Effect of hypobaric storage on quality, antioxidant enzyme and antioxidant capability of the Chinese bayberry fruits
South China Botanical Garden, Chinese Academy of Sciences
Food Science Institute, Zhejiang Academy of Agricultural Sciences
School of Life & Environmental Sciences, Wenzhou University
Background: The Chinese bayberry (Myrica rubra Sieb. and Zucc.) is a subtropical fruit native to China, with unique flavor, sweet and sour taste, and high nutrition and health values. The fruit is highly perishable and susceptible to mechanical injury, physiological deterioration and fungal decay once harvested. This study was to investigate the effect of hypobaric storage on the quality of Chinese bayberry fruit and then develop storage technology to prolong the supply of the fruit. Results: The fruit stored under hypobaric conditions exhibited lower decay, higher titratable acidity and total phenolics compared with those stored under normal atmospheric conditions. Hypobaric storage significantly reduced malonaldehyde accumulation, respiratory rate and maintained high catalase and peroxidase activities of Chinese bayberry fruit. Ferric reducing antioxidant power was also higher in the fruit stored under hypobaric condition than those under normal atmospheric conditions. Conclusion: Hypobaric storage improved the metabolism, antioxidant system and postharvest quality of Chinese bayberry fruit and provided an effective alternative method to prolong the storage life of this fruit.
The Chinese bayberries (Myrica rubra Sieb. and Zucc.) are
a subtropical fruit native to China. In terms of the unique
flavor, sweet and sour taste, attractive red color, and high
nutrition and health values; Chinese bayberries have been
cultivated in eastern and southern China for more than
2000 years and are being introduced to other countries.
The fruit mature in early summer season and are praised
as the precious southern Yangtze fruit of early summer
[1-3]. The Chinese bayberries contain abundant
anthocyanins, flavonoids and other phenolic compounds, with high
antioxidant capacity [3-5]. Unfortunately, the fruit are
highly perishable and susceptible to mechanical injury,
physiological deterioration and fungal decay, resulting in a
short postharvest life of 12 days at ambient temperature
. Some methods, including low temperature storage
[7,8], high oxygen atmosphere treatment [9,10], hot air
treatment [6,11], combined treatment of ethanol vapor
with hot air , have been used to investigate postharvest
physicochemical and physiological attributes and storage
life extension of the Chinese bayberry fruit. However,
due to the delicate nature of the fruit, poor handling
practices and inadequate storage facilities, the shelf life of the
Chinese bayberry is still short, which markedly limits its
market. As this fruit is further commercially developed, it
is important to develop effective storage methods to
prolong the shelf life.
Among these techniques for controlling postharvest
decay of fruit and vegetables, the use of sub-atmospheric
pressure exhibits a potential to store fresh Chinese
bayberries. Hypobaric storage can quickly remove heat and
reduce oxygen level . During storage, water spray
could be used to solve the problem of insufficient
environmental humidity . It has been reported that
hypobaric treatment delayed ripening of some climatic fruits
such as apples, avocados, bananas, mangoes, tomatoes,
apple, sweet cherry, asparagus, and peach [14-17]. With
the development of storage technology, different models
of hypobaric storage machine have been developed and
tested for the storage of fruit and vegetables. However,
little information is available in the literature about this
storage technology for Chinese bayberry fruit.The
objective of this present study was to investigate the effects
of different hypobaric storage treatments on postharvest
life and quality of the Chinese bayberry fruit. The
antioxidant enzyme activities and antioxidant capacity were
also evaluated. Finally, the optimal condition of
hypobaric storage to extend the shelf life of the Chinese
bayberry fruit was determined.
Results and discussion
Effect of hypobaric storage on fruit decay of Chinese
Chinese bayberries are highly perishable and susceptible to
mechanical injury, physiological deterioration and fungal
decay . The fruit stored under normal atmospheric
pressure (control) showed 37.5% decay after 6 days of storage,
but the fruit stored under 855, 555 and 155 kPa
exhibited 7.25, 5.0 and 6.25% decay, respectively. As shown in
Figure 1, the decay severity increased gradually with
increasing storage. After 15 days of storage, the decay
percentages of the Chinese bayberries stored under 101.3, 855,
555 and 155 kPa were 81.2, 31.25, 18.75 and 25%,
respectively. It was reported that low pressure treatment
discouraged commodity deterioration caused by bacteria and
fungi and was capable of killing many insects infesting
agricultural commodities . Romanazzi, et al.  reported
hypobaric treatment was effective in reducing decay of
sweet cherries, strawberries and table grapes. The present
study showed that hypobaric storage was an effective
method to reduce decay of the Chinese bayberry. Among
these four treatments, it was found that application of 555
kPa was the optimal to reduce the fruit decay.
Figure 1 Effects of hypobaric storage on fruit decay of the
Chinese bayberry. Fruits were stored at 10.5C and 8590%
relative humidity under different atmosphere pressures.
Effects of hypobaric storage on skin colour and pulp TSS
The Chinese bayberry fruit of most cultivars are red or
dark-red colour due to the presence of anthocyanins [3-5].
The major anthocyanin in the Chinese bayberry fruit was
identified to be cyanidin-3-glucoside which represented
more than 95% of the total anthocyanins . Skin colour is
an important index to evaluate the quality of the Chinese
bayberry fruit. As shown in Figure 2A and 2B, L* value
increased while a* value of the fruit decreased gradually
during storage. However, no significant (P<0.05)
differences in the skin colour were observed among these
different pressure treatments after 15 days of storage.
TSS content of the Chinese bayberry fruit decreased
during storage (Figure 2C) but no significant (P<0.05)
differences existed among the different pressure treatments.
Figure 2D presented TA content of the Chinese
bayberries. The TA content decreased gradually during
storage. By the end of storage, TA content decreased to 11.6,
5.0, 8.7 and 6.9% under 101.3, 855, 555 and 155 kPa
conditions, respectively. The changes in TA and TSS
contents could be associated with the metabolic activity and
respiratory rates of the fruits. Corey, et al.  reported
that respiration rate of lettuce decreased by 40% when
stored in a chamber at a pressure of 51 kPa. The results in
this study could be explained by a lower respiratory rate
(Figure 3A), which caused less depletion of sugars and
acids when fruit was stored at lower pressures.
Effects of hypobaric storage on respiration and ethylene
The respiratory rate of the Chinese bayberry fruit was
around 9 mg CO2 kg-1 h-1 before storage and decreased
gradually with increasing storage (Figure 3A). By the end
of storage, respiratory rates of the Chinese bayberry stored
under 101.3, 855, and 555 kPa conditions were 4.75,
3.72, and 3.16 mg CO2 kg-1 h-1, respectively. The
respiratory intensity of the fruit under hypobaric storage was
significantly (P<0.05) inhibited, as compared with under
normal atmospheric pressure. Ethylene production rates
of the Chinese bayberries decreased gradually, from 0.52
before storage to 0.042 L kg-1 h-1 by the end of storage
(Figure 3B). However, no significant differences in
ethylene production rates were observed between the
hypobaric storage and normal storage.
It was reported that hypobaric packaging reduced the
respiration rates of strawberry and curled lettuce . He
et al.  also reported that hypobaric storage conditions
could reduce greatly the ethylene production rate in both
lettuce and wheat. The removal of ethylene production
could delay senescence of fruits and vegetables and,
indirectly, reduce their susceptibility to pathogens . The
inhibition of the respiratory rate in the Chinese bayberry fruit
by hypobaric treatment can help to extend the shelf life.
Effect of hypobaric storage on malondialdehyde (MDA)
MDA is considered to be an indicator of membrane lipid
peroxidation caused by oxidative stress. As shown in
Figure 4, MDA content of the Chinese bayberry fruit
under normal pressure condition rose gradually during
storage. MDA contents of fruit stored under 101.3, 855,
555 and 155 kPa conditions after 15 days of storage
were 5.0810-3, 4.5110-3, 4.2110-3 and 4.5310-3 mol
g-1 on fresh weight (FW) basis, respectively, which
exhibited that hypobaric storage inhibited the accumulation of
MDA. Similar results were obtained by Li et al.  who
reported that hypobaric storage could reduce MDA
accumulation and retard senescence in asparagus.
Effect of hypobaric storage on antioxidant enzyme
The accumulation of reactive oxygen species, such as
superoxide, hydrogen peroxide, and the hydroxyl radical,
causes plant tissue damage and reduces the storage quality
and marketability of fruits and vegetables .
Antioxidative enzymes such as catalase (CAT) and peroxidase
(POD) play an important role to scavenge reactive oxygen.
As shown in Figure 5A and 5B, CAT activities of the Chinese
bayberry fruit tended to change differently at various
pressure conditions, but the fruit stored under the
hypobaric condition exhibited a higher activity than those stored
under the normal pressure condition. For POD activity,
hypobaric storage condition maintained a significantly
at 101.3 kPa, which indicated that hypobaric storage
was more effective to maintain phenolic content of
the fruit. In similarity with the change in total phenolic
content, reducing power of fruit rose gradually during
storage (Figure 6B). In this study, the fruit stored under
555 kPa showed the strongest ferric reducing
antioxidant power. However, ferric reducing antioxidant power
(FRAP) was not correlated well with the total phenolic
contents. It has been reported that different antioxidant
activity could be due to the difference in phenolic
Materials and methods
The Chinese bayberries (Myrica rubra Sieb. & Zucc., cv.
Dongkui) fruit were harvested manually from a
commercial orchard in Xianju county of Zhejiang Province,
China on June 28, 2010, and then transported to the
laboratory by a refrigerated car within 3 hours. Fruit were
selected for uniformity of shape and color and the
blemished and diseased fruit were discarded.
Fruit treatments were performed in a hypobaric storage
system with storage chambers whose pressure could be
set independently (Model XL-5, Xianlv Low-pressure
Fresh Keeping Equipment Co. Ltd., Shanghai, China).
Each replicate containing 2 kg fruits was put in a plastic
basket and placed into the hypobaric chamber. The
applied pressures were set to be 855, 555 and 155 kPa,
respectively. The normal atmospheric pressure (101.3
kPa) was used as control. These fruit were stored at
10.5C and 8590% relative humidity (RH). Fruit
samples were taken for analysis every 3 days in the storage
period of 15 days.
Evaluations of fruit decay and skin colour
Fruit decay was visually evaluated. Fruit with visible
mold growth with about 2% of the surface affected was
Figure 4 Effects of hypobaric storage on MDA content of the
Chinese bayberry fruit. Fruit were stored at 10.5C and 8590%
relative humidity under different atmosphere pressures.
(P<0.05) higher activity compared with the normal
atmospheric condition. Chen et al.  reported that hypobaric
storage maintained high CAT activity in peach. In this study,
application of hypobaric storage enhanced CAT and POD
activities and, thus, reduced membrane lipid peroxidation of
the Chinese bayberry (Figure 4).
Effects of hypobaric storage on total phenolic content
and total antioxidant capacity
Phenolic compounds including flavonoids and phenolic
acids are known to be responsible for antioxidant
capacity in fruits. The Chinese bayberries are rich in
phenolic compounds and exhibit high antioxidant activity
[3,4,24]. As shown in Figure 6A, total phenolic content
of the fruit increased during storage. However, no
significant differences were observed among these different
pressure treatments within the first 3 days of storage.
The total phenolic content of the fruit under the
hypobaric conditions after 3 days of storage rose rapidly, and
then reached 1.083, 0.999, and 1.134 mg g-1 FW when
fruit were stored at 855, 555 and 155 kPa after 15
days of storage, respectively, but only 0.892 mg g-1 FW
Figure 6 Effects of hypobaric storage on total phenolic content (A) and total antioxidant capacity (B) of the Chinese bayberry fruit.
Fruit were stored at 10.5C and 8590% relative humidity under different atmosphere pressures.
considered rotten. The severity of fruit decay was
expressed as percentage of fruit showing decay symptoms.
Skin colour of 20 fruit from each replicate was measured
using a colorimeter (Konica Minolta, CR-400, Japan) with
a 6-mm aperture size, which provided L* and a* values
according to the system established by the Commission
Internationale de LEclairage (CIE, International
Commission on Illumination). A reference white tile was used for
Measurements of respiration and ethylene production
Ten fruit were enclosed in 250 mL glass jars at 5C for 2 h
and then 2 mL of headspace gas were taken from each
jar. CO2 amount was measured by gas chromatography
(Rainbow, SP-9890, China) equipped with flame ionization
detector and a packed column (GDX-502, Zhonghuida
Inc., China). Ethylene concentration was analysed by gas
chromatography using a flame ionization detector.
Respiration and ethylene production rates were expressed as mg
CO2 and g per hour on fresh weight basis, respectively.
Measurements of total soluble solids and titratable
Fifty fruit from each treatment were taken. Juice was
obtained by a juicer (HR1861, Philips Co. Beijing, China),
followed by filtration through cheesecloth. The juice was
analyzed for total soluble solids (TSS) and titratable acidity
(TA). TSS concentration was determined by a portable
refractometer (Atago PAL-1, Japan) while TA content was
measured by titrating 20 mL of the juice to pH 8.2 using
0.1 mol L-1 NaOH.
MDA content determination
MDA content was determined according to the method
described by Li et al.  with some modification. Fruit
tissues (1 g) were extracted for 2 h with 5 mL of
trichloroacetic acid (10%). Three milliliters of 0.5%
thiobarbituric acid (TBA) in 10% trichloroacetic acid were
added to 1 mL of the extract. The solution was heated
in a boiling water bath for 20 min, then immediately
cooled, and finally centrifuged at 6000 g for 10 min to
clarify the solution. Absorbance was measured at 532
and 600 nm. MDA content was expressed as mol/g FW
by the method of Li et al. .
Enzymatic activity assay
Five grams of fruit tissues were homogenized in 25 mL of
100 mmol L-1 Tris-HCl buffer (pH 7.8) containing 2
mmol L-1 EDTA and 2 mmol L-1 1,4-dithiothreitol at 4C.
The homogenate was centrifuged at 15,000 g for 15 min
at 4C, and then the supernatant was collected for the
enzymatic activity assay. Protein was measured according to
the method of Bradford , using bovine serum albumin
(BSA) as the standard.
CAT was analyzed according to the method of Beers
& Sizer  with some modifications. The disappearance
of H2O2 was monitored by measuring the decrease in
absorbance at 240 nm of a reaction mixture containing
100 mmol L-1 Tris-HCl buffer (pH 7.8), 25 mmol L-1
H2O2, and 0.2 mL of crude enzyme extract. One unit of
enzymatic activity was defined as 0.01 change of
absorbance at 240 nm per minute. Specific CAT activity was
expressed as units per mg protein. POD activity was
assayed according to the method described by Yang
et al. . The reaction mixture (2 mL) consisted of 50
mmol L-1 sodium phosphate buffer (pH 6.5), 6 mmol L-1
guaiacol and 4.5 mmol L-1 H2O2 prior to the addation of
1 mL of crude enzyme extract. Increase in absorbance at
470 nm at intervals of 30 s was recorded. One unit of
enzymatic activity was defined as the amount of enzyme
that catalyzed the peroxidation of 1 mmol of guaiacol
per minute. Specific POD activity was expressed as units
per mg protein.
Total phenolic content determination
One gram of lyophilized fruit tissues was extracted with
25 mL of ethanol for 3 h. Total phenolic contents were
estimated colourimetrically using the Folin-Ciocalteu
method . The extract was appropriately diluted, and
then 1 mL of the dilution was oxidized with 0.5 mL of
Folin-Ciocalteau reagent. The reaction was neutralized
with 5 ml of 5% Na2CO3. The solution was immediately
diluted to a final volume of 25 mL with distilled water
and then mixed thoroughly. The absorbance was read at
765 nm after 1 hour of incubation in dark at 25C using
a spectrophotometer (Shimadzu UV-2550, Japan). Gallic
acid was used as a standard, and phenolic contents were
expressed as mg gallic acid equivalents (GAE)/g FW.
The ferric reducing ability of the Chinese bayberry was
measured according to the method of Benzie & Strain
. To prepare the FRAP reagent, a mixture of 0.3 mol
L-1 acetate buffer (pH 3.6), 10 mmol L-1
tripyridyltriazine (TPTZ), and 20 mmol L-1 ferric chloride (10:1:1,
v/v/v) was made. One gram of lyophilized fruit tissues
was extracted for 12 hours with 20 mL of ethanol. The
FRAP reagent (3.9 mL) was added to the extract solution
sample (0.1 mL) and then mixed thoroughly. The reaction
was then monitored for 10 min at 37C and the
absorbance was recorded at 593 nm on the Shimadzu UV-2550
spectrophotometer. The ferric reducing ability of the
Chinese bayberry fruit was expressed as mmol FeSO4 per
litre crude extract.
All samples were prepared and analysed in triplicate.
Statistical analysis was done with one-way analysis of
variance using the SAS statistical software package.
Previous studies indicated that hypobaric storage reduced
commodity respiration and prevented wilting and
senescence during storage [17,30]. In the present study,
application of hypobaric pressure to the Chinese bayberries
significantly reduced fruit decay and loss in total acids,
inhibited respiratory rate, decreased MDA accumulation
and maintained total phenolic content, antioxidant
capacity and CAT and POD activities. These data suggested
that hypobaric storage could be an effective technology in
maintaining postharvest quality and prolonging shelf life
of Chinese bayberry fruit.
The authors declare that they have no competing interests.
HC made a significant contribution to acquisition of data, and data analysis.
HY made a substantial contribution to data analysis and manuscript
preparation. HG made a significant contribution to experimental design and
data analysis. JL made a contribution to acquisition of data, and data
analysis. FT and XF participated in some experiments. YJ made a significant
contribution to experimental design, data analysis and manuscript revision.
All authors read and approved the final manuscript.
The work was supported by the National High Technology Research and
Development Program of China (863 Program) (grant No. 2012AA101606),
The International Cooperation Project of China (grant No. 2013DFA31450)
Special Fund for Agro-scientific Research in the Public Interest (grant No.
201303073) and The International Cooperation Project of the Science and
Technology Department of Zhejiang Province, China (grant No. 2011C14003).