Journal of

ETHNO-
PHARMACOLOGY
ELSEVIER Journal of Ethnopharmacology 55 (1996) 69 75

Antiinflammatory activity of extracts from A l o e vera gel

Beatriz Vfizquez "'*, Guillermo Avila a, David SeguraL Bruno Escalante b

~Laboratories of Pharmacology and Natural Products, Division of Investigation,
Escuela National de Estudios Projesionales Iztacala (E.N.E.P-I), Universidad National Autdnoma de Mbxico, Tlahwpantla,
Mbxico C.P. 54090. Mexico
bDepartment ~[" Pharmacology, Centro de lnvest~gaci6n y Estudios Avanzados, At. lnstituto Politdcnico National 2508,
Mbxico D.F. 07300, Mexico

Received 26 March 1994; revised 19 August 1996; accepted 30 August 1996

Abstract

We studied the effects of aqueous, chloroform, and ethanol extracts of Aloe vera gel on carrageenan-induced edema
in the rat paw, and neutrophil migration into the peritoneal cavity stimulated by carrageenan. We also studied the
capacity of the aqueous extract to inhibit cyclooxygenase activity. The aqueous and chloroform extracts decreased the
edema induced in the hind-paw and the number of neutrophils migrating into the peritoneal cavity, whereas the
ethanol extract only decreased the number of neutrophils. The antiinflammatory agents indomethacin and dexam-
ethasone also decreased carrageenan-induced edema and neutrophil migration. The aqueous extract inhibited
prostaglandin E 2 production from [~4C]arachidonic acid. The chemical tests performed in the aqueous extract for
anthraglycosides, reductor sugars and cardiotonic glycosides were positive. In the ethanol extract, the chemical tests
performed for saponins, carbohydrates naftoquinones, sterols, triterpenoids and anthraquinones were also positive. In
the chloroform extract, the chemical tests performed for sterols type A 5, and anthraquinones were positive. These
results demonstrated that the extracts of Aloe vera gel have antiinflammatory activity and suggested its inhibitory
action on the arachidonic acid pathway via cyclooxygenase.

Keywords: Aloe vera; lndomethacin; Dexamethasone; Inflammation: Cyclooxygenase

1. Introduction

A l o e vera T o u r n . ex Linn (syn: A l o e b a r b a d e n s i s

* Corresponding author. Laboratorios de Farrnacologia y Miller) (Liliaceae) is used with o t h e r species o f
Productos Naturales, Divisidn de Investigaci6n, E.N.E.P. lzta-
A l o e in the folk medicine o f M e x i c o a n d several
cala, Universidad Nacional Aut6noma de M6xico, Av. de los
Barrios s/n, Los Reyes Iztacala, Tlalnepantla, Estado de Mdx- o t h e r countries, for m a n y m e d i c a l a n d cosmetic
ico C.P. 54090. Mdxico. p u r p o s e s ( M o r t o n , 1961; D i e z - M a r t i n e z , 1981).

0378-8741/96/$15.00 ~G 1996 Elsevier Science Ireland Ltd. All rights reserved

PII $0378-8741 (96)01476-6
70 B. Vd=que: el al. ,' Journal o/Ethnopharmacology 55 (1996) 69 75

The fresh leaves of Aloe vera are used to obtain under reduced pressure, in a rotavapor. The dried
two components: (1) a bitter yellow juice (exudate) extracts were freshly dissolved in propylene glycol
with high content of 1,8 dihydroxyanthraquinone or saline solution before administration.
derivatives (aloe emodin) and their glycosides The aqueous extract was prepared with 1 g
(aloins), which are used for their cathartic effects powder in 20 ml of saline solution (0.45%), mixed
(Fairbairn, 1980) and (2) a mucilaginous gel from for 20 rain at 40°C and centrifuged at 100 x g for
the parenchymatous tissue, which has been used 10 rain; the supernatant was adjusted to pH 7.4.
for topical treatment of skin burns and wounds before administration and contained 40 mg/ml of
(Rovatti and Brennan, 1959). Some chemical con- extract material.
stituents with immunomodulatory and lectin-like
properties have been isolated from the extracts of 2.3. Animals
Aloe vera gel (Holdsworth, 1972; Winters et al.,
1981). These reports have stimulated research on Male Wistar rats (180 200 g) grown in our
the chemical, biochemical and pharmacological animal house and fed on standard chow diet and
properties of these compounds as well as on other water ad libitum were used for the experiments
medical uses (Grindlay and Reynolds, 1986). In (groups of 5 8) at 20 25°C.
spite of these data, there is no clear correlation
between the pharmacological properties of the 2.4. Antiinflammatory acti~ity
Aloe vera gel and its constituents.
The aim of this study was to investigate whether 2.4. I. Carrageenan-induced edema
the extracts of Aloe vera gel showed antiinflamma- The antiinflammatory activity of extracts was
tory activity and to analyze some of its chemical studied in groups of 6 8 rats. Edema was induced
constituents. according to the method described by Van Arman
et al. (1965). Briefly, 0.1 ml of 1% carrageenan
(type ll, Sigma, St Louis, MO) in sterile saline,
2. Materials and methods was injected into one hind paw, under the plantar
aponeurosis. A similar volume of saline solution
2.1. Preparation of the gel was injected into the other hind paw. The paw
volume was measured before injection of car-
Fresh Aloe vera leaves were collected (14 kg) rageenan or saline by the mercury displacement
from the Municipality of Apaxco, State of Mex- method (Van Arman et al., 1965) and the time
ico, in March, 1990. The plant was identified in the course of edema formation was followed over 8 h.
herbarium of the Department of Botany of the In separate groups of animals, either in-
Escuela Nacional de Estudios Profesionales Izta- domethacin (Sigma, St Louis, MO) 10 mg/kg or
cala, U.N.A.M., where a specimen was deposited dexamethasone (Merck-Sharp and Dohme) 0.5
under accession number RBP 9052. The gel was mg/kg were administered subcutaneously (s.c.) as
harvested from the green leaves, minced, homoge- standard drugs, saline solution and propylene gly-
nized, dried at 35°C and weighed. col as solvent controls. Different amounts of the
extracts or solvents were administered intraperi-
2.2. Preparation of extracts toneally (i.p.) 60 rain before carrageenan injection.
The volume increase (A volume) of the inflamed
The powder (l kg) was fractionated successively paw was estimated by subtracting the volume of
with n-hexane, benzene, ethyl acetate, chloroform, the contralateral paw. The antiinflammatory effect
acetone and 96% ethanol (Merck and Backer) in a of the drugs was evaluated as the degree of edema
Soxhlet extractor; to give the hexane (1.5 g), inhibition. Since the aqueous, chloroform and
benzene (2.0 g), chloroform (12.0 g), ethyl acetate ethanol extracts showed more antiinflammatory
(90.0 g), acetone (150.0 g) and ethanol (282.0 g) effect than the other extracts, the following studies
the solvent was evaporated at low tem]~erature, were done with these extracts.
 B. V{tzque: el al. ,' Journal O/Ethnopharmacology 55 (1996) 69 75 71

2.4.2. Neutrophil migration into peritoneal cavity method. Authentic prostaglandin standards were
Rats were injected with 3 ml of carrageenan used to identify retention times of prostaglandin
(100/~g/ml prepared in sterile saline solution) into E,. The concentration of the PGE 2 was estimated
the peritoneal cavity, and 4 h later the abdominal by densitometry after the autoradiography.
cavity was washed with 10 ml of phosphate
buffered saline solution containing 5 U/ml of 2.5. Ph vtochemieal analysis
heparin (Sigma, St Louis, MO) and 5% of bovine
serum albumin. Only 5 ml were withdrawn for cell Chemical tests were done on the Aloe vera gel
counts; the total cell counts were done in a Neu- for the detection of alkaloids, cardiac glycosides,
bauer chamber and differential cell counts were flavonoids, tannins, coumarins, anthraquinones,
performed by the technique reported by Souza saponins, sterols and triterpenes (Dominguez,
and Ferreira (1985). The results are expressed as 1973: Harborne, 1984). Thin layer chromatogra-
number of cells/ml of collected fluid. One hour phy of the extracts with biological activity was
before the carrageenan injection, the experimental performed on silica gel 60 G plates (Merck), with
groups were treated orally with 200 mg/kg of the ethyl acetate/methanol/water, 100:16.5:13.5 (v/v)
chloroform extract, 400 mg/kg of the aqueous or benzene/methanol 4:1 (v/v); the chro-
extract and 50, 200 and 600 mg/kg of the ethanol matograms were observed with an ultraviolet
extract. The control groups were also given saline lamp at 254--300 nm, to detect the separated
or propylene glycol by the oral route and the fractions which were marked and eluted from the
standard reference groups were treated subcuta- plate. The chemical tests were performed on the
neously with indomethacin (10 mg/kg) or dexam- eluted fractions.
ethasone (0.5 mg/kg).
2.6. Statistical ana@sis
2.4.3. Incubation of aqueous extracts with ram
seminal vesicles (RSV) microsomes Significant differences were assessed by Stu-
A RSV microsomal powder was prepared as dent's t-test for unpaired samples and values of
described previously by Laniado et al. (1989) and P < 0.05 were considered significant.
kept at - 7 0 ° C . The incubation was made ac-
cording to the method described, aqueous extract
(1, I0 and 100 /~g), 2 td ['4C]arachidonic acid 3. Results
(specific activity 58 mCi/mmol) obtained from
Amersham Corp., were incubated with the RSV The edema induced by carrageenan in the hind
cyclooxygenase preparation (500 Izg of protein), paw lasted for more than 8 h; the maximum effect
in the presence or absence of glutation (GSH) 0.1 was seen at the 4th hour (Fig. 1). The aqueous
mM for 10 rain. (Sigma, St Louis, MO). In con- and chloroform extracts showed antiinflammatory
trol experiments, incubations were done in the activity preventing the formation of edema after
presence of indomethacin (1 /lg). Arachidonic administration of carrageenan, i.e. carrageenan
acid metabolism was terminated by addition of 1 increased paw volume by 160 /L1 whereas car-
N citric acid to decrease pH to 4 5. Arachidonic rageenan administered concomitant with aqueous
acid metabolites were extracted with two volumes or chloroform extracts increased paw volumes to
of ethyl acetate. The ethyl acetate extract was 75 and 62 ILl, respectively (Fig. 1). Moreover, the
concentrated to dryness under nitrogen and resus- use of two well known antiinflammatory agents,
pended in 300 /L1 of methanol and applied to indomethacin and dexamethasone, also prevented
Silica Gel G plates (Merck). The plates were the carrageenan-induced increase in the paw vol-
developed in methanol/chloroform/acetic acid'wa- ume of the rat (Fig. 1). The aqueous chloroform
ter, 16:18:2:16 (v/v). For the separation of and ethanol extracts decreased the number of
prostaglandins, the thin layer chromatography neutrophils migrating into the peritoneal cavity
plate was examined by the autoradiographic (Fig. 2). The aqueous extract decreased migration
72 B. V/Izquez el al. Journal o/ Ethnot#~armacolo:(v 55 (1996) 69 75

300 * t~
..j 5

20o
tu~ 3
z©
100
i.t. o 2
O"-
_ ~
n~
0 I I t I I

1 2 3 4 5 ~ 0
SAL 50 200 600 DEX IND
TIME (hours) A. V E R A ( m g / K g )

Fig. I. Effects of aqueous and chloroform extracts of the Aloe Fig. 3. Dose-dependent effect of ethanol extract of the Aloe
~vra gel on the edema induced by carrageenan (100 pg in 0.1 cera gel on neutrophil migration. Increasing doses of Aloe vera
ml); administered intraplantarly, • • control group 0.1 ml (ethanol extract) were administered (v.o.) and compared with
(i.p.) propylene glycol: 10 mg/'kg indomethacin; • • vehicle control (SAL), dexamethasone (DEX) (0.5 mg/kg, s.c.)
0.5 mg/kg dexamethasone: • • 100 mg/kg (i.p.) aqueous or mdomethacin (10 mg/kg, s.c.). Values are the mean ±SE
extract of Aloe ~era gel: 7i 100 mg/kg (i.p.) chloroform t\)r the number of animals used ( n - 6). *P < 0.05 when com-
extract. Values are the mean ± SE for the number of animals pared vs. saline-treated.
used (n = 7). P < 0.05 when compared vs. treated groups.
k n o w n a n t i i n f l a m m a t o r y agents, d e x a m e t h a s o n e
by 28.6% and the chloroform extract by 42.9%. and i n d o m e t h a c i n , we observed that dexam-
The ethanol extract decreased the average neu- ethasone inhibited migration by 71.4% and in-
trophil migration in a dose-dependent fashion and d o m e t h a c i n by 36%.
the m a x i m u m effect was 48% and was o b t a i n e d I n c u b a t i o n of the RSV with 14C-AA, p r o d u c e d
with 600 mg/kg (Fig. 3). W h e n we used two well several radioactive metabolites. A c c o r d i n g to the
retention time of authentic p r o s t a g l a n d i n s stan-
dard, the m a i n metabolite was P G E > and incuba-
tion of the microsomes with i n d o m e t h a c i n gave a
significant decrease in the P G E : p r o d u c e d from
0 4 ~
6.2% to 2.3% of arachidonic acid converted/rag
I
p r o t e i n / m i n . This suggested that induced arachi-
u.I donic acid metabolism was mediated via cy-

z 1
I.i. o
O""
n,
i11
2-

1-
clooxygenase products. I n c u b a t i o n with 100 mg
of the A l o e z'era a q u e o u s extract also significantly
decreased the PGE~ p r o d u c t i o n fi'om 6.2% to
3.2% (Fig. 4).
,~ o- The phytochemical analysis of the A l o e vera gel
z SAL PG AQ CHL DEX a n d its extracts revealed the presence of an-
t h r a q u i n o n e s , sterols, sterols type A s, s a p o n i n s
Fig. 2. Effects of 400 mg/kg (AQ) aqueous and 200 rng/kg
chloroform extracts (CHL) of the Aloe vera gel, orally admin- and c a r b o h y d r a t e s (Table 1). The a q u e o u s a n d
istered, on neutrophil migration into the peritoneal cavity ethanol extracts had a pH of 5. The aqueous
induced by carrageenan (300 l~g). Saline solution (SAL) and extract showed high a m o u n t s of mucilagus com-
propylene glycol (PG) were used as vehicle control whereas p o n e n t s and pectins whereas in both extracts the
dexamethasone (0.5 mg/kg s.c.) was used as standard anti-
a n t h r a q u i n o n e s were either free or b o u n d to car-
intlammatory. Values are the mean +_SE for the number of
animals used (n = 8). *P < 0.05 when compared vs. saline- or bohydrates. The c h l o r o f o r m extract was only sol-
propylene glycol-treated groups. uble in propylene glycol at 30°C.
 B. Vdzquez et al. Journal o / E t h n o p h a r m a c o l o g v 55 (1996) 69 75 73

tory drug that inhibits phospholipase Az enzyme

z "~ which is responsible for arachidonic acid libera-
o o tion, the substrate for prostaglandin production
O O~ (Blackwell et al.. 1980). Thus, the antiinflamma-
tory action of these agents may be related to the
o ,~ inhibition of prostaglandins and leukotriene syn-
thesis. Further, the dose of dexamethasone used
0 produced a higher inhibition of neutrophil migra-
u,,I < tion than the effect produced by indomethacin,
supporting the possible role of leukotrienes in the
0 inflammation model.
CONTROL IND ALOE The antiedema effect of these two extracts cor-
Fig. 4, In vitro inhibitory effect of Aloe vera extract on the related with their ability to decrease the number
production of PGE.. RSV microsomes were incubated with of neutrophils migrating into the peritoneal
[~4C]arachidonic acid and glutation in the absence of inhibitor cavity. The effect on formation and neutrophil
(control) o1 in the presence of indomethacin 1 pg/ml (IND) or migration has been suggested as some of the
aqueous Aloe l'era extract 100 /~g/ml (Aloe). Each bar repre-
characteristics of the antiinflammatory agents
sents the mean ± S E of five experiments. * P < 0 . 0 5 when
compared vs. control. whose mechanism of action is related to in-
hibitory action of the arachidonic acid pathway
4. Discussion and conclusion (Higgs et al., 1979). Our results clearly demon-
strate that the aqueous extract inhibited in vitro
This study showed that the aqueous and chlo- conversion of arachidonic acid to PGE2 suggest-
roform extracts of the A l o e vera get contain com- ing the idea that the extract has cyclooxygenase
pounds with a potential to reduce carrageenan inhibitory properties. Since we are using exoge-
induced-edema. We found that at the doses used, nous arachidonic acid, thereby bypassing the re-
A l o e vera inhibited edema formation, by a per- lease of arachidonic acid by lipooxygenase activity
centage close to the inhibition produced by the and because the microsomes have high cyclooxy-
well established antiinflammatory agents, in- genase activity and negligible lipooxygenase activ-
domethacin and dexamethasone. Indomethacin, ity. we could suggest that the antiinflammatory
like most of the non-steroidal antiinflammatory effect of A l o e vera is related to cyclooxygenase
compounds, inhibits the biosynthesis of inhibition, rather than an effect on lipooxygenase
prostaglandins and this effect might explain its activity. However, we demonstrated that dexam-
antiinflammatory activity in carrageenan-induced ethasone in this model is an important antiinflam-
rat paw edema (Higgs et al., 1979). On the other matory agent, which suggested that probably both
hand, dexamethasone is a steroidal antiinflamma- arachidonic acid metabolite products,
prostaglandins and leukotrienes are involved in
the pathology of the inflammation. Thus the need
Table 1 to measure lipooxygenase metabolites on the exu-
Chemical groups identified in the extracts with antiinflamma- date of the edema and the effect of A l o e t,era on
tory activity of the Aloe t,era gel
these products, become necessary to discard inhi-
Extract Chemical class bition of the lipooxygenase products as one of the
A l o e cera mechanisms of action. PGE2 formation
Aqueous Anthraglycosides, reductor sugars, car- inhibition by the extract may account for the
diotonic glycosides, mucilagus and pectins antiinflammatory effects since PGE2 is the main
Chloroform Sterols type A 5 and anthraquinones
prostaglandin in the inflammatory exudate (Fer-
Ethanol Carbohydrates, naftoquinones, an-
thraquinones, saponins, sterns and triter- reira et al., 1974). The ethanol extract did not
penoids show an antiedema effect but reduced the migra-
tion of neutrophils. These results suggest the ab-
74 B. Vdzquez et al./ Journal of Ethnopharmacology 55 (1996) 69 75

sence in the ethanol extract of the compounds Acknowledgements

involved in the antiinflammatory effect.
A number of Japanese workers have found The authors wish to thank Drs Francisco
antiinflammatory compounds in Aloe species Posadas and Jose L. Reyes for their critical re-
other than Aloe vera. Fujita et al. (1979) de- view of this manuscript; to Leonor ZOfiiga and
scribed in vitro bradykininase and carboxypepti- Gabriel Martinez for expert technical assistance.
dase activities in Aloe arborescens while Yagi et This work was supported by a grant from Con-
al. (1982) reported in vitro antibradykinin activ- sejo Nacional de Ciencia y Tecnologia (CONA-
ity in Aloe saponaria; the faster breakdown of CyT), Mexico, Ref. Dl11-903672.
bradykinin might reduce pain and inflammation.
We have also demonstrated this activity of Aloe
t;era gel in the isolated rat ileum (results not References
shown). Chemical tests on the Aloe vera gel and
Blackwell, G.J.. Carnuccio, R., DiRosa, M., Flower, R.J..
its extracts revealed the presence of components Parente, L., and Persico, P. (1980) Macrocortin: a polypep-
with low molecular weights (Table 1). In the tide causing the anti-phospholipase effect of glucocorti-
aqueous and ethanol extracts, we identified gly- cold. Nature 287, 147 149.
cosides of anthraquinones while in the chloro- Diez-Martinez, S.D. (1981) La Zdbila. Comunicado No. 46
Sobre Recursos Bieticos Poteneiales del Pais. lnstituto Na-
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leaves had no inhibitory effect in carrageenan- Limusa, Mexico, pp. 281.
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 B, V~izquez el a/. /Journal ~/' Ethnopharmacoh~gy 55 (1996) 69 75 75

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