Active Beauty

Darkenyl™
Advanced hair pigmentation recovery
October 2018
‘Wrinkles mean you laughed,
Grey hair means you cared,
Scars mean you lived…’

Confidential and proprietary business information of Givaudan 2

Attitude Towards Ageing
Overall, what is your feeling about "ageing"? It is mainly …?

Positive

Neutral
Negative

Globally, ageing is more a neutral

topic than negative.
The brand education and the
global average data beauty movements in recent years
have shifted people’s mind to
Ageing is some extent.
mainly… It is no longer a negative thing by
default.

1712 respondents on 6 countries

 Attitude Towards Ageing
Ageing may be perceived as a positive evolution

% of people that consider ageing as a positive thing in different

countries.

USA Brazil France Germany China Japan

Question: Overall, what is your feeling about "ageing"? It is mainly …?

1712 respondents on 6 countries
Word Associated with Ageing
It is a mix of positive, negative and neutral elements
Question: What does “aging” mean to you? Please indicate the level to which you associate
the words below with “aging”. (% is the sum of “Completely” and “Very much”)

beauty
battle 80% mature
worry confidence
60%
Globally, loneliness experience
consumers see
40%
“ageing”
less social interaction natural
associated most
with: 20%

challenge freedom
# experience 0%
# mature
# change dependence happiness
# health problems
# natural
health problems quality time

less active sexy

look old charming

change authenticity
What makes you look older?
Most obvious signs of ageing

Confidential and proprietary business information of Givaudan 6

Which body areas is the main concern?
Among different body areas, which ones do you worry most
about "ageing"?

Aesthetic
Globally, consumers
worry most about facial
skin ageing. Hair ageing
is almost as worrisome as
health teeth ageing which is
associated more with
health issues.

Interesting to note, both

#1 and #3 concerns are
aesthetic orientated.
Top 3 body areas
Global average data

1712 respondents on 6 countries

 Hair whitening
Absence of colour in the hair shaft, resulting from the loss of
pigment (melanin) production in the hair follicle.

Canitie
Confidential and proprietary business information of Givaudan 8
White hair in our society

Between45 and 65
74% years old the incidence
of white hair reaches
74% of people.

Men are showing a

higher risk of white
78% 71% hair between 45 and 65
years old than women.

Hair whitening differs with

geographical and
ethnic origins with lower
incidence in Asian and
African populations rather
than Caucasians.
Confidential and proprietary business information of Givaudan 9
How Old is OLD?
What age do you start worrying about "ageing"?

25%
24%
22%
21%
19%
18%
16% 16%

11% skin ageing

8% hair ageing
5% 5%
3% 3%
2% 2%

I am not less than 20 20 30 40 50 60 older than

worried 60
about it

The trend is similar between skin ageing and hair ageing.

The majority of people start worrying about it between
30~50 y.o.
1712 respondents on 6 countries
 In which countries, people worry most about hair ageing?
Among different body areas, which ones do you worry most
about "ageing"?

74%
Global average 66%

59% 57%
54% 53% 53%

USA BR FR DE CN JP
(263) (287) (329) (357) (238) (238)

Japanese and Chinese see hair ageing most worrisome, above

the global average 59%.
1712 respondents on 6 countries
 How impactful are the signs of hair ageing?
What concerns consumers most

visible / external less visible / internal

Thinning and Hair color loss, More sensitive to Dry scalp Slower hair
hair loss white / grey hair aggression growth

59% 51% 40% 38% 35%

#1 #2 #3 #4 #5

Question: Here are some signs of hair ageing. Please indicate the level of the importance
to you or the impact on your life. 1712 respondents on 6 countries
How many consumers worry about hair color loss?
Are you currently bothered by "hair color loss, white/grey
hair"?

Over 60% of people in the world are currently bothered by

hair color loss…
1712 respondents on 6 countries
How many consumers try to tackle the issue?
Have you used any products to deal with “hair color loss,
white / gray hair” in the past 12 months?

66% 65%
62% 62%
Global average
57%
= 60%
50%

USA (166) BR (193) FR (178) DE (169) CN (185) JP (163)

60% of people in the world tried one (or more)

product(s) in the past year…
What are the main pain points with these products?
You said that you are “dissatisfied” or “very dissatisfied” with
the products that you have used. What are the reasons?

The effect does not last long / have to re-apply often 66%

Hair quality is worse (less shine, more break etc) 50%

Not effective in covering the white / gray hair 50%

Does not feel safe 38%

Does not look natural 37%

Bad / harsh / chemical smell 34%

It takes too much time 33%

The color is not homogeneous on the hair 29%

Not easy to use 24%

Expensive 20%

0% 10% 20% 30% 40% 50% 60% 70%

How does the hair get old?
Hair follicle
‘A mini organ’

• Regenerate completely
during the hair cycle.

• The bulge is the niche for

stem cells: Epidermal stem
cells (EpSCs) and
Melanocytes stem cells
(MSCs or
melanoblasts)

• Stem cells migrate to the

dermal papilla and help to
create new hair and provide
pigmentation to the
hair.

Confidential and proprietary business information of Givaudan 17

 Hair pigmentation
Closely linked to hair cycle

ANAGEN CATAGEN TELOGEN EARLY ANAGEN

Melanin produced by The melanocytes enter in The melanocytes are Bulge stem cells
active melanocytes is apoptosis and disappear eliminated. proliferate and migrate
transferred to cortical during telogen phase to the bulb.
keratinocytes

 Pigmentation of the  Stop of production  A signal is sent to  Melanoblast

entire hair shaft. of melanin. the bulge for starting differentiate into
a new anagen. active melanocytes.

Confidential and proprietary business information of Givaudan 18

Oxidative stress
Plays a major role in the process
of hair ageing

• ROS generated in scalp decrease

the differentiation of melanoblasts
into melanocytes resulting in a lack
of mature melanocytes in the bulb
region.

• ROS decrease the melanogenesis

at bulb level resulting in decrease
of melanin pigments transferred to
the cortical keratinocytes

Confidential and proprietary business information of Givaudan 19

What can you do about hair ageing?
It’s your choice

Hide it Wear it And now…

Reactivate
pigmentation
Confidential and proprietary business information of Givaudan 20
Darkenyl™
Smart active for
hair pigmentation recovery

A smart combination of:

• Taxifolin glucoside
a stabilised
antioxidant, acting as
a stimulator of stem
cells proliferation and
maintenance.

• N-acetyl-tyrosine
a precursor in melanin
synthesis pathway.

Confidential and proprietary business information of Givaudan 21

Darkenyl™
Advanced hair pigmentation recovery

1. Boosts proliferation of stem

cells into the bulge

2. Increases melanogenesis into

melanocytes of the dermal papilla

3. Reduces ROS damages on

melanocytes by activating
antioxidant defenses
4. Stimulates the recovery of hair
pigmentation.

Confidential and proprietary business information of Givaudan 22

The bulge
The stem cells factory of the
hair follicle

Confidential and proprietary business information of Givaudan 23

Taxifolin Glucoside stimulates stem cells proliferation
In vitro
Induction
(∆%) vs Cellular proliferation of ORSc
Hair follicle stem cells (ORSc) were control
treated for 24 hours with increasing
+30%
concentration of taxifolin glucoside 50
(0.04% = 1% Darkenyl™). ***
***
40
The proliferation of ORSc was
evaluated by measuring the
incorporation of. 30 ***

20
Results: **

10
Taxifolin glucoside significantly
increases:
0
 proliferation of the hair follicle 10ng/mL 0.002%
2μM 0.008%
10μM 0.04%
50μM
stem cells, up to +30% versus EGF Taxifolin Glucoside
control conditions.
 Bromodeoxyuridine (BrdU) into DNA of ** p<0.01 Student’s t-test
proliferating cells *** p<0.001 Student’s t-test

Confidential and proprietary business information of Givaudan 24

Taxifolin Glucoside stimulates hair follicle stem cells genes
In vitro

Hair follicle stem cells (ORSc) were

treated for 24 hours with 0.04% of
% vs Expression of genes involved in stem cells
untreated proliferation and maitenance
taxifolin glucoside (= 1% Darkenyl™).
700
The expression of various genes was
evaluated by RT-qPCR. 600

500

400
Results:
300
Taxifolin glucoside increases
significantly the expression of genes
200
involved in:

 proliferation of stem cells, 100

 hair follicle morphogenesis, 0

MKI67 PCNA WNT10B B catenin Keratin-15 VDR
 maintenance of hair follicle
stem cells’ phenotype. Proliferation Hair follicle Stem cell
morphogenesis maintenance

Confidential and proprietary business information of Givaudan 25

Hair follicle melanocytes
The pigmentary unit

Confidential and proprietary business information of Givaudan 26

Darkenyl™ stimulates melanogenesis
In vitro

A co-culture of normal human Melanogenesis stimulation

keratinocytes (NHK) and normal Melanin
(μg/mL/mg
human melanocytes (NHM) were proteins)
treated with Darkenyl™ at 0.01%
or Rapamycin (positive control), or +364%
30
left untreated for 72 hours. **

25
The melanin content in the cell *
supernatant was evaluated by 20
colorimetric assay (405nm).
15

10

Results: 5

 Darkenyl™ significantly 0
increases the melanogenesis Untreated Rapamycin 1µM Darkenyl™ 0.01%™
by +364 %. * p<0.05 Student’s t-test
** p<0.01 Student’s t-test

Confidential and proprietary business information of Givaudan 27

Oxidative stress
Main cause of hair ageing

Confidential and proprietary business information of Givaudan 28

Darkenyl™ activates antioxidant and pigmentation genes
Pro oxidative stress ex vivo test
In order to mimic an oxidative ∆Relative
stress occurring in ageing hair expression Expression of genes involved in pigmentation
vs oxidative
follicle, face lift skin explants from stress
and antioxidant activities
3 donors (average age 63) were control (%)
treated for 3 days repeatedly 180
during 1 hour with a mix of 9 mg ***
160
of hypoxanthin + 10 units of xantin **
oxidase. Then a treatment was 140
*** **
done with 1% of Darkenyl™. ** **
120
**
**
100
The expression of genes involved
*
in hair pigmentation, hair renewal 80 # #
and antioxidant defenses were 60 * ** **
measured by RT-qPCR. # **
40 * #
Results: 20

 Under oxidative stress, 0

MITF

HMOX1
CTNS

FST

GSS
HPS5
KRT5
EDN1

POMC

GLRX
AP3B1

KRT19
MYO5A

MAP1LC3A

MGST1
NRF2
TXN
Darkenyl™ significantly MC1R
stimulates the expression of
genes involved in antioxidant
defenses, renewal, and Pigmentation Renewal Antioxidant
pigmentation of the hair
follicle. # p<0.1 ANOVA Fisher
* p<0.05 ANOVA Fisher
** p<0.01 ANOVA Fisher
*** p<0.001 ANOVA Fisher

Confidential and proprietary business information of Givaudan 29

Darkenyl™ reduces ROS production
Pro oxidative stress ex vivo test

ROS score
Human hair follicles from a female (A.U) ROS accumulation in hair follicle
donor (56 years old) were treated 3.5
**
with 1% of Darkenyl™ or left 3
untreated. The hair follicles were -53%**
2.5
then exposed to an oxidative
stimulus (cumene hydroperoxide - 2
50µM) for 1 hour. 1.5 *

1
The ROS accumulation into the
hair follicle was evaluated using a 0.5
green fluorescent probe. 0
Untreated Untreated Darkenyl™ 1%
+ oxidative stress
* p<0.05 Student’s t-test
Results: ** p<0.01 Student’s t-test

 Under oxidative stress,

Darkenyl™ significantly
decreases by -53% the ROS
accumulation into the hair
follicle.

Confidential and proprietary business information of Givaudan 30

Darkenyl™ protects melanocytes against oxidative stress
Pro oxidative stress ex vivo test
%
pigmenting Protection of melanocytes/melanoblasts
Human hair follicles from a female cells
donor (56 years old) were treated 0.60%
with 1% of Darkenyl™ or left
0.50%
untreated. The hair follicles are
+189%**
then exposed to an oxidative 0.40%
stimulus (cumene hydroperoxide - 0.30%
50µM) for 1 hour.
0.20%
**
The pigmenting cells 0.10%
(melanocytes and melanoblasts)
quantification was obtained by 0.00%
Untreated Untreated Darkenyl™ 1%
immunostaining (in red on the
pictures below). + oxidative stress
** p<0.01 Student’s t-test

Results:

 Under oxidative stress,

Darkenyl™ significantly
protects the melanocytes and
melanoblasts by +189% into
the hair follicle

Confidential and proprietary business information of Givaudan 31

Reactivating pigmentation
In white hair

Confidential and proprietary business information of Givaudan 32

Darkenyl™ stimulates melanin production in white hair
Ex vivo
Melanin Boosting melanin content in white hair
content
(D% of
White human hair follicles in untreated)
anagen phase (growing) from 2
donors (35 and 53 years old) were
treated with 1% of Darkenyl™
for 72 hours or left untreated. +15%*
10%
The melanin content into the
hair follicles was evaluated using
melanin quantification with
Fontana Masson staining (black on
the pictures below) and image
analysis. 0%
Darkenyl™ 1%
* p<0.05 Student’s t-test

Results:

 Darkenyl™ significantly
increases by +15% the
production of melanin into
the hair follicles in 3 days.

Confidential and proprietary business information of Givaudan 33

Clinical test
Significant reduction of white hair in 4 months
Reduction of the white hair
proportion
Clinical test
Protocol:
 Double blind versus placebo
 2 groups of 22 volunteers
 Men (18 years old and above)
 With white hair
 One group of volunteers applied a
placebo, The second group applied the
hair lotion containing Darkenyl™ at 1%.
 The application was done in leave-on by
massage on the scalp, once a day for 4
months.

2 different measurements were performed:

• Scoring the white hair proportion,
• Quantifying the white hair density.
INCI:
AQUA/WATER, SODIUM BENZOATE, CITRIC ACID, SODIUM CITRATE, PPG-26
BUTETH-26, PEG-40 HYDROGENATED CASTOR OIL, FRAGRANCE,
BUTYLPHENYL METHYLPROPIONAL, D-LIMONENE, ALPHA-ISOMETHYL
IONONE, +/- 1% DARKENYL™

Confidential and proprietary business information of Givaudan 35

#1 Scoring of the white hair proportion
Protocol

Scoring of the white hair proportion

Pictures of the scalp were taken at T0 & after 4months.

A blind scoring was then performed to evaluate the

proportion of white hair (% of white hair on a parting area
on the scalp).

+ 4months

Confidential and proprietary business information of Givaudan 36

#1 Scoring of the white hair proportion
Results

Scoring of the proportion of white hair

(after 4months)

Results: ΔD0 (%)

Placebo Darkenyl™ 1%
0%
After 4 months, Darkenyl™
-2%
significantly decreases the
proportion of white hair by -4%

an average of -17% -6%

-8%

2.1 times more than the -10%

*

placebo. -12%
x2.1*
-14%
-17%
-16%

-18% ***
* p<0.05 Mixed ANOVA
*** p<0.001 Mixed ANOVA

Confidential and proprietary business information of Givaudan 37

#1 Scoring of the white hair proportion
Best respondent

T0 After 4 months
Results:

Darkenyl™ at 1%
The proportion of white

Volunteer 18
hair is visibly reduced,
with a reduction of
-56% for the
best respondent
(from 90% of white hair
at D0 down to 40%).
Darkenyl™ at 1%
Volunteer 7

No white roots effect was

observed during hair
growth!

Confidential and proprietary business information of Givaudan 38

#2 Quantifying the white hair density
Protocol

Quantifying the white hair density

Two days after shaving a 1cm2 area (at D0 and after 4

months), a picture of the area is taken.

The density of white hair (number of white hair per cm2) is

evaluated by counting the number of white hair on a
predefined area of the scalp of 0.7cm2.

All hair whose root is in the zone are counted.

+ 4months

Confidential and proprietary business information of Givaudan 39

#2 Quantifying the white hair density
Results

Absolute density of white hair

Results: White hair
(after 4 months)
density vs. D0

After 4 months of (nb/cm2)

0
Placebo Darkenyl™ 1%

treatment, Darkenyl™
significantly decreases the -1

number of white hair per -2

cm2 by -6.8, -3

3.4 times more -4

efficiently than the placebo. -5

x3.4#
-6

Equivalent to a reduction of -7
*
4080 white hair or -8

-35 white hair / day # p<0.1 Mixed ANOVA

* p<0.05 Mixed ANOVA

(the average scalp size

being 600cm2).

Confidential and proprietary business information of Givaudan 40

#2 Quantifying the white hair density
Best respondent

Results: T0 After 4 months

The density of white hair is

visibly reduced, with a
reduction of -55.7 white

Placebo
hair/cm2 after 4 months
of application for the
-2.9 white hair/cm2
best respondent,
Darkenyl™ at 1%

Equivalent to a reduction of
-33420 white hair or
-275 white hair / day
(the average scalp size
-55.7 white hair/cm2
being 600cm2).

Confidential and proprietary business information of Givaudan 41

Macroscopic clinical results summary
Examples from our best respondents

Results snapshot:

Volunteer #18 Volunteer #7 Volunteer #17

Criteria
(51 years old) (53 years old) (52 years old)

% of white hair -56% -41% -42%

Density of white -31.4 white -55.7 white -35.7 white

hair/cm2 hair/cm2 hair/cm2 hair/cm2
Total decrease of
white hair number on -18,800 white hair -33,420 white hair -21,420 white hair
scalp (600 cm2)

Confidential and proprietary business information of Givaudan 42

Macroscopic clinical results example
Volunteer #21 (61 years old)

After 4 months
T0 (Darkenyl™ 1%)

Confidential and proprietary business information of Givaudan 43

Macroscopic clinical results example
Volunteer #23 (62 years old)

After 4 months
T0 (Darkenyl™ 1%)

Confidential and proprietary business information of Givaudan 44

Macroscopic clinical results example
Volunteer #30 (58 years old)

After 4 months
T0 (Darkenyl™ 1%)

Confidential and proprietary business information of Givaudan 45

Summary
Advanced hair pigmentation
recovery

Confidential and proprietary business information of Givaudan 46

Darkenyl™
Advanced hair pigmentation recovery

1. Boosts proliferation of stem

cells into the bulge

2. Increases melanogenesis into

melanocytes of the dermal papilla

3. Reduces ROS damages on

melanocytes by activating
antioxidant defenses
4. Stimulates the recovery of hair
pigmentation.

Confidential and proprietary business information of Givaudan 47

Consumers benefits
Darkenyl™
Biological benefits:
 Stimulation of stem cells proliferation
 Stimulation of melanogenesis into hair follicles
 Activation of antioxidant defences in hair follicles
 Protection of melanocytes against oxidative stress
 Reactivates melanin production in grey/white hair
 Decrease of grey/white hair density
 Hair repigmentation
 Hair repigmentation action
 Prevent against hair ageing

Applications:
 Anti grey/white hair lotion,
 Anti white hair shampoo,
 Natural repigmenting hair mask,
 Anti grey hair for beard and mustache,
 Hair color recovery spray,
 Gel for premature grey/white hair.

Confidential and proprietary business information of Givaudan 48

Technical information
Darkenyl™

Glycerin (and) Water (and) Sodium

Metabisulfite (and) Larix Europaea
INCI Wood Extract (and) Glycine (and) Zinc
Chloride (and) Camellia Sinensis Leaf
Extract (and) Acetyl Tyrosine

White biotechnology,
Origin
Green technology and synthesis

Preservation Sodium Metabisulfite

Appearance Pale yellow liquid

Solubility Water soluble

Dosage 0.1%-1.0%

Can be added at the end of the

Processing formulation process under stirring
below 40°C and pH below 5.0.

Compliance

Confidential and proprietary business information of Givaudan 49

Thank you

Givaudan Active Beauty

global.cosmetic@givaudan.com

Confidential business and proprietary information of Givaudan, may not be copied or distributed to anyone without the express written permission of Givaudan
Givaudan Active Beauty Sales offices
Europe Asia Pacific
Givaudan France SAS Givaudan Singapore Pte Ltd
17-23 rue de la Voie des Bans 1 Pioneer Turn
FR-95100 Argenteuil (France) 627576 Singapore (Singapore)

Givaudan UK Ltd Givaudan Shanghai Ltd

Magna House 298 Li Shi Zhen Road
76-80 Church Street Pudong Zhang Jiang High Tech Park
Staines, TW18 4XR (United Kingdom) 201203 Shanghai (China)

North America Latin America

Givaudan Fragrances Corp. Givaudan do Brasil Ltda
40 W – 57th Street - Floor 17 Av. Engº Billings - 2185, Edificio 31, 1ºAndar - Jaguaré
NY 10019 - New York (United States) 05321-010 São Paulo - SP (Brazil)

global.cosmetic@givaudan.com

Confidential business and proprietary information of Givaudan. May not be copied or distributed without the express written permission of Givaudan. 51