:
: :
in vivo in vitro
-I -II ..
..
..
..
..
..
..
..
..
...
..
..
..
Olea europaea.L Oleaceae
..
Olea europaea.L ..
..
..
..
..
Olea europaea.L ..
-III ..
Olea europaea L ..
. . ..
. .
..
HPLC / DAD ..
..
tanin ...
..
..
. . . Sephadex
..
. . . HPLC
..
HPLC .
..
..
HPLC/ DAD ..
..
tannin ..
..
(C1) ..
HPLC-RP ..
..
..
.
-IV in vitro
.1 .
in vitro ..
1.1 DPPH
..
..
in vitro Chemlal
.
.OH .
Fe2+
..
..
..
...
.
in vitro ..
DPPH ..
(FRAP) .
..
in vitro Chemlal
.
.OH ..
Fe2+
. . . .
-V in vivo
.
.1.
..
. . ..
. . ..
. . ..
. . Glutathion
.
. 4.
..
. . .
. .
..
.
Glutathion .
. . . .
-VI .
-VII .
-VIII .
-ALA Alpha lipoic acid - CB Carotene bleaching -BHT Butyated hydroxytoluene -BSA Bovine serum albumin -(CH3)2Hg Dimethyl mercury -C2H5Hg+ Ethyl mercury cation
-CAT Catalase
-CDNB 1-chloro-2,4-dinitrobenzene -Ce2SO4/H2SO4 Cerium sulphate in /sulphiric acid -CH3Hg+ Methyl mercury cation
-DMPS Dimaval Unithiol- Sodium 2,3 - dimercapto propane sulfate -DHLA Dihydro lipoic acid
-DPPH Radical 1,1-diphenyl-2-picrylhydrazyl -DTNB Dithiobis(2-nitro benzoic acid) -EDTA Ethylene diamine tetra acetic acid -ENos Endothelial nitric oxide synthase
Ferric Reducing Antioxidant Power -FRAP
-GAE Galic acid equivalent -GLC Glutamate Cysteine Ligase -GPx Glutathion peroxidase -GR Glutathion reductase
-GSH Glutathion
-GST Glutathion S-transferase -GTP Guanosine tri phosphate -H2O2 Hydrogen Peroxide
-HCl Chlorure d'hydrogène
-Hep Human hepatoma
-Hg0 Elemental mercury
-Hg2+ Inorganic mercury cation -HgCl2 Mercury chloride
-HO.2 Hydroperoxyl
-HO-1 Heme Oxygenase-1
-HOCl Hypochlorous acid
-HPLC-DAD Hight performance liquid chromatography DiodeArray Detector -HPLC-RP Hight performance liquid chromatography Reverse phase
-HT Hydroxytyrosol
IC50 Inhibitor concentration of 50%
-ICAM-1 Intracellular adhesion molecule-1 -INos Inducible Nitric oxide
- Ip Intra peritoneal
-KLF-4 Kruppel like Factor 4 -LDH Lactate dehydrogenase -LDL Low density lipoprotein
LLC-PK1 Epilhelial cell line porcine kidney
-LPS Lipopoly saccharide
-LOO. Lipid peroxyl
-MDA Malondialdehyde
-MeHg Methyl mercury
-MPO Myeloperoxidase
-MT Metallothioneins
-NBT Nitro blue tetrazolium -NF- Nuclear factor- kappa beta -NMR Nuclear magnetic resonance -NO2 Nitrogen dioxide
- Nox NADPH Oxidase
-3-NO2-Tyr Nitrotyrosine
-NQO1 Quinone oxidoreductase1
-NrF2 Transcription nuclear factor erythroid Related 2 -O2.-
Superoxide -.OH Hydroxyl radical -ONOO- Peroxynitrite
-ROS Reactive oxygen species
-RNA Ribonucleic acid
-RNS Reactive nitrogen species
-SGLT1 Sodium dependent glucose transporter
-SH Thiol
-SOD Superoxide dismutase -TBA Thiobarbeturic acid -TLC Tin layer chromatography
-TE Trolox equivalent
-TAE Tannic acid equivalent
-THP-1 Human Promyelocytic cell line -TNF Tumor nicrosis factor
-TPTZ Tripyridyl triazine
.I
et al
,2007)
2+ . Hg MeHg GSH
Zalups and Lash ,1996 .
(Mahboob et al .,2001 ;
Huang et al .,1996) .
- 2. 2 O
2O H ., 1993
et al Lund
O2
H2
in vivo
myeloperoxidase (MPO)
(ROS) HOCl
(RNS) ONOO
et al. , 2007)
( Rooney , 2007)
dimaval unithiol- sodium 2,3 - dimercapto propane
(DMPS) sulfate (Georget al ., 2004)
(Gregus et al.,1992) alpha lipoic acid (ALA) )
(Goyer et al.,
selenoprotein-P (Sasakura and
Suzuki ,1998) .(Ou et al ., 1999)
Risher and Amler,2005 El-Shenawy and
Hassan ,2008 Spirulina fusiform
phycocyanin (Sharmaet al ., 2007 ) Ocinum sanctum
.OH 2005)
et al ., (Sharma
Schisandrin B Schisandra chinensis
(Stacchiotti et al.,2009) (Jemai et al . ,2008)
(Bouallaguiet al . , 2011) (Alarcon De la lastra et al.,
2001) Cioffi et al .,2009
(Deiana et al ., 2007) Olea europaea L
oleuropein secoiridoids
hydroxytyrosol tyrosol
.(Vissers et al ., 2004) Olea europaeaL
Olea europaea. L
leuropein o
.,2002) et al
(Andrikopoulos
.OH ., 2001) et al
(Gordon ydroxytyrosol
h O2
H2
Tuck
et al ., 2001 Soni et al.,2006)
HOCl
(Visioli et al ., 1998) Aruoma and Halliwell ,1987)
Olea europaeaL Olea europaea L
Leccino Ogliarola
Maiatica
Frantoio Coratina
Chemlal Sigoise
Cilento
Sigoise Chemlal
in vitro Olea L
europaea
in vivo Olea europaeaL
.1
No l,
2008
Stacchiotti et
al ., 2003 (Prozialeck and Edwards,2007)
.
- Hg0
et al (Clarkson
., 2003)
2+ - Hg
(Ozuah , 2000) Hgo
- (Brawnwald et al.,2001) -
-
HgCl CH3
(Rabenstein ,1978) Hg+
CH3
%
HgNO3
CH3
,1969) et al
(Aberg
+2 % Hg
HgCl2
(Clarkson ,1997) HgCl2
Hg+
CH3
(Edward and
Mc Bride, 1975) Staphylo cocci
E. Coli (Rowland et al .,
1975) (Yasutake et al., 1990)
% . (Clarkson, 1972)
Hg+2
Hg+
CH3
et al (Rabenstein
,1989) .
2Hg (Cys)
2Hg ) SH (G ,1973) and Am
(Rabenstein
Hg+
CH3
.,1990) et al
; Yasutake (Ballatori ,1991
GSH benstein ,1978)
a (R
HgCl CH3
Hg+
CH3
(Yasutakeet al,1989)
N- acetylcystein
Hg+
CH3
(Ballatori ,1991
(Richardson and Murphy, 1975) (Zalups and Ahmad,2005) N acetylcystein
(Brawnwald et al ., 2001)
) ., 1974 et al
Bryan Hg+2
Hg0
., 2001) et al
; Brawnwald ,. 1996
et al (Optiz .
(Clarkson, 2002) .
GSH
Dutczak and Ballatori ,1994 ;Ballatori ,1991) .(
Huang et al .,1996 Lash et al .,1998
Lundet al., 1993 et al .,2007)
.HgCl2
HgCl2
; ., 1996 et al
(Huang
Mahboobet al., 2001 (Hamilton et al ., 1997
oleic linoleic
linolenic arachidonic
L LO LOO L
LO LOO Larock ,1985
R )
.,1995 et al
Madhavi (
: (Madhaviet al.,1995)
oleic acid LH
Hamiltonet al.,1997 ; Stohs and Baghi ,1995
R Hg X
2Hg R , 1976 Lapert and Lednor
Hg - C R
H LH
L
R
2Hg R RHgX
LOO
Stohs and Baghi ,1995; Hamilton et al.,1997 (
Perottoni et al., 2004 .
HgCl2
.,1998 et al
Lash Hg2+
.
Zalups and Lash ,1994
Hg2+
) .,1985 et al
Berndt Zalups and Lash ,1997;
Hg2+
GSH
GSH GSH
Hg2+
(Lash and Zalups ,1996) GSH
Hayes and McLellah,1999
Hg(II) et al.,2007
Zalups and Lash ,1994
: Hayes and McLellah,1999
) Farina et al, (2013 Lund et al., (1993
Hg(II)
O2
H2
., 1991 et al
Lund
-2 O.
O2
H2
NO Yapet al.,2009
2 :
-
O.
O2
H2 .NO .,2009
et al Yap (
O2
H2
ONOO-
GSH
GSSG Yapet al.,2009
O2
H2
ONOO-
.,2009 et al
Yap
ICAM-1 integrins
selectins
(Go and Jones ,2011)
(Go and Jones ,2011)
Ksp-cadherin (Jiang et al .,2004)
ICAM-1 P-selectin
;Takada et al., 1993 (Kelly and Singer , 1993
Prozialeck and Edwards, 2007)
ICAM-1 Tipping and Timoshanko, 2005
myeloperoxidase ROS
HOCl
O2
H2
OH
- RNS ONOO .,2004)
et al (Liddell
ener et al.,2007
RNS (Liddell et al.,2004) ROS
NAD(P)H oxidase xanthine oxidase
·O2
O2
H2
O2
et al SOD Tamara
.,2008 HOCl
MPO (Liddellet al.,2004)
- 2.
O .,2008 et al
Tamara
E
Sharma et al, (2007) Spirulina fusiform
HgCl2
-carotene vitamin E LOO.
GPx selenodiglutathione
selenocysteine
dimethyl selenide
Sharma et al .,(2005) orientin
vicenin
Ocimum sanctum
.OH
Ocimum quercetin
O2
H2
.,2011 et al
Lagoa
O2
H2
.,2011 et al
Lagoa
B schisandrin Schisandra chinensis
B schisandrin
(Stacchiotti et al ., 2009) in vivo B
schisandrin GSH
.(Chiu et al.,2005 ;Chen and Ko,2010 (Stacchiotti et al .,2011)
schisandrin B (Augusti et al., 2008)
astaxanthin GPx Catalase
SOD staxanthin
a
.OH
Wu et al., 2006 ; Bernstein et al ., 2001 Goto et al
.,2001 astaxanthin
II Cu Fe III
Zhaoet al., 2005 vitamin E
in vitro in vivo
Valko et al .,2005 .
tocopheroxyl -tocopherol
C
Arita et al., 1998 Agarwal et al, (2010)
E
Olea europaea.L Oleaceae
Olea europaea .L Oleaceae
Scognamiglio et al ., 2012
Olea europaea .L
(Girre ., 2001) oleic
shikimate
phenyl propanoid Ryan and Robards, 1998
hydroxytyrosol tyrosol
secoiridoids
oleuropein (Granados-Principal et al.,2010
Olea europaea.L Oleaceae
(Granados-Principalet al.,2010
Olea europaea .L Olea europaea.L
(Bouaziz and Sayadi,2005) -
Oleuropeosides Verbascoside- Oleuropein
Flavonol- rutin
Catechin Flavan-3ols Tyrosol Hydroxytyrosol Vanillin Caffeic acid
(Fkiet al., 2005) Luteolin 7-O-glucoside Quercetin 3-O-glucoside Rutin
Quercetin Lutoelin Apigenin Chrysoeriol Oleuropein Hydroxytyrosol
(Rioset al., 2005 Hydroxytyrosol
tyrosol vanillic acid
vanillin p-coumaric
acid Ferulic acid 4Ethyl phenol
tyrosol Hydroxytyrosol acetate
Flavonoids Apegenin
Luteolin (Breneset al.,1999)
Secoiridoids- Oleuropein glucoside
- +pinoresinol +1-acetoxy pinoresinol
Tanet al., 2003 ; Ruiz Gutierrez et al., 2000 Bazotiet al., 2005
.
oleuropein glucosidase
oleuropein Corona et al .,2006
oleuropein
hydroxytyrosol metiloleoside
lipases Carrera-Gonzàlez et al ., 2013
oleuropein (Carrera-Gonzàlezet al., 2013
oleuropein oleuropein SGLT1
oleuropein
Edgecombe et al., 2000 .
hydroxytyrosol tyrosol
Mannaet al,(2000) Caco
2 hydroxytyrosol
. Corona et al .,(2006
hydroxytyrosol tyrosol
Caco 2
in vitro
Piskula and Terdo , 1998 .
(Miro casas et al . , 2003 )
in vitro
Hep G2
hydroxytyrosol hydroxytyrosol acetate
hydroxytyrosol tyrosol
>
%
Mateoset al.,2005 Caco - 2 hydroxytyrosol
O methylated cathecol O methyl transferase
Mannaet al.,2000
hydroxytyrosol GSH
Corona et al
.,2006
) ( Del Boccio et al , oleuropein
hydroxytyrosol oleuropein
oleuropein hydroxytyrosol
oleuropein hydroxytyrosol
Del Boccio , 2003
% hydroxytyrosol
C)
%
hydroxytyrosol sulfate conjugate homovanillic acid
homovanillic alcohol 3,4 dinydroxypheny aceticacid
3,4 dihydroxy phenyl
acetaldehyde tyrosol
(Tuck and Hayball , 2002)
(De la Torre Carbot et al.,2006)
hydroxytyrosol monoglucuronide hydroxytyrosol mono
sulfate tyrosol glucuronide
tyrosol sulfate homovanilicacid sulfate
LDL
(De la torre carbot et al . ,2006) .
hydroxytyrosol homovanilic alcohol
aldehyde
alcohol dehydrognase et al .,2001)
% hydroxytyrosol
hydroxytyrosol (Miro-Casas et al .,2003)
Visserset al, ( 2004) hydroxytyrosol
hydroxytyrosol
Olea europaea.L Olea europaea.L
2006 Farag et al,
ppm Kronaku
Olea europaea .L in vivo
in vitro
Mahmoudi et al., (2015) Chemlali
oleuropein hydroxytyrosol
catalase
MDA bisphenol A
-tocopherol
ferric nitrolo acetate
Deianaet al., 2007 Deiana et al, (2008
hydroxytyrosol homovanilic alcohol
PK1 -
2 LLC
2O H .
hydroxytyrosol glucuronides hydroxytyrosol hydroperoxides
7- ketocholesterol
ydroperoxides h
PK1 -
2 LLC
2O H Deiana
et al., 2011) Olea europaea .L
LDL
Olea europaea .L
LDL
intima LDL hydroxytyrosol
(Soniet al., 2006) ydroxytyrosol
2 h B hromboxane t
in vitro (Petroniet al.,1997 )
hydroxytyrosol eicosanoid
5- 12-
lipooxygenase Visioliet al, 2005)
(
ydroxytyrosol h
E2
rostaglandin p
nitric oxide synthase cyclooxygenase
J774 (Maiuriet al., 2005)
hydroxytyrosol vibroparahaemolyticus Vibrio cholerae
Salamonelle
typhi Haemophilus influenzae
Staphylococais aureus
(Bisignanoet al., 1999) Escherichia coli Candida albicans
kluyreromyces
marxianus Clostridium perfringens
Streptococcus mutans
Shigella sonnei Salmonella enterica
(Medina et al.,2006)
hydroxytyrosol
HL60 promyelocytic leukemia
HT29 colon
adenocarcinoma ( Della Ragioneet al, 2000)
lydroxytyrosol G1
(Fabianiet al., 2002) HL 60 (Visioliet al. , 2002)MPO
oleuropein hydroxytyrosol
Olea europaea L Olea europaea L
Coratina
Frantoio Leccino
Matiatica Ogliarala
Chemlal Sigoise
Olea europaea Chemlal
Sigoise
Cilento . . Quezel and Santa ,1963
Spermatophyte Embranchement
Angiosperme Sous embranchement
Dicotylédones Classe
Oléacées Famille
Olea(L) Genre
Olea europaeaL Espèce
Olea europaea L
lyophilisateur
ethyl acetate ethyl acetate
rpm
acetonitrile / n hexane Olea europaea RP-HPLC
DAD
(Cioffiet al.,2010) Acetonitrile ethyl acetate
Ethyl acetate
Acetonitrile n-hexane
Acetonitrile
5ml Methanol, Sonification Centrifugation 3000rpm/10min
Azote Dream 5ml Acetonitrile+ ml5 n-hexane
Sonification, Centrifugation 3000rpm/10min
Acetonitrile n-hexane
Ethyl acetate
Ethyl acetate
HPLC / DAD acetonitrile RP Agilent1200series HPLC
Agilent
Technologies, paloAlto,CA,USA) (G-1312A)
(G1329A) (DAD) 1315
OnYX monolithic(50x2 m m
C18, phenomenex , Italy) (A)
(Millipore, Bedford, MA, USA)
%0.1 TFA (B)
B % 0 B %30-0
B % 70-30 B % 90-70
B % 90 HPLC
278 nm .
Folin-Ciocalteu Condelliet al., (2015)
Folin-
Ciocalteu
%10 w/v
gallic acid
gallic acid Tanin
tanin McArt et al .,2006
BSA
% SDS
Triethanolamine
% ferric chloride
tannic acid
Cilento
Sigoise Chemlal
(Cioffi et al.,2010) EE
Ec EM
Cilento Sigoise
Chemlal
EE
Ec 29.40
4
EM
. . . Sephadex
Sigoise Chemlal
rpm
3 cm x 100 cm Sephadex
LH-20 25-100 pm,
pharmacia fine chemicals chemicals P1)
Pharmacia Fine Sigoise
Chemlal Silice F254)
(gel 60
: 25: 15) acetique (60
acid /
2O H / Butamol n
Petrolium ether
ChloroformX3
Petrolium etherX3
Methanol X3
° Sigoise
Chemlal C1
: Sigoise
:2) :18 (80
2O -H -MeOH CHCl3
: Sigoise
acid acetique O-
H2
nol- a nBut
8 8
: Chemlal
:2) :18 (80
2O -H -MeOH CHCl3
: Chemlal
acid acetique O-
H2
nol- a nBut
. . . HPLC
Shimadzu RID-10A Shimadzu LC-20AT promenence)
U6K Shimadzu RID-10A
(30cm x 7.8 mm) Water C-
Chemlal rpm
: 25: 15) acetique (60
acid /
2O H / Butamol : 2) n
: 18 O (80 H2
/ MeOH
3/ CHCl
C2
HPLC C2 S1
HPLC RP
Agilent1200series Agilent Technologies, paloAlto,CA,USA) binary (G-1312A) Auto sampler(G1329A)
(DAD) 1315 Diode Array
detectot OnYX monolithic(50x2 m m C18, phenomenex , Italy)
(Millipore, Bedford, MA, USA)Acidified milli Q Water (A)
%0.1 TFA (B)
B 0%
B %30-0 B %
70-30 B % 90-70
B % 90
. nm 278
. . . Chemlal
Folin-
Ciolteu Gallic acid
Singletonet al.,1999)
HPLC/ DAD RP
HPLC DAD
p-hydroxy benzoic acid Vanillic acid
Caffeic acid Gallic acid
Syringic acid p-Coumaric
acid Ferulic acid
Sinapic acid
Chrysoeriol Luteolin Hydroxytyrosol Tyrosol
Secoiridoids Oleuropein
Caffoyl glycoside phenylethanoid Verbascoside
gallic acid
Sigoise Frantoio
Leccino
Chemlal gallic acid
luteolin oleuropein
Leccino luteolin
Frantoio oleuropein
hydroxytyrosol Sigoise
vanillic acid Leccino
p-hydroxybenzoic acid
Chemlal Coratina
gallic acid hydroxytyrosol
luteolin
oleuropein verbascoside
6.94 GAE
Sigoise Coratina
Chemlal GAE
tannin tannin TAE
Leccino tannin
TAE
Chemlal TAE
tannin Sigoise
TAE
tannin Olea
europaea Dekdouk et al .,2015
Standard tannin
gallic acid tannic acid
(C1)
max 365=
)1 (
ppm
1H ppm 6.46=
H3
3 H
1H ppm 6.23=
H
A 1H
ppm 6.04=
H
A A
1H
= ppm
B
z) (J=8H 1H
= ppm
H B
- )
(J=8Hz
1H 7.28=
ppm -
B
1H) - (RMN d4
- MeOD (Hz)
(ppm)
H
luteolin
luteolin
= ppm
Hz) (J= 7.5
= ppm
H
Wood
I
= 336 nm
max
NaOH MeOH
I
OH
320-335nm OH
C-7
NaOAc MeOH
II
AlCl3
MeOH I
HCl AICI3
+ HCl AlCl3
38 nm -
B dihydroxy + HCl) (AICI3
MeOH I
49 nm
OH C-5
C
nm
II I
MeOH NaOH
AICI3
HCI /AICI3
NaOAc BO3 / NaOAc H3
C
Rf
Rf
) C1 (
luteolin
luteoline
HPLC-RP C2
S1
hydroxytyrosol oleuropein
leuropein o
tR
9.1= C2
leuropein o
ydroxytyrosol h
tR
= 0.7 S1
ydroxytyrosol h
oleuropein hydroxytyrosol
HPLC
tR
0.7= tR
9.1= ydroxytyrosol
h
oleuropein Cilento
a Sigoise b
Chemlal c
: HPLC-RP Cilento
a Sigoise b
Chemlal
a
b
c 1
2
2
1
Chemlal Folin-
Ciocalteu GAE
Olea europaea L Coratina
Frantoio Leccino
Matiatica Ogliarala
Chemlal
Sigoise RP-HPLC
DAD
. p-hydroxy benzoic acid
vanillic acid caffeic acid
gallic acid syringic acid
p-coumaric acid ferulic
acid sinapic acid
chrysoeriol luteolin
hydroxytyrosol tyrosol
secoiridoids
oleuropein Caffoyl glycoside phenylethanoid
verbascoside
hydroxytyrosol Sigoise oleuropein
glucosidase Briante et al.,
2002 hydroxytyrosol Auroraet al,2008
(Brahmi et al ., 2013 ; Bouaziz et al .,2006)
hydroxytyrosol chemlali
neb jmel
gallic acid hydroxytyrosol
luteolin oleuropein
verbascoside
Chemlali (Brahmiet al .,2013) neb jmel
tannin tannin
Leccino
Chemlal tannin
Sigoise
(Brahmi et al., 2013) Chemlali neb jmel
Joanaet al ., 2010
Sigoise Chemlal
secoiridoids
oleuropein hydroxytyrosol
luteolin
Chemlal (Bouaziz et al .,2005)
.Chemlali luteolin 7-
O-glucoside luteolin 7-O-rutinoside
apigenin7-O-glycoside rutin
luteolin
apigenin oleuropein
(Bouaziz et al
.,2005) Savournin et al .,2001
.
2000 Garciaet al., luteolin
apigenin 7-O-glucoside diosmetin
7-O-glucoside diosmetin
rutin flavan-3-ols
catechin
Olea europaea Villalonga
Alfafarenca Picual
Cornicarba
Blanqueta
Olea europaea L
Coratina Frantoio
Leccino Matiatica
Ogliarala
Chemlal Sigoise
. p-hydroxy benzoic acid
vanillic acid
caffeic acid gallic acid
syringic acid p-coumaric acid
ferulic acid sinapic
acid chrysoeriol
luteolin hydroxytyrosol
tyrosol secoiridoids
oleuropein verbascoside
gallic
acid Sigoise
Chemlal
hydroxytyrosol Sigoise
Chemlal tannin
Leccino Chemlal
tannin Sigoise
Sigoise Chemlal
oleuropein hydroxytyrosol
luteolin Chemlal
Chemlal tannin
hydroxytyrosol Sigoise
Chemlal
oleuropein Chemlal
in vitro
.OH Fe2+
.1 in vitro
1.1 DPPH
DPPH (Millelaet al., 2014)
DPPH
DPPH
Trolox Trolox
TE
(FRAP) FRAP
Russo et al, 2015
FRAP 300 mM
Acetate pH
20
mM O
6H2
FeCl3
mM TPTZ
40 mM HCl
FRAP tripyridyl °
triazine
3+- Fe Ferrous
tripyridyl triazine
2+- Fe
Trolox FRAP
Trolox
mg TE
(Padula et al.,2013) -carotene/linoleic acid
-carotene linoleic acid
Tween
° BHT -carotene
(AB-carotene after 180 min/Ainitial B-carotene)x100 (AA%)
Pearson
in vitro Chemlal
.OH
.OH FeSO4
O2
H2
OH
OH alicylate
2 s
2O H
4 6mM FeSO
1.5mM salicylate
mM
alicylate s
OH
OH Dose
- Log cid ascorbic
a
Smirnoff and Cumbes,1989 .
Fe2+
FeCl2
mM
errozine f
+ 5mM) Fe2
errozine f
Fe2+
Dose - Log EDTA
Huset al., 2006 .
1 - THP RPMI 1640
- 2Mml
glutamine streptomycine 0.1mg/ml
penicillin 100IU/ml mercaptoitanol
50µM 10%
°
CO2
% -
TNF cells
0.5x106
. Chemlal
Cytokine cytokine
° TNF-
ELISA KIT (Endogen,Rock,Ford ,IL ,USA)
pg/ml
LDH
LDH LDH
(Sigma) KIT
%
Pearson Coefficient P
(Graph Pad Prism 5 soft ware Sandiego, CA, USA in vitro
in vitro DPPH DPPH
DPPH
DPPH-H (P<0.05)
Coratina
Chemlal TE
. Sigoise
TE
TE n=3
:D,C,B,A) (P<0.05
(FRAP) Olea europaea Fe3+
Frantoio (P<0.05)
TE Sigoise
0 50 100 150 200 250
Chemlal Coratina Frantoio Leccino Maiatica Ogliarola Sigoise B
A
BC C
D C
D mg TE/g Extrait
(P<0.05) TE
Coratina
Chemlal Maiatica
(P<0.05) TE
TE TE
FRAP TE
n=3
,C,B,A) (P<0.05
%
% Ogliarola
n=3 (P<0.05) n=3
B,A) (P<0.05
0 50 100 150 200 250
Chemlal Ogliarola
0 5 10 15 20 25 30 35 40
Chemlal Coratina Frantoio Leccino Maiatica Ogliarola Sigoise
A A
A A
B
C
B B B
B B
A
% B
DPPH r=0.91
(r=0.61)
tannin DPPH
r=0.68
r= 0.63 tannin
Pearson Pearson p-hydroxy benzoic acid
verbascoside
r=0.93 r=0.52
tyrosol r=0.61
hydroxytyrosol
r=0.54 r=0.74
r=0.52 in vitro Chemlal
.OH Chemlal
.OH
IC50
±208.33 IC50
-2 -1.5 -1 -0.5 0 0.5 1 1.5
2 FRAP
BCBDPPH
.OH ) n=3
1 (P<0.0
AA MEC
Fe2+
Fe2+
50 . IC .
28
±2
50 EDTA IC
Fe2+
) n=3
1 (P<0.0
MEC
0 20 40 60 80 100 120
0 50 100 150 200 250 300
MEC AA
LDH THP-1
LPS
Chemlal LDH
(P<0.01) LDH
/ml g (P<0.01)
TNF-
/ml g
LPS
LDH ) TNF-
n=3 (P<0.01
in vitro
Coratina Chemlal
DPPH
in vitro R
phenoxyl
(Pietta,2000) (catechol)ortho-dihydroxy
B
4-oxo C
0 20 40 60 80 100 120
0 100 200 300 400 500 600 700 800 900 1000
Cont OEC leaves
10 ug/ml 100 1000 LPS OEC leaves
10 +LPS 100 + LPS 1000 + LPS ug/ml
(Croft , 2006) B OH
oxo (Croft ,2006 ; Bors et al. ,1990) (Rice-Evans et al
. ,1996) .
tannin DPPH
(Uchidaet al .,1987) tannin
galloylation
DPPH
- OH
2.
O galloylation
- OH
2.
O Pearson p-hydroxy benzoic acid
verbascoside tyrosol
hydroxytyrosol hydroxytyrosol
hydroxytyrosol
(Soniet al. ,2006) Chemlal
.OH
+
Fe2 .OH
Stohs and Bagchi .,1995b
+
Fe2
(Lloyd et al .,1997)
.OH
+
Fe2
GAE luteolin
secoiridoid oleuropein
. hydroxytyrosol
B (Burda and Oleszek,2001
C2-C3 oxo
oleuropein
.OH
et al
Chemlal
in vitro THP-1
LPS
THP-1 LPS
TNF-
LPS TNF-
THP-1 LPS
-2
O. .OH
.NO O2
H2
B - NF
(Schreck et al .,1991) TNF-
(Sanliogluet al.,2001) O2
H2
Oxidases NADPH
(Bokock,1995)
NADPH O2
-2
O.
O2
H2
O2
SOD LPS
NADPH Oxidase Rac
GTP ROS
TNF- (Sanlioglu et al .,2001)
TNF-
LPS
TNF- NF- B
luteolin
secoiridoid oleuropein
hydroxytyrosol
.NO
Zaslaver et al .,2005 hydroxytyrosol
tyrosol
1 -
2 THP
-
O.
O2
H2
B - NF
hydroxytyrosol ROS
.NO GSH
THP-1 LPS
Zhang et al .,2009 .
THP-1
NF- B et al TNF-
.,2010 .
et al ., 2004 hydroxytyrosol
O2
H2 -2
O.
ROS
in vitro
Coratina Chemlal
Sigoise
Frantoio Coratina
Chemlal Maiatica
Sigoise Ogliarola
Pearson p-hydroxy benzoic acid verbascoside
tyrosol hydroxytyrosol
in vitro Chemlal
.OH Fe2+
.OH Fe2+
LDH 1
- THP
TNF- LPS
Chemlal Sigoise
hydroxytyrosol Chemlal
in vitro
Chemlal hydroxytyrosol
Chemlal
in vivo
in vivo Chemlal
.1.
Albino libitum
.
(0.9%NaCl) Chemlal
(0.9%NaCl) (Sharma et al
.,2007) Iwata et al., 1973
.
Chemlal (Kaeidi et al., 2011)
o rmp .
. .
. .
(Fawcett and Scott,1960)
sera pak kitts De Jaffe (Popper ,1937 ; Seelig and Wust,1969)
.Seurobiokitts
. . (MDA)
%15KCL saccharose
%1 .
CAT SOD
GSH
GPx GST
GR
(MDA) MDA MDA
TBA (Ohkawa et al., 1979)
pH MDA
MDA
. (CAT) Catalase
CAT (Aebi ,1974)
O2
500 Mm H2
pH
O2
H2
nm
O2
H2
.
. (SOD)Superoxide dismutase
SOD (Bauchamp and Fridovich ,1971)
(NBT)
-2
O.
riboflavin
% OD
OD - OD
x
x
. . Glutathion
GSH DTNB
( Ellman,1959 )
. GPx
.,1984) et al
(Mohandas O2
H2
GR
NADPH NADP+
GR nmol NADPH
oxidized/min/g .,2007
et al Zhu . et GST
(Alin
al.,1985) nmol CDNB
GST
. 4.
(LDH) Lactate dehydrogenase (LDH) Kroblewski
Sigma
Wroblewski,1969
MPO MPO -
dianisidine
O2
H2
50mM sodium phosphate
hexadecyl trimethyl amonium bromide(pH6) rpm phosphate
dianisidine dihydrochloride o-
O2
H2
MPO MPO
O2
H2
.,1984) et al
(Bradley
NO Griess NO
nitrite
glycine Griess
0.1% N-(1-
) OPO4
naphthyl)ethylene diaminedihydrochloride,1%sulfanilamide,2.5%H3
nitrite
(Green et al.,1982)sodium nitrite
% pH
m x
400
(ANOVA) in vivo
P<0.05 P<0.01
(P<0.01) (P<0.01)
Chemlal (a)
(b) n=
P<0.01
Malondialdehyde.
(p < 0,01) MDA
(p < 0,01)
Chemlal MDA
n=
P<0.01
0 10 20 30 40 50 60 70 80
control CME HgCl2 Se+ HgCl2 CME + HgCl2 0
0.5 1 1.5 2 2.5 3 3.5 4
control CME HgCl2 Se + HgCl2 CME + HgCl2
mg/dl mg/dl
0 20 40 60 80 100 120
control CME HgCl2 Se+ HgCl2 CME+ HgCl2 nmol/g tissue
b a
CAT SOD
(p < 0,01) CAT
SOD
(p < 0,01) CAT
SOD
Chemlal CAT
(a)
SOD (b) n=
P<0.01
0 10 20 30 40 50 60 70 80 90 100
control CME HgCl2 Se+ HgCl2 CME+ HgCl2 0
10 20 30 40 50 60
control CME HgCl2 Se+ HgCl2 CME+ HgCl2 nmol/min/mg
U/mg protein
a
b
Glutathion (p < 0,01)
GSH
GPx GR
GTS
(p < 0,01) GSH
GPx 72.24
GR
GTS 80.38
55.21
0 5 10 15 20 25 30 35
control CME HgCl2 Se + HgCl2 CME + HgCl2
0 5 10 15 20 25 30
control CME HgCl2 Se + HgCl2 CME + HgCl2 0
2 4 6 8 10 12 14 16
control CME HgCl2 Se+ HgCl2 CME+ HgCl2 nmol/mg protein
nmol/ min/mg protein
nmol/ min/mg protein
**
**
a
b
c
Chemlal GSH
(a) GPx
(b) GR (c) GTS (d) n=
P<0.01
(p < 0,01) LDH
(p < 0,01) MPO
NO
LDH
(p < 0,01) MPO
NO
0 1 2 3 4 5 6
control CME HgCl2 Se + HgCl2 CME + HgCl2 µmol/ mg protein
d
Chemlal LDH
(a) MPO
(b) (c)NO n=
P<0.01
0 50 100 150 200 250
control CME HgCl2 Se+ HgCl2 CME+ HgCl2
0 10 20 30 40 50 60 70 80
control CME HgCl2 Se+ HgCl2 CME+ HgCl2 U/mg tissu
nmol/g tissu µg/g tissu
0 20 40 60 80 100 120
control CME HgCl2 Se+ HgCl2 CME+ HgCl2 a
b
c
x400
Chemlal x
A B
C D
A B C D
Stacchiottiet al ., 2003 (
Perottoniet al ., 2004
CAT SOD
MDA ROS
.OH ROS
.(Hayes and McLellan ,1999)
GSH SOD
O2
H2
OH
2 SOD
-
O.
GPx O2
H2
GSH
GSSG GPx
CAT O2
H2
GSH (Meister.,1988) GSH
GPx
GSSG GR
GSH
, 2004 al .
Perottoni et GSH .
GSH
Zalups and Lash ,1996 ; Quig ,1998
GSH
2 GPx
2O H
O2
H2
ROS GSH
ROS O2
H2
, 1991) et al.
(Lund
OH MDA
et al ., 2007 (Sharma et al .,
2007) ROS
et al., 2007
NO.
MPO NOS
L-argenine L-citrulline
NO.
,2010) et al
;Jung ,2010
et al (Breno
NOS
.,1999) et al
(Boucher NO
-2 - O
ONOO
Yapet al.,2009 MPO
Tipping and
Timoshanko , 2005 ; Takada et al .,1997 ; Kelly and Singer,1993 in vitro TNF-
LPS
THP-1 TNF-
et al
P-selectin E-selectin
ICAM-1 (Ghielli et al. , 2000)
MPO
(Borregaard and Cowland ,1997 ;Klebanoff,1999) MPO
Hoy et al .,
2002 MPO
RNS
ROS HOCl
., 2001) et al
(Radi O2
H2
(Taurog and
Dorris.,1992)
.OH
-2
O.
HOCl RNS
- HOCl ONOO
.,2000) et al
(Van Dallen
oxidase NAD(P)H
Xanthine oxidase
2 HOCl
-
O.
O2
H2
(Weiss ., 1989) ROS
HgCl2
2007) et al.,
(
MDA LDH
LDH LDH
ROS GSH
(Smith et al .,1986)
GSH/GSSG
CAT SOD
MDA
.(Sharma et al .,2007) ROS
OH ROS
caspase3
(Valkoet al., 2006) thioredoxin
(Das .,2005) thioredoxine
reductase metallothionein
(Hansen et al .,2006)
Hsp72 (Ait Aissa et al
.,2000 ; Guo et al .,2007) Hsp25
GSH
Hsp25
Hsp25 Stacchiottiet al.,2004
. ROS RNS
ROS RNS
MPO HOCl
GSH Schrauffstatter et al .,1990
HOCl GSH
(Winterbourn and Brennan.,1997)
LDH Weisset al.,1982
HOCl
progelatinase procollagénase
(Peppin and Weiss., 1986)
N-acetyl
hyaluronic acid N-acetyl glucosamine sulfate
proteoglycan
ONOO .,1991
et al Katrantzis RNS
2Cl NO
2 HOCl NO .,2003) et al
(Golpon MPO
caspase3 (Myzak and carr., 2002)
MPO
(Mathy-Hartert et al .,1995) ,1989
Weiss .
ROS
DMPS DMPS
: (Georg et al ., 2004)
(Greguset al .,1992)
et al
selenoprotein-P
(Sasakura and Suzuki., 1998) MDA
GPx O2
H2
GSH ., 1995
et al Madhavi
selenoprotein p Sasakura and Suzuki ,1998
Chemlal
ROS
GSH GPx
GR CAT
GTS SOD
.OH
in vitro Chemlal
.OH GAE
luteolin
secoiridoid oleuropein
hydroxytyrosol
in vitro OH (Heim et al.,2002
B C2-C3
oxo luteolin
RNS ROS
(Odonturaet al.,2005)
RNS ROS
,1999) et al.
(Nagata quercetin
catechin
O2
H2
GPx 9
- BL
SOD GPx
CAT
(Kaviarasan et al .,2008)
oleuropein hydroxytyrosol
oleuropein olenolic
ethanol secoiridoid
oleuropein
(SGLT1) Edgecombe et al ., 2000
glucosidases hydroxytyrosol oleuropein
(Corona et al . , 2006)
oleuropein
, 2002) et al .
(Andrikopoulas OH
, 2001) et al.
(Gordon
hydroxytyrosol O2
H2
, 2001) et al .
(Tuck
(Deiana et al. , 2008) LLC-PK1
hydroxytyrosol
2 MDA
2O H
(Deiana et al .,2011) tocopherol
hydroxytyrosol (Paiva-Martins et al .
. ,2003) .
NO MPO
RNS
MPO MPO
. in TNF-
vitro THP-1
LPS
TNF- RNA
et al
hydroxytyrosol
- OH ONOO l
HOC
MPO (Aruoma and Halliwell , 1987)
.
(Kim et al .,2004) luteolin
(Hendriks et al
.,2004) NOS
TNF- NF-
(Kim et al.,2007)
(Chen et al .,2007)AP-1 NF-
(Xagorari
et al., 2001) luteolin
TNF-
luteolin ortho-dihydroxy
B OH
C-5 A
(Odontura et al .,2005)
oleuropein RNS
NO .(De La Puerta., 2001)
hydroxytyrosol oleuropein
MPO
- OH ONOO et HOCL
(Visioli .
al . , 2002) hydroxytyrosol
molecule (Carluccio
.,2003) et al
hydroxytyrosol O2
H2 -2
O.
et al ,
2004 . oleuropein O-dihydroxy
Benavente-Garcia et al., 2000
hydroxytyrosol ortho-diphenolic
OH (Granados-Principalet al.,2010)
chemlal Olea europaea L
.
CAT SOD
MDA
NO.
MPO
MDA LDH
MDA Chemlal
ROS
.OH NO
MPO TNF-
in vitro THP-1
LPS
Olea europaea L
luteolin oleuropein
hydroxytyrosol Olea europaea L
Chemlal
Olea europaea L Coratina
Frantoio Leccino
Matiatica Ogliarala
Chemlal Sigoise
RP-HPLC DAD
p-hydroxy benzoic acid vanillic acid
caffeic acid
gallic acid syringic acid
p-coumaric acid ferulic acid
sinapic acid
chrysoeriol luteolin
hydroxytyrosol tyrosol
secoiridoids oleuropein
Caffoyl glycoside phenylethanoid
verbascoside gallic acid
Sigoise Chemlal
hydroxytyrosol Sigoise
Chemlal tannin
Leccino Chemlal
tannin Sigoise
Cilento Sigoise
Chemlal secoiridoids
oleuropein
hydroxytyrosol luteolin
Chemlal
Coratina Chemlal
Frantoio
Coratina Chemlal
Maiatica Sigoise
Ogliarola hydroxytyrosol
Chemlal
.OH Fe2+
LDH TNF-
LPS THP-1
Chemlal hydroxytyrosol
in vivo
Chemlal
.OH Fe2+
Chemlal ROS
.OH in vitro
GAE luteolin
secoiridoid oleuropein
hydroxytyrosol
NO MPO
RNS MPO
MPO .
TNF-
in vitro THP-1
LPS
luteolin secoiridoids
oleuropein hydroxytyrosol
Olea europaeaL
Olea europaea L Chemlal
Chemlal oleuropein
Chemlal Chemlal
oleuropein
in vitro in vivo
hydroxytyrosol
Chemlal Chemlal
in vitro in vivo
Olea europaea.L Olea europaea.L
luteolin oleuropein
hydroxytyrosol
(CLE)Chemlal (OE)
Coratina Chemlal
hydroxytyrosol DPPH
-carotene (CLE )
in vivo Chemlal
(CLE) .
(CLE)
superoxyde
dismutase catalase
glutathion peroxydase glutathion transferase
glutathion
réductase myeloperoxidase
LDH (CLE)
(CLE ) luteolin
oleuropein .hydroxytyrosol
Olea europaea.L
Olea europaea
Luteolin Hydroxytyrosol
Oleuropein
Résumé
Olea europaea L. contre la néphropathie induite par le chlorure du mercure chez les souris mâle Albinos wistar. La composition phénolique et les activités biologiques des extraits des feuilles et des Olea europaea L. des différentes variétés Algériennes et Italiennes ont été étudiées. Le contenu total des phénols et des tanins a été quantifiés dans les extraits.
En outre, 14 différents composés phénoliques ont été identifiés et leurs profils ont montré des différences quantitatives remarquables entre les extraits d'olive analysés
(OE). La lutéoline glycosylée identifiés dans
les feuilles d'oliviers de Chemlal (CLE). Les résultats ont montré que (OE) de Chemlal, entre les variétés Algériennes, et (OE) de Coratina, entre les espèces t révélés être les meilleurs inhibiteurs du DPPH, du blanchissement du carotène, et des bons réducteurs du fer. Bien que le (CLE) ait nettement montré des actions antioxydantes et anti-inflammatoires. L'étude in vivo a été conçue pour étudier l'effet protecteur possible de (CLE) contre le stress oxydatif rénal induit par le chlorure de mercure (HgCl2). Des souris Wistar albinos mâles adultes ont été exposés à (HgCl2; 5 mg / kg de poids corporel; ip) et (CLE; 200 mg / kg de poids corporel) pendant 10 jours par rapport à selinate de sodium (0,1 mg / kg) comme référence standard. Le traitement avec du HgCl2 a induit des dysfonctionnements rénaux par l'augmentation de l'urée et de la créatinine. La peroxydation des lipides a été élevée avec une diminution concomitante de glutathion et diverses enzymes antioxydantes, à savoir le superoxyde dismutase, le catalase, le glutathion peroxydase, le glutathion transférase
2a augmenté la production d'oxyde nitrique et de l'activité de la myéloperoxydase et la réduction de LDH. Le prétraitement avec (CLE) a montré une bonne protection comme en témoigne la correction fonctionnelle rénale, avec une amélioration du système antioxydant. Les réactions inflammatoires ont été modulées. L'examen histopathologique des tissus
2. Les résultats obtenus indiquent clairement que la réponse de (CLE) et de selinate de sodium sont comparables dans la plupart des paramètres. Ces résultats ont montré que l'extrait de feuilles d'olivier possédait des actions antioxydantes et anti-inflammatoire qui pourraient être attribuées aux principes bioactifs (lutéoline glycosylée, hydroxytyrosol, oleuropéine) agissant synergiquement. Les résultats suggèrent que Olea europaea peuvent représenter une source naturelle importante avec un potentiel antioxydant et anti-inflammatoire élevé et semblent être applicable à la fois à la santé et à l'alimentation médicale.
Mot Clés
Nephropathie , Chlorure du mercure ,Olea europaea L, Extraits des olives, Extraits des f Phénols, Lutéoline glycosylée ,Hydroxytyrosol, Oleuropein, Activité antioxydante , Activité anti-inflammatoire.
Abstract
This study investigate the protector effect of Olea europaea.L against nephropathy induced by chloride mercury in mouse male Albinos wistar.Phenolic composition and biological activities of olives and leaves extracts from Italian and Algerian Olea europaea L. cultivars were studied.Total phenolic and tannin contents were quantified in the extracts. Moreover 14 different phenolic compounds were identified , and their profiles showed remarkable quantitative differences among analysed olive extracts (OE). Glucosylated luteolin ,hydroxytyrosol and oleuropein are identified in the Chemlal olive leaves (CLE). Results showed that (OE)of Chemlal, between Algerian cultivars, and (OE) of Coratina, among Italian ones , and hydroxytyrosol, among identified phenolic compounds, were found to be the best inhibitors of the DPPH , and Carotene bleaching and a good iron reducers. Although (CLE) markedly provided an antioxidant and antiinflammatory actions. In vivo ,the study was designed to investigate the possible protective effect of (CLE) in mercuric chloride (HgCl2) induced renal oxidative stress. Adult male albino Wistar mice were exposed to (HgCl2; 5 mg/kg bwt;ip) and (CLE; 200 mg/kg bwt) for 10 days compared with sodium selinate (0.1 mg/kg) as standard reference. Treatment with HgCl2 induced renal dysfunction by increasing urea , creatinin . Lipid peroxidation was elevated and along with a concomitant decrease in glutathione and various antioxidant enzymes, namely superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase. HgCl2 intoxication increased nitric oxide production and myeloperoxidase and decreased LDH activities . Pretreatment with (CLE) exhibited a good protection as evidenced by the correction of renal functional, amelioration of the antioxidant system . The inflammatory reactions were modulated. Histopathological examination of the kidney tissues proved the protective effect of (CLE) against HgCl2. The obtained results clearly show that, the response of(CLE)and sodium selinate are comparable in most parameters. These findings showed that , the olive leaf extract possessed an antioxidant and an anti-inflammtory actions that could be attributed to the bioactive principles (glucosylated luteolin, hydroxytyrosol, oleuropein) synergistically acted. Results suggest that Olea europaea leaves and olives extracts can represent an important natural source with high antioxidant and anti- inflammatory potential and would seem to be applicable in both the health and medical food.
Key words
Nephropathy , Mercury chloride , Olea europaea, , Olive extracts, olive leaves extracts , Phenols ,Glucosylated luteolin ,Hydroxytyrosol, Oleuropein, Antioxydant
activity, Anti-inflammatory action
