Ciencias Exactas y Ciencias de la Salud

In the past years, abiotic stresses such as saline soaking and the exogenous application of phytohormones such as methyl jasmonate (MeJA) have been studied as an effective tool to improve the accumulation of bioactive phytochemicals in fresh products. While the application of selenium (Se), sulfur (S), and methyl jasmonate (MeJA) has been studied in mature crops of Brassicaceae family such as kale, information about their effects on carotenoid and phenolic compound content in sprouts is scarce. Moreover, bioactivity studies of kale extracts treated with Se, S, and MeJA have not been performed. Therefore, the present thesis evaluated, the effect of Se, S, and MeJA on the accumulation of glucosinolates, carotenoids and phenolic compounds in kale sprouts. On the other hand, the individual and combined effects of Se, S, and MeJA stress over the anti-inflammatory and antioxidant activity in an in vitro model of Inflammatory bowel disease (IBD). The present study was divided into three phases. In the first phase, the effects of Se, S, and MeJA on lutein, glucosinolate, and phenolic accumulation were assessed in kale sprouts. Red Russian and Dwarf Green kale were chamber-grown using different treatment concentrations of Se (10, 20, 40 mg/L), S (30, 60, 120 mg/L), and MeJA (25, 50, 100 µM). Sprouts were harvested every 24 h for 7 days to identify and quantify phytochemicals. The highest lutein accumulation occurred 7 days after S 120 mg/L (178%) and Se 40 mg/L (199%) treatments in Red Russian and Dwarf Green kale sprouts, respectively. MeJA treatment decreased the level of most phenolic levels, except for kaempferol and quercetin, where increases were higher than 70% for both varieties when treated with MeJA 25 µM. The most effective treatment for glucosinolate accumulation was S 120 mg/L in the Red Russian kale variety at 7 days of germination, increasing glucoraphanin (262.4%), glucoerucin (510.8%), 4-methoxy-glucobrassicin (430.7%), and glucoiberin (1150%). Results show that kales treated with Se, S, and MeJA could be used as a functional food for fresh consumption or as raw materials for different industrial applications. In the second phase of this study, it was assessed the individual and combined effects of Se, S, and MeJA on the content of carotenoids, phenolic compounds, and glucosinolates present in undigested extracts and intestinal-digested fractions of 7-day-old Red Russian kale sprouts. Also, the effect of undigested extract and digested kale on cellular antioxidant activity (CAA) and nitric oxide (NOx) production was evaluated. Kale sprouts treated with Se showed higher levels of total glucosinolates (53.2%) and lutein (149.3%) in the ethanolic extract, whereas phenolic content remained without significant changes. After in vitro digestion, a significant liberation of lutein was observed in all samples of kale, mainly the Se-enriched fraction (210.3%). On the other hand, phenolic compounds and glucosinolates suffered significant losses. Abiotic stress did not cause a significant change in phenolic content compared to control sprouts; however, Se treatment significantly preserved the concentration of glucoraphanin (70.8%%), glucobrassicin (58.2%), 4-methoxy-glucobrassicin (41.6%) and total glucosinolate content (63.2%) in kale compared with control sprouts. In correlation, the maximum percentage of CAA and NOx was achieved in the undigested extract and intestinal digested fraction from kale sprouts treated with selenium. Due to the best results of phases 1 and 2 were obtained with Se (40 mg/L) and S (120 mg/L) treatments, they were selected for the third phase of this research. In this study, phytochemical extracts from 7-days-old Red Russian kale sprouts treated with Se and S were encapsulated with maltodextrin by spray-drying to evaluate its ability to protect phytochemicals from degradation during the digestion process. Particle morphology, encapsulation efficiency, and storage stability were characterized. Mouse macrophages (Raw 264.7) and human intestinal cells (Caco-2) were treated with the digested fraction to assess the cellular antioxidant capacity, the production of nitric oxide (NOx), and the levels of inflammatory biomarkers such as cyclooxygenase (COX)-2, tumor necrosis factor (TNF)-α, and interleukin (IL)-1β, IL-2, IL-6, and IL-10. Se (89.4%) and S (85.8%) kale sprouts showed the highest encapsulation efficiency and spherical morphology. Digestion affected the content of compounds in both encapsulated and non-encapsulated extracts. However, encapsulation promoted the control of oxidative processes during storage, and the kale extracts germinated with S and Se showed less degradation of lutein (35.6%, 28.2%), phenolic compounds (20.3%, 25.7%) and glucosinolates (15.4%, 18.9%) compared to non-encapsulated samples, respectively. The encapsulate treated with S exerted the highest antioxidant capacity (94.2%) and anti-inflammatory activity by stimulating IL-10 by 88.9% and inhibiting COX-2 and NO production by 84.1% and 92.2%, respectively. Therefore, kale sprouts with enhanced levels of phytochemicals could be used as feedstock to produce processed foods or undergo further processing for extraction and purification of high-value health-promoting biomolecules. Based on these results, general strategies to yield a high accumulation of specific phytochemicals are presented in this work. With this information, stressed kale with high concentrations of Se or S can be used as a plant model of high-value biomolecule for the treatment of inflammatory diseases.

In this work, different contributions have been made to promote vetches as a viable option for human consumption, due to their potential as source of peptides with antioxidant activity and inhibition of the angiotensin-converting enzyme, as well to explore novel technologies to reduce non-nutritional factors which could limit their use. Chapter one provides useful information about the uses of pulses as source of bioactive peptides, as well as the enzymes used. This chapter is intended as an introduction to study the possible use of vetch as source of bioactive peptides. Chapter number two was conducted to review one of the pulses whose peptides have been shown to have various biological activities: common beans. With this background, it is possible to evaluate the potential of vetch as a novel source of bioactive peptides. Chapter three collects information on one of the most frequently found activities in bioactive peptides in pulses: the inhibition of the angiotensin-converting enzyme. In this chapter, information about the angiotensin-converting enzyme is provided. Chapter four was developed with the intention of testing innovative technologies to eliminate non-nutritional factors in vetches since these can limit their human consumption. The use of instant controlled pressure-drop (DIC), is positioned as a viable option for cooking and germination in the removal of non-nutritional factors. Chapter five covers the study of the antioxidant and inhibitory activity of the angiotensin-converting enzyme of the peptides obtained from the submerged fermentation of vetch protein isolate. The fungal strains isolated and characterized for this study were obtained from vetch protein isolate. Finally, a general conclusion about the experimental work is presented in Chapter six.

Ciencias Exactas y Ciencias de la Salud

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