Total synthesis of altissimacoumarin D, a small molecule sirtuin1 activator

Article abstract of DOI:10.21577/0103-5053.20180021

The total synthesis of the plant natural product altissimacoumarin D was achieved by the Mitsunobu alkylation of isofraxidin by geraniol. Isofraxidin was prepared from 2,4-dihydroxybenzaldehyde in five steps. The key reaction was the Knoevenagel condensat

Full text of DOI:10.21577/0103-5053.20180021

http://dx.doi.org/10.21577/0103-5053.20180021
J. Braz. Chem. Soc., Vol. 29, No. 5, 1157-1161, 2018
Short Report
Printed in Brazil - ©2018 Sociedade Brasileira de Química
Total Synthesis of Altissimacoumarin D, a Small Molecule Sirtuin1 Activator
a
b
a
a
Anna C. Silva, Hanae Benelkebir, Rosangela S. C. Lopes, Claudio C. Lopes and
,
b
A. Ganesan*
a
Departamento de Química Analítica, Instituto de Química,
Universidade Federal do Rio de Janeiro, CT, Bloco A, 508, 21949-900 Rio de Janeiro-RJ, Brazil
b
School of Pharmacy, University of East Anglia, Norwich Research Park, NR4 7TJ, Norwich,
United Kingdom
The total synthesis of the plant natural product altissimacoumarin D was achieved
by the Mitsunobu alkylation of isofraxidin by geraniol. Isofraxidin was prepared from
2
,4-dihydroxybenzaldehyde in five steps. The key reaction was the Knoevenagel condensation of
an ortho-hydroxybenzaldehyde with Meldrum’s acid under neutral conditions in water and one-pot
acid catalyzed cyclization to the coumarin.
Keywords: natural products, epigenetics, sirtuins, coumarins
+
Introduction
dinucleotide (NAD )-dependent catalytic mechanism. The
seven human sirtuins, Sirt1-7, are implicated in a number
of disease conditions and there is much interest in both
Historically, natural products are a rich source of
biologically active compounds including many of our current
15
activators and inhibitors of sirtuin enzymes. Although some
dietary natural products such as resveratrol, quercetin or
epigallocatechin gallate are reported as sirtuin modulators,
their promiscuous nature is non-ideal for use as chemical
probes and more selective molecules are needed. A recent
lead from natural product screening comes from Ailanthus
altissima (the tree of heaven), a plant used in Chinese
and Korean traditional medicine. From the bark extract,
four coumarins were identified that are Sirt1 activators
1-5
therapeutic agents. In the present day, natural products
continue to play a significant role in the development of small
6
molecule chemical probes for the new science of epigenetics.
Numerous natural products that inhibit specific epigenetic
enzyme-catalyzed structural modifications of nucleic acids
and histone proteins are known, such as laccaic acid (targeting
7
deoxyribonucleic acid (DNA) methyltransferase), anacardic
8
acid (targeting histone acetyltransferase), trichostatin A
9
16
(
targeting histone deacetylase), nauhoic acid (targeting
(Figure 1). At a concentration of 10 µM, the most active
10
lysine methyltransferase) and tripartin (targeting lysine
compound altissimacoumarin D significantly increased Sirt1
catalyzed deacetylation of a p53 peptide Sirt1 substrate
in vitro and decreased the transcriptional activity of p53 in
a cell-based reporter assay.
1
1
demethylase). The depsipeptide romidepsin, a histone
deacetylase inhibitor of bacterial origin, is a particularly
impressive case as it has progressed from discovery to
an approved epigenetic drug for the treatment of T-cell
Due to the interesting biological activity, we embarked
on the total synthesis of altissimacoumarin D. Our biogenetic
retrosynthesis (Figure 2) assumed that the likely origin of
altissimacoumarin D (1) is through the geranylation of
another coumarin natural product, isofraxidin (2). There
are several total syntheses of isofraxidin in the literature,
12,13
lymphoma.
The zinc-dependent histone deacetylases (HDACs) are
enzymes that hydrolyze acyl-lysine (chiefly acetyl-lysine)
residues in proteins and revert them to native lysine, usually
14
with a gene silencing effect as a consequence. An additional
class of sirtuin histone deacylases catalyze the same
reaction through an entirely different nicotinamide adenine
1
7
by Spath and Jerzmanowska-Sienkiewiczowa, Spath
1
8
19
20
and Dobrovolny, Rouessac and Leclerc, Chen and
21
Gao et al. from which we selected the Chen route as the
starting point for our studies due to its concise and simple
nature. In Chen’s four step synthesis, isofraxidin arose
*
e-mail: a.ganesan@uea.ac.uk
This paper is part of the PubliSBQ Special Issue “IUPAC-2017”
http://publi.sbq.org.br/).
(

1
158
Total Synthesis of Altissimacoumarin D, a Small Molecule Sirtuin1 Activator
J. Braz. Chem. Soc.
Figure 1. The structures of altissimacoumarin Sirt1 activators.
from the formation of a coumarin from an o-hydroxy
benzaldehyde derivative, which is in turn prepared by
the functionalization of resorcinol. Nevertheless, we
have substantially altered the reaction conditions for the
Knoevenagel reaction and subsequent coumarin cyclization
compared to the published procedure.
conditions, we achieved a high yield of the Knoevenagel
23
adduct 6 with Bigi’s procedure. This mild protocol with
Meldrum’s acid in the absence of extraneous acid or base
catalysis has the benefit of being environmentally green
as it is conducted in water without organic co-solvents.
While intermediate 6 was cyclized to coumarin 7 in 90%
yield under acid catalysis, we discovered that telescoping
the Knoevenagel condensation and cyclization in a one-
pot operation from aldehyde 5 to coumarin 7 was more
efficient. The final step in the isofraxidin synthesis was the
coumarin decarboxylation, for which our best yields were
Results and Discussion
Our synthesis began with the halogenation of
commercially available 2,4-dihydroxybenzaldehyde
2
4
(
3, Scheme 1) to give the dibrominated derivative 4.
obtained by refluxing 7 in pyridine and ethylene glycol.
Halogen displacement under van Koten’s conditions
with copper(I) chloride and sodium methoxide afforded
With isofraxidin in hand through a five step sequence
(58% overall yield), we introduced the geranyl sidechain.
Under Mitsunobu conditions at room temperature, the
reaction between isofraxidin and geraniol (Scheme 2)
provided altissimacoumarin D (1). Although the reaction
proceeded in a modest 54% yield, there was no attempt at
optimization as we obtained a sufficient quantity (100 mg)
of the natural product for biological studies. Our synthetic
2
,4-dihydroxy-3,5-dimethoxybenzaldehyde (5) in good
2
2
yield. At this stage, Chen’s Knoevenagel condensation
with diethyl malonate was attempted but resulted in poor
yields. Furthermore, the lack of experimental detail made
an effective reproduction difficult. The method used
1
9
in Rouessac’s synthesis, whereby a trimethoxylated
benzaldehyde was condensed with Meldrum’s acid and
zinc oxide, was also unsatisfactory when applied to
our aldehyde 5. After investigating alternative reaction
1
13
material had an excellent match with the H and C nuclear
magnetic resonance (NMR) spectroscopic data reported for
the natural product (Table 1).
Figure 2. Retrosynthesis of altissimacoumarin D (1) from resorcinol.

Vol. 29, No. 5, 2018
Silva et al.
1159
Scheme 1. (a) Br , EtOH, 30 min (98%); (b) NaOMe, CuCl, MeOH/DMF, reflux, 4 h (70%); (c) Meldrum’s acid, H O, 80 ºC, 2 h, (94%); (d) H SO ,
2
2
2
4
90 min (90%); (e) one-pot combination of (c) and (d) (90%); (f) Py, ethylene glycol, reflux, 3.5 h (94%).
Scheme 2. (a) Geraniol, Ph P, diisopropyl azodicarboxylate (DIAD), tetrahydrofuran (THF), 24 h (54%).
3
1
13
Table 1. Comparison of altissimacoumarin D H and C NMR chemical
Conclusions
shifts between the naturally isolated material and the synthetic sample
In summary, we report the total synthesis of the
Sirt1 activator altissimacoumarin D in six steps from
dC, natural dC, synthetic
dH, natural product
dH, synthetic sample
product
sample
2
,4-dihydroxybenzaldehyde. A noteworthy feature is
1
1
1
2
3
4
4
5
5
6
6
7
.59 (s)
1.52 (s)
1.60 (s)
16.4
16.4
.67 (s)
17.6
17.6
the new high-yielding route towards the intermediary
isofraxidin via Knoevenagel condensation in water
followed by cyclization to the coumarin carboxylic acid 7
in one-pot. Thermal decomposition of the pyridinium salt
of 7 effected decarboxylation to give isofraxidin in high
yield, after which geranylation afforded altissimacoumarin
D. The ready access to altissimacoumarin D will facilitate
its use as a starting material for the synthesis of the more
heavily oxygenated natural products in this family, while
the modular route facilitates the synthesis of analogues for
structure-activity relationship studies.
.69 (s)
1.63 (s)
25.6
25.6
.05, 2.08 (m)
.89 (s)
1.99 (m)
26.3
26.3
3.82 (s)
39.6
39.6
.03 (s)
3.96 (s)
56.3
56.2
.68 (d, J 7 Hz)
.06 (t, J 7 Hz)
.56 (t, J 7 Hz)
.34 (d, J 10 Hz)
.66 (s)
4.62 (d, J 7 Hz)
5.00 (t, J 8 Hz)
5.49 (t, J 8 Hz)
6.28 (d, J 8 Hz)
6.58 (s)
61.7
61.7
70.3
70.2
103.5
114.4
115.1
119.6
103.5
114.4
115.1
119.6
123.8
131.7
141.8
142.5
143.0
143.4
144.9
150.7
160.5
.61 (d, J 10 Hz)
7.54 (d, J 12 Hz)
1
1
1
1
1
1
1
1
1
23.8
31.7
41.8
42.5
42.9
43.5
44.9
50.7
60.6
Experimental
2,4-Dihydroxybenzaldehydewasobtainedcommercially
(Sigma-Aldrich Co., St. Louis, MO, USA) and used
without further purification. All reactions involving a
dry atmosphere were performed with a slight positive
pressure of argon. Analytical thin-layer chromatography

1
160
Total Synthesis of Altissimacoumarin D, a Small Molecule Sirtuin1 Activator
J. Braz. Chem. Soc.
was done on 0.25 mm silica gel 60 F254 plates, and
compounds visualized using UV irradiation or KMnO4.
Flash chromatography was carried out on columns packed
with silica gel 60 (0.040-0.063 mm particle sizes). Nuclear
magnetic resonance (NMR) spectra were obtained on a
Bruker Avance-III spectrometer. The splitting patterns are
reported as s (singlet), d (doublet), t (triplet), q (quartet),
and m (multiplet).
at 80 ºC and the reaction mixture left for 2 h to the point
where a precipitate was formed. The solid was filtered and
dried to give compound 6 as a white solid (0.323 g, 94%).
mp 259-261 °C; ¹H NMR (400 MHz, MeOD-d ) d 2.05
1
(s, 6H), 3.85 (s, 3H), 3.88 (s, 3H), 7.05 (s, 1H), 8.70 (s,
13
1H); C NMR (100 MHz, MeOD-d ) d 56.2, 60.8, 105.8,
1
109.5, 112.8, 134.2, 144.7, 145.9, 146.9, 149.6, 157.5,
164.2 (some carbon signals are coincident).
3
1
,5-Dibromo-2,4-dihydroxybenzaldehyde (4, CAS
16096-91-4)
7-Hydroxy-6,8-dimethoxy-2-oxo-2H-chromene-3-carboxylic
acid (7, CAS 149143-82-8)
To a stirred solution of 2,4-dihydroxybenzaldehyde
2.8 g, 20 mmol) in ethanol (40 mL) was added bromine
6.4 g, 40 mmol) dropwise at room temperature. After
Meldrum’s acid (0.16 g, 1 mmol) was added to a stirred
solution of aldehyde 5 (0.210 g, 1 mmol) in 2 mL of water
at 80 ºC. After two hours, the reaction mixture was cooled
(
(
o
stirring for 30 min the reaction mixture was poured into
distilled water (100 mL) and filtered. The filtrate was
washed with water and the residual bromine present in the
aqueous phase was quenched with a saturated solution of
sodium thiosulfate. Compound 4 was obtained as a white
powder (5.9 g, 98%), and the physical and spectroscopic
and acidified at 0 C to pH 1 with concentrated sulfuric
acid. After stirring for 1.5 h, the solution was poured
onto crushed ice and warmed to room temperature. The
crystals were filtered, washed with water, and dried to
give coumarin 7 as white crystals (0.25 g, 90%), and the
physical and spectroscopic data are in accordance with
2
5
19
19
data are in accordance with those previously reported.
those previously reported. mp 227-228 ºC (Lit. mp
25
mp 188-189 ºC (Lit. mp 200 ºC); ¹H NMR (400 MHz,
227-228 ºC); ¹H NMR (400 MHz, DMSO-d ) d 3.84 (s,
6
13
methanol-d ) d 7.88 (s, 1H), 9.71 (s, 1H); ¹³C NMR
6H), 4.63 (br s, 2H), 7.26 (s, 1H), 8.66 (s, 1H); C NMR
4
(
100 MHz, methanol-d ) d 103.1, 119.9, 132.0, 138.5,
(100 MHz, DMSO-d ) d 56.2, 60.8, 105.9, 109.5, 112.8,
4
6
1
54.1, 154.7, 196.2.
134.2, 144.7, 145.9, 146.9, 149.6, 157.5, 164.2.
2
1
,4-Dihydroxy-3,5-dimethoxybenzaldehyde (5, CAS
82427-46-9)
7-Hydroxy-6,8-dimethoxy-2H-chromen-2-one (2, isofraxidin,
CAS 486-21-5)
To a stirred solution of 4 (11.8 g, 40 mmol) in
a 2:5 solution of anhydrous methanol in anhydrous
dimethylformamide (70 mL) was added CuCl (0.5 g,
mmol) and NaOMe (21.6 g, 400 mmol). The reaction
mixture was heated under reflux at 100 ºC for 4 h. The
Coumarin 7 (0.32 g, 1.4 mmol) was refluxed for 3.5 h in
a solution of pyridine/ethylene glycol (1.35 mL/1.21 mL).
The solution was cooled, acidified with 2 M HCl and
extracted with dichloromethane. Solvent evaporation
afforded isofraxidin (2) as white crystals (0.25 g, 94%),
and the physical and spectroscopic data are in accordance
5
solvent was then evaporated in vacuo, distilled water
50 mL) was added and the pH adjusted to 2 using
1
9
19
(
with those previously reported. mp 148-149 ºC (Lit.
concentrated HCl. After 15 min the reaction mixture
was transferred into a separation funnel and the product
extracted using chloroform (5 × 20 mL). The organic
mp 144 ºC); ¹H NMR (400 MHz, CDCl ) d 3.86 (s, 3H),
3
4.01 (s,3H), 6.21 (d, J 9.0 Hz, 1H), 6.59 (s, 1H), 7.54 (d,
13
J 9.0 Hz, 1H); C NMR (100 MHz, DMSO-d ) d 56.2,
6
layer was washed with brine, dried with MgSO , filtered
60.8, 105.9, 109.5,112.8, 134.2, 144.5, 145.9, 146.9,
149.6, 164.2.
4
and the solvent evaporated in vacuo. Compound 5 was
obtained as a brown solid (5.5 g, 70%), and the physical and
spectroscopic data are in accordance with those previously
Altissimacoumarin D (1, CAS 1388627-67-5)
2
1
21
reported. mp 88-89 ºC (Lit. mp 87.5-89.8 ºC); ¹H NMR
To a stirring solution of isofraxidin (0.110 g, 0.31 mmol)
in anhydrous tetrahydrofuran (THF, 1.0 mL) was added
triphenylphosphine (0.081 g, 0.31 mmol) and geraniol
(0.05 mL, 0.29 mmol). DIAD (0.05 g, 0.25 mmol) in THF
(
(
400 MHz, DMSO-d ) d 3.86 (s, 6H), 7.29 (s, 1H), 8.68
6
s, 1H); ¹³C NMR (100 MHz, CDCl ) d 56.6, 60.9, 109.0,
3
1
12.9, 134.6, 141.3, 147.1, 151.7, 194.6.
(
0.5 mL) was added dropwise and the reaction mixture
5
-(2,4-Dihydroxy-3,5-dimethoxybenzylidene)-2,2-dimethyl-
,3-dioxane-4,6-dione (6)
Meldrum’s acid (0.16 g, 1 mmol) was added to a stirred
left stirring for 24 hours. The solvent was removed and
the crude product purified by preparative thin layer
chromatography with an eluent of 6% MeOH/CH Cl .
1
2
2
solution of aldehyde 5 (0.210 g, 1 mmol) in 2 mL of water
The product was extracted from the silica with a solution

Vol. 29, No. 5, 2018
Silva et al.
1161
of 10% MeOH/CH Cl , filtered and the solvent evaporated
in vacuo to afford altissimacoumarin D as a colourless
9. Yoshida, M.; Kijima, M.; Akita, M.; Beppu, T.; J. Biol. Chem.
1990, 265, 17174.
2
2
solid (0.10 g, 54%); mp 225-226 ºC; ¹H NMR (400 MHz,
10. Williams, D. E.; Dalisay, D. S.; Li, F.; Amphlett, J.; Maneerat,
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CDCl ) d 1.52 (s, 3H), 1.60 (s, 3H), 1.63 (s, 3H), 1.99 (m,
3
4H), 3.82 (s, 3H), 3.96 (s, 3H), 4.62 (d, J 7 Hz, 2H), 5.00
(
6
t, J 8 Hz, 1H), 5.49 (t, J 8 Hz, 1H), 6.28 (d, J 8 Hz, 1H),
13
.58 (s, 1H), 7.54 (d, J 12 Hz); C NMR (100 MHz, CDCl )
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3
d 16.4, 17.6, 25.6, 26.3, 39.6, 56.2, 61.7, 70.2, 103.5, 114.4,
1
1
15.1, 119.6, 123.8, 131.7, 141.8, 142.5, 143.0, 143.4,
44.9, 150.7, 160.5.
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9
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Supplementary Information
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1
13
Supplementary information ( H and C NMR spectra)
is available free of charge at http://jbcs.org.br as a PDF file.
1
5. Schiedel, M.; Robaa, D.; Rumpf, T.; Sippl, W.; Jung, M.; Med.
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We thank the Brazilian Science without Borders
program at CNPq-UFRJ for a scholarship toAnna C. Silva
and the COST Action TD0905 Epigenetics: From Bench
to Bedside for financial assistance.
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Submitted: August 18, 2017
Published online: February 9, 2018
This is an open-access article distributed under the terms of the Creative Commons Attribution License.