Molecules 25 00930 v2

Download as pdf or txt
Download as pdf or txt
You are on page 1of 29

molecules

Review
Extraction and Modification of Macroalgal
Polysaccharides for Current and
Next-Generation Applications
Madeleine Jönsson 1 , Leila Allahgholi 1 , Roya R.R. Sardari 1 , Guðmundur O. Hreggviðsson 2,3
and Eva Nordberg Karlsson 1, *
1 Biotechnology, Department of Chemistry, Lund University, Post Office Box 124, 221 00 Lund, Sweden;
madeleine.jonsson@biotek.lu.se (M.J.); leila.allahgholi@biotek.lu.se (L.A.);
roya.sardari@biotek.lu.se (R.R.R.S.)
2 Faculty of Life and Environmental Sciences, University of Iceland, Askja, IS-101 Reykjavík, Iceland;
gudmundo@matis.is
3 Matis Ohf, Vinlandsleid 12, IS-113 Reykjavik, Iceland
* Correspondence: eva.nordberg_karlsson@biotek.lu.se

Academic Editors: Christine Delbarre-Ladrat and Sylvia Colliec-Jouault 



Received: 31 January 2020; Accepted: 17 February 2020; Published: 19 February 2020

Abstract: Marine macroalgal (seaweed) polysaccharides are highly promising for next-generation
applications in several industries. However, despite the reported comprehensive potential of these
polysaccharides, commercial products are scarce on the market. Seaweed cultivations are increasing in
number and production quantity, owing to an elevated global trend of utilization interest in seaweed.
The extraction of polysaccharides from seaweed generally generates low yields, but novel methods
are being developed to facilitate and improve the extraction processes. Current areas of applications
for seaweed polysaccharides mainly take advantage of the physicochemical properties of certain
polysaccharides, such as gelling, thickening and emulsifying. However, many of the numerous
bioactivities reported are still only at research level and lack clinical evidence for commercialization.
It has been suggested the construction of smaller units may generate better defined molecules that
are more suitable for biomedical applications. Enzymatic modification is a promising tool for the
generation of more defined, targeted biomolecules. This review covers; structural differences between
the most predominant marine algal polysaccharides, extraction processes, modification alternatives,
as well as a summary of current and potential next-generation application areas.

Keywords: seaweed; composition; structure; extraction; enzymatic modification; application; bioactivity

1. Introduction
Marine macroalgae (seaweeds) are highly promising as biomass resources for next generation
biorefineries. Seaweeds include several species of macroscopic, multicellular marine algae. According
to Chapman and Chapman [1], “Seaweeds belong to a rather ill-defined assemblage of plants known
as the algae. The term ‘seaweed’ itself does not have any taxonomic value, but is rather a popular
term used to describe the common large attached marine algae found in the groups Chlorophyceae,
Rhodophyceae, Phaeophyceae or green, red and brown algae respectively”.
Currently, approximately 32 million tonnes (wet weight) of seaweed is harvested globally [2],
of which around one million tonnes are harvested wild, while the harvest of the remaining 31 million
tonnes involves farmed seaweeds—with the largest production countries being China, Indonesia and
the Philippines [2]. In Europe, wild harvests still dominate, with Norway being one of the leading
producers globally (0.16 million tonnes harvested wild in 2016) [2]. Interest in the cultivation of
seaweeds in Europe is however growing, although it is still at an early stage, with a total production of

Molecules 2020, 25, 930; doi:10.3390/molecules25040930 www.mdpi.com/journal/molecules


Molecules 2020, 25, 930 2 of 29

less than 2000 tonnes in 2016 (mainly large brown kelps, such as Saccharina latissima, Undaria pinnatifida,
and Alaria esculenta) [3]. The most important farmed seaweed species include the red seaweeds
Eucheuma spp. (10.2 million tonnes), Gracilaria spp. (3.9 million tonnes), Kappaphycus spp. (1.8 million
tonnes) and Porphyra spp. (also called nori, 1.2 million tonnes), and the brown seaweeds Saccarina
japonica (brown seaweed species, 8.0 million tonnes) and U. pinnatifida (also called wakame, 2.3 million
tonnes) [4].
Seaweed cultivation initiatives in Europe are driven both by the demand for sustainable biomass
resources for industrial applications, and the increasing attention on more sustainable food production
and consumption. Seaweeds are promising in these aspects, due to their high productivity (exceeding
terrestrial biomass), the possibilities of bulk-scale farming without the need for fertilizers (not competing
with agricultural land), combined with causing very limited harm to the seabed or to fishery
resources [4–6]. Seaweed farming requires no fresh water for cultivation and occupies no terrestrial
space. In addition, seaweeds have a high carbohydrate content (approximately 50% DW (dry weight)
in different polymeric forms [5,6]) and they contain essential amino acids [7]. Seaweeds are also known
to contain various compounds with bioactivity, including a number of their polysaccharides, which
have been investigated in numerous research articles, but are not yet established as industrial products.
The main components in seaweeds include: polysaccharides, minerals, proteins, small amounts of
lipids, polyphenols and pigments. The seaweed polysaccharides pose a challenge as industrial feedstock
because of their structural complexity, “unconventional” and heterogeneous sugar composition,
sulfation and other modifications. The possible product range from macroalgae may, however, surpass
other biomass of comparable bulk and ease of cultivation. Besides all lignocellulose sugars, hexoses
(glucose, mannose and galactose), and pentoses (xylose and arabinose), appreciable amounts of “rare
sugars” (sugar alcohols, deoxy sugars and sugar acids (uronates)) are present in macroalgae. These
recalcitrant macroalgal polysaccharides are relatively new to industry, but they need to be utilized to
ensure full valorisation of seaweed biomass. Added value derivatives that can be obtained by enzymatic
and microbial conversions include food and feed additives, such as modified polysaccharides and
oligosaccharides with reported health promoting effects. However, the most attractive unrealized
potential may be in unique, speciality chemicals and platform chemicals, based on properties and
possibilities due to their rare sugars, for the chemical and pharmaceutical industries.
The combined potential of underexploited biomass, the prospect of vastly increased global
production, novel added-value products, and the anticipated expanded product range from seaweed
biomass has led to a number of European projects involving both seaweed cultivation and the
development of technological platforms for fractionation, extraction and pre-processing. Continued
work in this field will help to overcome current challenges in terms of limited shelf-life of the harvested
material, and further developments in establishing biocatalytic refining processes will help to expand
the product portfolios that today are mainly confined to purified components from the biomass [8].
The main products of seaweeds are still the carbohydrate polymers alginate (brown seaweeds), agar
and carrageenan (red seaweeds), which are extracted and refined in bulk and have important uses as
hydrocolloids, emulsifiers and thickeners [1]. These compounds are produced by methodologies that
can be described as mechanical and/or physicochemical extraction and fractionation technology, and
hence they are basically unmodified seaweed components. Products (on a more industrial scale) based
on biocatalytic or microbial conversions of seaweed biomass are, in principle, limited to conversion to
energy via anaerobic digestion, which still has some challenges to overcome to become economically
viable [9]. Enzyme conversions are still mostly at research level [8] but hold a range of possibilities as
the necessary processes and tools are being developed at this stage.
In this review, we outline the main components of marine macroalgae, including brown, red,
as well as green seaweed species. Different protocols to extract and fractionate defined components
are shown, and modification possibilities of purified extracts are mentioned. Application areas of
seaweeds are also presented, showing both currently existing as well as novel possibilities.
Molecules 2020, 25, 930 3 of 29

2. Brown Seaweed Polysaccharides


Brown seaweed, Phaeophyceae, are widespread macro algae growing abundantly at northern
latitudes in marine cold waters [10], although some species are indigenous to fresh and brackish
waters. Brown algae contain a carotenoid pigment, fucoxanthin, providing the plant’s characteristic
brown color by suppressing other pigments. The sizes of brown algae plant bodies range from a few
millimeters up to some 70 m [11], indicating a variety in structure and composition between species
within the same algal class.
Brown macroalgae consist of a complex and dynamic cell wall rich in polysaccharides, such as
alginate (also alginic acid and algin), fucoidan (also fucoidin and fucan), laminarin (also laminaran),
and cellulose [12,13], but also containing polyphenols, proteins, glycoproteins, phlorotannins (sulfated
phenolic compounds), halogens such as iodine, and minerals, such as sodium, potassium, calcium
and magnesium [11,13]. The relative content of different compounds varies between species and is
highly dependent on seasonal, environmental and regional factors [14]. Additionally, while alginate
seems to constitute a significant part of all brown algae, laminarin and fucoidan appear to occur more
frequently in some species, such as Laminaria spp. and Fucus spp., than in others [11]. The presence of
free sugars, such as glucose, fructose and sucrose in brown algae is insignificant, since monomeric
sugars are immediately converted into d-mannitol and polysaccharides [14]. Glucose content, for
instance, is seasonally dependent, bound mainly in the beta-glucan, laminarin that acts as storage
polysaccharide in the cells.
Deniaud-Bouët et al. [12] suggest that the cell wall of brown algae consists of fucose containing
sulfated polysaccharides (FCSPs) as a major crosslinking glycan in the cell walls, interlocking the
cellulose microfibrils situated in layers parallel to the cell surface that constitute the framework of the
cell. They also suggest that short fragments of hemicellulose are intermediate links between cellulose
and the cross-linking FCSPs, although the composition of these proposed hemicellulose fragments is not
defined. These structures are embedded in a matrix of alginates. The cell wall matrix is postulated to
consist of mainly alginates, proteins, polyphenols (linked to both these compound groups), and iodine.
Cellulose and/or mixed linkage glucans, comprising only a minor fraction (<10% DW) [15], may also
have a structural role in the cell wall as microfibrils associated with fucoidan or alginate [12,16].
Brown algae appears to have emerged later in time than red algae, green algae and terrestrial plants.
No unicellular brown algae species have been found, and the development of the multicellularity and
plant-like structures of brown algae occurred isolated from other algae and plants. However, brown
algae have cell wall polysaccharides in common with plants (cellulose), animals (sulfated fucans) and
bacteria (alginate) [13]. Although brown algae comprise a wide variety of compounds, most focus
and research has been directed towards the utilization of algal polysaccharides, especially fucoidan,
alginate and laminarin.

2.1. Fucoidan
Fucoidans are categorized as fucose-containing sulfated polysaccharides (FCSPs), which
encompass the polysaccharides of various chain lengths, structures of branching, and degrees of
sulfation. The predominant component in fucoidan is fucose, but it also contains other monomers such
as galactose, mannose, xylose and residues of glucuronic acid [17,18]. Fucoidans are divided into two
subgroups; one group whose backbone structure constitutes α-1,3-l-fucopyranose residues, while the
second group is composed of alternating 1,3- and 1,4-linked α-l-fucopyranose residues [19] (Table 1).
The variation between fucoidans is reflected in the vast divergences in reported molecular weights;
from about 40 kDa up to 1600 kDa [20].
The fucoidan content appears to be higher during autumn than at any other time of the year [11,20].
Fucoidans are considered important components for regulating water and ion retention in extracellular
matrixes of the plant, to prevent desiccation and osmotic stress when exposed to low tides [13]. It is
unclear whether composition alterations correlate only with an increase in storage polysaccharides or
if the changes impact plant tissue structure as well [21].
tides [13]. It is unclear whether composition alterations correlate only with an increase in storage
The fucoidan
Polysaccharide
polysaccharides or content
ifStructure
the appears
1 to beplant
higher during autumn than [21].
at any other time of the year
polysaccharides or ifchanges impact
the changes tissue
impact plant structure
tissue as well
structure as well [21].
[11,20]. Fucoidans are considered important components for regulating water and ion retention in
extracellular
TableTable matrixes
1. Basic of
structures
1. Basic theofplant,
structures to preventderived
polysaccharides desiccation
of polysaccharides and
from from
derived marineosmotic stress (a)
macroalgae:
marine when
macroalgae: (a)exposed
alginate, to
(b)–(c)
alginate, low
(b)–(c)
tidesfucoidan,
[13].fucoidan,
It is(d)
unclear whether
(a) laminarin,
laminarin,
(d) composition
(e)–(f)(e)–(f) alterations
carrageenan, (g)–(h)
carrageenan, correlate
agar, agar,
(g)–(h) only
and and with an
(i)–(k)(i)–(k) increase
ulvanulvan
(where in storage
(i)–(j):(i)–(j):
(where
polysaccharides
ulvanobiuronic or acids
if theacids
ulvanobiuronic changes
and k:and k:impact
ulvanobiose). plant tissue structure as well [21].
ulvanobiose).
Molecules 2020, 25, 930 4 of 29
Polysaccharide
Table Structure
1. Basic structures
Polysaccharide of1 polysaccharides
Structure 1 derived from marine macroalgae: (a) alginate, (b)–(c)
fucoidan, (d) laminarin, (e)–(f) carrageenan, (g)–(h) agar, and (i)–(k) ulvan (where (i)–(j):
Table 1. Basic structures of polysaccharides derived from marine macroalgae: (a) alginate, (b)–(c)
ulvanobiuronic acids and k: ulvanobiose).
fucoidan, (d) laminarin, (e)–(f) carrageenan, (g)–(h) agar, and (i)–(k) ulvan (where (i)–(j): ulvanobiuronic
(a) (a)
acids and k: ulvanobiose).
Polysaccharide
Alginate Structure 1

Polysaccharide Structure 1

(a) D-mannuronic acid (M) and α-1,4-L-guluronic acid (G) residues forming GG, MM
β-1,4-
and M/G blocks

Alginate
Alginate

Alginate (b) (c)


β-1,4-β-1,4-
D-mannuronic acid (M)
D-mannuronic acidand
(M)α-1,4- L-guluronic
and α-1,4- acid (G)
L-guluronic acidresidues forming
(G) residues GG, MM
forming GG, MM
Alginate
and M/G
and blocks
M/G blocks

β-1,4-D-mannuronicacid
β-1,4-d-mannuronic (M)
acid and
(M) α-1,4-l-guluronic
and acid
α-1,4-L-guluronic (G)(G)
acid residues forming
residues GG,GG,
forming MMMM
and M/G blocks
R = SO3- or H R = SO3- or H
Fucoidan (b) (b) blocks
and M/G (c) (c)
Alternating 1,3- and 1,4-linked α-1,3-L-fucopyranose

α- L-fucopyranose
Fucoidan
(b) (c)

R = SO
R3=- or
SOH3- or H R = SO
R3=- or
SOH3- or H
Fucoidan
Fucoidan Alternating
Alternating 1,3-and
1,3- and 1,4-linked
1,4-linked α-1,3-
α- l-fucopyranose L-fucopyranose
α-1,3- L-fucopyranose
α-1,3-l-fucopyranose
Alternating 1,3- and 1,4-linked
α- L-fucopyranose
(d) α- L-fucopyranose
R = SO3- or H R = SO3- or H
Fucoidan Alternating 1,3- and 1,4-linked α-1,3-L-fucopyranose
Molecules 2019,
Molecules 24, x24,
2019, FOR PEER
x FOR REVIEW
PEER REVIEW 5 of532of 32
Laminarin α- L-fucopyranose

Laminarin (e) (e) (f) (f)


(d) (d)
Molecules 2019, 24, x FOR PEER REVIEW 5 of 32
Molecules 2019, 24, x FOR PEER REVIEW
β-1,3-d-glucopyranose backbone with branching β-1,6-d-glucopyranose unit
β-1,3-D-glucopyranose backbone with branching β-1,6-D-glucopyranose unit 5 of 32

(e) (f)
(d) (e) (f)

Laminarin κ-carrageenan: R1 =RSO


κ-carrageenan: −, R−2 = R3 = H
1 = 3SO 3 , R2 = R3 = H
Laminarin
Carrageenan
Carrageenan ι-carrageenan: R1 =RR
ι-carrageenan: 1 3==RSO
−, R2− = H
3 = 3SO 3 , R2 = H
µ -carrageenan: R1 =RSO
µ -carrageenan: −, R−2 = R3 = H
1 = 3SO 3 , R2 = R3 = H
θ-carrageenan: R1 =RH,
θ-carrageenan: 1 =R H,2 =RR2 3==RSO
3 = 3SO3
Carrageenan ν-carrageenan:
β-1,3-
ν-carrageenan:
β-1,3- R1 =RR
D-glucopyranose 1 3==
D-glucopyranose RSO
backbone−, R2− = H
3 = 3SO 3 , with
backbone branching
R2 = H β-1,6-
with branching D-glucopyranose
β-1,6-D-glucopyranose unit unit
Laminarin Alternating
κ-carrageenan: β-1,3-
Alternating R D
β-1,3--galactopyranose
1 = SO 3 , R2 = R3 = H andand
D-galactopyranose

λ-carrageenan:
λ-carrageenan: R1 =RH,1 =R H,2 =RR2 3==RSO

3 = 3SO3− κ-carrageenan: R1 = SO3−− , R2 = R3 = H
κ-carrageenan: R 1 = SO3 ,− R2 =− R3 = H
Carrageenan µ-carrageenan:
Alternating R1 =D-galactopyranose
α-1,4- SO3 −− , R2 = R3 = H 3,6-anhydro-α-
3,6-anhydro-α-
ι-carrageenan:
ι-carrageenan: R =3 =RSO
3 =3 SO
D1-galactopyranose
R=D1R-galactopyranose R2 = H
, R32 =, H
Alternating
µ -carrageenan: α-1,4-
R1 = SO R3 = H andand
D-galactopyranose
3 , R2 =
Carrageenan ν-carrageenan: R1 = R3 = SO− 3 , R2 = H −
ι-carrageenan: R11 ==RH,
θ-carrageenan: 3 =RSO2 =3−R
,R3 2==SO
H3
µ -carrageenan:
β-1,3- D-glucopyranose
R1 = backbone
SO3 ,−R2 = with R3 =−H θ-carrageenan: R1 = H, Runit 2 = R3 = SO3
λ-carrageenan:
β-1,3- 1 1== H,
-galactopyranose
RR R3 R
D-galactopyranose
ν-carrageenan: = 2SO= 3R,3R=2 =SOH3 branching β-1,6- D-glucopyranose
Alternating β-1,3-d-galactopyranose and
θ-carrageenan: R1 = H, R2 = R3 = SO3
Alternating
ν-carrageenan: R1 = R3 = SO3−, R2 and
α-1,4-d-galactopyranose =H 3,6-anhydro-α-d-galactopyranose
Alternating β-1,3- D -galactopyranose and
λ-carrageenan: R1 = H, R2 = R3 = SO3−
β-1,3-d-galactopyranose Alternating β-1,3-D-galactopyranose and
λ-carrageenan: R1 = H, R2 = R3 = SO3− 3,6-anhydro-α-D-galactopyranose
Alternating α-1,4-D-galactopyranose and
(g) Alternating
(g) α-1,4-D-galactopyranose and (h) 3,6-anhydro-α-
(h) D-galactopyranose
β-1,3-D-galactopyranose
β-1,3-D-galactopyranose
Agar
Agar
Agar
(g) (h)
(g) (h)
R = H or side chain substituents e.g., sulfate ester, methoxyl ether or pyruvic acid
Agar R = RH=or
Hside
Alternating chain substituents
orβ-1,3-d-galactopyranose
side chain e.g.,e.g.,
substituents sulfate
and ester,
sulfate methoxyl ether
Alternating
ester, methoxyl etheror pyruvic acidacid and
or pyruvic
β-1,3-d-galactopyranose
Agar 3,6-anhydro-α-1,4-l-galactopyranose α-1,4-l-galactopyranose
Alternating β-1,3- D -galactopyranose and
Alternating β-1,3-D-galactopyranose and Alternating β-1,3- D -galactopyranose
Alternating β-1,3-D-galactopyranose andand
3,6-anhydro-α-1,4- L-galactopyranose
3,6-anhydro-α-1,4- L-galactopyranose α-1,4- L-galactopyranose
α-1,4- L-galactopyranose

R = H or side chain substituents e.g., sulfate ester, methoxyl ether or pyruvic acid
R = H or side chain substituents e.g., sulfate ester, methoxyl ether or pyruvic acid
(i)
Alternating
(i) β-1,3-D-galactopyranose and (j) (j) β-1,3-D-galactopyranose and
Alternating
Alternating β-1,3-D-galactopyranose and Alternating β-1,3-D-galactopyranose and
3,6-anhydro-α-1,4-L-galactopyranose α-1,4-L-galactopyranose
3,6-anhydro-α-1,4-L-galactopyranose α-1,4-L-galactopyranose

(i) (j)
(i) (j)
Agar
Agar
R = H or side chain substituents e.g., sulfate ester, methoxyl ether or pyruvic acid
Molecules 2020, 25, 930 Alternating β-1,3-D-galactopyranose and Alternating β-1,3-D-galactopyranose and 5 of 29
R = H or side chain substituents e.g., sulfate ester, methoxyl ether or pyruvic acid
R = H3,6-anhydro-α-1,4- L-galactopyranose
or side chain substituents α-1,4-L-galactopyranose
e.g., sulfate ester, methoxyl ether or pyruvic acid
Alternating β-1,3-D-galactopyranose and Alternating β-1,3-D-galactopyranose and
Table
Alternating β-1,3-D-galactopyranose 1. Cont.
and Alternating β-1,3-D-galactopyranose and
3,6-anhydro-α-1,4-L-galactopyranose α-1,4-L-galactopyranose
(i)
3,6-anhydro-α-1,4-L-galactopyranose α-1,4- (j)
L-galactopyranose
1
Polysaccharide Structure

(i) (j)
(i) (j)

Alternating β-1,4-D-glucuronic acid


and α-1,4-L-rhamnopyranose
Alternating β-1,4-D-glucuronic acid Alternating α-1,4-L-iduronic acid and
Alternating
(from β-1,4-d-glucuronic
Ulva acid and
Alternating β-1,4-rigida)
D-glucuronic acid
α-1,4-L-rhamnopyranose
and α-1,4-L-rhamnopyranose
α-1,4-l-rhamnopyranose (from Ulva rigida) Alternating α-1,4-l-iduronic acid and
and α-1,4-L-rhamnopyranose Alternating α-1,4-L-iduronic acid and
Ulvan
Ulvan (from Ulva rigida) α-1,4-l-rhamnopyranose
(from Ulva (from Ulva armoricana)
armoricana)
Alternating α-1,4- L-iduronic acid and
α-1,4-L-rhamnopyranose
(from Ulva rigida) (k)α-1,4-L-rhamnopyranose
Ulvan (from Ulva armoricana)
Ulvan (from Ulva armoricana)
(k)
(k)

Alternating β-1,4-d-xylanopyranose β-1,4-D-xylanopyranose(as shown in Enteromorpha sp)


and α-1,4-l-rhamnopyranose
Alternating
1 Structures were drawn using BIOVIA Draw, based on structures from Synytsya et al. [22], Zhao et al. [23], Campo
and α-1,4-L-rhamnopyranose (as shown in Enteromorpha sp)
et al. [24], Tuvikene et al. [25], Pérez et al. [26], and Kidgell
Alternating et al.D-xylanopyranose
β-1,4- [27]
Alternating β-1,4-D-xylanopyranose
and α-1,4-L-rhamnopyranose (as shown in Enteromorpha sp)
2.2. Alginate
1 Structures were drawn using BIOVIA Draw, based on structures from Synytsya et al. [22], Zhao et
and α-1,4-L-rhamnopyranose (as shown in Enteromorpha sp)
al. [23], Campo et al. [24], Tuvikene et al. [25], Pérez et al. [26], and Kidgell et al. [27]
Alginates are the salts of alginic acid, which are linear unbranched copolymers comprised of
1 Structures were drawn using BIOVIA Draw, based on structures from Synytsya et al. [22], Zhao et

β-1,4-d-mannuronic
1 Structures were drawn acid (M)BIOVIA
using
al. [23], Campo et al. [24]
and et
, Tuvikene
α-1,4-l-guluronic
Draw, based
al. [25]
acid from
onetstructures
, Pérez
(G)Kidgell
al. [26], and
(Table
Synytsya 1). al.The
et al.et[27][22]size
, Zhaoofetthe copolymer,
M/G-ratio
al. [23], and theetproperties
Campo of integrated
al. [24], Tuvikene et al. [25]G-blocks affects
, Pérez et al. the Kidgell
[26], and functions,
et al. physical
[27] properties, mechanical
strength and biocompatibility of the compound [28,29]. The M/G-ratio of alginates varies between
algal species, harvest period, as well as between the structure and age of the algae tissue. The chelating
properties of alginate are dependent on the length of the compounds’ G-blocks (repetitive structure of
guluronic acid) [30]. Due to the rheological properties of alginates, these polysaccharides are widely
used in industries such as the pharmaceutical, medical technology, cosmetic, food, agricultural, textile
and paper industries [29].
Alginates are mainly used industrially for their physical characteristics such as their stabilizing,
thickening and emulsifying properties, but depending on specific properties, such as gel strength,
porosity or biocompatibility, they are expanding into applications such as biomaterials for tissue
engineering and bioprinting [31]. Alginates are analogues to pectin in terrestrial plants [11]. The current
and potential application of alginates will be further appraised in Section 7.
Alginates extracted from brown algae are categorized as phycocolloids, polysaccharides able to
form colloidal systems when dispersed in water [11]. Alginates are formed when alginic acid interacts
with present metal ions such as calcium, sodium or magnesium. Calcium alginates form gels which are
insoluble in water, while sodium and magnesium salts of alginic acid turn into water-soluble gels [32].
The glycosidic bonds between sugar monomers of alginate polymers can be exposed to cleavage in
both acidic and alkalic conditions, which causes degradation of the compound. Other factors affecting
the stability of the copolymer are temperature and free radicals generated by contaminants such as
polyphenols [29].
Molecules 2020, 25, 930 6 of 29

2.3. Laminarin
Laminarin is a β-1,3-glucan, consisting of β-1,3-d-glucopyranose units interspersed with
β-1,6-linked d-glucopyranose units forming branch-points or interchain residues (Table 1). It is
a comparatively short polymer consisting of about 20–25 monomers [14,33]. Laminarin is the
carbohydrate reserve of many brown seaweeds, corresponding to starch (α-1,4-glucans) in terrestrial
plants [10,14,34]. Interchain β-1,6-linkages contribute to the branching structure of laminarin and
determine the water solubility of the compound. Increased branching is associated with elevated
solubility in water [14]. Two different types of laminarin, G-type and M-type, have been explored, and
they simply differ in the reducing end of the polymer chain. Whilst laminarin G-type only contains
glucopyranose residues, the M-type is terminated with 1-O-linked d-mannitol [14,34]. The ratio
between M-type and G-type chains can be as high as 3:1 as observed in Laminaria digitata [35], while
M-types were reported to be absent in laminarin from Eisenia bicyclis [36].
Laminarin, and laminarin-derived oligosaccharides, can be expected to elicit the same
health-promoting responses in organisms as reported for other β-glucans, including prebiotic,
antioxidant, anticoagulant, hypocholesterolemic, and antimutagenic activity, and additionally,
they have reported anti-inflammatory and anti-cancer effects that appear to be confined to
β-1,3-(1,6)-glucans [37]. Immunomodulating activities vary between laminarins (Hreggviðsson
and Nordberg Karlsson, unpublished), depending on species, and may be related to varying content,
type (branchpoints or interchain) and spatial distribution of the β-1,6-linkages. The ratio between
β-1,6- and β-1,3-linkages in laminarin tends to vary between species; from a very low ratio in laminarin
from Laminaria hyperborea, which is composed mainly of β-1,3-linkages [38], to a moderate ratio of 1:7
in laminarin from L. digitata, and to a high range of 1:3 or 2:3 in laminarin from E. bicyclis [35,36].

3. Red Seaweed Polysaccharides


Red seaweed, Rhodophyta, is a group of mainly multicellular photosynthetic eukaryotic organisms,
reported to comprise more than 7000 species. They grow mostly in marine habitats, but up to 5% of
species can grow in freshwater environments, with greater numbers in warmer areas. The red color
is expressed due to the predominance of the pigments phycocyanin and phycoerythrin over other
pigments such as chlorophyll A and carotene. Red algae store sugars as floridean starch, a highly
branched amylopectin without amylose, which is accumulated outside the plastid in the cytosol of the
cells [39]. The cell wall of the seaweed is mainly composed of cellulose, carrageenan and agar, which
all have several industrial applications.

3.1. Carrageenan
Carrageenan is a high molecular weight sulfated polysaccharide, available in the cell wall of
red seaweed interacting with other bioactive compounds including proteins, lipids, pigments and
other polysaccharides. It is composed of alternating α-1,3-linked d-galactopyranosyl and β-1,4-linked
d-galactopyranosyl groups and 3,6-anhydrogalactose residues.
Molecules 2020, 25, 930 7 of 29

The three most important commercial types of carrageenan are kappa (κ), iota (ι) and lambda (λ)
carrageenan (Table 1), which differ in the amount and position of sulfate ester groups and the number
of 3,6-anhydrogalactose residues. κ-carrageenan is composed of 25–30% sulfate ester groups and
28–35% anhydrogalactose. The sulfate ester content in ι–carrageenan is similar to that of κ-carrageenan
(28–30%), but it contains a lower fraction of anhydrogalactose units (25–30%). The sulfate substitution
in λ-carrageenan is comparatively high (32–39%), but it contains no anhydrogalactose residues [24].
The backbone can be substituted by pyruvate ketal, which is only reported for the λ-family of
carrageenans [40].
The composition of carrageenan varies between species. For example, carrageenan from
Kappaphycus alvarezii (in the trade also called Eucheuma cottonii) is made of mainly κ-carrageenan,
whereas Eucheuma dendiculatum (trade name Eucheuma spinosum) consists of ι-carrageenan. Chondrus
crispus and Sarcothalia crispata contain both κ- and λ-carrageenan. The water solubility and gelation of
carrageenan depends on the amount of sulfate ester groups and associated divalent cations, such as
sodium and potassium, although λ-carrageenan can form a gel in the presence of trivalent ions [41].
Higher numbers of sulfate ester groups result in lower temperatures required for solubility and lower
gel strengths, which affects the application of carrageenan [42]. Further information about the source,
structure and properties of carrageenan are explained in a comprehensive review by Campo et al. [24]

3.2. Agar
Agar is a mixture of polysaccharides with gelling capacity, such as agarose and agaropectin,
extracted mostly from Gelidium and Gracilaria species [43]. Agarose is the major fraction of agar,
estimated to constitute 70% of agar polysaccharides. It is a high molecular weight polysaccharide
consisting of d-galactose and 3,6-anhydro-α-l-galactose units joined by β-1,3 and α-1,4 glycoside
linkages (Table 1) [26,44]. The agar backbone can be substituted with sulfate esters, methoxyl groups
and pyruvate ketals [44]. Agaropectin has a lower molecular weight and a higher amount of sulfate
ester groups than agarose. It has the same backbone as agarose, which, in agaropectin, is substituted
with various amounts of sulfate esters, d-guluronic acids and pyruvic acids.

4. Green Seaweed Polysaccharides


Green seaweeds are members of the Chlorophyta, which include the majority of the described
species of green algae [45,46]. Members of the Chlorophyta are inhabitants of both marine and freshwater
systems. Marine macroalgae (the focus in this review) from the classes of Chlorophyta in principle
involve the class Ulvophyceae [47] and mainly include candidates from the orders Ulvales-Ulotrichales,
Cladophorales, and Bryopsidales, including species that display large morphological and cellular
diversity [45,48,49]. Well known genera classified under Ulvophyceae include various edible species
of Ulva, Gayralia and Monostroma (classified under the order Ulvales-Ulvotrichales), Cladophora
(Cladophorales), Caulerpa and Codium (Bryopsidales), all of which have been used in human diets in
different parts of the world. Despite this traditional use, green seaweeds are not as well researched as
red and brown, and although having potential, they lack major industrial applications, such as those
seen for red and brown algae.
Green seaweeds contain interesting polysaccharides that are sulfated and/or carboxylated.
The backbones of the main polysaccharides differ in candidate species from different genera, but
roughly, they can be divided into two major groups that are classified as uronic acid rich or uronic acid
limited [22]. The uronic acid rich (sulfated) polysaccharides are represented by the ulvans that can
be found in various species of Ulva, Gayralia and Monostroma. The composition of ulvan is reported
to be dependent on species, season, growth conditions, as well as on the method of isolation [27].
Four different types of monosaccharides (rhamnose (Rha), xylose (Xyl), glucuronic acid (GlcA) and
iduronic acid (IdoA)) have, along with sulfate groups, been reported in different types of ulvans, which
are also described as consisting of several disaccharide fragments. All four types of monosaccharides
can occur in the backbone, while only glucuronic acids have, according to Synytsya et al. [22], also
Molecules 2020, 25, 930 8 of 29

been reported as side chains. The repeating bios-unit is often reported to consist of (uronic acid-Rha),
such as in Ulva sp., where the uronic acid is either glucuronic acid or iduronic acid. Repeating units
with alternating (Rha-Glc-Rha-xyl) structures have also been reported from Ulva pertusa [50]. The basic
structures of common ulvans are depicted in Table 1. The uronic acid limited polysaccharides occur,
for instance, in species of Codium and are reported as sulfated galactans, arabinopyranans and mannans.

5. Extraction of Seaweed Polysaccharides


Since seaweeds encompass high levels of water, extraction of polysaccharides from the biomass
should be preceded by water content determination. While traditional methods for determining
moisture level, such as thermogravimetric analyses, are primarily employed, new and faster techniques
have potential as alternatives. For instance, near-infrared spectroscopy has proved successful for the
fast and accurate analysis of water content in corn stover silage [51], and could be a useful tool also for
seaweed. The dry weight is further applied as the basis for yield calculations.
Processes for pretreatment and extraction are usually not distinctly separated when isolating
polysaccharides from seaweed, and different extraction methods are commonly combined with attempts
to increase yields. In this review, pretreatment and extraction refer to different processing steps
applied to macroalgae in order to solubilize the polysaccharides into a liquid medium. Pretreatment
methods include processes employed for swelling of macroalgal fibers and increasing pore sizes, or
reducing sizes of the biomaterial. The pretreatment of macroalgae consists of mainly mechanical (size
reduction, washing, beating, and sonication), thermal (microwave, steam explosion, wet oxidation,
and plasma-assisted), chemical (acid or alkali, peroxide), and biological pre-treatments [52]. However,
other methods such as liquid ammonia pretreatment have been performed as a promising method
for lignocellulosic biomass [53], and might also have potential for the pretreatment of macroalgae.
Traditionally, chemical extraction methods have predominantly been used to extract polysaccharides
from algal biomass, but have recently been accompanied by new techniques with the aims of
increasing extraction yield, using fewer solvents, operating at lower temperatures and decreasing the
extraction time (Table 2) [54,55].These efforts aim to reduce energy consumption, thereby increasing
the sustainability of the method. This review will further cover extraction processes of polysaccharides
from brown, red, and green seaweed.
Molecules 2020, 25, 930 9 of 29

Table 2. Overview of extraction methods.

Extraction Method Principle Advantages Disadvantages Ref.


Different procedures of acid or alkaline extraction
and hot or cold water extraction. Acidic or alkaline Long extraction time.
conditions are usually applied to facilitate High consumption of energy and water.
extraction, as hydrogen ions (H+ ) and hydroxyl Alcohol precipitation and recovery are
ions (OH− ) interfere with hydrogen linkages costly.
Conventional chemical
between polysaccharides. The conventional Well established methods. Chemical solvents may have health [56–61]
extraction
extraction procedures rely on the solubility hazards.
properties of target compounds and are often Acids and bases can cause degradation of
preceded by pretreatment steps where lipids, the target polysaccharide to compounds of
pigments, proteins and other impurities are smaller molecular size.
removed by solvents.
UAE acts by exposing the biomass to sound waves
of high frequencies larger than 20 kHz. Strong
ultrasound fields cause the implosion of vapor Short extraction time.
bubbles in liquids formed under high pressure Higher yields of extracted bioactives.
Ultrasound assisted extraction conditions. Implosion of these bubbles in near Simple method to operate. Degradation and structural changes in the
[54,62,63]
(UAE) proximity to liquid-solid borders, such as cell Operation at low temperatures. structure of polysaccharides.
walls, subject these solid surfaces to strong forces Relatively low amounts of solvent are
resulting in cell breakdown. UAE can be required.
performed at low temperatures which enable the
extraction of thermosensitive target compounds.
MAE utilizes non-ionizing electromagnetic
radiation with frequencies between 300 MHz and
300 GHz which cause the disruption of hydrogen
Short extraction time.
bonds and migration of dissolved ions. This
Microwave assisted extraction Relatively low amounts of solvent are Lower yield is achieved due to
enables the solvent to enter the cell matrix and [54,61,64,65]
(MAE) required. degradation.
facilitate the withdrawal of compounds of interest.
Higher quality of product.
Variables such as temperature, pressure, time and
algae/water ratio can be altered to optimize the
yield of the desired product.
Molecules 2020, 25, 930 10 of 29

Table 2. Cont.

Extraction Method Principle Advantages Disadvantages Ref.


Relatively low amounts of solvent are
required.
EAE operates by using enzymes for degradation of
Relatively low-cost technique.
the algal cell wall, thereby releasing target Extraction yield is dependent on optimum
Disruption of cell wall components is
compounds. Critical parameters, including pH, treatment time, pH and temperature
enzymatically performed.
temperature and treatment time, should be conditions of enzymes.
Enzyme assisted extraction Original efficacy of bioactives is preserved to
optimized for specific enzymes to maximize the Target compounds risk being degraded by [54,66–69]
(EAE) a high degree.
extraction result. Enzymes catalyzing degradative non-specific enzymes.
Potential of a higher yield of the target
reactions of cell wall structure compounds like Extraction efficiency is dependent on
compound.
cellulose, β-glucan and hemicellulose are usually enzyme properties.
The mild conditions applied to the sample
used to facilitate the extraction of target molecules.
during EAE are advantageous when
isolating sensitive bioactive compounds.
A supercritical fluid is a substance at a temperature
and pressure above its critical point, where gas
and liquid phases are indistinct. Altering the two
parameters can change the solubility of the fluid.
Carbon dioxide is commonly used as its critical
point is relatively low and thus requires less Use of non-toxic and non-flammable solvent.
energy input to become supercritical, compared to Supercritical CO2 is inexplosive, readily High pressure is needed to maintain the
Supercritical fluid extraction substances with higher critical points. Supercritical available, and can be removed easily from solvent in critical state, which can have
[54,70–72]
(SFE) fluids are characterized for their low viscosity and the final extract. negative effect on compounds.
high diffusivity which gives them better transport Does not cause degradation and structural Relatively high procurement costs.
properties than liquids. SFE is considered an disruptions in bioactive compounds.
environmentally friendly process as it does not
require the use of solvents. However, procurement
costs are high in relation to other extraction
methods. This concept is therefore predominantly
employed to extract highly valuable compounds.
Molecules 2020, 25, 930 11 of 29

5.1. Extraction of Brown Seaweed Polysaccharides


The available literature suggests a comprehensive number of methods for the extraction of
polysaccharides from macroalgae. However, the majority of these refer to modifications of existing
conventional methods. The development of extraction methods of fucoidan, one of the more difficult
polysaccharides to extract in good yield from brown algae, is reviewed by Ale et al. [18] with a historical
perspective (1913–1950).
A general, simplified flow chart of the extraction of brown macroalgal polysaccharides is
illustrated in Figure 1. For brown algae, a calcium compound (usually CaCl2 ) can be introduced to
the polysaccharide fraction during extraction to precipitate alginate, and thereby separate laminarin
from the alginate fraction, while fucoidan is present in various amounts in both fractions [73]. Sulfated
polysaccharides in the supernatant are often precipitated with ethanol, and further purification
steps—often repeated extractions in acid followed by precipitation, size-exclusive chromatography, or
filtration—can be applied to achieve pure fucoidan [59,60,74]. It has been suggested to always include
a benchmark procedure when employing new extraction methods for fucoidan, in order to get better
comparisons of fucoidan structures [75]. The extraction of laminarin is predominantly performed
at increased temperatures, since the water solubility of different forms of laminarin vary. Branched
forms tend to be more water soluble than non-branched. The typical extraction yields of laminarin are
presented in Table 3. Alginates, being industrially produced from brown macroalgae, are commonly
extracted by several ion-exchange procedures using either the ‘acid precipitation method’ or the
‘calcium precipitation method’ [76,77]. Simplified, water-insoluble divalent alginate gel or alginic acid
gel are swelled in acidic water prior to extraction. Sodium-containing compounds are then introduced
to convert the divalent alginate into water-soluble sodium alginate during extraction. Aqueous alginate
is highly viscous and must be diluted prior to clarification and filtration. The typical yield of alginate
extraction (Table 3) is usually higher than that of fucoidan and laminarin. However, different isolation
methods seem to affect the mannuronic/guluronic acid ratio (M/G ratio) of the extracts differently,
which impacts the gelling properties of alginate [78,79].

Table 3. Extraction yields from different extraction methods.

Polysaccharide Yield 1 Species Method Ref.


1.63% Padina tetrastromatica Conventional extraction (HCl) [74]
9.46% P. tetrastromatica Conventional extraction (hot water) [74]
3.51% Nizamuddinia zanardinii UAE [80]
4.44% (4.7%, CaCl2 ) Fucus evanescens UAE [81]
18.22% Fucus vesiculosus MAE [64]
16.08% (20.98%, HCl) Ascophyllum nodosum MAE [65]
5.58%, Alcalase
Fucoidan 4.80%, Cellulase
4.36%, Flavourzyme N. zanardinii EAE [82]
4.28%, Viscozyme
(5.20%, hot water)
1.5 ± 0.3% U. pinnatifida EAE [83]
3.02% (5.11%, EtOH) F. evanescens SFE [84]
1.26% (1.35%, SFE-EtOH)
S. japonica SFE [84]
(1.28%, EtOH)
0.57% (0.55%, SFE-EtOH)
Sargassum oligocystum SFE [84]
(0.65%, EtOH)
51.8% L. digitata Conventional extraction (HCl) [79]
13.47% Sargassum muticum Conventional extraction (HCl) [78]
Alginate
54% 2 Sargassum binderi UAE [85]
23.6 ± 1.2% U. pinnatifida EAE [83]
Molecules 2020, 25, 930 12 of 29

Table 3. Cont.

Polysaccharide Yield 1 Species Method Ref.


6.0 ± 0.7% S. latissima Conventional extraction (HCl + EtOH) [33]
19 ± 2.6% S. latissima Conventional extraction (HCl + NaOH) [33]
Laminarin 20% S. latissima Conventional extraction (hot H2 SO4 ) [33]
6.1 ± 1.6% S. latissima Conventional extraction (hot HCl) [33]
3.2 ± 0.9% U. pinnatifida EAE [83]
29.7 ± 1.9%
Gracilaria lemaneiformis Conventional extraction (hot water) [86]
Gel strength: 271 ± 38 g/cm2
Conventional extraction (alkali
25.8 ± 2.9% (Gel strength:
G. lemaneiformis pre-treatment with 5% NaOH + [86]
1761 ± 35 g/cm2 )
hot water)
Agar Conventional extraction
25.4 ± 1.7% (Gel strength:
G. lemaneiformis (alkali pre-treatment with 5% NaOH [86]
1913 ± 38 g/cm2 )
+Photobleaching + hot water)
29.7–34.6% (Gel strength: Gracilaria
Conventional extraction (hot water) [87]
72 g/cm2 ) vermiculophylla
15.03% (Gel strength: Conventional extraction (pre-treatment
G. vermiculophylla [87]
1064 g/cm2 ) with 7% NaOH + hot water)
46.43% E. cottonii Conventional extraction (hot water) [88]
37.02% E. cottonii Conventional Extraction (KOH) [88]
Furcellaria lumbricalis
76.3% Conventional extraction (hot water) [89]
Coccotylus truncates
F. lumbricalis
72.6% Conventional extraction (0.15 M NaOH) [89]
Carrageenan C. truncatus
F. lumbricalis
55.3% Conventional extraction (0.15 M KOH) [89]
C. truncatus
K. alvarezii
50–55% (for both species) 3 UAE [85]
Euchema denticulatum
28.65% Mastocarpus stellatus EAE [90]
1 Based on dried biomass. 2 Ultrasound extraction resulted in doubling the extraction yield from 28% to 54%.
3 Longer ultrasound treatment of up to 30 min did not further improve the extraction yield. Additionally, in this

study, degradation and structural modification were not observed in extracted polysaccharide.

In Table 3, a selection of different extraction approaches including novel techniques have been
compared with respect to their yields. However, although calculated yields are often used as measures
of efficacy of the extraction method, the structural integrity of the yielded compound is as important
for its function. Additionally, the estimated yield does not take into consideration the variation in
polysaccharide content between different species and other external factors, and the purity of the
extracted polysaccharide is usually not specified.
Molecules 2019, 24, x FOR PEER REVIEW 14 of 32

Figure 1.Figure
Basic1.flow
Basic flow
chartchart of extraction of
of extraction of polysaccharides
polysaccharides fromfrom
brownbrown
macroalgae.
macroalgae.
5.2. Extraction of Red Seaweed Polysaccharides
Carrageenan is extracted from several species of red seaweed including Hypnea, Gigartina,
Chondrus, Eucheuma, Kappaphycus, and Mastocarpus [91]. In conventional extraction, an alkali solution
is used to extract carrageenan, because in alkaline conditions some of the sulfate ester groups are
removed with the formation of 3,6-anhydro-D-galactose units, thus improving gel strength.
Molecules 2020, 25, 930 13 of 29

5.2. Extraction of Figure


Red Seaweed Polysaccharides
1. Basic flow chart of extraction of polysaccharides from brown macroalgae.

Carrageenan
5.2. Extraction is
of extracted
Red Seaweedfrom several species of red seaweed including Hypnea, Gigartina, Chondrus,
Polysaccharides
Eucheuma, Kappaphycus, and Mastocarpus [91]. In conventional extraction, an alkali solution is used to
Carrageenan is extracted from several species of red seaweed including Hypnea, Gigartina,
extract carrageenan, because in alkaline conditions some of the sulfate ester groups are removed with
Chondrus, Eucheuma, Kappaphycus, and Mastocarpus [91]. In conventional extraction, an alkali solution
the formation
is used to of 3,6-anhydro-d-galactose
extract carrageenan, because in units, thusconditions
alkaline improving gelof
some strength. Extraction
the sulfate parameters
ester groups are
such removed
as operationwith the formation of 3,6-anhydro-D-galactose units, thus improving gel strength.of the
time, temperature, concentration of alkali, and the ratio between the weight
seaweed and the
Extraction volume ofsuch
parameters the as
alkali solution
operation depend
time, on the seaweed
temperature, species
concentration and the
of alkali, desired
and quality
the ratio
betweencarrageenan
of extracted the weight of[25,91–94].
the seaweed and the volume of the alkali solution depend on the seaweed
species are
There and two
the desired quality ofextraction
conventional extracted carrageenan
procedures [25,91–94].
to extract refined carrageenan (RC) and
There are two conventional extraction procedures to extract
semi-refined carrageenan (SRC) (Figure 2a). In RC production, carrageenan refined carrageenan (RC) and
is solubilized insemi-
hot water
refined carrageenan (SRC) (Figure 2a). In RC production, carrageenan is solubilized in hot water
containing alkali, such as sodium hydroxide, and the remaining insoluble compounds are separated
containing alkali, such as sodium hydroxide, and the remaining insoluble compounds are separated
via filtration. The solution containing carrageenan is concentrated and carrageenan is subsequently
via filtration. The solution containing carrageenan is concentrated and carrageenan is subsequently
precipitated by alcohol, dried and milled.
precipitated by alcohol, dried and milled.

(a) (b)

2. Conventional
Figure
Figure extraction
2. Conventional extractionmethod
methodof of refined carrageenan(RC)
refined carrageenan (RC) and
and semi-refined
semi-refined carrageenan
carrageenan
(SRC)(SRC) (a). Alkaline
(a). Alkaline method
method forfor agarextraction
agar [95] (b).
extraction [95] (b).

In SRC production, carrageenan is not solubilized from the seaweed but forms a gel in the presence
of potassium chloride. In this method, the seaweed is initially boiled in hot potassium chloride solution.
Thereafter, compounds solubilized in the alkali solution including protein, carbohydrate and salts are
separated and the remaining seaweed is rinsed to remove residual alkali. The product which looks like
seaweed is dried and milled. This method is mainly applied to extract κ-carrageenan, but can also be
applied to produce semi-refined ι-carrageenan. The market for κ-carrageenan is significantly higher
than for ι-carrageenan due to the favorable properties of κ-carrageenan in many industries [91,95].
SRC is generally cheaper than RC, as no alcohol precipitation is used, hence alcohol recovery steps are
not needed. Different grades of refined carrageenan and semi-refined carrageenan, such as Philippine
natural grade (PNG) and processed Eucheuma seaweed (PES), have been developed to have a quality
suitable for human consumption whilst being less expensive than mere refined carrageenan [96].
Agar is typically extracted by solubilization in hot water. For some species such as Gelidium,
hot water is used directly after an initial cleaning step to extract agar, but for other species such
as Gracilaria, alkali treatment is essential before hot water extraction, to improve the gel strength
(Table 2b), otherwise the agar quality is too low for commercial applications. Similar to carrageenan,
the extraction time, temperature and alkali concentration have to be optimized for individual seaweed
Molecules 2020, 25, 930 14 of 29

species. Following alkali treatment, extensive rinsing is essential to neutralize residual alkali. Hot
water extraction can also be combined with pressure to improve the extraction yield [43,86,97].
Alkali treatment in the extraction of agar and carrageenan does not always cause an improvement
in the yield, but enhances the gel strength significantly, which can be observed in Table 3. During
the washing step, after alkaline treatment, massive amounts of by-product are produced, which are
currently poorly valorized. The study from Lebbar et al. [43] showed that the alkali waste contained low
molecular weight compounds belonging to the glycerol-galactoside family, which have the potential
for industrial applications.

5.3. Extraction of Green Seaweed Polysaccharides


The extraction of ulvan (the most well-known polysaccharide from green algae) has recently been
reviewed by Kidgell et al. [27], and hence only a brief summary is included here. In accordance with
the data found for other seaweed polysaccharide extractions (see Table 3), the quantitative yield was
shown to vary significantly, ranging from 2.7–40% of the dry weight (expressed as median% of algal
dry weight), and was dependent on both the applied process and the source of biomass.
The authors also highlighted that the main focus in published literature is on the physicochemical
composition of the polymer and its interaction with the cell wall, while less attention is generally given
to the risk of degradation and eventual co-extracted products. Factors of importance were identified to
be pH, temperature and duration of extraction [27].

6. Co-Extraction of Seaweed Byproducts


The extraction of seaweed cell wall polysaccharides will, in most of the extraction processes, result
in co-extraction of other polysaccharides (e.g., mixtures of alginate/fucoidans in brown seaweed or of
ulvan/glucuronan in green seaweed) [22], proteins from cell wall polysaccharides-protein complex, and
minerals which are adsorbed to the charged cell wall polysaccharides. These co-extracted byproducts
make the extraction process challenging, requiring more purification steps to obtain the pure target
polysaccharides. Sequential extraction of all seaweed compounds can be a solution to this, in order
to extract the target compound selectively and also take advantage of the extraction of multiple
compounds for different applications [98].
Many of the extraction processes described above result in products that in addition to the target
polysaccharide also contain minerals (unpublished data). Brown macroalgae have a high content of
ash, which mainly corresponds to different minerals, and have for example been reported as good
sources of sodium (Na) and potassium (K), and can be used in the place of salt in food. They are also
very good contributors of copper (Cu), zinc (Zn), molybdenum (Mo) and selenium (Se). Red and
green macroalgae contain high amounts of manganese (Mn) and green algae are a great source of iron
(Fe). Many macroalgae are also rich in iodine (I) [99]. An analysis of these minerals is thus important,
especially if the products are to be considered for consumption, to take into account recommended
daily allowances (RDAs) of minerals and trace elements [99,100]. Moreover, the absorption of heavy
and toxic metals by algae (e.g., arsenic (As), cadmium (Cd) and lead (Pb)), has raised concerns about
macroalgae consumption. Arsenic can occur in two forms: organic form (which has shown very low or
no toxicity), or inorganic form (which contrarily has presented high toxicity), making analysis of the
two forms important [101,102]. Generally, brown macroalgae accumulate As to a greater extent than
red and green algae. High amounts of Cd and Pb have also been reported in the brown and red algae
from some sites [99]. It can be concluded that the consumption of macroalgae should conform with the
tolerable daily intake (TDI) established by the World Health Organization (WHO).
Molecules 2020, 25, 930 15 of 29

7. Modification of Seaweed Polysaccharides

7.1. Fucoidan
The depolymerization of crude fucoidan is considered necessary for effective treatment in
biomedical applications; both due to the heterogeneity of fucose containing sulfated polysaccharides
(FCSP) and due to the large molecular sizes. Fucoidan of low molecular weight has namely demonstrated
better absorption and bioavailability compared with compounds of larger molecular weight [103].
The structural data of fucoidan suggest that there is no coherent basic structure of the polysaccharide, but
rather a large variation between species, with varying compound sizes and sulfate groups substituted
intermittently at C-2, C-3 and C-4. An example of fucoidan structure is presented in Table 1. Further
investigation of the grade and position of sulfation in combination with compound size and linkage
type is crucial for understanding the bioactivity of the compound [75].
Creation of oligosaccharides from sulfated polysaccharides is postulated as a way of controlling
the compound size and its bioactivity. Enzymatic degradation by hydrolysis, using glycosidases such
as endo- and exo-fucoidanases (also fucosidase, EC 3.2.1.44 and EC 3.2.1.51), can thus be valuable for
the generation of oligosaccharides of lower molecular weight. Three types (suggested denomination
‘Type 1-3, by Ale and Meyer [75]) of fucoidanases with different cleavage mechanisms are currently
identified, all classified under glycoside hydrolase 107 in the CAZy database (GH107, see www.cazy.org).
Type 1 catalyzes the endo-acting cleavage of α-1,4-glycosidic bonds, while Type 2 correspondingly
catalyzes the cleavage of α-1,3-glycosidic bonds. Type 3, however, includes α-l-fucosidase, which are
exo-acting enzymes that catalyze the degradation of α-l-fucosyl bonds at the non-reducing termini,
releasing fucose from the backbone of fucoidan [75]. These enzymes occur in different GH-families in
the CAZy database, of which the enzymes classified under GH29 are structurally related to the GH107
endofucoidanases [8].
Another important group of enzymes for the modification of fucoidan structure and bioactivity is
sulfatases (EC 3.1.5.6). Modifying the degree of sulfation, by sulfation or de-sulfation, is important
to control the effects of fucoidan and increase its biofunction [104]. Sulfatases are a uniform group
of enzymes with highly conserved sequences, structures, and mechanisms. However, four different
classes of enzyme are distinguished. Type I, formylglycine dependent sulfatase, comprises the majority
of the identified sulfatases to date. Type II consists of Fe(II) α-ketoglutarate-dependent alkylsulfatases
and Type III contains enzymes involved in Zn2+ - or Mn2+ -dependent metallo-β-lactamase activity.
Type II and III do not encompass any sulfatases active in carbohydrates. Finally, Type IV is another
potential sulfatase family member, with predicted marine polysaccharide activity [104].
The reported activity of sulfatases on polysaccharides and their specificity to oligosaccharides
suggest potential in the effective modification of marine sulfated compounds. Most recognized
sulfatases are constructed of one catalytic module, but dimodular and multimodular enzymes also
occur. Structural analyses indicate that sulfatases with human and bacterial origins have a similar
structure [104,105]. They act by catalyzing the hydrolytic desulfonation mechanism of sulfate esters
(CO-S) and sulfamates (CN-S).

7.2. Alginate
Within the pharmaceutical industry, alginic acid and alginates are primarily used as excipients in
formulations for oral and topical administration, due to their physiochemical properties. The primary
usage is for the assurance of controlled drug release, which is affected by systemic pH and temperature,
as well as the binding strength of alginate. G-residues of alginates have a stronger affinity to
divalent ions compared with M-residues, as they crosslink in the form of a pocket (denominated ‘the
egg-box’) surrounding the ion, generating alginates with significantly higher strength. Therefore,
the physiochemical properties are highly dependent on the M/G ratio of the polymer. The properties
of alginate products can accordingly be altered by enzymatically modifying the M/G ratio [106–108].
Molecules 2020, 25, 930 16 of 29

Four groups of alginate modifying enzymes have been characterized: mannuronan C-5-epimerases,
alginate lyases, alginate acetylases, and alginate deacetylases [106]. Mannuronan C-5-epimerases
are involved in the epimerization of mannuronic acid to guluronic acid, increasing the crosslinking
properties of alginate. Alginate lyases are well studied and have been demonstrated to catalyze alginate
degradation with different affinities to the four different bonds (M–M, G–G, M–G, G–M) of the polymer.
Most alginate lyases operate by endolytic modes, generating oligomers usually in the size range of two
to five monomers. However, some alginate lyases are not able to accommodate acetylated substrates in
their active site. Therefore, to facilitate or hinder enzymatic hydrolysis, alginate acetylases and alginate
deacetylases, respectively, can be utilized to modify the accessibility of the enzyme to the substrate.
Moreover, the degree of acetylation affects the water-binding properties and increases the viscosity of
the alginate.

7.3. Laminarin
Complex and highly potent structures could be engineered from laminarin as health-promoting
agents, if appropriate enzymes are accessed and used. Hydrolytic enzymes, including
endo-β-1,3-glucanases and -d-glucosidases (EC 3.2.1.6 and EC 3.2.1.39), as well as exo-β-1,3-glucanases
(EC 3.2.1.58), can be used for depolymerization reactions of the laminarin backbone structure, to generate
possibly more potent bioactive derivatives. Endo-acting enzymes act randomly by catalyzing the
hydrolysis reaction of the backbone structure into glucose and oligosaccharides, while exo-enzymes
catalyze hydrolysis of the reducing end into glucose. Although β-1,3-glucanases are a well-studied
group of enzymes, their β-1,6-linked side-chain analogues are less explored. Endo-acting enzymes can
be limited by steric hindrance from side groups in highly branched laminarins, resulting in reduced
enzyme activity [109].
Of perhaps greatest interest is the use of laminarin oligosaccharides as anti-inflammatory and
anticancer agents, relating to the potential effect of β-1,3-(1,6)-glucans. They bind primarily to the
receptor Dectin-1 [110], a type II transmembrane receptor expressed in high levels on blood and
splenic monocytes, neutrophils and alveolar and inflammatory macrophages, and at lower levels on
dendritic cells [110,111]. These properties are neither found in mixed β-1,3- and β-1,4-linked glucans,
1,6-glucans (pustulan), nor in 1,4-glucans [37]. The determinants of these important activities are,
however, not completely resolved, but they appear to relate to the size and content of β-1,6-linkages as
well as their type (branch points or kinks). β-1,3-linked oligosaccharides smaller than heptose do not
bind to Dectin-1. The smallest oligosaccharide binding to recombinant Dectin-1 is a β-1,3-d-glucan
oligomer, having a degree of polymerization of 7 (DP 7), with a single β-1,6-linked glucose unit [37].
Laminarin-derived oligosaccharides have also been shown to inhibit the proliferation of U937 leukemia
cells to a greater degree than the source laminarins from L. digitata and E. bicyclis [112]. A NMR
analysis indicated that the active β-glucan oligomers were larger than DP 8 with a ratio of 3:1
between β-1,3- and β-1,6-linkages. Oligosaccharides with a higher degree of branching derived
from E. bicyclis laminarin elicited a stronger reaction. Hreggviðsson et al. [113–115] have developed
efficient transglucosidases, which are highly promising as glucan engineering tools for making potent
bioactive laminarin oligosaccharides, as they can catalyze, depending on the enzyme, β-1,3-elongation,
β-1,6-branching or β-1,6-interchain kinking of β-1,3-glucan oligomers.

7.4. Carrageenan
There are many studies on enzymes hydrolyzing plant polysaccharides, but enzymes degrading
marine polysaccharides are not largely explored [116]. Indeed, enzymes acting on marine
polysaccharides must be adopted to accommodate the sulfate group of marine polysaccharides
which is not available in plant polysaccharides. Carrageenan is the most well-known sulfated marine
polysaccharide from red seaweed. Enzymes acting on carrageenan belong to different glycoside
hydrolase (GH) families listed in in the carbohydrate-active enzymes (CAZy) database. The enzymes
Molecules 2020, 25, 930 17 of 29

acting on κ-carrageenan (κ-carrageenase) and ι-carrageenan (ι-carrageenase) belong to the families


GH16 and GH82, respectively [117,118], while there is no specified GH family for λ-carrageenase.
κ-carrageenases are endo-acting enzymes that hydrolyze the β-1,4-linkage between
d-galactose-4-sulfate and 3,6-anhydro-d-galactose residues. The endo-hydrolase ι-carrageenase
cleaves the β-1,4-linkages between the d-galactose-4-sulfate and 3,6-anhydro-d-galactose-2-sulfate in
ι-carrageenan. λ-carrageenase is also an endo-acting enzyme hydrolyzing the glycoside linkage in
λ-carrageenan backbone-producing tetrasaccharides [119].

7.5. Agar
Agarases are enzymes that catalyze the hydrolysis of agarose. They can be intra-cellular or
extra-cellular and are isolated from different genera of bacteria available in seawater and marine
sediments, marine algae, marine mollusks, fresh water, and soil. Two types of agarase are classified
according to their cleavage pattern; α-agarase (E.C. 3.2.1.158) and β-agarase (E.C. 3.2.1.81) [120,121].
α-agarase hydrolyzes the α-1,3-linkages in agar backbone producing agarooligosaccharides (AOs) with
3,6-anhydro-l-galactopyranose as the reducing end [122], while β-agarase cleaves the β-1,4-linkages
producing neoagarooligosaccharides (NAOs) with d-galactose as the reducing end [120].
In addition to the production of oligosaccharides from agar with potential application in
pharmaceutical industries, agarases are also used for the recovery of DNA from agarose gel and
preparation of protoplasts of the seaweed to recover substances with biological activities, including
unsaturated fatty acids, vitamins, and carotenoids [120].

7.6. Ulvan
The enzymatic depolymerization mechanisms of ulvan, the major polysaccharide in green seaweed,
remain largely unknown [123]. Ulvan is degraded by ulvan lyases by β-elimination mechanisms
releasing oligosaccharides with unsaturated uronic acids at the nonreducing end. Some of the
characterized ulvan lyases are classified into the polysaccharide lyase family 24 (PL24), PL25 or PL28
in the CAZy database. To date, only a few ulvan lyases from marine bacteria have been isolated
and characterized, but have not been industrially employed further. Research on the isolation and
characterization of ulvan lyases is ongoing, but more investigation is needed to recognize the usability of
these enzymes. Additionally, the applicability of hydrolyzed products in the production of value-added
chemicals is investigated.

8. Current and Potential Novel Applications


Many coastal countries, especially Japan, Korea and China, have the culture of using macroalgae in
their food as ingredients in different soups, combined with fish or meat, or as a substitution for vegetables.
Examples of some edible macroalgae include Ulva spp., Enteromorpha spp., Caulerpa spp., Codium spp.,
Monostroma spp., Sargassum spp., Hydroclathrus spp., Laminaria spp., Undaria spp., Macrocystis spp., Porphyra
spp., Gracilaria spp., Eucheuma spp., Laurencia spp., and Acanthophora spp. [124].
Red algae have historically been consumed as feed [125], food, food ingredients and food
supplements. For example, Palmaria palmata, also called dulse or dillisk, which grows on the northern
coast lines of Atlantic and Pacific oceans, has been a part of the human diet in Iceland, Ireland and
Scotland for a long time, and its popularity is increasing in other western countries [126,127]. Porphyra
species including Porphyra tenera, Porphyra pseudolinearis and Porphyra yezoensis, also known as Nori,
are other well-known species grown mainly in Japan, the Republic of Korea, and China. These species
are reported as being among the most nutritious types of algae. They have a long history in the human
diet in many Asian countries and the consumption of these species has shown an increasing trend in
other countries as well [128,129]. In addition, red algae can have potential as a meat substitute due to
its relatively high protein content and amino acid composition [130].
In vitro studies of bioactive compounds in macroalgae have shown health promoting effects on
the human body. However, more in vivo trials are required in order to support the claims of the health
Molecules 2020, 25, 930 18 of 29

promoting effects of macroalgae. Additionally, sustainable and cost-effective pretreatment, extraction


and purification methods are needed with zero waste for industrial application [131]. Nutritional
compatibility, low production cost, constant accessibility, and enough quality are the main prerequisites
for commercialization [132]. Although seaweed as a whole product is emerging globally, extracted
polysaccharides from algae have received much research attention. This review focuses on the most
prominent polysaccharides in brown, red and green seaweeds.

8.1. Fucoidan
Fucoidan has been of interest in many applications due to its potential biomedical effects,
including anticarcinogenic, antimicrobial, anticoagulant, antiviral, immunomodulatory, and antioxidant
effects [133]. However, although the biomedical potential of fucoidan has been studied comprehensively
in vitro, and to some extent in vivo in non-human animal systems, only a few clinical studies on
humans have been performed. Studies on orally administrated fucoidan have indicated that they
are urinarily excreted. However, further studies on the absorption, distribution, metabolism and
excretion of fucoidan are required to understand its systemic effects [134]. A phase I clinical study
(ClinicalTrials.cov, NCT03422055) on tolerance, biodistribution and dosimetry of radio-labelled fucoidan
on healthy volunteers is currently ongoing. The study primarily aims to investigate the potential
adverse effects and serious adverse events after intravenous administration of the drug.
The anticarcinogenic properties of fucoidan have probably received the most attention so far and
have been studied in several in vitro systems to assess the mechanisms of action towards various types
of cancer. Several papers review the potential applications of fucoidan in cancer treatment, and these
also present which anti-cancer mechanisms have been studied to date [103,135,136]. Most studies
presented in the literature are performed in vitro or in vivo in non-human animal expression systems.
However, a phase II clinical study (ClinicalTrials.gov, NCT04066660), primarily investigating the
disease control rate of oligo-fucoidan as a dietary supplement for patients with advanced hepatocellular
carcinoma, is currently recruiting.
Although clinical evidence so far on the effectivity and bioavailability of fucoidan is scarce, several
dietary supplements are marketed as capsules, liquids and powders, with claimed indications such
as support of a healthy immune system, optimal cellular aging, a healthy digestive function, and a
healthy inflammation response.

8.2. Alginate
Alginate is currently used in a wide range of industries for applications such as feed stabilizer,
paper and welding rod coatings, and dye thickener in textile printing [77]. The polysaccharide is
also widely used in the food and drink industries due to its gelling, thickening, stabilizing and
emulsifying properties. The salts of alginic acid with monovalent cations form a viscose solution in
water. The viscosity of alginate solutions is affected by alginate concentration and molecular weight,
pH [137], and temperature [138]. Sodium alginate is not effective in acidic solutions because insoluble
alginic acid is formed. In mild acidic conditions, propylene glycol alginate (PGA) is preferred, since it
demonstrates a higher grade of stability. PGA, an emulsifier, stabilizer, and thickening agent, is an
ester form of alginate in which some of the carboxyl groups are esterified with propylene glycol, some
are neutralized with alkali, and some remain free. Alginate can form a hydrogel in the presence of
divalent cations due to the crosslinking of the carboxylate groups of G blocks in the polymer backbone,
constituting the egg-box model. This ability makes alginate a potential candidate for the production of
edible films and coatings for food packaging [139]. More information on the applications of alginate as
a food additive was published in a comprehensive review by Qin et al. [140]
In addition to its use in the chemical, food and feed industries, alginates are also used in
pharmaceutical applications. Although alginates are already utilized as excipients in drug delivery
systems, they are of interest as primary ingredients in some pharmaceutical applications. Several
clinical studies on human subjects focus on alginate applications for symptoms such as; obesity,
Molecules 2020, 25, 930 19 of 29

diabetes, dry eyes, gastroesophageal reflux, cystic fibrosis and topical disorders (clinicaltrials.gov,
search term ‘alginate’). In particular, studies linking alginate to decreased small intestinal fat absorption
and increased satiety have been conducted and have generated varying results [141–143].

8.3. Laminarin
Commercial applications of laminarin are limited, and so far the amount of research is inferior
to other polysaccharides of brown algae. However, similar to other β-1,3-(1,6)-glucans, laminarin is
reported to have a broad range of biological activities and to have similar potential for applications in
functional foods, nutraceuticals, pharmaceuticals, and cosmeceuticals [144]. Laminarin from brown
seaweed is used in agricultural applications where it has been shown to limit pathogen infections by
stimulating plant defense systems [145], and is commercialized as Vacciplant® [146]. Additionally,
unprocessed marine macroalgae have historically been used as agricultural nourishment and are today
commercialized as organic liquid fertilizer.
Since laminarin does not possess any chelating properties, its primary potential emerges within
medical and nutraceutical applications. It is suggested that laminarin has potential applicability as a
dietary fiber [33,147]. Dietary fibers, divided into soluble and insoluble fibers, are carbohydrates that
cannot be degraded by the endogenous enzymes of the body and are either fermented in the colon or
provide bulking. Laminarin from S. latissima has been demonstrated to evade hydrolysis in the upper
intestinal tract, while not being excreted with the feces of rat models, suggesting fermentation in the
colon. Laminarin has been shown to exert beneficial metabolic effects on gut health as a modulator
of intestinal metabolism through its effects on mucus composition, intestinal pH, and short-chain
fatty acid (SCFA) production, especially butyrate, by the gut microbiota [148]. However, although
studies suggest the prebiotic effects of laminarin, further studies on the digestibility, bioavailability
and biodistribution are required to better understand the systemic effects of the compound.
Laminarin may also be used as an immunomodulating and anti-cancer agent. These effects
are observed in related glucans and can be attributed to specific glycosidic β-1,3-(1,6)-linkage
patterns [149–151] that modulate the immune response [152] and inhibit cell proliferation in HT-29
colon cancer cells. On the basis of such properties, derivatives of insoluble β-1,3-(1,6)-glucans from yeast
are currently being developed as ‘imprime PGG’ by the company Biothera as intravenous agents against
cancer, affecting the innate immune system. Dietary products (Yestimun) developed from unmodified
yeast β-1,3-(1,6)-glucan and based on its immunomodulatory effects [153] are also commercialized.

8.4. Carrageenan
Carrageenan is used in a wide variety of applications in the food industry; mainly in dairy and
meat applications, based on its physiochemical properties as a thickening, gelling, and stabilizing agent.
Various types of carrageenan can show a broad range of gelling and emulsifying properties, such
as rigidity or elasticity, transparency or turbidity, toughness or tenderness, heat stability or thermal
reversibility, as well as low or high melting/gelling temperatures. These physiochemical properties are
affected by the chemical structure of carrageenan, the type and concentration of cations including salts
and proteins present in the solution, and the heating and cooling profile of the carrageenan solution.
In fact, ι-carrageenan can form an elastic gel in the presence of calcium salt, while κ-carrageenan forms
a brittle gel in the presence of calcium but forms a strong and rigid gel with potassium. On the other
hand, λ-carrageenan is not gel-forming in the presence of divalent salts, but develops a highly viscous
solution [154,155]. However, it exhibits gelation in the presence of trivalent ions [41].
Carrageenan has potential applications in the pharmaceutical industry because of its biological
activities including anticoagulant [156], antiviral [157,158], cholesterol-lowering [159], anti-tumor
and immunomodulatory [160–162], and antioxidant activities [163,164]. Degradation of carrageenan
polysaccharides into smaller compound structures could be a way of controlling the bioactivity.
Although oligosaccharides derived from carrageenan have reported antitumor [165], antioxidant
activity [166] and immune regulation activities [167], which make them very valuable in the
Molecules 2020, 25, 930 20 of 29

pharmaceutical industry, carrageenan also has various applications in bioethanol production, in


the textile industry, as a detergent additive and in the isolation of the protoplast of algae [168].
Recent applications of carrageenan in the pharmaceutical industry were summarized by
Li et al. [169], including as a polymer matrix in oral extended-release tablets, as a novel extrusion
aid for the production of pellets, and as a carrier/stabilizer in microparticle/nanoparticle systems.
Moreover, carrageenan is used in the production of nanoengineered injectable hydrogels in tissue
regeneration therapy [170]. Other potential applications of carrageenan include utilization as fertilizer,
and studies by Vera et al. [171] showed that oligo-carrageenan induces a broad range of protection
against pathogens in plants.

8.5. Agar
Agar is widely used as a gelling agent due to its unique physiochemical properties which enable
gel formation in water at temperatures around 38 ◦ C and facilitate melting at temperatures around
85 ◦ C [172]. In addition to food application, including bakery, confectionery, dairy products, canned
meat and fish products [173], agar is used in the preparation of bacteriological culture media [174].
Agarose, a fraction of agar with a lowest possible charge, is used for electrophoretic separation
in agarose gel electrophoresis [175], the preparation of porous beads for chromatographic protein
purifications, and mobility assay [176,177]. Agarose is also used as a support for the three-dimensional
cultures of human and animal cells [178,179].

8.6. Ulvan
Sulfated polysaccharides from marine seaweeds have recently received attention in the food,
cosmetics and pharmacology areas. Sulfated polysaccharides can originate from red (carrageenans) or
brown (fucoidans) macroalgae, but are also present in the form of ulvans in green seaweeds.
Ulvan has been reported to mimic glycosaminoglycans in animals, and hence mimic these
types of activities. For this purpose, sulfation is proposed as important [27], with the degree of
sulfation and sulfation patterns influencing the bioactivity. In addition, the molecular weight,
constituent sugars linkages and degree of branching are suggested as factors that influence bioactivity.
The bioactivities reported from green seaweeds, include antioxidant activity, shown from Ulva
pertusa [180], immunomodulating activity (from sulfated polysaccharides of e.g., Monostroma nitidum
and U. pertusa [181,182], antihyperlipidemic activity (reported from U. pertusa) [183], anticoagulant
activity from M. nitidum and anticancer from, for example, M. nitidum [181,184].
Recently, ulvans have also been found to influence plant immunity, including stimulation of the
inducible defenses involving recognition of microbes/pathogens. In this field, the sugar constituents
are reported as important, and rhamnose has for instance been shown to be important for triggering
plant defense in several different plants, including apple leaves, tomatoes, and thale cress (Arabidopsis
thaliana), as reviewed by Kidgell et al. [27].

9. Conclusions and Future Aspects


Although polysaccharides from marine macroalgae have comprehensively reported potential
in valuable biomedical applications, commercial products are scarce on the market. Many of the
numerous bioactivities reported are still only at research level, and more work remains before reaching
mature application. Today, marine macroalgal polysaccharides are mainly used industrially for their
distinctive physiochemical properties as additives or excipients. It is obvious that the extraction
processes of polysaccharides are crucial for the properties of the extractive. Strong acids and bases
can exert uncontrolled chemical degradation and introduce unwanted structural changes that alter
the bioactive properties of the polysaccharides. Yet, novel extraction methods are not optimized
for an industrial scale. Additionally, extraction yields are sometimes remarkably low, and require
improvements for industrial realization.
Molecules 2020, 25, 930 21 of 29

Polysaccharides exert a large structural variation depending on factors such as algae species,
harvest season, and growth geography, as well as extraction and purification methods, which is
problematic for commercialization. A few products from macroalgal polysaccharides are currently
marketed, although clinical evidence is scarce. One of the bottlenecks in commercializing the
bioactivities lies in the complex structure of the glycans, and enzymatic deconstruction into more
defined small units may be a way of achieving better defined molecules which are more suitable for
the applications. Enzymatic treatment, as mentioned, can be a useful tool for necessary modification of
the compound structure to achieve structures with bioactivity in humans. However, better knowledge
about structures and activities of both enzymes and bio-compounds are crucial.

Author Contributions: Conceptualization, E.N.K. and M.J.; investigation, E.N.K, L.A., M.J., and R.R.R.S.;
writing—original draft preparation, E.N.K, L.A., M.J., and R.R.R.S.; writing—review and editing, E.N.K, G.O.H.,
L.A., M.J., and R.R.R.S.; visualization, M.J.; funding acquisition, E.N.K. All authors have read and agreed to the
published version of the manuscript.
Funding: The authors wish to give thanks for the financial support for the projects Marine food resources for new
markets (Formas, grant 2018-01863), Macro cascade (Bio-Based Industries Joint Undertaking under EU Horizon
2020, grant agreement No 720755), ProSeaFood (Era-net SusFood2, grant agreement No 727473), and NordMar
Biorefine (Nordic Council of Ministers).
Conflicts of Interest: The authors declare no conflict of interest.

References
1. Chapman, V.J.; Chapman, D.J. Seaweeds and their Uses, 3rd ed.; Chapman and Hall Ltd.: London, UK, 1980.
[CrossRef]
2. FAO. FAO yearbook. Fishery and Aquaculture Statistics 2017; FAO: Rome, Italy, 2019.
3. Campbell, I.; Macleod, A.; Sahlmann, C.; Neves, L.; Funderud, J.; Øverland, M.; Hughes, A.D.; Stanley, M.
The Environmental Risks Associated With the Development of Seaweed Farming in Europe—Prioritizing
Key Knowledge Gaps. Front. Mar. Sci. 2019, 6. [CrossRef]
4. FAO. The Global Status of Seaweed Production, Trade and Utilization; Globefish Research Programme: Rome,
Italy, 2018; Volume 124, p. 120.
5. Jung, K.A.; Lim, S.-R.; Kim, Y.; Park, J.M. Potentials of macroalgae as feedstocks for biorefinery.
Bioresour. Technol. 2013, 135, 182–190. [CrossRef]
6. Field, C.B.; Behrenfeld, M.J.; Randerson, J.T.; Falkowski, P. Primary Production of the Biosphere: Integrating
Terrestrial and Oceanic Components. Science 1998, 281, 237–240. [CrossRef] [PubMed]
7. Harrysson, H.; Hayes, M.; Eimer, F.; Carlsson, N.-G.; Toth, G.; Undeland, I. Production of protein extracts
from Swedish red, green and brown seaweeds, Porphyra umbilicalis Kützing, Ulva lactuca Linneus, and
Saccharina latissima (Linneus), J.V. Lamoroux, using three different methods. J. Appl. Phycol. 2018, 30,
3565–3580. [CrossRef]
8. Hreggviðsson, G.O.; Nordberg-Karlsson, E.M.; Tøndervik, A.; Aachmanne, F.L.; Dobruchowska, J.M.;
Linares-Pastén, J.; Daugbjerg-Christensen, M.; Moneart, A.; Kristjansdottir, T.; Slettad, H.; et al. Biocatalytic
refining of polysaccharides from brown seaweeds. In Sustainable Seaweed Technologies—Cultivation,
Biorefinery, and Applications, 1st ed.; Torres, M.D., Kraan, S., Dominguez, H., Eds.; Elsevier: Amsterdam,
The Netherlands, 2020.
9. Milledge, J.J.; Harvey, P.J. Anaerobic Digestion and Gasification of Seaweed. In Grand Challenges in Marine
Biotechnology; Rampelotto, P.H., Trincone, A., Eds.; Springer: Cham, Switzerland, 2018; pp. 237–258.
[CrossRef]
10. Vijayaraghavan, M.R.; Sokhi, G. Phaeophyceae—An Ultrastructural and Histochemical Overview. Proc. Indian.
Natn. Sci. Acad. 1986, 4, 529–546.
11. Mautner, H.G. The Chemistry of Brown Algae. Econ. Bot. 1954, 8, 174–192. [CrossRef]
12. Deniaud-Bouët, E.; Kervarec, N.; Michel, G.; Tonon, T.; Kloareg, B.; Hervé, C. Chemical and enzymatic
fractionation of cell walls from Fucales: Insights into the structure of the extracellular matrix of brown algae.
Ann. Bot. 2014, 114, 1203–1216. [CrossRef]
Molecules 2020, 25, 930 22 of 29

13. Deniaud-Bouët, E.; Hardouin, K.; Potin, P.; Kloareg, B.; Hervé, C. A review about brown algal cell walls
and fucose-containing sulfated polysaccharides: Cell wall context, biomedical properties and key research
challenges. Carbohyd. Polym. 2017, 175, 395–408. [CrossRef]
14. Graiff, A.; Ruth, W.; Kragl, U.; Karsten, U. Chemical characterization and quantification of the brown algal
storage compound laminarin - A new methodological approach. J. Appl. Phycol. 2016, 28, 533–543. [CrossRef]
15. Kloareg, B.; Quatrano, R.S. Structure of the cell walls of marine algae and ecophysiological functions of the
matrix polysaccharides. Oceanogr. Mar. Biol. Annu. Rev. 1988, 26, 259–315.
16. Salmeán, A.A.; Duffieux, D.; Harholt, J.; Qin, F.; Michel, G.; Czjzek, M.; Hervé, C. Insoluble (1→3),
(1→4)-β-D-glucan is a component of cell walls in brown algae (Phaeophyceae) and is masked by alginates in
tissues. Sci. Rep. 2017, 7. [CrossRef] [PubMed]
17. Lim, S.J.; Mustapha, W.A.W.; Maskat, M.Y.; Latip, J.; Badri, K.H.; Hassan, O. Chemical Properties and
Toxicology Studies of Fucoidan Extracted from Malaysian Sargassum binderi. Food Sci. Biotechnol. 2016, 25,
23–29. [CrossRef] [PubMed]
18. Ale, M.T.; Mikkelsen, J.D.; Meyer, A.S. Important Determinants for Fucoidan Bioactivity: A Critical Review
of Structure-Function Relations and Extraction Methods for Fucose-Containing Sulfated Polysaccharides
from Brown Seaweeds. Mar. Drugs 2011, 9, 2106–2130. [CrossRef] [PubMed]
19. Jiao, G.; Yu, G.; Zhang, J.; Ewart, H.S. Chemical Structures and Bioactivities of Sulfated Polysaccharides from
Marine Algae. Mar. Drugs 2011, 9, 196–223. [CrossRef] [PubMed]
20. Fletcher, H.R.; Biller, P.; Ross, A.B.; Adams, J.M.M. The seasonal variation of fucoidan within three species of
brown macroalgae. Algal Res. 2017, 22, 79–86. [CrossRef]
21. Starko, S.; Mansfield, S.D.; Martone, T. Cell wall chemistry and tissue structure underlie shifts in material
properties of a perennial kelp. Eur. J. Phycol. 2018, 53, 307–317. [CrossRef]
22. Synytsya, A.; Čopíková, J.; Kim, W.J.; Park, Y.I. Cell Wall Polysaccharides of Marine Algae. In Springer
Handbook of Marine Biotechnology; Kim, S.-K., Ed.; Springer: Berlin, Germany, 2015; pp. 543–590.
23. Zhao, X.; Jiao, G.; Wu, J.; Zhang, J.; Yu, G. Laminaria japonica Aresch. and Ecklonia Kurome Okam. 昆布
(Kunbu, Kelp). In Dietary Chinese Herbs; Liu, Y., Wang, Z., Zhang, J., Eds.; Springer: Vienna, Austria, 2015.
24. Campo, V.L.; Kawano, D.F.; da Silva, D.B., Jr.; Carvalho, I. Carrageenans: Biological properties, chemical
modifications and structural analysis—A review. Carbohyd. Polym. 2009, 77, 167–180. [CrossRef]
25. Tuvikene, R.; Truus, K.; Robal, M.; Volobujeva, O.; Mellikov, E.; Pehk, T.; Kollist, A.; Kailas, T.; Vaher, M.
The extraction, structure, and gelling properties of hybrid galactan from the red alga Furcellaria lumbricalis
(Baltic Sea, Estonia). J. Appl. Phycol. 2010, 22, 51–63. [CrossRef]
26. Pérez, M.J.; Falqué, E.; Domínguez, H. Antimicrobial Action of Compounds from Marine Seaweed. Mar. Drugs
2016, 14, 52. [CrossRef]
27. Kidgell, J.T.; Magnusson, M.; deNys, R.; Glasson, R.K. Ulvan: A systematic review on extraction, composition
and function. Algal Res. 2019, 39. [CrossRef]
28. Zhao, X.; Li, B.; Xue, C.; Sun, L. Effect of molecular weight on the antioxidant property of low molecular
weight alginate from Laminaria japonica. J. Appl. Phycol. 2012, 24, 295–300. [CrossRef]
29. Brownlee, I.A.; Seal, C.J.; Wilcox, M.; Dettmar, P.W.; Pearson, J.P. Applications of Alginates in Food.
In Alginates: Biology and Applications; Rehm, B.H.A., Ed.; Springer-Verlag Berlin: Heidelberg, Germany, 2009;
pp. 211–228.
30. Miller, I.J. Alginate Composition of Some New Zealand Brown Seaweeds. Phytochemistry 1996, 41, 1315–1317.
[CrossRef]
31. Skjåk-Bræk, G.; Donati, I.; Paoletti, S. Alginate hydrogels: Properties and applications. In Polysaccharide
Hydrogels: Characterization and Biomedical Applications; Matricardi, P., Alhaique, F., Coviello, T., Eds.;
Pan Stanford Publishing Pte Ltd.: Singapore, 2015.
32. Domozych, D. Algal Cell Wall. In Encyclopedia of Life Sciences, 4th ed.; John Wieley & Sons Ltd.: Chichester,
UK, 2019; pp. 1–11.
33. Devillé, C.; Damas, J.; Forget, P.; Dandrifosse, G.; Peulen, O. Laminarin in the dietary fibre concept. J. Sci.
Food Agric. 2004, 84, 1030–1038. [CrossRef]
34. Kadam, S.U.; Tiwari, B.K.; O’Donnell, C.P. Extraction, structure and biofunctional activities of laminarin
from brown algae. Int. J. Food Sci. Tech. 2014, 50, 24–31. [CrossRef]
35. Read, S.M.; Currie, G.; Bacic, A. Analysis of the structural heterogeneity of laminarin by electrospray-
ionisation-mass spectrometry. Carbohydr. Res. 1996, 23, 187–201. [CrossRef]
Molecules 2020, 25, 930 23 of 29

36. Maeda, M.; Nisizawa, K. Fine structure of laminaran of Eisenia bicyclis. J. Biochem. 1968, 63, 199–206.
[CrossRef]
37. Adams, E.L.; Rice, P.J.; Graves, B.; Ensley, H.E.; Yu, H.; Brown, G.D.; Gordon, S.; Monteiro, M.A.;
Papp-Szabo, E.; Lowman, D.W.; et al. Differential high-affinity interaction of dectin-1 with natural or
synthetic glucans is dependent upon primary structure and is influenced by polymer chain length and
side-chain branching. J. Pharmacol. Exp. Ther. 2008, 325, 115–123. [CrossRef]
38. Nelson, T.E.; Lewis, B.A. Separation and characterization of the soluble and insoluble components of insoluble
laminaran. Carbohydr. Res. 1974, 33, 63–74. [CrossRef]
39. Viola, R.; Nyvall, P.; Pedersén, M. The unique features of starch metabolism in red algae. Proc. Biol. Sci. 2001,
268, 1417–1422. [CrossRef]
40. Amimi, A.; Mouradi, A.; Givernaud, T.; Chiadmi, N.; Lahaye, M. Structural analysis of Gigartina pistillata
carrageenans (Gigartinaceae, Rhodophyta). Carbohyd. Res. 2001, 333, 271–279. [CrossRef]
41. Running, C.A.; Falshaw, R.; Janaswamy, S. Trivalent iron induced gelation in lambda-carrageenan.
Carbohyd. Polym. 2012, 87, 2735–2739. [CrossRef] [PubMed]
42. Manuhara, G.J.; Praseptiangga, D.; Riyanto, R.A. Extraction and Characterization of Refined K-carrageenan
of Red Algae [Kappaphycus Alvarezii (Doty ex P.C. Silva, 1996)] Originated from Karimun Jawa Islands.
Aquat. Procedia 2016, 7, 106–111. [CrossRef]
43. Lebbar, S.; Fanuel, M.; Le Gall, S.; Falourd, X.; Ropartz, D.; Bressollier, P.; Gloaguen, V.; Faugeron-Girard, C.
Agar Extraction By-Products from Gelidium sesquipedale as a Source of Glycerol-Galactosides. Molecules 2018,
23, 3364. [CrossRef] [PubMed]
44. Lee, W.-K.; Lim, Y.-Y.; Leow, A.T.-C.; Namasivayam, P.; Ong Abdullah, J.; Ho, C.-L. Biosynthesis of agar in
red seaweeds: A review. Carbohyd. Polym. 2017, 164, 23–30. [CrossRef]
45. Leliaert, F.; Smith, D.R.; Moreau, H.; Herron, M.D.; Verbruggen, H.; Delwiche, C.F.; De Clerck, O. Phylogeny
and Molecular Evolution of the Green Algae. Crit. Rev. Plant. Sci. 2012, 31, 1–46. [CrossRef]
46. Sardari, R.R.R.; Nordberg Karlsson, E. Marine Poly- and Oligosaccharides as Prebiotics. J. Agric. Food Chem.
2018, 66, 11544–11549. [CrossRef]
47. Wichard, T.; Charrier, B.; Mineur, F.; Bothwell, J.H.; Clerck, O.D.; Coates, J.C. The green seaweed Ulva:
A model system to study morphogenesis. Front. Plant. Sci. 2015, 6. [CrossRef]
48. Mine, I.; Menzel, D.; Okuda, K. Morphogenesis in giant-celled algae. Int. Rev. Cell Mol. Biol. 2008, 266, 37–83.
[CrossRef]
49. Cocquyt, E.; Verbruggen, H.; Leliaert, F.; De Clerck, O. Evolution and Cytological Diversification of the Green
Seaweeds (Ulvophyceae). Mol. Biol. Evol. 2010, 27, 2052–2061. [CrossRef]
50. Cho, M.L.; You, S.G. Sulfated polysaccharides from green seaweeds. In Springer Handbook of Marine
Biotechnology; Kim, S.-K., Ed.; Springer: Berlin/Heidelberg, Germany, 2015; pp. 941–953. [CrossRef]
51. Zhang, M.; Zhao, C.; Shao, Q.; Yang, Z.; Zhang, X.; Xu, X.; Hassan, M. Determination of water content in
corn stover silage using near-infrared spectroscopy. Int. J. Agric. Biol. Eng. 2019, 12, 143–148. [CrossRef]
52. Maneein, S.; Milledge, J.J.; Vejby Nielsen, B.; Harvey, P. A Review of Seaweed Pre-Treatment Methods for
Enhanced Biofuel Production by Anaerobic Digestion or Fermentation. Fermentation 2018, 4, 100. [CrossRef]
53. Zhao, C.; Cao, Y.; Ma, Z.; Shao, Q. Optimization of liquid ammonia pretreatment conditions for maximizing
sugar release from giant reed (Arundo donax L.). Biomass Bioenerg. 2017, 98, 61–69. [CrossRef]
54. Kadam, S.U.; Álvares, C.; Twari, B.K.; O’Donnell, C.P. Extraction of biomolecules from seaweeds. In Seaweed
Sustainability—Food and Non-Food Applications; Tiwari, B.K., Troy, D.J., Eds.; Academic Press: San Jose, CA,
USA, 2015; Volume 1, pp. 243–269.
55. Sosa-Hernández, J.E.; Escobedo-Avellaneda, Z.; Iqbal, H.M.N.; Welti-Chanes, J. State-of-the-Art Extraction
Methodologies for Bioactive Compounds from Algal Biome to Meet Bio-Economy Challenges and
Opportunities. Molecules 2018, 23, 2953. [CrossRef] [PubMed]
56. Shiroma, R.; Uechi, S.; Taira, T.; Ishihara, M.; Tawata, S.; Tako, M. Isolation and Characterization of Fucoidan
from Hizikia fusiformis (Hijiki). J. Appl. Glycosci. 2003, 50, 361–365. [CrossRef]
57. Kim, W.-J.; Kim, S.-M.; Kim, H.G.; Oh, H.-R.; Lee, K.-B.; Lee, Y.-K.; Park, Y.-I. Purification and Anticoagulant
Activity of a Fucoidan from Korean Undaria pinnatifida Sporophyll. Algae 2007, 22, 247–252. [CrossRef]
58. Hahn, T.; Lang, S.; Ulber, R.; Muffler, K. Novel procedures for the extraction of fucoidan from brown algae.
Process. Biochem. 2012, 47, 1691–1698. [CrossRef]
Molecules 2020, 25, 930 24 of 29

59. Wang, C.-Y.; Chen, Y.-C. Extraction and characterization of fucoidan from six brown macroalgae. J. Mar.
Sci. Technol. 2016, 24, 319–328. [CrossRef]
60. Liu, X.; Liu, B.; Wei, X.L.; Sun, Z.L.; Wang, C.Y. Extraction, Fractionation, and Chemical Characterisation of
Fucoidans from the Brown Seaweed Sargassum pallidum. Czech. J. Food Sci. 2016, 34, 406–413. [CrossRef]
61. Khalil, H.; Lai, T.; Tye, Y.; Rizal, S.; Chong, E.; Yap, S.; Hamzah, A.; Fazita, M.; Paridah, M. A review of
extractions of seaweed hydrocolloids: Properties and applications. Express Polym. Lett. 2018, 12. [CrossRef]
62. Li, C.; Wang, C.; Wang, S.; Qian, G.; Zhu, Q.; Liu, Y.; Wang, W. Optimization of Ultrasonic-assisted Extraction
Technology of Sargassum fusiforme Polysaccharides and Evaluation of Their Antioxidant Activity. Food Sci.
Technol. Res. 2013, 19, 157–162. [CrossRef]
63. Rafiquzzaman, S.M.; Rahman, A.; Kong, I.-S. Ultrasonic-Assisted Extraction of Carrageenan. Seaweed
Polysacch. 2017, 75–81. [CrossRef]
64. Rodriguez-Jasso, R.M.; Mussatto, S.I.; Pastrana, L.; Aguilar, C.N.; Teixeira, J.A. Microwave-assisted extraction
of sulfated polysaccharides (fucoidan) from brown seaweed. Carbohyd. Polym. 2011, 86, 1137–1144. [CrossRef]
65. Yuan, Y.; Macquarrie, D. Microwave assisted extraction of sulfated polysaccharides (fucoidan) from
Ascophyllum nodosum and its antioxidant activity. Carbohyd. Polym. 2015, 129, 101–107. [CrossRef] [PubMed]
66. Lakmal, H.H.C.; Lee, J.H.; Jeon, Y.J. Enzyme-Assisted Extraction of a Marine Algal Polysaccharide, Fucoidan
and Bioactivities. In Polysaccharides - Bioactivity and Biotechnology; Ramawat, K.G., Mérillon, J.-M., Eds.;
Springer: Cham, Switzerland, 2015; pp. 1065–1077. [CrossRef]
67. Aðalbjörnsson, B.V.; Jónsdóttir, R. Enzyme-Enhanced Extraction of Antioxidant Ingredients from Algae.
In Natural Products from Marine Algae—Methods in Molecular Biology; Humana Press: New York, NY, USA,
2015; pp. 145–150.
68. Habeebullah, S.F.K.; Alagarsamy, S.; Sattari, Z.; Al-Haddad, S.; Fakhraldeen, S.; Al-Ghunaim, A.;
Al-Yamani, F. Enzyme-assisted extraction of bioactive compounds from brown seaweeds and characterization.
J. Appl. Phycol. 2019. [CrossRef]
69. Puri, M.; Sharma, D.; Barrow, C.J. Enzyme-assisted extracted of bioactives from plants. Trends Biotechnol.
2012, 30, 37–44. [CrossRef] [PubMed]
70. Michalak, I.; Górka, B.; Wieczorek, P.P.; Rój, E.; Lipok, J.; Łeska, B.; Messayasz, B.; Wilk, R.; Schroeder, G.;
Dobrzy’nska-Inger, A.; et al. Supercriticalfluid extraction of algae enhances levels ofbiologically active
compounds promoting plant growth. Eur. J. Phycol. 2016, 51, 243–252. [CrossRef]
71. Messyasz, B.; Michalak, I.; Ł˛eska, B.; Schroeder, G.; Górka, B.; Korzeniowska, K.; Lipok, J.; Wieczorek, P.;
Rój, E.; Wilk, R.; et al. Valuable natural products from marine and freshwater macroalgae obtained from
supercritical fluid extracts. J. Appl. Phycol. 2018, 30, 591–603. [CrossRef]
72. Espinosa-Pardo, F.A.; Martinez, J.; Martinez-Corre, H.A. Extraction of bioactive compounds from peach
palm pulp (Bactris gasipaes) using supercritical CO2 . J. Supercrit. Fluid. 2014, 93, 2–6. [CrossRef]
73. Rioux, L.-E.; Turgeon, S.L.; Beaulieu, M. Characterization of polysaccharides extracted from brown seaweeds.
Carbohyd. Polym. 2007, 69, 530–537. [CrossRef]
74. Rani, V.; Shakila, R.J.; Jawahar, P.; Srinivasan, A. Influence of Species, Geographic Location, Seasonal Variation
and Extraction Method on the Fucoidan Yield of the Brown Seaweeds of Gulf of Mannar, India. Indian J.
Pharm. Sci. 2017, 79, 65–71. [CrossRef]
75. Ale, M.T.; Meyer, A.S. Fucoidans from brown seaweeds: an update on structures, extraction techniques and
use of enzymes as tools for structural elucidation. RSC Adv. 2013, 3. [CrossRef]
76. Kimica. About Alginate—Manufacturing Process. Available online: https://kimica-algin.com/alginate/
process/ (accessed on 10 December 2019).
77. McHugh, D.J. Alginate. In A Guide to the Seaweed Industry; FAO: Rome, Italy, 2003.
78. Mazumder, A.; Løvstad Holdt, S.; De Francisci, D.; Alvarado-Morales, M.; Mishra, H.N.; Angelidaki, I.
Extraction of alginate from Sargassum muticum: process optimization and study of its functional activities.
J. Appl. Phycol. 2016. [CrossRef]
79. Fertah, M.; Belfkira, A.; Dahmane, E.M.; Taourirte, M.; Brouillette, F. Extraction and characterization of
sodium alginate from Moroccan Laminaria digitata brown seaweed. Arab. J. Chem. 2017, 10, 3707–3714.
[CrossRef]
80. Alboofetileh, M.; Rezaei, M.; Tabarsa, M.; You, S. Ultrasound-assisted extraction of sulfated polysaccharide
from Nizamuddinia zanardinii: Process optimization, structural characterization, and biological properties.
J. Food Process. Eng. 2018, 42, 1–13. [CrossRef]
Molecules 2020, 25, 930 25 of 29

81. Hmelkov, A.B.; Zvyagintseva, T.N.; Shevchenko, N.M.; Rasin, A.B.; Ermakova, S.P. Ultrasound-assisted
extraction of polysaccharides from brown alga Fucus evanescens. Structure and biological activity of the new
fucoidan fractions. J. Appl. Phycol. 2018, 30, 2039–2046. [CrossRef]
82. Alboofetileh, M.; Rezaei, M.; Tabarsa, M. Enzyme-assisted extraction of Nizamuddinia zanardinii for the
recovery of sulfated polysaccharides with anticancer and immune-enhancing activities. J. Appl. Phycol. 2019,
31, 1391–1402. [CrossRef]
83. Je, J.-Y.; Park, P.-J.; Kim, E.-K.; Park, J.-S.; Yoon, H.-D.; Kim, K.-R.; Ahn, C.-B. Antioxidant activity of enzymatic
extracts from the brown seaweed Undaria pinnatifida by electron spin resonance spectroscopy. LWT-Food
Sci. Technol. 2009, 42, 874–878. [CrossRef]
84. Men’shova, R.V.; Lepeshkin, F.D.; Ermakova, S.P.; Pokrovskii, O.I.; Zvyagintseva, T.N. Effect of pretreatment
conditions of brown algae by supercritical fluids on yield and structural characteristics of fucoidans.
Chem. Nat. Compd. 2013, 48, 923–926. [CrossRef]
85. Youssouf, L.; Lallemand, L.; Giraud, P.; Soulé, F.; Bhaw-Luximon, A.; Meilhac, O.; Lefèbvre D’Hellencourt, C.;
Jhurry, D.; Couprie, J. Ultrasound-assisted extraction and structural characterization by NMR of alginates
and carrageenans from seaweeds. Carbohyd. Polym. 2017, 166, 55–63. [CrossRef]
86. Li, H.; Yu, X.; Jin, Y.; Zhang, W.; Liu, Y. Development of an eco-friendly agar extraction technique from the
red seaweed Gracilaria lemaneiformis. Bioresour. Technol. 2008, 99, 3301–3305. [CrossRef]
87. Luz Arvizu-Higuera, D.; Rodriguez-Montesinos, Y.E.; Murillo-Alvarez, J.I.; Munoz-Ochoa, M.;
Hernandez-Carmona, G. Effect of alkali treatment time and extraction time on agar from Gracilaria
vermiculophylla. In Nineteenth International Seaweed Symposium; Borowitzka, M.A., Critchley, A.T., Kraan, S.,
Peters, A., Sjøtun, K., Notoya, M., Eds.; Springer: Dordrecht, The Netherlands, 2007; Volume 20, pp. 65–69.
88. Distantina, S.; Wiratni, W.; Fahrurrozi, M.; Rochmadi, R. Carrageenan properties extracted from Eucheuma
cottonii, Indonesia. World Acad. Sci. Eng. Technol. 2011, 78, 738–742.
89. Tuvikene, R.; Truus, K.; Vaher, M.; Kailas, T.; Martin, G.; Kersen, P. Extraction and quantification of hybrid
carrageenans from the biomass of the red algae Furcellaria lumbricalis and Coccotylus truncatus. Proc. Estonian
Acad. Sci. Chem. 2006, 55, 40–53.
90. Blanco-Pascual, N.; Alemán, A.; Gómez-Guillén, M.C.; Montero, M.P. Enzyme-assisted extraction of κ/ι-hybrid
carrageenan from Mastocarpus stellatus for obtaining bioactive ingredients and their application for edible
active film development. Food Funct. 2014, 5, 319–329. [CrossRef] [PubMed]
91. Mustapha, S.; Chandar, H.; Abidin, Z.Z.; Saghravani, R.; Harun, M.Y. Production of semi-refined carrageenan
from Eucheuma cotonii. J. Sci. Ind. Res. 2011, 70, 865–870.
92. Freile-Pelegrín, Y.; Robledo, D. Carrageenan of Eucheuma isiforme (Solieriaceae, Rhodophyta) from Nicaragua.
J. Appl. Phycol. 2008, 20, 537–541. [CrossRef]
93. Webber, V.; Matos de Carvalho, S.; Ogliari, P.J.; Hayashi, L.; Barreto, P.L.M. Optimization of the extraction
of carrageenan from Kappaphycus alvarezii using response surface methodology. Food Sci. Technol. 2012, 32,
812–818. [CrossRef]
94. Bono, A.; Anisuzzaman, S.M.; Ding, O.W. Effect of process conditions on the gel viscosity and gel strength of
semi-refined carrageenan (SRC) produced from seaweed (Kappaphycus alvarezii). J. King Saud Univ. Sci. 2014,
26, 3–9. [CrossRef]
95. Hernández-Carmona, G.; Freile-Pelegrín, Y.; Hernández-Garibay, E. Conventional and alternative
technologies for the extraction of algal polysaccharides. In Functional Ingredients from Algae for Foods
and Nutraceuticals; Domínguez, H., Ed.; Woodhead Publishing: Sawston, UK, 2013; pp. 475–516. [CrossRef]
96. Heriyanto, H.; Kustiningsih, I.; Sari, D.K. The effect of temperature and time of extraction on the quality of
Semi Refined Carrageenan (SRC). MATEC Web Conf. 2018, 154, 01034. [CrossRef]
97. Rao, A.V.; Bekheet, I.A. Preparation of agar-agar from the red seaweed Pterocladia capillacea off the coast of
Alexandria, Egypt. Appl. Environ. Microbiol. 1976, 32, 479–482. [CrossRef]
98. Torres, M.D.; Flórez-Fernández, N.; Domínguez, H. Integral Utilization of Read Seaweed for Bioactive
Production. Mar. Drugs 2019, 17, 314. [CrossRef]
99. Circuncisão, A.R.; Catarino, M.D.; Cardoso, S.M.; Silva, A.M.S. Minerals from Macroalgae Origin: Health
Benefits and Risks for Consumer. Mar. Drugs 2018, 16, 400. [CrossRef] [PubMed]
100. Mišurcová, L.; Machů, L.; Orsavová, J. Seaweed minerals as nutraceuticals. Adv. Food Nutr. Res. 2011, 64,
371–390. [CrossRef] [PubMed]
Molecules 2020, 25, 930 26 of 29

101. Wei, M.; Wanibuchi, H.; Morimura, K.; Iwai, S.; Yoshida, K.; Endo, G.; Nakae, D.; Fukushima, S.
Carcinogenicity of dimethylarsinic acid in male F344 rats and genetic alterations in induced urinary
bladder tumors. Carcinogenesis 2002, 23, 1387–1397. [CrossRef] [PubMed]
102. Yamamoto, S.; Konishi, Y.; Matsuda, T.; Murai, T.; Shibata, M.A.; Matsui-Yuasa, I.; Otani, S.; Kuroda, K.;
Endo, G.; Fukushima, S. Cancer induction by an organic arsenic compound, dimethylarsinic acid (cacodylic
acid), in F344/DuCrj rats after pretreatment with five carcinogens. Cancer Res. 1995, 55, 1271–1276.
103. van Weelden, G.; Bobinski, M.; Okła, K.; van Weelden, W.J.; Romano, A.; Pijnenborg, J.M.A. Fucoidan
Structure and Activity in Relation to Anti-Cancer Mechanisms. Mar. Drugs 2018, 17, 32. [CrossRef]
104. Helbert, W. Marine Polysaccharide Sulfatases. Front. Mar. Sci. 2017, 4, 1–10. [CrossRef]
105. Hanson, S.R.; Best, M.D.; Wong, C.-H. Sulfatases: Structure, Mechanism, Biological Activity, Inhibition, and
Synthetic Utility. Angew. Chem. Int. Ed. Engl. 2004, 43, 5736–5763. [CrossRef]
106. Ertesvåg, H. Alginate-modifying enzymes: biological roles and biotechnological uses. Front Microbiol. 2015,
6. [CrossRef]
107. Szekalska, M.; Puciłowska, A.; Szymańska, E.; Ciosek, P.; Winnicka, K. Alginate: Current Use and Future
Perspectives in Pharmaceutical and Biomedical Applications. Int. J. Polym. Sci. 2016, 1–17. [CrossRef]
108. Holdt, S.L.; Kraan, S. Bioactive compounds in seaweed: functional food applications and legislation.
J. Appl. Phycol. 2011, 23, 543–597. [CrossRef]
109. Becker, S.; Scheffel, A.; Polz, M.F.; Hehemann, J.-H. Accurate Quantification of Laminarin in Marine Organic
Matter with Enzymes from Marine Microbes. Appl. Environ. Microbiol 2017, 83. [CrossRef] [PubMed]
110. Brown, G.D.; Taylor, P.R.; Reid, D.M.; Willment, J.A.; Williams, D.L.; Martinez-Pomares, L.; Gordon, S.
Dectin-1 is a major beta-glucan receptor on macrophages. J. Exp. Med. 2002, 196, 407–412. [CrossRef]
[PubMed]
111. Taylor, P.R.; Brown, G.D.; Reid, D.M.; Willment, J.A.; Martinez-Pomares, L.; Gordon, S.; Wong, S.Y.C.
The β-glucan receptor, dectin-1, is predominantly expressed on the surface of cells of the monocyte/
macrophage and neutrophil lineages. J. Immunol. 2002, 169, 3876–3882. [CrossRef] [PubMed]
112. Pang, Z.; Otaka, K.; Maoka, T.; Hidaka, K.; Ishikima, S.; Oda, M.; Ohnishi, M. Structure of β-Glucan
Oligomer from Laminarin and Its Effect on Human Monocytes to Inhibit the Proliferation of U937 Cells.
Biosci. Biotechnol. Biochem. 2005, 69, 553–558. [CrossRef] [PubMed]
113. Dobruchowska, J.M.; Jonsson, J.O.; Fridjonsson, O.H.; Aevarsson, A.; Kristjansson, J.K.; Altenbuchner, J.;
Watzlawick, H.; Gerwig, G.J.; Dijkhuizen, L.; Kamerling, J.P.; et al. Modification of linear (β1→3)-linked
gluco-oligosaccharides with a novel recombinant β-glucosyltransferase (trans-β-glucosidase) enzyme from
Bradyrhizobium diazoefficiens. Glycobiology 2016, 26, 1157–1170. [CrossRef] [PubMed]
114. Jonsson-Wheat, J.O.; Hreggvidsson, G.O.; Fridjonsson, O.H.; Dobruchowska, J.M.; Kamerling, J.P. Glucan
Branching Enzymes and Their Methods of Use. US20160265013A1, 26 March 2014.
115. Hreggviðsson, G.H.; Dobruchowska, J.M.; Fridjonsson, O.H.; Jonsson, J.O.; Gerwig, G.J.; Aevarsson, A.;
Kristjansson, J.K.; Curti, D.; Redgwell, R.R.; Hansen, C.-E.; et al. Exploring novel non-Leloir
β-glucosyltransferases from proteobacteria for modifying linear (β1 → 3)-linked gluco-oligosaccharide
chains. Glycobiology 2011, 21, 304–328. [CrossRef]
116. Michel, G.; Czjzek, M. Polysaccharide-degrading enzymes from marine bacteria. In Marine Enzymes
for Biocatalysis—Sources, Biocatalytic Characteristics and Bioprocesses of Marine Enzymes; Trincone, A., Ed.;
Woodhead Publishing: Cambridge, UK, 2013; pp. 429–464. [CrossRef]
117. Barbeyron, T.; Henrissat, B.; Kloareg, B. The gene encoding the kappa-carrageenase of Alteromonas
carrageenovora is related to β-1,3-1,4-glucanases. Gene 1994, 139, 105–109. [CrossRef]
118. Barbeyron, T.; Gerard, A.; Potin, P.; Henrissat, B.; Kloareg, B. The kappa-carrageenase of the marine bacterium
Cytophaga drobachiensis. Structural and phylogenetic relationships within family-16 glycoside hydrolases.
Mol. Biol. Evol. 1998, 15, 528–537. [CrossRef] [PubMed]
119. Ohta, Y.; Hatada, Y. A Novel Enzyme, λ-Carrageenase, Isolated from a Deep-Sea Bacterium. J. Biochem. 2006,
140, 475–481. [CrossRef] [PubMed]
120. Fu, X.T.; Kim, S.M. Agarase: review of major sources, categories, purification method, enzyme characteristics
and applications. Mar. Drugs 2010, 8, 200–218. [CrossRef] [PubMed]
121. Ohta, Y.; Hatada, Y.; Miyazaki, M.; Nogi, Y.; Ito, S.; Horikoshi, K. Purification and Characterization of a
Novel α-Agarase from a Thalassomonas sp. Curr. Microbiol. 2005, 50, 212–216. [CrossRef] [PubMed]
Molecules 2020, 25, 930 27 of 29

122. Rochas, C.; Potin, P.; Kloareg, B. NMR spectroscopic investigation of agarose oligomers produced by an
α-agarase. Carbohyd. Res. 1994, 253, 69–77. [CrossRef]
123. Konasani, V.R.; Jin, C.; Karlsson, N.G.; Albers, E. A novel ulvan lyase family with broad-spectrum activity
from the ulvan utilisation loci of Formosa agariphila KMM 3901. Sci. Rep. 2018, 8, 14713. [CrossRef] [PubMed]
124. Sharma, S.D.; Pati, M.; Nayak, L. Uses of Seaweed and Its Application to Human Welfare: A Review. Int. J.
Pharm. Pharm. Sci. 2016, 8, 12–20. [CrossRef]
125. Makkar, H.; Tran, G.; Heuzé, V.; Giger-Reverdin, S.; Lessire, M.; Lebas, F.; Ankers, P. Seaweeds for livestock
diets: A review. Anim. Feed Sci. Technol. 2015, 212. [CrossRef]
126. Delaney, A.; Frangoudes, K.; Ii, S.A. Society and Seaweed: Understanding the Past and Present. In Seaweed
in Health and Disease Prevention; Fleurence, J., Levine, I., Eds.; Academic Press: San Diego, CA, USA, 2016;
pp. 7–40. [CrossRef]
127. Mouritsen, O.G. Seaweeds: Edible, Available & Sustainable; University of Chicago Press Chicago & London:
Chicago, IL, USA, 2013.
128. Hamid, N.; Ma, Q.; Boulom, S.; Liu, T.; Zheng, Z.; Balbas, J.; Robertson, J. Seaweed minor constituents.
In Seaweed Sustainability; Tiwari, B.K., Troy, D.J., Eds.; Academic Press: San Diego, CA, USA, 2015; pp. 193–242.
129. Venkatraman, K.L.; Mehta, A. Health Benefits and Pharmacological Effects of Porphyra Species. Plant. Food
Hum. Nutr. 2019, 74, 10–17. [CrossRef]
130. van der Weele, C.; Feindt, P.; van der Goot, A.J.; van Mierlo, B.; van Boekel, M. Meat alternatives: An integrative
comparison. Trends Food Sci. Technol. 2019, 88, 505–512. [CrossRef]
131. Ganesan, A.R.; Tiwari, U.; Rajauria, G. Seaweed nutraceuticals and their therapeutic role in disease prevention.
Food Sci. Hum. Wellness 2019, 8, 252–263. [CrossRef]
132. Wan, A.H.L.; Davies, S.J.; Soler-Vila, A.; Fitzgerald, R.; Johnson, M.P. Macroalgae as a sustainable aquafeed
ingredient. Rev. Aquacult. 2019, 11, 458–492. [CrossRef]
133. Shiroma, R.; Konishi, T.; Uechi, S.; Tako, M. Structural Study of Fucoidan from the Brown Seaweed Hizikia
fusiformis. Food Sci. Technol. Res. 2008, 14, 176–182. [CrossRef]
134. Kadena, K.; Tomori, M.; Iha, M.; Nagamine, T. Absorption Study of Mozuku Fucoidan in Japanese Volunteers.
Mar. Drugs 2018, 16, 254. [CrossRef] [PubMed]
135. Hsu, H.-Y.; Hwang, P.-A. Clinical applications of fucoidan in translational medicine for adjuvant cancer
therapy. Clin. Trans. Med. 2019, 8. [CrossRef] [PubMed]
136. Atashrazm, F.; Lowenthal, R.M.; Woods, G.M.; Holloway, A.F.; Dickinson, J.L. Fucoidan and Cancer:
A Multifunctional Molecule with Anti-Tumor Potential. Mar. Drugs 2015, 13, 2327–2346. [CrossRef]
[PubMed]
137. Lee, K.Y.; Mooney, D.J. Alginate: Properties and biomedical applications. Prog. Polym. Sci. 2012, 37, 106–126.
[CrossRef]
138. Pongsawatmanit, R.; Ikeda, S.; Miyawaki, O. Effect of temperature on viscoelastic properties of aqueous
alginate solutions. Thai J. Agric. Sci. 1998, 31, 583–593.
139. Senturk Parreidt, T.; Müller, K.; Schmid, M. Alginate-Based Edible Films and Coatings for Food Packaging
Applications. Foods 2018, 7, 170. [CrossRef]
140. Qin, Y.; Jiang, J.; Zhao, L.; Zhang, J.; Wang, F. Applications of Alginate as a Functional Food Ingredient.
In Biopolymers for Food Design; Grumezescu, A., Butu, A., Eds.; Academic Press: Cambridge, UK, 2018;
pp. 409–429.
141. Jensen, M.G.; Kristensen, M.; Astrup, A. Effect of alginate supplementation on weight loss in obese subjects
completing a 12-wk energy-restricted diet: A randomized controlled trial. Am. J. Clin. Nutr. 2012, 96, 5–13.
[CrossRef]
142. Houghton, D.; Wilcox, M.D.; Brownlee, I.A.; Chater, P.I.; Seal, C.J.; Pearson, J.P. Acceptability of alginate
enriched bread and its effect on fat digestion in humans. Food Hydrocoll. 2019, 93, 395–401. [CrossRef]
143. Odunsi, S.T.; Vázquez-Roque, M.I.; Camilleri, M.; Papathanasopoulos, A.; Clark, M.M.; Wodrich, L.;
Lempke, M.; McKinzie, S.; Ryks, M.; Burton, D.; et al. Effect of Alginate on Satiation, Appetite, Gastric
Function, and Selected Gut Satiety Hormones in Overweight and Obesity. Obesity 2010, 18, 1579–1584.
[CrossRef]
144. Mantovani, M.S.; Bellini, M.F.; Angeli, J.P.; Oliveira, R.J.; Silva, A.F.; Ribeiro, L.R. β-Glucans in promoting
health: Prevention against mutation and cancer. Mutat. Res. 2008, 658, 154–161. [CrossRef]
Molecules 2020, 25, 930 28 of 29

145. Aziz, A.; Poinssot, B.; Daire, X.; Adrian, M.; Bézier, A.; Lambert, B.; Joubert, J.M.; Pugin, A. Laminarin Elicits
Defense Responses in Grapevine and Induces Protection Against Botrytis cinerea and Plasmopara viticola.
Mol. Plant. Microbe Interact. 2003, 16, 1118–1128. [CrossRef]
146. UPL. Improve Overall Plant Health with Vacciplant®. Available online: https://us.uplonline.com/product-
details/vacciplant (accessed on 11 December 2019).
147. Seong, H.; Bae, J.-H.; Seo, J.S.; Kim, S.-A.; Kim, T.-J.; Han, N.S. Comparative analysis of prebiotic effects
of seaweed polysaccharides laminaran, porphyran, and ulvan using in vitro human fecal fermentation.
J. Funct. Foods 2019, 57, 408–416. [CrossRef]
148. Devillé, C.; Gharbi, M.; Dandrifosse, G.; Peulen, O. Study on the effects of laminaran, a polysaccharide from
seaweed, on gut characteristics. J. Sci. Food Agric. 2007, 87, 1717–1725. [CrossRef]
149. Zekovic, D.B.; Kwiatkowski, S.; Vrvic, M.M.; Jakovljevic, D.; Moran, C.A. Natural and modified
(1→3)-β-D-glucans in health promotion and disease alleviation. Crit. Rev. Biotechnol. 2005, 25, 205–230.
[CrossRef] [PubMed]
150. Dalonso, N.; Goldman, G.H.; Gernm, R.M. β-(1→3),(1→6)-Glucans: Medicinal activities, characterization,
biosynthesis and new horizons. Appl. Microbiol. Biotechnol. 2015, 99, 7893–7906. [CrossRef] [PubMed]
151. Park, H.-K.; Kim, I.-H.; Kim, J.; Nam, T.-J. Induction of apoptosis and the regulation of ErbB signaling by
laminarin in HT-29 human colon cancer cells. Int. J. Mol. Med. 2013, 32, 291–295. [CrossRef] [PubMed]
152. Bonfim-Mendonça, P.S.; Capoci, I.; Tobaldini-Valerio, F.K.; Negri, M.; Svidzinski, T. Overview of β-Glucans
from Laminaria spp.: Immunomodulation properties and applications on biologic models. Int. J. Mol. Sci.
2017, 18, 1629. [CrossRef]
153. Stier, H.; Ebbeskotte, V.; Gruenwald, J. Immune-modulatory effects of dietary Yeast Beta-1,3/1,6-D-glucan.
Nutr. J. 2014, 13. [CrossRef]
154. Distantina, S.; Rochmadi, R.; Fahrurrozi, M.; Wiratni, W. Synthesis of Hydrogel Film Based on Carrageenan
Extracted from Kappaphycus alvarezii. Mod. Appl. Sci. 2013, 7, 22–30. [CrossRef]
155. Qin, Y. Seaweed Hydrocolloids as Thickening, Gelling, and Emulsifying Agents in Functional Food Products.
In Bioactive Seaweeds for Food Applications; Qin, Y., Ed.; Academic Press: San Diego, CA, USA, 2018; pp. 135–152.
156. Pangestuti, R.; Kim, S.-K. Biological Activities of Carrageenan. In Advances in Food and Nutrition Research;
Kim, S.-K., Ed.; Academic Press: San Diego, CA, USA, 2014; Volume 72, pp. 113–124.
157. Carlucci, M.J.; Scolaro, L.A.; Noseda, M.D.; Cerezo, A.S.; Damonte, E.B. Protective effect of a natural
carrageenan on genital herpes simplex virus infection in mice. Antivir. Res. 2004, 64, 137–141. [CrossRef]
158. Eccles, R.; Meier, C.; Jawad, M.; Weinmüllner, R.; Grassauer, A.; Prieschl-Grassauer, E. Efficacy and safety
of an antiviral Iota-Carrageenan nasal spray: A randomized, double-blind, placebo-controlled exploratory
study in volunteers with early symptoms of the common cold. Respir. Res. 2010, 11, 108. [CrossRef]
[PubMed]
159. Panlasigui, L.; Baello, O.; Dimatangal, J.; Dumelod, B. Blood cholesterol and lipid-lowering effects of
carrageenan on human volunteers. Asia. Pac. J. Clin. Nutr. 2003, 12, 209–214. [PubMed]
160. Haijin, M.; Xiaolu, J.; Huashi, G. A κ-carrageenan derived oligosaccharide prepared by enzymatic degradation
containing anti-tumor activity. J. Appl. Phycol. 2003, 15, 297–303. [CrossRef]
161. Zhou, G.; Sheng, W.; Yao, W.; Wang, C. Effect of low molecular λ-carrageenan from Chondrus ocellatus on
antitumor H-22 activity of 5-Fu. Pharmacol. Res. 2006, 53, 129–134. [CrossRef] [PubMed]
162. Zhou, G.; Sun, Y.; Xin, H.; Zhang, Y.; Li, Z.; Xu, Z. In vivo antitumor and immunomodulation activities of
different molecular weight lambda-carrageenans from Chondrus ocellatus. Pharmacol. Res. 2004, 50, 47–53.
[CrossRef]
163. Sokolova, E.V.; Barabanova, A.O.; Homenko, V.A.; Solov’eva, T.F.; Bogdanovich, R.N.; Yermak, I.M. In Vitro
and Ex Vivo Studies of Antioxidant Activity of Carrageenans, Sulfated Polysaccharides from Red Algae.
Bull. Exp. Biol. Med. 2011, 150, 426–428. [CrossRef]
164. Rocha de Souza, M.C.; Marques, C.T.; Guerra Dore, C.M.; Ferreira da Silva, F.R.; Oliveira Rocha, H.A.;
Leite, E.L. Antioxidant activities of sulfated polysaccharides from brown and red seaweeds. J. Appl. Phycol.
2007, 19, 153–160. [CrossRef]
165. Hu, X.; Jiang, X.; Aubree, E.; Boulenguer, P.; Critchley, A.T. Preparation and In Vivo. Antitumor Activity of
κ-Carrageenan Oligosaccharides. Pharm. Biol. 2006, 44, 646–650. [CrossRef]
Molecules 2020, 25, 930 29 of 29

166. Yuan, H.; Zhang, W.; Li, X.; Lü, X.; Li, N.; Gao, X.; Song, J. Preparation and in vitro antioxidant activity
of κ-carrageenan oligosaccharides and their oversulfated, acetylated, and phosphorylated derivatives.
Carbohyd. Res. 2005, 340, 685–692. [CrossRef]
167. Xu, L.; Yao, Z.; Wu, H.; Wang, F.; Zhang, S. The immune regulation of κ-carrageenan oligosaccharide and its
desulfated derivatives on LPS-activated microglial cells. Neurochem. Int. 2012, 61, 689–696. [CrossRef]
168. Chauhan, P.S.; Saxena, A. Bacterial carrageenases: An overview of production and biotechnological
applications. 3 Biotech. 2016, 6. [CrossRef] [PubMed]
169. Li, L.; Ni, R.; Shao, Y.; Mao, S. Carrageenan and its applications in drug delivery. Carbohyd. Polym. 2014, 103,
1–11. [CrossRef] [PubMed]
170. Lokhande, G.; Carrow, J.K.; Thakur, T.; Xavier, J.R.; Parani, M.; Bayless, K.J.; Gaharwar, A.K. Nanoengineered
injectable hydrogels for wound healing application. Acta Biomat. 2018, 70, 35–47. [CrossRef]
171. Vera, J.; Castro, J.; Gonzalez, A.; Moenne, A. Seaweed polysaccharides and derived oligosaccharides stimulate
defense responses and protection against pathogens in plants. Mar. Drugs 2011, 9, 2514–2525. [CrossRef]
[PubMed]
172. Izydorczyk, M.; Cui, S.; Wang, Q. Polysaccharide Gums: Structures, Functional Properties, and Applications.
In Food Carbohydrates: Chemistry, Physical Properties and Applications, 1st ed.; Cui, S.W., Ed.; CRC Press:
Boca Raton, FL, USA, 2005.
173. Marcus, J.B. Food Science Basics: Healthy Cooking and Baking Demystified: The Science behind Healthy
Foods, Cooking and Baking. In Culinary Nutrition; Marcus, J.B., Ed.; Academic Press: San Diego, CA, USA,
2013; pp. 51–97.
174. Armisén, R.; Gaiatas, F. Agar. In Handbook of Hydrocolloids, 2nd ed.; Phillips, G.O., Williams, P.A., Eds.;
Woodhead Publishing: Cambridge, UK, 2009; pp. 82–107.
175. Serwer, P. Agarose gels: Properties and use for electrophoresis. Electrophoresis 1983, 4, 375–382. [CrossRef]
176. Armisén, R. Agar and agarose biotechnological applications. Hydrobiologia 1991, 221, 157–166. [CrossRef]
177. Ream, J.A.; Lewis, L.K.; Lewis, K.A. Horizontal Agarose Gel Mobility Shift Assay for Protein-RNA Complexes.
In Electrophoretic Separation of Proteins: Methods and Protocols; Kurien, B.T., Scofield, R.H., Eds.; Springer:
New York, NY, USA, 2019; pp. 363–370.
178. Aurelien, F.; Jon, C.; Steffen, L.; Esther, K.; Simon, T.; Maziar, M.; Ralf, T.; Shastri, V.P. Polysaccharide
hydrogels with tunable stiffness and provasculogenic properties via α-helix to β-sheet switch in secondary
structure. Proc. Natl. Acad. Sci. USA 2013, 110. [CrossRef]
179. Forget, A.; Pique, R.A.; Ahmadi, V.; Lu¨deke, S.; Shastri, V.P. Mechanically Tailored Agarose Hydrogels
through Molecular Alloying with β-Sheet Polysaccharides. Macromol. Rapid Commun. 2015, 36, 196–203.
[CrossRef]
180. Qi, H.; Zhao, T.; Zhang, Q.; Li, Z.; Zhao, Z.; Xing, R. Antioxidant activity of different molecular weight sulfated
polysaccharides from Ulva pertusa Kjellm (Chlorophyta). J. Appl. Phycol. 2005, 17, 527–534. [CrossRef]
181. Karnjanapratum, S.; You, S.G. Molecular characteristics of sulfated polysaccharides from Monostroma nitidum
and their in vitro anticancer and immunomodulatory activities. Int. J. Biol. Macromol. 2011, 48, 311–318.
[CrossRef]
182. Leiro, J.M.; Castro, R.; Arranz, J.A.; Lamas, J. Immunomodulating activities of acidic sulphated
polysaccharides obtained from the seaweed Ulva rigida C. Agardh. Int. Immunopharmacol. 2007, 7,
879–888. [CrossRef] [PubMed]
183. Qi, H.; Huang, L.; Liu, X.; Liu, D.; Zhang, Q.; Liu, S. Antihyperlipidemic activity of high sulfate content
derivative of polysaccharide extracted from Ulva pertusa (Chlorophyta). Carbohydr. Polym. 2012, 87, 1637–1640.
[CrossRef]
184. Mao, W.J.; Fang, F.; Li, H.Y.; Qi, X.H.; Sun, H.H.; Chen, Y.; Guo, S.D. Heparinoid-active two sulfated
polysaccharides isolated from marine green algae Monostroma nitidum. Carbohydr. Polym. 2008, 74, 834–839.
[CrossRef]

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).

You might also like

pFad - Phonifier reborn

Pfad - The Proxy pFad of © 2024 Garber Painting. All rights reserved.

Note: This service is not intended for secure transactions such as banking, social media, email, or purchasing. Use at your own risk. We assume no liability whatsoever for broken pages.


Alternative Proxies:

Alternative Proxy

pFad Proxy

pFad v3 Proxy

pFad v4 Proxy