Blog – KindofStephen https://kindofstephen.com A cosmetic formulator and skincare expert's blog on the latest skin care and cosmetic research, ingredients, and beauty news Wed, 24 Apr 2019 12:55:55 +0000 en-CA hourly 1 https://wordpress.org/?v=5.1.1 https://i1.wp.com/kindofstephen.com/wp-content/uploads/2017/06/cropped-profile-2.jpg?fit=32%2C32&ssl=1 Blog – KindofStephen https://kindofstephen.com 32 32 114122149 Seeing oxidation and reduction https://kindofstephen.com/seeing-oxidation-and-reduction/ Sat, 02 Mar 2019 18:37:26 +0000 https://kindofstephen.com/?p=3285 Why do our cosmetics change over time? Sometimes it’s because of oxidation, the loss of electrons. Ascorbic acid is easily oxidized, losing two electrons, becoming dehydroascorbic acid. Enzymes and antioxidants in our body can give dehydroascorbic acid back those two electrons, reducing it back into ascorbic acid.⁣⁣ Your foundation getting darker after it dries isn’t […]

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Why do our cosmetics change over time?

Sometimes it’s because of oxidation, the loss of electrons. Ascorbic acid is easily oxidized, losing two electrons, becoming dehydroascorbic acid. Enzymes and antioxidants in our body can give dehydroascorbic acid back those two electrons, reducing it back into ascorbic acid.⁣⁣

Your foundation getting darker after it dries isn’t oxidation, even though it’s often called that. It’s just the water or solvent in the foundation evaporating. If you’ve ever painted, you’ll know that wet paints tend to dry darker.

Methylene blue is a deeply blue organic dye. It is can be used in analytical chemistry as a redox (reduction-oxidation) indicator.⁣⁣
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In oxidizing environments, methylene blue is a bright blue, in reducing environments the methylene blue accepts electrons and becomes leukomethylene blue which is colourless.⁣⁣
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In the vial is an alkaline solution of glucose. Glucose is a reducing sugar, which means it can donate electrons. This creates a reducing environment for methylene blue, so the methylene blue is reduced to leukomethylene blue.⁣⁣
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When the vial is shaken, oxygen is dissolved into the solution – this oxidizes the leukomethylene blue causing it to lose electrons, turning it back into bright blue methylene blue. In turn, the oxygen is reduced to water.⁣

As the oxygen is consumed by the reaction, the glucose reduces the methylene blue – turning it back into the clear and colourless leukomethylene blue. In turn, the glucose is oxidized to gluconic acid.⁣
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In the video, you’re seeing excess oxygen dissipating out of the solution, as well as oxygen being consumed by the reaction.⁣

This experiment is repeatable by shaking in more oxygen, but won’t go on forever – eventually, all of the glucose will have been oxidized to gluconic acid and the glucose is needed to reduce the methylene blue.⁣

Oxidation was first observed with oxygen, hence its name. However, the modern definition of oxidation is the loss of electrons. Anything that can gain electrons, causing something else to lose electrons, is an oxidizing agent.⁣⁣
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Some common oxidizing agents that aren’t oxygen are elemental halogens like fluorine, chlorine, bromine, and iodine. Fluorine (F2) is such a strong oxidizer it can oxidize water into oxygen! ⁣⁣
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2F2 + 2H2O → 4HF + O2

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Yes Virginia, vegan collagen is real! https://kindofstephen.com/yes-virginia-vegan-collagen-is-real/ Fri, 04 Jan 2019 02:31:34 +0000 https://kindofstephen.com/?p=3210 When Algenist launched the Genius Liquid Collagen with “vegan collagen” my first thought was, “What? Only animals have collagen!”⁣ Well, you’re looking at a vial of collagen that has been produced by yeast.⁣⁣Collagen is the main structural protein in animals, there are over 28 types of collagen. Type I collagen makes up about 90% of […]

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When Algenist launched the Genius Liquid Collagen with “vegan collagen” my first thought was, “What? Only animals have collagen!”⁣

Well, you’re looking at a vial of collagen that has been produced by yeast.⁣

Collagen is the main structural protein in animals, there are over 28 types of collagen. Type I collagen makes up about 90% of the collagen found in humans.

Collagen gives our skin its strength, flexibility, structure, and durability. Collagen is a triple helix, made of three coils of amino acids wrapped around each other. This coiled structure allows collagen to be stretched without breaking.⁣ Check out an earlier post I wrote about collagen for more information.

Plants and microbes don’t normally make collagen, but turns out they can! With some help from science, of course.⁣

Vegan collagen is often produced from modified yeast and bacteria, scientists have been doing this for decades. Collagen can also be produced by modified plants, like the tobacco plant.⁣

In one method, 4 genes that encode for the building blocks of collagen were added into a yeast’s genetic structure. The human genes were expressed in the modified yeast and they started producing the building blocks of human collagen type I. These building blocks were collected and treated with pepsin (a digestive enzyme), which assembled them into collagen and broke down any material that didn’t form properly.⁣

Why make microbe or plant-based collagen? It’s often purer and it doesn’t rely on animals. Though it occurs rarely, animal collagen can cause foreign body or allergic reactions. Animal sources of collagen are fish, pigs, and cows.⁣

Collagen is useful as a moisturizer for the skin, but also has medical applications. Collagen is used as a material for cosmetic filler, as carriers in drug delivery, as sutures, and as scaffolds for tissue engineering. Collagen can also be modified and used for neuron regeneration, blood vessel repair, bone regeneration, wound healing, and more!⁣

M Nokelainen, High‐level production of human type I collagen in the yeast Pichia pastoris, Yeast, 2001. DOI: 10.1002/yea.730

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Paperview: Air quality and relative humidity in commercial aircrafts https://kindofstephen.com/paperview-air-quality-and-relative-humidity-in-commercial-aircrafts-an-experimental-investigation-on-short-haul-domestic-flights/ Thu, 15 Nov 2018 15:50:44 +0000 https://kindofstephen.com/?p=3143 Why is airplane air so dry? Outside air at a plane’s altitude is very cold and less dense. Cold air can hold less water compared to warmer air. At a plane’s flight height (around 10 km) the outside air can hold about 10% what it can at ground-level. Outside air is warmed, filtered, and then […]

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Why is airplane air so dry?

Outside air at a plane’s altitude is very cold and less dense. Cold air can hold less water compared to warmer air. At a plane’s flight height (around 10 km) the outside air can hold about 10% what it can at ground-level.

Outside air is warmed, filtered, and then cycled through the cabin air about 20 times an hour, depending on the plane model. Air inside the cabin is also filtered and recycled. The mix between outside air and recycled inside air is about 50/50.

Below, the solid black line shows the measured relative humidity inside an airplane’s cabin during a flight. You’ll see that about 15 minutes after take-off the relative humidity drops quickly to its low of about 10% – which is maintained until close to landing.

Why can’t they humidify the cabin air?

That does happen! But let’s go over some of the issues with humidifying a cabin.

As warm air cools, we know that the amount of water it can hold drops as well and that water it can’t hold anymore will condense on surfaces. We’ve all experienced this when we see condensation or “sweat” on a cold glass during a hot, humid day.

The outer layers of a plane are much, much colder than the inside of the plane and that change in temperature can cause condensation. That condensed water can be absorbed into the plane’s insulation which adds weight and more fuel costs, cause corrosion of metal parts, and even affect electrical wiring.

OK so, what about these humidified planes?

Newer plane models, like the Airbus A350 and Boeing 787, can be ordered with on-board humidifiers. They work by passing the cabin air over a water-moistened pad, as the water evaporates – it humidifies the air. That’s not all though, to prevent some of the mentioned problems above, the humidifying system is also paired with a drying system. The drying system warms incoming air and passes it through a silica gel – which absorbs excess water and dries the air. This reduces condensation as cabin air passes through to the outer areas of the plane – where water can cause problems.

These humidified areas aren’t available to everyone though. They’re often standard for areas that the crew members are in, but when it’s available to passengers it often comes with a premium. CTT, one manufacturer of these humidifying/drying systems, says that they’re often only found in business class and other luxury sections.

So what can you do about the dry air in a plane?

While the dry air can feel uncomfortable for our skin, it’s not likely to cause long-term problems.

Trans-epidermal water loss (TEWL) from the skin does seem to increase because of the low humidity, which can lead to dryer skin – especially on longer flights. One experiment found that TEWL increased from 13.0 ± 2.2 g/m²h at 60% humidity to 20.1 ± 4.9 g/m²h during a simulation of a 6-hour flight.

Applying a moisturizer before or during a flight could be of great help. Whether that’s a sheet mask, mist, lotion, balm, or potion. Or if you’re lazy like me and don’t feel like doing anything, your skin will naturally recover once you’re off the plane and back at a normal humidity.

Based on experiments with crew members, we tend to feel the low humidity the most in our eyes (10.8%), mouth and throat (7.0%), and nose (5.0%). Sinus congestion was also a common complaint, occurring in 29.0% of those surveyed.

Moisturizing eye drops (I like the Systane Ultra drops) can help with the eyes. A light-swipe of petrolatum (Vaseline) or another balm inside the nostrils can help with the nose. As for the mouth and throat, keep some water handy! You don’t need to guzzle gallons though, there’s little evidence that low humidity leads to your body becoming dehydrated. One experiment found that for a 6 hour flight, a 60 kg person would need about 450 ml of extra water.

Source: Giaconia, C., Orioli, A., & Di Gangi, A. Air quality and relative humidity in commercial aircrafts: An experimental investigation on short-haul domestic flights. Building and Environment (2013). DOI: 10.1016/j.buildenv.2013.05.006

Source: Lindgren, T., Norbäck, D., & Wieslander, G. Perception of cabin air quality in airline crew related to air humidification, on intercontinental flights. Indoor Air (2007). DOI: 10.1111/j.1600-0668.2006.00467.x

Hashiguchi, N., Takeda, A., Yasuyama, Y., Chishaki, A., & Tochihara, Y. Effects of 6-h exposure to low relative humidity and low air pressure on body fluid loss and blood viscosity. Indoor Air (2013). DOI: 10.1111/ina.12039

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Paperview: Sunscreen application to the face persists beyond 2 hours in indoor workers https://kindofstephen.com/sunscreen-office-indoors/ Wed, 24 Oct 2018 12:50:54 +0000 https://kindofstephen.com/?p=3109 What happens to your sunscreen throughout a work day? I often get this question, especially from people who work inside for most of the day. A group of researchers at Mahidol University in Thailand did an experiment that may provide us with some guidance. The researchers took 20 people (15 women) with mostly skin phototype […]

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What happens to your sunscreen throughout a work day? I often get this question, especially from people who work inside for most of the day. A group of researchers at Mahidol University in Thailand did an experiment that may provide us with some guidance.

The researchers took 20 people (15 women) with mostly skin phototype III and up. Skin phototype III means that they tan, but sometimes get mild burns.

The participants were asked to apply 1 gram of sunscreen to their face. The sunscreen was mixed with a blue fluorescent dye that would glow under UV light. This glow allowed the researchers to see the sunscreen on the skin and note changes in its brightness throughout the day.

The people only wore the sunscreen and were asked not to reapply. They weren’t allowed to use makeup or other skincare. They were also allowed up to 1 hour outside. The temperature outside was between 23 and 35 degrees Celsius throughout the day and described as humid. The indoor condition was inside the air conditioned Siriraj Hospital.

Every 2 hours, the researchers took a photo of the people and measured the glow of the sunscreen under a UV light. They looked at the cheeks, forehead, nose, moustache area, and the chin. They used a Visia device to help make sure the photos were consistent.

Sunscreen brightness reduction every 2 hours by percent. The bars indicate the range in measurement values.

The researchers found that the fluorescent glow on the people’s faces decreased the most in the first 2 hours after applying the sunscreen. On average the areas of the face were 16.3% less bright.

Between 2 hours and 4 hours after application, the brightness decreased by a further 7.4 percentage points on average. Between 4 hours and 8 hours, there was an average 4.5 percentage point decrease in brightness.

At the end of the day, there was about a 30% decrease in brightness on average compared to just after applying the sunscreen.

A 30% decrease in brightness in this experiment doesn’t necessarily mean a 30% decrease in sunscreen on the skin. There are ways to model this more accurately, but they did not have the tools in this experiment.

So what does this mean for you? At the end of the day, you’ll still have to use your best judgement.

If you pigment easily, are very concerned about photoaging, or have a family history of skin cancer – I think the best recommendation is to be on the safe side and reapply at least once around 2 hours.

If don’t care that much, consider the opposite, about 70% of the glow from the sunscreen still remained after 8 hours.

In either case, that first application is important. I’d recommend choosing a sunscreen with a high SPF and UVA protection and aiming for a 2 mg/cm² layer. An easy way to make it more likely you’ve applied that amount is to apply your sunscreen in two layers.

Source: Rungananchai, C., Silpa-archa, N., Wongpraparut, C., Suiwongsa, B., Sangveraphunsiri, V., & Manuskiatti, W. (2018). Sunscreen Application to the Face Persists Beyond 2 Hours in Indoor Workers: An Open Label Trial. Journal of Dermatological Treatment, 1–14. doi: 10.1080/09546634.2018.1530440

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A quick look at collagen https://kindofstephen.com/a-quick-look-at-collagen/ Thu, 20 Sep 2018 12:55:40 +0000 https://kindofstephen.com/?p=3050 Collagen, you’ve seen it in your skincare products and have probably eaten it at some point (Yay for artificially-coloured and jiggly gelatin). But what is it? Collagen is composed of a triple helix, three strands of proteins made up of joined amino acids wrapped around each other. The main amino acid constituents of these proteins […]

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Collagen, you’ve seen it in your skincare products and have probably eaten it at some point (Yay for artificially-coloured and jiggly gelatin). But what is it?

Collagen is composed of a triple helix, three strands of proteins made up of joined amino acids wrapped around each other. The main amino acid constituents of these proteins are glycine, proline, hydroxyproline, lysine, and hydroxylysine. The unique chemical structure of the amino acids helps form the shape and structure that their compounds make.

There are many types of collagen, which differ in their amino acid composition. Type I collagen is the most abundant in the human body, and Type I, III, IV, and others are found in our skin. In our body, multiple strands of collagen are found bundled together in fibrils.

You may have heard that ascorbic acid or Vitamin C is crucial in the formation of collagen, but how? Ascorbic acid is used in the conversion of proline to hydroxyproline along with oxygen, and alpha-ketoglutarate. The reaction is catalyzed or sped up by the enzyme prolyl hydroxylase and an iron. Similarly, it is needed in the hydroxylation of lysine to hydroxylysine by the enzyme lysyl hydroxylase.

Collagens are naturally glycosylated, meaning they have sugar molecules bound to them – they are found attached to the lysine and hydroxylysine molecules by the enzymes galactosyltransferase and glucosyltransferase. While this glycosylation is not fully understood, they seem important in forming and retaining the structure of the collagen. You may have heard of glycation or advanced glycation endproducts (AGEs), this happens when excessive sugar molecules are bound to the collagen non-enzymatically and can affect its structure, function, and flexibility.

The additional -OH (hydroxy) group on the hydroxyproline helps water molecules bind tightly to collagen. The coiled structure of collagen’s triple helix gives it impressive tensile strength and allows it to stretch when forces are applied. When too much force is applied the triple helix structure can become disorganized and damaged, no longer able to return to its triple helix form.

Experiments, where collagen was exposed to UV radiation in vitro, have shown that free radicals generated from the UV energy can cleave or break apart some of the bonds holding the amino acids together. When enough bonds are broken the triple helix structure can no longer be maintained and the collagen fibre loses its shape and function. Adding ascorbic acid to the solution of collagen, when it was exposed to UV, reduced some of the free radicals produced – leading to fewer bonds breaking and structure disruption. This may highlight one of the ways naturally present antioxidants in the skin help us defend against the environment.

N. Metreveli, L. Namicheishvili, K. Jariashvili, G. Mrevlishvili, A. Sionkowska. Mechanisms of the influence of UV irradiation on collagen and collagen-ascorbic acid solutions. International Journal of Photoenergy (2006), DOI: 10.1155/IJP/2006/76830

Duer Research Group. Collagen glycation and diabetes. Website, URL: https://www.ch.cam.ac.uk/group/duer/research/collagen-glycation-and-diabetes

A. Masic, L. Bertinetti, R. Schuetz, S.W. Chang, T.H. Metzger, M.J. Buehler, P. Fratzl. Osmotic pressure induced tensile forces in tendon collagen. Nature Communications (2015), DOI: 10.1038/ncomms6942

J.M. Waller, H.I. Maibach. ge and skin structure and function, a quantitativeapproach (II): protein, glycosaminoglycan, water, andlipid content and structure. Skin Research and Technology (2006), DOI: 10.1111/j.0909-752X.2006.00146.x

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The enzyme lysozyme and xanthan gum https://kindofstephen.com/the-enzyme-lysozyme-and-xanthan-gum/ Sun, 16 Sep 2018 19:45:54 +0000 https://kindofstephen.com/?p=3060 These are crystals of lysozyme, an enzyme which can break apart cell wall peptidoglycan of certain bacteria. It is part of our and other animals’ immune systems. Lysozyme is more effective against gram-positive bacteria, as gram-negative bacteria have additional cell membranes that make it harder for the lysozyme to reach the peptidoglycan. Lysozyme is often […]

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These are crystals of lysozyme, an enzyme which can break apart cell wall peptidoglycan of certain bacteria. It is part of our and other animals’ immune systems. Lysozyme is more effective against gram-positive bacteria, as gram-negative bacteria have additional cell membranes that make it harder for the lysozyme to reach the peptidoglycan.

Lysozyme is often used in the processing of xanthan gum. Xanthomonas campestris, a gram-negative bacteria, produce an exopolysaccharide which is a gooey, thick, and sticky slime. This slime may help the bacteria create a comfortable environment for itself and also act as camouflage from other organism’s immune systems.

The slime (aka xanthan gum) is used in cosmetics because it imparts viscosity, lubricity, and acts as a humectant water-binding film former. It also increases the yield stress of water, meaning things suspended in it don’t settle as fast. “Raw” xanthan gum can resemble snot and be cloudy.

In a series of processing steps, the Xanthomonas campestris’ cells are stripped of their membranes and broken apart – this can be done by heating in alkaline water then by treatment with lysozyme and protease. The xanthan gum becomes less gloopy and crystal clear.

Lysozyme can be sourced from a variety of things, but most commonly hen egg whites. It’s not often clear what the source of lysozyme is, so depending on the transparency of the supplier, it’s possible that products labelled ‘Vegan’ may have used animal lysozyme treated xanthan gum.

Plant-based lysozymes do exist, but their structures and functions often differ from animal lysozymes. Genetically modified organisms have been created to produce lysozymes more closely resembling animal lysozymes, but GMOs can be an issue for those who choose vegan products.

I first encountered this conundrum during a meeting with a supplier when they were promoting their vegan xanthan gum, which was slightly less clear than their regular grades. I’d always assumed xanthan gum was vegan, since it was made from bacteria. Sadly, the product line has been discontinued, but one of the largest chemical companies in the world recently launched a clear vegan xanthan gum that’s also GMO-free.

To see more images of lysozyme crystals, check out Dr. Kalju Kahn’s gallery created by students at UCSB.

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Paperview: Evaluation of the protection of a broad-spectrum SPF50+ sunscreen against DNA damage https://kindofstephen.com/paperview-evaluation-of-the-protection-of-a-broad-spectrum-spf50-sunscreen-against-dna-damage/ Wed, 22 Aug 2018 17:00:36 +0000 https://kindofstephen.com/?p=3000 Cyclobutane pyrimidine dimers (CPDs) are a form of DNA damage that is caused by UV exposure. CPDs interfere with base pairing during DNA replication – which can lead to mutations and cancer. UVB radiation is directly absorbed by DNA. The energy causes changes in the bonding of pyrimidine structures found in DNA leading to CPDs […]

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Cyclobutane pyrimidine dimers (CPDs) are a form of DNA damage that is caused by UV exposure. CPDs interfere with base pairing during DNA replication – which can lead to mutations and cancer.

UVB radiation is directly absorbed by DNA. The energy causes changes in the bonding of pyrimidine structures found in DNA leading to CPDs and pyrimidine-pyrimidone (6-4) photoproducts.

UVA on the other hand is poorly absorbed by DNA, but was also found to cause CPD formation in human skin. CPDs were found to remain longer in the skin when there was UVA exposure, leading to speculation that UVA may also suppress a repair mechanism.

Our cells do have DNA repair capabilities, where damaged DNA is excised and replaced – but these processes can be overwhelmed by an accumulation of damage.

Experiments have measured the amount of CPD formation in human skin when exposed to UVB. One study found that CPDs were formed even when there was no visible sunburn (0.5 sunburn dose). They also found CPDs in both the epidermis and dermis and these levels were elevated for about 10 days as the skin sloughed off.

These two images from the paper show (A) skin that was not exposed to UVB and (B) skin that was exposed to UVB. The brown staining of the cells indicates presence of CPDs.

The amount of CPDs found in both the epidermis and dermis increased as UVB exposure increased.

A recent experiment performed by Pierre Fabre (manufacturers of Avène) looked at the effect sunscreen had on the  formation of CPDs in human skin after UV exposure.

14 volunteers applied a sunscreen to their forearm and were exposed to UVB and UVA on skin protected by the sunscreen and also on unprotected skin. The area covered in sunscreen received 15 times the dose of UV to cause sunburn, whereas the unprotected skin received 2 times the dose.

After this exposure, their skin was blistered by vacuum and the contents of the blister were examined for CPDs using two different methods: immunostaining and spectrometry (HPLC-MS).

They found that the unprotected skin after exposure to UV had an elevated ratio of CPDs to normal DNA bases (90 CPD to 106 DNA bases). In comparison, the skin protected with the sunscreen had an amount of CPDs similar to unexposed skin and statistically significantly less than the unprotected skin (P < 0.001) – even though the area received more UV exposure. The CPD to normal DNA base ratio was not reported for the sunscreen protected and unexposed skin.

The sunscreen was not named, but it is SPF 50+, broad spectrum, and contained; Tinosorb M and S, Iscotrizinol, Avobenzone, and the antioxidant bis-ethylhexyl-hydroxydimethoxy benzylmalonate.

Preventing the formation of CPDs from reducing UV exposure is the most well-researched option, but there are other newer methods that are emerging – some of which are already available on the market.

Photolyase is a DNA repair enzyme that can be activated by the absorption of a photon and transfer an electron to the CPD, this can separate the CPD back into two normal pyrimidine bases – with the right timing. In humans, the photolyase enzyme no longer works, but there is some evidence that topical application of photolyase may reduce the formation of CPDs. An experiment where photolyase encapsulated in liposomes combined with light exposure was applied to human skin reduced the formation of CPDs by 40%-45% after exposure to UVB.

You can watch a lecture given by Aziz Sanzar about photolyase and DNA repair below. He won the Nobel Prize in Chemistry in 2015 for his work along with his colleagues Tomas Lindahl and Paul Modrich.

S.K. Katiyar, M.S. Matsui, H. Mukhtar, Kinetics of UV light–induced cyclobutane pyrimidine dimers in human skin in vivo: An immunohistochemical analysis of both epidermis and dermis, Photochemistry and Photobiology (2002), DOI: 10.1562/0031-8655(2000)0720788KOULIC2.0.CO2
J. Gwendal, T. Douki, J. Le Digabel, et al, Evaluation of the protection of a broad-spectrum SPF50+ sunscreen against DNA damage, Journal of the American Academy of Dermatology (2018), DOI: 10.1016/j.jaad.2018.05.570

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Urban particulate matter in air pollution penetrates into the barrier-disrupted skin and produces ROS-dependent cutaneous inflammatory response in vivo https://kindofstephen.com/urban-particulate-matter-in-air-pollution-penetrates-into-the-barrier-disrupted-skin-and-produces-ros-dependent-cutaneous-inflammatory-response-in-vivo/ Fri, 13 Jul 2018 05:00:03 +0000 https://kindofstephen.com/?p=2955 Anti-pollution or anti-particulate matter has become a huge buzzword in cosmetics. Pollution and particulate matter have been linked to many negative health effects (mainly cardiovascular) and while the link to skin health and acceleration of ageing are logical…does the data support it? There have a been a few correlational studies that have shown that people […]

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Anti-pollution or anti-particulate matter has become a huge buzzword in cosmetics. Pollution and particulate matter have been linked to many negative health effects (mainly cardiovascular) and while the link to skin health and acceleration of ageing are logical…does the data support it?

There have a been a few correlational studies that have shown that people living in areas with higher levels of pollution exhibit more signs of oxidative stress in skin lipids and some have even correlated it with increased wrinkling. But what’s the mechanism and can particulate matter even penetrate the skin?

A group of researchers from Seoul used an in vivo mouse and in vitro keratinocyte model to study this.

First was the collection of particulate matter from the air. To do this they set up a vinyl tarp on a rooftop near a busy intersection to collect dust. The particulate matter was then purified and separated to be used in the experiment. The majority of the particles ranged from 200 to 300 nm. Particulates found included: Naphthalene, biphenyl, acenaphthylene, acenaphthene, fluorene, dibenzothiophene, and 28 others identified.

For the in vitro portion of the experiment, cell cultures of human primary keratinocytes were performed with varying concentrations of the particulate matter. The cells absorbed the particulate matter, and the researchers found a concentration-dependent increase of inflammatory cytokine IL-8 and collagenase MMP-1. They also found that the addition of an antioxidant, n-acetyl cysteine, was able to suppress this effect.

In the in vivo portion of the experiment, the researchers used mice that did not produce melanin and divided them into two skin conditions: One with their skin intact, and another with barrier-damaged skin. To damage the skin barrier they stripped the skin 10 times with tape to remove layers of the stratum corneum. The particulate matter was applied 10 times over 2 weeks and included a skin penetration enhancer (DMSO).

While the in vitro results may be “scary”, the in vivo results were milder. Particulate matter was shown to penetrate into the intercellular space of the barrier-disrupted mice, but not the intact mice. Particulate matter was found in hair follicles of both, but there was no epidermal penetration of the particulate matter in the intact mice.

The researchers did find an increase in inflammation in the particulate matter treated skin compared to skin not exposed- whether or not the sin was intact or tape-stripped. However, the inflammation was much more severe in the tape-stripped group. The researchers also showed that intradermal n-acetyl cysteine was able to ameliorate the increase in inflammation caused by particulate matter, but they did not perform this portion of the experiment on the intact mice. It’s likely this same treatment will have a similar effect in the intact mice, but it is unknown.

The researchers also point out some issues with their own experiment: The concentration of particulate matter may not reflect the amount that a person would be exposed to and that their sampling of particulate matter had a high concentration of sulfur which may be unique to their location. It’s also important to remember that mice are not humans, and we may react differently.

While it’s likely that the addition of anti-inflammatories and antioxidants may help attenuate some of the potential inflammation caused by pollution and particulate matter, it’s unknown which chemicals and what combinations are most effective for humans. There’s also no standard measurement to gauge a protective effect so it is impossible to compare one product to another. Again, we see another case of the marketing being ahead of the science.

Source: Jin Seon-Pil, Li Zhenyu, Choi Eun Kyung, Lee Serah,
Kim Yoen Kyung, Seo Eun Young, Chung Jin Ho, Cho Soyun.Urban particulate
matter in air pollution penetrates into the barrier-disrupted skin and produces ROSdependent
cutaneous inflammatory response in vivo.Journal of Dermatological Science
https://doi.org/10.1016/j.jdermsci.2018.04.015

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Skin penetration of Vitamin C (Ascorbic acid): Part II https://kindofstephen.com/skin-penetration-of-ascorbic-acid-part-ii/ Wed, 13 Jun 2018 12:00:48 +0000 https://kindofstephen.com/?p=2627 “Applying 15% Vitamin C for three consecutive days creates a reservoir effect in the skin.” Firstly, I want to remind you that this study was done on pig skin – not humans. The way that ascorbic acid is stored and metabolized in pig skin may vary from human skin. Most animals, like pigs, are able […]

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“Applying 15% Vitamin C for three consecutive days creates a reservoir effect in the skin.”

Firstly, I want to remind you that this study was done on pig skin – not humans. The way that ascorbic acid is stored and metabolized in pig skin may vary from human skin.

Most animals, like pigs, are able to synthesize their own ascorbic acid from glucose, but humans cannot. It’s possible that this data from pigs will be similar to human data, but it’s also very possible that it won’t be. Neither has been proven yet. Presenting an assumption as truth is misleading – but often done in marketing.

I also want to remind you that the way that the ascorbic acid was applied to the skin was not the same way that we apply our skincare. In these experiments, the ascorbic acid solution was applied with a Hill Top Chamber, which occludes the skin, reducing evaporation and theoretically enhancing skin penetration.

For this part of the experiment, Pinnell and his group applied a 15% ascorbic acid solution at pH 3.2 to pig skin for 5 days with a Hill Top Chamber. After the 5th day, application of the ascorbic acid was stopped and ascorbic acid levels in the skin were monitored for an additional 5 days.

After the 3rd day of application of the ascorbic acid serum, the ascorbic acid levels in the skin do appear to reach a peak around 1100 pmol/mg. The deviation around the mean does appear to be reducing with each further day between the 3 subject pigs.

We do need to consider if this theoretical peak amount of ascorbic acid is reached in real-life situations. The living conditions of the pigs in the study were not described, so it’s possible that they were not exposed to natural daylight. It’s understood that UV exposure reduces the amount of ascorbic acid in the skin. UV increases the production of free radicals in the skin, and ascorbic acid is part of the natural antioxidants in the skin which help neutralize these free radicals.

In an experiment using human skin models, it was found that exposure to 16.9 joules/cm² (About 12 minimal erythemal dose equivalent) of UV reduced ascorbic acid levels in the skin model by almost ⅓. This was a higher amount of UV exposure the experimenters expected, they were also unable to detect dehydroascorbic acid in the skin. The study does have some issues which “may be explained by the high levels of ascorbate present in the [tissue] medium…added by the manufacturer to increase collagen synthesis”.

“Vitamin C remains in the skin for 3-4 days and doesn’t wash out”

This marketing claim may be due to some confusion of the term “washout”. In drug experiments a “washout period” refers to the period of time when treatment is stopped, it does not necessarily mean that the skin is washed out.

After applying the 15% ascorbic acid solution to the pig skin, they discontinued application and monitored ascorbic acid levels in the skin. Unfortunately, the methodology in this portion of the experiment isn’t explicitly described. It is unclear, for example, if the pig’s skin was washed each day. The washing procedure is described as “…at the end of the experiment, the formulation was washed vigorously from the skin with water.”

Because most of us use surfactant based cleansers to wash our skin, this data may not be as applicable as the pig’s skin was washed with only water. However, the pig’s skin was removed of stratum corneum before ascorbic acid measurement and the lower layers of skin are likely less affected by the washing and surfactant based-cleanser.

Based on this data, the half-life (the amount of time it takes for the detected ascorbic acid levels to drop by half) was estimated at around 4 days. But as mentioned above, it’s unclear what the living conditions of the pigs were and whether or not they were exposed to sunlight which reduces antioxidant levels in the skin.

Can Vitamin C derivatives increase levels of Vitamin C in skin?

The last portion of the Pinnell experiment looked at whether or not the topical application of Vitamin C derivatives could increase levels of Vitamin C as ascorbic acid in pig skin.

For 24 hours, solutions of dehydroascorbic acid, 10% ascorbyl-6-palmitate, 12% magnesium ascorbyl phosphate, and 15% ascorbic acid were applied to pig skin. Compared to control, only the 15% ascorbic acid solution created a statistically significant increase in ascorbic acid levels in the skin.

For the derivatives, there was no statistically significant difference between the application of the derivative and control (no application of derivatives or Vitamin C) – which implicates that, at least for pig skin, these specific derivatives do not convert to Vitamin C.

For the solutions of dehydroascorbic acid, pig skin levels of ascorbic acid were 7.51 ± 3.34 pmol/mg for 20 mM dehydroascorbic acid and 8.70 ± 2.13 pmol/mg for 1 M dehydroascorbic acid. Where no dehydroascorbic acid was applied levels of ascorbic acid were 9.24 ± 3.55 pmol/mg.

In conclusion…

It surprises me how influential this one study on ascorbic acid applied to pig skin has become in terms of marketing language for brands.

Even later studies with Dr. Pinnell as an author leave out that the data are collected from pig skin, “Following topical application, once the skin is saturated with L-ascorbic acid, it remains with a half-life of about 4 d (Pinnell et al, 2001).”

While this experiment is some of the best data we have in terms of ascorbic acid penetration based on formulation, the key point to remember is that human skin cannot be assumed to behave the same as pig skin.

So if you see a claim similar to “15% Vitamin C at pH 3.5 is the most effective concentration”, please imagine me beside you whispering “…for pigs”.

Edit: An error was made in the original version published, pigs can synthesize Vitamin C from glucose, but humans can not. Guinea pigs also cannot synthesize their own Vitamin C.

Edit: An error was made in the original version published, pmmol was corrected to pmol.

Source: Pinnell, S. R., Yang, H. , Omar, M. , Riviere, N. M., DeBuys, H. V., Walker, L. C., Wang, Y. and Levine, M. (2001), Topical L‐Ascorbic Acid: Percutaneous Absorption Studies. Dermatologic Surgery, 27: 137-142. DOI: 10.1046/j.1524-4725.2001.00264.x

Podda, M., Traber, M.G., Weber, C., Yan, L., Packer, L. (1998), UV-Irradiation Depletes Antioxidants and Causes Oxidative Damage in a Model of Human Skin, Free Radical Biology and Medicine, 24: 55-65. DOI: 10.1016/S0891-5849(97)00142-1

"Skin penetration of Vitamin C (Ascorbic acid): Part II" on KindofStephen.

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Skin penetration of Vitamin C (Ascorbic acid): Part I https://kindofstephen.com/skin-penetration-of-ascorbic-acid/ Thu, 24 May 2018 12:45:44 +0000 https://kindofstephen.com/?p=2584 Today I wanted to look at a research paper primarily led by Dr. Sheldon R. Pinnell. He is one of the founders of Skinceuticals and contributed much of the early research on the use of Vitamin C as ascorbic acid on skin. He and his group also discovered the synergistic effect of Vitamin C, Vitamin […]

"Skin penetration of Vitamin C (Ascorbic acid): Part I" on KindofStephen.

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Today I wanted to look at a research paper primarily led by Dr. Sheldon R. Pinnell. He is one of the founders of Skinceuticals and contributed much of the early research on the use of Vitamin C as ascorbic acid on skin. He and his group also discovered the synergistic effect of Vitamin C, Vitamin E, and Ferulic acid – which is commonly used in many products on the market today.

The data from this paper is often quoted in marketing material for Vitamin C serums, but one extremely important piece of information is often left out – the data was collected from pigs, white Yorkshire pigs to be exact.

Many people also have ethical concerns when it comes to the use of animals in cosmetic research. Synthetic and lab grown human skin equivalents are being researched and tested which will one day replace the use of animal as well as human testing in cosmetics.

It should be clear that human skin and pig skin are not the same, but they do have similar properties which is why it is often used in experiments. However, one should never assume that data from a pig can be assumed to be the same for a human. The movement and deposition of chemicals often differs between human and pig skin.

From my searches, I haven’t been able to find similar research performed on humans. This paper in particular has led to some of the often quoted “rules” about ascorbic acid.

“Ascorbic acid must have a pH below 3.5 for effective penetration.”

Pinnell and his group tested a 15% ascorbic acid solution adjusted to different pHs ranging from 2 to 5. The 15% ascorbic acid solutions also contained 2% zinc sulfate, 0.5% bioflavonoids, 1% hyaluronic acid, and 0.1% citrate.

While the control situation wasn’t described it’s likely either the vehicle (product without the ascorbic acid) or a water solution was applied to the skin. The control measurement shows that there is some inherent levels of ascorbic acid already present in the skin from the diet.

The test solutions were applied to the pig skin using a Hill Top Chamber. A Hill Top Chamber is a small and round disk which is placed on the surface of the skin, the product is placed in the chamber or a piece of fabric is soaked in the testing material, and the entire chamber is then sealed. This reduces loss of product from evaporation and is a common method of performing occlusive test patches.

The ascorbic acid solutions at pH 2.5, 3.0, 3.5, 4.0, and 5.0 were performed on three pigs, however the control, pH 2, and 4.5 were only performed on two pigs.

The Hill Top Chamber was soaked with 0.2 mL of the ascorbic acid solution then sealed for 24 hours. After this period of occlusion, the skin washed then stripped of the stratum corneum and then small pieces of the skin was removed and tested for ascorbic acid content.

As you can see from the data, the amount of ascorbic acid found in the skin was much higher in ascorbic acid solutions at pH 3.5, 3.0, 2.5, and 2.0. The researchers theorize that it is due to the pKa of ascorbic acid which is 4.2. When the pH of a solution containing ascorbic acid is lower than its pKa more of the ascorbic acid will be protonated. Protonated ascorbic acid is neutrally charged which may allow it to enter the skin more easily.

It’s important to notice the error bars on the amount of ascorbic acid absorbed at pH 2.0. There is considerable deviation from the mean in the results even though it was only tested on 2 subjects. More test subjects would provide a clearer idea of how much ascorbic acid would penetrate at pH 2 on an average population of pigs.

Statistical differences also weren’t calculated between the data points, for example it’s difficult to tell from the way that the data is presented if there is a change in ascorbic acid content between the control, pH 4.0, 4.5, and 5.0 – even if they look different on the graph. Likewise, it’s difficult to tell if there is an increase in ascorbic acid penetration between pH 3.0 and pH 2.5 – despite the trend with pH 2.0 pushing you towards that inference. It’s likely that there is a statistically significant difference between absorption between pH 3.5 and 3.0, but a larger study would provide us  with more confident answers.

So based on this data, many further studies and brands have assumed that a pH below 3.5 results in considerable more skin penetration of ascorbic acid on humans – despite these results being performed on pigs, and relative low strength of the study. If the reason why ascorbic acid is more easily absorbed into the skin is due to the pKa then this would likely hold true for humans as well.

This assumption is often presented as fact, which is misleading. It also doesn’t take into account other factors present in a cosmetic product, such as penetration enhancers. Encapsulation, surfactants, and solvents could increase (or decrease) the amount of ascorbic acid absorbed into the skin regardless of the product’s pH.

In this experiment, the stratum corneum was removed before measurements of ascorbic acid to test for deep penetration of ascorbic acid. It’s possible that some of the benefits conferred by topical application of ascorbic acid aren’t facilitated by deep penetration, the antioxidant and photoprotective effect of ascorbic acid may still occur when it is present in or on the stratum corneum. Other benefits like reduction of hyperpigmentation and an increase in collagen production are likely dependent on penetration past the stratum corneum.

Unfortunately I haven’t been able to find further studies on humans or otherwise to provide answers to these questions.

“Ascorbic acid serums must be at least 10% to be effective”

After the first experiment of testing 15% ascorbic acid with different pHs, Pinnell and his group tested how concentration of ascorbic acid affects skin penetration. This time they tested 7 ascorbic acid solutions with varying concentrations all at pH 3.2. The concentrations of the rest of the formulation are assumed to be the same as the previous experiment.

The ascorbic acid solutions were applied in the same manner, with a Hill Top Chamber for 24 hours, followed by washing, stripping, and then assessment.

The maximum amount of ascorbic acid penetration was seen when 20% ascorbic acid at pH 3.2 was used.

All concentrations were tested on 3 pigs, and there is quite a bit of deviation from mean between absorption among the 3 pigs tested. This makes it difficult to assess the true difference in absorption between a 10% and 15% ascorbic acid, and a 15% and 20% ascorbic acid.

Absorption also seemed to peak at 20%, the 25% ascorbic acid solution penetrated less than the 20%, and the 30% even less so. The researchers did not explore or hypothesize on why this occured, and I’ve been unable to find an answer in any later research as well.

While 20% ascorbic acid certainly led to the greatest increase in levels of ascorbic acid, the 5% solution still increased ascorbic acid levels in the pig skin by about 6 fold.

It’s very important to remember that the way that this experiment was performed does not mimic the way that ascorbic acid solutions are often applied to the skin. With the Hill Top Chamber, the solvent’s (in this case water) evaporation is reduced – whereas when we apply it to the skin the solvent evaporates. What this means is that the kinetics of ascorbic acid penetration into the skin may not be the same.

For example, if half of the solvent of a 10% ascorbic acid solution evaporates, it is equivalent to a 20% ascorbic acid solution – the total amount of ascorbic acid by mass is the same, but the concentration has changed. This may mean that we could see a different maximum absorption by concentration in an experiment where the solvent was allowed to evaporate the way that it is often applied.

Human clinical trials with “low” ascorbic acid concentrations, 3% ascorbic acid cream and a 5% ascorbic acid cream, were able to show statistically significant improvements on measurements of photodamage and photoageing in their study groups.

Another thing many people hold on to is the concept that their products must be working at “maximum efficiency”, unfortunately this is unrealistic and there’s going to be variations in the amount of ascorbic acid that penetrates your skin with each application – even the amount that you apply to your skin will vary each time. This is why good cosmetic studies are performed over a longer period of time.

For example, if we look at the 20% concentration, the pig skin concentration of ascorbic acid increased to about 1100 pmol of ascorbic acid per mg of pig skin, which is about 0.19 μg ascorbic acid per mg of pig skin. 1.0 mg of a 20% ascorbic acid (w/w) contains about 1135589.37 pmol of ascorbic acid, if that helps give you a sense of the “efficiency”. In these experiments, 200 μL or 0.2 mL solution was used in total for each application, which contains about 227117874.1767 pmol of ascorbic acid if we assume density of the solution (w/w) is 1.

Higher concentrations of ascorbic acid may lead to more irritation (measured by skin redness or erythema), but I haven’t found any studies that looked at this specifically.

Continued in Skin penetration of Ascorbic Acid: Part II

Source: Pinnell, S. R., Yang, H. , Omar, M. , Riviere, N. M., DeBuys, H. V., Walker, L. C., Wang, Y. and Levine, M. (2001), Topical L‐Ascorbic Acid: Percutaneous Absorption Studies. Dermatologic Surgery, 27: 137-142. DOI: 10.1046/j.1524-4725.2001.00264.x

"Skin penetration of Vitamin C (Ascorbic acid): Part I" on KindofStephen.

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