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Industrial Crops and Products 80 (2016) 93–100

Contents lists available at ScienceDirect

Industrial  Crops  and  Products

jo ur nal home p age: www.elsev ier .com/ locate / indcrop

he  green  generation  of  sunscreens:  Using  coffee  industrial
ub-products

.  Martoa,  L.F.  Gouveiaa,  B.G.  Chiarib,  A.  Paivac,  V.  Isaacb, P.  Pintoa, P.  Simõesc,
.J.  Almeidaa,  H.M.  Ribeiroa,∗

Research Institute for Medicines and Pharmaceutical Sciences (iMed.UL), Faculty of Pharmacy, University of Lisbon, Portugal
Faculdade de Ciências Farmacêuticas, UNESP – Univ Estadual Paulista, DFM – Laboratório de Cosmetologia – LaCos, São Paulo, Brazil
LAQV-REQUIMTE, Departamento de Química,Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Caparica, Portugal

 r  t  i  c  l  e  i  n  f  o

rticle history:
eceived 6 July 2015
eceived in revised form 4 November 2015
ccepted 9 November 2015
vailable online 6 December 2015

eywords:
pent coffee oil
reen coffee oil
upercritical fluid extraction
ater resistant

ickering emulsion sunscreen

a  b  s  t  r  a  c  t

Spent  coffee  grounds  and  green  coffee  defective  beans,  which  are  industrial  sub-products  of coffee  pro-
cessing,  have  a potential  use for cosmetic  applications,  due  to their  safety  and  high  content  in  lipids  that
present  interesting  physicochemical  properties.  Sunscreen  formulations  in  the  form  of  water-in-oil  emul-
sions might  be  a suitable  application  for these  sub-products  because  providing  a  higher  sun  protection
factor  (SPF)  for the  same  concentration  of  sunscreen  activities  than  oil-in-water  emulsions.

The  purpose  of  this  work  was  to  assess  the  biological  effects  of using  the  oil fraction  of  spent  coffee
grounds  extracted  with  supercritical  CO2 and  green  coffee  oil  in  the  development  of  new  generation  of
sunscreens  with  improved  sun  protection  performance.  The  oil  fractions  were  used  to prepare  w/o  sun-
screens involving  a cold  emulsification  process,  with  purified  water  as  disperse  aqueous  phase  and  TiO2

and  ZnO  particles  as  stabilizers.  The  sunscreens  were  characterized  in terms  of mechanical,  rheological
and  skin  adhesion  properties.  In  addition,  the  in  vitro  and  in  vivo  biological  properties  of the  formulations
were  evaluated,  including  safety  and  sunscreen  water  resistance  tests.

The  use  of  two  types  of  solid  particles  proved  to  be  useful  in the  developed  formulations,  ensuring
a  high  SPF  with  UVB/A  protection,  conferred  by TiO2 and  ZnO,  respectively.  Moreover,  the  emulsion
containing  35%  w/w  of  the  spent  coffee  grounds  oil fraction  presented  promising  characteristics  in  the
improvement  of water  performance  with  a  broad  spectrum  sun  protection  when  compared  to an  emulsion
containing  35% w/w  of  green  coffee  oil which  improved  the  SPF  in  physical  sunscreens.  The  formulations

are  industrial-scalable  and  suitable  for topical  use  according  to the  rheological,  mechanical  and  safety
assessment.

The use  of  spent  coffee  oil  in  cosmetic  industry  seems  to  be  a suitable  approach  for  the  valorisation  of
waste from  the  coffee  industry  and  presents  promising  characteristics  in the improvement  of  sunscreen
performance.

© 2015  Elsevier  B.V.  All  rights  reserved.
. Introduction

The efficacy of sunscreen products has been recognized as an
mportant public health issue and is usually expressed by the sun
rotection factor (SPF), which is calculated as the ratio between

he UV energy required to produce a minimal erythema dose of
rotected and unprotected skin (Dutra et al., 2004; Ribeiro et al.,
013).

∗ Corresponding author at: Research Institute for Medicines (iMed.ULisboa), Fac-
lty  of Pharmacy, Universidade de Lisboa Av. Professor Gama Pinto, 1649-003 Lisboa,
ortugal. Fax: +351 217946470.

E-mail address: hribeiro@campus.ul.pt (H.M. Ribeiro).

ttp://dx.doi.org/10.1016/j.indcrop.2015.11.033
926-6690/© 2015 Elsevier B.V. All rights reserved.
Avoiding sun exposure, covering the skin or applying sunscreens
with a high SPF are the main strategies strongly recommended to
prevent UV-induced cell damage. The UV filters can be divided in
two groups: (a) chemical filters which absorb UV radiation (UVR);
and (b) physical filters, such as, titanium dioxide (TiO2) which
reflect UVR (Ascenso et al., 2014).

Sunscreens are normally based on synthetic chemicals with high
capacity to absorb sun light at the region of UVB (320–290 nm)  and
UVA (400–320 nm)  spectrum. Several synthetic UV  filter molecules
(e.g. benzophenones, anthranilates, PABA derivatives, salicylates,

cinnamates and camphor derivatives) are available as photopro-
tective agents, but due to their harmful effects they are becoming
less popular. The main problem of the chemical sunscreen agents

dx.doi.org/10.1016/j.indcrop.2015.11.033
http://www.sciencedirect.com/science/journal/09266690
http://www.elsevier.com/locate/indcrop
http://crossmark.crossref.org/dialog/?doi=10.1016/j.indcrop.2015.11.033&domain=pdf
mailto:hribeiro@campus.ul.pt
dx.doi.org/10.1016/j.indcrop.2015.11.033


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4 J. Marto et al. / Industrial Cro

re the photoirritation, photosensitization and contact dermati-
is. Reducing the concentration of such chemicals in cosmetics is a
trategy to improve their quality, without affecting their properties
Serpone et al., 2007).

In recent years, the use of natural agents has been attracting sig-
ificant attention, due to their safety, multiple biological actions
n the skin and cost effectiveness. Consequently, an oily vehicle
ith antioxidant activity could be an excellent approach for its

mportant role on the product efficacy by improving the prod-
cts’ photoprotective activity (Calixto et al., 2011; Couto et al.,
009). In addition to bioactivity, natural products are, in general,
ot harmful for humans, not expensive, suitable to be used in a wide
ange of applications, and are obtained from renewable sources.
n this context, green coffee oil and spent coffee grounds oil have
risen as potential candidates to replace synthetic chemicals in sun-
creens since they are a rich source of antioxidants and polyphenols
Ribeiro et al., 2013). Phenolic compounds are excellent candidates
or the prevention of the harmful effects of UV radiation on the
kin. More specifically, flavonoids have photoprotection potential
ue to their UV absorbing capacity, ability to act as antioxidants
nd anti-inflammatory, and immunomodulatory agents (Saewan
nd Jimtaisong, 2013).

Coffee consumption is growing worldwide, being extremely
mbedded in the cultural habits of many countries, so the total
mport and export of coffee has been increasing. Spent coffee
rounds (SCG), which are the residue obtained after the treatment
f coffee with hot water or steam for extracting flavour substances,
an be used for industrial applications such as high quality biodiesel
roduction (Couto et al., 2009). Nevertheless, due to their high con-
ent in lipids, particularly fatty acids, SCG might also find a suitable
pplication in cosmetic products where these lipid compounds can
e used as valuable excipients. Therefore, the lipid fraction of SCG
xtracted with supercritical CO2 can be used in the development
f new, improved sunscreens. On average, a fifth of the Brazilian
offee production consists of defective beans, and several studies
ave been developed in order to find an alternative use for sub-
roduct, including the cosmetic application of the extracted oil
Preedy, 2014; Ribeiro et al., 2013).

Titanium dioxide has been incorporated in sunscreen formula-
ions for more than 25 years, being regarded as safe and effective,
hus bringing together two of the most desirable features in cos-

etic market (Renner, 2009). It is especially preferred by people
ith a high propensity for skin irritation, such as patients under-

oing oncological chemotherapy. Moreover, TiO2 particles are a
V-B filter, suitable for developing physical sunscreens combining
oth increasing stability and high SPF properties (Wang and Tooley,
011). The introduction of ZnO ensures an adequate protection in
he range of UVA. Due to its multifunctional nature, ZnO particles
ave been shown to be effective as antibacterial and antifungal
gent (Singh et al., 2012; Smijs and Pavel, 2011).

On the other hand, TiO2 particles can be also used as solid parti-
les for stabilization of surfactant-free emulsions stabilized by solid
articles (i.e. Pickering emulsions). This type of emulsions have

mportant advantages over the classical surfactant-based emul-
ions, such as higher resistance to coalescence due to an improved
tability, and a higher tolerability (Laredj-Bourezg et al., 2012). The
tabilization of emulsion droplets takes place by means of adsorp-
ion of solid particles at the surface of emulsion droplets. It can
e assumed that a stable water-in-oil (w/o) Pickering emulsion is

 function of particles concentration, pH and ionic strength. This
dsorption mechanism is quite different compared to surfactants
ince partial wetting of the solid particles surface by water and oil is

he reason of the strong anchoring of these particles at the water–oil
nterface. Few fully natural and biocompatible materials are avail-
ble for the effective stabilization of these emulsions since severe
 Products 80 (2016) 93–100

requirements must be simultaneously met, including insolubility in
both fluid phases and intermediate wettability (Folter et al., 2012).

The purpose of this work was  to develop and characterize a w/o
emulsion stabilized by physical sunscreens containing 35% of the
lipid fraction of spent coffee grounds extracted with supercriti-
cal CO2 and green coffee oil obtained from defective beans with
improved sunscreen performance. Sunscreen formulations might
be a suitable application for these types of sub-products because
w/o emulsions are water resistant and provide greater efficacy (a
higher SPF) for the same concentration of sunscreen actives than
their o/w counterparts (Couteau et al., 2012).

2. Material and methods

2.1. Materials

Spent coffee grounds oil (SCO) was  supplied by LAQV-
REQUIMTE – Departamento de Química, Faculdade de Ciências e
Tecnologia (Caparica, Portugal). Green coffee oil (GCO) was sup-
plied by Cooxupé – Cooperativa de Cafeicultores de Gauxupé,
(Minas Gerais, Brazil). Triethoxycaprylylsilane titanium dioxide
(mTiO2) (Unipure White LC 987) was a gift from Sensient (Milwau-
kee, USA). The starch used was aluminum starch octenylsuccinate
(ASt) (DryFlo® Plus) obtained from AkzoNobel (Amsterdam,
Netherlands). Zinc oxide (ZnO) (Tego® Sun Z 500) was obtained
from Evonik Industries AG (Essen, Germany). Purified water was
obtained by inverse osmosis (Millipore, Elix® 3).

2.2. Methods

2.2.1. Characterisation of the formulation ingredients
2.2.1.1. Wettability measurements. Contact angles of water, green
coffee oil and spent coffee oil on ZnO, mTiO2 and ASt in air atmo-
sphere were measured at room temperature by using ConAnXL—a
Microsoft Excel based workbook and add-in software (freely avail-
able upon request) as described in detail elsewhere (Marto et al.,
2015). All measurements were performed in triplicate.

2.2.1.2. Particle size distribution. Particle size distribution was
determined using a Malvern Mastersizer 2000 (Malvern Instru-
ments, UK), coupled with a Hydro S accessory. Data were expressed
in terms of relative distribution of volume of particles in the range
of size classes, and given as diameter values corresponding to per-
centiles of 10, 50 and 90. The Span value is a useful parameter to
characterize the particle size distribution broadness.

2.2.1.3. Natural oils.
2.2.1.3.1. Oil extraction. SCG oil was  obtained by supercriti-

cal CO2 extraction as described elsewhere (Ribeiro et al., 2013).
Supercritical fluid extraction can be an environmentally friendly
alternative to traditional organic solvent extraction processes
whereby extraction/separate recovery of oil and bioactive com-
pounds from agro-industrial residues can be done without their
degradation (Brunner, 2013). Mass transport is highly facilitated
owing to favourable transport properties (high mass and thermal
diffusivities coupled with low viscosities) and the solvation capac-
ity of the supercritical fluid is tuneable by changing the operating
conditions of pressure and temperature. The most commonly used
supercritical fluid, carbon dioxide, is non-toxic, non-flammable,
non-corrosive, relatively inert from a chemical point of view and
environmentally friendly. Its relatively low critical temperature

(304.3 K) allows extraction of thermolabile substances without
degradation.

The supercritical CO2 extraction of SCG oil was  done in a high
pressure extraction pilot unit. SCG were first dried in an oven at



J. Marto et al. / Industrial Crops and

Table  1
Qualitative and quantitative composition of the sunscreen formulations.

Ingredients Quantitative composition (%, w/w)

GCO sunscreen SCO sunscreen

Phase A
Green coffee oil 35 –
Spent coffee oil – 35

Phase B
Triethoxycaprylylsilane titanium dioxide 20 20
Zinc oxide 15 15
Aluminum starch octenylsuccinate 5 5

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Phase C
Purified water 25 25

78 K with air circulation until a moisture value of about 1%. Super-
ritical CO2 extractions were carried out at 55 ◦C and 250 bar using
.5 kg of dry SCG per batch with an average CO2 flowrate of 15 kg/h
or ca. 1 h of extraction.

GCO was obtained by cold pressed extraction. Briefly, green cof-
ee beans were pressed and come back to be pressed again for a
urther three times and then the oil is bottled with nitrogen.

2.2.1.3.2. SPF measurement. GCO and SCO were accurately
eighed (0.25 g), diluted with ethanol, followed by ultrasonication

or 5 min  and filtered through filter paper (WhatmanTM 42). The
bsorption spectra of samples solution were obtained in the range
f 290–320 nm (Hitachi U-2001, USA) every 5 nm,  using a standard

 cm quartz cell, and ethanol as the blank. Triplicates were made,
ollowed by the application of the Mansur equation (Eq. (1)).

PFspectrophotometric = CF × �320
290EE (�) × I(�) × Abs(�) (1)

here EE (�) is the erythemal effect spectrum; I (�) is the solar
ntensity spectrum; Abs (�) is the absorbance of sunscreen product;
F is the correction factor (=10) (Mansur et al., 1986). The values of
E × I are constants determined by Sayre et al. (1979).

.2.2. Sunscreen formulations—green coffee oil sunscreen (GCO
unscreen) and spent coffee oil sunscreen (SCO sunscreen)

Two formulations were developed (Table 1) based on their
acroscopic appearance, physical stability and SPF value. The

isperse aqueous phase was composed of purified water, the con-
inuous oil phase consisted of GCO or SCO. Solid particles (mTiO2,
nO and ASt) were firstly dispersed in the oil phase. The oil and
queous phases were then mixed using a high-speed homogenizer
UltraTurrax®, IKA-Werke GmbH & Co., KG, Germany) at room tem-
erature (cold process).

.2.3. Characterization of the sunscreen formulations

.2.3.1. Droplet size distribution. The emulsions were examined
 day after preparation, using an optical microscope (Olympus
X40, Japan) equipped with a video camera. The droplet size was
etermined using the image analysis software Olympus Stream
ssentials®. The size data was expressed in terms of relative size
istribution of particles (BS, 1993).

.2.3.2. Structural analysis of the sunscreen formulations.
2.2.3.2.1. Rheology studies. Shear rate vs shear stress measure-

ents were performed at 25 ◦C using a HAAKE RS-1 Rheometer,
quipped with automatic gap setting (Thermo ScientificTM,
altham, USA). Rotational viscosity was determined using a

35 mm cone geometry, with an angle of 2◦. Dynamic or shear
iscosity measurements were carried out by rotational shear exper-

ments, between 1 and 1000 Pa. Oscillation sweep tests were
erformed at frequencies ranging between 0.1 and 100 Hz. The
reep and recovery test was carried out with a shear stress of 1 Pa
or emulsions, allowing 360 s for creep and 360 s for relaxation. All
 Products 80 (2016) 93–100 95

tests were performed on samples of about 1 g, at 25 ± 0.5 ◦C. All
experiments were performed in triplicate using new samples for
each measurement.

2.2.3.2.2. Texture profile analysis (TPA). A Texture Analyzer
TA.XT Plus with Exponent 3.0.5.0 software (Stable Micro Systems
Ltd., Godalming, UK) was  used to examine hardness, elasticity,
compressibility, adhesiveness and cohesiveness of the emulsion.
A probe (P/10, 10 mm Delrin), which was twice depressed into
the sample at a defined rate (5 mm/s) to a desired depth (15 mm),
allowing 15 s of delay between consecutive compressions. Six repli-
cates were performed at 25 ◦C for each formulation.

2.2.3.3. In vitro SPF. The SPF was  assessed using the Optometrics
SPF-290S Analyzer (Optometrics Corporation, Essex, UK). The sam-
ples were prepared by spreading 110 mg  of each formulation over
a Transpore® tape (70.7 × 70.7 mm)  to obtain a film of 2 mg/cm2,
as specified by the European Regulation (EC, 2009). Each sample
was exposed to a xenon arc solar simulator, and the analyser per-
formed scans in 6 different spots on the Transpore® tape substrate.
Each scan takes a transmittance (T) measurement every 2 nm from a
wavelength ranging from 290 to 400 nm.  The Monochromatic Pro-
tection Factor (MPF) was  determined for the selected wavelengths
using Eq. (2). The SPF value was calculated using Eq. (3).

MPF  = 1
T

(2)

SPF =
∑400

290E�.B�
∑400

290
E�.B�
MPF�

(3)

where (E) is the spectral irradiance of terrestrial sunlight under
controlled conditions and (B) is the erythema effectiveness (Kale
et al., 2010).

2.2.3.4. In vitro sun product water resistance. The water resistance
of developed sunscreens was  measured using an improved in vitro
bath system. An amount of 2 mg/cm2 of sunscreen formulation
was dispensed onto the plate, and carefully applied with a rubber-
gloved finger. After drying for 15 min, the SPF of each sample was
determined using the SPF 290 analyzer (Optometrics SPF-290S Ana-
lyzer). The samples were immersed in the in vitro bath system
(29 ± 2 ◦C) and washed away by the water flow (150 rpm) during
20 min. The samples were allowed to air dry for 15 min and SPF
was measured again. The samples were immersed once more and
washed during 20 min. The samples were allowed to air dry for
15 min  and SPF was  measured to calculate the water resistance
retention (%WRR) of the sunscreens, as defined by Eq. (4).

%WRR  = SPFwet

SPFdry
× 100 (4)

where SPFdry and SPFwet are the SPFs before and after water immer-
sion, respectively (Ahn et al., 2008; COLIPA, 2005).

2.2.3.5. Skin adhesion properties. Skin adhesion measurements
were performed using Texture Analyzer TA.XT Plus (Stable Micro
Systems Ltd., Surrey, UK) equipped with a load cell of 5 kg, cylin-
der probe of 10 mm (P/10, 10 mm Delrin) and a measuring system
A/MUC (skin adhesion test rig), which holds the human skin. An
amount of 2 mg/cm2 of each sample was applied onto the skin.
The probe was  immersed in an in vitro bath system (29 ± 2 ◦C) and
washed away by the flow of water (150 rpm) during 40 min, in

order to simulate the water resistance method for sun protection
products (COLIPA, 2005). The area under the curve (AUC) was calcu-
lated from the force–distance plot as the work of skin adhesion per
square centimetre. The formulation given below was used to calcu-



9 ps and Products 80 (2016) 93–100

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Table 2
Contact angle of water, GCO and SCO on mTiO2, ZnO and ASt (mean ± SD, n = 3).

Contact angle (◦)

Samples Water Green coffee oil Spent coffee oil

3.2.1. Droplet size distribution
The use of a surfactant-free emulsion might be a strategy to

avoid skin irritations associated to this type of excipients (Marku
et al., 2012). In Pickering emulsions the droplet is stabilized by a

Table 3
Particle size distribution of the different solid particles proposed (mean ± SD, n = 6).

Solid Particles Particle size distribution (�m)
6 J. Marto et al. / Industrial Cro

ate the work of skin adhesion per square centimetre (mJ/cm2—Eq.
7)). Each experiment was carried out in triplicate.

orkofskinadhesion
(

mJ/cm2
)

= AUC
�r2

(7)

hereby �r2 is the surface area of the human skin that is in contact
ith the sunscreen formulations.

.2.3.6. Human repeat insult patch test (HRIPT). A safety evalua-
ion study was performed on emulsions, using the Marzulli and

aibach (1976) HRIPT protocol. In brief, the product was applied
n the back of 51 healthy volunteers that gave their prior informed
ritten consent. For the induction period, a series of nine patches

Finn Chamber standard) were performed over a period of 3 weeks.
eactions after patching were scored according to International
ontact Dermatitis Research Group (Fregert and Bandmann, 1975).

 2 weeks rest period was maintained without application of the
est material. During the challenge period, new patches were pre-
ared and fixed in the same manner as in the induction period, but
lso on the right side of the back (i.e.  a virgin site).

The protocol was approved by the local Ethical Committee and
espected the Helsinki Declaration and the AFSSAPS regulations on
erformed HRIPT studies on cosmetic products. The study was con-
ucted under the supervision of a dermatologist who  participated

n the evaluation of irritation/allergic reactions to the emulsions.

.2.3.7. In vivo sun product water resistance. The water resistance of
unscreens was tested on 3 subjects (Fitzpatrick skin type II). Pan-
llists cleanse their forearms using a mild cleanser and leave them
o air dry for 30 min  before starting the test. Initial cross-polarized
mages were taken after the sunscreens application (2 mg/cm2) on
he inner forearm (4 cm2).

The amount of each sunscreen formulation left before and after
ater bath immersion was quantified via cross polarized imaging

y means of the Visia® CA (Canfield Scientific, Faitfield, NJ). Panel-
ists immerse their forearms into a water bath system (29 ± 2 ◦C)
nd washed away by the flow of water (150 rpm) during 40 min
Ahn et al., 2008).

The RGB colour space of the raw bitmap images was  converted
o relative luminance using ImageJ®. From these images, average L
hanges for each sunscreen area were obtained from histograms.
kin whiteness was defined as the change in L value before and after
ater immersion, and the percentage of water resistance retention

%WRR) of the sunscreens was determined according to Eq. (8).

WRR  = Lwashedprotector − Lskin
Lprotector − Lskin

(8)

.2.4. Statistical analysis
The data are expressed as mean and standard devia-

ion (mean ± SD) of experiments. Statistical evaluation of data
as performed using one-way analysis of variance (ANOVA).

ukey–Kramer multiple comparison test (GraphPad PRISM 5 soft-
are, USA) was used to assess the significance of the difference

etween the groups (p < 0.05).

. Results and discussion

.1. Characterisation of the formulation ingredients

.1.1. Wettability measurements

In the surfactant-free system stabilized by solid particles, these

ill be preferentially wet by one of the liquids, with the more poorly
etting liquid becoming the disperse phase. Particle wettability at

he oil–water interface is quantified by the contact angle, �, that
mTiO2 106.5 ± 0.7 60.1 ± 1.9 9.7 ± 3.3
ZnO 100.2 ± 2.6 23.4 ± 0.1 13.1 ± 2.5
ASt 109.0 ± 0.4 10.6 ± 2.3 13.3 ± 4.0

the particle makes with it, which will determine the type of emul-
sion. Generally, if the water contact angle is <90◦, the solid surface
is considered hydrophilic and if the water contact angle is >90◦, the
solid surface is considered hydrophobic. Concerning the emulsions
stabilized by solid particles, if the contact angle, measured through
the aqueous phase, is <90◦ the emulsion will be o/w and, by con-
trast, if the contact angle is >90◦ the emulsion will be w/o  (Marku
et al., 2012). In this study, mTiO2, ZnO and ASt will stabilized more
effectively w/o  emulsions. All solid particles have a contact angle
with water >90◦, due to hydrophobic coatings, and simultaneously,
a contact angle with Green coffee oil (GCO) and Spent coffee oil
(SCO) <90◦ (Table 2). Thus, by combining these three particles it is
possible to obtain a stable w/o  emulsion.

3.1.2. Particle size distribution
Particle size distributions of mTiO2, ZnO and ASt showed that all

particles were larger than 100 nm (Table 3), thus complying with
the international “green” standards, whereby nanomaterials must
be avoided (COSMOS, 2013).

3.1.3. Natural oils
3.1.3.1. SPF measurement. Several studies have been developed in
order to find an alternative use for these defective coffee beans
or spent coffee ground, involving the characterization of their
lipid fractions. The GCO was extracted using mechanical pressing,
whereas the lipid fraction of SCG was extracted with supercritical
CO2, an environmentally friendly solvent that allows the extraction
and recovery of the oil at such conditions that no degradation of the
lipid composition oil occurs while avoiding the use of hazardous
organic solvents (Ribeiro et al., 2013).

The GCO and SCO have potential application in health care
products, including sunscreen formulations (Chiari et al., 2014).
In particular, due to the richness of polyphenols and flavonoids,
the pure GCO shows a SPF higher than 5, improving SPF and,
consequently, decreasing the concentration of chemical and/or
physical sunscreens in such formulations (Wagemaker et al., 2011).
In this study, GCO presented an SPF value of 5.03 ± 0.23 while
SCO presented only 1.57 ± 0.07, which is in line with the fact that
the coffee roasting process diminishes the content of polypheno-
lic compounds (Dai and Mumper, 2010; Speer and Kölling-Speer,
2006).

3.2. Characterization of the sunscreen formulations
Span d (0.1) d (0.5) d (0.9)

mTiO2 37.36 ± 1.38 0.14 ± 0.01 0.19 ± 0.01 7.12 ± 0.30
ZnO 19.51 ± 5.53 0.16 ± 0.01 0.57 ± 0.03 11.38 ± 3.74
ASt  1.00 ± 0.01 7.28 ± 0.01 13.52 ± 0.01 20.82 ± 0.01



J. Marto et al. / Industrial Crops and Products 80 (2016) 93–100 97

Table  4
Droplet size distribution of GCO and SCO sunscreen (mean ± SD, n = 625).

Formulation Droplet size distribution (�m)

Span d (0.1) d (0.5) d (0.9)

r
p
i
I
o
o
s
t
e
P
s
a
t
e
c
a
i
a

3
3
t
T
i
s

(
t
s
r

s
s
n
c
m
t
G
s

m
m
o
p
d

a
f
t
i
a
s
(

3
a
w
t

Table 5
Mechanical properties of the sunscreens extracted from the TPA mode (mean ± SD,
n  = 3).

Formulations Hardness g Adhesiveness
(|g s|)

Cohesiveness Compressibility
(g s)
GCO sunscreen 1.17 ± 0.02 3.01 ± 0.11 5.54 ± 0.28 9.49 ± 0.55
SCO sunscreen 0.90 ± 0.03 5.72 ± 0.10 8.87 ± 0.13 13.71 ± 0.34

eduction of the bare oil-water interface by adsorption of small
articles. The parameter to describe Pickering emulsions stability

s the contact angle of the adsorbed particles, as explained above.
nitially, a contact angle greater than 90◦ is required in order to
btain a w/o emulsion. In this assay we evaluated the influence
f the oil on Pickering emulsions droplet size distribution. Both
unscreen formulations showed a narrow droplet size distribu-
ion with a span value in the range of 0.9–1.2. The GCO sunscreen
xhibited lower mean droplet size than SCO sunscreen (Table 4).
revious studies demonstrated that coffee extracts have extremely
trong antioxidant properties than many other food products. This
pplies to both roasted and green coffee, although, the latter con-
ains even ten times higher concentration of polyphenols (Budryn
t al., 2013). Other study demonstrated that several polyphenols,
ould be adsorbed into the oil–water interface, present as water-
nd oil-insoluble particles and decrease the surface tension, provid-
ng very good stabilization of emulsions and, consequently, leading

 decrease in droplet size (Luo et al., 2011; Wagemaker et al., 2011).

.2.2. Structural analysis of the sunscreen formulations

.2.2.1. Rheology studies. The flow curves (Fig. 1(a)) showed that
he emulsions presented a shear-thinning and rheopetic behaviour.
his behaviour could influence the sunscreen performance, creat-
ng a uniform, impenetrable and protective semi-solid film over the
kin surface that is required to obtain an effective sunscreen.

In the range of 0–5 Pa, both emulsions were not disrupted
Fig. 1(b and c). Thus, the values of G’ and G” remained linear within
his region of linear viscoelasticity, which can therefore indicate the
uitable shear stress to be used in frequency sweep and creep and
ecovery tests.

The GCO sunscreen exhibited higher G’ and G” values than SCO
unscreen. The frequency sweep curves of GCO sunscreen (Fig. 1(b))
howed that in the range tested (0.1–100 Hz) there was  practically
o variation in the elastic and viscous moduli. Furthermore, vis-
oelastic behaviour was seen over the whole range, since G’ (elastic
odulus) was higher than G” (viscous modulus). The same rela-

ion is observed for SCO sunscreen, but the G’ and G” values for
CO sunscreen are higher than for SCO sunscreen, exhibiting high
tability.

In the creep and recovery test, both sunscreens suffered defor-
ation, shown by the compliance value (J), but SCO sunscreen was
uch more susceptible to this force (Fig. 1(c). In the recovery part

f this assay, when the shear stress was removed and the sam-
les could recover their former structure, the elastic part of the
eformation, was reversed.

In summary, the addition of GCO promoted an increase in the
pparent viscosity and the elastic modulus, when compared to the
ormulation prepared with SCO. Considering the results of the par-
icle size distribution (Table 4), the reduction in droplet size results
n an increase in the viscosity and storage modulus of the emulsions
nd the shear-thinning behaviour becomes even stronger, which
uggested an enhancement in emulsion stability and performance
Pal, 1996).
.2.2.2. TPA. The results of the calculations of textural parameters
re collected in Table 5. The adhesiveness is the only parameter,
hich differs between the two formulations (p < 0.05). Concerning

he emulsions adhesiveness, which is more a surface characteristic
GCO sunscreen 23.62 ± 1.76 39.76 ± 0.27 0.82 ± 0.04 29.87 ± 1.91
SCO sunscreen 25.23 ± 2.26 48.65 ± 0.53 0.83 ± 0.01 31.09 ± 1.86

and depends on a combined effect of adhesive and cohesive forces,
the presence of SCO caused an increase in this parameter.

According to Alves et al. (2003), during coffee roasting process
there are changes in the fatty acid composition, increasing the
trans-fatty acid levels, which increase the hydrophobic character
of the final formulation and, consequently enhance the adhesive
forces and water repellent performance (Shyr and Ou-Yang, 2015).
Other authors proved that spent coffee oil also contains more
hydrophobic compounds, such as fatty acids, fatty acid esters,
medium-chain paraffins and olefins than green coffee oil or other
vegetable oils (Kelkar et al., 2015).

Considering the hardness, cohesiveness and compressibility
results, the lipid type did not influence these parameters.

The fatty acid tails are hydrophobic because they are non polar
and the heads are hydrophobic because they are polar.

3.2.3. In vitro SPF
Both emulsions showed high values of SPF with a suitable

UVA/UVB ratio (Table 6). For GCO there was a synergistic increase
of SPF value (around 1.6 fold) when combined with physical fil-
ters as mTiO2 in GCO sunscreen. According to Wagemaker et al.
(2011) the oil extracted from unroasted beans is used by the cos-
metics industry due to its excellent properties for the human skin,
particularly maintaining skin-moisture due to its fatty acid com-
position. In addition, its ability to absorb UV radiation in the UVB
range allows its use in sunscreens as a SPF enhancer, allowing to
reduce the amount of physical and chemical filters (Chiari et al.,
2014).

3.2.4. In vitro sun product water resistance
The in vitro method measures the SPF following a defined water

immersion procedure. For a product to be considered water resis-
tant, the value for the lower 90% one-sided confidence limit has to
be greater than or equal to 50%. A ‘waterproof’ product should have
a %WRR exceeding 80% after two  immersions, while the %WRR of a
‘water resistant’ product should be more than 50% (COLIPA, 2005).

After the first immersion, the GCO sunscreen presented a %WRR
of 62.6, which decreased in the second immersion to 50.7%. In turn,
SCO sunscreen showed a value of 83.1% at first immersion and a
value of 80.8% of WRR  after the second immersion. Based on this
%WRR, it is possible to ensure the ‘water resistance’ mark for GCO
sunscreen and the ‘waterproof’ claim for SCO sunscreen. As men-
tioned previously, during the coffee roasting process the fatty acid
levels increase, enhancing the hydrophobicity of the SCO sunscreen
and, consequently, improving the effectiveness of water resistance
performance (Kelkar et al., 2015). Additionally, hydrophobic sun-
screen formulations can also serve as additional water barriers to
help mitigate the disruption in stratum corneum caused by constant
exposure to water (Shyr and Ou-Yang, 2015). In water resistant
products typically more polar lipophilic ingredients can be found.
Conversely, waterproofing sunscreens should contain more non-
polar ingredients, which is the case of SCO (Couteau et al., 2012).
3.2.5. Skin adhesion properties
Table 7 shows the skin adhesion properties of the formulations.

The force necessary to detach the GCO and SCO sunscreen samples
from the skin was  statistically higher than the one needed to detach



98 J. Marto et al. / Industrial Crops and Products 80 (2016) 93–100

eep an

t
a
T

Fig. 1. (a) Flow curves, (b) frequency sweeps and (c) cr
he blank. In our study, when the SCO was used, the work of skin
dhesion ability after water immersion was also increased (Table 7).
he type of lipid used in the preparation of water in oil emulsions
d recovery plots of GCO sunscreen and SCO sunscreen.
sunscreens influenced the skin adhesion ability of emulsions and,
consequently the water resistant performance. According to the lit-
erature (Draelos, 2011), the lipid material used in the preparation



J. Marto et al. / Industrial Crops and Products 80 (2016) 93–100 99

Table  6
In vitro and in vivo efficacy tests of the GCO and SCO sunscreen.

Formulations In vitrosun protection factor
(SPF)–optometrics SPF-290S Analyzer

In vitro sun product water resistance In vivosun product
water resistance

SPF Rating UVA/UVB UVA SPF after
1st
immersion

SPF after
2nd
immersion

%WRRa

after 1st
immersion

%WRRaafter
2nd
immersion

%WRRa

GCO sunscreen 82.3 ± 10.350+ 0.9 ± 0.1 71.1 ± 10.1 52.0 ± 14.2 42.1 ± 13.4 62.6 50.7 70.0 ± 2.2
SCO  sunscreen 51.9 ± 5.550 0.9 ± 0.1 44.2 ± 5.2 43.18 ± 6.88 41.95 ± 5.94 83.1 80.8 80.0 ± 3.6

a Water resistance retention.

Table 7
Skin adhesive properties of the formulations on dry skin and after 40 min  of water immersion (mean ± SD, n = 6).

Samples Work of skin adhesion (mJ/cm2) � of skin adhesion (mJ/cm2)a

GCO sunscreen on dry skin 0.05 ± 0.01 –
SCO  sunscreen on dry skin 0.05 ± 0.01 –
GCO  sunscreen after water immersion 0.01 ± 0.01 0.04
SCO  sunscreen after water immersion 0.04 ± 0.01 0.01

a Work of skin adhesion between sunscreens on dry skin and after water immersion.

F  and c
D  light

o
e
T
t
d
(
e
w
t
i
s

ig. 2. Histograms of skin whiteness resulting from sunscreens applied on dry skin
ark  grey–bare skin; grey–fresh sunscreen applications with 30 min  air drying; and

f w/o emulsions sunscreens influence the skin adhesion ability of
mulsions and consequently their water resistance performance.
he SCO extracted from roasted coffee is a brown viscous liquid due
o the presence of liposoluble Maillard reaction products separated
uring the oil extraction, enhancing its lipophilic characteristics
Speer and Kölling-Speer, 2006). Concerning the skin surface prop-
rties, it was demonstrated that dry skin is mostly lipophilic and

et skin is more hydrophilic, having higher surface energy. Thus,

he surface energy values of dry skin reflect its dominant lipophilic-
ty and more apolar materials, such as SCO, may also adhere to
kin.
ross-polarized images of two sunscreens applied to the volar forearm of a subject:
 grey - sunscreen after 40 min  water immersion.

3.2.6. HRIPT
Under the experimental conditions adopted, the repeated appli-

cations of the sunscreens under occlusive patch induced no
irritative reaction and the product has very good skin compatibility.
Moreover, the repeated applications induced no allergic reaction.

3.2.7. In vivo sun product water resistance

Human testing is considered to be the most acceptable method

for claiming water resistance. This method is a new in vivo screening
approach to measure WRR  using cross-polarized imaging. Although
it does not allow determining the exact SPF before and after the



1 ps and

i
t

f
e
a
a

4
t
i
G
h
b
u

4

b
t
p
A
p
a
b
w
t

i
u
d
p
p
m
b

t
i
t

A

n
S
a

R

A

A

A

B

B

B

00 J. Marto et al. / Industrial Cro

mmersion, it allows evaluating the amount of sunscreen lost due
o the action of water.

Skin whiteness results showed a similar behaviour as obtained
or water resistance results (Fig. 2). After water immersion, both
mulsions presented a range between 50 and 80% of whiteness with

 mean peak around 70% and 75% of whiteness for GCO sunscreen
nd SCO sunscreen, respectively.

GCO sunscreen and SCO sunscreen, when exposed to water for
0 min, revealed 70.0 ± 2.2% and 80.0 ± 3.6% of whiteness, respec-
ively (Table 6). In vivo WRR  values confirmed the results obtained
n in vitro assay, thus supporting the “water resistant” claim of the
CO and SCO sunscreens. Furthermore, these in vivo values were
igher than those obtained in vitro, which may  be explained by a
etter adhesion to human skin compared with the adhesive tape
sed for the in vitro assay.

. Conclusions

A novel sunscreen formulation with a high UVB/A protection,
iological activity and better tolerability was developed based on
he Pickering emulsions concept. The successful formulation was
ossible by combining natural and multifunctional compounds.
dditionally, the coffee oils studied in this work possess all of the
roperties required for sunscreens and may  be considered desirable
nd suitable ingredients for industrial applications. In addition to
eing a green product, it contains a series of lipophilic substances
ith important antioxidant characteristics, such as tocopherols,

hat protect the skin against UVB radiation.
When high oil contents are combined with a composition rich

n unsaturated acids, and compounds with high SPF value and high
nsaponifiable material content, the result is a product or ingre-
ient that is ideally suited for formulating high quality cosmetic
roducts able to promote moisture retention and simultaneously
rovide sun protection. All results revealed an excellent compro-
ise between stability, UV protection, rheological and mechanical

ehaviour, efficacy, safety and cosmeticity.
The use of spent and green coffee oil in cosmetic industry seems

o be a suitable approach to recycle and valorise wastes from coffee
ndustry. Moreover the coffee oil presented promising characteris-
ics in the improvement of sunscreen performance.

cknowledgments

This work was supported by the Fundaç ão para a Ciência e a Tec-
ologia, Portugal (UID/DTP/04138/2013 to iMed.ULisboa and grant
FRH/BDE/51599/2011), the strategic project UID/QUI/50006/2013
nd Laboratórios Atral S.A., Portugal.

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	The green generation of sunscreens: Using coffee industrial sub-products
	1 Introduction
	2 Material and methods
	2.1 Materials
	2.2 Methods
	2.2.1 Characterisation of the formulation ingredients
	2.2.1.1 Wettability measurements
	2.2.1.2 Particle size distribution
	2.2.1.3 Natural oils
	2.2.1.3.1 Oil extraction
	2.2.1.3.2 SPF measurement


	2.2.2 Sunscreen formulations—green coffee oil sunscreen (GCO sunscreen) and spent coffee oil sunscreen (SCO sunscreen)
	2.2.3 Characterization of the sunscreen formulations
	2.2.3.1 Droplet size distribution
	2.2.3.2 Structural analysis of the sunscreen formulations
	2.2.3.2.1 Rheology studies
	2.2.3.2.2 Texture profile analysis (TPA)

	2.2.3.3 In vitro SPF
	2.2.3.4 In vitro sun product water resistance
	2.2.3.5 Skin adhesion properties
	2.2.3.6 Human repeat insult patch test (HRIPT)
	2.2.3.7 In vivo sun product water resistance

	2.2.4 Statistical analysis


	3 Results and discussion
	3.1 Characterisation of the formulation ingredients
	3.1.1 Wettability measurements
	3.1.2 Particle size distribution
	3.1.3 Natural oils
	3.1.3.1 SPF measurement


	3.2 Characterization of the sunscreen formulations
	3.2.1 Droplet size distribution
	3.2.2 Structural analysis of the sunscreen formulations
	3.2.2.1 Rheology studies
	3.2.2.2 TPA

	3.2.3 In vitro SPF
	3.2.4 In vitro sun product water resistance
	3.2.5 Skin adhesion properties
	3.2.6 HRIPT
	3.2.7 In vivo sun product water resistance


	4 Conclusions
	Acknowledgments
	References