Study of vaccinium corymbosym blueberries

: characterization of extracts

Abstract

In Portugal, there has been a significant increase in blueberry production, which has led to an increasing need to find alternative markets in which to place the fruits, preferably through products with high commercial value. If one considers the consumers' preference for foods that are not only healthy but also 'health promoting' and that blueberries are perceived as being so, the incorporation of blueberries and their extracts into a product may be something that increases the product's intrinsic value and, therefore be an advantage when contemplating industrial application. The first step of this thesis aimed to select the raw matter to be used throughout the work. A study on four different cultivars and different ripening stages was carried out to select the blueberries that contained the higher levels of anthocyanins and other phenolics. The results showed that Goldtraube blueberries were the richest in anthocyanins (possessing both a higher content and a wider array of these compounds) as well as other phenolics, with later ripening stages being richer in anthocyanins. Therefore, Goldtraube blueberries collected at a later ripening stage were selected as the raw matter to be used. Afterwards, a solid-liquid extraction was optimized with the following conditions being identified as the most interesting: extraction of 10 g of blueberries with 100 mL acidified ethanol (0.01 % HC1), followed by 1 h incubation at 40 °C and a 15 min exposure to ultrasounds (35 kHz). The second stage of this thesis focused on the characterization of some of the biological potential of the extract starting with its antioxidant potential. In order to determine the extracts capacity to protect DNA from oxidative damage, an existing agarose gel electrophoresis method was improved to allow for quantitative measurement. The extract was able of reducing DNA degradation at a concentration range of 160 to 200 lig mL-1 (for an H202 and an H202/iron degradation system, respectively). Simultaneously, the extract's impact upon nosocomial agents was evaluated. Out of the eight pathogens tested the extract was only capable of inhibiting the growth of three (MSSA, MRSA and one E. coli strain) at the highest concentration of extract tested. However, when considering the antibiofilm and antiadhesive assays, the extract proved to be more effective as it was capable of inhibiting all microorganisms at some level. In an attempt to better understand the role of the different compounds in the observed activity, two research lines were explored: i) the effect of phenolic acids, present in blueberries, was evaluated, with the results demonstrating that they were only effective in inhibiting biofilm formation by Staphylococcus and had no effect upon E. coli. ii) the effect of a simulation of the extract, using the most abundant anthocyanin, against Staphylococcus (the most susceptible microorganisms to the extract). The results showed that, while the isolated compounds had some inhibitory effects, in most cases, the observed activity was significantly lower than that of the extract, hinting that synergies between the different compounds may play an important role in the observed inhibitions. As the extract demonstrated an interesting effect upon potential nosocomial agents, its potential effect upon food pathogens/contaminants as well as its effect upon possible beneficial bacteria such as probiotic microorganism, came into question. To provide an answer to this question the concentration capable of inhibiting the growth of four potential food pathogens was determined and afterwards, five different probiotics were incubated with the extract at the same concentration. No inhibition of probiotic growth was observed in the presence of extract. However, an increase in production/accumulation of organic acids (acetic, lactic and citric acids) as well as an anticipation of propionic acid production was observed. As the extract appeared to have a selective effect and the previous work demonstrated that it possessed an interesting antiadhesive effect, a study on the extract's effect upon probiotics and potential pathogens' adhesion was carried out. The results showed that the combined presence of extract and probiotic was more effective in inhibiting pathogen adhesion than the probiotic alone. The apparent synergy established between the extract and B. Bo was a particular example of a successful partnership as their combined action allowed for, in some cases, an almost complete inhibition of pathogen adhesion. Overall, the extract hereby proposed exhibited both antioxidant and antimicrobial capabilities properties that, when combined with their inherent colouring, makes it interesting to be used in food and cosmetic products.

Date of Award	7 Feb 2017
Original language	English
Awarding Institution	Universidade Católica Portuguesa
Supervisor	Maria Manuela Pintado (Supervisor), Rui M. S. C. Morais (Co-Supervisor) & Conceição Calhau (Co-Supervisor)