A Hyaluronic Acid 101

Hyaluronic acid is one popular ingredient in cosmetics at the moment. Touted to be the most potent humectant available, and an effective anti-aging active, are these claims actually backed by science? In this blog post, I went on a deep dive into all things hyaluronic acid. What is it, what does it do, and what should we know about this ingredient in the cosmetic products we purchase or formulate? Low molecular weight versus high molecular weight? I also broke down this debate and reviewed the available research about that conversation. Warning, this post is a wee bit technical, but if you’re looking to learn more about hyaluronic acid in cosmetics, we’ve got you covered!


What is hyaluronic acid?


Hyalruonic acid (HA) is a linear macromolecular mucopolysaccharide, essentially one huge unbranched molecule chain made up of lots of sugar units. Its chain is made up of alternatingly linked sugar (saccharide) units of glucuronic acid and N-acetylglucosamine. Also known as hyaluronan, it has a high molecular weight of roughly 105–107 Da. Below is what one unit of hyaluronic acid looks like structurally, a hyaluronic acid molecule contains thousands of these bad boys linked over and over again.


Hyaluronic acid (HA) is a naturally occurring polymer that plays a very important role in our biology and the biology of many other plants and animals out there. In fact, it has been evolutionarily conserved, all the way back to simple prokaryotes - a pretty good indicator that hyaluronic acid is one very important molecule. It’s a major element in the connective tissue in vertebrates (i.e. animals with spines) where it functions as scaffolding and support, is responsible for controlling things like tissue hydration, plays a role in cell signalling throughout the body, helps with wound healing and scarring, and is even found in the umbilical cords of babies, suggesting it may also help with tissue regeneration and growth.  Hyaluronic acid is found in most connective tissues in body fluids, from synovial fluid to the humor of the eye.


A brief history of hyaluronic acid


Hyaluronic acid has long received a lot of attention among researchers because of its versatility as a very functional biopolymer. First discovered from the eyes of cows in 1934 by Karl Meyer and John Palmer, who in 1950, solved the chemical structure of the molecule naming it hyaluronic acid (derived from ‘hyalos’, the Greek word for glass plus uronic acid). Initially, hyaluronic acid was isolated in its acid form, but eventually they saw that the molecule behaved more like a salt in biology (sodium hyaluronate).


In 1942, Ender Balazs patented the first application of hyaluronic acid as a substitute for egg white in bakery products. The first biomedical application happened in the late 1950s when it was used for vitreous replacement during eye surgery. For medical applications, Hyaluronic acid was initially isolated from the umbilical cord and shortly later, from rooster combs. Later, hyaluronic acid was isolated from other sources and the investigative interest grew. Today, hyaluronic acid widely used for many applications, from helping in drug delivery, for patients with osteoarthritis, as a dermal filler, the ‘gold standard’ for fillers treating wrinkles, hydrating skin and increasing volume, and most relevant to this post, for cosmetic products.

One of the sources of hyaluronic acid, rooster combs. Hyaluronic acid is also commonly produced via bacteria fermentation today.

Skin aging and hyaluronic acid


Over time and due to things like sun exposure and gravity, your skin gradually goes through an aging process; the skin, soft tissue and skeletal support in the face changes, first resulting in wrinkles. Over time, aging skin causes things like flattening of the epidermal-dermal interphase, a deterioration of the dermis, loss of elasticity, less blood flow from a loss of blood vessels, poorer collagen content, poorer hydration and melanocyte activity, and so on. Visually, on top of wrinkles, this can be seen as skin thinning, hyperpigmentation, and sagging.

 

One of the reasons why there’s so much interest in hyaluronic acid, especially for aging skin, is because it’s heavily linked with this whole process. One very important property of the molecule is that it can hold onto a LOT of moisture; roughly 6L of water in just 1g! As a result, on top of structural support, it’s also a very potent humectant that helps keep moisture in the skin, aiding to its suppleness. One of the reasons youthful skin seems so hydrated is because it contains lots of hyaluronic acid in the dermis. As we age, this content decreases and by the time of adulthood, the content is as low as 5% of this baseline.To add to all of this, as we lose the hyaluronic acid in our skin, this also affects tissue volume and elasticity, also linked with hydration. It’s no wonder why hyaluronic acid is so popular as a dermal filler and in ‘anti-aging’ cosmetic products. Applied topically, hyaluronic acid has been shown to act as a potent humectant, can reduce the appearance of wrinkles, and helps to provide smoothness and softening to the skin. As a formulator, this ingredient, if the budget allows for it, is one that I find I’m often reaching to because I find it, anecdotally, pretty darn effective, even after one application of the product at a percentage as low as 0.5%.

The decrease of epidermal hyaluronic acid content is a hallmark of skin aging on the molecular basis (adapted from Necas et al 6 ). 


High molecular weight vs low molecular weight hyaluronic acid

There’s a bit of confusion and debate about high and low molecular weight hyaluronic acid in cosmetics. While both are used in products, I often see low molecular weight hyaluronic acid being promoted as the superior option to formulators; it’s easier to incorporate into a formula and is better able to penetrate the skin because it’s so small. I’m just going to put it out there that I don’t think  that it’s this simple… from going down a bit of a rabbit hole researching hyaluronic acid, it seems that they might be missing a very important point about hyaluronic acid as a therapeutic agent. Here I’ll try to clarify both sides, but to do that, I think it’s best to start with how the different weights of hyaluronic acid act in nature.

As I mentioned above, hyaluronic acid plays an important role in wound healing and scar formation. In its natural form, hyaluronic acid exists at a very high molecular weight. Upon environmental changes, for example pH changes in response to something like injury, enzymes are stimulated to start to break down hyaluronic acid to lower molecular weight fragments. Different weights of hyaluronic acid have been shown to influence macrophages of the immune system, important actors in wound healing, in different ways. As hyaluronic acid is broken down to lower molecular weight molecules, they seem to stimulate macrophages to have a pro-inflammatory response, one of the first steps to wound healing. In contrast, high molecular weight hyaluronic acid is demonstrated to stimulate an anti-inflammatory response from macrophages. Although these are two extreme affects, macrophages are multifaceted and often exist within a spectrum of these extremes. As a result of these interactions, low molecular weight hyaluronic acid has been found to contribute to scar formation, which was minimized when higher molecular weight hyaluronic acid was found in the wounded area. All this suggests that the different weights of hyaluronic acid play an important role together in wound healing. Where high molecular weight hyaluronic acid favored tissue integrity, fragments (low molecular weight) signalled injury and initiated the inflammatory response. Because hyaluronic acid impacts the immune system in this way, understanding these interactions more is crucial in the context of using it as a therapeutic agent.

 

 

In a 2015 study by Rayahin and colleagues, they highlighted these interactions; different molecular weights of hyaluronic acid was demonstrated to affect macrophage expression differently.Their findings suggested that therapeutically applied hyaluronic acid can alter macrophages in a molecular weight-dependent manner. After examining the impacts of different molecular weights of hyaluronic acid, they concluded that high molecular weight hyaluronic acid (in the megadaltons range) had an anti-inflammatory effect from macrophages while the lower molecular weight (smaller than 12 sugar molecule chains)  hyaluronic acid stimulated an inflammatory response.Their results show that administration of high molecular weight hyaluronic acid is able to promote the resolution phase of wound healing, and removing this polymer is detrimental to the process. In contrast, administration of low molecular weight hyaluronic acid promoted inflammation and reduced its resolution. While this study offers a preliminary understanding of hyaluronic acid, it was simplified in a in-vitro (cell culture) model. Moving forward, in-vivo (i.e. in animal) studies will be important, considering our bodies are a whole lot more complex than a petri dish. Despite this, this study was the first to show that the molecular weight of hyaluronic acid can have these impacts on how macrophages act and should not be ignored, as I see it is most times in marketing about low molecular weight hyaluronic acid.

From this standpoint, it seems pretty straight forward that applying low molecular weight hyaluronic acid is probably not a good idea, and perhaps high molecular weight may be the better option. So why is low molecular weight so popular? One of the challenging characteristics of hyaluronic acid in its natural state for topical products is that these molecules are roughly 3000 nm in diameter. In contase, the intercellular space between your skin cells, which the molecule would have to penetrate to reach the deeper layers of your skin to have a profound impact, only 15 to 50 nm and just 6 to 10 nm at the hyaline membrane. This makes it impossible for high molecular weight hyaluronic acid to reach the deep layers of the dermis (note, this is actually a major challenge for formulators in general, actually getting the active ingredients to penetrate the skin, an idea that’s commonly confused by beauty bloggers. Does your cosmetic products penetrate your skin? Click here for our blog post all about this topic!).

Until recently, the only way to get hyaluronic acid to these deeper layers was by things like injecting across the skin barrier. Scientists at Forlle’d Laboratories in Japan managed to reduce the size of hyaluronic acid molecules all the way down to 5nm, allowing hyaluronic acid to cross the skin barrier when applied topically. After verifying that this very low molecular weight hyaluronic acid could indeed penetrate the skin, scientists conducted a clinical study to verify its effects. Summarizing the results, there was a significant moisturizing effect - after 2,4 and 8 weeks, the skin was significantly more hydrated than untreated skin. In addition, there was an improvement of skin elasticity after two weeks of treatment, which continued throughout the study. This study confirmed that smaller molecular weight hyaluronic acid topically had a greater skin penetration than higher molecular weight hyaluronic acid. Note, while these are usually the findings marketed by ingredient suppliers, this study happened one year before the  2015 study by Rayahin and colleagues above.

 

 

So what should formulators and brands be using? Low molecular weight hyaluronic acid that is better able to penetrate the skin but demonstrably may cause inflammation or skin irritation? Or high molecular weight hyaluronic acid that barely penetrates but doesn’t have the same inflammatory risks as its lower molecular weight counterpart? At this point, I’m going to say that more research is needed to clarify these questions. In my opinion, gathered from drawing ideas from the available research and my own anecdotal experience (i.e. opinion not fact :), my preference is the high molecular weight hyaluronic acid. While it’s really freaking tricky to incorporate into a formula - because of its high molecular weight, it absorbs a boat load of water, and if not incorporated carefully enough or if added at wee bit too high of a concentration, it can quickly turn your formula into a jelly mess that's virtually unworkable if your not careful - time and time again, I’m really impressed with how effective it is at making my skin look and feel great. While it may not be able to penetrate to the deep layers of the skin, it still has a very noticeable effect to the outer layers. In addition, the high molecular form of hyaluronic acid has a better hydrating effect than its low molecular counterpart, and keeping your skin hydrated is an important part of keeping it looking youthful. Regarding the better penetration of the low molecular weight version, I would say, don’t be fooled into thinking that it will be able to replace the lost hyaluronic acid in your dermis. Even with its low molecular weight, other components of a formula may affect it actually being able to penetrate, and even if it does, the effects will be transient (short-term), and for continued effects, it will have to be continually applied. Continual application of an ingredient that demonstrably promotes inflammation? Not something I’m particularly comfortable with, in light of my current understanding of the research.

And that’s a wrap! What do you think? Do you have a preference for high or low molecular weight hyaluronic acid in a formula? Have you used a product with hyaluronic acid that you loved? Join in on the conversation and share your thoughts below!

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Essendoubi M et al. (2016) Human skin penetration of hyaluronic acid of different molecular weights as probed by Raman spectroscopy. Skin res technol. 22(1):55-62.

Fakhari A and Berkland C (2013) Applications and Emerging Trends of Hyaluronic Acid in Tissue Engineering, as a Dermal Filler, and in Osteoarthritis Treatment. Acta Biomater. 9(7):7081-7092.

Jegasothy S et al. (2014) Efficacy of a New Topical Nano-hyaluronic Acid in Humans. J clin aesthet dermatol. 7(3):27-29.

Liu X, et al.(2014) Comparative Studies of Hyaluronan in Marketed Ophthalmic Products. Optom. Vis. Sci. 91:32–38.

Noble P. (2002) Hyaluronan and Its Catabolic Products in Tissue Injury and Repair. Matrix Biol. 21:25–29.

Rayahin et al. (2015) High and low molecular weight hyaluronic acid differentially influence macrophage activation. ACS Biomater Sci Eng. 2015 Jul 13; 1(7): 481–493.

Sokolowska M et al. (2014) Low Molecular Weight Hyaluronan Activates Cytosolic Phospholipase A2alpha and Eicosanoid Production in Monocytes and Macrophages. J. Biol. Chem. 289:4470–4488.

Stern R et al. (2007). The Many Ways to Cleave Hyaluronan. Biotechnol. Adv. 25:537–557.