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Toxicological Risk Assessment

26 important questions on Toxicological Risk Assessment

What is the mayor difference in toxicological and microbiological risk assessment?

In toxicological risk assessment the focus lays on the hazard characterization and not so much on the exposure assessment. This is because the toxins will not grow in your product. However there are many different toxins and characterizing which will be a danger for you food product is harder.

On what should your hazard identification be based?

On the critical effect and the target organ. If you protect the target organ (organ that shows adverse health effect at the lowest dose) you protect all the organs.

What is an invivo and a invitro test?

In vivo is on animals
In vitro is not on animals

What are the different kind of animal test that can be performed?

Acut test (single dose toxicity)
Short term (sub-acute) test (28 days)
Semi-chronic test (90 days)
Chronic test (6-12 months) (also carciogenicity).

What are the minimal requirements for testing a food ingredient?

In vitro genotoxicity
In vivo semi-chronic (90 days) tests
In vivo repdroductive and developmental toxicity tests
Chronic toxicity/carcinogenicity test

Why is acute toxicity not required?

Because normally you only eat a very low dose. Only sometimes tested for for instance a hot apple.

What are the test you need to perform in order to see if your ingredient is genotoxic?

In vitro tests recommended for food ingredients:
- bacterial and/or mammalian cell mutation tests (Ames, tk-assay)
- mammalian cell micronucleus test

In vivo follow up:
- mammalian erythrocyte or bone marrow micronucleus test
- transgenic rodent somatic and germ cell gene mutation assays
- in vivo Comet assay

Is a compound seen as genotoxic when the in vitro test is positive and then invivo test is negative?

No when the invivo test is negative the ingredient will be excepted.

How does the ames test with Salmonella typhimurium go?

The salmonella can only grow if there is histidine present. When a mutation occurs this will be different and then the salmonella is able to grow. In this way you can see if your toxin is genotixic.

How does the mouse lymphoma thymide kinase (tk) assay go?

You have a mouse lymphoma cells in which you only have one intact koppy of the tk gene.
Under the toxic conditions only that one cell with the intact tk gene can grow.
However when there are mutations more cells will be able to grow.

What do the large and the small colonies stand for in the mouse lymphoma thymidin kinase assay?

The large colonies are the point mutations.
The small colonies are chromosome damage.

What happens in a micronucleus assay?

The chromosomes will not be equally divided if the dna is damaged. It might even be so severe that a micro-nucleus is formed.

If you see this micro-nucleus it is a strong indication you have a genotoxic compound

What is a commet assay?

can be done with in vitro and in vivo cels.
Cels are put on a glass plate and if the DNA is damaged you will have smaller particles and they will diffuse faster then large particles. So if the DNA is damaged you will get a tail.

What is do you do after you realize your compound is not genotoxic?

You go on to step 2 and proceed whit the hazard assessment.
You wand a dose response curve for the target organ.

Why is a dose response curve essential in your risk assessment?

you need it in order to prove taht your chemical is causing the adverse health effect. If you dont get a dose responce curve it is not the case.
You need to get the NOAEL or the BMDL

What are the experiments done in order to find the critical effect and the target organ?

Animal experiments:

Macroscopic evaluation:

external picture,
position and weight of body organs,
edema,
tumors,
haemorrhages,
inflammations

Histopathology (to see if organs are dammaged):

light microscopy (especially important for kidneys)
histochemistry

Biochemical tests:

microsomal enzymes,
cholinesterase, etc.

Analytical chemical tests:

amount of dosed substance or its metabolites

What are the negative points of animal testing?

ethical considerations
time consuming and expensive
high to low dose extrapolation
in diet compound in complex food matrix
animal to human extrapolation

If you wand to reduce animal testing, on what should you work?

You should come up with an in vitro study for reproductive-toxicity.

What might be a possible in vitro reproductive-toxicity test?

Stem cells that would become heart cells are used. If they don't become beating heart cells you know that the the product might be reproductive toxic.

The nice thing is that you test it on human cels.

Compute models can recalculate it to a daily intake of human effect.

What are the effect you should know of the dose response curve?

Still based on animal experiments
Should focus on target organ and critical effect
Proves that adverse effect is due to the compound tested
route of administration
Defines NOAEL, LOAEL, Benchmark dose (lower confidence limit) = BMD(L)10
To be used as Point of Departure for defining safe levels (ADI, TDI) and risk assessment (MOE)

What is the difference between TDI and ADI?

ADI =Acceptable Daily Intake for avoidable contaminants like additives, pesticides

TDI = Tolerable Daily Intake for unavoidable contaminants like dioxins, mycotoxins, heavy metals

How do you do a risk assessment for a non-genotoxic chemical?

You determiner the NOAEL or LOAEL form the dose response curve.
They have to be divided by 100 since there is a inter species and a inter individual difference.

From this division the ADI or TDI is obtained.

And then you have to see that if the expected daily intake (EDI) is under the ADI or TDI

What are the disadvantages of the NOAEL?

NOAEL dependent on sample size more dose groups, more animals lower
NOAELs; better experimental designs are penalized
NOAEL limited to experimental dose
NOAEL differ across studies
Only one experimental data point is used

All in all less testing is rewarded because you get a higher NOAEL. This is not good!

What is the bench Mark Dose approach?

This is the improvement of the NOAEL.

(Statistical lower confidence limit of) the dose corresponding to a small increase in effect over background level BMDL BMD BMR NOAEL

How do you do a risk hazard assessment for genotoxic compounds?

There will NOT be a zero TDI!

There are 2 different products.
The genotoxics that can be avoidable and the ones that are not avoidable.

The ones that are avoidable are NOT allowed. So you can use a genotoxic chemical.

The ones that are not avoidable can have 3 values:

ALARA: As Low As Reasonable Achievable
Virtural safe dose (VSD)
Margin of Exposure (MOE) approach

What is a DALY?

An approch to make sure you can compare different methods.

It are the disability adjusted life years.
DALY = YLL + YLD

YLL: number of life years lost (deaths)
YLD = number of years with illness or disability

The question on the page originate from the summary of the following study material:

Risk assessment of food

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