FORENSIC TOXICOLOGY ANALYSIS OF CYANIDE

Arranged By :
1. Afri Elfida Hastanti (20190350005)
2. Anggi Nopita Sari (20190350028)
3. Sindi Novita Sari (20190350031)
4. Prili Pricilia (20190350032)
5. Ulfa Fauziah (20190350035)
6. Ajeng Dwi Maulisa (20190350040)
7. Dio Fahlevi Yokka Rajasa (20190350055)
8. Fadhilah Anwar Hidayat (20190350059)
9. Dwi Febriyani (20190350064)
10. Nahtia Rini Asrianti (20190350096)
11. Silvia Agatha Putriyana (20190350097)
12. Nuha Haifa Arifin (20190350102)
13. Rissa Latifa (20190350113)

DEPARTMENT OF PHARMACY
FACULTY OF MEDICINE AND HEALTH SCIENCE
MUHAMMADIYAH YOGYAKARTA UNIVERSITY
2021
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 Table of Contents
FOREWORD .................................................................................................................................. 4

CHAPTER 1 ................................................................................................................................... 5

1.1 BACKGROUND ...................................................................................................................... 5

1.2. PROBLEM FORMULATION ............................................................................................. 6

1.3. PURPOSE ............................................................................................................................ 6

CHAPTER 2 ................................................................................................................................... 8

2.1. Forensic ............................................................................................................................... 8

2.1.1. Definition of forensic toxicology .................................................................................. 8

2.1.2. Purpose of Toxicological Analysis ............................................................................... 9

2.1.3. The Task Of Forensic Toxicology .............................................................................. 10

2.1.4. Principles Of Forensic Analysis .................................................................................. 11

2.1.5. Stages Of Forensic Analysis ....................................................................................... 11

2.1.6. The Field of Forensic Toxicology........................................................................... 13

2.1.7. The Legal Basis of Forensic Analysis .................................................................... 14

2.1.8. Examples of Case Handled by Forensic Toxicology Analysis ............................... 15

2.1.9. Samples on Forensic Toxicology Analysis ............................................................. 15

2.1.10. Forensic Toxicological Analysis of the Scenario ................................................... 16

2.1.11. The Main Factor that Determines the Analysis Interpretation Results ..................... 18

2.2. Poison ................................................................................................................................. 19

2.2.1. Definition of Poisoning ............................................................................................... 19

2.2.2. Classification of Poison .............................................................................................. 19

2.2.3. The Pathways for The Entry of Poison Into The Body ............................................... 21

2.2.4. Factors Affecting the Action of the Poison ................................................................. 22

2.2.5. Toxic Materials in the Scenario .................................................................................. 27

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 2.2.6. Physical and Chemical Properties of Cyanide ............................................................ 27

2.2.7. Mechanism of Cyanide in the Body ............................................................................ 27

2.2.8. Characteristics of Cyanide Poisoning ......................................................................... 28

2.2.9. Early and Advanced Treatment in Cyanide Poisoning ............................................... 29

2.2.10. Analysis of Cyanide .................................................................................................. 30

CHAPTER 3 ................................................................................................................................. 32

3.1 Conclusion .......................................................................................................................... 32

3.2 Suggestion ........................................................................................................................... 32

References ..................................................................................................................................... 33

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 FOREWORD
We would like to thank Allah SWT who has given us blessings and mercies so we can
complete this paper entitled “FORENSIC TOXICOLOGY ANALYSIS OF CYANIDE”
at the right time.
The purpose of this paper is to find out and inform about descriptions, mechanisms, and
service work from forensic toxicology. This paper was compiled with help from various
parties. Both parties come from outside as well as from parties concerned themselves. Finally,
by the grace of Almighty, these papers can be resolved. In the preparation of this paper, we
compile multiple compilers, namely :
1. Mr. Hari Widada, M.Sc., Apt.

2. Our parents

3. Classmates

We would like to thank those who have helped us in every way so that the paper can be
resolved. May Allah reward all your kindness. Aamiin. Lastly, we realize that there are a lot
of mistakes and weakness in this paper, therefore we need constructive criticism and
suggestions from the readers.

Yogyakarta, 23rd April 2021

Authors

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 CHAPTER 1

INTRODUCTION

1.1 BACKGROUND

The term “toxin” means the toxic material of plants, animals, microorganisms, viruses,
fungi, or infectious substances, or a recombinant molecule, whatever its origin or method of
production,including any poisonous substance or biological product that may be engineered
as a result of biotechnology produced bya living organism or any poisonous isomer or
biological product, homolog, or derivative of such a substance.
Toxicology comprises a variety of disciplines, including analytical chemistry,
biochemistry, epidemiology, pathology, physiology and statistics. Forensic toxicology covers
those disciplines of forensic science concerned with toxic substances. The main fields of
forensic toxicology are chemical warfare agents, designer drugs, drug metabolism, drugs of
abuse, doping control, postmortem toxicology, toxicokinetics of poisons, and analytical
methods for toxic substances. A forensic toxicologist must interpret analytical data on toxic
chemicals and poisons, using this as the basis for accurately judging the toxicokinetic
implications of such substances.
In forensic toxicology, analytical results are always crucial to investigation of crimes
or accidents and, moreover, sometimes provide conclusive proof for criminal trials. Once
samples (e.g. drugs, poisons, victims’ body fluids, and so on) are given to forensic
toxicologists, it is expected that the analytical results from the samples will be promptly fed
back to the crime investigation section. For fast and precise analysis of toxic substances, the
development of new methods with high selectivity, sensitivity, rapidity, and robustness is
required. Studies on the absorption, distribution, metabolism, and excretion of these drugs are
also important, both as the scientific basis for detecting them in forensic samples and for better
understanding of their toxicokinetics.
Cyanide is a chemical compound which is toxic and is a type of poison the fastest
active in the body so can cause death in time a few minutes. Traditional cyanide known as
poison and has been used in mass murder, agent suicide especially among health workers and
laboratories, and as weapons of war. Hydrogen cyanide is also called formonitrile, being in
liquid form known as prussitic acid and tamarind hydrocyanic. Hydrogen cyanide is a liquid

5
colorless or it can be blue pale at room temperature. Is volatile and flammable. Hydrogen
cyanide can diffuses well with air and materials explosive.Hydrogen cyanide is very easy mix
with water so often and potassium cyanide in powder form and white.
Cyanide in low doses can found in nature and present in every product that we usually
eat or use. Cyanide can be produced by bacteria, fungi and algae. Cyanide is also found in
cigarettes, motor vehicle fumes, and food such as spinach, bamboo, beans, tapioca flour and
cassava. Apart from that it can also be found in some synthetic products. Lots of cyanide used
in industry, especially in manufacture of salts such as sodium, or potassium calcium cyanide.
Cyanide used by military NATO (North American Treaty Organization) is the liquid type,
namely hydrocyanic acid (HCN).

1.2. PROBLEM FORMULATION

Based on the above background, the problem in this paper is formulated as follows :
1. What are the definition of forensic toxicology ?
2. what are the duties and principles of forensic toxicology?
3. How is the step of the toxicological analysis ?
4. what are the toxic substance in this scenario?
5. How is the classification of toxins and routes of entry into the body?
6. what is the initial and subsequent treatment in cyanide poisoning?
7. what are the characteristics and mechanisms of cyanide in the body?
8. what are organic poison antidotes?

1.3. PURPOSE
Based on the formulation of the problem above, the objectives of this paper are formulated as
follows :
1. To find definition and of forensic toxicology
2. To know duties and principels of forensic toxicology
3. To know step of toxicological analysis
4. To know the toxic substance in this scenario
5. To know classification of toxins and rotes of entry in the body
6. To know initial and subsequent treatment in cyanide poisioning

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7. To know the characteristic and mechanisms of cyanide in the body
8. To know antidotes of organic poison

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 CHAPTER 2

THEORY AND DISCUSSION

SCENARIO
BG, a pharmacist who works in the forensic division of Polda Metro Jaya received a sample
to be analyzed. Samples were taken from the victim's body and evidence in the case of the
death of the visitor SB cafe. The victim died after drinking a coffee with symptoms of seizures
and the skin turned blue. Presupposition the murder case uses poisonous materials.
2.1. Forensic
Forensics (derived from Greek 'Forensis' which means debate or debate) is a field of
science that is used to assist the process of upholding justice through the process of applying
science (science). Forensic science (commonly abbreviated as forensic) is an application of
various sciences to answer questions that are important for a legal system that may be related
to criminal acts. However, in addition to its relation to the legal system, forensics generally
includes something or methods that are scientific (scientific in nature) and also rules which
are formed from the facts of various incidents, to introduce physical evidence (for example
corpses, carcasses), etc).
2.1.1. Definition of forensic toxicology
Forensic toxicology is one of the branches of forensic science. According to Saferstein
meant by Forensic Science is "the application of science to low ”, then in general Forensic
science can be understood as an application or utilization of science certain for law
enforcement and the judiciary. To better understand the meaning and scope of forensic
toxicology work, the better previously if you know more about the field of science toxicology.
The science of toxicology is a science that examines the work and harmful effects of chemicals
or poison to a biological mechanism organism. Toxins are potential compounds that harm an
organism. The toxic nature of a compound is determined by dose, the concentration of toxins
at the receptors, physical properties of toxic chemicals, bioorganism conditions or
bioorganism systems, exposure to organisms, and the shape of the effect. Toxicology forensics
devotes itself to the application or use of toxicology for the benefit of Justice. The main
occupation of forensic toxicology is doing a qualitative analysis as well quantitative of poisons
from physical evidence and translate the findings of his analysis into expression whether or
not the poison is present involved in a criminal act, which is alleged, as evidence in a criminal

8
act (forensic) at court. Results of analysis and interpretation of findings this analysis will be
loaded into a report following the laws and regulations. According to the Criminal Procedure
Code (KUHAP), This report can be referred to as a Certificate Expert or Certificate. So
forensic toxicology can be understood as the use of knowledge toxicology for law
enforcement purposes and Justice. Forensic toxicology is a science application which in
practice is strongly supported by various other basic science fields, such as chemistry analysis,
biochemistry, chemical instrumentation, pharmacology, toxicology, pharmacokinetics,
biotransformation. On the biomedical field, toxicologist will take care of that side effect
occurs in humans as a result of exposure drugs and other chemical substances, as well proof
of safety or danger forensic toxicology itself relating to the application of science toxicology
in various cases and problems where crime drugs and chemicals which can lead to medicolegal
consequences as well as for becoming evidence in court. The methods that can be used in this
forensic toxicology continue developing in different parts world. Discoveries regarding
clinical medicine and very laboratory test method assist in use specific methods, tools
required, as well as the interpretation of the results from testing these samples.
2.1.2. Purpose of Toxicological Analysis
According to the Society of Forensic Toxicologist, Inc. (SOFT), the field of forensic
toxicology includes:
a. Analysis and evaluation of causative toxins death,
b. Analysis of presence-absence alcohol content, illegal drugs in body fluids or breath
ones can lead to change behavior (decreased ability driving a motor vehicle roads, acts
of violence and crime as well as use dopping),
c. Drug analysis in the blood and urine in cases narcotics abuse, psychotropic and illegal
drugs other.
Another purpose of analysis forensic toxicology can make a reconstruction fiction an event
that has happened, until where the drug has been can result in a behavior change. Another
goal of toxicological analysis forensics is making a reconstructed conjecture of an event that
happened, to what extent the drug or poison can result in a change in behavior (decreased
ability driving, which may result in an accident fatality, or acts of violence and crime).

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2.1.3. The Task Of Forensic Toxicology
The main task of forensic toxicology is to carry out both qualitative and quantitative
analyzes of poisons from physical evidence and translate the findings of the analysis into the
expression of whether or not the poison was involved in the alleged criminal act as evidence
of a criminal (forensic) act in court. Results of analysis and interpretation of findings This
analysis will be published in a report that is in accordance with the laws and regulations.
According to the Criminal Procedure Code (KUHAP), this report can be referred to as a
Certificate of Experts or a Certificate.
In general, the task of a forensic toxicologist is to assist law enforcement, especially
in conducting a poison analysis both qualitatively and quantitatively and then translating the
results of the analysis into a report (letter, expert statement letter, or expert witness), as
evidence in a criminal act (forensic) in court. More specifically, forensic toxicology covers
the application of natural science in the analysis of poisons as evidence in a crime, with the
aim of detecting and identifying concentrations of toxic substances and their metabolites from
biological fluids and finally interpreting the analysis findings in an argument about the cause
of poisoning of a case.

analyzes and
evaluates
causative toxins
Dead

According to the
forensic toxicology
society America
"Society of forensic
toxicologists, inc.
SOFT "
analysis of drugs
analysis of the
in blood and urine
presence /absence
in cases narcotics,
of alcohol, drugs
psychotropic and
in body fluids or
drug abuse other
breath
forbidden

Picture 1.The Diagram of Task Of Forensic Toxicology

Source : Writer Analysis

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2.1.4. Principles Of Forensic Analysis

Individuality Exchange

Forensic
Analysis

Picture 2.The Diagram of Principles of Forensic Analysis

Source : Writer Analysis
• The Individuality Principle
The principle of individuality is attributed to Paul L Kirk (1963) and is regarded as the
building block for forensic science. Individuality implies that every entity, whether person or
object, can only be identical to itself and so is unique. No two objects whether natural or
artificial can be the same. Kirk claimed that forensic science aims to focus on the source of
two items (questioned and known, or mark and print), which are thought to have come from a
single source. As such, identification is concerned with establishing individuality from traces
left at a crime scene rather than the sameness of two things. This means identification can be
shown indirectly through the analysis of traces and samples e.g. no two fingerprints are the
same.
• The Exchange Principle
The exchange principle is attributed to Edmond Locard. The principle states that
whenever two objects or subjects interact, some sort of trace will be left behind. This is
generally at the crime scene. Trace materials include hairs, blood, fibres, and gunshot
residues. Locard suggested that there are many traces left behind and if interpreted properly
they provide the most valuable information.
2.1.5. Stages Of Forensic Analysis

Sample Confirmatory Interpretation

Screening Test
Preparation Test Of Analysis

Picture 3.The Diagram of Stages of Forensic Analysis

Source : Writer Analysis

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a. Sample Preparation
Specimens for toxicological analysis forensics is usually seen by a doctor, for example
on case of unnatural death specimens were collected by a forensic doctor at the time of
performing the autopsy. Specimens can be biological fluids, tissues, organs. In the doctor's
specimen collection forensics labeled each one wrap / container and seal it.
Several things that need to be taken into account in the sample preparation stage are :
1. Types and the biological properties of the specimen
2. Physicochemical of the specimen, as well as the purpose of the analysis.
Thus it will be designing or selecting a sample handling method, the number of samples to
be used, as well as choosing appropriate analysis method. Sample handling is necessary
gets special attention, for the most part the sample is biological material, so conclusively
possibly preventing the breakdown of the analyte from occurring.
b. Screening Test
Screening test to filter and recognize group of compounds (analytes) in the sample.
Right here analytes are classified according to their good physicochemical properties, its
chemical properties and pharmacological effects raised. The screening test should work
identifies the analyte class by degree reliability and high sensitivity, relatively cheap and
implementation is relatively fast. There is a test technique screening, namely: a) thin layer
chromatography (TLC) which is combined with the color reaction, b) technique
immunoassay. General immunoassay technique has high reliability and sensitivity
properties, and the process takes time relatively short, but the tools and materials of this
technique everything has to be imported, so this technique to be relatively inexpensive.
Compared with immunoassay, TLC is relatively cheaper, however the process requires a
relatively small amount of time longer.
c. Confirmatory Test
This test aims to confirm the identity of the analyte and determine levels. Most
confirmatory test slightly sensitive to the screening test, but should be more specific.
Generally a confirmation test using a combined chromatographic technique with other
detector techniques, such as: chromatography gas-mass spectrophotometry (GC-MS),
chromatography high-performance liquid (HPLC) with a diode-array detectors, liquid
chromatography - mass spectrophotometry (LC-MS), TLC-Spectrophotodensitometry, and

12
 techniques other. Besides doing a potential identification test positive analyte (screening
test result), in this test too the assay of the analyte is carried out. Data analysis quantitative
analytes will be of great use to forensic toxicologists in interpreting the results analysis,
with regard to answering the questions that arise both from investigators and judges in
connection with the case which is related.
d. Interpretation of Analysis
On the interpretation of the results of the analysis in case of death, a forensic
toxicologist sued capable answer specific questions such as: route the use of the toxicant,
what is the toxicant concentration which is set simply as a cause death or cause of
poisoning. Routing usage is usually obtained from various analyzes the specimen, where it
is generally the concentration higher toxicants were found in the route area usage. If a toxic
amount is found large in the digestive tract and liver, it can concluded that exposure by oral
route. Likewise, if the concentration is high found in the lungs or in the visceral organs
others indicate exposure by inhalation. New injection marks on body surfaces (such as
palms, arms, etc.), which are found in cases of death due to abuse narcotics, is an indication
of exposure through injection.
2.1.6. The Field of Forensic Toxicology
The field of forensic toxicology involves three main sub-disciplines: postmortem
forensic toxicology, human performance toxicology, and forensic drug testing. These
specialized fields offer a variety of exciting career paths. In postmortem forensic toxicology,
forensic toxicologists work with pathologists, medical examiners, and coroners to help
establish the role of alcohol, drugs, and poisons in the causation of a death. The forensic
toxicology laboratory identifies and quantifies the presence of drugs and chemicals in
biological fluids and tissues that are taken from the body during the autopsy. A wide array of
specimens may be encountered in postmortem toxicology investigations including blood,
urine, vitreous fluid from the eye, liver, brain, and other tissues, as well as hair and nails. Once
the testing is complete, a forensic toxicologist then interprets these findings. This information
helps a forensic pathologist determine the cause and manner of death.

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 2.1.7. The Legal Basis of Forensic Analysis
The legal and statutory bases in the health sector governing the identification process are:
a. In connection with the doctor's obligation to assist the judiciary, it is regulated in Article
133 of the Criminal Code
− In the case of an investigator to assist the interests of the judiciary in dealing with
a victim, whether injured, poisoned or dead, who is suspected of being an event
which constitutes a criminal act, he or she has the authority to submit a request for
expert information from an expert in judicial medicine or a doctor or other expert.
− Request for expert information as referred to in paragraph (1) shall be made in
writing, which in the letter clearly states for examination of wounds or examination
of a corpse and / or examination of a post-mortem.
− A corpse sent to an expert in judicial medicine or a doctor at the hospital must be
treated properly with full respect for the corpse and given a label containing the
identity of the corpse, carried out by affixing a position stamp placed on the big toe
or other part of the body of the corpse.
b. Health Law Article 79 of 1992
In addition to illnesses, the State Police of the Republic of Indonesia are also given
special powers to be assigned to certain civil servants at the Ministry of Health as
investigators as stipulated in Law no. 8 of 1981 concerning Criminal Procedure Law,
to carry out criminal investigations as regulated in this law.
Investigators are authorized to:
− Checking the accuracy of reports and information.
− Conducting an examination of the person who is suspected of doing it.
− Request information and evidence from a person or business entity.
− Perform examinations or letters or other documents.
− Checking or confiscating material or evidence
− Request expert assistance in carrying out investigative tasks
− To stop an investigation if there is insufficient evidence relating to a crime in the
health sector
c. The investigator's authority is exercised according to Law no. 8 of 1981 concerning
HAP

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 2.1.8. Examples of Case Handled by Forensic Toxicology Analysis
The following is a description of those cases generally in developed countries that require
inspection forensic toxicology, covering three major groups that are :
a. Death from poisoning, which includes: sudden death, death in prison, death by fire, and
medical death caused by side effects of drugs or medical mishandling,
b. Fatal or non-fatal accidents, which can threaten the safety of one's own life or others,
which are generally the result of the influence of drugs, alcohol, or whatever drugs,
c. Drug abuse and cases of poisoning associated with the use of medicine, food, cosmetics,
medical devices, and other chemical hazardous materials, which are not meet health
standards (cases pharmaceutical forensics).
2.1.9. Samples on Forensic Toxicology Analysis
Determine where the poison is it resides from history and clinical signs found on
external examination and in-depth examination. About the victim who died, were needed
residual and suitable poison information with abnormalities found in corpse. Determination
samples that need to be taken on adjusted toxicological examination with the type of poison
that entered into the body.
Better to take a sample fresh and complete at the time of the autopsy than then have to
do grave digs to retrieve the samples required and performed a toxicological analysis on body
tissue that is rotten or already preserved. The sampling principle in case of poisoning taken as
much as possible afterward we set aside for backup and for histopathological examination. In
general, the samples should be taken are:
a. The stomach and its contents;
b. The whole intestine and its contents with make a bulkhead with ties to the intestine at
any distance about 60 cm;
c. Blood
Blood is drawn from the heart is done separately from the right and the left 50 ml each.
Peripheral blood as much as 30-50 ml, taken from the communist iliac vein not blood
from the portal vein. On the surviving victim, blood is the most important ingredient,
2 blood samples were taken, each 5 ml, the first one was given 1% NaF preservative
and others without preservatives;
d. Liver, as a detoxification site, taken as much as 500 grams;

15
 e. Kidneys, both are taken in cases of heavy metal poisoning in particular or when the
urine is not available;
f. Brain, taken 500 grams. Special for chloroform poisoning and cyanide, made possible
by the brain consists of a lipoid tissue have the ability to retain poison even if it has
underwent sedation;
g. Urine, taken in whole. Because in general, the poison will excreted in the urine,
especially in the filter test for narcotics, alcohol, and poisoning stimulant;
h. Bile, taken for space excretion of various toxins;
i. In special cases :tissue around the injection, tissue muscle, fat under the skin walls
stomach, hair, nails, and brain fluid;
2.1.10. Forensic Toxicological Analysis of the Scenario
a. Materials and Instrumentation
• Materials :
Potassium cyanide (KCN), acetonitrile, sulfuric acid. All the solvents and reagents were
analytical grade. The internal standard (IS) solutotion was prepared by placing 1 mL of
acetonitrile (ACN) in a 100 mL volumetric flask and diluting it with water. This solution was
further diluted 1000- fold, to yield a working solution with a final concentration of 7.86 µg/mL
of ACN. Cyanide working solution were prepared by weighing 25.0 mg of KCN and diluting
with 0.1 N of NaOH in a 100 mL volumetric flask to obtain a solution with a concentrtion of
100 µg/mL of CN-. Then, two successive dilutions were performed, so the final concentration
of 10 and 1 µg/mL were achieved.
• Instrumentation
Cyanide analysis was performed on a Varian 450 Gas Chromatographer (GC) equipped with
a Nitrogen and Phosphorous Detector (NPD) (Varian, Palo Alto, CA, USA) with a VF 624
capillary column (30 m × 0.32 mm i.d., 1.8 µm Įům thickness) (Agilent Technologies, Palo
Alto, CA, USA). The headspace oven and syringe temperatures were set at 60°C. Samples
were heated for 10 min with continuous shaking at 500 rpm. The column oven temperature
was programed as follows: 30°C for 0.25 min, increased to 40°C (3°C/min) and then increased
to 150°C (40°C/min) and held for 1 min (total run time of 7.3 min). The injector and detector
temperature were 200 and 300°C, respectively. The injector operated in split mode (20:1), and
the carrier gas flow (nitrogen) was 2 mL/min, with pressure pulse (25 psi for 0.25 min).

16
 b. Methods
The analytical method used in the scenario is CG-MS. The CG-MS technique is that the
analyte is separated using gas chromatography and then its identity is ascertained using the
mass spectrophotometry technique. Previously, the analyte was isolated from the biological
matrix, then if necessary, it was derivatized. Isolates will be passed to the CG column based
on differences in the physicochemical properties of the toxicants and their metabolites. Then
in CG, there will be a toxic separation from the compounds in the group or their metabolites.
In the principle of CG separation, the retention index of the separate analyte is highly specific
for the compound. However, this is not sufficient for forensic toxicology analysis. Separate
analytes will enter mass spectrophotometry (MS), here depending on the fragmentation
method in MS, the analyte will be fragmented to produce a very characteristic mass spectrum
pattern for each compound. By combining the retention index and mass spectrum data, the
identity of the analyte can be identified and ascertained. From the results of CG-MS
separation, cyanide compounds were found in the sample. The cyanide is lethal because it
diffuses rapidly into the tissue and binds to the target organ within seconds. So that cells are
deprived of oxygen due to blocking mitochondrial electron transport and oxidative
phosphorylation. Therefore, the victim's body went into spasm and the skin turned blue as
mentioned in the scenario.
c. Sample Preparation
A pool of blood from seven different deceased victims was analyzed to confirm the absence
of cyanide and then used as blank matrix. Fire victims were the only exclusion criteria.
Calibrators and quality controls (QC’s) samples were prepared fortifying the pooled cyanide-
free blood samples with the proper amount of cyanide working solutions at concentration of
0.1 (low), 0.7 (medium) and 3.0 µg/mL (high).
Half milliliter of the calibrators, QC’s and case samples (blood, urine, stomach content and
hepatic exudate) were placed into a 20 mL headspace vial along with 0.5 mL of the IS working
solution. Finally, 50 µL of sulfuric acid was added, and the vials were immediately capped
and analyzed by the HSGC/NPD method.
d. Interpretation of Analysis
Cyanide is found in concentrations high enough both in the blood, can be used as sufficient
evidence for declared the poison as the cause of death. Cyanide at high concentrations (0.17-

17
2.22 mg / l, found in death from poisoning cyanide), stated to be the cause of poisoning.
Whereas at a very small concentration (0.004 mg / l in healthy people and 0.006 mg / l in
smokers). In small amounts cyanide is also absorbed and raised during smoking. Therefore,
detects cyanide in the blood at levels below toxic concentration, can still be tolerated as
without any toxic effects.
2.1.11. The Main Factor that Determines the Analysis Interpretation Results
The main factor that determines the results of the analysis (DFG, 1990, 1995), namely:
1. The technical level of analysis that produces data analysis. At this level, errors can be
caused by the factor of the analysis method. To get valid analysis data, it is necessary to
carry out validation of analysis procedures, as appropriate with conditions that are
regulated internationally (for example following the provisions of the validation procedure
analysis contained in the Pharmacopoeia International, USP, AOAC, etc.).
2. Biological level, the biological matrix variance of sample lets give away contribution of
errors to the analysis results. There are three steps you can take in evaluating the analytical
data from an angle view of the biological level, namely: control plausibility, longitudinal
evaluation, and transverse.
- Plausibility control includes:
▪ Extreme data control, this data is controlled based on medical data such as data
the analysis did not match the data that had been obtained from the human
population or highly far off statistically.
▪ Constellation control ie comparing from various analytical data, which were
obtained from different biological matrices but data series still has that parameter
interdependent. For example, comparing data for analysis of toxicants and their
metabolites in blood and in urine, data constellation generated is controlled by
nature pharmacokinetics of toxicants and their metabolites.
▪ Control trend data: analytical data obtained from one patient (victim) evaluated
against changes in time, it aims to find out the nature of biological changes (eg
elimination rate) occurring at these patients.
The goal of plausibility control is to search for analysis errors, wherefrom the level of
analytic techniques is not identified so that it is hoped that the analysis data will be obtained
which is valid.

18
 - Longitudinal analysis, this evaluation is based on pharmacokinetic (toxicokinetic)
properties and biotransformation reactions from toxicants and its metabolites.
Analytical data (toxic and its metabolites) from the same patient, that is obtained
from the time interval of collection of different samples (scrapings) compared to
one another. From the results of comparisons analysis data, based on characteristics
pharmacokinetics, it can be used as a basis to predict/control the concentration of
the actuals (time of poisoning). Further data can be used as a basis for estimating
the time the exposition took place.
- Transverse analysis, data analysis obtained from one patient compared with the
control group. Data from the control group maybe it can be concentration data
toxicants/drugs, which are taken from a time interval effect, such as the
concentration-time interval of the effect therapeutic, toxic, or “lethal concentration
intervals dose".
3. The level of nosology (science grouping disease), errors can result from an error in
diagnosing poisoning or may arise as a result of an error interpret pathological findings or
physiologic patient (victim). This mistake may appear because poisoning can show
pathological abnormalities.
2.2. Poison
2.2.1. Definition of Poisoning
Poisoning is an emergency that can damage cells and some body functions as a result
the entry of a substance or food that contains poison, comes from the material toxic
which is formed as a result of food spoilage and bacteria
2.2.2. Classification of Poison
a. Based on Sources
• Natural / artificial sources = Original toxins come from flora and fauna and
contamination of organisms with various toxins from industrial raw materials and
toxic synthetic materials
• The source is in the form of points, areas, and moves
• Domestic, commercial and industrial sources
b. Based on Form

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Classification based on the form of pollutants in the form of solid, liquid, and gas. As well as
pollutant size, density, and composition
c. Based on Physical and Chemical Properties
• Corrosive
• Radioactive
• Evaporative
• Explosive
• Reactive
d. Based Based on The Formation of Pollutants/Xenobiotics
• Pollutants are formed and come out of the source, called primary pollutants
• Pollutants acting in the environment are called secondary pollutants
• Secondary pollutants react to become tertiary pollutants.
e. Classification of Health Effects
• Fibrosis: excessive formation of connective tissue;
• Granuloma: acquired chronic inflammatory tissue;
• Fever: body temperature exceeds normal temperature;
• Asphyxia: a state of lack of oxygen;
• Allergy: excessive sensitivity;
• Cancer: malignant tumor; Mutant: a different generation from the parent gene
• Teratogenic: congenital defects
• Systemic poisoning: poisoning that affects the whole body
f. Basic Classification of Target Organ Damage
• Hepatoxic: toxic to the liver;
• Nephrotoxic: toxic to the kidneys;
• Neurotoxic: toxic to nerves;
• Hematotoxic: toxic to blood cells;
• Pneumotoxic: toxic to the lungs
g. Classification Based on Life/Death Poison

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2.2.3. The Pathways for The Entry of Poison Into The Body
There are four routes by which a substance can enter the body: inhalation, skin (or eye)
absorption, ingestion, and injection.

a. Inhalation

For most chemicals in the form of vapors, gases, mists, or particulates, inhalation is the major
route of entry. Once inhaled, chemicals are either exhaled or deposited in the respiratory tract.
If deposited, damage can occur through direct contact with tissue or the chemical may diffuse
into the blood through the lung-blood interface. Upon contact with tissue in the upper
respiratory tract or lungs, chemicals may cause health effects ranging from simple irritation to
severe tissue destruction. Substances absorbed into the blood are circulated and distributed to
organs that have an affinity for that particular chemical. Health effects can then occur in the
organs, which are sensitive to the toxicant.

b. Skin (or Eye) Absorption

Skin (dermal) contact can cause effects that are relatively innocuous such as redness or mild
dermatitis; more severe effects include destruction of skin tissue or other debilitating
conditions. Many chemicals can also cross the skin barrier and be absorbed into the blood
system. Once absorbed, they may produce systemic damage to internal organs. The eyes are
particularly sensitive to chemicals. Even a short exposure can cause severe effects to the eyes

21
or the substance can be absorbed through the eyes and be transported to other parts of the body
causing harmful effects.

c. Ingestion

Chemicals that inadvertently get into the mouth and are swallowed do not generally harm the
gastrointestinal tract itself unless they are irritating or corrosive. Chemicals that are insoluble
in the gastrointestinal tract fluids (stomach, small, and large intestines) are generally excreted.
Others that are soluble are absorbed through the lining of the gastrointestinal tract. They are
then trasported by the blood to internal organs where they can cause damage.

d. Injection

Substances may enter the body if the skin is penetrated or punctured by contaminated objects.
Effects can then occur as the substance is circulated in the blood and deposited in the target
organs.

Once the chemical is absorbed into the body, three other processes are possible: metabolism,
storage, and excretion. Many chemicals are metabolized or transformed via chemical reactions
in the body. In some cases, chemicals are distributed and stored in specific organs. Storage
may reduce metabolism and therefore, increase the persistence of the chemicals in the body.
The various excretory mechanisms (exhaled breath, perspiration, urine, feces, or
detoxification) rid the body of the chemical over a period of time. For some chemicals
elimination may be a matter of days or months; for others, the elimination rate is so low that
they may persist in the body for a lifetime and cause deleterious effects.
2.2.4. Factors Affecting the Action of the Poison
The toxicity of a substance usually depends on the following factors :
a. Factors Related to the Substance
− Form and Innate Chemical Activity
The form of a substance may have a profound impact on its toxicity especially for
metallic elements, also termed heavy metals. For example, the toxicity of mercury vapor
differs greatly from methyl mercury. Another example is chromium. Cr3+ is relatively
nontoxic whereas Cr6+ causes skin or nasal corrosion and lung cancer.

22
The innate chemical activity of substances also varies greatly. Some can quickly
damage cells causing immediate cell death. Others slowly interfere only with a cell's
function. For example:
• Hydrogen cyanide binds to the enzyme cytochrome oxidase resulting in
cellular hypoxia and rapid death.
• Nicotine binds to cholinergic receptors in the central nervous system (CNS)
altering nerve conduction and inducing the gradual onset of paralysis.
− Dosage
The dosage is the most important and critical factor in determining if a substance will be
an acute or a chronic toxicant. Virtually all chemicals can be acute toxicants if sufficiently
large doses are administered. Often the toxic mechanisms and target organs are different
for acute and chronic toxicity.
− Exposure Route
The way an individual comes in contact with a toxic substance, or exposure route, is
important in determining toxicity. Some chemicals may be highly toxic by one route but
not by others. Two major reasons are differences in absorption and distribution within the
body. For example:
• Ingested chemicals, when absorbed from the intestine, distribute first to the liver
and may be immediately detoxified.
• Inhaled toxicants immediately enter the general blood circulation and can
distribute throughout the body before being detoxified by the liver.
− Absorption
The ability to be absorbed is essential to systemic toxicity. Some chemicals are readily
absorbed and others are poorly absorbed. For example, nearly all alcohols are readily
absorbed when ingested, whereas there is virtually no absorption for most polymers. The
rates and extent of absorption may vary greatly depending on the form of a chemical and
the route of exposure to it. For example:
• Ethanol is readily absorbed from the gastrointestinal tract but poorly absorbed
through the skin.
• Organic mercury is readily absorbed from the gastrointestinal tract; inorganic lead
sulfate is not.

23
b. Factors Related to the Organism
Toxic responses can vary substantially depending on the species. Most differences between
species are attributable to differences in metabolism. Others may be due to anatomical or
physiological differences. For example, rats cannot vomit and expel toxicants before they are
absorbed or cause severe irritation, whereas humans and dogs are capable of vomiting.
Selective toxicity refers to species differences in toxicity between two species simultaneously
exposed. This is the basis for the effectiveness of pesticides and drugs. For example:
• An insecticide is lethal to insects but relatively nontoxic to animals.
• Antibiotics are selectively toxic to microorganisms while virtually nontoxic to
humans.
− Life Stage
An individual's age or life stage may be important in determining his or her response
to toxicants. Some chemicals are more toxic to infants or the elderly than to young adults.
For example:
• Parathion is more toxic to young animals.
• Nitrosamines are more carcinogenic to newborn or young animals.
− Gender
Gender can play a big role in influencing toxicity. Physiologic differences between
men and women, including differences in pharmacokinetics and pharmacodynamics, can
affect drug activity.
In comparison with men, pharmacokinetics in women generally can be impacted by
their lower body weight, slower gastrointestinal motility, reduced intestinal enzymatic
activity, and slower kidney function (glomerular filtration rate). Delayed gastric emptying in
women may result in a need for them to extend the interval between eating and taking
medications that require absorption on an empty stomach. Other physiologic differences
between men and women also exist. Slower renal clearance in women, for example, may result
in a need for dosage adjustment for drugs such as digoxin that are excreted via the kidneys.
In general, pharmacodynamic differences between women and men include greater sensitivity
to and enhanced effectiveness, in women, of some drugs, such as beta-blockers, opioids, and
some antipsychotics.

24
Studies in animals also have identified gender-related differences. For example:
• Male rats are 10 times more sensitive than females to liver damage from DDT.
• Female rats are twice as sensitive to parathion as are male rats.
− Metabolism
Metabolism, also known as biotransformation, is the conversion of a chemical from one
form to another by a biological organism. Metabolism is a major factor in
determining toxicity. The products of metabolism are known as metabolites. There are two
types of metabolism:
1. Detoxification
In detoxification, a xenobiotic is converted to a less toxic form. This is a natural defense
mechanism of the organism. Generally, detoxification converts lipid-soluble compounds to
polar compounds.
2. Bioactivation
In bioactivation, a xenobiotic may be converted to more reactive or toxic forms. Cytochrome
P-450 (CYP450) is an example of an enzyme pathway used to metabolize drugs. In the elderly,
CYP450 metabolism of drugs such as phenytoin and carbamazepine may be decreased.
Therefore, the effect of those drugs may be less pronounced. CYP450 metabolism also can be
inhibited by many drugs. The risk of toxicity may be increased if a CYP450 enzyme-inhibiting
drug is given with one that depends on that pathway for metabolism.
There is an awareness that the gut microbiota can impact the toxicity of drugs and other
chemicals. For example, gut microbes can metabolize some environmental chemicals and
bacteria-dependent metabolism of some chemicals can modulate their toxicity. Also,
environmental chemicals can alter the composition and/or the metabolic activity of the
gastrointestinal bacteria, thus contributing in a meaningful way to shape an individual's
microbiome. The study of the consequences of these changes is an emerging area of
toxicology.
− Distribution Within the Body
The distribution of toxicants and toxic metabolites throughout the body ultimately determines
the sites where toxicity occurs. A major determinant of whether a toxicant will
damage cells is its lipid solubility. If a toxicant is lipid-soluble, it readily penetrates cell
membranes. Many toxicants are stored in the body. Fat tissue, liver, kidney, and bone are the

25
most common storage sites. Blood serves as the main avenue for distribution. Lymph also
distributes some materials.
− Excretion
The site and rate of excretion is another major factor affecting the toxicity of a xenobiotic.
The kidney is the primary excretory organ, followed by the gastrointestinal tract, and the lungs
(for gases). Xenobiotics may also be excreted in sweat, tears, and milk. A large volume of
blood serum is filtered through the kidney. Lipid-soluble toxicants are reabsorbed and
concentrated in kidney cells. Impaired kidney function causes slower elimination of toxicants
and increases their toxic potential.
− Health Status
The health of an individual or organism can play a major role in determining the levels and
types of potential toxicity. For example, an individual may have a pre-existing kidney or
liver disease. Certain conditions, such as pregnancy, also are associated with physiological
changes in kidney function that could influence toxicity.
− Nutritional Status
Diet (nutritional status) can be a major factor in determining who does or does not
develop toxicity. For example:
• Consumption of fish that have absorbed mercury from contaminated water can result
in mercury toxicity; an antagonist for mercury toxicity is the nutrient selenium.
• Some vegetables can accumulate cadmium from contaminated soil; an antagonist for
cadmium toxicity is the nutrient zinc.
• Grapefruit contains a substance that inhibits the P450 drug detoxification pathway,
making some drugs more toxic.
− Circadian Rhythms
Circadian rhythms can play a role in toxicity. For example, rats administered an
immunosuppressive drug had severe toxicity in their intestines 7 hours after light onset
compared to controls and to other times in the day. The rats had changes in their
digestive enzyme activity and other physiological indicators at this dosing time.
c. Other Factors
− Presence of Other Chemicals
The presence of other chemicals, at the same time, earlier, or later may:

26
a. Decrease toxicity (antagonism)
b. Add to toxicity (additivity)
c. Increase toxicity (synergism or potentiation)
For example:
• Antidotes are used to counteract poisons' effects through antagonism (atropine
counteracts poisoning by organophosphate insecticides).
• Alcohol may enhance the effect of many antihistamines and sedatives.
• A synergistic interaction between the antioxidant butylated hydroxytoluene (BHT) and
a certain concentration of oxygen results in lung damage in the form of
interstitial pulmonary fibrosis.
2.2.5. Toxic Materials in the Scenario
Based on the symptoms mentioned after drinking coffee are seizures and the skin turns blue,
it can be suspected that the human died cause of cyanide poisoning.
2.2.6. Physical and Chemical Properties of Cyanide
• Hydrogen cyanide (AC) is a pale blue or colorless liquid below 78° F and a colorless
gas at higher temperatures. It has a bitter almond odor and is highly volatile and
flammable at room temperature.
• Sodium cyanide and potassium cyanide are white powders which may have a bitter
almond-like odor. In the presence of moisture, either can form hydrogen cyanide.
• Cyanogen compounds can generate cyanides. Cyanogen chloride (CK) is a colorless
liquefied gas that is heavier than air and has a pungent, highly irritating odor. It is
soluble in water and organic solvents.
2.2.7. Mechanism of Cyanide in the Body
Cyanide is very lethal because it can diffuse rapidly in tissues and bind to the target
organ in a few seconds. Cyanide can bind and inactivate several enzymes, especially those
containing iron in the form of Ferri (Fe3+) and cobalt. The resulting chemical combination
results in a loss of structural integrity and effectiveness of enzymes.
Cyanide can cause intracellular hypoxia through irreversible bonding with cytochrome
oxidase a3 inside the mitochondria. Cytochrome oxidase a3 plays an important role in
reducing oxygen to water through phosphorylation oxidation process. Cyanide bonding with
ferri ions in cytochrome oxidase a3 will result in resistance to terminal enzymes in the
27
 respiration chain, electron transport chain and forforylation oxidation process. Oxidative
phosphorylation is a process by which oxygen is used for the production of adenosine
triphosphat (ATP).
Disruption of this process will be fatal because it is important to synthesize ATP and
the ongoing cellular respiration. This low supply of ATP resulted in mitochondria being
unable to extract and use oxygen, so although oxygen levels in norml blood were not able to
be used to produce ATP. The result is a shift in metabolism in cells i.e. from aerobic to
anaerobic. Cessation of aerobic respiration also leads to the accumulation of oxygen in the
veins. In this condition, the problem is not in the delivery of oxygen but on the production and
utilization of oxygen at the cellular level. The result of this aerobic metabolism is a buildup
of lactic acid that will eventually lead to metabolic acidosis conditions.
Inhibition of cytochrome oxidase a3 is not the only mechanism that plays a role in
cyanide poisoning. There are several other mechanisms involved, including: inhibition of
carbonic anhydrous enzymes that play an important role in aggravating metabolic acidosis
conditions and bonding with methemoglobin which can be concentrated between 1%-2% of
hemoglobin levels. This cyanide bond causes this type of hemoglobin to be unable to transport
oxygen.
2.2.8. Characteristics of Cyanide Poisoning
Cyanide poisoning can occur when a person is exposed to cyanide, either through skin
contact, inhalation, or ingesting cyanide. When exposed to cyanide, the body's cells are
deprived of oxygen. As a result, the body's cells will experience damage and death. Symptoms
and complaints that occur when a person experiences cyanide poisoning can occur quickly.
Symptoms of cyanide poisoning will depend on the amount of cyanide inhaled or swallowed.
When exposed to large amounts, cyanide will cause damage to cells, tissues, and organs, in a
very short time. Some of the symptoms that can arise are:
- Seizures
- Difficulty breathing
- Loss of consciousness
- Low blood pressure (hypotension)
- Stop breathing
- Slow heart rate (bradycardia)

28
- Cardiac arrest and heart failure

Cyanide poisoning can also cause skin discoloration to redness. This is because oxygen
is trapped in the blood and cannot enter the body's cells. Meanwhile, when exposed to small
amounts of cyanide, complaints will generally arise, such as dizziness, nausea, vomiting, rapid
breathing, fast heart rate, weakness, fatigue, and headaches.
2.2.9. Early and Advanced Treatment in Cyanide Poisoning
The treatment of cyanide poisoning patients requires the enforcement of a quick and
precise diagnosis, in addition a prompt clinical decision is required to reduce the risk of
morbidity and mortility in patients. The level of risk of patients is strongly influenced by the
dose and duration of cyanide exposure in patients. In principle the management of cyanide
poisoning therapy can follow these steps:
a. Decontamination
Decontamination is adjusted to the exposure path, in general it can be categorized as
follows:
1. Inhalation
Move the patient to a smoke-free location and remove the patient's clothing.
2. Eyes and skin
Remove contaminated clothing, wash exposed skin with soap and or water, irrigate
eyes exposed to water or copy, remove contact lenses.
3. Gastrointestinal tract
Do not induce emesis, activated charcoal can be given if the patient is conscious
and still within 1 hour from exposure to cyanide. Isolate emesis can be administered
to aid the production of hydrogen cyanide.
b. Basic Life Support (BLS) and Advanced Cardiac Life Support (ACLS).
The main purpose of BLS is to protect the brain from irreversible damage due to
hypoxia, since the blood circulation will stop for 3-4 minutes. In cases of cyanide
poisoning where there is a decrease in utilization, 100% oxygen administration in
patients with nonrebreather masks or endotracheal tubes can help. This could help the
effectiveness of the use of antidotes by the mechanism of competition with cyanide to
the side of the cytochrome oxidase bond. According to the American Hearth
Association Guidelines in 2005, bls action can be abbreviated to ABC technique,
29
 namely airway (freeing airway), breathing (giving artificial breath), and circulation
(heart massage in shock condition). However, in 2010 the BLS action was changed to
CAB (circulation, breathing, airway).
c. Antidot Therapy
One of the keys to successful cyanide poisoning therapy is the use of antidotes as soon
as possible with empirical experience without having to know the patient's detailed
health condition in advance. In America there are two antidotes that have been
approved by the FDA, namely cyanide antidota kits that have been used for decades
and hydroxokobalamin approved in 2006. Cyanide antidote kit is a combination of 3
types of antidotes that work synergistically (amyl nitrite, sodium nitrite, and sodium
thiosulfate).
d. Supporting Therapy
Supporting therapy that can be done in patients is by:
1. Monitor the heart, respiratory and cardiovascular function of patients in the ICU
room.
2. Conduct laboratory tests to monitor gas levels in arterial blood, lactate levels in
serum, complete blood tests, blood sugar levels, cyanide levels in the blood and
electrolyte levels.
3. Monitoring and arrhythmia therapy.
4. Monitoring and therapy of side effects of antidot use.

2.2.10. Analysis of Cyanide

a. A strong acid (H2SO4 or HCl-HH) reflux distillation under vacuum is combined with
an air purge to liberate hydrogen cyanide (HCN) from both simple and complex
cyanides. The resulting HCN gas is collected and trapped in a weak NaOH scrubbing
solution. Thiocyanate breakdown must be < 1% when distilling samples containing
thiocyanate.
b. A weak acid (pH 4.5 to 6.0) reflux distillation under vacuum is combined with an air
purge to liberate hydrogen cyanide (HCN) from simple and easily dissociable
cyanide complexes. The acetate buffer uses zinc salts to precipitate iron cyanide as a
further assurance of the selectivity of the method. The resulting HCN gas is collected
and trapped in a weak NaOH scrubbing solution

30
c. Manual Gas Diffusion: Sample preparation is carried out using a microdiffusion cell.
The water, wastewater or extract sample is introduced in the outer chamber of the
microdiffusion cell and is buffered at pH 6 and placed in the dark for 6 hours of
diffusion. Free cyanide diffuses as HCN gas and is absorbed as CN- into the sodium
hydroxide solution located in the center chamber of the microdiffusion cell.
d. Colourimetry. Hydrogen cyanide is determined photometrically, based on the
reaction of cyanide with chloramine‐T to form cyanogen chloride. Cyanogen chloride
subsequently reacts with isonicotinic acid and 1,3-dimethylbarbituric acid to yield a
red coloured complex. The intensity of this colour is measured at 590-610 nm.
Pyridine and barbituric acid may also be used but with a preferred wavelength of
575-582 nm. Note that pyridine has significant human toxicity; Refer to SDS before
use.
e. Amperometric Detection. Hydrogen cyanide (HCN) gas diffuses through a
hydrophobic gas diffusion membrane into an alkaline acceptor stream where the
CNanion is captured and sent to an amperometric flow cell detector with a silver
working electrode. In the presence of cyanide, the silver electrode surface is oxidized
at the applied potential (Eapp = 0,0 V vs. the reference electrode). The anodic current
measured is proportional to the concentration of cyanide in the standard or sample
injected.
f. Ion Selective Electrode (ISE): CNin the alkaline distillate from the preliminary
treatment procedures can be determined potentiometrically using a CNion selective
electrode in combination with a double junction reference electrode and a pH meter
with an expanded millivolt scale, or a specific ion meter. This method can be used to
determine CN- concentration in the concentration range of approximately 0.05 to 10
mg/L CN- . Refer to APHA 4500 CN F for more details. This method has limited
sensitivity and is unsuitable for evaluation of aquatic life standards.

31
 CHAPTER 3
CLOSING
3.1 Conclusion
Forensic toxicology is the study of the application of toxicology, which is useful for
assisting judicial processes. Forensic toxicology is not only to identify/determine the
amount/quantity of drugs, poisons or substances in the human body but also can determine
the effects.
Judging from the symptoms in the scenario, cyanide poisoning is suspected due to other
symptoms such as seizures and a blue body caused by low blood pressure and hypoxia. In the
mirna case example on January 6, 2016, the autopsy found gastric bleeding due to a corrosive
substance that damaged the stomach which came from cyanide acid. Cyanide acid was found
in coffee samples.

3.2 Suggestion
We, as writers, realize that this paper has many mistakes and is very far from perfection.
Of course, the author will continue to improve the paper by referring to an accountable source
later. Therefore, the authors are looking forward to criticism and suggestions regarding the
discussion of this paper
Thus the material this time, I hope this article can be useful and can add to our
knowledge about everything.

32
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