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Chapter 5 - Fish Gelatin

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From: Gokhan Boran and Joe M. Regenstein, Fish Gelatin,

In Steve L. Taylor editor:
Advances in Food and Nutrition Research, Vol. 60,
Burlington: Academic Press, 2010, pp.119-144.
ISBN: 978-0-12-380944-5
© Copyright 2010, Elsevier Inc.
Academic Press.
 Author's personal copy

CHAPTER 5
Fish Gelatin
Gokhan Boran*,1 and Joe M. Regenstein†

Contents I. General Information on Gelatin 120

A. The parent molecule: Collagen 121
B. Collagen–gelatin conversion 124
C. The mechanism of gelation 126
D. The characteristics of gelatin 127
E. Sources of raw material used in gelatin
manufacturing 127
F. The gelatin market 128
G. Industrial applications of gelatin 128
II. Fish Gelatin 129
A. Common and potential sources 129
B. Properties of fish gelatin 130
C. Other factors affecting quality 135
D. Quality of fish gelatin compared to
mammalian gelatins 138
III. Methodological Challenges in Quality
Determination of Gelatin 139
IV. Conclusions and Suggested Readings 140
References 140

Abstract Gelatin is a multifunctional ingredient used in foods, pharmaceu-

ticals, cosmetics, and photographic films as a gelling agent, stabi-
lizer, thickener, emulsifier, and film former. As a thermoreversible
hydrocolloid with a narrower gap between its melting and gelling
temperatures, both of which are below human body temperature,

* Department of Food Engineering, Yüzüncü Yıl University, Van, Turkey

{
Department of Food Science, Cornell University, Ithaca, New York, USA
1
Corresponding author: Gokhan Boran, E-mail address: Gboran@yyu.edu.tr

Advances in Food and Nutrition Research, Volume 60 # 2010 Elsevier Inc.

ISSN 1043-4526, DOI: 10.1016/S1043-4526(10)60005-8 All rights reserved.

119
 Author's personal copy
120 Gokhan Boran and Joe M. Regenstein

gelatin provides unique advantages over carbohydrate-based gel-

ling agents. Gelatin is mostly produced from pig skin, and cattle
hides and bones. Some alternative raw materials have recently
gained attention from both researchers and the industry not just
because they overcome religious concerns shared by Jews and
Muslims but also because they provide, in some cases, technologi-
cal advantages over mammalian gelatins. Fish skins from a number
of fish species are among the other sources that have been com-
prehensively studied as sources for gelatin production. Fish skins
have a significant potential for the production of high-quality
gelatin with different melting and gelling temperatures over a
much wider range than mammalian gelatins, yet still have a suffi-
ciently high gel strength and viscosity. Gelatin quality is industrially
determined by gel strength, viscosity, melting or gelling tempera-
tures, the water content, and microbiological safety. For gelatin
manufacturers, yield from a particular raw material is also impor-
tant. Recent experimental studies have shown that these quality
parameters vary greatly depending on the biochemical character-
istics of the raw materials, the manufacturing processes applied,
and the experimental settings used for quality control tests. In this
review, the gelatin quality achieved from different fish species is
reviewed along with the experimental procedures used to deter-
mine gelatin quality. In addition, the chemical structure of collagen
and gelatin, the collagen–gelatin conversion, the gelation process,
and the gelatin market are discussed.

I. GENERAL INFORMATION ON GELATIN

Gelatin is a term used for a class of protein fractions that have no existence
in nature. Gelatin is derived from collagen, which is a natural structural
protein, predominantly found in the connective tissues of animals (Balian
and Bowes, 1977; Belitz and Schieberle, 2004; DeMan, 1999) although also
found in many other tissues. Collagen is the most ubiquitous of animal
proteins. Gelatin is one of the most widely used biopolymers and is added
to foods, drugs, cosmetics, photographic films, and other products,
including paints, matches, and fertilizers, as a gelling agent, foam stabi-
lizer, and structure enhancer (Gudmundsson, 2002; Karim and Bahat,
2009; Yang et al., 2007; Zhou and Regenstein, 2004). Gelatin is able to
form a high viscosity solution in warm water, which sets to a gel on
cooling. The chemical composition of gelatin is, in many respects, similar
to that of collagen, its parent molecule. Gelatin is, however, not composed
of one size of collagen fraction or peptide chain but is a combination of
many fractions varying in size, including the whole a-chain of the tropo-
collagen molecule (a trimer of around 330 kDa that aggregates to form the
larger collagen structures) and hydrolytic fragments of parts of the
 Author's personal copy
Fish Gelatin 121

a-chains of different lengths (Eastoe and Leach, 1977). Gelatin gels have
relatively lower melting temperatures compared to the gels of other gel-
ling agents (Williams, 2007). Gelatin gels generally have a melting temper-
ature below 35
C, that is, below human body temperature, which makes
gelatin unique in terms of its sensory aspects, especially flavor release,
which is particularly desired for some food applications (Baziwane and
He, 2003; Boran and Regenstein, 2009; Choi and Regenstein, 2000). Other
gelling agents such as starch, alginate, pectin, and agar are carbohydrates
and their gels cannot melt below body temperature as most have much
higher melting temperatures (Williams, 2007).
Gelatin is obtained from the skins and bones of pigs and cattle, but
mostly from pig skin. However, there are alternative raw materials that
can be used in gelatin manufacturing, including by-products from the
chicken and fish processing industries. Fish skins have received attention
from researchers as an alternative raw material having the potential for
the production of large amounts of high-quality gelatin. Therefore, recent
studies with fish skin gelatin have focused on the evaluation of different
fish species as an alternative raw material for gelatin production and the
quality of the extracted gelatins in comparison with commercial gelatins
from conventional sources (Boran and Regenstein, 2009).
In this review, gelatin production and processing, the raw materials
used in gelatin production, the technological attributes of gelatin, the
gelatin market, and the market specifically for fish gelatin are discussed.
In addition, the most critical factors affecting the quality of gelatin are
discussed. For this purpose, the chemical structure of collagen is reviewed
in detail to take a closer look at the possible factors affecting the properties
of the resultant gelatin. The conversion process of collagen into gelatin
and the gelation mechanism are discussed to show which driving forces
are involved in gelation, which factors might affect the solution–gel and
gel–solution transitions, and how extraction conditions might affect the
final product, gelatin. The methods being currently used to determine the
quality of gelatin are also reviewed.

A. The parent molecule: Collagen

Collagen is the most abundant protein in the animal body (DeMan, 1999).
Collagen is part of the connective tissue in muscles and many other
organs, including the skin, bones, teeth, and tendons. Collagen fibrils
normally have a regular periodicity of 64 nm when stained for micros-
copy, which can be increased to 400 nm with r tension (DeMan, 1999).
Collagen molecules are arranged head-to-tail, with a 35-nm gap between
molecules, and are found in larger structures having staggered bundles,
that is, adjacent collagen molecules are not aligned with each other.
Charged and uncharged residues are found to be periodically clustered
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122 Gokhan Boran and Joe M. Regenstein

along the sequence of collagen at about every 230 residues, which is

around 64 nm, although this distance may vary somewhat among differ-
ent tissue sources of collagen. This suggests that the collagen molecules
are aligned such that the maximum electrostatic and hydrophobic inter-
actions occur between different molecules (Fig. 5.1).
Collagen constitutes 20–25% of the total protein in mammals and has a
unique amino acid composition, which includes two modified amino
(imino) acids, hydroxyproline and hydroxylysine (Belitz et al., 2004).
Its molecular structure is mainly the multiple repetition of a ‘‘Glycine-X-Y’’
sequence, where ‘‘X’’ is often proline and ‘‘Y’’ is often hydroxyproline.
Collagen has a unique triple helix structure that is based on a special
helix of three polypeptide chains with high levels of imino acids. Each
polypeptide chain is left handed and has three amino acids per turn. These
three polypeptide chains, called a-chains, are supertwisted around one
another and form a superhelix that is right handed (Nelson and Cox, 2005).
The basic structural unit of the collagen superhelix is called tropocollagen.
It has a molecular weight of approximately 330 kDa, with a length of
approximately 300 nm and a diameter of 1.5 nm (Belitz et al., 2004).

Collagen molecule
1.5 nm
300 nm

64
nm
Collagen fibril

10−300 nm

FIGURE 5.1 Schematic representations of collagen molecules and a collagen fibril.

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Fish Gelatin 123

When hydrolyzed, the collagen can give three different fractions:

independent a-chains, which may have slightly different molecular
weights, a b-chain, that is, two a-chains linked to each other by covalent
bonds, and a g-chain, that is, the three a-chains linked to one another by
covalent bonds. These fractions differ in molecular size: a-chains corre-
spond to a molecular weight of 80–125 kDa, b-chains correspond to a
molecular weight of 160–250 kDa, and g-chains correspond to a molecular
weight of 240–375 kDa, which is very similar to the molecular size of
collagen (Imeson, 1997).
Collagen typically contains about 35% glycine, 11% alanine, and 21%
proline and hydroxyproline, the amount of which varies somewhat
among different species although the high content of proline and
hydroxyproline seems to be characteristics of collagen structure regard-
less of the source (Balian and Bowes, 1977). Hydroxyproline is a rare
amino acid that is absent in nearly all other proteins and so its presence
can be used to determine the presence of collagen in the presence of other
proteins, and with proper calibration (i.e., the conversion factor is at a
minimum species specific), the amount of collagen or gelatin (Engel and
Bachinger, 2005) in the mixture. Another protein containing hydroxypro-
line is elastin, but the amount of hydroxyproline in elastin is very low and
the amount of elastin in most tissues is also very low when compared to
that of collagen (Nelson and Cox, 2005).
Collagen is generally considered to be an incomplete protein since the
concentration of some essential amino acids is low in collagen and conse-
quently, in gelatin (Belitz et al., 2004; Nelson and Cox, 2005). However,
when eaten as part of a meal, the contribution of gelatin to the amino acid
ingestion needs to be considered.
The amounts of the aromatic and sulfur-containing amino acids are
low (0–0.6%) in collagen, that is, tryptophan and cysteine are mostly
absent in collagen (Balian and Bowes, 1977). Cysteine is usually absent
in collagen, therefore, there are usually no disulfide bonds involved in
collagen structure although there are some collagens that have cysteine
(Engel and Bachinger, 2005). For those, disulfide bonds are also involved
in the formation of intermolecular cross-links (disulfide bonds) and pro-
vide additional stabilizing effects for the structure. The structure of colla-
gen provides an explanation of why glycine is the most abundant amino
acid and why proline and hydroxyproline are found so often in collagen.
Only glycine residues, as the smallest of the hydrophobic amino acids,
can fit into the very tight central core between the individual a-chains,
while proline and hydroxyproline residues permit sharp twists of the
collagen helix allowing for the relatively low three amino acids per turn
(Nelson and Cox, 2005).
The collagen molecule is primarily stabilized by hydrogen bonds
between the backbone amino group of glycine and the backbone carboxyl
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124 Gokhan Boran and Joe M. Regenstein

group of a residue in the X position of a neighboring a-chain, which is

usually a proline. Proline in the Y position is generally hydroxylated
posttranslation into hydroxyproline, which also plays an important role
in the formation of intra- and intermolecular hydrogen bonds. Hydroxy-
proline is, therefore, important for both the structure of the collagen
molecule and of the collagen fibrils (Brinckmann, 2005).
During maturation or aging of the living animal, collagen fibers
strengthen and are further stabilized primarily by complex covalent
bonds. Lysine, hydroxylysine, and histidine residues are heavily involved
in the formation of these covalent bonds, that is, aldimine bonds between
lysine and lysine or hydroxylysine (Balian and Bowes, 1977; Belitz et al.,
2004; Engel and Bachinger, 2005; Eyre and Wu, 2005; Nelson and Cox,
2005) that lead to the formation of desmosine and isodesmosine, which
are unusual in that they involve the participation of four amino acids in
the reaction. This is possible because of the rigid geometry of the collagen
structures and the long times available for these reactions to occur.
All the fibril-forming collagen types (type I, II, III, V, XI, XXIV, and
XXVII collagens) are cross-linked through a mechanism based on the
reactions of aldehydes derived from some lysine (or hydroxylysine) side
chains. Histidine might also participate in the formation of a trivalent
cross-link by reacting with an aldimine bond formed between a lysine
aldehyde and hydroxylysine residue.
With respect to tissue source, type I collagen is the most widely
occurring collagen found in skin, tendon, bone, cornea, lung, and the
vasculature, while type II collagen has a more specific tissue distribution
being limited essentially to cartilage, while type III is found in relatively
elastic tissues such as embryonic skin, lung, and blood vessels (Hulmes,
2008). For most nonfibrillar collagens (e.g., type IV, VI, and VII collagens),
disulfide bonds may be the only source of intra- and intermolecular
covalent bonds. There are usually no lysine-mediated cross-links in
these collagens (Eyre and Wu, 2005). The best known nonfibrillar collagen
is type IV collagen, which is a basement membrane collagen that forms
specialized structures found at tissue boundaries, and in fat, muscle, and
nerve cells. Collagen VI, on the other hand, is important in maintaining
tissue integrity (Hulmes, 2008).

B. Collagen–gelatin conversion
There are several methods used by the industry to manufacture gelatin
from collagen. The main purpose of the gelatin production process is to
first remove unwanted materials that will interfere with the gelatin
extraction and then to convert collagen that is insoluble in water into
gelatin that is soluble in water, while obtaining maximum yield and
superior functional properties (Hinterwaldner, 1977). In general, gelatin
 Author's personal copy
Fish Gelatin 125

is obtained using a sequence of three types of processing steps: pretreat-

ments to remove noncollagen impurities and prepare the collagen for
extraction, one or more water extraction steps to convert collagen into
gelatin, and finally, a series of refinement and recovery processes to get a
highly purified dried gelatin. A limited amount of gelatin has been
marketed in liquid form to avoid the drying step (J.M. Regenstein, per-
sonal communication).
In the first step, raw materials are water washed to remove obvious
impurities and then treated with alkali and/or acid to weaken the colla-
gen structure by breaking intramolecular cross-linkages including cova-
lent and hydrogen bonds and to release other impurities. Some size
reduction may also be applied to increase the efficiency of the process.
In the second step, the actual water extraction is performed at warm
temperatures for an appropriate period of time. In the last step, extracted
gelatin is subjected to several separation methods, including filtration,
evaporation, and deionization, followed by drying and grinding
(Hinterwaldner, 1977).
Gelatins are classified according to whether an acid or an alkali is used
in the final preextraction step. If an acid solution is used as the final
solvent, type-A gelatin (acid process) is obtained. In case of alkali as the
final solvent, type-B gelatin (alkali process) is obtained (Hinterwaldner,
1977). Type-A gelatin’s isoelectric point is higher compared to that of
type-B gelatin, as a milder acid process does not remove the amide
nitrogen of glutamine and asparagine, therefore, the resulting gelatin’s
isoelectric point might be as high as 9.4. If a more severe acid treatment is
required, then some of the amide groups are hydrolyzed and the isoelec-
tric point would be similar to that of the original collagen molecule, which
generally lies between 6 and 8. Type-B gelatin’s isoelectric point might be
as low as 4.8, as the alkali process results in the loss of the amide groups
(Eastoe and Leach, 1977).
In the acid process, the bones and skins are treated in a vessel contain-
ing a dilute solution of acid for a predetermined period of time. Then, the
acid is washed out with cold water. In the alkali process, the deminer-
alized bones (demineralization is mostly done with acid solutions to
remove calcium and other salts from the bone to prepare the collagen-
rich bone material known as ossein) are placed in liming pits and soaked
in a lime suspension for longer than 60 days. For the hides or skins, a
caustic soda solution is used for a shorter period of time. After this
treatment, the raw material is washed thoroughly to remove any residual
lime. The acid pretreatment is mostly used for skin, while the alkali
pretreatment is mostly used for bones (Petersen and Yates, 1977).
The actual extraction method used for both acid and alkali pretreated
raw materials is similar. The main extraction step is done using hot water
at controlled temperatures, mostly higher than 40
C, and it is the most
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126 Gokhan Boran and Joe M. Regenstein

important step in gelatin production. In the industry, the extraction step is

usually multiple extractions performed with gradually increasing
temperatures beginning from 50 to 60
C and going up to the boiling
temperatures, usually in 5–10
C temperature increment. Gelatins are
collected so that the lower temperature fractions have minimal degrada-
tion and the higher temperature fractions have more variable molecular
weights (Hinterwaldner, 1977). The dilute gelatin solution from the
extraction process is clarified using lamellar separators (this equipment
is built as a set of plates or discs that are arranged at such an angle that the
solids can slide off into the sludge chamber, thus achieving clarification)
and filtered using self-cleaning centrifugal filters or cellulosic filters. After
that, gelatin solutions are deionized by passing through ion exchangers
and concentrated, usually in a multiple effect vacuum evaporator. The
concentrated solution is then sterilized by hot air in batch driers, and then
cooled or chilled to rapidly form a gel. Then, the gel obtained is extruded
to get gelatin noodles (i.e., continuous strands), followed by a final drying
and grinding process. After all these treatments, gelatin granules or
powder are obtained. Acid or alkali pretreatments designed to destroy
or weaken cross-linkages between the a-chains or between tropocollagen
molecules need to be adjusted not only in terms of concentration but also
treatment time to avoid extensive degradation of collagen, which might
result in lower quality gelatin. But at the same time, enough degradation
is needed to be able to get a higher yield and acceptable gel strength
(Holzer, 1996).

C. The mechanism of gelation

Hydrogen bonds certainly play an important role in gelation (Johns and
Courts, 1977). Gelation can be considered as gelatin regaining its colla-
gen structure, but this would not be exactly correct, because the conver-
sion of collagen into gelatin is an irreversible process although gelatin
can partially regain some collagen structure by recovering some cross-
linkages. The greater the amount of cross-linkages recovered, the higher
the gel strength and viscosity along with the melting and gelling tem-
peratures (Belitz et al., 2004). The concentration of a-chains and the
cooling rate are the most important factors affecting the final gelation.
At high gelatin concentrations intermolecular bond formation would
occur with multiple strands, while the same process is more likely to
occur as intramolecular bonds within a single strand at low concentra-
tions. Similarly, slow rates of cooling allow more intra- and intermolec-
ular cross-link formation, while rapid cooling does not allow that to
happen (Belitz et al., 2004).
 Author's personal copy
Fish Gelatin 127

D. The characteristics of gelatin

Gelatin is a gelling agent that is able to form thermoreversible gels, which
means that when the gelatin gel is heated, it softens and again turns into a
liquid. Then, it is able to return to the gel form when the solution is cooled
again (Stainsby, 1977). Being able to melt below human body temperature
makes its use very favorable in the food industry since gelatin is able to
melt and release flavor when it is taken into the mouth, which is desired
in terms of the sensorial properties of food products (Choi and
Regenstein, 2000). Another important characteristic of gelatin is that its
gel strength is relatively higher than most of the common gelling agents,
which are usually carbohydrates and obtained from vegetable sources
(Badii and Howell, 2006). The gap between melting and gelling tempera-
ture of gelatin gels is smaller than that of other gelling agents, which is
desired for some particular applications, that is, food applications includ-
ing jellies and custards (Jones, 1977).

E. Sources of raw material used in gelatin manufacturing

Pork skin is the most abundantly used raw material in gelatin production.
About 45% of the world’s total gelatin production is obtained from pork
skin, followed by bovine hides with almost 30% (Karim and Bahat, 2009),
and 23% of gelatin is obtained from bovine and porcine bones. Other
sources include chicken and fish, but these account for only 1.5% of the
world’s annual gelatin production. In Europe, pork skin is the most
abundantly used raw material for gelatin production, accounting for
around 80% of the total, followed by cattle skin with 15% of the total
gelatin production. The remaining 5% is from pork and cattle bones, fish,
and chicken.
Recent studies have shown that fish skin, especially, might be an
alternative source for gelatin production. Fish skin gelatins may provide
better alternatives for some applications because of their, at times, rela-
tively lower gel strengths and melting temperatures compared to that of
mammalian gelatins. These characteristics may be desired in some food
systems for ease of flavor release, leading to better sensory characteristics
(Boran and Regenstein, 2009; Choi and Regenstein, 2000). In addition,
obtaining valuable by-products from the fishery industry and reducing
waste have made it an attractive research topic. As the issues of sustain-
ability and the better use of harvested resources become more critical, the
need to use fish waste more productively will only increase. Many fish
species have been investigated as a raw material for gelatin extraction and
the properties of the gelatins obtained from these sources have also been
reported.
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128 Gokhan Boran and Joe M. Regenstein

The waste from fish processing after filleting can account for as much
as 75% of the total catch weight (Shahidi, 1995). It includes the heads, skin,
and scales, guts/internal organs, frames (bone rack with adhering meat),
and trim (pieces cut from the fillets during processing) (Regenstein, 2004).
About 30% of such waste consists of skin, bone, and scale with high
collagen content that could be used to produce collagen and gelatin
(Gómez-Guillén et al., 2002; Sadowska et al., 2003; Young and Lorimer,
1960). Very little work has been done on fish bone gelatin (Muyonga et al.,
2004), while a little more work has been done on scale gelatin. Fish scales
gelatin has been obtained from sardine (Harada et al., 2007), Asian carp
(Wang and Regenstein, 2009), and lizardfish (Wangtueai and Noomhorm,
2009).

F. The gelatin market

The world’s total gelatin production is close to 350,000 tons annually,
accounting for the market’s economic value being over US$ 2 billion.
Again, gelatin is used in many products, including foods, pharmaceuti-
cals, cosmetics, photographic films, paints, fertilizers, and many more, as
a gelling agent, stabilizer, and structure enhancer (Jones, 1977).

G. Industrial applications of gelatin

Gelatin’s largest single use is in food products, more specifically, water
gel desserts. Gelatin desserts might consist of several other gelling agents
(i.e., agar, carrageenan) along with gelatin; however, gelatin gels usually
melt at temperatures that are lower than the body temperature, which
makes gelatin favorable compared to other gelling agents in food applica-
tions. In some parts of the world, water dessert gels are made from
carrageenan and these have to be chewed as they do not melt in the
mouth. Thus, though both are called water dessert gels, they have very
different sensory properties.
Gelatin is used in dairy products as a stabilizer and as an ingredient to
modify the texture. It is used in yogurt, ice cream, and other dairy
products. Gelatin is added to yogurt to reduce syneresis and to increase
firmness. Gelatin is an ingredient compatible with milk proteins and
improves the sensory perception by not masking the product flavor as
much as some other gums (Jones, 1977). The use of different concentra-
tions of gelatin would provide the manufacturer with the possibility of
obtaining a wide range of textures in food products. Gelatin is widely
used in confectionery products, including soft gummy-type candies and
marshmallows. Gelatin is the main gelling agent in gummy-type candies.
Marshmallows usually contain about 3% gelatin, in which gelatin serves
as a stabilizer and whipping agent ( Jones, 1977).
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Fish Gelatin 129

Gelatin is also used in nonfood products, including drugs, cosmetics,

photographic films, and paper and paint products. Pharmaceutical gela-
tin accounts for a significant proportion of the total production and it is
used in the production of capsules, tablets, and pastilles (Wood, 1977).
Gelatin is used for both soft and hard capsules. The gelatin protects the
drugs during distribution and they are not released until after they are in
the stomach. Gelatin acts as a binding agent in tablets. It is also used for
tablet coating to reduce dusting, to mask unpleasant tastes, and to allow
printing and color coatings for product identification.
Gelatin has been used for photographic emulsions for more than
100 years. It is still the principal constituent of the binder in the most
commercial photographic films and papers (Kragh, 1977).

II. FISH GELATIN

In the last decade, gelatin extraction from fish skin has been intensively
investigated. The physicochemical, textural, rheological, and sensory
properties of extracted fish gelatin have also been studied in comparison
with mammalian gelatin. The overall results suggest that fish skin might
be an alternative raw material for high-quality gelatin production, elim-
inating religious concerns shared by the Jewish and Muslim communities
and also providing an alternative and more lucrative way to use some
fishery by-products (Boran and Regenstein, 2009), thus also improving
the overall sustainability of the fishing industry. Some of the fish species
investigated include Atlantic salmon (Arnesen and Gildberg, 2007), cod
(Gudmundsson and Hafsteinsson, 1997), sin croaker and short fin scad
(Cheow et al., 2007), Alaska pollock (Zhou and Regenstein, 2004), big eye
snapper and brown stripe red snapper (Jongjareonrak et al., 2006), yellow
fin tuna (Cho et al., 2005), Nile perch (Muyonga et al., 2004), black and red
tilapia (Jamilah and Harvinder, 2002), grass carp (Kasankala et al., 2007),
and silver carp (Boran and Regenstein, 2009).

A. Common and potential sources

Gelatin can be obtained from various marine and freshwater sources. The
species available for gelatin production are divided into three categories:
marine invertebrates, sea mammals, and fishes. Based on their living
environments, fishes are usually subdivided into four groups: hot-water
fish, warm-water fish, cold-water fish, and ice-water fish. Cold-water
fishes, such as pollock, cod, and salmon, account for a large part of the
commercial fish capture. They are often processed into skinned and
boneless fillets, leaving large amounts of fish skin, scales, and bones as
waste or as a raw material that is sometimes wasted, sometimes used for
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130 Gokhan Boran and Joe M. Regenstein

fish meal, occasionally for leather, and which would be appropriate for
gelatin production. These by-products, especially the skin, usually con-
tain a large amount of protein, most of which is collagen. Warm-water
fishes account for most freshwater fish aquaculture production, and
currently, many commercial fish gelatins come from these fish species.

B. Properties of fish gelatin

1. Physical attributes
a. Gel strength Gel strength is one of the most important quality char-
acteristics used in the gelatin industry to differentiate gelatins. As mea-
suring gel strength is very popular, there is a standard method. According
to the standard method (Gelatin Manufacturers Institute of America,
GMIA), gel strength must be measured at 10
C on a gelatin sample
prepared at 6.67% concentration of protein (w/v). Thus, if the gelatin is
contaminated with other proteins, the amount of actual gelatin evaluated
would be less that 6.67%. For practical applications, this does not matter
as it allows the user to evaluate the ‘‘gelatin’’ they are using. For scientific
purposes, however, this may underestimate the quality of the actual
gelatin extracted. Another concern is with the protein determination
itself. Because gelatin has a unique amino acid composition, a special
conversion factor is needed for the Kjeldahl test, and work has shown
that calibration of the Biuret or Lowry procedures may also require
adjustments for an accurate measurement (Zhou and Regenstein, 2006).
The methods for dissolving gelatin in water is not standardized and
there are variations in the procedures used, that is, different water tem-
peratures, different durations of sitting at the higher temperature before
cooling, with or without stirring, etc. Maturation time and temperature are
standardized and are generally followed by most investigators, that is,
16–18 h at 10
C. A particular jar is used for this measurement, called a
‘‘bloom jar’’ (Fig. 5.2), it requires about 155 ml of gelatin solution, which
corresponds to about 10 g of gelatin. However, this particular jar cannot be
regularly used in many scientific studies because it requires such a sub-
stantial amount of sample, which is often limited in scientific studies.
Therefore, many scientists use other containers that differ in size and
shape leading to significant differences in the results, making the data
incomparable among the different studies. Because of sample limitations,
in some cases, a lower percentage of gelatin is used, often 3.33%. The test
settings are also standardized: It is the force required for a 12.7-mm diame-
ter flat probe to penetrate 4 mm into the gel that is being lowered into the
sample at a speed of 1 mm/s. This force, expressed in grams, is then
reported as the gel strength. If done totally according to the official meth-
odology, the resulting gel strength may be called the ‘‘Bloom strength.’’
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Fish Gelatin 131

FIGURE 5.2 Standard ‘‘bloom jar’’ provided by Texture Technologies Corporation

(Scarsdale, NY).

There are different instruments that can be used to handle the probe and
the different instruments may give different results (Table 5.1).

b. Viscosity Viscosity is generally measured using tubular glass visc-

ometers as they are relatively inexpensive and easy to use compared to
expensive and complicated computer-controlled instruments. Although
advanced viscosity instruments might provide higher reproducibility and
accuracy, the tubular glass viscometers also give high precision. Com-
pared to gel strength, viscosity is not as well correlated with textural
properties. The viscosity is mostly affected by molecular weight
distribution.
Gelatin samples with high molecular weight fractions give high vis-
cosity but that does not necessarily mean that their gel strengths would
also be high. Gelatin samples from fish skin, for example, give unexpect-
edly high viscosity while giving low gel strength compared to that of pork
skin gelatin due to the carefully controlled extraction conditions and
consequently, the presence of higher molecular weight protein fractions
(Boran and Regenstein, 2009). Arnesen and Gildberg (2007) reported that
Atlantic salmon and Atlantic cod skin gelatin had higher viscosities than
pork skin gelatin while giving lower gel strengths than pork skin gelatin.
Generally, fish skin gelatins are expected to have a lower viscosity com-
pared to that of gelatins obtained from porcine and bovine sources with
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132 Gokhan Boran and Joe M. Regenstein

TABLE 5.1 Gel strength of a commercial gelatin measured using different instruments
and probes in either a standard bloom jar or a 15-ml capacity small plastic jar

Measurement details Average SD

Standard bloom jar, TAXT2 texture analyzer, spherical 242 5.7

probe
Standard bloom jar, TAXT2 texture analyzer, cylindrical 523 2.1
probe
Standard bloom jar, Stevens texture analyzer, spherical 213 1.2
probe
Standard bloom jar, Stevens texture analyzer, cylindrical 466 3.8
probe
Small plastic jar, TAXT2 texture analyzer, spherical probe 320 9.2
Small plastic jar, TAXT2 texture analyzer, cylindrical probe 814 13.6
Small plastic jar, Stevens texture analyzer, spherical probe 294 2.5
Small plastic jar, Stevens texture analyzer, cylindrical probe 746 9.6

SD: standard deviation.Same sample (Knox Gelatin, Kraft Foods Global, Inc., Glenview, IL, USA) used for
the measurements: 6.67% gelatin, dissolved at 50
C for 30 min in distilled water, matured at 4
C for 16–18 h.
The measurements are done at 4
C using the following settings: 4 mm penetration with 12.7 mm diameter
probe (either spherical or cylindrical) with 1 mm/s penetration speed. Gel strength is given as g force required
penetrating the probe onto the sample (N ¼ 3).

similar molecular weight distributions. However, most investigators do

not look at the molecular weight distribution beyond looking at the SDS-
PAGE electrophoretigram. Because the distribution of peptides can be so
great, much of it is ‘‘smeared’’ over the gel and cannot be easily quanti-
tated. To do more is a lot of work and would likely be different for each
preparation, so it has not been done critically.

c. Melting and gelling temperature Rheological methods have recently

gained importance and have found applications in the determination of
gelatin quality. Rheological measurements of both melting and gelling
temperatures give highly reproducible results. A temperature sweep test
is performed for this purpose. Heating or cooling is required to determine
the melting and gelling temperature, respectively. The gelatin gel sample
is prepared at a certain concentration and matured at a certain tempera-
ture for a certain period of time to standardize the procedure to be able to
discriminate the samples based on their chemical differences (Chiou et al.,
2006; Cho et al., 2006; Fernandez-Diaz et al., 2003; Kasankala et al., 2007).
Prior to rheological determinations of melting and gelling temperatures,
the droplet method was used as a standard method for determining the
melting temperature. However, the rheological methods have replaced
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Fish Gelatin 133

this older method (Wainewright, 1977), which was less precise and more
laborious. A drop of an organic solvent with a dye in it, originally carbon
tetrachloride, but more recently chloroform, with the recognition of the
toxicity of carbon tetrachloride, has been used. The temperature when the
droplet fell is taken as the melting temperature—but is it the beginning of
the fall, the middle or when it touches the bottom. This is not well spelled
out. Obviously, the rate of heating will also affect the results both because
of differences in heat penetration and in the accuracy of the reading;
slower being better but taking longer!
The rheological test measured the stress/strain as a function of time
and temperature. Again, an arbitrary heating rate is selected. The mid-
point of the transition is usually considered to be the melting or gelling
temperature. The amount of cooperativity of the system can be deter-
mined by the width of the curve (in degrees) usually measured from the
25% point of transition to the 75% point of transition. The greater the
cooperativity (or uniformity of the material), the smaller the width
observed.
Other rheological tests, including time sweep, frequency sweep, stress
sweep, and strain sweep, have also found applications in determination
of gelatin quality, as they allow researchers to discriminate the gelatin
gels according to their strength and elasticity. Stress and strain sweep
tests are used to determine the linear viscoelastic region of the gels.
Frequency sweep tests are useful to determine if the gelatin gels change
with the changing frequency of the stress applied. Time sweep tests are
used to determine if the gelatin gels’ viscoelastic properties change with
time at a controlled temperature and at a set level of stress applied.
The recent literature on fish gelatin includes some examples of these
tests used to make comparisons among gelatin samples from different
sources. Chiou et al. (2006) used temperature sweep tests to determine the
melting and gelling temperature of gelatin gels. They also used time
sweep tests to show the increasing elastic modulus at different tempera-
tures with an increasing concentration of glutaraldehyde added to the
gelatin gels. Gudmundsson (2002) used frequency sweep tests success-
fully to differentiate the gelatin gels based on their elastic moduli and the
temperature sweep tests to determine the melting temperature of blended
fish gelatin gels. Zhou and Regenstein (2007) used temperature sweep
tests to compare the melting temperatures of gelatin gels from different
sources. In another study, Zhou et al. (2006) used strain sweep and
frequency sweep tests to compare the gelatin gels from different sources
based on their viscoelastic properties. Recent studies gave good examples
of how rheological measurements had strong correlations with conven-
tional parameters, including gel strength and viscosity (Gilsenan and
Ross-Murphy, 2000; Gudmundsson, 2002; Zhou et al., 2006).
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134 Gokhan Boran and Joe M. Regenstein

2. Chemical characteristics
a. Amino acid composition Chiou et al. (2006) showed that differences in
the amino acid composition have significant effects on the melting and
setting temperatures of gelatin obtained from different sources. Accord-
ing to their results, the higher proline and hydroxyproline content of pork
gelatin correlated with stronger gels having higher gelling temperatures.
Proline and hydroxyproline are, however, not the only amino acids hav-
ing significant effects on gelatin structure. The content of glutamic acid,
aspartic acid, lysine, hydroxylysine, arginine, and histidine are also
important in cross-link formation and electrostatic interactions. As colla-
gen usually lacks cystine, there are no disulfide bonds in the collagen
structure. Collagen is mostly stabilized by hydrogen bonds formed
between side chains of the amino acids and water in addition to the
twisted structure enforced by the high content of proline and hydroxy-
proline along with intra- and intermolecular cross-links (Engel and
Bachinger, 2005).

b. Peptide size in relation to quality Collagen-containing tissues are

treated with acid and/or alkali followed by a heat treatment in the
presence of water to break the structure of the collagen fibrils irrevers-
ibly to obtain gelatin (Eastoe and Leach, 1977). While the molecular
weight of the collagen molecule is about 330 kDa, gelatin is generally
considered to be all collagen fractions with a molecular weight higher
than an arbitrary minimum of 30 kDa. When most fractions are below
30 kDa, the product is usually called a gelatin hydrolysate, as such
products are not able to form a gel, although these peptides are believed
to participate in gel formation (Eastoe and Leach, 1977). A heat treat-
ment of about 40
C breaks hydrogen and electrostatic bonds in newly
formed collagen molecules releasing single a-chains, but this is insuffi-
cient to break the cross-links and covalent bonds in the collagen struc-
ture of mature collagen (Eastoe and Leach, 1977). With treatments at
higher temperatures, on the other hand, those covalent bonds, including
intermolecular cross-links and peptide bonds, break down and there-
fore, smaller a-chain fractions are obtained (Eastoe and Leach, 1977).
The position of the bond breaks determines the molecular weight and
the number of polypeptide chains. As amino acid sequence and compo-
sition of collagens from different sources vary greatly, bond breaks
appear to be relatively random and these random bond breakdowns
are the main cause of molecular heterogeneity in gelatin (Eastoe and
Leach, 1977).
The raw materials used in gelatin production contain a variety of
substances that are the source of organic and/or inorganic impurities in
gelatin. Noncollagen protein fractions, lipids, nucleic acids, and other cell
 Author's personal copy
Fish Gelatin 135

components are among the organic impurities. Inorganic impurities

include naturally present minerals such as calcium, sodium, potassium,
and iron along with those derived from substances added for gelatin
preparation, that is, acid and/or alkali with their impurities (Eastoe and
Leach, 1977). Finally, commercial gelatin products contain a substantial
amount of water usually as the second largest component in the whole
and its amount varies greatly based on the drying process applied, the
nature of the raw material, and the temperature and relative humidity
of storage. Generally speaking, the water in gelatin is between 9%
and 14% with occasional samples outside of this range (Eastoe and
Leach, 1977).

C. Other factors affecting quality

There are several factors that significantly affect the properties of gelatin
(Cho et al., 2006). The raw materials used in gelatin manufacture have
obvious effects on gelatin, mostly originating from differences in the
amino acid composition of the collagen of the raw material. Also, varia-
tions in processing conditions such as extraction time, extraction temper-
ature, and concentration of acid or alkali dramatically affect the product
(Boran and Regenstein, 2009; Cho et al., 2006; Hinterwaldner, 1977; Zhou
and Regenstein, 2005). For example, longer extraction temperatures and/
or higher extraction temperatures cause excessive damage to the collagen
molecule and the resulting gelatins form weak gels and have low viscos-
ity. Similarly, excessive concentrations of acid and/or alkali can cause
degradation of collagen structure giving a gelatin with lower functional
values.

1. The effects of processing conditions

The extraction conditions greatly affect the quality of gelatin as shown
by many researchers. Therefore, optimization of extraction is of great
interest to obtain the best possible product by eliminating and/or mini-
mizing the excessive effects of extraction treatments causing extensive
damage to collagen molecules. The most important extraction para-
meters are acid and/or alkali treatments, more specifically, the concen-
tration of acid and/or alkali and duration of the treatment; extraction
temperature and duration, and the conditions of refining and drying
processes. A general flowchart of gelatin extraction from fish skin is
given later and it shows that those treatments greatly affect the quality
of the gelatin (Fig. 5.3).

a. Alkali and acid treatments The gel strength of gelatin is greatly influ-
enced by the concentration of acid and/or alkali, the duration of the acid
and/or alkali treatment, and possibly the treatment temperature. A study
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136 Gokhan Boran and Joe M. Regenstein

Fish skin
Washing (cold water)
Removal of impurities
Washing (cold water)
Storage until processing (−20 ⬚C)

Cutting skin

Alkali treatment
Washing (cold water) Filtration
Acid treatment
Washing (cold water) Filtration
Water extraction (over 40 ⬚C)
Filtration
Drying

Gelatin sheets

FIGURE 5.3 A general flowchart of gelatin extraction from fish skin.

by Gudmundsson and Hafsteinsson (1997) showed that high concentra-

tions of alkali or acid increased the gelatin yield while decreasing gel
strength. These results have also been confirmed by Zhou and Regenstein
(2005). Another study done by Cho et al. (2006) showed that alkali con-
centration up to 1.5% increased gelatin yield significantly. Zhou and
Regenstein (2004) confirmed that the concentrations of acid or alkali
have a significant effect on gelatin yield, gel strength, and viscosity.
Acid treatment is also important for the sensory aspects of gelatin,
appearance and smell, as the acid treatment effectively removes odors
and color that originate from the raw material (Boran and Regenstein,
2009; Zhang et al., 2007). Alkali treatment is, similarly, important and
responsible for removal of possible impurities from the raw material
and also for weakening the collagen structure, leading to higher yield
and superior quality. In addition, alkali treatment causes glutamine and
asparagine to lose their amine groups, converting them to glutamic and
aspartic acid residues, respectively, lowering the isoelectric point of col-
lagen (Johns and Courts, 1977). Therefore, both acid and alkali treatments
need to be optimized for pH, duration, and temperature of extraction.
Zhou and Regenstein (2005) also did their pretreatment at low tempera-
tures, which led to a much higher amount of the full a-chains. They
suggest that the low temperature may have prevented any native pro-
teases/collagenases from degrading the collagen prior to the extraction.
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Fish Gelatin 137

The main extraction step although normally done in water can be done
at using mild acid or alkali conditions. Acid or alkali treatments are useful
for a more effective extraction, increasing the yield and shortening the
extraction time. Zhou and Regenstein (2005) showed that acidic conditions
are more favorable for higher gelatin yield. However, acidic conditions
also cause low gel strength, which is not desired in most gelatin applica-
tions. The isoelectric point of collagen is around 6–6.5, depending on the
amino acid composition, specifically the content of the acidic and basic
amino acids of collagen, which vary both due to source and to processing
conditions, for example, due to the impact of processing on glutamine and
asparagine. The isoelectric point of purified collagen is difficult to measure
because collagen is difficult to isolate in its natural form as it is not readily
soluble in water at room temperature and when it is dissolved with the help
of heat treatment, collagen loses its natural state. Therefore, the isoelectric
point measured does not reflect the physiological isoelectric point of the
collagen, but many researchers agree on the approximate value of 7.0 for
the isoelectric point of collagen under physiological conditions (Johns and
Courts, 1977). Neutral extracts of untreated tissues of pork and rabbit skin,
for example, had isoelectric points in the range of pH 5.6 and 6.8, respec-
tively (Johns and Courts, 1977). A pH that is higher or lower than the
isoelectric point results in higher extraction yield as collagen is less tightly
bound at pH values different from its isoelectric point. The net charge of the
collagen molecule is zero at the isoelectric point where there are an equal
number of positive and negative charges on the molecule allowing it to
form the maximum number of intermolecular salt bonds and electrostatic
interactions, which strengthen and stabilize the structure of the collagen.
However, the isoelectric point also reflects in part the binding of other ions
that change the isoelectric point. The most accurate figure would be the
isoionic point, which would be the extrapolation of the isoelectric point to
pure water (where the isoelectric point measurement is difficult because
there is no charge). According to the application in which gelatin is used,
the effect of pH on gelatin needs to be carefully considered and the pH of
the extraction solution needs to be adjusted to get a high quality gelatin. For
example, as type-A gelatin has a higher isoelectric point, its use is favorable
in those applications that require low pH at which the gelatin would be
conducive to forming gel networks. Similarly, as type-B gelatin has a low
isoelectric point, it is used in those applications that require a higher pH at
which the gelatin is readily available for formation of the gel network.

b. Extraction temperature and duration Different temperatures and

times are used in gelatin manufacturing but most extractions are between
45 and 60
C. Temperatures from 50 until 80
C can promote intramolec-
ular bond formation between strands and consequently gelatin with
stronger gelling ability can be obtained (Djagny et al., 2001). Higher
 Author's personal copy
138 Gokhan Boran and Joe M. Regenstein

temperatures over 80
C, however, result in fracturing of intramolecular

chains giving gelatin having a weaker gelling ability. Lower extraction
temperatures, on the other hand, lead to low yields but a superior quality.
Similarly, longer extraction times give better yield, while the extracted
material suffers from low gel strength and viscosity due to excessive
damage to collagen fractions with longer heating and possibly extraction
of other proteins. Therefore, it is necessary to balance both extraction
temperature and duration of the extraction, to get the best possible out-
come. For this purpose, a few modern optimization studies have been
done on gelatin extraction from skins of different fish species (Boran and
Regenstein, 2009; Kasankala et al., 2007; Zhou and Regenstein, 2004).

c. Storage and transportation There are many other factors affecting

gelatin properties. Going into detail for each one of them is beyond the
scope of this chapter. To be brief, every processing step, especially if heat
is involved, has an effect on gelatin properties including yield, gel
strength, melting and gelling temperatures, and viscosity. Raw materials
are important with respect to purity and ease of processing. The actual
type of the acid and/or alkali used in the pretreatment and/or the
extraction is also important. Freshness and storage of raw materials, any
possible microbial contamination or the presence of microbial or meta-
bolic enzymes, the water content of gelatin, and transportation conditions
are other factors that can affect the quality of gelatin.

D. Quality of fish gelatin compared to mammalian gelatins

Arnesen and Gildberg (2007) studied the skins of Atlantic salmon and
Atlantic cod for gelatin production and reported that Atlantic salmon
skin gelatins had higher gel strength and gelling temperatures than Atlan-
tic cod skin gelatins. The gel strength of the salmon and cod were found to
be 108 and 71 g, respectively, while their gelling temperatures were 12 and
10
C, respectively. Arnesen and Gildberg (2007) also reported that the gel
strength of the gelatins obtained increased with storage time and higher
extraction temperature resulted in lower gel strength. Gudmundsson and
Hafsteinsson (1997) also studied cod skin as a raw material for gelatin
production, reporting that the proline and hydroxyproline content of cod
(a cold-water species) skin gelatin ( 18%) was lower compared to that of
tilapia (a warm-water species) skin gelatin ( 25%), resulting in relatively
lower gel strength and viscosity. According to their results, tilapia skin
gelatin gave 260 g bloom strength, while cod skin gelatin had 180 g bloom
under the best extraction conditions reported. Choi and Regenstein (2000)
compared various gelatin samples from different sources in terms of their
physicochemical and sensory properties and reported that Alaska pollock
gelatin had lower gel strength along with lower melting temperature
 Author's personal copy
Fish Gelatin 139

compared to that of pork skin gelatin. Alaska pollock gelatin melted at

24
C while the pork skin gelatin melted at 29
C (Choi and Regenstein,
2000). They also compared the sensory properties of gelatin gels prepared
from Alaska pollock and pork skin gelatins and reported that a low melting
temperature and gel strength might be useful in creating products with a
faster and stronger flavor release. Chiou et al. (2006) studied Alaska pollock
and Alaska pink salmon for gelatin production and the quality of the
gelatin obtained in comparison with pork skin gelatin. They reported that
Alaska pollock and Alaska pink salmon skin gelatins had lower melting
and gelling temperatures along with lower gel strength compared to that of
pork skin gelatin due to the lower proline and hydroxyproline content of
skin gelatins obtained from these fish species. They reported that the
pollock and salmon skin gelatins had gelling temperatures of 7 and 5
C,
respectively, while pork skin was reported to have a gelling temperature of
24
C, which was attributed to the high content of proline and hydroxypro-
line of the pork skin gelatin (Chiou et al., 2006).
Kasankala et al. (2007) studied grass carp skin as an alternative raw
material for gelatin production and reported that the hydroxyproline con-
tent of grass carp skin gelatin (11.3%) was slightly higher than that of
bovine skin gelatin (11.2%) and a little lower than that of pork skin gelatin
(13.2%). They also reported high gel strength, melting and gelling tempera-
tures for grass carp skin gelatin compared to that of gelatins obtained from
other fish species. According to their results, the carp skin gelatin had a
gelling temperature around 19
C and a melting temperature around 26
C,
which was a little lower than that of pork skin (25 and 31
C, respectively)
and bovine gelatins (21 and 30
C, respectively) (Kasankala et al., 2007).
Boran and Regenstein (2009) also reported similar results for skin gelatin
obtained from silver carp, another Asian carp species, that is, it had high gel
strength (600 g for optimized gelatin) possibly due to the high hydroxypro-
line content ( 11%). Therefore, it does appear that the assumption that
there is a strong connection between the content of hydroxyproline and
proline and the physicochemical properties of gelatins continues to hold
with the more recent research with fish gelatins.

III. METHODOLOGICAL CHALLENGES IN QUALITY

DETERMINATION OF GELATIN

Both commercial gelatin powders and those produced on a small scale for
research purposes have an amount of water that varies due to the differ-
ences in processing and drying methods (Eastoe and Leach, 1977). Water
content of gelatin is important for both ease and duration of storage, as
high water content favors microbial spoilage. In addition, higher water
containing gelatin formulations can be sold for less. The drying method is
 Author's personal copy
140 Gokhan Boran and Joe M. Regenstein

the major factor affecting the water content of gelatin products. Heat
drying and freeze drying are two of the most common methods used to
remove water from gelatin preparations. Heat drying is generally done at
low temperatures between 40 and 60
C for several hours to several days
(Hinterwaldner, 1977). Freeze drying is a much faster method compared
to heat drying and able to remove water while causing less damage to the
gelatin, but it is generally more expensive.
The gelatin powder obtained is generally not tested for its water
content, and even when determined, this information is not generally
included in the calculations when preparing samples for testing, that is,
the gelatin is simply weighed out. This might lead to a lack of agreement
between data from different sources. To prevent confusion and to get
comparable data, water content of gelatin samples should be determined
and included in the calculations to make sure that the actual gelatin
amount is the same in each sample being compared for their character-
istics. After maturation, making a direct comparison of two gelatin sam-
ples with different amounts of gelatin for gel strength would be erroneous
as the actual gelatin concentration of the samples is different.

IV. CONCLUSIONS AND SUGGESTED READINGS

Previous studies done on gelatin have shown that there are clear connections
between gelatin’s functional properties and the extraction conditions. While
higher extraction temperatures and durations result in higher yield, the
gelatin obtained is of poorer quality due to damage to the collagen fractions.
Similarly, higher acid and/or alkali concentrations result in higher yield
along with purer material, but the gelatin obtained lacks necessary func-
tional properties. Therefore, an optimization of manufacturing process of
gelatin is needed to get a final product with desired properties.
This chapter only covers general information on collagen and gelatin
and a limited introduction to fish gelatin. For a thorough understanding
of collagen and gelatin, additional sources should be consulted. For this
purpose, an excellent book of contributed chapters edited by Ward and
Courts (1977), ‘‘The Science and Technology of Gelatin,’’ covers almost
every area related to gelatin and is still a very useful source of information
for current collagen and gelatin producers and users, and for researchers.

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