Expression of a lipoxygenase encoding gene (BoLOX1) during postharvest senescence of broccoli

Lipoxygenases (LOX) belong to a large family of plant enzymes that catalyze the hydroperoxidation of polyunsaturated fatty acids. Most of them are expressed during senescence and contribute to membrane deterioration and biosynthesis of jasmonic acid, a known senescence enhancer. In this work, we cloned a fragment of a gene encoding a LOX from broccoli (BoLOX1). The analysis of the sequence revealed that BoLOX1 is closely related to other LOX from higher plants. Furthermore, we analyzed the expression of BoLOX1 and detected a larger increase during postharvest senescence. A slight increase of total lipoxygenase activity was also found during senescence. In other sets of experiments, broccoli heads were treated with plant hormones, such as cytokinin and ethylene, as a way to assess the effect of such compounds on the expression of BoLOX1. Cytokinin treatment delayed the increase of BoLOX1 expression and lipoxygenase activity whereas ethylene accelerated both processes. Also, several postharvest treatments were applied in order to delay senescence in broccoli florets and to evaluate their effects on BoLOX1 expression. Samples treated with modified atmosphere, hot air, UV-C or white light showed a delay in chlorophyll degradation and degreening. In most cases, the treatments also delayed the increase of BoLOX1 expression, reaffirming the relationship between the expression of this gene and senescence. However, treatments like modified atmospheres and visible light markedly increased lipoxygenase activity, which suggests a lack of correlation between BoLOX1 expression and lipoxygenase activity.     

1-MCP处理对采后青花菜内源激素变化的影响

以青花菜品种"曼陀绿"(Brassica olracea L. var. italica)为试材,研究了1-MCP处理对青花菜采后叶绿素含量、乙烯释放速率和内源玉米素(ZR)、赤霉素(GA)、生长素(IAA)、脱落酸(ABA)含量的影响。结果表明：1-MCP延缓了叶绿素降解,降低了组织对乙烯的敏感性,延缓了GA的分解,推迟了ABA的积累。经1-MCP处理的青花菜ABA/GA和ABA/IAA比值上升缓慢。     

Effects of postharvest ethanol vapor treatment on activities and gene expression of chlorophyll catabolic enzymes in broccoli florets

Postharvest yellowing of broccoli (Brassica oleracea L. Italica Group) is an important indicator of quality deterioration and occurs with chlorophyll (Chl) degradation. Postharvest ethanol vapor treatment could delay the yellowing of broccoli florets, through the suppression of Chl degradation. As the first step in identifying the mechanism of this delay, the effects of postharvest ethanol vapor treatment on activities and gene expression of the Chl catabolic enzymes in broccoli were determined. Broccoli branchlets were placed in a perforated polyethylene bag with or without (control) an ethanol pad and stored at 20 °C in darkness. The Chl contents of the control broccoli florets decreased remarkably after three days in storage, whereas the contents of the ethanol-treated broccoli showed no significant changes except at day one. Changes in chlorophyllase activity in the ethanol-treated broccoli were similar to those of the control until three days in storage, but then the activity tended to decease. Mg-dechelatase and Chl-degrading peroxidase activities, which increased greatly with senescence in the controls, remained unchanged in the ethanol-treated broccoli. BoCLH1 expression showed changes almost similar to those of chlorophyllase activity. BoPAO expression in the control broccoli increased greatly at day one in storage and was maintained at a high level until three days. In contrast, this expression in ethanol-treated broccoli was suppressed until two days. BoRCCR expression in the control broccoli increased until two days in storage, while the expression in the ethanol-treated broccoli showed no change during storage. These results show that postharvest ethanol vapor treatment suppressed the activities and gene expression of Chl catabolic enzymes, resulting in delayed yellowing of broccoli florets.     

UV-C treatment delays postharvest senescence in broccoli florets

Central broccoli heads (cv. de Cicco) were harvested and treated with UV-C light (4, 7, 10, or 14 kJ m⁻²). All treatments delayed yellowing and chlorophyll degradation at 20 °C but the irradiation dose of 10 kJ m⁻² allowed retaining the highest chlorophyll content yet had lower amounts of pheophytins than every treatment other than 7 kJ m⁻². This dose was selected to analyze the effect of UV-C on postharvest broccoli senescence at 20 °C. The UV-C treatment delayed yellowing, chlorophyll a and b degradation, and also the increase in pheophytins during storage. The activity of chlorophyll peroxidase and chlorophyllase was lower in UV-C treated broccoli. Instead, Mg-dechelatase activity increased immediately after the treatment, but after 4 and 6 d this activity was lower in UV-C treated florets than in controls. Treated broccoli also displayed lower respiration rate, total phenols and flavonoids, along with higher antioxidant capacity. The results suggest that UV-C treatments could be a useful non-chemical method to delay chlorophyll degradation, reduce tissue damage and disruption, and maintain antioxidant capacity in broccoli.     

1-Methyl cyclopropene extends postharvest life of spinach leaves

Senescence of detached spinach leaves either untreated or treated with 0.1 or 1.0 μL L^?1 1-MCP has been investigated. 1-MCP treated leaves had higher chlorophyll content and photosystem II potential quantum yield (Fv/Fm) and lower solute leakage than untreated leaves after storage in darkness at 23 °C for 6 d, indicating a delay of senescence. Ethylene production was increased in spinach supplemented with 1-MCP after 3 d storage and then declined to the rates of untreated leaves. 1-MCP treated spinach had higher ascorbic acid and glutathione concentrations, and a low oxidised/reduced ratio for both antioxidants. Accumulations of ammonium and protein degradation were reduced by 1-MCP. The results presented here indicate that inhibition of ethylene sensitivity can be successfully used to extend the postharvest life of spinach leaves.     

1-MCP对果实采后生理变化的影响

对1-MCP作用效应的影响因素进行阐述,同时从果实风味、硬度、可滴定酸、可溶性固形物、果皮叶绿素、采后生理病害、乙烯生成和呼吸速率等几个方面描述1-MCP对果蔬采后贮藏保鲜的作用效果.     

Ethylene control in cut flowers: Classical and innovative approaches

Ethylene-mediated premature floral senescence and petal or flower abscission affect postharvest longevity of several species used as cut flowers. Exposure to exogenous or endogenously produced ethylene can be controlled in several ways. These include the use of ethylene biosynthesis inhibitors or ethylene action inhibitors, and ethylene removal technologies. In addition, genetic modification can be very effective in controlling ethylene synthesis and perception. We review here the potential for applications of nanotechnology to control ethylene levels and postharvest management in the flower industry. Already, nanosponges have been shown to enhance efficacy of the ethylene inhibitor 1-MCP in several flower species. In carnation, 1-MCP included in nanosponges also allowed better control of Botrytis cinerea damage. However other applications are also considered based on successes in the use of this technology to increase agricultural production and decrease postharvest waste. Nano-metal based sensors could be used for detection of ethylene in the store and to label the product along the distribution chain. Furthermore, nanocomposites could be included as scavengers for ethylene removal in active packaging, and nanocatalysts could promote ethylene catalytic degradation in the warehouse. Nanoparticles could also be introduced into a new generation of packaging to control effects of gases and UV, and increase strength, quality and packaging appearance. This review highlights recent results on the use of nanotechnology sensu lato and potential application for cut flower vase life improvement, focusing on ethylene control strategies.     

The effect of CO2 and 1-methylcyclopropene on the regulation of postharvest senescence of mint,Mentha longifolia L.

The regulatory effects of 5 kPa CO₂ and of the ethylene action inhibitor, 1-methylcyclopropene (1-MCP) at 0.5 μmol/l on the senescence of harvested mint, Mentha longifolia L. were assessed. Visual parameters of senescence including yellowing, browning, decay and leaf abscission were recorded and scored on scales linking the onset and progression of senescence to marketability. The effects of plant age on the rate of postharvest senescence and on the efficacy of the CO₂ and 1-MCP treatments were also investigated. All experiments were repeated with and without the presence of exogenous ethylene. Two experimental formats were used, with 6 days storage at room temperature representing local market conditions, and 6 days cold storage at 1.5 °C followed by 4 days at room temperature representing export market conditions. Sprigs from old plants were no longer of marketable quality after 6 days storage at room temperature. Exogenous ethylene accelerated the onset of senescence causing unacceptably high rates of leaf abscission. Raised levels of CO₂ in a controlled atmosphere system were found to be more effective in inhibiting senescence without the presence of exogenous ethylene than pre-treatment with 1-MCP, and no additive effect was found. However in the presence of exogenous ethylene, a combined treatment with 1-MCP together with raised CO₂ levels resulted in a significant additive effect in nullifying the ethylene-induced leaf abscission. Respiration rates as measured by CO₂ production, and ethylene production, were recorded throughout all experiments. While CO₂ levels were not affected by any experimental treatment, ethylene production was elevated in mint sprigs exposed to an initial dose of gaseous 1-MCP, and was further increased under a combined treatment of 1-MCP together with 5 kPa CO₂. However in the presence of exogenous ethylene, CO₂ strongly suppressed the 1-MCP induced ethylene production.     

New degreening treatments to improve the quality of citrus fruit combining different periods with and without ethylene exposure

In the Mediterranean area, early-season citrus fruit reach acceptable internal maturity standards for marketing, while the fruit are still green. A degreening treatment is widely used as a postharvest practice to improve the external color. Nevertheless, the application of ethylene during this treatment can be associated with calyx senescence. The aim of this study was to evaluate new degreening treatments in order to reduce this disorder. ‘Clemenules’ mandarins and ‘Navelina’ oranges, harvested at different external colors, were submitted to different degreening treatments, combining periods with and without ethylene exposure. In both cultivars, the periods without ethylene exposure during degreening reduced the incidence of calyx disorders. To obtain a typical ‘Clemenules’ variety color with the lowest incidence of calyx alteration, the optimum degreening treatment was based on 72 h with ethylene and 48 h without ethylene when fruit were harvested with a color index of between −9 and −3. On the other hand, a treatment of 24 h with ethylene plus 48 h without ethylene is recommended for fruit with an external color index higher than −2. For ‘Navelina’ oranges, highly sensitive to calyx senescence during degreening, a treatment consisting of 24 h with ethylene plus 48 h without ethylene is recommended, which involves harvesting the fruit with a color index of over 0.     

Effects of ethylene, pollination, and ethylene inhibitor treatments on flower senescence of gentians

Flower senescence of the potted gentian (Gentiana scabra) ‘Shinbisei’ was investigated in relation to ethylene sensitivity and production. ‘Shinbisei’ flowers were used for all experiments except for those with inflorescences. Exposure to ethylene at 0.5 μL L⁻¹ or higher concentrations for 24 h markedly accelerated flower senescence, indicating that G. scabra flowers are highly sensitive to ethylene. Treatment with 0.2 or 0.5 mM silver thiosulfate complex (STS) and 2 μL L⁻¹ 1-methylcyclopropene (1-MCP), ethylene action inhibitors, and 50 mM α-aminoisobutyric acid, an inhibitor of 1-aminocyclopropane-1-carboxylate (ACC) oxidase, did not delay flower senescence. However, treatment with 1 mM l-α-(2-aminoethoxyvinyl) glycine, an inhibitor of ACC synthase, slightly delayed flower senescence. Pollination significantly accelerated petal senescence of G. scabra flowers. Ethylene production of petals, gynoecium, and stamens in unpollinated flowers slightly increased during senescence. Pollination significantly increased ethylene production of petals, gynoecium and stamens 1 day after pollination. To clarify whether 1-MCP delays senescence of cut gentian inflorescences, cut G. scabra ‘Yuki-hotaru’, G. scabra × Gentiana triflora ‘Aoi-kaze’, and G. triflora ‘Koharu’ inflorescences with various stages of flowers, including buds with colored petals, were treated with 2 μL L⁻¹ 1-MCP for 24 h. 1-MCP treatment delayed flower wilting of cut inflorescences of ‘Aoi-kaze’ and ‘Yuki-hotaru’ more than that of ‘Koharu’, suggesting that there is species variation in the effect of 1-MCP in delaying flower senescence of cut gentian inflorescences.     

UV-C复合热处理或1-MCP处理对西兰花保鲜效果的研究

以采后西兰花为试材,研究UV-C与热处理、1-MCP单独或复合处理对西兰花采后冷藏及货架期间品质的影响。结果表明,在冷藏(0℃)前,采用UV-C照射与热处理、1-MCP单独或复合处理,对西兰花在冷藏65 d和货架5 d期间的感官品质和营养成分均具有较好的保鲜效果。其中以0.25μL/L的1-MCP熏蒸20 h及复合UV-C照射600 s对西兰花的护绿、抑制花蕾开放以及保持VC和叶绿素含量的效果显著好于其他处理,感官品质评价总分较高;45℃热水处理5 min及复合UV-C照射处理对控制西兰花腐烂率的效果较好。     

Pheophytinase activity and gene expression of chlorophyll-degrading enzymes relating to UV-B treatment in postharvest broccoli (Brassica oleracea L. Italica Group) florets

Pheophytinase (PPH) activity and gene expression of chlorophyll (Chl)-degrading enzymes relating to UV-B treatment in postharvest broccoli (Brassica oleracea L. Italica Group) florets were determined. PPH is involved in the dephytylation of Mg-free Chl a, pheophytin (Phy) a. However, in vitro chlorophyllase (Chlase, EC.3.1.1.14) also uses Phy a as a substrate to produce pheophorbide (Pheide) a by dephytylation. For an accurate determination of PPH activity, the PPH protein fraction was separated from Chlase protein by ammonium sulfate precipitation. The protein precipitated by 45-60% saturated ammonium sulfate included a little bit of Chlase activity and was suitable for PPH determination. PPH activity in broccoli florets treated with a UV-B dose of 19 kJ m⁻² was repressed for the first 2 d of storage at 15 °C, whereas it increased gradually with senescence of control broccoli florets. The expression level of BoCLH1 was reduced in broccoli florets on day 4 of storage, while BoCLH2 and BoCLH3 were up-regulated with UV-B treatment. A high BoPAO expression level was found in senescent broccoli florets, and the up-regulation of this gene was delayed by UV-B treatment. The highest expression level of BoPPH was found in the control, and its expression was clearly repressed by UV-B treatment on day 2 of storage. We suggest that the up-regulation of Chl-degrading enzyme genes could be delayed by UV-B treatment, resulting in the suppression of floret yellowing in stored broccoli.     

1-Methylcyclopropene (1-MCP)-induced protein expression associated with changes in Tsai Tai (Brassica chinensis) leaves during low temperature storage

Leaf yellowing is an early symptom of senescence and greatly affects the commercial value of leafy vegetable products. The ethylene action inhibitor 1-methylcyclopropene (1-MCP) has been widely studied regarding its effects on senescence, but little is known on how it influences the protein expression profile of leafy vegetables in storage. In this study, by using a proteomic approach, changes in proteomic profile induced by 1-MCP in Tsai Tai (Brassica chinensis) leaves during storage under low but non-freezing temperature were investigated. Compared with the control, 1-MCP treatment inhibited leaf etiolation and reduced weight loss of Tsai Tai stored at 1 °C, indicating that it delayed senescence. Two dimensional electrophoresis revealed 16 differentially accumulated protein spots, 12 of which were successfully identified by mass spectrometry. A comparison between the control and the 1-MCP-treated Tsai Tai showed that 1-MCP enhanced levels of four proteins involved in photosynthesis, i.e. photosystem II protein, oxygen-evolving enhancer protein 2, chloroplast ribulose 1,5-bisphosphate carboxylase/oxygenase activase and high molecular weight RuBisCO, inhibited malate dehydrogenase, a key enzyme involved in the tricarboxylic acid cycle, downregulated gibberellin 2-beta-dioxygenase expression, a key enzyme involved in biosynthesis of active gibberellins, and suppressed glutathione s-transferase, an important scavenger for reactive oxygen species. These results imply that 1-MCP could maintain leaf capacity for carbon assimilation, inhibit the tricarboxylic acid cycle, induce biosynthesis of GAs and reduce levels of reactive oxygen species, which together may have contributed to inhibition of leaf yellowing and delay of senescence.     

1-MCP regulates ethylene biosynthesis and fruit softening during ripening of ‘Tegan Blue’ plum

To investigate the effects of postharvest application of 1-MCP on ethylene production and fruit softening, activities of ethylene biosynthesis and fruit softening enzymes were measured during postharvest ripening of plum (Prunus salicina Lindl. cv. Tegan Blue) fruit after being exposed to 1-MCP (0, 0.5, 1.0 or 2.0 μL L⁻¹) at 20 ± 1 °C for 24 h. Following the treatments, fruit were allowed to ripen at ambient temperature (20 ± 1 °C), and ethylene production in fruit, activities of ACS and ACO, ACC content and fruit softening enzymes (PE, EGase, exo-PG and endo-PG) in fruit skin and pulp were recorded at different intervals. Postharvest application of 1-MCP significantly delayed and suppressed the climacteric ethylene production with reduction in the activities of ethylene biosynthesis enzymes (ACS, ACO) and ACC content, and fruit softening enzymes (PE, EGase, exo-PG and endo-PG) in the skin as well as in pulp tissues. The reduction was more pronounced with increased concentrations of 1-MCP. 1-MCP treated fruit showed different rates of fruit softening and activities of ethylene biosynthesis enzymes in the skin and pulp tissues which warrant further investigation on regulation of gene expression related to these enzymes with the inhibitory effect of 1-MCP.     

Postharvest application of a new growth regulator reduces calyx alterations of citrus fruit induced by degreening treatment

Synthetic auxins, specially 2,4-dichlorophenoxyacetic acid (2,4-D), have been used as a postharvest treatment to retard calyx abscission, drying and browning, which occur as the result of degreening processes. Nevertheless, the use of 2,4-D is restricted by European Union (EU) legislation, which means that finding an alternative to this auxin in controlling calyx senescence is necessary. In the present research, the effect of postharvest application of a new growth regulator, S-ethyl-4-chloro-O-tolilooxytioacetate (HF-Calibra^®), currently registered for preharvest use, was used to investigate calyx senescence of early-season citrus varieties (‘Oronules’, ‘Clemenrubi’ and ‘Clemenules’ mandarins and ‘Navelina’ oranges) submitted to a commercial degreening treatment. After shelf-life, application of HF-Calibra^® treatment at different concentrations significantly reduced calyx changes. Although auxin treatments can retard color change, this delay had no negative effect from a commercial viewpoint since all the studied varieties reached a commercially acceptable color. The application of HF-Calibra^® at 10, 20, 40 or 60 mL/L affected neither weight loss nor sensory quality of the fruit (firmness, total soluble solids, titratable acidity, acetaldehyde and ethanol content and flavor). Postharvest application of HF-Calibra^® could be an alternative to 2,4-D to avoid calyx senescence of citrus fruit induced by a degreening treatment and to maintain fruit quality.     

Expression of genes encoding xyloglucan endotransglycosylase/hydrolase in ‘Saijo’ persimmon fruit during softening after deastringency treatment

Persimmon (Diospyros kaki Thunb.) fruit undergoes intensive cell wall modification during postharvest fruit softening. Xyloglucan metabolism is important in cell wall disassembly. We cloned cDNAs for two xyloglucan endotransglycosylase/hydrolase genes (DkXTH1 and DkXTH2) from ‘Saijo’ persimmon fruit treated with dry ice to remove astringency. In order to determine the ethylene dependence of XTH gene expression, fruit were exposed to 1-methylcyclopropene (1-MCP), an inhibitor of ethylene action, prior to removal of astringency. Ethylene production increased in mature control and 1-MCP-pretreated fruit after dry-ice treatment, and flesh firmness decreased to the same extent during dry-ice treatment in the control and 1-MCP-pretreated fruit. After dry-ice treatment, control fruit softened completely, but fruit firmness was maintained in 1-MCP-pretreated fruit. Accumulation of DkXTH1 mRNA was induced simultaneously with commencement of ethylene production in mature control fruit. Pretreatment with 1-MCP delayed accumulation of DkXTH1 mRNA. DkXTH2 expression also coincided with fruit softening but was intensified by 1-MCP treatment during the deastringency treatment. These results indicate that fruit softening was related to both DkXTH1 and DkXTH2 expression in ‘Saijo’ persimmons.