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Effect of nano-ZnO-packaging on chilling tolerance and pectin metabolism of peaches during cold storage
Scientia Horticulturae 225 (2017) 128–133
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Effect of nano-ZnO-packaging on chilling tolerance and pectin metabolism of peaches during cold storage
MARK
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Dong Li, Li Li, Zisheng Luo , Hongyan Lu, Yang Yue Zhejiang University, College of Biosystems Engineering and Food Science, Key Laboratory of Agro-Products Postharvest Handing Ministry of Agriculture, Zhejiang Key Laboratory for Agro-Food Processing, Hangzhou, 310058, People’s Republic of China
A R T I C L E I N F O
A B S T R A C T
Keywords: Peach Nano-ZnO Chilling injury Quality Cell wall Pectinase
The effects of nano-ZnO-based low-density polyethylene (NZLDPE)-packaging on chilling tolerance and pectin metabolism in postharvest “Hujingmilu” peaches were investigated. Peaches packed in low-density polyethylene (LDPE) and NZLDPE were stored at 2 °C for 40 d. Compared with the control, both packages alleviated the development of chilling injury, showed higher fruit firmness with lower browning index, electrolyte leakage, relative viscosity, and decay rate. NZLDPE-packaging performed better than LDPE-packaging because of the rapid formation of low O2 and high CO2. Moreover, NZLDPE-packaging inhibited pectin esterase and enhanced polygalacturonase and β-galactosidase, leading to the promotion of alkali soluble-pectin and water-soluble pectin, and the decrease of chelater-soluble pectin. These effects were of great benefit to the maintaining of cell wall structure and the degradation of calcium-pectate gel, which finally alleviated the chilling injury and therefore maintained good quality during chilling stress.
1. Introduction Peach is a popular fruit for consumers because of its juiciness and attractive taste. Peaches ripen and decay quickly at room temperature after harvest, which leads to a loss of sales value. Cold storage is a widely used technology for postharvest fruit and vegetables to delay senescence and extend shelf-life (Wang, 1990). Nevertheless, as a climacteric fruit, peach suffers chilling injury at 2–5 °C. Several technologies, such as treatment with hot air (Yu et al., 2016), glycine betaine (Shan et al., 2016), and oxalic acid (Jin et al., 2014), have been reported to alleviate chilling injury in peaches during cold storage. However, reliable investigations on the practical techniques to inhibit or alleviate chilling injuries are still limited. Chilling injury results in many macroscopic symptoms in peaches, including browning, wooliness, reddening and leatheriness, which are related to cell wall integrity and pectin metabolism (Lurie and Crisosto, 2005). Cao et al. (2009) indicated that chilling injury is partly due to biochemical modifications of cell wall polysaccharides. These processes involve hydrolytic enzymes such as pectin esterase (PE), polygalacturonase (PG) and β-galactosidase (β-Gal) (Fischer et al., 1991). Fruk et al. (2014) reported that the relatively higher PE and lower PG activities observed in peaches were response to chilling stress, which may lead to the formation of high-molecular-weight pectin with a low degree of esterification. This kind of pectin can combine with calcium
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Corresponding author. E-mail address: [email protected] (Z. Luo).
http://dx.doi.org/10.1016/j.scienta.2017.07.003 Received 7 March 2017; Received in revised form 29 June 2017; Accepted 3 July 2017 0304-4238/ © 2017 Published by Elsevier B.V.
to form calcium-pectate gel complexes, which then bind with free water and eventually lead to woolliness, a symptom of chilling injury (Brummell et al., 2004a). In recent decades, nanomaterials used in food packaging have been actively explored (Chaudhry et al., 2008). Nanocomposites enhanced barrier and mechanical properties, and possessed antimicrobial effect along with spore germination inhibition ability in comparison to general polymer (Panea et al., 2014). Various nanocomposites including nano-ZnO-coated polyvinyl chloride (PVC) (Li et al., 2011), nano-Ag2Obased low-density polyethylene (Zhou et al., 2011a), and nano-CaCO3based low-density polyethylene (Luo et al., 2014) applied in the preservation of fruit and vegetables have been reported. During cold storage, polyethylene with nano-Ag, nano-TiO2 and montmorillonite has been proven to be efficient for inhibiting ethylene production, reducing degradation of nutritional components, extending organoleptic characteristics, preventing physiologic changes, and thus delaying the ripening and extending shelf-life of harvested kiwifruit (Hu et al., 2011). Li et al. (2011) stated that nano-ZnO-packaging film preserved fresh-cut ‘Fuji’ apple through reduction of fruit decay rate, depreciation of malondialdehyde (MDA) and ethylene accumulation, maintaining of total soluble solids (TSS) and titratable acid (TA) levels and inhibition of polyphenol oxidase (PPO) and pyrogallol peroxidase (POD) activities. However, to the best of our knowledge, the effect of nano-ZnO-packaging on chilling tolerance of peaches during cold storage has not been
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2. Materials and methods
distilled water in a beaker were stirred slowly for 20 min and the electrolyte conductivity of leach liquor (C1) was measured with a conductivity meter. Then the leach liquor with discs was boiled for 10 min and cooled down to the room temperature. Electrolyte conductivity (C2) was measured. Each treatment was conducted for three biological replicates. The rate of electrolyte leakage was calculated as (C1/C2) × 100%.
2.1. Plant material and treatments
2.6. Measurement of relative viscosity
“Hujingmilu” peaches (Prunus persica L.) with 70–80% maturity (with a firmness of 16.1 ± 0.7 N, total soluble solids content of 11.2 ± 0.2%, and titratable acid content of 0.21 ± 0.02%) were harvested in Yuhang District of Hangzhou City and transferred to the laboratory in Zhejiang University immediately. The fruit were air cooled to 4 °C in 6 h. The first and second groups of peaches were packed into low-density polyethylene (LDPE)-packaging (6 bags) and NZLDPE-packaging (6 bags) with a size of 20 cm × 30 cm respectively, and then sealed and stored at 2 °C for up to 40 days. The third group of peaches were unpackaged and stored under the same condition as control. For each treatment, one bag of peaches was randomly taken every eight days during storage for subsequent analyses (totally 6 bags were used). And another three bags were used to observe the decay rate every eight days during the whole storage. Three biological replications were conducted.
Peach flesh from one bag was mashed and filtered using gauze. After centrifuged at 5000 × g for 15 min, 10 mL of supernatant was collected and the time supernatant (T1) and distilled water (T0) flowed through a capillary viscometer with 0.4 mm inner diameter at 25 °C were measured respectively. Each treatment was conducted for three biological replicates. The relative viscosity was presented as (T1-T0)/T0.
researched. The objective of this study was to investigate the effect of nano-ZnObased low-density polyethylene (NZLDPE)-packaging on chilling tolerance and pectin metabolism in postharvest “Hujingmilu” peaches during cold storage.
2.7. Measurement of fruit firmness A texture analyzer (TA-XT2i, Stable Micro Systems Ltd., Godalmin, UK) was used to determine the peach firmness. Three peaches form one bag were measured on opposite sides of the equator by a 5 mm diameter flat probe. The test speed was 1.5 mm s−1 and the distance was 8 mm. The maximum force (Nm) was recorded as peach firmness and the average values were calculated. Each treatment was conducted for three biological replicates.
2.2. Preparation of NZLDPE materials 2.8. Measurement of decay rate A NZLDPE masterbatch containing 56% (w/w) of LDPE granule (softening point 95 °C, density 920 kg m−3), 30% (w/w) of the commercial nano-ZnO (30 nm) and 14% (w/w) of cross-link reagent ester titanate was blended by a high-speed mixer for 1 h. After cooled down, the mixture was extruded to a NZLDPE masterbatch by a twin-screw extruder with a screw speed of 300 r min−1. Then, 38.5 kg of LDPE granule and 1.5 kg of masterbatch were blended for 0.5 h. The above nanocompounds were made into films of 40 μm thickness by a plastic extruder, and the films were made into NZLDPE bags of 20 cm × 30 cm using a heat sealer after cooling. LDPE bags with the same thickness and size were made under the same condition.
For each treatment, another three bags were used to observe the decay rate every eight days during the whole storage. Any peach with visible mould growth was considered decay. The decay rate (%) was calculated as (the amounts of decayed peaches × 100)/(the total amounts of peaches). Each treatment was conducted for three biological replicates. 2.9. Isolation of cell wall material (CWM) The CWM was prepared using the method described by Chen et al. (2015) with some modifications. One hundred grams frozen flesh pooled from six peaches was homogenized and kept boiling reflux with 500 mL 80% (v/v) ethanol for 0.5 h. After immediate filtration, the residue was collected and washed with 80% (v/v) ethanol for several times until no reducing sugar was detected in the filtrate. Then the residue was washed with acetone and incubated overnight with 90% (v/v) dimethylsulphoxide at 4 °C. The CWM was filtered, washed for several times with distilled water, dried in a vacuum oven and stored in a vacuum desiccator.
2.3. Measurement of in-packaging atmospheric composition Changes in CO2 and O2 concentrations were measured by injecting samples of headspace gases form the packages into a gas chromatograph with a thermal conductivity detector (GC-14A; Shimadzu, Suzhou, China). Each treatment was conducted for three biological replicates. 2.4. Measurement of browning index
2.10. Extraction of CWM constituents For each replicate, eight peaches from one bag were used for evaluation of browning index. Peaches were cut along the fruit suture and the browning scale (B) was divided into four degrees according to the percentage of browning area of flesh; 0: no browning; 1: B ≤ 25%; 2: 25% < B ≤ 50%; 3: B > 50%. The browning index was calculated based on the following formula:
Extraction of CWM constituents was performed according to the methods described by Chen et al. (2015) with some modifications. The water-soluble pectin (WSP), chelater-soluble pectin (CSP) and alkalisoluble pectin (ASP) were extracted consecutively on the CWM. Three hundred milligrams of CWM was added to 100 mL 50 mM sodium acetate buffer (pH 6.5), oscillated and extracted at ambient temperature for 3 h. This process repeated for 3 times and the filtrate contained WSP was collected. Similarly, CSP and ASP was extracted by 50 mM sodium acetate buffer (pH 6.5) containing 50 mM ethylene diamine tetraacetic acid and 100 mM KOH respectively.
Browing index(%) =
∑
(browning scale × the amount of peaches at this scale) total peach number ×3
× 100
2.5. Measurement of electrolyte leakage
2.11. Measurement of CWM constituents
Discs of peach peel from one bag were excised randomly using a 6 mm diameter stainless steel borer. Ten grams of discs with 20 mL
The CWM constituents were measured based on the method described by Bu et al. (2013). WSP, CSP and ASP extracts (5 mL) were 129
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added to 30 mL H2SO4 and heated for 1 h in boiling water. After cooling to room temperature, the solution was mixed with 5 mL of 0.15% (w/v) carbazole reagent and placed for 0.5 h in the dark. The absorbance at 530 nm was measured and galacturonic acid (GalA) was used to prepare a standard curve. The results were expressed as g kg−1 fresh weight. 2.12. Extraction of crude enzyme Enzyme extraction was carried out according to the method described by Meng et al. (2009). Five grams of fruit tissues were homogenized with 20 mL 0.15 M NaCl containing 10 g L−1 polyvinylpolypyrrolidone (PVPP). The homogenate was stirred for 20 min and centrifuged at 15,000 × g for 20 min. The supernatant was collected and used for assays of PE, PG and β-Gal activities. 2.13. The activity of PE The activity of PE was measured according to Pan et al. (2014) with some modifications. Five milliliters of crude extract was added to 20 mL of 1% (w/v) pectin in 100 mM NaCl, titrated with 20 mM NaOH to keep pH 7.4 at 37 °C for 0.5 h. Boiled enzyme extract was used as the control. One unit of PE activity was defined as the amount of PE that released 1 μmol of acid per minute. 2.14. The activity of PG The activity of PG was determined according to the method described by Bu et al. (2013) with a slight modification. Enzyme extract (0.5 mL) was added to 0.5 mL of 0.5% (w/v) polygalacturonic acid in 50 mM sodium acetate buffer (pH 4.4) and incubated at 37 °C for 1 h. Then 1 mL of 3,5-dinitrosalicylic acid (DNS) was added to the mixture and heated for 5 min in boiled water. After cooled down, 25 mL of distilled water was added to the mixture and the absorbance at 540 nm was measured. Boiled enzyme extract was used as the control. One unit of PG activity was defined as production of 1 μmol GalA per minute.
Fig. 1. The O2 (A) and CO2 (B) concentrations of ‘Hujingmilu’ peach in NZLDPE-packaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
2013). Meanwhile, previous study indicated that adding nano-ZnO to poly(butylene adipate-co-terephthalate) decreased the O2 transmission rate of films and thus improved their barrier (Venkatesan and Rajeswari, 2017). As showed in Fig. 1, the O2 concentrations decreased while CO2 concentration increased within both LDPE-packaging and NZLDPE-packaging during the cold storage. NZLDPE-packaging could reach the gas environment with lower O2 and higher CO2 more rapidly compared with LDPE-packaging, which is benefit on fruit cold storage (de Azeredo, 2009). The result was in accordance with the research reported by Luo et al. (2014).
2.15. The activity of β-Gal The activity of β-Gal was measured as described by Minas et al. (2014). Two hundred microliters of enzyme extract was added to 2 mL of 3 mM p-nitrophenylgalactopyranoside in 50 mM HAc-NaAc buffer (pH 5.0) and incubated at 37 °C for 1 h. After incubated, 2 mL of 400 mM Na2CO3 was added to stop the reaction and the absorbance at 410 nm was measured. Boiled enzyme extract was used as the control. One unit of β-Gal activity was defined as production of 1 μmol p-nitrophenyl per minute.
3.2. Changes of fruit firmness and decay rate Firmness is one of the important indicators of fruit quality after harvest. The decrease of firmness is positively correlated with degradation of the cell wall in fruits (Zhou et al., 2011b). A continuous decreased in fruit firmness of peaches was observed in all groups during cold storage (Fig. 2A). However, NZLDPE-packaging significantly inhibited the decrease of firmness (p < 0.05). On day 40, NZLDPEpacked peaches retained 12.9 N of firmness while there were only 11.6 N and 9.6 N of firmness in peaches packed in LDPE and control, respectively. Hu et al. (2011) reported that application of polyethylene with nano-Ag, nano-TiO2 and montmorillonite was capable of delaying kiwifruit softening during storage. Peaches are highly perishable and susceptible to fungal decay, which severely reduce the goods value (Rai and Paul, 2007). As shown in Fig. 2B, incipient decay of peaches without packaging was observed on day 16. LDPE-packaging and NZLDPE-packaging postponed the decay incidence for almost eight days, compared to control. The decay rate increased rapidly from day 24 to day 40. Both LDPE-packaging and NZLDPE-packaging alleviated the decay and NZLDPE-packaging performed better effects. This lower decay rate might be attributed to the relative higher barrier property of nano-packaging materials against O2 and H2O than the normal packaging, which kept the fruit in a lowmoisture surroundings, thus not favoring the growth of fungi (Yang
2.16. Statistical analysis The experiments were conducted in a completely randomized design. All statistical analyses were performed with SPSS (SPSS Inc., Chicago, IL, USA). The ANOVA was performed, and means were compared at a significance level of 0.05. 3. Result and discussion 3.1. Changes of O2 and CO2 concentrations in the packages Modified atmosphere packaging (MAP) is achieved by the natural interaction between the respiration rate of the product and the transfer of gases through the packaging material (Oliveira et al., 2015). High aspect ratio of particles in film can bring about a permeation barrier in the LDPE by hindering the penetration of gas molecules and increasing their average path length, which may result in the decreased transmission rate of CO2 and O2 of LDPE-packaging (Hosseinkhanli et al., 130
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Fig. 2. The fruit firmness (A) and decay rate (B) of ‘Hujingmilu’ peach in NZLDPEpackaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
et al., 2010).
3.3. Changes of browning index, electrolyte leakage and relative viscosity Browning in the internal flesh is one of symptoms of chilling injury in peaches (Brummell et al., 2004a). It is evident to be caused by PPO, which catalyzes the oxidation of polyphenols to form colored quinones (Yan et al., 2015). PPO and polyphenols are located in the cytoplasm and vacuoles in intact tissues respectively. The damage of cell membrane results in the mixing of the enzyme and substrates, and exposure to oxygen leads to the browning reaction (Degl'Innocenti et al., 2005). As shown in Fig. 3A, chilling symptoms appeared on day 16 and the browning index increased during the storage at 2 °C. Both packages inhibited the increase of browning in which NZLDPE-packaging was observed with greater effect. The browning index of peaches with NZLDPE-packaging was 35.79% lower than peaches with no packaging on day 40 (p < 0.05). Similarly, Li et al. (2011) reported nano-ZnOcoated packaging alleviated the increase of browning index of fresh-cut apple during cold storage. Cell membrane is a primary site for the development of chilling injury and electrolyte leakage is regarded as the indicator of damages to membranes (Janero, 1990; Jin et al., 2013). In present study, electrolyte leakage increased during the whole storage period while NZLDPEpackaging inhibited the rise in electrolyte leakage. The value in NZLDPE-packaging peaches was 27.64% and 11.43% lower than that in unpackaged peaches and LDPE-packed peaches, respectively (Fig. 3B). Similar result was reported by Jiang et al. (2014) on an herbal medicine named Gynura which packed with nano-powder-PE-packaging during storage at 0 °C. Changes of viscosity is due to the type of polymers in a crude flesh homogenate (Fruk et al., 2014). As shown in Fig. 3C, relative viscosity increased gradually during the cold storage for all groups. The value of peaches in NZLDPE-packaging reached 18.6% on day 40, which is significantly lower than that in unpackaged peaches (26.3%) and LDPE-
Fig. 3. The browning index (A), electrolyte leakage (B) and relative viscosity (C) of ‘Hujingmilu’ peach in NZLDPE-packaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
packaging peaches (21.7%) (p < 0.05). Previous research indicated that no high viscosity in mealy fruit was related to the reduction of the most easily extractable cell wall pectins (Brummell et al., 2004a). 3.4. Changes of ASP, CSP and WSP contents Pectins are one of the major components in the primary cell wall and in the central lamella. The loss of fruit firmness is associated with the disassembly of primary cell wall and central lamella structures (Chen et al., 2015). During postharvest storage, the levels of ASP and CSP, which are enriched for ionically and covalently bound pectins respectively, decreased whereas the level of WSP, which solubilizes in vivo but remains in the apoplast, increased. The changes of pectin contents caused the significant reduction of cell adhesion, and resulted in the dissolution of middle lamella (Carrington et al., 1993; Paniagua et al., 2014). In present study, ASP content declined gradually during the cold storage, while NZLDPE-packaging postponed this phenomenon. As shown in Fig. 4A, 11.32 g kg−1 of ASP was observed in peaches packed in NZLDPE compared with 9.01 g kg−1 and 10.08 g kg−1 in control and LDPE on day 40, respectively. Ketsa et al. (1998) suggested that fruit softening during ripening was accompanied by a rapid decline in ASP. 131
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Fig. 4. The ASP (A), CSP (B) and WSP (C) contents of ‘Hujingmilu’ peach in NZLDPEpackaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
Fig. 5. The PE (A), PG (B) and β-Gal (C) activities of ‘Hujingmilu’ peach in NZLDPEpackaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
The higher level of ASP in NZLDPE-packaging peaches might result in the higher fruit firmness during cold storage. In contrast, both packages accelerated the decrease of CSP content by 35.09% and 20.11% lower in peaches packed with NZLDPE-packaging and LDPE-packaging, respectively, than those with no packaging (Fig. 4B). It was reported that the CSP fraction produced a gel with calcium in the middle lamella, which caused woolliness of flesh, the main symptom of chilling injury (Zhou et al., 2000a). NZLDPE-packaging reduced the accumulation of the gel and therefore effectively alleviated the development of chilling injury. Similar result was found in the change profiles of ASP and CSP during chilling stress in zucchini fruit (Carvajal et al., 2015). In comparison to CSP, WSP was confirmed that it did not form a gel in vitro (Levaj et al., 2003). During storage, WSP content increased, accompanying with the softening of peaches. In addition, the NZLDPEpackaging accelerated the increase of WSP, which might be attributed to the degradation of CSP (Fig. 4C).
galacturonosyl residues, by destroying the Ca2+ cross-linkages, and leading to loss of texture. And it also plays an important role in determining the extent to which pectin is accessible to degradation by PG (Fischer and Bennett, 1991). PG catalyzes the hydrolysis of α(1 → 4) galacturonan linkages of in galacturonides and other polysaccharides, resulting in ripening-associated pectin degradation and fruit softening (Özkaya et al., 2016). As shown in Fig. 5A and B, PE activity increased rapidly in the first twenty-four days during the cold storage and then decreased slowly, while PG activity increased gradually during the whole storage. However, PE activity was much higher than PG activity during storage, which may lead to failure of normal pectin degradation. Pectin matrix can be deesterified without subsequent depolymerisation under the relatively higher PE and lower PG activities, which may lead to the formation of high-molecular-weight pectin with a low degree of esterification, and therefore cause woolliness (Fruk et al., 2014; Zhou et al., 2000b). In present study, both packages inhibited the increase of PE activity and promoted PG activity, which may reduce the accumulation of high-molecular-weight pectin, and therefore contributed to the alleviation of chilling injury of peaches (Fig. 5A and B). Compared with LDPE-packaging, NZLDPE-packaging had better effect on maintaining
3.5. Changes of PE, PG and β-Gal activities The degradation of pectin is closely related to the activities of pectinase. PE catalyzes demethylation of the C6 carboxyl group of 132
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fruit quality, which was possibly due to the rapid formation of higher CO2 and lower O2 contents in storage atmosphere. Zhou et al. (2000b) reported CA (10% CO2, 3% O2 controlled atmosphere storage) led to a high PG/PE ratio of activity in nectarines which coordinated both demethylation and cleavage of pectin during ripening. β-Gal can degrade galactose-containing cell wall polysaccharides, including pectin and hemicellulose. It removes β-(1 → 4)-linked galactose, releases free galactose, and results in the damage of cell wall (Biles et al., 1997). Brummell et al. (2004b) pointed out that the advanced stage of mealiness in peaches was not only correlated with low levels of endo-PG and high levels of PE activity, but also with reduced activities of β-Gal during cold storage. In present study, the activity of β-Gal decreased gradually while NZLDPE-packaging significantly maintained the high level of β-Gal activity. On day 40, there was 70.6 U g−1 of β-Gal activity remained in peaches in NZLDPE-packaging, compared with 50.6 U g−1 and 60.6 U g−1 of β-Gal activity in peaches with no packaging and in LDPE-packaging, respectively (Fig. 5C). Whether high activity of β-Gal contributed to the reduction of calcium-pectate gel complexes, as PG did to alleviate chilling injury, needs further investigation. 4. Conclusions In conclusion, NZLDPE-packaging, which formed the gas atmosphere with low O2 and high CO2 rapidly, was successfully applied in the cold storage of peaches at 2 °C. In comparison to LDPE-packaging, NZLDPE-packaging showed superior chilling tolerance of peaches during cold storage, through the maintaining of fruit firmness, reduction of browning and decay rate, and inhibition of rises in electrolyte leakage and relative viscosity. Meanwhile, NZLDPE-packaging retained ASP and WSP levels, and decreased CSP content by suppressing PE activity and enhancing PG and β-Gal activities. Maintaining cell wall structure and reducing high-molecular-weight pectin content with a low degree of esterification might alleviate chilling injury and extend storage. Therefore, NZLDPE-packaging might be an effective approach to the preservation of peaches in cold storage. However, the safety of nano-ZnO is still under investigated. Further study should focus on food safety in prior to industrialized application. Acknowledgements The research was financially supported by the National Key Technologies R&D Program of China (2015BAD16B06, 2017YFD0401304) and the National Natural Science Foundation of China (31371856). References Özkaya, Ö., Yildirim, D., Dündar, Ö., Tükel, S.S., 2016. Effects of 1-methylcyclopropene (1MCP) and modified atmosphere packaging on postharvest storage quality of nectarine fruit. Sci. Hortic. 198, 454–461. Biles, C.L., Bruton, B.D., Russo, V., Wall, M.M., 1997. Characterisation of β-galactosidase isozymes of ripening peppers. J. Sci. Food Agric. 75, 237–243. Brummell, D.A., Dal Cin, V., Lurie, S., Crisosto, C.H., Labavitch, J.M., 2004a. Cell wall metabolism during the development of chilling injury in cold-stored peach fruit: association of mealiness with arrested disassembly of cell wall pectins. J. Exp. Bot. 55, 2041–2052. Brummell, D.A., Dal Cin, V., Crisosto, C.H., Labavitch, J.M., 2004b. Cell wall metabolism during maturation, ripening and senescence of peach fruit. J. Exp. Bot. 55, 2029–2039. Bu, J., Yu, Y., Aisikaer, G., Ying, T., 2013. Postharvest UV-C irradiation inhibits the production of ethylene and the activity of cell wall-degrading enzymes during softening of tomato (Lycopersicon esculentum L.) fruit. Postharvest Biol. Technol. 86, 337–345. Cao, S., Zheng, Y., Wang, K., Rui, H., Tang, S., 2009. Effect of 1-methylcyclopropene treatment on chilling injury, fatty acid and cell wall polysaccharide composition in loquat fruit. J. Agric. Food Chem. 57, 8439–8443. Carrington, C., Greve, L.C., Labavitch, J.M., 1993. Cell wall metabolism in ripening fruit (VI. Effect of the antisense polygalacturonase gene on cell wall changes accompanying ripening in transgenic tomatoes). Plant Physiol. 103, 429–434. Chaudhry, Q., Scotter, M., Blackburn, J., Ross, B., Boxall, A., Castle, L., Aitken, R., Watkins, R., 2008. Applications and implications of nanotechnologies for the food sector. Food Addit.
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Effect of nano-ZnO-packaging on chilling tolerance and pectin metabolism of peaches during cold storage
MARK
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Dong Li, Li Li, Zisheng Luo , Hongyan Lu, Yang Yue Zhejiang University, College of Biosystems Engineering and Food Science, Key Laboratory of Agro-Products Postharvest Handing Ministry of Agriculture, Zhejiang Key Laboratory for Agro-Food Processing, Hangzhou, 310058, People’s Republic of China
A R T I C L E I N F O
A B S T R A C T
Keywords: Peach Nano-ZnO Chilling injury Quality Cell wall Pectinase
The effects of nano-ZnO-based low-density polyethylene (NZLDPE)-packaging on chilling tolerance and pectin metabolism in postharvest “Hujingmilu” peaches were investigated. Peaches packed in low-density polyethylene (LDPE) and NZLDPE were stored at 2 °C for 40 d. Compared with the control, both packages alleviated the development of chilling injury, showed higher fruit firmness with lower browning index, electrolyte leakage, relative viscosity, and decay rate. NZLDPE-packaging performed better than LDPE-packaging because of the rapid formation of low O2 and high CO2. Moreover, NZLDPE-packaging inhibited pectin esterase and enhanced polygalacturonase and β-galactosidase, leading to the promotion of alkali soluble-pectin and water-soluble pectin, and the decrease of chelater-soluble pectin. These effects were of great benefit to the maintaining of cell wall structure and the degradation of calcium-pectate gel, which finally alleviated the chilling injury and therefore maintained good quality during chilling stress.
1. Introduction Peach is a popular fruit for consumers because of its juiciness and attractive taste. Peaches ripen and decay quickly at room temperature after harvest, which leads to a loss of sales value. Cold storage is a widely used technology for postharvest fruit and vegetables to delay senescence and extend shelf-life (Wang, 1990). Nevertheless, as a climacteric fruit, peach suffers chilling injury at 2–5 °C. Several technologies, such as treatment with hot air (Yu et al., 2016), glycine betaine (Shan et al., 2016), and oxalic acid (Jin et al., 2014), have been reported to alleviate chilling injury in peaches during cold storage. However, reliable investigations on the practical techniques to inhibit or alleviate chilling injuries are still limited. Chilling injury results in many macroscopic symptoms in peaches, including browning, wooliness, reddening and leatheriness, which are related to cell wall integrity and pectin metabolism (Lurie and Crisosto, 2005). Cao et al. (2009) indicated that chilling injury is partly due to biochemical modifications of cell wall polysaccharides. These processes involve hydrolytic enzymes such as pectin esterase (PE), polygalacturonase (PG) and β-galactosidase (β-Gal) (Fischer et al., 1991). Fruk et al. (2014) reported that the relatively higher PE and lower PG activities observed in peaches were response to chilling stress, which may lead to the formation of high-molecular-weight pectin with a low degree of esterification. This kind of pectin can combine with calcium
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Corresponding author. E-mail address: [email protected] (Z. Luo).
http://dx.doi.org/10.1016/j.scienta.2017.07.003 Received 7 March 2017; Received in revised form 29 June 2017; Accepted 3 July 2017 0304-4238/ © 2017 Published by Elsevier B.V.
to form calcium-pectate gel complexes, which then bind with free water and eventually lead to woolliness, a symptom of chilling injury (Brummell et al., 2004a). In recent decades, nanomaterials used in food packaging have been actively explored (Chaudhry et al., 2008). Nanocomposites enhanced barrier and mechanical properties, and possessed antimicrobial effect along with spore germination inhibition ability in comparison to general polymer (Panea et al., 2014). Various nanocomposites including nano-ZnO-coated polyvinyl chloride (PVC) (Li et al., 2011), nano-Ag2Obased low-density polyethylene (Zhou et al., 2011a), and nano-CaCO3based low-density polyethylene (Luo et al., 2014) applied in the preservation of fruit and vegetables have been reported. During cold storage, polyethylene with nano-Ag, nano-TiO2 and montmorillonite has been proven to be efficient for inhibiting ethylene production, reducing degradation of nutritional components, extending organoleptic characteristics, preventing physiologic changes, and thus delaying the ripening and extending shelf-life of harvested kiwifruit (Hu et al., 2011). Li et al. (2011) stated that nano-ZnO-packaging film preserved fresh-cut ‘Fuji’ apple through reduction of fruit decay rate, depreciation of malondialdehyde (MDA) and ethylene accumulation, maintaining of total soluble solids (TSS) and titratable acid (TA) levels and inhibition of polyphenol oxidase (PPO) and pyrogallol peroxidase (POD) activities. However, to the best of our knowledge, the effect of nano-ZnO-packaging on chilling tolerance of peaches during cold storage has not been
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2. Materials and methods
distilled water in a beaker were stirred slowly for 20 min and the electrolyte conductivity of leach liquor (C1) was measured with a conductivity meter. Then the leach liquor with discs was boiled for 10 min and cooled down to the room temperature. Electrolyte conductivity (C2) was measured. Each treatment was conducted for three biological replicates. The rate of electrolyte leakage was calculated as (C1/C2) × 100%.
2.1. Plant material and treatments
2.6. Measurement of relative viscosity
“Hujingmilu” peaches (Prunus persica L.) with 70–80% maturity (with a firmness of 16.1 ± 0.7 N, total soluble solids content of 11.2 ± 0.2%, and titratable acid content of 0.21 ± 0.02%) were harvested in Yuhang District of Hangzhou City and transferred to the laboratory in Zhejiang University immediately. The fruit were air cooled to 4 °C in 6 h. The first and second groups of peaches were packed into low-density polyethylene (LDPE)-packaging (6 bags) and NZLDPE-packaging (6 bags) with a size of 20 cm × 30 cm respectively, and then sealed and stored at 2 °C for up to 40 days. The third group of peaches were unpackaged and stored under the same condition as control. For each treatment, one bag of peaches was randomly taken every eight days during storage for subsequent analyses (totally 6 bags were used). And another three bags were used to observe the decay rate every eight days during the whole storage. Three biological replications were conducted.
Peach flesh from one bag was mashed and filtered using gauze. After centrifuged at 5000 × g for 15 min, 10 mL of supernatant was collected and the time supernatant (T1) and distilled water (T0) flowed through a capillary viscometer with 0.4 mm inner diameter at 25 °C were measured respectively. Each treatment was conducted for three biological replicates. The relative viscosity was presented as (T1-T0)/T0.
researched. The objective of this study was to investigate the effect of nano-ZnObased low-density polyethylene (NZLDPE)-packaging on chilling tolerance and pectin metabolism in postharvest “Hujingmilu” peaches during cold storage.
2.7. Measurement of fruit firmness A texture analyzer (TA-XT2i, Stable Micro Systems Ltd., Godalmin, UK) was used to determine the peach firmness. Three peaches form one bag were measured on opposite sides of the equator by a 5 mm diameter flat probe. The test speed was 1.5 mm s−1 and the distance was 8 mm. The maximum force (Nm) was recorded as peach firmness and the average values were calculated. Each treatment was conducted for three biological replicates.
2.2. Preparation of NZLDPE materials 2.8. Measurement of decay rate A NZLDPE masterbatch containing 56% (w/w) of LDPE granule (softening point 95 °C, density 920 kg m−3), 30% (w/w) of the commercial nano-ZnO (30 nm) and 14% (w/w) of cross-link reagent ester titanate was blended by a high-speed mixer for 1 h. After cooled down, the mixture was extruded to a NZLDPE masterbatch by a twin-screw extruder with a screw speed of 300 r min−1. Then, 38.5 kg of LDPE granule and 1.5 kg of masterbatch were blended for 0.5 h. The above nanocompounds were made into films of 40 μm thickness by a plastic extruder, and the films were made into NZLDPE bags of 20 cm × 30 cm using a heat sealer after cooling. LDPE bags with the same thickness and size were made under the same condition.
For each treatment, another three bags were used to observe the decay rate every eight days during the whole storage. Any peach with visible mould growth was considered decay. The decay rate (%) was calculated as (the amounts of decayed peaches × 100)/(the total amounts of peaches). Each treatment was conducted for three biological replicates. 2.9. Isolation of cell wall material (CWM) The CWM was prepared using the method described by Chen et al. (2015) with some modifications. One hundred grams frozen flesh pooled from six peaches was homogenized and kept boiling reflux with 500 mL 80% (v/v) ethanol for 0.5 h. After immediate filtration, the residue was collected and washed with 80% (v/v) ethanol for several times until no reducing sugar was detected in the filtrate. Then the residue was washed with acetone and incubated overnight with 90% (v/v) dimethylsulphoxide at 4 °C. The CWM was filtered, washed for several times with distilled water, dried in a vacuum oven and stored in a vacuum desiccator.
2.3. Measurement of in-packaging atmospheric composition Changes in CO2 and O2 concentrations were measured by injecting samples of headspace gases form the packages into a gas chromatograph with a thermal conductivity detector (GC-14A; Shimadzu, Suzhou, China). Each treatment was conducted for three biological replicates. 2.4. Measurement of browning index
2.10. Extraction of CWM constituents For each replicate, eight peaches from one bag were used for evaluation of browning index. Peaches were cut along the fruit suture and the browning scale (B) was divided into four degrees according to the percentage of browning area of flesh; 0: no browning; 1: B ≤ 25%; 2: 25% < B ≤ 50%; 3: B > 50%. The browning index was calculated based on the following formula:
Extraction of CWM constituents was performed according to the methods described by Chen et al. (2015) with some modifications. The water-soluble pectin (WSP), chelater-soluble pectin (CSP) and alkalisoluble pectin (ASP) were extracted consecutively on the CWM. Three hundred milligrams of CWM was added to 100 mL 50 mM sodium acetate buffer (pH 6.5), oscillated and extracted at ambient temperature for 3 h. This process repeated for 3 times and the filtrate contained WSP was collected. Similarly, CSP and ASP was extracted by 50 mM sodium acetate buffer (pH 6.5) containing 50 mM ethylene diamine tetraacetic acid and 100 mM KOH respectively.
Browing index(%) =
∑
(browning scale × the amount of peaches at this scale) total peach number ×3
× 100
2.5. Measurement of electrolyte leakage
2.11. Measurement of CWM constituents
Discs of peach peel from one bag were excised randomly using a 6 mm diameter stainless steel borer. Ten grams of discs with 20 mL
The CWM constituents were measured based on the method described by Bu et al. (2013). WSP, CSP and ASP extracts (5 mL) were 129
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added to 30 mL H2SO4 and heated for 1 h in boiling water. After cooling to room temperature, the solution was mixed with 5 mL of 0.15% (w/v) carbazole reagent and placed for 0.5 h in the dark. The absorbance at 530 nm was measured and galacturonic acid (GalA) was used to prepare a standard curve. The results were expressed as g kg−1 fresh weight. 2.12. Extraction of crude enzyme Enzyme extraction was carried out according to the method described by Meng et al. (2009). Five grams of fruit tissues were homogenized with 20 mL 0.15 M NaCl containing 10 g L−1 polyvinylpolypyrrolidone (PVPP). The homogenate was stirred for 20 min and centrifuged at 15,000 × g for 20 min. The supernatant was collected and used for assays of PE, PG and β-Gal activities. 2.13. The activity of PE The activity of PE was measured according to Pan et al. (2014) with some modifications. Five milliliters of crude extract was added to 20 mL of 1% (w/v) pectin in 100 mM NaCl, titrated with 20 mM NaOH to keep pH 7.4 at 37 °C for 0.5 h. Boiled enzyme extract was used as the control. One unit of PE activity was defined as the amount of PE that released 1 μmol of acid per minute. 2.14. The activity of PG The activity of PG was determined according to the method described by Bu et al. (2013) with a slight modification. Enzyme extract (0.5 mL) was added to 0.5 mL of 0.5% (w/v) polygalacturonic acid in 50 mM sodium acetate buffer (pH 4.4) and incubated at 37 °C for 1 h. Then 1 mL of 3,5-dinitrosalicylic acid (DNS) was added to the mixture and heated for 5 min in boiled water. After cooled down, 25 mL of distilled water was added to the mixture and the absorbance at 540 nm was measured. Boiled enzyme extract was used as the control. One unit of PG activity was defined as production of 1 μmol GalA per minute.
Fig. 1. The O2 (A) and CO2 (B) concentrations of ‘Hujingmilu’ peach in NZLDPE-packaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
2013). Meanwhile, previous study indicated that adding nano-ZnO to poly(butylene adipate-co-terephthalate) decreased the O2 transmission rate of films and thus improved their barrier (Venkatesan and Rajeswari, 2017). As showed in Fig. 1, the O2 concentrations decreased while CO2 concentration increased within both LDPE-packaging and NZLDPE-packaging during the cold storage. NZLDPE-packaging could reach the gas environment with lower O2 and higher CO2 more rapidly compared with LDPE-packaging, which is benefit on fruit cold storage (de Azeredo, 2009). The result was in accordance with the research reported by Luo et al. (2014).
2.15. The activity of β-Gal The activity of β-Gal was measured as described by Minas et al. (2014). Two hundred microliters of enzyme extract was added to 2 mL of 3 mM p-nitrophenylgalactopyranoside in 50 mM HAc-NaAc buffer (pH 5.0) and incubated at 37 °C for 1 h. After incubated, 2 mL of 400 mM Na2CO3 was added to stop the reaction and the absorbance at 410 nm was measured. Boiled enzyme extract was used as the control. One unit of β-Gal activity was defined as production of 1 μmol p-nitrophenyl per minute.
3.2. Changes of fruit firmness and decay rate Firmness is one of the important indicators of fruit quality after harvest. The decrease of firmness is positively correlated with degradation of the cell wall in fruits (Zhou et al., 2011b). A continuous decreased in fruit firmness of peaches was observed in all groups during cold storage (Fig. 2A). However, NZLDPE-packaging significantly inhibited the decrease of firmness (p < 0.05). On day 40, NZLDPEpacked peaches retained 12.9 N of firmness while there were only 11.6 N and 9.6 N of firmness in peaches packed in LDPE and control, respectively. Hu et al. (2011) reported that application of polyethylene with nano-Ag, nano-TiO2 and montmorillonite was capable of delaying kiwifruit softening during storage. Peaches are highly perishable and susceptible to fungal decay, which severely reduce the goods value (Rai and Paul, 2007). As shown in Fig. 2B, incipient decay of peaches without packaging was observed on day 16. LDPE-packaging and NZLDPE-packaging postponed the decay incidence for almost eight days, compared to control. The decay rate increased rapidly from day 24 to day 40. Both LDPE-packaging and NZLDPE-packaging alleviated the decay and NZLDPE-packaging performed better effects. This lower decay rate might be attributed to the relative higher barrier property of nano-packaging materials against O2 and H2O than the normal packaging, which kept the fruit in a lowmoisture surroundings, thus not favoring the growth of fungi (Yang
2.16. Statistical analysis The experiments were conducted in a completely randomized design. All statistical analyses were performed with SPSS (SPSS Inc., Chicago, IL, USA). The ANOVA was performed, and means were compared at a significance level of 0.05. 3. Result and discussion 3.1. Changes of O2 and CO2 concentrations in the packages Modified atmosphere packaging (MAP) is achieved by the natural interaction between the respiration rate of the product and the transfer of gases through the packaging material (Oliveira et al., 2015). High aspect ratio of particles in film can bring about a permeation barrier in the LDPE by hindering the penetration of gas molecules and increasing their average path length, which may result in the decreased transmission rate of CO2 and O2 of LDPE-packaging (Hosseinkhanli et al., 130
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Fig. 2. The fruit firmness (A) and decay rate (B) of ‘Hujingmilu’ peach in NZLDPEpackaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
et al., 2010).
3.3. Changes of browning index, electrolyte leakage and relative viscosity Browning in the internal flesh is one of symptoms of chilling injury in peaches (Brummell et al., 2004a). It is evident to be caused by PPO, which catalyzes the oxidation of polyphenols to form colored quinones (Yan et al., 2015). PPO and polyphenols are located in the cytoplasm and vacuoles in intact tissues respectively. The damage of cell membrane results in the mixing of the enzyme and substrates, and exposure to oxygen leads to the browning reaction (Degl'Innocenti et al., 2005). As shown in Fig. 3A, chilling symptoms appeared on day 16 and the browning index increased during the storage at 2 °C. Both packages inhibited the increase of browning in which NZLDPE-packaging was observed with greater effect. The browning index of peaches with NZLDPE-packaging was 35.79% lower than peaches with no packaging on day 40 (p < 0.05). Similarly, Li et al. (2011) reported nano-ZnOcoated packaging alleviated the increase of browning index of fresh-cut apple during cold storage. Cell membrane is a primary site for the development of chilling injury and electrolyte leakage is regarded as the indicator of damages to membranes (Janero, 1990; Jin et al., 2013). In present study, electrolyte leakage increased during the whole storage period while NZLDPEpackaging inhibited the rise in electrolyte leakage. The value in NZLDPE-packaging peaches was 27.64% and 11.43% lower than that in unpackaged peaches and LDPE-packed peaches, respectively (Fig. 3B). Similar result was reported by Jiang et al. (2014) on an herbal medicine named Gynura which packed with nano-powder-PE-packaging during storage at 0 °C. Changes of viscosity is due to the type of polymers in a crude flesh homogenate (Fruk et al., 2014). As shown in Fig. 3C, relative viscosity increased gradually during the cold storage for all groups. The value of peaches in NZLDPE-packaging reached 18.6% on day 40, which is significantly lower than that in unpackaged peaches (26.3%) and LDPE-
Fig. 3. The browning index (A), electrolyte leakage (B) and relative viscosity (C) of ‘Hujingmilu’ peach in NZLDPE-packaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
packaging peaches (21.7%) (p < 0.05). Previous research indicated that no high viscosity in mealy fruit was related to the reduction of the most easily extractable cell wall pectins (Brummell et al., 2004a). 3.4. Changes of ASP, CSP and WSP contents Pectins are one of the major components in the primary cell wall and in the central lamella. The loss of fruit firmness is associated with the disassembly of primary cell wall and central lamella structures (Chen et al., 2015). During postharvest storage, the levels of ASP and CSP, which are enriched for ionically and covalently bound pectins respectively, decreased whereas the level of WSP, which solubilizes in vivo but remains in the apoplast, increased. The changes of pectin contents caused the significant reduction of cell adhesion, and resulted in the dissolution of middle lamella (Carrington et al., 1993; Paniagua et al., 2014). In present study, ASP content declined gradually during the cold storage, while NZLDPE-packaging postponed this phenomenon. As shown in Fig. 4A, 11.32 g kg−1 of ASP was observed in peaches packed in NZLDPE compared with 9.01 g kg−1 and 10.08 g kg−1 in control and LDPE on day 40, respectively. Ketsa et al. (1998) suggested that fruit softening during ripening was accompanied by a rapid decline in ASP. 131
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Fig. 4. The ASP (A), CSP (B) and WSP (C) contents of ‘Hujingmilu’ peach in NZLDPEpackaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
Fig. 5. The PE (A), PG (B) and β-Gal (C) activities of ‘Hujingmilu’ peach in NZLDPEpackaging during cold storage at 2 °C for 40 d. Values are presented as means ± SD (n = 3).
The higher level of ASP in NZLDPE-packaging peaches might result in the higher fruit firmness during cold storage. In contrast, both packages accelerated the decrease of CSP content by 35.09% and 20.11% lower in peaches packed with NZLDPE-packaging and LDPE-packaging, respectively, than those with no packaging (Fig. 4B). It was reported that the CSP fraction produced a gel with calcium in the middle lamella, which caused woolliness of flesh, the main symptom of chilling injury (Zhou et al., 2000a). NZLDPE-packaging reduced the accumulation of the gel and therefore effectively alleviated the development of chilling injury. Similar result was found in the change profiles of ASP and CSP during chilling stress in zucchini fruit (Carvajal et al., 2015). In comparison to CSP, WSP was confirmed that it did not form a gel in vitro (Levaj et al., 2003). During storage, WSP content increased, accompanying with the softening of peaches. In addition, the NZLDPEpackaging accelerated the increase of WSP, which might be attributed to the degradation of CSP (Fig. 4C).
galacturonosyl residues, by destroying the Ca2+ cross-linkages, and leading to loss of texture. And it also plays an important role in determining the extent to which pectin is accessible to degradation by PG (Fischer and Bennett, 1991). PG catalyzes the hydrolysis of α(1 → 4) galacturonan linkages of in galacturonides and other polysaccharides, resulting in ripening-associated pectin degradation and fruit softening (Özkaya et al., 2016). As shown in Fig. 5A and B, PE activity increased rapidly in the first twenty-four days during the cold storage and then decreased slowly, while PG activity increased gradually during the whole storage. However, PE activity was much higher than PG activity during storage, which may lead to failure of normal pectin degradation. Pectin matrix can be deesterified without subsequent depolymerisation under the relatively higher PE and lower PG activities, which may lead to the formation of high-molecular-weight pectin with a low degree of esterification, and therefore cause woolliness (Fruk et al., 2014; Zhou et al., 2000b). In present study, both packages inhibited the increase of PE activity and promoted PG activity, which may reduce the accumulation of high-molecular-weight pectin, and therefore contributed to the alleviation of chilling injury of peaches (Fig. 5A and B). Compared with LDPE-packaging, NZLDPE-packaging had better effect on maintaining
3.5. Changes of PE, PG and β-Gal activities The degradation of pectin is closely related to the activities of pectinase. PE catalyzes demethylation of the C6 carboxyl group of 132
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fruit quality, which was possibly due to the rapid formation of higher CO2 and lower O2 contents in storage atmosphere. Zhou et al. (2000b) reported CA (10% CO2, 3% O2 controlled atmosphere storage) led to a high PG/PE ratio of activity in nectarines which coordinated both demethylation and cleavage of pectin during ripening. β-Gal can degrade galactose-containing cell wall polysaccharides, including pectin and hemicellulose. It removes β-(1 → 4)-linked galactose, releases free galactose, and results in the damage of cell wall (Biles et al., 1997). Brummell et al. (2004b) pointed out that the advanced stage of mealiness in peaches was not only correlated with low levels of endo-PG and high levels of PE activity, but also with reduced activities of β-Gal during cold storage. In present study, the activity of β-Gal decreased gradually while NZLDPE-packaging significantly maintained the high level of β-Gal activity. On day 40, there was 70.6 U g−1 of β-Gal activity remained in peaches in NZLDPE-packaging, compared with 50.6 U g−1 and 60.6 U g−1 of β-Gal activity in peaches with no packaging and in LDPE-packaging, respectively (Fig. 5C). Whether high activity of β-Gal contributed to the reduction of calcium-pectate gel complexes, as PG did to alleviate chilling injury, needs further investigation. 4. Conclusions In conclusion, NZLDPE-packaging, which formed the gas atmosphere with low O2 and high CO2 rapidly, was successfully applied in the cold storage of peaches at 2 °C. In comparison to LDPE-packaging, NZLDPE-packaging showed superior chilling tolerance of peaches during cold storage, through the maintaining of fruit firmness, reduction of browning and decay rate, and inhibition of rises in electrolyte leakage and relative viscosity. Meanwhile, NZLDPE-packaging retained ASP and WSP levels, and decreased CSP content by suppressing PE activity and enhancing PG and β-Gal activities. Maintaining cell wall structure and reducing high-molecular-weight pectin content with a low degree of esterification might alleviate chilling injury and extend storage. Therefore, NZLDPE-packaging might be an effective approach to the preservation of peaches in cold storage. However, the safety of nano-ZnO is still under investigated. Further study should focus on food safety in prior to industrialized application. Acknowledgements The research was financially supported by the National Key Technologies R&D Program of China (2015BAD16B06, 2017YFD0401304) and the National Natural Science Foundation of China (31371856). References Özkaya, Ö., Yildirim, D., Dündar, Ö., Tükel, S.S., 2016. Effects of 1-methylcyclopropene (1MCP) and modified atmosphere packaging on postharvest storage quality of nectarine fruit. Sci. Hortic. 198, 454–461. Biles, C.L., Bruton, B.D., Russo, V., Wall, M.M., 1997. Characterisation of β-galactosidase isozymes of ripening peppers. J. Sci. Food Agric. 75, 237–243. Brummell, D.A., Dal Cin, V., Lurie, S., Crisosto, C.H., Labavitch, J.M., 2004a. Cell wall metabolism during the development of chilling injury in cold-stored peach fruit: association of mealiness with arrested disassembly of cell wall pectins. J. Exp. Bot. 55, 2041–2052. Brummell, D.A., Dal Cin, V., Crisosto, C.H., Labavitch, J.M., 2004b. Cell wall metabolism during maturation, ripening and senescence of peach fruit. J. Exp. Bot. 55, 2029–2039. Bu, J., Yu, Y., Aisikaer, G., Ying, T., 2013. Postharvest UV-C irradiation inhibits the production of ethylene and the activity of cell wall-degrading enzymes during softening of tomato (Lycopersicon esculentum L.) fruit. Postharvest Biol. Technol. 86, 337–345. Cao, S., Zheng, Y., Wang, K., Rui, H., Tang, S., 2009. Effect of 1-methylcyclopropene treatment on chilling injury, fatty acid and cell wall polysaccharide composition in loquat fruit. J. Agric. Food Chem. 57, 8439–8443. Carrington, C., Greve, L.C., Labavitch, J.M., 1993. Cell wall metabolism in ripening fruit (VI. Effect of the antisense polygalacturonase gene on cell wall changes accompanying ripening in transgenic tomatoes). Plant Physiol. 103, 429–434. Chaudhry, Q., Scotter, M., Blackburn, J., Ross, B., Boxall, A., Castle, L., Aitken, R., Watkins, R., 2008. Applications and implications of nanotechnologies for the food sector. Food Addit.
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