Introduction

The main factors that degrade flower quality and reduce vase life are relative humidity (RH), ethylene, wet or dry storage, and bacteria. Carnations and lily flowers are sensitive to ethylene, while bacteria are the main factors affecting the vase life of cut roses and gerbera flowers (Hoogerwerf et al. 1989). The presence of bacteria obstructs the stem xyle vessels and thereby decreases the water supply rate to the flowers (van Doorn et al. 1989). A number of bacteria are often present in the vase solutions used by growers, wholesalers, retailers, and consumers to hydrate cut flowers (Macnish et al. 2008). The vase life of cut gerbera flowers is often shortened due to bending of the scape, which precedes wilting of the ray petals (Zhong et al. 2012). Cut gerberas produced in Korean farms are fixed with a metal pin about 20 cm length from capitulum and then taped to prevent scape breaking and bending if the water supply is suspended after harvesting (Yoon et al. 1996). These required tasks increase in the material and labor costs. Consequently, any improvement in the production skills of gerberas and in the efficacy of the applied floral preservatives that prevents scape bending will be beneficial. The commercially available floral preservatives such as silver nitrate, aluminum sulfate, 8-hydroxyquinoline sulfate, and sodium hypochlorite are effective in extending the vase life of cut gerberas (Steinitz 1984). But the residues from these biocides were injurious to environment and human health (Damunupola and Joyce 2006).

On the other hand, chlorine dioxide (ClO₂) which is a greenish-yellow gas and is a single-electron-transfer oxidizing agent with a chlorine-like odor, is regarded as relatively safe as evidenced by its approval for use to sanitize surfaces of fruits and vegetables (US FDA 1998). ClO₂ has been recognized since the beginning of the century for its disinfecting properties; and has been approved by the US EPA for many applications including the widespread use of ClO₂ in the treatment of drinking water. Beyond this and numerous other aqueous applications, the sporicidal properties of gaseous ClO₂ were demonstrated in 1986 (Rosenblatt et al. 1987). Subsequent to these initial studies, it has been shown that gaseous CD is a rapid and effective sterilant active against bacteria, yeasts, molds, and viruses (Rosenblatt et al. 1987). There are previous reports that ClO₂ reduces significantly the number of bacteria on surfaces of vegetables and fruits (Pao et al. 2007) and that it removes or sanitizes the odor of drinking water (Aieta and Berg 1986). During vase life of cut roses, bacteria inhibition by ClO₂ was very effective for both of holding solution and pulsing solution, but the ClO₂ contents in holding solution for cut rose flowers were scavenged in only two to four days after treatment, meaning that environmental injurious residue materials did not remain in the holding solution after postharvest treatment (Lee and Kim 2014). Therefore, this study was conducted to know if ClO₂ can be applied to extend the postharvest life of cut gerbera flowers as existing preservatives.

Materials and Methods

Plant materials

The gerbera ‘Jenny’ investigated in this research was originally grown at a farm located in Miryang, Korea. It had been harvested before the experimentation and transferred to our laboratory. Gerbera jamesonii ‘Jenny’ was first bred in Korea, 2012. Gerbera ‘Jenny’ is characterized as a large double flower (12 cm in diameter) with red-ray and yellow-disk florets and mean peduncle length of 56 cm. In this study the cut gerbera flowers of 9-11 cm in floral diameter and 45 cm above in peduncle length were used as materials.

Experimental design

Expt. 1. The first experiment was conducted to investigate the applicability of ClO₂ as floral preservatives for cut gerbera flowers. Besides ClO₂, five commercialized biocides, namely, aluminum sulfate, silver nitrate, AVB, sodium hypochlorite, and 8-hydroxyquinoline sulfate were selected for treatment through a literature review (Jones and Hill 1993; Macnish et al 2008). The each peduncle was randomized for soaking in one of the following treated holding solutions: tap water; distilled water; ClO₂: 10 μL • L⁻¹; AVB (Chrysal, Netherlands): 1500 mg • L⁻¹; sodium hypochlorite: 50 mg • L⁻¹; silver nitrate: 10 mg • L⁻¹; aluminum sulfate: 500 mg • L⁻¹, and 8-hydroxyquinoline sulfate: 200 mg • L⁻¹ (Fig. 1). The experiment was run for 4 weeks from 27 February 2013 under the indoor conditions of air temperature of 20.1 ± 2℃, relative humidity (RH) of 47.8%, and light intensity of 16.2 μmol • m^-2s^-1 PPFD for 12 hours under the fluorescent lighting.

Expt. 2. The second experiment was conducted to identify the optimum concentration in ClO₂ as floral preservatives, where farm groundwater was tried as the control instead of tap water or distilled water. The peduncles were randomized for soaking in one of the following treated holding solutions: farm groundwater (control) and ClO₂ 2, 5, and 10 μL • L⁻¹. The experiment was run for four weeks from 19 April 2013 under the indoor conditions of 22.8 ± 5℃, RH of 52.1%, and light intensity of 16.3 μmol • m⁻²s⁻¹ PPFD for 12 hours under the fluorescent lighting.

In the experiments, each holding solution was filled 500 mL per bottle for just one peduncle and not replenished during experiment. 10 peduncles were repeated for each treatment.

Vase life and bacteria detection

In the experiments, vase life was recorded as the time (days) after treatment (day 0), when flowers started showing symptoms of ray petal wilting or curling, peduncle bending (≥ 90°) or breaking (Geraspolus and Chebli 1999), based on a definition for the vase life of cut flowers as extending until the ornamental value of the cut flower begins to depreciate (Park et al. 2004). The ClO₂ concentrations during two experiments were measured with a chlorine dioxide analyzer (Chlorine Dioxide HI 93738 ISM, Hanna instruments Inc., Woonsocket, USA). In the second experiment, the ClO₂ concentration was measured in the condition of peduncle presence or not. On day 6 after treatment, the number of bacteria in each holding solution was measured by using the dilution plate technique with nutrient agar (NA) medium. The NA medium was cultured for 3 days at 28℃ and the number of bacteria was counted per 24 hours (van Doorn et al. 1989). The uptake of holding solution by a flower and the fresh weight of a flower were measured every two days interval.

Statistical analysis

Data were analyzed separately for each extraction by a multifactor analysis of variance (ANOVA) using version 9.2 of the SAS statistical software package (Systat 9.2, Systat Software Inc., Richmond, USA). Duncan’s Multiple Range Test (DMRT P = 0.05) was used for comparisons of treatment means.

Results and Discussion

Applicability of ClO2 as floral preservatives

The vase life of cut gerbera flowers differed significantly according to the holding solution and ranged from 14.6 days in aluminum sulfate to 21.9 days in ClO₂. Among five commercialized biocides, ClO₂ (10 μL • L⁻¹) was significantly effective for extending the vase life of cut gerbera flowers, equally with sodium hypochlorite, silver nitrate, and 8- hydroxyquinoline sulfate (Table 1). This result coincides with previous research showing that ClO₂ 10 μL • L⁻¹ is effective for extending the vase life of Gerbera, Gypsophila, and Anthurium (Macnish et al. 2008). The treatment with the low pH holding solutions of aluminum sulfate (pH 3.5) and AVB (pH 2.7) afford a vase life approximately three days shorted than that with tap water (Table 1). It was considered that antimicrobial compounds depending on the hydrolysis of chlorine to hypochlorous acid for antibacterial activity may not be proper for use in acidified flower hydration solutions (Macnish et al. 2008).

On day 6, no bacteria were detected in ClO₂ or in any of commercialized biocides except for tap water (7.1 × 10³ CFU • L⁻¹) and distilled water (5 × 10² CFU • L⁻¹) (Table 1 and Fig. 1). In previous studies, aluminum sulfate, sodium hypochlorite, 8-hydroxyquinoline sulfate, and ClO₂ 10 μL • L⁻¹ decreased the buildup of bacteria and extended the longevity of cut flowers (Halevy and Mayak 1981; Macnish et al. 2008). Therefore it means that the antibacterial activity by biocides could be helpful for postharvest life of cut gerbera flowers.

For two weeks after the treatment, especially from day 2 to day 16, vase solution uptake was highest in aluminum sulfate and lowest in AVB (Fig. 2). But these two biocides had the shortest vase life of cut gerbera flowers among six biocides as shown in Table 1. Therefore it is not clear whether vase solution uptake is related to vase life in cut gerbera flowers or not. The fresh weight was reduced gradually after first two days taken the peak in all treatments regardless of the type of biocide (Fig. 3). Unlike vase solution uptake, the relative fresh weight rate was the lowest in aluminum sulfate and AVB which showed the lowest vase life. ClO₂ was relatively well in vase solution uptake and relative fresh weight rate compared to other five commercial biocides. As with other biocides except for aluminum sulfate and AVB, therefore, ClO₂ was more effective for extending the vase life of cut gerbera flowers. Applicable concentration of ClO₂

In the second experiment, the vase life ranged from 12.4 days in farm groundwater (control) to 15.4 days in ClO₂ 10 μL • L⁻¹. There were no significant differences in vase life among the three ClO₂ treatments: 2, 5, and 10 μL • L⁻¹ (Table 2). This result is similar to previous studies showing that ClO₂ 5 μL • L⁻¹ extended the vase life of cut roses (Koermer and Wldman 2002; Lee and Kim 2014) and the addition of low concentrations (i.e., 2 or 10 μL • L⁻¹) of aqueous ClO₂ to clean vase water significantly extended the display life of eight cut flower species by 0.9-3.4 days (Macnish et al. 2008).

In farm groundwater (control), 6.8 × 107CFU • L-1 of bacteria was detected, and no bacteria were detected in any of the three ClO₂ treatments (Table 2 and Fig. 4). This result coincides with previous researches revealing bacterial counts of 107-1011CFU • L⁻¹ in vase water. Vascular occlusions and associated wilting in flower species such as Dianthus and Rosa generally develop (van Doorn et al. 1985; van Doorn et al. 1995).

Referred to Table 1, therefore, there was no significant difference between tap water and farm groundwater in terms of bacterial contamination during postharvest period of cut gerbera flowers. We found that ClO₂ slightly increased vase solution uptake in high concentrations of 5 and 10 μL • L⁻¹, and delayed the decrease in relative fresh weight rate significantly at 10 μL • L⁻¹ (Fig. 5). The addition of ClO₂ to the vase solution improved the water uptake in a hybrid Limonium (Doi and Reid 1995) and vase solution uptake was directly associated with microbes in the holding solution (Kim et al. 1998). Therefore, the increase in vase solution uptake and fresh weight was affected by the antibacterial function of ClO₂.

The amount of ClO₂ in the holding solution dramatically decreased and almost disappeared on day 2 after treatment regardless of the concentration and was no different according to the existence of a peduncle in the solution, except for the condition without a peduncle in the solution of ClO₂ 10 μL • L⁻¹ (Fig. 6). However, the antibacterial effect was maintained up to day 6 after the treatment as mentioned in Fig. 4. This result was similar to the previous study showing that chlorine decreased more rapidly in solutions with initially lower chlorine concentration (Xie et al. 2008). In addition, this result verified that the effect of ClO₂ on extending the vase life of cut gerbera flowers was resulted from the early antibacterial function by ClO₂ in the range of 5 and 10 μL • L⁻¹ which was applicable concentration in the holding solution.

In conclusion, our study results provide a very important evidence for using ClO₂ as floral preservatives not to cause environmental pollution sources such as heavy metals or toxic chemical compounds which are generally produced from the existing commercial biocides for cut flowers.6