Synthesis and electrochemical studies of new antimalarial endoperoxides

Article abstract of DOI:10.1002/ejoc.200300165

Structural analogues of endoperoxides belonging to the family of G factors have been synthesized under Mannich-type conditions. The structures of the different diastereoisomers have been established from NMR spectroscopic data. Their cathodic peak potentials have been determined by thin layer electrochemistry under potentiostatic conditions, and compared to artemisinin. These endoperoxides were evaluated in vitro against Plasmodium falciparum and showed moderate to good activity. Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003.

Full text of DOI:10.1002/ejoc.200300165

FULL PAPER
Synthesis and Electrochemical Studies of New Antimalarial Endoperoxides
Fadia Najjar,[a] Michel Baltas,[a] Liliane Gorrichon,[a] Yolande Moreno,[a]
[b]
[c]
[a]
´
`
Theodore Tzedakis, Henri Vial, and Christiane Andre-Barres*
Keywords: Biological activity / Electrochemistry / Natural products / Peroxides
Structural analogues of endoperoxides belonging to the fam-
ily of G factors have been synthesized under Mannich-type
conditions. The structures of the different diastereoisomers
have been established from NMR spectroscopic data. Their
cathodic peak potentials have been determined by thin layer
electrochemistry under potentiostatic conditions, and com-
pared to artemisinin. These endoperoxides were evaluated
in vitro against Plasmodium falciparum and showed moder-
ate to good activity.
( Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim,
Germany, 2003)
Introduction
synthetic derivative of yingzhaosu (3),[5] BO7 (5),[6] a cis-
fused cyclopenteno 1,2,4-trioxane, or the cyclic peroxyke-
tals 6 prepared by Posner.[7] The molecule 1,2,4,5-tetraox-
ane 7 (WR148999), although quite different from 2, also
displays antimalarial activity. Recently, B. Meunier de-
scribed synthesis of ‘‘trioxaquines’’[8] 8, hybrid compounds
with both trioxane and 4-aminoquinoline moieties. In all of
those compounds exhibiting good activity (in the nanomo-
lar range) on chloroquine-resistant Plasmodium strains, the
endoperoxide framework seems to be vital for the activity
to occur.
Endoperoxides are important chemical frameworks in the
activity of a number of pharmacologically active com-
pounds. Examples include the natural products chondrillin
(1) and analogues, which show significant antitumor
activity,[1] artemisinin (qinghaosu, 2;[2] Figure 1) and
yingzhaosu (3),[3] both isolated from traditional Chinese
herbal treatments, which are potent antimalarial com-
pounds.
Malaria is a disease caused by parasitic protozoa belong-
ing to the Plasmodium genus, mostly Plasmodium falcipa-
rum. This disease affects some 200Ϫ300 million people
worldwide every year. The anti-malarial properties of non-
alkaloid compounds such as artemisinin 2 and its related
endoperoxides 3 have attracted considerable attention and
these compounds are widely used, as they are one of the few
effective treatments for multi-drug resistant P. falciparum
malaria. These compounds are active against the early form
of the malarial blood stages and induce rapid clearance of
blood parasitemia, and no resistance has been confirmed.
Nevertheless, since malaria parasites are rapidly developing
drug resistance, there is an increasing need for more acces-
sible and more effective drugs. To that goal, a number of
derivatives possessing the endoperoxide framework have
The key steps in the mechanism of antimalarial endo-
peroxide action are contact with haeme iron and electron
transfer (ET) resulting in the cleavage of the OϪO bond.
Oxygen-centered radical intermediates are formed first, and
then carbon-centered radicals, high valent iron, and other
reactive intermediates. All these species may then interact
with so far unidentified vital intercellular target molecules,
through undefined mechanisms, resulting in the death of
the parasite. The cleavage process has been intensively in-
vestigated in various model systems with haeme or non-
haeme iron.[8]
The naturally occurring G factors (G1, G2, G3) consti-
tute a family of six-membered unsaturated cyclic endo-
peroxides fused to another six-carbon ring deriving from
syncarpic acid. The G factors were first isolated from Euca-
been designed and synthesized, including arteflene (4),[4]
a
[a]
`
´
´ ˆ
Synthese et Physicochimie de Molecules d’Interet Biologique, lyptus grandis. They act as phytohormones and seem to be
´
UMR CNRS 5068, Universite Paul Sabatier,
involved in frost resistance and water loss reduction in these
species. They cannot be present in their physiological forms,
but rather in some inactive form, and are readily released
in response to damage to the plant or to a biological stimu-
lus.[9]
118 route de Narbonne, 31062 Toulouse cedex 04, France
Fax: (internat.) ϩ 33-5/61558255
E-mail: candre@chimie.ups-tlse.fr
[b]
[c]
´
´
Laboratoire du Genie Chimique, UMR CNRS 5503, Universite
Paul Sabatier,
118 route de Narbonne, 31062 Toulouse cedex 04, France
´
Dynamique Moleculaires des Interactions Membranaires,
In this paper we report the synthesis of G factor ana-
´
UMR CNRS 5539, Universite Montpellier 2,
cc107, Place E. Bataillon, 34095 Montpellier Cedex 5, France
logues 13Ϫ15, by a previously developed method,[10] and of
Eur. J. Org. Chem. 2003, 3335Ϫ3343
DOI: 10.1002/ejoc.200300165
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C. Andre-Barres et al.
FULL PAPER
Figure 1. Some examples of biologically active endoperoxides
their methylated derivatives 16Ϫ18, together with the deter- formed from reactions between syncarpic acid 9 and alde-
mination of their reduction potentials and evaluation of hydes 10, 11, and 12, in the presence of stoichiometric
their biological activity. Tertiary hydroxy methylated com- amounts of piperidine (Scheme 1). These Mannich bases
pounds were synthesized for two reasons: firstly, from anal- could be isolated in high yield and are stabilised by internal
ogy to Posner’s antimalarial cyclic peroxyketals,[6] which hydrogen bonding. In aqueous acid, rapid piperidine elim-
showed that antimalarial activity is higher when the hemi- ination occurs and the ene-one (in equilibrium with diene-
ketal position is methylated, lower when OH is free and ol) system spontaneously adds oxygen to give the endo-
decreases when OH is alkylated by a too hindered isopropyl peroxide structure. The speed of oxygen uptake is depen-
group, and secondly, in order to evaluate the effect of the dent on the substituents.
methyl group in mimicking the trioxane structure of arte-
misinin.
In view of the possible mode of action of these com-
The starting aldehydes for the endoperoxides 13 and 14
are commercially available, while for endoperoxide 15, 2-
methyl-3-[(tert-butyldiphenylsilyl)oxy]propanal (12) was
pounds, proper consideration of the role of electron transfer
synthesised from 2-methyl-1,3-propanediol in two steps
(ET) in systems possessing a peroxide linkage involved in
[monoprotection followed by Doe¨ring oxidation (Pyr·SO3,
their biological activity requires knowledge of the reduction
DMSO, Et3N, CH2Cl2)] in 75% overall yield.
potentials of the endoperoxides.[11]
For (Ϯ)-2-phenylpropionaldehyde (11), two dia-
Finally, biological results concerning the activity of the
stereomeric products 14a and 14b were obtained in an over-
synthesised peroxides against the human malarial parasite
all yield of 70% (60:40), and could be separated by chroma-
Plasmodium falciparum are discussed.
tography. The structure of each isomer was attributed with
the aid of NMR HSQC (Heteronuclear Single Quantum
Coherence) 1H-13C COSY spectra followed by HMBC
Results and Discussion
(Heteronuclear Multiple Bond Connectivity), giving com-
1
plete assignments of both H and 13C spectra. The relative
The synthesis of the target compounds was undertaken stereochemistry of each isomer and differentiation between
under Mannich-type conditions.[10] Mannich bases were methyl groups in each gem-dimethyl frame was determined
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Synthesis and Electrochemical Studies of New Antimalarial Endoperoxides
FULL PAPER
Scheme 1. Synthesis of endoperoxides 13Ϫ15 and methylated compounds 16Ϫ18
by NOESY spectroscopy. Diastereoisomer 14b was assigned in agreement with the observed NOE effect. Similar results
as the one displaying NOE effects (Figure 2) between the observed for compounds 15a and 15b suggested the same
C15 methyl group and each C11 and C13 methyl, while no conclusions.
effect was observed for diastereoisomer 14a. In this latter
Methylation of the tertiary hydroxyl group was under-
case, a trans relationship between the OH and Ph groups taken. Although this reaction is reported to be efficient in
was observed.
acidic medium (Hϩ/methanol) for other peroxides pos-
Treatment of aldehyde (Ϯ)-12 with syncarpic acid gave, sessing an adjacent tertiary hydroxyl group[13] it was unsuc-
after removal of the amine, a complex mixture that evolved cessful in the present case. Endoperoxide 13 could not be
slowly. After eight days, endoperoxides 15a and 15b were derivatised but rather underwent degradation. Methyl io-
isolated from the crude mixture in 60% yield (75:25) by dide was found to be inert under basic conditions (nBuLi,
silica gel chromatography (Table 1).
MeI). Gratifyingly though, under basic conditions[14] endo-
Similar experiments were undertaken in order to establish peroxide 13 reacted well with methyl triflate to produce the
the relative stereochemistry of endoperoxides 15a and 15b. methylated compound 16 in 70% yield. The same method-
HSQC, HMBC, and NOESY experiments confirmed the ology (BuLi/TfOMe) was used to methylate 14a, 14b, 15a,
same trans relationship between OH and CH2OSi(Ph)2tBu and 15b. All of these reactions afforded methylated com-
for compound 15a, and cis for compound 15b.
pounds Ϫ 17a, 17b, 18a, and 18b, respectively Ϫ in good
Modelling studies were performed on compounds 14a, yields (about 70%). NMR HSQC, HMBC, and NOESY
14b, 15a, and 15b. The optimal conformations obtained experiments were undertaken on the methylated series and
through energy minimization[12] showed spatial proximity confirmed the stereochemistry. NOE effects are observed
between the C11 and C15 methyl groups for 14b (Figure 3), between OMe and the C15 methyl group for compounds
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Figure 2. NOESY experiments for compounds 14a and 14b
Table 1. Yields in the synthesis of endoperoxides issued from syncarpic acid
Endoperoxides
R1
R2
Yield (diastereomeric ratio)
13
Me
Me
Ph
Me
Me
Ph
Me
85%
70% (60:40)
14a
14b
15a
15b
CH2OSi(Ph)2tBu
Me
60% (75:25)
CH2OSi(Ph)2tBu
17a and 18a, but not for 17b and 18b. Typical NOESY
Carbon felt ( France; reference CFT 3000 10), set
experiments are shown for compounds 18a and 18b in Fig- between two glass slides delimiting a compartment with a
ure 4.
volume of a few tens of µL, was used as working electrode.
The choice of carbon was justified by the high cathodic
overvoltage in DMF solvent (ca. 2 V). Because of the small
volume of the thin-layer cell, all the electroactive molecules
present in solution can be transformed during one scan.
There is no mixing between the solution of the thin layer
cell and the solution of the auxiliary electrode compart-
ment, so it is possible to examine the behaviour of products
appearing at the electrode during the potential scan. From
the amount of charge, determination of the number of elec-
trons exchanged during the electrode reaction is straightfor-
ward.
Electrochemical Studies
For consideration of the role of electron transfer in the
possible mode of action of cyclic endoperoxides, accurate
values for their reduction potentials are required. The elec-
trochemical behaviour of endoperoxides 13Ϫ18 was studied
and compared with that of artemisinin, by use of a thin
layer voltammetry method under potentiostatic conditions.
The method used is an original way to determine anodic
peak potentials (Ep) on small quantities and to obtain a
relative and comparative scale between these compounds
and artemisinin in the reduction step of the endoperoxide
function.
All the electrode potentials were measured with respect
to a saturated calomel electrode (SCE; Hg/Hg2Cl2/ClϪ satu-
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Synthesis and Electrochemical Studies of New Antimalarial Endoperoxides
FULL PAPER
The electrochemical cell was kept under an inert atmos-
phere (nitrogen at 1.5 atm) and all experiments were per-
formed in the absence of oxygen.
Current-potential curves were typically obtained with en-
doperoxides at a 0.01  concentration with use of 0.3
Ϫ
mol·LϪ1 of NEt4ϩClO4 as electrolyte in DMF solvent,
and are shown in Figure 5.
Table 2 shows the potential of various signals at the ca-
thodic part (first scan). For all compounds (scan rate r ϭ
0.5 mV·sϪ1), an intense cathodic signal appeared in the
Ϫ1.4 to Ϫ1.8 V range. Integration of the peak showed one
electron exchange, suggesting that a radical anion appears
at the electrode[15] according to the equation:
AB ϩ eϪ Ǟ Ao ϩ BϪ
It is also noteworthy that, for every compound studied,
a strong shift between the zero current potential (Ϫ0.1 to
Ϫ0.2 V) and the potential of the peak of reduction of the
endoperoxide (Ϫ1.4 to Ϫ1.8 V) was observed, indicating the
presence of slow redox systems (Table 2).
The inversion of the potential scan rate after the end of
the cathodic plotting for each endoperoxide was also re-
corded (Figure 5). It showed that there is no anodic signal
in the potential range from Ϫ1.7 to Ϫ1.2 V, in accordance
with the presence of slow redox systems. Several signals are
located at potentials higher than ca. Ϫ0.8 V, indicating that
Figure 3. Energetically optimal conformations through modelling
studies of compounds 14a and 14b
rated), immersed in a Luggin capillary located close to the reduction of the endoperoxide produces several by-
(3Ϫ4 mm) the working electrode and containing DMF and products, the oxidation of which is observed.
the electrolyte in large excess. The auxiliary electrode was
Supplementary reverse scanning provided cathodic
made of platinum. The electrochemical apparatus used was curves not including any signals in the range from Ϫ1.3 to
a Radiometer Voltalab PGZ 100 computerised potentiostat. Ϫ1.8 V; the absence of endoperoxide indicates the irreversi-
Figure 4. NOESY experiments for compounds 18a and 18b
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Figure 5. Current-potential curves obtained with carbon felt enclosed in a thin layer cell with various endoperoxides at a concentration
Ϫ
of 0.01  in 0.3  NEt4ϩClO4 in DMF; temp. ca. 20 °C; r ϭ 0.5 mV·sϪ1; plotted under 1.5 atm of N2 unless otherwise indicated.
Artemisin cathodic (1: VTL: 52 µL) and anodic parts (2: VTL: 54 µL); curves obtained separately; 13: VTL: 50 µL; plotted order 1, 2, 3.
14a: 1: residual anodic and cathodic curve, plotted separately in atmospheric air; VTL: 47 µL; 2: residual cathodic curve; VTL: 49 µL; 3:
VTL: 51 µL. 14b: VTL: 37µL. 15a: VTL: 58 µL. 15b: VTL: 70 µL. 16: 1: Residual cathodic curve; VTL: 39µL; 2: VTL: 39 µL. 17a: VTL: 57
µL. 17b: VTL: 48 µL. 18a: VTL: 59 µL
bility of the reduction process under these experimental (16, 17a, 18b) presented redox potentials similar to that of
conditions.
artemisinin, while for the others, the peroxide linkage was
The electrochemical results showed that the Ep values of easily cleaved. The Ep value of artemisinin found by thin
methylated endoperoxides (16Ϫ18b) are lower than those of layer voltammetry in our study is similar to that obtained
the hydroxylated molecules (13Ϫ15b). Some compounds by Workentin.[11] Nevertheless, we have to specify that com-
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Synthesis and Electrochemical Studies of New Antimalarial Endoperoxides
FULL PAPER
Table 2. Potentials of the peaks of the various endoperoxides
13Ϫ18b, measured on curves obtained by use of an electrochemical
thin layer cell and carbon felt as working electrode; all experimental
conditions are as indicated in the caption for Figure 5
Table 3. In vitro antimalarial activity of endoperoxides
Endoperoxides
IC50 (µ)[a]
Chloroquine
13
0.030
20Ϯ10
20Ϯ10
27Ϯ3
0.7Ϯ0.2
0.44Ϯ0.06
0.28Ϯ0.14
2.0Ϯ1.0
0.7Ϯ0.3
1.5Ϯ0.5
1.4Ϯ0.4
Endoperoxides
Ep /V vs. SCE (Ϯ10 mV)
(cathodic part: first scan)
14a
14b
15a
15b
16
17a
17b
18a
18b
Artemisin
13
Ϫ1.68
Ϫ1.50
Ϫ1.49; Ϫ2.16 (solvent)
Ϫ1.45
Ϫ1.60
14a
14b
15a
15b
Ϫ1.52
16
Ϫ1.76; Ϫ2.06 (solvent)
Ϫ1.68; Ϫ2.09 (solvent)
Ϫ1.55
Ϫ1.70
not performed
17a
17b
[a] IC50 values were considered acceptable when values did not vary
by more than a factor of three.
18a
18b
belonging to the family of G factors. The synthesis is easy
and modulable and gives an original route to the endo-
peroxide bridge; it was developed using syncarpic acid ob-
tained by synthesis, although this is a natural product that
can also be extracted from Myrtaceae. The formation of the
peroxide bridge is biomimetic and occurs through spon-
taneous oxygen uptake by precursors without any sensitizer,
under mild conditions and whatever the substituent. This
step is very clean, with good yields and no by-products.
This class of compounds possesses electrochemical proper-
ties approaching those of artemisinin, and clear antima-
larial activity over quite a large range of concentrations,
from 0.2 to 30 µ. Lipophilicity appears to be another rel-
evant parameter besides redox ability for antimalarial ac-
tivity in this series. In view of the urgent need for novel
classes of antimalarials, and the potential of endoperoxide-
based ones, this series of low-molecular-weight compounds
could be of interest for the future, especially if structural
modifications on substituents R1 and R2 were able to im-
prove the antimalarial activity.
parison between Ep values can be only qualitative and some
discrepancies in Ep values can be noted in the litera-
ture,[11,16] indicating the influence of the operating con-
ditions used for the Ep determination in irreversible systems
(technique, scan rate, solvent, electrolyte). It is also note-
worthy that in irreversible systems the potential of the peaks
can be overrated from the corresponding standard potential
E°. In fact, Workentin[11] showed that artemisinin un-
dergoes an irreversible dissociative reduction by cyclic vol-
tammetry with an anodic peak potential (Ep) that varies
with scan rate, being Ϫ1.68 V vs. SCE at 1 V/s. The stand-
ard potential E° of artemisinin was also determined by
these authors, for the first time, as Ϫ0.89 V vs. SCE in
DMF, so direct electrochemical reduction of artemisinin is
subject to a large activation overpotential. Consequently,
a rigorous comparison requires knowledge of the standard
potential E° of the dissociative reduction of these com-
pounds, the determination of which is the object of an on-
going study.
Biological Studies
The compounds were tested in vitro against the Nigerian
strain of P. falciparum. All of them exerted potent inhi-
bition on the growth of this human parasite. Inhibition was
complete and occurred in a narrow range of concentrations,
covering less than two on a log-scale of concentration. The
IC50 values reported in Table 3 indicate drug concentrations
reducing cell growth by up to 50% of that of untreated con-
trol after one cycle. They are in the range of 0.2 to 30 µ.
Compounds with high lipophilicity were the most active.
The activity of the methylated analogues 16, 17a, and 17b
was enhanced by an order of ten in relation to compounds
13, 14a, and 14b. Methylation of the tertiary hydroxyl group
of the quite lipophilic compounds 15a and 15b gave com-
pounds (18a and 18b) with similar activities.
Experimental Section
General Remarks: The solvents were dried by standard procedures,
distilled, and stored under argon until used. THF and Et2O were
freshly distilled on sodium/benzophenone. CH2Cl2 was distilled
from CaH2; DMF was dried and distilled from CaH2. Commer-
cially available aldehydes were used after purification. Artemisinin
was obtained from Aldrich and used as received. All reactions were
monitored by thin-layer chromatography (TLC) carried out on alu-
minium-backed 0.2 mm silica gel 60 plates with fluorescent indi-
cator UV254 and/or by developing the plate with a solution of phos-
phomolybdic acid. HPLC chromatography was carried out on a
JobinϪYvon apparatus on Merck 15µm or Amicon silica (6Ϫ35
µm). NMR spectra were recorded at 250 MHz and 400 MHz for
1H and 62.8 MHz and 100.6 MHz for 13C in CDCl3 solutions with
TMS as internal reference. Mass spectra were obtained on a Ner-
mag R10Ϫ10 mass spectrometer. Elemental analyses were per-
Conclusion
´
formed at the Ecole Nationale Superieure de Chimie de Toulouse.
We have synthesised a number of new endoperoxides pos-
1-Hydroxy-4,8,8,10,10-pentamethyl-4-phenyl-2,3-dioxabicyclo-
sessing a peroxyhemiketal or peroxyketal framework and [4.4.0]dec-5-ene-7,9-dione (14): A solution of syncarpic acid (9,
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FULL PAPER
0.198 g, 1.09 mmol) and piperidine (0.092 g, 1.09 mmol) in di-
chloromethane (10 mL) was added at 25 °C to a stirred solution of
(400 MHz, CDCl3): δ ϭ 7.57Ϫ7.61 (m, 4 H, Ar), 7.39Ϫ7.41 (m, 6
H, Ar), 7.12 (s, 1 H, ϭCH), 3.72, 3.65 (AB system, JAB ϭ 11.32 Hz,
2-methylpropanal (10, 0.148 g, 1.09 mmol) and piperidine (0.046 g, 2 H, CH2OSi), 1.36, 1.35, 1.33, 1.31, 1.02 (5s, 15 H, 5 ϫ CH3),
0.545 mmol) in dichloromethane (10 mL). After 40 min of stirring,
all volatile materials were removed under reduced pressure. The
solid residue was dissolved in dichloromethane and agitated vigor-
ously with an aqueous solution of 1  HCl /NH4Cl (saturated) for
10 min. The organic layer was then separated and dried (MgSO4),
and the solvents were evaporated under reduced pressure. The yield
of the residual product was nearly quantitative. It was then dis-
0.97 [s, 9 H, C(CH3)3] ppm. 13C NMR (100.64 MHz, CDCl3) δ ϭ
210.6 (C9), 197. 7 (C7), 140.4 (C5), 135.8; 135.6, 130.3, 130.2,
128.5, 128.2 (C-CH of phenyl), 132.5, 132.3 (C-CH of phenyl),
134.1 (C6), 97.5 (C1), 83.5 (C4), 67.2 (C16), 55.2 (C8), 51.4 (C10),
26.9 [C(CH3)3], 26.7 (C14), 23.8 (C13), 20.7 (C11), 19.4 (C15), 19.3
[C(CH3)3], 15.3 (C12) ppm. IR (neat): ν˜ ϭ 3541 (OϪH), 1688 (Cϭ
O), 1637 (CϭO), 1109 (SiϪO), 823 (OϪO) cmϪ1. MS (DCI /NH3):
solved in dry chloroform and left for 3 days under air. Evaporation m/z (%) ϭ 540 [MNH4]ϩ (100), 523 [M ϩ1]ϩ (4), 522 [M] (9).
of solvent gave a solid (mixture of 14a and 14b), which was purified
1-Methoxy-4,4,8,8,10,10-hexamethyl-2,3-dioxabicyclo[4.4.0]dec-5-
ene-7,9-dione (16): nBuLi (82 µL, 0.13 mmol) was added slowly,
under argon at Ϫ78 °C, to a stirred solution of 13 (35 mg,
by HPLC (eluent petroleum ether/CH2Cl2/ethyl acetate, 60:32:8),
yielding 14a and 14b (70%; 60:40).
1-Hydroxy-4,8,8,10,10-pentamethyl-4-phenyl-2,3-dioxabicyclo-
0.13 mmol) in THF (3 mL), and the mixture was allowed to warm
[4.4.0]dec-5-ene-7,9-dione (14a): White powder; M.p. 160Ϫ162 °C.
up to Ϫ20 °C. After 15 min, TfOMe (24.6 mg, 0.15 mmol) was ad-
1H NMR (250 MHz, CDCl3): δ ϭ 7.67 (s, 1 H, CϭCH), 7.34 (m, ded at Ϫ20 °C. The reaction mixture was stirred for 30 min and
5 H, Ar), 3.80 (s, OH), 1.64 (s, 3 H, CH3 in position 15), 1.41 (s, 3 was then extracted with diethyl ether. The extract was dried
H, CH3 in position 14), 1.37 (s, 3 H, CH3 in position 13), 1.24 (s,
3 H, CH3 in position 12), 0.74 (s, 3 H, CH3 in position 11) ppm.
(MgSO4) and concentrated to give 16 as an oil. Purification by
silica gel chromatography (eluent petroleum ether/ethyl acetate, 9:1)
1
13C NMR (50 MHz, CDCl3): δ ϭ 210.7 (C9), 198.6 (C7), 141.6 gave 16 (26 mg, 70%). H NMR (250 MHz, CDCl3): δ ϭ 7.35 (s, 1
(C5), 141.4, 128.6, 128.1, 125.6 (Ar); 132.4 (C6), 97.5 (C1), 82.3 H, C-CH), 3.44 (s, 3 H, OCH3), 1.45, 1.35, 1.32, 1.28, 1.27, 1.03
(C4), 55.0 (C8), 51.9 (C10), 26.7 (C14), 26.2 (C15), 24.3 (C13), 20.6
(6s, 18 H, 6 ϫ CH3) ppm. 13C NMR (50 MHz, CDCl3), δ ϭ 210.5
(C9), 198.9 (C7), 145.7 (C5), 128.2 (C6), 100.3 (C1), 78.8 (C4), 54.7
(O-CH3), 53.1 (C10), 25.9, 24.7, 23.8, 23.5, 21.6, 15.8 (6 ϫ CH3)
ppm. MS (DCI/NH3): m/z (%) ϭ 300 [MNH4ϩ] (100), 283 [M ϩ1]ϩ
(13), 210 (67).
ϩ
(C11), 15.0 (C12) ppm. MS (DCI/NH3): m/z (%) ϭ 348 [MNH4
]
(45), 365 (12), 269 (100), 304 (95), 313 (39). C19H22O5 (330.4):
calcd. C 69.07, H 6.71; found C 69.10, H 6.12.
1-Hydroxy-4,8,8,10,10-pentamethyl-4-phenyl-2,3-dioxabicyclo-
[4.4.0]dec-5-ene-7,9-dione (14b): White powder; M.p. 159Ϫ160 °C.
1-Methoxy-4,8,8,10,10-pentamethyl-4-phenyl-2,3-dioxabicyclo-
1H NMR (250 MHz, CDCl3): δ ϭ 7.46 (s, 1 H, CϭCH), 7.40 (m, [4.4.0]dec-5-ene-7,9-dione (17a): Same procedure as for 16; yield
5 H, Ar), 3.70 (s, OH), 1.87 (s, 3 H, CH3 in position 15), 1.39 (s, 6 80%. 1H NMR (250 MHz, CDCl3): δ ϭ 7.67 (s, 1 H, CϭCH), 7.30
H, CH3 in position 13Ϫ14), 1.36 (s, 3 H, CH3 in position 12), 1.13
(s, 3 H, CH3 in position 11) ppm. 13C NMR (50 MHz, CDCl3): 1.32, 1.17, 0.60 (4s, 12 H, 4 ϫ CH3) ppm. 13C NMR (50 MHz,
δ ϭ 210.6 (C9), 198.1 (C7), 141.9 (C5), 137.7, 129.4, 129.0, 126.4 CDCl3): δ ϭ 210.5 (C9), 199.3 (C7), 143.9 (C5), 141.9 (C6), 129.4,
(Ar), 131.5 (C6), 97.6 (C1), 82.6 (C4), 55.1 (C8), 51.9 (C10), 26.6, 128.4, 127.9, 125.4 (Ar), 100.3 (C1), 81.8 (C4), 54.8 (C8), 54.6 (O-
(m, 5 H, Ar), 3.53 (s, 3 H, OCH3), 1.61 [s, 3 H, CH-CH3(Ph)], 1.33,
24.0 (C13, C14), 23.3 (C15), 21.0 (C11), 15.1 (C12) ppm. MS (DCI/
NH3): m/z (%) ϭ 348 [MNH4ϩ] (4), 330 [M] (4), 299 (100).
C19H22O5 (330.4): calcd. C 69.07, H 6.71; found C 69.27, H 6.49.
CH3), 53.4 (C10), 26.4, 26.1, 24.9, 21.1, 15.4 (5 ϫ CH3) ppm.
1-Methoxy-4,8,8,10,10-pentamethyl-4-phenyl-2,3-dioxabicyclo-
[4.4.0]dec-5-ene-7,9-dione (17b): Same procedure as for 16; yield
67%. 1H NMR (250 MHz, CDCl3): δ ϭ 7.70 (s, 1 H, CϭCH), 7.44
(m, 5 H, Ar,), 3.44 (s, 3 H, OCH3), 1.85 [s, 3 H, CH-CH3(Ph)],
1.37, 1.34, 1.31, 1.15 (4s, 12 H, 4 ϫ CH3) ppm. 13C NMR (50 MHz,
CDCl3): δ ϭ 210.7 (C9), 198.6 (C7), 145.1 (C5), 138.2 (C6), 128.6,
129.1, 129.0, 126.2, 125.4 (Ar), 100.9 (C1), 81.9 (C4), 54.9 (C8),
53.5 (C10), 54.9 (O-CH3), 26.0, 24.8, 23.7, 21.7, 15.6 (5 ϫ CH3)
ppm. MS (DCI/NH3): m/z (%) ϭ 362 [MNH4]ϩ (46), 313 (100),
330 (21).
4-(tert-Butyldiphenylsilyloxymethyl)-1-hydroxy-4,8,8,10,10-penta-
methyl-2,3-dioxabicyclo[4.4.0]dec-5-ene-7,9-dione (15): A solution
of syncarpic acid (9, 0.198 g, 1.09 mmol) and piperidine (0.046 g,
0.545 mmol) in dichloromethane (10 mL) was added to a solution
of aldehyde 12 (0.356 g, 1.09 mmol) and piperidine (0.92 g,
1.09 mmol) in dichloromethane (10 mL). By the workup described
above, we obtained 15a and 15b (60%; 75:25) after purification (elu-
ent petroleum ether/diethyl ether, 7:3).
1
Compound 15a: White powder; M.p. 157 °C. H NMR (250 MHz,
4-(tert-Butyldiphenylsilyloxymethyl)-1-methoxy-4,8,8,10,10-
CDCl3): δ ϭ 7.76Ϫ7.58 (m, 4 H, Ar), 7.44Ϫ7.28 (m, 6 H, Ar), 7.22 pentamethyl-2,3-dioxabicyclo[4.4.0]dec-5-ene-7,9-dione (18a): Same
(s, 1 H, CϭCH), 3.84, 3.73 (2 H, AB system JAB ϭ 10.2 Hz, CH2-
procedure as for 16, yield 71%. 1H NMR (400 MHz, CDCl3) : δ ϭ
O-Si), 3.56 (s, 1 H, OH), 1.50 {s, 3 H, CH-CH3[CH2OSi(Ph)2]}, 7.67Ϫ7.59 (m, 4 H, Ar), 7.40 (s, 1 H, ϭCH), 7.39Ϫ7.32 (m, 6 H,
1.35, 1.33, 1.24, 0.67 (4s, 12 H, 4 ϫ CH3), 1.09 [s, 9 H, C-(CH3)3] Ar) 3.77, 3.67 (AB system, JAB ϭ 10.2 Hz, 2 H, CH2OSi), 3.43 (s,
ppm. 13C NMR (50.28 MHz, CDCl3): δ ϭ 210.6 (C9), 197.8 (C7), 3 H, CH3O), 1.47 1.27 1.24 1.19 (4s, 4 ϫ CH3), 1.06 [s, 9 H,
140.4 (C5), 135.8, 135.6, 129.9, 127.8 (C-CH of phenyl), 132.9,
C(CH3)3], 0.60 (s, 3 H, CH3) ppm. 13C NMR (100.64 MHz,
132.6 (C-CH of phenyl), 133.1 (C6), 97.2 (C1), 82.1 (C4), 65.7 CDCl3): δ ϭ 210.5 (C9), 198.5 (C7), 143.2 (C5), 135.9, 135.7, 130.1,
(CH2-O), 54.9 (C8), 51.5 (C10), 26.8 [C-(CH3)3], 19.3 [C-(CH3)3], 128.0 (C-CH of phenyl), 132.7, 133.2 (C-CH of phenyl), 129.8 (C6),
24.1, 20.8, 19.1, 15.1 (5 ϫ CH3) ppm. C30H38SiO6 (522.7): calcd.
C 68.93, H 7.33, O 18.36; found C 68.74, H 7.29. IR (KBr): ν˜ ϭ
100.3 (C1), 81.6 (C4), 65.7 (C16), 55.8 (C8), 55.0 (OCH3), 53.7
(C10), 27.0 [C(CH3)3], 26.1 (C13), 24.9 (C14), 21.6 (C11), 19.5
3525 (OϪH), 1690 (CϭO), 1635 (CϭO), 1113 (SiϪO), 824 (OϪO) [C(CH3)3], 19.1 (C15), 15.7 (C12) ppm. IR (neat): ν˜ ϭ 1691 (Cϭ
cmϪ1. MS (DCI /NH3): m/z (%) ϭ 540 [MNH4]ϩ (69), 344 (24),
330 (100).
O), 1640 (CϭO), 1108 (SiϪO), 823 (OϪO) cmϪ1. MS (DCI /NH3):
m/z (%) ϭ 540 [MNH4]ϩ (36), 522 [M] (2), 299 (100).
4-(tert-Butyldiphenylsilyloxymethyl)-1-hydroxy-4,8,8,10,10-penta-
methyl-2,3-dioxabicyclo[4.4.0]dec-5-ene-7,9-dione (15b): 1H NMR
4-(tert-Butyldiphenylsilyloxymethyl)-1-methoxy-4,8,8,10,10-penta-
methyl-2,3-dioxabicyclo[4.4.0]dec-5-ene-7,9-dione (18b): Same pro-
3342
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
Eur. J. Org. Chem. 2003, 3335Ϫ3343

Synthesis and Electrochemical Studies of New Antimalarial Endoperoxides
FULL PAPER
1
J. Mareda, P. Millason-Schmidt, G. Brunner, V. Burger, Helv.
cedure as for 18a; yield 74%. H NMR (250 MHz, CDCl3) : δ ϭ
Chim. Acta 1996, 79, 1475Ϫ1487. [2b] J. N. Cummings, P. Ployp-
7.69Ϫ7.62 (m, 4 H, Ar), 7.47 (s, 1 H, ϭCH), 7.45Ϫ7.35 (m, 6 H,
Ar), 3.71, 3.69 (AB system, JAB ϭ 11.1 Hz, 2 H, CH2OSi), 3.41 (s,
3 H, OCH3), 1.43, 1.36, 1.32, 1.30, 1.06 (5s, 5 ϫ CH3), 1.04 [s, 9
[2c]
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R.
A.
[2d]
H, C(CH3)3] ppm. 13C NMR (100.64 MHz, CDCl3): δ ϭ 210.6
[3]
(C9), 198.6 (C7), 143.7 (C5), 135.8, 135.7, 130.2, 130.1, 128.1,
128.0, (C-CH of phenyl), 132.6, 132.5 (C-CH of phenyl), 129.8
(C6), 100.7 (C1), 82.6 (C4), 67.2 (C16), 55.0 (C17), 54.9 (C8), 53.3
(C10), 26.9 [C(CH3)3], 26.1 (C14), 24.9 (C13), 20.8 (C11), 19.6
[4]
(C15), 19.3 [C(CH3)3], 15.7 (C12) ppm. IR (neat): ν˜ ϭ 1691 (Cϭ
[5]
O), 1638 (CϭO), 1102 (SiϪO), 819 (OϪO) cmϪ1. MS (DCI /NH3):
M. D. Bachi, E. E. Korshin, R. Hoos, A. Splizman, J. Hetero-
cyclic Chem. 2000, 37, 639Ϫ646.
m/z (%) ϭ 540 [MNH4]ϩ (100).
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In vitro Antimalarial Activity: The Nigerian strain of Plasmodium
falciparum was maintained in culture by the method of Trager and
Jensen.[17] The activity of the compounds against the in vitro
growth of P. falciparum was determined according to Desjardins et
al.,[18] by use of [3H]hypoxanthine incorporation as an assessment
of parasite growth. Stock drug solutions were prepared in 100%
dimethyl sulfoxide (DMSO) at concentration of 10Ϫ40 m and
were serially diluted to the appropriate concentration with com-
plete medium. Assays were performed in sterile 96-well microtiter
plates. The final volume in each well was 200 µL, consisting of 50
µL of complete medium (RPMI 1640 containing 25 m HEPES-
buffer, pH 7.4 and 10% ABϩ human serum) either without (con-
trols) or with drug and 150 µL of P. falciparum-infected erythrocyte
suspension (1.5% final hematocrit and 0.6% parasitemia). After
48 h incubation at 37 °C, 30 µL of complete medium containing
0.7 µCi [3H]hypoxanthine were added to each well. After 18 h at 37
°C, the cells were lysed by use of an automatic cell harvester and
the parasite macromolecules, including radioactive nucleic acids,
were harvested onto glass fibre filters. The filters were counted for
radioactivity in a liquid scintillation spectrometer. The radioac-
tivity background was obtained from incubation of non-infected
erythrocytes under the same conditions, and deducted. Each drug
was tested in triplicate at least twice with different batches of cells
and parasite growth was compared to that in control wells Ϫ rep-
resenting 100% parasite growth. Parasitic viability was expressed as
IC50, which is the drug concentration causing 50% parasite
growth inhibition.
Jefford, S. Kohmoto, D. Jaggi, G. Timari, J.-C. Rossier, M. Ru-
´
daz, O. Barbuzzi, D. Gerard, V. Berger, P. Kamalaprija, J. Ma-
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fields (CVFF) were used. Energetically optimised confor-
mations were determined by sampling structures during molec-
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the NVT method, with a 1fs time step. From the energy/time
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Acknowledgments
We thank Christiane Vidal for molecular modelling assistance, and
Sharon Wein and Marjorie Maynadier for drug testing. Financial
support by the MESR and CNRS is gratefully acknowledged.
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Received March 11, 2003
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Eur. J. Org. Chem. 2003, 3335Ϫ3343
www.eurjoc.org
 2003 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
3343