Food Packaging and Shelf Life 11 (2017) 4048

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Food Packaging and Shelf Life

journal homepage: http://www.elsevier.com/locate/fpsl

Evaluating the efcacy of moringa leaf extract, chitosan and

carboxymethyl cellulose as edible coatings for enhancing quality and
extending postharvest life of avocado (Persea americana Mill.) fruit
Samson Zeray Tesfaya,* , Lembe Samukelo Magwazaa,b
a
Discipline of Horticultural Science, School of Agricultural, Earth and Environmental Sciences, University of KwaZulu-Natal, Private Bag X01, Scottsville, 3209,
Pietermaritzburg, South Africa
b
Discipline of Crop Science, School of Agricultural, Earth and Environmental Sciences, University of KwaZulu-Natal, Private Bag X01, Scottsville, 3209,
Pietermaritzburg, South Africa

A R T I C L E I N F O A B S T R A C T

Article history:
Received 24 July 2016 This experiment was conducted to investigate a novel moringa leaf extract, together with commercially
Received in revised form 27 October 2016 available edible coatings, namely, chitosan and carboxymethyl cellulose (CMC), as postharvest
Accepted 17 December 2016 treatments to enhance shelf-life and improve the quality of Fuerte and Hass avocado fruit. Postharvest
Available online xxx treatment included a 2% moringa extract with an emulsier, two levels of chitosan (0.5, 1%), and CMC (0.5,
1%). Moringa extract with emulsier and moringa containing chitosan and CMC signicantly improved
Keyword: fruit quality of both cultivars. Fuerte fruit treated with the combination of CMC (1%) and 2% moringa had
Chitosan signicantly lower mass loss (1.78
0.08%), electrical conductivity (192.0
3.0 m/m) and respiration rate
Carboxymethyl cellulose (CMC)
(167.4
40.8 mg/kg/h) compared to the untreated control with respective values of 4.7
0.7%,
Edible coatings
290.0
5.0 m/m and 290.0
62.0 mg/kg/h. The same treatment had higher values for rmness
Antimicrobial
Antioxidant (50.0
4.25 N) and phytochemical characteristics, mainly mannoheptulose (5.7
0.6 g/kg), and lower
Polyphenol oxidase polyphenol oxidase (0.6
0.06 *1000 U/kg) and lipid peroxidation (0.89
0.06 nmol/g) in Fuerte fruit.
For Hass, similar results were also observed, where a combination of 2% moringa leaf extract with 1% of
CMC reduced mass loss almost by 50%, while mannoheptulose was maintained by 8-folds. The results
observed in this study showed that investigated edible coatings containing moringa leaf extract improve
fruit quality and shelf-life. It could therefore potentially be commercialized as a new edible coating for
future industry application.
2016 Elsevier Ltd. All rights reserved.

1. Introduction consequences to fruit quality and can lead to tremendous nancial

Avocado (Persea americana, Mill.) is an extremely perishable
fruit, with very high metabolic rate, resultant in a short postharvest
life of about three to ve weeks when stored under optimum
conditions (Yahia & Gonzalez-Aguilar, 1998). The postharvest
storage life of avocado fruit is limited by its climacteric ripening
pattern which exhibits high ethylene accumulation, stimulating
faster ripening as result of the high rate of respiration (Blakey,
Tesfay, Mathaba, Bertling, & Bower, 2012). Avocado fruit are also
considered to have a high postharvest mass loss, and this is mostly
due to moisture loss through transpiration, which has been shown
to contribute about 90% of the total fruit mass loss (Cutting &
Wolstenholme, 1992). Postharvest moisture loss has undesirable
* Corresponding author.
E-mail address: Tesfay@ukzn.ac.za (S.Z. Tesfay).
losses since the price of fruit is determined by its mass (Turner,
1997).
Loss of moisture from the fruit is predominantly due to water
pressure gradient (vapour pressure decit) between the less
saturated ambient atmosphere and the fruit, which is close to
saturation with water (Cutting & Wolstenholme, 1992; Ben-
Yehoshua, Fishman, Fang, & Rodov, 1994; Macnish, Joyce, &
Hetherington, 1997; Magwaza, Opara, Cronje, Landahl, & Terry,
2013a). The initial symptoms of excessive loss of moisture from an
avocado fruit are shriveling, which immediately appears on the
peel, and affected fruit loses its shine, softens and senesces.
Essentially, any postharvest treatment that reduces moisture loss
and maintain fruit turgidity and rmness has a potential to reduce
respiration and ripening.
Commercially, avocado, with the only exception of those
marketed as organic, are generally waxed to enhance appearance

http://dx.doi.org/10.1016/j.fpsl.2016.12.001
2214-2894/ 2016 Elsevier Ltd. All rights reserved.
 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048
and reduce moisture loss and the wax also functions as a medium
to apply fungicides thus, extending shelf-life (Kremer-Khne &
Duvenhage, 1997). Fruit supplied to the European Union market
are not waxed, resulting in major economic losses due to water loss
and short shelf-life (Kruger, 2013). Consumer preference of
unwaxed fruit also coincided with recent trends towards
organically produced fruit and vegetables. This increasing con-
sumer awareness for health and fruit quality, coupled with the
industry demand for environmentally friendly coatings for fresh
fruit have spurred a considerable interest among researchers to
develop edible, natural and food safe coatings for avocado fruit.
Edible coatings are an ecologically friendly substitutes applied
on fresh produce to reduce water transfer, gaseous exchange and
oxidation processes (Dhall, 2013). The composition of edible lms
and coatings can be categorised into different groups, namely,
hydrocolloids, lipids, proteins, polysaccharides and composite
material (Dhall, 2013). Another distinguished characteristics of
edible coatings are their high latent ability to be used as carriers of
postharvest active ingredients such as colorants, antimicrobial
compounds, spices, nutrients, avors, and anti-browning agents
that can reduce the risk of pathogens on food surface and prolong
produce postharvest life (Pranoto, Salokhe, & Rakshit, 2005).
Among edible coatings, chitosan and carboxymethyl cellulose
(CMC) have dominated the food industry. Retarding moisture loss
and extending shelf-life by using chitosan have been reported on
litchi (Dong, Cheng, Tan, Zheng, & Jiang, 2004). CMC have been
reported on avocado fruit (Maftoonazad & Ramaswamy, 2005). A
signicantly lower rate of respiration during storage of peaches
coated with chitosan compared to the untreated control was
reported by Li and Yu (2000). Banks et al. (1997) reported that
coating fruit with CMC (2 g/100 g) substantially increased the risk
of fermentation. In a later study, Salvador, Miranda, Aragon, and
Lara (1999) observed a longer storage life of 24 days at 310  C and
6 days at 2729  C on avocado fruit with chitosan.
Moringa (Moringa oleifera Lam., Moringaceae) is one of the most
useful trees in the tropics and subtropics of Asia and Africa. The
leaf, bark, sap, root, ower and seed extracts of moringa plants
possess antimicrobial and antioxidant activity, contributed by a
high concentration of phenolics, vitamins and carotenoids (Busani,
Julius, & Voster, 2012; Tesfay, Bertling, Odindo, Workneh, &
Mathaba, 2011a; Tesfay, Modi, & Mohammed, 2016). This identies
a vast potential for moringa plant parts to be used as an ingredient
in the development of different functional food products such as
edible coatings. Moringa plant is the most credible but cheap
alternative for not only coating fruit but also providing antimicro-
bial effects during postharvest storage. The addition of moringa
extracts into the edible coatings, namely corn starch and CMC have
been shown to slow down mass loss in citrus fruit (Adetunji et al.,
2012). Yousef, Abd El-Moniem, and Saleh, (2015) evaluated the
effect of moringa and other natural products on storability and
fruit quality properties of Fuerte avocado. However, there is
currently no research reporting the use of moringa leaf extract as
an edible coating material on Hass cultivar, the most important
commercial avocado variety. This study was therefore conducted
to investigate the efcacy moringa leaf extract, together with
commercial hydrophilic polysaccharide based edible coatings,
namely CMC and chitosan as a novel approach to enhancing quality
and shelf-life of Fuerte and Hass avocado fruit.
2. Materials and methods
2.1. Fruit samples
Avocado (Persea americana, Mill. cv. Fuerte and Hass) fruit
used in this study were harvested from Everdon Estates, a
commercial avocado farm of Westfalia fruit (Pty) LTD. The farm
41
is located in Howick, a cool subtropical area of KwaZulu-Natal
Province, South Africa (Latitude: 29 450 S, Longitude: 30 250 E),
typical of a mist-belt climate. Using dry matter as a maturity index,
Fuerte and Hass avocado fruit were harvested at commercial
maturity (Magwaza & Tesfay, 2015) when the mean dry matter
content in the esh was 19 and 20.8%, respectively. Harvested fruit
of Hass and Fuerte cultivars were packed in open display boxes
and transported immediately in a ventilated vehicle to the
Postharvest Laboratory of the University of KwaZulu-Natal.
2.2. Treatments and storage
A total of 360 Fuerte and Hass avocado fruit with an average
mass ranging from 191 to 210 g, were assigned to six coating
treatments (Untreated control, Moringa 2% (M), M + Chitosan 0.5%
(CN 0.5%), M + CN 1%, M + CMC 0.5%, M+ CMC 1%). Each treatment
consisted of 3 replicates of 20 fruit per replicate. Harvested fruit
were dipped in treatment solutions for 1 min and left for 30 min on
a laboratory bench top to dry at room temperature (21
1  C). In
the preliminary study on postharvest coatings, 2% had a positive
result when applied with other commercially found edible
coatings as compared to its different concentrations (Data not
presented). Fruit were then transferred to a cold room with a
delivery air of 5.5  C and RH of 95%
2%, simulating a shipment of
21 days. After which, fruit were removed from the cold room and
ripened in the laboratory at room temperature (21
1  C) and 60%
RH. Fruit sampling was taken every 7 days. Nine fruit were used for
physical as well as biochemical analysis. Destructive fruit sampling
was applied for biochemical analysis, where mesocarp tissue was
snap-shocked using liquid nitrogen and freeze-dried, ground and
stored at 20  C until further analysis.
2.3. Postharvest quality measurements
During the ripening stage at room temperature, fruit were
evaluated for various physico-chemical properties and quality
attributes, including time to ripen, rmness, respiration rate and
possibility of pathological and physiological disorders. At regular
intervals during storage, samples were taken for biochemical
analysis including electrical conductivity (EC), carbohydrates
concentration, polyphenol oxidase activity (PPO), lipid peroxida-
tion. At each interval, mesocarp of sampled fruit were snap frozen
using liquid nitrogen and freeze-dried using Virtis Benchtop freeze
drier system (ES Model, SP Industries Inc., Warmister, USA) for six
days at 0.015 kPa and 75  C. Lyophised mesocarp samples were
ground into ne powder and kept in a freezer at 20  C until
further biochemical analysis. In addition, samples of the peel were
taken for surface microstructural analysis using scanning electron
microscopy (SEM).
2.3.1. Determination of fruit rmness
Fruit rmness was determined every seven days during cold
storage using a hand-held rmness tester (Bareiss, Germany). Two
rmness measurements were taken in the equatorial region of
each fruit after rotating the fruit 180 . The rmness tester used
measures fruit rmness on a scale from 150 (very hard, unripe
fruit) to <60 (soft ready to eat fruit) (Magwaza & Tesfay, 2015).
2.3.2. Fruit CO2 evolution
Fruit CO2 evolution or respiration rate was measured every
seven days using an environmental gas monitor (EGM-1, PP
Systems, Hitchin, UK), a method heretofore dened by Tesfay,
Bertling, and Bower (2011b). Individual fruit were sealed for 10 min
in an airtight 1 L jar, where after the headspace CO2 concentration
was measured and the respiration rate calculated and expressed in
42 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048
mg/kg/h, after adjusting calculations taking into consideration
fruit mass, headspace and ambient CO2 concentration.
2.3.3. Determination of electrical conductivity
The EC from mesocarp tissue electrolyte leakage was deter-
mined using a multi-range conductivity meter (HI 9033, Hanna
Instruments, Johannesburg, South Africa) using a method by
Venkatarayappa, Fletcher, and Thompson (1984), with slight
modication. Briey, a mesocarp sample was taken from the
distal end of a half-cut fruit. An 11 mm thickness disc (2.02.5 g)
was cut from this piece of sample, washed three times in ultra-pure
water and placed for boiling in a tube containing 25 mL ultra-pure
water. This was then boiled and tubes continuously shaken for 3 h
tubes when the solution was ready for the EC analysis. The EC of
each sample was recorded before and after boiling, the electrolyte
leakage calculated as the EC*100 (m/m) using Eq. (1).
ECf  EC i
EC
n A532nm  A600nm
Where, ECi is initial EC reading; ECf is the nal EC reading and n is a
number of samples evaluated.
2.3.4. Determination of soluble sugars
To extract soluble sugars, freeze-dried mesocarp powder
(0.10 g) was added to 10 mL of 80% (v/v) ethanol and homogenised
for 1 min. The mixture was then placed for 60 min in a water bath at
80  C. Subsequently, the mixture was stored in a refrigerator at 4  C
overnight to facilitate the release of soluble sugars. The mixture
was then centrifuged at 4  C for 15 min at 12000g. The supernatant
was ltered through glass wool and the ltrate taken for drying
under vacuum in a GenVac1 concentrator (SP Scientic, Genevac
LTD., Suffolk, UK). Dehydrated samples were re-constituted using
2 mL ultra-pure water, and ltered through a 0.45 mm nylon
syringe lter into an HPLC vial. Sugars were analyzed using an
isocratic HPLC system equipped with a refractive index detector,
according to Liu, Robinson, Madore, Witney, and Arpaia (1999).
Sample extracts were injected into a Rezex RCM monosaccharide
Ca+ (8%) column of 7.8 mm diameter  300 mm (Phenomenex,
Torrance, CA, USA) with a Carbo-Ca2+ guard column of 3 mm  4
mm  (Phenomenex). The column temperature was kept at 80  C
using a thermoregulated column compartment. The mobile phase
was ultra-pure water at a ow rate of 0.6 mL/min. The
concentration and the presence of individual sugars (sucrose,
glucose, fructose, mannoheptulose, and perseitol) were deter-
mined by matching peak areas of samples with the peak areas and
concentration of the standard curves (0.051.25 mg/L; R2 = 0.99).
2.3.5. Extraction proteins and determination of polyphenol oxidase
activity
Proteins were extracted according to Kanellis and Kalaitzis
(1992), with slight modications. Freeze-dried, milled mesocarp
tissue (1.0 g DM) was extracted in 5 mL of 50 mM TrisHCl buffer
(pH 7.4) containing 0.2 M NaCl, 20 mM MgSO4, 1 mM Ethyl-
enediaminetetraacetic acid (EDTA), 5 mM mercaptoethanol,
0.5 mM phenylmethane sulfonyl uoride (PMSF), 10 mM leupeptin,
and 10% (v/v) glycerol. The samples were then homogenised using
the ultrasonic cell disrupter to extract free and membrane-bound
proteins. Subsequently, the mixture was allowed to stand on ice for
15 min and centrifuged at 20,000  g for 20 min. The supernatant
was used for enzyme assays after being ltered through
Miracloth1.
The enzymatic activity of polyphenol oxidase (PPO; EC 1.14.18.1)
activity was determined using an assay previously described by
Van Lelyveld, Gerrish, and Dixon (1984) with minor adjustments. A
sample of 100 mL of extracted protein was added to a mixture of
1.45 mL of 20 mM 4-methyl-catechol and 1.45 mL of 10 mM (pH
5.0) acetate buffer. Total activity of PPO was read in a
spectrophotometer at 420 nm and expressed as U/kg of the plant
tissue.
2.3.6. Determination of lipid peroxidation
Lipid peroxidation was determined using the quantity of
reaction mixture of malondialdehyde (MDA) with thiobarbituric
acid (TBA) and forms a TBAMDA complex as the end product
(Chong, Abdullah, Fadzillah, Lai, & Lajis, 2005). Briey, a 1 mL
extract of crude protein was added onto a test tube containing 1 mL
of 20% (w/v) TCA, 0.01% (w/v) BHT and 0.65% (w/v) TBA. The
mixture was vigorously agitated, incubated for 30 min at 95  C,
allowed to cool down in ice and centrifuged for 10 min at 3000g.
The absorbance of each sample was read at 532 and 600 nm with
an ultra-violet to the visible spectrophotometer. Lipid peroxidation
was determined as total MDA equivalent (nmol g1), calculated
using Eq. (2) described by Heath and Packer (1968).
1  
TotalMDA x103 2
155
Where A532nm and A600nm were an absorbance at 532 and 600 nm.
2.4. Scanning electron microscopic (SEM) fruit membrane image
analysis
Samples were sputter-coated with gold using an EIKO IB3 gold
coated and viewed using the SEM (Zeiss EVO LS15) according to
Talbot and White (2013) with slight modication. Briey, Avocado
mesocarp tissues were prepared for SEM image analysis. Tissues
were dissected and xated in 3% glutaraldehyde and immediately
washed with cacodylate buffer. Then after the tissue was
dehydrated using a different concentration of ethanol for 1 h.
The same tissue was further dried to critical point using freeze
drier. The dried samples were mounted on SEM stubs and coated
with sputter coater and the samples were ready for viewing.
2.5. Statistical analysis
The collected data was subjected to the analysis of variance
(ANOVA) using GenStat statistical software (GenStat1, 17.1 edition,
VSN International, UK) 17.1. Mean separation was performed using
Fischers least signicant difference (LSD) at 5% level of signi-
cance. Standard error values were calculated where a signicant
standard deviation was found at p  0.05 between individual
values.
3. Results and discussion
3.1. Fruit mass loss
Coating treatments had a signicant inuence on mass loss of
avocado fruit during postharvest storage. For both cultivars, higher
fruit mass loss (>5%) was observed in untreated control compared
to coated fruit (p < 0.01) (Fig. 1). Further, and perhaps very
important, was the rate of mass loss over the storage period. The
greatest mass loss was observed during the rst ten days of
postharvest storage. In fact, most moisture loss and thus mass loss
could be expected to occur during the initial cooling of the fruit
(Wills, McGlasson, Graham, & Joyce, 1998; Bower & Magwaza
2004). Fruit moisture loss was consistently declining for all
treatments during cold storage as well as during ripening stage at
ambient conditions.
The most efcient mass loss control was in the fruit coated with
a combination of moringa and 0.5 and 1% CMC. Untreated control
fruit continued losing water into the cold room atmosphere. This is
 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048
Control
Moringa 2%
6 M+CN 0.5%
Fuerte M+CN 1%
M+CMC 0.5%
M+CMC 1%
4 M+CMC 0.5%
2
Fruit mass loss (%)
0 2
6
Hass
4 0
2 Hass
0
0 7 14 21 28
Storage time (Days)
Fig. 1. Effect of edible coatings in avocado fruit mass loss during postharvest storage
time. M is 2% moringa leaf extract, CN is chitosan, and CMC is carbohymethyl
cellulose. Vertical bars represent standard error (SE) of the mean value (n = 3). The
SE refers to the standard deviation of each measured values from the mean value.
due to the cold room atmosphere continually being dried as it
passes through the cooling coil, and the resultant deciency of
water being replaced down the concentration gradient from the
fruit to the atmosphere (Bower & Magwaza, 2004). Observed
moisture loss was mostly due to moisture loss by transpiration,
which accounts for about 90% of total mass loss in fruit (Magwaza
et al., 2013a). This is in accordance with the literature stating that
moisture loss from the fruit results from vapour pressure decit
between the less saturated atmosphere and the fruit, which is close
to saturation with water (Cutting & Wolstenholme, 1992; Magwaza
et al., 2013a).
The differences between control and coating treatments
recorded in the current study could be due to the nature of edible
coating properties, which is hydrophilic, and it displays less
resistance to water loss than the lipophilic coatings (Kester &
Fennema, 1986). This observation has been anticipated in avocado
fruit tissues which accumulate high oil content that has a lipophilic
functional property which exhibits resistance to water loss
(Kruger, 2013). The results observed in the current study are also
in agreement with Maftoonazad and Ramaswamy (2005) who
showed that methyl cellulose retards moisture loss in avocado
fruit. Higher postharvest mass loss observed in the untreated
control fruit could also be explained by the removal and re-
organisation of natural wax on the fruit surface which is brushed
down and reorganised by standard commercial brushing and
washing (Magwaza et al., 2013b).
3.2. Mesocarp electrical conductivity
Both cultivars had low EC during the rst 14 d of cold storage
(Fig. 2). The fruit membrane only started to release more
electrolytes after 14 d and increased EC exponentially during
the shelf-life storage at ambient condition. These observations
suggest that regardless of the variety and coating treatments, EC of
avocado fruit increases with postharvest storage time. The coating
treatments had a statistically signicant effect on mesocarp EC and
this was consistent over storage time. Fruit coated with CMC and
43
Control
Moringa 2%
M+CN 0.5%
M+CN 1%
4
Fuerte M+CMC 1%
EC*100 (S/m)
1
3
2
1
0
0 7 14 21 28
Storage time (Days)
Fig. 2. Effect of postharvest edible coatings in avocado mesocarp electrolyte
leakage during storage time. M is 2% moringa leaf extract, CN is chitosan, and CMC is
carbohymethyl cellulose. Vertical bars represent standard error of the mean value
(n = 3). The SE refers to the standard deviation of each measured values from the
mean value.
chitosan containing moringa leaf extract had respective EC of
(170.0
3.0 m/m) and (210.0
5.0 m/m) which were signicantly
lower than that of the control fruit (292.0
5.0 m/m). The two
levels of CMC (0.5% and 1%) containing moringa leaf extract and the
2% moringa with emulsier retained fruit rmness longer than
other coating treatments. The observed retention of rmness may
suggest that these coating treatments resulted in better membrane
integrity which improved fruit quality and shelf-life. Similar
results were reported by Azad (2006) fruit treated any coating
material retains rmness and releases less EC. The observed
disrupted ion balance and electrolyte leakage is hypothesized to
have resulted from ultra-structural changes in the membranes
(Lyons, 1973). In addition, the relative EC of the mesocarp tissue in
the current study was gradually increasing during postharvest
storage, suggesting a continuous and a gradual loss of cell
membrane integrity.
3.3. Fruit respiration
Both Hass and Fuerte cultivars displayed a similar pattern of
fruit respiration (Fig. 3). The fruit were respiring at a slower rate for
the rst 14 d during cold storage and started to increase at a faster
rate towards the end of the cold storage. In the current study, a
gradual increase in the rate of respiration was observed during
postharvest storage, indicating that the rate of avocado fruit
respiration increases as the fruit ripens or senesces. For instance, in
Hass, coating treatments had a strong effect on fruit respiration
with untreated fruit displaying the fastest rate (290.0
62.0 mg/
kg.h) compared to fruit treated with different coatings. Coating
Hass avocado fruit with moringa 2% and the combination of CMC
and moringa signicantly reduced fruit respiration rate to
44 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048
Control
Moringa 2%
400 M+CN 0.5%
Fuerte M+CN 1%
M+CMC 0.5%
300
M+CMC 1%
200
CO production (mg/kg/h)
100
0 0
400
Hass
300
2
200
100
0 0
28
0 7 14 21
Storage time (Days)
Fig. 3. Effect of edible coatings on avocado fruit CO2 production during postharvest
storage time. M is 2% moringa leaf extract, CN is chitosan, and CMC is
carbohymethyl cellulose. Vertical bars represent standard error of the mean value
(n = 3). The SE refers to the standard deviation of each measured values from the
mean value.
(112.0
40.8 mg/kg.h) and (122.0
20.3 mg/kg.h), respectively. At
the last sampling date a similar trend was observed in Fuerte,
where these two treatments had the lowest respiration rate. There
were differences in fruit oxidation during the storage period. In
avocado, typical a climacteric fruit, the increase in respiration rate,
which is triggered by ethylene accumulation, is accompanied by a
complex of biochemical changes resulting in fruit softening. The
results observed herein are similar to those reported in other
studies on avocado fruit, where respiration have been shown to
increase with time and the rate reduced by coating treatments
(Jeong, Huber, & Sargent, 2002; Jeong, Huber, & Sargent, 2003).
3.4. Fruit rmness
In both Hass and Fuerte avocado cultivars, fruit rmness
decreased signicantly (p < 0.05) with storage period and these
trends were similar for both treated and control fruit, although the
rate of decrease was not the same (Fig. 4). In Hass, the highest
value of rmness was maintained by the moringa 2%
(55.0
4.25 N) and the combination of CMC 1% and moringa
(60.0
3.75 N). Upon removal from cold storage, the untreated
fruit had the lowest values of rmness (15.0
3.0 N) (Fig. 4). The
softness of fruit is typical of avocado fruit resulting from the
weakening of the cell wall structure, loss of membrane integrity,
hydrolysis of cellulose and hemicellulose as well as depolymerisa-
tion of pectin and starch (Seymour, Taylor, & Tucker, 1993). The
hydrolysis and depolymerisation of these components are
catalysed by the action of several enzymes, including polygalac-
turonase, pectin methylesterase, pectate lyase, Rhamnogalactur-
onase and b-galactosidase (Yaman & Bayoindirli, 2002; Payasi,
Mishra, Chaves, & Singh, 2009). Structural changes in the integrity
of cell wall are likely to result from the activities of cellulase (Pesis,
Fuchs, & Zauberman, 1978; Tucker & Laties, 1984) which decreases
Control
Moringa 2%
160 Fuerte
M+CN 0.5%
140
M+CN 1%
120
M+CMC 0.5%
100
M+CMC 1%
80
60
40
20
Firmness (N)
160 Hass
140
120
100
80
60
40
20
0
14 21 28
7
Storage time (Days)
Fig. 4. Effect of postharvest edible coatings in avocado fruit softness during storage
time. M is 2% moringa leaf extract, CN is chitosan, and CMC is carbohymethyl
cellulose. Vertical bars represent standard error of the mean value (n = 3). The SE
refers to the standard deviation of each measured values from the mean value.
cellular cohesion by cell disarrangement and degradation of pectin
(Awad & Young, 1979). The gaseous composition of lower O2 and
higher CO2 percentage in coated fruit is likely to limit the activities
of oxidising enzymes allowing the retention of rmness during
postharvest storage (Salunkhe, Boun, & Reddy, 1991). Our ndings
are therefore in agreement with this argument and that of Jeong,
Huber, and Sargent (2000) who stated that rmness of untreated
avocado fruit decreased faster whereas fruit treated with both 1-
MCP and wax retained rmness for a longer period. Loss of
rmness during marketing negatively affect fruit quality, hence the
ability of these coating treatments to reduce the rate of softening is
advantageous in extending shelf-life and reducing postharvest
losses. The coating effect on fruit rmness is not unique to avocado
but have been previously reported in other fruit, where fruit coated
with chitosan had reduced activities of the macerating enzymes
such as polygalacturonase, pectate lyase, and cellulase, thus,
retaining rmness of tomatoes (Reddy, Angers, Castaigne, & Arul,
2000) and peaches (Atkinson et al., 2012).
3.5. Polyphenol oxidase activity
PPO activity of mesocarp tissue of both fruit cultivars
signicantly increased (p < 0.05) with storage period for all
treatments. Starting from day seven till the end of the storage
period, PPO activity for the control fruit increased gradually
dominating over the treated fruit (Fig. 5). In Hass avocado fruit,
the moringa 2% and the combination of CMC 1% and moringa had
the least PPO activity of (0.73
0.07*1000 U/kg) and
(0.68
0.06*1000 U/kg) than the other treatments. The effect of
these treatments was similar in Fuerte avocado fruit. Browning as
a symptom of chilling injury in persimmon fruit involves oxidation
of phenolic substrates mediated by PPO (Toms-Barbern & Espn,
2001). We speculate that the maintenance of membrane integrity
in treated fruit contributed to the reduction in skin and esh
 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048 45

Control
Moringa 2%
Control M+CN 0.5%
4
Moringa 2% M+CN 1%
1.2
Fuerte M+CN 0.5%
Fuerte M+CMC 0.5%
M+CN 1% 3 M+CMC 1%
0.8 M+CMC 0.5%
M+CMC 1% 2
PPO activity *1000(U/kg)

0.4
1

Lipid peroxidation (nmol/g)

0.0
0

Hass 2.4
Hass
0.8
2.0

1.6

0.4 1.2

0.8

0.4
0.0
0 7 14 21 28 0.0
0 7 14 21 28
Storage time (Days)
Storage time (Days)
Fig. 5. Effect of postharvest edible coatings in avocado mesocarp polyphenol
oxidase (PPO) activity during storage time. M is 2% moringa leaf extract, CN is Fig. 6. Effect of postharvest edible coatings in avocado fruit mesocarp lipid
chitosan, and CMC is carbohymethyl cellulose. Vertical bars represent standard peroxidation during storage time. M is 2% moringa leaf extract, CN is chitosan, and
error of the mean value (n = 3). The SE refers to the standard deviation of each CMC is carbohymethyl cellulose. Vertical bars represent standard error of the mean
measured values from the mean value. 1 Unit = Change of absorbance/min at value (n = 3). The SE refers to the standard deviation of each measured values from
420 nm. the mean value.

Hass
20 Mannoheptulose Perseitol
16

12

4
Fructose Control
Soluble sugars (g/kg)

3 Moringa
M + CN 0.5%
2 M + CN 1%
1
M + CMC 0.5%
M + CMC 1%
0

4 Sucrose Glucose
3

0
7 14 21 28 35 7 14 21 28 35

Storage time (Days) Storage time (Days)

Fig. 7. Effect of postharvest edible coatings in avocado fruit carbohydrate accumulation during storage time. M is 2% moringa leaf extract, CN is chitosan, and CMC is
carbohymethyl cellulose. Vertical bars represent standard error of the mean value (n = 3). The SE refers to the standard deviation of each measured values from the mean value.
46 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048

browning by preserving cell membrane structure and membrane
integrity.
3.6. Lipid peroxidation
In both Fuerte and Hass avocado cultivars, lipid peroxidation
of fruit signicantly increased with storage period. There were
signicant (p < 0.05) differences among treatments. The control
fruit scored the highest lipid peroxidation than the treated fruit
(Fig. 6). The moringa 2% and CMC 1% with moringa leaf extracts had
the least fruit lipid peroxidation. The oxidation of polyunsaturated
fatty acids results in oxidants such as peroxide ions and
malondialdehyde (MDA). The lipid peroxidation increases with
the level of cold stress and fruit prone to chilling injury are
characterised by the high lipid peroxidation. High fruit lipid
peroxidation, accumulation of MDA is often taken as an indicator of
chilling injury (Wongsheree, Ketsa, & Doorn, 2009), further, affects
the fruit quality. The pattern of MDA content of avocado fruit
during cold postharvest storage followed by ripening at ambient
condition tended to increase in the control fruit than the treated
ones.
3.7. Soluble sugars
For both cultivars, signicant (p < 0.05) differences were
observed in concentrations of soluble sugars among the coating
treatments (Figs. 7 and 8). For instance, mannoheptulose which is a
dominant soluble sugar in avocado fruit signicantly decreased
with storage period from average of 11.8 g/kg (for Hass) and 17.6 g/
kg (for Fuerte) at the beginning of the experiment to less than 6 g/
kg (for both cultivars) upon ripening. At the end of the storage
period, control fruit, from both cultivars, had the lowest
concentration of mannoheptulose. The treatments had an effect
on the dominantly produced C7 sugars than the hexoses (Figs. 7
and 8). Avocado fruit is known to produce the two dominant C7
sugars, D-mannoheptulose and perseitol and they also produce
hexoses as the relatively smaller amount. It is also reported that D-
mannoheptulose is known to delay the onset of ripening or else
improves fruit shelf-life. The fruit treated with CMC as well as
chitosan containing moringa extracts retained these C7 sugars as
compared to control. A similar trend was reported by Bertling and
Bower (2005) who observed a decrease in the content of C7 sugar
in avocado fruit during ripening.
3.8. Time to ripen
Fruits were also evaluated for rate of fruit ripening. For Hass
avocado, the untreated control fruit had the shortest shelf-life,
ripening in 6 d compared other coating treatments which ripened
after 8 d (Fig. 9a). A similar trend was also observed with Fuerte
avocado fruit (Fig. 9b). In both cultivars, fruit treated with edible
coats containing moringa leaf extract improved the fruit quality
and resulted in extended shelf-life for more than 9 d during
postharvest storage. This could be attributed to treatments effect

Fuerte
20 Mannoheptulose
Perseitol
16

12

0
Fructose
4
Control
Soluble sugars (g/kg)

3 Moringa
M +CN 0.5%
2 M + CN 1%
1
M + CMC 0.5%
M + CMC 1%
0
Glucose
4 Sucrose
3

0
7 14 21 28 35 7 14 21 28 35

Storage time (Days) Storage time (Days)

Fig. 8. Effect of postharvest edible coatings in avocado fruit carbohydrate accumulation during storage time. M is 2% moringa leaf extract, CN is chitosan, and CMC is
carbohymethyl cellulose. Vertical bars represent standard error of the mean value (n = 3). The SE refers to the standard deviation of each measured values from the mean value.
 S.Z. Tesfay, L.S. Magwaza / Food Packaging and Shelf Life 11 (2017) 4048
10 Fuerte
8 This research was supported by the South African Avocado
6 Growers Association (SAAGA) and the National Research Founda-
4 tions competitive support for unrated researchers grants (CSUR:
Time to ripen (Days)
2
0 Africa for providing fruit for entire research.
Hass
12
10
8 the online version, at http://dx.doi.org/10.1016/j.fpsl.2016.12.001.
6
4
2
0 Majolagbe, O. N., Ogundele, B. A., Aina, J. A., & Adetunji, J. B. ([95_TD$DIFF]2012).
Control Moringa 2% M+CN 0.5% M+CN 1% M+CMC 0.5% M+CMC 1%
Coating treatments
Fig. 9. The effect of edible coating treatments on the time to reach ripening at room
temperature. M is 2% moringa leaf extract, CN is chitosan, and CMC is
carbohymethyl cellulose. There were 20 avocados at the start of the experiment
under ambient condition. The SE refers to the standard deviation of each measured
values from the mean value.
on fruit quality attributes by reducing fruit moisture loss,
respiration rate, membrane electrolyte leakage as well as by
retaining energy reserve, mainly mannoheptulose than untreated
fruit (Blakey, Bower, & Bertling, 2009; Tesfay, Bower, & Bertling,
2010).
3.9. Peel microstructural analysis
The fruit membrane image of control and fruit treated with
CMC 1% with moringa extract were also viewed under SEM over
the shelf-life period. The result showed that the treated fruit
retained membrane integrity than untreated fruit (Supplementary
data). The onset of ripening was delayed, improves the fruit shelf-
life beyond 6 d, while the untreated fruit reached the ready-to-eat
softness in 6 d. This SEM result further supported our ndings that
fruit treated with the coating material and moringa leaf extracts
had less respiration, moisture loss and high rmness.
4. Conclusions
Overall, the study demonstrated that Fuerte and Hass
avocado fruit coated with moringa and a combination of moringa
+1% CMC had lower rates of respiration, higher values of rmness
compared with the uncoated control and this was observed
consistently during the entire storage period. The results also
showed that reduced rate of respiration, moisture and rmness
loss resulted in improved fruit quality and shelf-life. Furthermore,
fruit treated with moringa and a combination of moringa +1% CMC
had the highest concentration of C7 sugars, mannoheptulose, and
perseitol, conrming their vital role in delaying fruit ripening
process. In conclusion, the study reported the potential of this
novel edible coating containing moringa leaf extracts to improve
avocado fruit quality during the storage period. Although moringa
and a combination of moringa and CMC preserved fruit quality and
extended shelf life of Hass and Fuerte avocado fruit, further
research on the effect of these treatments on sensory evaluation
still need to be conducted before these treatments are recom-
mended for commercial application.
47
Acknowledgments
105978). The authors are grateful to Mr Lynton Freese and Mr Sipho
Sekhune, Westfalia packhouse in Howick, KwaZulu-Natal, South
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
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