Applying of sodium nitroprusside ( SNP ) on postharvest ‘ Nam Dok Mai No . 4 ’ mango fruits delay ripening and maintain quality

The aim of this study was to evaluate the concentrations of sodium nitroprusside (SNP), a nitric oxide donor, on postharvest ripening and quality of mango fruit cv. Nam Dok Mai No.4. The fruits were dipped in 1 or 2 mM of SNP solution for 30 min while control fruits were dipped in tap water for 30 min. After treatment, all fruits were stored for 18 days at 13°C and RH 90%. Both SNP doses showed significantly reduced ethylene production and respiration rate, maintained the fruit firmness, decreased the changes in total soluble solids and titratable acidity, and reduced fruit weight loss. The treatments of SNP also delayed color development in fruit peel and pulp. SNP-treated fruits showed higher L* and hue angle values and lower a* and b* values in comparison with control fruits. No significant differences were found between 1 and 2 mM SNP treatments. SNP applying on dose of 1mM to postharvest ‘Nam Dok Mai No.4’ mango fruits and then stored at 13°C changed ethylene production and respiration rates, maintained firmness pulp, decreased weight loss and slowed the changes in fruit peel and pulp color, total soluble solids (TSS) and titratable acidity (TA),.


INTRODUCTION
Nitric oxide (NO) is recognized as a biological messenger in plants.It is a highly reactive gaseous free radical and is soluble in water and lipid (Hayat et al., 2010).NO can mediate various pathophysiological and developmental processes, including the expression of defence-related genes, programmed cell death, stomatal closure, seed generation and root development (Neill et al., 2003).Optimum NO levels could delay the climacteric phase of many tropical fruits and prolong the post-harvest shelf life of a wide range of horticultural crops by preventing ripening and senescence (Singh et al., 2013).NOfumigated fruits reduced ethylene production due to binding of NO with 1-aminocyclopropane-1-carboxylic acid (ACC) and ACC oxidase to form a stable ternary complex, thus limiting ethylene production (Tierney et al., 2005).Sodium nitroprusside (SNPa NO donor) solution *Corresponding author.E-mail: apiradee.uth@kmutt.ac.th,Tel: +66 81 699 0955.Fax: +66 2 470 7729.Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0 International License at 1 mM was able to inhibit pectin solubilization/depolymerization, delay the increase in total phenolic content, and lower DPPH radical scavenging activity in plum fruits applied at postharvest during 120 days at 2°C (Li et al., 2007).Peach fruits treated with 1 mM SNP prolonged shelf life by reducing ethylene production rate, increasing firmness and antioxidant activity potential of the enzymes such as superoxide dismutase, and catalases (Sis et al., 2012).
Thailand is one of the most important countries in Asian continent in exporting mangos and the cv.Nam Dok Mai is a well known cultivar, with a fibreless, delicious taste and sweet aroma.However, as a fleshy fruit, the losses resulting in senescence, desiccation, physiological disorders, mechanical injuries and microbial spoilages can occur at any point from harvest to utilization (Wu, 2010).Up to 30% of mango products are lost due to improper post-harvest operations (Malik et al., 2005).Mango storage time is limited to 2 to 3 weeks in normal air at 10 to 15°C depending on the cultivar (Yahia, 1999).Application of 1-MCP has potential for the commercial control of ripening and softening of harvested mangos.However, the use of appropriate 1-MCP concentration still requires being investigated further (Sivakumar et al., 2011).The treatment of NO on mango fruits was mostly reported in gas (fumigation with gaseous NO).In practice, the gaseous NO treatment requires postharvest infrastructures, therefore the dipping with SNP solution will be more suitable than gaseous treatment (Sis et al., 2012).The SNP reaction releases NO and cyanide (CN) which is known as a toxic chemical agent.Free CN radical may bind and inactivate tissue cytochrome oxidase and prevent oxidative phosphorylation (Friederich and Butterworth, 1995).Minimum absorbed lethal dose (LD 50 ) of CN in humans was estimated to be 1.4 to 3.0 mg/kg (Way, 1981).Therefore, using SNP in postharvest management should be cautious for human safety in term of the concentration and also ventilation during treatment.Up to this time, data on applying of NO on Thai mangos are still limited, and in particular no information has been reported on SNP treatment on 'Nam Dok Mai' mangos.The aim of this study was to determine different concentrations of SNP on the effects of physiological and biochemical properties of 'Nam Dok Mai No.4' mango fruits during cold storage.

MATERIALS AND METHODS
'Nam Dok Mai No.4' mango fruits at the mature stage were purchased from the exporting company in Bangkok, Thailand.The fruit were selected for uniform, size and color, besides of disease and mechanical damage.SNP (HIMEDIA company, Indiamolecular weight: 297.95 g mol⁻¹) was dissolved in distilled water before using.In the preliminary study, we found that 0.5 mM SNP did not have clear effect on mango quality during storage, and 4 mM SNP caused the damage to mango fruit skin).In this research, the fruits were dipped in 1 or 2 mM of SNP solution for 30 min at the ambient temperature.Control samples were dipped in water following the same procedure as fruits treated in SNP.Each sample fruit were composed by 3 fruits x 3 replications.After treatment all fruits were stored at refrigerated chambers at 13°C for 18 days with RH 90%.

Measurements of ethylene production and respiration rate
Fruits were put in a closed plastic box, and then incubated for 1 hour at the storage condition.A 1 mL gas sample from each plastic box was injected into the gas chromatograph (Shimadzu GC-8A and Shimadzu GC-14B, Japan) to measure CO₂ and C₂H₄ concentrations.The fruit respiration rate was expressed as mg CO₂ kg⁻¹ h⁻¹ and ethylene production was expressed as µL C₂H₄ kg⁻¹ h⁻¹.

Measurements of quality
Fruit firmness was measured using the texture analyser (TA.XT plus, UK) and the results were expressed in Newtons (N).The color changes in the fruit peel and pulp were measured using a colorimeter (Konica Minolta CR-400, Japan).It ranges from 0 to 360° (h=0°: red; h=90°: yellow; h =180°: green; h=270°: blue).Fruit weight losses were expressed as a percentage of weight loss relative to the initial weight.Total soluble solids (TSS) were measured using the digital refractometer (Atago, PAL-1, Japan) and the results expressed as °Brix.To measure titratable acidity (TA), fruit juice was titrated against 0.1 N NaOH until pH 8.1, and the results were expressed as % of citric acid.

Statistical analysis
All statistical analyses were performed with SPSS software ® .The experiment was carried out in a completely randomized design with three replicates for each treatment.All parameters were measured at day 0 and then at three-day intervals until 18 days.The data were expressed as the mean±SE.Differences between SNPtreated and the control fruits of the same day were compared using Fisher's least significant differences (LSD) F-test with p<(0.05).

Respiration rate and ethylene production
Respiration rate in all fruits increased from day 0 and reached the respiration peaks at day 12 then decreased.Both SNP treatments significantly decreased respiration rates at days 3, 6 and 12.The rate of respiration in untreated fruits was 1.5-fold higher than in treated fruits (Figure 1a).The SNP treatments significantly inhibited ethylene production during cold storage.Ethylene production in the control treatment was 2-fold higher than in the SNP treatment at day 3 and kept that rate until the last day of storage (Figure 1b).The treatment of 1 mM SNP was more effective than 2 mM SNP in the inhibition of respiration rate and ethylene production in mango fruits during cold storage.

Firmness, weight loss, TSS and TA
The firmness in the control fruits reduced slightly from day 3 to day 12 then decreased sharply to day 18, while the treated fruits exhibited the maintenance of fruit firmness.Fruit treated with 1 or 2 mM SNP was significantly firmer than untreated fruit during cold storage except day 6 or days 6 and 12, respectively (Figure 2a).The SNP treated fruits exhibited significantly lower weight loss than untreated fruits during cold storage.Significant decrease in weight loss in 1 mM SNP was observed throughout the storage period, while 2 mM SNP showed significant differences in weight loss only at later periods (days 12, 15 and 18) (Figure 2b).Fruit treated with SNP exhibited lower values in TSS and higher values in TA during cold storage.The significant differences between treated and untreated fruits in TSS were found on days 6 and 18 and in TA on days 9 and 15, respectively (Figures 2c and d).At 1 mM SNP was more effective than 2 mM SNP in decreasing the changes in TSS and TA with no significant difference.

Fruit color
Treatment with SNP inhibited the changes in mango peel and pulp color during storage, especially at the later periods.Significantly lower a* and b* values in the fruit peel were measured at days 9, 12, 15, 18 and days 9 and 12, respectively.The treatment of 1 mM SNP seemed to be more effective than 2 mM SNP in decreasing the changes in the hue angle of fruit peel.Fruits treated with 1 mM SNP significantly decreased the change in hue angle from days 12 to 18 while 2 mM SNP had only one period of significant difference (day 18) compared with untreated fruits.Significant differences in fruit pulp color were found at days 15 and 18, with higher values in L* and hue angle and lower values in b* in SNP treated fruits compared with the control fruits (Table 1).

DISCUSSION
Mangos are climacteric fruits and their ripening is characterized by the burst of ethylene production and a respiration climacteric followed by a series of biochemical changes (Reddy and Srivastava, 1999).In our studies, SNP treatments inhibited ethylene production in mango fruits during cold storage.The inhibition of ethylene may suppress the ripening process including maintaining the firmness pulp, suppressing the increase in TSS (on day 6 and day 8), suppressing the decrease in TA (on day 9 and day 15).In addition, the fruits treated with 1 mM SNP exhibited the significantly lower weight loss during storage.During ripening process of mango fruits, the peel color changes from green to yellow, and the pulp color changes from white to yellow.These changes are associated with the changes in the values of L*, a*, b*, and hue angle.In this research, SNP treatments decreased the color development during mango ripening due to inhibiting the increase in a* and b*, and the decrease in h angle in fruit peel, and inhibiting the increase in b* values and the decrease in L* values and h angle in fruit pulp.These effects resulted in delaying mango fruit ripening.
The effects of NO on postharvest fruits were pronounced in some papers.NO fumigation at 60 μL.L⁻¹ effectively suppressed ethylene formation and respiratory rate, reduced weight loss, maintained firmness and delayed changes in peel color and TSS in papaya fruits during 20 days of storage (Li et al., 2014).Post-harvest exposure of 'Amber Jewel' plums to NO gas with 10     μL.L⁻¹ delayed ripening by 3 to 4 days at 21±1°C by suppressing respiration and ethylene production, delaying the color development, maintaining the firmness and decreasing changes in TA in plum fruits (Singh et al., 2009).NO fumigation was reported to reduce respiration in 'Kensington Pride' mango fruits during ripening at 21°C (Zaharah and Singh, 2011).Similarly, NO treatment was found to reduce respiration in plums (Singh et al., 2009), peaches (Flores et al., 2008) and strawberries (Zhu and Zhou, 2007).Our results also showed that SNP treatment at 1 mM inhibited the respiration rate and weight loss during storage.The maintenance of fruit weight may be due to a suppression of respiration rate in the SNP treated fruit.NO affects the mitochondrial activity in plant cells and reduces total cell respiration due to its inhibitory effect on the cytochrome leading to decreasing respiration (Hayat et al., 2010).
The delay of the ripening and the maintenance of fruit quality in SNP treated fruits may be explained by suppressing biochemical changes in ethylene or respiration pathway corroborating that SNP can regulate ethylene dependent or independent pathway by activity enzymatic or genes expression (Cheng et al., 2009).In our research, the treatment with 1 mM SNP was more effective than 2 mM SNP in inhibiting ethylene production and maintaining the quality of 'Nam Dok Mai' mango fruits during storage at 13°C (Figure 3).The exogenous The color was expressed as the values of L*, a*, b* and hue angle (h).The values of L* describes the lightness increasing from black to white.The values of a* and b* describe the color position.Negative a* values indicate green while positive values indicate magenta.Negative b* values indicate blue and positive values indicate yellow.h describes the hue angle.

Figure 1 .
Figure 1.Effect of SNP treatment on respiration rate (a) and ethylene production (b) of 'Nam Dok Mai No.4' mango fruit during cold storage at 13°C for 18 days.Data represent the means ± SE, n = 3. Vertical bars represent SE of the means and are not visible when the values are smaller than the symbols.

Figure 2 .
Figure 2. Effect of SNP concentrations on the changes of firmness (a), weight loss (b), TSS (c) and TA (d) of 'Nam Dok Mai No.4' mango fruit during cold storage at 13°C for 18 days.Data represent the means ± SE, n = 3. Vertical bars represent SE of the means.

Table 1 .
Effect of SNP treatment on the color changes of 'Nam Dok Mai No.4' mango fruits during storage at 13°C.
means ± SE, n = 3.Values followed by the different letters within the same column indicate significant differences among treatments at p ≤ 0.05 (LSD test).

Figure 3 .
Figure3.The peel and pulp color in mango fruits treated with SNP at 1 and 2 mM and control fruits on day 18 of storage at 13°C.