Tomorrow s Vision Today! ABS Viola Tricolor Extract Efficacy Data Code: 10346PF INCI Name: Hydrolyzed Viola Tricolor Extract CAS #: 9015-54-7 EINECS #: 310-296-6 Type of Study Hydration Capacity Results The results indicate that ABS Viola Tricolor Extract PF may be used to increase the level of epidermal hydration when compared to the placebo over the course of 24 hours. Compared to the placebo, ABS Viola Tricolor Extract PF may increase epidermal by more than 10%. Aquaporin-3 Synthesis Increase in CD44 Receptor Sites The results indicate that ABS Viola Tricolor Extract PF is capable of significantly increasing the production of aquaporin-3 by human keratinocytes when compared to the biological control and EGF. Therefore it may be possible to increase osmosis between the dermis to the stratum corneum to preserve moisture. A 2% concentration of ABS Viola Tricolor Extract PF was observed to increase the proliferation of CD44 receptors by 43% compared to the negative control and by 7% more than the positive control, EGF. The use of ABS Viola Tricolor Extract PF as a means to increase the prevalence of the CD44 receptor appears to be dose dependent. Anti-Wrinkle activity Assay The results indicate that using 2% ABS Viola Tricolor Extract PF twice daily for 28 days is effective for decreasing the number of wrinkles presents in the outer eye area while also minimizing the total wrinkle surface and wrinkle length. One may therefore use ABS Viola Tricolor Extract PF in anti-wrinkle products to decrease the overall appearance in wrinkles. Version#2/12-9-15/ Form#82 info@activeconceptsllc.com +1 (704)- 276-7100 Fax: +1 (704)- 276-7101
Tomorrow s Vision Today! Longterm Hydration Capacity Assay Production of Hyaluronan Synthase The results indicate that ABS Viola Tricolor Extract PF may be used to effectively hydrate the epidermis for an extended length of time. Compared to the placebo, ABS Viola Tricolor Extract PF is capable of hydrating the skin by more than 10% after a period of 14 days, and almost 20% after a period of 28 days. The results indicate that ABS Viola Tricolor Extract PF is capable of increasing mrna expression for hyaluronan synthase-2, and the mrna expression is increased by almost 60%. Thereby increasing the synthesis of hyaluronic acid to increase moisture retention within the skin. Version#2/12-9-15/ Form#82 info@activeconceptsllc.com +1 (704)- 276-7100 Fax: +1 (704)- 276-7101
Hydration Capacity Assay info@activeconceptsllc.com +1 (704)-276-7100 Fax: +1 (704)-276-7101 Abstract: The hydration activity of ABS Viola Tricolor Extract PF was determined using 2% within a base lotion. Measurements of epidermal hydration were taken with a Corneometer after a single application to the calf regions on the subject s legs. Materials and Methods: The assay utilized 25 female subjects between the ages of 25 and 49 years of age with an average of 33 years. Inclusion in the assay was based on epidermal hydration levels of subjects being less than or equal to 65 is the numerical quantification for dry skin. A CM 820 Corneometer (Courage & Khazaka, Germany) was used in the assay. The instrument consisted of a probe that quantitatively measured the electric capacitance of the skin based on the ability of intercellular water levels of the stratum corneum to conduct electrons. The following formulation was used in the study: Base Lotion Test Formula: TRADE NAME INCI SUPPLIER Percent (%) Lanol 1688 Cetearyl octanoate Seppic 5.0 Isopropyl Palmitate Isopropyl Palmitate - 5.0 ABS Viola Tricolor Extract Aqua & Hydrolyzed Viola Active Concepts LLC 2.0 PF Tricolor Extract & Leuconostoc/Radish Root Ferment Filtrate Sepigel 305 Polyacrylamide/C 13-14 Seppic 2.0 Isoparaffin/Lauret-7 Montanox 60 Polysorbate 60 Seppic 1.7 Sorbitan sesquioleate Sorbitan sesquioleate - 1.3 Leucidal Liquid Leuconostoc/Radish Root Active Micro 4.0 Ferment Filtrate Technologies LLC Deionized Water Aqua - q.s. 100 Two zones (5cm 2 ) were measured on the calves and the hydration of the skin in each zone was determined based on 3 measurements per zone. 2µl/cm 2 of the test formula was randomly applied and massaged into the area using a gloved finger. Each zone was treated with the test formula containing 2% ABS Viola Tricolor Extract PF or the placebo (Cetaphil Moisturizing Lotion). The hydration of the zones was measured after 24 hours under conditions which included a room temperature of 22 C ± 2 C with a relative humidity between 40% and 60%. Results were obtained based on a ratio between the variations in hydration levels compared to the placebo. use of its 1chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the infringement of any patent. We make no warranty of any kind, expressed or implied, other than that the material conforms to the applicable standard specification. Page 1 of 2 Version#1/06-24-15
Hydration Capacity Assay info@activeconceptsllc.com +1 (704)-276-7100 Fax: +1 (704)-276-7101 Results: Treatment Day 14 Day 28 2% ABS Viola Tricolor Extract PF 15 20 (in Base Lotion) Placebo (Cetaphil Moisturizing Lotion). 3 3 Figure 1: Hydration Results Comparing 2% ABS Viola Tricolor Extract PF in Base Lotion and the Placebo. Discussion: The results indicate that ABS Viola Tricolor Extract PF may be used to increase the level of epidermal hydration when compared to the placebo over the course of 24 hours. Compared to the placebo, ABS Viola Tricolor Extract PF may increase epidermal by more than 10%. use of its 2chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the infringement of any patent. We make no warranty of any kind, expressed or implied, other than that the material conforms to the applicable standard specification. Page 2 of 2 Version#1/06-24-15
Aquaporin-3 Synthesis info@activeco nceptsllc.com Phone: +1-704 -276-710 0 Fax: +1-704-276-7101 Abstract: Aquaporin-3 is a synthesized by human keratinocytes as a means to enhance the moisture of our skin by increasing osmosis between the dermis and the stratum corneum. Materials and Methods: The production of aquaporin-3 was observed via a comparison between the biological control, 30 ng/ml of EGF and ABS Viola Tricolor Extract PF at concentrations of 0.25% and 0.50% respectively. Human Keratinocytes were cultured in KSFM medium and incubated for a period of 24 hours at a constant temperature of 37 C with CO 2 levels limited to 5%. Image analysis was performed with BIO Profil software (Bio1D, Vilber-Lourmat, France). The visible bands were then quantified using densitometry. Results: Treatment Level of Aquaporin-3 (%) Control 104 EGF (30 ng/ml) 150 0.25% ABS Viola Tricolor Extract PF 137 0.5% ABS Viola Tricolor Extract PF 156 Figure 1: Aquaporin-3 production comparing ABS Viola Tricolor Extract PF, the untreated control and EGF. Discussion: The results indicate that ABS Viola Tricolor Extract PF is capable of significantly increasing the production of aquaporin-3 by human keratinocytes when compared to the biological control and EGF. Therefore it may be possible to increase osmosis between the dermis to the stratum corneum to preserve moisture. 1 use of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 1 of 1 Version #1/06-23-15
Increase in CD44 Receptor Sites info@activeco nceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Abstract: CD44 is the cellular receptor with specificity for Hyaluronic Acid. The effects of ABS Viola Tricolor Extract PF on the increase in the CD44 receptor were determined using the flow cytometry method on human keratinocytes. Materials and Methods: Immunolabeling of human keratinocytes was performed after a 72-hour incubation period. The keratinocytes were then incubated for an additional 45 minutes at 4 C with the anti-cd44 immunoglobulin. A second antibody with a luminescent flag (Alexa Fluor-488, Interchim, Franc, Cat. No. A11017) was used to identify the immunolabeled cells. The cells were then sorted using a FA Vantage cell sorter (Becton Dickson, USA). They were passed in front of the argon laser (emission 488nm, adsorption 430 nm) at the rate of 500 cells/second. A quantification of fluorescence was performed on 10 cells. Results: Treatment CD44 Receptor Increase (%) Control 100 EGF 136 0.5% ABS Viola Tricolor Extract PF 104 1% ABS Viola Tricolor Extract PF 132 2% ABS Viola Tricolor Extract PF 143 Figure 1: Results of the increase in CD44 following treatment with the test materials. use 1of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 1 of 2 Version #1/06-24-15
Increase in CD44 Receptor Sites info@activeco nceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Discussion: A 2% concentration of ABS Viola Tricolor Extract PF was observed to increase the proliferation of CD44 receptors by 43% compared to the negative control and by 7% more than the positive control, EGF. The use of ABS Viola Tricolor Extract PF as a means to increase the prevalence of the CD44 receptor appears to be dose dependent. use 2of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 2 of 2 Version #1/06-24-15
In-vivo Anti-wrinkle Activity info@activeco nceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Abstract: The anti-wrinkle effects of ABS Viola Tricolor Extract PF were examined using a panel of 25 female subjects. Silicone mold replicas of test subjects outer eye area were used to determine a decrease in total wrinkle surface, total wrinkle length and total number of wrinkles. Materials and Methods: Silicone Molds were made of the test subject s outer eye area before the trial began and again after 28 days of treatment. A lotion was formulated which contained 2% ABS Viola Tricolor Extract PF, subjects were asked to apply the lotion to the outer eye area of their faces twice daily for a period of 28 days. The anti-wrinkle effects of ABS Viola Tricolor Extract PF were determined using a profilometer equipped with an image analyzer. Measurements were taken at a room temperature of 22 2 C with a relative humidity between 40 and 60% Base Lotion Test Formula: Trade Name INCI Supplier Percent (%) Lanol 99 Isononyl isononanoate Seppic 7.0 Montanov 202 Arachidyl alcohol/behenyl Seppic 5.0 Alcohol/Arachidyl glucoside Lanol 1688 Cetearyl Octanoate Seppic 4.0 Propylene glycol Propylene Glycol 4.0 ABS Viola Tricolor Extract PF Aqua & Hydrolyzed Viola Tricolor Active Concepts LLC 2.0 Extract & Leuconostoc/Radish Root Ferment Filtrate Simulsol 165 PEG-100 Stearate Glyceryl Stearate Seppic 2.0 Glycerol Glycerin - 2.0 Glyceryl Stearate Glyceryl Stearate - 1.0 Cetyl Alcohol Cetyl Alcohol Lambert Riviere 1.0 Isohexadecane Isohexadecane Lambert Riviere 1.0 Leucidal Liquid Leuconostoc/Radish Root Active Micro 4.0 Ferment Filtrate Technologies LLC Deionized Water Aqua - q.s. 100 use 1of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 1 of 2 Version #1/06-24-15
In-vivo Anti-wrinkle Activity info@activeco nceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Results: Characteristic Percent (%) Deacrease Number of wrinkles 8 Total wrinkle surface 15 Total length 15 Figure 1: Mean changes in wrinkle characteristics using 2% ABS Viola Tricolor Extract PF. Discussion: The results indicate that using 2% ABS Viola Tricolor Extract PF twice daily for 28 days is effective for decreasing the number of wrinkles presents in the outer eye area while also minimizing the total wrinkle surface and wrinkle length. One may therefore use ABS Viola Tricolor Extract PF in anti-wrinkle products to decrease the overall appearance in wrinkles. use 2of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 2 of 2 Version #1/06-24-15
Long Term Hydration Capacity Assay info@activeco n ceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Abstract: The effects of ABS Viola Tricolor Extract PF on long-term effects epidermal hydration were studied using 2% in an emulsified lotion. The effects were measured with a Corneometer after the lotion was applied twice daily for 28 days. Test subjects consisted of 20 females between the ages of 37 and 62 years of age. The test site was a 5cm 2 region on the calves of the subjects. Materials and Methods: The assay utilized 25 female subjects between the ages of 25 and 49 years of age with an average of 33 years. Inclusion in the assay was based on epidermal hydration levels of subjects being less than or equal to 65 is the numerical quantification for dry skin. A CM 820 Corneomater (Courage & Khazaka, Germany) was used in the assay. The instrument consisted of a probe that quantitatively measured the electric capacitance of the skin based on the ability of intercellular water levels of the stratum corneum to conduct electrons. The following formulation was used in the study: Base Lotion Test Formula: TRADE NAME INCI SUPPLIER Percent (%) Lanol 1688 Cetearyl octanoate Seppic 5.0 Isopropyl Palmitate Isopropyl Palmitate - 5.0 ABS Viola Tricolor Extract PF Aqua & Hydrolyzed Active Concepts LLC 2.0 Viola Tricolor Extract & Leuconostoc/Radish Root Ferment Filtrate Sepigel 305 Polyacrylamide/C 13-14 Seppic 2.0 Isoparaffin/Lauret-7 Montanox 60 Polysorbate 60 Seppic 1.7 Sorbitan sesquioleate Sorbitan sesquioleate - 1.3 Leucidal Liquid Leuconostoc/Radish Active Micro 4.0 Root Ferment Filtrate Technologies LLC Deionized Water Aqua - q.s. 100 Two zones (5cm 2 ) were measured on the calves and the hydration of the skin in each zone was determined based on 3 measurements per zone. 2 l/cm 2 of test formula was randomly applied and massaged into the area using a gloved finger. Each zone was treated with the test formula containing 2% ABS Viola Tricolor Extract PF or the placebo (Cetaphil Moisturizing Lotion). The hydration of the zones was measured after 14 and 28 days under conditions which included a room temperature of 22 C 2 C with a relative humidity between 40 and 60%. Results were obtained based on a ratio between the variations in hydration levels compared to the placebo. 1 use of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 1 of 2 Version #1/06-24-15
Long Term Hydration Capacity Assay info@activeco n ceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Results: Treatment Percent Increase in Hydration Day 14 Percent Increase in Hydration Day 28 2% ABS Viola Tricolor Extract PF 15 20 (in Base Lotion) Placebo (Cetaphil Moisturizing Lotion) 3 3 Figure 1: Skin hydration results when comparing 2% ABS Viola Tricolor Extract PF in base lotion to the Placebo. Discussion: The results indicate that ABS Viola Tricolor Extract PF may be used to effectively hydrate the epidermis for an extended length of time. Compared to the placebo, ABS Viola Tricolor Extract PF is capable of hydrating the skin by more than 10% after a period of 14 days, and almost 20% after a period of 28 days. 2 use of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 2 of 2 Version #1/06-24-15
Production of Hyaluronan Synthase-2 info@activeco nceptsllc.com Phone: +1-704 -276-7100 Fax: +1-704-276-7101 Abstract: Hyaluronan is a protease vital for the synthesis of hyaluronic acid. RT-PCR of human keratinocytes was used to determine the effect of ABS Viola Tricolor Extract PF on the expression of mrna coding for Hyaluronan Synthase-2 (HAS2). Materials and Methods: Human Keratinocytes were treated with 1% ABS Viola Tricolor Extract PF for a period of 24 hours at the constant temperature of 37 C with CO 2 levels limited to 5%, compared with untreated human keratinocytes (control) kept for a period of 24 hours at the constant temperature of 37 C with CO 2 levels limited to 5%. The total RNA was then extracted from the cells and reverse transcriptase was used to obtain the DNA sequence analyzed by PCR. -actin mrna was used as the internal standard. Band intensity was quantified by image analysis using the Bio-profil system ad Bio-1D software. Results are expressed as a ration between the band intensity for the analyzed gene and -actin. Results: Treatments Percent of mrna Untreated Control 100 1% ABS Viola Tricolor Extract PF 152 Figure 1: Increase in MRNA comparing ABS Viola Tricolor and Placebo Discussion: The results indicate that ABS Viola Tricolor Extract PF is capable of increasing mrna expression for hyaluronan synthase-2, and the mrna expression is increased by almost 60%. Thereby increasing the synthesis of hyaluronic acid to increase moisture retention within the skin. use 1of its chemical products since the conditions of use are beyond our control. Statements concerning the possible use of our products are not intended as recommendations to use our products in the Page 1 of 1 Version #1/06-24-12/Form#5