May 31, 2010

Mandom is the first in the industry to discover that “lowmolecular weight hyaluronic acid improves rough skin”
Confirmation of its deep penetration into the stratum corneum and the promotion ofpolymeric hyaluronic acid production

Mandom Corporation (headquarters: Osaka City, President & CEO: Motonobu Nishimura; hereinafter referred
to as “Mandom”) has discovered that low molecular weight (LMW) hyaluronic acid enhances the recovery time ofbarrier strength in rough skin. Mandom has proposed that LMW hyaluronic acid deeply penetrates the stratum
corneum, acting on cells within the dermis to promote the production of polymeric hyaluronic acid.
The results of this research were presented at the “130th Annual Meeting of the Pharmaceutical Society of Japan,”which was held between March 28 and 30.

<Background of Research>
In the body, hyaluronic acid surrounds cells, collagen, and other nutrients, resulting in skin flexibility. Hyaluronic
acid also possesses excellent moisture retention capacity as 1 g of hyaluronic acid can hold an estimated 6 liters of water.
Furthermore, it is widely used as a moisturizer in cosmetics.
Hyaluronic acid is composed of two repeating sugars, N-acetylglucosamine and glucuronic acid (Figure 1).
Polymeric hyaluronic acid contains many of these repeating subunits while low molecular weight hyaluronic acids
contains a small number of such repeats.
It has been reported that LMW hyaluronic acid has higher penetrability into the stratum corneum and better
sustained moisturizing effect than polymeric hyaluronic acid. Research on its effect when applied to the skin and its
impact on hyaluronic acid production have not been reported. Therefore, Mandom has been advancing research aimed toelucidate the effect of LMW hyaluronic acid on the skin.
Owing to these investigations, Mandom has become the first in the industry to discover the improved effectexhibited by LMW hyaluronic acid on rough skin. We propose that LMW hyaluronic acid penetrates deep into thestratum corneum, acts on cells in the dermis and promotes the production of polymeric hyaluronic acid.

Contact

mandom corp.
Public Relations Div.
mail: press@mandom.co.jp

URL: https://www.mandom.co.jp/en/

Please contact us in Japanese or English.

-1-Stratum corneumStratum granulosum
Stratum spinosum

Epidermis

Stratum basale
N-acetylglucosamine
Glucuronic acid

DermisFigure 1. Structure of hyaluronic acid
Figure 2. Structure of skin

<Improvement effect of low molecular weight hyaluronic acid on rough skin>
To evaluate moisture barrier strength, we applied LMW hyaluronic acid solution, polymeric hyaluronic acid solution,
and water twice per day (morning and night; three days) to the arms of subjects (age range, 20-40+; having provided
informed consent) where rough skin (*2) was induced by stripping tape (*1).
In areas where only LMW hyaluronic acid was applied, moisture barrier strength recovered 2-fold compared to areas
where water was applied (Figure 3). Therefore, LMW hyaluronic acid contributed to the recovery of moisture barrierstrength.
*1 Tape stripping: A method using cellophane tape to remove stratum corneum and reduce barrier strength.

Low  Moisture barrier strength High*2 Rough skin: Skin with decreased moisture barrier strength.
Water
Polymeric hyaluronic acid
LMW hyaluronic acidTwo-fold

Number of days elapsed

Figure 3. Recovery of barrier function by low molecular weight hyaluronic acid

We performed several experiments to elucidate the mechanism driving the “Improved effect on rough skin”
demonstrated by low molecular weight hyaluronic acid.
<Elucidating the mechanism (1): Low molecular weight hyaluronic acid penetrates deep into the stratum
corneum>
We performed tape stripping on model skin, induced rough skin, and demonstrated the penetrability of low
molecular weight hyaluronic acid. Not only did LMW hyaluronic acid reach the stratum corneum and epidermis, butslight transport to the dermis (Figure 4) was observed. In a similar experiment, polymeric hyaluronic acid did not
penetrate the stratum corneum, the uppermost layer of the skin.

-2-Epidermis

DermisFigure 4. Penetration of LMW hyaluronic acid in the rough skin model
Photo of skin after LMW hyaluronic acid wasapplied
Sample has been stained to distinguishbetween epidermis and dermis

Photo visualizing LMW hyaluronic acidGreen area indicates LMW hyaluronic acid.

<Elucidating the mechanism (2): Acting on cells in dermis and promoting hyaluronic acid production>

Gene expression levels

Fibroblasts present in the dermis are representative of cells that synthesize hyaluronic acid. Addition of LMW
hyaluronic acid solution to a culture solution of these fibroblasts increased the gene expression of HAS1, the polymeric
hyaluronic acid synthase, and TGF-β1, the controller of HAS1 (Figure 5). Although HAS1, HAS2 and HAS3 areenzymes known to synthesize hyaluronic acid, HAS1 is a known synthesizer of hyaluronic acid.

Not added
LMW hyaluronic acid

Figure 5. Impact of hyaluronic acid on gene expressionHAS1, HAS2, HAS3: Hyaluronic acid synthases
TGF-β1: A gene that increases HAS1 expression

By adding LMW hyaluronic acid, we confirmed that the production of polymeric hyaluronic acid increases infibroblasts. Therefore, we now understand that the addition of LMW hyaluronic acid promotes the production ofhyaluronic acid with higher molecular weight (Figure 6).
These results suggest that LMW hyaluronic acid increases the activity of hyaluronic acid synthases and enhances theproduction of polymeric hyaluronic acid.

-3-Large

Size of molecule

Small

LMW hyaluronic acid added
Low  Hyaluronic acid level  HighLow  Hyaluronic acid level  HighNot added

Large

Size of molecule

Small

Figure 6. Promotion of polymeric hyaluronic acid production by LMW hyaluronic acid
Production level and molecular weight ofhyaluronic added with LMW HA addition

Production level and molecular weight of hyaluronic acid withLMW HA additionindicates the level of polymeric hyaluronic acid production; an increase was observed with LMW
hyaluronic acid addition.

*The above results suggest that LMW hyaluronic acid contributes to the recovery of barrier function in the skin bypenetrating the rough skin and promoting the synthesis of polymeric hyaluronic acid (i.e., LMW hyaluronic acid is“Hyaluronic acid that increases the hyaluronic acid content in the skin”, which penetrates rough skin and produces
polymeric hyaluronic acid in the skin). We therefore expect LMW hyaluronic acid to increase polymeric hyaluronic
acid in the skin in an effort to recover barrier function, and elevate the moisture retention capacity of the skin, toprovide subsequent flexibility in the skin.

Mandom will continue its strive to create products to achieve “ideal skin” that will allow people to feel
comfortable; this will be performed by studying the skin’s ingredients to achieve better skin.
We also plan to apply the findings of this research to our company’s cosmetic products for women, namely thewarm-moisturizing skincare series, Barrier Repair.

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