Chapter 1

i) Discuss the factors determine the toxicity

 Toxicity is influenced by various factors , each playing a critical role in
determining the extent of a substance's harmful effects on an
organism.
o Genetic status, such as acetylator phenotype and P450
polymorphisms, can significantly impact how a substance is
metabolized, with certain genotypes leading to slower or faster
processing of toxins.
o Body mass is crucial in dosage calculations, as larger
individuals may require higher doses for the same effect, while
smaller ones are more susceptible to overdose.
o Sex differences also contribute, with variations in drug-
metabolizing enzyme (DME) expression and hormone levels
affecting toxicity.
o Age and maturity determine metabolic rates and organ function,
influencing how toxins are processed and excreted.
o Health and well-being, including psychological and
immunological status, can alter susceptibility, as a compromised
immune system or mental stress can exacerbate toxic effects.
o Diet plays a role by affecting the absorption and metabolism of
substances.
 Whether a substance is toxic depends on multiple factors: the
substance itself, type of organism exposed to the toxin,
amount of substance [dose], route of exposure, and when
more than one chemical is present.
o The inherent toxicity of a substance is determined by its
chemical structure and reactivity.
o Different organisms may exhibit varying sensitivities to the
same toxin due to physiological and metabolic differences.
o The dose is a critical determinant, with higher amounts generally
leading to greater toxicity.
o The route of exposure (ingestion, inhalation, dermal contact)
influences the rate and extent of absorption.
 o When multiple chemicals are present, their interactions can be
simply additive, synergistic (enhanced effect), potentiative (one
substance increases the effect of another), or antagonistic (one
substance reduces the effect of another).

ii) Explain the dose response curve

• The mathematical relationship between dose and measured

response.
• Plot: %age response vs log dose
• Dose is related to body weight e.g. mg/kg, mg/g
• Relationship is SIGMOIDAL.

• Lethal Dose for 50% Mortality (LD50)

– Statistically derived single dose  death in 50% of a given population
of organisms.
– Under a defined set of experimental conditions. [form, entry route]
– Not an exact value.
– May vary from species to species.
– Only use in a comparative sense i.e. compare toxicity of one
substance vs another
– Gives no information on sub-lethal toxicity.
 To gain a true, meaningful picture of the toxicology of a compound
need to consider slopes of the curves.
 Where flatter slope is more likely to  more deaths at doses smaller
than the LD50.

 Therapeutic Index of drugs: LD50/ED50

TD50/ED50
 Require this value to be as high as possible.
Alternatives to the LD50 test
 New OECD Guidelines:
 OECD: Organisation for Economic Co-operation and Development:
Environmental Health & Safety Programme  responsible for
reviewing guidelines for chemical toxicity test considering new
scientific data.
 420 which does not require death as endpoint
 423 which requires deaths < 3
 425 which requires deaths < 3

 Key to EU classification:
T* = very toxic
T = toxic
H = harmful
U = unclassified
 OECD Test 420
 Normally start with a sighting study
o Uses a single animal per dose
o Starting dose based on information from literature
o Use same doses as Test 420.
 Key outcomes
 A =  2 deaths
 B =  1 with toxicity or no death
 C- = no toxicity
  OECD Test 423
 Same doses as OECD 420 test
o Uses 3 animals/step [not 5]
 Key outcomes
 No further testing required
 Same dose 3 more animals
 Next higher or lower dose to 3 additional animals

 OECD Test 425

 Uses a maximum of 5 animals
 A sequential dosing method
 First animal given dose a step below LD 50
 Monitor for 48 hrs before next step

 Key outcomes
 Survives –> next animal increase dose by factor of 3.2
 Dies –> next animal reduce dose by similar factor
 Progression is by a series of up and down step

iii) Interaction of Xenobiotics

 The interactions between multiple chemicals can significantly
influence the overall toxic effects on an organism.
 o In additive interactions, the combined effect of two chemicals
equals the sum of their individual effects, such as when two
organophosphate insecticides are used together (2 + 3 = 5).
o Synergistic interactions occur when the combined effect
exceeds the sum of their individual effects, exemplified by the
heightened liver toxicity observed when ethanol and carbon
tetrachloride (CCl4) are combined (2 + 3 = 10).
o Potentiation is observed when a non-toxic compound enhances
the toxicity of another compound, such as phenobarbital
increasing the toxicity of paracetamol (0 + 3 = 10).
o Antagonism occurs when one compound reduces the toxicity of
another, as seen with pyrazole decreasing methanol toxicity (4 +
6 = 5).
Chapter 2
1. Alternatives to animal tests
 Toxicity testing employs a range of methodologies to ensure
comprehensive assessment and safety.
 In vitro techniques, including the use of isolated enzymes,
organelles, micro-organisms, tissue culture cells, isolated organs,
and advanced artificial systems like organs-on-chips and 3D human
cell-derived models, allow for detailed investigation of biochemical
pathways and organ-specific responses.
 Lower forms of animal and plant life provide initial toxicity data and
insights into environmental impacts.
 Computational methods, such as quantitative structure-activity
relationship (QSAR) models and other in silico approaches, facilitate
rapid and cost-effective toxicity prediction based on chemical
structures.
 Additionally, human-patient simulators offer advanced modeling of
human responses to toxins, integrating pharmacokinetics and
pharmacodynamics in a controlled and ethical manner.

2. Bacterial assay for mutation

 Using bacterial systems in toxicity testing offers several advantages.
 o Large populations of bacteria can be easily cultured and
treated, facilitating high-throughput screening.
o The extensive knowledge of bacterial genetics allows for
precise manipulation and understanding of genetic responses
to toxins.
o Bacterial assays are not only quick and inexpensive but also
well-validated, ensuring reliable and reproducible results.
o Additionally, specialized bacterial strains have been developed
with enhanced sensitivity to detect specific toxic effects,
further increasing the efficacy and accuracy of these assays in
toxicity testing.

AMES test [reverse mutation test]

 The Ames test is a widely used assay to assess the mutagenic

potential of chemical compounds.
 It utilizes auxotrophic strains of the bacterium Salmonella
typhimurium that have lost the ability to synthesize histidine due
to specific mutations.
 These bacteria are exposed to the chemical being tested, and the
ability of the chemical to induce mutations is assessed by measuring
the number of colonies that regain the ability to grow in a histidine-
free medium.
 Several features enhance the sensitivity of the test:
o Mutations in the bacterial cell wall allow chemicals to easily
enter the cells.
 o uvrB mutation eliminates the bacteria's ability to repair DNA
damage.
o The presence of the plasmid pKM101 stimulates the
conversion of DNA damage into inheritable mutations.
o The test is an indirect method of assessing mutagenicity by
observing changes in the bacterial phenotype, specifically the
reversion to histidine synthesis, which indicates that mutations
have occurred.

Chapter 3
1. Lethal synthesis
 Inappropriate metabolism of relatively safe xenobiotics into more toxic
molecules is a significant concern in toxicology.
 This often arises due to the broad specificity of xenobiotic-metabolizing
enzymes, which can convert benign substances into harmful ones.
 These enzymes exhibit tissue specificity, leading to localized toxin
production within specific organs.
 Although all tissues contain xenobiotic-metabolizing enzymes, the liver
is the most crucial organ for this process due to its central role in
metabolism.
 The liver is often the primary site of hepatotoxicity, as it is exposed to
the highest concentrations of both the xenobiotics and their toxic
metabolites.
 o Understanding and monitoring hepatic enzyme activity essential
in assessing the safety and potential toxicity of various
substances.

2. Briefly elaborate paracetamol toxicology and its treatment

 Paracetamol, a widely available non-narcotic analgesic, acts primarily
on peripheral tissues and is safe at therapeutic doses.
 However, overdose can lead to hepatotoxicity and liver necrosis.
 The major route of paracetamol metabolism is by conjugation,
removing about 60-90% of the drug, with only 5% remaining free after
each pass through the liver.
 Toxicity arises from an alternative metabolic pathway facilitated by
cytochrome P450 enzymes, which produce the toxic metabolite NAPQI.
 In overdose situations, glutathione, which normally detoxifies NAPQI,
becomes depleted, allowing NAPQI to bind to liver proteins and cause
necrosis.
 Treatment for paracetamol toxicity includes:
o Inhibit P-450 using Cimetidine.
o Increase glutathione synthesis.
o Provide sulphate for conjugation using methionine.
o Provide glutathione alternative with n-acetyl cysteine.

3. Screening and assessment

 Screening for alcohol and drug abuse should be a routine part of
physical examinations for individuals over 60, utilizing the "Brown
Bag Approach" where patients bring all their medications for review.
 Re-screening should occur if physical symptoms indicative of
substance abuse arises or during major life transitions.
 Direct questions should be asked, linking inquiries to medical
conditions of health concerns and avoiding stigmatizing language
[drug addict].
 Special assessments must include
 o Evaluations of functional abilities using tools like Activities of
Daily Living [ADLs], Instrumental Activities of Daily Living [IADLs],
and the SF-36.
o Assessments of comorbid physical and psychiatric conditions,
such as affective disorders and suicide risk.
o Special attention should be given to sleep disorders and cognitive
impairments.
 For dementia, tests like the
Orientation/Memory/Concentration Test and the Folstein
Mini-Mental Status Exam (MMSE) are used.
 For delirium is assessed with the Confusion Assessment
Method (CAM).
 Other cognitive impairments, including those resulting
from falls, accidents, or Wernicke-Korsakoff syndrome,
should also be considered to provide a comprehensive
evaluation of the older adult’s health status.

4. Morphine [narcotic analgesics]  acts chiefly in the CNS.

 Main effects of morphine on the CNS:
o Depression, leading to analgesia, respiratory depression (decrease
in sensitivity of the respiratory center to PCO 2), depression of cough
reflex, sleep).
o Excitation, leading to vomiting, miosis (pupil constriction),
convulsions (very rare).
o Changes of mood – euphoria (sense of well-being) or dysphoria.
o Tolerance and dependence (psychological and physical)
o Smooth muscle stimulation: Gastrointestinal muscle spasm (with
constipation) and biliary tract spasm.
o Cardiovascular system: Dilation of resistance and capacitance
vessels.
o Other effects: Sweating, histamine release, pruritus, piloerection,
antidiuretic effect.

Entrepreneurship questions
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Toxicology questions
- 5 factors that influence toxicity
- Factors that reduce free toxicants and about cytochrome p450
- Morphine effects
- AMES test