SUPPLEMENTARY MATERIAL
Hepatoprotective and neuroprotective tocopherol analogues isolated from the peels
of Citrus unshiu Marcovich
Kyeong-Hwa Seoa, Dae-Young Leeb, Dong-Sung Leec, Hee-Cheol Kangd, Hak-Soo
Kime, Youn-Chul Kimcand Nam-In Baeka*
aGraduate School of Biotechnology and Department of Oriental Medicine
Biotechnology, Kyung Hee University, Yongin 446-701, Republic of KoreabDepartment of Herbal Crop Research, National Institute of Horticultural and Herbal
Science, RDA, Eumseong 369-873, Republic of KoreacCollege of Pharmacy, Wonkwang University, Iksan 570-749, Republic of KoreadR&D Center, GFC Co., Ltd., Yongin 446-908, Republic of KoreaeSEOULCOSMETICS, Co., Ltd, R&D center, Incheon 405-821, Republic of Korea
Three tocopherol analogues methoxytocopherol (1), α-tocopherol (2)
and γ- tocopherol (3),were isolated from the peels of Citrus unshiu
Marcovich. The protective effects of the isolated compounds against
tert-butyl hydroperoxide (t-BHP)-induced hepatotoxicity in human
liver-derived HepG2 cells and glutamate-induced oxidative stress in
HT22-immortalized hippocampal cells were evaluated. Compounds 1-3
were significantly protective in HepG2, cells with EC50 values of 21.22
± 2.01, 25.21 ± 2.11, and 25.25 ± 1.21 μM, respectively, and in HT22
cells, compounds 1-3 had EC50 values of 20.62 ± 1.36, 6.44 ± 1.65, and
9.52 ± 1.54 μM, respectively.
Keywords:Citrus unshiu, hepatoprotective, HepG2, HT22,
methoxytocopherol, neuroprotective, tocopherol
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Structures and Protective effects of Compounds 1-3
Table S1. Protective effects of compounds 1-3, isolated from the peels of Citrus unshiu
Marcovich, on glutamate-induced oxidative injury in HT22 cells and t-BHP-
induced hepatotoxicity in HepG2 cells
Figure S1. Structures of compounds 1-3 from the peels of Citrus unshiu.
Figure S2.Effects of compounds 1-3 on t-BHP-induced oxidative injury in HepG2 cells.
Cells were pretreated with compounds 1-3 for 12 h and then incubated for 6
h with t-BHP (50 μM). Curcumin (20 μM) was used as a positive control.
Data are presented as the means ± S.D. of three independent experiments;
*P < 0.05 vs. control.; #P < 0.05 vs. t-BHP.
Figure S3. Protective effects of compounds 1-3 on glutamate-induced oxidative
neurotoxicity in HT22 cells. Cells were treated with compounds 1-3 for 4 h
and then incubated for 12 h with glutamate (5 mM). Trolox (100 μM) was
used as a positive control. Data are presented as the means ± S.D. of three
independent experiments; *P < 0.05 vs. control.; #P < 0.05 vs. glutamate.
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Table S1. Protective effects of compounds 1-3, isolated from the peels of Citrus unshiu
Marcovich, on glutamate-induced oxidative injury in HT22 cells and t-BHP-induced
hepatotoxicity in HepG2 cells
Compound HT22 (EC50, μM) HepG2 (EC50, μM)
Positive control Trolox (22.42 ± 3.23) Curcumin (8.78 ± 2.41)
1 20.62 ± 1.36 21.22 ± 2.01
2 6.44 ± 1.65 25.21 ± 2.11
3 9.52 ± 1.54 25.25 ± 1.21
The HT22 and HepG2 cells were treated with 1-3 for 4 h and then incubated for 12 h
with glutamate (5 mM). Next, the cells were incubated for 6 h with t-BHP (50 μM). The
resulting half-maximal effective concentrations (EC50) are expressed as the percentage
of viability compared to the control. Trolox and curcumin were used as positive
controls. Data represent the mean values ± SD of three experiments.
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Figure S1. Structures of compounds 1-3 from the peels of Citrus unshiu.
Figure S2. Effects of compounds 1-3 on t-BHP-induced oxidative injury in HepG2
cells. Cells were pretreated with compounds 1-3 for 12 h and then incubated
for 6 h with t-BHP (50 μM). Curcumin (20 μM) was used as a positive
control. Data are presented as the means ± S.D. of three independent
experiments; *P < 0.05 vs. control.; #P < 0.05 vs. t-BHP.
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Figure S3. Protective effects of compounds 1-3 on glutamate-induced oxidative
neurotoxicity in HT22 cells. Cells were treated with compounds 1-3 for 4 h
and then incubated for 12 h with glutamate (5 mM). Trolox (100 μM) was
used as a positive control. Data are presented as the means ± S.D. of three
independent experiments; *P < 0.05 vs. control.; #P < 0.05 vs. glutamate.
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