Synthesis and cytotoxic activity of geranylmethoxyhydroquinone derivatives

Article abstract of DOI:10.4067/s0717-97072012000300005

The new synthetic geranyl-2,4-methoxyhydroquinone 1 and the known geranyl-4,5-methoxyhydroquinone 2 were prepared by Electrophilic Aromatic Substitution (EAS) reactions between geraniol and 1,3,5-trimethoxyphenol using BF3·Et2O as a catalyst. Furthermore,

Full text of DOI:10.4067/s0717-97072012000300005

J. Chil. Chem. Soc., 57, Nº 3 (2012)
SYNTHESIS AND CYTOTOXIC ACTIVITY OF GERANYLMETHOXYHYDROQUINONE DERIVATIVES
EVELYN BAEZA MATURANAa, KAREN CATALÁN MARÍNa, JOAN VILLENA GARCÍAb, HÉCTOR CARRASCO
ALTAMIRANOc, MAURICIO CUELLAR FRITISd AND LUIS ESPINOZA CATALÁNa*.
aDepartamento de Química, Universidad Técnica Federico Santa María, Valparaíso, Chile.
bCentro de Investigaciones Biomédicas, Facultad de Medicina, Universidad de Valparaíso, Centro Regional de Estudios en Alimentos Saludables,
CREAS, Valparaíso, Chile.
cDepartamento de Ciencias Químicas, Universidad Andrés Bello, Viña del Mar, Chile.
dFacultad de Química y Farmacia, Universidad de Valparaíso, Valparaíso, Chile.
(Received: October 3, 2011 - Accepted: April 8, 2012)
ABSTRACT
The new synthetic geranyl-2,4-methoxyhydroquinone 1 and the known geranyl-4,5-methoxyhydroquinone 2 were prepared by Electrophilic Aromatic
Substitution (EAS) reactions between geraniol and 1,3,5-trimethoxyphenol using BF ∙Et O as a catalyst. Furthermore, the new geranylmethoxyhydroquinones
derivatives (3-6) were obtained by chemical transformations of 1 and 2. The compou3nds2have been evaluated for their cytotoxic activities against PC-3 human
prostate cancer cell line, MCF-7 and MDA-MB-231 human breast cancer cells lines and dermal human fibroblasts DHF. IC50 values for compounds 1 and 5 ranged
in the 80 µM level.
Keywords: Geranylmethoxyhydroquinone, synthesis, cytotoxic activity
INTRODUCTION
Polyprenylated 1,4-benzoquinones and hydroquinones such as
ubiquinones, plastoquinones, and tocopherols are widespread in plants
and animals, in which they play important roles in electron transport,
photosynthesis, and as antioxidants1,2. Prenyl benzoquinones have been also
isolated from brown algae of the order Fucales3-6, sponges7-10, alcyonaceans11,
gorgonaceans12, and ascidians belonging to the genus Aplidium13-18. These
substances present a terpenoid portion ranging from one to nine isoprene units.
On the other hand, different studies of the structure activity relation (SAR) in
a series of nonmethoxylated and methoxylated prenylated quinones with side
chains containing from one to eight isoprene units reported that the optimum
length of the side-chain is two isoprene units and in the para-position relative
to the methoxy-group19, 20. Additionally, these authors informed that all tested
quinones (3-Demethylubiquinone Q2 and its synthetic derivatives) have
inhibited of JB6 Cl41 cell transformation and p53 activity and induction of
apoptosis, AP-1 and NF-kB activity.
Scheme 1. Synthesis of 2,4,5-trimethoxyphenol 9 (a) MCPBA/CH2Cl2, r.t.
1.87 g, 86%; (b) CH3OH/Et3N, r.t. 2.45 g, 78%.
The next objective consisted in the coupling reaction between geraniol
with 2,4,5-trimethoxyphenol (9). For this purpose the strategy of Electrophilic
Aromatic Substitution (EAS), was used21-23. Nevertheless, in this reaction
was not possible to obtain the wished compound 10 (scheme 2) and
unexpectedly two compounds were obtained: 2,4-dimethoxy-5-((E)-3’,7’-
dimethylocta-2’,6’-dienyl)phenol (1) (15% yield) and 4,5-dimethoxy-2-((E)-
3’,7’-dimethylocta-2’,6’-dienyl)phenol (2) (13% yield). The mass difference
corresponds to geraniol not reacted, geraniol decomposition and not reacted
2,4,5-trimethoxyphenol.
Due to the importance that showed by these compounds in the mentioned
biological activities, in this work we describe the synthesis and cytotoxic
activity of geranylmethoxyhidroquinones derivatives (1-6). The new
synthetic geranyl-2,4-dimethoxyhydroquinone 1 and known geranyl-4,5-
dimethoxyhydroquinone 2 were synthesized using the strategy of Electrophilic
Aromatic Substitution (EAS) reactions, according to the reported protocol21-23
.
The geranylmethoxyhidroquinones derivatives (3-6) are synthetic analogs of 1
and 2, while the known compound 5 is analogous to the marine natural product
verapliquinone A. All the compounds were evaluated in vitro against various
human cancer cells lines in order to analyse the influence of the molecular
structure on the cytotoxic activity.
Figure 1. Structure of synthesized compounds.
RESULTS AND DISCUSSION
Chemistry
Scheme 2. Synthesis of geranylmethoxyhydroquinone derivatives. (a)
dioxane/BF3∙Et2O, N2, r.t.: 1, 79.1 mg 15% and 2 65.3 mg 13% yield.
The formation of compounds 1 and 2 was not possible to explain it by the
EAS strategy, probably the coupling reaction ocurred from a different pathway,
competing with the EAS. In the presence of BF3, the compound 9 afforded
the allyl aryl ethers 9a wich presumably undergoes Claisen rearrangement27-28
giving the dienones 9b y 9c. The dienones recover the aromaticity through the
formation and later loss of a formaldehyde molecule. The mechanism which
was proposed to explain the formation of compounds 1 and 2 is showed in
scheme 3.
Our first objective was the preparation of compound (9) from commercially
available asaronaldehyde (7). The first step involves a Baeyer Villiger
oxidation reaction and subsequent saponification of formate 824, according to
the synthetic route shown in scheme 1.
e-mail: luis.espinoza@usm.cl
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J. Chil. Chem. Soc., 57, Nº 3 (2012)
Scheme 3. Proposed reaction mechanism for the formation of compounds 1 and 2.
In the 1H-NMR spectrum of geranyl-2,4-dimethoxyphenol 1 the signals at
δ 3.82 ppm (s, 3H, OCH ), 3.76 (s, 3H, OCH3) of the two methoxyl groups, δ
3.25 ppm (d, J = 7.0, 2H,3H-1’) of the point of coupling, 5.49 ppm (s, 1H, OH)
of the phenol and two aromatic hydrogens at δ 6.68 ppm (s, 1H, H-6), 6.54 (s,
1H, H-3) were mainly observed. In the 13C-NMR spectrum the presence of the
methoxyl groups at δ 56.8 ppm, 56.1 ppm, the signals of two aromatic carbons
at δ 99.3 ppm (C-3) and 112.8 ppm (C-6) also were confirmed. These data also
spectral data20. This compound showed the existence of an hydroxyl group at
δ 4.93 ppm and the presence of two aromatic hydrogens at δ 6.62 ppm (s, 1H,
H-5); 6.45 ppm (s, 1H, H-6). In addition, the point of coupling was confirmed
by the presence of the signal at δ 3.30 (d, J = 7.0 Hz, 2H, H-1’). While in the
13C-NMR spectrum the signals at δ 113.7 (C-3), 101.2 (C-6) of two aromatic
carbons and 29.6 (C-1’) corroborate the molecular structure.
For the preparation of 2,4-dimethoxy-5-((E)-3’,7’-dimethylocta-2’,6’-
dienyl)phenyl acetate derivative 3, the geranyl-2,4-dimethoxyphenol 1 was
acetylated with acetic anhydride and dimethylaminopyridine (DMAP) as
catalyst, then the compound 3 was obtained with a 55% of yield (scheme 4).
The molecular structure was corroborated for the signals at δ 2.31 ppm, 20.7
ppm, 169.2 ppm of the acetyl group, in addition for comparison of their spectral
data with geranyl-2,4-dimethoxyphenol 1.
The preparation of 2,4,5-trimethoxy-1-((E)-3’,7’-dimethylocta-2’,6’-
dienyl)benzene derivative 4, was carried out by reaction of methylation with
(CH3)2SO4 in slightly basic conditions of the compound 1 or 2 with a 40 %
yield (scheme 4). 1H-NMR spectrum showed three singlets at δ 3.87 ppm 3.82
ppm and 3.80 ppm, the signals corresponding to the methoxyl groups. These
signals also were observed in the 13C-NMR spectrum at δ 56.3 ppm, 56.7 ppm
and 56.6 ppm.
3
were corroborated by 2D HMBC J heteronuclear correlations between H-1’
and the carbon signals at δ 151.8 (C-4), 112.8 (C-6), 136.0 (C-3’) ppm and
2J with δ 122.8 (C-2’), 120.9 (C-5) ppm (figure 2a). The E-geometry of the
trisubstituted C2’-C3’ double bond was established from gs-sel-1H 1D-NOESY
experiments (figure 2b), where H-1’ showed a long range interaction with the
methyl group in position 10’ (1.70 ppm). In addition, the configuration of the
C2’-C3’ double bond was compared with the chemical shifts of C-10’ reported
for similar compounds19.
The
2-methoxy-5-((E)-3’,7’-dimethylocta-2’,6’-dienyl)-1,4-benzoqui-
none compound 5, was prepared by oxidation reaction of geranyl-4,5-dimeth-
oxyphenol 2 with nitrate of cerium and ammonium (CAN), and was obtained
with a 15 % yield (scheme 4). The molecular structure was confirmed for the
presence of the signals at δ 6.46 ppm (t, 1H, H-6) and 5.92 ppm (s, 1H, H-3)
Figure 2. Structure of compound 1. (a) HMBC correlations; (b) NOE
correlations.
1
of the quinonic hydrogens in the H-NMR spectrum and the existence in the
Nevertheless the 1H-NMR spectrum of 2 was compared with the reported
13C-NMR spectrum of two carbonyl carbons at δ 182.4 ppm (C-1), 187.6 ppm
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J. Chil. Chem. Soc., 57, Nº 3 (2012)
(C-4), in addition to comparison with the spectral data reported for the com-
pound20.
gel (Merck 200-300 mesh) was used for C.C. and silica gel plates HF-254 for
TLC. TLC spots were detected by heating after spraying with 25% H2SO4 in
H2O.
Finally the 4,5-dimethoxy-2-((E)-3’,7’-dimethylocta-2’,6’-dienyl)phenyl
acetate derivative 6, was prepared using the same protocol of synthesis for
the derivative 3 with a 70 % yield (scheme 4). 1H-NMR and 13C-NMR spectra
showed the signals corresponding to the acetyl groups at δ 2.29 ppm, 20.8 ppm
and 169.8 ppm.
Chemistry
General procedure for the Electrophilic Aromatic Substitution (EAS),
synthesis of compounds 1, 2
BF3.Et2O (0.08 g, 0.6 mmol) was gradually added at room temperature to
a solution of 2,4,5-trimethoxyphenol (0.3278 g, 1.8 mmol) and geraniol (0.3 g,
1.8 mmol) in freshly distilled 1,4-dioxane (20 mL). The mixture was stirred at
room temperature under a nitrogen atmosphere for 24 h, when the completion
of the reaction was verified by TLC. The mixture was poured onto crushed ice
(about 30 g) and the organic layer was extracted with EtOAc (3 × 30 mL). The
combined organic phase was washed with 5% NaHCO3 (30 mL), then with
water (2 × 20 mL) and dried over anhydrous Na SO4, filtered and evaporated.
The crude residue was redissolved in CH Cl2 (25 mL) and chromatographed
on silica gel with petroleum ether/EtOA2c mixtures of increasing polarity
(19.8:0.2→16.0:4.0 for 1 and 19.8:0.2→13.0:7.0 for 2).
2,4-dimethoxy-5-((E)-3’,7’-dimethylocta-2’,6’-dienyl)phenol 1: colorless
1
viscous oil, 79.1 mg (15%). H-NMR: 6.68 (s, 1H, H-6); 6.54 (s, 1H, H-3);
5.49 (s, 1H, OH); 5.28 (t, J= 7.0 Hz, 1H, H-2’); 5.11 (t, J= 7.0 Hz, 1H, H-6’);
3.82 (s, 3H, OCH3); 3.76 (s, 3H, OCH3); 3.25 (d, J= 7.0, 2H, H-1’); 2.08 (m,
4H, H-5’ and H-4’); 1.70 (s, 3H, H-10’); 1.67 (s, 3H, H-8’); 1.60 (s, 3H, H-9’).
13C-NMR: 144.1 (C-1), 140.1 (C-2), 99.3 (C-3), 151.8 (C-4), 120.9 (C-5),
112.8 (C-6), 56.8 (OCH3), 56.1 (OCH3), 27.7 (C-1’), 122.8 (C-2’), 136.0 (C-
3’), 39.8 (C-4’), 26.8 (C-5’), 124.3 (C-6’), 131.4 (C-7’), 25.7 (C-8’), 17.7 (C-
9’), 16.1 (C-10’). IR (cm-1): 2924, 1602, 1512, 1194, 1042. MS (m/z, %): M+
290 (< 1%), 288 (13), 287 (28), 246 (43), 245 (9), 219 (12), 203 (15), 178 (39),
177 (70), 175 (9), 163 (17), 161 (31), 149 (10) 124 (16), 123 (100), 122 (21),
69 (36).
Scheme 4. Synthesis of geranylmethoxyhydroquinone derivatives.
(a) Ac2O/CH2Cl2/DMAP, r.t. for 3, 30 mg 55%; for 6, 30 mg 70% yield
(b) (CH3)2SO4/K2CO3, acetone, r.t. for 4 from 1, 37 mg, 40% whereas for
compound 4 from 2 the same yield was obtained (c) CAN/acetonitrile/water,
r.t. 20 mg 15% yield.
Antiproliferative activity
The cytotoxicity of the compounds was evaluated in vitro against three
different cancer cell lines: PC-3 human prostate cancer, MCF-7 and MDA-
MB-231 human breast cancer and one non-tumoral cell line, dermal human
fibroblasts (DHF) by using sulforhodamine dye assay. This conventional
colorimetric assay was set up to estimate the IC50 values which represents the
concentration of a drug that is required for 50% inhibition in vitro after 72 h
of treatment with the test compounds. Five serial dilutions (from 12.5 to 100
µM) for each sample were evaluated in triplicate. The results obtained from
these assays are shown in Table 1. The compounds 2-4 and 6 did not affect
the viability of the cells lines studied, however the compounds 1 and 5 showed
IC50 values with inhibitory activity, ranged in the 80 µM level for breast cancer
cell line MCF-7. Moreover the compound 5 showed IC50 values with inhibitory
activity, ranged in the 80 µM level for breast cancer cell line MDA-MB-231,
these values may be due to the presence of a quinone moiety. In the case of
the compound 1 the cytotoxicity could be influenced by the position of the
hydroxyl group in the ring, orto to the methoxyl group. Furthermore, it was
demonstrated by the results obtained for the acetylated derivative 3 and the
meta position of the hydroxyl group in the compound 2, which did not display
cytotoxic activity. Moreover, compound 1 and 5 showed some selectivity for
the cancer cells versus fibroblast cells because the IC for human fibroblast
were on 100 µM, which could provide an approach50to obtain compounds
with potential less toxicity in normal human cells. The increased citotoxicity
induced by compounds 1 and 5 on human breast cancer cell lines was due
probably to the presence of a quinone moiety and orto position of the hydroxyl
group on the structure of these compounds versus the compounds 2, 3, 4 and 6.
4,5-dimethoxy-2-((E)-3’,7’-dimethylocta-2’,6’-dienyl)phenol 2: colorless
1
viscous oil, 65.3 mg (13%). H-NMR: 6.62 (s, 1H, H-5); 6.45 (s, 1H, H-6);
5.30 (t, J= 7.0 Hz, 1H, H-2’); 5.07 (t, J= 7.0 Hz, 1H, H-6’); 4.93 (s, 1H, OH);
3.82 (s, 6H, OCH ); 3.30 (d, J= 7.0, 2H, H-1’); 2.09 (m, 4H, H-5’ and H-4’);
1.78 (s, 3H, H-103’); 1.68 (s, 3H, H-8’); 1.60 (s, 3H, H-9’). 13C-NMR: 148.4
(C-1), 117.3 (C-2), 113.7 (C-3), 142.8 (C-4), 148.3 (C-5), 101.2 (C-6), 55.9
(OCH ), 56.6 (OCH3), 29.6 (C-1’), 121.9 (C-2’), 138.6 (C-3’), 39.6 (C-4’), 26.4
(C-5’)3, 123.7 (C-6’), 132.0 (C-7’), 25.6 (C-8’), 17.7 (C-9’), 16.2 (C-10’). IR
(cm-1): 2924, 1619, 1512, 1451, 1195. MS (m/z, %): M+ 290 (40), 221 (4), 207
(4), 205 (15), 190 (5), 175 (6), 168 (12), 167 (100), 166 (8), 69 (9).
Synthesis of geranylmethoxyhidroquinones derivatives (3–6)
2,4-dimethoxy-5-((E)-3’,7’-dimethylocta-2’,6’-dienyl)phenyl acetate 3
To geranyl-2,4-dimethoxyphenol 1 (0.0485 g, 0.16 mmol) dissolved in
dichloromethane (10 mL), acetic anhydride (0.02 g, 0.02 mL, 0.16 mmol) and
dimethylaminopyridine (DMAP, 2 mg, 0.016 mmol) were added. The mixture
was stirred at room temperature during 2 hours. The organic phase was washed
with water and after it was dried over anhydrous Na2SO4. The solution was
evaporated to dryness to afford the crude reaction product, which gave 30 mg
(55 % yield) of compound 3 after column chromatography (eluent to hexane/
ethyl acetate 19.8:0.2→17.8:2.2), colorless viscous oil. 1H-NMR: 6.80 (s, 1H,
H-6); 6.57 (s, 1H, H-3); 5.30 (t, J= 7.0 Hz, 1H, H-2’); 5.12 (t, J= 7.0 Hz, 1H,
H-6’); 3.77 (s, 3H, OCH3); 3.76 (s, 3H, OCH3); 3.30 (d, J= 7.0, 2H, H-1’); 2.31
(s, 3H, CO2CH3); 2.09 (m, 4H, H-5’ and H-4’); 1.70 (s, 3H, H-10’); 1.67 (s,
3H, H-8’); 1.60 (s, 3H, H-9’). 13C-NMR: 144.6 (C-1), 137.7 (C-2), 106.1 (C-3),
151.2 (C-4), 128.2 (C-5), 114.4 (C-6), 56.0 (OCH3), 56.6 (OCH3), 28.0 (C-1’),
122.0 (C-2’), 136.6 (C-3’), 39.7 (C-4’), 26.7 (C-5’), 124.2 (C-6’), 131.4 (C-7’),
26.7 (C-8’), 17.7 (C-9’), 16.1 (C-10’); 20.7 (CO2CH3); 169.2 (CO2). IR (cm-1):
2929, 1768, 1768, 1510, 1215, 1203, 1040. MS (m/z, %): M+ 332 (87), 291
(20), 290 (94), 221 (39), 207 (16), 189 (40), 175 (18), 168 (14), 167 (100), 161
(52), 154 (13), 129 (20), 69 (18).
EXPERIMENTAL
General
Unless otherwise stated, all chemical reagents purchased (Merck or
Aldrich) were of the highest commercially available purity and were used
without previous purification. IR spectra were recorded as thin films in a
Nicolet Impact 420 spectrometer and frequencies are reported in cm-1. Low
resolution mass spectra were recorded on a Shimadzu QP-2000 spectrometer at
70eV ionising voltage and are given as m/z (% rel. int.). 1H, 13C (DEPT 135),
sel. 1D 1H NOESY, 2D HSQC and 2D HMBC spectra were recorded in CDCl3
solutions and are referenced to the residual peaks of CHCl3 at δ 7.26 ppm and
δ 77.0 ppm for 1H and 13C, respectively, on a Bruker Avance 400 Digital NMR
spectrometer, operating at 400.1 MHz for 1H and 100.6 MHz for 13C. Chemical
shifts are reported in δ ppm and coupling constants (J) are given in Hz. Silica
2,4,5-trimethoxy-1-((E)-3’,7’-dimethylocta-2’,6’-dienyl)benzene 4
To 1 or 2 (90 mg, 0.3 mmol) dissolved in acetone (10 mL), potassium of
carbonate (0.04 g, 0.3 mmol) and dimethyl sulphate (0.04 g, 0.3 mmol) were
added. The mixture was stirred at room temperature during 24 hours. After
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J. Chil. Chem. Soc., 57, Nº 3 (2012)
filtration, the solution was evaporated to dryness. Dilution with diethyl ether
was followed by washing with NaOH solution (5%) and the organic phase was
dried over anhydrous Na2SO4. The solution was evaporated to dryness to afford
the crude reaction product, which gave 37 mg (40 % yield) of compound 4 after
column chromatography (eluent to hexane/ethyl acetate 19.8:0.2→18.2:1.8),
dryness. Dilution with EtOAc was followed by washing with HCl solution
(5%) (2 × 10 mL) and the organic phase was dried over anhydrous Na2SO4. The
solution was evaporated to dryness to afford the crude reaction product, which
gave 2.45 g (78 % yield) of compound 9 after column chromatography (eluent
1
to hexane/ethyl acetate 19.8:0.2→14,0:6,0), colorless viscous oil H-RMN:
1
colorless viscous oil. H-NMR: 6.71 (s, 1H, H-6); 6.52 (s, 1H, H-3); 5.29 (t,
6.58 (s, 1H, H-6), 6.44 (s, 1H, H-3), 3.87 (s, 3H, OCH3), 3.84 (s, 6H, OCH3).
J= 7.0 Hz, 1H, H-2’); 5.11 (t, J= 7.0 Hz, 1H, H-6’); 3.87 (s, 3H, OCH3); 3.82
(s, 3H, OCH3); 3.80 (s, 3H, OCH ); 3.27 (d, J= 7.0, 2H, H-1’); 2.08 (m, 4H,
H-5’ and H-4’); 1.71 (s, 3H, H-130’); 1.67 (s, 3H, H-8’); 1.61 (s, 3H, H-9’).
13C-NMR: 143.0 (C-1), 147.6 (C-2), 98.2 (C-3), 151.4 (C-4), 121.9 (C-5),
113.9 (C-6), 56.3 (OCH3), 56.7 (OCH3), 56.6 (OCH3), 27.6 (C-1’), 122.7 (C-
2’), 136.0 (C-3’), 39.8 (C-4’), 26.8 (C-5’), 124.3 (C-6’), 131.4 (C-7’), 25.7
(C-8’), 17.7 (C-9’), 16.0 (C-10’). IR (cm-1): 2921, 1511, 1459, 1202,1037. MS
(m/z, %): M+ 304 (100), 235 (36), 221 (20), 205 (22), 204 (52), 203 (13), 190
(13), 189 (24), 182 (62), 69(11).
13C-RMN: 104.8 (C-3), 100.1 (C-6).
Cell lines
The experimental cell lines MCF-7, MDA-MB-231 and PC-3 were
obtained from American Type Culture Collection (Rockville, MD, USA).
MCF-7, MDA-MB-231, PC-3 and DHF cells were grown in DMEM-F12
containing 10% FCS, 100 U/ml penicillin, 100 µg/ml streptomycin and 1 mM
glutamine at 37°C under a humidified 5% CO2.
Cell growth inhibition assay
2-methoxy-5-((E)-3’,7’-dimethylocta-2’,6’-dienyl)-1,4-benzoquinone 5
The colorimetric assay using sulforhodamine B (SRB) following an
adaptation of the described method for Skehan25-26 was used. Cells were seeded
in 96-well microtiter plates, at 5x103 cells per well in aliquots of 100 µL of
medium, and they were allowed to attach to the plate surface by growing in
drug-free medium for 18 h. Afterwards, compounds samples were added in
aliquots to achieve a final concentration of 12.5, 25, 50 and 100 µM. Stock
solution of compounds was prepared in ethanol and the final concentration of
this solvent was kept constant at 1%. Control cultures received 1% ethanol
alone. After 72 h exposure, the cytotoxicity was measured by the SRB dye
assay. Cells were fixed by adding cold 50% (wt/vol) trichloroacetic acid (TCA,
25 µL) and incubated for 60 min at 4 ºC. Plates were washed with deionized
water and dried; SRB solution (0.1% wt/vol in 1% acetic acid, 50 µL) was
added to each microtiter well and incubated for 30 min at room temperature.
Unbound SRB was removed by washing with 1% acetic acid. Plates were air-
dried and bound stain was solubilized with Tris base (100 µL, 10 mM). Optical
densities were read on an automated spectrophotometer plate reader at a single
wavelength of 540 nm. Values shown are the % viability vs. Ctrl + SD, n=four
independent experiments in triplicate.
To geranyl-4,5-dimethoxyphenol 2 (0.1416 g, 0.5 mmol) dissolved in
20 mL acetonitrile/water (1:1), CAN (0.1725 g, 0.3 mmol) was added. The
mixture was stirred at -5 ºC during 20 minutes. Dilution with diethyl ether
was followed by washing with water and the organic phase was dried over
anhydrous Na2SO4. The solution was evaporated to dryness to afford the crude
reaction product, which gave 20 mg (15 % yield) of compound 5, after column
chromatography (eluent to hexane/ethyl acetate 19.8:0.2→17.0:3.0), colorless
viscous oil. 1H-NMR: 6.46 (t, 1H, H-6); 5.92 (s, 1H, H-3); 5.15 (t, J= 7.0 Hz,
1H, H-2’); 5.07 (t, J= 7.0 Hz, 1H, H-6’); 3.81 (s, 3H, OCH ); 3.13 (d, J= 7.0,
2H, H-1’); 2.07 (m, 4H, H-5’ and H-4’); 1.69 (s, 3H, H-10’);31.61 (s, 3H, H-8’);
1.60 (s, 3H, H-9’). 13C-NMR: 182.4 (C-1), 158.7 (C-2), 107.7 (C-3), 187.6
(C-4), 149.6 (C-5), 130.3 (C-6), 56.2 (OCH3), 27.3 (C-1’), 117.8 (C-2’), 140.1
(C-3’), 39.6 (C-4’), 26.4 (C-5’), 130.3 (C-6’), 131.9 (C-7’), 25.7 (C-8’), 17.7
(C-9’), 16.1 (C-10’). IR (cm-1): 3018, 2929, 1675, 1651, 1606, 1458. MS (m/z,
%): M+ 274 (< 1%), 247 (3), 221 (20), 205 (29), 168 (15), 167 (100), 166 (8),
161 (6), 69 (11).
4,5-dimethoxy-2-((E)-3’,7’-dimethylocta-2’,6’-dienyl)phenyl acetate 6
Table 1. Cytotoxicity (IC50 µM +/- SD) of geranylmethoxyhydroquinone
derivatives 1-6.
To geranyl-4,5-dimethoxyphenol 2 (0.0367 g, 0.13 mmol) dissolved
in dichloromethane (10 mL), acetic anhidric (0.01 g, 0.01 mL, 0.13 mmol)
and dimethylaminopyridine (DMAP, 1.5 mg, 0.013 mmol) were added. The
mixture was stirred at room temperature during 2 hours. The organic phase was
washed with water and after it was dried over anhydrous Na2SO4. The solution
was evaporated to dryness to afford the crude reaction product, which gave
30 mg (70 % Yield) of compound 6 after column chromatography (eluent to
hexane/ethyl acetate 19.8:0.2→17.0:3.0), colorless viscous oil. 1H-NMR: 6.70
(s, 1H, H-6); 6.56 (s, 1H, H-3); 5.22 (t, J= 7.0 Hz, 1H, H-2’); 5.10 (t, J= 7.0 Hz,
1H, H-6’); 3.84 (s, 3H, OCH3); 3.83 (s, 3H, OCH3); 3.17 (d, J= 7.0, 2H, H-1’);
2.29 (s, 3H, CO CH3); 2.07 (m, 4H, H-5’ and H-4’); 1.69 (s, 3H, H-10’); 1.67
(s, 3H, H-8’); 12.59 (s, 3H, H-9’). 13C-NMR: 146.9 (C-1), 124.8 (C-2), 112.3
(C-3), 141.8 (C-4), 147.5 (C-5), 106.1 (C-6), 56.0 (OCH ), 56.1 (OCH3), 28.2
(C-1’), 121.8 (C-2’), 136.8 (C-3’), 39.7 (C-4’), 26.7 (C-5’3), 124.1 (C-6’), 131.5
(C-7’), 25.7 (C-8’), 17.7 (C-9’), 16.2 (C-10’), 20.8 (CO2CH3), 169.8 (CO2). IR
(cm-1): 2922, 1764, 1658, 1620, 1513, 1205, 1174, 1014. MS (m/z, %): M+ 332
(87), 291 (20), 290 (94), 221 (39), 207 (16), 189 (40), 175 (18), 168 (14), 167
(100), 161 (52), 154 (13), 129 (20), 69 (18).
MDA-
MB-231
Compound
PC-3
MCF-7
DHF
1
2
3
4
5
6
> 100
> 100
> 100
> 100
> 100
> 100
82.2 +/- 6.5
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
84.2 +/- 8.9
> 100
86.2 +/- 7.1
> 100
> 100
1.34 +/-
0.31
Daunorubicin 0.36+/-0.12 0.19 +/- 0.11 0.32 +/- 0.09
CONCLUSIONS
In summary, we have prepared the new synthetic geranyl-2,4-
methoxyhydroquinone and known geranyl-4,5-methoxyhydroquinone
2,4,5-trimethoxyphenyl formate 8
1
2. These compounds were obtained unexpectedly by Electrophilic
Aromatic Substitution (EAS) coupling reactions between geraniol with
2,4,5-trimethoxyphenol. Furthermore, the geranylmethoxyhydroquinone
derivatives (3-6) were obtained of the chemical transformations of the
compounds coupling 1 and 2. The compound 5 showed cytotoxic activity
against cell line MCF-7 and MDA-MB-231 with IC50 values of 84.2 and 86.2
µM respectively. The compound 1 showed some selectivity against cell line
MCF-7 with an IC value of 82.2 µM. The increased citotoxicity induced by
compounds 1 and 550 on cell lines can be probably due to the presence of a
quinone moiety and position of the hydroxyl group on the structure of these
compounds versus the compounds 2, 3, 4 and 6.
To 2,4,5-trimethoxybenzaldehyde 7 (2.02 g, 10 mmol) dissolved in
dichloromethane (70 mL), metachloroperoxibenzoic acid (MCPBA, 3.30 g, 19
mmol) and sodium bicarbonate (1.82 g, 21 mmol) were added. The mixture
was stirred at room temperature during 2 hours. The organic phase was washed
with water (2 × 10 mL) and after it was dried over anhydrous Na SO . The
solution was evaporated to dryness to afford the crude reaction prod2uct 4which
gave 1.87 g (86 % Yield) of compound 8 after column chromatography (eluent
1
to hexane/ethyl acetate 19.8:0.2→14,0:6,0), colorless viscous oil H-RMN:
8.28 (s, 1H, CHO), 6.85 (s, 1H, H-6), 6.78 (s, 1H, H-3), 3.91 (s, 3H, OCH3),
3.85 (s, 6H, OCH3). 13C-RMN: 161.3 (CHO), 122.1 (C-6), 101.8 (C-3).
2,4,5-trimethoxyphenol 9
ACKNOWLEDGEMENTS
To 2,4,5-trimethoxyphenyl formate 8 (3.68 g, 17 mmol) dissolved in
methanol (50 mL) triethylamine (4 mL) were added. The mixture was stirred
at room temperature during 30 minutes. Then, the solution was evaporated to
The authors thank to Programa Incentivo a la Iniciación Científica (PIIC
QUI-2010 for E.B.M. and grant Nº 13.11.36) of the Dirección General de
1222

J. Chil. Chem. Soc., 57, Nº 3 (2012)
Investigación y Postgrado (DGIP) from the Universidad Técnica Federico
Santa María and grant CT-Val Nº FB/30 LE /10, for financial support.
14. Targett, N. M.; Keeran, W. S.; J. Nat. Prod. 1984, 47, 556.
15. Guella, G. I.; Mancini, Pietra, F.; Helv. Chim. Acta 1987, 70, 621.
16. Faulkner, D. J.; Nat. Prod. Rep. 1993, 10, 497.
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