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Scientia Horticulturae 222 (2017) 193–202

Contents lists available at ScienceDirect

Scientia Horticulturae
journal homepage: www.elsevier.com/locate/scihorti

Effects of preharvest applications of natural antimicrobial products on MARK


tomato fruit decay and quality during long-term storage
Carmela Anna Miglioria, Luca Salvatib, Luigi Francesco Di Cesarec, Roberto Lo Scalzoc,

Mario Parisid,
a
Consiglio per la ricerca in agricoltura e l'analisi dell'economia agraria, Centro di ricerca Ingegneria e Trasformazioni agroalimentari (CREA-IT), Sede di Torino, Strada
delle cacce 73, 10135, Torino, Italy
b
Consiglio per la ricerca in agricoltura e l'analisi dell'economia agraria, Centro di ricerca Agricoltura e Ambiente (CREA-AA), Sede di Roma, Via della Navicella 2-4,
00184, Roma, Italy
c
Consiglio per la ricerca in agricoltura e l'analisi dell'economia agraria, Centro di ricerca Ingegneria e Trasformazioni agroalimentari (CREA-IT), Sede di Milano, Via
Venezian 26, 20133, Milano, Italy
d
Consiglio per la ricerca in agricoltura e l'analisi dell'economia agraria, Centro di ricerca Orticoltura e Florovivaismo (CREA-OF), Sede di Pontecagnano, Via Cavalleggeri
25, 84098, Pontecagnano, Italy

A R T I C L E I N F O A B S T R A C T

Keywords: The effects of preharvest sprays of Thyme essential oil, Propolis and Chitosan on postharvest quality and decay of
Solanum lycopersicum L a “long-storage” tomato, called “Vesuviano”, stored at room temperature for 120 days, were investigated.
GRAS substances Postharvest fruit quality [number of swollen-healthy fruits (SF), withered-healthy fruits (WF) and rotten fruits
Fruit senescence (RF)], organoleptic-related indexes (dry matter, soluble sugars, organic acids, volatiles) and health-related
Phytochemicals
compounds (total carotenoids and phenols) were investigated at 40, 80 and 120 days post-harvesting (T40, T80,
Total solids content
T120). Propolis and Chitosan were able to reduce rotten fruits starting from T80, while the effect of Thyme was
Aromatic compounds
evident as early as T40. Furthermore Chitosan delayed fruit senescence (as expressed by SF/WF ratio) during the
long-storage period. All treatments did not affect the overall postharvest quality, nevertheless some compounds
(such as total soluble sugar for Chitosan and Thyme; total carotenoids and flavonols for Chitosan; total organic
acids, 2-(E)-hexenal, 2-isobutylthiazole and terpenes for Propolis), were better retained than the control during
postharvesting period. Among the three natural fungicides, Chitosan was most effective in reducing fruit
senescence, maintaining a good quality of the fruits over a long-term.

1. Introduction quality (Baldwin et al., 2000). The most important health-related


compounds in tomato fruit are carotenoids with lycopene being the
Tomato (Solanum lycopersicum L.) is one of the most produced and most representative (Shi and Maguer, 1999). Epidemiological studies
extensively consumed vegetable crops in the world (Sacco et al., 2015). have highlighted a strong association between consumption of carote-
Fresh fruits and tomato-based foods provide basic organoleptic fea- noids-rich foods and prevention of different cancers such as cervical,
tures, as well as a wide variety of nutrients and health-related breast, colon, prostate, rectal and stomach (Dorais et al., 2008).
phytochemicals. Sugars and organic acids include the majority of the Phenolic compounds, such as naringenin chalcone, quercetin-type
total dry matter content of tomato fruit. The most abundant sugars are flavonols and hydroxycinnamic acids, are another important class of
fructose and glucose, while major organic acids are citric and malic, phytochemical compounds of tomato fruit (Giovannelli et al., 1999).
with predominance of the first acid (Davies and Hobson, 1981). Earlier Their potential benefit for human health, likely due to the antioxidant
studies have shown that the level of sugars and acids affect not only activity, is widely acknowledged (Siracusa et al., 2012).
tomato taste attributes, but also the overall flavour (Hobson and Despite benefits that can be derived from the crop, postharvest
Bedford, 1989). A relevant contribution to tomato flavour is also given losses make its production in most parts of the world unprofitable.
by volatiles: hexanal, 2(E)-hexenal and 2-isobutylthiazole are consid- Postharvest losses in tomatoes can be as high as 25–42% globally and
ered as characteristic tomato compounds (Dirinck et al., 1976). More- are due to physiological disorders, physical injury and fungal infections
over, terpenes have been demonstrated to contribute to product’s (Arah et al., 2015). Fruits are susceptible to attacks of several


Corresponding author.
E-mail address: mario.parisi@crea.gov.it (M. Parisi).

http://dx.doi.org/10.1016/j.scienta.2017.04.030
Received 30 January 2017; Received in revised form 21 April 2017; Accepted 25 April 2017
0304-4238/ © 2017 Elsevier B.V. All rights reserved.
C.A. Migliori et al. Scientia Horticulturae 222 (2017) 193–202

pathogenic fungi [such as Alternaria alternata (Fr.:Fr.) Keissl., Botrytis fermentation-senescence markers (ethanol).
cinerea (Pers), etc.] making them unsuitable for consumption by
producing mycotoxin, in addition to causing rots (Ibrahim and Al- 2. Material and methods
Ebady, 2014). Although public opinion strongly demands for a reduc-
tion of synthetic agents, fungicides are still the primary means of 2.1. Field trial, treatments and storage conditions
controlling postharvest diseases. Negative effects of chemical fungicides
call for alternative products that reduce losses due to postharvest decay. The study was performed adopting a previously described accession
Among naturally derived substances, i.e. GRAS (Generally Recognized of “Vesuviano” landrace (PV-ISCI 10) (Ruggieri et al., 2014). Tomato
as Safe), essential oils, propolis and chitosan were recently considered plants were grown in open field, according to the traditional practices,
with the aim to controlling biological spoilage and extending the on the slopes of Vesuvio volcano (Massa di Somma, 40° 50′ 0″ N, 14° 22′
storage life of different fresh commodities (Tripathi and Dubey, 2004). 0″ E, 175 m a.s.l.). Transplant was performed in single rows with a
Plant-derived essential oils are considered non-phytotoxic com- density of 5.0 plants m2. A single drip irrigation was applied in support
pounds and are potentially effective as natural pesticides for crop of rainfall along the growing season (115 mm). This experiment
protection (Pane et al., 2013). They contain complex mixtures of included three preharvest treatments: Chitosan (Chitoplant®, Agritalia
secondary metabolites, which are biologically active, endowed with − Villa Saviola, Italy), Propolis (Propoli®, Serbios − Badia Polesine,
antimicrobial, allelopatic, antioxidant and bioregulatory properties Italy) and Thyme (Thymus vulgaris essential oil, A.C.E.F. − Fiorenzuola
(Antunes and Cavaco, 2010). d’Arda, Italy). These substances were diluted by water to give final
Propolis is a natural resinous substance obtained from leaf buds and concentrations of 0.3%, 0.2% and 1% (water emulsion) respectively,
bark of poplar and conifer trees. It has antibiotic, antibacterial and and were compared to untreated control (water). The commercial
antifungal activity (Tripathi and Dubey, 2004). Application of propolis product Chitoplant® was an aqueous solution containing 95% of
has shown positive effects on reducing decay and extending the shelf chitosan (47–65 kDa), 2.5% of boron and 2.5% of zinc; Propoli® was
life of different vegetables and fruits (Yang et al., 2016). a propolis glycolic extract with a flavonoid content (expressed as
Chitosan is a cationic polysaccharide produced by alkaline N- galangin) of 20 mg ml−1, while Thyme was Thymus vulgaris essential
deacetylation of chitin and it has been reported to have plant protective oil containing 57.4% of thymol and 2.8% of carvacrol. The above-
and antifungal properties on different crops (Pichyangkura and mentioned natural antimicrobial substances were spread on the plants
Chadchawan, 2015). (at 5, 15, 25 and 35 days before the harvest) until all fruits were wet to
The efficacy of postharvest applications of different natural anti- runoff.
microbial substances on reducing postharvest losses have been docu- Field trials were carried out in a completely randomized experi-
mented in earlier studies (Aminifard and Mohammadi, 2012; Reddy mental design with three replicates and 30 plants for each replicate. At
et al., 2000; Ordóñez et al., 2011). Conversely, studies regarding the harvest (July 31, 2013), successively mentioned as T0, ripe bunched
preharvest use are reported for few fresh commodities excluding tomato fruits were harvested taking care not to detach them from peduncles.
crop (Romanazzi et al., 2002, 2012, 2013; Saavedra et al., 2016; For each experimental plot, 8 kg of fruits (320 in number, approxi-
Tezotto-Uliana et al., 2014; Meng et al., 2008). An improved investiga- mately) were selected based on uniformity of size and colour and
tion on preharvest uses depend on varieties or landraces belonging to absence of physical injury or lesions caused by pathogen. These
each fruit or vegetable species, since they differ in term of shelf lives quantities were subdivided in two wooden boxes, each containing
extension and storage condition (temperature, relative humidity) can 4 kg approximately of fresh product, used for quali-quantitative market
be very diversified. Therefore the effectiveness of GRAS substances assessment and chemical-physical analysis. The overall samples (with-
adopted to control postharvest decay and quality can be severely out any packaging) were then transferred in an unconditioned and well-
affected by varying background conditions. ventilated shed up to 120 days after harvest, where temperature and
Several cherry-like tomato landraces showing high drought toler- relative humidity were recorded, every 60 min, by a wireless data
ance are traditionally cultivated in Mediterranean countries, and logger (EL-USB-2 model, Lascar Electronics Ltd. − United Kingdom)
especially in southern Italy, under non-irrigated conditions. Peculiar (Supplementary Fig. 1).
textural properties, such as high firmness and thickness of skin, allow
an extended shelf life of the fruits, which after the harvest are stored in 2.2. Postharvest decay assessment
the typical hung-shaped appearance for 3–4 months under no-condi-
tioned atmosphere (“long-storage tomatoes”) (Siracusa et al., 2012; In order to evaluate the decay rate, both treated and untreated fruits
Mercati et al., 2015). The most important and profitable “long-storage” of each experimental plot were visually observed at 40 (September 10,
landrace is cultivated on the slopes of the Vesuvio volcano (Campania 2013), 80 (October 20, 2013) and 120 (November 30, 2013) days post
region) and is called “Vesuviano” or “Pomodorino del piennolo del harvest (thereafter mentioned as T40, T80 and T120). At these time
Vesuvio” (labeled as PDO − Protected Designation of Origin; Reg. points fruits were classified in three categories: swollen-healthy fruits
1238, 2009; Ercolano et al., 2008, 2014). This high-value product is (SF) (ripe fruits without abiotic/biotic damages and showing high
commercialized throughout the Italian peninsula and in other European firmness), withered-healthy fruits (WF) (wrinkled/senescent fruits
countries starting from winter to early spring, when high-quality fresh without abiotic/biotic damages and showing low firmness) and rotten
tomatoes are not available on the markets. However, despite the fruits (RF) caused by physiological or biological alterations
prolonged shelf life, unprofitable fruit rots and quality decay are (Supplementary Fig. 2). Number of fruits falling in these classes were
currently observed during the typical storage of “Vesuviano” landrace. recorded at each time point and the relative incidence (%) was
Therefore the adoption of means to limit these losses are highly expressed as cumulative number of each category on number of fruits
desirable. at harvest (T0). RF were discarded each time as waste product, while
Based on these premises, the objectives of this study were (i) to WF and SF represented the commercial yield.
examine the effects of preharvest spraying of propolis, chitosan and A preliminary report (Carrieri et al., 2016) provided technical
thyme essential oil for controlling natural decay and (ii) to evaluate details and specific analysis on post harvest decay microorganisms.
postharvest quality during the typical prolonged storage at room
temperature by measurement of dry matter, soluble sugars, organic 2.3. Analytical methods
acids, by the main health-related compounds (phenols and total
carotenoids), and evaluating volatile substances both for tasting 2.3.1. Tomato samples
(hexanal, 2(E)-hexenal, 2-isobutylthiazole and total terpenes) and for For each treatment, healthy tomato fruits were sampled in duplicate

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C.A. Migliori et al. Scientia Horticulturae 222 (2017) 193–202

(around 600 g/sample) and within 12 h from harvesting, they were significant peaks. The clean coloured organic layer was filtered
quickly frozen in a forced-air blast tunnel (Thermo-Lab, Lodi, Italy) at (0.45 μm regenerated cellulose filter) and analyzed by HPLC (Jasco-
−50 °C. Subsequently, sampled fruits were divided in two aliquots, one Italy, Lecco, Italy models PU980, diode array detector mod MD-2010,
stored at −80 °C and the other subjected to lyophilisation. Dry matter the used column was a C30 YMC Carotenoid 0,8 ml/min, 35 °C, YMC
content was calculated by the ratio lyophilized product weight vs fresh Europe, Dinslaken, Germany) using the methodology described by
one. The frozen material, before analysis, was homogenized in a blade Ishida et al. (2001). The concentration of total carotenoids, obtained
mill blender, sieve of 1 mm and kept at −20 °C. by the sum of β-carotene and all-trans lycopene, the major carotenoids
in fresh tomato fruits, was calculated from the experimental peaks area
2.3.2. Soluble sugars and organic acids by analytical interpolation, using standard calibration curves and were
The extraction was carried out on 500 mg of lyophilized sample, expressed as mg/100 g dw.
homogenized with 10 ml deionised water and separated by centrifuga-
tion. The supernatant was filtered (0.45 μm nylon filter), diluted and 2.3.5. Extraction-concentration and analysis of volatile substances
analyzed via high-performance liquid chromatography (Jasco-Italy, The volatile fraction from tomatoes samples was extracted and
Lecco, Italy models PU980, RI 930 refractive index detector, the used concentrated by a combined microwave-resin-solvent and the extracts
column was a Biorad Aminex HPX-87C, 0,7 ml/min, 85 °C, Bio-Rad obtained were analyzed by GC/MS (Agilent Mod. 6890N and MS
Laboratories, Milano, Italy) under the conditions described by Forni 5973N, Agilent Technologies Italia, Milano, Italy, the used column
et al. (1992). As for organic acid analysis, the extraction was performed was a capillary DB-1, 60 m, 0.25 mm 0.25 μm film thickness, J & W
as for soluble sugars and the HPLC analysis (Jasco-Italy, models PU980, Scientific, Folsom, California, USA), following an already validated
UV1570 detector setted at 214 nm, the used column was a GL Sciences protocol (Migliori et al., 2012). The volatile components were identi-
Inertsil ODS-3 C18, 0,6 ml/min, 30 °C, Microcolumn, Monza, Italy) was fied using the retention times of chromatographic peaks, by comparing
performed as described by Lo Scalzo et al. (2012). their mass spectra with those in a commercial library (Wiley 7 n. 1
Library, Mass Spectral Data Base, Hewlett-Packard, Vienna, Austria)
2.3.3. Phenols and by using commercial standards when available. For quantitative
Analysis of phenol content was performed according to Van der Rest analysis, standard solutions of the components at known concentrations
et al. (2006), with some modifications. For the extraction, 300 mg of were used to calculate the response factors. For the components whose
lyophilized powder was dissolved in 10 ml of a 1:1 v/v mixture of EtOH standards were not commercially available, the internal standard
and 0.12 N HCl. The mixture was vortexed for 1 min and subjected to procedure was followed, by using solutions of methyl palmitate. The
shaking for 2 h. The mixture was centrifuged at 25000g for 5 min at hexanal, 2(E)-hexenal and 2-isobutylthiazole were considered as char-
4 °C, and an aliquot (400 μl) was diluted with an equal volume of 80% acteristic tomato volatile compounds, according to Dirinck et al.
EtOH plus 50 μl of 50% CH3COOH prior to HPLC injection. The HPLC (1976). The level of total terpenes, given by the sum of 2,3-epoxiger-
analysis was performed using a JASCO system equipped with a diode anial, neral, geranial, β-damascenone, nerylacetone, β(Z)-ionone, (E, E)-
array detector (MD-910 JASCO). The pump (PU-980 JASCO) was pseudoionone have been reported and discussed.
coupled to a ternary gradient unit (LG-1580-02 JASCO). The data were
evaluated using a software-management system for chromatographic 2.3.6. Ethanol
data (ChromNAV, Jasco). The separation was performed by reversed- The ethanol measurement was performed by a static headspace GC-
phase chromatography using an ODS-3 Lichrosorb 250 × 4 mm col- FID (DANI Instruments, Milano, Italy, GC Mod. 6400 equipped with an
umn. The flow rate was 0.7 ml/min, the injection volume was 30 μl, HSS 86.50 headspace apparatus, the used column was a wide-bore DB-
and the oven temperature was 45 °C. The mobile phase consisted of 5% WAX, 30 m, 0.32 mm 0.25 μm film thickness, J & W Scientific, Folsom,
CH3COOH in water (solvent A) and methanol/water/CH3COOH California, USA. The carrier gas was helium pure for gas-chromato-
(90:5:5) (solvent B). The gradients were as follows: (A/B): 95/5 for graphy 5.0) method, already validated on other plant materials (Lo
0–3 min, from 95/5 to 70/30 in 5 min, 70/30 for 20 min, from 70/30 to Scalzo et al., 2007). In this case, the used material was represented by
35/65 in 7 min, 35/65 for 5 min, from 35/65 to 95/5 in 10 min, and homogenized frozen tomatoes, added with 10% NaCl w/w and condi-
95/5 for 12 min. The total analysis time was 59 min. The peaks were tioned at 80 °C for 20 min in gas-tight 20 ml vials. The dosage was made
identified by direct comparison with commercial standards and by applying an external standard methodology.
inspection of spectral and chromatographic properties with respect to
relevant data reported in the literature. The main compound observed 2.3.7. Statistical analysis
were chlorogenic acid, rutin, two rutin derivatives, tentatively identi- A quantitative analysis based on Chi-square contingency tests was
fied as rutin-O-hexoside and rutin-O-pentoside, from chromatographic used to evaluate the effect of four treatments (Control, Thyme, Propolis
data of previous works (Moco et al., 2006; Van der Rest et al., 2006; and Chitosan) on the number of collected fruits falling into three (RF,
Vallverdú-Queralt et al. 2010), and chalconaringenin, whose presence SF and WF) or two [RF, (SF + WF)] commercial categories over time
in tomato is well known. Quantification was based on the calibration (T40-T80-T120).
curves of external standards. The levels of total flavonols (mg/100 g The effectiveness of natural substances on fruit senescence delaying
dw), resulting from the sum of contents of rutin and the two rutin was investigated considering changes in the SF/WF ratio over different
derivatives, were reported. time points. This variable was analyzed using a Kruskal-Wallis non-
parametric analysis of variance with significance tested at p ≤ 0.05,
2.3.4. Total carotenoids adopting two designs: (a) four comparison groups (1 = control,
These lipophilic compounds were extracted under controlled con- 2 = Thyme, 3 = Propolis, 4 = Chitosan) or (b) three comparison
ditions (in darkness at 0–1 °C to avoid sample decomposition) from 10 g groups [1 = control, 2 = (Thyme + Propolis), 3 = Chitosan]. In both
frozen homogenized sample, using 100 ml n-hexane/acetone/ethanol cases, the post-hoc Median Test was applied testing for significance at
(2:1:1 v/v/v) solution, added with 1 mg/ml butylated hidroxytoluene p ≤ 0.05 level.
(BHT), as described by Shi et al. (1999). The mixture was separated by All chemical determinations were performed in triplicate for each
centrifugation at 5000 rpm for 10 min at low temperature. As the replicate thesis. Data were subjected to the analysis of variance
method followed made only one extraction, it was validated by (ANOVA) and the comparison of the average values was carried out
laboratory trials with subsequent extractions, checking the colour of using the Tukey test. Statistical differences at P ≤ 0.05 were considered
the pellet, which remained uncoloured after the first solvent treatment. significant; standard deviation were also reported. Statistical analysis
Chromatograms of the second and subsequent extracts gave no was performed using Statistica version 6.0 (Tulsa, Oklahoma, US).

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C.A. Migliori et al. Scientia Horticulturae 222 (2017) 193–202

Table 1
Chi-square contingency table analysis showing comparison between three treatments and
untreated control for three and two fruit commercial categories under three time-points.

(1) (2)
Comparision T RF vs WF vs SF RF vs (WF + SF)

χ2 p d.f. χ2 p d.f.

Thyme vs control 40 20.25 0.0270 10 18.74 0.0021 5


80 35.04 0.0001 10 24.61 0.00016 5
120 25.53 0.0044 10 23.49 0.00027 5

Chitosan vs control 40 8.90 0.5354 10 6.89 0.2286 5


80 20.63 0.02383 10 10.05 0.0738 5
120 30.80 0.0006 10 18.17 0.0027 5

Propolis vs control 40 11.14 0.3465 10 9.92 0.0774 5


80 19.25 0.0371 10 12.01 0.0347 5
120 31.78 0.0004 10 16.10 0.0066 5

Legend: (1) T = days post-harvest; (2)


RF = rotten fruits; WF = withered-healthy fruits;
SF = swollen-healthy fruits.

percentages of SF were detected likewise for Thyme (39.6%), Propolis


(40.9%) and Chitosan (44.9%) in respect to the control (34.7%)
(Fig. 1C). Results of the statistical analysis highlighted a great influence
at T120 (χ2 = 74.1; d.f. = 22; p < 0.0001) and at T80 (χ2 = 51.1;
d.f. = 22; p = 0.0004). Grouping all commercial fruits (WF + SF) and
comparing them against rotten fruits, a valuable effect of the treatments
was observed over the three time points [T40 (χ2 = 21.5; d.f. = 11;
p = 0.028), T80 (χ2 = 29.8; d.f. = 11; p = 0.002), T120 (χ2 = 43.6;
d.f. = 11; p < 0.0001)].
In order to detect the most effective treatment capable to reduce
rotting fruits, a Chi-square contingency test was finally run comparing
each thesis against the untreated control, for each of the time points,
using separately three (RF vs WF vs SF) and two variables [RF vs (SF
+ WF)]. By considering three homogeneous variables (RF vs WF vs SF)
by treatments (Thyme, Propolis and Chitosan), the overall impact on
the proportion of the RF, WF and SF fractions was significant against
the control at T80 and T120 and significant at 40 days post-harvesting
for Thyme only (Table 1). Pooling together WF and SF fractions against
RF, all treatments impacted significantly the proportions of commercial
categories at T120. Propolis was effective also at T80 and Thyme was
effective at both 40 and 80 days post-harvesting.
Fig. 2 shows the time-course of the SF/WF ratio by treatment. For all
treatments, fruit senescence negatively impacted the level of the SF/WF
ratio over time. However, high values of the SF/WF ratio were observed
for Chitosan in respect to the control at each point in time (2.60 vs 1.99
at T40; 2.17 vs 1.36 at T80; 1.59 vs 0.99 at T120, respectively).
The Kruskal-Wallis test run on four single treatments (untreated

Fig. 1. Postharvest time-course of three fruit commercial categories (RF, WF and


SF).RF = rotten, WF = withered-healthy and SF = swollen-healthy at three time-points
(A = 40; B = 80; C = 120 days post-harvesting) as affected by preharvest treatments
with Thyme, Propolis, Chitosan and water (control). Data correspond to the mean ± SE
of three independent replicates.

3. Results

3.1. Postharvest decay

The effect of three preharvest treatments on fruits number falling


into three commercial categories (RF, WF and SF) is shown in
Fig. 1(A–C). Despite a relevant difference in the incidence of both
rotten and healthy fruits (Fig. 1A), a Chi-square contingency test
showed no significant difference among treatments at T40
(χ2 = 24.75; d.f. = 22; p = 0.309). At T80 the tomato samples treated
with Thyme, Propolis and Chitosan showed higher incidences of SF
Fig. 2. Postharvest time-course of SF/WF ratio as affected by preharvest treatments with
than the untreated control (53.0%, 55.1% and 52.6% vs 43.6%,
Thyme, Propolis, Chitosan and water (control).WF = withered-healthy fruits;
respectively) (Fig. 1B), as well as a lower percentages of rotten fruits SF = swollen-healthy fruits. Data correspond to the mean ± SE of three independent
(16.3%, 18.8% and 17.2% vs 24.5%, respectively). At T120, high replicates.

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Table 2
Postharvest changes in dry matter, soluble sugars and organic acids content of “Vesuviano” tomatoes treated with Thyme, Propolis, Chitosan and water (control) during preharvest.

Treatment Time- Dry matter SOLUBLE SUGARS (g/100 g d.w.) ORGANIC ACIDS (g/100 g d.w.)
point (1) (%)
Glucose Fructose Total change % Fructose/ Malic Acetic Citric Total
(2)
amount glucose amount

Control T0 10.8 ± 0.1 aA 17.8 ± 1.7 bA 19.7 ± 0.2 aA 37.5 bA - 1.1 aB 1.3 ± 0.4 cD 0.1 ± 0.0 cD 5.3 ± 0.0 bA 6.7 bA
T40 9.4 ± 0.4 aA 18.7 ± 0.8 bA 18.5 ± 0.5 abA 37.2 bA −0.8 1.0 aB 1.6 ± 0.2 aC 0.4 ± 0.1 bC 4.3 ± 0.3 aB 6.4 aB
T80 8.9 ± 0.1 bA 14.2 ± 1.8 bB 21.6 ± 2.1 aA 35.8 bA −4.5 1.5 aA 2.2 ± 0.3 aB 0.7 ± 0.2 cB 4.1 ± 0.4 aB 7.2 aA
T120 9.8 ± 0.3 aA 12.6 ± 3.8 aC 21.2 ± 4.9 aA 33.8 aB −9.9 1.7 bA 2.5 ± 0.2 bA 1.1 ± 0.2 bA 2.9 ± 0.5 cC 6.4 bB

Thyme T0 10.6 ± 0.5 aA 21.6 ± 0.7 aA 22.4 ± 0.1 aA 44 aA - 1.0 aB 1.9 ± 0.4 aC 0.1 ± 0.0 bD 5.9 ± 1.7 aA 7.9 aA
T40 9.7 ± 0.5 aA 22.1 ± 0.5 aA 20.4 ± 0.5 aA 42.5 aA −3.4 0.9 aB 1.3 ± 0.5 bD 0.5 ± 0.0 aC 4.7 ± 0.05 aB 6.5 aB
T80 10.2 ± 0.3 aA 17.1 ± 1.9 aB 21.7 ± 1.1 aA 38.9 aB −11.6 1.3 bA 2.1 ± 0.5 aB 0.9 ± 0.1 bB 3.2 ± 0.7 bC 6.2 bB
T120 8.7 ± 0.4 bB 13.1 ± 1.3 aC 21.4 ± 4.7aA 34.6 aC −21.4 1.6 bA 2.4 ± 0.5 bA 1.1 ± 0.2 bA 3.2 ± 0.3 bC 6.7 bB

Propolis T0 9.8 ± 0.3 aA 22.6 ± 5.1 aA 24.5 ± 3.3 aA 47.1 aA - 1.1 aC 1.2 ± 0.08 cD 0.1 ± 0.0 bD 6.4 ± 0.6 aA 7.8 aA
T40 10.0 ± 1.2 aA 18.2 ± 0.8 bB 17.8 ± 0.2 bC 36.0 bB −23.6 1.0 aC 1.7 ± 0.4 aC 0.4 ± 0.1 bC 4.7 ± 0.6 aB 6.8 aB
T80 9.4 ± 0.8 aA 17.6 ± 2.0 aB 21.3 ± 1.5 aB 38.9 aB −17.4 1.2 bB 2.3 ± 0.09 aB 0.9 ± 0.2 bB 4.0 ± 0.2 aC 7.3 aA
T120 8.2 ± 0.2 bB 11.1 ± 1.7 aC 21.4 ± 1.5 aB 32.5 bC −31.0 1.9 aA 2.9 ± 0.2 aA 1.2 ± 0.1 aA 3.7 ± 0.2 aD 7.9aA

Chitosan T0 10.3 ± 0.0 aA 18.6 ± 1.5 bB 20.6 ± 1.9 aA 39.3 bB - 1.1 aC 1.5 ± 0.5 bB 0.2 ± 0.0 aC 5.9 ± 0.0 aA 7.6 aA
T40 10.1 ± 0.6 aA 21.0 ± 0.2 aA 21.1 ± 1.2 aA 42.1 aA +7.1 1.0 aC 1.7 ± 0.1 aB 0.4 ± 0.0 bB 4.0 ± 0.4 bB 6.2 bB
T80 10.2 ± 0.3 aA 17.6 ± 1.7 aB 21.8 ± 0.7 aA 39.4 aB +0.3 1.2 bB 2.3 ± 0.2 aA 1.0 ± 0.1 aA 3.3 ± 0.4 bC 6.7 bB
T120 9.4 ± 0.0 aA 11.8 ± 1.6 aC 19.4 ± 0.5 aB 31.1 bC −20.9 1.6 bA 2.3 ± 0.2 bA 1.1 ± 0.0 bA 2.9 ± 0.8 cD 6.3 bB

For each treatment, different capital letters indicate significant differences between the storage time points (p ≤ 0.05). For each storage time point, different lower-case letters indicate
differences between the treatments (p ≤ 0.05). Data correspond to the mean ± SD of three independent replicates.
Legend: (1) T0 = harvest; T40-T80-T120 = 40-80–120 days postharvest; (2) percentage change respect to T0.

values for Thyme (44.0 g/100 g dw) and Propolis (47.1 g/100 g dw) at
harvest (T0), instead no significant variation was observed in Chitosan-
treated sample compared with the Control. Considering the time-course
of total sugar contents, pre-harvest treatments with Chitosan resulted in
better retention than the Control and the other natural products both at
T40 and T80 (+7.1 and +0.3%, respectively). The fructose/glucose
ratio increased along the storage period, without strong differences
among treatments.
As concern organic acids, both malic and acetic acids reached the
highest values at T120 for all theses. By contrast, citric acid constantly
decreased for all treatments during the postharvest period (from 5.9 g/
100 g dw to 3.2 g/100 g dw). As shown in Table 1, preharvest
treatments with Thyme, Propolis and Chitosan positively affected total
content of organic acids at harvest in respect to the control (7.9, 7.8, 7.6
and 6.7 g/100 g dw, respectively) due to the higher amount of citric
acid (5.9, 6.4 and 5.9 g/100 for the three treatments against 5.3 g dw of
the control). Moreover the malic acid content in Thyme-treated fruits
Fig. 3. Postharvest changes in total carotenoids concentration of “Vesuviano” tomatoes was increased at T0 respect to the untreated fruits (1.9 and 1.3 g/100 g
treated with Thyme, Propolis, Chitosan and water (control) during dw, respectively). A good retention of total organic acids were noticed
preharvest.T0 = harvest; T40-T80-T120 = 40-80-120 days postharvest. Data correspond up to T120, as a result of pre-harvest treatments with Propolis, in
to the mean ± SD of three independent replicates.
relation to a slower decrease in citric acid contents and stronger
increases of malic acid in respect to the control (Table 2).
control, Thyme, Chitosan and Propolis) showed no significant effects on
the SF/WF ratio for each time point (T40-T80-T120). By adopting a less
stringent assumption that include three homogeneous groups (1 = con- 3.3. Total carotenoids
trol, 2 = Thyme + Propolis, 3 = Chitosan), the Kruskal-Wallis test was
non-significant (T40, p = 0.11; T80, p = 0.09; T120, p = 0.15). By The time-course of total carotenoids (from T0 to T120) showed
contrast, the median test applied on the same dataset was significant to significant increases up to T120 in Control, Thyme- and Propolis-
the alternative hypothesis that at least one group median was different treated fruits (Fig. 3).
from the population median of at least one other group at both T80 For each treatment, different capital letters indicate significant
(χ2 = 6; d.f. = 2; p = 0.049) and T120 (χ2 = 6; d.f. = 2; p = 0.049). differences between the storage time points (p ≤ 0.05). For each
storage time point, different lower-case letters indicate differences
between the treatments (p ≤ 0.05). Data correspond to the
3.2. Dry matter, soluble sugars and organic acids mean ± SD of three independent replicates.
Legend: T0 = harvest; T40-T80-T120 = 40–80–120 days post-har-
The dry matter content of tomato fruits was around 10% with vest
negligible changes among the different treatments at T0 (Table 2). A different pattern was found adopting preharvest treatments with
Anyway, good retains were found up to T120 in Chitosan-treated fruits Chitosan on “Vesuviano” tomatoes. This biopolymer induced a weak
(10.3% at T0 and 9.4% at T120) as well as happened in the Control decrease between T0 and T40, with no significant variations being
(10.8% and 9.8% at T0 and T120, respectively). The total soluble sugars detected for T80 and T120 (175.5 and 179.9 mg/100 g dw).
content, given by the sum of glucose and fructose, reached the highest Considering the differences among the treatments and the control

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Table 3
Postharvest changes in single polyphenols content of “Vesuviano” tomatoes treated with Thyme, Propolis, Chitosan and water (control) during preharvest. All values are expressed in mg/
100 g dw.

(1)
Treatment Time-point Chlorogenic acid Chalconaringenin FLAVONOLS

Rutin-O-hexoside Rutin-O-pentoside Rutin Total Flavonols

Control T0 10.0 ± 1.5 aA 4.7 ± 2.4 aA 9.1 ± 3.2 aC 16.0 ± 2.5 aB 45.4 ± 4.8 aC 70.5 ± 9.2 aB
T40 2.7 ± 0.4 aB 3.1 ± 1.1 bB 11.4 ± 3.1 aB 20.6 ± 5.2 aA 72.1 ± 17.9 aA 104.0 ± 26.2 aA
T80 0.8 ± 0.2 bD 0.2 ± 0.1 bD 7.7 ± 1.6 cC 8.1 ± 1.8 cC 23.9 ± 6.7 cD 39.7 ± 10.0 cC
T120 2.0 ± 0.5 aC 0.3 ± 0.0 aC 20.7 ± 5.9 aA 16.3 ± 5.3 aB 61.5 ± 15.9 aB 98.5 ± 26.9 aA

Thyme T0 9.3 ± 2.5 aA 1.4 ± 0.3 cB 7.0 ± 2.4 cC 10.3 ± 2.2 cC 27.4 ± 3.9 cC 44.7 ± 8.1 cC
T40 1.8 ± 0.1 bB 1.8 ± 0.28 cA 5.1 ± 0.5 dD 13.8 ± 3.0 bA 37.0 ± 5.5 dB 56.0 ± 8.9 dB
T80 0.9 ± 0.2 bC 0.5 ± 0.3 aC 9.0 ± 0.3 bB 12.1 ± 1.1 bB 43.7 ± 7.2 bA 64.8 ± 8.6 bA
T120 1.5 ± 0.4 bB 0.2 ± 0.08 aD 16.2 ± 1.5 bA 7.9 ± 1.1 dD 40.7 ± 2.2 cA 64.8 ± 4.2 dA

Propolis T0 8.0 ± 0.9 bA 4.3 ± 1.6 aB 8.5 ± 3.5 bD 13.0 ± 3.5 bB 38.9 ± 8.2 bB 60.4 ± 15.1 bB
T40 2.6 ± 0.4 aB 5.8 ± 2.8 aA 9.4 ± 1.6 bC 16.0 ± 2.9 bA 57.8 ± 9.2 bA 83.2 ± 13.4 bA
T80 0.8 ± 0.2 bD 0.3 ± 0.1 bC 10.5 ± 3.9 aB 13.4 ± 2.7 aB 44.0 ± 8.8 bB 67.9 ± 15.2 bB
T120 1.4 ± 0.4 bC 0.3 ± 0.1 aC 16.2 ± 2.1 bA 8.8 ± 1.5 cC 40.4 ± 4.4 cB 65.5 ± 8.0 cB

Chitosan T0 7.5 ± 0.9 bA 2.0 ± 0.7 bA 7.7 ± 2.5 cC 15.4 ± 2.8 aA 43.6 ± 6.4 aB 66.7 ± 10.2 aB
T40 1.9 ± 0.21 bB 1.4 ± 0.2 cB 7.0 ± 1.1 cC 15.1 ± 1.8 bA 51.4 ± 5.4 cA 73.6 ± 7.7 cA
T80 1.1 ± 0.0 aD 0.6 ± 0.3 aC 10.9 ± 2.7 aB 14.1 ± 2.2 aA 51.0 ± 7.9 aA 76.0 ± 11.9 aA
T120 1.7 ± 0.4 bC 0.3 ± 0.1 aC 19.5 ± 4.9 aA 11.6 ± 2.8 bB 51.7 ± 15.2 bA 82.8 ± 22.9 bA

For each treatment, different capital letters indicate significant differences between the storage time points (p ≤ 0.05). For each storage time point, different lower-case letters indicate
differences between the treatments (p ≤ 0.05). Data correspond to the mean ± SD of three independent replicates.
Legend: (1) T0 = harvest; T40-T80-T120 = 40-80–120 days postharvest.

for each time-point, a significant higher level of total carotenoids at dw) and Propolis (779 vs 483 μg/100 g dw). As for the time course of 2-
harvest (126,5 mg/100 g dw) was detected in Chitosan-treated fruits (E)-hexenal contents, no significant differences were observed for the
than other thesis and the control (80.9 mg/100 g dw for Thyme, control, while higher values were detected at T0 than T120 by adopting
88.7 mg/100 g dw for Propolis and 91.6 mg/100 g dw for the control). three preharvest treatments with GRAS substances. Regarding 2-
However, for the following time-points, no notable differences were isobuthylthiazole (responsible of the “wine-like” aroma of tomato
found between the control and Chitosan treatment. Finally, preharvest fruits), the use of Thyme strongly increased the content of this volatile
treatments with Propolis or Thyme resulted in lower carotenoids compound at T0 which was more than double compared to the
content than the control at T40-T80-T120 time-points. untreated control (937 vs 449 μg/100 g dw). The effects of three
preharvest treatments at T40 on the contents of 2-isobuthylthiazole
3.4. Phenolic compounds were similar to what happened for hexanal; a better retention between
T40 and T80 was observed for Thyme- and Propolis-treated fruits
As shown in Table 3, chlorogenic acid significantly decreased during compared with the control.
the storage at room temperature and the most relevant decreases were As for the terpenes contents, peaks of content were detected at T120
found in all samples from T0 to T40. Furthermore, all preharvest for Thyme (998 mg/100 g dw), at T80 for Propolis (1135 mg/100 g dw)
treatments did not affect significantly the decreasing rate of chlorogenic and at T40 for Chitosan (847 mg/100 g dw). For the latter treatment a
acid up to T120. Regarding the chalconaringenin contents no signifi- lower variability than other treatments were also observed over the
cant differences were found at T0 between untreated control (4.3 mg/ time (Table 4).
100 g dw) and Propolis treatment (4.7 mg/100 g dw). Moreover, In the same Table, ethanol, that can be considered an index of
despite an overall decrease over time in the concentration of this anaerobic metabolism (Park et al., 1994), was also reported. At T0, a
flavonoid, significant increases were observed from T0 and T40 for significant higher content of this alcohol was found in the control than
Thyme and Propolis treatments (from 1.4 to 1.8 mg/100 g dw and from the other treatments (29 mg/100 g dw vs 21, 19 and 15 mg/100 g dw of
4.3 to and 5.8 mg/100 g dw, respectively). The level of total flavonols Thyme, Propolis and Chitosan, respectively). Moreover, except for the
at T0, as sum of rutin and its two derivatives, resulted about the same, control, in all treated samples strong increases were detected moving
around 70 mg/100 g dw, with a significant lower content in Thyme- from T0 to T40, with the highest variation in Chitosan-treated fruits
treated fruits (44.7 mg/100 g dw). Starting to T40, rutin was well (174 mg/100 g dw). Starting to 40 days post-harvesting, no clear
retained in Chitosan-treated fruits; a notable increase in rutin-O-hexo- behaviour was observed for the control, while relevant decreases in
side content was also detected up to T120. Moreover, moving from T0 ethanol contents were observed in all tomatoes treated with GRAS
to T120 a better retention of rutin-O-pentoside was found for the same substances. As a general rule, significant highest values of alcohol
treatment than the control and the other treatments. According to these contents were always found in Chitosan-treated fruits, until to the end
results, a clear increasing in total flavonols content was observed in of the storage.
Chitosan-treated tomatoes up to the end of storage (from 66.7 to
82.8 mg/100 g dw). 4. Discussion

3.5. Volatile substances Alternative strategies aiming to control postharvest decay and to
reduce dependency on synthetic fungicides are currently investigated in
Taking into account the evolution of characteristic volatile com- tomato and other perishables.
pounds during the postharvest period (Table 4), highest values of Several in vitro studies have highlighted the inhibitory activities of
hexanal occurred at T80 in untreated control (1006 μg/100 g dw) or at Propolis on postharvest decay-causing fungi and bacteria of different
T40 in the other treatments. Moreover, the content of this aldehyde was fruits and vegetables (Basim et al., 2006; Tosi et al., 1996). The
higher at T120 than T0 for the untreated control (895 vs 435 μg/100 g effectiveness of this natural substance has been also demonstrated in

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Table 4
Postharvest changes in characteristic volatile compounds, total terpenes and ethanol contents of “Vesuviano” tomatoes treated with Thyme, Propolis, Chitosan and water (control) during
preharvest. All values are expressed in μg/100 g dw, with the exception of ethanol, that is expressed as mg/100 g dw.

(1)
Treatment Time-point Hexanal 2-(E) hexenal 2-isobutylthiazole Terpenes Ethanol

Control T0 435 ± 35 dC 371 ± 6 bA 449 ± 73 bD 713 ± 2 aB 29 ± 1 aA


T40 843 ± 99 bB 325 ± 35 cA 761 ± 49 cC 781 ± 2 cB 16 ± 3 cC
T80 1006 ± 141 aA 356 ± 63 bA 891 ± 114 bB 629 ± 10 cC 24 ± 11 bB
T120 895 ± 38 aB 324 ± 21 aA 982 ± 84 aA 877 ± 15 bA 3 ± 1 cD

Thyme T0 932 ± 128 aB 309 ± 18 cB 937 ± 164 aB 634 ± 31 bB 21 ± 5 bB


T40 1281 ± 49 aA 437 ± 38 aA 1122 ± 116 aA 928 ± 5 bA 98 ± 6 bA
T80 864 ± 56 bB 290 ± 23 cB 879 ± 62 bB 587 ± 8 cB 23 ± 7 bB
T120 864 ± 21 aB 256 ± 13 bC 874 ± 38 bB 998 ± 28 aA 5 ± 3 bC

Propolis T0 482 ± 14 cD 458 ± 16 aA 345 ± 17 cD 594 ± 33 bC 19 ± 14 cC


T40 1227 ± 172 aA 385 ± 48 bB 953 ± 101 bB 1108 ± 17 aA 99 ± 49 bA
T80 983 ± 98 aB 399 ± 33 aB 1106 ± 141 aA 1135 ± 36 aA 26 ± 19 bB
T120 779 ± 132 bC 242 ± 20 bB 655 ± 63 dC 839 ± 54 bB 2 ± 1 dD

Chitosan T0 597 ± 26 bB 426 ± 22 aA 371 ± 17 cC 732 ± 63 aA 15 ± 3 dC


T40 1194 ± 45 aA 417 ± 51 aA 981 ± 41 bA 847 ± 104 bA 174 ± 52 aA
T80 617 ± 69 cB 192 ± 32 dB 754 ± 40 cB 785 ± 50 bA 39 ± 10 aB
T120 592 ± 40 cB 175 ± 11 cB 750 ± 74 cB 657 ± 5 cB 6 ± 1 aD

For each treatment, different capital letters indicate significant differences between the storage time points (p ≤ 0.05). For each storage time point, different lower-case letters indicate
differences between the treatments (p ≤ 0.05). Data correspond to the mean ± SD of three independent replicates.
Legend: (1) T0 = harvest; T40-T80-T120 = 40-80–120 days postharvest.

vivo studies performed on some perishables (such as grapefruit, pear, in the leakage of intracellular electrolytes and proteinaceous constitu-
apple and strawberry), however few studies have investigated the use of ents (Palma-Guerrero et al., 2008). Bhaskara Reddy et al. (2000)
Propolis in controlling postharvest diseases in tomato crop (Bakeer demonstrated that stem scar application of chitosan inhibits develop-
et al., 2016; Yang et al., 2016; Ordónez et al., 2011). Our results ment of blackmold rot [Alternaria alternata (Fr.) Keissl.] of tomatoes
highlight a positive effect on reducing rotten fruits starting to T80; and reduces production of pathogenic factors by the fungus, such as cell
anyhow preliminary results by Carrieri et al. (2016) reported no wall-degrading enzymes (polygalacturonase, pectate lyase and cellu-
efficacy on fungal pathogen at 200 days post-harvesting, hypothesizing lase), organic acids, and host specific toxins responsible for fungal
a moderate effect over time. Bankova et al. (2002) suggested that penetration and host tissue damage.
composition and biological activities of propolis depend on many Besides antifungal activity, Chitosan has the potential for inducing
factors such as the geographical origin as well as the collection time defense-related enzymes (Bautista-Baños et al., 2006) and phenolics in
and plant source. plants (Romanazzi et al., 2017). Recently, Zhang et al. (2014) suggest
Regarding the effect of Thyme reducing the incidence of fruit rots, that one of the molecular mechanisms involved in the enhancing of
this antimicrobial substance showed a relevant efficacy through the resistance to gray mold (Botrytis cinerea Pers.) in cherry tomato fruit
entire storage period of “Vesuviano” tomato fruits. Moreover, a was likely associated with activation of the mitogen-activated protein
satisfactory control on postharvest fungal decay [caused by Alternaria kinase (MAPK) signaling pathway by Chitosan.
alternata (Fr.) Keissl., Pennicillium spp, Aspergillus spp and Fusarium spp] The high values of the SF/WF ratio in Chitosan-treated fruits may
was also found up to 200 days post-harvesting, by Carrieri et al. (2016). indicate a delaying of fruit senescence over time. Several studies
Unlike to in vitro studies, few studies have been performed in vivo reported that Chitosan coating provides a semi-permeable film around
conditions to evaluate the fungicidal properties of Thyme in controlling the fruit surface, which modifies the internal atmosphere by reducing
postharvest spoilage of tomatoes (Soylu et al., 2010). Furthermore, the oxygen and/or elevating carbon dioxide levels, which decreases the
investigations did not assess the antimicrobial effects on very extended fruit respiration level and metabolic activity, and hence delays the fruit
periods of storage (a few months). Vitoratos et al. (2013) reported the ripening and senescence processes (El-Ghaouth et al., 1992; Tezotto-
inhibitory effects of Thyme on spore germination of Penicillium italicum Uliana et al., 2014). A slower fruit softening, due to low polygalactur-
(Wehmer) and Penicillium digitatum [Pers. (Sacc.)] on tomatoes stored at onase activity induced by Chitosan, can explain a greater resistance to
22 °C for 6 days. Other postharvest fungal pathogens [Aspergillus flavus fungal infection, as reported by different authors (Stevens et al., 2004;
(Link) and Aspergillus niger (Tiegh.) Sacc.] were satisfactorily controlled Ruoyi et al., 2005).
by this essential oil, as reported by Ibrahim and Al-Ebady (2014). Most The quality of the tomato fruits is also an important index
chemical components of essential oils are terpenoids, including mono- evaluating the storage effect by different treatments adopted. Quality
terpenes, sesquiterpenes, and their oxygenated derivatives. Among composition of the “Vesuviano” fruits at harvest was in agreement with
them, the most active antimicrobial compounds are terpenes. Thymol the results reported by other authors for the same landrace (Ercolano
and other phenolic compounds (eugenol and carvacrol) may inactivate et al., 2008; Carli et al., 2011; Ruggieri et al., 2014). Moreover, the
essential enzymes, reacting with the cell membrane or disturbing time-course of total soluble sugars, organic acids and main volatile
genetic material functionality (Davidson, 2001). Finally, Zambonelli compounds observed in stored fruits, resembled the behaviour of
et al. (2004) correlated the fungicidal activity of different commercial “Penjar” tomato, a long-storage landrace spreads in Spain and showing
thyme essential oils with their thymol contents. similar fruit morphology to “Vesuviano” tomato (Casals et al., 2011;
Regarding the Chitosan, its use resulted in significant reduction in Missio et al., 2015). The long shelf life of the “Penjar” varietal type is
rotten fruit percentages (especially from T80) of “Vesuviano” tomatoes controlled by the ripening mutant “alcobaça” (alc), that is an allele of
and these findings appear to agree with extended antifungal activity the non-ripening gene (nor) (Casals et al., 2012).
over the time, observed by Carrieri et al. (2016). The mechanism by As far as the effect of natural fungicides, the decreasing trend in
which chitosan affects the growth of pathogens may be related to the total soluble sugars detected in Propolis-treated tomatoes suggested the
ability of this GRAS substance to interfere with the negatively charged occurrence of fermentation and senescence processes and this beha-
residues of macromolecules exposed on the fungal cell surface, resulting viour could be ascribed to utilization of these carbohydrate as

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substrates for fruit respiration. Conversely, reduced rates of respiration, position are available in literature. Anyway, Baldwin et al. (1995)
due to coating film around the fruit surface, could explain the good found that the use of edible coatings on citrus fruit resulted in an
retention of soluble sugars up to T80 in “Vesuviano” tomatoes by increase in desirable flavour compounds after storage, as compared
Chitosan treatments. Our results are in agreement with those of Zhang with uncoated fruits. This finding seems in accordance with data here
et al. (2014) that reported a lower decreasing of soluble sugars in cherry discussed, especially if considering the terpenes amounts which were
tomatoes coated with chitosan + zinc (5% + 2%) solution and stored well retained over the time in Thyme- and Propolis-treated fruits. Since
for 24 days at 20 °C with 85% R.H. The higher total sugar contents up to the terpenes are positively related to good organoleptic features, our
T80 in Thyme-treated fruits than the control were in accordance with results highlighted the maintaining of a better quality in “Vesuviano”
Tzortzakis et al. (2011), that described the behaviour of cherry tomato tomato fruits during the storage by preharvest treatments with Propolis
fruits treated with eucalyptus (Eucalyptus globulus L.) and cinnamon and Thyme essential oil.
(Cinnamomum zeylanicum Blume) essential oils and stored at 13 °C for
10 days. 5. Conclusion
Interestingly, a high amount of total organic acid content was
detected in Propolis-treated tomatoes at T0 and a good retention was Results of the present study indicated that preharvest applications of
observed until to 120 days post-harvesting. These results were in Thyme, Chitosan or Propolis reduced decay incidence during the long-
agreement with those reported by Zahid et al. (2013) on dragon fruits storage period of “Vesuviano” tomatoes under room temperature.
treated with ethanolic extract of propolis at 0.5% and stored for 20 days Among the three natural fungicides, Chitosan was the most efficacy
at 20 ± 2 °C and 80 ± 5% RH. After this treatment the authors to reduce fruit senescence processes and maintain a good overall quality
observed higher levels of titratable acidity in the fruits than the of the fruits over the time. However, further studies are required to
untreated control. Increased contents of total of organic acids were investigate different aspects of chitosan application (regarding its
also detected at harvest in “Vesuviano” tomatoes treated with Thyme: solvent, concentrations, molecular weight and deacetylation degree)
anyhow the inducing effect seemed no so extended in time. Similar as well as its antimicrobial activity on the major fungal pathogen
results were reported by Perdones et al. (2016), studying the effects of causing fruit rotting during the extended storage of “Vesuviano” and
oregano essential oil incorporated into film-forming dispersions based other similar tomatoes.
on biopolymers (chitosan and/or methylcellulose) on tomato fruits.
The positive effect of Chitosan treatments on carotenoids retention Acknowledgements
until to the end of storage is in agreement to the results reported by
Zhang et al. (2014) on tomato. By treating dragon fruits with 1.0% The authors wish to thank “Azienda Agricola Casa Barone” for
conventional Chitosan or submicron Chitosan dispersions at 600 nm hosting the experimental field and Dr. Francesco Di Dato, Dr. Riccardo
and 1000 nm, Ali et al. (2013) observed a good retention of total Riccardi and Mr. Raffaele Perreca for assistance in merceological
phenols, total flavonoids and lycopene during storage at 10 ± 2 °C and assessment of the fruits. This research was partially supported by the
80 ± 5% RH for 28 days. Campania Region (Italy) − Department of Agriculture (grants
In our study a steady increase in flavonols contents in “Vesuviano” “Programma delle attività di collaudo e sperimentazione del Centro
tomatoes was detected until to the end of storage as resulting of Orticolo Campano- IV annualità”).
preharvest treatments with Chitosan. Badawy and Rabea (2009),
reported decreased activity of polyphenol oxidase (PPO) and a con- Appendix A. Supplementary data
sequent enhancing of phenolic compounds by Chitosan in wounded
tomatoes fruit stored at different temperatures until to 21 days. The Supplementary data associated with this article can be found, in the
authors, hypothesized that the effects of this biopolymer on gray mold online version, at http://dx.doi.org/10.1016/j.scienta.2017.04.030.
(Botrytis cinerea Pers.) may be associated with direct fungitoxic proper-
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