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Effect of 1-methylcyclopropene on shelf life, visual quality and nutritional quality of netted melon Y Shi, BL Wang, DJ Shui, LL Cao, C Wang, T Yang, XY Wang and HX Ye Food Science and Technology International published online 4 February 2014 DOI: 10.1177/1082013214520786 The online version of this article can be found at: http://fst.sagepub.com/content/early/2014/02/04/1082013214520786
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Original Article
Effect of 1-methylcyclopropene on shelf life, visual quality and nutritional quality of netted melon Y Shi1, BL Wang1, DJ Shui2, LL Cao1, C Wang1, T Yang1, XY Wang2 and HX Ye1
Abstract The effects of 1-methylcyclopropene (1-MCP) on shelf life, fruit visual quality and nutritional quality were investigated. Netted melons were treated with air (control) and 0.6 ml l1 1-MCP at 25 C for 24 h, and then stored at 25 C or 10 C for 10 days. 1-MCP significantly extended the shelf life, inhibited weight loss and delayed firmness decline of melon fruits. Ethylene production was also inhibited and respiration rate was declined. 1-MCP retarded 1-aminocyclopropane-1-carboxylic acid (ACC) increases and inhibited ACC synthase and ACC oxidase activity. Moreover, 1-MCP treatment reduced the decrease in total soluble solids and titratable acidity, as well as the decrease of the content of sugars (sucrose, fructose and glucose). These results indicated that 1-MCP treatment is a good method to extend melon shelf life and maintain fruit quality, and the combination of 1-MCP and low temperature storage resulted in more acceptable fruit quality.
Keywords Netted melon, 1-methylcyclopropene (1-MCP), shelf life, ethylene, firmness, sugars Date received: 26 June 2013; accepted: 23 December 2013
INTRODUCTION Netted melon (Cucumis melo var. reticulatus Naud.) is characterized by having fine, uniform net, a round shape and green pulp. It is famous for its special flavor and pleasant aroma. Netted melon is a climacteric fruit that produces moderate-to-high amount of ethylene. Melons can be subject to water loss and softening, and even rot after harvest, which can cause considerable economic loss. Postharvest rot of melon is mainly caused by physiology senescence, mechanical injury and pathogens. Cold storage is used in marketing of fruits and vegetables although its use alone does not maintain qualities of some products for a time long enough to cover marketing distances (Alves et al., 2005). On the other hand, in some areas, especially developing countries, cooling facilities are not always available, which restricts the application of cold Food Science and Technology International 0(0) 1–13 ! The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav DOI: 10.1177/1082013214520786 fst.sagepub.com
storage. Therefore, storage of netted melon is typically limited to 2–4 days at room temperature. The main problem in extending melon postharvest life is the high rate of respiration and senescence. The short postharvest life of netted melon is usually considered to be associated with the high ethylene production rate and carbon dioxide production during storage. An alternative method to extend the shelf life of fruit products is to treat them with 1-methylcyclopropene (1-MCP). 1-MCP has been reported to be an antagonist of ethylene action that blocks physiological action of ethylene (Khan and Singh, 2009). 1-MCP is a gas which competes with ethylene for binding the hormone cell membrane receptor (Sisler and Serek, 1997). 1-MCP has some advantages compared with the classic 1
Department of Horticulture, Zhejiang University, Hangzhou, Zhejiang, China 2 School of Vegetable Research, Wenzhou Academy of Agricultural Science, Wenzhou, Zhejiang, China Corresponding author: BL Wang, Department of Horticulture, Zhejiang University, Hangzhou, Zhejiang 310058, China. Email: [email protected]
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Food Science and Technology International 0(0) ethylene inhibitors: it has got a non-toxic mode of action, it leaves a negligible residue; it is active at very low concentrations and finally it is non-toxic for humans and the environment (Luo et al., 2007). Exposure of 1MCP inhibited ethylene action, and thus improved the fruit quality in climacteric fruits such as apples, avocados, bananas, stonefruits, mangos and tomatoes (Blankenship and Dole, 2003), as well as in plums (Khan and Singh, 2009), persimmons (Harima et al., 2003), broccolis (Yuan et al., 2010) and loquats (Cai et al., 2006). But 1-MCP concentration required to inhibit ethylene action varies greatly with stage of maturity and crop species (Harima et al., 2003). 1-MCP also reduced chilling injuries in persimmons (Salvador et al., 2004), and retarded enzyme activities involved in browning and oxidation (Wang et al, 2009). Active concentrations of 1-MCP varied widely among fruits of different species, even different varieties within. Concentration and time of 1-MCP exposure play an important role in fruit ripening, as reported in tomato (Guille´n et al., 2007), persimmon (Harima et al., 2003), avocado (Jeong et al., 2002) and melon (Gal et al., 2006). 1-MCP was effective in postharvest storage of melon (de Melo et al., 2008; Gal et al., 2006; Li et al., 2011b) although inconsistent results were obtained on the most effective concentration of 1-MCP in these reports. As for commercial usage, it is important to establish the optimum 1-MCP dose to achieve the maximum efficiency. Several research were reported about the effect of 1MCP application on melons. In ‘Galia’ melon (Ergun et al., 2005; Gal et al., 2006) and ‘Hami’ melon (Bi et al., 2003; Li et al., 2011a, 2011b), it is found that 1-MCP significantly extended melon shelf life, improved external qualities and nutritional qualities of melon fruits. And 1-MCP is also proved to be effective in improving fruit qualities of cantaloupe (Amaro et al., 2013) and ‘Yujinxiang’ melon (Ma et al., 2012). Among the research on netted melon, much research exists on the application effect of 1-MCP, whereas, there are not any published papers on the cultivar of netted melon in this research. The aim of this paper was to determine the most appropriate concentration of 1-MCP to extend shelf life of netted melon, and the experimental results demonstrated that the most suitable 1-MCP concentration is 0.6 ml l1. Then we studied the effects of 0.6 ml l1 1-MCP treatment of netted melons on the changes in visual quality, physiological responses and nutritional quality during postharvest storage at both 25 and 10 C.
trays filled with commercial container medium ‘Farfard’. Then one-month-old seedlings were transplanted into the greenhouse. Netted melon was cultivated from March to June in 2012. All plants were vertically trained to a single main vine and fruits were allowed to set on similar node positions to guarantee their uniformities in fruit size and similar growing conditions. Female flowers were hand-pollinated and tagged at anthesis, and one fruit per plant was allowed to set. Fruits were harvested at 45 days after pollination (DAP). Melon fruits at commercial harvest maturity of uniform size, free from visual symptoms of any disease or mechanical injury were harvested. The bulked fruits were transported to the laboratory immediately after harvest. The fruits were randomly selected for different treatments. Then the fruits were arranged randomly into different treatment groups. Methods 1-MCP and storage treatments. Melon fruits were treated with air (control), 0.1, 0.2, 0.4, 0.6, 0.8 and 1 ml l1 1-MCP in different 240 l air-tight plastic bag at 25 C for 24 h in the dark. After treatment, all the experimental fruits were stored either at ambient temperature of 25 C or at 10 C where relative humidity (RH) was maintained at 60%. All experimental groups were stored according to randomized complete block design. The shelf life was then determined. The concentration of 0.6 ml l1 1-MCP was selected for further study since shelf life of fruits were maximally extended under this treatment of concentration. First and foremost, intact melon fruits were used for analysis of weight loss, ethylene production and respiration rate. Then, firmness and total soluble solid contents were measured after fruits were cut into two halves. Finally, some of the edible pulp tissue of melons was cut into pieces and stored in 80 C for analysis of 1-aminocyclopropane-1-carboxylic acid (ACC) content, ACC synthase (ACS) activity, ACC oxidase (ACO) activity, titratable acidity and sugar components. Shelf life. According to Bi et al. (2003), shelf life of melon fruits was determined according to the levels of lesion appearance on the peel. Melons with more than two lesions (diameter 2 cm) were considered to have reached the end of their shelf life. Fruits were observed daily for appearance of lesions.
MATERIAL AND METHODS Materials Seeds of netted melon (Cucumis melo var. reticulatus Naud. cv. Zheyong No. 2) were sown in seedling raising
Ethylene production and respiration rate. Ethylene production and respiration rate were assayed every day after treatment. Five melon fruits from each treatment were used for these measurements. Each fruit was
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Shi et al. sealed in a 14 l plastic bucket for 1 h. Gas samples were collected with a syringe for measuring ethylene concentration using a gas chromatograph model SP6800 (Lunan Chemical Engineering Instrument Co. Ltd, Shandong, China) equipped with a flame ionization detector fitted with a GDX-502 column held at 90 C, and detection temperature of 140 C. External standard ethylene in a given concentration was adopted in quantification of ethylene in this experiment. According to principles of chromatography, gas concentration ratio of standard and tested sample ethylene matches peak area of standard ethylene and tested sample ethylene and gas concentration of sample ethylene will be calculated. Ethylene production was expressed as ml kg1 h1. The CO2 concentration was measured using an ICA 40 system (International Controlled Atmosphere Ltd., UK) and the results were expressed as mg kg1 h1 of CO2 evolved. The gas sample withdrawn from the same volume of container without melon fruits was taken as control. ACC content and activities of ethylene biosynthesis enzymes. ACC content was evaluated using five fruits per treatment every other day after treatment. ACC content was measured in fruit pulp tissue by following the method described by Lizada and Yang (1979) with minor modification. The fruit pulp tissue (5–6 g) was homogenized in a glass pestle and mortar using 300 mg white quartz sand with 10 ml distilled water at 2 1 C temperature. Following centrifugation at 10,000 g for 20 min at 4 C, 0.5 ml supernatant was mixed with 0.1 ml HgCl2 (50 mM) in two sets, one with and the other without 0.1 ml ACC (100 mM) and the final volume 1.8 ml was made with distilled water. The glass test tubes were sealed with a rubber septum and both test tubes were placed on ice, 0.2 ml (5% NaOCl þ saturated NaOH 2:1, v/v) solution was injected into the sealed tubes and vortexed for 5 s. After 25 min incubation on ice, the tubes were again vortexed and 1 ml gas sample was taken for ethylene measurements, expressed as pmol g1 fresh weight (FW). After the ethylene concentrations of the gas samples generated from both the test samples and the ACC solutions were determined, the proportions of ACC solutions that were transformed into ethylene in the reactions were calculated. Then the ACC concentrations in the test samples were calculated since it is assumed that there are the same transformation proportions for both the test samples and the ACC solutions. 1-Aminocyclopropane-1-carboxylic acid synthase (ACS) activity was measured using five fruits per treatment every other day after treatment. ACS activity was measured in pulp tissue by following the method of Mathooko et al. (1993) with some modification. Pulp tissue (10 g) was homogenized with 5 ml K-phosphate
buffer (0.5 M, pH 8.5) containing 5 mM pyridoxal phosphate and 5 mM dithiothreitol in the presence of 5% polyvinylpyrrolidone (PVP) in a glass pestle and mortar using 400 mg white quartz sand (50 þ 70 mesh, Sigma-Aldrich, Australia) at 2 1 C. Contents were centrifuged at 14,000 g for 30 min at 4 C and 2 ml of supernatant was mixed with 1 ml of 500 mM S-adenosyl methionine (SAM) in a glass reaction tube. The reaction tube was sealed with a rubber septum, incubated for 30 min at 30 C, and transferred to an ice bath. Using a syringe through the stopper, 0.1 ml of HgCl2 (50 mM) and 0.3 ml of NaOCl (5%) and saturated NaOH (2:1, v/v) were added into the reaction tube. The reaction tube was incubated in ice for a further 2.5 min and a 1 ml gas sample was taken from the head space and injected into the gas chromatograph for ethylene estimation, as described earlier. ACS activity was expressed as nmol ACC g1 FW h1. ACO activity was measured using five fruits per treatment every other day after treatment. ACO activity was measured in pulp tissue by following the method of Mathooko et al. (1993) with some modifications. Pulp tissue (2 g) was homogenized in a glass pestle and mortar using 200 mg white quartz sand in 5 ml extraction buffer consisting of 0.1 M Tris-HCl (pH 7.2), 10% (w/v) glycerol and 30 mM sodium ascorbate in the presence of 5% PVP. The homogenized mixture was centrifuged at 14,000 g for 30 min. All the steps in the enzyme extraction were carried out at 2 1 C. The enzyme activity was measured in 2 ml reaction mixture containing 1.8 ml of the supernatant solution, 0.1 ml ACC (40 mM) and 0.1 ml FeSO4 (1 mM). The reaction tube was sealed with a rubber septum and incubated at 30 C for 60 min. One milliliter of gas taken from the headspace was injected into the gas chromatograph for ethylene estimation, as described earlier. ACO activity was expressed as nmol C2H4 g1 FW h1. Weight loss and firmness. Five melon fruits of each treatment were weighted every other day to determine the weight loss. Weight loss (%) was calculated by equation (1).
Weight lossð%Þ ¼ ðWx W0 Þ=W0 100
ð1Þ
where W0 was the weight at 0 day and Wx was the weight at a certain day after storage. After melons were lengthways cut into two halves, firmness measurements were made on both of the equator and polar of the pulp tissue 1 cm from the seed cavities on the cut surfaces. Firmness was measured via five fruits per treatment every other day and was expressed as Newton (N). The assay was made using a TA-XT2i texture analyzer (Stable Micro Systems, England) 3
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Food Science and Technology International 0(0) with a probe of 5 mm in diameter, a penetration depth of 5 mm and a rate of penetration 1 mm/s.
RESULTS AND DISCUSSION
Total soluble solid (TSS) contents and titratable acidity (TA). TSSs were determined on the juice obtained from melon edible pulp tissue using a digital hand-held refractometer (Atago PAL-1, Tokyo, Japan). TSS was expressed as Brix. TA was measured by following a previous method (Guille´n et al., 2007) with modifications. The juice was also obtained from fruit edible pulp tissue. TA was determined by potentiometric titration with 0.1 N NaOH up to pH 8.1, using 1 ml of diluted juice in 25 ml distilled H2O and results are expressed as gram of citric acid equivalent per 100 g1 FW. Five fruits from each treatment were used for analysis of these parameters.
In the present study, shelf life of netted melon under different concentrations of 1-MCP was monitored to determine the most effective 1-MCP dose. Netted melons treated with 0.6 ml l1 1-MCP exhibited the longest shelf life, approximately seven (25 C) or nine (10 C) days (Figure 1), which is significantly longer than that of control or other treatments, including those treated with 0.8 or 1 ml l1 1-MCP. When stored under room temperature (25 C), the shelf life of melon fruits was prolonged for 4 days under 0.6 ml l1 1-MCP treatment when compared with the control. And the shelf life of melons stored under 10 C was also 4 days longer than that of the control. Therefore, under both temperatures, 25 C and 10 C, 0.6 ml l1 was the most effective dose of 1-MCP to extend fruit shelf life. The results are consistent with a previous report (de Melo et al., 2008), in which postharvest application of 1-MCP extended shelf life of orange-flesh melons from 15 days to 21 days under room temperature storage. In this experiment, as it is shown from fruit appearance, higher concentration of 1-MCP (0.8 and 1 ml l1) did not produce better effect. It is probably because 1-MCP reached its saturating concentration after 0.6 ml l1. When 1-MCP permanently binds to a sufficient number of ethylene receptors, higher doses of 1-MCP will not maintain longer fruit shelf life.
Sugar components. Sugars were determined using five fruits from each treatment every other day as previously described (Komatsu et al., 1999; Lowell et al. 1989). Samples of mesocarp tissues (ca. 1 g FW) were homogenized for 5 min in 5 volumes of extracted solution (ethanol:chloroform:water ¼ 12:5:3, l/l). Water and chloroform were then added to bring the final E:C:W ratio to 10:6:5. Subsequent separation of the chloroform layer allowed removal of lipids and pigments. The remaining aqueous-alcohol phase was evaporated to dryness in a vacuum at 50 C and re-dissolved in 1 ml distilled water. Analysis of soluble sugars was performed by high performance liquid chromatography (Shimadzu Co., Kyoto, Japan) with an NH2 column at 30 C using 65% acetonitrile solution (1 ml min1) as a mobile phase and a refractive index detector (Shimadzu RID-10 A). Peaks were quantified by using corresponding analytical software. As to the quantification of sugar content, the procedures are divided into three steps in the following. Firstly, several sorts of dilution of sugar solution (glucose, fructose and sucrose) were made by dissolving the required anount of authentic standard in deionized water (dH2O). Secondly, three standard sugar solutions will be analyzed in the ideal chromatography. Finally, the concentration of sucrose, glucose and fructose were quantified by comparison between standard sugar solution and sample solution. Statistical analysis. Statistical analysis was performed using the SPSS package program version 11.5 (SPSS Inc. Chicago, IL). Data were analyzed using one-way analysis of variance (ANOVA) model. The variants were storage time and different treatments. The means were compared using the least significant differences (LSDs) test at a significant level of 0.05. The values were reported as means with standard deviations.
Shelf life
Ethylene production and respiration rate Ethylene plays an important role in fruit ripening and senescence. Many ripening processes, including fruit softening, weight loss, peel degreening, aroma generating and change of sugar content, are highly dependent on ethylene action. Netted melon exhibited typical climacteric changes during storage. Ethylene accumulation is a necessity as to the full accomplishment of some ripening procedure which is closely related to fruit quality in climacteric fruits (e.g. cell wall degradation reflected in fruit softening). The problem emerges when this ethylene production is excessive or too prolonged in time; then detrimental effects appear such as fruit rot, mainly because of extreme softening or the fruit that breeds cracking of it and production of smelly scents rather than agreeable aroma. There are many tools to inhibit onset of autocatalytic ethylene production among which, the effectiveness of 1-MCP in extending storage time and improving quality in many kinds of fruit has been widely proven (Blankenship and Dole, 2003). For instance, postharvest application of 1-MCP after onset of ripening completely halted subsequently softening in Charentais melon (Nishiyama et al., 2007) and in ‘Galia’ melon,
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onset of ethylene climacteric was delayed by 10 days compared with the control (Gal et al., 2006). It is reported that ethylene production in 1-MCPtreated melons was significantly lower than that in control during the first 6 days of storage (Li et al., 2011a). In addition, 1-MCP treatment delayed onset of ethylene climacteric and subsequently retards other ripening processes in persimmon (Harima et al., 2003) and tomato (Guille´n et al., 2007). This is consistent with the present work. In this study, ethylene production exhibited typical climacteric changes under both ambient (25 C) and low temperature (10 C) storage conditions. Ethylene peaks in 1-MCP-treated fruits were delayed for 3 days (25 C) or 2 days (10 C) as compared to control (Figure 2). The maximum ethylene production of 1-MCP-treated fruits was reduced by 35.2% and 35.5%, respectively, under 25 C and 10 C when compared with the control. 1-MCP occupied ethylene receptors so that ethylene cannot bind and elicit its action (Blankenship and Dole, 2003). There is an optimum concentration of 1-MCP for each crop. The effectiveness of the low concentration of 1-MCP in the present study may be due to high accessibility and affinity of 1-MCP to the receptors. Although binding of 1-MCP to ethylene receptor is irreversible, plant recover their ethylene sensitivity due to turnover or production of new receptors (Sisler and Serek, 1997). It is observed from Figure 2 that in the middle period of storage (day 4), new receptors probably are being synthesized since the autocatalytic ethylene production quickly increased, and then the process of fruit ripening continued. In this research, the trends of respiration rates in terms of 1-MCP treated and untreated fruits distinguished a lot. Between the two figures, respiration
rates of untreated fruits rose dramatically to the peak at first, and then delined during the storage period. Whereas, both of respiration rates of 1-MCP-treated fruits maintained a steady growth. Meanwhile, it is worth to notice that, under different storage temperatures, respiration rates in terms of 1-MCP treated and untreated fruits have a clear difference. Particularly, for one, respiration rate of 1-MCP-treated fruits in 25 C was far lower than that of untreated fruits in 2–7 days; for another, respiration rate of 1-MCP-treated fruits in 10 C was under that of untreated fruits in 4–7 days. In ‘Galia’ melon (Gal et al., 2006), onset of ethylene and respiration climacterics are dramatically delayed and both of the peak values are reduced, which are consistent with results of our present work. However, 1-MCP treatment had no effect on apricot (Dong et al., 2002). The distinct results in apricots might be due to fruit species, maturity, cultivar or some other unknown factor (Blankenship and Dole, 2003). Respiration rate of produce is a useful guide to assess the potential storage life of produce. Respiration is energy-consuming and causes loss of sugars and of weight, which are both important causes of fruit quality deterioration. Therefore, 1-MCP-treated fruits with lower respiration rate have a higher capacity of storage than the nontransformed control fruits as the present study shows. Storage temperature In this research, netted melons were stored under both 10 C and 25 C to examine the effect of temperature on efficacy of 1-MCP. Our results showed that ethylene production and respiration rate of melons stored in 10 C were greatly lower than that in room temperature (25 C) (Figure 2), indicating 10 C storage obviously 5
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delayed the fruit senescence, postponed the ethylene peaks and reduced the respiration rate. In the experiment of kiwifruit, the extent of ethylene inhibition caused by 1-MCP is related to temperature, namely, low temperature storage resulted in better 1-MCP effects (Kim et al., 2001), which is consistent with our results. However, applying lower temperature to attain a better postharvest storage is not completely true, since it has been reported by Miccolis and Saltveit (1995) that, ‘for certain melon varieties, chilling injury induced symptoms were much serious in fruits when stored in 7 C for 1–2 weeks than when stored under 15 C’. On the other hand, low temperature storage is an effective method to preserve melons, but cold storage facilities are not always available. Therefore, postharvest 1-MCP application can be very helpful in extending fruit shelf life and maintaining fruit postharvest qualities, especially where low temperature chain transportation is unavailable.
ACC, ACS and ACO Ethylene biosynthesis is initiated from the amino acid, methionine, via S-adenosylmethionine (Adomet) through the intermediate 1-aminocyclopropane-1carboxylic acid (ACC) that the biosynthesis of this intermediate is catalyzed by the enzyme ACC synthase (ACS). Next, the enzyme ACO catalyzes the conversion of ACC to ethylene (Ezura and Owino, 2008). The biosynthesis of ethylene may also be affected by 1-MCP due to its competitive binding to ethylene receptors. Our results showed that 1-MCP treatment significantly inhibited ACC accumulation during the storage under both 25 C and 10 C, although lower ACC content was found out under storage at 10 C when compared with that at 25 C. According to Khan and Singh (2009), ACC content of 1-MCP-treated plum were suppressed. Consistent results were also found out in mango, in which the ACC increase was significantly delayed by
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Figure 3. Effect of 1-MCP treatment on ACC content, ACS activities and ACO activities in pulp of netted melon fruits during storage at both 25 C (a, c, e) and 10 C (b, d, f). Each value is the mean of five fruits. Vertical bars represent standard deviation of the mean.
1-MCP (Wang et al., 2009). The reduction of ACC content in 1-MCP-treated melon may be resulted from reduced ACS and ACO activity in the fruit. In this study, similar patterns of ACC contents and ACS activity were detected in both treated and untreated fruits. As it is shown in Figure 3, distinct decline of
ACS activity was found in 1-MCP treated melons. At the end of storage, ACS activity in 1-MCP treated melons was 27.2% (25 C) and 41.7% (10 C) lower than that of control, correspondingly. We also found that during storage, ACO activity in melons increased and then declined, which were consistent with the 7
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pattern of ethylene production. Moreover, ACO activity in 1-MCP treated melons were constantly lower than that of control, and the peaks were 55.6% (25 C) and 42.3% (10 C) lower than that of control correspondingly during storage. Therefore, it is concluded that the decline of ACC content by 1-MCP may be resulted from the decreased ACS and ACO activity. In ‘Hami’ melon, 1-MCP also markedly inhibited the accumulation of ACC and maintained lower activities of ACS and ACO (Li et al., 2011a). Meanwhile in plums, a reduction in ACS and ACO activity and ACC content in 1-MCP-treated fruits during storage at ambient or cold temperature was also reported (Khan and Singh, 2009). Therefore, reduced climacteric ethylene production in 1-MCPtreated fruit, may be ascribed to the reduced activity of ethylene biosynthesis enzymes ACS and ACO. Several ACS and ACO isoenzymes have been identified in melon tissues and both enzymes are encoded by multigene family (Lasserre et al., 1996; Miki et al., 1995; Yamamoto et al., 1995). In other species, the
adjustment of the expression of the main ethylene biosynthetic genes, ACS and ACO, has also been mentioned (Ezura and Owino, 2008). It should be noted that the different patterns in ACS and ACO activity during melon storage may be due to the different levels of ACS and ACO transcriptions, which was reported by Pathak et al. (2003). Consequently, it can be concluded that the suppression of ethylene perception by 1-MCP in melons is mainly attributed to its feedback suppression of ACS and ACO activity. Fruit weight and firmness Fruit weight and firmness are two important attributes that affect melon visual quality and are also closely related to fruit shelf life. In this study, weight loss of both 1-MCP-treated fruits and the control fruits increased during the storage period. In ‘Galia’ melon (Gal et al., 2006), 1-MCP treatment caused a reduction of weight loss. Alves et al. (2005) also found that in ‘Charentais’ melon, control fruit lost 7% of weight
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Figure 5. Effect of 1-MCP treatment on total soluble solid (TSS) content and titratable acidity (TA) in pulp of netted melon fruits during storage at both 25 C (a, c) and 10 C (b, d). Each value is the mean of five fruits. Vertical bars represent standard deviation of the mean.
while 1-MCP-treated fruit lost only 5% of weight at ambient temperature. However, contradictory results were also found in fruits of other species. In apricot (Fan et al., 2000) and orange (Porat et al., 1999), no effect was observed after 1-MCP treatment, while in avocado (Jeong et al., 2002) and Chinese kale (Sun et al., 2012), weight loss in the fruits were delayed by 1-MCP treatment. In addition, 1-MCP application significantly reduced the weight loss in both storage conditions. The main cause of weight loss is consumption of respiration substrates, mainly sugars, and this was confirmed by the similarity in changes of weight loss and respiration rate in present study. Softening is one of the most sensitive ripening processes to ethylene. Ethylene plays a major role in regulating softening of climacteric fruit, including Charentais melon (Ayub et al., 1996; Hadfield et al., 2000). In ‘Galia’-type melon (Ergun et al., 2005; Gal et al., 2006), 1-MCP maintained fruit firmness when compared to the control. In ‘Hami’ melon, decrease in firmness was significantly inhibited by 1-MCP (Li et al., 2011a). In this study, the decrease of pulp firmness was obviously reduced in 1-MCP treated melons
when contrasted with the control fruits during the 8 days of storage (Figure 4). At the end of the experiment, firmness values of 1-MCP treated melon were 49.6% and 26.4% higher, respectively, under 25 C and 10 C when compared with the control. Fruit firmness was reduced in both treated and untreated fruits when stored at the lower temperature (10 C). While in fruits of other species, firmness was better maintained by 1-MCP in apricots (Fan et al., 2000), peaches (Kluge and Jacomino, 2002) and plums (Dong et al., 2002, 2001) as well. TSS and TA In netted melon, accumulation of sugars (especially sucrose) is one of the essential factors that determine fruit quality and commodity values (Lingle and Dunlap, 1987). In ‘Galia’ melon, 1-MCP did not affect TSS content after prolonged storage and shelflife simulation. However, in the present study, less reduction of TSS content was monitored in 1-MCPtreated fruits than the control fruits. 1-MCP treatment effectively inhibited the reduction of TSS contents of 9
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melon fruits (Figure 5). After 8 days of storage, TSS of fruits treated with 1-MCP were reduced by approximately 14.19% (25 C) and 10.72% (10 C), respectively, as compared to 22.71% and 13.03% in untreated fruits. Consistent results were observed in 1-MCP treated ‘Hami’ melon (Li et al., 2011b), TSS of 1-MCP-treated fruits is higher than that of the control. In fresh-cut ‘Hami’ melon (Guo et al., 2011) and fresh-cut cantaloupe (Amaro et al., 2013), TSS of the fruit are also better maintained better after
1-MCP treatment. Since respiration was lower in 1-MCP-treated fruit, less sugar is consumed when comparing with control. Therefore, it is possible that it is not 1-MCP that influences sugar content, but the concomitant low respiration rate. As it has been mentioned, during 2–7 days storage and 4–7 days storage of 25 C and 10 C respectively, respiration rate of 1-MCP-treated fruits was obviously lower that of control fruits, which tells TSS content of 1-MCP-treated fruits in 2–8 days storage was evidently less than that of
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Shi et al. control fruits. The reduction of TSS loss by 1-MCP treatment was higher in melons stored at 10 C than those at 25 C. However, Cohen and Hicks (1986) found that storage temperature did not influence soluble sugar content after 2, 5 or 9 days at 5, 12.5 and 20 C. Moreover, the effect of 1-MCP on soluble sugars also depends on the crop (Blankenship and Dole, 2003). There are different 1-MCP effects in TSS in other species. Soluble solids were higher in 1-MCP-treated pineapple (Selvarajah et al., 2001), papaya (Hofman et al., 2001) and kiwifruit (Boquete et al., 2004). However, soluble solids were reduced in 1-MCP-treated strawberries (Tian et al., 2000). In other commodities, such as persimmons, apricots and plums, TSSs were not affected by 1-MCP (Dong et al, 2002; Salvador et al., 2004; Watkins et al., 2000). As it can be observed in Figure 5, 1-MCP treatment significantly inhibited TA reduction in netted melon fruits. In addition, reduction of TA in fruit stored in 25 C was higher than those stored in 10 C during storage period. In previous reports, 1-MCP prevented acidity loss in tomatoes (Wills and Ku, 2002), delayed TA loss in plums (Dong et al., 2002) and loquats (Cai et al., 2006). In contrast, 1-MCP did not affect TA in apricots (Dong et al., 2002) or apples (Mir et al., 2001) during storage at different temperatures. Sucrose, glucose and fructose In this study, patterns of sugar metabolism in the melon fruit were greatly influenced by 1-MCP during storage. In netted melon, sugars that contribute to the TSSs were mainly sucrose, fructose and glucose. Changes in the contents of these sugar components were shown in Figure 6. Sucrose was the predominant sugar in melon pulp tissue and in both treated and untreated fruits, sucrose content exhibited constant decrease during postharvest storage. However, 1-MCP treatment reduced the sucrose content up to a certain degree. At the end of storage, sucrose content in 1-MCP treated melons were 5.27% and 2.15% higher, respectively, under 25 C and 10 C than that of control. Moreover, higher sucrose content was observed in fruits stored at 10 C when compared with those stored at 25 C. Fructose and glucose are two important sugar components in melon since they make up the components of sucrose, the main sugar found in melon fruit. It can be observed in Figure 6 that changing curves of both fructose and glucose were similar to that of sucrose, and that 1-MCP also effectively reduced the diminution of the contents of these sugars. This is consistent with the results of a report on ‘Yujinxiang’ melon (Ma et al., 2012). However, under 10 C storage, as to different sugar components, 1-MCP treatment showed significant effects from
different time: day 4 for sucrose, day 6 for fructose and day 2 for glucose.
CONCLUSIONS A volume of 0.6 ml l1 1-MCP significantly extended melon shelf life, reduced ethylene production and respiration rate, and retarded the changes in parameters related to fruit ripening, including fruit weight loss, firmness, TSS content, acidity and the main sugar contents. In conclusion, 1-MCP treatment is a good method to extend melon shelf life and maintain fruit quality, no matter fruits were stored under room temperature (25 C) or lower temperature (10 C), and the combination of 1-MCP and low temperature storage resulted in more acceptable fruit quality. ACKNOWLEDGMENTS We are thankful to the 985-Institute of Agrobiology and Environmental Sciences of Zhejiang University for providing access to analytical instruments.
CONFLICT OF INTEREST None declared.
FUNDING This work was funded by Major Scientific and Technological Key Program of Ningbo (2010C10012), Major special breeding program of Science and Technology Department of Zhejiang Province (2012C12903-2-7) and National Science and Technology Support Program of China (2013BAD201300).
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avocado, custard apple, mango and papaya fruit to 1methylcyclopropene. Australian Journal of Experimental Agriculture 41(4): 567–572. Jeong J, Huber DJ and Sargent SA. (2002). Influence of 1methylcyclopropene (1-MCP) on ripening and cell-wall matrix polysaccharides of avocado (Persea americana) fruit. Postharvest Biology and Technology 25(3): 241–256. Khan AS and Singh Z. (2009). 1-MCP application suppresses ethylene biosynthesis and retards fruit softening during cold storage of ‘Tegan Blue’ Japanese plum. Plant Science 176(4): 539–544. Kim HO, Hewett EW and Lallu N. (2001). Softening and ethylene production of kiwifruit reduced with 1-methylcyclopropene. Acta Horticulturae 553: 167–170. Kluge RA and Jacomino AP. (2002). Shelf life of peaches treated with 1-methylcyclopropene. Scientia Agricola 59(1): 69–72. Komatsu A, Takanokura Y and Moriguchi T. (1999). Differential expression of three sucrose-phosphate synthase isoforms during accumulation in citrus fruit (Citrus unshiu Marc). Plant Science 140(2): 169–178. Lasserre E, Bouquin T, Hernandez JA, Bull J, Pech JC and Balague C. (1996). Structure and expression of three genes encoding ACC oxidase homologs from melon (Cucumis melo L.). Molecular and General Genetics 251(1): 81–90. Lingle SE and Dunlap JR. (1987). Sucrose metabolism in netted muskmelon fruit during development. Plant Physiology 84(2): 386–389. Li XW, Cao SF, Zheng YH and Sun AP. (2011a). 1-MCP suppresses ethylene biosynthesis and delays softening of ‘Hami’ melon during storage at ambient temperature. Journal of the Science of Food and Agriculture 91(14): 2684–2688. Li XW, Jin P, Wang J, Zhu X, Yang HY and Zheng YH. (2011b). 1-Methylcyclopropene delays postharvest ripening and reduces decay in Hami melon. Journal of Food Quality 34(2): 119–125. Lizada MC and Yang SF. (1979). A simple and sensitive assay for 1-aminocyclopropane-1-carbroxylic acid. Analytical Biochemistry 100(1): 140–145. Lowell CA, Tomlinson PT and Koch KE. (1989). Sucrose metabolizing enzymes in transport tissue and adjacent sink structures in developing citrus fruit. Plant Physiology 90(4): 1394–1402. Luo ZS, Xu XL, Cai ZZ and Yan M. (2007). Effects of ethylene and 1-methylcyclopropene (1-MCP) on lignification of postharvest bamboo shoot. Food Chemistry 105(2): 521–527. Mathooko FM, Kubo Y, Inaba A and Nakamura R. (1993). Regulation by carbon dioxide of wound-induced ethylene biosynthesis in tomato pericarp and winter squash mesocarp tissues. Postharvest Biology and Technology 3(1): 27–38. Ma WP, Cao JK, Ni ZJ, Tian WN, Zhao YM and Jiang WB. (2012). Effects of 1-methylcyclopropene on storage quality and antioxidant activity of harvested ‘‘Yujinxiang’’ melon (Cucumis Melo L.) fruit. Journal of Food Biochemistry 36(4): 413–420. Miccolis V and Saltveit ME. (1995). Influence of storage period and temperature on the postharvest characteristics
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Sisler EC and Serek M. (1997). Inhibitors of ethylene responses in plants at the receptor level: recent developments. Physiologia Plantarum 100(3): 577–582. Sun B, Yan H, Liu N, Wei J and Wang Q. (2012). Effect of 1-MCP treatment on postharvest quality characters, antioxidants and glucosinolates of Chinese kale. Food Chemistry 131(2): 519–526. Tian MS, Prakash S, Elgar HJ, Young H, Burmeister DM and Ross GS. (2000). Responses of strawberry fruit to 1-methylcyclopropene (1-MCP) and ethylene. Plant Growth Regulation 32(1): 83–90. Wang BG, Wang JH, Feng XY, Lin L, Zhao YM and Jiang WB. (2009). Effects of 1-MCP and exogenous ethylene on fruit ripening and antioxidants in stored mango. Plant Growth Regulation 57(2): 185–192. Watkins CB, Nock JF and Whitaker BD. (2000). Response of early, mid and late season apple cultivars to postharvest application of 1-methylcyclopropene (1-MCP) under air and controlled atmosphere storage conditions. Postharvest Biology and Technology 19(1): 17–32. Wills RBH and Ku VVV. (2002). Use of 1-MCP to extend the time to ripen of green tomatoes and postharvest life of ripe tomatoes. Postharvest Biology and Technology 26(1): 85–90. Yamamoto M, Miki T, Ishiki Y, Fujinami K, Yanagisawa Y, Nakagawa H, Ogura N, Hirabayashi T and Sato T. (1995). The synthesis of ethylene in melon fruit during the early-stage of ripening. Plant and Cell Physiology 36(4): 591–596. Yuan G, Sun B, Yuan J and Wang Q. (2010). Effect of 1methylcyclopropene on shelf life, visual quality, antioxidant enzymes and health-promoting compounds in broccoli florets. Food Chemistry 118(3): 774–781.
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Effect of 1-methylcyclopropene on shelf life, visual quality and nutritional quality of netted melon Y Shi, BL Wang, DJ Shui, LL Cao, C Wang, T Yang, XY Wang and HX Ye Food Science and Technology International published online 4 February 2014 DOI: 10.1177/1082013214520786 The online version of this article can be found at: http://fst.sagepub.com/content/early/2014/02/04/1082013214520786
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Original Article
Effect of 1-methylcyclopropene on shelf life, visual quality and nutritional quality of netted melon Y Shi1, BL Wang1, DJ Shui2, LL Cao1, C Wang1, T Yang1, XY Wang2 and HX Ye1
Abstract The effects of 1-methylcyclopropene (1-MCP) on shelf life, fruit visual quality and nutritional quality were investigated. Netted melons were treated with air (control) and 0.6 ml l1 1-MCP at 25 C for 24 h, and then stored at 25 C or 10 C for 10 days. 1-MCP significantly extended the shelf life, inhibited weight loss and delayed firmness decline of melon fruits. Ethylene production was also inhibited and respiration rate was declined. 1-MCP retarded 1-aminocyclopropane-1-carboxylic acid (ACC) increases and inhibited ACC synthase and ACC oxidase activity. Moreover, 1-MCP treatment reduced the decrease in total soluble solids and titratable acidity, as well as the decrease of the content of sugars (sucrose, fructose and glucose). These results indicated that 1-MCP treatment is a good method to extend melon shelf life and maintain fruit quality, and the combination of 1-MCP and low temperature storage resulted in more acceptable fruit quality.
Keywords Netted melon, 1-methylcyclopropene (1-MCP), shelf life, ethylene, firmness, sugars Date received: 26 June 2013; accepted: 23 December 2013
INTRODUCTION Netted melon (Cucumis melo var. reticulatus Naud.) is characterized by having fine, uniform net, a round shape and green pulp. It is famous for its special flavor and pleasant aroma. Netted melon is a climacteric fruit that produces moderate-to-high amount of ethylene. Melons can be subject to water loss and softening, and even rot after harvest, which can cause considerable economic loss. Postharvest rot of melon is mainly caused by physiology senescence, mechanical injury and pathogens. Cold storage is used in marketing of fruits and vegetables although its use alone does not maintain qualities of some products for a time long enough to cover marketing distances (Alves et al., 2005). On the other hand, in some areas, especially developing countries, cooling facilities are not always available, which restricts the application of cold Food Science and Technology International 0(0) 1–13 ! The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav DOI: 10.1177/1082013214520786 fst.sagepub.com
storage. Therefore, storage of netted melon is typically limited to 2–4 days at room temperature. The main problem in extending melon postharvest life is the high rate of respiration and senescence. The short postharvest life of netted melon is usually considered to be associated with the high ethylene production rate and carbon dioxide production during storage. An alternative method to extend the shelf life of fruit products is to treat them with 1-methylcyclopropene (1-MCP). 1-MCP has been reported to be an antagonist of ethylene action that blocks physiological action of ethylene (Khan and Singh, 2009). 1-MCP is a gas which competes with ethylene for binding the hormone cell membrane receptor (Sisler and Serek, 1997). 1-MCP has some advantages compared with the classic 1
Department of Horticulture, Zhejiang University, Hangzhou, Zhejiang, China 2 School of Vegetable Research, Wenzhou Academy of Agricultural Science, Wenzhou, Zhejiang, China Corresponding author: BL Wang, Department of Horticulture, Zhejiang University, Hangzhou, Zhejiang 310058, China. Email: [email protected]
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Food Science and Technology International 0(0) ethylene inhibitors: it has got a non-toxic mode of action, it leaves a negligible residue; it is active at very low concentrations and finally it is non-toxic for humans and the environment (Luo et al., 2007). Exposure of 1MCP inhibited ethylene action, and thus improved the fruit quality in climacteric fruits such as apples, avocados, bananas, stonefruits, mangos and tomatoes (Blankenship and Dole, 2003), as well as in plums (Khan and Singh, 2009), persimmons (Harima et al., 2003), broccolis (Yuan et al., 2010) and loquats (Cai et al., 2006). But 1-MCP concentration required to inhibit ethylene action varies greatly with stage of maturity and crop species (Harima et al., 2003). 1-MCP also reduced chilling injuries in persimmons (Salvador et al., 2004), and retarded enzyme activities involved in browning and oxidation (Wang et al, 2009). Active concentrations of 1-MCP varied widely among fruits of different species, even different varieties within. Concentration and time of 1-MCP exposure play an important role in fruit ripening, as reported in tomato (Guille´n et al., 2007), persimmon (Harima et al., 2003), avocado (Jeong et al., 2002) and melon (Gal et al., 2006). 1-MCP was effective in postharvest storage of melon (de Melo et al., 2008; Gal et al., 2006; Li et al., 2011b) although inconsistent results were obtained on the most effective concentration of 1-MCP in these reports. As for commercial usage, it is important to establish the optimum 1-MCP dose to achieve the maximum efficiency. Several research were reported about the effect of 1MCP application on melons. In ‘Galia’ melon (Ergun et al., 2005; Gal et al., 2006) and ‘Hami’ melon (Bi et al., 2003; Li et al., 2011a, 2011b), it is found that 1-MCP significantly extended melon shelf life, improved external qualities and nutritional qualities of melon fruits. And 1-MCP is also proved to be effective in improving fruit qualities of cantaloupe (Amaro et al., 2013) and ‘Yujinxiang’ melon (Ma et al., 2012). Among the research on netted melon, much research exists on the application effect of 1-MCP, whereas, there are not any published papers on the cultivar of netted melon in this research. The aim of this paper was to determine the most appropriate concentration of 1-MCP to extend shelf life of netted melon, and the experimental results demonstrated that the most suitable 1-MCP concentration is 0.6 ml l1. Then we studied the effects of 0.6 ml l1 1-MCP treatment of netted melons on the changes in visual quality, physiological responses and nutritional quality during postharvest storage at both 25 and 10 C.
trays filled with commercial container medium ‘Farfard’. Then one-month-old seedlings were transplanted into the greenhouse. Netted melon was cultivated from March to June in 2012. All plants were vertically trained to a single main vine and fruits were allowed to set on similar node positions to guarantee their uniformities in fruit size and similar growing conditions. Female flowers were hand-pollinated and tagged at anthesis, and one fruit per plant was allowed to set. Fruits were harvested at 45 days after pollination (DAP). Melon fruits at commercial harvest maturity of uniform size, free from visual symptoms of any disease or mechanical injury were harvested. The bulked fruits were transported to the laboratory immediately after harvest. The fruits were randomly selected for different treatments. Then the fruits were arranged randomly into different treatment groups. Methods 1-MCP and storage treatments. Melon fruits were treated with air (control), 0.1, 0.2, 0.4, 0.6, 0.8 and 1 ml l1 1-MCP in different 240 l air-tight plastic bag at 25 C for 24 h in the dark. After treatment, all the experimental fruits were stored either at ambient temperature of 25 C or at 10 C where relative humidity (RH) was maintained at 60%. All experimental groups were stored according to randomized complete block design. The shelf life was then determined. The concentration of 0.6 ml l1 1-MCP was selected for further study since shelf life of fruits were maximally extended under this treatment of concentration. First and foremost, intact melon fruits were used for analysis of weight loss, ethylene production and respiration rate. Then, firmness and total soluble solid contents were measured after fruits were cut into two halves. Finally, some of the edible pulp tissue of melons was cut into pieces and stored in 80 C for analysis of 1-aminocyclopropane-1-carboxylic acid (ACC) content, ACC synthase (ACS) activity, ACC oxidase (ACO) activity, titratable acidity and sugar components. Shelf life. According to Bi et al. (2003), shelf life of melon fruits was determined according to the levels of lesion appearance on the peel. Melons with more than two lesions (diameter 2 cm) were considered to have reached the end of their shelf life. Fruits were observed daily for appearance of lesions.
MATERIAL AND METHODS Materials Seeds of netted melon (Cucumis melo var. reticulatus Naud. cv. Zheyong No. 2) were sown in seedling raising
Ethylene production and respiration rate. Ethylene production and respiration rate were assayed every day after treatment. Five melon fruits from each treatment were used for these measurements. Each fruit was
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Shi et al. sealed in a 14 l plastic bucket for 1 h. Gas samples were collected with a syringe for measuring ethylene concentration using a gas chromatograph model SP6800 (Lunan Chemical Engineering Instrument Co. Ltd, Shandong, China) equipped with a flame ionization detector fitted with a GDX-502 column held at 90 C, and detection temperature of 140 C. External standard ethylene in a given concentration was adopted in quantification of ethylene in this experiment. According to principles of chromatography, gas concentration ratio of standard and tested sample ethylene matches peak area of standard ethylene and tested sample ethylene and gas concentration of sample ethylene will be calculated. Ethylene production was expressed as ml kg1 h1. The CO2 concentration was measured using an ICA 40 system (International Controlled Atmosphere Ltd., UK) and the results were expressed as mg kg1 h1 of CO2 evolved. The gas sample withdrawn from the same volume of container without melon fruits was taken as control. ACC content and activities of ethylene biosynthesis enzymes. ACC content was evaluated using five fruits per treatment every other day after treatment. ACC content was measured in fruit pulp tissue by following the method described by Lizada and Yang (1979) with minor modification. The fruit pulp tissue (5–6 g) was homogenized in a glass pestle and mortar using 300 mg white quartz sand with 10 ml distilled water at 2 1 C temperature. Following centrifugation at 10,000 g for 20 min at 4 C, 0.5 ml supernatant was mixed with 0.1 ml HgCl2 (50 mM) in two sets, one with and the other without 0.1 ml ACC (100 mM) and the final volume 1.8 ml was made with distilled water. The glass test tubes were sealed with a rubber septum and both test tubes were placed on ice, 0.2 ml (5% NaOCl þ saturated NaOH 2:1, v/v) solution was injected into the sealed tubes and vortexed for 5 s. After 25 min incubation on ice, the tubes were again vortexed and 1 ml gas sample was taken for ethylene measurements, expressed as pmol g1 fresh weight (FW). After the ethylene concentrations of the gas samples generated from both the test samples and the ACC solutions were determined, the proportions of ACC solutions that were transformed into ethylene in the reactions were calculated. Then the ACC concentrations in the test samples were calculated since it is assumed that there are the same transformation proportions for both the test samples and the ACC solutions. 1-Aminocyclopropane-1-carboxylic acid synthase (ACS) activity was measured using five fruits per treatment every other day after treatment. ACS activity was measured in pulp tissue by following the method of Mathooko et al. (1993) with some modification. Pulp tissue (10 g) was homogenized with 5 ml K-phosphate
buffer (0.5 M, pH 8.5) containing 5 mM pyridoxal phosphate and 5 mM dithiothreitol in the presence of 5% polyvinylpyrrolidone (PVP) in a glass pestle and mortar using 400 mg white quartz sand (50 þ 70 mesh, Sigma-Aldrich, Australia) at 2 1 C. Contents were centrifuged at 14,000 g for 30 min at 4 C and 2 ml of supernatant was mixed with 1 ml of 500 mM S-adenosyl methionine (SAM) in a glass reaction tube. The reaction tube was sealed with a rubber septum, incubated for 30 min at 30 C, and transferred to an ice bath. Using a syringe through the stopper, 0.1 ml of HgCl2 (50 mM) and 0.3 ml of NaOCl (5%) and saturated NaOH (2:1, v/v) were added into the reaction tube. The reaction tube was incubated in ice for a further 2.5 min and a 1 ml gas sample was taken from the head space and injected into the gas chromatograph for ethylene estimation, as described earlier. ACS activity was expressed as nmol ACC g1 FW h1. ACO activity was measured using five fruits per treatment every other day after treatment. ACO activity was measured in pulp tissue by following the method of Mathooko et al. (1993) with some modifications. Pulp tissue (2 g) was homogenized in a glass pestle and mortar using 200 mg white quartz sand in 5 ml extraction buffer consisting of 0.1 M Tris-HCl (pH 7.2), 10% (w/v) glycerol and 30 mM sodium ascorbate in the presence of 5% PVP. The homogenized mixture was centrifuged at 14,000 g for 30 min. All the steps in the enzyme extraction were carried out at 2 1 C. The enzyme activity was measured in 2 ml reaction mixture containing 1.8 ml of the supernatant solution, 0.1 ml ACC (40 mM) and 0.1 ml FeSO4 (1 mM). The reaction tube was sealed with a rubber septum and incubated at 30 C for 60 min. One milliliter of gas taken from the headspace was injected into the gas chromatograph for ethylene estimation, as described earlier. ACO activity was expressed as nmol C2H4 g1 FW h1. Weight loss and firmness. Five melon fruits of each treatment were weighted every other day to determine the weight loss. Weight loss (%) was calculated by equation (1).
Weight lossð%Þ ¼ ðWx W0 Þ=W0 100
ð1Þ
where W0 was the weight at 0 day and Wx was the weight at a certain day after storage. After melons were lengthways cut into two halves, firmness measurements were made on both of the equator and polar of the pulp tissue 1 cm from the seed cavities on the cut surfaces. Firmness was measured via five fruits per treatment every other day and was expressed as Newton (N). The assay was made using a TA-XT2i texture analyzer (Stable Micro Systems, England) 3
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RESULTS AND DISCUSSION
Total soluble solid (TSS) contents and titratable acidity (TA). TSSs were determined on the juice obtained from melon edible pulp tissue using a digital hand-held refractometer (Atago PAL-1, Tokyo, Japan). TSS was expressed as Brix. TA was measured by following a previous method (Guille´n et al., 2007) with modifications. The juice was also obtained from fruit edible pulp tissue. TA was determined by potentiometric titration with 0.1 N NaOH up to pH 8.1, using 1 ml of diluted juice in 25 ml distilled H2O and results are expressed as gram of citric acid equivalent per 100 g1 FW. Five fruits from each treatment were used for analysis of these parameters.
In the present study, shelf life of netted melon under different concentrations of 1-MCP was monitored to determine the most effective 1-MCP dose. Netted melons treated with 0.6 ml l1 1-MCP exhibited the longest shelf life, approximately seven (25 C) or nine (10 C) days (Figure 1), which is significantly longer than that of control or other treatments, including those treated with 0.8 or 1 ml l1 1-MCP. When stored under room temperature (25 C), the shelf life of melon fruits was prolonged for 4 days under 0.6 ml l1 1-MCP treatment when compared with the control. And the shelf life of melons stored under 10 C was also 4 days longer than that of the control. Therefore, under both temperatures, 25 C and 10 C, 0.6 ml l1 was the most effective dose of 1-MCP to extend fruit shelf life. The results are consistent with a previous report (de Melo et al., 2008), in which postharvest application of 1-MCP extended shelf life of orange-flesh melons from 15 days to 21 days under room temperature storage. In this experiment, as it is shown from fruit appearance, higher concentration of 1-MCP (0.8 and 1 ml l1) did not produce better effect. It is probably because 1-MCP reached its saturating concentration after 0.6 ml l1. When 1-MCP permanently binds to a sufficient number of ethylene receptors, higher doses of 1-MCP will not maintain longer fruit shelf life.
Sugar components. Sugars were determined using five fruits from each treatment every other day as previously described (Komatsu et al., 1999; Lowell et al. 1989). Samples of mesocarp tissues (ca. 1 g FW) were homogenized for 5 min in 5 volumes of extracted solution (ethanol:chloroform:water ¼ 12:5:3, l/l). Water and chloroform were then added to bring the final E:C:W ratio to 10:6:5. Subsequent separation of the chloroform layer allowed removal of lipids and pigments. The remaining aqueous-alcohol phase was evaporated to dryness in a vacuum at 50 C and re-dissolved in 1 ml distilled water. Analysis of soluble sugars was performed by high performance liquid chromatography (Shimadzu Co., Kyoto, Japan) with an NH2 column at 30 C using 65% acetonitrile solution (1 ml min1) as a mobile phase and a refractive index detector (Shimadzu RID-10 A). Peaks were quantified by using corresponding analytical software. As to the quantification of sugar content, the procedures are divided into three steps in the following. Firstly, several sorts of dilution of sugar solution (glucose, fructose and sucrose) were made by dissolving the required anount of authentic standard in deionized water (dH2O). Secondly, three standard sugar solutions will be analyzed in the ideal chromatography. Finally, the concentration of sucrose, glucose and fructose were quantified by comparison between standard sugar solution and sample solution. Statistical analysis. Statistical analysis was performed using the SPSS package program version 11.5 (SPSS Inc. Chicago, IL). Data were analyzed using one-way analysis of variance (ANOVA) model. The variants were storage time and different treatments. The means were compared using the least significant differences (LSDs) test at a significant level of 0.05. The values were reported as means with standard deviations.
Shelf life
Ethylene production and respiration rate Ethylene plays an important role in fruit ripening and senescence. Many ripening processes, including fruit softening, weight loss, peel degreening, aroma generating and change of sugar content, are highly dependent on ethylene action. Netted melon exhibited typical climacteric changes during storage. Ethylene accumulation is a necessity as to the full accomplishment of some ripening procedure which is closely related to fruit quality in climacteric fruits (e.g. cell wall degradation reflected in fruit softening). The problem emerges when this ethylene production is excessive or too prolonged in time; then detrimental effects appear such as fruit rot, mainly because of extreme softening or the fruit that breeds cracking of it and production of smelly scents rather than agreeable aroma. There are many tools to inhibit onset of autocatalytic ethylene production among which, the effectiveness of 1-MCP in extending storage time and improving quality in many kinds of fruit has been widely proven (Blankenship and Dole, 2003). For instance, postharvest application of 1-MCP after onset of ripening completely halted subsequently softening in Charentais melon (Nishiyama et al., 2007) and in ‘Galia’ melon,
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onset of ethylene climacteric was delayed by 10 days compared with the control (Gal et al., 2006). It is reported that ethylene production in 1-MCPtreated melons was significantly lower than that in control during the first 6 days of storage (Li et al., 2011a). In addition, 1-MCP treatment delayed onset of ethylene climacteric and subsequently retards other ripening processes in persimmon (Harima et al., 2003) and tomato (Guille´n et al., 2007). This is consistent with the present work. In this study, ethylene production exhibited typical climacteric changes under both ambient (25 C) and low temperature (10 C) storage conditions. Ethylene peaks in 1-MCP-treated fruits were delayed for 3 days (25 C) or 2 days (10 C) as compared to control (Figure 2). The maximum ethylene production of 1-MCP-treated fruits was reduced by 35.2% and 35.5%, respectively, under 25 C and 10 C when compared with the control. 1-MCP occupied ethylene receptors so that ethylene cannot bind and elicit its action (Blankenship and Dole, 2003). There is an optimum concentration of 1-MCP for each crop. The effectiveness of the low concentration of 1-MCP in the present study may be due to high accessibility and affinity of 1-MCP to the receptors. Although binding of 1-MCP to ethylene receptor is irreversible, plant recover their ethylene sensitivity due to turnover or production of new receptors (Sisler and Serek, 1997). It is observed from Figure 2 that in the middle period of storage (day 4), new receptors probably are being synthesized since the autocatalytic ethylene production quickly increased, and then the process of fruit ripening continued. In this research, the trends of respiration rates in terms of 1-MCP treated and untreated fruits distinguished a lot. Between the two figures, respiration
rates of untreated fruits rose dramatically to the peak at first, and then delined during the storage period. Whereas, both of respiration rates of 1-MCP-treated fruits maintained a steady growth. Meanwhile, it is worth to notice that, under different storage temperatures, respiration rates in terms of 1-MCP treated and untreated fruits have a clear difference. Particularly, for one, respiration rate of 1-MCP-treated fruits in 25 C was far lower than that of untreated fruits in 2–7 days; for another, respiration rate of 1-MCP-treated fruits in 10 C was under that of untreated fruits in 4–7 days. In ‘Galia’ melon (Gal et al., 2006), onset of ethylene and respiration climacterics are dramatically delayed and both of the peak values are reduced, which are consistent with results of our present work. However, 1-MCP treatment had no effect on apricot (Dong et al., 2002). The distinct results in apricots might be due to fruit species, maturity, cultivar or some other unknown factor (Blankenship and Dole, 2003). Respiration rate of produce is a useful guide to assess the potential storage life of produce. Respiration is energy-consuming and causes loss of sugars and of weight, which are both important causes of fruit quality deterioration. Therefore, 1-MCP-treated fruits with lower respiration rate have a higher capacity of storage than the nontransformed control fruits as the present study shows. Storage temperature In this research, netted melons were stored under both 10 C and 25 C to examine the effect of temperature on efficacy of 1-MCP. Our results showed that ethylene production and respiration rate of melons stored in 10 C were greatly lower than that in room temperature (25 C) (Figure 2), indicating 10 C storage obviously 5
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Figure 2. Effect of 1-MCP treatment on ethylene production and respiration rate of netted melon fruits during storage at both 25 C (a, c) and 10 C (b, d). Each value is the mean of five fruits. Vertical bars represent standard deviation of the mean.
delayed the fruit senescence, postponed the ethylene peaks and reduced the respiration rate. In the experiment of kiwifruit, the extent of ethylene inhibition caused by 1-MCP is related to temperature, namely, low temperature storage resulted in better 1-MCP effects (Kim et al., 2001), which is consistent with our results. However, applying lower temperature to attain a better postharvest storage is not completely true, since it has been reported by Miccolis and Saltveit (1995) that, ‘for certain melon varieties, chilling injury induced symptoms were much serious in fruits when stored in 7 C for 1–2 weeks than when stored under 15 C’. On the other hand, low temperature storage is an effective method to preserve melons, but cold storage facilities are not always available. Therefore, postharvest 1-MCP application can be very helpful in extending fruit shelf life and maintaining fruit postharvest qualities, especially where low temperature chain transportation is unavailable.
ACC, ACS and ACO Ethylene biosynthesis is initiated from the amino acid, methionine, via S-adenosylmethionine (Adomet) through the intermediate 1-aminocyclopropane-1carboxylic acid (ACC) that the biosynthesis of this intermediate is catalyzed by the enzyme ACC synthase (ACS). Next, the enzyme ACO catalyzes the conversion of ACC to ethylene (Ezura and Owino, 2008). The biosynthesis of ethylene may also be affected by 1-MCP due to its competitive binding to ethylene receptors. Our results showed that 1-MCP treatment significantly inhibited ACC accumulation during the storage under both 25 C and 10 C, although lower ACC content was found out under storage at 10 C when compared with that at 25 C. According to Khan and Singh (2009), ACC content of 1-MCP-treated plum were suppressed. Consistent results were also found out in mango, in which the ACC increase was significantly delayed by
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Figure 3. Effect of 1-MCP treatment on ACC content, ACS activities and ACO activities in pulp of netted melon fruits during storage at both 25 C (a, c, e) and 10 C (b, d, f). Each value is the mean of five fruits. Vertical bars represent standard deviation of the mean.
1-MCP (Wang et al., 2009). The reduction of ACC content in 1-MCP-treated melon may be resulted from reduced ACS and ACO activity in the fruit. In this study, similar patterns of ACC contents and ACS activity were detected in both treated and untreated fruits. As it is shown in Figure 3, distinct decline of
ACS activity was found in 1-MCP treated melons. At the end of storage, ACS activity in 1-MCP treated melons was 27.2% (25 C) and 41.7% (10 C) lower than that of control, correspondingly. We also found that during storage, ACO activity in melons increased and then declined, which were consistent with the 7
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pattern of ethylene production. Moreover, ACO activity in 1-MCP treated melons were constantly lower than that of control, and the peaks were 55.6% (25 C) and 42.3% (10 C) lower than that of control correspondingly during storage. Therefore, it is concluded that the decline of ACC content by 1-MCP may be resulted from the decreased ACS and ACO activity. In ‘Hami’ melon, 1-MCP also markedly inhibited the accumulation of ACC and maintained lower activities of ACS and ACO (Li et al., 2011a). Meanwhile in plums, a reduction in ACS and ACO activity and ACC content in 1-MCP-treated fruits during storage at ambient or cold temperature was also reported (Khan and Singh, 2009). Therefore, reduced climacteric ethylene production in 1-MCPtreated fruit, may be ascribed to the reduced activity of ethylene biosynthesis enzymes ACS and ACO. Several ACS and ACO isoenzymes have been identified in melon tissues and both enzymes are encoded by multigene family (Lasserre et al., 1996; Miki et al., 1995; Yamamoto et al., 1995). In other species, the
adjustment of the expression of the main ethylene biosynthetic genes, ACS and ACO, has also been mentioned (Ezura and Owino, 2008). It should be noted that the different patterns in ACS and ACO activity during melon storage may be due to the different levels of ACS and ACO transcriptions, which was reported by Pathak et al. (2003). Consequently, it can be concluded that the suppression of ethylene perception by 1-MCP in melons is mainly attributed to its feedback suppression of ACS and ACO activity. Fruit weight and firmness Fruit weight and firmness are two important attributes that affect melon visual quality and are also closely related to fruit shelf life. In this study, weight loss of both 1-MCP-treated fruits and the control fruits increased during the storage period. In ‘Galia’ melon (Gal et al., 2006), 1-MCP treatment caused a reduction of weight loss. Alves et al. (2005) also found that in ‘Charentais’ melon, control fruit lost 7% of weight
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Figure 5. Effect of 1-MCP treatment on total soluble solid (TSS) content and titratable acidity (TA) in pulp of netted melon fruits during storage at both 25 C (a, c) and 10 C (b, d). Each value is the mean of five fruits. Vertical bars represent standard deviation of the mean.
while 1-MCP-treated fruit lost only 5% of weight at ambient temperature. However, contradictory results were also found in fruits of other species. In apricot (Fan et al., 2000) and orange (Porat et al., 1999), no effect was observed after 1-MCP treatment, while in avocado (Jeong et al., 2002) and Chinese kale (Sun et al., 2012), weight loss in the fruits were delayed by 1-MCP treatment. In addition, 1-MCP application significantly reduced the weight loss in both storage conditions. The main cause of weight loss is consumption of respiration substrates, mainly sugars, and this was confirmed by the similarity in changes of weight loss and respiration rate in present study. Softening is one of the most sensitive ripening processes to ethylene. Ethylene plays a major role in regulating softening of climacteric fruit, including Charentais melon (Ayub et al., 1996; Hadfield et al., 2000). In ‘Galia’-type melon (Ergun et al., 2005; Gal et al., 2006), 1-MCP maintained fruit firmness when compared to the control. In ‘Hami’ melon, decrease in firmness was significantly inhibited by 1-MCP (Li et al., 2011a). In this study, the decrease of pulp firmness was obviously reduced in 1-MCP treated melons
when contrasted with the control fruits during the 8 days of storage (Figure 4). At the end of the experiment, firmness values of 1-MCP treated melon were 49.6% and 26.4% higher, respectively, under 25 C and 10 C when compared with the control. Fruit firmness was reduced in both treated and untreated fruits when stored at the lower temperature (10 C). While in fruits of other species, firmness was better maintained by 1-MCP in apricots (Fan et al., 2000), peaches (Kluge and Jacomino, 2002) and plums (Dong et al., 2002, 2001) as well. TSS and TA In netted melon, accumulation of sugars (especially sucrose) is one of the essential factors that determine fruit quality and commodity values (Lingle and Dunlap, 1987). In ‘Galia’ melon, 1-MCP did not affect TSS content after prolonged storage and shelflife simulation. However, in the present study, less reduction of TSS content was monitored in 1-MCPtreated fruits than the control fruits. 1-MCP treatment effectively inhibited the reduction of TSS contents of 9
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Figure 6. Effect of 1-MCP treatment on sucrose, fructose and glucose contents in pulp of netted melon fruits during storage at both 25 C (a, c, e) and 10 C (b, d, f). Each value is the mean of three replicates. Vertical bars represent standard deviation of the mean.
melon fruits (Figure 5). After 8 days of storage, TSS of fruits treated with 1-MCP were reduced by approximately 14.19% (25 C) and 10.72% (10 C), respectively, as compared to 22.71% and 13.03% in untreated fruits. Consistent results were observed in 1-MCP treated ‘Hami’ melon (Li et al., 2011b), TSS of 1-MCP-treated fruits is higher than that of the control. In fresh-cut ‘Hami’ melon (Guo et al., 2011) and fresh-cut cantaloupe (Amaro et al., 2013), TSS of the fruit are also better maintained better after
1-MCP treatment. Since respiration was lower in 1-MCP-treated fruit, less sugar is consumed when comparing with control. Therefore, it is possible that it is not 1-MCP that influences sugar content, but the concomitant low respiration rate. As it has been mentioned, during 2–7 days storage and 4–7 days storage of 25 C and 10 C respectively, respiration rate of 1-MCP-treated fruits was obviously lower that of control fruits, which tells TSS content of 1-MCP-treated fruits in 2–8 days storage was evidently less than that of
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Shi et al. control fruits. The reduction of TSS loss by 1-MCP treatment was higher in melons stored at 10 C than those at 25 C. However, Cohen and Hicks (1986) found that storage temperature did not influence soluble sugar content after 2, 5 or 9 days at 5, 12.5 and 20 C. Moreover, the effect of 1-MCP on soluble sugars also depends on the crop (Blankenship and Dole, 2003). There are different 1-MCP effects in TSS in other species. Soluble solids were higher in 1-MCP-treated pineapple (Selvarajah et al., 2001), papaya (Hofman et al., 2001) and kiwifruit (Boquete et al., 2004). However, soluble solids were reduced in 1-MCP-treated strawberries (Tian et al., 2000). In other commodities, such as persimmons, apricots and plums, TSSs were not affected by 1-MCP (Dong et al, 2002; Salvador et al., 2004; Watkins et al., 2000). As it can be observed in Figure 5, 1-MCP treatment significantly inhibited TA reduction in netted melon fruits. In addition, reduction of TA in fruit stored in 25 C was higher than those stored in 10 C during storage period. In previous reports, 1-MCP prevented acidity loss in tomatoes (Wills and Ku, 2002), delayed TA loss in plums (Dong et al., 2002) and loquats (Cai et al., 2006). In contrast, 1-MCP did not affect TA in apricots (Dong et al., 2002) or apples (Mir et al., 2001) during storage at different temperatures. Sucrose, glucose and fructose In this study, patterns of sugar metabolism in the melon fruit were greatly influenced by 1-MCP during storage. In netted melon, sugars that contribute to the TSSs were mainly sucrose, fructose and glucose. Changes in the contents of these sugar components were shown in Figure 6. Sucrose was the predominant sugar in melon pulp tissue and in both treated and untreated fruits, sucrose content exhibited constant decrease during postharvest storage. However, 1-MCP treatment reduced the sucrose content up to a certain degree. At the end of storage, sucrose content in 1-MCP treated melons were 5.27% and 2.15% higher, respectively, under 25 C and 10 C than that of control. Moreover, higher sucrose content was observed in fruits stored at 10 C when compared with those stored at 25 C. Fructose and glucose are two important sugar components in melon since they make up the components of sucrose, the main sugar found in melon fruit. It can be observed in Figure 6 that changing curves of both fructose and glucose were similar to that of sucrose, and that 1-MCP also effectively reduced the diminution of the contents of these sugars. This is consistent with the results of a report on ‘Yujinxiang’ melon (Ma et al., 2012). However, under 10 C storage, as to different sugar components, 1-MCP treatment showed significant effects from
different time: day 4 for sucrose, day 6 for fructose and day 2 for glucose.
CONCLUSIONS A volume of 0.6 ml l1 1-MCP significantly extended melon shelf life, reduced ethylene production and respiration rate, and retarded the changes in parameters related to fruit ripening, including fruit weight loss, firmness, TSS content, acidity and the main sugar contents. In conclusion, 1-MCP treatment is a good method to extend melon shelf life and maintain fruit quality, no matter fruits were stored under room temperature (25 C) or lower temperature (10 C), and the combination of 1-MCP and low temperature storage resulted in more acceptable fruit quality. ACKNOWLEDGMENTS We are thankful to the 985-Institute of Agrobiology and Environmental Sciences of Zhejiang University for providing access to analytical instruments.
CONFLICT OF INTEREST None declared.
FUNDING This work was funded by Major Scientific and Technological Key Program of Ningbo (2010C10012), Major special breeding program of Science and Technology Department of Zhejiang Province (2012C12903-2-7) and National Science and Technology Support Program of China (2013BAD201300).
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