International Food Research Journal 23(6): 2363-2369 (December 2016)

Journal homepage: http://www.ifrj.upm.edu.my

Extraction of phenolic antioxidants from four selected seaweeds obtained

from Sabah
1
Fu, C.W.F., 2Ho, C.W., 3Yong, W.T.L., 4Abas, F., 1Tan, T.B. and 1,*Tan, C.P.
1
Department of Food Technology, Faculty of Food Science and Technology, Universiti Putra
Malaysia, 43400 UPM Serdang, Selangor, Malaysia
2
Department of Food Science and Nutrition, Faculty of Applied Sciences, UCSI University, No. 1,
Jalan Menara Gading, UCSI Heights, Cheras 56000, Kuala Lumpur, Malaysia
3
Biotechnology Research Institute, Universiti Malaysia Sabah, Jalan UMS, 88400 Kota Kinabalu,
Sabah, Malaysia
4
Department of Food Science, Faculty of Food Science and Technology, Universiti Putra
Malaysia, 43400 UPM, Serdang, Selangor, Malaysia

Article history Abstract

Received: 24 August 2015 Algal have attracted attention from biomedical scientists as they are a valuable natural
Received in revised form: source of secondary metabolites that exhibit antioxidant activities. In this study, single-
7 March 2016 factor experiments were conducted to investigate the best extraction conditions (ethanol
Accepted: 10 March 2016
concentration, solid-to-solvent ratio, extraction temperature and extraction time) in extracting
antioxidant compounds and capacities from four species of seaweeds (Sargassum polycystum,
Keywords
Eucheuma denticulatum , Kappaphycus alvarezzi variance Buaya and Kappaphycus alvarezzi
variance Giant) from Sabah. Total phenolic content (TPC) and total flavonoid content (TFC)
Seaweeds assays were used to determine the phenolic and flavonoid concentrations, respectively, while
Antioxidants Single-factor 2,2-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and 2,2-diphenyl-1-picylhydrazyl
experiments (DPPH) radical scavenging capacity assays were used to evaluate the antioxidant capacities of
Total phenolic content assay all seaweed extracts. Results showed that extraction parameters had significant effect (p < 0.05)
Total flavonoid content on the antioxidant compounds and antioxidant capacities of seaweed. Sargassum polycystum
assay portrayed the most antioxidant compounds (37.41 0.01 mg GAE/g DW and 4.54 0.02 mg
2,2-azinobis-3-
CE/g DW) and capacities (2.00 0.01 mol TEAC/g DW and 0.84 0.01 mol TEAC/g DW)
ethylbenzothiazoline-
6-sulfonic acid radical amongst four species of seaweed.
scavenging capacity assay
2,2-diphenyl-1-picylhydrazyl
radical scavenging capacity
assays All Rights Reserved

Introduction recent years due to the demand caused by abalone

farmers (Vasquez, 1999) the development of new
For centuries, seaweed has been used in the products such as organic fertilisers and use for human
preparation of salads, soups and also as low-calorie food (Alejandro et al., 2008).
foods in Asia (Jimnez-Escrig and Snchez-Muniz, In recent years, seaweed products have received
2000). Japanese are the main consumers of seaweed special attention as a source of natural antioxidants
with an average consumption of 1.6 kg (dry weight) (Lim et al., 2002) and some of them possess biological
per year per capita (Dhargalkar and Pereira, 2005). activity of potential medicinal value (Satoru et al.,
Most Europeans and Americans use processed 2003). Natural antioxidants are perceived to be safe
seaweed as additives in their food preparation by consumers because they are naturally found in
(Boukhari and Sophie, 1998). However, in India, plant materials and have been used for centuries
seaweeds are exploited mainly for the industrial (Frankel, 1996). Natural antioxidants have shown
production of phycocolloids such as agar-agar, to play a significant role in preventing a number
alginate and carrageenan; and not as cookery item of chronic diseases such as heart disease, cancer,
or for recovering beneficial biomolecules. In 1978, Alzheimers and Parkinsons diseases (Weinreb et
seaweed cultivation was introduced in Sabah and al., 2004).
had increasingly become an economically important Several researchers have reported the antioxidant
natural resource for Malaysia, particularly for Sabah. properties of both brown and red seaweeds from
The interest for seaweed escalates tremendously in across the globe (Heo et al., 2005). Some active

*Corresponding author.
Email: tancp@upm.edu.my
Tel: +603-8946-8418; Fax: +603-8942-3552
2364 Fu et al./IFRJ 23(6): 2363-2369

antioxidant compounds from marine algae were time. After the extractions, seaweed extracts were
identified as phylopheophylin in Eisenia bicyclis filtered by a glass funnel with Whatman No. 1 filter
(Cahyana, Shuto and Kinoshita, 1992), phlorotannins paper (Whatman International, England). The clear
in Sargassum kjellamanianum (Yan et al. 1996) and solution of crude extract was collected in a light-
fucoxanthinin in Hijikia fusiformis (Yan et al., 1999). protected amber bottle (50 mL) for analysis without
Furthermore, there are evidences available to show further treatment. All extractions were carried out in
the potential protective effects of seaweed against replicates.
oxidative stress in target tissues and lipid oxidation
in foods (Rajamani et al., 2011). Factor 1: Ethanol concentration
Therefore, the main objective of this study was to 10 mL of ethanol and deionised water were mixed
evaluate the effect of extraction conditions (ethanol according to the ethanol concentration set in 5 levels
concentration, solid-to-solvent ratio, extraction (0, 25, 50, 75 and 100 %, v/v), added to 1 g of each
temperature and extraction time) in extracting sample. They were then placed in a water bath shaker
antioxidant compounds as well as antioxidant at 40 C at 150 rpm for 2 h.
capacities of the four selected seaweeds (Sargassum
polycystum, Eucheuma denticulatum , Kappaphycus Factor 2: Solid-to-solvent ratio
alvarezzi variance Buaya and Kappaphycus alvarezzi An amount of ethanol and deionised water (best
variance Giant) and determine the best extraction ethanol concentration obtained from section Factor
conditions for the seaweeds. 1) was added to each sample according to the solid-
to-solvent ratio set in 5 levels (1:10, 1:15, 1:20, 1:25
Materials and Methods and 1:30, w/v). They were then placed in a water bath
shaker at 40 C at 150 rpm for 2 h.
Seaweed cultivation and collection
Sargassum polycystum (SP) and Eucheuma Factor 3: Extraction temperature
denticulatum (ED) were commercially farmed An amount of ethanol and deionised water (best
seaweed in Semporna, Sabah. They were harvested ethanol concentration obtained from section Factor
at week 6 (maturity stage). Kappaphycus alvarezii 1) were added to each sample according to the best
variance Giant (KAG) and Kappaphycus alvarezii solid-to-solvent ratio obtained from section Factor 2.
variance Buaya (KAB) were tissue cultured seaweed, They were then placed in a water bath shaker at 5
grown in Universiti Malaysia Sabah (Kota Kinabalu, different temperatures (25, 35, 45, 55 and 65C) at
Malaysia). 1.0 g of explants was cultured in-vitro 150 rpm for 2 h.
for 10 - 12 weeks, producing 50.0 g of seedlings to
acclimatize in the open sea. They were harvested at Factor 4: Extraction time
week 16 (maturity stage). Seaweeds were cleaned An amount of ethanol and deionised water (best
under running water and air-dried for 2 days. Then, ethanol concentration obtained from section Factor
they were placed in oven at 60 C until they were 1) were added to each sample according to the best
completely dry. Dried seaweed were packed and solid-to-solvent ratio obtained from section Factor
delivered to Universiti Putra Malaysia (Serdang, 2. They were then placed in a water bath shaker at
Malaysia) for future analysis. the best temperature of each sample obtained from
section Factor 3 at 150 rpm for a range of time set in
Sample preparation 5 levels (1, 2, 3, 4 and 5 h).
500 g of dried seaweeds were ground in a
laboratory grinder (Mikro-Feinmuhle-Culatti. MFC Total phenolic content (TPC) assay
grinder, Janke and Kunkel GmbH and Co., Staufen,. Total phenolic content (TPC) was determined
Germany) with a particle size of 0.08 mm. Powdered using Folin-Ciocalteu (F-C) assay (Lim et al., 2007)
samples were then vacuum-packed and stored in dark 500 L of crude extracts obtained from extraction
for further research. were added into Eppendorf falcon tubes (2 mL)
followed by 500 L of Folin-Ciocalteus reagent
Sample extraction (diluted 10 times with water). After 4 min, 400 L
1 g of powdered sample of each species of of 7.5% (w/v) sodium carbonate were added. The
seaweeds was accurately weighed into conical flasks blank was prepared by replacing 500 L of sample
(50 mL). The extraction processes were carried out with 500 L of deionised water. Subsequently, the
by varying the experiment parameters for ethanol falcon tubes were vortexed for 10 s with vortex mixer
concentration, solid-to-solvent ratio, temperature and (VTS-3000L, LMS, Japan). They were incubated
 Fu et al./IFRJ 23(6): 2363-2369 2365

in the dark environment at room temperature for 5, Secomam, France). Both the crude extracts and
2 h. Absorbance was measured against the blank negative control were carried out in triplicate. Trolox
reagent at 765 nm using UV light spectrophotometer solution was used to calibrate the standard curve.
(Model XTD 5, Secomam, France). Each extract The mean SD results of triplicate analyses were
was analyzed in triplicate and TPC were expressed expressed as mol trolox equivalent per 100 g dried
as gallic acid equivalent (GAE) in mg per 100 g dry sample (mol TEAC/100 g dried sample).
weight (DW).
ABTS radical scavenging capacity (%) = [1 (Ao /
Total flavonoid content (TFC) assay A1)] 100 % (1)
The determination of flavonoids was based on
the procedures described in the study (Ozsoy et Where Ao is A734 of the crude extract; A1 is A734 of
al., 2008) with slight modifications. 50 L of crude negative control in ethanolic ABTS solution.
extract added to 250 L of deionised water, followed
by the addition of 15 L of 5% sodium nitrite in 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical
Eppendorf falcon tubes (2 mL). After 6 min, 30 L scavenging capacity assay
of 10% aluminium chloride hexahydrate was added Antioxidant capacity was determined by
into the mixture and was allowed to stand for further measuring the scavenging activity of the radical,
5 min. Then, 100 L of 1 M sodium hydroxide and 2-diphenyl-1-picrylhydrazyl (DPPH) based on the
55 L of deionised water were added. The blank was method (Saha et al., 2004) with slight modifications.
prepared by replacing the 50 L sample with 50 L 25 L of undiluted crude extract was added to 975
of deionised water. The falcon tubes were mixed L of ethanolic DPPH in the Eppendorf falcon
thoroughly by using a vortex mixer (VTS-3000L, tubes and vortexed for 1 min using the vortex mixer
LMS, Japan) for 10 s. Then, absorbance readings (VTS-3000L, LMS, Japan). They are allowed to
were immediately taken at 510 nm using the UV stand in a dark environment at room temperature
light spectrophotometer (Model XTD 5, Secomam, for 30 min. Absorbance was measured at 517 nm
France). Each extract was analyzed in triplicate and using UV light spectrophotometer (Model XTD 5,
TFC were expressed as catechin equivalent (CE) in Secomam, France). Absolute ethanol was used as
mg per 100 g dry weight (DW). blank. Absorbance of negative control (25 L of
absolute ethanol and 975 L of ethanolic DPPH) and
2,2-azinobis-3-ethylbenzothiazoline-6-sulfonic acid absorbance of blank were also measured at 517 nm.
(ABTS) radical scavenging capacity assay Both sample and negative control were analyzed in
Antioxidant capacity was determined by triplicate. Trolox solution was used to calibrate the
measuring the scavenging activity of the radical standard curve. The mean SD results of triplicate
2,2-azinobis-3-ethylbenzothiazoline-6-sulfonic acid analyses were expressed as mol trolox equivalent
(ABTS) based on the method (Surveswaran, 2007) per 100 g dried sample (mol TEAC/100 g dried
with slight modifications. 10 mL of 7 mM ABTS sample). The capability to scavenge the DPPH
solution and 10 mL of 2.45 mM potassium persulfate radicals was calculated by using the equation below.
(K2S2O8) solution were transferred into a 100 mL
light protected amber bottle. The solution were DPPH radical scavenging capacity (%) = [1 (Ao
mixed by vortex mixer (VTS-3000L, LMS, Japan) / A1)] 100 % (2)
for 10 s and allowed to stand in a dark environment
at room temperature for 16 h to give a dark blue Where Ac is A517 of the crude extract; A1 is A517 of
solution. This solution was diluted with 95% ethanol negative control in ethanolic DPPH solution.
until the absorbance was equilibrated to 0.7 ( 0.02)
at 734 nm. 975 L ABTS solution with equilibrated Statistical analysis
absorbance of 0.7 0.02 was added to 25 L of the The experimental results were analyzed with
undiluted extract in an Eppendorf falcon tube (2 Microsoft Office Excel 2007 (version 12.0, Microsoft
mL). Negative control was prepared by replacing Corp., USA) and Minitab statistical software (Version
25 L of undiluted crude extract with 25 L of 95% 16, Minitab Inc., USA). Every measurement of each
ethanol whereas blank was prepared by using 95% assay was performed in triplicate, and every sample
ethanol solely. The reaction was allowed to occur was duplicated. All values were expressed as the
at room temperature for 6 min and the absorbance means standard errors (SE) of six measurements
at 734 nm was immediately recorded against blank (n=6) and the calculations were performed using
using the UV light spectrophotometer (Model XTD Microsoft Office Excel 2007. One-way analysis
2366 Fu et al./IFRJ 23(6): 2363-2369

of variance (ANOVA) with Tukeys test was used

to determine the significant differences (p < 0.05)
between the means.

Results

From Figure 1, it could be seen that the amount

of phenolic compounds increased as the ethanol
concentration increased until a peak was reached,
and then it decreased slightly. However, the highest
antioxidant content from each species was obtained
with different ethanol concentrations. SP, KAB and
KAG achieved a maximum TPC value of 23.58 mg
GAE/100 g DW, 23.65 mg GAE/100 g DW and 18.48
mg GAE/100 g DW at a 50% ethanol concentration,
Figure 1. Effects of ethanol concentration towards (a)
respectively; ED achieved a maximum of 10.08 mg
TPC, (b) TFC, (c) ABTS and (d) DPPH of Sargassum
GAE/100 g DW at a 75% ethanol concentration. The polycystum (SP), Kappaphycus alvarezzi variance Buaya
trend for the TFC value is about the same as for TPC; (KAB), Kappaphycus alvarezzi variance Giant (KAG) and
it increased as the ethanol concentration increased, Eucheuma denticulatum (ED)
and then decreased after a peak was reached. It is
obvious that flavonoids in KAG were significantly
higher than in the other species (3.1 mg CE/g DW).
Antioxidant capacities of all seaweeds species were
significantly affected by the ethanol concentration
as shown in Figure 1. The trend exhibited by both
assays agrees well with the TPC and TFC results.
Figure 2 showed a significant effect (p < 0.05)
of the solid-to-solvent ratio on TPC, TFC, ABTS
and DPPH for the four seaweeds. In a preliminary
test, a ratio of 1:5 was used, but no results were
obtained. The samples absorbed the solvent and
expanded during the extraction, forming a thick and
viscous semisolid mass. This could be attributed
to insufficient solvent to penetrate the sample
and therefore, no extraction occurred. Hence, it is
concluded that solid-to-solvent ratio of 1:5 is too low Figure 2. Effects of solid-to-solvent ratio towards (a)
to extract phenolics in the samples, so this ratio was TPC, (b) TFC, (c) ABTS and (d) DPPH of Sargassum
not included in this experiment. At a solid-to-solvent polycystum (SP), Kappaphycus alvarezzi variance Buaya
ratio of 1:10, the TPC and TFC reached a maximum (KAB), Kappaphycus alvarezzi variance Giant (KAG) and
for all four seaweeds. Both TPC and TFC for the Eucheuma denticulatum (ED)
four seaweeds decreased at ratios greater than 1:10.
included in the range of extraction temperature used
According Figure 2, the radical scavenging capacities
in this study. ABTS was not significantly affected
of ABTS and DPPH were significantly affected (p <
by temperature (as shown in Figure 3); while DPPH
0.05) by the solid-to-solvent ratio. At the lower ratio
presented increasing trend and peaked at 65C.
of 1:10, both ABTS and DPPH showed significantly
From Figure 4, it is obvious that each of the
high radical scavenging capacities for all four
seaweed had a different optimum extraction time
seaweeds. This trend agreed with the results from the
for phenolic compounds. SP showed the highest
antioxidant compound assay performed earlier.
TPC (37.41 mg GAE/g DW) at 2 hours; KAB
Figure 3 showed an increasing trend for TPC and
had an optimum (34.43 mg GAE/g DW) time of 4
TFC, and reached a peak at 65C for all seaweeds.
hours; KAG showed the highest TPC value (25.4
However, a preliminary test, a temperature of 75C
mg GAE/g DW) at 5 hours, and ED peaked (12.1
was used to extract phenolics. It caused a significant
mg GAE/g DW of TPC) at 3 hours. In a preliminary
decline in both the amount of antioxidant compounds
test, we used a 6 hours extraction time for KAG. A
and the antioxidant capacity. Therefore, 75C was not
significant decrease was observed, and so 6 hours of
 Fu et al./IFRJ 23(6): 2363-2369 2367

the extraction of more water soluble polyphenols.

Studies show that an ethanol and water mixture
extracts flavonoids (Spigno et al., 2007), catechin,
rutin and quercetin (Angela and Meireles, 2008). The
ethanol concentration affects extraction significantly,
whereby low ethanol concentration would favour
impurities extraction (Chirinos et al., 2007) while
high ethanol concentration tends to extract lipid
components (Wang et al., 2008). Hence, different
samples should have their best ethanol concentration
to extract maximum phenolics. Results showed in this
experiment can be explained by the different type and
structure of phenols contained in each species (Zhang
et al., 2008). It was believed that the highly active
phenolic compounds present in SP, KAB and KAG
Figure 3. Effects of extraction temperature towards (a) were balanced between polar and non-polar because
TPC, (b) TFC, (c) ABTS and (d) DPPH of Sargassum both ABTS and DPPH reached a maximum at 50%
polycystum (SP), Kappaphycus alvarezzi variance Buaya
ethanol concentration. On the other hand, ED reached
(KAB), Kappaphycus alvarezzi variance Giant (KAG) and
a maximum at 75% ethanol concentration, which
Eucheuma denticulatum (ED)
indicated that it contains moderately polar active
phenolic compounds. SP, KAB and KAG presented
50% as the best ethanol concentration; while ED
showed 75 % as the best ethanol concentration.

Effects of solid-to-solvent ratios

Evaluating effects of solid-to-solvent ratios is
imperative in an industry viewpoint to ensure
efficient and economic phenolics extraction. In the
preliminary test, ratio of 1:5 was tested. As portrayed
in the results, a ratio of 1:10 was the best for all of the
samples. Nonetheless, when the ratio was increased,
the amount of extracted phenolics in the extract
remained the same but was diluted with the extra
solvent added. The decreases in ABTS and DPPH can
be explained by the decreased values of TPC and TFC.
Figure 4. Effects of extraction time towards (a) TPC, (b) Dilution by excessive solvent affects the antioxidant
TFC, (c) ABTS and (d) DPPH of Sargassum polycystum capacity significantly. In addition, the lesser total
(SP), Kappaphycus alvarezzi variance Buaya (KAB),
phenolic compounds present in the extract, the lower
Kappaphycus alvarezzi variance Giant (KAG) and
the antioxidant capacity it possessed. It was reported
Eucheuma denticulatum (ED)
that the antioxidant activity of a plant extract often
extraction time was not included in this experiment. originates from phenolic compounds (Amarowicz et
Figure 4 presented that the trend for the antioxidant al., 2000). A solid-to-solvent ratio of 1:10 was chosen
capacities is almost the same as that for the amount as the best condition to extract the highest amount of
of antioxidant compounds extracted. antioxidant compounds and capacity from SP, KAB,
KAG and ED.
Discussion
Effects of extraction temperature
Effects of ethanol concentrations 65C was the best extraction temperature for
The nature of the solvent used determines the all four species of seaweeds. In a preliminary test,
types of phenols extracted from the plant material 75C were tested, but a sharp decrease occurred.
(Liyana-Pathirana and Shahidi, 2005). A dual It was believed that phenolics were degraded at
solvent system is more desirable than a mono- that temperature. Increasing temperature promotes
solvent system (Wang et al., 2008) because it analyte solubility. This is mainly because incubation
creates a moderately polar medium which enhances in hot water weakens the cellular constituents of the
2368 Fu et al./IFRJ 23(6): 2363-2369

Table 1. Best extraction condition (ethanol concentration, solid-to-solvent ratio,

extraction temperature and time) for 4 selected seaweeds

seaweeds, releasing more bound phenols into the after a particular time (Pinelo et al., 2006). Results
solvent (Spigno et al., 2007). Furthermore, a higher of antioxidant compounds and antioxidant capacities
extraction temperature reduces solvent viscosity and were compatible; this is likely because the phenolic
surface tension, thus, accelerating the extraction compounds extracted are active. Prolonged extraction
process and increasing the diffusion coefficient. time leads to the decomposition of active compounds
Additionally, studies showed that the rate of recovery (Liyana-Pathirana and Shahidi, 2005) due to long
of thermally stable antioxidants at an elevated exposure to the environment (i.e., temperature, light
temperature (up to 65C) was greater than the rate and oxygen) (Lafka, Sinanoglou and Lazos, 2007),
of decomposition of less soluble phenolics (Liyana- increasing the chance that the phenolic compounds
Pathirana and Shahidi, 2005). Despite an increasing become oxidized, which decreases the antioxidant
in the amount of antioxidant compounds extracted capacity. Furthermore, undesirable reactions such as
at a higher temperature, Figure 3 shows that ABTS enzymatic oxidation and polymerization might be
does not significantly change during extraction favoured by the extended extraction time (Biesaga
at high temperature. This is likely because the and Pyrzynska, 2013). The best extraction times were
bioavailability of phenolics or bioactive compounds set as follows: for SP (2 h), KAB (4 h), KAG (5 h)
was negatively affected by the relatively high and ED (3 h).
temperature. Nevertheless, the antioxidant capacity
of the sample could experience thermal destruction Conclusions
(Spigno et al., 2007), in turn reducing its antioxidant
activities, therefore resulting in almost no change The best extraction conditions (ethanol
in ABTS. Nevertheless, DPPH was significantly concentration, solid-to-solvent ratio, extraction
increased for all four seaweeds. DPPH is known to temperature and time) for four selected seaweeds were
react well with low molecular weight compounds successfully identified by single-factor experiments.
(Paixo, 2007). Furthermore, DPPH radicals reacted However, Sargassum polycystum possessed the most
with phenolic compounds even at high temperatures. antioxidant compounds and capacities amongst the
It is concluded that the four seaweeds contain a high four species. The results obtained from this study are
proportion of heat-resistant low molecular weight important in the development of industrial extraction
active phenolic compounds. processes of phenols from seaweed. Purification and
identification of the phenolic components in seaweed
Effects of extraction time can be done to identify phenolic compounds that are
Extraction time is determined purely by the responsible for the antioxidant characteristics.
molecular size, quantity and chemical structure of
the phenolic compounds in the sample (Chirinos et Acknowledgment
al., 2007). Different species of seaweeds contain a Financial support of this work by Universiti
different composition of bioactive compounds as Putra Malaysia through research funding is gratefully
well as of phenolic compounds. For instance, some acknowledged.
phenols require a longer extraction time because
the phenols are bound with fiber (Benjama and References
Masniyom, 2011). Phenols that are tightly bound to
cell-wall polymers may need a longer extraction time Alejandro, H.B., Daniel, A.V., Mara, C.H.G. and Pirjo, H.
compared than free phenolic compounds. Therefore, 2008. Opportunities and challenges for the development
a different optimum extraction time resulted for each of an integrated seaweed-based aquaculture activity in
Chile: determining the physiological capabilities of
of the four seaweeds. The time required for the solvent
Macrocystis and Gracilaria as biofilters. Journal of
to interact with the solid material is critical for solute Applied Phycology 20: 571-557.
recovery. According to Ficks second law of diffusion, Amarowic,z R., Naczk, M. and Shahidi, F. 2000.
final equilibrium is attained between the solution Antioxidant activity of various fractions of non-tannin
concentration in the solid matrix and the solvent phenolics of canola hulls. Journal of Agriculture and
 Fu et al./IFRJ 23(6): 2363-2369 2369

Food Chemistry 48: 2755-2759. Pinelo, M., Arnous, A. and Meyer, A.S. 2006. Upgrading of
Angela, A. and Meireles, A. 2008. Extracting Bioactive grape skins: Significance of plant cell-wall structural
Compounds for Food Products: Theory and components and extraction techniques for phenol
Applications. CRC Press. release. Trends in Food Science and Technology 17:
Benjama, O. and Masniyom, P. 2011. Nutritional 579-590.
composition and physicochemical properties of two Rajamani, K., Manivasagam, T., Anantharaman, P.,
green seaweeds (Ulva pertusa and U. intestinalis) from Balasubramanian, T. and Somasundaram, S.T. 2011.
the Pattani Bay in Southern Thailand. Songklanakarin Chemopreventive effect of Padina boergesenii
Journal Science Technology 33(5): 575-583. extractson ferric nitrilotriacetate (Fe-NTA)-induced
Biesaga, M. and Pyrzynska, K, 2013. Stability of bioactive oxidativedamage in Wistar rats. Journal of Applied
polyphenols from honey during different extraction Phycology 23: 257-263.
methods. Food Chemistry 136: 46-54. Saha, K., Lajis, N.H., Israf, D.A., Hamzah, A.S., Khozirah,
Boukhari and Sophie, 1998. Anyone for algae? UNESCO S. and Khamis, S. 2004. Evaluation of antioxidant and
Courier 51(7/8): 31-32. nitric oxide inhibitory activities of selected Malaysian
Cahyana, A.H., Shut,o Y. and Kinoshita, Y. 1992. medicinal plants. Journal of Ethnopharmacology 92:
Pyropheophytin as an antioxidative substance from the 263-267.
marine algae, Arame (Eisenia bicyclis). Bioscience, Satoru, K., Noboru, T., Hiroo, N., Shinji, S. and Hiroshi,
Biotechnology and Biochemistry 56: 1533-1535. S. 2003. Oversulfation of fucoidan enhances its anti-
Chirinos, R., Rogez, H., Campos, D., Pedreschi, R. angiogenic and antitumor activities. Biochemistry
and Larondell, Y. 2007. Optimization of extraction Phamacology 65: 173-179.
conditions of antioxidant phenolic compounds from Spigno, G., Tramelli, L. and DeFaveri, D.M. 2007.
mashua (Tropaeolum tuberosum Ruiz and Pavon) Effects of extraction time, temperature and solvent on
tubers. Separation and Purification Technology 55: concentration and antioxidant activity of grape marc
217-225. phenolics. Journal of Food Engineering 81: 200-208.
Dhargalkar, V.K. and Pereira, N. 2005. Seaweed: promising Surveswaran, S., Cai, Y., Corke, H. and Sun, M. 2007.
plant of the Millennium. Science and Culture 71: 60- Systematic evaluation of natural phenolic antioxidants
66. from 133 Indian medicinal plants. Food Chemistry
Frankel, E.N. 1996. Antioxidants in lipid foods and their 102: 938-953.
impact on food quality. Food Chemistry 57: 51-55. Vasquez, J.A. 1999. The effect of harvesting of brown
Heo, S.J., Park, E.J., Lee, K.W. and Jeon, Y.J. 2005. seaweeds: a social, ecological and economical
Antioxidant activities of enzymatic extracts from important resource. World Aquaculture 30: 19-22.
brown seaweeds. Bioresource Technology 96: 1613- Wang, J., Sun, B., Cao, Y., Tian, Y. and Li, X. 2008.
1623. Optimisation of ultrasound-assisted extraction
Jimnez-Escrig, A. and Snchez-Muniz, F. 2000. Dietary of phenolic compounds from wheat bran. Food
fibre from edible seaweeds: chemical structure, Chemistry 106: 804-810.
physicochemical properties and effects on cholesterol Weinreb, O., Mandel, S., Amit, T. and Youdim, M. 2004.
metabolism. Nutrition Research 20: 585-598. Neurological mechanisms of green tea polyphenols
Lafka, T.I., Sinanoglou, V. and Lazos, E.S. 2007. On in Alzheimers and Parkinsons diseases. Journal of
the extraction and antioxidant activity of phenolic Nutrition and Biochemistry 15: 506-516.
componds from winery wastes. Food Chemistry 104: Yan, X.J., Chuda, Y., Suzuki, M. and Nagata, T.
1206-1214. 1999. Fucoxanthin as the major antioxidant in
Lim, S.N., Cheung, P.C.K., Ooi, V.E.C. and Ang, P.O. Hijikia fusiformis. Bioscience, Biotechnology and
2002. Evaluation of antioxidative activity of extracts Agrochemistry 63: 605-607.
from a brown seaweed, Sargassum siliquastrum. Yan, X.J., Li, X.C., Zhou, C.X. and Fan, X., 1996.
Journal of Agricultural and Food Chemistry 50: 3862- Prevention of fish oil rancidity by phlorotannins
3866. from Sargassum kjellmanianum. Journal of Applied
Lim, Y.Y., Lim, T.T. and Tee, J.J. 2007. Antioxidant Phycology 8: 201-203.
properties of several tropical fruits: A comparative Zhang, Y., Li, S. and Wu, X. 2008. Pressurized liquid
study. Food Chemistry 103: 1003-1008. extraction of flavonoids from Houttuynia cordata
Liyana-Pathirana, C. and Shahidi, F. 2005. Optimization Thunb. Separation and Purification Technology 58:
of extraction of phenolic compounds from wheat 305 - 310.
using response surface methodology. Food Chemistry
93: 47-56.
Ozsoy, N., Can, A., Yanardag, R. and Akev, N. 2008.
Antioxidant activity of Smilax excelsa L. leaf extracts.
Food Chemistry 110: 571-583.
Paixo, N., Perestrelo, R., Marques, J.C. and Camara, J.S.
2007. Relationship between antioxidant capacity and
total henolic content of red, ros and white wines.
Food Chemistry 105: 204-214.