phar 303 midterm

Definition of toxicology

Definition of toxicology

the study of the adverse effects of xenobiotics on biological and ecological systems

Definition of xenobiotics

any substance that is foreign to a biological system

definition of toxin

substance produced by an organism that can cause harmful effects, like snake venom

definition of toxicant

man-made substance that can cause harmful effects, like DDT

toxic substance can be classified based on why they exist

chemical warfare agents, pesticides, household poisons, drugs, etc.

Toxicokinetics

• how the toxicant is absorbed, metabolized, and distributed

• ADME

• strictly speaking, looks at the movement of molecules inside the body and how we analyse that with different models

Toxicodynamics

How the toxicant exerts its effects

How does the BBB keep us safe

• most capillaries allow lipid- and water-soluble molecules through, but the BBB has tight junctions so only lipid-soluble molceules can get through but everything else has to be actively transported

• the brain is therefore only exposed to lipid-soluble compounds

Oocyte barrier to toxins

dynamic barrier that changes due to a variety of factors

Placenta as a barrier to toxins

the placenta is NOT a barrier! anything in the blood can pass to the fetus

Metabolism of toxicants

• mainly in the liver

1. hydrolysis

2. reduction

3. oxidation

4. conjugation

Hydrolysis

Breaking down complex molecules by the chemical addition of water. Can sometimes make even more toxic metabolite

oxidation

• performed by P450s

The skin as a barrier

• toxicants can cross the skin by diffusion

• the more lipophilic, the more rapidly it can cross

Special barriers to toxins

• the BBB

• the eyes

• the blood-testes barrier

• the blood-granulosa cell barrier

• the placental barrier (not true)

primary site of metabolism

the liver

Overview of metabolism of toxic compounds

• lipophilic -> hydrophilic

• biologically active -> non-biologically active

epoxide hydrolase

• makes a more reactive metabolite

• example of hydrolysis

ADH and ALDH

• alcohol is metabolized into acetaldehyde by ADH

• acetaldehyde is metabolized into acetic acid by ALDH (rate limiting_

• acetic acid is excreted quickly

• acetaldehyde accumulation bad

what performs oxidation

CYTP450s

CYTP450s

18 families of P450 genes, 43 subfamilies with multiple enzymes

conjugation

• adds a group to the compound to make it more hydrophilic

• 6 majors conjugation reactions

• ex. glutathione reaction: will react with an electrophile to make it less toxic as it will no longer be able to react with anything

volume of distribution

Vd = dose/concentration

high serum C means low Vd; it's not in the body's compartments

amount of toxicant variable

So

Vd variability

• if a toxicant is very polar, it will stick only to the plasma, so its Vd will be based on the 4.5L of standard plasma volume

• if it accumulates fat, the Vd can grow

warfarin Vd variability example

• very polar therefore low Vd

• does not leave the plasma easily

First-order elimination

the rate of elimination is a constant fraction of the dose

Zero order elimination kinetics

the rate of elimination is a constant amount

importance of knowing order of elimination for toxicokinetics

you can know how long the toxicity will last ex. DDT has a half life of 5 years, so it can stick around for the lifespan of an individual

factors affecting elimination rate

• lipid solubility

• metabolism

• high volume of distribution!!

how to identify a target molecule responsible for toxicity:

toxicant must:

• reach an effective concentration at the target site

• react with target and adversely affect function

• alter target in a way that is mechanistically related to toxicity

reaction types

• non-covalent binding

• covalent binding

non-covalent binding

reversible and common

• ex. membrane receptors, ion channels, enzymes

covalent binding

usually irreversible

• permanently alters endogenous molecules

• the toxicant is usually electrophilic and targets nucleotphiles

• action by hydrogen abstraction, electron transfer, enzymatic reactions

• modifying endogenous molecule can lead to autoimmunity

graded dose response curve

the dose response curve of an individual

quantal dose response curve

the dose response curve of an entire population (average)

dose response curve

• sigmoidal

• quantal or quantitative

how to make probit units

• perform log10 transformation on the DRC to produce a straight line

• add 5 to 0 to avoid negative numbers (probit units = 5+nSD)

how to use probit units

what's the point of probit units

compare different DRC

probit unit range

1-10, with 5 being 50% response

comparing dose response curves

threshold in toxicology

the concentration above which exposure leads to toxic effects governments and organizations need to decide safe thresholds

zero threshold

applied to anti-cancer drugs assuming that there is a linear relationship between the 0 point and the point where we see an effect, no dose is safe

NOAEL

no observed adverse effect level

the highest concentration at which we do not see a statistically significant effect

LOAEL

lowest observed adverse effect (NOAEL)

POD

point of departure

where we start seeing an effect, but it is not significant

can be the NOAEL, LOAEL, or BMD10 depending on the data you have to figure out RfD

MOST IMPORTANT for RfD calculation and based on data

using BMD is preferred

RfD

reference dose in mg/kg/day

= NOAEL/(UF * MF)

an estimate of the maximum acceptable oral dose of a toxic substance

can use NOAEL, BMD, or calculated point of departure

UF

uncertainty factor

• accounts for things we can't be sure about by being extra precautious

• can be reduced through further investigation

• ex. intra or interspecies variation

MF

modifying factor

• accounts for variations or inconsistencies in data

• ex. variation in concentration in

ED50

the point at which 50 percent of the test organisms show a negative effect from a toxin.

LD50

the amount of a chemical that kills 50% of the animals in a test population

hormesis

• beneficial effects produced by low doses of a toxic substance, effects change as dose changes.

• leads to funky DRCs

example of hormesis

vitamins

• too little = death

• too much = death

• U-shaped curve

non-monotonic dose response curves

• example of hormesis

• sometimes chemicals are toxic at low doses but not high

• sometimes chemicals have varying toxicity at different levels (like vitamins)

• difficult to determine safe threshold

• ex. endocrine disrupting chemicals

low dose effects

decreasing doses of toxins trigger different responses

toxicant interactions

• additive

• syngergistic

• potentiation

• antagonism

additive interaction

effects of more than one substance add up (2 + 2 = 4)

synergism

effects of more than one substance work together (2 + 2 = 10)

potentiation

substance that has no toxic effect enhances the toxicity of another substance that is toxic (0 + 2 = 10)

antagonism

occurs when the effects are less than additive; the toxins make each other less toxic (2 + 2 = 3) ex. Calcium and Selenium

ways toxins can alter biological systems

• can alter pH

• uncouplers of oxphos

• solvents and detergents can destabilize membranes

• space occupiers (oxalic acid in gout)

• basic receptor stuff

hazard vs. risk

Hazard = an event that could cause serious harm Risk = the probability that the hazard could happen Hazard x exposure = risk

hazard x exposure =

risk

risk = f(??)

risk = f(hazard, exposure)

exposure =

exposure = intensity x frequency x duration

parameters of total exposure

intensity, duration, frequency

exposure pathway

route of a toxin from its source to its end point

5 components of exposure pathway

1. source ex. chemical spill in water

2. transport mechanism ex. movement of contaminated water into well

3. point of exposure ex. well

4. route of exposure ex. eating, drinking, breathing, touching

5. a receptor population ex. the owners of the well

ingestion and inhalation are examples of which part of the exposure pathway?

exposure route

types of exposure data

• quantitative personal dosimeter measurements

• quantitative ambient measurements in vicinity

• quantitative surrogate of exposure

• residence or employment in proximity of source

• residence or employment in general area

best types of exposure data

directly measures data very close to the affected population

quantitative surrogate of exposure

replacement way of measuring levels ex. estimating quantities of drinking water consumed if measuring exposures to lead in water

which exposure data would be the best approximation of an actual exposure level?

water in your private well showing a reading of 1.3ppm of Copper

issues in measuring exposure

• factors influencing biodistribution

• time activity patterns make sampling hard

• homogenous vs heterogenous samples, like dirt

• mixtures (there can be synergism between toxins)

how long to sample?

depends on minimum time needed to obtain sufficient amounts for lab analysis

must consider:

• sensitivity of procedure

• contaminant concentration

• periodicity

5 major sampling media

• air

• water

• dirt

• food

• tissues

particulates and the lungs

0.5-5.0 microns in size can get trapped in the lungs

the smaller, the worse

how to measure air contaminants

through a filter (not good for volatiles) or through an absorbent (to capture gases)

fine particulate matter

2.5µm in size or less is significant indicator of air quality

air quality and industry

strong correlation between air pollution and industry locations

principles of sampling soil

• a soil test is only as good as its sample

• it may not be homogenous like air

• need a lot of different samples according to soil type, slope, crop, history, fertilizer, etc.

soil sampling grid system

need to collect 20 to 25 samples following a grid system to ensure good distribution

distribution of water samples

more homogenous than soil, but downstream effects are not homogenous

water sampling techniques

• surface-water grab technique

• continuous monitoring (requires device implanted in water)

• entrapment procedures

per capita consumption rate analysis

• measure how much food people are eating

• measure per capita consumption rate (production + import)/population

Food and Nutrition Surveillance Group

• produces national food and nutrition-related surveys to facilitate the dissemination of results

• build idetary assessment tools

• helps toxicologists get a sense of what people are eating

tolerable daily intake (TDI)

estimated quantity of a contaminant that we can be exposed to over a lifetime without posing a significant health risk

• if something is eaten frequently, a lower TDI for toxins in that food ex. rice or bread

Orchratoxin example

• produced by mold and grows on many crops

• carcinogen

• can't have 0 levels of Ochratoxin because it'll show up eventually, but we have lower tolerable limits for it in infant food than in adult food for exmaple

Medical vs. forensic tissue testing

medical:

• does not require patient consent

• identity of specimen is presumed

• screening result is sufficient for medical decisions

• results are used for medical evaluation

forensics:

• subject must consent to be tested

• identity of the specimen must be proved

• only confirmed results can be considered positive

• results are used for legal action

sampling tissue examples

done in medical or forensic contexts, experimental studies, environmental studies, workplace sampling, doping testing/urine sampling

toxins to look for in dopings

• anabolinc-androgenic steroids (AAS)

• hGH

• erythropoietin

• blood transfusions

urine testing limitations

• timing is important because of variation in how long drugs stick around

• ex. marijuana can be detected for up to 30 days but opiates only last 2 days

steps for measuring toxins in samples

1. extraction

2. separation

3. identification

extraction from samples

• make the sample homogenous

• concentrate the sample

• can involve physical crushing, blending, and chemical techniques

separation of samples

involves separating out your chemical of interest

• concentrate and purify the sample

• relies on the principle that compounds interact differently with different stationary and mobile phases

• chromatography babyyy

HPLC (high performance liquid chromatography)

popular method for chromatography

• stationary phases: silica + mobile

• modify the stationary phase depending on what you want to separate

reverse phase chromatography

uses a nonpolar card with a polar solvent to separate out something hydrophobic

anion exchange

a form of ion-exchange chromatography in which negatively charged ions (anions) are removed by a positively charged resin

• add ammonium group

cation exchange

a form of ion-exchange chromatography in which positively charged ions (cations) are removed by a negatively charged resin

• add carboxyl group