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Grupo 2 Ghanghas 2020 FRI Rice
Grupo 2 Ghanghas 2020 FRI Rice
To cite this article: Neeraj Ghanghas, Mukilan M. T., Shikha Sharma & Pramod K. Prabhakar
(2020): Classification, Composition, Extraction, Functional Modification and Application of Rice
(Oryza�sativa) Seed Protein: A Comprehensive Review, Food Reviews International, DOI:
10.1080/87559129.2020.1733596
Article views: 48
CONTACT Pramod K. Prabhakar pramodkp@niftem.ac.in Department of Food Science and Technology, National
Institute of Food Technology Entrepreneurship and Management, Kundli, India
© 2020 Taylor & Francis
2 N. GHANGHAS ET AL.
Introduction
Rice (Oryza sativa) is one of the most important cereal crops cultivated across more than 100
countries across the globe. Rice is considered as the staple food, it feeds more than half of the
world population. India is the second-largest producer of rice in the world after china with an
output of 165.02-million-ton paddy in the year 2017 and also is the second largest in term of
total consumption of milled rice with consumption of 97.35 million ton.[1] It is mainly
consumed in boiled/cooked form. The two major sub-classes of Oryza sativa are japonica
which is sticky and short grain while Indica is non-sticky and long grain. Japonica is cultivated
in the areas of South-East Asian upland, temperate East Asia and South Asian high elevations,
while Indica is cultivated mainly in low-land areas of tropical Asia.[2]
Rice is nutritious and shows some unique functional properties which include hypo-
allergenicity and flavour-carrying capability.[3] White rice is most widely consumed while
there are also other cultivars such as red rice, black rice and brown rice which contain colour
pigments. The name of these varieties is due to the colour of their kernel (red, purple or black),
the deposition of the anthocyanins in various layers (aleurone, pericarp and seed coat) is
responsible for the different colour of the kernel.[4] According to Paine et al.,[5] golden rice is
the genetically modified rice variety which produces carotenoids in the endosperm, conferring
yellow colour to the grain. Suzuki et al.[6] reported that the black rice has higher protein,
vitamins and minerals which give it nutritional advantages over other common rice. Rice is
considered as the queen among the cereals due to its nutritional quality and higher
digestibility.[7] Rice seed protein is classified into four fractions, i.e. albumin, globulin, prola-
min and glutelin based on solubility in various solvents.[8] The seed storage proteins in rice
grain are present as protein bodies (PB-I & PB-II), these two protein bodies possess different
structure and composition.[9] Rice proteins have a sound amino acid profile comprising
essential amino acids such as threonine, leucine and phenylalanine along with sulphur-rich
amino acids like methionine and cysteine which are crucial for the human body. Rice proteins
contain high amount of methionine and phenylalanine as compared to casein. Amino acid
profile of foods is not only important in nutritional point of view but is also related to the
physical and chemical properties of the food product.[10–12] Rice and other sources of plant
proteins are abundantly available and are inexpensive with exceptional nutritive and func-
tional properties. The functional properties (solubility, foaming properties, oil and water
absorption, emulsification properties, etc.) of rice proteins are influenced by various para-
meters such as pH, salt & sugar concentrations; these parameters can be varied to modify and
exploit the functional properties of rice proteins for applications in different food
formulations.[13,14] Protein stability and functionality are crucial parameters that decide its
application in food and allied industries. The trend has shifted to protein modification in
recent years; researchers are exploring various conventional and novel treatments such as
micro-fluidization, high hydrostatic pressure treatment, enzymatic modification, chemical
modification, fermentation, ultrasonic treatment, microwave treatment, etc., for protein
modification. Protein modification helps to attain desired stability and functionality.
Rice has historically been a major part of an infant’s diet, it is generally among the first
solid foods introduced to a child post the weaning period and this is due to its superior
digestibility and hypoallergenic characteristics.[15] Rice proteins are also incorporated in
infant food supplements and baby foods due to their complete amino acid profile, they
have been shown to possess functional bioactive and anti-oxidative compounds that can
FOOD REVIEWS INTERNATIONAL 3
aid in treating cardiovascular ailments, gut functioning and allergies; thus acting as a novel
food ingredient.[16] They also show potential to act as filling and binding agents that
provide volume, texture, and mouth-feel to meat products such as sausages and salamis.
Protein films derived from rice have also been developed and tested for their mechanical
durability, vapour transmission, concentration effects and overall usage feasibility.[17]
Figure 1. Rice milling process; commercial rice milling systems. Source: [18].
Sotelo et al.[27] reported that polishing brown rice eliminates protein, ash, fat, and fibre
by 13%, 50%, 69%, and 66%, respectively, and these losses were calculated considering the
relative chemical composition and the percentage of each fraction (brown rice 100%, white
rice 91% and bran 9%) of rice grain.
different amount of total protein content 7.4–11.3 g/100 g,[29] 7.16–10.85 g/100 g[4] in
different varieties of rice and this significant difference among protein content of various
rice varieties are possibly due to environmental factors, nitrogen fertilizers, genetic traits
and production system.[30] Wei et al.[31] reported that the total protein of the seven
different rice cultivars determined by the Bradford method was found to be in the range
of 87.9–92,7 mg/g dry weight, while these values were significantly higher than those
obtained using Kjeldahl method (58.2–73.74 mg/g dry weight) as presented in Table 2.
The differences in values obtained from the two methods may be due to the coomassie
brilliant blue staining in Bradford method or the interference of SDS used in the extrac-
tion buffer with protein staining in Bradford method, the other possible reason might be
the interaction between protein and dyestuff.[31] The protein content in rice varies with
the geographical location. Verma & Srivastav[7] studied the six aromatic and two non-
aromatic rice accessions procured from the different parts of India and reported that the
protein content of the rice accessions is in the range of 7.74–10.72% (Table 3).
Table 2. Amount of total protein and low molecular weight proteins (mg/g dry weight).
Total protein Albumin
Cultivar Bradford Kjeldahl +Globulin Prolamin
Ilmibyu 90.8 ± 1.3ab 58.22 ± 0.01a 10.70 ± 0.27ab 1.31 ± 0.01a
Ilpumbyu 95.7 ± 1.3 c 63.20 ± 0.04 c 11.98 ± 0.21b 1.80 ± 0.11b
Saechuchungbyu 92.6 ± 1.6bc 72.35 ± 0.07 f 11.08 ± 0.49bc 2.20 ± 0.08 c
Chuchungbyu 87.9 ± 1.1a 61.12 ± 0.01b 9.94 ± 0.39a 1.64 ± 0.04b
Junambyu 89.9 ± 1.2ab 66.91 ± 0.06e 11.65 ± 0.76bc 1.85 ± 0.10b
Dongjinbyu 91.7 ± 1.6b 64.71 ± 0.01d 10.54 ± 0.62ab 1.60 ± 0.17b
Nampyungbyu 92.7 ± 3.4bc 73.74 ± 0.10 f 11.10 ± 0.40bc 1.84 ± 0.21b
Data are expressed as mean ± standard deviation (n = 3). Difference letters in the same column indicate significant
differences (p < 0.05). Low molecular proteins such as albumin, globulin and prolamin were analyzed by fractionation.
Source: [31].
FOOD REVIEWS INTERNATIONAL 7
Table 3. Protein content and distribution of protein fractions in 12 varieties of rice grown in North
Vietnam.
Total protein recovered
Glutelin
Cultivar Protein content (N × 5.95) Albumin (%) Globulin (%) Prolamin (%) (%)
Tam 10.50 7.00 10.90 8.60 73.50
Du 9.20 8.80 9.40 7.80 74.00
Gie 8.80 10.50 8.30 7.60 73.60
Sai Duong 10.60 4.00 12.50 6.40 77.10
Nep Mua 10.80 8.20 10.50 6.00 75.30
Bau Chau Qui 9.70 7.10 7.60 6.50 78.80
Cuom Chau Qui 8.70 6.20 10.20 7.90 75.70
Tran Chau Lun 9.60 5.90 8.30 8.10 77.70
NN5 7.80 5.40 9.00 7.00 78.60
NN8 8.90 5.80 6.00 8.20 80.00
NN756 7.00 7.40 7.10 5.50 80.00
NR2151 10.60 8.90 11.20 9.50 70.40
Average 9.35 7.11 9.35 7.34 76.23
Source: [32].
PB-I accounts for prolamin and has a spherical lamellar structure; PB-II has no lamellar
structure and is devoid of prolamin subunits, glutelin and globulin are the main compo-
nents of PB-II.[9]All rice bran products including proteins are reported as nontoxic by
various researches and panels such as the Cosmetic Ingredient Review (CIR) expert
panel.[38]
The protein fractions in rice seed are heterogeneous in nature, this could be attributed
to cultivar differences and method of protein extraction used.[39] The distribution of
protein fractions and thus total protein content in rice flour are different compared to
bran. Studies by Ju et al.[40] have reported that flour contains 4.5%, 13.1%, 79.7% and 2.6%
of albumin, globulin, glutelin and prolamin, respectively.
Fractional weights
Adebiyi et al.[41] reported that the molecular masses of albumin, globulin, glutelin and
prolamin fractions were distributed in the range of 30–45, 20–66, 10–66 and 10–53 kDa,
respectively, whereas earlier studies conducted by Hamada,[42] resulted in different values,
their research arrived on rice bran albumin, globulin, glutelin and prolamin to be in the
range 10–100, 10–150, 33–150, and 25–100 kDa, respectively. A similar study by
Amagliani et al.[43] showed that rice albumin was resolved into a wide range of subunits,
with molecular weight (MW) ranging from about 13 to 110 kDa. Shibuya & Iwasaki,[44]
reported Albumin to be between 10 – 200 kDa and globulin to be between 16 and 130
kDa. Various other studies on rice bran and flour protein also report different relative
molecular mass values for the various protein fractions, these discrepancies are assumed to
be caused by the heterogeneous nature of polypeptides in rice flour and bran.
Albumin
Albumin constitutes 4% to 6% of seed protein in rice, around 35% of rice bran, 4% to 8%
of the endosperm and is a minor protein type along with prolamin.[41] Albumin content in
bran is seen to be nearly 6 times greater as compared to normal rice, with albumin
yielding eight peptides upon hydrolysis.[45] Digesting rice albumin with trypsin shows the
presence of 24 peptides whereas 28 are predicted from its amino acid composition. It is
8 N. GHANGHAS ET AL.
seen to possess a major glycoprotein fraction that is a 60 kDa monomer with an isoelectric
point of 6.54.[46] The relative molecular mass values determined by size-exclusion HPLC
were 10–100 kDa[42] however the same was reported to range from 10 to 200 kDa in an
earlier study by Shibuya & Iwasaki,[44] using conventional size-exclusion chromatography.
Albumins are generally extracted by solubilising them with distilled water (400–600 mL)
by shaking or stirring and subjecting it to centrifugation (3500–4000 g for 15 min.) to
allow its separation. They are water soluble and coagulate upon heating like albumins of
other cereals, possessing minimal disulphide cross links and sufficient net charge. They
have been cited to possess appreciable antigenic properties.[35] Ju et al.[40] reported that
their studies yielded 97% protein extraction out of which albumin’s share was reported to
be 4.5% with an Isoelectric pH of 4.1 and enthalpy of 2.88 J/g. Denaturation temperatures
for albumin were reported to be 75.7 degrees and 73 degrees by Ju et al.,[40] and Adebiyi
et al.,[41] respectively.
Albumin hydrolysates are also found to exhibit the highest Angiotensin-converting
enzyme (ACE) inhibiting activity amongst the rice protein fractions, thus possessing high
nutraceutical utility by aiding in the relief of hypertension.[16,47] Functional bioactive
peptides and compounds have been derived from albumin proteins, peptide compounds
such as “oryzatensin” obtained from the ‘RA5ʹ fraction of albumin show bio-regulatory
activity in the human gut by influencing contraction of the ileum.[48] Trypsin hydrolysed
denatured albumin (MW 800 to 2100Da) shows high antioxidant activity, surpassing the
activity of its native Rice Bran Protein (RBP) and hydrolysate forms.[49] Albumin proteins
also hold the minimal allergenic proteins found in rice, such as the 14–16 kDa protein that
inhibits serum IgE binding activity which is seen in the endosperm albumin, these hail
from the wheat α-amylase-trypsin inhibitor family.[50] However, these have not been
observed in RBPs and this may be attributed to the fact that rice bran proteins would
undergo structural disintegration/denaturation during the extraction process that gener-
ally employs alkaline solvents and enzymes. Albumins also possess the highest phytate
content amongst the four, which can act as an anti-nutritional factor that must be
removed before usage in food formulations
Globulin
Globulins are salt soluble proteins and are the second most abundant rice seed protein
fraction. They possess good antigenic properties[35] and have an isoelectric point of 4.3. It is
extracted by solubilizing it with NaCl (400 mL to 500 mL for 3 to 4 h) and centrifuging at
4000 g for 15 to 20 min.[41] Globulin fractions are seen to have molecular weights of 10–150
kDa.[42] They are found in crystalline Protein Bodies that range from 2–3 μm in diameter.
Most of these can be seen in the sub-aleurone layer.[9] It is predominantly found in rice bran
and polish and accounts for 8–10% of total endosperm proteins.[37] The globulin fraction is
characterized by its lack of aggregation, rice globulins are argued to be made of polypeptide
chains of different sizes linked by inter-chain disulphide bonds.[42] Adebiyi et al.[41] reported
that Rice globulin is seen to possess a structure similar to oat globulin, its solubility
characteristics are seen to be influenced by the presence of random coil structure, anti-
parallel chain of intra-molecular β-sheet structure which is absent in albumin. Globulin also
seems to have a relatively low denaturation temperature of 60 degrees. The rice globulin
fraction is composed of two polypeptides of 23–27 kDa and 16 kDa.[51] The 23–27kDa
polypeptides, termed a-globulin, is structurally homologous to wheat grain glutenin.[52]
FOOD REVIEWS INTERNATIONAL 9
Globulin contains the highest amount of the sulphur-containing amino acids cysteine and
methionine but has a lower level of lysine content as compared to albumin and globulin.
Hamada[42] had earlier reported that cultivars do not contribute to a significant differ-
ence in the amount of globulin proteins that are extracted from bran. More recent studies
(shown in Table 4) however show high discrepancy and they attribute this difference to
both the extraction methods used and the cultivar taken for the study. Spectroscopic
studies conducted on rice globulin show that it exists as predominantly α-helical struc-
tures, and is influenced by pH, ionic strength, by some protein structure perturbants and
heat treatments. Covalent and non-covalent forces are seen to be involved in protein
stabilization while hydrophobic and disulphide interactions influence aggregation upon
heating.[51]
Glutelin
Glutelins are the alkali-soluble protein fraction of rice seed; they are made of two subunits
with molecular weights of 30–35 kDa (α) and 19–25 kDa (β) respectively. Glutelins are
generally considered the primary seed storage protein present in rice unlike other cereals
where prolamin holds this value. It is poorly soluble in water but is readily solubilizes in
alkali and acidic conditions (pH >10 and pH <4, respectively). The hydrophobicity of
insoluble glutelins may be decreased by use of several dissociating agents such as sodium
dodecyl sulfate (SDS), sodium stearate, and cetyltrimethylammonium bromide or other
surfactants.[42] The bran and endosperm are argued to contain 11–27% and 66–78%,
respectively.[56,57] It is mainly found in Protein Body II (PB II) and contains a good
amount of lysine, just behind rice albumin. The molecular mass peak was observed at
a range of 11–52 kDa when analysed using MALDI-TOF mass spectrometry.[41]
The presence of Glutelin as the major protein fraction makes it highly suited towards
alkaline extraction. Dilute alkali solvents like 0.1 M NaOH or 0.1 M KOH are employed to
extract the glutelin fraction out, most food industries, however, employ this method to
obtain purified rice starch. They are also strongly aggregated with disulfide linkages.
Glutelin is said to be present at around 75–81%, 79–83% and 22–45%, in brown rice,
milled rice and rice bran, respectively.[43] Ju et al.[40] reports that glutelin has an Isoelectric
pH of 4.8, which they determined by measuring the supernatant’s turbidity post extraction
from defatted rice flour. Chrastil and Zarins[58] have reported that purified glutelin
subunits, the α (acidic) and β (basic) polypeptides can be segregated into 9 and 5 bands,
respectively, with isoelectric pH of 5.0–8.0 and 8.0–11, respectively.
Prolamin
Prolamins are alcohol soluble protein fractions. They are a minor protein fraction in rice
seed, made of heterogeneous polypeptides that are more evenly distributed across the rice
10 N. GHANGHAS ET AL.
Alkali extraction
Alkali extraction method is the most widely used method of protein extraction. The
proteins are exposed to alkali for dissolving the protein in the preparation of protein
isolate/concentrate.[65] Nashef et al.[66] reported that disulfide containing proteins having
different properties and structures when treated with alkali resulted in similar types of
products but possessing different activation energies. The extreme pH condition during
alkali extraction process changes the native structure of the proteins and also affects the
FOOD REVIEWS INTERNATIONAL 11
nutritional value of the protein. De Groot & Slump[65] studied the severe alkali treatment
on the nutrition value and amino acids composition of proteins and reported the
destruction of cystine and lysine amino acids. Lysine is the limiting amino acid in the
rice proteins therefore alkali extraction method should be used at optimum conditions to
prevent the loss of nutritive value of protein. Paraman et al.,[67] in his research, optimized
the alkali extraction condition of the rice protein with 65.9% yield and 86.9% protein
content. For protein extraction, a suspension of one kg of rice flour in 8 L of de-ionized
water was prepared and homogenized for 5 min. The pH of the suspension was adjusted
to 11 by 1 M NaOH solution and stirred for 3 h at 40° C. The solution was centrifuged for
15 min at 3840 × g to separate the solubilised protein; the supernatant was collected
separately while the same procedure was repeated on the residue to extract the additional
protein. The pH of the supernatant collected was adjusted to the iso-electric pH (4.5) to
precipitate the protein. After pH adjustment, the solution was kept undisturbed for 1 h at
4° C and protein precipitate was recovered by centrifugation for 20 min at 5000 × g. The
de-ionized water (pH 4.5) in w/v ratio of 1:4 was used for washing of protein precipitate.
The pH of the protein precipitate thus obtained was adjusted to 7.0, followed by freeze
drying and storage at 5°C.[67]
12
Cultivars
Amino acids Keu Nak Ohd Wha Tam Du Gie SD NM BCQ CCQ TCL NN5 NN8 NN 176 IR-2151
Cys 3.60 2.80 2.70 3.20 1.00 1.40 2.40 2.00 1.40 1.12 1.89 1.37 2.22 2.31 1.85 2.01
Asp 10.00 9.60 9.80 10.40 8.00 9.20 9.00 10.20 8.67 9.15 8.75 9.00 7.80 8.06 8.75 9.86
Glu 26.60 26.70 26.40 27.00 20.00 15.00 21.10 16.00 17.15 19.33 17.21 18.00 16.40 19.21 16.65 18.20
Ser 5.40 5.80 5.60 5.60 3.00 3.60 5.90 5.90 6.00 3.99 4.50 3.00 5.50 5.15 6.00 6.18
Gly 5.80 4.90 4.60 4.40 4.50 4.90 4.33 5.00 4.65 4.00 4.80 4.01 4.33 4.27 4.85 4.66
His 3.50 3.40 3.80 3.60 1.00 1.80 2.50 2.00 1.46 1.79 1.80 1.38 2.05 2.44 1.95 2.05
Arg 3.90 3.80 4.00 3.80 7.20 8.15 9.30 8.77 8.25 7.11 9.00 8.66 8.25 7.21 8.11 7.85
Thr 4.10 4.20 4.10 3.90 3.40 3.48 4.43 4.00 3.37 3.15 4.01 4.00 3.35 3.15 3.67 4.02
Ala 5.00 5.50 5.80 5.80 4.50 5.11 6.50 6.01 6.02 4.80 5.33 4.61 5.00 5.11 4.99 5.77
Pro 8.80 7.90 7.20 6.60 3.00 3.90 4.33 6.90 4.00 3.77 4.15 4.22 3.81 3.00 4.15 3.47
Tyr 0.30 0.30 0.20 0.20 4.50 4.00 3.60 4.12 4.70 3.80 5.01 4.30 3.99 4.33 3.03 4.11
Val 6.10 6.50 6.70 6.50 5.30 5.70 5.76 7.00 7.30 5.22 6.15 5.00 6.28 6.90 5.11 5.38
Met 0.80 1.50 1.30 1.20 1.40 1.03 1.77 1.60 1.35 1.58 1.38 1.19 1.61 1.39 1.47 1.42
Ile 3.60 3.70 3.90 3.90 5.40 4.25 5.00 4.82 4.54 4.15 5.35 4.84 4.10 4.29 5.10 5.01
Leu 6.90 7.50 7.70 7.90 8.82 8.67 8.19 8.40 8.71 8.00 8.63 8.75 8.57 8.65 8.00 8.32
Phe 4.30 3.90 4.10 4.10 4.00 4.73 5.01 5.81 4.60 1.18 4.35 5.00 4.15 5.11 4.33 4.22
Trp 0.00 0.30 0.20 0.20 1.00 1.61 1.52 1.70 1.72 1.18 1.37 1.31 1.62 1.12 1.33 1.34
Lys 1.30 1.60 1.80 1.70 4.30 4.67 4.95 3.39 4.71 3.10 3.95 4.17 3.80 4.00 5.10 4.25
Keu: Keumhobyu; Nak: Nakdongbyu; Ohd: Ohdaebyu; Wha: Whaseongbyu; SD: Sai Duong; NM: Nep Mua; BCQ: Bau Chau Qui; CCQ: Cuom Chau Qui; TCL: Tran Chau Lun. Source: [31].
FOOD REVIEWS INTERNATIONAL 13
at 20° C. The protein fractions were precipitated by adjusting the pH of supernatant to the
iso-electric point of each protein fraction, except prolamin which was precipitated by
acetone. The iso-electric pH determined by Ju et al.,[40] using turbidity measurement and
reported that albumin and glutelin precipitate at pH 4.1 and 4.8, respectively, while
globulin precipitate at both 4.3 and 7.9 pH. After the precipitation, proteins were washed
twice by distilled water and freeze dried after adjustment of pH to 7.0.[40]
Enzyme-assisted micro-fluidization
Xia et al.[74] reported that rice protein can be effectively extracted from broken rice by
applying micro-fluidization treatment to wet-milled broken rice, followed by density-based
separation. Microfluidization treatment disrupts the protein-starch agglomerate, and further
enzyme treatment removes the starch residue and increases protein purity. In this method,
distilled water is added to broken rice in solid to solvent ratio of 1:20 (w/v) and colloid
milled (wet milling) for 30 min. After colloid milling, the slurry is processed by two pass
microfluidization at 100 MPa. This is carried out to disrupt the protein-starch agglomerate.
The microfluidized slurry was centrifuged for 10 min at 8000 × g, followed by decantation of
the supernatant thus obtained. Two different layers, one rich in protein and the other in
starch were observed in the precipitate due to their difference in density. The supernatant
protein layer was scrapped cautiously and re-dispersed with 10 times (w/v) distilled
water.[74] For increasing the purity of the protein-rich fraction enzymatic method was
employed as discussed in previous section.
those foods depends. The functional properties depend on various factors such as extrac-
tion method used, treatment employed, food pH, concentration of salt & sugar in the
foods, and also on the other components present in the food such as carbohydrates &
lipids.[13,14]
Solubility
Rice protein isolate has a very low solubility in the pH range of 3 to 9 and above pH 9 it
shows a slight increase in solubility.[11] According to Romero et al.,[75] rice protein
concentrate shows poor solubility (25–55%) over a wide range of pH (2 to 10) and
minimum solubility at iso-electric point (pH 4.5). Glutelin is a high molecular weight
protein and it is composed of subunits which are bound by the di-sulphide linkages
therefore it is soluble only in dilute acid or alkali. The high percentage of glutelin fraction
(about 80%) of rice protein results in the low solubility of the rice protein isolate. Shih &
Daigle[11] described that rice protein alone shows very limited solubility in water but
shows significant increase in the solubility in the presence of xanthan gum. The portion of
rice seed from which the protein is extracted significantly effects the solubility. In the pH
4–7 the Rice Bran Protein solubility is higher than brown rice protein and white rice
protein while at the pH value >7 or <4, solubility of white rice protein is reported to be
higher than brown rice protein and rice bran protein.[76] This difference in solubility is
due to the Glutelin which is the major component of the protein extracted from both
Brown Rice and White Rice. Solubility of rice protein is one of the limiting factors for its
application in food. Wang et al.[12] reported solubility of rice protein increases signifi-
cantly compared to the control sample when rice protein is freeze milled. According to
Wang et al.,[12] freeze milling exerts a profound mechanical energy on the Rice Protein
which results in the unfolding of the rice protein. The unfolded conformation exposes the
functional groups to the solvent and this strengthens the interactions of water and protein.
Pietrysiak et al.[77] used direct steam injection (DSI) process to exploit the potential
application of pea rice (PR) protein isolate blend and reported significant increase in
the solubility of DSI-PR. Solubility of PR control sample was <4% while the solubility DSI-
PR at pH 3, 6, 9 were <14%, 14.9% and 50.1%, respectively.
Foaming properties
Foaming property of proteins is crucial in the production of different types of foods.
Foams are two-phase system having air cells separated by a continuous liquid layer. For
application in a variety of foods, proteins should form stable foams effectively and rapidly
over the pH range of the food and also at low concentration.[78] According to Tang
et al.,[79] as a prerequisite to have foam capacity, the proteins should solubilize in the
aqueous phase and rapidly unfold to form a cohesive layer of protein around gas/air
droplets. Diffusion, rapid changes in the conformation and succeeding rearrangement at
air–water interface is crucial to obtain protein-based foams[12] and for this, flexible protein
molecules with few secondary and tertiary structures are required[79] and for the devel-
opment of stable foam, the formation of continuous intermolecular polymer of protein
molecules around the air bubbles is important. This is because stable foam formation
requires this intermolecular cohesiveness and elasticity.[79] The foaming capacity (FC)
16 N. GHANGHAS ET AL.
depends on the pH, salt concentration, and sucrose concentration. The increase of the pH
from 5 to 11 has resulted in 3.65-, 5.52-, and 4.21-fold increase in the FC of brown rice
protein, white rice protein and rice bran, respectively. The FC of BRP, RBP and WRP
remains unaffected up to 12% sucrose concentration but above 12% FC declines signifi-
cantly (P < .05). The increase in salt concentration from 0.4% to 2%, significant (P < .01)
increase in FC of Rice Bran Protein was reported, while for the Brown Rice Protein and
White Rice Protein, the foam only significantly (P < .05) increased at low salt concentra-
tion and above 0.8% FC remains constant.[76] Khan et al.[80] reported that un-stabilized
rice bran protein isolate (Un-PI) forms relatively dense and stable foams as compared to
microwave stabilized rice bran protein isolate (MW-PI), dry heat stabilized rice bran
protein isolate (DH-PI) and parboiled rice bran protein isolate (PAR-PI). Foaming
capacity and foaming stability are maximum for Un-PI (15.7 mL, 84 min) followed by
MW-PI (10.5 mL, 74 min), DH-PI (10.4 mL, 71 min) and PAR-PI (9.2 mL, 56 min).[80]
Chandi and Sogi[81] reported FC of rice bran protein concentrate (RBPC) extracted from
HBC 19 rice cultivar (5.2%) is higher than the casein (3.95%) at pH 5 and at neutral pH,
the FC of both casein (14.25%) and RBPC of HBC 19 (8.10%) has increased. The FC of
Casein decreased to 10.15% while FC of RBPC of HBC 19 increased to 10.03% when the
pH was shifted from 7 to 9. The possible reason for the lower FC at acidic pH might be the
isoelectric point (pH 4.5) of the rice bran protein.
Emulsifying properties
Emulsification properties are defined in terms of emulsifying activity index, emulsifying
capacity and emulsifying stability, and these properties are crucial for food applications
such as the development of various traditional and novel foods. Although the main
emulsifying agents are proteins, but its emulsifying activity within the food can alter
due to the presence of carbohydrates and might result in the change in quality of
food.[82,83] Proteins and other amphoteric molecules facilitate the formation of stable oil
droplets through the development of interfacial membranes and improve the oil droplet
dispersion in the immiscible phase of the emulsion by preventing the coalescence of
droplets.[82] Emulsifying activity index is defined as the protein’s ability to induce creation
of the newly formed dispersed particles in emulsions, whereas emulsion capacity denotes
the maximum amount of oil that is emulsified by standard amount of protein under
specific conditions. Emulsifying stability is measured as the amount of cream/oil separated
from emulsion during a specific period of time at definite temperature and gravitational
field.[78,83] Emulsion formation depends on various factors such as homogenizer type,
degree of homogenization, volume fraction of oil, pH, and volume of dispersion, type of
oil and also on the starch incorporated into the food.[82,83] Cao et al.[76] studied three types
of rice proteins, i.e. brown rice protein, white rice protein, and rice bran protein at
different conditions of pH, salt and sugar and reported that the white rice protein has
18.52% higher emulsion capacity at pH 3–11 than that of rice bran protein. At pH 11, the
three types of rice protein – brown rice protein, white rice protein and rice bran protein
showed maximum emulsifying volume of 44.32%, 47.06%, and 42.93%, respectively.
According to Cao et al.,[76] white rice protein (mainly glutelin) is a better emulsifier
than the other two types of rice protein in strong acidic and alkali conditions because of
the improvement in protein solubility due to the breakdown of disulphide bonds. The low
FOOD REVIEWS INTERNATIONAL 17
Water absorption
Water-absorption capacity and water-holding capacity determine the ability of a food
product to imbibe and hold water in its matrix. This property is especially important to
food substances that need high water retention; baked foods rely on high water absorption
capacity to minimize moisture loss post packaging. WAC also maintains freshness, overall
product mouth feel and texture.[81] Water absorption capacities are generally determined
by dissolving 0.5–1 g of the protein in 5–10 mL of distilled water, allowing it to rest,
centrifuging at around 3000 g and measuring the volume of the supernatant.[84] Chandi &
Sogi[81] carried out water absorption studies of Rice bran protein concentrates from
various rice varieties including Basmati and found them to have good WAC enabling its
usage in products that need high water retention. Their studies also showed that water
absorption and Nitrogen solubility index of proteins are related in some manner, casein
with the lowest Nitrogen solubility index is seen to have the least water solubility.
Parboiling has a negative impact on water absorption, supposedly due to conformational
changes induced by processing.
Zhau et al.[85] reports that the water holding capacity of rice endosperm protein and
rice dreg protein are 2.81 g/g and 3.02 g/g, respectively, these values are lower than that of
Soy protein isolates but however are well under the values stated by Aletor et al.[86] as
recommended for viscous foods. Cao et al.[76] have also outlined in their studies that
Brown Rice Protein and White Rice Protein have low water-binding values of 1.96 and
1.78 mL/g, respectively, followed by Rice Bran Protein with (3.54 mL/g). Water absorption
in the range of 1.49–4.72 g/g is deemed as essential for viscous food systems like gravies,
curries and soups,[86] thus rice protein is seen as a component that can surely serve the
purpose of water retention in recipes. Rice bran protein, in particular, can be used as an
ingredient for achieving the same.
Patil & Khan,[84]’s studies suggest that Parboiling seems to aid oil absorption, contrary to
its effect on water absorption, the unfolding of protein structure and exposure of more
hydrophobic groups allows the physical entrapment of oil, making it clear that heat
treatment has a significant effect on absorption capabilities as it can directly affect the
protein’s structural conformation. Rice protein and its hydrolysates are seen to have lower
Oil absorption capacities as hydrolysis breaks protein chains, exposing internal hydro-
phobic groups. Good oil absorption properties of ingredients are critical to food systems
like sausages, nuggets and batters.[59,87] Rice bran proteins possess good oil absorption
characteristics with a value of 3.83 mL/g followed by Brown rice protein and White rice
protein with 2.93 mL/g and 2.56 mL/g, respectively.[76] RBP’s oil absorption values are
seen to be superior to both casein and soy protein isolates.[81]
Protein modification
As discussed in previous section, the rice protein has shown different functional properties
at different pH, salt & sugar concentration, etc., which are due to the structural and
conformational modification of protein caused by the external factors. The structure and
nature of protein plays a crucial role in deciding the protein functionality. During food
processing, various intrinsic and extrinsic factors of food results in modification in the
native structure and conformation of protein, thus leading to alteration in it is the way of
interacting with other food components. The changes occurred during processing may
have either desired or undesired effects on the functionality as well on the nutritive value
of proteins. For application in food model, one should have clear understanding of the
protein structure and its correlation with the specific functional property. The better
understanding of molecular basis for functional property of protein requires the compre-
hensive knowledge of the forces responsible for protein structure and its interaction.[88]
The structure – function relationship of protein largely depends on the covalent cross-
links naturally present. Altering the naturally present cross-links or introducing new
cross-links can result in the modification of the functional properties.[89] Various chemi-
cal, physical, and enzymatic methods have been employed for the protein modification
with the aim of enhancing nutritive values and functionality of protein. The chemical
modification involves the derivatization of few amino acids and side chains. Several
reactive side chains are present in the primary structure of protein and chemically
modifying them can result in functional enhancement of protein. Its aim is to change
the non-covalent forces responsible for protein conformation to obtain desired functional
enhancement.[90,91] Enzymes such as papain,[92] trypsin,[93] pronase,[94] etc., can also be
used for altering or extending the protein functionality. Enzymatic modification involves
the incorporation of inter-molecular or intra-molecular cross-links, or partial hydrolysis
of polypeptide chain, or introduction of specific group to the proteins.[91]
In recent studies various novel physical methods such as microfluidization,[74] high-
pressure processing,[95] freeze milling,[13] high-intensity ultrasound,[96] electron beam
irradiation,[97] radiofrequency,[98] extrusion cooking,[99] etc., have been explored for the
functional enhancement of proteins. Effect on the nutritive value and functionality of the
rice protein due to the different methods employed for protein modifications have been
studied by researcher. Few of the recent studies on rice protein modification using
different methods and their effect on protein functionality are depicted in Table 7. For
FOOD REVIEWS INTERNATIONAL 19
Table 7. Effect of various protein modification methods on the functionality of rice protein.
Protein
S. No. Modification method source Effect on functionality of protein References
[100]
1 Controlled enzymatic Rice Solubility and emulsification enhancement
hydrolysis endosperm
[97]
2 Electron beam Rice seed Improves solubility and emulsification
assisted enzymolysis
[101]
3 Glycosylation, Rice Enhancement of solubility and emulsifying properties
deamidation, endosperm
proteolysis
[13]
4 Freeze milling Rice seed Improved Foaming, emulsification and solubility
[102]
5 Glutaminase Rice seed Improved structural and sensory properties
treatment
[98]
6 Radio frequency Rice bran Improvement in absorption and emulsifying properties while
the negative effect on other functional properties
protein modification, the method needs to be selected carefully considering its final
application, effect on nutritive value, undesirable effect on the other functional properties,
toxic by-products which might develop during the processing or due to the reaction with
the other components of food, etc.
Meat products
The various functional and nutritional advantages possessed by rice protein have paved its
way for the application in food formulation and food processing. The strong antioxidant
property of rice protein hydrolysates (RPH) is effective in improving the shelflife of the
meat products by preventing the lipid oxidation. Zhou et al.[109] studied the effect of
addition of RPH on the lipid peroxidation in the cooked ground beef and reported that
RPH can be used in food models to inhibit the lipid oxidation.
In another study, it is reported that fortification of beef patties with rice protein has not
only increased protein content but also reduces cooking loss, and improved the redness
value which is desirable according to consumer point of view. Also, rice protein is found
to show better antioxidant properties in comparison to pea protein isolate and lentil
flour.[110] Rice protein is a natural antioxidant and can be used to replace the artificial
antioxidants.
Baking
Stabilized or parboiled rice bran has appealing characteristics to be used in food formula-
tion. For example, the granulated powder of bran having a cream colour with tasteless and
FOOD REVIEWS INTERNATIONAL 21
properties.[17] The best properties were obtained for rice bran protein films with lower protein
and glycerol concentrations and with the addition of phenolic extract without the presence of
MMT clay. Solubility was mainly influenced by protein concentration, parameters such as
luminosity and opacity of the films also followed a similar trend. However, in case of mechanical
properties, glycerol concentration was the main factor. Phenolic extract addition affected film
opacity, tensile strength and elongation. The major challenge associated with protein or poly-
saccharide films is that they have brittle structure at low relative humidity or low strength at high
relative humidity; thus, modification of film strength and water sensitivity is generally achieved
by changing processing procedures and varying the composition of the mixture by adding
components that strengthen and promote film setting.[118] Studies employing starch and protein
from broken rice to make edible films with appreciable characteristics were also carried out by
Dias et al.[119] They were able to arrive upon films whose properties were similar to solely starch-
based films but possessing remarkably higher vapour permeability. It is also noted that addition
of sorbitol yields better mechanical rigidity and lowers water permeability, but glycerol addition
confers the opposite effect.
Protein-based coating is an alternative way of extending the shelf life of food products.
The rice protein possesses excellent barrier properties and can be used as edible coating
material. Using rice protein extracted from the by-products of rice milling for food
coating may provide us an economical and sustainable solution. da Silva et al.[120] have
reported that rice protein-based coating with suitable plasticizer can be used to preserve
the internal quality and extending the shelf life of eggs. The rice protein-based coating
blocks the pores of shell surface thus resulting in preserving the quality of albumen and
yolk. The use of sorbitol as plasticizer instead of glycerol and propylene glycol is efficient
in maintaining and controlling the increase in the pH of albumen.
Proteins and various polysaccharides have been studied for the encapsulations of bioactives
such as drugs, carotenoids, polyphenols, vitamins, etc. Encapsulation has numerous advan-
tages such as decrease core material reactivity; prevent core material from physical and
chemical damage during processes, controlled release of drugs and bioactive compounds,
etc. Wang et al.,[13] in their study fabricated oil droplets having modified rice protein shell and
core of soybean oil core and studied its potential application for oral delivery of lipophilic
bioactive. The rice protein has extra advantageous compared to soy and dairy protein because
of its hypoallergenicity and other health benefits. The β-carotene encapsulated in these oil
droplets having modified rice protein shell enabled the partial release of β-carotene during the
simulated gastric digestion, while during the succeeding stimulated intestinal digestion
encapsulation enabled the controllable zero-order release rate of β-carotene.
Conclusion
Rice seed protein holds immense potential to act as a healthy, economic and widely
available protein source for consumption; it can act as an excellent novel ingredient and
a quality alternative for existing cereal and animal-based protein options. Its excellent
nutritional content and functional properties make it a highly utilisable option in various
food formulations. Rice seed protein is of excellent quality, possessing appreciable biolo-
gical value, digestibility and efficiency ratios that are comparable to and even exceed those
of casein and soy protein. They also have a sound amino acid profile comprising of
essential amino acids such as threonine, leucine and phenylalanine along with sulphur-
FOOD REVIEWS INTERNATIONAL 23
rich amino acids like methionine and cysteine which are critical for the human body. Rice
seed protein’s hypoallergenicity is a unique trait that enables its incorporation in infant
food formulations and supplements for treating ailments like Cow Milk Protein allergy
and cardiovascular relief. It is also seen to possess bioactive compounds that aid in body
regulation and gut relaxation along with excellent antioxidative properties. Studies have
also highlighted that rice protein derived films can act as sufficiently strong edible
packaging with good barrier properties; it can also act as filler material in meat products.
Moreover, the usage of rice protein extracted from the milling by-products of paddy
would significantly reduce the environmental load caused by them and effectively increase
the value of these underutilised products that are either burnt, used as fodder or fuel. The
main hurdles in the commercialisation of rice protein are the difficulty in efficiently
extracting the protein fractions without causing excessive modification at large scales
and the time required to achieve extraction. Moreover, its lower overall protein content
and the internal distribution of protein fractions make it difficult to find a suitable
solubilising solvent for extraction and its incorporation in food.
Acknowledgments
The support from Department of Food Science and Technology, NIFTEM and NIFTEM Knowledge
Centre is greatly acknowledged.
Disclosure statement
Author and co-authors declare no conflict of interest.
ORCID
Neeraj Ghanghas http://orcid.org/0000-0002-0683-2065
Pramod K. Prabhakar http://orcid.org/0000-0002-1967-6575
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