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Avena sativa L.
E. Kpeli Akkol
a,
*
, I. Sntar
a
, I. Erdogan Orhan
a
, H. Keles
b
, A. Kan
c
, G. oksari
d
a
Department of Pharmacognosy, Faculty of Pharmacy, Gazi University, Etiler, 06330 Ankara, Turkey
b
Department of Pathology, Faculty of Veterinary Medicine, Afyon Kocatepe University, 03030 Afyonkarahisar, Turkey
c
Vocational School of Technical Sciences, Program for Food Technologies, Seluk University, 42070 Konya, Turkey
d
Department of Field Crops, Faculty of Agriculture, Seluk University, 42070 Konya, Turkey
a r t i c l e i n f o
Article history:
Received 9 April 2010
Received in revised form
15 December 2010
Accepted 24 January 2011
Keywords:
Avena sativa
Wound healing
Antioxidant
Total phenol and avonoid
a b s t r a c t
Avena sativa L. (Poaceae) has been reported to have traditional utilization against skin diseases and
inammation. Therefore, in this study, the n-hexane, ethyl acetate, ethanol, and water extracts of A. sativa
were investigated for their wound healing and antioxidant activities. Total phenol and avonoid contents
of the extracts were established spectrophotometrically. For the wound healing activity, linear incision
and circular excision models on rats and mice were evaluated with a standard ointment Madecassol
.
Antioxidant activity was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, ferrous
ion-chelating, and ferric-reducing antioxidant power (FRAP) assays. Signicant wound healing activity
was observed with the ointment formulation of the ethanol extract at 1% concentration. The histo-
pathological examination results also supported the outcome of both linear incision and circular excision
wound models. All of the extracts exerted low antioxidant activity in the applied assays. The present
study provides a scientic evidence for the traditional usage of A. sativa in the management of wound
healing.
2011 Published by Elsevier Ltd.
1. Introduction
Avena sativa L. (Poaceae), known as common oat, is a cereal
grain species used also as food. It has been cultivated in Anatolian
peninsula sinceancient times whose cultivationwas recordedbythe
Greek scientist Galenus in the 17th century. The major cultivation
area of oat is the Marmara region and internal Anatolia inTurkey. In
some folk medicine systems belonging to different parts of the
world, it has been reported to be used traditionally against some
diseases. For instance; in the southern Appalachia region of the
United States, A. sativa was recorded to have utilization against
chickenpox, poison ivy, and other rashes externally (Cavender,
2006). In traditional medicine of Lebanon, its alcoholic macerate
was reported to be used as antirheumatic and antineuralgic (El
Beyrouthy et al., 2008). Ballabh et al. (2008) reported use of oat
against kidney and urinary disorders in traditional medicine of clod
desert Ladakhinthe remotest regionof the Indiansubcontinent. Oat
is an important crop plant containing high levels of proteins, lipids,
vitamins, dietary bers, and minerals as well as a food plant for both
humans and animals (Gajdosova et al., 2007). A. sativa was reported
to possess a variety of biological activities such as angiotensin-I
converting enzyme inhibitory (Cheung et al., 2009), anti-inam-
matory and anti-itchy (Sur et al., 2008), anti-HIV (Shun et al., 2008),
and cardioprotective activities (Ryan et al., 2007).
The aim of the present study was to investigate in vivo wound
healing activity of A. sativa to clarify its traditional use in a scientic
platform along with its in vitro antioxidant activity. The n-hexane,
ethyl acetate, ethanol, and water extracts of the plant were tested in
mice and rats for their wound healing activity using in vivo linear
incision and circular excision wound models, which have been used
together for conrmation of the wound healing activity. Addition-
ally, antioxidant activity was conducted in the extracts to exploit its
relation to wound healing activity. Antioxidant activity of the
extracts was determined using three different in vitro methods; 2,2-
diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, ferrous ion-
chelating, and ferric-reducing antioxidant power (FRAP) assays at
250, 500, and 1000 mg/ml concentrations. Total phenol and avo-
noid contents of the extracts were established spectrophotomet-
rically by FolineCiocalteau and AlCl
3
methods, respectively.
Abbreviations: ABTS, 2,2
0
azinobis-(3-ethylbenzthiazoline-6-sulphonic acid);
DPPH, 2,2-diphenyl-1-picrylhydrazyl; FRAP, Ferric-reducing antioxidant power;
S.E.M, Standard error mean.
* Corresponding author. Tel.: 90 312 2023185; fax: 90 312 2235018.
E-mail address: esrak@gazi.edu.tr (E.K. Akkol).
Contents lists available at ScienceDirect
Journal of Cereal Science
j ournal homepage: www. el sevi er. com/ l ocat e/ j cs
0733-5210/$ e see front matter 2011 Published by Elsevier Ltd.
doi:10.1016/j.jcs.2011.01.009
Journal of Cereal Science 53 (2011) 285e290
[280]
2. Experimental
2.1. Plant material
Cultivated sample of A. sativa was obtained from the experi-
mental farm of Seluk University, in Konya province, Turkey, at the
owering stage in June, 2009.
2.2. Preparation of plant extracts
After the plant material was driedinshade at roomtemperature, it
was weighedaccurately(200g) ina digital balancer (MettlereToledo)
and powdered in a mechanical grinder. Then, the powdered material
was subjected to successive extraction starting from n-hexane
(2 L 2), ethyl acetate (3 L 2), ethanol (3 L 2), and water (3 L 1).
After the corresponding organic phases in each extraction were
lteredthroughregular lter paper, they were evaporatedinvacuotill
dryness to give the crude extracts. The water extract was nally
lyophilized. The extract yields (w/w) were calculated as follows: The
n-hexaneextract; 2.09%, theethyl acetate extract; 25.98%, theethanol
extract; 15.61%, the water extract; 29.47%.
2.3. Wound healing activity tests
2.3.1. Animals
Male, SpragueeDawley rats (160e180 g) and Swiss albino mice
(20e25 g) were purchased fromthe animal breeding laboratories of
Experimental Animal Research Center of Gazi University (GDAM)
(Ankara, Turkey).
The animals were left for 3 days at room conditions for accli-
matization. They were maintained on standard pellet diet and
water ad libitum throughout the experiment. A minimum of six
animals were used in each group. Throughout the experiments,
animals were processed according to the suggested international
ethical guidelines for the care of laboratory animals under the audit
of Gazi University Commission of Animal Ethics.
2.3.2. Preparation of test samples for wound healing activity
Incision and excision wound models were employed to evaluate
the wound healing activity. For the in vivo wound models, test
samples were prepared in an ointment base (vehicle) consisting of
glycol stearate, 1,2-propylene glycol, liquid parafn (3:6:1) in 1%
concentration. 0.5 g of each test ointment was applied topically on
the wounded site immediately after the wound was created by
a surgical blade.
Animals of the vehicle group were treated with the ointment
base only, whereas animals of the reference drug group were
treated with 0.5 g of Madecassol
), and
vehicle were topically applied once a day throughout 9 days.
Animals of the negative control group were not treated with any
material. All the sutures were removed on the 9th post wound day.
On the 10th day, all the animals were killed under anesthesia. One
linear-paravertebral incised skinwas measured using a tensiometer
(Zwick/Roell Z0.5, Germany) for its tensile strength, the other
incised skin was sent for histopathological examination (Lodhi
et al., 2006; Suguna et al., 2002).
Tensiometer measures the breaking strength in N (Newtons),
which is called tensile strength.
Tensile strengthTS of extract %
TS
extract
TS
vehicle
100
TS
vehicle
Tensile strength of reference %
TS
reference
TS
vehicle
100
TS
vehicle
Tensile strength of vechicle %
TS
vechicle
TS
negative
100
TS
negative
2.3.3.2. Circular excision wound model. A circular excision wound
model was used to monitor wound contraction and wound closure
time. Each group of animals (six animals in each) was anaesthetized
by 0.01 cc Ketalar
Digital Image
Analyze System) and graded as mild (), moderate () and severe
() for epidermal or dermal re-modeling. Re-epithelization or
ulcus in epidermis; broblast proliferation, mononuclear and/or
polymorphonuclear cells, neovascularization and collagen deposi-
tions in dermis were analyzed to score the epidermal or dermal re-
modeling. At the end of the examination, obtained results were
combined and staged for wound healing phases as inammation,
proliferation, and re-modeling in all groups.
Table 1
Effects of the extracts of A. sativa on linear incision wound model.
Materials Statistical
mean S.E.M.
a
(mm
2
)
(Tensile
strength %
b
)
Vehicle 14.86 1.81 12.1
Negative control 13.26 2.19 e
c
n-Hexane extract 14.31 2.03 e
Ethyl acetate extract 17.42 1.14 17.2
Ethanol extract 19.21 1.87 29.3
Water extract 17.91 1.21 20.5
Madecassol
0 19.14 1.89 19.60 2.09 19.86 1.22 20.05 1.03 20.28 1.68 19.25 1.26 19.21 1.44
2 17.22 1.40 17.30 1.56 17.01 2.24 16.29 1.69 16.05 1.10 17.04 2.37 15.29 2.09
(0.5) (1.2) (5.4) (6.8) (1.0) (11.2)
4 14.31 1.82 16.43 2.99 14.08 2.14 13.32 1.50 12.21 1.54 12.46 1.26 11.29 2.76
(12.9) (1.6) (6.9) (14.7) (12.9) (21.1)
6 12.53 1.07 13.42 1.62 12.27 1.38 10.62 1.63 9.96 1.32 11.10 1.17 8.43 1.64
(6.63) (2.1) (15.2) (20.51) (11.4) (32.7)*
8 8.77 1.37 10.78 1.54 8.80 1.43 7.53 0.36 6.46 0.11 8.51 0.94 3.90 1.71
(18.6) e
c
(14.1) (26.3) (2.9) (55.5)**
10 4.58 0.91 4.96 1.19 4.63 0.17 3.49 0.66 3.09 0.70 3.98 0.86 1.03 0.18
(7.7) e (23.8) (32.5)* (13.1) (77.5)***
12 2.50 0.06 2.76 0.17 2.59 0.19 1.76 0.26 1.48 0.03 2.24 0.16 0.00 0.00
(9.4) e (29.6) (40.8)* (10.4) (100.0)***
*p < 0.05.
***p < 0.001.
a
Standard error meaning.
b
Percentage of the contraction values: The vehicle group was compared to the negative control group; Extracts and the reference material were compared to vehicle group.
c
No activity.
Table 3
Wound healing processes and healing phases of the vehicle, negative control, A. sativa extracts and Madecassol
administered animals.
a
Groups Wound Healing Processes Healing Phases
S U RE FP CD MNC PMN NV I P R
Vehicle / / /
Negative Control / e / / e
n-Hexane extract / e e
Ethyl acetate extract / / / /
Ethanol extract e / / / / / / / /
Water extract / / / / / / /
Madecassol
e / / / / / /
a
HE and VG stained sections were scored as mild (), moderate () and severe () for epidermal and/or dermal re-modeling. S: Scab, U: Ulcus, RE: Re-epithelization,
FP: Fibroblast proliferation, CD: Collagen depositions, MNC: Mononuclear cells, PMN: Polymorphonuclear cells, NV: Neovascularization, I: Inammation phase, P: Proliferation
phase, R: Re-modeling phase.
E.K. Akkol et al. / Journal of Cereal Science 53 (2011) 285e290 287
samples were mixed with 750 mL of FolineCiocalteaus reagent and
sodium carbonate in test tubes. The tubes were then vortexed and
incubated at 40
C for 30 min. Afterward, absorption was measured
at 760 nm using a Unico 4802 UVevisible double beam spectro-
photometer (USA). Total avonoid content was calculated by the
aluminum chloride colorimetric method (Woisky and Salatino,
1998). To sum up, a number of dilutions of quercetin were
obtained to prepare a calibration curve. Then, the same dilutions
from the samples were also prepared and separately mixed with
95% ethanol, 10% aluminum chloride, 1 M sodium acetate and
distilled water. Following incubation for 30 min at room tempera-
ture, absorbance of the reaction mixture was measured at a wave-
length of 415 nm with a Unico 4802 UVevisible double beam
spectrophotometer (USA). The total phenol and avonoid contents
of the extracts were expressed as gallic acid and quercetin equiv-
alents (mg/g extract), respectively.
2.6. Statistical analysis of the data
The data on percentage wound healing was statistically
analyzed using one-way analysis of variance (ANOVA). The values
of p 0.05 were considered statistically signicant. Histopathologic
data were considered to be nonparametric; therefore, no statistical
tests were performed.
3. Results
3.1. Results of wound healing activity
The wound healing activity of the n-hexane, ethyl acetate,
ethanol, and water extracts of A. sativa was evaluated on rats and
mice by linear incision and circular excision wound models to
conrm the claimed folkloric usage of the plant on a scientic base.
The histopathological changes formed by these extracts were also
assessed. The results of the measurements of tensile strength were
shown in Table 1. Tensile strength of the animals treated with the
ethanol extract of the plant demonstrated the highest value (29.3%,
p < 0.05) on day 10. Topical application of the extract on the inci-
sion wound model expressed a signicant increase in wound
tensile strength as compared to other extract treated groups and
the control groups. On the other hand, reference drug Madecassol
administered animals. Skin sections show the hematoxylin & eosin (HE) stained
epidermis and dermis in A, and the dermis stained with Van Gieson (VG) in B. The
original magnication was 100 and the scale bars represent 120 mm for gures in A,
and the original magnication was 400 and the scale bars represent 40 mm for B. Data
is representative of 6 animals per group. 1) Vehicle group, 10 day old wound tissue
treated with only vehicle, 2) Negative Control group, 10 day old wound tissue,
untreated group, 3) n-Hexane extract group, 10 day old wound tissue treated with n-
Hexane extract, 4) Ethyl acetate extract group, 10 day old wound tissue treated with
ethyl acetate extract, 5) Ethanol extract group, 10 day old wound tissue treated with
Ethanol extract, 6) Water extract group, 10 day old wound tissue treated with water
extract, 7) Reference group, 10 day old wound tissue treated with Madecassol
.
Arrows pointing events during wound healing; s: scab, u: ulcus, re: re-epithelization, f:
broblast, c: collagen, mnc: mononuclear cells, pmn: polymorphonuclear cells, nv:
neovascularization.
E.K. Akkol et al. / Journal of Cereal Science 53 (2011) 285e290 288
was seen in the other groups. In experimental groups, best re-
modeling, particularly, re-epithelization was detected in the
Madecassol