Volume 12, No 2
International Journal of Radiation Research, April 2014
Protective effect of ferulic acid on ionizing
radiation induced damage in bovine serum albumin
K. Mishra#, H. Ojha#, S. Kallepalli, A. Alok, N. Kumar Chaudhury*
ChemicalRadioprotectorandRadiationDosimetryResearchGroup,DivisionofRadiationBiosciences,Instituteof
NuclearMedicineandAlliedSciences,Delhi‐110054,India
#Contributedequally
ABSTRACT
► Original article
*Correspondingauthor:
Dr.N.KumarChaudhury,
Fax:+911123919509
E‐mail:
[email protected]
Received: Feb. 2013
Accepted: May 2013
Int. J. Radiat. Res., April 2014;
12(2): 113-121
Background: Ionizing radia on causes deleterious effects on living system
mainly due to oxida ve damages of macromolecules and protein is the major
target due to its abundance. The aim of this study was to inves gate the
effects of ionizing radia on induced changes in the molecular proper es of
bovine serum albumin (BSA); its secondary and ter ary structures,
degrada on, cross linking and radioprotec ve role of ferulic acid, a natural
an oxidant on these radia on induced changes. Materials and Methods: This
study was carried out to inves gate the gamma radia on induced oxida ve,
structural damage of BSA and radioprotec ve efficacy of ferulic acid through
SDS‐PAGE, DTNB assay, DNPH assay, FOX assay methods. Hydroxyl radical
scavenging capacity of ferulic acid was es mated using 2‐deoxy ribose assay.
Further, radia on induced changes in the anisotropy and excita on state
life mes of BSA were examined. Results: SDS –PAGE data suggested that the
loss of protein was linearly dependent on the radia on dose.
Gamma‐irradia on of BSA caused the forma on of protein carbonyls,
hydroperoxides and loss of thiols . Ferulic acid protected the radia on induced
loss of protein as well as reduced various oxida ve damages. Ferulic acid
protected the protein from radia on induced damages in a concentra on
dependent manner. Conclusion: The results provide insight into radia on
induced molecular changes in the protein. Ferulic acid protected the BSA from
oxida ve modifica on caused by radia on sugges ng that ferulic acid
possesses strong an radical proper es. Ferulic acid is known to protect DNA,
the prime target of radia on and further its ability to protect protein
sugges ng its ability to protect different biomolecules and therefore can be a
good candidate for development radioprotector.
Keywords: BSA, protein carbonyl, radioprotection, ferulic acid.
damages, organ dysfunction and systemic
failures.Oxidativedamagestobiomoleculeshave
been implicated in the pathogenesis of
variousdiseasessuchascancer,atherosclerosis,
Ionizingradiationgeneratesfreeradicalsbya
rheumatoid arthritis, infectious diseases and
variety of mechanism through radiolysis of
diabetes mellitus. It is believed that in many
water in cells and target all macromolecules
cases higher oxidative stress and occurrence of
DNA, proteins and lipids. As a result a plethora
disease are correlated (1, 2). Free radicals
of damages, collectively called as oxidative
mediatedoxidativedamagesofproteinsresultin
damages in these biomolecules occur.
their structural and functional impairment and
Accumulation of radiation induced such
increases proteolytic susceptibility (3).Oxidation
oxidative damages can lead to cell death, tissue
INTRODUCTION
Mishra et al. / Protective effect of ferulic acid on ...
ofaminoacidsinprotein,formationofcarbonyls
MATERIALSANDMETHODS
and hydroperoxides etc. lead to structural
changes, aggregation, cross linking and
Materials
fragmentation in proteins (4‐6). Therefore,
Bovine serum albumin (BSA, fraction V,
protein damage may implicate in initiation and
approximately 99%; protease free and
propagationofseveralpathologicalconditions.
essentially g‐Globumin free), Guanidine
Although many synthetic and natural
hydrochloride (GuHCl), trichloroacetic acid,
compounds
were
investigated
for
xylenol orange (o‐cresosulphononaphalene‐
radioprotection considering DNA as a critical
3,3‐bis‐(sodiummethyliminodiacetate)),
tris
biomolecules in living cells (7,8) but very few
(hydroxymethyl) aminomethane (TRIS),
studies were attempted for protein damage. acrylamide,
tetramethylethylenediamine
Several studies have suggested the use of (TEMED),
b‐mercaptoethanol,
coomassie
natural dietary antioxidants as therapeutic or
brilliant blue, bromophenol blue, 2, 4‐dinitro
prophylacticagentsagainstvariousdiseasesand
phenyl‐hydrazine (DNPH) and 5, 5’‐dithio‐bis
also as a countermeasure to radiation induced
(2‐nitrobenzoic acid), were purchased from
effectsonlivingsystem(7,9,10).
Sigma chemical company (USA). Ferulic acid
Ferulic acid (4‐ hydroxyl‐3‐ methoxy
was from Acros Organics. Methanol, sodium
cinnamic acid) (FA) is a natural phenolic
dihydrogen phosphate dihydrate, anhydrous
compound produced from metabolism of
disodiumhydrogenphosphate,sodiumchloride
amino acid (phenyl alanine and tyrosine)
(NaCl), hydrogen peroxide, ethanol, ethyl
throughshikimicpathway.Presenceofextended
acetate, butyrate hydroxyl toluene (BHT) and
conjugated aromatic structure along with
ferroussulphatewerepurchasedfromEMerck
electron donating ability of hydroxyl and
Germany. 2‐deoxy ribose was purchased from
methoxy substituted groups, carboxylic group
Himedia (Mumbai, India). Milli Q grade (18
with unsaturated C‐C double bond provides
Megaohm)waterwasusedforthepreparation
strongantioxidantpropertiestoferulicacid (11).
of different reagents. (Millipore Corp USA,
Ogiwara etal.reported that ferulic acid scaveng‐
modelElix3).
esfreeradicalsviz.,hydroxylradical,superoxide
and various oxygen centered and nitrogen cen‐
Samplepreparation
tered radicals (12). Khanduja et al reported
BSA stock solutions (2mg/ml and 10 mg/ml)
antiapoptotic potential of ferulic acid in normal
were prepared in 0.1M phosphate buffer (pH
humanperipheralbloodmono nuclearcells (13).
7.6) and concentrations were quanti ied
In murine peripheral blood leukocytes both in
spectrophotometrically at 278 nm using
pre‐irradiation and post radiation administra‐
extinction coef icient, e1% 6.8. Stock solution of
tion of ferulic acid lowers gamma irradiation
ferulicacid(10mM)waspreparedinphosphate
induced dicentric aberration on bone marrow
bufferofpH7.6.Themolarextinctioncoef icient
and lipid peroxidation in cultured lymphocytes
offerulicacidinphosphatebufferat310nmwas
(14‐18). Therefore, above mentioned studies
determined
as
18000
M‐1cm‐1 (20)
suggested the potential of ferulic acid as
and used for
spectrophotometrically
therapeutic agent againstvariousailmentssuch
estimation of concentration. The working
asdiabetes,cancer,alzheimerandparkinson (19)
concentrations of ferulic acid were prepared by
aswellasinradioprotection.
furtherdilutionin0.1Mphosphatebuffer.
The aim of the present study was to
Biochemical assays viz., carbonyl and
investigatetheradioprotectiveef icacyofferulic
thiol determination, the working concentration
acidagainstgammaradiationinducedoxidative
of BSA was 5 mg/ml while for hydroperoxide
damage of bovine serum albumin (BSA) using
assay it was 1 mg/ml. The samples were
SDS PAGE and biochemical assays viz., 2,
prepared by mixing BSA (10mg/ml and
4‐dinitrophenylhydrazine (DNPH) 5, 5’‐Dithio‐
2mg/ml) and phosphate buffer (pH 7.6) in the
ratio1:1(v/v).Ferulicacidworkingsolutions20
bis(2‐nitrobenzoicacid(DTNB)andFOXassays.
Int. J. Radiat. Res., Vol. 12 No. 2, April 2014
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Mishra et al. / Protective effect of ferulic acid on ...
µMand200µMwerepreparedandmixedin1:1
used to quantify the level of protein carbonyls.
volumeratiowithBSAtoobtain inalconcentra‐
Protein of the samples was quanti ied by
tionsof10µMand100µMrespectively.
measuringabsorbanceat278nm.Theamountof
protein carbonyl groups were expressed in
Gammairradiation
μmolescarbonylpermgofBSA.
BSA solution was prepared in borosilicate
glassvialsandirradiatedatdifferentdosesupto
DTNBassay
toamaximumof1200Gy(doserate,1.4kGy/h)
The cystein/thiol content of proteins was
using 60Co gamma ray irradiator, Model GC
determined by Ellman’s reagent as described
5000, BRIT, Mumbai, India,). Protein samples
earlier (22). Brie ly 2 ml of each sample was
mixed with 100 μl of DTNB (5mM) and
containing10µMand100µMferulicacidwere
incubated for 30 minutes at room temperature.
irradiatedat400Gyand1000Gy.
Absorbance of each sample was measured at
412 nm against blank. Concentration of thiols
SDS‐PAGE
was determined using extinction co‐ef icient of
SDS‐PAGE was performed according to the
method of Laemmli (21). Equal volume of the 13600M‐1cm‐1.
protein samples (1mg/ml) were loaded in each
lane, resolved on a 12 % separation and 5%
FOXassay
stacking gel and stained with Coomassie
Protein hydroperoxides of the samples were
Brilliant Blue. Image was taken using Gel DOC
determinedbyFOXreagentasdescribedearlier
(22). Fox reagents were prepared by mixing
system (Model GelDoc XR, Biorad, USA). Bands
acidi ied ferrous sulphate (5 mM), water and
were quantitated by intensity densitometry
xylene orange (5 mM) (1:2:2). Brie ly, 200 μl of
using (quantity one) software. The D0 doses
were calculated from the semi log plot of
each sample was mixed with 100 μl of 10%
radiation dose versus fraction of BSA remained
trichloroaceticacid(TCA).Sampleswerekeptat
intact. The equation for linear regression
ice temperature for 30 minutes. The samples
wasy=ae ‐bx.whereyisequaltothefractionof
werestoredat0°Covernight.Sampleswerethen
BSA remain intact a and b are intercept and
centrifugedat10000rpmfor5minutes.Protein
slope respectively. D0 dose is de ined as dose
pellets were washed thrice with 3 mM BHT
responsiblefor37%damageinBSAofitsinitial
(prepared in methanol). Samples were then
concentration.(8).
dissolved in 500 μl of guanidium hydrochloride
(6M).25μlofabovementionedfoxreagentwas
Proteincarbonylationassay added to the sample and left for 30 minutes at
Carbonyl content of protein was analysed
roomtemperature.Absorbancewasmeasuredat
spectrophotometrically as described elsewhere
560 nm against blank containing guanidium
(22).Brie ly 200μlof eachsamplewasincubated
hydrochloride. The concentrations of the
with200μlof10mMofDNPH(preparedin2N
hydroperoxides equivalent to the hydrogen
HCl) and incubated in dark for 1hour at room
peroxide were determined using the standard
temperature and vortexed after every 10
plotofhydrogenperoxide(0‐50μM)preparedin
minutes. The samples were precipitated by ice
guanidiumhydrochloride(22).
cold 20 % solution of TCA and centrifuges at
Determinationofhydroxylradicalscavenging
10000 rpm for 10 minutes. The pellets were
byferulicacid
washed three times with ethanol/ethyl acetate
Hydroxyl radical estimation was performed
(1/1; v/v) to remove excess of DNPH reagent.
using 2‐deoxyriobose assay. 2 mM solution of
The resulting protein pellet was dissolved in
2‐deoxyribosewaspreparedin0.1Mphosphate
600 µl of 6M GuHCl. Samples were mixed and
buffer (pH 7.4). Different concentrations of
theabsorbancewasmeasuredat370nmagainst
ferulic acid10μM and 100μM were mixed and
blank (GuHCl) as reference. The molar
solutions were irradiated at 400 and 1000Gy. 1
absorption coef icient of 22,000 M‐1cm‐1 was
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Int. J. Radiat. Res., Vol. 12 No. 2, April 2014
Mishra et al. / Protective effect of ferulic acid on ...
ml each of TBA (1% prepared in 0.5 N NaOH)
andTCA(10%)wasaddedto1mlofthesample.
The samples were heated at 95°C for 30 min in
water bath. The samples were cooled to room
temperature and absorbance was measured at
531 nm. Extinction co‐ef icient of MDA (1.56
×105 M‐1cm‐1) was used to calculate the MDA
concentrationineachsample(10).
Statisticalanalysis
Origin 7.0 statistical software was used for
statisticalanalysis.Allthevalueswerereported
as mean ± SD of three or more experiments.
These mean values were compared using ONE
WAY ANOVA using tukey test and signi icant
valuesof<0.05wasconsideredsigni icant.
induced BSA degradation by scavenging free
radicals as evident from SDS‐PAGE by the
presence of intact band. The extent of
degradation is dependent on both the radiation
dose and concentration of protein (25). Cho et al
has showed that at 0.13 % and 0.41 % BSA
concentration, the radiation effect was
signi icant and clear aggregated pattern of
proteinbandsappeared(26).Inthepresentstudy,
1mg/ml BSA (0.1 %) the SDS pro ile shows
radiationeffectofFA.
Proteincarbonylation
Carbonylcontentofproteinwasmeasuredusing
spectrophotometer in all sample by
dinitrophenylhydrazine
(DNPH)
labelling
( igure 2). Radiation enhances the oxidation of
proteins and causes formation of carbonyl
groups. Protein carbonylation decreased in the
RESULTSANDDISCUSSION
presence of ferulic acid in irradiated solutions.
GelElectrophoresisofBSA
Gamma irradiation of BSA at 400 Gy increased
SDS‐PAGEisasensitivemethodandhasbeen
the carbonyl content from 1.05 µmol CO/mg to
used to monitor gamma irradiation mediated
2.5 µmol CO/mg of BSA while at 1000 Gy
protein damage. Gamma irradiation may cause
carbonylcontentincreasedfurtherto4.21µmol
breakage of the polypeptide chains through
CO/mg of BSA .In presence of ferulic acid (10
oxidative damage of amino acids in peptide
µM) protein carbonyl formation decreased to
chainsandproteindamagecanbevisualizedby
2.13 µmol CO/mg in samples irradiated at 400
the formation of fragmented bands and
Gy and to 2.76 µmol CO/mg in samples
smearing of gel (23, 24). Fig1a shows radiation
irradiatedat1000Gy.Similarlyferulicacid(100
induceddamageofBSAupto1200GyusingSDS
µM)decreasedthecarbonylcontentto2.4and2
PAGE.Densitometryofbandsindicateslinearity
µmol CO/mg of BSA at 400 Gy and 1000 Gy
in the loss of protein with increasing dose of respectively.
radiation.Radiationdosesof1000Gyand1200
Gamma‐irradiation caused increase in the
Gy caused about 60 % damage in protein
carbonylcontentofBSA,(27)inirradiatedsample
( igure1a).TheD0dosefortheBSAdamagewas
carbonylcontentincreasedupto140and300%
calculated from the slope of semi log plot and
at 400 and 1000 Gy respectively. Ferulic acid
the value obtained as 1250 Gy ( igure 1b). The
100 µM decreased carbonyls level to almost
SDSpro ilein igure1cshowedprotectiveeffect
un‐irradiated control . While this decrease was
offerulicacidattwoconcentrations(10and100
relativelylessat10µMFA,viz.,120and160%at
μM) against gamma radiation doses of 400 and
400 Gy and 1000 Gy respectively. Decrease in
1000Gy.Therefore,itisinferredthatferulicacid
carbonyl content suggesting radioprotective
hasabilitytoreduceradiationinducedprotein
effectsofFA.
degradation/loss.
SDS‐PAGEpro ileofirradiatedBSAwasfound
DTNBassay
to be in agreement with reported observations
BSA contains thiol groups in the form of
(24, 25). Radiation causes breakage of covalent
cysteineresidues;oxidationofthesethiolgroups
bonds, the effect of which is manifested in the
is used as the indicator of protein modi ication.
SDS pro ile by the gradual disappearance of DTNBreactswithreducedthiolsresultinginfor‐
major band. Ferulic acid prevents the radiation
mation of 5‐thio‐2‐nitrobenzoic acid which is
Int. J. Radiat. Res., Vol. 12 No. 2, April 2014
116
Mishra et al. / Protective effect of ferulic acid on ...
a
C
b
d
Figure 1. Representa ve SDS‐PAGE profile of BSA irradiated
at different dose of radia on with or without ferulic acid. The
concentra on of BSA was 1mg/ml. a) PAGE profile of BSA
irradiated at different doses without ferulic acid. Lane 1 to 7
represents radia on dose 0, 200, 400, 600, 800, 1000 and
1200 Gy respec vely.
b) Semilog plot of densitometric data obtained from figure
1a.
c) Representa ve gel image of BSA irradiated at 400 and
1000 Gy in presence of ferulic acid10 μM and 100 μM respec‐
vely. The lane descrip ons are: Lane1, BSA (control); lane 2,
BSA+ ferulic acid (10 μM); lane 3, BSA + ferulic acid (100 μM);
lane 4, BSA+ 400 Gy; lane 5, BSA+ ferulic acid (10μM) +
400Gy; lane 6, BSA+ ferulic acid (100μM)+ 400Gy ; lane 7,
BSA + 1000 Gy ; lane 8, BSA+ ferulic acid (10μM) + 1000Gy ;
lane 9, BSA+ ferulic acid (100μM) + 1000Gy. M denotes mo‐
lecular weight markers (Medium range protein marker, Ban‐
galore Genie, India).
d) Rela ve quan ta on of remaining BSA in each well using
gel image shown in figure 1c.
quanti ied at 412 nm spectrophotomerically.
Figure 3showsthechangesinthethiolcontent
FOXassay
ofirradiatedandcombination(irradiationwith
FOX assay was performed to assess the
different concentrations ferulic acid) samples.
radiation induced protein peroxide generation.
Thethiolcontentofproteindecreasedfrom14.7
Carbon centered radicals of protein participate
± 1 μM to 1.5 ± 0.3 µM in irradiated solution.
in the formation of peroxyl radical upon
Ferulicacidat10µMand100µMprotectedthiol
irradiation. Onceperoxylradicalisformedfrom
contentofBSAfromradiationinduceddamages
sidechainaminoacid,itpropagatesfurtherand
in a concentration dependent manner and
damages other amino acids resulting in
protectedthelevelofthiolsupto3±0.2µMand
formation of hydroperoxides. Figure 4 shows
4.1±0.4µMrespectively.Radiationdose(1000
increase in the yield of protein hydroperoxides
Gy) did not further decrease the thiol content.
when BSA solution was irradiated at 400 and
Ferulic acid on the other hand protected thiol
1000 Gy. Radiation dose of 400 Gy produced
damageevenat1000Gy.
about 4.5 µM of hydroperoxides. Ferulic acid
Itisreportedthatthiollosscanbecorrelated
decreased the production of hydroperoxides to
(27)
to loss of activity of protein . In the present
3.7 and 3.3 µM at 10 µM and 100 µM
study, ferulic acid provided signi icant
respectively. Radiation dose of 1000 Gy
protection against radiation induced oxidation
produced around 6 µM of hydroperoxides.
of thiols groups in a concentration dependentFerulic acid decreased the production of
manner.
hydroperoxidesto5.4 and4.3µM at10μMand
117
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Mishra et al. / Protective effect of ferulic acid on ...
100µMrespectively.Ferulicaciddecreasedthe
formation of hydroperoxides in concentration
dependent manner signifying the role of FA as
potentialantioxidant.
Figure 2. Irradia on (400 Gy and 100 Gy) induced changes in the carbonyl forma on of BSA. Two different concentra on 10
μM and 100 μM of ferulic acid were used. ferulic acid decreased the radia on induced carbonyl forma on in
concentra on dependent manner. For details see material and methods. The data is the mean ± SD of three independent
experiments. *control vs radia on, # radia on vs ferulic acid treated group. The data was significant at the p < 0.05.
Figure 3. Irradia on (400 Gy and 1000 Gy) induced loss of protein thiols. Two different concentra on of ferulic acid, 10 and
100 μM were used. Ionizing radia on caused decrease in the thiols forma on. Ferulic acid protected the loss of thiole moie es.
The data is the mean ± SD of three independent experiments. *control vs radia on, # radia on vs ferulic acid treated group.
The data was significant at the p value of < 0.05.
Int. J. Radiat. Res., Vol. 12 No. 2, April 2014
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Mishra et al. / Protective effect of ferulic acid on ...
Figure 4. Irradia on (400 Gy and 1000 Gy) induced forma on of protein hydroperoxides. All other experimental details were
similar to figure 5. The data is the mean ± SD of three independent experiments. *control vs radia on, # radia on vs ferulic acid
treated group. The data was significant at the p value of <0.05
Hydroxylradicalscavengingassay
proteinbackboneatthesiteofa‐carbonperoxyl
Hydroxyl radical (˙OH) scavenging potential
radicals via amidation pathway or through
offerulicacidwasinvestigatedbyusing2‐deoxy
oxidation of glutamine side chains, resulting in
ribose degradation assay ( igure 5). Hydroxyl
the formation of a peptide in which the
radicals generated by radiolysis of water by
N‐terminal amino acids is blocked by an
ionizingradiationistheprimarydamagingagent
α‐ketoacyl derivative (27, 28). Similarly protein
peroxide and breakage of thiol bonds are also
forallbiomoleculesinlivingsystem.Antioxidant
related to reaction of OH radicals with protein,
scavenges various free radicals including
thereby inhibiting the OH radicals FA may
hydroxyl radicals. Ionizing radiation enhances
reduce irradiation mediated protein damage.
generationofmelonaldehyde(MDA)production
TheOHradicalscavengingdataissupplemented
in dose dependent manner. Radiation dose of
withouranotherrecentlyreportedstudywhere
400Gyand1000Gyproduced13μMand19μM
itwasobservedthatferulicacidscavengesDPPH
MDA respectively. Presence of ferulic acid
freeradicalquitesigni icantlywithEC50valueof
decreasedtheproductionofMDAto23%and54
22±2.2µM(29).
% at 10uM and 100uM respectively at 400 Gy.
Similarly, ferulic acid in decreased the MDA
Bindingofferulicacidwithserumprotein
concentrationto18and11μMat10μMand100
Besides radical scavenging, binding of
μMrespectivelyinBSAirradiatedat1000Gy.
antioxidantwithproteinmayaffectthestability
Hydroxyl radical (‐OH)‐mediated protein
oxidationhasreceivedmostattentionduetoits
ofprotein.Itisreportedinourearlierstudythat
importance as hydroxyl radicals are highly
ferulic acid binds non‐covalently to bovine
reactive free radicals to protein. Hydroxyl
serumalbuminwithbindingconstant(Ka)40.15
± 0.02 x104 M (30). Hydrogen bonding and
radical upon reacting with protein may lead to
electrostatic forces play a vital role in
formation of reactive carbonyl groups.
stabilization of ferulic acid‐BSA complex. It was
Tryptophan, histidine and cystein are most
furtherobservedthroughcirculardichroismand
susceptible to the hydroxyl radical mediated
thermal melting study that in the presence of
damage. Protein carbonyl derivatives may also
ferulic acid melting temperature (Tm) of BSA
be generated through oxidative cleavage of
119
Int. J. Radiat. Res., Vol. 12 No. 2, April 2014
Mishra et al. / Protective effect of ferulic acid on ...
dependingonthepropertiesoftheantioxidant.
increasedfrom341KfornativeBSAto348Kin
the presence of ferulic acid (10 µM). Therefore,
ferulic acid on binding with BSA increases the
proteinstability.SimilarlyKapooretal. (31)have
CONCLUSION
reported that non‐covalent binding of another
antioxidant curcumin with bovine serum
Role of oxidized products of proteins as the
albumin has shown inhibition of radiation
source of causing toxicity or cell death in the
induced protein damage. Since ferulic acid also
system has been implicated. Further this study
interacts with BSA, binding of ferulic acid with
provides insight into the ferulic acid mediated
BSA may be implicated in case of ferulic acid
radioprotectionandantiradicalef icacyofferulic
mediated radioprotection. Salvi et al. (32)
acid.Thisstudysuggestedthatbothef icientROS
reportedstructuraldamagetoproteinssuchas
scavenging ability and binding interaction with
lysozyme, human serum albumin (HSA) and
BSAplayedanimportantroleintheradioprotec‐
beta‐lactoglobulin A induced by free radicals.
tive ef icacy of ferulic acid. The results can help
Authors have reported that out of four
indesigningtheapplicationsforferulicacidasa
well‐known antioxidants (quercetin, melatonin,
radiation protector even as for food
Trolox, and chlorogenic acid), melatonin and
preservation.
chlorogenic acid, which did not bind to any of
the three proteins under study, showed
scavenging and protective activities well
correlated with the amount of free radicals ACKNOWLEDEGEMENT
generated.Trolox,whichbindsonlytoHSA,was
K.
Mishra
is
thankful
to UGC, Delhi for the
a better protector of HSA than of the two other
senior
research
fellowship.
The work was funded
proteins, indicating that its antioxidant capacity
through a DRDO NBC project (1.29), Govt. of
is increased by a shielding effect. These results
India.
demonstrate that binding of an antioxidant to a
protein may potentiate protection or damage
Figure 5. Radia on induced genera on of hydroxyl radical and scavenging by ferulic acid. Ferulic acid decreased the
radia on induced genera on of hydroxyl radical in concentra on dependent manner. 2‐deoxyribose degrada on assay were
used to es mate the hydroxyl radical scavenging by ferulic acid. The data is the mean ± SD of three independent
experiments. *control vs radia on, # radia on vs ferulic acid treated group. The data was significant at the p < 0.05.
Int. J. Radiat. Res., Vol. 12 No. 2, April 2014
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