54 ANTI-AGEING
Figure 3: Observation of lawn grass after seven days
extremely pure (>99.9%). The appearance of the product is shown in Figure 4.
Why is the AOH active ingredient so named? Turfgrass occasionally grows in the shape of a ring, exhibiting thicker growth than the surrounding grass; thereafter, mushrooms form within the same circle (Figure 5). This phenomenon is called ‘fairy rings’ and occurs on lawns in parks and golf courses. According to Western legend, fairies make these circles and dance inside them.3, 4, 5 The first scientific paper on ‘fairy rings’ was published in 1675. That paper was introduced to Nature in 1884 and yet, the cause of the phenomenon remained a mystery for a long time.6 Approximately 140 years later, Professor Kawagishi discovered a substance in the mushrooms responsible for the formation of ‘fairy rings’. Since AOH is a type of substance that forms
fairy rings, the AOH active ingredient was named Repista from the scientific name of the mushroom, Lepista sordida. The scientific name is Lepista, with the initial letter L. However, because AOH has effects similar to retinol, such as activating cells and promoting the production of hyaluronic acid, the initial letter R was used for its product name.
Cell-activation effect Since the AOH active ingredient is water-soluble and likely to remain in the stratum corneum or penetrate the epidermis, its effect on normal human epidermal keratinocytes (NHEKs) was examined. The 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay was used to evaluate the effects of the AOH active ingredient on NHEK cell proliferation. NHEK cells were treated with the AOH active
ingredient and incubated at 37°C for 48 hours. As shown in Figure 6, the AOH active ingredient showed a significant increase in cell viability at various concentrations of the AOH active ingredient (0.04–0.3 %).9 When 0.3% of the AOH active ingredient was
tested, NHEK cell proliferation increased by 10% (p< 0.05) compared with the control, implying the AOH active ingredient has a cell-activating effect.
Effect on skin barrier function The increase in cell numbers with the addition of the AOH active ingredient is evidence that it
PERSONAL CARE January 2023
Figure 4: Product appearance of the AOH active ingredient
has some effect on NHEKs. To comprehensively analyze the action of the AOH active ingredient, a DNA microarray was performed using NHEKs targeting approximately 150 genes involved in skin ageing and disease.9 The results showed increased expression
of transglutaminase 1 (TGM1) and involucrin (IVL), both of which promote stratum corneum maturation (Figure 4). IVL and TGM1 are important substrates and enzymes for cornified envelope composition that contribute to the physical and chemical toughness of the stratum corneum. In addition, the expression levels of claudin
1 (CLD1) and desmocollin 1 (DSC1) were also increased (Figure 7). CLD1 and DSC1 are important for tight junction formation and intercellular adhesion. These results suggest that AOH enhances cell-cell interactions and strengthens adhesion, thereby improving the barrier function of the skin. Furthermore, the expression levels of keratin
1 (KLT1) and keratin 10 (KLT10) were significantly increased (Figure 7). KRT1 and KRT10 may contribute to the enhancement of the epidermal cytoskeleton. Along with KRT10, which encodes a fibrin protein and fibrillogenesis pair, KRT1 is involved in cytoskeleton formation in the upper epidermal cells of the skin and plays an important role in maintaining the mechanical strength of these cells. Loss of KRT1 not only causes epidermal detachment but also affects the expression of pro-inflammatory cytokines such as interleukin-18.10
In the epidermis, the expression
of hyaluronan synthase (HAS3), which is involved in hyaluronic acid production, also increased (Figure 7). These results suggest the AOH active ingredient acts on a wide range of epidermal functions such as promoting epidermal turnover, differentiation, maturation, and the metabolism of old skin cells. Subsequent clinical trials revealed that the application of
a lotion containing the AOH active ingredient for eight weeks increased stratum corneum water content and decreased transepidermal water loss, providing clinical evidence to corroborate the DNA microarray results.6
Safety evaluation of the AOH active ingredient AOH, the active ingredient of the AOH active ingredient, is considered safe because it is a natural organic compound that is found in very small amounts in edible plants. However, since AOH is a new cosmetic ingredient, we decided to conduct all the necessary safety tests for the development of cosmetic ingredients, including the mutagenicity test (Ames test), skin irritation test, skin photo-irritation test, and skin sensitization tests, namely the direct peptide reactivity assay (DPRA), human cell line activation test (h-CLAT), and KeratinoSens method, according to the alternative animal testing methods listed in the Organization for Economic Co-operation and Development for Test Guidelines.
Figure 5: Fairy ring formation of mushrooms
TABLE 1: PRODUCT INFORMATION Trade name
Repista Package
Storage conditions INCI name CAS No.
Recommended dosage Usage
1 Kg (liquid) Room temperature
Aza-Oxohypoxanthine, Sodium Hyaluroxide, Water
1403820-30-3 0.3%
Skincare, Makeup, Body care, Hair care
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