Preharvest sprays and their effects on the postharvest quality of fruit

Purpose of review: This paper reviews studies on the effects of preharvest spray treatments on the postharvest quality and storage potential of fruits, with the objective of summarising the main effects in each case and identifying major topics requiring further research. Findings: The literature survey shows that most of the studies on preharvest sprays have considered either calcium or growth regulator treatments. Calcium applications are generally reported to delay ripening, decrease postharvest rots and alterations, and extend the keeping period, but their effects are partially dependent on the calcium source and formulation used, and phytotoxicity has also been occasionally observed. Preharvest sprays with growth regulators such as aminoethoxyvinylglycine, gibberellins or polyamines have also been studied and have shown promising potential for delaying ripening and improving storage potential or particular quality traits. Directions for future research: Although some common effects have been identified on fruit physiology for a particular treatment, a certain degree of variability across fruit types or cultivars has been observed in all cases. The suitability and the particular conditions of each treatment should be assessed and adjusted for each fruit type. In addition, because fruit metabolism is complex and strictly regulated, improved keeping potential may be contradicted by detrimental effects on eating quality, meaning that treatment effects should be evaluated as a whole. A third aspect worthy of more intense research efforts involves effects on key quality attributes such as aroma or bioactive compound contents, or on other traits relevant for quality preservation such as fruit cuticles.


Introduction
Fruit condition at harvest is essential for postharvest performance of produce. This entails an appropriate maturity stage, but also involves other aspects such as nutritional status and content of particular minerals. For this reason, preharvest spraying with certain compounds has become a widely used practice during on-tree development of some economically relevant fruit species. This paper reviews published reports on the effects of such treatments on postharvest quality and storage potential of fruit.
Since a number of postharvest alterations arise at least partially from mineral deficiency, many preharvest sprays are aimed at supplementing the fruit with a higher content of that particular mineral element. Because of its impact on different aspects related to fruit quality, calcium has been particularly used for preharvest treatment of these commodities, and the object of intensive research efforts. However, this is not the only feasible approach for the modulation of Lara / Stewart Postharvest Review 2013, 3:5 2 fruit quality attributes prior to harvest. Postharvest quality of fruit produce can also be manipulated by preharvest application of a range of different compounds, including plant growth regulators such as gibberellins, polyamines, ethyleneinhibiting or -releasing chemicals, fungicides or chitosan. Among these, the effects of preharvest applications of fungicides and the ethylene antagonist 1-methyl-cyclopropene (1-MCP) on postharvest quality are reviewed elsewhere in this issue, and will not be covered here.
Although some common effects are usually recognisable for a given preharvest spray treatment (Tables 1 and 2), some variability may exist across fruit species or even cultivars. This means that it is necessary to confirm the suitability or to optimise the application protocol on a case-by-case basis. On the other hand, the beneficial effects on a given attribute may be counteracted by a detrimental influence on another trait.
Furthermore, most published studies on the influence of preharvest sprays on postharvest quality have targeted the extension of storage potential and marketing possibilities through the preservation of the usual commercial quality attributes or the control of the incidence of decay and physiological disorders. However, some major traits contributing to the sensory quality of fruit, such as aroma, have been largely overlooked, and would need intensive work in order to assure optimal quality of fresh fruit reaching the consumers.

Calcium sprays
The divalent calcium cation (Ca 2+ ) is required for several key physiological processes related to ripening-related changes, including those in cell wall structure, membrane integrity and functionality, activity of particular enzymes, or signal transduction. Calcium deficiency in fruit produce can result in physiological disorders of considerable economic relevance such as cracking, vitrescence or bitter pit [1*]. Therefore, calcium treatments have the potential to delay fruit ripening and senescence, and to show beneficial effects on a wide range of attributes related to quality and storability of produce. Different procedures have been used successfully for postharvest calcium treatment of fresh and minimallyprocessed fruit [2*]. Yet calcium applications can also be undertaken prior to commercial harvest, in order to provide an extra supply of the mineral before the deficiency symptoms appear. Because calcium uptake from the soil and its movement to aerial plant organs are limited, direct spray applications onto the plant canopy are preferable, as they often allow effective increase of calcium content in the fruit [3]. Nevertheless, this may not necessarily be the case in all instances: cantaloupe melon fruit did not benefit from preharvest applications of either amino acid-chelated or mannitolcomplexed calcium, while treated honeydew fruit displayed higher calcium concentrations associated with improved firmness and marketability [4]. Similarly, no consistent effects on calcium content in fruit have been reported occasionally for calcium-sprayed apples [5,6]. In other cases, phytotoxic effects have been observed [7,8], which indicates the need to optimise treatment conditions individually for each species or cultivar.
While the chloride salt is the most frequently used source of calcium for these preharvest sprays (Table 1), some studies have also been undertaken in which different calcium sources or formulations were applied and compared in relation to their suitability for increasing calcium content in the fruit or for extending the keeping period after harvest [4,6,[9][10][11][12][13][14][15][16][17]. In some cases, the calcium formulation has been shown to influence the efficiency of treatment, particularly regarding the incidence of physiological alterations or decay. In addition to source or formulation, season-to-season variability in the effectiveness of preharvest calcium applications has also been observed occasionally for apple [18,19] and kiwifruit [12].
Many studies have addressed the effects of preharvest calcium applications on the standard attributes generally used to CaCl 2 1% Delayed softening and increased storage life potential; decreased incidence of low temperature breakdown. [26] CaCl 2 0.25 to 1.5% Increased calcium in the flesh; no effect on firmness or pitting incidence. [68] CaCl 2 0.8% Delayed softening; phytotoxic effects. Diverse (commercial Ca 2+ -containing products) Increased calcium content; altered antioxidant power and total phenols and ascorbic acid contents (formulation-dependent). Increased firmness (season-dependent); no effects on SSC or TA. [12] . [34] Plum CaCl 2 1.6 kg/ha Reduced postharvest decay in soft cultivar; no effect in firm cultivars. Non-significant effects on soluble solids or firmness. [71] Rambutan Chelated calcium 5.63 mg/L Decreased decay incidence and severity, weight loss and browning. Increased peel thickness. No effects on TA, SSC or ascorbic acid content. [28] Strawberry CaCl 2 0.4% Increased firmness. Less sweet taste; necrotic brown spots after storage. [7] CaSO 4 0.04 to 0.2% Ca 2+ Non-significant. [72] Sweet cherry Ca(OH) 2 0.7% Reduced cracking incidence; higher firmness, SSC and calcium content both in skin and flesh. [25] CaCl 2 0.5% Higher SSC and phenolics content; reduced decay and cuticular fractures. Non-significant effects on TA, colour or firmness. Increased weight loss during storage.

L/ha
Non-significant for cantaloupe fruit. Improved firmness, marketability and calcium content in honeydew fruit (no effect on sugars or taste). [4] Olive CaCl 2 0.65% Delayed calcium loss, softening and pectin solubilisation.
No effects on colour change, respiration or ethylene production rates. [33] *β-Gal, β-galactosidase; β-Xyl, β-xylosidase; ADH, alcohol dehydrogenase; AFase, α-L-arabinofuranosidase; PDC, pyruvate decarboxylase; PL, pectate lyase; PME, pectinmethylesterase 5 evaluate commercial quality of fruit, such as firmness, titratable acidity (TA), soluble solids content (SSC) or postharvest rots and alterations. Calcium sprays have been generally reported to delay ripening as indicated by respiration rates or ethylene production, to increase fruit firmness and TA both at harvest and after storage, and to decrease the incidence of postharvest decay (Table 1). In a few instances, significant changes in antioxidant capacity or the content of antioxidant compounds such as phenols and ascorbic acid have also been found [12,20,21]. These treatments have also been shown to prevent to a large extent the occurrence of commercially relevant physiological and storage disorders. For example, preharvest calcium sprays reportedly reduced the incidence of bitter pit, lenticel blotch pit, scald and internal breakdown in apple [9,10,18,19,[22][23][24], of cork spot in pear [22], and of cracking in sweet cherry [21,25]. These treatments also help prevent or reduce chilling injury and internal browning in susceptible fruit species such as kiwifruit [26], peach [17], pear [27] and rambutan [28].
Whereas reported effects of preharvest calcium sprays on firmness, decay and alterations appear to be quite general, their influence on other quality indicators such as SSC, SSC/ TA ratios, colour or weight loss have been observed to be much more erratic, non-significant or even contradictory (Table 1). A part of this variability may be related to genotypic differences among fruit species or cultivars, or to the calcium concentration or formulation used. In other cases, though, these discrepancies may prove more difficult to ascribe; the same calcium source and concentration applied to the same fruit species resulting in clearly different effects (compare, for example, references [7] and [29]).
Although improved firmness retention is frequently cited as a major general effect of preharvest calcium applications, the biochemical basis for delayed firmness loss in treated fruit has received less attention. However, some information is available for apple, blueberry, kiwifruit, olive, peach, nectarine and pepper. Delayed softening has been observed to arise from delayed pectin solubilisation and matrix glycan breakdown in treated fruit [15,16,20,[30][31][32][33][34]. Indeed, exogenous calcium can favour the formation of non-covalent cross-links between polyuronides through calcium bridges, thus preventing the dissolution of the middle lamella and reinforcing the 6 cell wall structure. Accordingly, calcium applications often result in increased yields of the chelator-soluble fraction of pectins, comprised mainly of the non-covalently bound cell wall polyuronides ( Figure 1A), associated with better retention of total uronic acids. In addition to directly reinforcing the cell wall structure, calcium may also improve firmness retention through the modulation of some cell wallmodifying enzyme activities, as treated apple and peach fruit have been found to display lower levels of PME, PL, PG, β-Gal, AFase or β-Xyl activity [15,16,30,31]. The effects of calcium on flesh firmness and cell wall composition are not simply related to its electrostatic properties or to its properties as a divalent cation, but must involve some specific effect, as treatment of apple fruit with strontium chloride (SrCl 2 ) failed to mimic the effects of a similar treatment with CaCl 2 on cell wall properties or β-Gal activity [35].
The objectives of preharvest calcium applications have been focused fundamentally on the prevention of physiological alterations and on the extension of shelf life as indicated by the usual commercial quality attributes such as firmness, SSC or TA. In contrast, little information has been reported to date on the effects of calcium treatments on fruit aroma. This attribute has been largely disregarded, even though it is a major contributor to sensory quality and consumer acceptance of fruit. In the case of apple, for example, the usual practice of harvesting the fruit before reaching full ripeness, aimed at obtaining higher firmness levels and thus better storage potential, often leads to deficient aroma as the production of related volatile compounds develops with maturity stage. In this context, a recent report has shown the potential of preharvest calcium sprays for improving this important quality trait [36**]; treated 'Fuji Kiku-8' apples not only displayed higher firmness and TA, but also showed increased production of aroma-related volatile compounds ( Figure 1B), and particularly of the impact compounds contributing to the characteristic aroma. Improved emission of key volatile compounds was the result of the enhancement of major enzyme activities providing the necessary precursors for the final reaction in the biosynthetic pathway.

Growth regulator sprays
Preharvest sprays with certain growth regulators can also be applied with the aim of modifying the ripening process of fruit, or of modulating the development of a particular attribute with influence on storage potential or commercial appeal ( Table 2). Although some common effects of each particular compound on fruit physiology can be identified in each case, the results of such applications on postharvest quality have shown a certain degree of variability across fruit types or even cultivars.

Ethylene-inhibiting and -releasing compounds
Aminoethoxyvinylglycine (AVG) acts as a competitive inhibitor in the conversion of S-adenosylmethionine (SAM) to the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). Owing to this capacity to block reversibly the ethylene biosynthesis pathway, pre-and postharvest AVG applications have been tested as a means to delay ripening and to enhance storage potential of climacteric fruits. In general, significant delays in fruit ripening have been reported to result from preharvest AVG sprays, with significant decreases in ethylene production and higher firmness levels in treated fruit ( Table 2) resulting in extended shelf life.
The literature survey, though, also indicates that some other practical benefits of these treatments are species-specific: preharvest AVG applications have been shown to improve uniformity of maturity stage at harvest in peach [37], but not in melon [38]. No consistent influence on the control of decay incidence has been demonstrated either. Furthermore, sprays may be not a suitable method for AVG application in all cases: for example, no effects of AVG treatment were found in melon fruit when the compound was applied as a spray, whereas fruit firmness was increased significantly when it was directly injected to the soil into the root zone [39].
However, in spite of the generally beneficial effects of AVG applications on storage potential as indicated by the usual standard quality parameters, it should be kept in mind that treatment effects should be evaluated in all instances as a whole, since extension of storability could be counteracted by undesirable effects on sensory quality and consumer acceptance of produce. A preharvest AVG treatment has been reported to decrease the content of bitterness-contributing phenolics in olive [40], which suggests that eating quality was improved. Nevertheless, ripening delays will typically result in delayed development of the characteristic flavour, which is strictly dependent on maturity stage [36**]. Since flavour is, together with texture, a major attribute contributing to consumer acceptance of the main fruit commodities with commercial importance, insufficient development of this important quality trait could result in detrimental effects on sensory quality. For example, total flavour-contributing ester production by 'Redchief Delicious' apple peel tissue after cold storage was reduced by 44% in response to preharvest AVG sprays [41], arising from a decreased supply of alcohol precursors for ester biosynthesis. Similarly, preharvest AVG application had a negative impact on the biosynthesis of aroma volatiles by 'Delbarde Estivale' apple fruit, particularly of those having the most impact on the characteristic aroma [42], also as a result of impaired precursor supply (Figure 2). Although no sensory analyses were undertaken in either study, it is apparent that the eating quality of produce must have been compromised, and thus that treatment benefits in terms of storage potential probably did not compensate for the loss of sensory quality.
In contrast to AVG, ethephon penetrates into tissues and decomposes to ethylene, phosphate and chloride ion in aqueous solutions above pH 4-5, and thus such treatments are expected to show opposite effects to those of AVG on fruit physiology. As an ethylene-liberating compound, preharvest [74] Increased incidence of internal browning disorders; no effect on IEC.
[77] Decreased production of ethylene, TA and flavour-contributing volatile esters. No effect on SSC, respiration rates or AAT* activity. [41] Higher firmness and delayed colour changes; lower ethylene and CO 2 production. Minor effects on TA and SSC. Decreased production of aroma-related volatile esters associated with lower LOX, HPL, PDC and ADH* activities. [42] Melon Lower ethylene production. No effect on firmness, SSC, decay incidence or uniformity of fruit maturity.
[82] Increased peel puncture resistance; delayed colour changes; decreased SSC and decay. No effect on juice content, TA or SSC/TA ratio. [83] Mandarin Decreased TA and juice content. Increased ascorbic acid content, SSC and SSC/TA ratio. [48] Mango Higher TA, ascorbic acid and total chlorophyll contents; lower SSC, SSC/TA ratios, total carotenoid content, and amylase and peroxidase activities. [49] Plum Higher firmness, TA, and resistance to flesh compression and penetration; delayed colour changes; lower SSC. [78,79] Sour cherry Enhanced firmness and storage potential. [84] Sweet cherry Higher firmness and TA; delayed softening and fruit maturation; decreased PG* and cellulase activities (cultivar-dependent). No effect on SSC, β-Gal* or β-glucosidase activities. [50] Firmer, heavier and larger fruit, better preservation of pedicels. No effect on colour or SSC.
[86] Tangerine Increased peel puncture resistance; delayed colour changes; decreased SSC and decay. No effect on juice content, TA or SSC/TA ratio. [83] ... Table 2 continues on page 8 Lara / Stewart Postharvest Review 2013, 3:5 8 ethephon applications can be used to promote fruit ripening or to aid harvest by stimulating abscission. Although reported research has demonstrated limited effect on the usual indicators of fruit quality [43,44], preharvest ethephon treatments can also facilitate postharvest operations by improving uniformity of fruit maturity at harvest [45] or by helping with the removal of troublesome surface structures such as glochids of cactus pear [46].

Gibberellins
Gibberellic acid (GA 3 ) is a pentacyclic diterpene acid which promotes plant cell growth and elongation. It is considered a 'juvenile' plant growth regulator, and as such has been observed to delay ripening and senescence in some fruits, and to improve certain quality characteristics (Table 2). GA 3 sprays are actually a common horticultural practice in some production areas to control colour changes or to delay rind senescence in citrus fruit, or to increase fruit firmness in sweet cherry. Since gibberellins are actively synthesised in seeds, they are also used on seedless grapes to increase berry size.
Studies on the effects on preharvest GA 3 sprays on postharvest fruit quality has shown generally delayed ripening in treated fruit, both climacteric and non-climacteric. Commonly observed effects include retention of higher firmness levels and delayed colour changes (Table 2). Additionally, treat-ment-related changes in cuticular wax morphology have been found for cactus pear [47], and higher ascorbic acid content was also observed for mandarin [48] or mango [49]. Little research has been published on the biochemical mechanisms underlying improved firmness retention, but decreased polygalacturonase and cellulase activities were demonstrated for GA 3 -treated sweet cherry fruit, with some cultivar-dependent variability [50].

Polyamines
Because polyamine (PA) and ethylene biosynthesis pathways share the precursor SAM, these cationic aliphatic amines have been targeted as potential antagonists of ethylene production. All three major polyamines found in plants (putrescine, spermidine and spermine) have an impact on fruit ripening-related events, and reported research has shown delaying effects of preharvest PA applications on the onset of ethylene production, with a concomitant extension of shelf life in a number of fruit species (Table 2). Ripening-delaying effects of preharvest PA sprays, though, are dependent to some extent on the specific PA applied. For example, spermine increased ascorbic acid content in mango, whereas spermidine and putrescine decreased it. In spite of this, general effects on fruit quality were similar, with enhanced firmness and delayed colour changes [51]. Such dependence on the specific polyamine compound used for the application has Compound Fruit Effect on postharvest quality Reference
In contrast to experiments with calcium, which usually comprise several applications throughout on-tree fruit development, reported preharvest PA treatments have been implemented as a single application at a given time point prior to harvest. These treatments have been shown to delay ethylene production arising from lessened SAM decarboxylase (SAMDC), 1-aminocyclopropane-1-carboxylate oxidase (ACO) or 1-aminocyclopropane-1-carboxylate synthase (ACS) expression or activity [52][53][54], leading to deferred ripening-related changes while retaining acceptable quality of produce during subsequent storage and shelf life.

Other growth regulators
Compounds such as jasmonates or salicylic acid play key roles in plant responses to environmental stresses, being in-volved in signal transduction in some biochemical pathways which lead to the biosynthesis of defence compounds such as phenolics and alkaloids. Accordingly, preharvest sprays with these substances have been shown to induce disease resistance or to enhance stress responses in fruit. For example, preharvest jasmonate sprays with methyl jasmonate (MJ) or propyl dihydrojasmonate (PDJ) were reported to induce many transcriptional changes in peach fruit [55]. Unlike most published studies on the effects of preharvest growth regulator sprays on postharvest quality of fruit, in this work the application was undertaken at three different developmental stages. Observed results included a complex set of transcriptional changes apparently resulting from an overlap between ripening and stress responses, with inhibited ethylene production and up-regulated defence-related pathways. In relation with postharvest quality, treatments resulted in increased fruit firmness associated with down-regulation of PG and Exp3 gene expression. Ripening-related colour changes and increase in SSC were also delayed.
For sweet cherry, a single preharvest spray with either 0.2 mM MJ or 2 mM salicylic acid, undertaken 3 days before harvest, was significantly more effective than a postharvest treatment with the same substances in enhancing resistance to infection by Monilinia fructicola, with reduced lesion diameters in comparison with the postharvest applications [56]. These preharvest sprays increased defence-related enzyme activities such as β-1,3-glucanase, phenylalanine ammonialyase and peroxidase, showing a good potential as a strategy for the control of postharvest decay.

Miscellaneous sprays
Preharvest sprays with other substances have also been occasionally reported to affect postharvest quality of some fruit species. Urea and KNO 3 applications were found to increase juice content, soluble solids and SSC/TA ratios of mandarin fruit after storage for 30 days [48]. Interestingly, ascorbic acid content was also increased in treated fruit, showing that not only eating, but also nutritional quality could benefit from this practice. For grape berries, a treatment comprised of three preharvest sprays with N-(2-chloro-4-pyridyl)-Nphenylurea (CPPU) led to higher firmness and juice content, and to decreased weight loss and percentage of unmarketable fruit after being kept one week at ambient temperature [57].
Although very scarcely, the influence of preharvest sprays with mineral elements other than calcium has also been explored. Boron sprays strongly suppressed browning disorders in 'Conference' pears throughout 4 months of cold storage under controlled atmosphere, apparently arising from reduced membrane leakage and increased content of ascorbic acid, the antioxidant properties of which may protect fruit tissues from these disorders [58]. In mandarin, preharvest sprays with potassium led to higher rind firmness and juice acidity, while other eating quality attributes such as juice content, SSC or SSC/TA ratios were unaffected [59], which disagrees with research by El Otmani et al. [48], maybe due to the different 10 potassium concentration used. However, peroxidase activity was lower in treated fruit during storage at 4 o C, suggesting that treatment induced some tolerance to chilling injury. These reports indicate some potential for the prevention of physiological storage disorders, which is worthy of further research.
Finally, preharvest sprays with edible coatings such as chitosan, oligochitosan or sucrose have also been studied [60][61][62][63]. Besides beneficial effects on the eating quality of fruit, these treatments have been generally shown to increase fruit resistance to decay through the modification of enzyme activities related to the antioxidant status of the fruit, such as superoxide dismutase, poliphenol oxidase, peroxidase or phenylalanine ammonia-lyase.

Conclusion
Review of the preharvest spray literature indicates good potential for some treatments to modulate postharvest quality and marketing possibilities of fruit produce, together with the need for further research. Because of the variability across species and cultivars, treatment conditions should be studied and optimised specifically for each particular case. In addition, treatment effects need to be evaluated as a whole, paying particular attention to quality traits so far disregarded, but yet largely relevant for the eating quality of produce, such as aroma.