Impaginato 389 Adv. Hort. Sci., 2018 32(3): 389-398 DOI: 10.13128/ahs-23229 Postharvest control of Aspergillus niger in mangos by means of essential oil S. Javadpour 1, A. Golestani 2, S. Rastegar 2 (*), M.M. Dastjer 2 1 Food and Cosmetic Health Research Center, Hormozgan University of Medical Sciences, Bandar Abbas, Iran. 2 Department of Horticultural Science, College of Agriculture, University of Hormozgan, Hormozgan, Iran. Key words: Aspergillus niger, essential oil, mango, postharvest. Abstract: The use of essential oil as an alternative mean to synthetic fungicides has been considered in the past years for management of the postharvest decay of fruits in order to ensure more safe and long storage life of these per- ishable commodities. Aspergillus niger is one of the most dangerous fungal pathogen which can cause postharvest diseases in fresh mangos. The aim of this study was to assess the effectiveness of essential oil from four aromatic plants (Thymus vulgaris, Salvia mirzayanii, Artemisa persica, and Rosmarinus officinalis) in comparison to fungicide ‘Mancozeb’ against A. niger under in vitro and in vivo conditions. After inoculation of mango fruits with an isolate of A. niger followed by curative treatments with essential oil, the main physical and chemical attributes of mangoes were determined under postharvest condi- tion. The in vitro results showed that colonies of A. niger were totally inhibited by application of essential oil of T. vulgaris (at all the tested concentrations) and A. persica (1500 μl/l). While, S. mirzayanii showed the lowest effect at 1000 μl/l if compared with the other essential oils. The results of the in vivo experiments showed that treatments with T. vulgaris and S. mirzayanii essen- tial oil had significant (P<0.05) effects in preventing fruit decay at 1000 μl/l after 10 days of storage, while, R. officinalis essential oil significantly (P<0.05) reduced deterioration of mango fruits at 500 μl/l, followed by A. persica. Rosemary also showed the highest fruit firmness in comparison with other treatments. Also, the essential oils maintained higher chlorophyll content. The results of this work showed that application of essential oil on mangos assur- ance both a significant preservation on their quality attributes by controlling, at the same time, decaying caused by A. niger during the postharvest phase. 1. Introduction Postharvest diseases are among the major causes of losses of mangos (Mangifera indica L.) fresh produce throughout the supply chain. The inci- dence of the postharvest diseases can also affect the quality of mangos limiting their shelf life up to 3-4 days. In literature is reported that about 17-37% of fresh mangos is wasted after harvesting and marketing (Madan (*) Corresponding author: rastegarhort@gmail.com Citation: JAVADPOUR S., GOLESTANI A., RASTEGAR S., DASTJER M.M., 2018 - Postharvest control of Aspergillus niger in mangos by means of essential oil. - Adv. Hort. Sci., 32(3): 389-398 Copyright: © 2018 Javadpour S., Golestani A., Rastegar S., Dastjer M.M. This is an open access, peer reviewed article published by Firenze University Press (http://www.fupress.net/index.php/ahs/) and distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: All relevant data are within the paper and its Supporting Information files. Competing Interests: The authors declare no competing interests. Received for publication 14 May 2018 Accepted for publication 12 September 2018 AHS Advances in Horticultural Science Adv. Hort. Sci., 2018 32(3): 389-398 390 and Ullosa, 1993). Sharma et al. (1994) have reported that about 17.7% of this fresh produce is lost during the storage and marketing. Mango decay caused by the plant pathogenic fungus Aspergillus niger is one of the most dangerous postharvest diseases, leading t o t h e l o s s e s o f f r u i t q u a l i t y d u r i n g s t o r a g e (Duamkhanmanee, 2008). It well known from published reports as more negative effects associated to use of chemical fungi- cides for controlling postharvest diseases have been reported on the human’s health and environment (Wightwick et al., 2010). Furthermore, consumers believe that fruits not treated (or minimally-treated) with fungicides are safer for fresh consumption (Du Plooy et al., 2009). In the past 20 years there has been a great interest in using essential oils (EOs) to control postharvest diseases, such increasing shelf life of stored fruits (Tripathi and Dubey, 2004). Several studies have also reported on the antifungal activity of Thymus vulgaris against different strains of Colletotrichum gloeosporioides, Rhizopus stolonifer, P e n i c i l l i u m d i g i t a t u m ( A b d o l a h i e t a l . , 2 0 1 0 ; S e l l a m u t h u e t a l . , 2 0 1 3 ) , A s p e r g i l l u s f l a v u s , A s p e r g i l l u s n i g e r , A s p e r g i l l u s f u m i g a t u s , a n d Alternaria alternata (Kumar et al., 2008). Different species of Salvia are yet used as antimicrobial agents (Fiore et al., 2006; Kelen and Tepe, 2008), however Salvia mirzayanii is an endemic plant which grows only in some parts of Iran, and thus there is no an exhaustive information regarding its effect on A. niger (Rechinger, 1986). Centeno et al. (2010) have reported that extracts from Rosmarinus officinalis and T. vulgaris could have a significant effect on the control of fungal decaying. Previous studies have also confirmed the interesting antimicrobial activity of R. officinalis EO against spoilage and pathogenic food- related fungi (Abdolahi et al., 2010). De Sousa et al. (2013), for instance, detected the strong effect of the Origanum vulgare and R. officinalis EOs in controlling A. flavus. On the other hand, fewer papers report issues regarding to postharvest control of A. niger on mango fruits using EOs as an alternative mean to syn- thetic fungicides. This study was focused to evaluate the effectiveness of EOs derived from four aromatic plants (T. vulgaris, S. mirzayanii, Artemisa persica and R. officinalis) to in vitro and in vivo suppress the growth of one pathogenic strain of A. niger by pre- serving the mango fruit quality attributes under postharvest condition. 2. Materials and Methods Plant material and extraction of essential oil S. mirzayanii and A. persica samples were collect- ed from Lar region of Fars Province, Iran (Lat. 27°41’ 3’’ N and Long. 54°2’ 10’’E). A T. vulgaris sample was collected from Geno region of Hormozgan Province, Iran (Lat. 25° 38’ 37.9’’ N and Long. 57° 46’ 28’’ E) and R. officinalis samples from Kerman Province, Iran (Lat. 30° 17’ 2.1’’ N and Long. 57° 5’ 0.1’’ E). Samples were harvested in vegetative stage (before flower- ing). The leaves of samples were cut into small pieces and shade-dried at room temperature. The material was then ground to fine powder. The 80 g of plant material were subjected to extraction of EOs by hydro-distillation method for 6 h using a Clevenger’s apparatus (Moghaddam et al., 2011). The EOs were separately collected, dehydrated using sodium sul- phate (Na2SO4), and finally stored in a dark bottle at 4°C until tested. In vitro experiments Fungi were isolated from mango and their identity was confirmed The Aspergillus niger (PTCC 5010) was supplied by Iranian research organization for science and tech- nology (IROST). Culture of the pathogen organism was maintained on potato dextrose agar (PDA) medi- um. Stock cultures were grown at 25°C for 7 days to allow for sufficient sporulation. Antifungal effects of EOs were carried out by the Solution Method (SM) according to Pitarokili et al. (1999). Inhibitory effects of EO extracted from S. mirzayanii, A. persica, R. officinalis and T. vulgaris were determined by in vitro antifungal assays. To measure the direct fungal inhi- bition of each EO on mycelial growth of A. niger, three different concentrations of them (1000, 1200 and 1500 μl/l) were added to potato dextrose agar (PDA; provided by Scharlau) media before solidifica- tion into Petri dishes (8 cm diameter) at 45-50°C. Fungal disks with 5 mm diameter were placed on the middle of Petri dishes and incubated at 25°C for 10 days. Three replicates per each treatment (4 EO × 3 concentrations) including control plate without EO were prepared. Inhibition percentage was deter- mined at the end of incubation time by the index: IP = (dc-dt)/dc ×100 IP = inhibition percentage, dc = mycelium diameter in the control plate, and dt = in the EOs-treated plate. Javadpour et al. - Postharvest control of Aspergillus niger in mangos 391 In vivo experiments Mangos cv. Halily were harvested from Minab (Lat. 27°07’51’’ N and Long. 57°05’13’’ E) at the maturity stage of development. Fruit surface was before disinfected with 2% sodium hypochlorite for 3 min, and then artificial inoculations were done by puncturing fruit surface (4 mm deep and 2 mm wide for each inoculation point) with a sterile needle on two sides of each fruit with 40 μl of a conidial sus- pension containing 106 UFC/ml of A. niger that it has been sprayed above each wound. After one-day of incubation at 25°C to allow conidia germination into fruit tissue, fruits were treated with 500 and 1000 μl/l EO of each plant in comparison to 0.5 and 1.0 mg/l mancozed. After the treatment (curative), all fruit trials, including the control (fruits were inoculat- ed with conidial suspension without the treatment with essential oils), were placed into boxes and kept at 25°C for 1, 2, and 3 weeks. Decay percentage of fruit was calculated as the number of decayed fruit/ total number of fruit at each replication* 100 (El- Anany et al., 2009). Physical-chemical analysis Firmness of each fruit was measured at two points of the equatorial region by using a texture analyzer with a 5 mm probe (Lurton, Taiwan) with units expressed in kg/cm2. Surface color was mea- sured on each fruit at two opposite sides using a chromameter (CR 400, Minolta) which provided CIE L*, a*, and b* values. L* is color lightness (0= black and 100= white). The a* scale shows in the maximum the red (+a*) and in the minimum the green color (- a*) while the b* ranged from yellow (+b*) to blue (- b*). The content of ascorbic acid (AA) expressed as mg/100 g fruit weight was determined as described by Molla et al. (2011). Aliquots of 10 ml of each sam- ple was homogenized in 100 ml of extraction buffer containing 3% metaphosphoric acid. Aliquots of 10 mL of homogenate was titrated against standard dye 2,6-diclorophenol indophenols to a faint pink color. The method proposed by Lichtenthaler (1987) was u s e d t o d e t e r m i n e t h e t o t a l c h l o r o p h y l l a n d carotenoids content of fruit. The ‘Total Soluble Solids’ (TSS) content was determined at 20°C using a digital refractometer, and expressed as °Brix. The pH of fruit juice was measured using a Jenway 3320 pH meter calibrated by pH 4 and 7 buffer solutions. The ‘Titratable Acidity’ (TA) was determined by titration of 5 mL extract with 0.1 mol L−1 sodium hydroxide at pH 8.1 and expressed as percent citric acid (Molla et al., 2011). Weight loss, fruit firmness, surface color c h a n g e , c o n t e n t o f A A , t o t a l c h l o r o p h y l l , a n d carotenoids, and TSS, TA and pH were determined after 1, 2, and 3 weeks of storage at 25°C. These characteristics was done with 3 replicates (3 large fruits for 1 replicate). Statistical analysis T h e e x p e r i m e n t w a s c o n d u c t e d i n a randomized factorial designed whit essential oils treatment and storage time as the two factors. Data were submitted to one-way analysis of variance (ANOVA) using SAS version 16.0 and means were separated by the Duncan test at P<0.05 (n=3). 3. Results and Discussion Aspergillus niger mycelia inhibition In vitro experiments showed that mycelia growth of A. niger was significantly suppressed (P<0.05) when treated with the different concentrations of each EO (Fig. 1). The fungal growth was totally inhib- ited (IP= 100%) by all the concentrations of T. vul- garis EO and with 1500 μl/l A. persica EO. On the other hand, S. mirzayanii EO showed lower effect (IP= 72%) than other EOs when tested at 1000 μl/l concentration. O u r f i n d i n g s a r e i n a g r e e m e n t t o t h e o n e s described by Kohiyama et al. (2015) who reported that T. vulgaris EO was able to control the growth of A. flavus. Similar observations on the prevention of Fig. 1 - Inhibitory effect of the Thymus vulgaris, Salvia mirzaya- nii, Artemisia persica and Rosmarinus officinalis EOs tested at 1000, 1200 and 1500 μl/l on mycelia growth of Aspergillus niger cultures incubated at 25°C for 10 days. Bars indicate the SD of the mean. Different letters indica- te significant differences in mean values (p<0.05). Adv. Hort. Sci., 2018 32(3): 389-398 392 different pathogenic fungi by using EOs have been reported in the previous studies, such as those con- ducted by Tripathi and Dubey (2004) and Pawar and Thaker (2006). Boubaker et al. (2016) reported the antifungal activity of four Thymus species EOs against Penicillium digitatum, Penicillium italicum and Geotrichum citriaurantii. Pawar and Thaker (2006) s h o w e d t h a t C i n n a m o m u m z e y l a n i c u m , C i n n a - m o m u m z e y l a n i c u m , C i n n a m o m u m c a s s i a , Cymbopogon citratus and Syzygium aromaticum were the best plant sources for EOs extraction show- ing a noticeable inhibitory effect against A. niger. It was also reported that the mycelial growth of A. niger was inhibited by application of 2.5 and 3.0 μg/ml of Citrus sinensis oil (sweet orange) in Potato D e x t r o s e B r o t h a n d A g a r W a t e r m e d i u m , respectively (Sharma and Tripathi, 2008). Modifications on fungal structures induced by the EOs afore-quoted might be due to interactions of their components (Carvacrol, thymol, eugenol, vanillin and etc.) with cell wall synthesis, which could affect fungal growth and its morphology (Rasooli et al., 2006; Rao et al., 2010). Some researchers have stated that some phenolic compounds present in the EOs could affect the plasma membrane and the cellu- lar organelles, such as mitochondria of the fungi by decreasing the lipid and saturated fatty acid levels and increasing the unsaturated fatty acids, resulting in the leakage of Ca2+, Mg2+ and K+ (Sharma and Tripathi, 2008). In addition, the existence of the hydroxyl groups and the aromatic nucleus could be a other important factor for the EOs antimicrobial activity (Numpaque et al., 2011). Fruit decay suppression As shown in figure 2, the decay percentage of fruits increased with the storage time, variable from 1 to 3 weeks of incubation. The percentage of decay significantly decreased (P<0.05) with increasing of the concentration of T. vulgaris and A. persica EOs after 3 weeks of storage. After one week, no significant dif- ferences were observed between control and treated fruits (data not shown). After two weeks, significant differences were found among treatments with R. officinalis EO at 500 μl/l and T. vulgaris at 1000 μl/l. At the end of experiment, after two weeks, the maxi- mum level of decay was related to the control fruits (70%), and the minimum one was attributed to R. officinalis (500 μl/l) and A. persica (1000 μl/l) EOs, reaching 12% and 13.3% decay, respectively. T h e s e d a t a a g r e e w i t h t h o s e o b t a i n e d b y Ramezanian et al. (2016) who showed the possibility of using Z. multiflora and T. vulgaris EOs to control postharvest citrus Alternaria decay (black rot). In addition, Elshafie et al. (2015) reported that O. vul- gare EO can control the brown rot of peach. Jhalegar et al. (2015) addressed their study on the influence of lemon grass, eucalyptus, clove and neem EOs against P. digitatum and P. italicum in ‘Kinnow’ man- darin. These authors showed that the decay rot dur- ing storage was less in the treated fruits than in the control ones. Duamkhanmanee (2008) reported that 4000 ppm lemon grass EO could control anthracnose by C. gloeosporioides decay of mangos. Phenolic compounds such as Carvacrol and thymol (Rao et al., 2010) contained in EOs have a lipophilic molecular structure, therefore it interfer with membrane-cat- alyzed enzymes and cell wall, causing the cell death of microbes (Shirzad et al., 2011). Many researchers believe that the type and the amount of phenolic compounds present in the oil can determine the anti- fungal activity of the EOs (Tripathi and Dukey, 2004). Weight loss The weight loss of fruit was increased strongly during the early weeks, but this increase was gradual throughout the storage period (Fig. 3). After two weeks of storage, the highest and lowest weight loss was observed in the control samples and those treat- ed with 1000 μl/l S. mirzayanii (12.7% and 9.8% respectively). During the storage, the main mango weight loss (13.2%) was found in the control fruits, while the lowest one (10.8%) was observed in R. officinalis treated fruits at 500 μl/l concentrations. The mechanism of EOs for reducing physiological Fig. 2 - Suppressive effect of the Thymus vulgaris, Salvia mir- zayanii, Artemisia persica and Rosmarinus officinalis EOs tested at 500 and 1000 μl/l on mangos decay after 10 and 15 days of storage compared to ‘mancozed’ (0.5 and 1.0 mg/l). Bars indicate the SD of the mean. Different let- ters indicate significant differences in mean values (p <0.05). Javadpour et al. - Postharvest control of Aspergillus niger in mangos 393 cell wall components. Breakdown and the enzymatic degradation of insoluble protopectins into more sim- ple soluble pectin can be associated with softening (Willats et al., 2001). Ramezanian et al. (2016) found that the EOs reduced the activity of polygalactur- o n a s e a n d g a l a c t o s i d a s e , w h i c h a r e s o f t e n i n g enzymes in the cell wall components, and maintained orange fruit firmness through the storage. The results obtained in the present study agree with those of Maqbool and Alderson (2010), who showed that by the application of lemongrass oil (0.05%) and Cinnamon oil (0.4%), the firmness of banana and p a p a y a f r u i t s w a s m a i n t a i n e d d u r i n g s t o r a g e . However, Tzortzakis (2007) reported that eucalyptus and cinnamon EOs had no effect on the tomato and strawberry firmness. Surface color change The results related to the changes in the fruit color (in terms of L*, a* and b*) of the treated mango showed that the lightness of the fruits peel was decreased throughout the storage time (Table 1). The fruits treated with 1000 μl/l R. officinalis EO retained higher L* over other treatments and control samples, after three weeks of storage. The highest and lowest L* was found in R. officinalis and A. persica at 1000 μl/l concentration, respectively. The results showed that a* was significantly decreased during the storage. However, the fruits treated with EOs maintained a higher a* than did the control ones. At the end of the storage, the lowest a* (7.46) and the highest a*(11.73) were found in the control and R. officinalis treated fruits at 1000 μl/l concentra- loss in weight might be related to the reduction of ethylene production and the respiration rate. Also, EOs cover the peel of fruit, creating the water barrier between the fruit and the environment, thereby reducing water exchange (Morillon et al., 2002). This agrees with previous studies showing the efficacy of EOs in reducing the weight loss of cherries and grapes (Serrano et al., 2005). Similarly, Du Plooy et al. (2009) reported that the use of Mentha spicata and L i p p i a s c a b e r r i m a E O s r e d u c e d w e i g h t l o s s i n ‘Valencia’ oranges. Fruit firmness A continuous decline in mangos firmness was observed throughout storage (Fig. 4). However, the fruits treated with EOs showed higher firmness than the control ones. In each stage of storage, no signifi- cant difference was identified in the firmness of fruits, at different concentrations of EOs. After 3 weeks of storage, the firmness of control fruits was around 3.06 kg/cm2, while the treated fruits were sig- nificantly firmer (P<0.05). In this stage, fruits treated with mancozeb showed no significant difference, as compared with those treated with 500 μl/l T. vulgaris EO. However, R. officinalis in both concentrations showed the highest firmness, as compared with other treatments. Firmness, as one of the fruits properties, is a com- plex sensory attribute that also includes crispiness and juiciness; it is important in determining the acceptability of horticultural crops. It has been accepted that the loss of fruit firmness throughout the storage is mainly due to the depolymerization of Fig. 3 - E f f e c t o f t h e T h y m u s v u l g a r i s , S a l v i a m i r z a y a n i i , Artemisia persica and Rosmarinus officinalis EOs tested at 500 and 1000 μl/l on mangos weight loss after 1, 2, and 3 weeks of storage compared to ‘mancozed’ (0.5 and 1.0 mg/l). Bars indicate the SD of the mean. Different let- ters indicate significant differences in mean values (p <0.05). Fig. 4 - E f f e c t o f t h e T h y m u s v u l g a r i s , S a l v i a m i r z a y a n i i , Artemisia persica and Rosmarinus officinalis EOs tested at 500 and 1000 μl/l on mangos firmness after 1, 2, and 3 weeks of incubation compared to ‘mancozed’ (0.5 and 1.0 mg/l). Bars indicate the SD of the mean. Different let- ters indicate significant differences in mean values (p <0.05). 394 Adv. Hort. Sci., 2018 32(3): 389-398 tion, respectively. The results also showed that b* was increased during the storage time, but the trend in fruits treated by EOs was slower than that in the control. At the end of storage, S. mirzayanii and R. officinalis, at 500 μl/l concentrations, showed the minimum b* value (40.7 and 40.6), respectively. However, control and mancozeb samples showed the maximum b* value (56.6 and 55.9), respectively. The results obtained in the present study showed that EOs treatment could have better litheness with lower a* and b*, as compared to mancozed and control groups. Ramezanian et al. (2016) showed the effect of the Zataria multiflora and T. vulgaris EOs on the black rot of ‘Washington Navel’ orange fruit. They found that the best color was related to zataria at 300 μl/l and thyme EOs at 400 μl/l concentrations. In agreement with our findings, Marjanlo et al. (2009) showed the effect of Cumin EO on the postharvest quality of strawberries, finding that the essential oil treated fruits maintained a higher L* during storage in com- parison with the controls. AA content Ascorbic acid (vitamin C) content was gradually decreased during storage; however, its strength was lower in the treated samples (Table 2). Different con- centrations of the EOs significantly maintained ascor- bic acid content, as compared to the control. Overall, the most (14 mg/100 g/1) and the least (9 mg/100 Table 1 - Effect of the Thymus vulgaris, Salvia mirzayanii, Artemisia persica and Rosmarinus officinalis EOs tested at 500 and 1000 μl/l on mangos color change ± SD after 1, 2, and 3 weeks of storage compared to ‘mancozed’ (0.5 and 1.0 mg/l) In each character, different letters indicate significant differences in mean values (p<0.05). Testing index Treatment (concentration) Storage time Week 1 Week 2 Week 3 L* Control 63.3±0.43 a 47.5±0.56 n 46.2±0.56 n Mancozeb 0.5 mg/l 64.9±0.5 a 59.8±0.48 c-g 54.1±0.59 j-m Mancozeb 1 mg/l 55.3±.23 f-l 55.9±0.75 f-l 56.7±0.76 f-l Thymus vulgaris 500 μl/l 62.1±0.33 a-d 54.8±0.46 i-m 51.4±0.36 mn Thymus vulgaris 1000 μl/l 60.6±0.54 b-f 57.8±0.49 e-j 51.8±0.66 mn Salvia mirzayanii 500 μl/l 54.6±0.32 i-m 51.9±0.58 l-n 56.1±0.54 f-k Salvia mirzayanii 1000 μl/l 59.9±0.33 c-g 58.4±0.44 d-g 49.7±0.38 mn Artemisa persica 500 μl/l 59.7±0.43 c-g 56.9±0.58 f-l 48.7±0.65 n Artemisa persica 1000 μl/l 57.9±0.23 e-j 58.8±0.75 d-g 41.4±0.65 o Rosmarinus officinalis 500 μl/l 59.8±0.19 c-g 60.7±0.66 a-d 54.1±0.39 j-m Rosmarinus officinalis 1000 μl/l 62.2±0.25 a-d 65.3±0.76 a 59.5±0.53 c-g a* Control 16.4±0.87 12.6±0.87 g-i 7.4±0.87 l Mancozeb 0.5 mg/l 17.2±0.87 15.8±0.88 a-e 8.1±0.98 l Mancozeb 1 mg/l 16.8±0.99 16.1±0.98 a-d 8.6±0.99 kl Thymus vulgaris 500 μl/l 16.4±0.67 16.1±0.76 b-f 8.6±0.76 kl Thymus vulgaris 1000 μl/l 17.2±0.89 15.3±0.87 a-c 11.2±0.85 ij Salvia mirzayanii 500 μl/l 17.1±0.78 16.7±0.88 c-f 11.1±0.76 ij Salvia mirzayanii 1000 μl/l 17.6±0.77 15±0.68 e-g 8.4±0.87 kl Artemisa persica 500 μl/l 17.1±0.77 14.2±0.87 f-g 10.8±0.97 ij Artemisa persica 1000 μl/l 17.4±0.69 13.5±0.88 f-h 11.6±0.97 h-j Rosmarinus officinalis 500 μl/l 16.8±0.90 15.8±0.98 a-e 11.7±0.98 ij Rosmarinus officinalis 1000 μl/l 14.6±1.02 16.3±0.96 a-d 10.1±1.03 jk b* Cntrol 25.5±0.78 ij 34.7±1.02 e 56.6±2.2 a Mancozeb 0.5 mg/l 20.7±1.03 l-n 30±1.03 fg 55.9±3.2 a Mancozeb 1 mg/l 21.5±0.85 l-n 32.9±1.07 ef 46.7±2.9 bc Thymus vulgaris 500 μl/l 20.8±0.87 mn 25.1±0.84 h-j 46.7±3.8 bc Thymus vulgaris 1000 μl/l 20.6±0.78 mn 30.6±1.06 fg 49.4±3.5 b Salvia mirzayanii 500 μl/l 17.0±0.78 ab 30.7±0.94 fg 40.6±2.6 d Salvia mirzayanii 1000 μl/l 23.8±1.03 i-l 27.3±0.99 gh 48.3±3.5 bc Artemisa persica 500 μl/l 19.4±0.86 n 26.6±0.95 hi 47.8±3.6 bc Artemisa persica 1000 μl/l 20.8±0.98 l-n 30.8±1.04 fg 47.6±3.2 bc Rosmarinus officinalis 500 μl/l 20.7±1.03 l-n 24.4±0.90 i-k 40.7±2.8 d Rosmarinus officinalis 1000 μl/l 22.8±0.99 j-m 23.4±1.05 i-l 45.5±3.5 c 395 Javadpour et al. - Postharvest control of Aspergillus niger in mangos g/1) amount of ascorbic acid content was detected in the fruits treated with S. mirzayanii at the concentra- tion of 500 μl/l and control after three weeks of stor- age, respectively. In general, fruits treated with man- cozeb showed lower ascorbic acid content in compar- ison with those treated with EOs throughout the storage. In agreement with our findings, Geransayeh et al. (2012) showed that the vitamin C content of grapes was decreased significantly during the storage; how- ever, a higher vitamin C amount was observed in the samples treated with T. vulgaris EO. Our results were nevertheless in contrast with those of Marjanlo (2009) who did not detect any significant difference in the amount of ascorbic acid in strawberry fruits treated by the Essential Oils. Carotenoids and total chlorophyll content Analysis of the variance of carotenoids content revealed a significant difference (p< 0.05) between t r e a t m e n t s ( T a b l e 2 ) . T h e c o n c e n t r a t i o n o f carotenoids content was low at the initial time of storage and then significantly increased during stor- age. At the end of the process, control fruits showed the highest content of carotenoids (1.94 mg 100 g-1). R. officinalis and T. vulgaris, tested at 500 μl/l show- ing 1.09 and 1.15 (mg 100 g-1) as carotenoids content, respectively, were the lowest one, as compared with other treatments. The total chlorophyll content was gradually decreased to a lower concentration in all Table 2 - Effect of the Thymus vulgaris, Salvia mirzayanii, Artemisia persica and Rosmarinus officinalis EOs tested at 500 and 1000 μl/l on the ascorbic acid, carotenoids and total chlorophyll content ± SD in mangos after 1, 2, and 3 weeks of storage compared to ‘mancozed’ (0.5 and 1.0 mg/l) In each character, different letters indicate significant differences in mean values (p<0.05). Active metabolite Treatment (concentration) Storage time (week) Week 1 Week 2 Week 3 Ascorbic acid Control 12.3±3 cd 11±2 cd 9.5±4 d Mancozeb 0.5 mg/l 13.3±4 c 12.6±3 cd 11±3 cd Mancozeb 1 mg/l 13.5±4 c 12.3±3 cd 11±2 cd Thymus vulgaris 500 μl/l 14.8±7 bc 13.6±4 c 12±4 cd Thymus vulgaris 1000 μl/l 14.5±4 bc 13.7±4 c 12±5 cd Salvia mirzayanii 500 μl/l 17.4±4 a 16.4±4 b 14±5 bc Salvia mirzayanii 1000 μl/l 13.3±6 c 12.9±3 cd 12±4 cd Artemisa persica 500 μl/l 15.9±7 bc 14.8±2 bc 12±5 cd Artemisa persica 1000 μl/l 13.4±4 c 12.6±5 cd 11±5 cd Rosmarinus officinalis 500μl/l 17.5±4 a 17±5 b 13±3 c Rosmarinus officinalis 1000 μl/l 15.8±3 bc 14.5±4 bc 11±c 2 d Carotenoids Control 0.85±0.03kn 1.64±0.06 b 1.94±0.03 a Mancozeb 0.5 mg/l 0.69±0.04 m-o 1.57±0.03 b 1.43±0.03 b-e Mancozeb 1 mg/l 0.74±0.03 m-o 1.52±0.04 bc 1.48±0.08 b-d Thymus vulgaris 500 μl/l 0.74±0.07 m-o 1.12±0.05 g-j 1.15±0.03 f-i Thymus vulgaris 1000 μl/l 0.55±0.03 o 1.28±0.08 c-g 1.91±0.06 a Salvia mirzayanii 500 μl/l 0.69±0.08 m-o 1.26±0.05 d-h 1.39±0.07 b-f Salvia mirzayanii 1000 μl/l 0.7±0.06 m-o 1.1±0.06 g-k 1.53±0.03 bc Artemisa persica 500 μl/l 0.56±0.05 o 0.85±0.04 l-n 1.31±0.04 c-g Artemisa persica 1000 μl/l 0.6±0.06 o 0.87±0.08 j-m 1.22±0.03 e-h Rosmarinus officinalis 500 μl/l 0.52±0.04 o 0.93±0.05 i-m 1.09±0.04 g-l Rosmarinus officinalis 1000 μl/l 0.5±0.07 o 1.01±0.03 h-l 1.16±0.05 f-i Total chlorophyll Control 2.72±0.2 f 1.19±0.02 gh 0.03±0.002 i Mancozeb 0.5 mg/l 3.98±0.4 e 1.31±0.02 g 0.53±0.03 h Mancozeb 1 mg/l 3.44±0.2 f 1.72±0.02 g 0.31±0.02 h Thymus vulgaris 500 μl/l 4.86±0.5 de 2.5±0. 6 f 0.11±0.01 h Thymus vulgaris 1000 μl/l 4.33±0.2 de 2.66±0. 5 f 0.11±0.02 h Salvia mirzayanii 500 μl/l 5.43±0.3 d 3.55±0.2 f 0.16±0.01 h Salvia mirzayanii 1000 μl/l 7.62±0.2 bc 3.47±0.2 f 0.15±0.02 h Artemisa persica 500 μl/l 9.61±0.2 a 2.58±0.7 f 0.16±0.03 h Artemisa persica 1000 μl/l 8.38±0.3 b 2.35±0.5 f 0.22±0.02 h Rosmarinus officinalis 500 μl/l 8.69±0.2 ab 3.27±0.8 f 0.16±0.04 h Rosmarinus officinalis 1000 μl/l 8.19±0.4 b 3.3±0.6 f 0.32±0.5 h 396 Adv. Hort. Sci., 2018 32(3): 389-398 treatments throughout the storage (Table 2); howev- er, the highest reduction was observed in the control fruits. At the end of storage, the minimum chloro- phyll content (0.03) was recorded in the control, while the highest was found in mancozeb with 0.5 mg/l concentration. TSS, TA and pH A gradual increase in TSS percentages was deter- mined in all treatments (Table 3). Generally, the fruits treated with EOs had lower TSS percentages than the control fruits throughout the storage. However, the treated fruits did not show any signifi- cant difference in TSS, as compared with the con- trols. These results were in agreement with those in other studies (Marjanlo et al., 2009). Nevertheless, they are in discordance with Rabiei et al. (2011), who reported that thyme EOs treatment had a significant effect on the pH of apples. As shown in Table 3, TA values were also gradually decreased during storage. At the end of storage, the maximum TA values (0.27%) were observed in A. persica (1000 μl/l), while the minimum (0.10%) was in the control fruits. Among the EOs treatments, R. officinalis (1000 μl/l) resulted in the lowest acidity (0.17%), this was fol- lowed by T. vulgaris (0.16%) with 1000 μl/l concen- tration during storage; however, no significant differ- ence was observed between the treatments. These results are in agreement with those reported by Maqbool and Alderson (2010), who showed that the Table 3 - Effect of the Thymus vulgaris, Salvia mirzayanii, Artemisia persica and Rosmarinus officinalis EOs tested at 500 and 1000 μl/l on Total Soluble Solids (TSS), pH, and Titratable Acidity (TA) ± SD in mangos after 1, 2, and 3 weeks of incubation compared to ‘mancozed’ (0.5 and 1.0 mg/l) In each character, different letters indicate significant differences in mean values (p<0.05). Quality parameter Treatment Storage time (week) Week 1 Week 2 Week 3 TSS Control 8.9±3 c 9.6±3 b 9.9±2 a Mancozeb 0.5 mg/l 8.06±3 cd 9.23±5 b 9.5±3 b Mancozeb 1 mg/l 8.33±4 c 9.13±5 bc 9.5±2 b Thymus vulgaris 500 μl/l 8.96±5 c 9.63±4 b 9.7±2 ab Thymus vulgaris 1000 μl/l 8.83±4 c 9.76±4 ab 9.8±4 ab Salvia mirzayanii 500 μl/l 8.7±4 c 9.26±2 b 9.3±3 b Salvia mirzayanii 1000 μl/l 8.33±3 c 9.26±5 b 9.6±2 b Artemisa persica 500 μl/l 8.66±3 c 9.3±3 b 9.6±3 b Artemisa persica 1000 μl/l 8.1±5 cd 9.53±5 b 9.7±4 b Rosmarinus officinalis 500 μl/l 7.5±3 d 9±4 bc 9.2±2 b Rosmarinus officinalis 1000 μl/l 8.66±4 c 9.53±5 b 9.5±3 b pH Control 1.15±0.03 cd 2.89±0.2 c 4.75±1.4 a Mancozeb 0.5 mg/l 1.06±0.02 cd 2.38±0.4 c 4.3± 2 ab Mancozeb 1 mg/l 1.09±0.02 cd 2.59±0.3 c 4.3± 2 ab Thymus vulgaris 500 μl/l 1.06±0.02 cd 2.51±0.2 c 4.18± 2. 2 ab Thymus vulgaris 1000 μl/l 1.26±0.02 cd 2.54±0.3 c 4.31±1.8 ab Salvia mirzayanii 500 μl/l 0.99±0.02 d 2.74±0.2 c 4.24±1.9 ab Salvia mirzayanii 1000 μl/l 1.12±0.02 cd 2.26±0.2 c 4.42±2 ab Artemisa persica 500 μl/l 1.03±0.02 cd 2.45±0.3 c 4.48±3 ab Artemisa persica 1000 μl/l 1.08±0.02 cd 2.17±0.2 c 4.47±1.4 ab Rosmarinus officinalis 500 μl/l 1.02±0.02 cd 1.76±0.1 cd 4.36±1.2 ab Rosmarinus officinalis 1000 μl/l 1.07±0.02 cd 2.52±0. 5 c 4.56±2.1 ab TA Control 0.208±0.03 c 0.196±0.01 c 0.10±0.009 d Mancozeb 0.5 mg/l 0.254±0.02 bc 0.255±0.01 bc 0.23±0.02 bc Mancozeb 1 mg/l 0.27±0.02 bc 0.234±0.02 bc 0.23±0.02 bc Thymus vulgaris 500 μl/l 0.328±0.02 b 0.26±0.03 bc 0.23±0.02 bc Thymus vulgaris 1000 μl/l 0.308±0.01 b 0.262±0.02 bc 0.16±0.01 cd Salvia mirzayanii 500 μl/l 0.384±0.02 b 0.267±0.03 bc 0.25±0.02 bc Salvia mirzayanii 1000 μl/l 0.352±0.03 b 0.299±0.03 bc 0.24±0.02 bc Artemisa persica 500 μl/l 0.288±0.02 bc 0.277±0.02 bc 0.23±0.02 bc Artemisa persica 1000 μl/l 0.405±0.02 b 0.307±0.01 b 0.27±0.02 bc Rosmarinus officinalis 500 μl/l 0.471±0.02 a 0.302±0.02b 0.26±0.02 bc Rosmarinus officinalis 1000 μl/l 0.328±0.01 b 0.261±0.01 bc 0.17±0.01 cd Javadpour et al. - Postharvest control of Aspergillus niger in mangos 397 maximum reduction of the TA values was observed in the control fruits of bananas and papayas. Data relat- ed to the changes in the pH of fruits during storage revealed a significant increase in all treatments (Table 3). In each time, a higher pH value was found in the control groups. These results were in line with t h o s e r e p o r t e d b y D u P l o o y e t a l . ( 2 0 0 9 ) a n d Tzortzakis (2007), showing no significant differences between the pH of control and treated fruits 4. Conclusions The present study proves that the T. vulgaris, A. persica, R. officinalis and S. mirzayanii EOs could be employed under postharvest condition to control a pathogenic isolate of A. niger on mango fruits. The effectiveness of these EOs was more than mancozeb. So that, the EOs here tested could be used as a nat- ural fungicide to control an isolate of A. niger during postharvest mangos. 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