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Advisor(s)
Abstract(s)
Devido à crescente preocupação com as doenças cardiovasculares, estando apontadas como
uma das principais causas de morte a nível mundial, e sabendo que a hipercolesterolemia é um
fator de risco ao seu desenvolvimento, a implementação de um estilo de vida saudável é uma
das medidas principais para prevenção e redução dos níveis de colesterol, sendo por vezes
necessário a integração de alimentos funcionais a esta dieta de modo a potenciar a diminuição
do colesterol.
Neste trabalho, estudou-se a alga Fucus vesiculosus, que é um recurso abundante na costa
portuguesa, pretendendo-se avaliar as suas atividades biológicas, e para além disto tentar
entender quais os compostos bioativos responsáveis por essas atividades. De modo a enriquecer
o extrato de F. vesiculosus em compostos bioativos, fez-se um fracionamento do mesmo, através
da tecnologia de membranas usando membranas de ultrafiltração, operando em modo de
diafiltração, na qual foram obtidas diferentes frações.
Caracterizaram-se os extratos e as frações através da sua análise em cromatografia líquida
de elevada resolução (HPLC-DAD), cromatografia líquida acoplada a espectrometria de massa
(LC-MS), e foi ainda feita a determinação dos fenóis totais, dos taninos, das proteínas, dos
glúcidos e do manitol. Relativamente às atividades biológicas, foram quantificadas a capacidade
antioxidante, a inibição da acetilcolinesterase (AChE), da 3-hidroxi-3-metilglutarul coenzima A
redutase (HMGR) e determinada a citotoxicidade em células do carcinoma humano
hepatocelular (Hep-G2).
Dos extratos estudados, o que demonstrou melhores resultados em termos de atividade foi
o extrato da decocção a uma pressão de 2 bar (AD) obtendo-se uma concentração de extrato
que origina 50% de atividade antioxidante de 0.447 mg extrato seco/mL solução, uma
concentração de extrato que origina 50% inibição da AChE de 8.226 mg extrato seco/mL solução
e da HMGR de 1.590 mg extrato seco/mL solução.
Para as frações obtidas através da tecnologia de membranas, as frações que ganharam
atividade biológica em relação ao extrato inicial, foram a do permeado da membrana de maior
limite de exclusão molecular (MWCO), C35_P, e a do concentrado da membrana de menor MWCO
(C25_C). Tendo em conta as pequenas diferenças observadas entre estas duas frações, optou-se
por aprofundar o estudo na fração do permeado da membrana de maior MWCO (C35_P), para a
qual se identificaram os componentes por LC-MS verificando-se um enriquecimento da fração
em manitol e em derivados do floroglucinol, designados por florotaninos.
Em termos da citotoxicidade tanto o extrato da decocção (AD) como o permeado da
membrana de maior MWCO (C35_P) não demonstraram toxicidade para as células Hep-G2,
apresentando mais de 100% de viabilidade celular para a maioria das concentrações estudadas.
Concluiu-se que o extrato da F. vesiculosus, apresentou uma atividade antioxidante
promissora, assim como a sua possível aplicação no tratamento/prevenção em doenças
cardiovasculares e neurodegenerativas.
Due to the growing preoccupation with cardiovascular diseases, which are pointed as one of the main causes of death worldwide, and knowing that hypercholesterolemia is a risk factor, the implementation of healthy habits is one of the important behaviours that should be acquired to prevent and reduce the cholesterol levels. Sometimes it is necessary to integrate functional foods in this healthy diet in order to enhance the lowering of cholesterol. In this work, the Fucus vesiculosus alga, which is an abundant resource in the Portuguese coast, was studied in order to evaluate their biological activities, and to try to understand which are the bioactive compounds. In order to obtain a bioactive compounds enrichment, the F. vesiculosus extract was fractionated by membrane technology, using ultrafiltration membranes operating in diafiltration mode, generating different fractions with different characteristics. The characterization of the extracts and fractions was made by high performance liquid chromatography (HPLC-DAD) and liquid chromatography-mass spectrometry (LC-MS) analysis. It was also determined the content of total phenols, of tannins, of proteins, of carbohydrates and of mannitol. Regarding the biological activities, the antioxidant activity, inhibition of acetylcholinesterase (AChE) and 3-hydroxy-3-methylglutarul coenzyme A reductase (HMGR) were assessed, and it was also determined the cytotoxicity in hepatocellular carcinoma cells (Hep-G2). The extract that presented the best results in terms of bioactivity was the decoction at a pressure of 2 bar (AD) extract, with an extract concentration that originates 50% antioxidant activity of 0.447 mg dry extract/mL solution, an extract concentration that originates 50% AChE inhibition of the 8,226 mg dry extract/mL solution and the HMGR of 1,590 mg dry extract/mL solution. For the fractions obtained by the membrane technology, the ones that show a gain in biological activity, compared with the initial extract, were the permeate of the large molecular weight cut-off (MWCO) membrane, C35_P, and the concentrate of the smaller MWCO membrane, C25_C. Considering the small differences between these two fractions, it was decided to extend the study of the permeate of the large MWCO membrane (C35_P), by identifying by LC-MS the compounds that were enriched, and they were the mannitol and phloroglucinol derivates, called phlorotannins. In terms of cytotoxicity, the decoction extract (AD) and the permeate of the membrane with larger MWCO (C35_P) showed no toxicity to Hep-G2, presenting more than 100% cell viability for most of the studied concentrations. In conclusion, the extract of F. vesiculosus present a promising antioxidant activity, as well as a possible application in the treatment/prevention cardiovascular and neurodegenerative diseases.
Due to the growing preoccupation with cardiovascular diseases, which are pointed as one of the main causes of death worldwide, and knowing that hypercholesterolemia is a risk factor, the implementation of healthy habits is one of the important behaviours that should be acquired to prevent and reduce the cholesterol levels. Sometimes it is necessary to integrate functional foods in this healthy diet in order to enhance the lowering of cholesterol. In this work, the Fucus vesiculosus alga, which is an abundant resource in the Portuguese coast, was studied in order to evaluate their biological activities, and to try to understand which are the bioactive compounds. In order to obtain a bioactive compounds enrichment, the F. vesiculosus extract was fractionated by membrane technology, using ultrafiltration membranes operating in diafiltration mode, generating different fractions with different characteristics. The characterization of the extracts and fractions was made by high performance liquid chromatography (HPLC-DAD) and liquid chromatography-mass spectrometry (LC-MS) analysis. It was also determined the content of total phenols, of tannins, of proteins, of carbohydrates and of mannitol. Regarding the biological activities, the antioxidant activity, inhibition of acetylcholinesterase (AChE) and 3-hydroxy-3-methylglutarul coenzyme A reductase (HMGR) were assessed, and it was also determined the cytotoxicity in hepatocellular carcinoma cells (Hep-G2). The extract that presented the best results in terms of bioactivity was the decoction at a pressure of 2 bar (AD) extract, with an extract concentration that originates 50% antioxidant activity of 0.447 mg dry extract/mL solution, an extract concentration that originates 50% AChE inhibition of the 8,226 mg dry extract/mL solution and the HMGR of 1,590 mg dry extract/mL solution. For the fractions obtained by the membrane technology, the ones that show a gain in biological activity, compared with the initial extract, were the permeate of the large molecular weight cut-off (MWCO) membrane, C35_P, and the concentrate of the smaller MWCO membrane, C25_C. Considering the small differences between these two fractions, it was decided to extend the study of the permeate of the large MWCO membrane (C35_P), by identifying by LC-MS the compounds that were enriched, and they were the mannitol and phloroglucinol derivates, called phlorotannins. In terms of cytotoxicity, the decoction extract (AD) and the permeate of the membrane with larger MWCO (C35_P) showed no toxicity to Hep-G2, presenting more than 100% cell viability for most of the studied concentrations. In conclusion, the extract of F. vesiculosus present a promising antioxidant activity, as well as a possible application in the treatment/prevention cardiovascular and neurodegenerative diseases.
Description
Trabalho final de mestrado para obtenção do grau de mestre em Engenharia Química e Biológica
Keywords
Algas marinhas Marine algae Fucus vesiculosos Membranas Membranes Ingredientes bioactivos Bioactive ingrediente Doenças cardiovasculares Cardiovascular diseases
Citation
LOPES, Ana Margarida Marques - Recuperação de compostos bioativos da alga Fucus Vesiculosus por ultrafiltração/diafiltração e avaliação da ação biológica em doenças cardiovasculares. Lisboa: Instituto Superior de Engenharia de Lisboa, 2020. Dissertação de mestrado.
Publisher
Instituto Superior de Engenharia de Lisboa