We’re introducing our new “test of the month” feature, to share more about the science, regulations and commercial aspects behind our key tests.
This month, at the height of summer here in the Northern Hemisphere – while we all enjoy the longer days and hopefully some sunshine! – where better to start than our human-relevant, animal-free test for phototoxicity? Read on to discover more about the test, why it’s important, and how we can help you to obtain critical data to support your new product development and safety assessment processes!
What’s phototoxicity and why test for it?
Phototoxicity is easily overlooked in safety assessment, but it’s a critically important factor for ingredients and products to be used in contact with the skin, as well as for some chemicals in the systemic circulation. Also known as photo-irritation, phototoxicity is chemically induced skin irritation that requires light for activation. The skin reaction can be caused either by topical exposure, or by a chemical reaching the skin from the inside via the bloodstream, for example following ingestion. This all means that, even when chemicals give a non-irritant result in standard skin irritation testing (such as OECD TG439), potential photo-activation could be overlooked.
It’s also possible that multiple ingredients within a formulation could interact to cause photo-irritation, so the endpoint is important to assess for finished products, too. One of the latest trends in the cosmetics industry is a move towards “suncare-as-skincare”. We’ve moved beyond the inclusion of SPF in skin creams, to a situation where many suncare products also contain active ingredients for healthy skin, and users often apply multiple products together.
The same concept extends to occupational exposure, particularly for outdoor workers in agriculture and heavy industry, especially where PPE requirements may not be strictly observed or enforced.
Given the more complex chemistries in contact with our skin in today’s world, the potential for phototoxicity is significantly increased, so the availability of relevant tests for this endpoint is timely.
Traditional tests
Phototoxicity tests assess the ability of the chemical to cause skin irritation in the absence and presence of UV light. The testing was traditionally performed on animals, but the first in vitro phototoxicity test was the 3T3 Neutral Red Uptake method, which has been in use for many years and documented in OECD Test Guideline 432. However, the test has several drawbacks:
Firstly, it lacks human relevance, because it uses mouse-derived cells cultured in medium containing foetal bovine serum (FBS).
Secondly, it lacks physiological relevance, since items are in direct contact with cells, therefore not taking skin absorption and metabolism into account.
Thirdly, the test poses practical problems for chemicals with limited aqueous solubility, since the item must be dissolved in cell culture medium for the test.
The latest human-relevant test
To address limitations of the 3T3 Neutral Red Uptake method, scientists started to assess the ability of human reconstructed skin models to predict the phototoxicity of chemicals. This type of test system is highly human-relevant and physiologically relevant, simulating real-life exposure to ingredients and formulations by topical exposure including a skin barrier. The approach also overcomes solubility issues and therefore can be applied to a wide range of items.
Following extensive validation, the reconstructed human epidermis method was adopted as a new OECD Test Guideline (TG498) in July 2023. XCellR8 was proud to support pre-validation activities for the test.
Here’s an overview of the method:
- 5 concentrations of the test item are applied to the surface of the skin model, in triplicate.
- Controls are also assessed (the negative control is water or sesame oil and the positive control is chlorpromazine).
- The dosed skin models are incubated overnight in a cell culture incubator, then one set is exposed to a defined dose of UVA while a second set is left in the dark.
- All items and controls are removed by washing and the skin models are once again incubated overnight.
- The viability of the skin models is assessed by the MTT method. (MTT is a formazan salt that is converted by intracellular enzymes to a purple metabolite, which is quantified using a spectrophotometer). In skin models that display an irritant response, either with or without UV exposure, the cells’ ability to convert MTT is compromised.
- A range of acceptance criteria must be satisfied for the test to results to be valid.
- A validated prediction model is used to determine the phototoxic potential of the test item, which is predicted to have a phototoxic potential if one or more test concentrations of the (+UVA) part of the experiment reveal a decrease in viability exceeding 30% when compared with identical concentrations of the (-UVA) part of the experiment.
- Prediction of phototoxicity is supported if, in addition, the (+UVA) induced reduction in tissue viability shows a dose response relationship.
You can find more information about the method in our handy test overview, and all the tiny details in OECD TG 498.
The test in practice at XCellR8
At XCellR8, we’re proud to provide the reconstructed human skin model method to assess the phototoxic potential of ingredients and formulations, for the cosmetics and personal care sectors and beyond. It’s an important member of our portfolio of tests that support our mission “to accelerate the world’s transition to 100% animal-free testing, through our scientifically advanced and ethical approach”. As with all our tests, we’ve assessed the method against our unique animal product free scale. The test falls into Level 5, in the green zone, supporting brands who want to comply with expectations for totally cruelty-free and vegan testing.
Interested to test your ingredients or products for phototoxicity?
To celebrate our “test of the month”, we’re offering special discounts on our phototoxicity test throughout the month of August. Discounts vary according to the number of items to test. So, whether you’re trying out this test for the first time, or already familiar with the method, feel free to get in touch with our friendly, expert scientists, who will be happy to chat about your projects and how we can make your summer extra sunny, whatever the weather!
Email info@x-cellr8.com
Call +44 (0)1925 607 134.
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