ESTiG - Dissertações de Mestrado Alunos
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- Recovery of added value molecules from bio-residues: aesculus hippocastanum L. as a source of saponins and sterolsPublication . Abismail, Abderrahmane; Reis, Filipa S.; Barros, Lillian; Mokkadem, YassinIt is estimated that about 80% of the world population living in developing countries depends on herbal treatments as primary health and traditional medicine source. Nowadays, even in the most developed countries, plants continue to be of great importance both as sources of nutrients and bioactive compounds. These compounds can exert beneficial functions in our body and are often incorporated into nutraceutical/pharmaceutical formulations. However, and regarding the use of wild species, it is essential to consider that they are a finite natural resource, and their excessive harvest can have negative impacts on ecosystems. In this way and promoting concepts that are currently fundamental in developing technologies and products, namely the circular economy and sustainability, bio-residues have been identified as an excellent source of interesting bioactive compounds. Aesculus hippocastanum L., commonly known as horse-chestnut, is a large tree native to southeastern Europe and widely used as an ornamental and landscape plant in parks and gardens worldwide. Regarding its fruits/seeds, most of them end up in landfills, making these plants a source of bio-residues with no economic value. Horse chestnut seeds are not edible due to the bitter taste conferred by the bioactive compounds present, namely pentacyclic triterpene saponins. Saponins are compounds of high interest since they have multiple bioactivities, being recognized for their anti-inflammatory, hypocholesterolemic, immunomodulatory and even antitumor potential. Although there is an increasingly widespread tendency to adopt environmentally-friend extraction methodologies to obtain interesting compounds, the fact is that conventional technologies are still widely used. Frequently because of their practicality and lower cost, as well as because extraction yields are sometimes lower with green methodologies. According to the abovementioned, the present work aimed to investigate the real difference between conventional techniques (Soxhlet) and more innovative and greener techniques (ultrasound-assisted extraction) for extracting saponins. With this, it is intended to implement the extraction these valuable compounds, which is not performed in our laboratory and is of great interest given their bioactive potential. In addition, since the unsaponifiable fraction of chestnuts seeds can be a source of phytosterols, the chemical characterization of this fraction was also the subject of study. A. hippocastanum seeds (pulp, bark, and skin) were extracted by Soxhlet and ultrasound-assisted extraction (UAE), and to optimise the extraction of saponins from the seed samples the response surface methodology (RSM) was used. The saponins profile of the samples were initially characterised by ultrafast liquid chromatography coupled to a photodiode array (UFLC-PDA), using aescin as standard. Then, and after obtaining the optimal extraction points by RSM, the bioactivity of the A. hippocastanum pulp, skin and bark was evaluated. It was possible to detect saponins, namely aescin, on the samples obtained from both methods, and it was possible to identify the different aescin isoforms all along. For the optimisation, time, solvent/solid ratio, and concentration of solvents (ethanol and methanol) were tested in Soxhlet extraction; for extraction by UAE, time, ultrasonic power, and concentration of solvents (ethanol, and methanol) were tested. The polynomial models were fitted to the experimental data, statistically validated, and used to determine the optimal extraction conditions. Generally, the extraction of the target compounds was greatly affected by all the independent variables. After obtaining the optimal points for the extraction of saponins, both Soxhlet and UAE were performed for the pulp, skin, and bark of A. hippocastanum. The bioactivity of these extracts was evaluated, namely the antimicrobial activity against human pathogens, cytotoxicity in human tumor cells and non-tumor cells, and anti-inflammatory potential. Regarding the antimicrobial properties, gram-positive bacteria, and Aspergillus brasiliensis seemed to be more sensitive pathogens to this type of extracts. However, since no relationship between the type of extraction or solvents was observed, this could be due to possible synergistic effects between saponins and other bioactive compounds. Concerning the cytotoxic and anti-inflammatory activities, generally, the most promising values were obtained for the bark. These results can prove the synergistic effects previously inferred. Overall, this work was based on the recovery of added-value molecules from bio-residues currently without any application or economic value, promoting the recycling of materials and sustainable development, in addition to being a comprehensive study of the methodologies for extracting bioactive compounds of interest. The results of this study could be exploited by companies interested particularly in saponin-rich natural ingredients, such as the pharmaceutical, cosmetic, and food industries.