Cell wall modification in 1-methylcyclopropene-treated post-climacteric fresh-cut and intact papaya fruit

Papaya is a climacteric fruit in which ripening is greatly regulated by ethylene often associated with stress responses such as wounding. The changes in cell wall compositions in papaya fruit at an advanced stage of ripening under stress conditions including chilling temperature of 5°C and wounding employed as fresh-cut and how these changes were affected by an ethylene action inhibitor of 1-methylcyclopropopene (1-MCP) were examined in the study. The recovery of ethanol-insoluble solids, total soluble sugars, water-soluble polyuronides, neutral hemicelluloses, and neutral sugars of rhamnose, arabinose, mannose and glucose were not affected by 1-MCP or fresh-cut processing. The fresh-cut processing, however, caused a higher loss of total polyuronides and the neutral sugar galactose while increasing the recovery of chelator-soluble polyuronides. Few significant differences due to 1-MCP application were recorded in the recoveries of alkali-soluble polyuronides, hemicellulosic polyuronides extracted with 4% KOH, and the neutral sugar xylose. Modifications of cell wall polyuronides and hemicelluloses in ripe fresh-cut papaya fruit exhibited mostly similar patterns to those in intact ripe papaya fruit under the chilling temperature of 5°C while minimally affected by 1-MCP.     

Responses of strawberry fruit to 1-Methylcyclopropene (1-MCP) and ethylene

1-Methylcyclopropene (1-MCP), a competitive inhibitorof ethylene action, binds to the ethylene receptor toregulate tissue responses to ethylene. In this work,we investigated the effects of 1-MCP and exogenousethylene on ripening, respiration rate, ionicconductivity and peroxidase activity in strawberryfruit. Strawberry fruit can ripen without exogenousethylene treatment, but exogenous ethylene inducessecondary ripening processes. Results indicated thatstimulation of respiration by ethylene wasdose-dependent. Fruit colour development and softeningwere slightly accelerated by ethylene, but changes insoluble solid content were not. 1-MCP may/may notaffect the respiratory rise induced by exogenousethylene dependent on fruit maturity. Cycloheximide(CHI) reduced the ethylene-induced respiratoryincrease. Combinations of 1-MCP and CHI reducedrespiration more than CHI alone. 1-MCP and CHI did notinfluence the primary respiratory change in nonethylene-treated fruit. This indicates that ethyleneinduced respiratory increase may involve an ethylenereceptor in early harvested fruit, but not in laterharvested fruit. Exogenous ethylene stimulatedrespiration by regulating new respiratory enzyme(s)synthesis in strawberry fruit. Ethylene induced anionic leakage increase, and this was positivelycorrelated to fruit water loss and peroxidaseactivity. These results suggest that non-climactericfruit, such as strawberry, may have different ethylenereceptor(s) and/or ethylene receptor(s) may havedifferent regulatory functions. It may be thesecondary effect of ethylene to stimulate respirationin strawberry. Non-climacteric fruit ripening may berelated to the development of active oxygen species(AOS) induced by postharvest stress.     

Papaya fruit softening, endoxylanase gene expression, protein and activity

Papaya ( Carica papaya L.) cell wall matrix polysaccharides are modified as the fruit starts to soften during ripening and an endoxylanase is expressed that may play a role in the softening process. Endoxylanase gene expression, protein amount and activity were determined in papaya cultivars that differ in softening pattern and in one cultivar where softening was modified by the ethylene receptor inhibitor 1-methylcyclopropene (1-MCP). Antibodies to the endoxylanase catalytic domain were used to determine protein accumulation. The three papaya varieties used in the study, 'Line 8', 'Sunset', and 'Line 4-16', differed in softening pattern, respiration rate, ethylene production and showed similar parallel relationships during ripening and softening in endoxylanase expression, protein level and activity. When fruit of the three papaya varieties showed the respiratory climacteric and started to soften, the level of endoxylanase gene expression increased and this increase was related to the amount of endoxylanase protein at 32 kDa and its activity. Fruit when treated at less than 10% skin yellow stage with 1-MCP showed a significant delay in the respiratory climacteric and softening, and reduced ethylene production, and when ripe was firmer and had a 'rubbery' texture. The 1-MCP-treated fruit that had the 'rubbery' texture showed suppressed endoxylanase gene expression, protein and enzymatic activity. Little or no delay occurred between endoxylanase gene expression and the appearance of activity during posttranslational processing from 65 to 32 kDa. The close relationship between endoxylanase gene expression, protein accumulation and activity in different varieties and the failure of the 1-MCP-treated fruit to fully soften, supported de novo synthesis of endoxylanase, rapid posttranslation processing and a role in papaya fruit softening.     

Papaya Fruit Softening: Role of Hydrolases

Papaya (Carica papaya L.) cultivars show a wide variation in fruit softening rates, a character that determines fruit quality and shelf life, and thought to be the result of cell wall degradation. The activity of pectin methylesterase, β-galactosidase, endoglucanase, endoxylanase and xylosidase were correlated with normal softening, though no relationship was found between polygalacturonase activity and softening. When softening was modified by 1-MCP treatment, a delay occurred before the normal increase in activities of all cell wall activities except endoxylanase which was completely suppressed. Significant cell wall mass loss occurred in the mesocarp tissue during normal softening, but did not occur to the same extent following 1-MCP treatment. During normal softening, pectin polysaccharides and loosely bound matrix polysaccharides were solubilized and the release of xylosyl and galactosyl residues occurred. Cell wall changes in galactosyl residues after 1-MCP treatment were comparable to those of untreated fruit but 1-MCP treated fruit did not soften completely. The changes in the cell wall fractions containing xylosyl residues in 1-MCP treated fruit showed less solubilization and a higher association of xylosyl residues with the pectic polysaccharides. The results indicated that normal modification of cell wall xylosyl components during ripening did not occur following 1-MCP treatment at the color-break stage, this was associated with the failure of these fruit to fully soften and a selective suppression of endoxylanase activity. The results support a role for endoxylanase in normal papaya fruit softening and its suppression by 1-MCP lead to a failure to fully soften. Normal papaya ripening related softening was dependent upon the expression and activity of endoglucanase, β-galactosidase and endoxylanase.     

香蕉一个Ⅲ类酸性几丁质酶基因与果实成熟关系的研究

为了解Ⅲ类酸性几丁质酶基因(MaCHⅢ)与香蕉果实采后成熟过程的相互关系,对经乙烯和1-甲基环丙烯(1-MCP)处理的巴西香蕉果实采后乙烯释放量、Ⅲ类酸性几丁质酶基因(MaCHⅢ)表达以及几丁质酶活性进行了测定.结果显示:(1)乙烯催熟处理的香蕉果实,乙烯释放量比对照处理的果实提前15 d达到高峰;1-MCP处理的香蕉果实,乙烯生物合成和果实成熟明显受到了抑制.(2)外源乙烯加速了MaCHⅢ基因的下调表达和Ⅲ类酸性几丁质酶活性的下降,MaCHⅢ表达量和Ⅲ类酸性几丁质酶活性分别在采后第3天和第4天下降到最小值.(3)1-MCP处理使MaCHⅢ基因呈现上调表达,Ⅲ类酸性几丁质酶活性上升,MaCHⅢ基因表达量和Ⅲ类酸性几丁质酶活性分别在采后18 d和25 d达到高峰.研究表明,MaCHⅢ基因可能与香蕉果实采后成熟呈负相关.     

Differential Suppression of Ethylene Biosynthesis and Receptor Genes in ‘Golden Delicious’ Apple by Preharvest and Postharvest 1-MCP Treatments

Harvista?, a sprayable formulation of 1-methylcyclopropene (1-MCP), has recently been developed for preharvest use on horticultural products, whereas SmartFresh? is a widely used 1-MCP treatment for products after harvest. The effects of Harvista? on apple fruit ripening when sprayed at different maturities and on expression patterns of ethylene biosynthesis and receptor genes during storage have been investigated. Harvista? applied to on-tree maturing apple fruit at an average starch pattern index of 2.5 resulted in a higher at-harvest firmness value compared with those treated at a starch pattern index of 1.5 and 3.5. This indicates that the timing of the Harvista? application is critical. An application of Harvista? led to better postharvest fruit firmness retention as well as reduced ethylene production. In addition, both preharvest and postharvest 1-MCP treatments resulted in contrasting responses in the expression patterns of two ethylene biosynthesis genes and in differentially suppressing effects on four ethylene receptor genes. Furthermore, the combined application of Harvista? + SmartFresh? resulted in greater fruit firmness retention and longer ethylene suppression. The expression profiles of these genes during on-tree fruit maturation prior to Harvista? application were also characterized. Different regulation patterns of receptor genes could contribute to differential effects by 1-MCP treatments. The potential roles of Harvista? to manipulate the ripening process as well as the molecular mechanism influencing 1-MCP treatment efficacy are discussed.     

Responses of banana fruit to treatment with 1-methylcyclopropene

Experiments were conducted to determine levels of 1-methylcyclopropene (1-MCP) exposure needed to prevent ethylene-stimulated banana fruit ripening, characterise responses of ethylene-treated fruit to subsequent treatment with 1-MCP, and to test effects of subsequent ethylene treatment on 1-MCP-treated fruit softening. Fruit softening was measured at 20°C and 90% relative humidity. One hour exposure at 20°C to 1000 nl 1-MCP/l essentially eliminated ethylene-stimulated ripening effects. Exposure for 12 h at 20°C to just 50 nl 1-MCP/l was similarly effective. Fruit ripening initiated by ethylene treatment could also be delayed with subsequent 1-MCP treatment. However, 1-MCP treatment only slowed down ripening of ethylene-treated fruit when applied at 1 day after ethylene and was ineffective when applied 3 or 5 days after ethylene treatment. The ripening response of fruit treated with 1-MCP and subsequently treated with ethylene varied with interval time between 1-MCP and ethylene treatments. As time increased, the response of 1-MCP-treated fruit to ethylene was enhanced. Responses to 0.1, 1, 10 or 100 µl ethylene/l concentrations were similar. Enzyme kinetic analysis applied to 1-MCP effects on ethylene-induced softening of banana fruit suggested that 1-MCP inhibition is by noncompetitive antagonism of ethylene binding.     

Delay ripening of ‘Qingnai’ plum (Prunus salicina Lindl.) with 1-methylcyclopropene

‘Qingnai’ plum fruit were treated with 0, 250, 500 or 1000 nL L⁻¹ of 1-methylcyclopropene (1-MCP) for 6 h and stored at 20 °C. The fruit firmness, peel color, chlorophyll content, titratable acidity (TA), respiration rate and ethylene production, chlorophyllase, pectin methylesterase (PME) and polygalacturonase (PG) activities were monitored during postharvest ripening of ‘Qingnai’ plums. ‘Qingnai’ plums without 1-MCP treatment soften very rapidly at room temperature after harvest, showing a continuing decrease in hue angle, chlorophyll content, TA and increase in chlorophyllase, PME and PG activities during postharvest storage. In contrast, the 1-MCP-treated fruits showed reduced ethylene production and respiration rate and delayed softening, which was associated with the reduction in the activity of PME and PG. The 1-MCP treatment also significantly inhibited the chlorophyllase activity and peel color development in ‘Qingnai’ plums during postharvest ripening at 20 °C. These results suggest that 1-MCP treatment may be useful for maintaining the fruit quality and extending the postharvest shelf-life of ‘Qingnai’ plums.     

Regulatory mechanisms of ethylene biosynthesis in response to various stimuli during maturation and ripening in fig fruit (Ficus carica L.).

In order to obtain a greater uniformity of maturation, the growth of the fig fruit (Ficus carica L.) can be stimulated by the application of either olive oil, ethrel/ethephon or auxin. The three treatments induce ethylene production in figs. In this study, we investigated the regulatory mechanisms responsible for oil, auxin and ethylene induced ethylene production in figs. The ethylene production in response to olive oil, auxin, and propylene treatments and during ripening were all induced by 1-methylcyclopropene (1-MCP) and inhibited by propylene indicating a negative feedback regulation mechanism. Three 1-aminocyclopropane-1-carboxylic acid (ACC) synthase genes (Fc-ACS1, Fc-ACS2 and Fc-ACS3) and one ACC oxidase gene (Fc-ACO1) were isolated and their expression patterns in response to either oil, propylene or auxin treatment in figs determined. The expression patterns of Fc-ACS1 and Fc-ACO1 were clearly inhibited by 1-MCP and induced by propylene in oil treated and ripe fruits indicating positive regulation by ethylene, whereas Fc-ACS2 gene expression was induced by 1-MCP and inhibited by propylene indicating negative regulation by ethylene. The Fc-ACS3 mRNA showed high level accumulation in the auxin treated fruit. The inhibition of Fc-ACS3 gene by 1-MCP in oil treated and in ripe fruits suggests that auxin and ethylene modulate the expression of this gene by multi-responsive signal transduction pathway mechanisms. We further report that the olive oil-induced ethylene in figs involves the ACC-dependent pathway and that multiple ethylene regulatory pathways are involved during maturation and ripening in figs and each specific pathway depends on the inducer/stimulus.     

Low-temperature-modulated fruit ripening is independent of ethylene in 'Sanuki Gold' kiwifruit

Fruit ripening in response to treatments with propylene, 1-methycyclopropene (1-MCP), and low temperature was characterized in 'Sanuki Gold' kiwifruit, Actinidia chinensis Planch. Propylene treatment immediately induced rapid fruit softening, increased AC-PG (polygalacturonase) and AC-EXP (expansin) mRNA accumulation, and stimulated an increase in the soluble solid concentration (SSC) and a decrease in titratable acidity (TA). After 3?d exposure to propylene, ethylene production and AC-PL (pectate lyase) mRNA accumulation were observed. 1-MCP treatment after 24?h exposure to propylene eliminated AC-PG mRNA accumulation and suppressed continued changes in SSC and TA. Application of 1-MCP at the start of the treatment, followed by continuous propylene exposure, markedly delayed fruit softening, and the expression of the cell wall-modifying genes, and changes in the SSC and TA, indicating that kiwifruit become insensitive to ethylene at least for 3?d following 1-MCP exposure. Surprisingly, significant fruit softening, mRNA accumulation of AC-PG, AC-PL, and AC-EXP, and decreased TA were observed without ethylene production in intact fruit stored at low temperature for 1 month, but not in fruit stored at room temperature. Repeated 1-MCP treatments (twice a week) failed to inhibit the changes that occurred in low temperature storage. These observations indicate that low temperature modulates the ripening of kiwifruit in an ethylene-independent manner, suggesting that kiwifruit ripening is inducible by either ethylene or low temperature signals.     

Effect of eco-safe compounds on postharvest quality preservation of papaya (<Emphasis Type="Italic">Carica papaya</Emphasis> L.)

Early ripening and susceptibility to microbial infection are major postharvest problems in papaya fruits. Being a tropical climacteric fruit, low-temperature storage is not successful in papaya. In this study, we demonstrate the effect of aqueous salicylic acid (1 and 2 mM), nitric oxide (1 and 2 mM), and calcium chloride (1 and 2%) to enhance the papaya shelf life at the ambient conditions with reduced disease incidence. Calcium chloride 2% was the most effective for maintaining postharvest quality. The fruits had good firmness and maintained TSS, acidity, total chlorophyll, free radical scavenging activity and ascorbic acid on the 6th day during ambient storage. Moreover, the weight loss, yellow color development and disease incidence were minimum in calcium chloride 2%, followed by 1% solution of calcium chloride. The nitric oxide (2 mM) maintained higher antioxidant capacity and total phenol content in fruits that was followed by 1 mM salicylic acid during storage. The result suggests that application of calcium chloride 2% could be an easy and effective technique for extending the shelf life without impairing fruit quality during storage.     

Effect of post-harvest heat treatment on proteome change of peach fruit during ripening

The extracted proteins from the heat-treated peach fruit (dipped in hot water at 48°C for 10min and then stored at room temperature (20°C-25°C) for up to 6 days) were used for proteomic analysis in order to understand the response of post-harvest peach fruit to heat treatment during ripening stage at proteomic level. After two dimensional gels electrophoresis (2-DE) was conducted, more than 600 protein spots were detected. Among them, 35 differently expressed spots (P<0.05) were selected to be excised and analyzed using MALDI-TOF/TOF, and finally 30 protein spots were confidently identified according to NCBI database. The results demonstrated that among the thirty protein spots expressed particularly induced by heat treatment, 43% were related to stress response, 17% to cell structure, 13% to protein fate, 7% to glycolytic pathway, 3% to ripening and senescence and 17% to unclassified. All of them are involved in the regulation of peach fruit development and ripening. All these indicated that the self-defense capability of peach fruit was improved by heat treatment. The study will enable future detailed investigation of gene expression and function linked with peach fruit ripening.     

1-甲基环丙烯采后处理对樱桃番茄果实成熟过程的影响

研究了不同浓度(0、0.035、0.07和0.11μL/L)的乙烯受体竞争性抑制剂1-甲基环丙烯(1-MCP)采后处理对绿熟期樱桃番茄的乙烯合成、果实软化、果实色素(叶绿素、茄红素、β-胡萝卜素)含量消长的影响.0.07 μL/L及其以上浓度的1-MCP降低了前期乙烯合成,同时推迟了乙烯释放高峰,但0.035 μL/L浓度的1-MCP处理并不能抑制内源乙烯合成.1-MCP显著延迟了果实软化和叶绿素降解,但并不影响这两个过程的启动.茄红素合成的启动和积累均受到了1-MCP抑制,而1-MCP并不推迟β-胡萝卜素合成的启动,只抑制其积累.这些结果提示了乙烯调节成熟生理过程的不同机制.对于绿熟期的樱桃番茄,0.07～0.11μL/L的1-MCP是实用的有效处理浓度.1-MCP有效浓度可能用于了解果实的乙烯受体水平和乙烯敏感性.     

Inhibition of the ethylene response by 1-MCP in tomato suggests that polyamines are not involved in delaying ripening, but may moderate the rate of ripening or over-ripening

Ethylene initiates the ripening and senescence of climacteric fruit, whereas polyamines have been considered as senescence inhibitors. Ethylene and polyamine biosynthetic pathways share S-adenosylmethionine as a common intermediate. The effects of 1-methylcyclopropene (1-MCP), an inhibitor of ethylene perception, on ethylene and polyamine metabolism and associated gene expression was investigated during ripening of the model climacteric fruit, tomato (Solanum lycopersicum L.), to determine whether its effect could be via polyamines as well as through a direct effect on ethylene. 1-MCP delayed ripening for 8 d compared with control fruit, similarly delaying ethylene production and the expression of 1-aminocyclopropane-1-carboxylic acid (ACC)-synthase and some ethylene receptor genes, but not that of ACC oxidase. The expression of ethylene receptor genes returned as ripening was reinitiated. Free putrescine contents remained low while ripening was inhibited by 1-MCP, but increased when the fruit started to ripen; bound putrescine contents were lower. The activity of the putrescine biosynthetic enzyme, arginine decarboxylase, was higher in 1-MCP-treated fruit. Activity of S-adenosylmethionine-decarboxylase peaked at the same time as putrescine levels in control and treated fruit. Gene expression for arginine decarboxylase peaked early in non-treated fruit and coincident with the delayed peak in putrescine in treated fruit. A coincident peak in the gene expression for arginase, S-adenosylmethionine-decarboxylase, and spermidine and spermine synthases was also seen in treated fruit. No effect of treatment on ornithine decarboxylase activity was detected. Polyamines are thus not directly associated with a delay in tomato fruit ripening, but may prolong the fully-ripe stage before the fruit tissues undergo senescence.