 10. Alcohol’s movement through the circulatory system After alcohol is ingested it moves down the esophagus into the stomach – where about 20% of the alcohol is absorbed through the stomach lining into the portal vein. Remaining 80% is absorbed into the blood through the lining of the small intestines Once in the blood it is transported to the liver, heart, lungs, back to heart to be distributed to all parts of the body What does Alcohol do to the Body?
  • 11. 1. Blood rich in carbon dioxide is pumped from the heart into the lungs through the pulmonary arteries. (Arteries are blood vessels carrying blood away from the heart; veins are blood vessels carrying blood to the heart.) 2. In the lungs, CO2 in the blood is exchanged for O2. 3. The oxygen-rich blood is carried back to the heart through the pulmonary veins. 4. This oxygen-rich blood is then pumped from the heart to the many tissues and organs of the body, through the systemic arteries. 5. In the tissues, the arteries narrow to tiny capillaries. Here, O2 in the blood is exchanged for CO2. 6. The capillaries widen into the systemic veins, which carry the carbon-dioxide-rich blood back to the heart. Circulatory/Respiratory System
  • 13.
  • 14. During period of alcohol absorption, alcohol concentration within arterial (oxygenated) blood will be considerably higher than in venous (deoxygenated) blood. Blood drawn for testing comes from the veins in the arm. The true level of alcohol may not be accurate during the early phases of absorption. Alcohol levels will equalize as absorption continues into later and final stages.
  • 16. Breathalyzer Test Widely used until 1990 A device for collecting and measuring the alcohol content of alveolar breath
  • 17. A subject blows into a mouthpiece that leads to a metal cylinder which causes a piston to rise to a height that exposes two vent holes. When collection of alveolar breath is complete the piston drops back into place trapping the collected ~52.5 ml or 1/40 of 2.100 ml of alveolar breath. Breathalyzer Test
  • 18. This trapped alveolar breath can then be analyzed as it passes into a glass ampoule containing 3 ml of 0.025% potassium dichromate and 0.025% silver nitrate in sulfuric acid and water. Any alcohol present in the sample dissolves in the dichromate solution and is oxidized to acetic acid. Breathalyzer Test
  • 19. The oxidation process destroys the potassium dichromate – the extent of which is measured by the Breathalyzer and relates to the quantity of alcohol passed into the ampoule. This is a spectrophotometer specially designed to measure the absorption of light passing through the potassium dichromate solution at a single wavelength. Breathalyzer Test
  • 20. REACTION: 2K2Cr2O7 + 3C2H5OH + 8H3SO4 Potassium Ethyl Sulfuric dichromate alcohol acid 2Cr2(SO4)3 + 2K2SO4 + 3CH3COOH + 11H2O Chromium Potassium Acetic Water sulfate sulfate Acid Breathalyzer Test
  • 21. Potassium dichromate Yellow in color and absorbs visible light wavelengths of 420 nm The Breathalyzer indirectly determines the quantity of alcohol consumed by measuring the absorption of light by potassium dichromate before and after its reaction to alcohol Breathalyzer Test
  • 22. Newer Technology - Infrared Designed to minimize operator error and free of chemical reagents Operates on infrared light absorption and/or fuel cells Infrared light absorption uses filters selected to a wavelength of infrared light at which alcohol will absorb. Greater alcohol concentration decreases intensity of light measured by a photoelectric detector
  • 23. Infrared light signal is proportional to concentration of alcohol present in captured breath sample Processed by an electronic microprocessor and % BAC is displayed in a digital readout on a card for permanent record A second simultaneous infrared wavelength is also used to check for acetone or other chemical interferences and warns operator of the presence of an “interferant” Newer Technology - Infrared
  • 24. Fuel cell converts a fuel and an oxidant into an electrical current. Breath alcohol = the fuel Atmospheric oxygen (O2) = oxidant Alcohol is converted into acidic acid within the fuel cell which generates a current that is proportional to the quantity of alcohol present in the breath. Device is microprocessor controlled which reduces operator error and provides a printout for permanent record Newer Technology – Fuel Cell
  • 25. Newer Technology – Fuel Cell
  • 26. Newer Technology Both techniques utilize a slope detector which insures that the sample administered is from deep long breaths which relate to the true BAC of the subject being tested
  • 27. Field Sobriety Tests Normally performed to ascertain the degree of the suspect’s physical impairment and whether or not an evidential test is justified. Roadside breath tests Preliminary and non-evidential Probable cause for later tests
  • 28. Psychophysical tests – reliable and effective Horizontal gaze nystagmus, Involuntary jerking of eye as it moves to side as intoxication increases the ability of the eye to follow an object as it moves to the side without jerking decreases. If BAC is at .10%, eye jerking will begin at 45° angle as BAC increases jerking will begin at increasingly small angles Walk and turn – a divided attention task Tests subject’s ability to comprehend and simultaneously carry out more than two instructions. This ability decreases with increasing consumption of alcohol One leg stand – a divided attention task Field Sobriety Tests
  • 29.
  • 30. Analysis of Blood for Alcohol Gas Chromatography Alcohol is separated from other volatiles in blood. Compares resultant alcohol peak area to ones obtained with known blood-alcohol standards Can be calculated with a high degree of accuracy Oxidation of alcohol to acetaldehyde in the presence of the enzyme alcohol dehydrogenase ad a coenzyme nicotin-amide-adenine dinucleotide (NAD). As oxidation proceeds, NAD NADH. The extent of conversion is related to alcohol concentration.
  • 31. Collection and Preservation of Blood – Live Suspect Must be drawn under medically accepted conditions by a qualified person Must use a sterile needle or lancet Cleanse with a non-alcoholic disinfectant such as: Aqueous benzalkonium chloride (Zepiran) Aqueous mercurie chloride Providone-iodine (Betadine)
  • 32. Seal blood sample in an airtight container with an anticoagulant (EDTA or Potassium oxalate) and preservative (sodium fluoride) Place in refrigerated storage for delivery to toxicology ASAP If procedures are not strictly adhered to, then BAC decreases and never increases – so sloppy police and lab work benefits the suspect. Collection and Preservation of Blood – Live Suspect
  • 33. Requires added precautions Must account for bacterial action so collect from a number of different body sites. Each sample is kept in separate airtight containers with an anticoagulant, sodium fluoride, and refrigerated. Collect urine and vitreous humor fluids – both usually do not suffer from postmortem ethyl alcohol production to any significant extent. Collection and Preservation of Blood – Postmortem Samples
  • 34. Alcohol and the Law A BAC of .08% or .10% in most states in equivalent to intoxication for most drivers Federal standard for bus and truck operators is .04%
  • 35. Implied consent comes with obtaining a license to operate a motor vehicle Driver has a choice – submit to test or be subject to loss of license for a given period of time. Schmerber v. California Court ruled against defendant – the Fifth Amendment only prohibits self-incrimination. Being compelled to furnish “physical” evidence such as fingerprints, photographs, measurements, and blood samples are not protected by the Fifth Amendment Alcohol and the Law
  • 36. Role of the Toxicologist First attempt to narrow down type of toxic substance based on victim’s symptoms, postmortem pathological exam, exam of victim’s personal effects, nearby presence of empty drug containers or household chemicals Next, use general screening procedures Test for small quantities that may be remaining Must be aware that the body always processes substances – few substances enter and completely leave the body in the same chemical state – most are metabolized – so test for the metabolite Must also assess for substance’s toxicity
  • 37. Toxicological Techniques 90% of drugs analyzed is for alcohol and cocaine The toxicologist must devise an analytical scheme that will successfully detect, isolate, and specifically identify a toxin Large number of drugs are either acidic (H+) or alkaline (OH-) By controlling the pH of a water solution into which blood, urine, or tissues are dissolved, the toxicologist can control the type of drug that will be recovered.
  • 38. Acidic Drugs Acid drugs are extracted from an acidified water solution with organic solvents such as chloroform. Barbiturates Acetylsalicylic acid (aspirin)
  • 39. Alkaline (Basic) - Drugs Basic drugs are extracted from an alkaline water solution with organic solvents Phencyclidine Methadone Amphetamines Cocaine
  • 40. Sample Acidic Drugs Basic Drugs Extraction at appropriate pH SCREENING TEST Immunoassay Gas Chromatography Thin-Layer Chromatography CONFIRMATION TEST Gas Chromatography-Mass Spectrometry
  • 41. Heavy Metal Poisons Rarely encountered Arsenic, bismuth, antimony, mercury, and thallium Extraction into HCL solution – insert a copper strip (Reinsch Test) – positive presence of a heavy metal will develop a silvery or dark coating on the copper. Confirm with atomic absorption spectrophotometry, emission spectroscopy, or X-Ray differaction
  • 42. Carbon Monoxide – A Commonly Encountered Poison CO binds with hemoglobin carboxyhemoglobin Methods to determine concentration of CO in the blood: Spectrophotometric methods examine bloods visible spectrum to determine the relative amount of carboxyhemoglobin to oxyhemoglobin or total hemoglobin Treat a volume of blood with a reagent to liberate the CO then measure with gas chromatography. Measured as percent saturation – 50-60% saturation is considered fatal when combined with .2% BAC 35- 45% saturation maybe fatal
  • 43. Smokers usually have an 8-10% saturation In fire evaluation – determination of victim’s condition prior to fire. If victim was alive at time of fire they would have a high % CO saturation If victim was dead at time of fire they would have low % CO saturation since they could not breath in any of the fumes Carbon Monoxide – A Commonly Encountered Poison