Investigation of antifungal activity of nanoencapsulation of Thyme vulgaris essential oil against botrytis cinerea in red shahroodi grape (Vitis vinifera CV. Red)

Fresh grapes (Vitis vinifera L.) show severe lesions at the post-harvest stage and during the storage period. Decreasing the quality of grapes in the post-harvest stage limits its consumption and commercialization. Some methods such as using bio-control agents, natural antimicrobials, physical methods, disinfectants agents ,GRAS, (ozone, ethanol, acetic acid) have been used to control Botrytis cinerea after harvest. Encapsulation of the essential oils will increase their ability by increasing the effectiveness of the essential oils. Currently, chitosan has been interested for encapsulating bioactive compounds, as they are generally known to be safe and possess superior biological properties such as biodegradability, biocompatibility and non-toxicity. The aim of this study was to encapsulate thyme essential oil in chitosan nanogels to enhance and maintain its antifungal effect against B. cinerea in vitro and in vitro on Shahroodi red grape.

For the purpose of this study, chitosan nanogels were first prepared and the infrared spectrum of chitosan-meric acid nanogels was measured using FT-IR430 infrared spectrophotometer at 20 ° C. The morphology of chitosan-meric acid nanogels and encapsulated essential oil was analyzed by SEM. Release test was then performed to determine the release rate of thyme essential oil encapsulated in chitosan-meric acid nanogels. Effect of chitosan-meristic acid nanogels at three levels of 0, 150 and 300 µl/L, pure thyme essential oil and thyme essential oil encapsulated in chitosan-meristic acid nanogels at three levels of 0, 75 and 150 µl / L in vitro and on the shelf life of grape fruits was studied under modified atmospheric conditions during 72 days storage at 2 to 4 °C. During storage, some traits such as firmness by manual penetrometer, electrical conductivity of fruit tissue, some components of fruit skin color and soluble solids were assessed by a refrectometer.

 Infrared spectroscopy (FTIR) results confirmed the successful coupling between chitosan amin groups and carboxylic acid-meristic acid groups and scanning electron microscopy image showed that the particle size of chitosan-meric acid nanogel was than 100 nm. The Comparison of the particle size in the present study with the previous studies on the size of chitosan-meric acid nanogels was smaller and more uniform. These differences could be related to several reasons, transform the long chitosan chain into smaller fragments by initial sonication, the important role of ultrasound in the reduction of the particle size and passing the nanogels through the filter. Release test showed that the diffusion of thyme essential oil from chitosan-meric acid nanogels has a two-step process. The chitosan-meric acid nanogels prepared in this study have hydrophilic (chitosan polymer) and hydrophilic (meristic acid fatty acid chain) regions, which led to the gradual release of thyme essential oil due to their hydrophobic nature. The results of the infected packaged berries confirmed that with increasing concentration of thyme essential oil, the antifungal effect was also increased. Evaluation of the effects of essential oil and nanogels - essential oils on grape berries infected with pathogenic spores showed the highest number of spores in control treatment (10.125 × 105 per ml) and the lowest number in NE2 (1.375 × 105 per ml) were observed. Coating treatments of chitosan-meric acid nanogel and thyme essential oil encapsulated in chitosan-meric acid nanogels showed better results at higher concentrations, but in the case of essential oil, lower spores were observed. The lowest electrical conductivity and discoloration, the highest L * component and chroma index were observed in thyme essential oil treatment with concentration of 75 µl/l. The results showed that the lower concentrations of the essential oil in the control of botrytis cinerea was better than the pure essential oils, whereas in the experiments on the storage of grape fruit, the treatment of thyme essential oil with a concentration of 75 µl/l It showed the most favorable result. Based on the results of the present study, the effect of essential oil nano-gel and essential oil alone on the shelf life after harvesting of grapes packed with polyethylene film confirmed that the essential oil performance was better than the essential oil nanogel. The lower performance of the essential oil nanogel than that of the essential oil alone can be due to inhibition of the essential oil diffusion through the packaging film while, the encapsulated essential oil release slower than the essential oil alone, it is likely to have an effect longer than expected in this study for the storage period of the grape. Considering this case and the antimicrobial capability of essential oil nanogel, it is necessary to investigate the effect of essential oil nanogel compared to essential oil on longevity of grapes for a longer of storage period.