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ReGenistem® (Glycyrrhiza – liquorice extract)

ReGenistem® (Glycyrrhiza – liquorice extract)

Liquorice root extract is a common ingredient found in many skin lightening cosmeceuticals and is also used in the treatment of a wide variety of diseases even outside the scope of dermatology due to its anti-oxidant, anti-inflammatory, antiviral, antimicrobial and anticarcinogenic properties.

Liquorice extract is obtained from the root of the glycyrrhia glabra linnera which is native to the Mediterranean region with striated stalks of about one metre.

Liquorice

Liquorice root

The primary antioxidant and anti-inflammatory compounds found in liquorice root are the glycosides – glycyrrhizin and glycyrrizinic acid, flavonoids and saponins.

Glabridin (a polyphenolic flavonoid) is one of the main components of liquorice extract. Glabridin works by three main mechanisms:

  • Inhibition of tyrosinase activity leading to reduced melanin production
  • Inhibition of superoxide anion production. Superoxide anion is a reactive oxygen species (ROS). These are harmful oxygen molecules generated by UV rays and pollution. ROS attacks and reacts with stable molecules within skin cells causing irreversible damage to cells which trigger wrinkles and lessening the skins natural ability to repair itself. This is known as oxidative stress. Glabridin thereby inhibits irreversible skin cell damage by preventing free radical production.
  • Inhibition of cyclooxygenase activity thereby reducing and preventing inflammation.
  • Up-regulation of proteins responsible for the synthesis of collagen type 1 and collagen type 3. The up-regulation of these collagen producing proteins leads to anti-aging properties such as improving skin elasticity and firming up dull, lagging skin.

Studies have shown that glabridin prevents ultraviolet B induced pigmentation, prevents damage to skin cells and exerts anti-inflammatory effects by inhibiting superoxide anion and cyclooxygenase activity.

These mechanisms contribute to skin lightening, anti-inflammatory and anti-aging properties alongside soothing and regenerative characteristics.

Other active agents of liquorice extract include liquitrin and isoliquertin. Liquitrin is a glycoside containing flavonoid that induces skin lightening by dispersing melanin. In a study conducted in 20 Egyptian women showed that topical Liquitrin (1g/day) for four weeks was both safe and effective in the treatment of melasma.

In a double blind placebo controlled study, 2.5% of liquorice extract was evaluated as treatment of melasma in 93 females. Comparison between formulation treated group and placebo treated group indicated significant differences for overall response to treatment. The improvement in active treatment group was 93% while the corresponding value in placebo treated group was 4%. This clinical trial indicated that 2.5% liquorice extract was effective as treatment for melasma in a 28 days course with minimal side effects.

ComplexiLight research team have elected ReGenistem® as source for liquorice extract. ReGenistem® is a sustainable skin brightening active which is produced using a patented process that elicits secondary metabolites from meristem cultures obtained from liquorice differentiating ReGenistem® from other liquorice based ingredients because the technology utilizes calli that takes no longer than a month to grow. Utilization of the meristematic culture technology reduces the dependency of liquorice plant harvests, providing a more uniform, reliable skincare active.

In-vitro Efficacy Studies
ReGeniStem® Brightening was evaluated in a tanned epidermal melanin assay, comparing its ability to inhibit melanogensis against standard ingredients commonly found in skin lightening products. These skin-lightening ingredients include hydroquinone, kojic acid, and arbutin. A water-based liqourice extract was also included in the assay to compare to ReGeniStem® Brightening.

ReGeniStem® Brightening Melanin Assay

Regenistem

Melanin assay comparing ReGeniStem® Brightening to common skin brightening ingredients. ReGeniStem® Brightening showed a statistically significant change in melanin compared to untreated skin

The results from the tanned epidermal melanin assay showed ReGeniStem® Brightening outperformed all three of the common skin lightening ingredients and the water based liquorice extract.

ReGeniStem® Brightening at 0.01% and 0.02% showed a statistically significance change in melanin compared to untreated skin.

An additional assay was performed to measure tyrosinase inhibition. The tyrosinase enzyme is a key contributor to melanin synthesis and the mechanism of focus. Inhibition of tyrosinanse leads to a lighter, brighter skin tone. ReGeniStem® Brightening was compared to two common actives known for their ability to inhibit tyrosinase activity: kojic acid and arbutin. The results show ReGeniStem® Brightening performed comparably to kojic acid and arbutin in inhibiting tyrosinase activity.

Tyrosinase

Tyrosinase inhibition study of ReGeniStem® Brightening compared to common skin lightening ingredients.

The results from the tyrosinase inhibition study, in conjunction with the results from the tanned epidermal melanin assay, provide firm justification to further evaluate ReGeniStem® Brightening in an in-vivo panel study.

In-vivo Skin Brightening Study
A preliminary 5-person, 22-day skin brightening study was initiated to evaluate the efficacy of ReGeniStem® Brightening in reducing the colour of skin. Skin was irradiated with a 2 minimal erythema dose (MED) on the back of panellists. ReGeniStem® Brightening was incorporated into a lotion base at 0.2%, 0.4%, 1%, and 2% use levels. Kojic acid, the positive control of the study, was also incorporated into a lotion base at 0.5% and 1% use levels. The base lotion served as the placebo in the study. The lotions were applied daily on the panellist back over the 22 day study, with measurements made on Day 8 and Day 22. The treated skin was measured with a Minolta chromameter and measured for the L* values of skin colour. A greater change in L* values indicates lighter skin tone.

In-viva Skin Brightening Study

In-vivo skin brightening study with ReGeniStem® Brightening compared to kojic acid, a common skin-lightening ingredient

The results show ReGeniStem® Brightening at the lower concentrations (0.2%, 0.4%) performed comparably to kojic acid at 0.5%. ReGeniStem® Brightening at the higher concentrations (1%, 2%) performed comparably to kojic acid at 1%. Based on the results, ReGeniStem® Brightening has shown the ability to brighten the colour of skin and help even skin tone. Additionally, the results justify evaluating ReGeniStem® Brightening at a 1% and 2% use level for a larger, in-vivo study.

In-vivo Skin Brightening Study

An 84-person, 60-day study was initiated to evaluate the efficacy of ReGeniStem® Brightening in improving skin brightening. The 84-panelist pool was divided into three 28-person groups, with each group applying one of three products on their entire face. The products were a lotion containing 1% ReGeniStem® Brightening, a lotion containing 2% ReGeniStem® Brightening, and a control lotion (the base lotion). Each group contained an evenly distributed population of gender and ethnicities: 25% African-American, 25% Asian, 25% Caucasian, and 25% Hispanic men and woman. Panellists applied the lotion on their face twice daily, once in the morning and once before bed. Measurements were made on Day 30 and Day 60 of the study with the Canfield VISIA CR system using visible light and analysed using the Vaestro Image Analysis Toolkit. The skin brightening study was initiated in late spring and concluded in the middle of summer.

ReGeniStem Day 0 - Day 60

Figure 1

A panellist treated with 1% ReGeniStem® Brightening with a 0.55% improvement in skin brightening over baseline via Vaestro Image Analysis Toolkit

The results from the 84-person in-vivo study showed ReGeniStem® Brightening improved skin brightening at use levels as low as 1%. The panelist from figure 1 was measured to have a 0.55% improvement in skin brightening, a value which, when initially read, would indicate very little improvement. However, when correlating the measurement to the images generated from the study, a 0.55% measurement does show a visible improvement in overall skin tone.

ReGeniStem Day 0 - Day 60 fig 2

Figure 2

A panellist treated with 1% ReGeniStem® Brightening with a 1.10% improvement in skin brightening over baseline via Vaestro Image Analysis Tookit

ReGeniStem Day 0 - Day 60 fig 3

Figure 3

Panellist treated with 1% ReGeniStem® Brightening with a 2.00% improvement in skin brightening over baseline via Vaestro Image Analysis Toolkit

The panellist from figure 3 was measured to have a 2.00% improvement in skin brightening. The effects of ReGeniStem® Brightening are prominent, with the active improving the overall skin tone with lighter and more balanced skin. In addition, ReGeniStem® Brightening helped to target the large blemish on their cheek by lightening the pigmentation and helping to blend it in with their natural skin tone.

In-vivo Skin Brighteness Study with ReGeniStem® Brightening

Change in Skin Brightness

Mean improvement in skin brightening with ReGeniStem® Brightening. ReGeniStem® Brightening showed a statistically significant improvement in skin brightening compared to the control.

Across the entire in-vivo study, ReGeniStem® Brightening showed a statistically significant improvement in skin brightness of the panellist evaluated, with a mean improvement in skin brightening of 1.00% when utilizing ReGeniStem® Brightening at a 1% use level. In addition, we see a dose dependent response in improving skin brightness, with a 1.20% improvement in skin brightening when utilizing ReGeniStem® Brightening at a 2% use level.

Additional In-vitro Data

The images generated from the in-vivo study showed an improvement in overall skin tone, while targeting imperfections and hyperpigmentation very effectively. In addition, there are signs there are anti-aging benefits such as helping to reduce the appearance of fine lines and wrinkles. A protein assay was performed on ReGeniStem®Brightening to discover any additional benefits which the novel active may provide the skin.

 

In-vitro Protein Assay of ReGeniStem® Brightening: Type-III Collagen

In-vivtro Protein Assay

Results of in-vitro protein assay of ReGeniStem® Brightening for Type-III Collagen. ReGeni- Stem® Brightening at 0.005% and 0.01% showed a statistical significant upregulation of Type-III Collagen compared to untreated skin

In-vivtro Protein Assay 2

Results of in-vitro protein assay of ReGeniStem® Brightening for Type-I Collagen. All ingredients evaluated showed a statistical significant upregulation of Type-I Collagen compared to untreated skin. TGF-Beta is included in the study as a positive control for Type-I collagen synthesis

The results showed ReGeniStem® Brightening upregulated proteins responsible for the synthesis of Collagen Type-I and Collagen Type-III. The upregulation of these collagen producing proteins could lead to anti- aging properties, such as improving skin elasticity and firming up dull, lagging skin.

Additional Clinical Images

In-vivo Skin Brightening Study – 2% ReGeniStem® Brightening:

ReGeniStem Day 0 - Day 60 fig 4

Figure 4

Panellist treated with 2% ReGeniStem® Brightening with a 0.22% improvement in skin brightening over baseline via Vaestro Image Analysis Toolkit

ReGeniStem Day 0 - Day 60 fig 5

Figure 5

Panellist treated with 2% ReGeniStem® Brightening with a 1.57% improvement in skin brightening over baseline via Vaestro Image Analysis Toolkit.

Figure 6

ReGeniStem Day 0 - Day 60 fig 6

Figure 6

Panellist treated with 2% ReGeniStem® Brightening with a 3.40% improvement in skin brightening over baseline via Vaestro Image Analysis Toolkit

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