Human cancer cell proliferation inhibition by a pentapeptide isolated and characterized from rice bran

Article abstract of DOI:10.1016/j.peptides.2010.05.018

Food-derived bioactive peptides promote functional activity against diseases and present as nutraceutical agents. The purpose of our research was to isolate and fully characterize peptide(s) derived from rice bran having anti-cancer properties. Gastrointestinal juices resistant peptide fractions were initially generated from heat stabilized de-fatted rice bran from which <5 kDa fraction was shown to inhibit proliferation of cancer cells. Based on these published findings the <5 kDa peptide fraction was selected for further characterization to obtain single pure peptide(s) with enhanced anti-cancer properties. Purification and characterization from the fraction was done employing chromatography and mass spectrometric techniques. Cancer cell viability was measured using a cell titer assay that uses a tetrazolium dye [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl) -2H-tetrazolium, inner salt; (MTS)] and the electron coupling reagent, phenazine methosulfate. Ion-exchange chromatography elutes that showed anti-cancer properties were further purified to liberate pure peptide. The pure peptide at 600-700 μg/mL dose caused 84% inhibition to colon cancer cells (Caco-2, HCT-116) growth, 80% to breast cancer cells (MCF-7, MDA-MB-231) growth and 84% to liver cancer cells (HepG-2) growth. Mass spectrometry analysis and de novo sequencing revealed the sequence of Glu-Gln-Arg-Pro-Arg for the peptide with a molecular mass of 685.378 Da. A novel pentapeptide was isolated from rice bran to possess cancer growth inhibitory properties on colon, breast, lung and liver cancer cells. This peptide could serve as a nutraceutical agent against cancer.

Full text of DOI:10.1016/j.peptides.2010.05.018

Peptides 31 (2010) 1629–1634
Contents lists available at ScienceDirect
Peptides
journal homepage: www.elsevier.com/locate/peptides
Human cancer cell proliferation inhibition by a pentapeptide isolated and
characterized from rice bran
Arvind Kannana, Navam S. Hettiarachchya,∗, Jackson O. Layb, Rohana Liyanageb
a Department of Food Science, University of Arkansas, 2650 N Young ave, Fayetteville, AR 72703, USA
b Department of Chemistry and Biochemistry, University of Arkansas, 119 Chemistry Building, Fayetteville, AR 72701, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 6 April 2010
Received in revised form 21 May 2010
Accepted 24 May 2010
Available online 1 June 2010
Food-derived bioactive peptides promote functional activity against diseases and present as nutraceu-
tical agents. The purpose of our research was to isolate and fully characterize peptide(s) derived from
rice bran having anti-cancer properties. Gastrointestinal juices resistant peptide fractions were initially
generated from heat stabilized de-fatted rice bran from which <5 kDa fraction was shown to inhibit pro-
liferation of cancer cells. Based on these published findings the <5 kDa peptide fraction was selected for
further characterization to obtain single pure peptide(s) with enhanced anti-cancer properties. Purifica-
tion and characterization from the fraction was done employing chromatography and mass spectrometric
techniques. Cancer cell viability was measured using a cell titer assay that uses a tetrazolium dye [3-
(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt;
(MTS)] and the electron coupling reagent, phenazine methosulfate. Ion-exchange chromatography elutes
that showed anti-cancer properties were further purified to liberate pure peptide. The pure peptide at
600–700 ␮g/mL dose caused 84% inhibition to colon cancer cells (Caco-2, HCT-116) growth, 80% to breast
cancer cells (MCF-7, MDA-MB-231) growth and 84% to liver cancer cells (HepG-2) growth. Mass spec-
trometry analysis and de novo sequencing revealed the sequence of Glu-Gln-Arg-Pro-Arg for the peptide
with a molecular mass of 685.378 Da. A novel pentapeptide was isolated from rice bran to possess cancer
growth inhibitory properties on colon, breast, lung and liver cancer cells. This peptide could serve as a
nutraceutical agent against cancer.
Keywords:
Rice bran
Peptide
Anti-cancer
Characterization
© 2010 Elsevier Inc. All rights reserved.
1. Introduction
Heat stabilized de-fatted rice bran; a co-product of rice process-
ing that is underutilized could be a source of bioactive peptides.
A number of food-derived bioactive peptides have been isolated
from enzyme hydrolysates derived from several food products.
Usually the amino acids in bioactive peptides range from 3 to
16 amino acids and depending upon the nature of their activities
amino acids like Val, Leu, Pro, His, Tyr, Glu, or Asp have been shown
to predominate [18]. Precise characterization of peptides employ-
ing isolation and initial fractionation of the hydrolysates from the
food source followed by purification and characterization using
proteomic tools becomes important in ascribing the peptide to its
bio-functional role, sometimes resulting in more enhanced bioac-
tivity than the hydrolysates from which they were derived from.
As an example, soy protein hydrolysates are a rich source of bioac-
tive peptides including ACE-1 inhibition [3] that were originally
separated using membrane fractionation methods [5].
Nam et al. (2005) [16] studied extracts derived from pigmented
rice bran to show anti-cancer properties. Oryzatensin, an ileum-
contracting bioactive peptide obtained from rice albumin was
shown to have a sequence of amino acids Gly-Tyr-Pro-Met-Tyr-Pro-
Leu-Pro-Arg. This peptide was shown to have immunostimulatory
role that was mediated by histamine release [22]. Based on
recent reports enzymatic hydrolysates prepared from rice bran
was able to show growth inhibitory effects to certain types of
cancer cells in vitro [8,9]. The significance of utilizing enzymatic
hydrolysates for characterization of bioactivities from cereal com-
ponents such as bran is to utilize the proteins that are highly
complexed within rice bran that otherwise limit protein extrac-
tion efficiency [23]. Bandyopadhyay et al. (2008) [2] prepared
protein isolate from full fat rice bran by alkaline hydrolysis and
isoelectric precipitation; obtained peptides from partial hydroly-
sis with papain under controlled conditions and studied functional
properties. From an industrial perspective utility to proteins or par-
tially hydrolyzed protein components from rice bran, hydrolyzed
protein has been produced and exploited as vegetable protein
[6].
∗
Corresponding author at: 2650 N young ave, Fayetteville, AR 72704, USA.
Tel.: +1 479 575 4779; fax: +1 479 575 6936.
E-mail address: nhettiar@uark.edu (N.S. Hettiarachchy).
0196-9781/$ – see front matter © 2010 Elsevier Inc. All rights reserved.
doi:10.1016/j.peptides.2010.05.018

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A. Kannan et al. / Peptides 31 (2010) 1629–1634
With limited literature information about rice bran protein
of the eluates obtained after ion-exchange chromatography (IEC)
was determined using the MTS assay [8].
hydrolysates and peptides exhibiting functional properties, more
research is needed particularly to evaluate potential nutraceutical
attributes of rice bran peptides such as its ability to arrest major
disease progression states.
2.4. Preparative HPLC purification of IEC eluate showing
anti-cancer activity
In this context, and based on earlier published findings that rice
bran peptide fractions have the ability to cause growth arrest in
certain cancer cell type proliferation [8,9], the objectives involved
separation of the <5 kDa fraction from rice bran peptide hydrolysate
by pressure-driven membrane based separation (ultrafiltration),
purification from the <5 kDa fraction to obtain single peptide(s)
that show significant anti-cancer effects, further characterization
to determine accurate mass and amino acid sequence of the pep-
tide(s). The <5 kDa peptide fraction was subjected to ion exchange
and HPLC purification to obtain single pure peptide(s). This was
followed by evaluation of single pure peptides for enhanced cancer
cell anti-proliferation activity. The pure peptide that showed cancer
cell anti-proliferation activity was subjected to mass spectrometry
analysis for determining the accurate molecular mass, amino acid
analysis and tandem mass spectrometry for determining the amino
acid sequence of the pure peptide.
Preparative scale peptide-specific column (Biopore Prep ID 22 x
L 250 mm part # 34955) was used to separate peptides from the
IEC eluates that showed better anti-proliferative activity and the
absorbance of the eluate monitored at 215 nm. The gradient from
solvent A (1.2 mL TFA/1000 mL de-ionized water) to solvent B (0.1%
TFA in Acetonitrile:water 50:50) was varied from 100% solvent A
to 100% solvent B over 80 min at 2 mL/min flow rate monitored
at 215 nm. The peaks were collected and tested for cancer cell
anti-proliferative activity, and the peak that showed such effects
was fully characterized using mass spectrometry and amino acid
sequencing.
2.5. Amino acid analysis
A modified method of AOAC 982.30a (1990) was used for
hydrolyzing purified peptides. 10 mg of peptide samples were
hydrolyzed in 10 mL of 6.0 N HCl under vacuum at 150 ◦C for 12 h
and evaporated under nitrogen at 60 ◦C. Sodium diluent buffer
pH 2.2 (1 mL) was added to the dried peptide, filtered and the
filtrate was analyzed for amino acids. The peptides were pre-
treated with performic acid prior to hydrolysis to preserve cysteine
and methionine, while alkali hydrolysis was conducted to deter-
mine tryptophan (AOAC 982.15, 2000). Amino acid analysis of
the filtrate was conducted on an automated amino acid ana-
lyzer (Beckman 6300, Beckman Instruments, Inc. Palo Alto, CA)
at a flow rate of 0.67 mL/min (0.44 mL/min for buffer solutions
and 0.23 mL/min for ninhydrin solution). Sodium citrate buffers
(pH 3.3, 4.3 and 6.3) were used as eluents. The amino acid con-
tents (in g/100 g sample) were quantified by comparing them with
amino acid profiles from external amino acid standards as follows
2. Materials and methods
2.1. Materials
HDRB (Heat stabilized De-fatted Rice Bran) was obtained from
Riceland foods (Stuttgart, AR), Human colon (Caco-2), breast (MCF-
7), liver (HepG-2), and lung (A549) cancer cell lines including
growth media were purchased from ATCC, USA. Media supplements
including fetal bovine serum, gentamycin, were purchased from
Hyclone, USA. MTS kit was purchased from Promega USA. Prepar-
ative liquid chromatography system LC-8A was purchased from
Shimadzu, USA. Sephadex G-75 resin from Pharmacia biotech AB,
Uppsala Sweden, biopore C-18 preparative HPLC column, amino
acid analyzer from Beckman Coulter, Bruker Reflex III (Bruker
Daltonics GMBH, Bremen, Germany) and Bruker Ultraflex II time-
of-flight mass spectrometers at the Statewide mass spectrometry
facility, University of Arkansas. All other chemicals purchased were
of HPLC grade and purchased from Sigma, MO, USA.
(Peaksample/Peakstandard) × Concentrationstandard × MWstandard
.
2.6. Cancer cell culture and MTS anti-proliferative assay
Human colon, Caco-2 and liver, HepG2 cancer cells were cul-
tured in DMEM (Dulbecco’s modified eagles medium) in the
presence of 10% fetal bovine serum, supplemented with 1 mM
l-glutamine, Sodium pyruvate, 1 mM, Sodium bicarbonate and
50 ␮g/mL gentamycin. Human breast, MCF-7 cell line was cul-
tured in Eagle’s Minimum essential medium with the following
supplements: 2 mM l-glutamine, Earle’s BSS adjusted to contain
1.5 g/L sodium bicarbonate, 0.1 mM non-essential amino acids,
1 mM sodium pyruvate and 0.01 mg/mL bovine insulin, 90%, fetal
bovine serum, 10% and 50 ␮g/mL gentamycin. Human lung A-549
cancer cells were cultured in Ham’s F12K medium with 2 mM l-
glutamine adjusted to contain 1.5 g/L sodium bicarbonate, 90%;
fetal bovine serum, 10% and 50 ␮g/mL gentamycin at 37 ◦C in a 5%
CO2 incubator. All cells were cultured at 37 ◦C in a humidified 5%
CO2 incubator.
2.2. Sample preparation
Ground and sieved HDRB was treated with alcalase under opti-
mized conditions to generate peptide hydrolysates, which in turn
was passed through simulated gastrointestinal juices and fraction-
ated using ultrafiltration fractionation columns to generate definite
molecular sized fractions as described by Kannan et al. [8]. The
<5 kDa fraction previously shown to promote growth inhibitory
effects on cancer cells was subjected to purification and charac-
terization employing the following methods.
2.3. Ion-exchange chromatography
A sephadex G-75 ion exchange resin was packed into a glass
column and equilibrated with 10 mM phosphate buffer, pH 8.0.
10 mL of <5 kDa peptide hydrolysate (∼1 mg/mL protein concen-
tration) was loaded onto the column at 1 mL/min flow rate. The
elution was started by washing the unbound hydrolysate eluted
with 10 mM phosphate buffer. After washing, the hydrolysate was
eluted using 10 mM phosphate buffer containing 50 mM NaCl fol-
lowed by elution with 10 mM phosphate buffer containing 100 mM
NaCl. The eluates were collected, concentrated in an Amicon con-
centrator with buffer exchange and stored at 4 ◦C until used for
anti-cancer activity determination. The anti-proliferative activity
Cell proliferation inhibition was assayed using the phenazine
methosulfate 3-(4,5-dimethyl thiazole-2-yl)]-2,5-diphenyl tetra-
zolium bromide (MTS) mix-based cell titer assay. The cells were
transferred onto flat 96-well bottom plates following trypsinization
(0.1% trypsin–0.53 mM ethylene diamine tetra acetic acid (EDTA)
solution) from cell culture flasks used earlier for culture of cells.
Approximately 1000 cells per well were used for growth onto
96 well flat bottom plates. The cells were allowed to attach and
grow for 24–36 h. After 24–36 h old media were replaced with
fresh media, and samples of bran peptide fraction/pure peptide
were treated with the cells. After 24 h exposure of the samples to

A. Kannan et al. / Peptides 31 (2010) 1629–1634
1631
the cell monolayers, the phenazine methosulfate 3-(4,5-dimethyl
thiazole-2-yl)]-2,5-diphenyl tetrazolium bromide (MTS) mix was
added at a final volume of 20 ␮L/100 ␮L medium and then incu-
bated for an additional 60 min under the same conditions. The
reaction was terminated by adding 10% SDS, and the absorbance
of formazan measured at 490 nm. Saline was used as a viability
(untreated) control. Genistein was used as a positive control. All
assays were performed in triplicates and the results expressed as
mean values standard error. The percentage of cell inhibition was
calculated from ratio of experimental values compared with con-
trols as follows: (A490 Experiment/A490 Control) × 100.
Fig. 1. The anti-proliferactive activity of IEC eluted fraction in human cancer cell
lines by MTS assay.100 mM: 100 mM NaCl eluate; 50 mM: 50 mM NaCl eluate. Values
are means trials SE. Values not connected by same letters are significantly different
(P < 0.05). Untreated (negative control): saline.
2.7. Mass spectrometry characterization of pure peptide
For preliminary intact mass determination, 1.0 ␮L of the HPLC
purified peptide was mixed with 1.0 ␮L saturated HCCA and spotted
on Bruker MTP 384 ground stainless steel MALDI target. MALDI-TOF
(time of flight) mass spectra were acquired on a Bruker Reflex III
(Bruker Daltonics GMBH, Bremen, Germany) and Bruker Ultraflex
II time-of-flight mass spectrometers operated in the positive-ion
reflectron mode. High resolution exact mass of the same peptide
was obtained by using MALDI Ionspec 9.4 T ion cyclotron reso-
nance (ICR) mass spectrometer. For the exact mass measurements
2,5 dihydrobenzoic acid (DHB) was used as the MALDI matrix and
followed exactly the same spotting technique.
from 100% solvent A to 100% solvent B over 80 min at 2 mL/min
flow rate monitored at 215 nm. Fig. 2 shows the HPLC profile of
50 mM IEC eluates. The peaks were collected at 10 min intervals and
tested for cancer cell anti-proliferative activity. It was found that
the 60–70 min fraction showed anti-proliferative activity and this
fraction was further isolated in HPLC. Further purification form the
60–70 min fraction resulted in a single peptide which was tested
for anti-proliferative action on the cancer cell lines.
3.3. Cancer anti-proliferative activity of pure peptide
2.8. MALDI fragmentation
The pure peptide at nearly 1000 ␮g/mL showed 84% inhibi-
tion on colon cancer cells, 80% on breast cancer cells and 84%
on liver cancer cells significantly different from untreated con-
trol. On breast and liver cancer cells the pure peptide showed
80% and 85% inhibitions respectively, while on lung cancer cells
there was 69% inhibition (Fig. 3). The results show that the peptide
bears strong cancer cell anti-proliferative activities better than the
<5 kDa peptide fraction from which it was purified. Furthermore,
dose–response pattern of the pure peptide, as depicted in Fig. 3
revealed that, from 600 ␮g/mL to 1000 ␮g/mL maximum inhibition
was achieved on most cancer cells after a 24 h time period. There is
an increase in inhibition pattern as the dosage increases and seems
to saturate off beyond 600 ␮g/mL. The dosage for the pure peptide
is similar to what was observed with the <5 kDa fraction on most
cancer cell lines as per the earlier studies [8] except there is a 24 h
response time for significant anti-cancer activity.
Fragmentation of intact peptide ions was performed to obtain
sequence information using MALDI post source decay studies.
MALDI-PSD fragmentation of these ions was analyzed using the
“Lift” mode in Bruker Ultraflex II (Bruker Daltonics GMBH, Bre-
men, Germany) time-of-flight/time-of-flight mass spectrometer
(TOF/TOF MS).
2.9. Data analysis and de novo sequencing
Fragmentation pattern obtained by MALDI-TOF-TOF was inter-
preted using Bruker Biotools software, which uses de novo
sequencing algorithm to determine the best sequence for the
observed fragmentation pattern.
2.10. Data analysis
3.4. Characterization of the peptide
Experimental data were analyzed using JMP 7.0 statistical soft-
ware with the least significant differences between samples being
P < 0.05.
Characterization involved determination of amino acid analysis,
mass and amino acid sequencing of the peptide. The results of mass
and amino acid sequencing are shown in Figs. 4 and 5 respectively.
3. Results
3.4.1. Amino acid analysis
3.1. Anti-proliferative activity of ion-exchange chromatography
eluates
Amino acid analysis revealed the predominance of 3 amino
acids suggesting the anti-cancer peptide could be a short peptide.
The amino acids identified were arginine (2.65 nmol/mL), proline
(0.9 nmol/mL), and glutamic acid (1.5 nmol/mL).
The <5 kDa fraction was subjected first to IEC and the peaks
were eluted using 50 mM and 100 mM NaCl. On colon and liver
cancer cells, the 50 mM NaCl eluate showed nearly 75% inhibition
by the MTS dye assay. On breast cancer cells there was 68% inhibi-
tion while on lung cancer cells there was 60% inhibition (Fig. 1). The
eluate showing promising anti-proliferative activities on cancer cell
lines were selected for further purification.
3.4.2. MALDI characterization of peptide
Matrix assisted laser desorption ionization time of flight mass
spectrometry (MALDI-TOF-MS) analysis was performed on the
HPLC purified peptide. Fig. 4 shows the MALDI-TOF-MS spectrum
of the purified fraction and confirms the homogeneity of the frac-
tion. The accuracy of the mass was confirmed by MALDI-ICR (ion
cyclotron resonance)-MS where the mass of the ion was measured
to 10 ppm accuracy. By means of MALDI–TOF MS, single protonated
molecular ions (M + H+) of the intact peptide were located at m/z
685.378 suggesting the possibility of a short peptide.
3.2. Reverse phase HPLC purification
The 50 mM IEC eluate was subjected to HPLC in a preparative.
The gradient from solvent A (1.2 mL TFA/1000 mL de-ionized water)
to solvent B (0.1% TFA in Acetonitrile:water 50:50) was varied

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A. Kannan et al. / Peptides 31 (2010) 1629–1634
Fig. 2. HPLC profile of 50 mM IEC eluate. 50 mM NaCl eluate from IEC subjected to HPLC. Peaks were collected every 10 min of the run and analyzed for anti-cancer activity.
60–70 min peak (arrowhead) showed anti-cancer activity, which was further isolated in HPLC.
3.4.3. Amino acid sequencing of the peptide
peptide to homogeneity, and mass spectrometry helped decipher
the accurate mass and the amino acid sequence of the peptide.
Since the 50 mM ion-exchange chromatography eluate showed
cancer cell inhibitions better than the 100 mM eluates, the 50 mM
eluate possibly had the pool of <5 kDa peptides that contribute
towards the anti-proliferative effect on the cancer cells. Ion-
exchange chromatography has been the most popular tool used
especially to separate peptides from protein hydrolysates in the
past decade. Moreover bioactive peptides separated out in ion-
exchange columns have been subjected to an additional reverse
phase HPLC step to get pure peptides amenable for mass spec-
trometry and sequence studies. Hence the 50 mM IEC eluate
was subjected to HPLC purification using a peptide-specific col-
umn.
Several peptides from food sources have been isolated to purity
using this technique [12] where a mixture of solvents, usually
acetonitrile and water with ion pairing reagents like trifluo-
roacetic acid aids in formation of a polar to non-polar gradient.
The 60–70 min eluate obtained from the HPLC run showed anti-
proliferative activities, and further a peptide was isolated from this
elute and characterized for significant anti-proliferative activities
to cancer cells. The eluate suggested the presence of both polar and
non-polar amino acids evident from the retention time, which was
also confirmed by amino acid analysis. Although there were several
peaks eluting throughout the run time, these did not show anti-
proliferative activity as opposed to the 60 min fraction. It would be
interesting to test and characterize these peaks for other bioactiv-
ities.
Further tandem (MS–MS) was performed on the predominant
peak by isolating the ion inside the mass spectrometer to enable
post source decay (PSD) fragmentation [20] to obtain amino acid
sequence (Fig. 5). Three amino acids were identified from the C-
terminus end as Arg-Pro-Arg. The N-terminal amino acids when
matched against a database showed a high prediction of Glu-Gln
amino acids in the N-terminus end (Fig. 5: Inset). The de novo
sequencing thus revealed the peptide to have the amino acid
sequence as Glu-Gln-Arg-Pro-Arg (EQRPR).
4. Discussion
The study shows characterization of a peptide from rice bran
to have anti-proliferative activities on multiple cancer cell lines,
signifying that a natural co-product of rice is a potential source
of bioactive anti-cancer peptides. IEC and HPLC helped purify the
The activity of bio-functional peptides is based on their inher-
ent amino acid composition and sequence. The size of active
sequences may vary from two to twenty amino acid residues,
and many peptides are known to have multifunctional proper-
ties [13], e.g., peptides from the sequence 60–70 of ␤-casein show
immunostimulatory, opioid and angiotensin I converting enzyme
(ACE)-inhibitory activities. Here we found the possibility of a tetra
to penta peptide bearing anti-cancer properties against several
types of cancer cell lines. Other studies have also produced short
peptides possessing biological activities like immunomodulatory,
ACE-inhibitory, hypocholesterolemic and anti-oxidative properties
[14,15,17,18,21]. The peptides range from di-, tri- to penta-peptides
[19] isolated from ␤-lactoglobulin by tryptic hydrolysis. The shorter
the peptide chains, more the solvent accessibility, exposing the
functional groups to impart functional properties. Moreover pres-
ence of proline signifies more accessibility to solvent owing to
its predominance in turns of a polypeptide chain. Occurrences of
proline residue in bioactive peptides are not uncommon and are
also thought to be a major player to contribute to bioactivity of
Fig. 3. Anti-proliferative activity of doses of pure peptide at 24 h of exposure on
cancer cells by MTS assay. Values are means trials SE. Values not connected by
same letters are significantly different (P < 0.05). Negative (viability control): saline;
Positive (inhibitory control): Genistein (400 ␮g/mL).
Fig. 4. MALDI profile of the pure peptide obtained after HPLC purification. x-axis:
mass to charge ratio (m/z); y-axis: intensity. Molecular mass of the peptide was
identified to be 685.378 Da. Inset: Purified peak at 62 min elution from HPLC.

A. Kannan et al. / Peptides 31 (2010) 1629–1634
1633
Fig. 5. Tandem MS–MS PSD fragmentation of peptide 685.37 Da.
Inset: de novo sequencing of peptide matched for probable sequence based on exact mass of 685.378 Da.
Peptide was de novo sequenced by using the MS spectra monoisotopic mass of the amino acids. Inset: de novo sequencing of peptide matched for probable sequence based
on exact mass of 685.378 Da. Three amino acids were identified from the C-terminus end as Arg-Pro-Arg. The N-terminal amino acids when matched against the database
showed a high prediction of Glu-Gln amino acids in the N-terminus end. De novo sequencing revealed the amino acid sequence as Glu-Gln-Arg-Pro-Arg (EQRPR).
the peptide [1]. Precise sequence information can help peptides to
be synthesized in vitro, and also in large scale where the finished
product can be suitable for ingredient formulation functioning as a
nutraceutical agent.
with bioactivity against human cancers and promote further study
of value added co-products from cereal grains.
References
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from the <5 kDa rice bran hydrolysate resulted in identifying the
cancer anti-proliferative component within rice bran. The presence
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milk proteins have been possible [24]. Lunasin, an anti-cancer pep-
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