COMBINATION TREATMENT WITH NATURAL ANTIMICROBIALS

Control of postharvest decay through a combination of antimicrobial agents possessing multiple mechanisms of action is a thrust area of current research; the reason is being the inability of these molecules to serve as stand-alone treatments for achieving commercially desirable decay control. Entrapment of the agents in edible coatings is an effective way to achieve this objective. Now-a-days, the application of edible films for food packaging is becoming very popular in the food industry since they are considered to be a smart approach to reduce fruit decay in the marketing chain; thus, research on the incorporation of EOs into edible films has commenced actively. The edible films with added EOs may lead to a major advantage since the polymeric matrices of coating retard the diffusion of antimicrobial agents resulting in this method being more beneficial than spray applications (Sanchez- Gonzales et al., 2010). Moreover, they can be incorporated into the film in smaller amounts (Ponce et al., 2008) and the fruit coating can also reduce weight loss in the produce during storage and transportation. As an instance, a combination of Bacillus subtilis HFC103 strain with candelilla wax is a novel method to prolong shelf life of strawberry (Oregel-Zamudio, 2017). In a similar fashion, carvacrol and methyl cimiamate vapors incorporated to strawberry puree helped to maintain firmness and brightness of strawberries as compared with the untreated strawberries during storage at 10°C for 10 days. Natural antimicrobial vapors also increased the total soluble phenolic content and antioxidant activity of fruits at the end of the storage period (Peretto et al., 2014).

In another study, the combined efficiency of chitosan and Mentha piperita L. essential oil (MPEO), and the synergistic effect of these two compounds on controlling mango (var. Tomy Atkins) was reported. The application of coatings of 5 or 7.5 mg/mL chitosan and MPEO (0.6 or 1.25 mL/mL) mixtures resulted in synergistic interactions and decreased anthrac- nose severity in mangoes spiked with Colletotrichum strains over 15 days of storage at 25°C. The reduction in disease severity was in a level comparable with application of fungicides thiophanate-methyl (10 mg a.i./mL) and difenoconazole (0.5 mg a.i./mL) (de Oleiveira et al., 2017). In a similar study by Guerreiro et al. (2015), edible coatings based on sodium alginate (2%) and pectin enriched with citral (0.15%) and eugenol (0.1%)extended shelf-life of strawberries. Limonene and peppermint oil incorporated into acylated chitosan coupled with Tween 80 helps to create bioactive edible coatings suitable for extending the shelf life of fresh strawberries during storage (Vu et al., 2011).

Multilayer coating techniques have been used as a good alternative to overcome challenges of coating with a hydrophobic substance. Hence, more than two layers of film material are used in the layer-by-layer technique to bond physically and chemically to each other. In this method, fruits and vegetables are dipped into different solutions that contain oppositely charged polyelectrolytes, and the excess of coating material from the fruit's surface is allowed to be removed by a diying step between each dipping step. Coatings manufactured with this technique have been proven to be successful in papaya, pineapple, and cantaloupe.

Incorporation of different antimicrobial agents, namely, natamycin, grape extract, and pomegranate extract into chitosan matrix showed significantly higher antimicrobial effect against the mesophilic bacteria and yeast and mold. As a general result, antimicrobial active packaging based on the combination of chitosan coating with antimicrobial agents increased the shelf life of fresh strawberry compared with uncoated fruit (Duran et al., 2016). The nonvolatile bio actives present in plants like Moringa (Moringa oJeifera) also has high-antimicrobial activity. A study investigated the potential of edible carboxyl methylcellulose (CMC) containing moringa leaf and seed extracts as a novel postharvest treatment for maintaining storage quality and controlling diseases in “Hass” and “Gem” avocado fruit. The study also investigated the antifungal activity of methanolic and ethanolic moringa plant extracts. Briefly, 1% CMC was blended with 2% of moringa leaf (MLE) or seed extract (MSE). After the fruit was dipped in either CMC +MLE or CMC +MSE, it was stored at 5.5°C (95% RH) for 21 days. After cold storage, fruits were stored at ambient conditions (21±1°C) and 60% RH to simulate retail conditions. Postharvest quality attributes such as ethylene production, respiration rate, and fruit firmness were measured. Both coatings were also tested against postharvest fungi in reference to potato dextrose agarose. Coated fruit had a lower mass loss, ethylene production and respiration rate compared with the uncoated fruit (Tesfay et al., 2017).

Mexican oregano or cinnamon incorporated chitosan is also reported to have high antifungal effects against A. niger and P digitatum. Applications of bergamot EO to chitosan edible films were reported to affect film transparency (Sanchez-Gonzales et al., 2010), while the incorporation of thyme and clove oils into the chitosan edible films showed adequate gloss and transparency (Hosseini et al., 2009). However, the combination of EOs with chitosan films could lead to films that are less resistant to breakage and are deformable, all of which are aspects used more with cinnamon EO than thyme and clove oils. These interactions could affect the release of the added antimicrobial agent (Hosseini et al., 2009) and indirectly the antimicrobial response of the composite films. The possibility of using edible coatings with EOs added is more practicable for certain fruit, such as citrus, avocado, mango, etc., since they are normally coated with wax or something similar, while other fruit species, such as peaches, cherries, table grapes, and strawberries, are highly perishable and very prone to damage caused by handling. In this case, other systems should be applied as active packaging that has been developed for the table grapes, where eugenol or thymol is distributed on sterile gauze inside the bag, thus avoiding direct contact with the berries. Berries treated with the latter method and stored for 56 days at 1°C and 2 days at 20°C, showed a significant reduction of microbial spoilage and lower loss of quality in terms of sensory, nutritional and functional properties than the control berries (Valverde et al., 2005).

EOs were incorporated in sachets and effectively inserted into the MAP as shown by Sellamuthu et al. (2013) and Cindi et al. (2015). This system helps to trap the active components of the volatiles within the packaging during storage, transportation, or even at the display on the market shelf. An active packaging consisted of a label with cinnamon EO incorporated into it and attached to plastic packaging to extend the shelf-life of “Calanda” peach fruit. After 12 days of storage at room temperature, the reduction of infected fruit in the active label packaging was to the tune of 84.9%, compared to control. In addition, the sensory analysis of the treated peaches showed that the most positive descriptors were not significantly different from the optimum quality level before storage (Montero-Prado et al., 2011). Similarly, thyme oil sachets inserted in polyethylene terephthalate punnets and sealed with chitosan boehmite nanocomposite lidding film significantly reduced the incidence and severity of M. laxa in naturally infected “Kakawa” peaches. The chitosan boehmite nanocomposite lidding film, maintained the active components (thymol 56.43%, caryophyllene 9.47%, and b-linalool 37.6%) of thyme oil within the punnet during low temperature storage at 0.5°C and thereafter under the simulated market shelf conditions for 3 days at 15°C and at 75% RH (Cindi et al., 2015). The EOs can also be incorporated into low-density polyethylene film (LDPE) with the objective to develop an antifungal activity, for example, thyme oil (5%) was directly incorporated into LDPE used for avocado packaging (Pillai et al., 2016). In this regard, the results indicated that there was no alteration in the water vapor transmission characteristics and that it resulted in effective inhibition of C. gloeo- sporioides mycelial growth. The antifungal activity was substantial due to the release of thyme oil components, such as thymol, caryoplyllene, and carvacrol.

NANOFORMULATIONS

With the advancement in nanotechnology, nanoparticles of chitosan, silver, zinc oxide, etc., are being looked into for controlling postharvest decay in fruits. Treatments with zinc oxide in their nanoforms decreased the microbial load during fruit storage (Sogvar et al., 2016). Utility of unilamellar nanovesicles (liposome) containing D-limonene against two fruit rotting fungi (Botrytis cinerea and Penicillium chrysogenum) was evaluated on the extended shelf life and enhanced food safety of blueberries treated with D-limonene and liposomes (Umagiliyage et al., 2017). These liposomal nanoparticles were created by thin lipid film hydration followed by sonication. Application of liposomes with 50 mM concentration of limonene suppressed germination of B. cinerea conidia, and 2.2 and 2.8 log10 reductions for P chrysogenum. In vivo study of liposome coatings on blueberries also revealed protection against microbial growth even after nine weeks of storage at 4°C with 60% reduction in the treated samples at the end of 9 weeks. The results of this study can benefit the food industry through both enhancement of food safety and extending the shelf life of blueberries, further highlighting the commercial applications of liposomes

Chitosan nanoformulations are more effective against Fusarium solani and Aspergillus niger than high molecular weight chitosan used to prepare the nanoformulation. Applications of chitosan nanoformulations at 1%, 1.5%, or 2% similarly inhibited the growth of Colletotrichum musae and

C. gloeosporioides, and at 1% effectively controlled anthracnose of banana, papaya, and dragonfruit (Zahid et al., 2012). The effectiveness of the lowest dose of 1% was attributed by the authors to the use of nanoformulations. Anusuya et al. (2016) suggested that preharvest sprays of nanoformulations of hexenal assisted in retention fruits for 3-4 weeks longer in the orchard itself besides extending shelf-life under storage conditions without the loss of quality of fruits.

 
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