NAB Rhodiola Extract Your Partner in the Fight Against Urban Stress

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Personal Care NAB Rhodiola Extract Your Partner in the Fight Against Urban Stress Key Product Attributes Adaptogenic plant extract that helps protect skin against the stresses of modern urban life Enhances the resistance of skin cells to UV-radiation, reactive oxygen species, and pollution Protects key skin components against the harmful effects of free radicals

Introduction Modern life comes with a variety of stress factors: UV radiation, pollution, sleep deprivation, poor diet, and emotional stress, to name but a few. What all these factors have in common is that they, through different physical, chemical and biological mechanisms, can lead to the formation of reactive oxygen species (ROS). 1,2 These ROS can damage proteins and lipids and have been implicated in several diseases as well as the general process of aging. 2 As our first layer of protection against the environment our skin is particularly sensitive to this process of oxidative stress. Our skin is constantly under attack from the outside, but also from toxins generated within, often, as the result of unhealthy lifestyles. Making matters worse, the skin s natural defenses, such as antioxidant production and cell damage repair processes, decline as we get older, further accelerating the aging of the skin. 2 What our skin needs to mitigate the visible effects of aging due to the stresses of modern life is protection against both the causes and effects of oxidative stress. This is exactly what NAB Rhodiola Extract offers. In Vitro Auto Climate Control for the Skin Human cells can only survive within a narrow range of temperatures, with 37 o C representing and optimal temperature. Deviations from this temperature can induce the expression of specialized proteins which help the cells to survive. 4 To evaluate the ability of NAB Rhodiola to protect against thermal stress, a method using cultured human keratinocytes exposed to hot or cold temperatures was performed. Keratinocyte Viability with Thermal Stress Cell Viability (%) 12 1 8 6 4 2 NAB Rhodiola Extract Untreated (No Stress) Untreated (Stress) NAB Rhodiola 1 mg/ml NAB Rhodiola 2 mg/ml NAB Rhodiola Extract is an extract from the root Rhodiola rosea, which originates from the Arctic regions of Eastern Siberia and the Tibetan mountains. Studies demonstrated that Rhodiola is an adaptogenic plant, which means it has developed natural defense mechanisms, such as increased production of powerful antioxidants, against the harmful effects of extreme environmental conditions. 3 Data shows that NAB Rhodiola Extract enhances the skin s resistance to stress and thus promotes skin cell vitality. Specifically, NAB Rhodiola Extract: Boosts the activity of endogenous antioxidants in skin cells Protects the skin against harmful effects of UV radiation Helps protect skin lipids against oxidative stress Helps protect against degradation of squalene on skin exposed to environmental pollutants Reduces the expression of inflammatory cytokines Can help protect skin against exposure to extreme temperatures Heat Stress Figure 1 shows that keratinocytes treated with 1% of NAB Rhodiola Extract provides a protective effect against both hot (15-2 min utes of hyperthermia @ 48 C) and cold (15-17 hours of hypothermia @ 4 C) thermal stress as indicated by the increase in viability of the cells. NAB Rhodiola Extract 2% provides a protective effect against hot stress. Denotes (p<.5) difference from the Untreated (with stress) sample. Inflammaging Cold Stress Recent scientific studies have provided new insights into the important role of low-grade inflammation in skin aging. Specifically, pro-inflammatory cytokines, such as Interleukin-8 (IL-8) and Interleukin-6 (IL-6) can cause skin damage by activating dermal fibroblasts to secrete Matrix metalloproteinases (MMPs). These MMPs are well known for their role in photo-aging, through their ability to degrade extra cellular matrix (ECM) proteins such as collagens and elastin, which can ultimately lead to the formation of wrinkles 5,6 In vitro data presented in Fig. 2 demonstrate the effect of NAB Rhodiola on reducing the expression of inflammatory cytokines in human keratinocytes challenged with Phorbol Myristate Acetate (PMA), a well known activator of inflammation. 2 Personal Care NAB Rhodiola Extract

IL-6 pg/ml 2 15 1 % Fibroblast Damage 3 23 15 5 8 Control NAB Rhodiola 1 μg/ml Control UV+ (1KJ/m2) NAB Rhodiola Extract 1 mg/ml IL-8 pg/ml 2 15 1 5 Control NAB Figure 3: Relative amount of DNA damage in normal human skin fibroblasts as determined through a Comet assay. 8 Shown are amount of DNA damage in culture medium (left), in medium irradiated with 1 kj/m 2 UV (center), and medium with 1mg/mL NAB Rhodiola Extract irradiated with same UV dose (right). The asterisk indicates that NAB Rhodiola Extract significantly minimizes UV induced DNA damage relative to untreated control (P<.5). The images show the raw data on which the quantitative data is based. Damaged DNA fragments are separated through electrophoresis and visualized using a fluorescent stain resulting in the appearance of a Comet tail. Figure 2: Expression of IL-6 (top) and IL-8 ( bottom) in normal human keratinocytes in culture medium (control), medium with 15 µm PMA, and medium with 1 µg/ml NAB Rhodiola Extract and PMA. The asterisk indicates NAB Rhodiola Extract significantly reduces the increased expression of both IL-6 and IL-8 induced by PMA (P<.5) Protecting Human Fibroblasts from UV radiation It is well documented that UV radiation can accelerate skin aging by either directly damaging DNA or through the generation of reactive oxygen species (ROS). 7 Figure 3 shows that NAB Rhodiola Extract significantly minimizes UV induced DNA damage in human fibroblasts in vitro relative to untreated control. Antioxidant activity (Gluthathione) A critical antioxidant that is naturally produced in the skin is Glutathione. Glutathione is a tripeptide that is normally found in its reduced form (GSH) but is sensitive to ROS itself leading to its inactive oxidized form (GSSG). The ratio of GSH to GSSG is therefore a marker of oxidative stress and is known to decrease with age. 9 Figure 4 shows the effect of NAB Rhodiola Extract on the GSH levels in human Keratinocytes (HaCaT) in vitro under UV stress. The results show that NAB Rhodiola Extract significantly outperforms both control and Resveratrol (positive control) in maintaining GSH levels. 3 Personal Care NAB Rhodiola Extract

Relative GSH level (AU) 1.5 1.2.9.6.3. Control UV (1.5J/cm2) UV + Resveratrol 1µM UV+ 1µg/mL NAB Rhodiola Extract Figure 4: GSH levels in normal human skin keratinocytes in culture medium, medium irradiated with UV 1.5J/cm2, medium with 1µM Resveratrol irradiated with same UV dose, and medium with NAB Rhodiola Extract irradiated with same UV dose. Table 2 shows that NAB Rhodiola Extract helps to inhibit the degradation of squalene, in the presence of artificial pollution. Sample Squalene (unexposed) Squalene (exposed) 98 Squalene + NAB Rhodiola Extract (exposed) 95 Table 2: Squalene degradation percentages after exposition to artificial pollution (gasoline motor and ozone generator); and with NAB Rhodiola.ANOVA, p<.5 Technical Specifications INCI Name Water/Pentylene glycol/rhodiola rosea root Extract No preservative added Recommended Use Level 1-3% Appearance light-dark amber liquid Lipid Peroxidation Just like proteins, skin lipids are also sensitive to the harmful effects of ROS. Specifically, lipid peroxidation has been established as a marker of cellular injury caused by oxidative stress. 1 The effect of NAB Rhodiola Extract on lipid peroxidation was determined in a lipid bilayer system by monitoring the presence of Malondialdehyde, one of the end products of lipid peroxidation. are listed in Table 1, showing that NAB Rhodiola Extract significantly inhibits lipid peroxidation relative to control and outperforms the positive control (Quercetin). Group % lipid peroxidation 1 Quercetin 4µg/mL (positive control) 18 NAB Rhodiola Extract 1 mg/ml 6 Table 1: Lipid peroxidation determined through photometry assessment of Malondialdehyde levels. Each result is the average of three separate evaluations. Anti-pollution Squalene, a major component of sebum, is highly prone to photooxidation and as such is a reliable oxidative stress marker. 11 In vivo studies have shown that those living in polluted areas tend to have decreased levels of squalene on the skin. 12 The anti-pollution benefits of NAB Rhodiola Extract were evaluated by monitoring the degradation of squalene in the presence of artificially generated pollution from a gasoline motor and ozone generator. NAB Rhodiola Extract Personal Care 4

References 1. Koohgoli R, Hudson L, Naidoo K, Wilkinson S, Chavan B, Birch-Machin MA. Bad air gets under your skin. Exp Dermatol. 217; 26(5): pp384-7. 2. Naidoo K, Birch-Machin MA. Oxidative Stress and Aging: The Influence of Environmental Pollution, Sunlight and Diet on Skin. Cosmetics 217; 4(1): pp4. 3. Wagner H, Nörr H, Winterhoff H. Plant adaptogens. Phytomedicine. 1994; 1(Jun 1): pp63-76. 4. Russotti G, Brieva TA, Toner M, Yarmush ML. Induction of tolerance to hypothermia by previous heat shock using human fibroblasts in culture. Cryobiology 1996; 33(5): pp567-58. 5. Franceschi C., Campisi J. Chronic inflammation (inflammaging) and its potential contribution to age-associated diseases. J Gerontol A Biol Sci Med Sci 214; June 69 (S1): pp4-9. 6. Countryman NB, Pei Y, Yi Q, Spandau DF, Travers JB. Evidence for Involvement of the Epidermal Platelet-Activating Factor Receptor in Ultraviolet-B-Radiation-Induced Interleukin-8 Production J. Invest. Dermat. 2; 115: pp267-272. 7. Shah P, He Y. Molecular Regulation of UV-Induced DNA Repair Photochem Photobiol. 215 91(2): pp254 264. 8. Singh NP, McCoy MT, Tice RR, Schneider EL. A Simple Technique for Quantification of Low Levels of DNA Damage in Individual Cells. Exp. Cell Research 1988; 175: pp184-91. 9. Weydert CJ, Cullen JJ. Measurement of superoxide dismutase, catalase and glutathione peroxidase in cultured cells and tissue. Nat Protoc. 21: 5(1); pp51-66. 1. Oxford Biomedical Research Colorimetric Microplate Assay for Lipid Peroxidation 11. Pham D-M, Boussouira B, Moyal D, Nguyen, L. Oxidization of squalene, a human skin lipid. A new and reliable marker of environmental pollution studies. Int. J. Cosmet. Sci. 215; 37: pp357-65. 12. Lefebvre MA, Pham DM, Boussouira B, Qiu H, Ye C, Long X, Chen R, Gu W, Laurent A, Nguyen QL. Consequences of urban pollution upon skin status. A controlled study in Shanghai area. Int J Cosmet Sci. 216; 38(3): pp217-23. Review and follow all product safety instructions. The statements made in these materials have not been evaluated by the U.S. Food and Drug Administration or any other regulatory authority. Lonza s products are not intended for use to diagnose, treat, cure or prevent any disease. All product information corresponds to Lonza s knowledge on the subject at the date of publication, but Lonza makes no warranty as to its accuracy or completeness and Lonza assumes no obligation to update it. Product information is intended for use by recipients experienced and knowledgeable in the field, who are capable of and responsible for independently determining the suitability of ingredients for intended uses and to ensure their compliance with applicable law. Proper use of this information is the sole responsibility of the recipient. This information relates solely to the product as an ingredient. It may not be applicable, complete or suitable for the recipient s finished product or application; therefore republication of such information or related statements is prohibited. Information provided by Lonza is not intended and should not be construed as a license to operate under or a recommendation to infringe any patent or other intellectual property right. No claims are made herein for any specific intermediate or end-use application. All trademarks belong to Lonza or its affiliates or to their respective third parties and are used here only for informational purposes. 218 Lonza 5 Personal Care NAB Rhodiola Extract

USA Lonza Consumer Care 7 Tyler Place South Plainfield, NJ 78 Tel +1 98 561 52 Switzerland Lonza Ltd Muenchensteinerstrasse 38 42 Basel Tel +41 61 316 81 11 www.lonza.com www.lonza.com/personalcare Personal Care NAB Rhodiola Extract 3/18