Impregnación supercrítica de extractos naturales en la preservación de alimentos

Abstract

Active packaging is one of the most innovative alternatives for food preservation in the food industry. Supercritical Solvent Impregnation (SSI) of natural extracts is one of the most pioneering approaches among the different techniques that have been employed to develop active packaging. This thesis intends to determine the most suitable conditions to produce food-grade multilayer active PET/PP films (polyethylene terephthalate/polypropylene) impregnated with a particular olive leaf extract (OLE) that provides the packaging material with antioxidant and antimicrobial properties. The process should guarantee that the mechanical properties of the untreated polymer remain relatively unaltered, so that it can be used for food preservation purposes. To be used as an active substance, the olive leaf extract must have high bioactivity levels. For this reason, by means of the reaction with the reagent 2,2-diphenyl-1-picrylhydrazyl (DPPH assay), the most favourable conditions to obtain the extract were determined according to their antioxidant capacity. Subsequently, the extract’s antibacterial capacity against Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella enteritidis was investigated, as well as the phenolic profile analysed by means of UHPLC-ESI-QToF-MS. In order to determine the most convenient conditions for the impregnation of OLE, several parameters such as pressure (100–400 bar), temperature (35 and 55 °C), depressurization rate (1 and 100 bar/min), time (5 and 30 min, and 1, 2, 5 and 22 h), the co-solvent percentage (1–7%) (v/v) in the impregnation vessel and the ratio active substance/polymer (0.1–1) have been evaluated. After being impregnated, the active properties acquired by the films were studied and compared with those of the initial extract. The results revealed that despite the low level of impregnation obtained, if compared to the impregnation levels reached when other active substances were used, the antioxidant capacity of the films obtained is high enough as to play a role in food preservation. Regarding the chemical characterization, oleuropein and luteolin-7-glucoside were the main phenolic compounds in both the extracts and the impregnated matrices. 400 bar, 35 °C, 1 hour and 100 bar/min of depressurization were established as the best impregnation conditions for bath mode. It was observed that the matrix had a higher concentration and a wider variety of OLE compounds. Regarding co-solvent characteristics, the highest AO capacity and polyphenol loading were found in films impregnated when 7% co-solvent and a 1/1 OLE/polymer ratio were used, where c.a. 5 mg AO/g film and 60 μg total polyphenols/g film were achieved. Under such conditions, the antimicrobial capacity of impregnated OLE films against E. coli, S. aureus and S. enteritidis was evaluated. A decrease in growth inhibition with respect to that found in crude OLE was observed, except in the case of P. aeruginosa, against which inhibition increased. Although the bioactivity of the film after impregnation was evident, the integrity of the film after impregnation must be ensured. Supercritical conditions, despite using relatively low temperature, apply high pressures that may damage polymer structure. The impact that pressure, temperature, CO2 flow and the presence of OLE had on the properties of the films were evaluated for films produced either by batch mode (BM) or by semi-continuous mode (SM). The process temperature changed some of the thermal properties of the films. While at 35 °C the crystallinity of PP layer decreased, at 55 °C PET’s glass transition temperature (Tg) went up. This was due to the CO2 sorption by each polymer under those conditions. With regards to pressure, high levels affected only the PP layer, decreasing its crystallinity. Higher pressure levels favoured impregnation when BM was used, whereas different impregnation trends were found when SM was employed. Although the films’ physical properties were not compromised after their impregnation, the CO2 stream used for SM slightly weakened the impregnated films. Overall, none of the impregnation conditions in this study affected the integrity of the impregnated films. According to the results obtained, 400 bar and 35°C using BM were the most favourable conditions to produce films with a highly antioxidant properties while their structure modifications could be considered as negligible. This type of active packaging does not necessarily have to be compatible with all kinds of food. The study of migration into food simulants is used as a previous step to their implementation to actual food. This would let us to evaluate what kinds of food are more appropriate for active packaging. This study demonstrated that lipid-rich food would be the best to extract impregnated compounds, followed by food with a pH higher than 4.5, such as vegetables and some fruits. Therefore, active packaging should serve its purpose more successfully with those products, while those with pH < 4.5 would hardly extract any impregnated compounds from active films. Based on these results, and as a final step to prove the efficacy of impregnated films for food preservation, two packaging assays were carried out with real food. In the first one, the antioxidant capacity provided by the wrapping impregnated film was assessed by evaluating lipid oxidation of roasted peeled sunflower seeds during storage. The samples wrapped up with impregnated films showed significantly lower peroxide values during storage than the sunflower seeds that were unpacked or wrapped up with non-impregnated films. In the second experiment, the antimicrobial capacity of the films was proved for cherry tomato preservation. The tomatoes wrapped up with impregnated films showed lower microbial growth and exhibited a 20-day longer shelf life than tomatoes packed with non-impregnated films. It was, therefore, demonstrated that active packaging could be implemented using supercritical technology in combination with natural extracts. This provided the resulting films with their intrinsic characteristics, such as the antioxidant and antimicrobial capacities that play an active role in food preservation.