Postharvest Biology and Technology 91 (2014) 7277

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Postharvest Biology and Technology

journal homepage: www.elsevier.com/locate/postharvbio

Chitosan applications pre- or postharvest prolong raspberry shelf-life

quality
Jaqueline Visioni Tezotto-Uliana , Gabriela Possati Fargoni, Gabriela Maria Geerdink,
Ricardo Alfredo Kluge
Universidade de So Paulo, Escola Superior de Agricultura Luiz de Queiroz, 13418-900 Piracicaba, SP, Brazil

a r t i c l e

i n f o

Article history:
Received 17 September 2013
Accepted 31 December 2013
Keywords:
Rubus idaeus
Edible coating
Postharvest life
Decay
Ethylene
Anthocyanins

a b s t r a c t
Raspberries are fruit with high metabolism that makes them very perishable, impairing their storage
and shelf-life. Chitosan coatings have the potential to improve their postharvest life by reducing water
loss, respiration rate and decay incidence. The purpose of this work was to study the effect of different
concentrations of chitosan, applied pre- or postharvest, on the retention of quality attributes of fresh
raspberries. The chitosan concentrations tested were 0 (control), 0.5, 1.0 or 2.0%. The postharvest treatment was applied immediately after harvest, dipping the fruit in the solutions for 5 min. The pre-harvest
treatment was done with one hand-spray application per week for three weeks, starting when the fruit
were just turning pink. In both experiments the fruit were stored at 0 C and 90% RH. Pre- or postharvest
use of chitosan at 1 or 2% was effective in maintaining titratable acidity and retarding respiration and
ethylene production, weight loss and decay incidence. Application by both means resulted in the highest
chitosan concentrations accelerating a reduction of ascorbic acid contents. Firmness was maintained only
when the fruit were treated pre-harvest at 2%. Thus, application of chitosan at 1 or 2% postharvest and
2% pre-harvest was able to retain key raspberry quality attributes for 15 and 12 days, respectively.
2014 Elsevier B.V. All rights reserved.

1. Introduction
Recent research on the characteristics of cultivation, marketing
and nutritional properties of berries have attracted the interest of growers, researchers and consumers, especially because of
their high value and benets for human health when consumed
fresh. However, a common problem with berries is their high perishability which results in a fast ripening period and senescence,
hampering storage and marketing (El Ghaouth et al., 1991; Han
et al., 2004). The postharvest life of berries is generally determined
by their susceptibility to water loss, softening, mechanical injuries,
and especially to the presence of pathogens such as Botrytis cinerea
and Rhizopus sp (Mass, 1981; Reddy et al., 2000).
Various studies have proposed techniques to control pathogens
while preserving the quality of these fruit, such as modied atmospheres with high CO2 , cooling, heat and osmotic treatments,
irradiation and edible coatings (Mass, 1981; Brown, 1922; Vicente
et al., 2005; Fan et al., 2009; Castello et al., 2010; Velickova et al.,
2013). Edible coatings have been of increasing interest because of
their capacity to reduce respiration and transpiration rates, and
increase storage periods, rmness retention and decay control
(Debeaufort et al., 1998; Vu et al., 2011; Velickova et al., 2013).

Among the various coatings, chitosan has been tested in berries.

It is a cationic polysaccharide, with high molecular weight, obtained
from alkaline deacetylation of chitin, and is found abundantly in
nature, constituting the exoskeleton of crustaceans and insects and
the structure of the fungi (El Ghaouth et al., 1991). These characteristics allow chitosan to be considered a non-toxic, biodegradable
and biocompatible product (Azevedo et al., 2007). The advantages
of chitosan also include the relative low cost and that its use as
a food additive has been approved by the United States Food and
Drug Administration (USFDA) (Knorr, 1986).
Other studies have shown that the post harvest use of chitosan
helps to control diseases in fresh raspberries (R. idaeus L.) (Zhang
and Quantick, 1998; Han et al., 2004), but there has been no investigation into the pre-harvest use of this product on the raspberry
quality. Furthermore, there is a need for studies on the most effective concentration. This kind of information is extremely important
for growers to obtain the best results with chitosan. Therefore, the
purpose of this work was to study the effect of different concentrations of chitosan, applied pre- or post harvest, on the retention of
key quality attributes of raspberry.
2. Materials and methods
2.1. Fruit material

Corresponding author. Tel.: +55 19 34294136; fax: +55 19 34294100.

E-mail addresses: jaqueline.tezotto@usp.br, jaque vt@hotmail.com
(J.V. Tezotto-Uliana).
0925-5214/$ see front matter 2014 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.postharvbio.2013.12.023

Autumn Bliss raspberries (R. idaeus L.) were obtained from a

commercial orchard in Ibina, SP, Brazil, from three year old plants

J.V. Tezotto-Uliana et al. / Postharvest Biology and Technology 91 (2014) 7277

cultivated on an espalier system. The experiments were performed

in the 2011/2012 growing season during the summer.
For both pre- and postharvest treatments, the fruit were handpicked when they had reached full maturity (pinkish surface color).
Harvested fruit were selected for uniform color, shape and size
of the fruit, and absence of mechanical or pathogen injuries. The
harvested fruit were directly placed in commercial plastic containers (perforated polyethylene terephthalate, with 120 g fruit and no
modied atmosphere) which were set in polystyrene boxes containing ice sheets, and immediately transported to the laboratory
at So Paulo University (USP), Piracicaba, SP, Brazil.
2.2. Preparation of chitosan solutions
The irradiated chitosan powder (Opco Fenix pharmaceutical
industry, Brazil) extracted from crab and shrimp shells, was characterized as a food-grade, odorless and tasteless powder with low
deacylation.
The concentrations tested in this study were 0, 0.5, 1.0 and 2.0%
(w/v). The chitosan was previously dissolved using 30 m/L of acetic
acid (5%) per gram of chitosan, and then the volume was completed with deionized water. For homogenization, the solutions
were heated and stirred for 3 h, then neutralized with 1.0 N NaOH
to pH 5.5 and ltered.
2.3. Treatments
The postharvest treatment was done in the laboratory immediately after harvest. The raspberries were randomly divided into
four groups of 4.2 kg (35 plastic containers) each and dipped in 10 L
of the chitosan solution for 5 min. The fruit were allowed to dry for
40 min on absorbent paper at room temperature, placed back in the
plastic containers and then stored at 0 1 C and 90 5% RH, for 15
days.
The pre-harvest treatment was applied once a week for three
weeks between when the fruit were turning pink and the harvest point. For each concentration, in each weekly application,
20 L of chitosan solution was used, hand-sprayed in rows with 30
plants until complete coverage, totaling 13.33 L m2 (considering
one plant per 0.15 m2 ). The fruit were harvested the day following the last application, and after transportation, were stored at
0 1 C and 90 5% RH, for 12 days. The following assessments
were conducted with four replicates of 120 g each.
2.4. Respiration rate and ethylene production

73

in the same four packages, each containing 35 raspberries, from

the beginning to the end of the experiment. Decay incidence was
visually inspected, counting the number of fruit that showed signs
of fungal growth and mealiness (a condition of extreme softness
and oozing) and the results were expressed in %. To determine
weight loss, the raspberries were weighed at the harvest (day 0)
and thereafter each day, the results being expressed as percentage
loss of the initial total weight. For the following analyses, the fruit
were removed from cold storage and held at room temperature
(25 C, 65% RH) 24 h before analysis to simulate shelf-life. Firmness
was determined by a horizontal attening technique, representing
a ratio between applied force and measured attened area of the
fruit, this measured according to the smaller and larger diameters
of the presumed elliptical gure formed and the result expressed
in newtons (Calbo and Nery, 1995). The pectin solubilization percentage was determined by washing the samples with ethanol,
complexing with EDTA, pH adjustment and pectinase, according to
McCready and McCoomb (1952), and the determination was performed colorimetrically using the method described by Bitter and
Muir (1962), with alterations, reducing the fruit mass from 5 to 2 g
in each sample, and the aliquot used for the quantication, from
1 to 0.2 mL. The ascorbic acid content was determined by titration
with DCFI and results express in mg 100 g1 of fruit (Carvalho et al.,
1990). The titratable acidity was determined from 10 g pooled juice
sample diluted with 90 mL distilled water, titrated with 1 N NaOH
to pH 8.1 and expressed in % citric acid. The anthocyanin contents
were determined according to Lees and Francis (1972) and was
expressed in g 100 g1 of fruit. The pulp masses were reduced from
100 to 10 g and homogenized in a 10 mL ethanol and HCl solution.
The color index (CI) to express the intensity of the red color was
determined using a colorimeter (model CR-400, Konica Minolta
Inc., Japan), calculated as CI = 100 a/(L b). The CI ranged from
35 to 75, with a greater CI value indicating a more intense red color
of the fruit.
2.6. Statistical analysis
Data analyses were performed by GLM (general linear model)
using SAS statistical software 9.2 (SAS Institute, Cary, NC, USA).
Multiple comparisons among the treatments with signicant differences tested were conducted by using Tukey test (P < 0.05).
3. Results
3.1. Respiration rate and ethylene production

The respiratory rate and ethylene production analyses were performed on the harvest day, the following day and thereafter every
two days. For the analysis, eight raspberries were placed into 80 mL
glass asks and hermetically sealed for 30 min at the same temperature and relative humidity conditions of the cold storage. A 0.5 mL
sample of internal atmosphere was collected through a silicone septum tted in each ask lid and measured by ame ionization gas
chromatography using a gas chromatograph (model Trace GC Ultra,
Thermo Electron Corporation, MA, USA), which was equipped with
two ame ionization detectors (FID), two injectors, two Porapack N
columns and one methanator set to 350 C. The results were determined considering the chromatographic values, fruit mass, ask
volume and the time it remained closed. The respiratory rate was
expressed in mg kg1 h1 of CO2 and ethylene production in L
kg1 h1 of C2 H4 .

We hypothesized that the use of chitosan as coating could reduce respiration

and ethylene production fresh raspberries and thus extend their postharvest life.
The data revealed that the postharvest use of chitosan at 2% reduced the respiration
of the fruit when compared to the control from the rst day after harvest (Fig. 1a).
This behavior also occurred with the fruit treated with chitosan at 1% starting from
the ninth day. When the application was made pre-harvest, the fruit treated with
1 and 2% had lower respiratory rates from the harvest day, and during the storage
period the three chitosan concentrations reduced the respiratory rate (Fig. 1b).
Regarding ethylene, we found that the higher the chitosan concentration used,
the lower was the ethylene production. In the postharvest treatments, the three
concentrations resutled in lower production compared to the control from the rst
day after harvest (Fig. 1c), while in the pre-harvest treatments, trhis only occurred
with the 1 and 2% treatments (Fig. 1d). We also observed that between harvest
and the rst day of storage, the respiratory rate and ethylene production were
respectively reduced more than 80 and 65% in both pre- and postharvest treatments, but then showed an increase through the evaluation period, related to the
cold temperature during storage. Thus, the data revealed that the application of 1
or 2% chitosan pre-or postharvest was effective in slowing respiration and ethylene
production.

2.5. Physicochemical and biochemical quality

3.2. Decay incidence and weight loss

Qualitative analyses were carried out at harvest day (day 0), and
thereafter every three days. Decay and weight loss were measured

One of the main benets from the chitosan application was decay control,
which was observed in both the pre- and postharvest applications. The postharvest
concentrations of 0.5, 1 and 2% reduced decay incidence by 75.5, 80.9 and 88.9%,

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J.V. Tezotto-Uliana et al. / Postharvest Biology and Technology 91 (2014) 7277

Respiration rate (mg CO2 kg-1 h-1)

80

80
a

70

Postharvest

60

60

50

50

40

40

30

30

20

20

10

10

Pre-harvest

0
0

11

13

15

14
Ethylene production (LC2H4 kg-1 h-1)

70

11

11

14
c

12

12

10

10

0
0

11

13

15

Days after harvest

Days after harvest

0%

0.5%

1%

2%

Fig. 1. Respiration and ethylene production of Autumn Bliss raspberries treated with chitosan postharvest (a and c) or pre-harvest (b and d) and cold stored at 0 C and 90%
of RH. Vertical bars represent the standard error (n = 4).

Decay rate (%)

40
35

40

Postharvest

35

30

30

25

25

20

20

15

15

10

10

0
0

Weight loss (%)

Pre-harvest

12

15
6

12

6
9
Days after harvest

12

0
0

12

15

Days after harvest

0%

0.5%

1%

2%

Fig. 2. Decay and weight loss of Autumn Bliss raspberries treated with chitosan postharvest (a and c) or pre-harvest (b and d), and cold stored at 0 C and 90% of RH. Vertical
bars represent the standard error (n = 4).

J.V. Tezotto-Uliana et al. / Postharvest Biology and Technology 91 (2014) 7277

0.9

1.2

Postharvest

1.1

Pre-harvest

1.0

0.8
Firmness (N)

75

0.9

0.7

0.8
0.6

0.7

0.5

0.6
0.5

0.4

0.4

0.3

0.3
0

12

15

12

12

12

6
9
Days after harvest

12

Pectin Solubilization (%)

35

100
80

25
20

60

15

40

10
5
0

12

15

Titratabel acid (%)

1.9
1.8

20
2.2

2.1

1.7

1.9

1.6

1.8
1.5
1.7
1.4

1.6

1.3

1.5

1.2

1.4
0

Ascorbic acid (mg 100g-1)

30

32
30
28
26
24
22
20
18
16
14
12

12

15
32

h
30
28
26
24
22
20
18
0

6
9
12
Days after harvest
0%

15
0.5%

1%

2%

Fig. 3. Physicochemical analyses of Autumn Bliss raspberries treated with chitosan postharvest (a, c, e and g) or pre-harvest (b, d f and g), and cold stored at 0 C and 90%
of RH. Fruit were removed from the cold storage and held at room temperature (25 C, 65% RH) 24 h before the analysis to simulate shelf-life. Vertical bars represent the
standard error (n = 4).
respectively, at the 12th day, although the treatments extended the postharvest
life until the 15th day after harvest (Fig. 2a). In the pre-harvest application, the 1
and 2% concentrations reduced decay incidence by 13.98% and 27.96% respectively,
extending the postharvest life until the 12th day (Fig. 2b).
We found that the weight loss was slightly reduced by all pre- and postharvest treatments, but were not statistically different compared to the control (Fig. 2c
and d).
3.3. Firmness, titratable acidity and ascorbic acid
Firmness maintenance is one of the most important physical attributes in maintaining quality of raspberries. The postharvest application of chitosan was not
effective in this aspect, and all treatments suffered an average loss of 45% in rmness

(Fig. 3a). However, when the treatment was applied pre-harvest, the fruit treated
with 2%chitosan remained rmer than the untreated and 0.5% treated fruit at from
day 0 to 9 (Fig. 3b).
Pectin solubilization analysis was performed in order to conrm the rmness
results. In the postharvest application the control fruit had higher solubilized pectin
percentages than the other treatments in the last two days of analysis. However, the
different concentrations of chitosan showed the same behavior, with an average of
26.3% of the pectin solubilized by the 15th day (Fig. 3c). In the pre-harvest application,
the fruit treated with 2%chitosan differed from the others at the harvest and ninth
days, although throughout the experimental period, fruittreated with 0.5% had the
lowest solubilization (Fig. 3d). Thus, we observed that the postharvest application
of chitosan did not prevent the loss of rmness in raspberries, but pre-harvest use
at 2% was able to slow this loss.

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J.V. Tezotto-Uliana et al. / Postharvest Biology and Technology 91 (2014) 7277

75

Index color

70

75

Postharvest

70

65

65

60

60

55

55

50

50

45

45

40

40

35

35
0

Anthocyanins content (g 100g-1)

Pre-harvest

12

15

12

6
9
Days after harvest

12

45

45
c

40

40

35

35

30

30

25
25

20

20

15

15

10

10

5
0

12

15

0.5%

1%

Days after harvest

0%

2%

Fig. 4. Index color and anthocyanin content of Autumn Bliss raspberries treated with chitosan postharvest (a and c) or pre-harvest (b and d), and cold stored at 0 C and
90% of RH. Fruit were removed from the cold storage and held at room temperature (25 C, 65% RH) 24 h before the analysis to simulate shelf-life. Vertical bars represent the
standard error (n = 4).
There was a decrease in titratable acidity until the sixth day in the postharvest
application, regardless of the chitosan concentration (Fig. 3e). From this day, the
fruit treated with 2% chitosan retained constant acidity. With the other treatments,
acidity kept decreasing, but the 1% treatment had showed the lowest decrease in the
12th and 15th days. In the pre-harvest treatment, we observed a decrease in acidity
during the entire period, but the fruit treated with 2% chitosan had the highest
acidity percentage compared to the other treatments (Fig. 3f). Thus, the use of 1 and
2% chitosan postharvest, and 2% pre-harvest were the most effective in maintaining
the acidity of raspberries.
The ascorbic acid contents of all fruit with postharvest applications were
reduced from day 0 to 3 in direct correlation with the chitosan concentration
(Fig. 3 g). Thereafter, the control fruit and those which received 0.5% maintained
constant ascorbic acid contents, whereas fruit treated at 1 and 2% continued to show
a decline, presenting 66.7 and 48.2% of the initial value, respectively at the 15th day.
In the pre-harvest experiment, we also observed a decrease in ascorbic acid content;
however, untreated fruit showed the smallest reduction (Fig. 3 h). Thus, we found
that the use of chitosan accelerated the reduction of the ascorbic acid content.
3.4. Color index and anthocyanin contents
Chitosan interacts with the main pigment of raspberry (Massa and Miniati,
1993), changing its color, so we studied the CI and anthocyanin contents.
Our results showed that its use in the 1 and 2% postharvest treatments resulted
in the smallest CI, indicating less darkening which is linked to better maintenance
of freshness (Fig. 4a). However, pre-harvest use had no inuence on the CI, which
increased by an average of 61.7% during the experimental period (Fig. 4b).
The anthocyanin contents followed a similar pattern, with the 1 and 2% postharvest application resulting in the lowest content (Fig. 4c), and the pre-harvest
application having no affect the pigment content (Fig. 4d). Thus, we conclude that
chitosan maintains the color and anthocyanin contents of the raspberries when
applied postharvest at 1 or 2%.

4. Discussion
The use of chitosan slowed respiration rates and ethylene
production of raspberries regardless of the application form. Several studies have shown that chitosan has excellent selective

permeability to the respiratory gases, acting as a barrier to the passage of O2 (Elsabee and Abdou, 2013). This control of gas exchange
between the fruit and environment reduces the respiration and the
action of ACC oxidase and synthase enzymes, which besides being
key enzymes of ethylene biosynthesis, are greatly inuenced by
the presence of O2 (Noh, 2005). In this study we found that the
application of c1 and 2% chitosan, both pre and postharvest, might
have formed a barrier around the fruit and reduced respiration and
ethylene production, which is directly correlated to the retention
of other fruit quality attributes.
Weight loss reduction from the use of chitosan is also related
to the formation of a selective barrier around the surface of the
fruit, which reduces moisture loss to the environment and reduces
respiration, the main metabolic processes that lead to water loss
(Han et al., 2004; Hong et al., 2012).
Decay was inuenced by both the chitosan concentration and
time of treatment. According to recent studies, the chemical structure of chitosan inhibits the growth of fungi and bacteria through
electrostatic forces between the protonated amino groups (NH2 )
of the chitosan, and the negative charges or phosphoryl groups
present on the cell surface of the microorganisms (Elsabee and
Abdou, 2013). Another reason for the benecial effects of chitosan
may be the reduction of polygalacturonase production by the fungi,
limiting its ability to colonize the fruit tissue (El Ghaouth et al.,
1997). In our study, the postharvest application possibly left a
greater amount of residue on the surface of the raspberries than
the pre-harvest, and in both applications, the highest chitosan concentration probably provided the greater the number of NH2 groups
and consequently had lower decay incidence.
The postharvest use of chitosan did not prevent rmness loss
of raspberries unlike that observed in most of the fruit treated
with pre-harvest applications, such as strawberry (Reddy et al.,

J.V. Tezotto-Uliana et al. / Postharvest Biology and Technology 91 (2014) 7277

2000), guava (Hong et al., 2012) and cherry (Martinez-Romero et al.,

2006). According to these authors, rmness maintenance is due to
respiratory rate reduction which decreased release of free radicals
and intensied pectinase activity, factors of cell wall degradation
and fruit softening (Kon and Schwimmer, 1977). In the postharvest
treatment, this process has probably not occurred due to persistent humidity around the fruit after application of the chitosan.
The difculty of drying the raspberries completely after dipped
in chitosan solutions was also reported by Han et al. (2004), who
suggested that spraying the chitosan solution might be the best
alternative.
The use of higher concentrations of chitosan minimized acidity loss in both experiments. This result is consistent with those
observed by Zhang and Quantick (1998) and Han et al. (2004) and
the other results from this study, because the higher percentage
of citric acid in the fruit treated with chitosan is related to lower
respiration rates and less fungal infestation, which results in longer
postharvest life.
The results observed for ascorbic acid content were not
expected. We predicted that the ascorbate contents of raspberries
would decrease through the experimental period, and that the use
of chitosan would minimize the loss rather than increase it. Since
chitosan might reduce the amount of O2 available for oxidative
reactions, one possible reason for this result is that the ascorbic
acid has reacted with other oxidizing agents present in the fruit
or with metallic ions that perhaps were in the chitosan solutions,
or has been involved in cell division during the ripening process
(Smirnoff, 1996). Therefore, more research is necessary to afrm
the real reasons for the ascorbate reduction in raspberries due to
the use of chitosan.
The postharvest application of higher concentrations of chitosan
was effective in reducing darkening and maintaining the anthocyanin contents, attributes directly related to freshness. The color
change in the raspberries is due to enzyme activity, especially
polyphenol oxidase and peroxidase, and the synthesis of anthocyanins. Activity of these enzymes is inuenced by the presence
of O2 inside the fruit, so when chitosan form a barrier around it,
enzyme activity will be reduced and consequently the darkening
of the fruit (Kang et al., 2005). It is possible that anthocyanin synthesis was reduced by the reduction in gas metabolism resulting
from the chitosan barrier. In addition, the positive charges present
in the coating may have stabilized the anthocyanins, helping to
maintain the raspberry color (Han et al., 2004). In the pre-harvest
treatment we did not observe this behavior, and this may be related
to the fruit metabolism of this fruit which might have not been
reduced enough to modify enzyme activity and the anthocyanin
synthesis.
In conclusion, we have observed that the postharvest application of 1 or 2% chitosan and 2% in the pre-harvest treatment
are able to retain key raspberry quality attributes for 15
and 12 days, respectively. Future studies should focus on the
properties of the chitosan coating, so that its use can be recommended.
Acknowledgement
The authors would like to thank the So Paulo Research
Foundation (FAPESP) for granting the scholarships. Grant n
2010/02601-3.

77

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