Impaginato 221 Adv. Hort. Sci., 2023 37(2): 221­229 DOI: 10.36253/ahsc­13608 Impact of exogenous pre and posthar­ vest salicylic acid applications on MD2 pineapple quality D.M. Cano­Reinoso 1, C. Wibowo 2 (*) 1 Department of Agrotechnology, Faculty of Agriculture, Jenderal Soedirman University, Purwokerto, Jl dr. Suparno, Karangwangkal 53123, Indonesia. 2 Department of Food Science and Technology, Faculty of Agriculture, Jenderal Soedirman University, Purwokerto Jl dr. Suparno, Karangwangkal 53123, Indonesia. Key words: Antioxidant, browning, concentration, enzyme, translucency. Abstract: Salicylic acid (SA) is a natural plant compound that has been proven to enhance the quality of fruits; therefore, its impact on pineapple should be further studied, especially in the most marketable hybrids. This study aimed to evaluate the effect of SA treatments on MD2 pineapple quality. The experiment consisted of two parts with applications pre and postharvest following the next treatments, control: No use of SA, and 5, 7 and 9 mM of SA. The total soluble solids, total acidity, ascorbic acid content, respiration rate, together with the severity and incidence of internal browning and flesh translucency were deter­ mined, after 40 days of cold storage. The treatment using 9 mM of SA in pre and postharvest delivered the best results, having the most elevated ascorbic acid (526.75 mg kg­1) and total acidity (0.8%), the lowest severity and incidence outcomes of internal browning and flesh translucency (0% in both cases), with the most reduced respiration rate values during postharvest. In conclusion, SA treatments with concentrations of 9 mM applied in pre and postharvest on MD2 pineapple can improve its quality after 40 days of cold storage. 1. Introduction Pineapple is a fruit characterized by its rich source of sugars, organic acids, fibers, minerals, vitamins, flavonoids, and carotenoids (De Ancos et al., 2017). These are essential food properties for healthy human nutri­ tion. Nowadays, low acid hybrids like MD2 are the most exported by the industry worldwide (Hossain, 2016; Cano­Reinoso et al., 2022 a). This hybrid is known for its bright­gold colour, sweeter taste, high ascorbic acid (AsA) content, and uniform size (Bin Thalip et al., 2015; Cano­Reinoso et al., 2022 a). Nevertheless, MD2 is susceptible to physiological disorders like flesh translucency and internal browning, which are major problems that negatively impact its quality (Chen and Paull, 2017; Paull and Chen, 2018). (*) Corresponding author: condro.wibowo@unsoed.ac.id Citation: CANO­REINOSO D.M., WIBOWO C., 2023 ­ Impact of exogenous pre and postharvest salicylic acid applications on MD2 pineapple quality. ­ Adv. Hort. Sci., 37(2): 221­229. Copyright: © 2023 Cano­Reinoso D.M., Wibowo C. This is an open access, peer reviewed article published by Firenze University Press (http://www.fupress.net/index.php/ahs/) and distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: All relevant data are within the paper and its Supporting Information files. Competing Interests: The authors declare no competing interests. Received for publication 27 August 2022 Accepted for publication 9 March 2023 AHS Advances in Horticultural Science https://doi.org/10.36253/ahsc-13608 http://www.fupress.net/index.php/ahs/ http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/ Adv. Hort. Sci., 2023 37(2): 221­229 222 Currently, the use of natural compounds to deal with these disorders has become a trend (Lu et al., 2011; Goñi et al., 2017). Because of the reduced neg­ ative impact these compounds cause to the environ­ ment and human health, multiple studies have been implemented by producers and growers (Ponce et al., 2011; Goñi et al., 2017). In this context, salicylic acid (SA) has been investigated as a potential treat­ ment to decrease physiological disorders due to its positive impact on fruit metabolism (Hayat et al., 2010; Goñi et al., 2017). This secondary metabolite has been proved to enhance ion uptake and trans­ port, disease resistance, ripening delay, and control postharvest quality and shelflife of horticultural prod­ ucts (Asghari and Aghdam, 2010; Goñi et al., 2017). For instance, SA has demonstrated outstanding results in reducing the fruit softening rates and the degrading of the sugar and acid content during postharvest (Asghari and Aghdam, 2010; Goñi et al., 2017). Also, SA can mitigate the cell wall degrading and polygalacturonase (PG) enzyme activity, phe­ nomena highly associated with translucency and internal browning in pineapple (Goñi et al., 2017; Paull and Chen, 2018; Cano­Reinoso et al., 2021). Previous studies in pineapple fruit demonstrated that postharvest SA treatments (in solutions with concentrations between 5 and 9 mM), reduced the internal browning severity and translucency occur­ rence without affecting the total soluble solids (TSS) and total acidity (TA) negatively (Lu et al., 2010, 2011; Cano­Reinoso et al., 2022 b). Besides, it was determined that in pineapple SA could cause a posi­ tive effect on the antioxidant content with a reduc­ tion of the peroxidase (POD), polyphenol oxidase (PPO) and phenylalanine ammonia­lyase (PAL) enzyme activities, concomitantly with a low respira­ tion rate (Lu et al., 2010, 2011; Cano­Reinoso et al., 2022 b). However, despite the previous information described currently, there are no sufficient studies on pineapple fruit that document the impact of SA, especially complementing postharvest with prehar­ vest applications. For example, concerning the pre­ harvest stage only two experiments have reported the use of SA, obtaining contradictory results, and mostly implementing just low concentrations (around 2 mM), like in Lu et al. (2011) and Cano­Reinoso et al. (2022 b). On top of that, some of these former researches have mainly focused on acid hybrids. Therefore, this issue open questions concerning the determination of the exact impact of SA on low acid hybrids like MD2, primordially with a complementary administration, including pre and postharvest. As a result, this study aims to evaluate the impact of exogenous pre and postharvest SA applications on MD2 pineapple quality. 2. Materials and Methods Preparation of the experiment This research was implemented in pineapple fields in Lampung, a place located in south Sumatra Indonesia, from January to March 2020, employing the MD2 pineapple hybrid. In order to fulfil the objective of the research, the experiment was divid­ ed into two parts. The first part regarding preharvest applications of SA, while the second part concerned the postharvest administration of this natural com­ pound. Preharvest treatments implementation Regarding the first part of the experiment, a com­ pleted randomized block design was used with four replications, having 20 fruits each of the replications. The fruits were harvested between 144­147 days after flowering, when it has been determined that MD2 can expose the most optimal quality character­ istics for commercial consumption (Bin Thalip et al., 2015; Ding and Syazwani, 2016). Besides, for this part of the research, four rows in each block were pre­ pared with a width and length of 0.4 and 3.75 m, respectively. Pineapple plants were organized in two lines of ten plants with a separation of 0.25 m. For this stage the treatments implemented were; con­ trol: No use of SA, 5, 7 and 9 mM of SA. Furthermore, the administration of SA was done using sprayings with their respective solutions until the fruits were wet to runoff. SA solutions were mixed with 1% (v/v) of ethanol and 0.01% (v/v) of tween 20 (emulsifier) before their application. The sprayings were performed at eight, six, four and two weeks before harvest at night, in the fruit shell and crown. Previous experiment of Cano­Reinoso et al. (2022 b) demonstrated that two applications prior to harvest (six and three weeks) can cause a positive effect on pineapple quality. Therefore, this research aimed to increase the frequency of application pre­ harvest in order to enhance the influence on the fruit quality. Moreover, the fruit shell and crown were selected as adequate spots to be used based on the findings about mineral mobility in pineapple plants Cano‐Reinoso and Wibowo ‐ Salicylic acid application on MD2 pineapple 223 described by Vásquez­Jiménez and Bartholomew (2018) and Murai et al. (2021). They demonstrated that through these plant structures there was miner­ al assimilation when foliar fertilizations were carried out after flowering. The soil, where the pineapple plant was cultivat­ ed, was fertilized previously with 200 kg ha­1 Di­ ammonium Phosphate, 1000 kg ha­1 of K2SO4 and 200 kg ha­1 Kieserite crystal. Three months after plating 700 kg ha­1 of Urea were administered by sprayings; also, 700 kg ha­1 of (NH₄)₂SO₄, 1000 kg ha­1 of K2SO4, 170 kg ha­1 of MgSO4, 60 kg ha­1 FeSO4, 60 kg ha­1 ZnSO4, were applied in intervals of 30 days. Besides, borax was sprayed in doses of 30 kg ha­1 at flower induction. Climatological conditions were determined where the plants were cultivated with a weather sta­ tion (LSI Lastem; equipped with a CR6 data logger from Campbell Scientific; Italy). An average of 70.60% of relative humidity (RH), 22.43°C, 10.18 w m­1 of solar radiation, and rainfall of 353.10 mm were detected. The physical and mineral composition of the soil of the first part of the experiment is present­ ed in Table 1. Postharvest treatments implementation Concerning the second part of the experiment, randomly ten fruits per replication were selected at harvest, organized according to their respective treatments and replications inside a cold storage dur­ ing 40 days (8°C and 90% RH), and analyzed in inter­ vals of eight days. The treatments implemented in this stage of the research were; control: No use of SA, 5, 7 and 9 mM of SA. All the treatments, including the control, received fungicide and waxing applica­ tions in postharvest; those materials, following that order, were administrated in dipping applications for ten seconds, just after the dipping on SA. The fungi­ cide product used was Prochloraz in doses of 2 cc l­1, while the waxing material employed was Sta­Fresh 2952 in doses of 74 g l­1. Furthermore, the SA concen­ trations were dissolved in a water container of 25 L; Besides, in this case SA was also mixed with 1% (v/v) of ethanol and 0.01% (v/v) of tween 20 (emulsifier). The dipping in SA was done for five minutes. This dip­ ping time and the concentrations implemented were selected based on Lu et al. (2011) and Cano­Reinoso et al. (2022 b). They proved that postharvest dipping applications around five minutes should have a mini­ mum concentration of 2 mM to cause a positive impact on pineapple fruit. Finally, the summary of the treatments implemented pre and postharvest in this research are presented in Table 2. Fruit quality evaluation Total soluble solids (TSS) and total acidity (TA) in the fruit The total TSS and TA content was calculated in every fruit selected of every treatment from each replication, employing the method described in Shamsudin et al. (2020). TSS was measured using a hand­held refractometer (MASTER­53 α; Atago; Japan), while TA was determined by titration to pH 8.1 with 0.1 M NaOH employing phenolphthalein indicator and expressed as a percentage of citric acid. Table 1 ­ Physical and mineral composition of the soil in the first part of the experiment The N, P, K, Ca, Mg and Na represent the available mineral con­ tent in the soil. Properties Content Texture Clay (%) 9.00 Loam (%) 36.56 Sand (%) 50.42 Chemical properties pH (H20) 7.62 C (%) 4.60 N (mg kg­1) 853.00 P (mg kg­1) 5.29 K (mg kg­1) 9.68 Ca (mg kg­1) 76.39 Mg (mg kg­1) 109.00 Na (mg kg­1) 12.84 Table 2 ­ Summary of the treatments implemented in pre and postharvest stage of the experiment * Preharvest salicylic acid (SA) was sprayed at 8, 6, 4 and 2 weeks before harvest until fruits were wet to runoff. Postharvest SA was administrated with dipping of 5 minutes, and comple­ mented with fungicide and waxing applications. Pre and posthar­ vest SA solutions were mixed with: 1% (v/v) ethanol, and 0.01% (v/v) tween 20 (emulsifier). Treatment Description A Control: No use of SA B 5 mM SA pre/postharvest C 7 mM SA pre/postharvest D 9 mM SA pre/postharvest Adv. Hort. Sci., 2023 37(2): 221­229 224 Fruit respiration rate The respiration rate was calculated in a fruit selected from eight until 40 days of cold storage in each replication of every treatment implemented. A similar method previously reported in Bhande et al. (2008) and Cano­Reinoso et al. (2022 b) was imple­ mented. For this case, changes in the CO2 concentra­ tion were measured in a sealed glass container hav­ ing 31 cm height x 24 cm wide with 9 l of capacity. The device used for this procedure was a portable AZ7788A carbon dioxide detector (CO2 range: 0­5000 mg kg­1, 10­95 % RH, 0­50°C; AZ Instrument Corp; Taiwan). Moreover, before beginning the procedure, the fruit weight was calculated using a weighing scale, as described in Shamsudin et al. (2007). Once finished this process, the CO2 detector and the fruit were arranged inside the container, avoiding any air introduction or scape. The changes in the CO2 con­ centration were measured during one hour in every fruit, and after that, the respiration rate was deter­ mined using the following expression: (1) Where, respiration of the fruit (RF) is the respiration rate in ml CO2 kg­1·· h­1, FCO2 is the CO2 gas concentra­ tion in ml l­1, h is the storage time in hours, Δt the time difference between two CO2 gas measurements, V is the free volume of the container in l and FW is the weight of the fruit in kg. The free volume of the container was obtained as the total volume of the container minus the volume occupied by its content at the moment of the measuring, using a water dis­ placement method as described in Bhande et al. (2008). Figure 1 shows a picture of the arrangements carried out to implement the respiration rate method of this experiment already described. Ascorbic acid (AsA), translucency and internal brown‐ ing incidence determination These variables were measured in each of the fruits per replication of every treatment. The AsA content was calculated using dye, 2,6­dichlorophe­ nol­indophenol titration method described in Ding and Syazwani (2016) and Ojukwu and Nwobi (2017). First, diluted pineapple juice was pipetted into a coni­ cal flask and mixed with glacial acetic acid, titrating the solution until faint permanent pink colour. Next, the titrated value was recorded; then, the titration was repeated, boiling and cooling with distilled water for a blank and a standard ascorbic acid solution. The titrations obtained were repeated twice, and the average value was calculated and expressed as mg kg­1 fruit fresh weight. On the other hand, the translucency internal browning incidence was obtained accounting number of fruits affected from the total examined, since har­ vest until 40 days of cold storage. The results are expressed in percentage. Statistical analysis The statistical analyses were performed using SPSS Version 22.0 software (SPSS Inc., Chicago, IL, USA). All data were analysed by an analysis of vari­ ance of one­way (ANOVA). Mean significant differ­ ences at p<0.05 were determined by Duncan’s multi­ ple range tests. 3. Results Fruit physicochemical quality The TSS, TA and TSS/TA ratio did not show signifi­ cant differences after 40 days of cold storage in the implemented treatments. The TSS and TA content was on average 15 and 0.70%, respectively; mean­ Fig. 1 ­ Picture showing the respiration rate method employed. 1) CO2 detector used, 2) Cable of electricity to activate the detector introduced by the top of the container, 3) Fruit and 4) Sealed container. Cano‐Reinoso and Wibowo ‐ Salicylic acid application on MD2 pineapple 225 while, for the TSS/TA the mean values ranged around 21 (Table 3). For these variables, high SA concentra­ tions were not clearly associated with more elevated mean results. On the other hand, the AsA results did not demonstrate significant differences in this experi­ ment after 40 days of cold storage. Nevertheless, it is important to notice that the treatment using 9 mM of SA in pre and postharvest obtained the highest mean value (526.75 mg kg­1), which was linked to the more superior content of TA (0.80%) (Table 3). Moreover, in figure 2 it is possible to observe the trend of AsA through the postharvest time of the experiment. AsA continually elevated and reduced its level, with the treatment employing 9 mM of SA in pre and postharvest having the most inferior peak change, especially between 32 and 40 days of cold storage. Furthermore, in the case of the respiration rate, this variable exposed significant differences after 40 days of cold storage. The control treatment (no use of SA) provided the highest value, while the treat­ ment utilizing 9 mM of SA in pre and postharvest had the most reduced one (8.24 and 6.32 ml CO2 kg­1·h­1, respectively). Figure 3 shows the respiration rate trend during postharvest. In this figure, it is evi­ denced that the treatments employing a higher SA concentration, like the treatments with 7 and 9 mM of SA in pre and postharvest, had a steadier trend and lower change during cold storage. On the other hand, the control treatment and the one using 5 mM of SA in pre and postharvest suffered a remarkable increase, primordially in 16 days, suggesting a repre­ sentative metabolic change at that moment in the fruit. On top of that, high concentrations of SA were associated with a lower respiration rate and more elevated AsA content at harvest. Internal browning and flesh translucency in the fruit The internal browning and flesh translucency pro­ vided significant differences in the results obtained. Regarding the internal browning, this had its most elevated severity and incidence in the treatments using 5 and 7 mM of SA in pre and postharvest (3.75 and 6.25%, respectively), while the control treatment Fig. 2 ­ Effect of the treatments implemented on the ascorbic acid (AsA) content during 40 days of cold storage. A) con­ trol [No use of salicylic acid (SA)], B) 5 mM SA pre/postharvest, C) 7 mM SA pre/postharvest, and D) 9 mM SA pre/postharvest. Values are the mean four repli­ cates, and vertical bars represent ± SE. Fig. 3 ­ Effect of the treatments implemented on the fruit respi­ ration rate during 40 days of cold storage. A) control [No use of salicylic acid (SA)], B) 5 mM SA pre/postharvest, C) 7 mM SA pre/postharvest, and D) 9 mM SA pre/posthar­ vest. Values are the mean four replicates, and vertical bars represent ± SE. Table 3 ­ Impact of the treatments implemented on the fruit quality characteristics after 40 days of storage * Mean values ± SE in each column followed by the same lower­case letters are not statistically different by Duncan’s multiple range test, and Kruskal­Wallis test (for the internal browning and translucency data) (p < 0.05). Treatment TSS (%) TA (%) TSS/TA AsA (mg Kg­1) Respiration rate (ml CO2 Kg­1 h­1) Browning (%) Translucency (%) Severity Incidence Severity Incidence A 14.80 ± 0.22 a 0.70 ± 0.07 a 21.73 ± 1.85 a 496.25 ± 62.86 a 8.24 ± 0.40 a 0.00 b 0.00 b 1.85 ab 16.67 a B 15.40 ± 0.50 a 0.67 ± 0.07 a 24.10 ± 3.65 a 402.50 ± 71.81 a 8.23 ± 0.92 a 3.75 a 6.25 a 0.64 b 8.33 b C 15.80 ± 0.29 a 0.75 ± 0.05 a 21.43 ± 1.53 a 492.50 ± 65.97 a 6.83 ± 0.65 ab 3.75 a 6.25 a 0.35 b 4.17 b D 15.15 ± 0.31 a 0.80 ± 0.06 a 19.13 ± 1.23 a 526.75 ± 23.47 a 6.32 ± 0.53 b 0.00 b 0.00 b 0.00 b 0.00 c 226 Adv. Hort. Sci., 2023 37(2): 221­229 and the one employing 9 mM of SA in pre and postharvest did not expose representative symptoms after 40 days of cold storage (0% for the severity and incidence in both cases) (Table 3). Moreover, this lowest browning severity and incidence outcome was related to the most inferior respiration rate and the highest AsA content at harvest, primordially in the treatment utilizing 9 mM of SA in pre and posthar­ vest. Concerning the flesh translucency, the out­ comes of this experiment also revealed that this pre­ vious treatment is the one causing the lowest severi­ ty and incidence (0% in both cases), while the control treatment was the least efficient in this aspect (1.85 and 16.67%, respectively) (Table 3). Similar to the internal browning results, the most elevated AsA content and lowest respiration rate at harvest in the treatment employing 9 mM of SA in pre and posthar­ vest were linked to the most inferior translucency incidence and severity. 4. Discussion and Conclusions Typically, MD2 pineapple displays 12% or higher values for TSS during cold storage (Paull and Chen, 2018; Cano­Reinoso et al. , 2021). The values obtained in this research were higher than 14%, especially for those using SA applications. TSS repre­ sents the sucrose content in pineapple fruit primor­ dially; also, a change in its content commonly is asso­ ciated with modifications in the cell wall invertase (CWI) activity (Saradhuldhat and Paull, 2007; Paull and Chen, 2018). The results of this experiment infer that the pre and postharvest treatments employed did not cause a representative impact on the sucrose content and the sugar metabolizing enzymes, essen­ tially a negative impact, to cause a change in the TSS level at the end of the cold storage. In the case of the TA, due to its low acid proper­ ties, MD2 has evidenced TA values among 0.4­0.7% at harvest, which is much lower than acid hybrids like smooth cayenne (Ding and Syazwani, 2016; Paull and Chen, 2018). Nevertheless, a small increase in this range has been reported during cold storage of pineapple due to the rise of the citric acid content (Mandal et al., 2015; Ding and Syazwani, 2016). Citric acid is considered a source for several metabolizing process in fruits, like antioxidant scavenging and res­ piration; also, it is the main acid influencing the TA measuring in pineapple (Paull and Chen, 2018; Yang et al., 2019). Therefore, in concomitance with the fruit decay, this acid could has speed up its accumula­ tion as a reflect of this physiological deterioration, influenced by the need of a scavenger agent to cope with this condition. This fact can explain why despite the lack of significant differences, in mostly all the treatments, TA had higher values than the minimal recommended for MD2, after 40 days of cold stor­ age. On top of that, as also the control treatment (no use of SA) had an ideal TA outcome, it could be sug­ gested that the waxing formula used was ideal to decrease the fruit metabolism in order to avoid a high consumption of citric acid as a substratum of several physiological process, generating the respec­ tive TA content. Furthermore, a small increase in the TSS and TA content has been observed by SA applications in pineapple (Lu et al., 2011; Mandal et al., 2015). Moreover, some studies have demonstrated that pre and postharvest implementations of SA tend to sup­ press the TSS and TA degrading, especially during postharvest, like in mango and grape (Champa et al., 2014; Hong et al., 2014). This information could explain why the TSS and TA values were in the opti­ mal content for consumption or slightly superior in almost all the treatments employing SA. Besides, MD2 has more elevated TSS/TA ratios than other hybrids, especially because of its lower acid content, ranging between 20­30 in mostly all cases. (Chen et al., 2009; Ding and Syazwani, 2016). Overall, values between this range were observed in the results of this experiment (Table 3). On the other hand, regarding AsA, this is consid­ ered a powerful plant antioxidant and one of the most representative in pineapple fruit (Kongsuwan et al., 2009; Noichinda et al., 2017). This information infers that the treatment utilizing 9 mM of SA in pre and postharvest can provide a more elevated antioxi­ dant production, as it impacts the AsA level positively and the previous described TA content associated with citric acid. MD2 delivers AsA values as minimal as 300 mg kg­1 at harvest, much higher than pineap­ ple acid hybrids (Lu et al., 2014; Paull and Chen, 2018). This value tends to increase during posthar­ vest to maintain optimal physiological conditions, essentially when a stress factor affects the fruit (Lu et al., 2011; Mandal et al., 2015). The variability of the AsA exposed in figure 2 could be associated with the activity of the ascorbic peroxidase (APX) and mon­ odehydroascorbate reductase (MDHAR) enzymes. For example, APX is an optimal scavenger of H2O2, which is generated with the progress of the fruit Cano‐Reinoso and Wibowo ‐ Salicylic acid application on MD2 pineapple 227 senescent. However, this enzyme can cause the oxi­ dation of AsA into monodehydroascorbate (MDHA). Therefore, MDHA have to be recycled into AsA by the glutamic acid pathway employing MDHAR as cofac­ tor, in order to maintain an ideal level of AsA to cope with impact of the fruit decay. As a result, these two enzymes can be used as an indicator of the AsA con­ tent and metabolization (Gallie, 2013; Akram et al., 2017). Based on that, the treatment working with 9 mM of SA in pre and postharvest could be considered the one causing more influence on these enzymes, catalyzing their activities during cold storage. Similar patterns in the AsA production during postharvest has been reported in pineapple by Lu et al. (2010) and Cano­Reinoso et al., (2022 b), where higher SA concentrations were associated with a more superior AsA content. In the case of the respiration rate, values between 5 and 20 ml CO2 kg­1·h­1 have been reported for pineapple in previous studies (Lu et al., 2011; Hu et al., 2012; Cano­Reinoso et al., 2022 b). The outcomes obtained in this experiment provided results among that recommended range. On the other hand, SA has been associated with an increase in antioxidant scav­ enger enzymes like catalyze (CAT) and superoxide dismutase (SOD), which, together with the APX, reduced the impact of the fruit decay symptoms dur­ ing postharvest, eliminating singlet of oxygen­derived from reactive oxygen species (ROS), essentially the already mentioned H2O2 (Goñi et al., 2017; Noichinda et al., 2017). On top of that, Lu et al. (2011) and Cano­Reinoso et al. (2022 b) demonstrated that SA solutions with concentrations higher than 5 mM, essentially with postharvest implementations, can generated a low respiration rate, with a similar trend pattern of this experiment. Besides, they determined that this low respiration rate was associated with a more superior activity of the CAT, SOD and APX, especially between 16 and 24 days of cold storage. That period of time was regarded as the moment when more ROS were produced in the fruit; there­ fore, the higher activity of these enzymes to cope with the damage caused. According to that, in this experiment the treatments using 7 and 9 mM of SA in pre and postharvest could be suggested as the ones causing a more elevated scavenger activity of CAT, SOD, APX, generating less ROS, decreasing the respiration rate, and promoting an ideal fruit physio­ logical condition. Regarding the internal browning, this physiologi­ cal condition in fruits is associated with a high activity of phenylalanine ammonia­lyase (PAL) and polyphe­ nol oxidase (PPO) enzymes during postharvest, con­ comitant with an elevated level of ROS and low antioxidant content (Youryon et al., 2018; Paull and Chen, 2019). This finding suggests that SA solutions with concentrations of 5 and 7 mM applied pre and postharvest can catalyze PAL and PPO activity, increasing the ROS content, creating higher browning symptoms in the flesh. In the case of the treatment employing 9 mM of SA in pre and postharvest, its lowest internal browning severity and incidence was linked to the lowest respiration rate and the most elevated antioxidant content. This fact infers that those high concentrations of SA (9 mM), on the con­ trary, can inhibit the PAL and PPO activity, reduce the ROS level, generating positive effect on the fruit metabolism. On the other hand, these results contradict the outcomes of Lu et al. (2011) and Mandal et al. (2015), where concentrations of 5 mM or higher applied pre and postharvest caused the lowest inter­ nal browning incidence and severity, as in this experi­ ment the control treatment, produced positive results in this matter. Furthermore, despite having a high respiration rate, the control treatment delivered an ideal AsA and TA content. This situation exposes that the fruits of this treatment were harvested with optimal physiological characteristics, necessary to delay any internal browning symptoms until the last day of cold storage. Besides, some authors have sug­ gested that MD2 pineapple, due to its higher antioxi­ dant content, could extend its postharvest shelflife up to 45­50 days, depending on the fruit quality con­ dition collected from the field (Paull and Chen, 2018, 2019). Conversely, translucency is a physiological disor­ der of pineapple fruit characterized by water soaking symptoms, being MD2 a susceptible hybrid (Chen and Paull, 2017; Paull and Chen, 2018). Furthermore, the low calcium content in the fruit and possible low assimilation of Ca2+ into the cell wall matrix have been linked to this physiological disorder’s exhibition (Paull and Chen, 2015, 2018). These facts suggest that SA applications employed in pre and posthar­ vest, essentially at high concentrations like in the treatment working with 9 mM of SA in pre and postharvest, could enhance the calcium ion assimila­ tion and concentration in flesh tissues, reducing the exhibition of translucency. These results agree with Adv. Hort. Sci., 2023 37(2): 221­229 228 the findings of Mandal et al. (2015), where imple­ menting SA with concentrations of 5 mM after 15 days of cold storage got to reduce translucency inci­ dence, compared to treatments without SA. On top of that, similar with the internal browning, the lowest translucency severity and incidence was associated with the most inferior respiration rate and highest AsA content. As a result, after analyzing all the vari­ ables measured in this experiment, it is possible to suggest that this treatment could be considered as the more appropriate to obtain an ideal pineapple quality after the 40 days of cold storage. Finally, the current available information about the SA effect on pineapple exposes that low prehar­ vest concentrations, between 2 and 5 mM, mixed with more elevated ones in postharvest, provide the best results regarding the fruit quality (Lu et al., 2010, 2011; Cano­Reinoso et al. , 2022 b). Nevertheless, this experiment got to demonstrated that elevated concentrations of SA employed prehar­ vest could also benefit the fruit. For example, a more superior calcium uptake, and AsA level and recycling can be encourage with those high SA concentrations (primordially between 7 and 9 mM). Because of that, future experiments in pineapple should be focus on these concentrations, prioritizing the impact on the fruit mineral status and antioxidant content. Also, deeper researches concerning the determination of the ideal pre harvest frequency of SA application should be done, as former studies still lack on this matter. These preliminary results demonstrated that four preharvest applications can be more beneficial than two, complementing former experiments. In conclusion, SA treatments applied pre and postharvest affected the MD2 pineapple quality. The treatment employing SA solutions with a concentra­ tion of 9 mM pre and postharvest provided the best results, essentially because it had the lowest and reg­ ular postharvest trend and outcome of respiration rate, and the highest value of AsA and TA content. Also, this treatment obtained the most reduced severity and incidence of internal browning and flesh translucency after 40 days of cold storage. Acknowledgements The authors would like to express their sincere thanks to PT Great Giant Pineapple and their research department in Lampung Indonesia for the technical support during the research period. 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