Short Communication
Skin Pharmacol Physiol 2010;23:113–116 Received: March 30, 2009
DOI: 10.1159/000265682
Pre-Treatment with Aloe vera Juice
Does Not Enhance the in vitro Permeation
of Ketoprofen across Skin
L. Ballam C.M. Heard
Welsh School of Pharmacy, Cardiff University, Cardiff, UK
Key Words
Aloe vera ⴢ Ketoprofen ⴢ Transdermal delivery
Abstract
Background/Aims: The potential of pre-treating skin with
Aloe vera juice as a penetration enhancer was evaluated in
vitro using ketoprofen as model permeant. Methods: To ex-
cised porcine skin mounted in Franz diffusion cells was ap-
plied either: (1) commercial Aloe vera; (2) commercial Aloe
vera followed by massaging; (3) previously boiled commer-
cial Aloe vera; (4) water (negative control); (5) tea tree oil (pos-
itive control). After 1 h, the pre-treatment was removed and
the skin dosed with a saturated solution of ketoprofen in
polyethylene glycol 400; the appearance of drug in the re-
ceptor phase was then monitored by HPLC. Results: No sta-
tistically significant differences in the transdermal delivery
of ketoprofen were observed between water and all the Aloe
vera pre-treatments (p 1 0.05). The tea tree oil pre-treatment
was significantly different to all others (p ! 0.05). Conclu-
sion: Aloe vera appears to have no value as a penetration
enhancer when used as a pre-treatment, although the data
indirectly support the mechanism of action proposed previ-
ously, work when used ‘within-vehicle’. Handling household
products containing Aloe vera appears not to leave the user
at elevated risk of subsequent absorption of exogenous
chemicals. Copyright © 2009 S. Karger AG, Basel
© 2009 S. Karger AG, Basel
1660–5527/10/0232–0113$26.00/0
Fax +41 61 306 12 34
E-Mail karger@karger.ch Accessible online at:
www.karger.com www.karger.com/spp
Accepted after revision: August 20, 2009
Published online: December 14, 2009
Introduction
Aloe vera juice, obtained from the succulent leaves of
Aloe barbadensis Miller, has been recognised since an-
cient times as a natural healing agent [1]. By virtue of its
purported ‘skin-friendly’ properties, it is to be found in
an increasing number of household products, including
washing up liquid and the inner linings of disposable
gloves. Delivering drugs into or across the skin is an im-
portant and commonly used route for treatment of a wide
range of conditions. Due to the generally low permeabil-
ity of the skin to most therapeutic agents, various meth-
ods of chemical penetration enhancement have been in-
vestigated to facilitate drug molecule transport across the
skin [2]. However, concerns about toxicity from the use
of chemical enhancers have led to interest in the use of
natural products, and Aloe vera is of potential interest due
to its skin compatibility.
The potential of Aloe vera as a penetration enhancer
was claimed in two United States patents [3, 4]. More re-
cently, a selective mechanism for skin penetration en-
hancement by Aloe vera was proposed based upon size
exclusion and the pull effect [5], involving the application
of infinite doses of saturated solutions of caffeine, colchi-
cine, mefenamic acid, oxybutynin, and quinine applied
to excised porcine skin. Pre-treating skin can be effective
in allowing enhancing processes to modulate the skin
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Charles M. Heard
Welsh School of Pharmacy, Cardiff University
University of Pittsburgh
Cardiff, CF10 3XF (UK)
Tel. +44 2920 875 819, Fax +44 2920 874 149, E-Mail heard @ cf.ac.uk
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barrier prior to dosing with drug. Furthermore, with
Aloe vera featuring in an ever growing list of domestic
products, it is important to know if the use of such prod-
ucts is liable to increase the risk of subsequent absorption
of other exogenous compounds. In the current work, ex-
cised porcine skin membranes were subjected to a num-
ber of pre-treatments involving Aloe vera juice, and the
effects of the permeation of ketoprofen examined.
Materials and Methods
Materials
Aloe vera juice was purchased from HealthWize (Wilmington,
Del., USA). Ketoprofen, tea tree oil, potassium phosphate and PBS
sachets were purchased from Sigma-Aldrich (Poole, UK). High
vacuum grease was obtained from Dow Corning (Barry, UK) and
HPLC grade acetonitrile was purchased from Fisher Scientific
(Loughborough, UK). All other reagents were of analytical grade
or equivalent. Freshly excised ‘large white’ pig ears, obtained from
a local abattoir prior to steam cleaning, were cleaned under run-
ning water and the hairs removed using electric clippers. Full
thickness skin was removed from the dorsal side of the ear, cut
into approximately 2 ! 2 cm sections and carefully inspected us-
ing a magnifying lens to ensure no holes or other imperfections
were present, prior to being stored in sealed aluminium foil at
–20 ° C until required (⬃24 h).
Skin Permeation Experiments
In vitro skin permeation was modelled using Franz-type dif-
fusion cells incorporating pig ear skin as a model for human skin
[6]. Donor variability was eliminated by equal distribution of the
skin between experiments, with at least ears from 2 animals used
per treatment. The cell flanges were greased and the skin placed,
stratum corneum uppermost, between the two cell chambers,
which were then clamped. The receptor chambers were filled with
receptor phase (degassed PBS, pH 7.4) and microstirrer bars add-
ed. The sampling arms were occluded using glass caps and the
complete cells placed on a magnetic stirrer plate (Variomag, Day-
tona Beach, Fla., USA) submersed in a Clifton Unstirred water
bath at 37 ° C. After 15 min, the pre-treatment step was per-
formed.
Several procedures were examined for pre-treatment with
Aloe vera. Firstly, the Aloe vera juice was applied directly from the
original container. Secondly, Aloe vera was rubbed into the skin
by 20 circular motions with a glass rod immediately after applica-
tion. Thirdly, Aloe vera had been boiled for 30 min on a hot plate
and cooled to room temperature before application. Pre-treat-
ment with de-ionised water and tea tree oil, Melaleuca alternifolia
[7], served as negative and positive controls, respectively. In each
case, 500 ␮l of solution was used and removed by aspiration after
1 h. The cells were then dosed with 500 ␮l of ketoprofen in PEG-
400 saturated at 32 ° C with ketoprofen. The donor chambers were
occluded with parafilm. The whole receptor phase was removed
at pre-set times over 48 h and 1 ml samples retained at –20 ° C for
analysis. The diffusion cells were refilled with fresh receptor
phase, equilibrated to 37 ° C, at each time point.
114 Skin Pharmacol Physiol 2010;23:113–116
HPLC Analysis of Ketoprofen
Ketoprofen was quantified using a Hewlett Packard 1100
HPLC system, fitted with a Kingsorb C18 column, 150 ! 4.6 mm,
5 ␮m (Phenomenex, Macclesfield, UK). The mobile phase was
acetonitrile 45% v/v, potassium phosphate buffer (pH 1.5) 55%
v/v. The flow rate was 1.0 ml ⴢ min–1 and ultraviolet detection car-
ried out at 258 nm. Standard solutions were prepared in receptor
phase solution over the range 1–500 ␮g ⴢ ml–1 for ketoprofen and
calibration curves were constructed, providing an R 2 value of
60.999. The retention time of ketoprofen was 6.1 min and the
limit of detection was 0.03 ␮g ⴢ ml–1. The assay was robust, with
coefficients of variation 0.8% (intra-day) and 0.9% (inter-day).
Data Analysis
The cumulative amounts of ketoprofen permeated over the
48-hour period (␮gⴢcm–2) were plotted against time and a least
squares regression was fitted to the linear section of the curve
(between 6–24 h) using Microsoft Excel 2000 to provide steady-
state flux (Jss); dividing Jss by the donor concentration gave per-
meability coefficient (kp). One-way analysis of variance (ANO-
VA) test was employed and followed by Tukey’s post-hoc tests
(p ! 0.05) to identify any significant differences between treat-
ments using Instat 3 for Macintosh.
Results and Discussion
Typical permeation profiles were produced in every
case (not shown) and the main permeation parameters
are listed in table 1. Water comprises the bulk of Aloe vera
juice and thus is the logical comparator (negative control)
when examining the effects of Aloe vera as a potential
penetration enhancer. Water as a pre-treatment provided
a mean steady state flux of ketoprofen of 5.18 ␮g ⴢ cm–2 ⴢ
h–1. When Aloe vera juice was used to pre-treat the skin,
the flux was slightly higher at 7.7 ␮g ⴢ cm–2 ⴢ h–1; however,
table 2 shows that the difference was statistically insig-
nificant (p 1 0.05). Massaging the Aloe vera into the skin
can assist in loading a solution into the upper layers of the
skin and also mimics the massaging of a formulation into
the skin, as is commonly carried out in clinical practice,
e.g. when applying a cream. This can result in higher de-
livery. However, the flux obtained was 5.39 ␮g ⴢ cm–2 ⴢ h–1
and again statistically insignificant compared to water
(table 2). A portion of the Aloe vera juice was boiled in
order to de-activate any proteolytic and other enzymes
present that are potentially capable of modulating skin
and transdermal delivery. When used as a pre-treatment,
a flux of 5.12 ␮g ⴢ cm–2 ⴢ h–1 was observed – again statisti-
cally not different to pre-treatment with water. Similarly,
consideration of cumulative permeation at 24 h revealed
no statistical differences (table 3). Regarding the positive
control, the flux of ketoprofen was ⬃7.5 times greater
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Ballam/Heard
University of Pittsburgh
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Table 1. Steady-state flux (Jss), permeability coefficient (kp), lag time and cumulative permeation after 24 h (mass, Q24; percentage,
%24) data for the in vitro permeation of ketoprofen across porcine skin with different pre-treatments (n = 5)

Pre-treatment Dose Jss SD kp Lag time SD Q24 SD %24

␮g ␮g ⴢ cm–2 ⴢ h–1 !10–4 cm ⴢ h–1 h ␮g ⴢ cm–2

Water 24,683 5.19 1.31 2.10 3.09 0.44 77.2 13.0 0.50
Aloe vera 24,683 7.70 1.80 3.12 3.24 0.36 95.5 19.4 0.87
Aloe vera, rubbed 24,683 5.39 2.26 2.19 4.12 0.59 65.7 21.2 0.59
Aloe vera, boiled 24,683 5.12 4.42 2.07 4.19 0.96 69.2 14.1 0.58
Tea tree oil 24,683 38.4 11.20 15.5 3.09 0.30 529 140.0 3.11

Table 2. Statistical analysis of steady-state flux data for the in vit- Table 3. Statistical analysis of cumulative permeation, Q24, data
ro permeation of ketoprofen across porcine ear skin subjected to for the in vitro permeation of ketoprofen across porcine ear skin
different pre-treatments subjected to different pre-treatments

Comparison Mean q p value Comparison Mean q p value

difference difference

Water vs. AV –2.515 1.025 >0.05, NS Water vs. AV –18.300 0.6345 >0.05, NS
Water vs. AV rubbed –0.2090 0.08522 >0.05, NS Water vs. AV rubbed 11.500 0.3988 >0.05, NS
Water vs. AV boiled 0.07000 0.02854 >0.05, NS Water vs. AV boiled 8.000 0.2774 >0.05, NS
Water vs. tea tree oil –33.175 13.527 <0.001*** Water vs. tea tree oil –451.80 15.666 <0.001***
AV vs. AV rubbed 2.306 0.9402 >0.05, NS AV vs. AV rubbed 29.800 1.033 >0.05, NS
AV vs. AV boiled 2.585 1.054 >0.05, NS AV vs. AV boiled 26.300 0.9119 >0.05, NS
AV vs. tea tree oil –30.660 12.501 <0.001*** AV vs. tea tree oil –433.50 15.031 <0.001***
AV rubbed vs. AV boiled 0.2790 0.1138 >0.05, NS AV rubbed vs. AV boiled –3.500 0.1214 >0.05, NS
AV rubbed vs. tea tree oil –32.966 13.441 <0.001*** AV rubbed vs. tea tree oil –463.30 16.065 <0.001***
AV boiled vs. tea tree oil –33.245 13.555 <0.001*** AV boiled vs. tea tree oil –459.80 15.943 <0.001***

One-way analysis of variance: a p value <0.0001 is considered
extremely significant (***). Variation among column means is
significantly greater than expected by chance.
Tukey-Kramer multiple comparisons test: if the value of q is
>4.232 then the p value is <0.05.
Water = Negative control; AV = Aloe vera; AV rubbed = Aloe
vera rubbed into skin; AV boiled = Aloe vera boiled beforehand;
tea tree oil = positive control.
One-way analysis of variance: a p value <0.0001 is considered
extremely significant (***). Variation among column means is
significantly greater than expected by chance.
Tukey-Kramer multiple comparisons test: if the value of q is
>4.232 then the p value is <0.05.

cipient. Therefore, thermodynamic activity and donor

when the skin was pre-treated with tea tree oil, delivering
some 3% of the applied dose of ketoprofen after 24 h (ta-
ble 1). Against this control, all ketoprofen fluxes from
Aloe vera and water pre-treatments were statistically sig-
nificantly different (p ! 0.05), delivering approximately
0.6% of the applied dose after 24 h. Table 1 shows the lag
time data, and although some small differences are ap-
parent, ANOVA analysis revealed no statistical differenc-
es within the data set (p = 0.0673, considered not quite
significant; post-tests were not calculated).
All cells were dosed using the same saturated solution
of ketoprofen in PEG400, which is a non-enhancing ex-
concentrations were the same, and any differences ob-
served would be attributable solely to modulation of the
skin barrier function by the Aloe vera pre-treatment. Tea
tree oil is comprised of a mixture of lipophilic organic
terpenoids, mainly terpinen-4-ol, and provided substan-
tial enhancement by virtue of modulating the skin bar-
rier, e.g. lipoidal domains of the stratum corneum [8],
although 1,8,-cineole has been proven to readily perme-
ate skin [9], possibly as a consequence of leaching skin
lipids. It was recently reported that up to 2–4% of tea tree
oil permeated skin [10]. On the other hand, Aloe vera is
largely water in composition and generally immiscible
with skin lipids, although elevated hydration of the skin
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Aloe vera Does Not Enhance Permeation Skin Pharmacol Physiol 2010;23:113–116 115
of Ketoprofen across Skin
University of Pittsburgh
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is known to increase permeability [11] via phase changes
in the lipid domains and corneocyte swelling [12]. It is
possible that with a pre-treatment time of 1 h the skin in
each case, with the exception of tea tree oil, will have be-
come equally hydrated with potentially no further en-
hancement possible, despite the solutes contained within
Aloe vera.
Permeation enhancement by Aloe vera was previously
observed for certain molecules from infinite doses of sat-
urated solutions applied directly onto the skin, without
any pre-treatment [5]. The results were in part attributed
to interactions between the solute drug and the complex
array of phytochemicals within Aloe vera, i.e. within the
liquid vehicle, as suggested by the high amounts of other
Aloe vera components permeating through to the recep-
tor phase. In the current work, the Aloe vera pre-treat-
ment was removed after 1 h, thus the ketoprofen was not
interacting with Aloe vera phytochemicals in the same
manner, and this may go some way to explaining the lack
of enhancement observed in the current work. However,
it is also possible that the PEG-400 may have re-solu-
bilised the enhancing compounds within Aloe vera re-
maining in the skin after pre-treatment, restoring the
barrier function to its pre pre-treatment state. Either way,
the skin was rendered no more permeable to ketopro-
fen – a fact that may be extrapolated to in-use scenarios.
The positive control, tea tree oil, a volatile/essential oil,
demonstrated clear penetration enhancement proper-
ties.
As a ‘natural’ product, the precise constituents of Aloe
vera are subject to variability depending upon a wide
range of factors, including soil, location and local climate.
Therefore, it is feasible that Aloe vera from different
sources may provide somewhat different results; further-
more, other potential penetrants may behave differently
to the ketoprofen used in this model.
In terms of risk exposure, the current data suggest that
handling products containing Aloe vera would not leave
the individual pre-disposed to enhanced ingress of exog-
enous chemicals due to compromised skin barrier.

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