Pectin Basics Review
Pectin Basics Review
Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol
Review
a r t i c l e i n f o a b s t r a c t
Article history: Pectins are a diverse family of biopolymers with an anionic polysaccharide backbone of ␣-1,4-linked d-
Received 21 September 2016 galacturonic acids in common. They have been widely used as emulsifiers, gelling agents, glazing agents,
Received in revised form 2 December 2016 stabilizers, and/or thickeners in food, pharmaceutical, personal care and polymer products. Commercial
Accepted 16 December 2016
pectin is classified as high methoxy pectin (HMP) with a degree of methylation (DM) >50% and low
Available online 24 December 2016
methoxy pectin (LMP) with a DM <50%. Amidated low methoxy pectins (ALMP) can be obtained through
aminolysis of HMP. Gelation of HMP occurs by cross-linking through hydrogen bonds and hydrophobic
Keywords:
forces between the methyl groups, assisted by a high co-solute concentration and low pH. In contrast,
Pectin
Polysaccharide
gelation of LMP occurs by the formation of ionic linkages via calcium bridges between two carboxyl
Rheology modifier groups from two different chains in close proximity, known as the ‘egg-box’ model. Pectin gels exhibit
Extraction Newtonian behaviour at low shear rates and shear-thinning behaviour when the shear rate is increased.
Hydrogels An overview of pectin from its origin to its physicochemical properties is presented in this review.
Gelling mechanisms © 2016 Elsevier Ltd. All rights reserved.
Hydrogen bonding
Hydrophobic interactions
Ionic interactions
Rheology
Contents
1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
2. Occurrence of pectin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
3. Chemical structure of pectin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
3.1. Homogalacturonan (HGA) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
3.2. Rhamnogalacturonan I (RG-I) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
3.3. Rhamnogalacturonan II (RG-II) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
3.4. Analytical tools for pectin macromolecules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
4. Sources of pectin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
5. Safety status and classification of pectins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122
6. Extraction and purification of pectin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 123
6.1. De-esterification of pectin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 124
6.2. Other feasible extraction additives and methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 125
∗ Corresponding authors at: Wee Sim Choo is from School of Science, Monash University Malaysia, 47500 Bandar Sunway, Selangor Darul Ehsan, Malaysia. David James
Young is from Faculty of Science, Health, Education and Engineering, University of the Sunshine Coast, Maroochydore DC, Queensland 4558, Australia. Xian Jun Loh is from
Institute of Materials Research and Engineering (IMRE), Agency for Science, Technology and Research (A*STAR), 2 Fusionopolis Way, Innovis, #08-03, Singapore 138634,
Singapore.
E-mail addresses: [email protected] (W.S. Choo), [email protected] (D.J. Young), [email protected] (X.J. Loh).
http://dx.doi.org/10.1016/j.carbpol.2016.12.033
0144-8617/© 2016 Elsevier Ltd. All rights reserved.
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 119
Fig. 1. [A] Pectin with covalently ␣-1,4-linked d-galacturonic acid (GalA) units, [B] The structure of cell wall. Pectins are often associated with other cell wall components such
as cellulose or hemicellulose in plant tissues; and [C] Pectins are most abundant in middle lamella and primary cell walls, and greatly decreased or are absent in secondary
cell walls towards the plasma membrane (Sticklen, 2008).
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 121
3. Chemical structure of pectin the pectin backbone comprises RGI domain with HG being linked as
its side chain (Vincken et al., 2003); and (c) the pectin backbone is
Although it has been more than 210 years since the discov- composed of alternating, perpendicularly linked-HGA strands and
ery of pectins, their chemical and structural properties are still the a RGI domain (Yapo, 2011a).
subject of investigation due to the inhomogeneity of this polymer
family. To begin with basic structure, pectin is assembled by at
least 17 different monosaccharides, of which GalA (Fig. 2Ai) is the
most plentiful, followed by l-arabinose, d-galactose, l-rhamnose
and others (Kaya, Sousa, Crépeau, Sørensen, & Ralet, 2014). These
3.1. Homogalacturonan (HGA)
monosaccharides can be interconnected through 20 different link-
ages (Kaya et al., 2014). Within GalA monomers, the carboxylic
HGA is a linear homopolymer of ␣-1,4-linked GalA and is known
groups or hydroxyls may be methyl-esterified (Fig. 2Aii) and/or
as the “smooth region” (Fig. 2Bi). It is an abundant and widespread
O-acetyl-esterified (Fig. 2Aiii), respectively. O-acetyl-esterification
domain of pectin, accounting for approximately 60–65% of total
occurs predominantly at the O-3 position and occasionally at the
pectin amount (Voragen et al., 2009). HGA appears to be synthe-
O-2 position.
sized in the Golgi apparatus and then transferred to the middle
Native pectins in plants are composed of polysaccharide
lamella and primary cell walls (Willats et al., 2001). It is then
domains: homogalacturonan (HGA), rhamnogalacturonan I (RG-I),
deposited in a form that has 70–80% of GalA units methyl-esterified
rhamnogalacturonan II (RG-II), xylogalacturonan, and apiogalac-
and could be O-acetyl-esterified at O-2 or O-3 (Mohnen, 2008;
turonan. The latter three have been classified as substituted HGA
O’Neill, Albersheim, & Darvill, 1990). The amount of GalA units
(Caffall & Mohnen, 2009; Harholt et al., 2010; Yapo, 2011a). The
present in a HGA chain is estimated to be around 100–200 units,
deposition and the way in which these domains are joined to one
however, this range could be an underestimate (Bonnin, Dolo, Le
another remains a matter of debate. Currently, it is thought that
Goff, & Thibault, 2002). The smooth region can sometimes be joined
the polysaccharide domains are covalently linked, and to a greater
by one or two ␣-1,2-linked l-rhamnopyranose units and most of the
or lesser extent, ionically cross-linked with other pectin strands to
pectins have this structure. In addition, GalA units may be substi-
form pectic networks that branch throughout the primary cell walls
tuted at the C-2 or C-3 positions with residues of xylose or apiose,
(Fig. 2B) (Harholt et al., 2010; Willats, McCartney, & Knox, 2003).
producing domains known as xylogalacturonan or apiogalacturo-
There are three models proposed: (a) the pectin backbone consists
nan, respectively (Ovodov, 2009). These biosynthetic modifications
of alternating HGA and RGI domains (Schols & Voragen, 1996); (b)
alter the functional properties of the HGA domain.
Fig. 2. [A] Forms of galacturonic acids (GalA) found in pectin: (i) GalA, (ii) Methylated GalA and (iii) O-Acetylated GalA; and [B] Pectin chain comprising of covalently linked
(i) homogalacturonan (HGA), (ii) rhamnogalacturonan I (RG-I) and (iii) rhamnogalacturonan II (RG-II). The diagram shown here is intended only to illustrate some of the
major domains found in most pectins rather than indicate definitive structures.
122 S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139
3.2. Rhamnogalacturonan I (RG-I) & Schols, 2000; Limberg et al., 2000). Pectin is an extremely complex
and structurally diverse group of polymers. The chemical structure
RG-I consists of up to 100 or more repeating units of the dis- of pectin varies between plants, tissues and even within a cell wall
accharide ␣-1,2-linked-l-rhamnose-␣-1,4-d-GalA (Fig. 2Bii). The (Willats, Knox, & Mikkelsen, 2006). Although considerable research
rhamnose residues account for >30% of a pectin (Renard, Crépeau, has been devoted to its chemical structure, there is much to be
& Thibault, 1995). Within these rhamnose residues, 50–78% is esti- done to correlate the functionalities and interactions of each divi-
mated to be RGI (Renard et al., 1995). The GalA residues of RG-I are sion of pectin to its unique biomechanical properties. This requires
not methyl-esterified; however, the GalA residues of RG-I could be thoughtful planning and the application of new analytical tools.
O-acetyl-esterified (Caffall & Mohnen, 2009). In most cases, 20–80%
of rhamnose residues in this domain are substituted at the C-4 posi-
4. Sources of pectin
tion with neutral sugar side chains (Mohnen, 2008). Attachment of
neutral sugar side chains to the C-2, C-3 and/or C-4 positions of
In spite of its availability in a large number of plant species, com-
rhamnose residues are also possible. The neutral sugars side chains
mercial sources of pectin are limited. Historically, apple pomace
are predominantly galactose and arabinose, forming galactan, ara-
has been the major source, but there has been an increasing use of
binan and arabinogalactans. Other sugars such as glucose, mannose,
citrus peel in recent years (Brejnholt, 2009). Today, commercially
fucose, xylose, and glucuronic acid are found covalently linked to
available pectin are mostly extracted from citrus peel (85.5%), fol-
the backbones as side chains. The composition and size of neutral
lowed by apple pomace (14.0%) and to a smaller extent, sugar beet
sugar side chains can be a single glycosyl residue up to 50 or more,
pulp (0.5%) (Ciriminna et al., 2015; Staunstrup, 2009). Citrus peel
resulting in a large and highly variable family of polysaccharides
and apple pomace are available in copious amounts as remain-
with a range of glycosidic linkages (Voragen et al., 2009). The highly
ders from juice and essential oil production (Masmoudi et al.,
branched nature of RG-I has led to the name “hairy region”.
2008; Nussinovitch & Hirashima, 2013; Shalini & Gupta, 2010),
while sugar beet pulp is obtained from the sugar industry (Yapo,
3.3. Rhamnogalacturonan II (RG-II)
Robert, Etienne, Wathelet, & Paquot, 2007). Other feasible sources
are shown in Table 1. Different plant species have different pectin
RG-II is not structurally related to RG-I, despite their names
content and the pectins extracted from each plant species have
running in sequence and the fact that it is also included in the
different physicochemical properties.
hairy region (Fig. 2Biii). It is a branched pectic domain contain-
ing a HGA backbone. RG-II is a highly compact homopolymer of
around nine ␣-1,4-linked GalA units (of which some are methyl- 5. Safety status and classification of pectins
esterified) with four structurally different polymeric side chains
attached. These side chains contain eleven rare sugars including The Joint FAO/WHO Expert Committee on Food Additives
apiose, 2-O-methyl-l-fucose, 2-O-methyl-d-xylose, 3-C-carboxy- (JECFA) has recommended pectin (Codex Alimentarius No. 440) as
5-deoxy-l-xylose (aceric acid), 3-deoxy-d-manno-octulosonic acid a safe additive with no limit on acceptable daily intake. Pectin is
and 3-deoxy-d-lyxoheptulosaric acid (Yapo, 2011b). A fascinat- not digested by humans as there are no enzymes in vivo that are
ing feature of RG-II is that it appears to be the only major pectic able to degrade the molecule (HansUlrich & Frank, 2012). However,
domain that does not have significant structural diversity or mod- certain bacteria of the gut flora such as Bacteroides, Bifidobacterium,
ulation of its fine structure. It is thought that the ends of RG-II Clostridium, Erwinia, Escherichia, and Eubacterium strains are able
are linked glycosidically to the HGA domains (Yapo, 2011a). It is a to use pectin as substrates for fermentation (Dongowski, Lorenz, &
highly conserved and widespread domain isolated from cell walls Anger, 2000; HansUlrich & Frank, 2012). Pectin is quite stable under
by endopolygalacturonase cleavage, indicating its covalent attach- the acidic condition of the stomach, although a de-esterification
ment to HGA. Interestingly, RG-II crosslinks two pectin molecules of a small extent may occur. Without the fermentation process,
(apiosyl residues) within the cell wall by borate ester links. pectin would pass almost unchanged through the digestive sys-
tem. Increasing consumer awareness of a healthy lifestyle and
3.4. Analytical tools for pectin macromolecules the emerging trend to produce functional food have made pectin
popular. It has been reported that pectin has numerous positive
There are multiple analytical tools capable of exploring the influences on health (Endress, 1991) including improving colonic
fine structures and functionalities of the pectin macromolecule health (Min et al., 2015), lowering of cholesterol and serum glucose
such as Fourier transform infrared spectroscopy (FT-IR) (Coimbra, levels (Jones et al., 2015; Zhu et al., 2015), reducing cancer propen-
Barros, Barros, Rutledge, & Delgadillo, 1998), FT-Raman spec- sity (Concha, Weinstein, & Zúñiga, 2013; Wang, Chen, & Lü, 2014),
troscopy (Bichara et al., 2016), gas chromatography (GC) (Huisman, and stimulating the immune response (Bernard et al., 2015).
Oosterveld, & Schols, 2004; Walter & Sherman, 1983), high perfor- Commercial pectin is very heterogeneous. The degree of methyl-
mance liquid chromatography (HPLC) (Levigne, Ralet, & Thibault, esterification (also known as degree of methylation) (DM) of GalA
2002; Voragen, Schols, & Pilnik, 1986), gas chromatography mass units is used to classify pectin (Deuel, 1943). DM is a percentage
spectroscopy (GC–MS) (Savary & Nuñez, 2003), electrospray- which expresses the molar ratio of methyl-esters present to GalA
ionization mass spectroscopy (ESIMS) (Ishii, Ichita, Matsue, Ono, & units (includes both free GalA and substituted GalA). It is the major
Maeda, 2002), ion-exchange chromatography (IEC) (Bonnin et al., parameter affecting gelling, influencing surface tension and emul-
2002; Kravtchenko, Voragen, & Pilnik, 1992b; Ralet, Bonnin, & sion formation (Müller-Maatsch, Caligiani, Tedeschi, Elst, & Sforza,
Thibault, 2001), polysaccharide analysis using carbohydrate gel 2014). There are two types of pectin classified according to DM:
electrophoresis (PACE) (Goubet, Ström, Dupree, & Williams, 2005), low methoxy pectin (LMP) and high methoxy pectin (HMP). HMP
capillary electrophoresis (CE) (Ström, Ralet, Thibault, & Williams, forms gels in high co-solute concentration (55–75%) and acidic (pH
2005; Zhong, Williams, Goodall, & Hansen, 1998) and nuclear mag- 2.50–3.50) systems (Oakenfull & Scott, 1984) while LMP forms gels
netic resonance (NMR) spectroscopy (Cardoso, Silva, & Coimbra, in the presence of cations over a broad pH range (D. Gidley, Morris,
2002). Combinations of these methods with enzymatic finger- Murray, Powell, & Rees, 1980; Hoefler, 1991). Aside from the typical
printing is a powerful suite to elucidate the complex structure LMP and HMP, there is another type of commercial pectin, viz ami-
of pectin (Daas, Arisz, Schols, De Ruiter, & Voragen, 1998; Daas, dated low methoxy pectin (ALMP). ALMP is obtained from ammonia
Meyer-Hansen, Schols, De Ruiter, & Voragen, 1999; Daas, Voragen, de-esterification of HMP (Fig. 3A). The difference between LMP and
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 123
Table 1
Pectin content (%) of commercial and other feasible raw materials used for pectin production.
Commercial Sources
Apple pomace 4.60–20.92 Canteri-Schemin, Fertonani, Waszczynskyj, and Wosiacki (2005), Min et al.
(2011)
Citrus peel Orange peel 10.90–24.80 Kaya et al. (2014), Koubala et al. (2008),
Grapefruit peel 21.60–28.00 Venzon et al. (2015)
Lemon peel 20.90–30.60
Lime peel 9.00–33.60
Sugar beet pulp 4.10–24.96 Lv, Wang, Wang, Li, and Adhikari (2013), Yapo et al. (2007)
ALMP is that there are primary amide groups in the chemical struc- agents may selectively extract pectin with different solubility from
ture, replacing methyl-ester groups on carboxyl groups in HMP. the same starting material (Yapo, 2009a,b). This selectivity, how-
Degree of amidation (DAm) is a percentage which expresses the ever, is less evident when the extraction does not occur in the above
molar ratio of primary amide group present to GalA units (includ- sequence, especially when acid is used first or when these agents
ing both free GalA and substituted GalA). Most commercial ALMP are used individually to extract pectin from the same starting
contains a DAm of 15–25. materials (Yapo, 2009a,b). Acids are generally the highest yield-
Deuel (1943) also proposed calculating the degree of acetyl- ing extracting agents (Yapo, 2009a,b). Another advantage of using
esterification (DA). DA is a percentage which expresses the molar acid extraction is that the pectin obtained are usually enriched with
ratio of acetyl-esters present to GalA units (includes both free GalA GalA units (Rao & Silva, 2006).
and substituted GalA). DA is also an important determinant of On a commercial scale, mineral acids are used to extract pectin.
gelling capacity of pectin; however, it is not as significant as DM Mineral acids are cheaper and more efficient in terms of produc-
in commercial pectin classification. Recently, another popular way tion yield. Commonly used minerals acids include hydrochloric
to classify pectin has been developed, the degree of blockiness (DB). acid, nitric acid and sulphuric acid (Fishman, Chau, Hoagland, &
Daas et al. (1999) introduced DB as a percentage expressing the total Hotchkiss, 2006; Iglesias & Lozano, 2004; Mesbahi, Jamalian, &
amount of non-esterified mono-, di- and tri- GalA liberated from Farahnaky, 2005; Pagán, Ibarz, Llorca, Pagán, & Barbosa-Cánovas,
degradation of pectin by endopolygalacturonase relative to the 2001; Rao & Silva, 2006). Pretreatment such as washing, blanching,
total amount of non-esterified GalA present in the pectin. Through or drying increase the stability of raw materials during trans-
DB, the pattern of methyl-esterification can be known. Moreover, portation and storage. These processes inactivate enzymes such
the pattern of methyl-esterification can radically alter gel forma- as pectin methyl-esterase (PME) and bacteria that would other-
tion and the properties of the gels formed. Guillotin et al. (2005) wise rapidly degrade pectin. To prevent fermentation, peels are
have further introduced the expression “absolute degree of block- dried to 10–12% moisture level. The fresh or dried raw materials are
iness” (DBabs ), which takes the original total GalA units (includes then extracted in demineralized water that has been acidified to pH
both free GalA and substituted GalA) into account. DBabs is defined 1.5–3.0 at 50–100 ◦ C for approximately three hours (Nussinovitch
as the amount of non-esterified mono-, di- and tri- GalA liberated & Hirashima, 2013). The extract containing pectin is separated from
from degradation of pectin by endopolygalacturonase relative to the raw materials by filtration or centrifugation. Polyvalent cations
the total amount of GalA present in the pectin (Fig. 3B). such as aluminum or copper salts are also used to precipitate pectin.
These metal ions are then removed by acidified alcohol washes, fol-
lowed by a wash in alkaline alcohol to neutralize the product. The
6. Extraction and purification of pectin precipitates obtained are washed with alcohol again and pressed
dry. Pectin can be further de-esterified in an alcohol suspension. The
Pectin can be extracted via water, buffers, chelating agents, final steps involve drying and milling to yield powdered pectin, fol-
acids, and/or bases. When used successively in the above- lowed by blending with different production batches to standardize
mentioned order (from water to bases), these different chemical
124 S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139
Newtonian behaviour (Fig. 4Aii), but only up to a certain con- and deduced that this was due to an increase in these hydrophobic
centration. Pectin dispersions at concentrations up to 3% behave interactions.
like Newtonian liquids and subsequent increases in concentration Recent studies have demonstrated the importance of the pattern
results in shear thinning behaviour (i.e. a decrease in viscosity with of methylation on gelation of HMP (Löfgren et al., 2005; Slavov et al.,
increase in shear rate) (Fig. 4B). It should be noted that the actual 2009). Pectins gel at a faster rate when they possess a block distribu-
pectin concentration at which the solution transforms from New- tion of methyl esterified-GalA units, relative to pectin with the same
tonian to shear thinning behaviour is dependent on the molecular DM, but with a random distribution of methyl esters (Löfgren et al.,
weight of the pectin used. There are many studies reporting the 2005) (Fig. 5B). Pectins with randomly distributed methyl esters
observation of an increase in pseudoplasticity with an increase in have considerably lower storage moduli (G’) relative to pectins with
pectin concentration (Muhammad et al., 2014; Nascimento et al., a higher DB and therefore the latter form stronger gels.
2016; Sato, Oliveira, & Cunha, 2008; Sousa et al., 2015). This pseu- Sucrose is often used as an additive for HMP gel preparation
doplastic behaviour is magnified with increasing concentration but it can be replaced by other co-solutes such as glucose, fruc-
(characterized by increasing slope at higher shear rate) (Fig. 4B). tose or polyols such as glycerol (Evageliou, Richardson, & Morris,
While at near-zero shear rate, the viscosity of pectin increases with 2000; Tsoga, Richardson, & Morris, 2004a). Hydrophobic interac-
increasing concentration (Fig. 4B). This phenomenon is common for tions depend largely on the molecular geometry of the co-solutes
polysaccharides in which the zero shear viscosity value becomes and the interaction with neighboring water molecules (Matubayasi,
higher with an increase in polymer concentration (Muhammad 1994; Tsoga, Richardson, & Morris, 2004b). Oakenfull and Scott
et al., 2014). It is also noteworthy that the Newtonian plateau limit (1984) deduced that the function of sugar in the formation of gels
is shifted to a low shear rate region with increasing pectin concen- by HMP is to stabilize junction zones by promoting hydrophobic
tration. interaction between methyl-ester groups. The effect of sucrose can
be explained by the concept of “preferential hydration” introduced
7.2. Pectin gelation by Lee and Timasheff (1981). “Preferential hydration” describes the
heterogeneous distribution of solvent around the macromolecules.
Gelling is the formation of a three-dimensional (3D) network Sucrose at a concentration of 50% by weight increases hydrophobic
of polymer chains with solvent and solutes trapped within. (Loh interaction between two methyl esters by 67% compared to water
et al., 2014; Loh, Nam Nguyen, Kuo, & Li, 2011; Nam Nguyen, Kuo, alone (Oakenfull & Scott, 1984).
& Loh, 2011; Wu, Loh, Wu, Lay, & Liu, 2010) The conditions required pH is also an important gelation factor. Pectin is an anion
for gelation and the properties of the gel ultimately depend on the polysaccharide and lowering the pH protonates carboxylic groups,
molecular structure, the intermolecular forces which hold the net- reducing electrostatic repulsions along and between pectin chains
work together and the nature of the junction zones at which the (Thakur et al., 1997). Moreover, at low pH, non-dissociated car-
polymer molecules are cross-linked (Axelos & Thibault, 1991a). The boxylic groups form inter- and intramolecular hydrogen bonds
junction zones in polysaccharide gels are complex and the molec- with secondary alcohol groups.
ular structures are held together by a large number of individually The formation of HMP gels is time-dependent (Seshadri, Weiss,
weak interactions such as electrostatic interactions and hydrogen Hulbert, & Mount, 2003) (Fig. 5C). In this study, a native pectin mix-
bonds. HMP and LMP have different gelation mechanisms, although ture (1.15% apple pectin, 41.40% sucrose, pH 1.50) at time zero was
the gel characteristics are governed by the same macromolecular highly viscous in nature. This solution was then subjected to stress
properties such as the composition, size and conformation of the at different gelation times and the magnitude of the stress response
polymers (Axelos & Thibault, 1991a). to the oscillatory sweeps increased as gelation time increased,
indicating that the solution became more elastic in nature. The
7.2.1. High methoxy pectin (HMP) frequency dependence of the storage modulus (G’) became simul-
HMP gels are obtained at low pH in the presence of a high taneously less pronounced, corresponding to a change from viscous
sucrose concentration or similar co-solute. A high sucrose con- to a rubbery behavior. At t = 225 min, G’ was higher than G”, indi-
centration reduces water activity which is necessary to promote cating that an elastic gel was formed. This rheological data can be
chain–chain interactions rather than chain-solvent interactions; explained in terms of network formation. Initially, there is no net-
while low pH protonates carboxylate residues, minimizing elec- work formed and each pectin molecule behaves very much like
trostatic repulsion (Thibault & Ralet, 2003). Walkinshaw and Arnott a single, dispersed molecule. As the pectin dispersion is sheared,
(1981) inferred from X-ray diffraction (XRD) studies that HMP gels the molecules relax quickly due to their high molecular mobility.
are stabilized by intermolecular hydrogen bonds and hydropho- As a consequence, there is no extensive stress buildup, particu-
bic bonding between methyl esters. This finding was subsequently larly at low oscillation frequencies. As time progresses, the pectin
confirmed by Oakenfull and Scott (1984). Fig. 5A schematically molecules begin to associate by the formation of hydrogen bond
illustrates the gelation of HMP. and with the help of hydrophobic interactions. The resulting 3D
Hydrogen bonds between pectin molecules are favored by the network restricts the freedom of molecules to react to the super-
conformation of adjacent GalA units. Individual hydrogen bonds imposed flow. As a result, the relaxation time increases, which is
are relatively weak, but a large number of them confer significant indicated by a shift of the storage modulus from the viscous region
thermodynamic stability to the gel. Hydrogen bonding is the main to the rubber plateau.
interaction that sustains the HMP gel structure, but it is insuffi- The gelation of HMP is also highly dependent on temperature,
cient to overcome the entropic barrier to gelation. Hydrophobic as demonstrated in a study by da Silva, Gonçalves, and Rao (1995)
interactions are essential for HMP gelation. The contribution of (Fig. 5D). The authors performed gelation of HMP at a tempera-
hydrophobic interactions to the free energy of junction zones in ture of 5 ◦ C and found that gelation was slow and resulted in a
HMP (with DM = 70%) is half that arising from hydrogen bonding weak gel. They attributed this to the lack of hydrophobic interac-
(Oakenfull & Scott, 1984). The unfavourable interaction between tion at low temperature. The pseudo-equilibrium G’ increased for
the non-polar methyl esters of pectin and water molecules result temperatures up to 30 ◦ C and then decreased. It was inferred that
in this hydrophobic effect (Thakur, Singh, Handa, & Rao, 1997). hydrogen bonds and hydrophobic interactions are best facilitated
These methyl esters coalescence, thereby reducing the surface at an intermediate temperature range and contribute to a network
area exposed to water and therefore overall entropy is decreased. with higher elasticity. The structure of HMP-sugar gels is reported
Guimarães et al. (2009) have reported that HMP is more viscous to be irreversible on heating (Rao & Silva, 2006). Evageliou et al.
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 127
(2000) reported an unusual “thermal annealing” behaviour of HMP instead of hydrogen bonds and hydrophobic interactions (Axelos
when a gel was heated and cooled. This behaviour, however, was & Thibault, 1991b) (Fig. 6 A). This is known as the “egg-box” model
pH-dependent. At pH 4.7, a HMP (of DM = 70%, 0.5 wt%) gel in the (Gidley, Morris, Murray, Powell, & Rees, 1979; Grant, Morris, Rees,
presence of 65 wt% sucrose was fully reversible on heating with no Smith, & Thom, 1973; Morris, Powell, Gidley, & Rees, 1982) and
detectable thermal hysteresis. At pH 3.0 and 3.5, there was a slight is characterized as junction zones formed by the ordered, side-
reduction in G’ and G” on heating. However, stability at 30 ◦ C was by-side associations of GalA, whereby specific sequences of GalA
observed to be virtually constant despite the pH difference. monomers in adjacent chains are linked intermolecularly through
electrostatic and ionic bonding of carboxyl groups, forming struc-
7.2.2. Low methoxy pectin (LMP) tures like an egg-box (Braccini & Pérez, 2001). The “egg-box” model
was first introduced to explain the gelation mechanism of alginate.
LMP gels are stabilized by ionic cross-linkages via calcium Although the “egg-box” model can explain the gelation of alginate
bridges between two carboxylates from two different chains, (Low, Chee, Kai, & Loh, 2015), it is only an approximation for the
Fig. 5. [A] HMP gelation is governed by intermolecular hydrogen bonds and hydrophobic interactions between methyl esters; [B] Gelation time for 0.75% of citrus pectin (i)
A, (ii) B and (iii) C with DM 74%, 72% and 82%, respectively, in the presence of 60% sucrose at pH 3. The gelation time was plotted as a function of (iv) degree of blockiness (DB).
Figure adapted from Löfgren, Guillotin, Evenbratt, Schols, and Hermansson (2005); [C] Frequency sweeps on apple pectin dispersion at pH 1.5, 22 ◦ C: (i) storage moduli and
(ii) loss moduli. Figures adapted from Seshadri et al. (2003); and [D] Storage moduli (G’) as a function of ageing time for 1% HMP from citrus in the presence of 60% sucrose at
pH 3 at several ageing temperatures: 5 ◦ C (shaded-triangle), 10 ◦ C (shaded-square), 15 ◦ C (open square), 20 ◦ C (shaded-circle), 30 ◦ C (open circle), 50 (open triangle). Figure
adapted from da Silva et al. (1995).
128 S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139
Fig. 5. (Continued)
mechanism of gelation of LMP. The latter is better described as a gel. Fraeye et al. (2009) further observed that the gel strength of
shifted “egg-box”, since one of the chains is slightly shifted with LMP-calcium gels was better explained by the pattern of methyla-
respect to the other. The “egg-box” models for the gelation of algi- tion than by the degree of methylation. G’ increased as DB increased
nate and pectin have been revisited by Braccini and Pérez (2001); from 0 to 40%, indicating that stronger gels were formed when
Fang et al. (2008); and Ventura, Jammal, and Bianco-Peled (2013). the NMG residues were distributed blockwise (Fig. 6Bii). The same
A two-stage process has been suggested with an initial dimeriza- correlation is true for the DBabs (Fig. 6Biii).
tion of two macromolecules and subsequent aggregation of these Fraeye et al. (2009) reported that the gel strength of
dimers (Axelos & Thibault, 1991b; Rao & Silva, 2006). The junc- LMP-calcium gels increases with increasing GalA concentration.
tions are formed between unbranched, non-esterified galacturonan However, it was suggested that this is also affected by the distri-
blocks bound together by coordinating calcium ions. The intra- bution of non-methylated GalA available for egg-box formation.
and intermolecular interactions of the polymer are modified by Dobies, Kozak, and Jurga (2004) reported that the strength of LMP-
the charge density and distribution of the non-methyl-esterified calcium gel is stronger with a higher concentration of calcium
carboxyl groups. The new conformation changes the placement of ions, as evidenced by the increase in G’ values with calcium con-
carboxyl groups to be outward facing from the polymer backbone, centration. An increase in concentration of calcium ions allows
facilitating inter-chain hydrogen bonding (Gilsenan, Richardson, & additional cross-linking of non-methylated GalA, generating a
Morris, 2000; Mansel et al., 2015). In general, egg-boxes formed denser and more elastic gel network (Basak & Bandyopadhyay,
between two neighbouring chains are stabilized predominantly by 2014; Ngouémazong et al., 2012). The storage moduli of pectin
electrostatic interactions, followed by hydrogen bonds and then by solutions also increase with increasing concentration of divalent
van der Waals interactions. iron ions (Mierczyńska et al., 2015) (Fig. 6C). Addition of calcium
Formation of cooperative egg-boxes is only possible when or iron ions increases pseudoplasticity, providing an alternative
stretches of non-methylated GalA units of a minimum length are to higher concentrations of thickening agents in fluids (Fig. 6Ci
present. The “egg-boxes” are stable only when there are at least six and ii). Conversely, addition of magnesium ions causes a decrease
consecutive carboxyl groups on the interior of each participating in G’, suggesting a weakening of the pectin gel by magnesium
chain (Braccini & Pérez, 2001; Powell, Morris, Gidley, & Rees, 1982). (Mierczyńska et al., 2015) (Fig. 6Ciii). It could be assumed that
The occurrence of methyl ester groups in the primary backbone lim- divalent iron ions could probably bind to pectin in an egg-box like
its the extent of such junction zones leading to formation of the gel. model; however there is sparse information on the interaction of
As DM decreases, LMP-calcium mixtures transform from liquid-like pectin with metal ions other than calcium.
to gel-like with G’ increasing, hence increasing gel strength with Commercially, LMP is usually dispersed in aqueous calcium
decreasing DM (Fraeye et al., 2009) (Fig. 6Bi). With decreasing DM, solutions at about 70 ◦ C and then cooled slowly. At high temper-
there is a higher probability of the number of sequences of non- ature, gelation of LMP is dominated by the formation of egg-box
methylated GalA residues being long enough to form “egg-boxes”. junction zones via calcium bridges, supported by hydrophobic
Formation of a higher amount of “egg-boxes” results in a stronger interactions. On cooling, hydrogen bonding increases, supported
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 129
Fig. 6. (Continued)
solution depended on the DM of each pectin and was adjusted so that in the final
gel the stoichiometric ratio R = 2[Ca2+ ]/[COO− ] was equal to 0.2. Figure adapted
from Fraeye et al. (2009); [C] Storage moduli of apple pectin with the addition of
(i) calcium hydroxide [Ca(OH)2 ], (ii) iron lactate [C6 H10 FeO6 ] and (iii) magnesium
Fig. 6. [A] LMP gelation mechanism is governed by ionic cross-linking via calcium hydroxide [Mg(OH)2 ]. Figure adapted from Mierczyńska, Cybulska, Sołowiej, and
(divalent) ions between two carboxyl groups from two different chains in close Zdunek (2015); [D] (i) Gelation of LMP via hydrogen bonds in the presence of acid,
proximity; [B] Effect of (i) degree of methylation (DM), (ii) degree of blockiness (DB) (ii) Evolution of storage moduli for 1 and 2% acid LMP gels and time resolved small-
and (iii) absolute degree of blockiness on storage moduli of pectin de-esterified angle X-ray scattering (SAXS) for the corresponding gels recorded at four different
by sodium hydroxide (symbol: rhombus), pectin de-esterified by Aspergillus times after mixing (tw ). Figures adapted from Mansel et al. (2015); and [E] Storage
aculeatus pectin methyl-esterase (PME) (symbol: square) and pectin de-esterified moduli (G’) as a function of time for 1% pectin samples at pH 3 with the absence of
by tomato PME (symbol: triangle). Data points placed between brackets represent salt (open symbol), and the addition of 0.05 M LiCl (light gray), 0.05 M NaCl (dark
pectin-calcium mixtures that did not exhibit gel character. The final GalA concen- gray) and 0.05 M KCl (black). The temperature from the left to the vertical dotted line
tration was fixed at 43 mM. The calcium concentration of the added calcium chloride was decreased from 40 ◦ C to 5 ◦ C over time, and kept constant at T = 5 ◦ C throughout
the time frame investigated. Figure adapted from Ström, Schuster, and Goh (2014).
130 S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139
Fig. 6. (Continued)
ence of sugar can also promote inter-chain interactions, as in HMP 21 helices are highly extended and reduction in temperature and/or
gelation. However, high concentrations of sugar will affect the pH can promote a conformational transition to the more compact
gelling of LMP adversely as the dehydrating effect of sugar favours 31 helices. This transition is reversible. Gelation occurs by dimer-
hydrogen bonding and decreases cross-linking by calcium ions (Fu ization of (antiparallel) three-fold helices, with more extensive
& Rao, 2001). aggregation at very low pH where the chains become essentially
Recent studies have reported new ways to make LMP gels via uncharged (Gilsenan et al., 2000). Protonation of carboxyl groups
acid-induced (Gilsenan et al., 2000; Ström et al., 2007) and mono- appears to promote conformational ordering and association by
valent ion-induced gelation (Ström et al., 2014; Wehr, Menzies, & two different mechanisms: (a) suppression of electrostatic repul-
Blamey, 2004). Pectin has a conformational transition below a cer- sion, and (b) allowing the carboxyl groups to act as hydrogen-bond
tain pH that depends on both the temperature and pH and induces donors (Gilsenan et al., 2000).
aggregation and eventually gelation. At low pH and/or low tem- Calcium-pectin interactions are essentially cooperative at low
perature, the conformation of pectin are predominantly 31 helices ionic strength; however, if the DM is too low, pectin can be precipi-
(Gilsenan et al., 2000). At high pH and/or high temperature, the tated out of solution instead of forming a stable gel. The number
solution conformation are predominantly 21 helices, with only lim- of dissociated free carboxyl groups is relatively low at a pH of
ited conversion to the 31 form on cooling (Gilsenan et al., 2000). The about 3 (i.e. below the pKa value of 3.5). Therefore, the formation
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 131
of typical egg-box-junction zones is limited with more interac- calcium. Monovalent ions such as sodium, potassium and lithium
tions between undissociated carboxyl groups via hydrogen bonds induce a stronger acid gel for LMP by permitting both hydrogen and
instead. The hydrogen bonds formed between protonated and ionic bonding (Ström et al., 2014) (Fig. 6E).
unprotonated carboxyl groups are weakened by protonation and
so acid-induced pectin gels are not stable at very low pH (Gilsenan
et al., 2000; Mansel et al., 2015) (Fig. 6Di). As gelation proceeds,
small-angle X-ray scattering (SAXS) data indicates that the entan-
7.2.3. Amidated low methoxy pectin (ALMP)
gled pectin solution starts to aggregate and form cross-links with
Löfgren, Guillotin, and Hermansson (2006) have demonstrated
time (Schuster, Cucheval, Lundin, & Williams, 2011), but not at low
that ALMP and LMP appear very similar in gel microstructure and
pH (Mansel et al., 2015) (Fig. 6Dii). In the early stages of network
kinetic behaviour at pH 3 and pH 7. ALMP shows rapid gel formation
build-up, pectin molecules in solution are far apart. The polymers
with G’ > G” throughout, with and without 30% sucrose (Fig. 7A). The
then cluster, thereby lowering the effective concentration of the
presence of sucrose influences neither the kinetic behaviour nor
solution. As more clusters nucleate, grow and connect, a stronger
the microstructure of the gels, but strongly increases the storage
network is slowly formed, as reflected by the increase in G’. Acid-
modulus. ALMP generally has reduced sensitivity towards calcium
induced gels have less junction zones than pectin-calcium gels,
(Löfgren et al., 2006). However, some studies have suggested that
highlighting the importance of hydrogen bond formation (Mansel
amidation might have little influence on sensitivity towards cal-
et al., 2015).
cium (Capel, Nicolai, Durand, Boulenguer, & Langendorff, 2006). By
Ström et al. (2014) have reported that gelling of LMP at low pH
comparison, ALMP strongly favours acid-induced gelation (Capel
may be assisted by monovalent ions up to a critical limit (Wang, Lee,
et al., 2006; Lootens et al., 2003). The hydrogen bonds between
Wang, & Huang, 2007). Interestingly, ionic cross-linking by diva-
amide groups help to promote gel formation at low pH. Gels from
lent ions is inhibited by monovalent ions such as sodium, which
ALMP are pH sensitive and thermally reversible. They can be heated
tie up the free carboxyl groups. However, monovalent ions can also
and solidified again on cooling, while conventional pectin-gels
be beneficial, improving the solubility of LMP in the presence of
remain liquid under the same conditions (Chen et al., 2014).
Fig. 7. [A] G’ and G” during gel formation of 8% pectin G (non-amidated pectin) at (a) pH 3; (b) pH 7. Normalized G’ values during gel formation of 0.8% pectin G and pectin
C30 (amidated pectin) at (c) pH 3; (d) pH 7. The initial 9 min involves a temperature decrease from 80 to 25 ◦ C. Figure reprinted from Löfgren et al. (2006); and [B] Effect of
(i) sonication intensity and (ii) sonication time on the development of storage moduli of apple pectin gels. Figures adapted from Seshadri et al. (2003).
132 S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139
Table 2
Molecular weight and rhamnose content (%) of commercial pectin.
Apple 63,000–81,000 2.30 12.60 Schmidt, Schmidt, Kurz, Endreß, and Schuchmann (2015),
Citrus 38,000–162,000 1.40 5.10 Yapo et al. (2007), Kravtchenko, Voragen et al. (1992a),
Sugar beet 20,200–90,100 5.40 6.80–32.90 Leroux, Langendorff, Schick, Vaishnav, and Mazoyer (2003)
Table 3
Young’s modulus of high methoxyl pectin (HMP), low methoxyl pectin (LMP) and amidated low methoxyl pectin (ALMP) gels (Vithanage et al., 2010).
7.3. Intrinsic and extrinsic factors ing extraction or purification and are therefore not usually present
in significant amounts in commercial pectins.
The gelation process is affected by both intrinsic and extrinsic
parameters. Intrinsic factors include the number and distribution 7.4. Comparison in rheology of HMP, LMP and ALMP
pattern of free carboxyl groups, the molecular weight and types of
pectin while extrinsic factors include pectin concentration, calcium Comparing the three commercially available pectins, HMP
concentration, pH, ionic strength and temperature (Gigli, Garnier, shows the highest apparent viscosity, followed by LMP and ALMP
& Piazza, 2009). (G. Vithanage, Grimson, Wills, Harrison, & Smith, 2010). Young’s
Comparing commercial pectin from different sources; apple modulus is used to define the force displacement of matter. It is
pectin produces a more elastic-viscous gel and citrus pectin pro- determined by calculating the slope from the plot of stress versus
duces a more elastic-brittle gel. By contrast, sugar beet pectin is strain. Vithanage et al. (2010) found that Young’s modulus for HMP
less effective as a gelling agent. However, it possesses emulsify- gel was the highest, followed by LMP and ALMP (Table 3). In other
ing properties (Chen, Fu, & Luo, 2016) and is capable of forming words, HMP gel requires a higher force to break its structure com-
covalently cross-linked hydrogels and so is of commercial value pared to LMP and ALMP gels, indicating that hydrogen bonding in
(May, 1990). Gelation of pectin is influenced by molecular weight HMP pectin is stronger than the ionic bonding (with calcium ions)
because the rigidity of the gel is determined by the number of effec- in LMP and ALMP. Young’s modulus is temperature-dependent
tive junctions formed per chain (Axelos & Thibault, 1991a). The (Table 3).
lower the molecular weight, the less junction zones per molecule HMP gels demonstrate the broadest range of elastic behaviour
can be formed, decreasing the extent of cross-linking and thereby in puncture and strain sweep tests, followed by LMP and ALMP
weakening the gel. Although molecular weight is mainly affected gels (Vithanage et al., 2010). Comparing LMP and ALMP gels, the
by the origin of the pectin, handling methods such as extraction strength of the former is reported to be greater than that of the
also affect the molecular weight of the polymer significantly. For latter; however, ALMP gels exhibit a higher storage modulus than
example, ultrasound may be used to increase the yield of pectin LMP gels (Vithanage et al., 2010). The G’ and G” values for ALMP gel
extraction. However, Seshadri et al. (2003) found that the rheo- increase with increasing temperature, forming a more rubbery elas-
logical properties of pectin that had been treated with ultrasound tic medium; whereas HMP and LMP gels show a decrease of G’ and
were inferior. As sonication intensity and time was increased, gel G” with increasing temperature, forming a more liquidly medium
strength was reduced. As shown in Fig. 7Bi and ii, G’ decreased as (Vithanage et al., 2010). This is postulated to be due to the forma-
sonication intensity and time increased. This result was attributed tion of stable junctions by hydrogen bonding through amide groups
to the reduction in the molecular weight of pectin by cavitational in ALMP. At lower temperature, hydrogen bonds are favoured, rein-
promoted chain degradation. forcing the junction zones; while at higher temperatures, the loss of
The presence of sugar monomers such as rhamnose, whose hydrogen bonding is compensated for with reinforced hydrophobic
dimensions are not compatible with the geometry of the junction interactions (Alonso-Mougán, Meijide, Jover, Rodrıı́guez-Núñez, &
zones formed by GalA monomers, obstructs the molecular orien- Vázquez-Tato, 2002). The sol/gel transition of LMP is sensitive to
tation necessary for junction-zone formation (Axelos & Thibault, the ionic strength of the medium, while the viscoelastic properties
1991a). Rhamnose influences the conformation of the polymer in of the gel structure is time-dependant and resistant to moderate
solution and ultimately its gelling properties (Oakenfull, 1991). This temperature (retained up to 60 ◦ C) (Fu & Rao, 2001; Gigli et al.,
sugar is most abundant in rhamnogalacturonan, disturbing the reg- 2009).
ularity of the galacturonan backbone and forming the “hairy region”
which limits inter-chain association. This could further explain 7.5. Rheology of mixed pectin systems
the reason sugar beet pectin exhibited the poorest gelling prop-
erties of the three commercial pectins, as it possesses a relatively Synergistic mixed gels are also of interest to industry. Syner-
high rhamnose content (Table 2). However, it should be noted that gism can lead to different microstructures to that of pure gels with
the regularity and frequency of the rhamnose interruptions to the improved texture and enhanced rheological properties that result
pectin chain are different in pectins from different plant species in cost savings during manufacturing. Synergistic effects can be
or even within the same species. The presence of neutral sug- achieved by mixing HMP, LMP and/or ALMP to produce a strength-
ars also limit inter-chain association (Kravtchenko, Voragen et al., ened mixed gel. The G’ of mixed HMP/LMP gel is higher than that of
1992a) (Table 2). However, these branches are often removed dur- HMP or LMP (Löfgren & Hermansson, 2007). Fig. 8A demonstrates
that G’ values for mixed HMP/LMP are considerably higher than
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 133
for HMP gels. The mixed gel exhibits similar rheological behaviour Polysaccharides and proteins are two major components of
to HMP gels of higher sugar concentration. The strong aggregation food products. They are often used to control structure, texture,
of LM pectin in the presence of sucrose and calcium contributes and stability. Textural and structural properties depend largely
to a large influence on the storage modulus of the mixed gels, on these biopolymers forming organized structures. The mixture
increasing the storage modulus by about thirty times (based on of different components or even the same class of biopolymer
60% sucrose) (Löfgren & Hermansson, 2007). By comparison, the (carbohydrate–carbohydrate or protein-protein) can lead to phase
addition of HMP in a mixed gel based on 30% sucrose increased the separation. There are two types of phase separation: segregative
storage modulus five times (Löfgren & Hermansson, 2007). When which results in an unmixingof the two phases; and associative
ALMP is mixed with HMP, it is possible to obtain high viscosity and which leads to complex formation of the two biopolymers. Associa-
reduced pseudoplasticity (Sato et al., 2008). tive phase separation has been widely applied to stabilize products
in the food, cosmetic, pharmaceutical, medical and biotechnolog-
Fig. 8. [A] Storage moduli during gel formation in mixed 0.8% HMP/0.4% LMP compared to 1.2% HMP gels. A, B, and HMC are HMP while C30 and LMC are LMP. (a), (b) and
(c) were gels formed in the presence of 60% sucrose while (d), (e) and (f) were gels formed in the presence of 45% sucrose. Figure reprinted from Löfgren and Hermansson
(2007); [B] (i) Temperature dependences of a fixed dynamic moduli (frequency of 1.2 rad/s) for acid aqueous mixtures of pectin-chitosan at the compositions indicated. (ii)
Frequency dependences of storage modulus and loss modulus at different temperatures (stages) of the gel-forming process for pectin-chitosan mixtures at compositions
indicated. Figures adapted from Nordby, Kjøniksen, Nyström, and Roots (2003); [C] (i) The complex viscosity, *, storage modulus G’, and loss modulus G” versus angular
frequency for -lacglobulin-pectin coaservates at composition 5:1 prepared at a concentration of sodium chloride = 0.02 M. (ii) The complex viscosity, *, storage modulus G’,
and loss modulus G” versus angular frequency for 2% pectin solution prepared at a concentration of sodium chloride = 0.02 M. (iii) The storage modulus G’ versus frequency
curves for -lacglobulin-pectin coaservates at different compositions prepared at a concentration of sodium chloride = 0.11 M. Figures adapted from Xiaoyong Wang et al.
(2007); and [D] Temperature sweep of low-fat spread with varying ratios of fish gelatin and pectin. Figure reprinted from Cheng, Lim, Chow, Chong, and Chang (2008).
134 S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139
Fig. 8. (Continued)
ical industries. In the pectin-casein system, for example, pectin interconnected gel-like network structure. The typical viscoelas-
can prevent aggregation of casein micelles or be the cause of it tic behaviour of pectin solution (Fig. 8C) has been observed in
(Maroziene & de Kruif, 2000), changing the microstructure stabil- coacervates with similar pectin content, indicating that the elas-
ity and rheology of dairy products. Pectin and milk protein are tic behaviour of -lacglobulin-pectin coaservates is primarily due
generally not considered compatible at neutral pH; however, the to the interactions between -lacglobulin molecules and pectin
interaction can be mediated by calcium bridging between pectin chains. Sadahira, Rodrigues, Akhtar, Murray, and Netto (2016)
and casein micelles. deduced that egg white and pectin complexes increased continu-
Agoda-Tandjawa, Durand, Gaillard, Garnier, and Doublier (2012) ous phase viscosity and enhanced foam stability by slowing liquid
demonstrated that the presence of LMP does not modify the drainage. When incorporated with fish gelatin in a low-fat spread,
viscoelasticity and microstructural properties of microfibrillated an increase in pectin content could also increase the consistency
cellulose suspensions but it does increase the shear-thinning and melt ability of the low-fat spread (Cheng et al., 2008) (Fig. 8D). It
behaviour and thixotropic characteristic of the suspensions. They was also predicted that the low-fat spread sample with more pectin
suggested that this outcome was due to the viscosifier effect of content would perform better with ‘melt in the mouth’ character-
Nordby et al. (2003) demonstrated that a pectin-chitosan mix- istics and a better instant in-mouth flavour release effect.
ture at different ratios at very low pH (≈1.7) behaves more elastic
at lower temperatures (Fig. 8Bi). At temperatures below the gel 8. Modification of pectin structure
temperature, G’ is always larger than G”, indicating that the pectin-
chitosan mixture is in a solid-like state. At temperatures above Pectin formulations can be manipulated to achieve gels, 3-
the gel temperature (42–50 ◦ C), G” is always larger than G’, indi- D matrices, films and micro-/nano-particles. However, there can
cating that the mixture is in a more viscous-like state (Fig. 8Bii). be a lack of reproducible performance due to the large diversity
Khondkar, Tester, Hudson, Karkalas, and Morrow (2007) used phos- of molecular structures, leading to problems in quality control
phate cross-linking between starch and pectin to prepare gels with and quality assurance (Günter et al., 2014). Solutions to this
greater elasticity and better microstructure. complication fall into two categories: the development of new
Wang et al. (2007) have demonstrated that increasing the technologies for pectin isolation, purification, and characteriza-
amount of pectin in a protein-polysaccharide mixture favours tion and the modification of pectin macromolecules (Liu, Fishman,
the formation of stronger gels. The significantly higher G’ than & Hicks, 2007). Chen et al. (2014) recently published a thorough
G” indicates that -lacglobulin-pectin coaservates form a highly review on chemical modifications of pectins detailing substitutions,
S.Y. Chan et al. / Carbohydrate Polymers 161 (2017) 118–139 135
Chen, J., Liu, W., Liu, C.-M., Li, T., Liang, R.-H., & Luo, S.-J. (2014). Pectin Fraeye, I., De Roeck, A., Duvetter, T., Verlent, I., Hendrickx, M., & Van Loey, A.
modifications: A review. Critical Reviews in Food Science and Nutrition, 55(12), (2007). Influence of pectin properties and processing conditions on thermal
1684–1698. pectin degradation. Food Chemistry, 105(2), 555–563.
Chen, H.-m., Fu, X., & Luo, Z.-g. (2016). Effect of molecular structure on emulsifying Fraeye, I., Doungla, E., Duvetter, T., Moldenaers, P., Van Loey, A., & Hendrickx, M.
properties of sugar beet pulp pectin. Food Hydrocolloids, 54(Part A), 99–106. (2009). Influence of intrinsic and extrinsic factors on rheology of
Cheng, L. H., Lim, B. L., Chow, K. H., Chong, S. M., & Chang, Y. C. (2008). Using fish pectin–calcium gels. Food Hydrocolloids, 23(8), 2069–2077.
gelatin and pectin to make a low-fat spread. Food Hydrocolloids, 22(8), Fu, J. T., & Rao, M. A. (2001). Rheology and structure development during gelation
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