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Tomorrow’s Vision… Today!®
Truth + Fiction + Path to SustainabilityMeristemic Cells + Brand Differentiation + Phyto-Biotics
CONTENTS
1. Our Environment + Our Footprint + Our Responsibility
2. Sustainability in the Cosmetic + Personal Care Markets
4. A New Approach: Brand Differentiation + Sustainability
3. Meet the Stem Cell
5. Phyto-Biotics Product Line
OUR ENVIRONMENT + OUR FOOTPRINT
On average, American women use 12 personal care products a day, and men average six products daily. That means an adult is likely to be exposed daily to
126 unique chemical ingredients in personal care products alone.1
Environmental, social and economic impact of the personal care industry
Research shows the most significant footprints in the cosmetic industry lie with packaging and ingredients
The cumulative effects of wasteful and irresponsible habits associated with the personal care industry can significantly contribute to global warming
Reference: Good Sustainability Practice (GSP) for the Cosmetics Industry. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2010) 1-35.
OUR NEGATIVE IMPACT
"Our global footprint now exceeds the world's capacity to regenerate by about 30 percent.“2
FACTS:
Extinction rates are rising by a factor of up to 1,000 above natural rates. Every hour, three species disappear. Every day, up to 150 species are lost. Every year, between 18,000 and 55,000 species become extinct. The cause: human activities. 3 ~ Ahmed Djoghlaf
Every day, an estimated 100 plant and animal species are lost to deforestation. 4
Many river systems approach the fate of those in China, where chiefly because of pollution 80 percent of the 50,000 kilometers of major channels can no longer support fish of any kind. 5
OUR RESPONSIBILITY
“The social, economic and environmental pillars of sustainability all have a place at the heart of the life cycle of cosmetic and personal care products. The
integration of these elements is not a marketing tool, it represents a genuine commitment to sustainability … at many levels of the business model.”
~ Bertil Heerink - Director-General of Cosmetics Europe
What this means?
Personal Care and Cosmetic companies need to think and act sustainably
Includes sourcing, producing, manufacturing, packaging, and distributing
Attention to health and well-being of consumers
Reference: Good Sustainability Practice (GSP) for the Cosmetics Industry. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2010) 1-35.
WHAT IS SUSTAINABILITY?
Development that meets the needs of the present without compromising the ability of future generations to meet their own needs.
Three Pillars of Sustainability
Environmental
Economic
Social
Sustainability takes an honest look at how a company’s business practiceseffect each of these pillars on an individual scale.
Reference: Good Sustainability Practice (GSP) for the Cosmetics Industry. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2010) 1-35.
ACHIEVING SUSTAINABILITY
Creating and maintaining a company that generates a profit
Use available resources wisely(employees, materials, money)
Reduce business’s effects on the environment
Integrate and remain a successful part of the community and how it operates
Manage, train and develop aconscientious staff
Social
Environment Economic
Bareable Equitable
Viable
Sustainable
Reference: Ten Steps to Sustainability; All you need to know and do for a successful start. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2011) 1-15.
PERSONAL CARE MARKET + SUSTAINABILITY
The personal care and cosmetic industries directly affect all three pillars of sustainability
Consume natural resources at an unsustainable rate Diminishing biodiversity
Initiatives, specifically by the European Trade and Cosmetic Association have been taken to reduce negative impacts and increase positive impacts
Evaluate operations to identify the social, economic and environmental impacts at every stage in the business value chain:
Raw materials Product design + development Production Packaging Transport + retailing Use End of life
Reference: Ten Steps to Sustainability; All you need to know and do for a successful start. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2011) 1-15.
BENEFITS OF SUSTAINABILITY
Reduce costs + sheds light on areas of wasted resourced
Inspires product development
Maintain/ increase sales
Fosters competitive position in the market + remain competitive in the future
Employee Motivation + Employee Satisfaction =Increased Productivity + Improved Company Climate
Boosts company reputation and community perception
Promotes + nurtures supplier and retailer relationships
Viewed as fiscally responsible leading to enhanced investments
Reference: Good Sustainability Practice (GSP) for the Cosmetics Industry. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2010) 1-35.
A FACE-LIFT FOR THE STEM CELL
“The new age of anti-aging is how Cosmetic Design is describing plant stem cell technology, saying plant stem cell extracts are efficacy in a jar”
~ Eric Perrier/LVMH
The demand for stem cell technologies that promise potent anti-aging benefits has grown exponentially in the last decade
Cosmetic market has seen many incarnations of stem technology Plant stem cells Ingredients to activate endogenous adult stem cells
Plant stem cell technology facilitates the development of eco-friendly and sustainable cosmetic products. Technological advances allow cosmetic scientists to select, extract and improve natural molecules.
A FACE-LIFT FOR THE STEM CELL
WHAT ARE STEM CELLS?
Non-specialized cells capable of self-renewal…
What this means…
Each new cell has the potential to either remain a stem cell or become another type of cell with a more specialized function
Specialized cells define cell differentiation
Cellular Plasticity:
A specific characteristic of stem cells
The cell’s ability to move from an undifferentiated state toa specific cell type
Types of Plasticity:
Pluripotent-cells that can transform from a generic plant or animal cell into many different cell types
Totipotent-cells that can transform into any cell type
STEM CELL DIFFERENTIATION
Meristem Cells
• Pluripotent cells found in plants
• Have the ability to replicate beyond Hayflick’s Limit
• No ethical issues surround the use of plant stem cells (in contrast to embryonic stem cells)
• Currently two approaches to stem cells: Stimulation of adult stem cell proliferation The use of plant stem cells
STEM CELLS + CURRENT TECHONOLOGY
Meristemic Cell Technology in Cosmetics
Most popular form of stem cell technology
Said to slow aging by defending against extrinsic stress
This technology uses non-differentiated cells from simple cell extracts
Real problem for “anti-aging” cosmetic products… No specific activity No cosmetic benefits
STEM CELLS + COSMETIC MARKET TRENDS
The Green Approach: Plant Cell Cultures
Molecules of interest are highly concentrated in the meristem cells (buds)
In culture, all cells are dedifferentiated cells with meristematic character
Only requires a small amount of plant tissue to find active substance
Completely eco-friendly, protects biodiversity, no harvesting wild plants or crops
Full access to natural substances plus a higher presence of key plantmolecules
Numerous plants can be grown in culture in the presence of bacteria and yeast to enhance the production of secondary metabolites
PLANT CELL CULTURES + MARKET TRENDS
A NEW APPROACH:THE TRUE PATH TO SUSTAINABILITY
A sustainable and eco-friendly product line
• The use of biotic stress induces secondary metabolites: Allows for the sustainable production of extracts precisely
standardized for specific actives
• The use of secondary metabolites: Essential for the plant to interact with its environment Allows for adaptation, defense & ultimately survival in less than ideal
conditions
Biotic Stress
Damage caused by living organisms prompts plant to excrete stress responses
Examples: Bacteria, Fungi, Viruses, & Insects
Abiotic Stress
A negative impact on plants due to extreme environmentalconditions and/or pathogenic stress
Examples: Extreme Heat/ Cold, Intense UV Radiation, Flood/ Drought, Salicylate Production
A NEW APPROACH:BRAND DIFFERENTIATION + SUSTAINABILITY
Efficacious + Eco-Sustainable Extracts
Continued destruction of plants poses a major extinction threat
Secondary metabolites have no fundamental role in the maintenance of a plant’s life process
Isolating these organic compounds through solvent extraction for specific benefits
Numerous plants can be grown in culture in the presence of bacteria and yeast to enhance the production of secondary metabolites
Allows for the differentiation of plant stem cell trough pathogenic stress
A NEW APPROACH:BRAND DIFFERENTIATION + SUSTAINABILITY
PHYTO-BIOTICS PRODUCT LINE
Use biotic stress embodied by sustainable practice of co-culturing plant cells with Leuconostoc or Pseudomonas sp.
This approach induces cellular differentiation, or finely customized materials, standardized for specific activity and cosmetic skin benefits
Phyto-Biotics Product Line promotes brand differentiation & a means ofincluding patented, potent and efficacious actives Cosmetic benefit claims link to the product not the active Avoid intellectual property infringement
Phyto-Biotics Acai®
• Code - 16587
• INCI Name – Euterpe Oleracea Fruit Extract
• INCI Status – Approved
• Suggested Use Levels – 1.0-10.0%
• Suggested Applications – Anti-aging, Anti-wrinkle, Soothing, Antioxidant, ATP Synthesis, Increases Cellular Metabolism, Moisturizing, Decrease in Moisture Regression
PHYTO-BIOTICS PRODUCT LINE
Addresses:
Intrinsic + Extrinsic Aging
Dehydrated Skin
How:
Antioxidant protection
Increased ATP Synthesis
Enhanced Cellular Metabolism
Potent moisturizing benefits that continue to moisturize the skin even after application has ceased
PHYTO-BIOTICS ACAI®
Why is Acai of Interest?
This tree is known for its ability to survive in an extremely hot environment
Tree evolved by producing secondary metabolites to survive
What Secondary Metabolite Helps Acai Trees Survive?
Ferulic Acid (C10H10O4):
Found in the plants cell wall components
Readily forms a resonance stabilized phenoxy radical
Acts as a potent antioxidant & ROS scavenger
PHYTO-BIOTICS ACAI®
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
In-vivo Moisturization AssayProtocol
• DermaLab Corneometer was used to measure moisture levels on the volar forearms
• Corneometer is an instrument that measures the amount of water within the skin
• The presence of moisture in the skin improves conductance & results in higher readings than dry skin
• Baseline readings were taken on day one of the study (T=0)
Graph 1. Average moisturization levels measured at each testsite.
Protocol
• DermaLab Corneometer was used to measure moisture levels on the volar forearms.
• Corneometer is an instrument that measures the amount of water within the skin
• The presence of moisture in the skin improves conductance & results in higher readings than dry skin
• Baseline readings were taken on day one of the study
20
40
60
80
100
120
Pe
rce
nt
(%)
Dif
fere
nce
Experimentalvs. UntreatedControl
Experimental vs. Base Lotion
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
In-vivo Moisturization Assay
Comparative Moisture Results Between Test Sites
0
t = 24 T= 1 week T= 2 weeks T= 3 weeks T= 4 weeks
Graph 2. Comparative moisture analysis between testsites
Protocol
• Genetically uniform, shoot-based clonal lines of Acai were isolated & co-cultured
• Inoculated with Leuonostoc sp. for 30 days
• Ferulic acid was extracted from both inoculated & un-inoculated
• Absorbance was measured at 333nm
• Improvements of Ferulic Acid content in the co-cultured Acai palms compared to the control
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
0-1 0-4 0-5 0-24 OM-1 OM-3 OM-8
Tota
lFe
rulic
Aci
dC
on
ten
t(m
g/g
FW)
Control Inoculated
Reference: Fu-Hsiang, L, et al. Ferulic Acid Stabilizes a Solution of Vitamin C & E & doubles its Photoprotection of Skin. Journal of Investigative Dermatology (2005)125;826-832.
In-vitro Ferulic Acid Assay
Improvements in Ferulic Acid Content in Acai Co-Cultured with Leuconostoc sp.
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
Graph 3. Compared to the control, Ferulic Acid content increases when Acai is co-cultured with Leuconostocsp.
Protocol
• Purpose of study was to evaluate the effect of dosage on skin protection using known antioxidant solutions
• Skin is pretreated with 75, 150 &250mL of 15.0% Vitamin C, 1.0%Vitamin E and 0.5% Ferulic Acid
• Skin is irradiated with solar-stimulated radiation at a MED
• Sunburn protection is dosedependent
0
5
10
15
20
25
30
1 2
SBC
/mm
skin
3
xMedControl
15.0% Vit C, 1.0% Vit E, 0.5% FA, 150mL
4 5
15.0% Vit C, 1.0% Vit C, 0.5% FA, 250mL
15.0% Vit C, 1.0% Vit E, 0.5% FA, 75mL
Reference: Fu-Hsiang, L, et al. Ferulic Acid Stabilizes a Solution of Vitamin C & E & doubles its Photoprotection of Skin. Journal of Investigative Dermatology (2005)125;826-832.
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
In-vitro Ferulic Acid Assay
Effect of Dosage on Skin Photo-Protection
Graph 4. Effects of dosage on photo-protection when using vitamins C, E and Ferulic Acid.
Protocol
• Purpose of this study was to compare photo-protection by topical antioxidant formulations
• Skin was pretreated with vehicle, 0.5% Ferulic Acid, 15.0% Vitamin C + 1.0% Vitamin E, , 15.0% Vitamin C + 1.0% Vitamin E + 0.5% Ferulic Acid
• Skin is irradiated with solar-stimulated radiation 2x – 10x minimal erythema dose (MED) at 2 x MED intervals
• Evaluations 1 day later
Reference: Fu-Hsiang, L, et al. Ferulic Acid Stabilizes a Solution of Vitamin C & E & doubles its Photoprotection of Skin. Journal of Investigative Dermatology (2005)125;826-832.
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
0
5
10
15
20
25
30
35
2 4 8 10
Co
lori
met
er(%
)In
cre
ase
6
Minimal Erythema Dose
In-vitro Ferulic Acid Assay
Colorimeter Measurements of Photoprotection by Antioxidant Solutions
Control Vehicle Ferulic Acid Vitamin C + Vitamin E Ferulic Acid + Vitamin C + Vitamin E
Graph 5. Colorimeter measurements of photoprotection by antioxidant solutions
Protocol
• Purpose of this study was to compare visual and antioxidant photo-protection by topical antioxidant formulations
• Skin was pretreated with vehicle, 0.5% Ferulic Acid, 15.0% Vitamin C + 1.0% Vitamin E, , 15.0% Vitamin C + 1.0% Vitamin E + 0.5% Ferulic Acid
• Skin is irradiated with solar-stimulated radiation 2x – 10x minimal erythema dose (MED) at 2 x MED intervals
• Evaluations 1 day later
Reference: Fu-Hsiang, L, et al. Ferulic Acid Stabilizes a Solution of Vitamin C & E & doubles its Photoprotection of Skin. Journal of Investigative Dermatology (2005)125;826-832.
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
0
5
10
15
20
25
30
35
40
2 4 8 10
SBC
/m
msk
in
6
Minimal ErythemaDose
In-vitro Ferulic Acid Assay
Visual Erythema + Antioxidant Photo Protection by Antioxidant Solutions
Control Vehicle Ferulic Acid Vitamin C + Vitamin E Ferulic Acid + Vitamin C + Vitamin E
Graph 6. Visual erythema + antioxidant photoprotection by antioxidant solutions
Protocol
• Trolox used as a positive control
• Solution were prepared at three concentration as a reference
• Fluorescent measurements were takenevery two minutes for two hours
• Phyto-Biotics Acai® showed antioxidant activity at 0.025% concentrations
Graph 5. The results indicated that Ferulic Acid provides comparable antioxidant activity toTrolox.
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
In-vitro ORAC Assay
Protocol
• Human fibroblasts are allowed to grow to confluency in complete DMEM.
• 1.0%, 0.1% and 0.01% of Phyto-Biotics Acai® was used
• Incubated with fibroblasts for 24 hours
• Decrease in IL-6 production, indicates a reduce inflammatory environment
• Decrease signs of aging & reduceformation of lines & wrinkles
Graph 6. Decrease in IL-6 production when using Phyto-Biotics Acai® indicates a reduced inflammatoryenvironment.
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
In-vitro IL-6 ELISA Assay
Protocol
• Measures cell mediated cytotoxicity,cell proliferation & metabolic activity
• Human fibroblasts were seeded into a 96 well tissue culture with complete DMEM
• Concentrations tested were 1%, 0.1% & 0.01%
• The fibroblasts were incubated for 24 hours
• .01% Phyto-Biotics Acai® increased cell metabolism
Graph 7. Cellular metabolism of Phyto-Biotics Acai® treated in fibroblasts expressed in terms of percentcontrol
EFFICACY TESTING + PHYTO-BIOTICS ACAI®
In-vitro Cellular Viability Assay
Phyto-Biotics Acai®
• Code - 16587
• INCI Name – Euterpe Oleracea Fruit Extract
• INCI Status – Approved
• Suggested Use Levels – 1.0-10.0%
• Suggested Applications – Anti-aging, Anti-wrinkle, Soothing, Antioxidant, ATP Synthesis, Increases Cellular Metabolism, Moisturizing, Decrease in Moisture Regression
PHYTO-BIOTICS PRODUCT LINE
PHYTO-BIOTICS PRODUCT LINE
Phyto-Biotics Perilla
• Code – 40600
• INCI Name – Perilla Frutescens Extract
• INCI Status – Approved
• Suggested Use Levels – 1.0-10.0%
• Suggested Applications – Anti-aging, Anti-wrinkle, Soothing, Antioxidant, ATP Synthesis, Increases Cellular Metabolism, Increases Moisturization
Addresses:
Intrinsic + Extrinsic Aging
Dehydrated Skin
How:
Antioxidant protection
Increased ATP Synthesis
Reduces Appearance of Wrinkles
Enhanced Cellular Metabolism
Potent moisturizing benefit
PHYTO-BIOTICS PERILLA
Brand Distinction Using Perilla
Of particular interest – Perilla frutescens
This Chinese basil is a popular medicinal treatment
Can flourish in the most desolate conditions
Its secondary metabolites provide:
Potent antioxidant properties
Soothing properties
Protection from inflammation caused by free radicals
PHYTO-BIOTICS PERILLA
What Secondary Metabolite Protects Perilla?
Rosmarinic Acid (C18H16O6):
Ester of caffeic acid & 3,4 dihydroxyphenyllactic acid
Its four phenolic hydrogens contribute to its ability to control free radical oxidation
Contains two catechol (1,2 dihydroxybenzene) rings, giving it a quality of polarity
Thus can form intermolecular hydrogen bonds between the free hydrogen of its hydroxyl & its phenoxyl radical
Significantly improving its radical stability
PHYTO-BIOTICS PERILLA
0.0
50.0
100.0
150.0
200.0
250.0
T = 24 Hours T = 1 Week T = 2 Weeks T = 3 Weeks T = 4 Weeks
Mo
istu
riza
tio
n(µ
Sie
men
s)
2.0% Phyto-Biotics Perilla in Base Lotion
Base Lotion
Untreated
Protocol
• DermaLab Corneometer was usedto measure moisture levels on thevolar forearms
• Corneometer is an instrument that measures the amount of water within the skin
• The presence of moisture in theskin improves conductance & results in higher readings than dry skin
• Baseline readings were taken on day one of the study
Graph 1. Average moisturization measured at eachsite
In-vivo Moisturization Assay
Average Increase in Moisture Levels
EFFICACY + PHYTO-BIOTICS PERILLA
Protocol
• DermaLab Corneometer was used to measure moisture levels on the volar forearms
• Corneometer is an instrument that measures the amount of water within the skin
• The presence of moisture in the skin improves conductance & results in higher readings than dry skin
• Baseline readings were taken on day one of the study
0.0
20.0
40.0
60.0
80.0
100.0
120.0
140.0
160.0
T = 24 Hours T = 1 Week T = 2 Weeks T = 3 Weeks T = 4 Weeks
Graph 2. Comparative moisture analysis between testsites
Pe
rce
nt
(%)
Dif
fere
nce
2.0% Phyto-Biotics Perilla + Base Lotion vs. Untreated
2.0% Phyto-Biotics Perilla in Base Lotion vs. Base Lotion
In-vivo Moisturization Assay
Comparative Moisture Results Between Test Sites
EFFICACY + PHYTO-BIOTICS PERILLA
Protocol
• Genetically uniform, shoot-based clonal lines of Perilla frutescens were isolated & co-cultured
• Inoculated with Pseudomonas, sp for 30 days
• Rosmarinic acid was extracted from both inoculated & un-inoculated
• Absorbance was measured at 333nm
• Improvements of Rosmarinic Acid content in the co-cultured Perilla frutescens compared to the control
0
1
2
3
4
5
6
7
0-1 0-4 0-5 0-24 OM-1 OM-3 OM-8
Tota
lPh
en
olic
Co
nte
nt
(mg
/gFW
)
ClonalLines
EFFICACY + PHYTO-BIOTICS PERILLA
In-vitro Rosmarinic Assay
Improvements in Rosmarinic Acid in Perilla frutescens Co-Cultured with Pseudomonas sp.
Control Inoculated
Graph 3. Improvements in Rosmarinic Acid in Perilla frutescensco-cultured with Pseudomonassp.
Reference: Mi Yoo, Sun and Ran Kang, Jeong et al. Antioxidant Effects of Rosmarinic Acid on Human Skin Melanoma Cells Treated with Hydrogen Peroxide.Journal of Korean Society of Applied Biological Chemistry (2009) 52(3), 247-251
Protocol
• To evaluate antioxidant effect of Rosmarinic Acid (RA) on cultured human skin melanoma cells injured by ROS
• Cytotoxicity & antioxidant effects were analyzed by an XTT assay after cells were treated with H2O2
• Rosmarinic Acid increased cell adhesion activity & DPPH-radical scavenging activity in cells treated with H2O2
• Rosmarinic Acid showed antioxidant effects on ROS, such as H2O2
0
20
40
60
80
100
120
LDH
Act
ivit
y(%
of
Co
ntr
ol)
Protective Effects of Rosmarinic Acid on H2O2
EFFICACY + PHYTO-BIOTICS PERILLA
In-vitro Rosmarinic Assay
Control 90nMH2O2 100 Mg/ mL 130 Mg/mL
Graph 4. The protective effects of Rosmarinic Acid againstROS.
Reference: Mi Yoo, Sun and Ran Kang, Jeong et al. Antioxidant Effects of Rosmarinic Acid on Human Skin Melanoma Cells Treated with Hydrogen Peroxide.Journal of Korean Society of Applied Biological Chemistry (2009) 52(3), 247-251
Protocol
• Trolox used as a positive control
• Solution were prepared at three concentration as a reference
• Fluorescent measurements were taken every two minutes for two hours
• Phyto-Biotics Perilla showed antioxidant activity at 0.1% concentrations
10
20
30
40
50
60
70
An
tiox
idan
tC
apac
ity
(nM
TE)
0
200nM Trolox 12.5nM Trolox Phyto-Biotics Perilla 0.1%
Graph 5. The results indicated that Phyto-Biotics Perilla providescomparable antioxidant activity to Trolox.
EFFICACY + PHYTO-BIOTICS PERILLA
In-vitro ORAC Assay
Protocol
• Human fibroblasts are allowed to grow to confluency in complete DMEM
• 1.0%, 0.1% and 0.01% of Phyto-biotics Perilla was used
• Incubated with fibroblasts for 24 hours
• A decrease in IL-6 production, indicates a reduce inflammatory environment
• Decrease signs of aging & reduce formation of lines & wrinkles
-80
-70
-60
-50
-40
-30
-20
-10
0
Phyto-Biotics Perilla1.0%
Phyto-Biotics Perilla0.1%
Phyto-Biotics Perilla0.01%
Perc
ent
(%)
Dec
reas
e
Graph 6. Decrease in IL-6 production when using Phyto-BioticsPerillaindicates a reduced inflammatoryenvironment.
EFFICACY + PHYTO-BIOTICS PERILLA
In-vitro IL-6 Elisa Assay
Protocol
• Measures cell mediated cytotoxicity, cell proliferation & metabolic activity
• Human fibroblasts were seeded into a 96 well tissue culture with complete DMEM
• Fibroblasts were incubated for 24 hours
• 0.10% & 0.01% concentrations of Phyto-Biotics Perilla increased cell metabolism
104
106
108
110
112
114
116
118
Via
bili
ty(%
of
Co
ntr
ol)
Phyto-Biotics Perilla 0.10% Phyto-Biotics Perilla 0.01%
Graph 7. Cellular metabolism of Phyto-Biotics Perilla treated in fibroblasts expressed in terms of percentcontrol..
EFFICACY + PHYTO-BIOTICS PERILLA
In-vitro Cellular Viability Assay
PHYTO-BIOTICS PRODUCT LINE
Phyto-Biotics Perilla
• Code – 40600
• INCI Name – Perilla Frutescens Extract
• INCI Status – Approved
• Suggested Use Levels – 1.0-10.0%
• Suggested Applications – Anti-aging, Anti-wrinkle, Soothing, Antioxidant, ATP Synthesis, Increases Cellular Metabolism, Increases Moisturization
PHYTO-BIOTICS PRODUCT LINE
Phyto-Biotics Quercus
• Code – 16588
• INCI Name – Quercus Alba Bark Extract
• INCI Status – Approved
• Suggested Use Levels – 1.0-10.0%
• Suggested Applications – Anti-aging, Anti-wrinkle, Soothing, Antioxidant, ATP Synthesis, Increases Cellular Metabolism, Increases Moisturization, Improves Skin Density
Addresses:
Intrinsic + Extrinsic Aging
Dehydrated Skin
Sagging Skin + Wrinkles
How:
Antioxidant Protection
Enhances ATP Synthesis + Cellular Metabolism
Improves Skin Density
Potent Moisturizing Benefit
PHYTO-BIOTICS QUERCUS
Quercus – The Herbal Soother
Quercus alba, or White Oak bark, was traditionally used for its astringent & anti-inflammatory effects
Has high constituents of phenolic compounds, tannins & quercin
Potent astringent properties:
Helps absorb toxins
Sooth irritated/swollen skin
Can improve appearance of problem skin
PHYTO-BIOTICS QUERCUS
What Secondary Metabolite Protects Quercus?
Tannic Acid (C72H52O46): Ester of caffeic acid & 3,4 dihydroxyphenyllactic acid
Its four phenolic hydrogens contribute to its ability to control free radical oxidation
Contains two catechol (1,2 dihydroxybenzene) rings,giving it a quality of polarity
Thus can form intermolecular hydrogen bongs between the free hydrogen of its hydroxyl & its phenoxyl radical
Significantly improving its radical stability
PHYTO-BIOTICS QUERCUS
Protocol
• DermaLab Corneometer was used tomeasure moisture levels on the volarforearms
• Corneometer is an instrument that measures the amount of water within the skin
• The presence of moisture in the skin improves conductance & results in higher readings than dry skin
• Baseline readings were taken on dayone of the study
Graph 1. Average moisturization levels measured at each testsite.
0.00
50.00
100.00
150.00
200.00
250.00
300.00
T = 24 Hours T = 1 Week T = 2 Weeks T = 3 Weeks T = 4 Weeks
2.0% Phyto-Biotics Quercus + Base Lotion
Base Lotion
Untreated
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vivo Moisturization Assay
Average Increase in Moisture Levels
0.00
20.00
40.00
60.00
80.00
100.00
120.00
140.00
160.00
Pe
rce
nt
(%)
Dif
fere
nce
2.0% Phyto-Biotics Quercus+ Base Lotion vs. Untreated
2.0% Phyto-Biotics Quercus+ Base Lotion vs. Base Lotion
Protocol
• DermaLab Corneometer was used tomeasure moisture levels on the volarforearms
• Corneometer is an instrument that measures the amount of water within the skin
• The presence of moisture in the skin improves conductance & results in higher readings than dry skin
• Baseline readings were taken on day one of the study
T = 24 Hours T = 1 Week T = 2 Weeks T = 3 Weeks T = 4 Weeks
Graph 2. Comparative moisture analysis between testsites
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vivo Moisturization Assay
Comparative Moisture Results Between Test Sites
60.00
65.00
70.00
75.00
80.00
85.00
T = 0 T = 1 Week T = 2Weeks
T = 3Weeks
T = 4Weeks
Am
ou
nt
of
Co
llag
en
Experimental (2.0% Phyto-Biotics Quercus+ Base Lotion)
Base Lotion
Untreated
Protocol
• 10 M/F subjects, ages 23-45. High Resolution Ultrasound Skin imaging was used on the volar forearms
• The intensity of the signals arereceived refer to a color scale
• Dark colors represent areas of skin with low reflection-no or little change in skin density
• Bright colors represent areas with strong reflections or substantial change in skin density
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vivo Skin Density Assay
Average Results Per Test Site
Graph 3. Skin density measured via a high resolution ultra sound which reports collagen level at each test site.
Protocol
• 10 M/F subjects, ages 23-45. High Resolution Ultrasound Skin imaging was used on the volar forearms
• The intensity of the signals arereceived refer to a color scale
• Dark colors represent areas of skin with low reflection-no or little change in skin density
• Bright colors represent areas with strong reflections or substantial change in skin density
-0.10
Graph 4. Percent Difference between test site collagen levels over 4 weeks.
-0.05
0.00
0.05
0.10
0.15
0.20
T = 0 T = 1 Week T = 2 Weeks T = 3 Weeks T = 4 Weeks
Perc
en
t(%
) D
iffe
ren
ce
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vivo Skin Density AssayCollagen Ultrasound
Experimental vs. Base Lotion Treatment
Protocol
• Genetically uniform, shoot-basedclonal lines of Quercus alba wereisolated & co-cultured
• Inoculated with Leuconostoc, sp for 30 days
• Tannic acid was extracted from both inoculated & un-inoculated
• Absorbance was measured at 333nm
• Improvements of Tannic Acid content in the co-cultured Quercus alba compared to the control
0
1
2
3
4
5
6
Tota
lTan
nic
Aci
dC
on
ten
t (m
g/gF
W)
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vitro Tannic Acid Assay
Improvements in Tannic Acid Content in White Oak Bark Co-Cultured with Leuconostoc sp.
AC81912 AC61712 AC51112 AC12513 AC22413 AC30813 AC42813
Control Inoculated
Graph 5. Improvements in Tannic Acid in Quercus alba co-cultured with Leuconostocsp.
Protocol
• Trolox used as a positive control
• Solution were prepared at three concentration as a reference
• Fluorescent measurements weretaken every two minutes for twohours
• Phyto-Biotics Quercus showed antioxidant activity at 0.5% concentrations
0
50
100
150
200
250
200nM Trolox 12.5nM Trolox 0.5% Phyto-Biotics™ Quercus
An
tiox
idan
tC
apac
ity
(nM
TE)
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vitro ORAC Assay
Graph 6. The results indicated that Tannic Acid providescomparable antioxidant activity to Trolox.
Protocol
• Human fibroblasts are allowed to grow to confluency in complete DMEM
• 0.1% and 0.01% of Phyto-BioticsQuercus was used
• Incubated with fibroblasts for 24 hours
• Decrease in IL-6 production, indicatesa reduce inflammatory environment
• Decrease signs of aging & reduce formation of lines & wrinkles
-90
-70
-50
-30
-10
Phyto-Biotics Quercus 0.10% Phyto-Biotics Quercus 0.01%
Perc
ent
(%)
Dec
reas
e
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vitro IL-6 Elisa Assay
Graph 7. Decrease in IL-6 production when using Phyto-BioticsQuercusindicates a reduced inflammatoryenvironment.
Protocol
• Measures cell mediated cytotoxicity, cell proliferation & metabolic activity
• Human fibroblasts were seeded into a 96 well tissue culture with complete DMEM
• Concentrations tested were 0.01%
• The fibroblasts were incubated for24 hours
• Phyto-Biotics Quercus at a concentration of 0.01% increased cell metabolism
80
90
100
110
120
Control Phyto-Biotics Quercus 0.01%
Graph 8. Cellular metabolism of Phyto-Biotics Quercus treated in fibroblasts expressed in terms of percentcontrol..
Via
bili
ty(%
of
Co
ntr
ol)
EFFICACY + PHYTO-BIOTICS QUERCUS
In-vitro Cellular Viability Assay
PHYTO-BIOTICS PRODUCT LINE
Phyto-Biotics Quercus
• Code – 16588
• INCI Name – Quercus Alba Bark Extract
• INCI Status – Approved
• Suggested Use Levels – 1.0-10.0%
• Suggested Applications – Anti-aging, Anti-wrinkle, Soothing, Antioxidant, ATP Synthesis, Increases Cellular Metabolism, Increases Moisturization, Improves Skin Density
REFERENCES
1. Living Planet Report. World Wide Fund for Nature. Ropress. Gland, Switzerland. October 2008. http://www.yoursharemakesadifference.com/ysmad_statistics.html (2.14.2014)
2. Dojoghlaf, Ahmed. U.N. Convention on Biological Diversity. Message to the Board. (2007) http://www.yoursharemakesadifference.com/ysmad_statistics.html (2.02.14)
3. Problem: Extinction of Plant and Animal Species. The National Wildlife Federation. 2010. http://www.yoursharemakesadifference.com/ysmad_statistics.html (2.02.14)
4. Wilson, E.O. The Creation – An Appeal to Save Life on Earth. W.W. Norton & Company, Inc. New York, New York. 2006.
5. Good Sustainability Practice (GSP) for the Cosmetics Industry. Colipa – European Trade Association. Colipa – The European Cosmetics Association. Brussels. (2010) 1-35.
6. Ten Steps to Sustainability; All you need to know and do for a successful start. Colipa – European TradeAssociation. Colipa – The European Cosmetics Association. Brussels. (2011) 1-15.
7. Fu-Hsiang, L, et al. Ferulic Acid Stabilizes a Solution of Vitamin C & E & doubles its Photoprotection of Skin. Journal of Investigative Dermatology (2005)125;826-832.
8. Mi Yoo, Sun and Ran Kang, Jeong et al. Antioxidant Effects of Rosmarinic Acid on Human Skin Melanoma Cells Treated with Hydrogen Peroxide. Journal of Korean Society of Applied Biological Chemistry (2009) 52(3), 247-251
Truth + Fiction + Path to SustainabilityMeristemic Cells + Brand Differentiation + Phyto-Biotics
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