Chilling Stress Upregulates α-Linolenic Acid-Oxidation Pathway and

Article pubs.acs.org/JAFC

Chilling Stress Upregulates α‑Linolenic Acid-Oxidation Pathway and Induces Volatiles of C6 and C9 Aldehydes in Mango Fruit Velu Sivankalyani,† Itay Maoz,†,‡ Oleg Feygenberg,† Dalia Maurer,† and Noam Alkan*,† †

Department of Postharvest Science of Fresh Produce, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem, Rehovot 76100, Israel

‡

S Supporting Information *

ABSTRACT: Mango-fruit storage period and shelf life are prolonged by cold storage. However, chilling temperature induces physiological and molecular changes, compromising fruit quality. In our previous transcriptomic study of mango fruit, cold storage at suboptimal temperature (5 °C) activated the α-linolenic acid metabolic pathway. To evaluate changes in fruit quality during chilling, we analyzed mango “Keitt” fruit peel volatiles. GC−MS analysis revealed significant modulations in fruit volatiles during storage at suboptimal temperature. Fewer changes were seen in response to the time of storage. The mango volatiles related to aroma, such as δ-3-carene, (Z)-β-ocimene, and terpinolene, were downregulated during the storage at suboptimal temperature. In contrast, C6 and C9 aldehydes and alcoholsα-linolenic acid derivatives 1-hexanal, (Z)-3-hexenal, (Z)-3-hexenol, (E)-2-hexenal, and nonanalwere elevated during suboptimal-temperature storage, before chilling-injury symptoms appeared. Detection of those molecules before chilling symptoms could lead to a new agro-technology to avoid chilling injuries and maintain fruit quality during cold storage at the lowest possible temperature. KEYWORDS: mango fruit, chilling injuries, volatiles, α-linolenic acid-oxidation pathway, oxylipins

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INTRODUCTION Postharvest cold storage is widely used to prolong the storage of fresh produce.1 Mango is consumed worldwide for its delicious taste and aroma and nutritional qualities.2 Fruit aroma components are affected by postharvest techniques, such as cold, heat, irradiation, atmospheric composition, and chemical applications.3 The volatile constituents and their concentrations in mango fruit peel and pulp have been studied in various cultivars4−6 in response to various postharvest conditions, such as heat vapor treatment,7 methyl jasmonate (MeJA) application,8 intact mango fruit compared to infested with fruit fly,9 various stages of fruit ripening,10,11 and hot water dipping or fungicide treatments.12 Moreover, a reduction in aroma volatile production in “Kensington Pride” mango pulp was observed in response to CI.13 Similarly, lowtemperature storage reduce the aromas of different fruits, including mandarin,14 peach,15 and tomato.16 Cold storage is considered the most effective method for prolonging the shelf life of fresh produce. However, tropical fruits such as mango (Mangifera indica) are sensitive to lowtemperature storage.2 Mango fruit is stored at 10−12 °C; storage at lower temperature causes chilling injury (CI) and reduced fruit quality.17 In mango, CI symptoms include lenticel discoloration (red and black spots) and pitting on the peel (Figure 1), reduced aroma and flavor, and more.17 The mango peel is more susceptible to CI than the pulp.18 In our previous work, “Keitt” mango fruit were stored at various temperatures (12, 8, and 5 °C). Mango fruit stored at optimal temperature (12 °C) showed very minor CI symptoms, whereas storage at 5 °C led to severe CI symptoms, including black spots and pitting. Accumulation of those CI symptoms was correlated to severe lipid peroxidation.17 The transcriptomic analysis of “Keitt” mango fruit in response to © XXXX American Chemical Society

Figure 1. Representative pictures of chilling injuries development on “Keitt” mango fruit stored at 5 °C for 3 weeks.

suboptimal temperature storage revealed the activation of several pathways.17 The aim of this manuscript was to characterize the effect of optimal and suboptimal cold storage on volatile constituents and concentrations in “Keitt” mango fruit peel. Transcriptomic evaluation of mango fruit peel17 suggested that storage at suboptimal temperature upregulates the genes of the αlinolenic acid-oxidation pathway, which leads to the synthesis of JA and C6 and C9 aldehydes volatiles before visible CI symptoms appear. Early detection of these volatiles would indicate that the fruit suffers from cold-stress and that the storage temperature should be elevated. In this manner, a dynamic cold storage could be obtained in order to maintain fruit quality for longer periods of time under cold storage. Received: October 5, 2016 Revised: December 26, 2016 Accepted: December 28, 2016

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DOI: 10.1021/acs.jafc.6b04355 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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Figure 2. Upregulation of the α-linolenic acid-oxidation pathway genes in response to cold storage. (A) α-Linolenic acid-oxidation pathway based on the KEGG mapper. Genes circled in red were significantly upregulated during cold storage at 5 °C compared to 12 °C. (B) Heat map of relative expression of genes in the α-linolenic acid-oxidation pathway at two different storage temperatures (5 and 12 °C) at different sampling times (2, 7, and 14 days) compared to harvest time. Z-scores represent rescaled log fold change values. Abbreviations, transcript identifications, and expression profiles are described in Supporting Information Table S1.

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storage at 5 or 12 °C on days 2, 7, 14 and 19, and on days 1 and 7 of additional shelf life storage at 20 °C were sampled and used for GCMS analysis. The optimal (12 °C) and suboptimal (5 °C) temperatures were chosen based on preliminary results. Peel tissue (2 g) of each sample and its replicates was immediately collected in a 20 mL amber vial (LaPhaPack, Langerwehe, Germany) prepared in advance with 5 mL of 20% (w/v) NaCl (Sigma-Aldrich, St Louis, MO), 0.6 g NaCl, and 50 μL of 10 ppm S-2-octanol (Sigma-Aldrich) added as an internal standard. Samples were stored at −20 °C until analysis. On the day of analysis, samples were prewarmed for 1 h at 30 °C on an orbital shaker at 250 rpm. GC−MS Analysis of Mango Volatiles. A solid-phase microextraction (SPME) holder (Agilent, Palo Alto, CA) assembled with fused silica fiber (Supelco, Bellefonte, PA) coated with polydimethylsiloxane (50/30 μM thickness, 1 cm) were used to absorb the volatile compounds. The fiber was desorbed at 250 °C for 3 min in the splitless injection mode to Agilent gas chromatograph series 7890A fitted with an Agilent HP-5MS fused silica capillary column (30 mm long ×0.25 mm ID × 0.25 μm film thickness), coupled to a 5975C MS detector (Agilent). The analytical conditions were adjusted as follows: temperature program, 40 °C for 2 min, raised at 10 °C/min to 150 °C, then at 15 °C/min to 220 °C for 5 min; injector temperature, 250 °C. Sequence total run time was 22.667 min with helium as the carrier gas adjusted to a flow rate of 0.7941 mL/min in splitless mode, with ionization energy of 70 eV. Identification and Quantitation of Aroma Volatiles. Compounds were tentatively identified by use of the NIST mass spectral library (version 5) with Chemstation version E.02.00.493. The retention times of a series of straight-chain alkanes (C5−C20) (Sigma-Aldrich) were used to calculate the retention indices (RIs) for all identified compounds, and the identities were confirmed by comparison of their linear RIs with the Kovats RIs of published data.4,6,12 Volatile compounds were semiquantitated relative to the internal standard. The concentration of volatiles was expressed as μg/ gFW. Statistical analysis. Data are presented as mean ± SE and analysis involved one-way ANOVA. P < 0.05 was considered statistically significant by tukey’s test with Sigma Stat 3.5 (Systat Software Inc., San Jose, CA). Principal component analysis (PCA), volatiles were selected with P < 0.05 by Kruskal−Wallis Test, correlation analysis and heatmapping for the volatile metabolites were performed using the webbased MetaboAnalyst 3.0 (http://www.metaboanalyst.ca).26

MATERIALS AND METHODS

Chemicals. NaCl, S-2-octanol, and straight-chain alkanes (C5−C20) were purchased from (Sigma-Aldrich, USA). Deionized water was collected from the Milli-Q water purification system by Millipore Corp. (Saint-Quentin, France). All other chemicals used were of analytical grade. Fruit Material and Suboptimal Temperature Storage. Mango fruit (Mangifera indica L., cv. Keitt) were obtained 2 h after harvest from a commercial orchard (Mor Hasharon storage house, Israel) and transported (1 h) to the Agricultural Research Organization (Israel). Uniform, unblemished fruit weighing 424 ± 16 g were selected, washed with tap water, and air-dried. On the same day (day of harvest), six biological replicates, each with 10 fruit, were stored at 5 and 12 °C for 19 days in the cold-storage rooms, and a further 7 days at 20 °C (shelf life storage) and physiological and pathological parameters were evaluated.17 The temperature in the cold-storage room was monitored by a DAQ tool (double-strand wire logger/dataacquisition control system; T.M.I. Barak Ltd., Ramat-Gan, Israel). Fruit core temperature was monitored using a MicroLite data logger LITE5032P-EXT-A (Fourier Technologies, Ramat-Gan, Israel), by inserting the probe to 5 cm depth near the fruit calyx. The experiments were repeated in three consecutive seasons2013, 2014 and 2015 and showed similar physiological and pathological results.17 The presented is the experiment with cv. Keitt in 2014. RNA-Sequencing (RNA-Seq). Total RNA from the peel tissue of mango fruit at harvest and during storage for 2, 7, and 14 days at 5 or 12 °C was extracted, cDNA library were prepared, and the RNA-Seq protocol using the Illumina Hiseq2000 system (San Diego, CA) were followed as described previously.17 Cleaned sequences were mapped to a reference mango transcriptome19 using the Bowtie2 software.20 Transcripts abundance were calculated by the RSEM software package.21 Differentially expressed transcripts were evaluated by EdgeR package22 of the Bioconductor R packages.23 Following a false discovery rate (FDR). Following the Trinity protocol24 the expression was normalized. Transcripts of upregulated clusters were annotated with the Kyoto Encyclopedia of Genes and Genomes (KEGG). Upregulated transcripts with KEGG orthology descriptions were mapped to their associated KEGG pathways. Heat-mapping of upregulated genes was performed on normalized data using the R package “FactomineR”.25 Sample Preparation for GC−MS Analysis. Mango fruit peel tissue (2 mm deep) was randomly sliced from six fruits for each replicate, and three biological replicates (each biological replicate had two technical replicate) for each time point as harvest and after of cold B

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RESULTS

Chilling Injuries and Activation of the α-Linolenic Acid Metabolic Pathway. To better understand and characterize the dynamics of transcript expression, RNA deep sequencing was conducted using Illumina HiSeq 2000 on samples extracted from the peel part of “Keitt” mango fruit stored at 12 or 5 °C for 2, 7, and 14 days, as described previously.17 To identify chilling-related pathways that are activated during cold storage, the upregulated clusters were BLAST against the KEGG database (http://www.genome.jp/ kegg/), identified, and collated to the upregulated pathways. The α-linolenic acid-oxidation pathway was activated in mango in response to chilling stress. The key genes of the αlinolenic acid metabolic pathway, such as those encoding 13Slipoxygenase (LOX), allene oxide synthase (AOS), allene oxide cyclase (AOC), and 12-oxophytodienoate reductase (OPR), were significantly upregulated at 5 °C compared to fruit on the day of harvest and those stored at 12 °C (Figure 2). The relative expression of each transcript during fruit storage at 5 or 12 °C after 2, 7, and 14 days of storage in comparison to fruit on the day of harvest is represented in a heat map, where upregulated and downregulated genes are presented in red and green, respectively (Figure 2B). Most of these genes were upregulated after 2 days of cold storage at 5 °C. The mango transcriptome revealed 10 LOX genes, of which four 13S-LOX genes were upregulated. Two LOX genes LOXi (comp20944) and LOXiv (comp17114)were upregulated 2.1- and 4.3-fold, respectively, after 2 days of cold storage at 5 °C vs 12 °C. LOXii and LOXiii (comp31741 and comp39918) were upregulated 2.3- and 2.5-fold, respectively, after 7 days of storage at 5 °C vs 12 °C (Figure 2B). Focusing on the MeJA biosynthesis pathway, downstream of LOX, three AOS isoform-encoding genes (comp11945, comp26483, comp29600) were upregulated 4.2-, 3.1-, and 4.6-fold, respectively, in response to 2 days of cold storage at 5 °C vs 12 °C storage (Figure 2B). AOC (comp13545) was upregulated at 5 °C vs 12 °C at all-time points, with a maximum 9.2-fold increase after 7 days in cold storage. OPR (comp18454) was upregulated at 5 °C vs 12 °C at all-time points, with a maximum 2.4-fold increase after 7 days of cold storage. In addition to the upregulation of transcripts related to the αlinolenic acid metabolic pathway during storage at suboptimal temperature (Figure 2), also various transcripts related to secondary metabolites modification and the release of natural aroma volatiles were intensely modified, mostly down-regulated during storage at suboptimal temperature. Those transcripts were 12 transcripts of glycosyl hydrolases, 10 transcripts of alcohol dehydrogenases, 19 transcripts of SAM-methyltransferases, one transcript of terpene synthases, 5 transcripts of carotenoid cleavage dioxygenases, and 7 transcripts of carboxylesterases. Global Volatile Changes during Cold Storage. Using GC−MS, we monitored the profile of “Keitt” mango peel volatiles at harvest, and on 2, 7, 14, and 19 days of storage at 5 and 12 °C. The change in volatile profile among the samples was determined by the spatial scattering of total fruit volatiles, represented by principal component analysis (PCA). According to their spatial scattering, the samples could be divided into three distinct groups: harvest, stored at 5 °C, and stored at 12 °C (Figure 3A). The volatile profile of the “harvest” group (blue cloud) was closely related to the profile of the “12 °C storage” group (the optimal temperature; red circle). However,

Figure 3. Overview of changes in mango fruit volatiles in response to cold storage. (A) PCA presenting spatial scattering of total fruit volatiles at harvest (blue X) and on days 2, 7, 14, and 19 of cold storage at 5 °C (green plus sign) or 12 °C (red triangle). (B) Biplot of PCA analysis described in (A) of tentative identified volatiles.

the volatile profile of the “5 °C storage” group (green circle) was distinct from the other two groups. The duration of cold storage had only a minor effect on the change in the total volatile profile. Overall, 43 different putative volatiles were detected from peel tissue of mango fruit cv. Keitt. Among these, 17 tentative volatiles were significantly altered during storage at the different temperatures (Figure 3B). The changes in each compound (presented as a vector arrow) determined the differences in their total spatial distribution, presented as a biplot of PCA (Figure 3B). Figure 3B shows the 9 compounds that were elevated in fruit stored at 5 °C ((Z)-3-hexenal and (Z)-3hexenol, (E)-2-hexenal, 1-hexanal, nonanal, octanal, and C

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Journal of Agricultural and Food Chemistry heptanal), as reflected by the right-pointing arrows. In addition, 11 compounds were elevated at harvest or during storage at 12 °C (copaene, β-selinene, β-caryophyllene, α-pinene, δ-3-carene, α-humulene, (E)-3-carene-2-ol, unknown monoterpene, (Z)-βocimene, α-phellandrene, and terpinolene), as reflected by the left-pointing arrows (Figure 3B). A correlation analysis of all 43 compounds showed strong coactivation among various monoterpenes and sesquiterpenes, among alcohols (1-butanol, 3-methyl-1-butanol, and 2-methyl1-butanol), and among the aldehydes and their alcohol derivatives (1-hexanal, (Z)-3-hexenal, (Z)-3-hexenol, (E)-2hexenal, nonanal, and octanal) (Figure 4).

Figure 5. Heat map of the relative concentrations of volatiles in the different samples. The tentative identified volatile concentration was monitored at harvest (control) and on days 2, 7, and 14 of cold storage (5 and 12 °C).

shelf life (20 °C) showed that all of these compounds increased significantly after 2 days of cold storage at 5 °C and remained elevated until fruit transfer to 20 °C (Figure 6). The transfer of mango fruit to the higher temperature of 20 °C (shelf life) was accompanied by a sharp decrease in the concentrations of C6 and C9 aldehydes in fruit stored at 5 °C and a slight increase for fruit stored at 12 °C (Figure 6).

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DISCUSSION Cold storage is considered the most effective method of prolonging the shelf life of fresh produce, because it maintains fruit quality and inhibits fruit deterioration.1 The subtropical mango fruit is considered to be susceptible to cold storage. Mango fruit are stored at 10−12 °C. Storage at lower temperatures for long periods leads to CI symptoms, decreased fruit flavor and quality, and, ultimately, fruit loss. GC−MS analysis of “Keitt” mango fruit volatiles at harvest and during cold storage at the optimal 12 °C and the chilling temperature of 5 °C showed 43 different compounds, most of them having been previously detected in mango.4,6,10 Using PCA, we showed that the overall volatile profile in fruit stored at 12 °C is similar to the volatile profile of fruit at harvest. In contrast, a significant shift in the volatile profile was observed in fruit that were stored at 5 °C. This shift was correlated to several volatiles (Figure 3B). Correlation analysis showed that volatiles of a similar nature were coexpressed at the different temperatures, hinting at common transcriptional regulation of those compounds (Figure 4). Aroma volatiles extracted from the pulp of “Kensington Pride” mango fruit, mainly monoterpenes and sesquiterpenes, were significantly reduced during storage at suboptimal temperature.13 Interestingly, various transcripts related to secondary metabolites modification and the release of natural aroma volatiles were down-regulated during storage at suboptimal temperature. These transcriptional changes led to a reduction in several aroma volatile compounds such as δ-3carene, (Z)-β-ocimene, and terpinolene in the peel of “Keitt”

Figure 4. Correlation heat map of coexpression between different volatiles (tentatively identified, 1 represents perfect coexpression, 0 represents no correlation, −1 represents perfect negative coexpression).

Specific Changes in Volatiles during Cold Storage. Ten tentative volatiles were significantly regulated by storage temperature. The relative concentrations of those volatiles in the different samples are presented in a heat map (Figure 5). Interestingly, chilling stress increased the volatiles of C6 and C9 aldehydes and their alcohol derivatives, which are the oxidative products of the α-linolenic acid and oxylipin pathways. The significantly increased compounds at 5 °C compared to harvest and storage at 12 °C were as follows: 1-hexanal, (Z)-3-hexenal, (Z)-3-hexenol, (E)-2-hexenal, nonanal, heptanal, and octanal. The volatiles δ-3-carene, (Z)-β-ocimene, and terpinolene were found at higher concentrations in fruit at harvest and stored at 12 °C (Figure 5) than in fruit stored at 5 °C. Closer evaluation of the concentrations of C6 and C9 aldehydes (1-hexanal, (Z)-3hexenal, (Z)-3-hexenol, (E)-2-hexenal, and nonanal) during 19 days of cold storage (5 or 12 °C) and an additional 7 days of D

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Figure 6. Changes in C6 and C9 volatile concentrations in cold storage. Volatiles’ concentrations were monitored at harvest and during cold storage at 5 °C (gray) and 12 °C (black) for 26 days. At day 19 the fruit was transfer to shelf-life conditions at 20 °C. (A) 1-hexanal; (B) (Z)-3-hexenol; (C) (Z)-3-hexenal; (D) Nonanal; (E) (E)-2-hexenal. The concentrations of the volatiles are presented in μg/g FW of fruit peel, and the values are means ± SE.

transcript of PLC (comp37611) and four transcripts of PLD (comp18965, comp24723, comp25980, and comp17158) were significantly upregulated.17 LOX is a key gene in the response to chilling, as it initiates the first step of α-linolenic acid oxidation and the synthesis of MeJA.34−36 Here, we show that four LOX transcripts were upregulated in response to chilling (Figure 2). Downstream of LOX in the JA biosynthesis pathway is AOS, which is an important enzyme in the defense response to wounding37 that acts as a key enzyme in oxylipin metabolism.38 Here we show that three mango AOS transcripts were activated in response to chilling (comp11945, comp26483, and comp29600). The oxidation and degradation of α-linolenic acid also led to the release of oxylipins, such as the volatile alkenes C6 and C9.39 This reaction is mediated by HPL, which was consistently expressed during storage at 12 and 5 °C. The resultant C6 and C9 oxylipins, which are known as green leaf volatiles, are released mainly in response to wounding and biotic stress.39 Interestingly, in our study, the GC−MS analysis showed that the same oxylipin volatiles of C6 and C9 alkenes (1-hexanal, (Z)-3-hexenal, (Z)-3-hexenol, (E)-2-hexenal, and nonanal) are the major upregulated compounds in response to chilling in mango peel. Interestingly, suboptimal temperature storage did not elevate the same oxylipin volatiles in mango pulp.13 (E)-2-

mango fruit stored at suboptimal temperature. This supports previous observations of compromised fruit aroma and taste in chilled mango fruit27 and suggests that the loss of aroma is due to down-regulation of major transcripts and followed by a decrease in volatiles such as δ-3-carene, (Z)-β-ocimene, and terpinolene. Lipid peroxidation is considered a main characteristic of fruit suffering from CIs.28 An elevation in lipid peroxidation was observed by luminescence and MDA biochemical analysis in “Keitt” mango fruit after 14 days of storage at 5 °C, just before the visual CI symptoms appeared.17 However, the transcripts in the metabolic pathway of fatty-acid oxidation of α-linolenic acid metabolism were activated after 2 days of storage at 5 °C (Figure 2). In this regard, the oxidation of α-linolenic acid is connected not only to fatty acid oxidation, but also to much more basic defense responses to abiotic stressJA synthesis and oxylipin signalingwhich further activated the fruit defense response. MeJA is a major compound regulating the global plant response to abiotic stress. MeJA is also known to activate chilling resistance in various fruits.29−32 The plant defense response to wounding involves phospholipase C (PLC) and phospholipase D (PLD), which cause the release of α-linolenic acid, activation of LOX, and synthesis of JA.33 In the mango fruit’s response to chilling, one E

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jasmonate; HPL, hydroperoxide lyase; FPKM, kilobase per million reads mapped; KEGG, Kyoto Encyclopedia of Genes and Genomes; MDA, malondialdehyde; LOX, lipoxygenase; AOS, allene oxide synthase; AOC, allene oxide cyclase; OPR, 12-oxophytodienoate reductase; PCA, principal component analysis; PLC, phospholipase C; PLD, phospholipase D

Hexenal is an important green leaf volatile that has been suggested as a signaling molecule inducing abiotic-stress-related transcription factors such as WRKY40 and WRKY6.40,41 The volatiles (E)-2-hexenal, (Z)-3-hexenal and (E)-3-hexenal have been shown to have antibacterial42,43 and antifungal44 activity, and they have been suggested to act as intra- and interplant volatile signals.39 When leaves are damaged by herbivores or pathogens, they begin to form these volatiles, some of which diffuse into the leaves or are vaporized to reach other leaves and induce defense responses. This characteristic of intra- and interplant volatile signals could be very beneficial for application during postharvest cold-chain transportation in small chambers. Interestingly, application of these compounds has already been considered to improve the shelf life and safety of processed fruit.45 Indeed, fruits such as apple, pear, cherry, peach, grape, banana, and tomato have shown enhanced shelf life and quality in response to the application of hexanal.46−48 These applications probably increase the natural defense response in the harvested fruit, thereby helping the fruit cope with various postharvest abiotic and biotic stresses. This study showed that the harvested mango fruit responds to abiotic stress, such as chilling temperature storage, by activating the α-linolenic acid pathway, which ends in the synthesis of JA and the oxylipin volatiles of C6 and C9 aldehydes and alcohols. The release of these volatiles could be detected long before the visible CI symptoms could be observed. Thus, detection of these volatiles might indicate that the fruits are starting to suffer from chilling stress, and ideally, the temperature should be elevated to avoid chilling and to prolong storage.

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(1) McGlasson, W.; Scott, K.; Mendoza, D., Jr The refrigerated storage of tropical and subtropical products. Int. J. Refrig. 1979, 2, 199−206. (2) Sivakumar, D.; Jiang, Y.; Yahia, E. M. Maintaining mango (Mangifera indica L.) fruit quality during the export chain. Food Res. Int. 2011, 44, 1254−1263. (3) El Hadi, M.; Zhang, F.-J.; Wu, F.-F.; Zhou, C.-H.; Tao, J. Advances in fruit aroma volatile research. Molecules 2013, 18, 8200. (4) Pandit, S. S.; Chidley, H. G.; Kulkarni, R. S.; Pujari, K. H.; Giri, A. P.; Gupta, V. S. Cultivar relationships in mango based on fruit volatile profiles. Food Chem. 2009, 114, 363−372. (5) de Jesus Benevides, C. M.; de Almeida Bezerra, M.; Pereira, P. A. P.; de Andrade, J. B. HS-SPME/GC-MS Analysis of VOC and multivariate techniques applied to the discrimination of brazilian varieties of Mango. Am. J. Anal. Chem. 2014, 5, 157. (6) Pino, J. A.; Mesa, J.; Munoz, Y.; Marti, M. P.; Marbot, R. Volatile components from Mango (Mangifera indica L.) cultivars. J. Agric. Food Chem. 2005, 53, 2213−2223. (7) Singh, S. P.; Saini, M. K. Postharvest vapour heat treatment as a phytosanitary measure influences the aroma volatiles profile of mango fruit. Food Chem. 2014, 164, 387−395. (8) Lalel, H.; Singh, Z.; Tan, S. The role of methyl jasmonate in mango ripening and biosynthesis of aroma volatile compounds. J. Hortic. Sci. Biotechnol. 2003, 78, 470−484. (9) Carrasco, M.; Montoya, P.; Cruz-lopez, L.; Rojas, J. C. Response of the fruit fly parasitoid Diachasmimorpha longicaudata (Hymenoptera: Braconidae) to mango fruit volatiles. Environ. Entomol. 2005, 34, 576−583. (10) Lalel, H. J. D.; Singh, Z.; Tan, S. C. Aroma volatiles production during fruit ripening of ’Kensington Pride’ mango. Postharvest Biol. Technol. 2003, 27, 323−336. (11) Lebrun, M.; Plotto, A.; Goodner, K.; Ducamp, M. N.; Baldwin, E. Discrimination of mango fruit maturity by volatiles using the electronic nose and gas chromatography. Postharvest Biol. Technol. 2008, 48, 122−131. (12) Dang, K. T. H.; Singh, Z.; Swinny, E. E. Impact of postharvest disease control methods and cold storage on volatiles, color development and fruit quality in ripe ‘Kensington Pride’ mangoes. J. Agric. Food Chem. 2008, 56, 10667−10674. (13) Nair, S.; Singh, Z.; Tan, S. C. Aroma volatiles emission in relation to chilling injury in ‘Kensington Pride’ mango fruit. J. Hortic. Sci. Biotechnol. 2003, 78, 866−873. (14) Tietel, Z.; Lewinsohn, E.; Fallik, E.; Porat, R. Importance of storage temperatures in maintaining flavor and quality of mandarins. Postharvest Biol. Technol. 2012, 64, 175−182. (15) Raffo, A.; Nardo, N.; Tabilio, M. R.; Paoletti, F. Effects of cold storage on aroma compounds of white- and yellow-fleshed peaches. Eur. Food Res. Technol. 2008, 226, 1503−1512. (16) Díaz de León-Sánchez, F.; Pelayo-Zaldívar, C.; Rivera-Cabrera, F.; Ponce-Valadez, M.; Á vila-Alejandre, X.; Fernández, F. J.; EscalonaBuendía, H. B.; Pérez-Flores, L. J. Effect of refrigerated storage on aroma and alcohol dehydrogenase activity in tomato fruit. Postharvest Biol. Technol. 2009, 54, 93−100. (17) Sivankalyani, V.; Noa, S.; Feygenberg, O.; Hanita, Z.; Dalia, M.; Alkan, N., Transcriptome dynamics in mango fruit peel reveals mechanisms of chilling stress. Front. Plant Sci. 2016, 7.10.3389/ fpls.2016.01579 (18) Farooqi, W.; Sattar, A., Jr.; Daud, K.; Hussain, M. In Studies on the postharvest chilling sensitivity of mango f ruit (Mangifera indica L.),

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jafc.6b04355. Table S1. Concentrations of all volatile compounds found in mango peels at harvest, during cold storage at 5 and 12 °C for 19 days, and after 7 additional days of shelf life at 20 °C. Table S2. Abbreviations and the upregulated transcripts of the α-linolenic acid-oxidation pathway. Table S3. Identified aroma related transcripts of mango fruit peel (PDF)

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REFERENCES

AUTHOR INFORMATION

Corresponding Author

*Tel.: 972-3-9683605. Fax: 972-3-9683622. E-mail: noamal@ volcani.agri.gov.il. ORCID

Noam Alkan: 0000-0001-7163-6514 Funding

This manuscript is contribution number 766/16 from the Agricultural Research Organization, the Volcani Center, Israel. This research was supported by the Chief Scientist of the Israeli Ministry of Agriculture and Rural Development (Grant No. 430-0544-14). Notes

The authors declare no competing financial interest.

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ABBREVIATIONS USED CI, chilling injury; GC−MS, gas chromatography−mass spectrometry; MeJA, methyl jasmonate; MeJA, methyl F

DOI: 10.1021/acs.jafc.6b04355 J. Agric. Food Chem. XXXX, XXX, XXX−XXX

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DOI: 10.1021/acs.jafc.6b04355 J. Agric. Food Chem. XXXX, XXX, XXX−XXX