Journal of Food Engineering 109 (2012) 337346

Contents lists available at SciVerse ScienceDirect

Journal of Food Engineering

journal homepage: www.elsevier.com/locate/jfoodeng

Review

Isolation and valorisation of vegetable proteins from oilseed plants:

Methods, limitations and potential
Ivo M. Rodrigues a,, Jorge F.J. Coelho b, M. Graa V.S. Carvalho b
a
b

Departamento de Cincia e Tecnologia Alimentar, Escola Superior Agrria, Instituto Politcnico de Coimbra, Bencanta, 3040-316 Coimbra, Portugal
Departamento de Engenharia Qumica, Faculdade de Cincias e Tecnologia, Universidade de Coimbra, Rua Slvio Lima, 3030-790 Coimbra, Portugal

a r t i c l e

i n f o

Article history:
Received 21 June 2011
Received in revised form 17 October 2011
Accepted 22 October 2011
Available online 30 October 2011
Keywords:
Oilseed
Rapeseed
Meal valorisation
Protein

a b s t r a c t
Oilseed plants used for the production of vegetable oil are also natural sources of vegetable proteins, which
have great nutritional value. This manuscript presents a literature review of the production and the application of vegetables proteins obtained from oilseed plants, in particular rapeseed. Some general concepts
related to proteins are overviewed followed by the discussion of the characteristics and composition of
oilseeds and oilseed meal. The techniques and methodologies used to extract and isolate plant proteins
and their formulation for specic applications are also reviewed, including measures to minimize the presence of antinutritional or toxic compounds in the isolates. A valorisation of protein isolates, in particular
from rapeseed, with regard to their nutritional and functional properties is presented as well. It is possible
to produce innumerous added-value proteins products from industrial by-products of oilseed oil production with applications in food and pharmaceutical industries, developing and applying innovative technologies providing environmentally sustainable solutions for agriculture and agro-industry.
2011 Elsevier Ltd. All rights reserved.

Contents
1.

2.
3.
4.

5.

Proteins general remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1.1.
The biological function of proteins from oilseed plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1.2.
The structure characteristics of proteins from oilseed plants. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1.3.
The classification of proteins from oilseed plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1.4.
The definition of food proteins from oilseed plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Oilseeds as a source of vegetable oils and proteins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Rapeseed mealprotein composition and utilization limitations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Valorisation of oilseed meals. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4.1.
Alternative technologies for oilseed meals valorisation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4.2.
Production technology of protein isolates and their functional properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Conclusions and outlook . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Proteins general remarks

Proteins are extremely complex polymers based on up to 20
different amino acids (Fig 1). The amino acids are connected
through amide bonds, also known as peptide bonds. They are made
up of 5055% carbon, 67% hydrogen, 2023% oxygen, 1219%
nitrogen and 0.23% sulfur (Belitz and Grosch, 1999; Damodaran,
2000).

Corresponding author. Tel.: +351 239 802 940; fax: +351 239 802 979.
E-mail address: ivorod@esac.pt (I.M. Rodrigues).
0260-8774/$ - see front matter 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.jfoodeng.2011.10.027

337
337
338
339
340
340
340
342
343
344
345
345

The structural and functional differences in the proteins are the

result of the sequence in which the amino acids are connected,
their size and type and the size of the peptide chain. Therefore,
in theory, there is a limitless number of proteins with unique
properties.

1.1. The biological function of proteins from oilseed plants

The biological functions of the proteins enable their classication as enzymatic catalysts, structural proteins, contractile proteins (myosin, actin, tubulin, hormones (insulin), carrier proteins

338

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

Fig. 1. Twenty amino acids, grouped according to the character of their side chain or R group. Source: Adapted from Huret (2008).

(hemoglobin, transferrin), antibodies (immunoglobulin), reserve

proteins (avalbumin, seed proteins) and protective proteins
(toxins) (Coultate, 1984; Damodaran, 2000).
1.2. The structure characteristics of proteins from oilseed plants
From the structural point of view protein structure is divided
into four levels: primary, secondary, tertiary and quaternary structures (Fig 2).

The primary structure is dened by a linear sequence, in which

the amino acids are bonded together by a covalent bond, through
the peptide bond. This bond is the result of the condensation of
the carboxylic group in the amino acid i with the amide group of
the amino acid i + 1, with the elimination of a water molecule.
The chain size and the order in which the amino acids bind together determines the physical, chemical, structural, biological
and functional properties of a protein. The secondary structure is
related to the spacial disposition of the amino acids sequence in

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

339

Fig. 2. Protein structures. Source: Adapted from Darryl (2010).

certain segments of the peptide chain. They can be usually found in

two secondary periodical structures, such as helicoidal structures
and stem-like structures. The tertiary structure is related to the
spatial disposition occurring when a linear peptide chain, with segments originating from several secondary structures, folds back on
itself to achieve a compact tridimensional form. The formation of
tertiary structures assumes the optimization of hydrophobic, electrostatic and Van der Walls interactions, and also hydrogen bonds
between the several groups of the protein, to reduce the free energy of the molecule. The hydrophobic areas are located, in the
majority of cases, inside the structure, while the hydrophilic areas,
specially the charged ones, are located in the proteins water interface. The quaternary structure refers to the spatial disposition of
proteins with more than one peptide chain. Many biologically
important proteins are dimmers, trimers or tetramers, etc. Any of
these complex quaternaries can be formed by identical sub-unities
(homogeneous) or distinct ones (heterogeneous). The formation of
quaternary structures is largely due to the proteinprotein specic
interactions, none of which are covalent bonds, being hydrogen

bonds or hydrophobic and electrostatic interactions (Damodaran,

2000).
1.3. The classication of proteins from oilseed plants
Proteins are divided into two sub-categories: homoproteins,
formed only by amino acids and heteroproteins conjugated proteins formed by aminoacids and nonprotein components called
prosthetic groups. Some examples of homoproteins are albumine,
queratine and insuline and of heteroproteins are glycoproteins
(ovalbumin, j-casein), phosphoproteins (b and a caseins, and
phosphorylases), lipoproteins (proteins in the yolk of the egg),
metalloproteins (hemoglobin, myoglobin and several enzymes)
and nucleoproteins (ribosomes) (Belitz and Grosch, 1999; Damodaran, 2000).
Proteins can also be classied by their shape as globular or
brosis proteins. The former are spherical or ellipsoidal, resulting
from the enrolling on itself, while the second are stem-like
shaped, formed by linear peptide chains enrolled. The majority

340

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

of enzymes are globular proteins, while the brosis proteins have

a structural function (Belitz and Grosch, 1999; Damodaran,
2000).
1.4. The denition of food proteins from oilseed plants
All biologically synthesized proteins can be used as food proteins.
From a practical point of view, food proteins can be dened as those
which are easily digested, non-toxic, nutritionally adequate, functionally useful and abundant. Traditionally, food proteins are present in milk, meat (including sh and poultry), eggs, cereals,
legumes and oilseeds. Since the functional properties of the proteins
are related to their structural and physico-chemical characteristics,
their behavior in food can only be improved if their physical, chemical, nutritional and functional properties are known, in addition to
the changes undergone during processing (Damodaran, 2000).
2. Oilseeds as a source of vegetable oils and proteins
Oilseeds plants are plants that containing seeds or fruits with a
high level of oils and other food fat, used as an energy reserve
(Samouco, 1998). Oilseeds also possess reasonably balanced quantities of carbohydrates, fats and proteins (Yfera, 1998) and are
characterized not only by their rich composition in oils, but also
by their high level of proteins. Oil can be extracted from the seeds
of these by plants using the appropriate technology, which can
then be used in human food and in the production of biodiesel,
resulting in a high protein fat-free meal. This meal can be valorised
and used in human or animal food (Duran et al., 1980).
The most produced oilseeds worldwide are, in decreasing order,
soybean, rapeseed, cotton, peanut and sunowers, amongst others
(FAO, 2009).
In 2007/2008 soybean production represented more than 50% of
the total (220 out of 393.2 million tons produced), the main producers being the USA (33.5% of total production), followed by Brazil and Argentina (26.6% and 21.8% respectively). These three
nations make up approximately 82% of worldwide soybean production (FAO, 2010).
The soybean, belonging to the family of the Fabaceae, is used in
both animal and human food. As a component in the latter, it is
widely used in vegetarian diets. The oil extracted from the soybean
is used in the production of biodiesel. However, other oilseeds, such
as the rapeseed and sunower tend to be used as the prime materials for biodiesel production, essentially due to their higher level
of oil (Fig 3).
China, Canada and India are the main producers of rapeseed,
and together are the responsible for more than half of worldwide
production in 2007 (56.3%) (FAO, 2010).
Rapeseed is an herbaceous plant belonging to the Cruciferae
family, also known as rape (Ratnayake and Daun, 2004; Shahidi,

1990). Rapeseed oil is extracted from its seeds, used in human

food, but also in the production of biodiesel. Rapeseed leaves are
also used in animal feed. Thus, rapeseed is farmed in many countries for its high level in lipids and medium level of proteins.
Rapeseed and Canola are commonly used names for the cultures
included in the Brassica napus L., Brassica rapa L. (ex. Brassica
campestris L.) and Brassica juncea L. (Ratnayake and Daun, 2004).
Rapeseeds are among the oilseeds with the highest levels of oil
between 45% and 50%. The oil is composed largely of oleic acid,
about 58%, a level similar to that found in olive oil, and 10% of linoleic acid, similar to the level found in soybean oil. Its fatty acids
content is lower than peanut or palm oils, but higher than that
found in soybean, sunower, corn and cotton oils. However, also
erucic acid (whish is a fatty acid described as being medium toxic
to humans) in high doses (Ratnayake and Daun, 2004; Shahidi,
1990), although it can be used as a food additive in small doses
(Temple-Heald, 2004). Rapeseeds also consist of approximately
2530% proteins, and 2530% carbohydrates (d.b.) (Shahidi, 1990).
Proteins and carbohydrates are also the main components of rapeseed meal, the designation given to the product originating from
the oil extraction process. There are also components present at
lower levels such as vitamins and minerals, which are important
from a nutritional point of view, and antinutritional compounds
such as glucosinolates and phytates. Glucosinolates are compounds
responsible for the large reduction in the nutritional level of the
rapeseeds used in animal feed. In order to reduce the risk of consuming rapeseed in its natural state, several varieties have been genetically modied to reduce the levels of erucic acid and glucosinolates
(Shahidi, 1990). In Canada, these are named Canola, a portmanteau
word based on Canadian oil, low acid and registered in 1979 by the
Western Canadian Oilseed Crushers Association (Oplinger et al.,
1989). Canola oil is considered to be one of the healthiest oils available in the market, due to its low level of saturated fat, high level of
monounsaturated fat, and the presence of omega-6 and omega-3
fatty acids. It has a light avor, ideal for cooking and salads (Canola
Council of Canada, 2008; McDonald, 2004).
The increase of oilseed production to satisfy the needs of biodiesel is inevitable, resulting in an increase of agricultural and industrial by-products with a high potential for their valorisation.
However, it will be essential to develop innovative technologies,
providing environmentally sustainable solutions for agriculture
and agro-industry.
3. Rapeseed mealprotein composition and utilization
limitations
The use of vegetable proteins, especially those originating from
oilseeds and cereals, has increased considerably in recent decades.
The vegetable proteins from these plants have been used as alternatives to animal proteins in human nutrition, as functional agents

Fig. 3. Approximate composition of some oilseeds. Source: Adapted from Yfera (1998).

341

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

and bioactive components not only in food but also in cosmetics

and pharmaceuticals (Zivanovic et al., 2010). Vegetable proteins
are usually presented in the form of protein concentrate or protein
isolate, although sometimes its use is limited by its inadequate
properties, such as its solubility (Moure et al., 2006).
Rapeseed meal comprises about 40% (d.b.) protein (Salunkhe
et al., 1992), with an amino acid composition similar to other cereals. Compared to soybean meal, rapeseed presents a lower level of
proteins (Table 1) and lower digestibility, due to the high level of
ber and the presence of glucosinolates. For this reason commercial value of rapeseed meal is signicantly lower and is used as
food only for ruminant animals due to their more complex digestive system. In addition, the rapeseed growth is increased to satisfy
the biodiesel production needs, leading to high amount of rapeseed
meals as byproducts with high potential valorisation.
The main proteins in rapeseed meal are globulins (those soluble
in a saline solution) which comprise approximately 70% of the total, and albumins (those soluble in water). The globulins are also
called cruciferines and the albumins are also known as napines
(Salunkhe et al., 1992; Schwenke et al., 1998; Krause and
Schwenke, 2001). Beside these two reserve proteins, there is also
oleosine, a structural protein associated with lipidic compounds
(Ghodsvali et al., 2005).
The globulins, with a molar mass of 300,000 g/mol, are composed of six sub-unities and form a quaternary structure similar
to that found in storing proteins, but with some structural differences. The polypeptide chains which form the sub-unities are only
partially linked by disulphuret bridges and present a pH level in
the highest isoelectric point, of about 7.0 (Krause and Schwenke,
2001), compared to the normal level of 4.0 (Salunkhe et al., 1992).
The albumins are included in a protein group, which are characterized by their low molar mass, of about 12,00014,000 g/mol.
They are water soluble and present a pH level in its highest isoelectric point, superior to 10. The albumins have a helix structure (typical rigid protein), formed by intra- and intermolecular disulphuret
bonds (Krause and Schwenke, 2001).
The amino acids composition of the rapeseed proteins is quite
similar to that encountered in other seeds. Rapeseed proteins are
rich in glutamic acid, aspartic acid, leucin and prolinase (Shahidi,
1990) and contain considerable amounts of all the essential amino
acids, phenylalanine, isoleucine, leucine and tryptophan. When
compared with soybean meal, as shown Table 2, rapeseed meal
contains similar levels of riboavin, thiamine, biotin and folic acid,
higher levels of niacin and lower levels of pantothenic acid (Shahidi, 1990).
The recuperation of proteins present in rapeseed meal, making
it available for use in human food, is important due to its balanced
amino acids composition when compared with the amino acids required in the diet, which translates into a high biological value. It
possesses acceptable functional properties for its use in the formulation of several foods and a similar biological value to the one
found in casein and superior to other vegetable sources such as
soybean and wheat (Ghodsvali et al., 2005).
Carbohydrates are in lower proportion in rapeseeds and are
mostly amyloid compounds, cellulose, pectins and arabinogalac-

Table 1
Rapeseed meal versus soybean meal composition (d.b.) (Pirie and Swaninathan,
1975).

Rapeseed meal
B. napus
B. campestris
Soybean meal

Proteins(N6.25)

Oil

Nitrogen-free
extract

Fiber

Ash

43.1
41.1
50.4

2.3
2.0
0.5

36.9
37.8
35.4

10.7
11.6
6.9

7.0
7.5
6.8

Table 2
Amino acid composition of proteins in rapeseed/canola and soybean (g per 100 g of
protein) (Shahidi, 1990).
Amino acid

Rapeseed and canola

Soybean

Alanine
Arginine
Aspartic acid
Cysteine
Glutamic acid
Glycine
Histidine
Isoleucine
Leucine
Lysine
Methionine
Phenylalanine
Proline
Serine
Threonine
Tryptophan
Tyrosine
Valine

4.3
5.8
7.0
1.7
17.5
4.9
2.7
4.0
7.0
5.8
1.9
3.8
6.0
4.6
4.5
1.3
3.1
5.0

4.3
7.2
11.7
1.6
18.7
4.2
2.6
4.5
7.8
6.4
1.3
5.0
5.1
5.1
4.0
1.3
3.2
4.8

tan. Cellulose is found mostly in hull, together with hemicellulose

and lignin (Shahidi, 1990).
From the carbohydrates of low molecular weight, monosaccharides and oligosaccharides, about 48% are ethanol soluble (Shahidi,
1990). The main component is sucrose, with smaller amounts of
rafnose and stachyose. Some digestive problems which lead to
atulence in both humans and animals are linked to the oligosaccharide derivatives of sucrose (for example stachyose and rafnose). These problems result from the fact that the human
digestive system does not have the enzymes responsible for the
hydrolysis of these sugars (a-galactosidade), and therefore they
are anaerobically fermented in the large intestine with gas release
(Shahidi, 1990).
The mineral composition of rapeseed meal is dependent on the
mineral composition of the soil where it is planted, and, therefore,
highly variable. The minerals present in rapeseed include phosphorus and potassium (Table 3). The majority of the phosphorus is
found in the form of phytic acid, which in turn, can interfere in
the absorption of calcium, zinc and iron (Kirk-Othmer, 1997).
Rapeseed also contains high levels of glucosinolates, compounds responsible for the signicant reduction of the nutritional
level of the rapeseeds used in animal feed, erucic acid, which
depending on the efciency of the oil extraction process can also
be present in the defatted meal, and phytates, salts derived from
the phytic acid. These compounds have toxic and antinutritional
characteristics, so their removal is essential to enable their use in
human food. On this matter, Ghodsvali et al. (2005) studied the
application of membrane technology, ultraltration and dialtration, to prepare canola protein isolates with low levels of glucosinolates and phytates.

Table 3
Mineral and vitamins composition of rapeseed meal (Shahidi, 1990).
Mineral

Composition
(mg/kg)

Vitamin

Composition
(mg/kg)

Calcium
Copper
Iron
Magnesium
Manganese
Phosphorus
Potassium
Selenium
Zinc
Sulfur

6300
5.7
141
5100
49.2
10,200
12,100
1.1
68.2
8500

Choline
Folic acid
Niacin
Pantothenic acid
Riboavin
Thiamin
Biotin

6700
2.3
160
9.5
3.7
5.2
1.1

342

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

Fig. 4. General structure of glucosinolates; R = aliphatic, aromatic or heterocyclic

group.

Fig. 5. Chemical structure of erucic acid.

The general structure of the glucosinolates (Fig. 4) is characterized by a group of b-D-thioglucose, a portion of sulphonated oxime
and a lateral chain derived from either the methionine, the phenylalanine or the tryptophan (Graser et al., 2000). There are more
than 120 glucosinolates in nature, with different lateral, aliphatic,
aromatic or indole chains, which results in a high variability in biological activity and its polarity (Chen and Andreasson, 2001).
Glucosinolates are hydrolysated by the thioglucosidases, called
myrosinases, producing a vast array of degradation products, nitriles, isothicyonates, oxazolidine-2-thiones and thiocyanates with
diverse biological activity (Halkier and Gershenzon, 2006; Kliebenstein et al., 2001). These compounds limit not just the consumption
of food due to their reduced palatability, but they also interfere in
the functional activity of the thyroid, damaging vital organs and
interfering with metabolic processes (Burel et al., 2001; Tripathi
and Mishra, 2006).
Erucic acid (Fig 5) is an omega-9 monounsaturated fatty acid
(22:1 n-9) abundant in rapeseed, erisim and mustard seeds. Erucic
acid is also known as cis-13-docosenoic acid, and the isomer trans
is known as brassidic acid. This compound presents a level of toxicity for humans when taken in high doses, as it affects the growth,
functioning and morphology of several organs, such as the heart
and skeletal muscles. Due to these adverse effects, the Alimentarius Codex Committee considers rapeseed oil, with an erucic acid
composition equal or superior to 5% of the total fraction of fatty
acids, unsuitable for use in food (Vianni and Braz-Filho, 1996).

The phytates, also known as phytic acid (Fig. 6) are chemical

compounds used by plants to store phosphorous. Considered antinutritional compounds due to the fact that, in addition to interacting with basic residues of the proteins, they reduce the
bioavailability in the organism of bivalent minerals, such as calcium, iron, magnesium, manganese, cuprum and zinc, making
them biologically unavailable for absorption (Liener, 1994). When
phytates interfere with the proteins, the solubility of the latter is
reduced, affecting their functional properties, besides reducing
the nutritional value of the product. The enzymatic digestion of
the protein is reduced because phytates inhibit the action of the
digestive enzymes, pepsin, trypsin and a-amylase (Liener, 1994).
As exposed in the previous sections, there is an enormous portfolio of high value compounds that can be extracted from oil seed
plants. However, in order to incorporate these compounds in food
formulations, suitable methods of extraction, isolation and purication need to be developed.

4. Valorisation of oilseed meals

After the oil extraction from the oilseeds, the resultant byproduct is a defatted meal. The oil is used in human food and in a biodiesel production. Defatted meal, formed essentially by proteins
and carbohydrates, has a high nutritional level. Therefore, it is
important to proceed to its valorisation, ensuring a better global
sustainability of biodiesel industry from oilseeds, particularly from
the rapeseed. The extraction, concentration and isolation of the
protein and carbohydrates fractions from defatted meal is essential
to the valorisation of the products.
Protein fraction of rapeseed defatted meal can be valorised in
protein concentrates and protein isolates using techniques capable
of reducing or eliminating the compounds responsible for the loss
of its nutritional value, such as the glucosinolates, phytates and
erucic acid (Aherne et al., 1976; Badawy et al., 1994; Brand et al.,
2007; Fenwick et al., 1983; Liener, 1994; Tripathi and Mishra,
2006). This way, it will be possible to use it as a supplement in food
for human or animal consumption and also to attain, by protein
hydrolysis, of increased valued products such as the functional
peptides and the essential amino acids.
Carbohydrates fraction can be hydrolyzed, by a chemical or an
enzymatic way in fermentable sugars which, with a further action
of several types of microorganisms, may lead to a large number of
added-value products. The bioethanol production through the
alcoholic fermentation of those sugars is an example.

Fig. 6. Chemical structure of phytic acid and its conformations (a) one axial position and ve equatorials positions and (b) ve axial positions and one equatorial position.
Source: Adapted from Quirrenbach et al. (2009).

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

4.1. Alternative technologies for oilseed meals valorisation

The denomination of protein concentrates or isolates is assigned to the obtained extracts, depending on their protein concentration on a dry base. They are protein concentrates or isolates
when their concentration level reaches 65% or 90% respectively
(Kirk-Othmer, 1997; Moure et al., 2006).
Protein concentrates are usually obtained through the application of techniques which involve, at a rst stage, the extraction
of the non-protein material in the meal, essentially formed of carbohydrates, thus increasing the concentration of protein fraction in
the resulting product. The non-protein material extraction techniques encompass the use of alcoholic aqueous solutions (60
80%) and acid water at a pH 4.5 (to minimize the loss of protein
fraction solubilized) (Kirk-Othmer, 1997; Lewis and Gradison,
1996). Protein concentrates obtained in this way retain most of
the ber content in the seed, which is favorable in the case of
the soybean, due to its application in cereal mixtures and meatsubstitute products in human food. In the rapeseeds case, the presence of bers contributes to its lower digestibility, as it presents
higher levels of ber content than those of the soybean.
Protein isolates are obtained through the solubilization of the
protein, as mentioned above, in saline solutions and/or in an environment where the pH is far from the isoelectric point, promoting,
the concentration through ultraltration and dialtration membranes or the precipitation in an environment where the pH is
close to the isoelectric point of the solubilized proteins.
Proteins are quite complex structures, with distinct forms and
shapes, presenting hydrophobic and hydrophilic areas, which
determine their behavior in solution, and consequently its success
or failure in their separation. Therefore, the solubility of a protein
depends on the properties of the groups that are at a surface level
in the molecule, from the kind of solvent, the temperature and the
pH (Lewis and Gradison, 1996). The isoelectric point of the proteins
is determined by its amino acid constitution. If the protein has a
high level of basic amino acids (for example, lysine and arginine)
its isoelectric point would be higher than 7.0. If its amino acid constitution is predominantly composed of amino acids with acid residues (for example aspartic acid or glutamic acid) its isoelectric
point would be lower than 7.0.
The extraction and isolation of proteins involves solubilization
and precipitation techniques based on properties such as solubility
and the isoelectric point of the protein molecules (Lewis and Gradison, 1996).
The solubilization of the proteins contained in several vegetable
materials, namely in oilseeds (such as soybean, sunower, rapeseed), in their meals, or in watermelon and pumpkin seeds, has
been studied. The extraction has been carried out in water or in
NaC1 solutions with widely varying pH levels (controlled by the
addition of NaOH or HC1 solutions), and by changing of the solid/solution ratio, temperatures and extraction times. Several patents have been registered in recent decades. Cameron and Myers
(1983) patented a method of rapeseed extraction in water of 10
15% (w/v) of protein material with constant stirring during 10
60 min at a temperature of 1535 C. Intended of making only
water, Murray and Mills (1999) extracted the oilseed meals with
a NaC1 0.5 M solution, at a pH of 5.36.2 and a temperature between 5 and 35 C, with constant stirring for 1060 min. Within
the same range of pH, temperature and extraction times, Milanova
et al. (2003), registered a new patent for the extraction of proteins
from the canola meal in a saline solution with a concentration of
0.150.6 M and a solid/solution ratio of 13% (w/v). Within the
aforementioned conditions, Quanhong and Caili (2005), studied
the extraction of proteins in pumpkin seeds, and concluded that
the quantity of solubilized protein reached a maximum point to
a solid/solution ratio of 3.31% (w/v), in a NaC1 solution with

343

0.73 M, during 18.1 min of extraction time. As for the extraction

of sunower proteins, Pickardt et al. (2009), concluded that it
was in NaC1 solutions up to 2.8 M, with a pH between 5.6 and
7.4, at room temperature and a solid/solution ratio of 5% (w/v), that
the best compromise between a higher efciency in the extraction
and a minor interaction of the extracted proteins with the phenolic
acids was reached, thus producing better quality isolates in terms
of color.
Other authors developed their investigations, mapping the effect of NaC1 concentration with pH in the efciency of protein
extraction. Moure et al. (2001), obtained extraction yields of between 55% and 60% in a NaC1 0.5 M solution and pH 11, considerably higher than those obtained when the pH of the solution was
4.5, in the protein extraction of Rosa rubiginosa seeds, using a solid/solution ratio of 5% (w/v). Yields of protein higher than 87% in
cowpeas are also obtained when subjected to a NaC1 0.15 M solution with pH of 9.9, in a solid/solution ratio of 10% (w/v), during 2 h
at 35 C, with constant stirring (Mune et al., 2008).
However, there is a multitude of references related to the solubilization of vegetable proteins in alkaline solutions, usually of
NaOH, with concentrations up to 5%, at various temperatures,
and with extraction times ranging between 30 and 60 min.
Bora and Ribeiro (2004) solubilized 83% of the existing proteins
in defatted macadamia in a pH 12 solution, while with a pH of 2
failed to reach 53%, keeping the solid/solution ratio of 20% (w/v),
stirring for 60 min at room temperature.
The conditions achieved by Wani et al. (2008) which resulted in
the most efcient extraction of proteins from watermelon seeds,
consisted of a NaOH solution of 1.2%, a solid/solution ratio of
1.4% (w/v), and a period of 15 min at a temperature of 50 C.
The extraction of proteins from the seeds of red peppers, however, is most efcient with a solution of pH 8.8, used in a solid/
solution ratio of 4.8% (w/v), at a temperature of 31 C for 20 min
(Durmus and Evranuz, 2010). Table 4 resume the extracting conditions more applied in the productions of proteins isolates from
vegetable sources.
Pedroche et al. (2004) and Ghodsvali et al. (2005) follow a similar method to extract proteins from rapeseed and canola, using
solutions with a pH between 9.5 and 12 and a solid/solution ratio
of 510% (w/v). These conditions are preferable when the objective
is to reduce the phytate content of the protein isolates, because
with pH 12 their solubility is minimal. The proteinphytate interaction depends on the pH and the contents in terms of divalent
ions. In an acidic pH, proteins are positively charged, while the
phytic acid has a negative charge, resulting in the formation of
phytate-protein complexes. The presence of calcium ions in excess
favors the dissociation of this complex. With an alkaline pH, both
the protein and the phytic acid are negatively charged, therefore
the proteinphytate interaction only occurs in the presence of multivalent ions, forming proteinmineralphytate complexes. To
avoid the formation of these complexes, it is necessary to remove
these ions, using chelators, such as the EDTA (Diosaday et al.,
1989; Serraino and Thompson, 1984).
With regards to glucosinolates, the deactivation of the myrosinases with resource to steam prevents its decomposition without
altering the bioactive properties in the rapeseed protein concentrates and improves the sensorial attributes of the sausage industry
where it is applied in replacement of the casein (Yoshie-Stark et al.,
2006).
Once the protein material is solubilized it is necessary to recover it. Such a task can be carried out by promoting the precipitation of the proteins or their concentration, and then subjecting
them to centrifugation and drying (conventional) or to lyophilization to obtain protein powder.
The precipitation of the proteins can be achieved with water,
where the protein solution is diluted, accompanied by a reduction

344

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

Table 4
Extracting conditions to prepare proteins isolates from vegetable sources.
Vegetable material

Extraction solution

pH

Temperature (C)

Time (min)

Solid/solution ratio (%w/v)

References

Rapeseed
Oil seed meal
Rosa rubiginosa seeds
Canola meal
Pumpkin seeds
Sunower meal
Cowpeas
Macadamia
Rapeseed meal
Canola meals
Watermelon seeds
Red pepper seeds
Rapeseed meal
Canola meals

Water
NaCl 0.5 M
NaCl 0.5 M
NaCl 0.150.6 M
NaCl 48%(0.681,37 M)
NaCl 13 M
NaCl 00.5 M
HCl, NaCl 0.5 M and NaOH
NaOH 0.2%
Water, NaOH
NaOH 0.31.5%
Water, NaOH
NaOH 0.2%
Water, NaOH

5.36.2
4.511
5.36.2

3.27.4
7.011.0
2.012.0
12
9.512.0

1535
535
3060
535
35
1545
35
Room temp.

30
4060
3050

30

1060
1060
90
10
1030
60
120
60
60
15
525
2060
60
15

1015
37
5
13
310
5
10
20
10
5.6
1.43.3
3.310
10
5.6

Cameron and Myers (1983)

Murray and Mills (1999)
Moure et al. (2001)
Milanova (2003)
Quanhong and Caili (2005)
Pickardt et al. (2009)
Mune et al. (2008)
Bora and Ribeiro (2004)
Pedroche et al. (2004)
Ghodsvali et al. (2005)
Wani et al. (2008)
Durmus and Evranuz (2010)
Pedroche et al. (2004)
Ghodsvali et al. (2005)

7.09.0
12
9.512.0

in the temperature (515 C), aiding formation of the protein

aggregates (Cameron and Myers, 1983). However, this dilution increases water consumption and the need to address this leads to
difculties in the solid/solution separation and therefore in the
recuperation of the material precipitated. An alternative is to use
acid solutions with a pH adjusted by HC1 or H2SO4 concentrated
solutions, up to a value below the pH in the isoelectric point of
the proteins in the solution. This is the most common processes,
namely when the extraction is promoted in alkaline solutions.
In terms of rapeseed proteins, Pedroche et al. (2004), advocated
an isoelectric precipitation at a pH of 4.3, while Ghodsvali et al.
(2005) obtained a pH between 3.5 and 7.5. Bora and Ribeiro
(2004) used a pH of 5 to precipitate the protein material solubilized from macadamia.
Protein concentration, after its solubilization, is performed
essentially by the use of membrane technology, namely ultraltration and dialtration. Murray and Mills (1999), Milanova (2003)
and Moure et al. (2001) are some of the investigators who used
these technologies, usually associated with protein extraction
through saline solutions.
Murray and Mills (1999) also applied a cooling stage as a removal operation of the lipid fraction which is solubilized at the
time of the extraction, in order to increase the protein concentration in the solution between solubilization and precipitation. The
fat removal process also involves decantation, ltration or centrifugal operations after the cooling of the solution.
Membrane technology has been successfully used in the recovery of solubilized proteins from several vegetable sources, including rapeseed meal (Barker et al., 2002; Ghodsvali et al., 2005).
The use of ultraltration and dialtration enables protein concentrates or isolates with minor concentrations of soluble solids to be
obtained, due to the fact that as they are molecules of lower molecular mass, they are not retained in the pores of the membranes.
This feature is important as it may lead to a signicant reduction
in the compounds responsible for the economical and nutritional
devaluation present in the rapeseed meal, such as glucosinolates,
phytates and polyphenols.
Globulins are the main protein component in the rapeseed
meal. They are soluble proteins in saline solutions and they have
pH 7.0 on the isoelectric point. The techniques of solubilization
go through the extraction in an alkaline aqueous medium, of a
pH superior to 7.0, conjugated or not with an increase of the salt
concentration in the solution. The albumins, minority fraction of
protein in the rapeseed meal, are characterized by their low molecular weight. They are water soluble and they present a pH 12.0 on
the isoelectric point. Thus, the studied strategies for the solubilization of the protein material in defatted rapeseed meal, go through
the usage of alkaline solutions with pH between 7 and 12, eventu-

ally with the increase of ionic force promoted by the addition of

NaC1 in certain concentrations.
Other factors to consider in the optimization of the extraction
process are temperature, operation time, solid/solution ratio and
stirring. Temperature rise favors the protein solubilization, however if too high, it may cause its thermal denaturation and precipitation. For that reason, according to the consulted references, the
preferential use of room temperature or slightly higher prevails.
Temperatures above 60 C are not commonly studied. With regards
to the extraction time, it varies unanimously between 10 and
60 min, with constant stirring and a solid/solution ratio between
5% and 15% (w/v).
The precipitation of proteins can be achieved by resorting to
several techniques, such as temperature decrease, pH adjustment
up to its isoelectric point, addition of apolar solvents to reduce
the attraction between groups with polar surface and water, alteration of the ionic force (by diminution of the salt concentration) or
by thermal denaturation (Lewis and Gradison, 1996). All these
techniques aim to decrease the solubility of the protein molecules
in the solution, but none of them enable proteins in a solid state to
be obtained. Therefore, it will always be necessary to conduct solid/liquid separation operations, such as centrifugation and ltration, promoting the drying operation to obtain the nal product.

4.2. Production technology of protein isolates and their functional

properties
Some denitions of the functional properties have been described. However, none of them describes entirely the functional
properties of the proteins, due to their great diversity. The inuence of the molecular properties on the structure and function of
the proteins, their nutritional value (amino acid composition,
inhibitors presence, nutritional compounds and protein quality)
and their organoleptic and biological properties, such as therapeutic and antioxidant functions (Moure et al., 2006) should all be
considered.
The purication of vegetable proteins involves a series of physicalchemical treatments, which affect the nutritional value and
the functional properties of the nal product in both food and nonfood products. When added to food, proteins give desirable functional properties, such as solubility, viscosity, foam formation,
emulsication and water and oil retention capability. Proteins also
perform an important role in the nutritional, sensorial, physical
chemical and organoleptic (color, texture, avor and so on) properties of the food, while the adherence properties and lm formation
have great potential in applications on an industrial scale (Moure
et al., 2006).

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

As described before, the production of protein isolates implies

the solubilization of the protein in saline aqueous solutions, of neutral or alkaline pH and posterior recuperation through precipitation
techniques (isoelectric point), concentration and separation (ultraltration) before drying. Factors like the pH, the presence or absence
of salts and respective concentration, ionic force of the medium, and
electrostatic repulsions inuence both the protein extraction efciency and their functional properties. High temperature and
lengthy treatments reduce the nutritional quality of the isolates.
Processing in an alkaline medium triggers a set of undesirable chemical reactions like amino acid racemization, lysinoalanine formation
through reaction of the lysine with the desidroalanine produced by
the degradation of the cysteine and serine, and a reduction in digestibility and a loss of essential amino acids (Moure et al., 2006).
Products obtained by ultraltration present better functional
properties than those obtained by isoelectric precipitation, particularly in terms of emulsifying properties. Besides, the glucosinolates
and phytates are efciently removed from the isolates without
altering functional properties. The drying technology also inuences the functional properties of the proteins (Moure et al., 2006).
The majority of the native proteins do not present the desirable
functional properties of the food industry, and therefore they are
subjected to modications capable of increasing their nutritional
value and their functional properties, especially in terms of solubility (Panyam and Kilara, 1996). Modications that aim to preclude
degradation reactions, removal of toxic or inhibitors compounds,
and also the incorporation of nutrients or additives, present considerable advantages. Therefore, the protein isolates can be subjected to chemical, enzyme or thermal treatments in order to
strengthen their functional properties. Chemical modications
can be promoted through alkylation, oxidation, acylation, esterication and amides formation. The enzymatic changes aim to break
the peptide chains, producing peptides with size, charge and surface properties desirable, in order to increase the solubility and removal of antinutritional compounds (Moure et al., 2006).
Rapeseed protein isolates partially hydrolyzed with alcalase,
produced by isoelectric precipitation, present better functional
properties, (water and oil absorption, whipping, foam formation
and stability, emulsifying capacity and emulsion stability), than
the original isolates, although such properties decrease as the degree of hydrolysis increases (Vioque et al., 2000).
Pedroche et al. (2004) hydrolyzed rapeseed protein isolates
with alcalase and obtained peptides with high inhibitory activity
of the angiotensin converting enzyme (ACE), whose activity is related to high blood pressure and other cardiac diseases. Also Marczak et al. (2003) isolated four angiotensin converting enzyme (ACE)
inhibitory peptides from rapeseed meal, using pepsin, trypsin,
chymotrypsin, pancreatin, subtilisin and thermolysin enzymes.
Other physical treatment, such as use of high pressures, unfolding and the exposure of the hydrophobic sites increases the functional properties of proteins. The addition of sugar improves the
whipping capacity of oilseed proteins, and also their combination
with polysaccharides promotes the production of biopolymers
with active surfaces which promote a higher stability under heat
or pressure, in food formulations like foams, emulsions or dispersions. The coprecipitation of proteins from different vegetable
sources blends or with whey proteins shows higher nutritional
value and better functional properties when compared with
individual isolates per se (Moure et al., 2006).

5. Conclusions and outlook

The total use of the oilseeds meals implies the valorisation of
their protein fraction, creating value-added products destined for
the food, cosmetic and pharmaceutical industries.

345

The valorisation of these proteins involves extraction and isolation techniques which encompass the solubilization in saline solutions with NaC1 concentrations up to 5 M and/or alkaline solutions
of pH up to 12.5. The isolation presupposes precipitation techniques of the protein fraction based on the isoelectric point of
the proteins in solution, encouraged by the decrease of the pH to
3.5, with the addition of concentrated solutions of mineral acids
like hydrochloric acid and sulfuric acid. Those which have been
precipitated, after being centrifuged and/or ltrated, are dried, thus
producing protein isolates. Another way of achieving the isolates is
through the use of membrane technology, namely ultraltration
and dialtration as a way of concentrating the solubilized proteins
by ionic force changes (saline solutions) or in alkaline solutions.
This technique is preferable when the solubilization is provided
in saline solutions, since it allows the production of isolates with
a lower salt content. It is also of particular interest in the production of rapeseed protein isolates, since it allows a reduction in low
molecular weight compounds associated to their digestive, nutritional and sensorial depreciation, such as glucosinolates, phytates
and phenolic compounds, thereby granting an extra asset to these
products.
The functional properties of the protein isolates can be increased through chemical, physical and enzymatic modications,
enhancing the solubility, emulsion and foam capacity and stability,
increasing the nutritional value, producing bioactive peptides and
isolate amino acids amongst others, and thus providing compounds of increased value.
It is possible, through appropriate techniques and methodologies to enhance the oilseeds meal, specially rapeseed, which as a
byproduct of the oil extraction industry is characterized by a low
commercial value when compared with, for example, soybean.

References
Aherne, F., Bowland, J., Christian, R., Hardin, R., 1976. Performance of myocardial
and blood seral changes in pigs fed diets containing high or low erucic acid
rapeseed oils. Canadian Journal of Animal Science 56, 275284.
Badawy, I., Atta, B., Ahmed, W., 1994. Biochemical and toxicological studies on the
effect of high and low erucic acid rapeseed oil on rats. Nahrung 38, 402411.
Barker, L.D., Martens, R.W., Murray, E.D., 2002. Production of Oilseed Protein Isolate.
Int. Patent WO 02089597.
Belitz, H.D., Grosch, W., 1999. Food Chemistry, second ed. Springer-Verlag, Berlin.
Bora, P.S., Ribeiro, D., 2004. Inuence of pH on the extraction yield and functional
properties of macadamia (Macadamia integrofolia) protein isolates. Food Science
Technology International 10 (4), 263267.
Brand, T.S., Smith, N., Hoffman, L.C., 2007. Anti-nutritional factors in canola
produced in the Western and Southern Cape areas of South Africa. South
Africa Journal of Animal Science 37, 4550.
Burel, C., Boujard, T., Kaushik, S.J., Boeuf, G., Mol, K.A., Van der Geyten, S., Darras,
V.M., Khn, E.R., Pradet-Balade, B., Qurat, B., Quinsac, A., Krouti, M., Ribaillier,
D., 2001. Effects of rapeseed meal-glucosinolates on thyroid metabolism and
feed utilization in rainbow trout. General and Comparative Endocrinology 124
(3), 343358.
Cameron, J.J., Myers, C.D., 1983. Rapeseed Protein Isolate. Patent Application
Publication US 4 418 013.
Canola Council of Canada, 2008. Canola Oil: The Truth! <http://www.canolacouncil.
org> (July 2010).
Chen, S., Andreasson, E., 2001. Update on glucosinolate metabolism and transport.
Plant Physiology and Biochemistry 39, 743758.
Coultate, T.P., 1984. Alimentos Qumica de sus componentes. Editorial Acribia SA,
Zaragaza.
Damodaran, S., 2000. Aminocidos, pctidos y protenas. In: Fennema, O.R. (Dir),
Qumica de los Alimentos. Editorial Acribia SA, Zaragoza, pp. 381511.
Darryl, L., 2010. Glossary of Genetic Terms. National Human Genome Research
Institute.
<http://www.genome.gov/Glossary/resources/protein.pdf>
(July
2010).
Diosaday, L.L., Rubin, L.J., Tzeng, Y.M., 1989. Production of Rapeseed Protein
Materials. Patent Application Publication US 4 889 921.
Duran, M.L., et al., 1980. Enciclopdia Luso-Brasileira de Cultura, 14th ed., vols. 14
and 17. Editorial Verbo, Lisboa.
Durmus, E.F., Evranuz, O., 2010. Response surface methodology for protein
extraction optimization of red pepper seed (Capsicum frutescens). LWT Food
Science and Technology 43, 226231.
FAO (2009). Perspectivas Alimentarias anlisis de los mercados mundiales.
<http://www.fao.org/docrep/012/ak341s/ak341s00.pdf>, (July 2010).

346

I.M. Rodrigues et al. / Journal of Food Engineering 109 (2012) 337346

FAO (2010). Food and Agricultural commodities production. <http://faostat.fao.org/

site/339/default.aspx> (July 2010).
Fenwick, G.R., Heaney, R.K., Mullin, W.J., 1983. Glucosinolates and their breakdown
products in food and food plants. Critical Reviews in Food Science and Nutrition
18, 123201.
Ghodsvali, A., Khodaparast, M.H.H., Vosoughi, M., Diosady, L.L., 2005. Preparation of
canola protein materials using membrane technology and evaluation of meal
functional properties. Food Research International 38, 223231.
Graser, G., Schneider, B., Oldham, N.J., Gershenzon, J., 2000. The methionine chain
elongation pathway in the biosynthesis of glucosinolates in Eruca sativa
(Brassicaceae). Archives of Biochemistry and Biophysics 378, 411419.
Halkier, B.A., Gershenzon, J., 2006. Biology and biochemistry of glucosinolates.
Annual Review of Plant Biology 57, 303333.
Huret, J. L. 2008. Amino Acids (reminder). Atlas of Genetics and Cytogenetics in
Oncology
and
Haematology.
<http://AtlasGeneticsOncology.org/Educ/
AminoAcidsEngID30066ES.html> (July 2010).
Kirk-Othmer, 1997. Encylclopedia of Chemical Technology, 4th ed., vol. 22. John
Wiley & Sons, New York.
Kliebenstein, D.J., Kroymann, J., Brown, P., Figuth, A., Pedersen, D., Gershenzon, J.,
Mitchell, O.T., 2001. Genetic control of natural variation in Arabidopsis
glucosinolate accumulation. Plant Physiology 126, 811825.
Krause, J.P., Schwenke, K.D., 2001. Behaviour of a protein isolate from rapeseed
(Brassica napus) and its main protein components globulin and albumin at
air/solution and solid interfaces, and in emulsions. Colloids and Surfaces B:
Biointerfaces 21, 2936.
Lewis, M.J., Gradison, A.S., 1996. Separation Process in the Food and
Biotechnology Industries: Principles and Applications. Woodhead Publishing
Ltd., Cambridge.
Liener, I.E., 1994. Implications of antinutritional components in soybean foods.
Critical Reviews in Food Science and Nutrition 34 (1), 3134.
Marczak, E.D., Usui, H., Fujita, H., Yang, Y., Yokoo, M., Lipkowski, A.W., Yoshikawa,
M., 2003. New antihypertensive peptides isolated from rapeseed. Peptides 24,
791798.
McDonald, B.E., 2004. Food uses and nutritional properties. In: Gunstone, F.D. (Ed.),
Rapeseed and Canola Oil. Blackwell Publishing, Oxford, pp. 131153.
Milanova, R. Murray, E.D., Westdal, P.S., 2003. Protein Extraction from Canola Oil
Seed Meal. Patent 6992173. <http://www.patentstorm.us/patents/6992173/
description.html> (July 2010).
Moure, A., Sineiro, J., Dominguez, H., 2001. Extraction and functionality of
membrane-concentrated protein from defatted Rosa rubiginosa seeds. Food
Chemistry 74, 327339.
Moure, A., Sineiro, J., Domnguez, H., Paraj, J.C., 2006. Functionality of oilseed
protein products: a review. Food Research International 39, 945963.
Mune, M.A.M., Minka, S.R., Mbome, I.L., 2008. Response surface methodology for
optimisation of protein concentrate preparation from cowpea [Vigna
unguiculata (L.) Walp]. Food Chemistry 110, 735741.
Murray, E.D., Mills, E., 1999. Oilseed protein extraction. Patent Application
Publication US 6 005 076.
Oplinger, E.S., Hardman, L., Gritton, E.T., Doll, J.D., Kelling, K.A., 1989. Canola
(Rapeseed), University of Minnesota, St. Paul, MN 55108. <http://
www.hort.purdue.edu/newcrop/afcm/canola.html>, July 2010.

Panyam, D., Kilara, A., 1996. Enhancing the functionality of food proteins by
enzymic modication. Trends in Food Science and Technology 7 (4), 120125.
Pedroche, J., Yust, M., Megas, C., Lqari, H., Alaiz, M., Girn-Calle, J., Milln, F., Vioque,
J., 2004. Utilisation of rapeseed protein isolates for production of peptides with
angiotensin I-converting enzyme (ACE)-inhibitory activity. Grasas y Aceites 55
(4), 354358.
Pickardt, C., Neidhart, S., Griesbach, C., Dube, M., Knauf, U., Kammerer, D.R., Carle, R.,
2009. Optimisation of mild-acidic protein extraction from defatted sunower
(Helianthus annuus L.) meal. Food Hydrocolloids 23, 19661973.
Pirie, N.W., Swaninathan, M.S., 1975. International Biological Programme: Food
Protein Sources, vol. 4. CUP Archive, Cambridge.
Quanhong, L., Caili, F., 2005. Application of response surface methodology for
extraction optimization of germinant pumpkin seeds protein. Food Chemistry
92, 701706.
Quirrenbach, H.R., Kanumfre, F., Rosso, N.D., Filho, M.A.C., 2009. Comportamento do
cido ftico na presena de Fe(II) e Fe(III). Cincia e Tecnologia de Alimentos 29
(1), 2432.
Ratnayake, W.M.N., Daun, J.K., 2004. Chemical composition of canola and rapeseed
oils. In: Gunstone, F.D. (Ed.), Rapeseed and Canola Oil. Blackwell Publishing,
Oxford, pp. 3773.
Salunkhe, D.K., Adsule, R.N., Chavan, J.K., Kadam, S.S., 1992. World Oilseeds:
Chemistry, Technology and Utilization. Springer, New York.
Samouco, R., 1998. Dicionrio de Agronomia. Pltano Edies Tcnicas, Lisboa.
Schwenke, K.D., Dahme, A., Wolter, T., 1998. Heat-induced gelation of rapeseed
proteins: effect of protein interaction and acetylation. Journal of American Oil
Chemists Society 75 (1), 8387.
Serraino, M.R., Thompson, L.U., 1984. Removal of phytic acid and protein-phytic
acid interactions in rapeseed. Journal of Agriculture and Food Chemistry 32, 38
40.
Shahidi, F., 1990. Canola and Rapeseed: Production, Chemistry, Nutrition, and
Processing Technology. Van Nostrand Reinhold, New York.
Temple-Heald, C., 2004. High erucic oil: its production and uses. In: Gunstone, F.D.
(Ed.), Rapeseed and Canola Oil. Blackwell Publishing, Oxford, pp. 111130.
Tripathi, M.K., Mishra, A.S., 2006. Glucosinolates in animal nutrition: a review.
Animal Feed Science and Technology 132, 127.
Vianni, R., Braz-Filho, R., 1996. cidos graxos naturais: importncia e ocorrncia nos
alimentos. Qumica Nova 19 (4), 400407.
Vioque, J., Snchez-Vioque, R., Clemente, A., Pedroche, J., Milln, F., 2000. Partially
hydrolyzed rapeseed protein isolates with improved functional properties.
Journal of the American Oil Chemists Society 77 (4), 447450.
Wani, A.A., Kaur, D., Ahmed, I., Sogi, D.S., 2008. Extraction optimization of
watermelon seed protein using response surface methodology. LWT Food
Science and Technology 41, 15141520.
Yoshie-Stark, Y., Wada, Y., Schott, M., Wasche, A., 2006. Functional and bioactive
properties of rapeseed protein concentrates and sensory analysis of food
application with rapeseed protein concentrates. LWT Food Science and
Technology 39, 503512.
Yfera, E.P., 1998. Qumica de los Alimentos. Editorial Sintesis, Madrid.
Zivanovic, I., Vastag, Z., Popovic, S., Popovic, L., Pericin, D., 2010. Hydrolysis of hullless pumpkin oil cake protein isolate by pepsin. International Journal of
Biological and Life Sciences 6 (1), 3034.