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EVALUATION OF THE ANTIGENOTOXIC POTENTIAL OF
GARLIC SULFUR COMPOUNDS IN HEPG2 CELLS
C. Belloir, M.H. Siess, C. Daurat and A.M. Le Bon
INRA, UMR Toxicologie Alimentaire, 17 rue Sully, 21065 Dijon cedex, France
Introduction
Numerous epidemiological studies have reported that consumption of garlic reduces the risk of
cancers in humans1. Organosulfur compounds present in high amounts in garlic would account for its
anticarcinogenic activity. However only few reports are available on the antigenotoxic properties of
garlic constituents in human cells. The aim of the present study was to assess the antigenotoxic
potential of a major constituent of garlic, diallyl sulfide (DADS) in a human cell line, HepG2 cells. The
protective activities of two metabolites of DADS, allicin2 (DADSO) and allyl mercaptan3 (AM), were
also determined.
S
Diallyl disulfide (DADS)
S
O
Allicin (DADSO)
S
S
SH
Allyl mercaptan (AM)
Methods
In order to study the mechanisms of action (modulation of drug metabolizing enzymes, scavenging of
ultimate genotoxic compounds), two experimental protocols were performed :
i) a pre-treatment protocol in which HepG2 cells were first incubated with the sulfur compound for 20
hr then were treated with the indirect-acting genotoxic compound (aflatoxin B1 (AFB1) or
benzo(a)pyren (BaP) or nitrosodimethylamine (DMN)) for 20 hr;
ii) a co-treatment protocol in which HepG2 cells were simultaneously incubated with the sulfur
compound and the direct-acting genotoxic compound (hydrogen peroxyde (H2O2) or 4nitrosoquinoline oxide (4-NQO) or methylmethane sulfonate (MMS)) for 4 hr.
After the treatments, cells were harvested. DNA damage was immediately evaluated using the comet
assay4. The tail moment as defined by Olive et al5 was recorded as parameter for DNA damage.
Experimental protocol
Seeding
0
24
48
72 hours
Pre-treatment study
Co-treatment study
Comet assay
Sulfur compound
Genotoxic compound
Results
PRE-TREATMENT STUDY
AFB1
CO-TREATMENT STUDY
DMN
BaP
H2O2
MMS
25
DADS
Olive tail moment
6
4
*
3
*
*
*
2
*
*
*
4
1
20
4
*
*
20
12
*
*
15
0
0
G
5
25
50
100 µM
*
*
C
G
5
25
50
100 µM
5
*
3
*
*
0
C
G
5
25
15
*
8
10
4
5
0
0
50
100 µM
C
G
5
25
50
100 µM
*
4
6
*
4
50
100 µM
6
G
5
25
50
100 µM
C
G
5
25
50
C
100 µM
*
8
G
5
25
50
*
*
12
*
*
4
10
*
6
4
5
*
15
25
50
*
*
25
50
100 µM
*
5
0
100 µM
14
*
G
10
0
*
*
*
4
0
C
G
5
25
50
100 µM
C
G
10
5
*
20
8
*
100 µM
*
*
*
15
6
8
3
6
*
10
4
2
2
*
2
0
C
C
20
5
5
*
100 µM
10
0
25
50
4
1
5
25
*
2
G
5
15
2
C
G
*
20
2
1
C
*
6
3
0
Olive tail moment
*
5
4
AM
*
2
2
C
Olive tail moment
*
16
10
0
DADSO
*
6
4-NQO
4
2
1
0
0
C
G
5
25
50
100 µM
2
5
0
0
2
0
C
G
5
25
50
100 µM
0
C
G
5
25
50
100 µM
C
G
5
25
50
100 µM
C
G
5
25
50
100 µM
C
G
5
25
50
100 µM
C = control cells G = cells + genotoxic compound
= cells + genotoxic compound + sulfur compound (5-100 µM)
* : significantly different to values for samples treated with genotoxic compound alone (P 0.05, Mann-Whitney-U-test)
 Pre-treatment of HepG2 cells with DADS or DADSO inhibited the
genotoxicity of aflatoxin B1. DADS and AM significantly reduced DNA
damage induced by benzo(a)pyrene. AM afforded protection against
dimethylnitrosamine genotoxicity in a concentration-dependent
manner.
 In co-treatment studies, AM and, to a lower extent, DADSO and
DADS were shown to inhibit hydrogen peroxide genotoxicity.
DNA damage provoked by methylmethane sulfonate was significantly
prevented when this genotoxic compound was incubated together with
DADS or DADSO. AM and DADSO strongly reduced 4nitrosoquinoline oxide genotoxicity.
Conclusion
The present study showed that DADS and two metabolites, DADSO and AM, afford protection
against DNA damage induced by direct- and indirect-acting genotoxic compounds. These sulfur
compounds could act through different mechanisms of action, i.e. modulation of enzymes
involved in activation/detoxification of genotoxic compounds and scavenging of ultimate species.
These findings support the assumption that sulfur compounds could be responsible for the
chemopreventive properties of garlic in humans.
References
1. Fleischauer,A.T. and Arab,L., 2001. J; Nutr., 131, 1032-1040.
2. Teyssier C. et al., 1999. Drug. Metab. Disp., 27, 835-841.
3. Egen-Schwind C. et al., 1992. Planta Medica, 58, 301-305.
4. Singh N.P. et al., 1988. Exp. Cell. Res., 175, 184-191.
5. Olive P.L. et al., 1990. Radiat. Res., 122, 86-94.
This project was partly financed by the European
Quality of Life and Management of Living Resources