Enzyme-enhanced extraction of antioxidant ingredients from red algae Palmaria palmata

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Abstract

The effect of various protease and carbohydrase treatments on the extraction of polyphenols and other antioxidant ingredients from the red algae Palmaria palmata (dulse) was investigated. In addition, the relative contribution of different fractions to the overall antioxidant capacity of the hydrolysate was evaluated. Considerable differences were observed both in total phenolic content (TPC) and antioxidant activities of the hydrolysates evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, oxygen radical absorbance capacity (ORAC) and ferrous ion-chelating ability assays. All the proteases tested had significant enhancing effect on the extraction of polyphenols and other active components compared to carbohydrases and cold water extraction (control). The Umamizyme extract had the highest TPC and consequently exhibited the strongest scavenging capacity against DPPH and peroxyl radicals. Further fractionation of the Umamizyme extract revealed that the crude polyphenol fraction possessed the highest peroxyl radical scavenging activity, whereas the crude polysaccharide fraction was more effective for chelating ferrous ions. The data from this study suggest the potential of protease treatment to improve value-added utilization of dulse extracts as antioxidants in functional foods and nutraceuticals.

Introduction

Marine macroalgae have a long history of use as food and folk medicine in Asia, but are much less common in Europe and North America, as a part of the diet. Only a few species are harvested for direct human consumption in limited coastal regions. Palmaria palmata (dulse) is one of the most widely distributed edible seaweeds in Iceland and was an important source of food supply when food variety was scarce in earlier times according to documentaries (Kristjánsson, 1980). Nowadays, dulse is mainly consumed as snacks and sold in health stores, but there is growing awareness that it should be utilized more as a source of important nutrients and functional ingredients (Plaza, Cifuentes, & Ibanez, 2008). It has been reported that dulse has the second highest protein content of all common seaweeds, after Porphyra tenera (Nori) (Galland-Irmouli et al., 1999). The essential amino acids (EAAs) can represent between 26 and 50% of the total amino acids (Galland-Irmouli et al., 1999, Morgan et al., 1980). Dulse is also high in iron and many other easily assimilated minerals and trace elements as well as a good source of dietary fibers and vitamins (MacArtain et al., 2007, Morgan et al., 1980).

In recent years, there is increasing interest in the search for naturally occurring compounds with antioxidant activity as alternatives to synthetic products. Aquatic plants are also a rich source of natural antioxidants (Duan, Zhang, Li, & Wang, 2006). Previous studies have shown that P. palmata contains several classes of hydrophilic antioxidant components including l-ascorbic acid, glutathione (GSH), polyphenols as well as MAAs (Yuan et al., 2005b, Yuan et al., 2009). However, the high degree of structural complexity and rigidity of the algal cell wall is a major obstacle to the efficient extraction of the intracellular bioactive constituents (Deniaud, Quemener, Fleurence, & Lahaye, 2003). Conventional water and solvent extraction have several drawbacks such as low selectivity, low extraction efficiency, solvent residue and environmental pollution (Herrero, Cifuentes, & Ibanez, 2006). As an alternative technology, enzyme-assisted extraction has attracted considerable interest. The application of hydrolytic enzymes has shown a great potential to improve the extraction yield, enhance the release of secondary plant metabolites and preserve the bioactive properties of the extracts (Li, Smith, & Hossain, 2006). Enzymatic extraction has also been reported to increase the extractability of bioactive compounds from several brown algae (Heo et al., 2005, Siriwardhana et al., 2008). These cell wall-degrading enzymes help to weaken or disrupt the cell wall structure, break down complex interior storage materials, thereby facilitating the release of the intracellular bioactive compounds from algal biomass. Furthermore, the hydrolytic breakdown of high-molecular-weight (HMW) polysaccharides and proteins may contribute to enhanced antioxidant activities (Siriwardhana et al., 2008).

P. palmata is one of the most abundant seaweed species along the coastline of Iceland, which has so far not been explored as a source of valuable antioxidant ingredients. Therefore, the aim of the present study was to investigate the efficiency of enzyme-assisted extraction of polyphenols and other antioxidant components from P. palmata. The potential of enzyme treatments to improve the antioxidant activity of the extract were assessed by three in vitro assays based upon different reaction mechanisms. The antioxidant tests were also carried out on three subfractions from Umamizyme extract, namely crude polyphenol, crude polysaccharide and an LMW aqueous fraction to estimate their relative contribution to the overall antioxidant capacity of the hydrolysate.

Section snippets

Algal materials

The red algae (Rhodophyta) P. palmata (Linnaeus) Kuntze, was collected in Hvassahraun coastal area nearby Hafnarfjordur, southwestern Iceland on October 12th, 2007. The samples were carefully rinsed with tap water. The small cut pieces (about 2 cm × 2 cm) were freeze-dried, ground into powder and passed through a 1.0 mm sieve. The ground powders were stored in tightly sealed polystyrene containers at −20 °C prior to extraction.

Chemicals

Fluorescein sodium salt (FL),

Proximate composition of P. palmata

Proximate composition analyses showed that the dried P. palmata sample contained approximately 4.1 ± 0.4% moisture, 61.5 ± 2.4% carbohydrate, 20.5 ± 0.4% crude protein, 0.4 ± 0.4% crude fat and 13.5 ± 0.5% ash. The crude protein content was within the range for P. palmata (8–35%) as reported earlier by Morgan et al. (1980) and Galland-Irmouli et al. (1999). Significant seasonal fluctuations of protein content in P. palmata were previously reported by Galland-Irmouli et al. (1999) and Fleurence

Conclusions

The results of the present study showed that enzyme-assisted extraction was effective in enhancing the recovery of polyphenols and other hydrophilic antioxidant compounds from P. palmata. Proteases were more effective than carbohydrases and Umamizyme extract exhibited the greatest scavenging activity against DPPH and peroxyl radicals. The use of different in vitro antioxidant tests verified the complexity of the antioxidant effects of the seaweed hydrolysates. Crude polyphenols contributed

Acknowledgement

This work was financed by AVS R&D Fund of Ministry of Fisheries in Iceland and Icelandic Research Fund and in collaboration with Hollusta úr hafinu ehf. The author Tao Wang would like to thank United Nations University-Fisheries Training Programme for a PhD scholarship granted.

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