International Journal of Cosmetic Science, 2016, 38, 512–523
12320
Microneedle-mediated intradermal delivery of epigallocatechin-3-
gallate
A. Puri, H. X. Nguyen and A. K. Banga
Department of Pharmaceutical Sciences, College of Pharmacy, Mercer University, Atlanta GA 30341, USA
Received 22 October 2015, Accepted 20 March 2016
Keywords: epigallocatechin-3-gallate, formulation, microneedles, penetration, rheology, skin barrier
Abstract
OBJECTIVE: Epigallocatechin-3-gallate (EGCG) is the physiologi-
cally most active and abundant flavanol, accounting for 50–80%
of green tea catechins. It is an anti-inflammatory, antioxidant,
chemopreventive and skin photoprotective agent. However, light
sensitivity and low permeability of EGCG across the stratum cor-
neum due to its high molecular weight as well as strong binding to
the lipid bilayers in the skin make it difficult to be used as a key
ingredient in cosmetic products. This study aimed to formulate a
photostable hydrogel of EGCG with good rheological properties for
dermal application and investigate the effect of skin microporation
using maltose microneedles on its permeation through dermatomed
porcine skin.
METHODS: Effect of L-glutathione on photodegradation of EGCG
was investigated by exposing samples to ultraviolet irradiation for
1 h using a solar simulator. Hydrogels with varying concentrations
of Carbopol 980 (0.5–2% w/v) as an gelling agent were prepared,
and their rheological properties were evaluated using a rheometer.
Skin microporation was confirmed by assessing the skin resistance,
transepidermal water loss and calcein imaging of the microchan-
nels created by the microneedles. Permeation of EGCG from aque-
ous solution as well as the rheologically optimized hydrogel
through the dermatomed porcine skin (untreated and microneedle
treated) was evaluated using static vertical Franz diffusion cells.
RESULTS: L-glutathione acting as a co-antioxidant and photostabi-
lizer significantly (P < 0.05) reduced the degradation of EGCG from
21.53
2.78% to 1.0
0.68% after 1 h of ultraviolet irradiation.
Rotational and oscillatory rheological tests indicated that the hydro-
gel containing 1.5% Carbopol 980 is acceptable for topical applica-
tion in terms of flow behaviour, elasticity, spreadability, structural
stability and thixotropy. Microneedle-treated skin showed significant
enhancement (P < 0.05) in the delivery of EGCG to viable epidermis
and dermis from the aqueous solution (38.67
2.96 lg cm2) as
well as hydrogel (24.60
2.62 lg cm2) in comparison with the
untreated skin (24.16
2.11 and 15.62
0.24 lg cm2 for aque-
ous solution and hydrogel, respectively).
CONCLUSION: Addition of glutathione in EGCG formulations signifi-
cantly reduces its photodegradation. Skin microporation with maltose
microneedles facilitates the penetration of EGCG across the stratum
corneum into the deeper skin layers – viable epidermis and dermis.
Correspondence: Ajay K. Banga, Department of Pharmaceutical
Sciences, College of Pharmacy, Mercer University, Atlanta, GA 30341,
USA. Tel.: 678 547 6243; fax: 678 547 6423; e-mail: Banga_ak@
mercer.edu
512 © 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
doi: 10.1111/ics.
Re  sume 
OBJECTIF: Epigallocatechine-3-gallate (EGCG) est le flavanol phy-
siologiquement le plus actif et abondant, ce qui represente 50–80%
des catechines du the vert. C’est un agent anti-inflammatoire, anti-
oxydant, chimiopreventif et photoprotecteur de la peau. Cependant,
sa sensibilite a la lumiere et sa faible permeabilite a
 travers la cou-
che cornee en raison de son poids moleculaire eleve ainsi qu’une
forte liaison aux bicouches lipidiques de la peau, rendent difficile
son utilisation comme un ingredient cle dans les produits cosme-
tiques. La presente etude visait a  formuler un hydrogel photostable
de l’EGCG avec de bonnes proprietes rheologiques pour une applica-
tion cutanee et a  etudier l’effet de la microporation de la peau en
utilisant des micro-aiguilles de maltose pour sa permeation a tra-
vers la peau de porc dermatomee.
METHODES: L’effet de la L-glutathion sur la photodegradation de
l’EGCG a ete etudie en exposant des echantillons a  un rayonne-
ment ultraviolet pendant 1 h en utilisant un simulateur solaire.
Des hydrogels avec des concentrations variables de Carbopol 980
(0,5–2% p/v) comme agent gelifiant ont ete prepares et leurs pro-
prietes rheologiques ont ete evaluees en utilisant un rheometre.
La microporation de la peau a ete confirmee par l’evaluation de
la resistance de la peau, la perte insensible en eau et de l’image-
rie calceine des microcanaux crees par les microaiguilles. La
permeation d’EGCG a  partir d’une solution aqueuse, ainsi que de
l’hydrogel rheologiquement optimise a travers la peau de porc
dermatomee (non traitees et traitees aux microaiguilles) a ete
evaluee en utilisant des cellules de diffusion de Franz verticales
statiques.
RESULTATS: Le L-glutathion agissant en tant que co-antioxydant
et de photostabilisation, reduit de maniere significative (P < 0,05)
la degradation de l’EGCG de 21,53
2,78 a  1,0
0,68% apres
1 h d’irradiation par les ultraviolets. Les essais rheologiques rota-
tionnelles et oscillatoires ont indique que l’hydrogel contenant
1,5% Carbopol 980 est acceptable pour une application topique en
termes de comportement d’ecoulement, l’elasticite, l’etalement, la
stabilite structurelle, et thixotropie. La peau traitee aux micro-
aiguilles a montre une amelioration significative (P < 0,05) dans la
fourniture de l’EGCG a  l’epiderme viable et au derme a  partir de la
solution aqueuse (38,67
2,96 lg cm2), ainsi que de l’hydrogel
(24,60
2,62 lg cm2) par rapport a  la peau non traitee cutane
(24,16
2,11 et 15,62
0,24 lg cm2 pour une solution
aqueuse et d’un hydrogel, respectivement).
CONCLUSION: L’addition de glutathion dans les formulations de
EGCG reduit de maniere significative sa photodegradation. La
microporation de la peau avec les micro-aiguilles au maltose facilite
Intradermal delivery of epigallocatechin-3-gallate
 travers la couche cornee dans les cou-
la penetration de l’EGCG a
ches profondes de la peau: epiderme et du derme viables.
Abbreviations
EGCG: epigallocatechin-3-gallate
UV: ultraviolet
TEA: triethanolamine
RP-HPLC: reverse-phase high-performance liquid chromatography
CPB: citrate phosphate buffer
LVR: linear viscoelastic region
TEWL: transepidermal water loss
PPI: pore permeability index
SD: standard deviation
Introduction
Green tea, obtained as an extract from the leaves of Camellia sinen-
sis, is one of the most popular beverage consumed worldwide [1–
4]. Polyphenolic compounds known as catechins account for 30%
dry weight of green tea. Of all catechins, EGCG is the most abun-
dant, constituting about more than 50% polyphenolic fraction
[2,3,5–8]. It is also pharmacologically the most significant and
potent green tea catechin and has been reported to have antioxi-
dant, anti-inflammatory and anti-cancer properties [7,9,10].
Topical application of EGCG has been reported to protect the
skin against ultraviolet (UV) radiations and prevent photoageing
due to inhibitory effect on apoptosis of epidermal keratinocytes [3],
attenuation of UV-B-induced hydrogen peroxide production that
further inhibits the phosphorylation of mitogen-activated protein
kinase signalling pathways [5], reduction in expression and activ-
ity of matrix metalloproteinases preventing collagen degradation
[11], production of antioxidant enzymes and depletion of oxidative
enzymes [12]. Topical use of EGCG has also been demonstrated to
reduce the number of skin tumours (malignant as well as non-
malignant) in animal models [4,8,10,13]. Moreover, EGCG has
also been found to have exceptionally high preventive effect
against photocarcinogenesis in terms of tumour incidence, number
and size when applied topically [14]. It inhibits inflammation
[11,15] and immunosuppression caused by carcinogenic agents.
Mechanistically, this occurs due to induction of interleukin-12, an
immunoregulatory cytokine [14], apoptosis or cell growth arrest
by the activation of killer caspases, suppression of nuclear factor
kappa Beta activation and, overall, altered expression of cell cycle-
regulating proteins by EGCG [2]. Besides photoprotective and
chemopreventive effects, topical EGCG has also been explored for
other indications such as vitiligo [16], rosacea [17], antibacterial
infections [18] and telangiectasias [19].
Many topical green tea extract-based skincare cosmetic products
are available in the market such as face creams, lotions, soaps,
moisturizers and hydrogel masks [20]. VEREGENâ is a 15% green
tea ointment, approved by the United States Food and Drug Admin-
istration (US FDA) for the treatment of external perianal and geni-
tal warts [20]. However, topical formulations or products of EGCG,
the most active and potent catechin in green tea, as the active
ingredient are presently not available in the market.
© 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
International Journal of Cosmetic Science, 38, 512–523
A. Puri et al.
Many studies in the literature report low skin permeation of
EGCG (soluble in water, log P of 3.1, unstable in light) due to the
presence of gallate group that results in its high molecular weight
(458.40 Da) and also increases its binding to the lipid bilayers of
the stratum corneum [11,20–23]. An ideal antioxidant must, how-
ever, diffuse across the outermost barrier of skin, that is stratum
corneum, and reach the viable epidermis and dermis where the UV
radiations penetrate and have damaging effects [11,20,24]. An
in vivo study performed in human volunteers to investigate the per-
cutaneous penetration of EGCG from o/w emulsion and hydrogel
formulations showed diffusion of EGCG into the stratum corneum
[11]. Various strategies have been investigated to improve the pen-
etration of EGCG across stratum corneum such as emulsification
[20], use of terpenes as penetration enhancers [23] and by encap-
sulation in liposomes [21].
Microneedles are micron-sized structures designed to perforate
the stratum corneum and enable delivery of drugs into the epider-
mis or dermis [25]. Microporation using microneedles is a mini-
mally invasive technique that creates aqueous microchannels in
the upper layers of skin, enabling drugs to bypass the stratum cor-
neum and reach the deeper layers [26]. Microneedles range from
25 to 2000 lm in length and are a patient compliant means of
delivering therapeutic agents, in both micro- and macromolecules,
intra- as well as transdermally [27]. Different kinds of microneedles
have been previously demonstrated to enhance the cutaneous
delivery of 5-aminolevulinic acid [28], botulinum toxin A [29],
DNA vaccine [30], recombinant HIV-1 CN54gp140 [25] and
nanoparticles [26].
The overall objective of this study was to formulate a photo-
stable hydrogel of EGCG and assess its delivery through der-
matomed porcine ear skin using maltose microneedles. EGCG has
been reported to degrade markedly in the presence of light [31,32].
However, as photostability is an essential requirement for an effec-
tive topical antioxidant, one of our objectives was to reduce the
degradation of EGCG in light and formulate a photostable hydrogel.
Further, as evaluation of the rheological properties is essential to
formulate a topical product with good elasticity, spreadability,
structure stability, adhesiveness and thixotropic behaviour and is
also a determinant of diffusion and release of the drug from the for-
mulation, various rheological tests were conducted while formulat-
ing the hydrogel. Skin was treated with microneedles to create
microchannels before the hydrogel was applied to assess the perme-
ation and retention of EGCG in skin. To our knowledge, this study
would be the first one to report the topical delivery of EGCG
through skin, microporated using maltose microneedles, and pro-
vide a detailed picture of the distribution of EGCG in the different
skin layers.
Materials and Methods
Materials
EGCG and L-glutathione (reduced) were purchased from MedChem
Express LLC (Princeton, NJ, USA) and Sigma–Aldrich (St. Louis,
MO, USA), respectively. Citric acid, disodium hydrogen phosphate
and triethanolamine (TEA) were obtained from Fisher Scientific
(Fair Lawn, NJ, USA). Carbopol 980NF was procured from Lubrizol
Corporation (Wickliffe, Ohio, USA). Methanol and ethanol were
obtained from Pharmco-Aaper (Brookfield, CT, USA), and Fluore-
softâ (0.35%) from Holles Laboratories Inc. (Cohasset, MA, USA).
A local slaughter house (Atlanta, GA, USA) provided the porcine
513
Intradermal delivery of epigallocatechin-3-gallate A. Puri et al.

ear skin. Maltose microneedles (array of 27 identical needles,

500 lm long) were purchased from Elegaphy Inc. (Tokyo, Japan).
Photodegradation studies
Effect of UV irradiation on 1% (w/v) solution of EGCG in citrate
phosphate buffer (CPB) prepared using disodium hydrogen phos-
phate (Na2HPO47H2O, 126 mM), citric acid (49 mM) and pH
adjusted to 5.5 with o-phosphoric acid was studied. Photodegrada-
tion was evaluated with and without the addition of 0.5% w/v L-
glutathione to the EGCG solution using solar simulator (Newport,
Oriel Instruments, Stratford, CT, USA) equipped with a xenon–
ozone-free 150-Watt lamp. Solutions (50 lL) were placed in wells
(surface area, 0.5 cm2) and exposed to solar simulator for 1 h at
150 W. After exposure, samples were diluted 10 times with CPB,
vortexed and analysed using high-performance liquid chromatogra-
phy (HPLC) method. Results were reported as mean
SD (n = 3).
Rheological evaluation
Rheological analysis was performed using a rheometer (Rheoplus/
32 V3.62, Anton Paar Germany GmbH, Germany). Parallel-plate
spindle (PP 25/S) with the diameter of 24.987 mm was used, and
a gap of 100 lm was maintained between the plates. Rotational
and oscillatory tests were then performed on the hydrogels at a
temperature of 32°C to assess the flow behaviour and structural
stability of the gel formulations.
Flow curves
Gels were sheared at an increasing rate of 0–100 s1, and rheo-
grams of viscosity versus shear rate were plotted. Shear stress
values were also recorded. The data were fitted by the Herschel–
Bulkley model [35]:
s ¼ sy þ Kcn

Formulation of hydrogels
Hydrogels were prepared using different concentrations of Carbopol
980 NF as a gelling agent. Carbopol, in varying concentrations
(0.5, 1, 1.5, 2% w/v), was added to the solution of EGCG (1% w/v)
and glutathione (0.5% w/v) in CPB. The resulting mixture was
slowly stirred overnight at room temperature on a magnetic stirrer
(VWRâ, Radnor, PA, USA). After 24 h, the mixture was neutral-
ized with TEA to form a transparent gel matrix with pH of about
5.5 [33]. The pH was measured using a Denver InstrumentTM
Model 215 pH meter (Fisher Scientific, NJ, USA). TEA was preferred
over other bases such as sodium hydroxide for neutralization as it
forms gels with better transparency [34]. The symbols used for the
different hydrogel formulations have been mentioned in Table I.
where s represents shear stress (Pa), sy the yield stress (Pa), K the
flow consistency index (Pa.sn), c the shear rate (upto 100 s1) and
n the flow behaviour index (dimensionless). The values of yield
stress, flow consistency index and flow behaviour index were
recorded for all the hydrogels.
Amplitude sweep
The samples were exposed to an increasing strain of 1–100% at a
constant angular frequency of 10 rad s1. The values of the oscil-
latory shear responses, storage modulus (G0 ) and loss modulus (G″)
were plotted in a logarithmic scale against the increasing strain
(log scale). This test enabled determination of the linear viscoelastic
region (LVR) that was subsequently used to choose a strain value
to be applied for the frequency sweep and thixotropy oscillation
tests.

Frequency sweep
Table I Flow properties of hydrogels as calculated by Herschel–Bulkley Hydrogels were exposed to an increasing angular frequency (0.1–
model 100 rad s1) at a constant strain (1% for F0.5 and 5% for F1,
F1.5 and F2) as determined from the LVR of amplitude sweep
curves. The values of storage modulus (G0 ) and loss modulus (G″)
Carbopol Yield Flow were plotted against the increasing frequency using a log–log plot.
Formulation concentration stress consistency Flow
symbols (% w/v) (Pa) index (Pa.sn) index (n)
Thixotropy oscillation
An oscillatory thixotropy test was used to evaluate the structure
F0.5 0.5 8.05 2.71 0.42 breakdown and rebuilding behaviour of the hydrogel formulations
F1 1 32.92 7.98 0.40 upon application of high shear. Setting a constant strain (1% for
F1.5 1.5 39.21 56.29 0.23 F0.5 and 5% for F1, F1.5 and F2) as per the LVR of amplitude
F2 2 192.46 335.24 0.12
sweep curves, the gels were exposed to cyclic changes in shear
rate: 10 s1 for 100 s followed by 3000 s1 for 15 s and finally at
10 s1 for 100 s. The structure recovery ratio was recorded for all
the gel formulations.
Table II Specifications of the test groups evaluated in the in vitro perme-
ation study Preparation of skin samples
Full-thickness porcine ear skin, obtained from a local slaughter
Group code EGCG formulation Microneedle treatment house, was cleaned thoroughly using DI water and stored at
80°C till further use. For the experimental study, skin was
allowed to thaw at room temperature and washed with DI water,
SOL Solution No
SOL MN Solution Yes
and hair were cut carefully with scissors. It was then dermatomed
GEL hydrogel – F1.5 No using Dermatom 75 mm (Nouvag AG, Switzerland) to obtain skin
GEL MN hydrogel – F1.5 Yes pieces, approximately 0.6 mm thick. Thickness was measured
using a thickness gauge (Cedarhurst, NY, USA). The dermatomed

514 © 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
International Journal of Cosmetic Science, 38, 512–523
Intradermal delivery of epigallocatechin-3-gallate
skin was then cut into appropriate sizes (2 9 2 cm2) for mounting
on the vertical Franz diffusion cells.
Microneedle insertion and visualization of microchannels
An array of 500-lm long and sharp-tipped solid maltose micronee-
dles was used for treatment in the skin permeation studies. Each
array comprised of 3 straight parallel rows of 27 identical micro-
needles [36,37]. Pore uniformity, transepidermal water loss and
skin resistance were measured to confirm successful microporation
of skin and characterize the microchannels before using the micro-
needles for in vitro permeation studies.
Pore uniformity
Microneedles were inserted for 1 min in the skin, and microchan-
nels were visualized by calcein imaging and pore uniformity was
determined using Fluoropore image analysis tool. Fluoresoftâ solu-
tion (0.35%) was applied on the microporated skin for 1 min
after which the dye was wiped off with alcohol swabs and kim-
wipes. The protocol described previously by us was followed [38].
A fluorescent image was then taken two-dimensionally that
showed the distribution of fluorescent intensity within as well as
around each pore. This was further converted into pore perme-
ability index (PPI) which represented the calcein flux for each
channel.
Transepidermal water loss measurement
The objective of TEWL measurement was to assess the effect of
microneedle treatment on the barrier integrity of the porcine ear
skin as microneedles disrupt the stratum corneum and increase the
TEWL in comparison with the untreated skin [36,38]. TEWL was
thus measured before and after microneedle treatment, non-inva-
sively using a VapoMeter (Delfin Technologies Ltd, Kuopio, Finland)
in the ambient conditions of humidity and temperature after
mounting the skin on vertical Franz diffusion cells (PermeGear,
Hellertown, PA, USA) containing CPB in the receptor compart-
ment. The probe was placed still on the skin for 10 s, and a stable
value was displayed on the screen [36]. The TEWL values obtained
by pre- and post-microneedle treatment were recorded and com-
pared (n = 4).
Skin resistance evaluation
Skin resistance was measured before and after microneedle treat-
ment using silver/silver chloride electrodes, Agilent 33220A,
20 MHz function/arbitrary waveform generator and 34410A 6 ½
digit multimeter (Agilent Technologies, CA, USA). Skin was
mounted on the vertical Franz diffusion cells, and CPB was added
in the receptor as well as in the donor. Silver chloride electrode
was placed in the donor and silver in the receptor. Load resistor
(RL) was connected in series with the skin, and the drop-in voltage
across the complete circuit (VO) and across the skin (VS) was
recorded as displayed on the multimeter. Skin resistance (RS) was
calculated using the formula [39]:
RS ¼ VS RL =ðVO  VS Þ
where RL and VO were 100 kΩ and 100 mV, respectively.
Also, measurement of skin resistance of the intact skin (before
microneedle treatment) enabled us to assess the integrity of skin
barrier and select suitable skin samples for the permeation study.
Skin pieces with resistance lower than 17 kΩ cm2 were not
included in the study.
© 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
International Journal of Cosmetic Science, 38, 512–523
A. Puri et al.
In vitro skin permeation studies
Permeation of EGCG (n = 4), through dermatomed porcine ear
skin, with and without microneedle treatment was studied in vitro
using static vertical Franz diffusion cells that provided an effective
diffusion area of 0.64 cm2. Temperature of the receptor compart-
ment was maintained at 37°C using a water bath and an in-built
water circulation jacket around the receiver cells. This enabled
maintaining the temperature of the skin surface at 32°C through-
out the study. The cells were washed prior to the experiment and
filled with 5 mL CPB. Skin was mounted on the Franz cells with
the dermis facing the receptor and stratum corneum towards the
donor. For the microporation study, microneedles were inserted,
and TEWL as well as skin resistance was evaluated after mounting
the skin as described earlier. Formulations applied in the donor
chamber included 100 lL of EGCG solution in CPB (10 mg mL1)
and 100 mg of 1% EGCG (w/v) rheologically optimized hydrogel.
The donor chambers were covered with parafilm to avoid any
chances of oxidation of EGCG during the study [10,21]. The recep-
tor buffer (300 lL) was withdrawn at pre-determined time points
and replaced immediately with 300 lL fresh buffer solution. Sam-
ples were analysed using HPLC.
Extraction recovery study
The objective of this study was to determine the efficiency of the
procedure to be employed for the extraction of EGCG from der-
matomed porcine ear skin after separation of stratum corneum as
achieving 100% drug extraction is not practically feasible due to
chemical interactions between the drug and skin components.
Dermatomed porcine skin was tape stripped twenty times using
D-Squame Stripping DiscsD (Dallas, TX, USA) to separate the
stratum corneum from epidermis and dermis. After removal of
stratum corneum, epidermis and dermis were cut into small pieces.
Skin pieces were weighed, and 50 mg of skin was placed in
separate vials. EGCG standard solutions (100, 200, 500, 1000 and
2000 lg mL1) were prepared in ethanol, and 50 lL of each of
the solutions was added into respective vials containing the
weighed amount of skin (n = 3). The vials were first centrifuged at
13 148 g for 15 min at room temperature so as to ensure that all
the skin pieces were at the bottom of the vial and in complete con-
tact with ethanol to maximize drug absorption into skin layers and
then kept overnight in an incubator at 37°C. Ethanol (500 lL)
was then added to each vial and vortexed for 30 s to remove any
drug present on the skin surface or adhering to the walls of the
vials. Ethanol solutions were analysed by HPLC to calculate the
amount of EGCG actually taken up by the skin. The skin pieces
were then removed and placed individually from each vial in a 6-
well plate (Becton Dickinson, Franklin Lakes, NJ, USA), and the fol-
lowing extraction protocol was performed. Methanol (2 mL) was
used as the extraction solvent and added to each well. The plates
were kept on the shaker (New Brunswick Scientific Co. Inc., Edison,
NJ, USA) at 150 rpm for 4 h. Samples were then filtered through
0.22 lm syringe filters (Celltreat Scientific Products, Shirley, MA,
USA) and analysed using HPLC.
Extraction of EGCG from skin
After the 24-h permeation study, donor formulations were removed
first with two dry Q tips, and the skin was then cleaned properly
with two CPB-soaked Q tips and kimwipes. To measure the amount
515
Intradermal delivery of epigallocatechin-3-gallate
of EGCG retained in the skin layers, stratum corneum was sepa-
rated from the underlying epidermis and dermis using tape strip-
ping. Adhesive tapes, D101-D-Squame Stripping DiscsD, were
applied onto the permeation area of skin, one by one, pressed five
times manually with a finger, removed quickly with forceps and
collected in 6-well plates. The first two tapes were discarded to pre-
vent the overestimation of EGCG in stratum corneum due to any
drug remaining on the skin surface. Subsequent tapes 1–5, 6–10,
11–15 and 16–20 were collected separately. The remaining skin
(viable epidermis and dermis) was cut into small pieces using a sur-
gical scissors and placed in 6-well plate. Further, the extraction
protocol as described in the recovery study was followed. Similar
procedure was performed using blank dermatomed porcine skin
(n = 4) to investigate any background noise as well as interference
at the retention time of EGCG due to components of skin.
Quantitative analysis
Reverse-phase high-performance liquid chromatography (RP-HPLC)
based on UV detection was used for the quantitative estimation of
EGCG. Analysis was carried out using Waters Alliance 2695 sepa-
ration module (Milford, MA, USA) coupled with a 2996 photodiode
array detector. Gradient elution was performed on Waters Xbrid-
geTM C18, 50 9 4.6 mm, 3.5 lm (Waters Corporation, Milford,
MA, USA) column at a flow rate of 1.0 mL min1 and column
temperature of 35°C after injecting 30 lL of sample. The chro-
matographic conditions were as follows: methanol (phase A) and
0.1% v/v trifluoroacetic acid in DI water (phase B) from 5% to
45% A in 6 min, 45% to 80% A (6–6.5 min), 80% to 5% A (6.5–
8.5 min) and at 5% A up to 12 min for equilibration. The run time
was 12 min, and the retention time of EGCG was around 5.3 min.
Drug standards were prepared in CPB, and the detection wave-
length was 274 nm. The limit of detection and quantification was
0.74 lg mL1 and 2.25 lg mL1, respectively, and linearity was
observed in the concentration range of 0.1–50 lg mL1
(R2 = 0.998).
Data analysis
Data were analysed using Microsoft Excel Worksheets and the SPSS
software package version 21.0 (IBM, USA). Statistical analysis was
carried out using Student’s t-test (Graph pad software), and
P < 0.05 was considered for significant difference.
Results and discussion
Photodegradation studies
An ideal cutaneous photoprotective agent must possess adequate
stability upon exposure to solar irradiation [32]. Photodecomposi-
tion of such agents would depreciate their protective activity
against UV radiations [31]. Photochemical behaviour of EGCG has
been explored previously, and literature reports marked degrada-
tion of EGCG when exposed to sunlight [11,31,32]. Photodegrada-
tion of EGCG was tested in CPB (pH 5.5) which was further used
as the vehicle for its hydrogel formulation as EGCG has been
reported to exhibit sufficient chemical stability in aqueous solutions
of low pH, and it was also close to the pH of skin that would ren-
der it safe for dermatological application [10,11,31,32]. The study
investigated the effect of addition of 0.5% w/v L-glutathione on the
photodecomposition of EGCG.
516 © 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
A. Puri et al.
Exposure of 1% EGCG solution to solar radiations (300 W cm2)
was observed to degrade 21.53
2.78% EGCG after 1 h. Forma-
tion of EGCG dimers, the major oxidation products [40] upon pho-
tolysis of EGCG, has been reported earlier [32]. However, addition
of 0.5% L-glutathione significantly reduced the decomposition of
EGCG to 1.0
0.68%, when exposed to similar conditions of solar
irradiation (P < 0.05).
Different strategies have been previously investigated for photo-
stabilization of EGCG. These include addition of co-antioxidants
[31,32] and UV filters or sunscreen agents [31] to the EGCG for-
mulations. Vitamin E and butylated hydroxyl toluene were found
to enhance photodegradation of EGCG [31], whereas ascorbic acid
and alpha-lipoic acid successfully imparted photostabilization [32].
Also, addition of a UV-B filter, benzophenone-4, was found to sig-
nificantly stabilize EGCG in the presence of light [31].
The present study investigated the potential of glutathione, a
water-soluble antioxidant, as a photostabilizing agent for EGCG,
and it was found to significantly reduce its photolysis. A possible
explanation for the same can be attributed to the lower reactivity
and redox potential of glutathione that ranges from 0.35 V to
0.04 V [41] as compared to that of EGCG (0.43 V) [32] which pre-
vents the oxidation of EGCG and provides photoprotection. The free
sulfhydryl group present in glutathione provides nucleophilicity
due to which it acts as a reducing agent and itself transforms to
the oxidized form [42], protecting EGCG from oxidative degrada-
tion. Glutathione is a physiologic antioxidant [43], and its presence
in the formulation might provide additional antioxidant and photo-
protective activity as well besides stabilizing EGCG in the presence
of light.
Rheological evaluation of hydrogels
Aqueous polymers are commonly employed as thickening agents in
topical pharmaceutical and cosmetic products to improve their rhe-
ological properties [44,45]. Evaluation and knowledge of rheologi-
cal parameters of topical dosage forms are essential as they tend to
influence the microstructure responsible for the diffusion and
release of drugs from the formulation and can also be an indirect
measure of the same. Rheology can thus be used as an optimiza-
tion tool for topical drug delivery of dermatological formulations in
terms of physical stability, spreadability, thixotropy, time for which
the formulation stays on skin and drug release rates [44].
In this study, we used a carbomer polymer, Carbopol 980 NF, as
the gelling agent which is an acrylic acid polymer (anionic) cross-
linked with divinyl glycol [46]. It forms a gel in an aqueous med-
ium upon neutralization with organic amines or other bases such
as sodium hydroxide. Carbomers are widely used in the pharma-
ceutical products due to their thermal stability, mucoadhesive prop-
erty, lower concentrations resulting in high viscosity and good
compatibility with many active ingredients, and the resultant gels
produced have a characteristic flow behaviour, are aesthetically
elegant and patient compliant as well [44,47]. Carbomer grades
containing no residual benzene are toxicologically preferred, and
thus, their use is encouraged in topical formulations [48]. There-
fore, Carbopol 980NF, that is polymerized in co-solvent system of
cyclohexane and ethyl acetate, was used as a gelling agent in this
study. Our aim of characterizing the flow behaviour of EGCG
hydrogels as a function of Carbopol concentration was to determine
an optimum concentration that would produce physically stable
gels with good spreadability as well as structure recovery potential
after being sheared at high shear rates.
International Journal of Cosmetic Science, 38, 512–523
Intradermal delivery of epigallocatechin-3-gallate A. Puri et al.

(a) TAU (c) Rheoplus

3
10
F1.0
180 000
F1 Pa G''
cP F1.5 G'
2
10
140 000 F0.5 F1.5
120 000 F2 G'' G''
1
G'
100 000 10 F2
η 80 000
G''
G'
60 000 G'
0
10 F0.5
40 000
G''
20 000
G'
–1
0 10
0 10 20 30 40 50 60 70 80 90 1/s100 0.1 1 10 rad s–1 100
.
Shear rate (γ ) Angular frequency (ω )
Anton Paar GmbH Anton Paar GmbH

(b) CSD (d) 3ITT - OSC

3
10
F0.5
1000 F2
G'
Pa Pa G'
G''
100 G''
F1
G' 10 2
F1.5
G' G' 10 G'
G''
G''
F1.5 1 F1
G'
G'' 101 G'' G'
G'' 0.1
G''
F2
F0.5
G' 0.01
G'
0
G''
10 0.001 G''
1 10 % 100 0 20 40 60 80 100 120 140 160 180 s 200
Strain (γ ) Time (t)
Anton Paar GmbH Anton Paar GmbH

Figure 1 Comparative rheograms of hydrogels with different concentrations of Carbopol 980 corresponding to the following rheological tests: (a) flow curves, (b)
amplitude sweep, (c) frequency sweep and (d) thixotropy oscillation. F0.5: EGCG hydrogel with 0.5% (w/v) Carbopol; F1: EGCG hydrogel with 1% (w/v) Carbopol;
F1.5: EGCG hydrogel with 1.5% (w/v) Carbopol; F2: EGCG hydrogel with 2% (w/v) Carbopol; g: viscosity (cp); G0 : storage modulus (Pa); G0 0 : loss modulus (Pa).

Flow curves
The flow curves obtained for different hydrogels are shown in
Fig. 1a. Data were fitted and compared using the well-known Her-
schel–Berkley model applied using Rheoplus software. The parame-
ters (yield stress, consistency and flow index) obtained for all the
hydrogels are mentioned in Table I. All the gels showed pseudo-
plastic or shear-thinning behaviour as the values of flow index
were observed to be less than 1 and the viscosities decreased with
increasing shear rate. Pseudoplasticity is a desirable characteristic
for topical semisolid formulations as it eases application over larger
area utilizing a lesser dosage amount [49]. Further, with the
increase in percentage of Carbopol, viscosity, yield stress and con-
sistency values were found to increase. This depicted better entan-
glement of the polymeric chains and, thus, gel strength with
higher percentage of gelling agent. Minimum stress that should be
applied for the material to actually begin to flow is referred to as
yield stress and is an important rheological parameter for topical
gels. It signifies the extent of cross-linking, strength and stability of
the microgel structure, which was observed to increase with the
increase in percentage of Carbopol in this study [44].
Amplitude sweep
Amplitude or strain sweep experiments were carried out at a con-
stant oscillating frequency of 10 rad s1 and increasing strain of
1–100%. The oscillatory shear responses, storage modulus (G0 ) and
loss modulus (G″) were monitored. The storage or elastic modulus
is a representative of the elastic storage of energy and is therefore a
measure of rigidity of the material. The viscous or loss modulus, on
the other hand, is a measure of dissipation of viscous energy and is
large for predominantly viscous samples [47,50]. As shown in
Fig. 1b, both storage and loss modulus curves were observed for all
the formulations, confirming their viscoelastic nature. Moreover, at
each concentration of Carbopol, storage modulus was observed to
be greater than the loss modulus, indicating the samples being
more elastic than viscous which is a characteristic property of an
ideal gel [51]. It was also observed that an increase in concentra-
tion of Carbopol from 0.5% to 2% resulted in an increase in G0 and
G″. Thus, the gels F1.5 and F2 were better structured, more elastic
and viscous as compared to F0.5 and F1. A crossover point (G0 = G
″ = 2.42 Pa) at strain of 90% was observed for F0.5 that showed
its structural breakdown. However, no structural breakdown was
observed for F1, F1.5 and F2. Amplitude sweep curves were also
used to determine the LVR for all the formulations so as to obtain
the strain values at which the other investigative tests could be
conducted without disruption of the gel structure. Strain of 1% for
F0.5 and 5% for F1, F1.5 and F2 was obtained beyond which
changes in the structure of the gel were evident as depicted in
Fig. 1b. Polymers used in the formulations must also possess good

© 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie 517
International Journal of Cosmetic Science, 38, 512–523
Intradermal delivery of epigallocatechin-3-gallate
adhesiveness to be able to make contact with the surface and
spread. Adhesiveness is a combination of liquid-like properties that
enable the formation of good molecular contact when pressure is
applied as well as solid-like properties that provide resistance to
any applied stress after the bond between polymer and surface is
formed. Elastic modulus of less than 105 Pa is an indicator of good
adhesiveness of a formulation as it indicates ability to dissipate
energy due to viscous contributions as well as to deform to make a
good contact with the surface [51]. In our study, all the gels had
an elastic modulus of less than 1000 Pa, thus fulfilling the crite-
rion for good adhesion to skin surface.
Frequency sweep
These tests are useful to investigate the variations in responses to
the polymeric gels with the changes in applied frequency and are
mostly applicable for polymeric gel networks with imperfections as
these gels show variable performance with changes in frequency
[51]. As depicted in Fig. 1c, the measured storage and loss moduli
were observed as almost constant for all the formulations, indepen-
dent of the increase in frequency from 0.1 to 100 rad s1. This
ensured the formation of gels with no imperfections in the network.
Moreover, in concordance with the amplitude sweep tests, the
rheograms clearly showed an increase in G0 and G″ values with an
increase in concentration of Carbopol (0.5–2%). Thus, it was again
confirmed that the gels with 1.5% and 2% Carbopol were better
structured, more elastic and viscous in comparison with 0.5% and
1% Carbopol gels. However, the highest elasticity and viscosity
were observed for F2.
Thixotropy oscillation
The hydrogels were exposed to varying oscillatory shear rates of
10 s1 followed by 3000 s1 and finally 10 s1 at constant strain
(a)
(c)
Figure 2 (a) Fluorescent image of microneedle-treated skin with (b) pore permeability index values and (c) histogram generated using Fluoropore software.
518 © 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
A. Puri et al.
(1% for F0.5 and 5% for F1, 1.5 and F2). The changes in the stor-
age and loss moduli were recorded and have been illustrated in
Fig. 1d. Complete structural breakdown was observed for F0.5,
whereas the structure recovery ratios for F1, F1.5 and F2 were
100.52
0.44%, 108.30
1.91% and 111.15
1.62%, respec-
tively. Statistically insignificant difference (P > 0.05) was observed
between F1.5 and F2. However, structural recovery of F1.5 and F2
gels was found to be markedly (P < 0.05) higher than F0.5 and F1.
According to the amplitude and frequency sweep tests, F2 exhib-
ited the highest elasticity and gel strength in comparison with
F0.5, F1 and F1.5. However, thixotropy oscillation test provided a
better picture and depicted insignificant difference between F1.5
and F2 formulations in terms of thixotropic behaviour and struc-
ture recovery ability after being sheared at a high rate. Thus, over-
all, due to being well-structured, possessing high elasticity, physical
stability, better spreadability and structure recovery ability after
breakdown, both F1.5 and F2 were found to be rheologically
acceptable for application to skin [52]. However, according to the
inactive ingredients guide provided by the US FDA, 1.75% Car-
bomer polymer has been incorporated in the approved topical gel
formulations. Hence, F1.5 was selected for the in vitro permeation
studies [53].
Characterization of microchannels
Pore uniformity
After insertion of maltose microneedles, calcein was applied, and
due to its hydrophilic nature, it was taken up by the aqueous
microchannels. The fluorescence hence produced was captured by
the fluorescence microscope as shown in Fig. 2a,b. Fluoropore soft-
ware was used to analyse the fluorescent images, and it provided
the PPI value of 12.7
6.14 for the 73 micropores created by the
(b)
International Journal of Cosmetic Science, 38, 512–523
Intradermal delivery of epigallocatechin-3-gallate A. Puri et al.

microneedles on the basis of the fluorescent intensity in and
around each pore. A histogram was also generated depicting rela-
tively uniform distribution of pores (Fig. 2c). The results obtained
were similar to those reported previously by us for pores created
using maltose microneedles [37].
TEWL measurement
As shown in Fig. 3, an increase in TEWL was observed after treat-
ment with microneedles (22.2
2.24 g m2 h1) as compared to
the base value of pre-treatment (8.78
1.22 g m2 h1)
(P < 0.05). Normal, intact or unbreached skin generally loses very
small amount of water. However, the loss is more in case of com-
promised skin (disrupted stratum corneum) [38]. Thus, an increase
in TEWL is a measure to assess the formation of microchannels
using microneedles as it is an indicator of the barrier integrity of
skin. Our results for TEWL, substantiated with the visualization of
pores by calcein imaging, confirmed the formation of micropores in
the dermatomed porcine skin that was to be used for in vitro per-
meation studies.
Skin resistance
Intact skin offers resistance to microneedle insertion. Therefore,
greater force is required to overcome the resistive force and porate
the skin. Skin disruption by microneedles results in a significant
drop-in electrical resistance. The resistance of dermatomed porcine
skin as measured before microneedle treatment was found out to
be 30.07
14.03 kΩ cm2. However, it reduced 35 times to
0.82
0.18 kΩ cm2 after microneedle treatment (P < 0.05) as
shown in Fig. 3, confirming the formation of microchannels and
also corroborating the results of calcein imaging and TEWL mea-
surement. Similar results have also been reported earlier in the
literature [54,55].

Skin resistance TEWL

50.00 30.00
45.00 *
*
SKIN RESISTANCE (KΩ cm–2) ± SD

25.00
40.00

TEWL (g m–2 h–1) ± SD

35.00
20.00
30.00
25.00 15.00
20.00
10.00
15.00
10.00
5.00
5.00
0.00 0.00
PRE-MICRONEEDLE INSERTION POST-MICRONEEDLE INSERTION
Groups

Figure 3 Transepidermal water loss (TEWL) and skin resistance values, pre- and post-microneedle insertion in dermatomed porcine skin. n = 4. *Statistical
difference from other group (Student’s t-test, P value < 0.05).

30.00
y = 0.2449x – 1.2797
AVERAGE AMOUNT OF EGCG EXTRACTED

R² = 0.9919
25.00
FROM SKIN (μg) ± SD

20.00

15.00

10.00

5.00

0.00
0.00 20.00 40.00 60.00 80.00 100.00
AMOUNT OF EGCG ADDED IN SKIN (μg)

Figure 4 Extraction recovery of EGCG from viable epidermis and dermis.

© 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie 519
International Journal of Cosmetic Science, 38, 512–523
Intradermal delivery of epigallocatechin-3-gallate A. Puri et al.

solution which was explained due to strong chemical interaction of

In vitro skin permeation studies
Permeation of EGCG from aqueous solution as well as rheologically
optimized hydrogel (F1.5) through dermatomed porcine skin with
and without microneedle treatment was evaluated (n = 4). The
codes for different test groups have been defined in Table II.
No EGCG was detected in the receptor compartment even after
24 h in all the test groups. This indicated the inability of EGCG to
permeate across the skin passively as well as through the micro-
needle-treated skin. The results were consistent with the previous
studies carried out to investigate the skin permeation of EGCG
[10,20,21,23]. Fang et al. [21] encapsulated EGCG in a liposomal
formulation consisting of anionic surfactants and ethanol and eval-
uated its permeation with aqueous solution as a control using mice
skin. The authors reported ‘zero flux’ of EGCG from the aqueous
EGCG with the lipid bilayers of the skin. A slight increase in flux
(0.17
0.04 nmol cm2 h1) was observed with the liposomal
formulation which might have been observed as mice skin is highly
permeable [56]. This could hence be one of the reasons for no per-
meation of EGCG observed through microneedles as porcine skin is
less permeable than the mice skin used in other studies [21]. Also,
it may be possible that any small amount of EGCG if permeated
could have been below the limit of detection of the HPLC method
(0.74 lg mL1).
Another study was performed to explore the effect of a- terpineol
on skin permeation of EGCG in rats using subcutaneous microdialy-
sis [23]. No EGCG was found to permeate through a- terpineol-trea-
ted skin also that was explained due to its high molecular weight,
the presence of galloyl groups rendering it lipophilic and resulting

45 SOL SOL MN GEL GEL MN

AVERAGE AMOUNT OF EGCG (μg cm–2) ± SD

40

35

30

25

20

15

10

0
TAPES 1–5 TAPES 6–10 TAPES 11–15 TAPES 16–20 TOTAL

TAPE STRIPS

Figure 5 Distribution of EGCG in different layers of stratum corneum of dermatomed porcine skin using tape stripping technique. SOL: EGCG solution without
microneedle treatment; SOL MN: EGCG solution with microneedle treatment; GEL: EGCG hydrogel containing 1.5% (w/v) Carbopol without microneedle treat-
ment; GEL MN: EGCG hydrogel containing 1.5% (w/v) Carbopol with microneedle treatment.

*
AVERAGE AMOUNT OF EGCG (μg cm–2) ± SD

45.00
40.00
35.00 *
30.00
25.00
20.00
15.00
10.00
5.00
0.00
SOL SOL MN GEL GEL MN

TEST GROUPS

Figure 6 Average amount of EGCG in viable epidermis and dermis of dermatomed porcine skin after 24 h permeation. SOL: EGCG solution without micronee-
dle treatment; SOL MN: EGCG solution with microneedle treatment; GEL: EGCG hydrogel containing 1.5% (w/v) Carbopol without microneedle treatment; GEL
MN: EGCG hydrogel containing 1.5% (w/v) Carbopol with microneedle treatment. n = 4. *Statistical difference from other group (Student’s t-test, P value
< 0.05).

520 © 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
International Journal of Cosmetic Science, 38, 512–523
Intradermal delivery of epigallocatechin-3-gallate
in strong binding to the lipid bilayers of stratum corneum and
retention in skin [23]. Yoshino et al. [20] reported 90% binding of
EGCG to the skin tissues and further explained that the gallate moi-
ety in its structure increases the number of phenolic groups as well
as its molecular weight and enhances its binding to skin. Our
observations of undetectable levels of EGCG in the receptor com-
partment were thus in concordance with the above-mentioned
studies. However, we also determined the amount of EGCG in the
stratum corneum and underlying layers (viable epidermis and der-
mis).
Recovery studies
As shown in Fig. 4, the amount of EGCG added to the skin (epider-
mis and dermis) was plotted against the amount of EGCG
extracted/recovered with the method used for extraction as
explained earlier. The amounts of EGCG obtained in the epidermis
and dermis after the permeation experiment were corrected using
the equation obtained from the correlation plot in Fig. 4. The
extraction efficiency determined was low (24–29%) for the method
used in the study. However, these results were similar to those
reported by Zillich et al., where 19% of extraction efficiency for
EGCG from porcine skin was observed using a 90 : 10 (% v/v) mix-
ture of methanol–water mixture containing 0.2 g L1 vitamin C as
the extraction solvent. This method showed maximum recovery for
all the tea catechins except for EGCG that was explained due to its
oxidative instability or strong chemical binding with the skin tis-
sues – lipids and proteins [57] which might also be the same rea-
son for low recovery of EGCG observed in this study as well.
Extraction of EGCG from skin
The amount of EGCG was analysed separately in stratum corneum
and underlying layers (viable epidermis and dermis) using the tape
stripping technique. Twenty tape strips have been demonstrated to
completely remove the stratum corneum from porcine skin in the
literature [24,58,59]. No background noise as well as interfering
peaks at the retention time of EGCG was observed in the blank
skin.
The total amount of EGCG retained in the stratum corneum of
the different test groups, SOL, SOL MN, GEL and GEL MN, was
found to be 28.59
12.40, 29.62
2.37, 8.71
1.07 and
8.67
0.21 lg cm2, respectively (Fig. 5). There was no signifi-
cant difference between the total amount of EGCG deposited in the
stratum corneum of the intact skin and microneedle-treated groups
for both the solution (SOL vs. SOL MN) and gel formulation (GEL
vs. GEL MN) (P > 0.05). Distribution of EGCG in the different layers
of stratum corneum has also been illustrated in Fig. 5. The tapes
1–5, 6–10, 11–15 and 16–20 were pooled separately so as to
achieve a quantifiable amount of drug. As shown in the bar graph
representation, the amount of EGCG gradually reduced from the
outer layers (tapes 1–5) to the deeper layers (tapes 16–20) of the
stratum corneum with statistically significant difference between
the drug amount in these layers (P < 0.05). This trend was
observed for all the test groups. However, no significant difference
was observed in the drug distribution in different layers of stratum
corneum between SOL and SOL MN as well as GEL and GEL MN
groups. Therefore, microneedles did not affect the retention or dis-
tribution of EGCG in the stratum corneum.
Figure 6 shows the amount of EGCG analysed in viable epider-
mis and dermis for all the test groups after being corrected using
© 2016 Society of Cosmetic Scientists and the Societe Francßaise de Cosmetologie
International Journal of Cosmetic Science, 38, 512–523
A. Puri et al.
the equation obtained from the recovery study (Fig. 4) accounting
for the efficiency of the extraction procedure. The amount of EGCG
observed in the passive permeation test groups, SOL and GEL, was
24.16
2.11 and 15.62
0.24 lg cm2, respectively. Significant
(P < 0.05) increase in the deposition of EGCG in epidermis and der-
mis was observed in the microneedle-treated groups, SOL MN
(38.67
2.96 lg cm2) as compared to SOL and GEL MN
(24.60
2.62 lg cm2), in comparison with GEL group. This
showed enhancement in delivery of EGCG to the deeper skin layers
with the use of microneedles which are ideally the sites of action
for its antioxidant, photoprotective and chemopreventive activities.
The enhanced skin delivery of EGCG by maltose microneedle treat-
ment was supported by the measurement of TEWL, skin resistance
and PPI that depicted successful piercing of the stratum corneum
by the microneedles into the underlying layers and creating chan-
nels for the movement of EGCG across the barrier layer of the skin.
Also, as reported in our earlier studies, maltose microneedles with
500 lm length have been observed to create about 150- to 160-
lm-deep microchannels [36,38]. Due to aqueous solubility, EGCG
was able to pass through the hydrophilic microchannels, but at the
same time due to lipophilicity rendered by the gallate group and
very high binding capacity to the skin tissues [20,21,23], it was
restricted to epidermis, dermis and stratum corneum and was not
observed to reach the receptor. Moreover, Jackson et al. [60]
reported high binding affinity of EGCG for collagen due to hydrogen
bonding as well as hydrophobic bonding of the galloyl group with
the collagen network. This could thus be one of the possible rea-
sons for EGCG binding to the dermis.
Similar results have been reported earlier where encapsulation
in liposomes [21] and a-terpineol [23] enhanced the skin deposition
of EGCG mainly due to its binding to lipid bilayers of stratum cor-
neum. However, these studies did not analyse the drug separately
in stratum corneum and underlying layers. The present investiga-
tion highlighted specifically the delivery of EGCG to deeper skin lay-
ers besides stratum corneum. However, significantly lower amount
of EGCG was observed in the stratum corneum, epidermis and der-
mis from the gel formulation when compared to the solution (GEL
vs. SOL and GEL MN vs. SOL MN) (P < 0.05). This would have
possibly been due to high viscosity and entrapment of drug parti-
cles in the three-dimensional gel matrix that tends to decrease the
drug release from the formulation itself.
Our results for enhancement in skin deposition of EGCG with the
use of microneedles are consistent with various other studies in
which microneedle devices have been reported to enhance intrader-
mal delivery of 5-aminolevulinic acid [28], botulinum toxin A [29],
DNA vaccine [30], recombinant HIV-1 CN54gp140 [25] and
nanoparticles [26].
Conclusion
This study showed significant enhancement in the delivery of EGCG
from photostable aqueous solution as well as hydrogel to deeper
skin layers (viable epidermis and dermis) with the use of maltose
microneedles. L-glutathione (0.5% w/v) was used as the photostabi-
lizer as photostability is a prerequisite for an ideal and efficient topi-
cal antioxidant and photoprotective agent. EGCG hydrogel
formulated using 1.5% w/v Carbopol as a gelling agent was found
to be rheologically stable and acceptable for topical application. In
the in vitro permeation studies using dermatomed porcine skin,
EGCG was not detected in the receptor compartment. However, its
delivery to the deeper skin layers from the solution as well as gel
521
Intradermal delivery of epigallocatechin-3-gallate
formulation was significantly enhanced with the use of maltose
microneedles in comparison with the untreated skin. As epidermis
and dermis are the ideal sites for the pharmacological effects of
EGCG, enhancement in its intradermal delivery mediated by micro-
needles seems to have a bright future for its use as an anti-photoa-
geing and chemopreventive agent in cosmetic products.
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International Journal of Cosmetic Science, 38, 512–523