Nabifarkhani Cherry Nanocoposite Food Science
Published on: Mar 3, 2016
Transcripts - Nabifarkhani Cherry Nanocoposite Food Science
Sweet cherry due to early season fruit ripening and excellent
quality is one of the most popular fruit by consumers. The
main quality indexes are skin colour, which is related to
fruit ripening and affected by anthocyanin concentration
(Serrano et al. 2005a), and the ratio of total soluble solids
to total acidity (TSS/TA) at harvest time. Both parameters,
together with the absence of stem browning determine con-
sumer acceptance (Crisosto et al. 2003). TSS varied from
11 to 25 0
Brix, depending on cultivar and is mainly due to
glucose and fructose and less to the presence of sucrose and
sorbitol. In sweet cherry depending to the cultivar total acid-
ity ranged between 0.4–1.5% and malic acid is the main
organic acid (Esti et al. 2002; Bernalte et al. 2003). Colour
is one of the most important indicators of maturity and
quality of fresh, stored, and processed cherries fruit (Drake
et al. 1982). In cherries, colour is mainly affected by the
concentration and distribution of different anthocyanins in
the skin (Gao and Mazza 1995). The major anthocyanins
in sweet cherries include the 3-O-glucoside and 3-O-rutinoside
(D-rhamnosyl D-gluco pyranose) of cyanidin, with peonidin-
Effect of nano-composite and Thyme oil (Tymus
Vulgaris L) coating on fruit quality of sweet cherry
(Takdaneh Cv) during storage period
, Mehdi Sharifani1
, Amir Daraei Garmakhany2
, Ebrahim Ganji Moghadam3
Department of Horticulture Science, Gorgan University of Agricultural sciences and Natural Resources, Gorgan, Iran
Department of Food Science and Technology, Toyserkan Faculty of Industrial Engineering, Bu-Ali Sina University, Hamadan, Iran
Khorasan Razavi Natural Resource Agricultural Research Centre, Mashhad, Iran
Faculty of Chemistry, Tehran University, Tehran, Iran
© 2015 The Authors. Food Science & Nutrition published by Wiley Periodicals, Inc. This is an open access article under the terms of
the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium,
provided the original work is properly cited.
HPLC method, nano composite, storage
periods, sweet cherry, thyme oilthyme oil
Amir Daraei Garmakhany, Department of
Food Science and Technology, Toyserkan
Faculty of Industrial Engineering, Bu-Ali Sina
University, Beheshti Ave., Bahri Esfahani Ave.,
Toyserkan City, 6581869819, Hamadan, Iran.
Mehdi Sharifani, Department of Horticulture
Science, Gorgan university of Agricultural
sciences and Natural Resources, Gorgan,
No funding information provided.
Received: 18 December 2014; Revised: 25
January 2015; Accepted: 11 February 2015
Sweet cherry is one of the most appreciated fruit by consumers since it is an
early season fruit and has an excellent quality. In this study effect of active
nano composite formed from chitosan (as a matrix material), nano cellulose
ﬁber (1% concentration) and Thyme oils (Tymus Vulgaris L) at 1% concentra-
tion on fruits quality was investigated. Treated fruits were stored at 1°C for
5 weeks and changes of different qualities attributes including weight loss, total
acidity, TSS, anthocyanin, total sugar and malic acid content (by high perfor-
mance liquid chromatography (HPLC) method) were measured each week. Results
showed that nano composite and Thyme oil signiﬁcantly affect fruit’s water
retention and so decrease fruit weight loss and preserve anthocyanin (P < 0.05).
None of applied treatments had any signiﬁcant effects in comparison with control
in regard to acidity while total sugar content and TSS signiﬁcantly affected by
treatment compared to control samples. Result of HPLC analysis showed that
there was no signiﬁcant difference between different treatment and control sample
in term of malic acid concentrations during storage period but increase storage
time lead to increase malic acid concentration in all treatments. For conclusion
it can be Saied that fruits coating with nano-composite, lead to increase fruit
shelf life, better appearance and prevents fungal growth that may be due to
creation of an active packaging by these compounds.
2 © 2015 The Authors. Food Science & Nutrition published by Wiley Periodicals, Inc.
N. Nabifarkhni et al.Inﬂuences of nano-composite and Thyme oil
3-O-rutinoside and glucoside, as well as pelargonidin-3-O-
rutinoside occurring in much lower amounts (Goncalves
et al. 2004). Sweet cherry fruit deteriorate rapidly after harvest
and in some cases do not reach consumers at optical quality
after transport and marketing. Postharvest loss of quality in
cherries is characterized by bruising of the skin, softening
and loss of acidity (Bernalte et al. 2003), drying and brown-
ing of the stem (Kappel et al. 2002) and by fungal diseases
caused mainly by Monilinia fructicola, Botrytis cinerea (Ganji
Moghadam and Bosari 2009) and Penicillium expansum.
This fungal spoilage can lead to great economic losses, al-
though the occurrence of rots and their inﬂuence on sweet
cherry quality have been reported to be dependent on cultivar
and ripening stage at harvesting time (Esti et al. 2002; Kappel
et al. 2002). Several pre and postharvest technologies have
been used to control decay, but the postharvest use of
chemicals as fungicides is restricted in most countries and
consumers demand agricultural commodities without pesti-
cide residues (Wilcock et al. 2004). Among these technologies,
the use of edible coatings is traditionally used to improve
food appearance and conservation. They act as barriers dur-
ing processing, handling and storage, and do not solely retard
food deterioration enhancing its quality, but are safe due
to natural biocide activity, or to the incorporation of anti-
microbial compounds (Petersen et al. 1999). Different com-
pounds have mainly been used as edible coatings to prevent
weight loss, including wax, milk proteins, celluloses derivatives,
lipids, starch, zein, alginate, and chitosan (Cha and Chinnan
2004). Chitosan, obtained by deacetylation of chitin, might
be an ideal preservative coating for fresh fruit because of
its ﬁlm-forming and biochemical properties (Muzzarelli 1986).
Unlike other coating materials, chitosan is known to be
antifungal to several fungi, including Botrytis cinerea And
Rhizopus stolonifer (El Ghaouth et al., 1992), to induce
chitinase, a defense enzyme (Mauch et al. 1984), and to
elicit phytoalexin (pisatin) accumulation in pea (Pisum sa-
tivum L.) (Kendra and Hadwiger 1984). Furthermore, chitosan
is a by-product from the seafood industry, appears to be a
safe material as indicated by toxicological studies (Hirano
et al. 1990). Since chitosan can form a semi permeable ﬁlm
(Bai et al. 1988), coating fruit with chitosan may modify
the internal atmosphere of the tissue and consequently delay
ripening. Essential oils are natural colorless compounds con-
sisting of alcohols, aldehydes and esters that have fewer odors
and its molecular weight is lower than water. Essential oils
are volatile compounds that can be used as ﬂavoring, an-
tioxidant and antimicrobial agents in foods (Omid Beigi
2005). Also it can be used as edible coatings for fruits and
vegetables to increase the shelf life of agricultural products
(Park et al. 2002). According to their antioxidant role, they
may be used to prevent enzymatic browning reaction (Nicoli
et al. 1994). Some of the study reported the beneﬁcial effects
of natural essential oils on quality of different fruits
including grapes, avocados and cherries (Serrano et al. 2005b).
According to the introduction the aim of this study was
investigation the alteration of quality attributes and reduction
of decays in cherry fruit by use of edible coatings (nano
composite) formed from chitosan and nano cellulose and
Thyme oils (Tymus Vulgaris L) during storage periods.
Materials and Methods
In this study more than 20 kg sweet cherry fruits from
a commercial orchard located in Mashhad city of Iran
was collected. Applied nano composite was made from
chitosan (1% concentration) as the matrix material and
nano-cellulose (0.1% concentration). Thyme oil was dis-
solved in ethanol 25% and used in 1% concentration.
This project was conducted in three treatments; Control,
nano composite, Thyme oil. Treated fruits were stored
at a temperature of 1°C and 90% RH for 6 weeks and
each week quality attributes of fruits were measured.
The method to prepare the coating solutions was devel-
oped by Alikhani et al. (2012) with minor modiﬁcations.
The chitosan solution (1.0%) containing 0.5% acetic acid
as a solvent was stirred by a magnetic stirrer at room
temperature for 1 h to obtain complete dispersion. Then
the thyme oil, mixed with tween 80 (0.2%), to help dis-
tribute and completely incorporate the thyme oil, was
added to the chitosan solution and then stirred using a
magnetic stirrer for 30 min. The ﬁnal solution was cen-
trifuged for 10 min at 2268 g and the supernatant obtained
was used to prepare the edible coating.
Total soluble solids (Brix) and titratable acid
Total soluble solids (0
BX) were determined according to
AOAC (2005) using hand refrectometer at room tem-
perature. Titratable acid was determined by titration of
Figure 1. Effect of time and coating on titratable acid content in cherry
3© 2015 The Authors. Food Science & Nutrition published by Wiley Periodicals, Inc.
Inﬂuences of nano-composite and Thyme oilN. Nabifarkhni et al.
fruits juice by NaOH (0.1 N) until reach the pink color
and express as the mli eqi of malic acid per gram of
Fruit weight loss
Fruits weight loss for each treatment was calculated by
the following equation at each storage periods:
is initial weight of fruit samples and w2
of fruit samples at each storage periods.
Extraction and measurement of total sugar
(mg/100 g of fresh tissue)
Total sugar was extracted using Omokolo et al. (1996)
method. In this method, 40 mg of fresh fruits was taken
and mixed with 5 mL of ethanol (80%) for 10 min in a
hot water bath (70°C). The resulting extracts were centri-
fuged (1000 g) 15 min and obtained supernatant was trans-
ferred to a beaker, this process repeated 4 times on remained
the residual tissue. The extract was concentrated by heating
to 1/5 of its volume. The resulting aqueous phase was cen-
trifuged (10000 g) for 10 and clear upper phase was removed
and used for determination of soluble sugars.
Total soluble sugar was determined with McCready
et al. (1950) method. For determination of total sugar,
0.2 mL (200 mL) from produced extract mix with 3 mL
Antrun for 20 min in hot water bath (100°C). After cool-
ing the absorbance of each sample was measured at 620 nm.
Anthocyanin measurement (M/g of fresh
The amount of anthocyanin was measured by Wanger
(1979) method. For determination of anthocyanin 1 g of
fresh cherry fruit juice was taken and mixed with 10 mL
acidiﬁed methanol and kept in the dark place (4°C) for
24 h. Then, the extract was centrifuged (4000 g) for
10 min and absorbance of the supernatant was read in
520 nm using a spectrophotometer (S 2000 uv/vis).
Weight loss (%)=
Figure 2. Typical HPLC curve for malic acid determination.
Figure 3. Effect of time and coating on (A) weight loss (B) total soluble
solid and (C) soluble sugar content in cherry fruit.
4 © 2015 The Authors. Food Science & Nutrition published by Wiley Periodicals, Inc.
N. Nabifarkhni et al.Inﬂuences of nano-composite and Thyme oil
Malic acid measurement by HPLC method
Malic acid was quantiﬁed by high performance liquid chro-
matography (HPLC). The analysis was carried out using
a Shimadzu chromatograph, model LC-10 Ai (Shimadzu
Corp., Kyoto, Japan), equipped with an Ultra Violet detec-
tor (UV- SPD-6AV). A Shimadzu column C18 (Shim-pack
SCR-101H, separating column 7.9 mm×30 cm), was oper-
ated at 25°C for organic acids. Mobile phase was consisted
of isocratic and acidic dionized water with sulfuric acid
(pH = 2) at a ﬂow rate of 1 mL/min. The malic acid
content of the samples was detected via PID detector
(214 nm). The quantiﬁcation of malic acid was performed
using calibration curve obtained from the standard com-
pound. All samples were examined in duplicate. The sen-
sitivity of system was equal 3 and the temperature of
separating column was 25°C. The injection was carried out
using 20 µL volume of the sample with application of
Hamilton syringe. Before injection the samples passed
through the ﬁlter with pores diameter of 0.45 µ. The con-
centration of malic acid calculated using standard curve,
retention time and the space under the curve of each
This experiment was conducted in a completely randomized
factorial design. The ﬁrst factor consists of edible coatings:
control, nano composite, and thyme oil respectively. The
second factor is the storage time (0, 7, 14, 21, 28 and
35 days of storage) respectively. Results of this study were
analyzed using SAS (2001) software and means comparison
was performed by Duncan’s multiple range test (95%).
All the experiments and treatment were done with three
Results and Discussion
Based on the results (Fig. 1), the total acidity decreased
during the storage period. Nano composite and thyme
oil lead to preserve organic acids in cherry fruit signiﬁ-
cantly. Reduction of respiratory rate, lead to reduction
of the amount of organic acids consumption. Finidokht
et al. (2013) showed that chitosan by decrease respiration,
reduces the amount of organic acids consumption in cherry
fruit, while increase storage time lead to more consume
organic acids. They stated that the chitosan in short time
storage periods preserve organic acids while in long time
storage periods their protection properties will decrease.
Previous studies showed that chitosan had positive effect
on organic acids perseveration in tomato and strawberry
fruit (Kittur et al., 1998; Lydakis and Aked 2003).
Malic acid analysis using HPLC
Results of malic acid analysis using HPLC method showed
that the concentration of malic acid in control samples
was preserved with an increasing trend from 1st week to
3rd of storage periods (Fig. 2). In the ﬁrst week of storage
the malic acid concentration was 4.9 gr/lit and in 3rd week
of storage the concentration increased to 6.76 g/lit. However
the malic acid concentration at 6th week was 7.16 g/lit.
In 3rd week of storage period, application of essential
oil treatment caused a concentration of malic acid reached
to 5.67 g/lit and in the 6th week of storage period malic
acid concentration was 6.87 g/lit. This result revealed the
effect of essential oil on preservation of fruit water con-
tent. Higher malic acid concentration was due to the less
evaporation rate from fruit surface by using essential oil
treatment. There was no difference between control and
nano-composite treatment in term of malic acid concen-
trations until 3rd week of storage. The concentration of
Table 1. ANOVA of weight loss, TA, TSS, Total sugar and anthocyanin in sweet cherry as a function of storage time and coating type.
Surce of variation Df Weight loss (%) TA (mleq/g) TSS (%) Total sugar
Type of cover 2 14.3** 0.004* 1.28* 0.01* 0.454**
Time 5 27.98** 0.009** 3.05** 0.03** 6.35**
Type of cover × time 10 1.33** 0.003ns
Superscripts ns, * and ** means not signiﬁcant, signiﬁcant at 0.05 and 0.01 level respectively.
Figure 4. Effect of time and coating on antocyanin content of cherry fruit.
5© 2015 The Authors. Food Science & Nutrition published by Wiley Periodicals, Inc.
Inﬂuences of nano-composite and Thyme oilN. Nabifarkhni et al.
malic acid for nano-composite and control treatment was
7.3 and 7.16 g/lit in 6th week of storage respectively.
The ﬁnal conclusion of malic acid analysis among the
applied treatments indicated that 3 weeks of storage is
the most acceptable storage period for essential oil and
nano composite treatments, whereas there was no signiﬁ-
cant difference between applied treatments in term of of
malic acid concentration during storage period. It might
be offered that malic acid concentration is not a useful
criterion to judge the efﬁciency of treatments. Water status
of fruit, turgor pressure, antioxidant activity, sugar and
anthcyanin content are other crucial criteria’s which affect
judgment on efﬁciency of the treatments.
Weight loss, total soluble solid (TSS) and
Based on the results (Fig. 3A) the greatest weight loss
was related to control samples and nano composite and
thyme oil showed lower trend in weight loss than con-
trol samples which is in agreement with the results of
Finidokht et al. (2013) and Shamloo et al. (2013). They
stated that coated cherries with chitosan had lower water
loss than uncoated cherries. It can be concluded that
nano composite and thyme oil treatments by creation
a physical barrier against moisture loss from fruits skin,
prevent surface dehydration, fruits shrinkage and decrease
respiration during storage. Also the higher water loss
in uncoated Cherry than coated ones was due to an
increase in respiration and evaporation (Finidokht et al.
2013). Further research for example; Ebrahimpur
Komeleh et al. (2008) reported that Hindi clove oil at
concentration of 500 ppm, lead to the lowest percentage
of weight loss while control samples (uncoated) and
treated samples by cumin and clove oil fumigation has
the greater weight loss.
Results showed that TSS gradually increased during
storage (Fig. 3B) that may be due to activity of hydrolytic
enzymes or increase of water loss during storage period
(Alonso et al. 2003). At the end of storage period, the
lowest (16.5 0
BX) and highest (17.630
BX) amount of TSS
was related to nano composite and control samples re-
spectively. Type of coating agent and storage time had
signiﬁcant effect on amount of fruit TSS during storage
(Table 1). Our result in term of TSS was in agreement
with other studies (Vesal talab and Gholami; 2012). They
study the gradual increase in the amount of TSS during
storage of grape fruit treated with clove oil and clove
extract compared to control.
As can be seen from Figure 3C, nano composite and
thyme oil treated samples had lower content of soluble
sugar than control sample. Total sugar content increased
during storage period that may be due to the dehydration
and decomposition of organic acids (used as an energy
source) in the fruit (vesal talab and Gholami; 2012).
As can be seen in Figure 4, anthocyanins increase during
storage period. At the end of storage period, the lowest
(1.89 mol/g) and highest (2.63 mol/g) amount of antho-
cyanin was related to nano composite and control samples
respectively. Anthocyanin results were in agreement with
the results of Shoja et al. (2011). They showed that an-
thocyanins increases during storage in blood orange.
Applied treatments (nano composite and thyme oil) lead
to preserve TSS, anthocyanins and total sugar content
and reduce fruit weight loss in comparison with control
samples (P < 0.05) while there are no signiﬁcant differ-
ences between applied treatments (nano composite and
thyme oil) and control in regard to titrable acidity and
malic acid concentration. Results of this study showed
that application of edible coatings can be used to increase
fruit shelf life, better appearance of fruits and prevents
fungi growth that may be due to creation a sort of active
packaging by these compounds.
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