Effects of ethylene on the postharvest quality of inflorescences of Oncidium varicosum ‘ Samurai ’

Oncidium varicosum is an orchid largely distributed in South America, whose inflorescences with bright yellow flowers are of rare beauty. One of the objectives of the postharvest physiology of flowers is the study of the factors related with the quality loss of cut flowers. The ethylene performs a very important function on the senescence of flowers because it induces abscission of flowers buds and open flowers causing precocious wilting and loss of quality. The sensitivity to ethylene has been reported as variable in function of floral species, exposition period and regulator concentration. The present research evaluated the postharvest of cut inflorescences of O. varicosum ‘Samurai’, after application of ethylene (Ethrel: 1, 10 and 100 L L -1 ). The experiment was performed as complete randomized design with factorial scheme composed of two factors: four postharvest treatments and four dates of evaluation. The inflorescences were maintained in the refrigerated ambient. The results demonstrated that the exposition to ethylene caused physiological alterations such as reduction of relative water content of the flowers, decrease of the soluble carbohydrates contents of the petals, increase of the respiration rate and flowers abscission. The concentration of 100 L L -1 of Ethrel reduced the decorative life of flowers in seven days in relation to the control without product application, while at 1 μL L -1 , the flowers presented lower sensitivity to product, with results similar to the control.


INTRODUCTION
The orchids of genus Oncidium are economically important and contains more than 750 species.Most of these species grow in South America, as 105 species of the orchid genera are found in Brazil, being the genus hybrids for cutting are commercialized.
Ethylene is a simple molecule composed of two carbon atoms symmetrically linked by a double bond and it naturally occurs in gaseous form.It is, furthermore, a plant growth regulator involved in the regulation of a wide range of different physiological processes, including germination, growth, floral initiation and opening, leaf and floral senescence as well as organ abscission and fruit ripening (Yoo et al., 2009).
There are significant differences in ethylene sensitivity between species and even cultivars of the same species (Serek et al., 2006b;Scariot et al., 2008).The response and sensitiveness to ethylene are different according to the growth stage, variety and the plant organ perception (Ciardi and Klee, 2001;Jones et al., 2001).For example, petals of orchids (Phalaenopsis) and Hibiscus (Çelikel and Reid, 2002), wilt in response to ethylene, in other species, such as wax flower (Chamelaucium uncinatum) (Macnish et al., 2000), ethylene induces petal or flower abscission.Huang et al., (2007) reported through the gene expression standard analysis of ethylene receptor in Oncidium "Gower Ransey" inflorescences, pointing out that their senescence is associated with the increase of ethylene receptors.
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 (Scariot et al., 2014).Among the main biochemical and metabolic alterations described during the orchid flower senescence are related the anthocyanin synthesis and breakdown; ethylene production; increase of leaf respiration; RNA synthesis; synthesis and activation of various enzymes; transport of organic and inorganic substances; hydrolysis of structural and reserve molecules (Serek et al., 2006a).Lin (1999) observed that the shelf life of Oncidium gender is reduced after harvest, and this is more pronounced due to its branched-inflorescence, which exposes the flowers to damages.Huang and Paull (2009) observed that for Oncidium "Gouver Ramsey" orchid, the floral buds were more sensitive to the exogenous ethylene than the proper flower.The present study aimed to evaluate the ethylene application effect over the postharvest conservation of O. varicosum "Samurai" orchid inflorescences.

MATERIALS AND METHODS
O. varicosum ("Samurai") inflorescences were taken in the morning when 30% of the orchids were open, in a commercial production area in Atibaia (SP).Subsequently, each stalk was placed in a closed plastic tube with 10 ml of distilled water and was wrapped in micro-perforated plastic.The transport of the flowers to the Vegetal Physiology Laboratory at FCAV-UNESP (Jaboticabal, São Paulo) was completed using a refrigerated vehicle.Once in the laboratory, the floral stalks were standardized, labeled, weighed and sprayed with Ethrel (2-chloroethylphosphonic) at concentrations of 0 μL L -1 (control treatment), 1, 10 and 100 μL L -1 .Two applications were done in 30 min interval.
Experimental delineation was entirely random, in a factorial scheme formed by two factors including the following: four postharvest treatments and four evaluation dates.Three repetitions for each treatment were used, with three inflorescences in each repetition.The inflorescences were kept in a refrigerated environment at 20±1.1°C (88% HR), and lighted for 12 h.Every four days it was evaluated: i) Relative water content: obtained from 10 flowers in each repetition.Flowers were weighed and immersed in distilled water (to be hydrated) during 4 h.Subsequently, the orchids were weighed for a second time and were taken to a drying stove at 70°C (Kramer, 1983).ii) Color evaluation of the inflorescences: completed using a colorimeter (Minolta CR-400b), which determined the following values: L (100 = white; 0 = black), a* (positive = red; negative = green), and b* (positive = yellow; negative = blue).Chromaticity was calculated using equations according to the Konica (2007).iii) Soluble carbohydrates of flowers from Oncidium inflorescences: extracted in ethanol using a phenol-sulfuric method (Robyt and White, 1987).iv) The amount of CO2 (respiratory activity) produced: quantified by a gas analyzer (model PBI-DANSENSOR 9900) when the samples were removed from the atmosphere inside the containers.
The vase life was evaluated in a separate batch of inflorescences and it was considered finished when 50% or more flowers in a specific inflorescence lost their ornamental quality.The obtained data were submitted to variance analysis through an F test, and the averages were compared using a Tukey test (P≤ 0.05), in which the differences between the two treatments were considered significant when they were higher than the sum of two standard deviations (Shamaila et al., 1992).

RESULTS AND DISCUSSION
After 8 days of vase life (DVL) the Ethrel at the concentration of 100 μL L -1 proportioned the greater reduction of relative water content (RWC) (77.57%) (Figure 1).At 11 DVL there was an intense and significant RWC reduction (48.93%) in the inflorescences maintained at the concentration of 10 μL L -1 of Ethrel , what points out that treatments even with concentrations 10 times lower can reduce the O. varicosum 'Samurai' orchid shelf life.The water balance alterations after exogenous ethylene application are related with the first signs for flower senescence (Goliás and Kobza, 2003).
In the Figure 2 is observedthat a reduction of the inflorescence brightness during the evaluation period, especially in the treatment at 100 μL L -1 . The Ethrel spray in the 100 μL L -1 made the inflorescences browning what was statistically significant at 4 and 8 DVL.The flower browning is a senescence indicative and may be related to water stress and low carbohydrate levels, caused by the phenol oxidation, mainly the leuco-anthocyanin, which react with other cell compounds producing brown precipitate (Reid, 2002).In hybrids of Dendrobium the discoloration of the flowers it was also related to a possible increase in the pH of the vacuole of the cells or to the effect of the polyphenol oxidase and peroxidase  , 24 h) (Almasi et al., 2012).The obtained results for the chromaticity parameter showed that the color intensity was drastically reduced in the highest Ethrel concentration treatment (100 μL L -1 ) at 8 DVL, which differed from the others.It is observed in the Figure 3 a significant reduction of the soluble carbohydrate content in the inflorescence along the days of vase life.The reduction of soluble carbohydrate content in the inflorescences is an indicative of vegetal senescence, when using its carbon reserves to maintain the respiration to repair the degrading processes of the metabolism.At 4 DVL, the inflorescences sprayed with Ethrel in the 100 μL L -1 concentration presented soluble carbohydrate content in the petals of 12.58 g of glucose per 100 g -1 of fresh mass, and more than 50% of the flowers old, culminating in a shelf life end.Orchid flowers are particularly sensitive to ethylene (Halevy and Mayak, 1981), as the ethylene production is an autocatalytic process, greater quantities of this substance can be produced once they are put together with other senescent flowers (Hew, 1994).The Ethrel concentrations of 1 and 10 μL L -1 and control presented soluble carbohydrate content similar to those from 11 DVL, what can indicate that the exposition to ethylene concentrations of 1 and 10 μL L -1 , for the experimental conditions, were less damaging.Arditti (1992) related that the carbohydrate content is variable among the different genders and species of orchid flowers.Aranda orchid generally maintain a high level of sugars in the sepals, while in Cymbidium, the sugar content decreases with age.Huang and Paull (2009) found that flower buds of Oncidium "Gouver Ramsey" are extremely sensitive to ethylene.The vase live of flower buds was significantly reduced by their exposure to 0.03 nL L -1 ethylene concentration.The buds were more sensitive than the flowers, since it was necessary five days to the ethylene treatment result in any effect on buds, and seven days to the flowers.
It is verified in the Figure 4 that the inflorescences sprayed with Ethrel in the concentration of 100 μL L -1 presented an intense respiration peak (265 mg of CO 2 kg -1 h -1 ) at 4 DVL, sharp decline at 8 DVL which coincided with the end of the flower longevity.These results agree with those published by Chadwick et al. (1980) and Harkema and Woltering (1982), saying that the exposure to high ethylene concentration may cause early flower senescence in orchids, and also flower buds, flowers and leaves abscission and early wilting.The application of the exogenous ethylene (10 μL L -1 , 24 h) in Dendrobium hybrids showed distinct variances in ethylene sensitivities and degrees of deterioration.The hybrids in the sensitive group exhibited the utmost hyponasty, weight loss and degradation of anthocyanin content in sepals and petals (Almasi et al., 2012).
For Oncidium "Goldiana" flowers, the ethylene production started after a latent period of 100 hours post harvesting and an increase in the climacteric period that formed a peak at 11 days (Hew and Yong, 2004).Ketsa and Thampitakorn (1995)  of 70% of flower buds.
The concentration of 100 μL L -1 led to a greater physiological response of the flowers, causing a pronounced reduction of the relative water content, respiratory rate increase, soluble carbohydrate content reduction, flower browning, intense flower drops and longevity reduction.

Conclusion
Oncidium varicosum "Samurai" presented a varied sensitivity to different exogenous ethylene concentrations (1, 10 and 100 μL L -1 ).The concentration of 100 μL L -1 of Ethrel reduced the decorative life of flowers in seven days in relation to the control without product application, while at 1 μL L -1 , the flowers presented lower sensitivity to product, with results similar to the control.

FWFigure 3 .
Figure 3. Contents of soluble carbohydrates (g of glucose 100 g -1 FW) of cut inflorescences of O. varicosum "Samurai" treated with Ethrel.Data shown are mean ± standard deviation.

Figure 4 .
Figure 4. Respiratory activity of cut inflorescences of O. varicosum "Samurai" treated with Ethrel.Data shown are mean ± standard deviation.