Substantiation of anti-pollution claims

For some years now, air pollution has been a highly topical issue, also gaining ground in the cosmetic industry. More and more companies advertise solutions with new ingredients that are being touted as a revolution to fully protect our largest organ against air pollution and many "old familiar" ingredients are suddenly taking on new functions in this regard. This inevitably raises the questions how can anti-pollution claims be justified and secured with today's technology? How do test strategies for anti-pollution formulations look like in this respect?

Air pollution has long been considered as a reason for serious health problems, especially when someone's exposed to certain substances over a long period of time primarily by inhalation, but not limited to.

According to the World Health Organisation (WHO) the concentration of pollutants in the air exceeds the recommended limits many times over and is the reason for millions of deaths all over the world [1].

Although ambient pollution can be caused by natural disasters like volcanic outbursts, the dramatic increase of pollution levels in the last decades has its roots in human activities.

An increasing number of publications refer to the harmful effects of air pollution on the human’s largest and most versatile organ, the skin. Our skin is the outermost interface between inside of the body and the environment and is often exposed to pollutants as well as other environmental factors like UV-irradiation.

What impact a short-term and long-term exposure to pollutants really has on our skin has not yet been conclusively clarified, as there are no convincing long-term studies up to now.

However, in this article the possibilities of testing cosmetic products on their efficacy with regard to anti-pollution claims will be examined in more detail.

Definition pollution

According to the EPA (environmental Protection Agency) Air Pollution consists mainly of fine particles and gases (VOC, volatile organic components). Pollutants (PM, particulate matter) are present as a mixture of a suspension consistent of solid and liquid particles which are classified according to their size.

PM10 (coarse particle) can only enter the nasal cavity and are mainly disposed by vibrissae. PM2.5 (fine particles) are of lower size and much more dangerous because they can penetrate deeply into the respiratory tract, remain there and harm the lung sustainable. PM0.1 (ultrafine particles) are considered as ultrathin particles which can easily enter the respiratory system and can even reach different organs spread through the blood stream.

There they can cause micro inflammation which can lead to a chain reaction and promote for instance heart attacks or other serious diseases. In addition, because of their small size paralleled by a high surface area pollutants are prone to absorb components like metals (Fe, Co, Ni) or polycyclic aromatic hydrocarbons.

Those connections can induce cell signalling cascades triggered by PM or penetrate directly into the skin. The most problematic key components of air pollution are gases, PM, ozone, cigarette smoke, polycyclic aromatic hydrocarbons (PAHs) and different Nano particles.

Skin damage through AP

The impact of pollutants on human skin can be divided into short-term and long-term effects. Whether and when these effects occur depends of course on exposure time and dose.

Some of the so far reported short-term effects are accumulation of toxins on the skin, irritation, (micro) inflammation, oxygen deficiency, clogging of pores, impairment of skin barrier, dryness / lack of moisture, excessive desquamation, altered metabolism as well as protein, lipid, and DNA oxidation.

Not every effect is directly noticeable, but a persistent dryness can for instance lead to itchy and rough feeling skin. Moreover skin can look slightly pale/dull over time.

It has also been shown that pollutants affect the metabolism of isolated human cells which in turn resulted in highly increased oxidative stress levels and consequently cell damage and reduced cell activity leading to cell death [4].

As a long-term effect, a connection between a high exposure to air pollution and signs of premature skin ageing had been reported by several researchers. PM seem to be the reason for accelerated formation of wrinkles. One mechanism could be the activation of enzymes which initiate and increase the collagen degradation process.

PM has also been linked to impurities and acne, allergies, age spots (hyperpigmentation), sensitivity, vitamin E deficiency, mutations and even skin cancer. Some facial occurrences linked to PM have also been reported like dark circles around the eye, sagging of eyebrow, increased fine lines, thinner lips and blemishes which could appear due to trapped toxins in pores.

However, some of those assumptions need to be validated in a scientific manner in further studies.

The main problem, according to unanimous opinion, is the formation of oxygen radicals (ROS) and the oxidative stress in skin cells which is dramatically increased by PM.

What can anti-pollution products really do to prevent or fight these problems?

The main objectives of current products in the market are: detoxification (cleaning), rebalance and protection which do not differ fundamentally from conventional cosmetics and their claims.

However, given the large number of different pollutants in the air or to which our skin could be exposed to in different areas, the choice of working ingredients for a functional anti-pollution formulation is not trivial.

The following points should always be considered while developing anti-pollution products:

  • Climate
  • Industrial activity
  • Urban, rural area
  • Traffic
  • Skin type
  • Ethical aspects
  • Pollutant composition
  • Indoor pollutants in the area.

Thus, an anti-pollution product should be developed with respect to the region in which it will be used and depending on the specific fine dust composition that prevails there.

Substantiation of AP claims

Parallel with the increasing cosmetic interest in Anti-Pollution, more and more ingredients have been explored and many tests have been developed and offered to support anti-pollution claims. There is a growing number of pollutants which are believed to be detrimental to the skin, but which ones are suitable to investigate the effect of a product?

First of all, one needs to have an overview about the skin parameters that are reported to be altered by pollutants. It has been found in recent research that several so called biomarkers were altered upon exposure to different pollutants.

Without product (left), with product (right)

Some biomarkers are usually increased relatively quickly after pollution treatment like:

  • pH
  • Lactic acid content
  • Sebum production and secretion
  • Advanced glycation End products (AGE)
  • Malondialdehyde (MDA) and squalene monohydroperoxide (SQOOH) (marker for lipid peroxidation)
  • Protein oxidation
  • Cellular oxidative stress levels (radicals)
  • Interleukin-1a (IL), IL-6. IL-8 (inflammatory markers)
  • Tissue oxygenation
  • Matrix Metalloproteinases (1, 2, 3, 9, 12)
  • Cell survival & proliferation
  • DNA damage

In contrast there are some downregulated biomarkers upon AP treatment:

  • Vitamin E content
  • Cholesterol content
  • Skin lipid level (triglycerides, wax esters, free fatty acids. squalene, the ratio among themselves).

Some Markers had been reported to be both, increased or reduced depending on the pollutant used:

  • Antioxidant and detoxifying enzymes (activity)
  • Transcutaneous oxygen partial pressure (O2 and CO2)
  • Cytokeratin 10, Fillagrin, Loricrin
  • Elastin and Collagen content
  • Adenosine triphosphate (ATP).

All of these markers can be addressed to evaluate the efficacy of a product or formulation against distinct particle mater treatment.

Anti-Pollution testing strategies

With the ever-increasing awareness and interest of consumers in the functioning of cosmetic ingredients, there is a need for information and even a demand for scientific evidence that companies face today, especially when it comes to relatively new topics like anti-pollution.

There is a constantly growing number of offered test strategies for efficiency testing both in vitro and in vivo. Unfortunately, there is still a lack of generally accepted standards and some methods are not reliable, reproducible, or even not sensitive enough for testing certain kinds of pollutants.

In addition, one should always take into consideration that many pollutants are not suitable to in vivo testing upon ethical issues. However, some frequently used methods for targeted AP-claims have been described in the literature:

  • Heavy metal analysis
  • Particle visualisation
  • Lipid peroxidation
  • Protein oxidation
  • Measurement of the O2 partial pressure
  • Skin gloss determination
  • Skin colour measurement
  • Hyperspectral analysis.

As mentioned before while developing a product one should consider where the product will be used (region) and what kind of pollutant prevails there. This is essential for the use of corresponding pollutants in subsequent tests. Here we are going to shed light on some of the mentioned tests.

Lipid peroxidation

One of the most frequent used approaches is the assessment of Lipid peroxidation (LP) of sebum and skin barrier lipids which can be done both in vitro and in vivo. As pollutants are mainly considered to trigger enormous amounts of oxidative stress to skin cells, LP quantitation is a useful test to assess oxidative stress which is quickly detectable after pollutant exposure.

Moreover the analysis can be done relatively easy with already established techniques like LC-MC or GC-MS. Alternatively “ready to use kits” are available to quantify for instance MDA via colorimetric or fluorometric assays.

An in vivo method where skin of volunteers (back and volar forearm) is exposed to cigarette smoke to mimic pollutant exposure has been developed and reported [2], because it has been well documented that cigarette smoke is able to oxidise sebum.

As a consequence increased levels of squalene monohydroperoxide (SQOOH) as well as malondialdehyde emerge. Elevated levels of SQOOH have already been linked to higher comedogenicity and therefore upcoming skin impurities.

After exposure of the skin to cigarette smoke sebum is removed from the subject’s skin by swabbing technique and analysed by LC-MS or GC-MS. It is reported by the developers that the method can assess short and long term effects, where with long term effects a maximum of 4 days is intended which is not really a long term exposure compared to the everyday situation.

However, another questionable point is of ethical nature. Is it really justifiable to expose human skin to distinct cigarette smoke to assess efficacy of cosmetic products where such an approach can also be done reliably in vitro? In contrast the method seems to need a relatively low number of volunteers and appears to be reproducible.

3 step strategy

A three step strategy has been reported to evaluate the efficacy of a product on molecular and cellular levels. As a model of pollutants carbon microparticles are used. Firstly, a skin barrier test is carried out. The volar forearm of subjects is treated with a test product and subsequently pollutants are applied.

After a standardised rinsing protocol remaining pollutants are assessed via imaging. Secondly, lipid peroxidation is quantified in vitro and in vivo, as a consequence of high level oxidative stress triggered by pollutants.

Thirdly, the Nrf2 pathway which cells activate to fight oxidative stress is assessed. A three step strategy seems to be more powerful as only heavy metal or lipid peroxidation tests, but the use of only carbon microparticles (1 pollutant) does not reflect the everyday situation as there is a wide variety of different pollutants which have different impacts on skin, cells and even molecular events [3].

Combination of relevant pollutants in the three step analysis would increase the expressiveness distinctly.

A standardized approach with different pollutants

Another test protocol has been published where test products efficacy is tested based on combined exposure of pollutants like cigarette smoke, ozone, infrared, UVR and blue light under standardised lab conditions in specifically developed chambers and by utilising an ozone generator.

Monitored is the anti-oxidative capacity by detecting and quantitation of arising ROS, the anti-inflammatory potential via assessment IL-6, anti-aging potential by detecting and quantifying MMP-1 as well as the evaluation of collagen and elastin integrity.

The combination of different pollutants and the possibility of standardised conditions are an advantage compared to other methods, but there is still only in vitro data from isolated cells and skin models.

Additionally, an in vivo evaluation of lipid peroxidation via squalene or MDA assessment and quantifying carbonylated proteins is offered, too. It must be positively emphasised that in this approach the certified urban dust “NIST 1649b” can be used which is consistent of most of relevant pollutants found in the air.

The physical parameters skin elasticity, hydration, barrier function, pigmentation and micro relief can be assessed too, but again, the so called long term effect considers only 5 days which per se cannot reflect the incurrence of wrinkles or decrease in skin elasticity [5].

Particle visualisation

A different approach, which we use at dermatest is to test the protection or cleansing efficacy of products by particle visualization. This can even be done in the facial area of subjects (or on hair).

A certified test dust like NIST 1649b and many others (depending on what pollutants prevailing in the area the anti-pollution product is intended to be used) can be applied to subject’s skin.

The “urban dust” consists of many common pollutants and different sizes reflecting an everyday situation very well. Firstly, following a protective approach, a testing product is applied onto a defined area on the forearm (or face) of volunteers (fig 1).

After incubation time a defined amount (15mg) of pollutants is applied to the skin via a specific sedimentation cylinder for 15 minutes. After sedimentation, samples from the encumbered skin area are taken using a specific sampling system where the uppermost skin layers are removed (similar to tape stripping).

The samples are subsequently analysed by electron-microscopy and the particles are visualised and quantified. Particles passing the protective skin “shield” of a product can be counted and assessed even concerning their size.

But this can only be done with the limitation that particles aggregate while sediment, which is well known. However, with that kind of method one can exactly evaluate the protective function of the product defending the skin against particles trying to attach.

The other way is to apply the dirt particles first in a defined area and then use the (cleansing) product of interest. After washing the skin area with a standardised protocol, which should meet the everyday situation, skin samples are taken and pollutant residues are detected and quantified via electron microscopy.

Here it can be assessed how effective a product cleans the skin from pollutants.

Conclusion

There is a very wide variety of pollutant molecules acting at different cellular levels to trigger the most common problem of air pollution, oxidative stress. Currently there are no international agreements on standardisation of anti-pollution tests but it seems logically to use certified dusts to meet the highest criteria for real pollutants and to reflect the everyday situation best.

Even it is broadly accepted that pollutants are detrimental to skin, it is not well known what kind of pollutants (or their combinations) cause which skin problem and there is no agreement on which bio markers are most suitable to reliably investigate the efficacy of a product.

Therefore the best strategy today from our point of view seems to be to stop the pollution molecules at first contact with the skin barrier and that is the reason why we decided to conduct the particle visualisation approach.

While developing an AP product, one has to take the discussed points into consideration and choose the most suitable and powerful test for claim substantiation.

It should not be forgotten, that the manufacture of cosmetic products itself is involved in anthropogenic environmental pollution in many ways, for instance, when one considers raw material extraction and water pollution. Minimising these is also a complex but necessary goal.


References

1. WHO. Ambient air pollution: Health impacts 2018. http://www.who.int/airpollution/ambient/health-impacts/en/.
2. Bielfeldt S, Böhling A, Laing S, Hoppe C, Wilhelm KP. Environmental Skin Protection Strategies – a New clinical Testing Method Employing a Cigarette Smoke Pollutant Model, SOFW Journal 142, 11;10-17; 2016
3. Cosmetic business 30-Aug-2018 A full range of tests for anti.pollution claims substantiation
4. Curpen S et al. A novel method for evaluating the effect of pollution on the human skin under controlled conditions, Skin Res Technol. 2020 Jan;26(1):50-60
5. https://www.cidp-cro.com/cidp-research-innovation/innovative-protocols

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