an active shelter against pollution - · pdf file•non-radicals: related nonradical...

V.16

AN ACTIVE SHELTER AGAINST POLLUTION

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

THE SKIN IS CONSTANTLY EXPOSED

• The skin is an interface and thus is constantly exposed to environmental agents:

• These aggressors induce the generation of free radicals and other highly reactive

chemical species that damage skin cells and other components.

Air pollution

Free

radicals

Reactive species are involved in the development of the

extrinsic signs of skin aging and accelerating intrinsic aging.

Sun radiation

Smoke Damage to cells and

structural components

of the skin. Reactive

chemical

species

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

FREE RADICALS AND REACTIVE SPECIES

Reactive species: • Free radicals: species with unpaired electrons

• Non-radicals: related nonradical species

Reactive oxygen species

Reactive nitrogen species

Reactive carbonyl species

ROS

RNS

RCS

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

ROS AND RNS EFFECTS

• ROS and RNS react with carbohydrates, proteins, glucosaminoglycans, DNA and

lipids.

• Can cause extensive injury and eventually cell death.

• Can give rise to other reactive species, further amplifying the damage.

ROS RNS

ROS and RNS can induce numerous harmful effects in cells

and other components of the skin.

Superoxide

(O2·-)

Hydroxyl

(OH·)

Singlet oxygen

(O21Δg)

Ozone

(O3) Hydrogen peroxide

(H2O2)

Nitric oxide

(NO·)

Nitrogen dioxide

(NO2·)

Peroxynitrite

(ONOO-)

Free radicals

Non-radicals

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

DAMAGE TO BIOMOLECULES BY REACTIVE SPECIES

• Lipid peroxidation (or autooxidation)

Free radicals + lipids lipid peroxides.

Causes the leakiness of membranes and an impairment of proteins.

• Protein nitration

Peroxynitrite (RNS) + tyrosine (proteins) 3-nitrotyrosine.

Compromises phosphorylation of proteins thus inactivating enzymes and

receptors, which are essential for the regulation of cellular mechanisms.

N H 2

O H

O

H O N H 2

O H

O

H O

O 2 N

Tyrosine 3-nitrotyrosine

peroxynitrite

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

REACTIVE SPECIES ORIGINATING FROM THE ENVIRONMENT

• Air pollution:

Originated mainly from fossil fuel combustion, but also

from cigarette smoke.

Includes SOx and NOx, CO, ozone, heavy metals, organic

aerosols…

Organic aerosols contain polycyclic aromatic

hydrocarbons, such as pyrene.

• Pyrene gets activated by visible/UVA light and increases

ROS.

• UV radiation:

Generation of ROS in the skin, inducing inflammation,

extracellular matrix breakdown and DNA damage.

Reactive species originate from air

pollution and sun radiation.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

REACTIVE SPECIES ORIGINATING FROM ENDOGENOUS SOURCES

• Mitochondria:

Major source of endogenous reactive species.

ROS are a by-product of the normal aerobic

metabolism (oxidative phosphorylation).

• Other sources:

Endoplasmic reticulum (cytochrome).

Peroxisomes (oxidative enzymes).

RNS

Reactive species are formed physiologically and play

important roles in biological systems at low concentrations.

ROS

ROS

RNS

At low levels, reactive species have physiological roles

(defense from microorganisms, cell signaling…).

And indeed they are necessary…

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

OXIDATIVE IMBALANCE LEADING TO AGING

ROS

RNS

Antioxidants

Oxidative imbalances produce stress that damages the skin

and cause the appearance of signs of aging.

• Oxidative and nitrosative stress results in damage to skin cells and other

components, leading to premature aging.

• A topical application of antioxidants is an effective preventive strategy.

OXIDATIVE STRESS

NITROSATIVE STRESS

• Endogenous antioxidants:

Enzymes: catalase, superoxide dismutases.

Molecules: vitamins C, E, coenzime Q10.

• Are reduced with age.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.


Chromane with a dual scavenging capacity for ROS and RNS to

protect the skin from oxidative damage.

• Scavenger of peroxynitrite and singlet oxygen.

• Inhibited peroxidation of lipids.

• Better antioxidative profile compared with other

antioxidants. Superior stability in a formulation.

• Prevented DNA damage induced by UV radiation and

by the atmospheric pollutant pyrene.

• Maintained cell viability in presence of oxidative

stress.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

LIPOCHROMAN® synthetic molecule EFFICACY

IN VITRO MOLECULAR-LEVEL EFFICACY

• Scavenging of RNS

• Singlet oxygen quenching

• Inhibition of peroxidation of membrane lipids

• Protection of essential oils from lipid peroxidation

• Antioxidative power

IN VITRO CELLULAR-LEVEL EFFICACY

• Shield against cellular oxidative stress

• Defense against photo-oxidative damage

• Protection against pollution components

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

SCAVENGING OF RNS

• Tyrosine and peroxynitrite were incubated with LIPOCHROMAN® synthetic molecule.

• The ability to scavenge peroxynitrite was monitored by measuring the formation of

3-nitrotyrosine by HPLC.

LIPOCHROMAN® synthetic molecule

prevented tyrosine nitration.

The active ingredient was able to block

3-nitrotyrosine formation by up to 94.4%.

3-n

itro

tyro

sine f

orm

ati

on (

%)

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

SINGLET OXYGEN QUENCHING

• Singlet oxygen was generated in the presence of different concentrations of

LIPOCHROMAN® synthetic molecule.

• The amount of singlet oxygen was measured by its near-IR emission to assess the

kinetics of its decay and calculate the quenching rate constant (kq).

LIPOCHROMAN® synthetic

molecule showed scavenging

properties toward singlet oxygen.

The active ingredient reduced the

lifetime of singlet oxygen. Its kq was

of (1.21 ± 0.08) x 108 M-1s-1, close

to α–tocopherol kq of 1.22 x 108 M-1s-1.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

INHIBITION OF LIPID PEROXIDATION

• Microsomal membranes were incubated with LIPOCHROMAN® synthetic molecule or

other antioxidants. Lipid peroxidation was induced by FeSO4 and ascorbic acid.

• The formation of lipid peroxides was assessed by using the thiobarbituric acid reactive

substances (TBARS) test.


molecule efficiently reduced

lipid peroxidation.

COMPOUND IC50 (µM)

LIPOCHROMAN® synthetic molecule 1.04 ± 0.13

Idebenone 11.5 ± 0.65

Kinetin > 1000

Lipoic acid > 1000


inhibited lipid peroxidation at lower

concentrations than other antioxidants.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

PROTECTION OF ESSENTIAL OILS FROM PEROXIDATION

The ingredient blocked peroxide

formation in both essential oils,

maintaining the initial levels along 2

weeks.


protected essential oils with time.

• Essential oils were mixed with potassium iodide and 0.01% LIPOCHROMAN® synthetic

molecule and kept at 60 ºC for 2 weeks.

• The amount of iodine liberated by the peroxidation of the oils was measured by titration

and the peroxide values were calculated.

Pero

xid

e v

alu

e (

mEq/k

g)

Pero

xid

e v

alu

e (

mEq/k

g)

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

ANTIOXIDATIVE POWER (I)

The active ingredient showed a higher AP

compared with that of other common

antioxidants. When stored in solution for 24 and

48 h it maintained its antioxidative properties.

• The antioxidative power (AP) method was used to compare LIPOCHROMAN® synthetic

molecule with other antioxidants while also assessing its stability in solution.

AP (

AU

) at

t 0

AP tr

0 h 1,130,000 0.20

24 h 1,310,000 0.19

48 h 1,147,000 0.17

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

ANTIOXIDATIVE POWER (II)

• The AP method was used to compare the properties of LIPOCHROMAN® synthetic

molecule with BHT alone or in a formulation.


showed better antioxidative

properties than BHT and remained

stable in a formulation.

AP (

AU

) at

RT


showed a 82-fold higher AP and 24-fold

greater reactivity compared with BHT.

In a formulation it had a higher AP with

time, even if stored at 40 ºC.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

SHIELD AGAINST CELLULAR OXIDATIVE STRESS (I)

• Human dermal fibroblasts (HDFa) were exposed to oxidative stress by the addition of

hydrogen peroxide (H2O2). Cell viability was assessed by the calcein-AM and the neutral

red uptake methods.

Pretreatment with the ingredient preserved cell viability in a

dose-dependent manner.

Non-treated

non-exposed

Non-treated

H2O2-exposed

LIPOCHROMAN®

synthetic molecule

(1µg/ml) + H2O2 ₂ ₂

₂ ₂ ₂ ₂

Cell v

iabilit

y (

%)

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

SHIELD AGAINST CELLULAR OXIDATIVE STRESS (II)

• HDFa were pretreated with different antioxidants and exposed to oxidative stress by the

addition of H2O2. Cell viability was assessed by the calcein-AM test.

LIPOCHROMAN® synthetic molecule was more effective than

resveratrol, vitamin E and ferulic acid against oxidative stress.

Cell v

iabilit

y (

%)

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

DEFENSE AGAINST PHOTO-OXIDATIVE DAMAGE (I)

• Human melanocytes were incubated with LIPOCHROMAN® synthetic molecule for 2 h

and irradiated with UVA (1.0 J/cm2) at 4 ºC.

• The DNA breaks induced were analyzed by the alkaline comet assay.

***p<0.001

DN

A d

am

age (

χ2 O

TM

)

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

DEFENSE AGAINST PHOTO-OXIDATIVE DAMAGE (II)

Non-treated

non-irradiated

Non-treated

UVA-irradiated


molecule (1 µg/mL) + UVA


molecule (10 µg/mL) + UVA LIPOCHROMAN® synthetic

molecule (50 µg/mL) + UVA

LIPOCHROMAN® synthetic molecule showed a photoprotective

capacity against DNA damage induced by UVA radiation.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

DN

A d

am

age (

χ2 O

TM

)

PROTECTION AGAINST POLLUTION COMPONENTS

• Human keratinocytes were incubated with LIPOCHROMAN® synthetic molecule in the

presence of 4 µM pyrene and irradiated with UVA/visible light to photoactivate pyrene.

The DNA breaks induced were analyzed by the alkaline comet assay.


protected skin cells from the

genotoxic effect of pollutants.

Non-treated +

pyrene + light


molecule (30 µg/mL) +

pyrene + light

The ingredient reduced DNA

damage by -99.3%, representing

significant protection.

***p<0.001

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

CONCLUSIONS

has a dual scavenging capacity: for ROS and for RNS.

inhibited tyrosine nitration up to -94.4% and showed a potent singlet oxygen scavenging

activity.

reduced peroxidation of membrane lipids and of essential oils.

presented a better antioxidative power compared with other well-known antioxidants, such as

BHT, even in a final formulation.

protected dermal fibroblasts against oxidative stress induced by ROS, increasing cell viability

up to 173.2% with respect to H2O2-exposed cells.

provided shelter from the ROS induced by UV irradiation or by air pollutants, reducing DNA

damage by up to -72.2% (UVA) and -99.3% (pyrene), in skin cells.

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

DESCRIPTION Chromane that protects biomolecules and cellular functions from the damage caused by the

reactive species generated from air pollution or UV radiation, as well as from endogenous sources,

consequently preventing the premature signs of aging.

APPEARANCE Powder.

INCI Dimethylmethoxy Chromanol. Preservative free.

PROPERTIES LIPOCHROMAN® synthetic molecule is a scavenger of ROS and RNS that inhibits protein inactivation

and DNA damage and prevents cell death due to oxidative stress.

APPLICATIONS LIPOCHROMAN® synthetic molecule can be incorporated into lipophilic-based cosmetic formulations

to avoid deterioration of skin.

DOSAGE 0.01-0.05%.

pH Not applicable.

TECHNICAL INFORMATION

© 2

014

The L

ubri

zol Corp

ora

tion.

All R

ights

Rese

rved.

AIMTEC® and LIPOCHROMAN® are owned by The Lubrizol Corporation.

The other tradenames and trademarks used herein belong to their respective and lawful owners.

Note: Graphs and photographs of efficacy tests are available for customer use provided that the final product contains the same concentration of active as the formulations in our tests. Customers must

request written permission for use of the graphic material and/or ingredient tradenames to Lipotec. Customers are responsible for compliance with local and international advertising regulations.

The specific situation of the trademark in each country may vary and we recommend that you contact us for updated information.

Disclaimer:

While the claims and supporting data provided in this publication are believed to be reliable and they are presented free and for guidance only, there are no warranties of any kind made as to its

accuracy, suitability for particular applications, how the product(s) will perform in combination with other substance or in the user’s process or the results obtained. All expressed and implied warranties

are disclaimed. Lipotec MAKES NO WARRANTIES, EXPRESS OR IMPLIED, INCLUDING, BUT NOT LIMITED TO, THE IMPLIED WARRANTIES OF MERCHANTABILITY AND FITNESS FOR A PARTICULAR PURPOSE. The

recipient is solely responsible for ensuring that products marketed to consumers comply with all relevant laws and regulations and assumes all risk and liability of any use or handling of any materials.

Recipient of this publication agrees to indemnify and hold harmless each entity of the LIPOTEC organization for any and all actions arising from recipient’s use of any claims or information in this

publication, including, but not limited to, use in advertising and finished product label claims, and not present this publication as evidence of finished product claim substantiation to any regulatory

authority. Nothing contained herein is to be considered as permission, recommendation, nor as an inducement to practice any patented invention without permission of the patent owner.


an active shelter against pollution - · pdf file•non-radicals: related nonradical...

Documents