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Postharvest Biology and Technology 53 (2009) 117–122
Contents lists available at ScienceDirect
Postharvest Biology and Technology
journa l homepage: www.e lsev ier .com/ locate /postharvbio
ssential oil amended coatings as alternatives to synthetic fungicides in citrusostharvest management
ilma du Plooya, Thierry Regnierb,∗, Sandra Combrinckb
John Bean Technologies, South Africa (Pty) Ltd, PO Box 891, Brackenfell, 7561, South AfricaDepartment of Chemistry, Tshwane University of Technology, PO Box 56208, Arcadia, Pretoria, 0001, South Africa
r t i c l e i n f o
rticle history:eceived 7 February 2009ccepted 18 April 2009
eywords:itrus
a b s t r a c t
A new approach to the control of postharvest pathogens, while maintaining fruit quality, has been imple-mented by the application of essential oil amended coatings to citrus. This approach eliminates the needfor synthetic fungicides, thereby complying with consumer preferences, organic requirements and reduc-ing environmental pollution. In vitro studies indicated that the essential oils and some of the terpenoidcomponents tested were active against Penicillium digitatum. In a series of subsequent semi-commercial
enicilliumippiapearmintostharvest
and commercial trials, Mentha spicata and Lippia scaberrima essential oils, as well as pure (d)-limoneneand R-(−)-carvone were incorporated into a variety of commercial citrus coatings. These amended coat-ings were applied postharvest to ‘Tomango’ oranges in the absence of the standard fungicide dip. Excellentdisease control was achieved with the amended coatings, while measured quality parameters indicatedthat overall fruit quality was maintained. Moreover, moisture loss was decreased significantly in fruittreated with essential oil enriched coatings. The efficacy of amended coatings as a viable alternative or
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supplement to existing fr. Introduction
Synthetic fungicides such as Imazalil, thiabendazole,yrimethanil (Smilanick et al., 2008), prochloraz (Danderson,986), and guazatine (FAO report, 1997) are generally used onacklines as the first line of defense against postharvest pathogens.ip solutions such as Imazalil, prepared in large tanks of at least500 L volume, are maintained for several days before the residue isisposed (Altieri et al., 2005). Toxic waste disposal is a costly exer-ise and hazardous waste poses serious environmental problems.urthermore, fungal pathogens have shown a concerning trendf resistance against these fungicides (Wild, 1983; El-Goorani etl., 1984), thereby shortening the life-span of protective productsEckert et al., 1994). Compounds that could equal or improveathogen control, yet avoid or minimize disposal problems woulde extremely valuable. The large volumes of citrus treated annually
nternationally, as well as the increasing demand for organicallyroduced fruit, emphasize the need to replace synthetic fungicides
ith safer and biodegradable alternatives (Wisniewski et al., 2001).Significant fruit losses after export result from decay caused byenicillium digitatum (green mould) (Porat et al., 2000). Recently,esearchers have shown an interest in the application of non-toxic
∗ Corresponding author. Tel.: +27 012 382 6126; fax: +27 021 382 6286.E-mail address: [email protected] (T. Regnier).
925-5214/$ – see front matter © 2009 Elsevier B.V. All rights reserved.oi:10.1016/j.postharvbio.2009.04.005
otection strategies was demonstrated in a commercial trial.© 2009 Elsevier B.V. All rights reserved.
alternatives or supplements to synthetic fungicides. Plant extracts,including essential oils, have been investigated as alternative mea-sures against pathological breakdown (Klieber et al., 2002; Ahmedet al., 2007). Regnier et al. (2008) proved that modification offruit coatings using essential oil of Lippia scaberrima, containing(d)-limonene, R-(−)-carvone and 1,8-cineole as main constituents(Combrinck et al., 2006), was an effective in vivo control measureagainst two mango postharvest spoilage pathogens.
In this study, in vitro investigations of essential oils and selectedterpenoids against a guazatine/Imazalil-resistant strain of P. digi-tatum were followed by semi-commercial and commercial citrustrials using supplemented coatings. ‘Tomango’ was selected for thecommercial trial since the rind is particularly prone to physiologicalbreakdown, resulting in the development of postharvest disorders.The cultivar is largely seedless with a smooth rind texture and usedprimarily for juicing. This paper shows that the pre-storage appli-cation of commercial coatings enriched with essential oils to citrusas a pathogen control measure, is feasible.
2. Materials and methods
2.1. Essential oils
Essential oil of L. scaberrima, harvested from the Wolmaransstadregion (North West Province, South Africa), was isolated by steamdistillation as described by Combrinck et al. (2006). Pure (d)-
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imonene, R-(−)-carvone and 1,8-cineole were obtained fromigma–Aldrich (Pty) Ltd. (Johannesburg, South Africa). Oil fromentha spicata (spearmint) was purchased at a later stage fromolistic Emporium (Johannesburg, South Africa), and tested sep-rately against a control, to confirm the role of R-(−)-carvone in thenhibition of P. digitatum.
.2. Pathogen isolation and toxic medium
P. digitatum was isolated from symptomatic ‘Valencia’ orangesbtained post-packline from Fort Beaufort Packhouse (Easternape, South Africa). The strain was purified and preserved at 24 ◦Cn malt extract agar (MEA, Oxoid, Johannesburg, RSA). In vitro expo-ure to 1000, 3000 and 5000 �L/L concentrations of guazatine andmazalil was used to confirm fungicide resistance. Spore suspen-ions (106 spores/mL) were prepared after 8 d incubation, by addingterile 1/4-strength Ringer’s solution to colonised Petri dishes. Then vitro fungitoxicities of the oils were determined according to the
ethod described by Regnier et al. (2008).
.3. In vivo inoculation (preventative and curative)
Commercial coatings used in all the in vivo trials were providedy John Bean Technologies (Pty) Ltd South Africa. Freshly harvested
Valencia’ fruit were surface sterilized and prepared for inocula-ion by inflicting a deep rind wound as described by Plaza et al.2004). Each of the 10 treatments consisted of 20 fruit and wasarried out in duplicate. This protocol was repeated in a separaterial. Curative treatments were executed by first dipping fruit in apore suspension of the pathogen for 3 min and drying overnightefore applying the appropriate treatment. Preventative treatmentsere done by first applying the specific treatment to wounded
ruit, leaving the fruit to dry overnight at ambient temperature21 ◦C) and then exposing treated fruit to the pathogen by dip-ing into the spore suspension for 3 min. In each case, controlreatments consisted of spraying the wounded fruit with sterileater only, before or after inoculation as relevant. Both curative andreventative treatments consisted of spraying with a commercialoating (Carnauba Tropical®) or a coating amended with L. scaber-ima essential oil (2500 �L/L). In addition, the spray applicationf commercial chemicals, 1000 �L/L thiabendazole or a mixture of000 �L/L Imazalil/guazatine (1:1), were included for comparison.reated fruit were placed, together with a beaker of distilled water,nto black plastic bags to ensure high relative humidity (±85% RH).ruit were stored for 6 d at ambient temperature (21 ◦C) in sealedags before opening the bags to allow development of the pathogen.he effects of treatments were evaluated after 4 d, and data wereecorded as the percentage of diseased fruit per treatment (Vero etl., 2002).
.4. Semi-commercial trials (hand dipping)
The semi-commercial trial (August 2007) consisted of untreatedValencia’ oranges obtained from Crocodile Valley (Nelspruit,
pumalanga Province, South Africa). Four treatments, each con-isting of three cartons (count 60), were applied at Citrusesearch International, Nelspruit. The control treatment consistedf untreated fruit washed at 40 ◦C. A standard commercial treat-ent was prepared by submersion of the fruit into a solution of
rochloraz at 40 ◦C, followed by coating with QuickDry Poly®. Forhe essential oil amended coatings, fruit were washed at 40 ◦C,
ipped by hand into QuickDry Poly® amended with Lippia essentialil (2000 and 3000 �L/L), respectively. No commercial fungicidesere included in the latter treatments. Treated fruit were air-dried,acked into commercial cartons, weighed and stored at 10 ◦C for 5eeks, followed by 1 week at 25 ◦C. Fruit quality parameters eval-nd Technology 53 (2009) 117–122
uated were weight loss, juiciness, degree Brix (TSS), pH, titratableacidity and taste. Weight loss (expressed as a percentage) was calcu-lated on the basis of a comparison of initial and final weights. Onekilogram of fruit was randomly selected from each carton, juiced(Braun Model 4161, Spain) and the volume recorded. Total solublesugar was measured using a pocket refractometer (PAL-3, AtagoInstruments, Tokyo, Japan) and pH was determined using a hand-held meter (pH Spear, Eutech Instruments, Oaklon, Vernon Hills,IL, USA). Titratable acidity was calculated by titrating a 20 mL juicevolume against a standardized NaOH solution to a phenolphthaleinendpoint. A panel consisting of six nonsmoking individuals familiarwith the taste and quality of ‘Valencia’ fruit was used for sensoryevaluation and identification of any off-flavours. Upon terminationof the trial, ten fruit from each treatment were removed from coldstorage and allowed to reach room temperature for 6 h. Thereafter,fruit were quartered and presented in random order to each pan-elist on a white plate. Each individual assessed at least one samplefrom each treatment at the same session. The parameters evaluatedwere juiciness, aroma, freshness, presence of flavours associatedwith fermentation and colour. Ratings were given according to thehedonic scale commonly used in the food industry (Srinivasa et al.,2004).
2.5. Commercial trial
Commercial coatings from the StaFresh series (SF865MS, abbre-viated MS1, SF875MS, abbreviated MS2) and the organicallyacceptable Carnauba Tropical® formulation, were used. The mid-season coating MS1, rather than a high-shine coating, was usedto apply a lower content of solids on the rind-sensitive fruit.A control of MS2 was applied as this was used by the pack-house for their export consignments. Approximately 2.4 tonnes ofuntreated, freshly harvested ‘Tomango’ fruit were obtained fromR10 Citrus in Letsitele (Limpopo Province, South Africa) and pro-cessed in the packhouse on the same day in May 2008. The sortedfruit were divided into seven experimental and two control treat-ments. Each treatment consisted of nine cartons (average 15 kgsize-sorted fruit per carton). The control treatments were done bydipping the fruit in a mixture of Imazalil (500 �L/L) and guaza-tine (1000 �L/L), followed by spray application of MS1 and MS2,respectively. The experimental treatments consisted of CarnaubaTropical® or MS1, amended with 2500 �L/L Lippia oil, spearmintoil, limonene or R-(−)-carvone, respectively, without the inclusionof synthetic fungicides. In this trial the treatment consisting of Car-nauba Tropical® amended with R-(−)-carvone was omitted. Treatedfruit were packed into commercial cartons, weighed and stored at10 ◦C. All cartons were re-weighed after 30 d in cold storage fol-lowed by 1 week at room temperature (22 ◦C). Disease incidencewas evaluated by visual inspection of the entire content of eachcarton. Weight loss was recorded and three cartons of each treat-ment were evaluated for overall quality. Ten fruit were randomlyselected from each carton, halved and the pulp colour (La*b*) deter-mined using a CR-400/410 chromameter (Konica Minolta, Narich,Johannesburg, South Africa). An additional 3 kg of fruit was ran-domly selected from each carton and juiced. The juice was weighedand used to determine pH, total soluble sugar, titratable acidity asdescribed above. Five factory workers involved with routine juic-ing, were used to assess the taste of the juice. The presence orabsence of off-flavours was recorded. To determine the shelf-life ofthe remaining fruit, the cartons were again left at room temperaturefor another 30 d before re-evaluation as described above.
2.6. Statistical analyses
Analysis of variance (ANOVA) among averages was per-formed using one-way ANOVA (single factor, and two factor
W. du Plooy et al. / Postharvest Biology a
Table 1Effects of toxic media (MEA incorporating different concentrations of inhibitors) onmycelial growth inhibition of Penicillium digitatum after six days incubation at 23 ◦C.
Inhibitor Inhibition (%) at concentration (�L/L) of
500 1000 2000 3000
Lippia scaberrima 52 Bd 62 Cc 73 Cb 100 AaMentha spicata 57 Ab 100 Aa 100 Aa 100 AaR-(−)-carvone 60 Ab 100 Aa 100 Aa 100 Aa(d)-Limonene 23 Cc 30 Db 34 Db 50 Ba1,8-Cineole 58 Ad 70 Bc 83 Bb 100 Aa
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verages (n = 10) followed by the same upper-case letter or lower-case letter did notiffer significantly within a column (upper-case) or row (lower-case), respectivelyt P ≤ 0.05.
ithout replication). After applying the least significant differ-nce (LSD) test, differences of P ≤ 0.05 were considered to beignificant.
. Results
.1. In vitro assays
The toxic medium study revealed that spearmint oil and R-(−)-arvone completely inhibited the mycelial growth of P. digitatum atoncentrations of 1000 �L/L and higher (Table 1). At a concentrationf 500 �L/L, spearmint oil and R-(−)-carvone caused approximately0% inhibition of the fungus. L. scaberrima oil and 1,8-cineole wereffective at 3000 �L/L, but fungal inhibition decreased to 73 and3%, respectively at a concentration of 2000 �L/L. (d)-Limonene was
neffective against the pathogen with only 50% inhibition achievedt 3000 �L/L.
.2. In vivo inoculation (preventative and curative)
Virulence of the isolate was confirmed by the presence ofypical lesions on all control fruit (Table 2). Preventative dip-reatment with the Carnauba Tropical® coating supplementedith the essential oil (2500 �L/L), reduced lesion development
ompletely when compared to the inoculated control. The unsup-lemented coating applied before inoculation, provided only 50%ontrol. Fruit treated preventatively with thiabendazole or theixture of Imazalil/guazatine alone yielded, correspondingly, 15
nd 5% decayed fruit. Similar results (15 and 10%, respectively)ere obtained for the curative treatment using these commercial
ungicides. In the curative treatments, the addition of essential oil
o Carnauba Tropical® coating reduced the occurrence of decayy 63% when compared to fruit treated with unsupplementedoating.able 2ffects of curative or preventative applications of oil amended coatings on the controlf Penicillium digitatum after six days at ambient temperature (21 ◦C) in sealed bags,ollowed by four days in an open bag.
reatment Percentage diseased fruit
Preventativetreatment
Curativetreatment
istilled water (control) 100 Aa 100 Aaarnauba Tropical® alone 50 Bb 80 Baarnauba Tropical® plus Lippia scaberrima oil 0 Eb 5 Eahiabendazole alone 15 Ca 15 Camazalil/guazatine alone 5 Db 10 Da
verages (n = 10) followed by the same upper-case letter or lower-case letter did notiffer significantly within a column (upper-case) or row (lower-case), respectivelyt P ≤ 0.05.
nd Technology 53 (2009) 117–122 119
3.3. Semi-commercial trials (hand dipping)
Untreated control fruit had the greatest weight loss(8.29 ± 2.38%) compared to the commercial treatment(5.47 ± 0.16%) and the treatments using amended coatings. Amere 2.49 ± 0.25% weight loss was recorded for the coatingamended with 2000 �L/L essential oil, while the 3000 �L/L treat-ment further reduced the weight loss to 1.84 ± 0.15%. The juiciness,expressed as mL/kg, followed a similar trend with untreated fruityielding 475 ± 7.3, the commercial treatment yielding 564 ± 5.7and 2000 and 3000 �L/L amended coatings yielding 557 ± 2.2and 554 ± 4.1, respectively. The values obtained for titratableacidity, total soluble sugars and pH did not differ significantlybetween treatments (data not shown). In addition, no differencesin olfactory parameters were indicated by the panelists betweenthe various treatments. In particular, no off-flavours or unfamiliarodours could be identified by any of the panelists.
3.4. Commercial trial using ‘Tomango’
All fruit used in the commercial trial had some form of pro-phylactic applied, and disease incidence therefore was low in alltreatments. Only a single fruit completely infected with P. digitatumwas found in one box for the treatments: Carnauba supplementedwith Lippia oil, Carnauba supplemented with spearmint oil andMS1 amended with R-(−)-carvone. However, despite the expo-sure of the remaining fruit to the high spore load present, not asingle co-infected fruit could be found. Furthermore, some fruitpresenting wounds with clear signs of pathogen stasis were foundin treatments of MS1 amended with limonene and spearmint.These fruit appeared to have become infected following injury sus-tained during packing. In addition, no phytotoxic reactions wereobserved, although a few examples of fruit with oleosis were ran-domly found, as well as fruit displaying signs of chafing and othersuperficial blemishes, but none of these were infected. At the ter-mination of the trial (30 d cold storage followed by 60 d at ambienttemperature), fruit treated with supplemented coatings exhibitedsignificantly lower weight loss than the controls treated with syn-thetic fungicides and coatings. Carnauba Tropical® and MS1, bothsupplemented with spearmint oil, as well as MS1 supplementedwith R-(−)-carvone, displayed the least weight loss (6.82, 8.49 and8.46%, respectively), but these values did not differ significantlyfrom those of the other amended coatings (Fig. 1A). Values obtainedfor moisture loss and juiciness were well correlated (Fig. 1A and B)and closely represented the inverse of one another for all the treat-ments. No significant differences were found between the pH anddegree Brix values (Fig. 1C and D) as well as between the titratableacidity and pulp colour values (data not shown) for all the treat-ments. Juice obtained from fruit treated with essential oil amendedcoatings could not be distinguished from that obtained from fruittreated with the conventional treatments.
4. Discussion
The use of citral (Klieber et al., 2002) and alicine from gar-lic (Obagwu and Korsten, 2003) are examples of the potentialefficacy of plant extracts for the control of P. digitatum. Manystudies have documented the in vitro efficacy of essential oilsagainst fruit postharvest spoilage pathogens (Klieber et al., 2002;Bakkali et al., 2008). However, application of these plant sec-
ondary metabolites in the packhouse environment was previouslyfound to be ineffective (Plaza et al., 2004). Some success has beenachieved using essential oils as volatiles to combat decay of fruitand vegetables (Tabanca et al., 2007; Tunc et al., 2007; Serrano etal., 2008). The incorporation of essential oils into fruit coatings,
120 W. du Plooy et al. / Postharvest Biology and Technology 53 (2009) 117–122
F s (B),a aubaC S1 wito e dip f
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ig. 1. Effect of essential oil amended coatings on moisture loss (A), juicinesfter storage. Carn + Lim = Carnauba Tropical® with limonene; Carn + Lippia = Carnhem + MS1 = standard fungicide dip followed by coating with MS1; MS1 + Lim = Mil; MS1 + Carvone = MS1 with pure R-(−)-carvone; Chem + MS2 = standard fungicid
rimarily applied to retain moisture, is gaining popularity. Thedvantage of using coatings amended with essential oils, ratherhan vapour, is that there is closer contact between the essentialils and fruit surfaces, allowing exposure of each fruit to similaroncentrations of inhibitor over a longer period. Amiri et al. (2008)pplied different formulations amended with eugenol oil (Euge-ia caryophylata) to two apple cultivars and successfully reducedhe disease incidence after cold storage. Trans (isomerized) jojobail was applied by Ahmed et al. (2007) as a coating for ‘Valencia’ranges. They effectively maintained fruit quality for up to 60 dsing concentrations of 20–30% of (trans)-jojoba oil. Recently ouresearch group successfully achieved pathogen control, withoutny observed physiological breakdown, by applying commercial
oatings amended with L. scaberrima essential oil to mango fruitRegnier et al., 2008). This positive outcome prompted the exten-ion of the application of supplemented coatings to citrus.The antifungal role of R-(−)-carvone against Penicillium citrinums indicated by Saleh et al. (2006), and against mango pathogens
pH (C) and total soluble sugar (degree Brix) content (D) of ‘Tomango’ fruitTropical® with Lippia oil; Carn + Spear = Carnauba Tropical® with spearmint oil;h limonene; MS1 + Lippia = MS1 with Lippia oil; MS1 + Spear = MS1 with spearmintollowed by coating with MS2.
(Regnier et al., 2008), was confirmed by the in vitro results obtainedin this study against P. digitatum (Table 1). We later found thatspearmint oil, which contains more than 80% R-(−)-carvone, wasequally effective in inhibiting fungal growth. L. scaberrima oil,containing only 34% R-(−)-carvone, was less effective than R-(−)-carvone and spearmint oil as an inhibitor. (d)-Limonene wasineffective at controlling P. digitatum, emphasizing the poor effi-cacy of the terpenoid against fruit pathogens as previously observedby Regnier et al. (2008). Findings by Eckert and Ratnayake (1994)have shown that germination of Penicillium spores is stimulatedwhen exposed to volatiles released by wounded citrus fruit. Drobyet al. (2008) found that (d)-limonene, a known major constituentof orange oil, stimulates germ tube elongation in green and blue
moulds.For total protection of the fruit against Penicillium rot, fromthe tree to the consumer, the fungicide applied must be effectiveregardless of the timing of micro-injuries. In both pre- and posthar-vest environments P. digitatum is a well-known wound pathogen
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Droby et al., 2008). Therefore, both curative and preventativereatments with coatings supplemented with L. scaberrima wereompared in this study and found to be superior to the standardommercial application of synthetic fungicides. The coating alone,lthough serving as a protective layer, is clearly not sufficient toimit decay (Table 1) and the addition of fungicide, whether natu-al or synthetic, is imperative. A further general observation washat amended coatings yielded fruit exhibiting no shrivelling orrowning even after ten days of storage.
In the semi-commercial trial involving hand-dipping of ‘Valen-ia’ fruit, an encouraging result was the significant reduction ineight loss of fruit treated with essential oils compared to that
f the standard control treatments. In all cases, the addition of. scaberrima oil maintained the tested quality parameters of theruit following storage. Although the hand-dipping trial gave a goodndication of the in vivo efficacy of the essential oils, the results
ay differ to those obtained when adapting the coating meth-ds applied to a commercial setting where spray application isone.
‘Tomango’ was selected as the cultivar for the commercial trialsecause the rind sensitivity is well-suited to reveal any phytotoxiceactions, displays physiological stress and allows the detectionf any infection due to ineffective postharvest protection. Resultsbtained for this cultivar can now be extrapolated to more robustarieties. Essential oil amended coatings are currently being eval-ated by our research group for application in major South Africanxport commodities ‘Valencia’ and lemons (‘Eureka’).
(d)-Limonene was included as a terpenoid for coating supple-entation, because it naturally occurs in citrus rind (Droby et
l., 2008). Although they found that limonene is able to stimu-ate fungal growth, our in vivo results using this terpene did noteveal significant differences between this treatment and the oth-rs with regard to disease incidence and overall quality. Spearmintil was used for supplementation, since the in vitro results (Table 1)btained at a later stage indicated that this oil was more effectivehan L. scaberrima oil against P. digitatum. Spearmint oil is commer-ially available in large quantities, is relatively inexpensive and cane purchased with a consistent R-(−)-carvone content rendering itighly suitable as a coating supplement.
The three isolated infections observed and associated with aigh spore load, suggest that sufficient protection is provided byhe essential oil amended coatings. Furthermore, fruit display-ng wounds with clear signs of pathogen stasis confirmed theungicidal activity of the oils and their components. The essen-ial oils and individual terpenoids appeared to enhance the abilityf the coatings to control gas exchange, thereby reducing mois-ure loss and increasing juiciness. Essential oils could therefore bextremely valuable to prevent shrivelling during prolonged exporteriods, extend shelf-life and preserve the outer appearance of theruit. ‘Tomango’ is an important juicing variety, with the pH andugar content of the fruit being of importance. The organolepticarameters measured compared favourably with those of the con-rol treatments as reflected by the pH, degree Brix (Fig. 1C and) and titratable acidity values. No significant colour differencesf the pulp were discernable between the different treatments.lthough the fruit treated with spearmint oil had a strong odour
mmediately after treatment, no spearmint odour was discernableollowing cold storage. The absence of off-flavours is encouraging,ince consumers are particularly sensitive to any hint of fermenta-ion.
The moisture retention observed may be related to the affinity of
dded essential oils to those terpenoid components present in theitrus rind, thereby improving film formation. The coating selecteds of primary importance, since the solids composition and contentf the wax and the emulsification systems applied are determinatearameters for the compatibility of the coating with the essentialnd Technology 53 (2009) 117–122 121
oil. Moreover, the layer uniformity and structural integrity of theamended coating on the fruit surface are also dependent on thecoating chemistry.
In conclusion, the innovative method of coating application pro-posed in this paper not only promotes fruit moisture retentionwhile maintaining fruit quality, but completely eliminates the useof fungicide. The exclusion of fungicide eradicates the need to dis-pose of toxic waste, thereby contributing significantly to a reductionin environmental pollution and the carbon footprint resulting fromcitrus production.
Acknowledgements
We thank Dr. K. Lezar from Citrus Research International, Pack-house Crocodile Valley Packhouse and R10 for the use of theirfacilities. We also thank Dr. John Mildenhall for providing the Peni-cillium digitatum isolate, Marius du Preez and Wilma Augustynfor technical assistance, and the National Research Foundation forfinancial support.
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