LWT - Food Science and Technology 60 (2015) 495e501

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LWT - Food Science and Technology

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Effect of Aloe vera gel coating on antioxidant capacity, antioxidant

enzyme activities and decay in raspberry fruit
Hamid Hassanpour*
Department of Horticulture, Faculty of Agriculture, Urmia University, Urmia, Iran

a r t i c l e i n f o

a b s t r a c t

Article history:
Received 3 November 2013
Received in revised form
1 July 2014
Accepted 30 July 2014
Available online 12 August 2014

Native populations of raspberry fruits (Rubus spp.) were coated with Aloe vera gel and were then assayed
for the antioxidant capacity, total anthocyanin, total phenol, antioxidant enzyme activities and postharvest quality after 8 days storage at 4
C, relative to a control group. These berries, coated with Aloe
vera gel, showed a higher antioxidant capacity, total anthocyanin and total phenol than those of the
controls (non-treated) group. The treated fruits exhibited less incidence of decay during storage at 4
C
than the control group. Thus postharvest life (as affected by fungal decay) was longer for berries treated
with Aloe vera gel than for the control fruit. However, total soluble solid, titratable acidity and pH were
predominantly inuenced by storage periods. Aloe vera gel treatments could reduce the natural decay
that happens over time. The activities of antioxidant enzymes, including glutathione peroxidase (GSHPOD), glutathione reductase (GR), superoxide dismutase (SOD), ascorbate peroxidase (AsA-POD) and
guaiacol peroxidase (G-POD) were enhanced. The nonenzyme components such as reduced glutathione
(GSH) and oxidized glutathione (GSSG) were also increased by Aloe vera gel. In conclusion, raspberry
fruits treated with Aloe vera gel maintained higher levels of antioxidant capacity, total phenol, total
anthocyanin and antioxidant enzymes during storage periods.
2014 Elsevier Ltd. All rights reserved.

Keywords:
Postharvest life
Raspberry
Total anthocyanin
Total phenol

1. Introduction
Berry fruit, including raspberries, have been reported to contain
high phenol and anthocyanin content (Heinonen, Lehtonen, &
Hopia, 1998). Native raspberry (Rubus spp.) constitutes a good
source of natural antioxidant substances. Thus, various antioxidants found in berry fruit provide signicant health benets. Interest in the role of antioxidants in human health has promoted
research in the field of horticulture and food science to evaluate
vegetable and fruit antioxidants and to measure how their content
and activity can be maintained or even improved through breeding,
cultural practices, and postharvest handling and processing (AyalaZavala, Wang, Wang, & Gonzalez-Aguilar, 2004). It is great interest
to investigate changes in antioxidant status during postharvest
storage of horticultural crops. Raspberry fruit is very valuable
nutritious, but because of being naturally highly perishable, the
storage time is restricted to just very short periods. To obtain the
optimum avor, shape and color, the berries should be harvested
for fresh consumption at a mature phase (Haffner, Rosenfeld,
Skrede, & Wang, 2002).

* Tel.: 98 09143094910; fax: 98 04262237718.

E-mail addresses: phhassanpour@gmail.com, ha.hassanpour@urmia.ac.ir.
http://dx.doi.org/10.1016/j.lwt.2014.07.049
0023-6438/ 2014 Elsevier Ltd. All rights reserved.

For centuries, Aloe vera has been used for its medicinal and
therapeutic properties (Eshun & He, 2005). The two major liquid
sources of Aloe vera are a yellow latex (exudates) and clear gel
(mucilage), both of which proceed from the large leaf parenchymatic cells (Ni, Turner, Yates, & Tizard, 2004). The oral ingestion
of the Aloe vera gel juice has been shown to be effective in treating
ulcerous, gastrointestinal, kidney and cardiovascular problems, in
addition has also been used to diminish the cholesterol and triglyceride levels in blood (Reynolds & Dweck, 1999). In addition,
anti-inammatory and antibiotic actions have been reported, as
well as intervention in the treatment of certain diseases (diabetes,
cancer, allergy, AIDS) (Eshun & He, 2005; Reynolds & Dweck, 1999).
Edible coatings are traditionally used to improve food appearance and conservation due to their environmentally friendly nature, since they are obtained from both animal and vegetable
agricultural products (Petersen et al., 1999). Generally, coatings can
be classied according to their nature, into protein, lipid, and
polysaccharide-based, alone or in combination. They act as barriers
to moisture and oxygen during processing, handling, and storage,
and not only retard food deterioration, but also improve safety, due
to their natural biocide activity or to the incorporation of antimicrobial compounds (Cha & Chinnan, 2004). The Aloe vera gel was
able to prolong the shelf life (SL) of sweet cherry and table grapes
through a delay in the parameters related to deterioration from the

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H. Hassanpour / LWT - Food Science and Technology 60 (2015) 495e501

Native populations of raspberry (Rubus spp.) grown naturally

near the Caspian sea in Iran, were hand-harvested at a commercially mature stage, sorted to eliminate damaged and unripe fruit,
and selected for uniform size and color. A total of 180 fruits was
divided into four groups and were treated with Aloe vera gel.
Treatment was performed at 20
C by immersing the fruits for
5 min in a solution of Aloe vera gel diluted 1:3 (gel 2), 1:1 (gel 3) and
3:1 (gel 4) with distilled water. The fruits immersed in distilled
water were served as the control (gel 1). Following treatment, all
fruits were air-dried for 30 min before storage at 4
C in darkness
for 8 days. Ten samples of both treated and control fruit were isolated after 0, 2, 4, and 8 days, and were immediately analyzed. Four
replicates of each treatment were used for analyses. The characteristics such as fruit decay, antioxidant capacity, antioxidant
enzyme activities, total anthocyanin, total phenol, ascorbic acid and
internal qualities were evaluated during the storage time.

the tissue was ground to a fine powder under liquid nitrogen by

cold mortar and pestle and 1 g of the resultant powder was added
to 10 ml of methanol containing HCl (1%, v/v) and held at 0
C for
10 min. The slurry was centrifuged at 17,000  g for 15 min at 4
C
and then the supernatant was used. Absorption was measured in a
Shimadzu spectrophotometer at 530 and 700 nm in buffers at pH
1.0 and 4.5, using A [(A530A700) pH1.0  (A530A700) pH4.5] with
molar extinction coefcients of cyanidin-3-O-glucoside for raspberry fruit juice. Results were expressed as mg of cyanidin-3-Oglucoside equivalents per 100 g of FW.
The antioxidant activity of the raspberry fruit was evaluated by
free radical 2, 2-dipheynl-1-picrylhydrazyl (DPPH) method
(Hassanpour, Hamidoghli, Hajilo, & Adlipour, 2011). For the determination of free radical scavenging activity, samples were extracted
with methanol. Then, they were centrifuged at 15,000  g for
10 min. The supernatants were concentrated under reduced pressure at 40
C. The dried extracts were dissolved in methanol. Fifty
microliters of the diluted extracts (concentrations 2e20 mg ml1)
were added to 1 ml of 6  105 mol L1 DPPH in methanol. The
mixture was shaken and left at room temperature for 30 min; the
absorbance was measured spectrophotometrically at 515 nm.
Methanol was used as an experimental control. The percent of
reduction of DPPH was calculated according to the following
equation:
% inhibition of DPPH (Abs control  Abs sample)/Abs
control  100; Abs control is the absorbance of DPPH solution
without the extract.
The extracts for the total phenolic content assay were obtained
by extracting 1 g of homogenized berries (Polytron PT 1200E Homogenizer) with 10 ml of methanol at ambient temperature for 1 h
with constant shaking. The solution was filtered, and the residue
was repeatedly extracted with 10 ml of methanol for 1 h. Finally, the
extracts were combined and diluted to 100 ml with methanol.
Then, total phenolic compounds were determined using the
FolineCiocalteu reaction (Singleton, Orthofer, & Lamuela-Raventos,
1999), with Gallic acid as the standard. Results were expressed as
mg Gallic acid equivalents per 100 g FW.
L-ascorbic
acid was determined by using 2,
6dichlorophenolindophenol- dye method, and the results were
expressed as mg ascorbic acid equivalent 100 g1 FW (Deepa,
Charanjit, Balraj, & Kapoor, 2006).

2.3. Fungal decay incidence

2.6. PAL enzyme activity assay

Fungal decay was visually inspected after 2, 4 and 8 days of

storage at 4
C. Raspberry fruit showing surface mycelia development was considered decayed. Fungal decay incidence is the mean
proportion (percentage) of fruit that showed any fungal decay (i.e. a
level of >0).

Measurement of phenylalanine ammonia-lyase (PAL) activity

was performed at 290 nm, according to the methods of Yao and
Tian (2005). PAL activity was evaluated as nmol cinnamic acid
h1 mg1 protein. Protein content was determined according to
Bradford (1976) with bovine serum albumin (BSA) as standard.

2.4. Chemical analysis

2.7. Antioxidant enzyme measurements

Twenty fruits from each replicate were wrapped in cheesecloth

and squeezed with a hand press, and the juice was analyzed for
total soluble solids (TSS) and titratable acidity (TA). TSS was
determined by refractometer (Digital ABBE, Bellevue, Washington,
U.S.A). TA was determined by diluting each 2 ml aliquot of raspberry juice in 20 ml of distilled water and then titrated to pH 8.2 by
using 0.1 M NaOH. pH value was indicated by pH meter (HBJ-260).

2.7.1. Glutathione peroxidase (GSH-POD, EC 1.11.1.9) and

glutathione reductase (GR, EC 1.6.4.2)
Fruit tissue (4 g fresh weight) was homogenized in 4 ml of 0.1 M
TriseHCl buffer (pH 7.8) containing 2 mM EDTA-Na, and 2 mM DTT.
The homogenate was centrifuged at 20,000  g for 30 min at 4
C,
and the supernatant was used for the GSH-POD and GR assays. The
activity of GSH-POD enzyme was measured using the method of
Tappel (1978). The reaction mixture contained 0.1 M TriseHCl
buffer (pH 7.9), 0.4 mM EDTA, 1.0 mM NaN3, 1.0 mM H2O2, 1.0 mM
glutathione (GSH), 0.15 mM NADPH, 1 unit of glutathione reductase, and 100 ml enzyme extract. The total reaction volume was
1.0 ml. The reaction was started by adding H2O2. GSH-POD enzyme
activity was measured by the rate of NADPH oxidation at 340 nm

point of view of quality and safety (Martnez-Romero et al., 2006;

Valverde et al., 2005).
The aim of this research was to assess the effects of Aloe vera gel
on raspberry's functional properties during postharvest cold storage, and subsequent SL, and then to get a better understanding of
the effects of Aloe vera on maintenance of the compounds that are
benecial to human health.
2. Materials and methods
2.1. Chemicals and reagents
Folin-Ciocalteu phenol reagent, gallic acid, sodium carbonate,
sodium ascorbate, 2,6-dichlorophenolindophenol, Hydrogen
peroxide and ascorbic acid were purchased from Merck Co.
(Darmstadt, Germany). Dithiothreitol (DTT), glutathione (oxidized
form), glutathione (GSH, reduced form), glutathione reductase,
guaiacol, nitro blue tetrazolium (NBT), 2,2-diphenyl-1picrylhydrazyl (DPPH), disodium ethylenediaminetetraacetatic
acid (EDTA), bovine serum albumin (BSA) and cinnamic acid were
purchased from SigmaeAldrich Chemie GmbH, Steinheim,
Germany.
2.2. Plant materials

2.5. Total anthocyanin, phenolic compounds, ascorbic acid and

antioxidant activity
Analysis of total anthocyanin content in fruit juice was determined using the pH differential method (Wrolstad, 1976). Some of

H. Hassanpour / LWT - Food Science and Technology 60 (2015) 495e501

via a spectrophotometer. GR enzyme activity was assayed according to Smith, Vierheller, and Thorne (1988) at 340 nm. The reaction
was started by adding GSSG (oxidized glutathione) and the rate of
oxidation was calculated using the extinction coefcient of NADPH
(6.22 mM1 cm1).
2.7.2. Superoxide dismutase (SOD)
Fruit tissue (4 g) was pulverized in a cold mortar and pestle with
4 ml potassium phosphate buffer (0.1 mol L1, pH 7.4) containing
1 mmol L1 EDTA and 2 mmol L1 DTT. The homogenate was
filtered through four layers of miracloth and centrifuged at
12,000  g for 10 min at 4
C. The supernatant was used for
assaying the SOD enzyme activity. Total SOD enzyme activity was
assayed photochemically (Monk, Fagerstedt, & Crawford, 1987;
Thayer, 1990). One unit of SOD was expressed as the amount of
enzyme, which produced a 50% inhibition of NBT reduction under
assay conditions.
2.7.3. Ascorbate peroxidase (ASA-POD) and guaiacol peroxidase (GPOD)
Fruit tissue (4 g) was ground in a cold mortar and pestle with
4 ml potassium-phosphate buffer (0.1 mol L1, pH 7.3) containing
1 mmol L1 EDTA and 2 mmol L1 DTT. The homogenate was
centrifuged at 12,000  g for 10 min at 4
C. The supernatant was
used for the AsA-POD, and G-POD assays. AsA-POD activity was
measured according to the method of Amako, Chen, and Asada
(1994). The reaction was started via adding H2O2. The G-POD
enzyme assay mixture containing 0.1 mol L1 phosphate buffer (pH
6.1), 4 mmol L1 guaiacol as donor, 3 mmol L1 H2O2 as substrate
and 1.0 ml crude enzyme extract. The total reaction volume was
3.0 ml. The rate of change in absorbance at 420 nm was measured,
and the level of enzyme activity was expressed as the difference in
absorbance (OD).
2.8. Non-enzyme component measurements
2.8.1. Glutathione (GSH) and oxidized glutathione (GSSG)
Raspberry fruit samples (4 g) were homogenized in 8.0 ml ice
cold, degassed 7.57 mM sodium ascorbate solution with cold
mortar and pestle under N2 at 0
C. The homogenate was filtered
through four layers of miracloth and centrifuged at 30,000  g for
15 min at 0
C. The supernatant was deproteined by incubation in a
water bath at 100
C for 3 min and then centrifuged at 15,000  g
for 15 min at 0
C. The supernatants were used for the GSH and
GSSG assays. GSH and GSSG enzyme were assayed using the
method of Castillo and Greppin (1988). GSSG was calculated by
subtraction of GSH from total glutathione.
2.9. Statistical analysis
Experiments were performed according to a factorial design.
Analysis of variance (ANOVA) of data was performed in this
experiment using the Statistical Analysis System (Software Version
9.1 SAS). In the case of a significant F-value, the means were
compared by Duncan's Multiple Range test at p < 0.01 significance
level in SAS.

497

of the control fruits after 8 days storage at 4
C (Table 1). The decay
incidence was reduced in the treated fruit, during storage at 4
C,
compared to the control, and no difference was found between
different gel levels. Overall, the higher rate of fruits decay was
found in untreated raspberry (control) compared with those
treated with Aloe vera gel. The antifungal activity of Aloe vera has
been reported against postharvest fruit pathogens, such as Penicillium digitatum, Penicillium expansum, Botrytis cinerea and Alternaria alternata and was based on the suppression of germination
and the inhibition of mycelial growth (Serrano et al., 2006).
In addition, the inhibitory effects of Aloe vera gel have been also
found in Aspergillus niger, Cladosporium herbarum and Fusarium
moniliforme (Ali, Shalaby, Elgamai, & Mousa, 1999), although the
specific mechanism of action is still unknown. Moreover, the
reduction of the growth of 17 bacteria by A. vera gel has been
proven (Reynolds & Dweck, 1999). Some individual components
found in A. vera gel, such as saponins, acemannan and anthraquinones derivatives, are known to have antibiotic activity, and could
be responsible for its antibacterial activity (Serrano et al., 2006).
3.2. Total soluble solids, titratable acidity, pH and ascorbic acid
content
The TSS, TA and ascorbic acid levels in raspberry fruits decreased
in all treatments after 8 days of storage (Table 1). Compared with
the control samples, the Aloe treated fruits had higher TSS, TA and
ascorbic acid levels, and the difference was significant (p < 0.01).
Depletion of TSS in the fruit could be explained by a high metabolism of the fruits and senescence processes. Martnez-Romero
et al. (2006) observed that sweet cherry fruits treated with Aloe
vera gel maintained higher levels of TSS. The results obtain in this
study are in agreement with their results (Table 1). Organic acids of
fruit are substrates that are consumed by respiration during storage
(Ancos, Gonzalez, & PilarCano, 1999). At harvest, fruits had the
highest titratable acidity but, its content gradually decreased. The
declining of TSS and TA resulted in decreasing TSS to TA ratio
(Table 1). Titratable acidity of berries treated with Aloe vera gel had
not signicant difference with untreated fruits after 8 days of
storage (Table 1). Thus, at the end of storage, no signicant differences were observed among the treatments, and this is in agreement with results of Serrano et al. (2006). The change in pH of
raspberry fruits is shown in Table 1. In general, the pH of raspberry
fruits increased after 8 days of storage, while no signicant differences were observed between coated and uncoated samples.
Ascorbic acid is primarily regulated by ascorbic acid oxidase and
phenoloxidase, whose activities are inuenced by the oxygen
contents in the storage condition (Zhou et al., 2008). In this study, A.
vera coating was more effective in the retention of TSS, TA and
ascorbic acid levels because of the lowest gas permeability of A. vera
coating that inhibited the respiratory rates and retarded the overall

Table 1
Effect of different Aloe vera gel treatment on the quality attributes of raspberry
(Rubus spp.) initially and at the end of the cold storage period at 4
C.
Treatments

Decay (%)

TSS (%)

TA (%)

TSS/TA

pH

ASA
(mg/100 g FW)

7.93a

2.16a

3.67b

3.46b

108.1a

6.7b
7.22a
7.13a
7.17a

1.19b
1.29b
1.25b
1.20b

5.63a
5.59a
5.70a
5.98a

3.84a
3.86a
3.75a
3.78a

3. Results and discussion

3.1. Fungal decay incidence
Raspberry fruit is highly perishable, due to the relatively high
water content, high metabolic activity and the susceptibility to
microbial molds and rots. Fungal decay increased continuously
during storage and resulted in deterioration of about 22.54 percent

At harvest
0.0c
After 8 days of storage
control
22.54a
25% A. vera 13.96b
50% A. vera 10.55b
75% A. vera 10.55b

63.56c
85.56b
79.52b
81.25b

Values within a column followed by the same letter are not signicantly different at
p 0.05 (Duncan's Multiple Range test).

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H. Hassanpour / LWT - Food Science and Technology 60 (2015) 495e501

metabolic activities of raspberry fruits during storage. Valverde

et al. (2005) and Martnez-Romero et al. (2006) have found that
grapes and sweet cherries coated with Aloe vera gel showed
retardation of the ripening process by delaying evolution of the
parameters related to organoleptic quality, as well as by a reduction
of fruit decay, with high acceptance from the sensory panel.
3.3. Total anthocyanin
Anthocyanins are regarded as important antioxidants in berry
fruits (Mullen et al., 2002). Total anthocyanin content was signicantly affected by gel treatment and the storage period. As observed
in Fig. 1A, after 8 days of storage time the berries treated with 50
and 75% gel maintained the highest level of anthocyanin as
compared with other treatment and control fruits. As storage
advanced, the levels of anthocyanin increased in both control and
treated berries, the increase being signicantly higher in the Aloe
vera-treated fruits than in the control (Fig. 1A). Increases in
anthocyanin content during storage have been previously reported
for raspberries, strawberries, low bush blueberries and high bush
blueberries (Kalt, Forney, Martin, & Prior, 1999). The increase in the
total amount of anthocyanin during storage time may be due to the
continued biosynthesis of phenolic compounds after harvest,
related to the ripening processes. Also, it is probable that Aloe vera
gel upregulate anthocyanin production in raspberry by stimulating
gene expression of enzymes in the anthocyanin biosynthetic
pathway, such as PAL.
Anthocyanins are a group of phenolic compounds responsible
for the red-blue color of many fruit and vegetables, and provide
benecial effects for human health (Garca-Alonso, Rimbach, RivasGonzalo, & Pascual-Teresa, 2004). The antioxidant capacity of
anthocyanin may be one of their most signicant biological properties (Wang, Cao, & Prior, 1996). Therefore, it is important to
maintain higher levels of these compounds during storage and
shelf life period. Ayala-Zavala, Wang, Wang & Gonz'alez-Aguilar
(2005) found that the strawberries treated with methyl jasmonate showed the highest anthocyanin content at the end of the
storage period. Also, our results in agreement with the results of
Serrano et al., (2006) in table grapes coated with A. vera gel, and
Ayala-Zavala et al., (2005) in strawberries treated with methyl
jasmonate.
3.4. Total phenol
Total phenol of raspberry fruits was affected by storage time and
different concentrations of Aloe vera gel (Fig. 1B). After 8 days of
storage time, the fruits treated with 50 and 75% gel had the highest
level of total phenol as compared with other treatment and control
fruits (Fig. 1B). Our results in agreement with the results of Serrano
et al. (2006) in Crimson Seedless table grapes coated with A. vera
gel. The application of Aloe vera gel as an edible coating in raspberry
fruits imparted benecial effects in terms of maintenance of total
phenol during storage (Langmead, Makins, & Rampton, 2004). The
evolution of total phenolics of fruits during storage could be
different; depending on the species, cultivar, temperature, and
climactic and environmental conditions during the growth period
(Kalt, 2005). Thus, total phenolics of raspberry increased during
storage at 4
C, these Increases have been previously reported for
raspberries, strawberries, low bush blueberries and high bush
blueberries without coatings (Kalt et al., 1999). The accumulation of
phenolic compounds shown in raspberry fruits treated with A. vera
coating may be promoted by phenylalanine ammonia lyase (PAL)
activity. Meng, Li, Liu, and Tian (2008) observed an increase in PAL
activity during storage in table grapes treated with chitosan. Also,
raspberry titratable acidity decreases during storage, and organic

Fig. 1. Total anthocyanins (A), Total phenolics (B) and Antioxidant capacity (C) values
in raspberry fruit treated with Aloe gel and stored for 8 days at 4
C. Gel 1: 0% Aloe vera
(control); gel 2: 25% Aloe vera; gel 3: 50% Aloe vera; gel 4: 75% Aloe vera. Vertical bars
represent standard error.

acids, through interconversion with carbohydrates, may provide

carbon skeletons for the synthesis of phenolics, including both
anthocyanin and non-anthocyanin phenolics (Kalt et al., 1999).
3.5. Antioxidant capacity
In our study, antioxidant capacity increased in raspberry fruits
treated with the different concentrations of Aloe vera gel. The high
antioxidant capacity was found in raspberry fruits that were treated
with 50 and 75% and followed by 25% and control (Fig. 1C). Several
previous studies have shown that berries are a good source of
natural antioxidants (Heinonen et al., 1998; Wang et al., 1996).
These natural plant antioxidants include phenolic compounds and
anthocyanins (Hassanpour et al., 2011). Serrano et al. (2006)
observed that Aloe vera gel treated grapes increase antioxidant
activity. Also, Hu, Hu, and Xu (2005) demonstrated that Aloe vera
gel may increase the resistance of tissues to decay through
enhancing their antioxidant system and their free radical

H. Hassanpour / LWT - Food Science and Technology 60 (2015) 495e501

499

scavenging capability. It has been reported that the oral ingestion of

Aloe vera was highly effective as an anti inammatory due to its
antioxidant properties (Langmead et al., 2004). A large number of
compounds have been reported in the composition of Aloe vera gel
(Ni et al., 2004), but it is thought that aloe-emodin is one of the
main components that contributes to antioxidant activity. Moreover, it has been recently postulated that extracts from Aloe vera
have greater antioxidant activity than butylated hydroxy toluene or
R-tocopherol (Hu et al., 2005). In this sense, higher antioxidant
capacity observed in Aloe vera treated raspberry fruits as well as its
maintenance during storage could be attributed to the presence of
these compounds (Serrano et al., 2006). In addition, the synthesis of
both anthocyanin and non-anthocyanin antioxidants may have
contributed to the increase of antioxidant activity in raspberry
fruits (Kalt et al., 1999).
3.6. PAL enzyme activity
As shown in Fig. 2, the activity of PAL in raspberry fruits treated
with Aloe vera gel increased when compared with the control
(p < 0.05), showing that Aloe vera gel treatment triggered the key
enzymes of the secondary metabolites biosynthetic pathways in
raspberry fruits. Biosynthesis of phenols and avonoids in plants
perform via the shikimate-phenylpropanoid-avonoids pathways
(Tsai, Harding, Tschaplinshi, Lindroth, & Yuan, 2006). PAL as a key
enzyme in the phenylpropanoid pathway, catalyzes conversion of
rez-Balibrea, Moreno, and
phenylalanine to trans-cinnamic acid. Pe
Garca-Viguera (2011) suggested that the activity of PAL, a key
regulatory enzyme responsible for the phenylpropanoid metabolism, induced by SA treatment, could favor the phenols biosynthesis gene expression (Jahangir, Abdel-Farid, Kim, Choi, &
Verpoorte, 2009), triggering the accumulation of these secondary
metabolites in broccoli sprouts. The results showed that Aloe vera
gel treatment signicantly enhanced PAL activity in raspberry
fruits.
3.7. Antioxidant enzymes
3.7.1. Glutathione-peroxidase (EC 1.11.1.9) and glutathione
reductase (EC 1.6.4.2)
GSH-POD and GR activities of raspberry fruits stored for 8 days
were varied among treatments and storage durations as shown in
Fig. 3. After 8 days of storage, raspberry fruit from 50% Aloe vera gel
treatment had the highest activities. Raspberry fruits from the
control and 25% Aloe vera gel treatment had lower activities
compared with those from 25% to 75% Aloe vera gel treatment. After

Fig. 2. PAL enzyme activity in raspberry fruits treated with Aloe gel and stored for 8
days at 4
C. Gel 1: 0% Aloe vera (control); gel 2: 25% Aloe vera; gel 3: 50% Aloe vera; gel
4: 75% Aloe vera. Vertical bars represent standard error.

Fig. 3. Superoxide dismutase (SOD), Glutathione reductase (GR) and Glutathione

peroxidase (GSH-POD) activity in raspberry fruits treated with Aloe gel and stored for 8
days at 4
C. Gel 1: 0% Aloe vera (control); gel 2: 25% Aloe vera; gel 3: 50% Aloe vera; gel
4: 75% Aloe vera. Vertical bars represent standard error.

8 days' storage, samples from the control treatment had the lowest
GSH-POD and GR activities. In this study, both GR and GSH-POD
activities reduced in raspberry fruits stored for 8 days. GR
enzyme activity rst increased during the storage period, and then
reduced after 8 days of storage. 50% Aloe vera gel had the highest GR
activity. GR is a ubiquitous NADPH dependent enzyme. It adds
hydrogen ion to the oxidized glutathione to regenerate reduced
glutathione. In contrast, GSH-POD activity reduced during the
storage period. This enzyme utilizes reduced glutathione to eliminate hydrogen peroxide and convert it to harmless water. The activity of GSH-POD is dependent on the availability of the reduced
ascorbate and GSH that are maintained by enzymes, such as GR
using NADPH as an electron donor (Roxas, Lodhi, Garrett, Mohan, &
Allen, 2000).
3.7.2. Superoxide dismutase (EC 1.15.1.1)
SOD enzyme activities of raspberry fruits stored for 8 days were
varied among treatments and storage durations as shown in Fig. 3.
SOD activities reduced with storage time. SOD activity of raspberry
fruits remained steady during the first 3 days of storage and then
decreased during the rest of storage. The entire raspberry fruits
treated with Aloe vera gel had higher SOD enzyme activities than

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H. Hassanpour / LWT - Food Science and Technology 60 (2015) 495e501

those from control fruit. SOD enzyme, a class of metal containing

proteins, catalyze the dismutation reaction of superoxide radical
anions into H2O2 and molecular oxygen (Scandalios, 1993). Three
different types of SOD enzyme exist according to their metal
cofactor: Cu/Zn-SOD, Mn-SOD, and Fe-SOD (McKersie et al., 1993).
However, in this study, we measured only the total SOD enzyme
activity in raspberry fruits treated with Aloe vera gel. The results
showed that SOD activities reduced in all treatments during
storage.
3.7.3. Ascorbate peroxidase (EC 1.11.1.11) and guaiacol peroxidase
(EC 1.11.1.7)
ASA-POD and G-POD activities of raspberry fruits treated with
Aloe vera gel increased initially and then decreased during storage
(Fig. 4). After 8 days of storage, 50% Aloe vera gel had the highest
ASA-POD activities in raspberry fruit; while 75% Aloe vera gel
treatment had the highest G-POD activities. The control treatment
had the lowest ASA-POD and G-POD activities. ASA-POD, involved
in the detoxification of H2O2, uses two molecules of ascorbate to
reduce H2O2 to water (Noctor & Foyer, 1998). Since the biological
function of G-POD and ASA-POD is removal of H2O2 and other hydroperoxides, higher activities of G-POD and ASA-POD in treated
raspberry fruits with Aloe vera gel would be benecial for protection against lipid peroxidation and DNA hydroperoxides in the
fruits (Chaudiere & Ferrari-Iliou, 1999).
3.8. Non-enzyme component measurements
3.8.1. Reduced glutathione, and oxidized glutathione
The amounts of GSH enzyme varied from 50.32 to
22.01 mmol kg1 fresh berries and GSSG enzyme ranged from 24.32
to 10.02 mmol kg1 fresh weights as shown in Fig. 5. Initial GSH and
GSSG activities of raspberry fruits were 22.01 and 10.02 mmol kg1
fresh weights, respectively. During the 8 days of storage, GSSG and
GSH activities of raspberry fruits increased and then decreased.

Fig. 5. Oxidized glutathione (GSSG) and reduced glutathione (GSH) content in raspberry fruits treated with Aloe gel and stored for 8 days at 4
C. Gel 1: 0% Aloe vera
(control); gel 2: 25% Aloe vera; gel 3: 50% Aloe vera; gel 4: 75% Aloe vera. Vertical bars
represent standard error.

After 8 days of storage, 50% Aloe vera gel had the highest GSSG and
GSH activities on raspberry fruits (Fig. 5). The important function of
GSH enzyme is the maintenance of cellular redox status
(Rennenberg, 1980). The primary oxidation product of GSH is its
disulfide, GSSG, which can be reduced back to GSH by glutathione
reductase at the expense of NADPH (Ric de Vos, Kraak, & Bino,
1994). The result showed that 50% Aloe vera gel enhanced the increase of GSH enzyme in raspberry fruits during storage. This
reduced form of glutathione serves as a substrate for DHAR and
reacts directly with free radicals, including hydroxyl, to prevent the
deactivation of enzymes by oxidation of the essential thiol group
(Ziegler, 1985).
4. Conclusions
In general, this study showed that raspberry fruits treated with
Aloe vera gel had higher antioxidant capacity, enzyme activity and
less decay than control fruit. These results suggest that Aloe vera gel
treatments may be a useful non-chemical way of maintaining
raspberry fruit quality and extending their postharvest life. Further
investigation is needed to elucidate the underlying relationship
between Aloe vera gel treatment and antioxidant capacity in raspberry fruits.
Acknowledgments
I would like to thank Debbie Bright for critical reading of the
manuscript.
References

Fig. 4. Ascorbate peroxidase (AsA-POD) and Guaiacol peroxidase (G-POD) activity in

raspberry fruits treated with Aloe gel and stored for 8 days at 4
C. Gel 1: 0% Aloe vera
(control); gel 2: 25% Aloe vera; gel 3: 50% Aloe vera; gel 4: 75% Aloe vera. Vertical bars
represent standard error.

Ali, M. I. A., Shalaby, N. M. M., Elgamai, M. H. A., & Mousa, A. S. M. (1999). Antifungal
effects of different plant extracts and their major components of selected Aloe
species. Phytotherapy Research, 13, 401e407.
Amako, K., Chen, G. X., & Asada, K. (1994). Separate assay specific for ascorbate
peroxidase and guaiacol peroxidase and for the chloroplastic and cytosolic

H. Hassanpour / LWT - Food Science and Technology 60 (2015) 495e501

isozymes of ascorbate peroxidase in plants. Plant and Cell Physiology, 35,
497e504.
Ancos, B. D., Gonzalez, E., & PilarCano, M. (1999). Differentiation of raspberry varieties according to anthocyanin composition. Zeitschrift fr Lebensmitteluntersuchung und-Forschung A, 208, 22e38.
Ayala-Zavala, F. J., Wang, S. Y., Wang, C. Y., & Gonzalez-Aguilar, G. A. (2004). Effect of
storage temperatures on antioxidants capacity and aroma compounds strawberry fruit. Food Science and Technology, 37, 687e695.
Ayala-Zavala, J. F., Wang, S. Y., Wang, C. Y., & Gonzalez-Aguilar, G. A. (2005). Methyl
jasmonate in conjunction with ethanol treatment increases antioxidant capacity, volatile compounds and postharvest life of strawberry fruit. European
Food Research and Technology, 221, 731e738.
Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of
microgram quantities of protein utilizing the principle of protein-dye binding.
Analytical Biochemistry, 72, 248e254.
Castillo, F. J., & Greppin, H. (1988). Extracellular ascorbic acid and enzyme activities
related to ascorbic acid metabolism in Sedum album L. leaves after ozone
exposure. Environmental and Experimental Botany, 28, 232e233.
Cha, D. S., & Chinnan, M. (2004). Biopolymer-based antimicrobial packaging: a review. Critical Reviews in Food Science and Nutrition, 44, 223e237.
Chaudiere, J., & Ferrari-Iliou, R. (1999). Intracellular antioxidants: from chemical to
biochemical mechanisms. Food and Chemical Toxicology, 37, 949e962.
Deepa, N., Charanjit, K., Balraj, S., & Kapoor, H. C. (2006). Antioxidant activity in
some red sweet pepper cultivars. Journal of Food Composition and Analysis, 19,
572e578.
Eshun, K., & He, Q. (2005). Aloe vera: a valuable ingredient for the food, pharmaceutical and cosmetic industries: a review. Critical Reviews in Food Science and
Nutrition, 44, 91e96.
Garcia-Alonso, M., Rimbach, G., Rivas-Gonzalo, J. C., & Pascual-Teresa, S. (2004).
Antioxidant and cellular activities of anthocyanins and their corresponding
vitisins studies in platelets, monocytes, and human endothelial cells. Journal of
Agricultural and Food Chemistry, 52, 3378e3384.
Haffner, K., Rosenfeld, H. J., Skrede, G., & Wang, L. (2002). Quality of red raspberry
(Rubus idaeus L.) cultivars after storage in controlled and normal atmospheres.
Postharvest Biology and Technology, 24, 279e289.
Hassanpour, H., Hamidoghli, Y., Hajilo, J., & Adlipour, M. (2011). Antioxidant capacity
and phytochemical properties of cornelian cherry (Cornus mas L.) genotypes in
Iran. Scientia Horticulturae, 129, 459e463.
Heinonen, M., Lehtonen, P., & Hopia, A. (1998). Antioxidant activity of berry and
fruit wines and liquors. Journal of Agricultural and Food Chemistry, 46, 25e31.
Hu, Q., Hu, Y., & Xu, J. (2005). Free radical-scavenging activity of Aloe vera (Aloe
barbadensis Miller) extracts by supercritical carbon dioxide extraction. Food
Chemistry, 91, 85e90.
Jahangir, M., Abdel-Farid, I. B., Kim, H. K., Choi, Y. H., & Verpoorte, R. (2009). Healthy
and unhealthy plants: the effect of stress on the metabolism of Brassicaceae.
Environmental and Experimental Botany, 67, 23e33.
Kalt, W. (2005). Effects of production and processing factors on major fruit and
vegetables antioxidants. Journal of Food Science, 47, 4639e4644.
Kalt, W., Forney, C. F., Martin, A., & Prior, R. L. (1999). Antioxidant capacity, vitamin
C, phenolics, and anthocyanins after fresh storage of small fruits. Journal of
Agricultural and Food Chemistry, 47, 4638e4644.
Langmead, L., Makins, R. J., & Rampton, D. S. (2004). Antiinammatory effects of
Aloe vera gel in human colorectal mucosa in vitro. Alimentary Pharmacology &
Therapeutics, 19, 521e527.
Martnez-Romero, D., Alburquerque, N., Valverde, J. M., Guillen, F., Castillo, S.,
Valero, D., et al. (2006). Postharves sweet cherry quality and safety maintenance by Aloe vera treatment: a new edible coating. Postharvest Biology and
Technology, 39, 93e100.
McKersie, B. D., Chen, Y., de Beus, M., Bowley, S. R., Bowler, C., Inze, D., et al. (1993).
Superoxide dismutase enhances tolerance of freezing in transgenic alfalfa
(Medicago sativa L.). Plant physiology, 103, 1155e1163.
Meng, X. H., Li, B. Q., Liu, J., & Tian, S. P. (2008). Physiological responses and quality
attributes of table grape fruit to chitosan preharvest spray and postharvest
coating during storage. Food Chemistry, 106, 501e508.

501

Monk, L. S., Fagerstedt, K. V., & Crawford, R. M. (1987). Superoxide dismutase as an

anaerobic polypeptide. Plant physiology, 85, 1016e1020.
Mullen, W., McGin, J., Lean, M. E. J., MacLean, M. R., Gardner, P., Duthie, G. G., et al.
(2002). Ellagitannins, avonoids, and other phenolics in red raspberries and
their contribution to antioxidant capacity and vasorelaxation properties. Journal
of Agricultural and Food Chemistry, 50, 5191e5196.
Ni, Y., Turner, D., Yates, K. M., & Tizard, I. (2004). Isolation and characterization of
structural components of Aloe era L. leaf pulp. International Immunopharmacology, 4, 1745e1755.
Noctor, G., & Foyer, C. (1998). Ascorbate and glutathione: keeping active oxygen
under control. Annual Review of Plant Physiology and Plant Molecular Biology, 49,
249e279.
rez-Balibrea, S., Moreno, D. A., & Garca-Viguera, C. (2011). Improving the
Pe
phytochemical composition of broccoli sprouts by elicitation. Food Chemistry,
129, 35e44.
Petersen, K., Nielsen, P. V., Lawther, M., Olsen, M. B., Nilsson, N. H., & Mortensen, G.
(1999). Potential of biobased materials for food packaging. Trends in Food Science and Technology, 10, 52e68.
Reynolds, T., & Dweck, A. C. (1999). Aloe vera leaf gel: a review update. Journal of
Ethnopharmacology, 68, 3e37.
Rennenberg, H. (1980). Glutathione metabolism and possible biological roles in
higher plants. Phytochemistry, 21, 2771e2781.
Ric de Vos, C. H., Kraak, H. L., & Bino, R. J. (1994). Ageing of tomato seeds involves
glutathione oxidation. Physiologia Plantarum, 92, 131e139.
Roxas, V. P., Lodhi, S. A., Garrett, K. D., Mohan, J. R., & Allen, R. D. (2000). Stress
tolerance in transgenic tobacco seedlings that overexpress glutathione Stransferase/glutathione peroxidase. Plant and Cell Physiology, 41, 1229e1234.
Scandalios, J. G. (1993). Oxygen stress and superoxide dismutes. Plant Physiology,
101, 7e12.
Serrano, M., Miguel, J., Guillen, F., Castillo, S., Martinez-Romero, D., & Valero, D.
(2006). Use of Aloe vera gel coating preserves the functional properties of table
grapes. Journal of Agricultural and Food Chemistry, 54, 3882e3886.
Singleton, V. L., Orthofer, R., & Lamuela-Raventos, R. M. (1999). Analysis of total
phenols and other oxidation substrates and antioxidants by means of Folin
Ciocalteu reagent. Methods in Enzymology, 299, 152e178.
Smith, I. K., Vierheller, T. L., & Thorne, C. A. (1988). Assay of glutathione reductase in
crude tissue homogenates using 5, 5-dithiobis (2-nitrobenzoic acid). Analytical
Biochemistry, 175, 408e413.
Tappel, A. L. (1978). Glutathione peroxidase and hydroperoxidase. Methods in
Enzymology, 52c, 506e513.
Thayer, W. S. (1990). Superoxide-dependent and superoxide-independent pathways for reduction of nitroblue tetrazolium in isolated rat cardiac myocytes.
Archives of Biochemistry, 276, 139e145.
Tsai, C. J., Harding, S. A., Tschaplinshi, T. J., Lindroth, R. L., & Yuan, Y. (2006). Genome
wide analysis of the structural genes regulating defense phenylpropanoid
metabolism in Populus. New Phytologist, 172, 47e62.
Valverde, J. M., Valero, D., Martnez-Romero, D., Guillen, F., Castillo, S., & Serrano, M.
(2005). A novel edible coating based on Aloe vera gel to maintain table grape
quality and safety. Journal of Agricultural and Food Chemistry, 53, 7807e7813.
Wang, H., Cao, G., & Prior, R. L. (1996). Total antioxidant capacity of fruits. Journal of
Agricultural and Food Chemistry, 44, 701e705.
Wrolstad, R. E. (1976). Color and pigment analysis in fruit products. Station Bull. 621.
Agric. Exp. Sta. Oregon Sta. University.
Yao, H. J., & Tian, S. P. (2005). Effects of a pre and post-harvest application of salicylic
acid or methyl jasmonate on inducing disease resistance of sweet cherry fruit in
storage. Postharvest Biology and Technology, 35, 253e262.
Zhou, R., Mo, Y., Li, Y., Zhao, Y., Zhang, G., & Hu, Y. (2008). Quality and internal
characteristics of Huanghua pears (Pyrus pyrifolia Nakai, cv. Huanghua) treated
with different kinds of coatings during storage. Postharvest Biology and Technology, 49, 171e179.
Ziegler, D. M. (1985). Role of reversible oxidation-reduction of enzyme thiols
disulfides in metabolic regulation. Annual Review of Biochemistry, 54, 305e329.