Inhibitory Activities of Ferulic and Caffeic Acid Alkyl Esters
J. Agric. Food Chem., Vol. 54, No. 15, 2006 5377
pure compounds. The caffeic acid esters 3-6 were prepared by Fischer
esterification as described below for ferulate esters (Figure 1) (17).
NMR (CDCl3) δ 167.4, 147.9, 146.7, 144.6, 127.0, 123.0, 115.6, 114.7,
109.3, 70.5, 55.9, 27.8, 19.1.
1
The structures of caffeic esters were confirmed by H and 13C NMR
1
n-Octyl Ferulate (21). H NMR (CDCl3) δ 7.62 (1H, d, J ) 15.0
spectral studies (18-20).
Hz), 7.19 (1H, dd, J ) 8.0, 2.0 Hz), 7.03 (1H, d, J ) 1.8 Hz), 6.91
(1H, d, J ) 8.4 Hz), 6.29 (1H, d, J ) 15.9 Hz), 4.18 (2H, t, J ) 6.6
Hz), 3.92 (3H, s, OMe), 1.67 (2H, m), 1.29 (10H, br s), 0.88 (3H, t, J
) 6.6 Hz). 13C NMR (CDCl3) δ 167.4, 147.9, 146.7, 144.6, 127.0,
123.0, 115.6, 114.7, 109.3, 64.6, 55.9, 31.8, 29.2, 29.18, 26.0, 22.6,
14.1.
Synthesis of Ferulic Acid Esters. Ferulic acids esters (15-27) were
synthesized according to the reported procedure (Figure 1) (17). The
ferulic acid (1 mM) and corresponding alcohols (1 mM) were stirred
with dry toluene (2 mL). The catalyst, p-toluenesulfonic acid (catalytic
amounts), was added to the reaction mixture,which was stirred for 4 h
at 120 °C. It was then extracted with saturated NaHCO3 solution and
the organic layer was dried over anhydrous Na2SO4. The solvent was
evaporated under reduced pressure, and the resulting syrupy mass was
purified on a silica gel column with n-hexane/EtOAc (9:1 and 8:2 v/v)
as the mobile phase. Fifteen fractions (15 mL each) were collected.
Evaporation of fractions eluted with hexane/EtOAc (8:2 v/v) gave pure
compounds. The structures of ferulic acid esters were confirmed by
1H and 13C NMR spectral experiments (21, 22).
1
Isooctyl Ferulate (22). H NMR (CDCl3) δ 7.62 (1H, d, J ) 15.0
Hz), 7.19 (1H, dd, J ) 8.0, 2.0 Hz), 7.05 (1H, d, J ) 2.0 Hz), 6.98
(1H, d, J ) 8.0 Hz), 6.32 (1H, d, J ) 15.0 Hz), 5.01 (1H, m), 3.98
(3H, s, OMe), 1.73 (1H, m), 1.45 (1H, m), 1.28 (8H, br s), 1.24 (3H,
d, J ) 6.0 Hz), 0.92 (3H, t, J ) 6.0 Hz). 13C NMR (CDCl3) δ 167.0,
147.8, 146.7, 144.4, 127.1, 123.0, 116.1, 114.6, 109.2, 71.0, 55.9, 36.0,
31.7, 29.1, 25.4, 22.6, 20.1, 14.1.
1
Dodecyl Ferulate (23). H NMR (CDCl3) δ 7.62 (1H, d, J ) 15.9
1
Hz), 7.07 (1H, dd, J ) 8.4, 2.1 Hz), 7.03 (1H, d, J ) 2.1 Hz), 6.91
(1H, d, J ) 8.1 Hz), 6.29 (1H, d, J ) 15.9 Hz), 4.19 (2H, d, J ) 6.6
Hz), 3.92 (3H, s, OMe), 1.68 (4H, m), 1.26 (14H, br s), 0.88 (3H, t, J
) 6.6 Hz). 13C NMR (CDCl3) δ 167.3, 147.8, 146.7, 144.6, 127.0,
123.0, 115.6, 114.6, 109.2, 64.6, 55.9, 31.8, 29.6, 29.5, 29.3, 29.2, 28.7,
26.0, 22.6, 14.1.
n-Propyl Caffeate (3). H NMR (acetone-d6) δ 8.26 (2H, s, OH),
7.53 (1H, d, J ) 16.2 Hz), 7.15 (1H, d, J ) 2.1 Hz), 7.04 (1H, dd, J
) 8.1, 2.1 Hz), 6.86 (1H, d, J ) 8.1 Hz), 6.28 (1H, d, J ) 16.2 Hz),
4.09 (2H, t, J ) 6.6 Hz), 1.67 (2H, m), 0.95 (3H, t, J ) 7.5 Hz). 13C
NMR (acetone-d6) δ 167.2, 148.4, 146.1, 145.3, 127.5, 122.3, 116.2,
115.6, 115.0, 65.9, 22.6, 10.5.
1
Eicosyl Ferulate (26). H NMR (CDCl3) δ 7.59 (1H, d, J ) 15.9
1
Isobutyl Caffeate (6). H NMR (acetone-d6) δ 8.30 (2H, s, OH),
Hz), 7.06 (1H, dd, J ) 8.4, 2.1 Hz), 7.02 (1H, d, J ) 2.0 Hz), 6.96
(1H, d, J ) 8.4 Hz), 6.27 (1H, d, J ) 15.9 Hz), 4.16 (2H, t, J ) 6.6
Hz), 3.90 (3H, s, OMe), 1.67 (2H, m), 1.23 (34H, br s), 0.86 (3H, t, J
) 6.0 Hz). 13C NMR (CDCl3) δ 167.4, 147.9, 146.7, 144.6, 127.0,
123.0, 115.6, 114.7, 109.2, 64.6, 55.9, 31.9, 29.7, 29.5, 29.4, 29.3, 28.8,
26.0, 22.7, 14.1.
7.54 (1H, d, J ) 16.2 Hz), 7.16 (1H, d, J ) 1.8 Hz), 7.05 (1H, dd, J
) 8.1, 1.8 Hz), 6.86 (1H, d, J ) 8.1 Hz), 6.29 (1H, d, J ) 16.2 Hz),
3.92 (2H, d, J ) 6.6 Hz), 1.96 (2H, m), 0.95 (6H, d, J ) 6.9 Hz). 13
C
NMR (acetone-d6) δ 167.2, 148.4, 146.0, 145.3, 127.4, 122.3, 116.1,
115.5, 115.0, 70.4, 28.4, 19.1.
1
Isopentyl Caffeate (8). H NMR (acetone-d6) δ 8.30 (2H, s, OH),
7.50 (1H, d, J ) 15.9 Hz), 7.15 (1H, d, J ) 2.1 Hz), 7.04 (1H, ddd, J
) 8.1, 2.1, 0.3 Hz), 6.86 (1H, d, J ) 8.1 Hz), 6.27 (1H, d, J ) 15.9
Hz), 4.17 (2H, t, J ) 6.9 Hz), 1.65 (1H, m), 1.58 (2H, m), 0.93 (6H,
d, J ) 6.6 Hz). 13C NMR (acetone-d6) δ 167.2, 148.5, 146.1, 145.3,
127.4, 122.3, 116.1, 115.5, 115.0, 62.8, 38.1, 25.6, 22.5.
RESULTS
The major hydroxycinnamic acids present in cereals are
ferulic acid, p-coumaric acid, and sinapic acid (1). The anecdotal
reports on the health benefits of these compounds are primarily
due to their antioxidant capacity alone. Therefore, we have
synthesized a series of alkyl esters of caffeic and ferulic acids
to study the impact of alkyl chain lengths of these esters and
their tumor cell proliferation, COX enzyme, and lipid peroxi-
dation inhibitory activities. In tumor cell proliferation inhibitory
assays, caffeic acid esters were more active than the ferulates.
For example, dodecyl (10) and hexadecyl (11) caffeates inhibited
colon cells by 89% and 88%, respectively, at 20 µg/mL (Figure
2A). A similar trend was observed for these compounds on all
other cell lines tested; dodecyl (10) and hexadecyl (11) caffeates
gave 82% and 75% inhibition on breast tumor cells; 80% and
77% inhibition on lung tumor cells; 79% and 52% on CNS
tumor cells; and 67% and 72% inhibition on AGS tumor cells
(Figure 2B-E). The IC50 values observed for compounds 10
were 4.35 (breast), 5.37 (lung), 2.46 (colon), 8.09 (CNS), and
7.57 (AGS) µg/mL; and for compound 11, 9.65 (breast), 8.62
(lung), 5.78 (colon), 12.04 (CNS), and 16.55 (AGS) µg/mL.
However, caffeates 3-8 showed only 40-50% inhibition on
colon cancer cell lines at 20 µg/mL. A similar growth inhibitory
effect was also observed for breast and lung cancer cell lines.
These compounds, 3-8, were also less effective on gastric and
CNS tumor cell lines as indicated by <30% growth inhibition.
Among ferulate esters, octyl (21) and dodecyl (23) esters of
ferulic acids were the most active and inhibited the growth of
breast tumor cells by 78% and 86%; of lung tumor cells by
27% and 77%; of colon tumor cells by 80% and 93%; of CNS
tumor cells by 48% and 79%; and of AGS tumor cells by 81%
and 87%, respectively (Figure 2A-E). The IC50 values
observed for compound 21 were 17.05 (breast), 11.0 (lung),
4.29 (colon), 14.63 (CNS), and 18.67 (AGS) µg/mL. Similarly,
dodecyl ferulate (23) exhibited IC50 values of 7.45, 7.76, 3.5,
12.21, and 13.46 µg/mL on breast, lung, colon, CNS, and AGS
1
Hexadecyl Caffeate (11). H NMR (acetone-d6) δ 8.31 (2H, br s,
OH), 7.52 (1H, d, J ) 16.2 Hz), 7.15 (1H, d, J ) 2.1 Hz), 7.03 (1H,
dd, J ) 8.1, 2.1 Hz), 6.85 (1H, d, J ) 8.1 Hz), 6.27 (1H, d, J ) 16.2
Hz), 4.13 (2H, t, J ) 6.6 Hz), 1.67 (2H, m), 1.27 (26H, br s), 0.86
(3H, t, J ) 6.9 Hz). 13C NMR (acetone-d6) δ 167.3, 148.5, 146.1, 145.3,
127.5, 122.3, 116.2, 115.6, 115.0, 64.5, 32.5, 28.9, 26.5, 23.2, 14.2.
1
Octadecyl Caffeate (12). H NMR (acetone-d6) δ 8.31 (2H, br s,
OH), 7.52 (1H, d, J ) 16.2 Hz), 7.15 (1H, d, J ) 2.1 Hz), 7.03 (1H,
dd, J ) 8.1, 2.1 Hz), 6.85 (1H, d, J ) 8.1 Hz), 6.27 (1H, d, J ) 16.2
Hz), 4.13 (2H, t, J ) 6.6 Hz), 1.67 (2H, m), 1.27 (30H, br s), 0.86
(3H, t, J ) 6.9 Hz). 13C NMR (acetone-d6) δ 167.3, 148.6, 146.2, 145.3,
127.4, 122.3, 116.2, 115.6, 115.0, 64.5, 32.5, 30.4-28.9, 26.5, 23.2,
14.2.
1
Eicosyl Caffeate (13). H NMR (acetone-d6) δ 8.45 (1H, s, OH),
8.21 (1H, s, OH), 7.58 (1H, d, J ) 15.9 Hz), 7.20 (1H, d, J ) 2.1 Hz),
7.09 (1H, dd, J ) 8.1, 2.1 Hz), 6.91 (1H, d, J ) 8.1 Hz), 6.32 (1H, d,
J ) 15.9 Hz), 4.18 (2H, t, J ) 6.6 Hz), 1.72 (2H, m), 1.32 (34H, br s),
0.92 (3H, t, J ) 6.9 Hz). 13C NMR (acetone-d6) δ 167.3, 148.5, 146.1,
145.3, 127.5, 122.3, 116.2, 115.6, 115.0, 64.5, 32.5, 30.4-28.9, 26.5,
23.2, 14.2.
Isopropyl Ferulate (16). 1H NMR (CDCl3) δ 7.59 (1H, d, J ) 15.9
Hz), 7.06 (1H, dd, J ) 8.4, 2.0 Hz), 7.00 (1H, d, J ) 2.1 Hz), 6.91
(1H, d, J ) 8.4 Hz), 6.27 (1H, d, J ) 15.9 Hz), 5.13 (1H, m), 3.91
(3H, s, OMe), 1.30 (6H, d, J ) 6.0 Hz). 13C NMR (CDCl3) δ 166.8,
147.8, 146.7, 144.4, 127.0, 123.0, 116.1, 114.6, 109.2, 67.6, 55.9, 21.9.
1
n-Butyl Ferulate (17). H NMR (CDCl3) δ 7.62 (1H, d, J ) 15.0
Hz), 7.19 (1H, dd, J ) 8.0, 2.0 Hz), 7.05 (1H, d, J ) 2.0 Hz), 6.98
(1H, d, J ) 8.0 Hz), 6.32 (1H, d, J ) 15.0 Hz), 4.22 (2H, t, J ) 6.0
Hz), 3.98 (3H, s, OMe), 1.73 (2H, m), 1.45 (2H, m), 1.0 (3H, t, J )
6.0 Hz). 13C NMR (CDCl3) δ 167.4, 147.9, 146.7, 144.6, 127.0, 123.0,
115.6, 114.7, 109.3, 64.3, 55.9, 30.8, 19.2, 13.7.
1
Isobutyl Ferulate (18). H NMR (CDCl3) δ 7.62 (1H, d, J ) 15.0
Hz), 7.19 (1H, dd, J ) 8.0, 2.0 Hz), 7.05 (1H, d, J ) 2.0 Hz), 6.98
(1H, d, J ) 8.0 Hz), 6.32 (1H, d, J ) 15.0 Hz), 4.01 (2H, d, J ) 6.0
Hz), 3.98 (3H, s, OMe), 2.0 (1H, m), 0.97 (6H, d, J ) 6.0 Hz). 13C