Synthesis, stability, and antimalarial activity of new hydrolytically stable and water-soluble (+)-deoxoartelinic acid

Article abstract of DOI:10.1021/jm020244p

(+)-Deoxoartelinic acid (13), a new hydrolytically stable, water-soluble, and potent non-acetal-type antimalarial drug candidate, was successfully prepared from artemisinic acid by using sulfur ylide and photooxygenative cyclization in seven steps. This c

Full text of DOI:10.1021/jm020244p

4940
J . Med. Chem. 2002, 45, 4940-4944
Syn th esis, Sta bility, a n d An tim a la r ia l Activity of New Hyd r olytica lly Sta ble a n d
Wa ter -Solu ble (+)-Deoxoa r telin ic Acid
Mankil J ung,*,† Kyunghoon Lee,† Howard Kendrick,‡ Brian L. Robinson,§ and Simon L. Croft‡
Department of Chemistry, Yonsei University, Seoul 120-749, Korea, Department of Infectious and Tropical Diseases, London
School of Hygiene and Tropical Medicine, Keppel Street, London WC1E 7HT, U.K., and Centre for Tropical Antiprotozoal
Chemotherapy, Northwick Park Institute for Medical Research, Watford Road, Harrow, Middlesex, HA1 3UJ , U.K.
Received J une 3, 2002
(+)-Deoxoartelinic acid (13), a new hydrolytically stable, water-soluble, and potent non-acetal-
type antimalarial drug candidate, was successfully prepared from artemisinic acid by using
sulfur ylide and photooxygenative cyclization in seven steps. This compound showed superior
in vitro antimalarial activity against the chloroquine-resistant K1 strain of Plasmodium
falciparum and higher suppression (98.7%) than arteether in vivo against Plasmodium chabaudi
infected mice. (+)-Deoxoartelinic acid also showed remarkable stability with a half-life of 258.66
h, 23 times more stable than clinically useful arteether in simulated stomach acid, and improved
solubility, 4 times more soluble than artemisinin in water.
In tr od u ction
received attention owing to their better bioavailability
and acid stability and their reduced neurotoxicity
compared to acetal-type analogues. Deoxoartemisinin
(2), prepared from either artemisinin (1) or artemisinic
acid (6) by J ung et al.,18 is the first non-acetal-type
analogue of artemisinin and shows more antimalarial
activity than artemisinin (1) both in vitro and in vivo.19
Thus, we designed hydrolytically stable and water-
soluble deoxoartelinic acid 13 as a C-12 non-acetal type
that is expected not only to be considerably more stable
in simulated stomach acid than clinically useful acetal-
type drugs but also to be active against P. falciparum
in vitro and in vivo. In this paper, we report the first
preparation of deoxoartelinic acid 13 and its exception-
ally high in vitro and in vivo antimalarial activity, acid
stability, and water solubility.
Artemisinin (1), a sesquiterpene lactone with the first
natural 1,2,4-trioxane, isolated from Artemisia annua
L, and its derivatives have been important as antima-
larial drugs having the most effective activity against
multidrug-resistant forms of Plasmodium falciparum.1
Artemisinin has been the subject of a number of
reviews1-10 because of its novel molecular structure,
clinically useful antimalarial activity, and interesting
in vitro anticancer activity.11-14 The rapid action,
powerful effect, and good tolerance of artemisinin and
its derivatives may be attributed to the 1,2,4-trioxane
moiety in these molecules (Chart 1). Such a novel
chemical structure from natural resources and interest-
ing bioactivities has encouraged chemists and pharma-
cologists to attempt further exploration.
Most semisynthetic derivatization has involved re-
placing the C-12 acetal functionality, forming the first
generation ether derivatives, most of which were poorly
soluble and hydrolytically unstable in simulated stom-
ach acid. Furthermore, evidence that acetal-type ana-
logues at C-12 are more neurotoxic in animal studies
than non-acetal-type analogues is also emerging15 and
may thus lead to the future abandonment of the
currently clinically used acetal-type analogues (arte-
misinin, artemether, arteether, artesunate, and artelinic
acid). Artelinic acid (5) was prepared to improve on
artesunate (4), which has poor stability in aqueous
solution because of the facile hydrolysis of ester linkage,
but in previously in vitro antimalarial tests against the
cloned D6 and W2 strains of Plasmodium falciparum,
it was 1.4-2.1 times less potent than artemisinin.16,17
Although many derivatives of artemisinin have been
prepared and show good antimalarial activities,2 most
of them possess an acetal group at the C-12 position.
Non-acetal-type analogues of deoxoartemisinin recently
Ch em istr y
Since the preparation of 12-n-butyldeoxoartemisinin
(3c) as the first hydrolytically stable non-acetal type
analogue20 containing a C-C bond at C-12, a series of
non-acetal-type derivatives including a few of heteroaryl
and unsaturated substituents at C-12 have been
prepared.21-29 Because direct introduction of the C-C
bond at C-12 of artemisinin for the preparation of novel
analogues may cause destruction of the biologically
essential endoperoxide, photooxygenative cyclization20
of the dihydroartemisinol derivative (11) has been
utilized as a key step.30 Deoxoartelinic acid 13 was
prepared from more abundant artemisinic acid (6) as a
useful chiral synthon in seven steps as shown in Scheme
1. Thus, reaction of dihydroartemisinyl aldehyde (9),
prepared from artemisinic acid (6) by a known proce-
dure31 (yield 88%), with trimethylsulfur iodide as a
sulfur ylide gave the epoxide 10 (12R/S ) 7/1) (yield
76%). The ring opening of epoxide 10 with 4’-vinylbenz-
ylmagnesium chloride afforded alcohol 11 (yield 81%)
with introduction of the C-C bond at C-12. Photooxy-
genative cyclization, as previously mentioned,18 of al-
cohol 11 provided the 4’-vinylhomobenzyldeoxoartemisi-
nin 12 (yield 35%) with natural â-configuration (J )
* To whom correspondence should be addressed. Phone: +82-2-
2123-2648. Fax: +82-2-364-7050. E-mail: mkjung@alchemy.yonsei.ac.kr.
† Yonsei University.
‡ London School of Hygiene and Tropical Medicine.
§ Northwick Park Institute for Medical Research.
10.1021/jm020244p CCC: $22.00 © 2002 American Chemical Society
Published on Web 09/10/2002

(+)-Deoxoartelinic Acid
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 22 4941
Ch a r t 1
Sch em e 1a
F igu r e 1. Kinetics of hydrolysis and decomposition of acetal
(C-O) type and non-acetal-type prodrugs of artemisinin in
simulated stomach acid (0.01 N HCl, pH 2.0, 37 °C).
form.32 Therefore, we are still in urgent need of acid-
stable and water-soluble antimalarial drugs from the
series for oral and parenteral application. In bioavail-
ability studies of orally administered arteether, for
example, one should consider the effect of gastric pH
and emptying time in the design of the study. To
examine the effect of molecular structure of acetal- and
non-acetal-type prodrugs of artemisinin on the stability
in simulated stomach acid (pH 2.0, 37 °C), 12-(n-butyl)-
deoxoartemisinin20,34 (3c), deoxoartemsinin18,19 (2), and
artelinic acid16 (5) have been prepared as comparative
references by known procedures from either artemisinin
or artemisinic acid.16,18-20 To measure the stability of
deoxoartelinic acid 13 and the other prodrugs in simu-
lated stomach acid, the disappearance of prodrugs (1
mg/mL) on 0.01 N HCl, pH 2.0 at 37 °C, was followed
by HPLC analysis. All prodrugs tested were found to
disappear with pseudo-first-order kinetics, with a half-
life of 10.98, 13.11, 23.5 h for arteether, artelinic acid,
and artemisinin, respectively, and of 186.89, 244.97, and
258.66 h for 12-(n-butyl)deoxoartemisinin, deoxoarte-
misinin, and deoxoartelinic acid, respectively (Figure 1
and Table 1). In general, it was found that the half-
lives of non-acetal-type analogues were 15-23 times
longer than those of acetal-type analogues under simu-
lated stomach acid condition (Table 1). In the simulated
stomach acid, artemisinin (1), arteether (3b), and arte-
linic acid (5) were found to be unstable and to easily
undergo hydrolysis. On the other hand, deoxoartelinic
acid 13 and its non-acetal-type derivatives were found
to be very stable and to possess long half-lives under
the same conditions, showing sufficient stability for oral
administration. Deoxoartelinic acid 13 and its sodium
salt are 4 and 6 times more soluble in water (5.0 mg/
a
Reagents and conditions: (a) CH2N2, Et2O, room temperature
for 3 h; (b) NaBH4, NiCl2‚6H2O, MeOH, 0 °C for 30 min; (c) DIBAL-
H, CH2Cl2, -78 °C for 1 h; (d) (CH3)3S+I-, KOH, DMSO, room
temperature for 2 h, 76%; (e) Mg, 4-vinylbenzy chloride, Et2O, 0
°C for 12 h, 81%; (f) (i) O2, rose bengal, 500 W tungsten lamp,
CH2Cl2/CH3CN (1/9), -40 °C for 4 h, (ii) TFA, O2, CH3CN, -40 °C
for 2 h, 35%; (g) NaHCO3, KMnO4, acetone, room temperature for
1 h, 1 M HCl, room temperature for 5 h, 83%.
10.9 Hz) in a single step. No R-diastereomer at C-12 was
detected. Although the yield for the photooxidative
cyclization is low, this step represents the shortest
synthetic route to compound 12. Direct oxidation of 12
with KMnO4 afforded deoxoartelinic acid 13 (yield 83%)
in a single step.31,40
Sta bility
Artemisinin is stable in neutral solvent heated to 150
°C.32 However, being a hemiacetal, dihydoroartemisinin
(3a ) is chemically more vulnerable than its parent
compound artemisinin (1), a lactone itself. Artemether,
arteether, artesunate, and artelinic acid, which are
acetal-type prodrugs under clinical use or trials are
susceptible to moisture and acidic conditions.32,33 From
published data on other arteether and artesunate
analogues, it would appear that any derivative at C-12
of the type -OCH2CH2R and -O(CdO)CH2R may not
have sufficient stability in aqueous solution to be
clinically useful in intravenously injectable dosage

4942 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 22
J ung et al.
Ta ble 1. Stability of Acetal (C-O) Type and Non-Acetal (C-C)
Type Prodrugs of Artemisinin in Simulated Stomach Acid (1
mg/mL, 0.01 N HCl, pH 2.0, 37 °C)
compared with artemisinin and artelinic acid and shows
higher suppression in vivo than either arteether or
artelinic acid. Deoxoartelinic acid is 23 times more
stable than clinically useful arteether and artelinic acid
and is 10 times more stable than artemisinin in simu-
lated stomach acid. Deoxoartelinic acid is 4 times more
soluble than artemisinin in water. Thus, deoxoartelinic
acid deserves further investigation as a new anti-
malarial drug candidate. Tests for bioavailabilty and
antimalarial activity from oral administration will be
performed and published in due course.
compound
half-life (h)
Acetal (C-O) Type
artemisinin (1)
arteether (3b)
artelinic acid (5)
23.50
10.98
13.11
Non-Acetal (C-C) Type
deoxoartemisinin (2)
244.97
258.66
186.89
deoxoartelinic acid (13)
12-(n-butyl)-deoxoartemisinin (3c)
Ta ble 4. In Vivo Antimalarial Activity of Deoxoartelinic Acid,
Arteether, and Chloroquine (10 mg kg-1 day-1 ip × 4) against
P. chabaudi in Mice
Ta ble 2. In Vitro Antimalarial Activity of Antimalarial Drugs
ED50 (ng/mL)
clone of P. falciparum
deoxoartelnic acid arteether chloroquine
antimalarial drug
3D7
K1
survival, mice
survival rate (%)
5/5
100
3/5
60
0/5
0
deoxoartelinic acid (13)
artemisinin (1)
artesunate (4)
artelinic acid (5)
arteether (3b)
1.1
2.9
0.2
4.0
0.2
8.0
0.6
0.7
0.6
1.4
0.9
Exp er im en ta l Section
Ch em istr y. NMR spectra were obtained on a Bruker AC250
spectrometer using Me4Si as an internal standard. The GC-
MS and direct mass were operated on an HP 5980II GC-HP
5988 and J MS-700 M station spectrometer in EI mode.
Infrared spectra were taken on a Nicolet Impact 400 spec-
trometer. Specific rotations were recorded on a Rudolph AP
III-589 polarimeter.
chloroquine
220
mL for deoxoartelinic acid, 7.4 mg/mL for sodium salt
of deoxoartelinic acid) when compared with artemisinin
(1.2 mg/mL) and have the same solubility as artelinic
acid.
P r ep a r a tion of Dih yd r oa r tem isin yl Ep oxid e (10) fr om
Dih yd r oa r tem isin yl Ald eh yd e (9). Dihydroartemisinyl al-
dehyde (9) (184 mg, 0.84 mmol) was mixed with trimethylsul-
fur iodide (375 mg, 1.67 mmol) and KOH (94 mg, 1.67 mmol)
in dry DMSO (10 mL), and this solution was stirred at room
temperature. After 2 h, ice water (50 mL) was added to the
reaction mixture and extracted with CHCl3 (30 mL × 3). The
extract was dried over MgSO4 and concentrated in vacuo to
give crude products, which were purified by a silica gel column
(hexane/ethyl acetate ) 5:1 as eluent) (Rf ) 0.71) to give
dihydroartemisinyl epoxide 10 (150 mg) in 76% yield as a
colorless oil: 1H NMR (CDCl3, 250 MHz) δ 5.12 (1H, s), 2.87-
2.78 (1H, m), 2.71-2.62 (1H, m), 2.58-2.53 (1H, m), 2.49 (1H,
bs), 1.96-1.64 (4H, m), 1.61 (3H, s), 1.59-1.40(5H, m), 1.07
(3H, d, J ) 5.9 Hz), 1.03-0.91 (2H, m), 0.89 (3H, d, J ) 6.3
Hz), 0.85-0.82 (1H, m); 13C NMR (CDCl3, 63 MHz) δ 135.7,
120.4, 49.6, 44.9, 42.3, 38.7, 37.5, 37.2, 35.9, 28.1, 27.2, 27.0,
26.2, 24.1, 20.2, 15.5; IR νmax 2920, 1454, 1382, 1214, 909, 822,
751 cm-1; EIMS m/z 234 ([M]+), 216 ([M - H2O]+), 201, 159,
145, 131, 105, 79, 55, 41, 29, 23, 14; HRMS (EI, 70 eV) m/z
234.1987 (obsd), 234.1985 (calcd for C16H26O).
P r ep a r a tion of 12-(p-Vin yl)h om oben zyld ih yd r oa r te-
m isin yl Alcoh ol (11) fr om Dih yd r oa r tem isin yl Ep oxid e
(10). Under a nitrogen atmosphere, the Grignard reagent,
prepared by 4’-vinylbenzyl chloride (183.2 mg, 1.2 mmol) and
magnesium metal, was slowly added dropwise to a solution of
dihydroartemisinyl epoxide 10 (130 mg, 0.55 mmol) in dry
diethyl ether (10 mL) at 0 °C. The mixture was stirred at 0 °C
in an ice bath in the dark. After 12 h, the mixture was
neutralized with 5% aqueous HCl and further diluted with
cooled H2O (50 mL). The mixture was extracted with diethyl
ether (20 mL × 3) and was washed with brine (10 mL × 2).
The extract was dried over MgSO4 and concentrated in vacuo
to give crude products, which were purified by a silica gel
column (hexane/ethyl acetate ) 5:1 as eluent) (Rf ) 0.42) to
give 12-(p-vinyl)homobenzyldihydroartemisinyl alcohol 11 (157
Biologica l Eva lu a tion
Deoxoartelinic acid 13 was tested in vitro against the
chloroquine-sensitive 3D7 strain and the chloroqine-
resistant K1 strain of P. falciparum (Table 2). Although
deoxoartelinic acid 13 possesses lower antimalarial
activity against 3D7 than artesunate (4), it is identical
to artesunate against the chloroquine-resistant K1
strain. Deoxoartelinic acid 13 shows 4 and 2 times more
activity than artelinic acid (5) against 3D7 and K1
strains of P. falciparum, respectively. Compound 13
shows almost the same level of activity as that of
arteether (3b) against the K1 strain. Deoxoartelinic acid
13 was subjected to further testing in mice (Table 3).
The results of the in vivo tests showed that there was a
98.7% suppression of parasitaemia in the Plasmodium
chabaudi infected mice with deoxoartelinic acid and that
the ED50 and ED90 values were both less than 10 mg/
kg. The survival rate of the five P. chabaudi infected
mice treated with deoxoartelinic acid is 100% compared
to 60% with arteether treatment (Table 4). Therefore,
deoxoartelinic acid is stimulating much interest as a
new and potent non-acetal-type antimalarial drug can-
didate.
In conclusion, (+)-deoxoartelinic acid, a new and more
active antimalarial drug candidate, was successfully
prepared from more abundant artemisinic acid by using
sulfur ylide and photooxygenative cyclization. (+)-
Deoxoartelinic acid shows superior antimalarial activity
in vitro against chloroquine-resistant malaria when
Ta ble 3. Blood Schizontocidal Activity (% Infected Erythrocytes) of Deoxoartelinic Acid and Chloroquine (10 mg kg-1 day-1 ip × 4)
against P. chabaudi in Mice
mouse
1
2
3
4
5
group mean
% control
% suppression
98.7
deoxoartelinic acid (5)
0.5
0.5
2.9
0.5
0.3
0.9
1.3
chloroquine
65.0
71.3
62.1
82.4
81.3
72.4
100

(+)-Deoxoartelinic Acid
J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 22 4943
mg) in 81% yield as a colorless foam: [R]25 +120.8° (c 0.48,
44.5, 37.7, 36.8, 34.6, 34.1, 31.6, 30.4, 26.3, 25.1, 24.9, 20.4,
13.2; IR νmax 3424, 2903, 2864, 1693, 1611, 1428, 1377, 1291,
1184, 1128, 1016, 944, 751 cm-1; HRMS (EI, 70 eV) m/z
416.2206 (obsd), 416.2199 (calcd for C24H32O6).
D
CHCl3); 1H NMR (CDCl3, 250 MHz) δ 7.35 (2H, d, J ) 7.8 Hz),
7.19 (2H, d, J ) 7.9 Hz), 6.75 (1H, dd, J ) 10.8, 17.6 Hz), 5.74
(1H, d, J ) 17.6 Hz), 5.21 (1H, d, J ) 10.9 Hz), 5.17 (1H, s),
3.91-3.86 (1H, m), 2.79-2.58 (2H, m), 2.48 (1H, bs), 1.92-
1.81 (3H, m), 1.78-1.64(2H, m), 1.62 (3H, s), 1.56-1.46 (3H,
m), 1.41-1.36 (2H, m), 1.30-1.24 (2H, m), 0.99-0.91 (2H, m),
0.88 (6H, d, J ) 6.7 Hz), 0.85-0.83 (1H, m); 13C NMR (CDCl3,
63 MHz) δ 142.2, 136.8, 135.4, 131.0, 128.7, 126.4, 120.7, 113.1,
72.2, 43.7, 42.4, 42.1, 40.4, 40.1, 37.7, 35.8, 32.8, 27.8, 26.9,
26.0, 24.0, 19.9, 10.7; IR νmax 3394, 2920, 2864, 1601, 1510,
1443, 1377, 990, 904, 822 cm-1; HRMS (EI, 70 eV) m/z
352.2752 (obsd), 352.2766 (calcd for C25H36O). Because of the
tendency of the styryl system to polymerize, compound 11 was
used immediately in the next step.
Sta bility Tests of Ar tem isin in An a logu es in Sim u la ted
Stom a ch Acid . Sa m p le P r ep a r a tion a n d Mea su r em en t
of Sta bility. A 100 µL portion of a 1.0 mg artemisinin prodrug
stock solution (in acetonitrile) was added to 1.0 mL of a freshly
prepared 0.01 N HCl aqueous solution (preheated to 37 °C).
The resulting mixture was sealed to prevent water evaporation
and maintained at 37 °C in a water bath. Acetal-type samples
(1, 3b, 5) of the reaction mixture were taken at time intervals
of 0, 200, 400, 600, 800, 1000, 1200, 1400 min, and non-acetal-
type samples (2, 3c, 13) were taken at 2, 4, 6, 8, 10, 12, and
14 days. The samples were stored in dry ice-acetone bath and
analyzed (in triplicate) as quickly as possible. HPLC with UV
detector (Waters 486 tunable absorbance detector, Millipore)
was used to identify the decomposition of the prodrugs and
quantitation. All prodrugs were detected with a wavelength
of 210 nm except 250 nm for aromatic compounds 5 and 13.
The column was a Nova-Pak C-18 (4 µm particle size, 15.0-
3.9 cm length) column used with a mobile phase (1.0 mL/min),
which was completely deoxygenated by flowing argon gas. Two
mobile-phase systems were used, where system 1 (for the assay
of n-butyldeoxoartemisinin (tr ) 5.53 min) consisted of 0.1 M
ammonium acetate with 80% acetonitrile in water. System 2,
used for deoxoartelinic acid (tr ) 9.83 min) consisted of 0.1 M
ammonium acetate with 30% acetonitrile in water. Two
internal standards were used: arteether (tr ) 3.45 min) for
HPLC system 1 and artelinic acid (tr ) 9.97 min) for system
2.
Biology. In Vitr o An tim a la r ia l Stu d ies. Cultures of the
3D7 chloroquine sensitive strain of P. falciparum35 and K1
chloroquine resistant36 were maintained in RPMI 1640 me-
dium (Sigma, U.K.), 37 °C, 5% CO2 in 5% hematocrit in A+
erythrocytes. Test compounds were tested in a 96-well plate
format using synchronized ring stage cultures prepared at 1%
parasitemia. An amount of 100 µL of synchronized culture was
added per well. Drugs were tested in triplicate over a 3-fold
dilution series from a final top drug concentration of 30 µg
/mL. Standard drug was chloroquine diphosphate. Control
wells were infected erythrocytes with no drug, and blank wells
were uninfected A+ erythrocytes. After 24 h of incubation at
37 °C, 5% CO2, 20 µL of [3H]-hypoxanthine was added to all
wells (0.1 µCi /well)37,38 and plates were shaken for 1 min and
then incubated for a further 24 h. The plates were freeze-
thawed rapidly, harvested onto a 96-well glass fiber filter plate
(Canberra Packard), and dried at 42 °C. Incorporation of
radioactive hypoxanthine was measured using a Canberra
Packard TopCount scintillation counter. Results were analyzed
using the Microsoft Excel based MsXlfit (IDBS, U.K.) to
calculate ED50 values.
P r ep a r a tion of 12-(p-Vin yl)h om oben zyld eoxoa r tem i-
sin in (12) fr om 12-(p-Vin yl)h om oben zyld ih yd r oa r tem i-
sin yl Alcoh ol (11). A pale-pink solution of alcohol 11 (120
mg, 0.34 mmol) and rose bengal (5 mg) in CH3CN/CH2Cl2 (9/
1, 25 mL) was irradiated with white light (500 W tungsten
lamp) at -40 °C under oxygen. After 4 h, TLC analysis
indicated that the majority of the starting material had
disappeared. The mixture was poured onto a saturated NaH-
CO3 solution (50 mL), and products were extracted into diethyl
ether (20 mL × 3). The sensitizer remained in the aqueous
phase. The combined ether extracts were washed with brine
(20 mL × 2) and dried with MgSO4. Removal of solvent under
reduced pressure left a colorless foam. This was dissolved in
CH3CN (10 mL), and the resultant solution was cooled to -40
°C and was followed by in situ treatment of acidic catalyst
TFA. The mixture was stirred at this temperature under an
oxygen atmosphere for 2 h. The reaction mixture was quenched
with saturated NH4Cl solution (10 mL), and the products were
extracted with diethyl ether (20 mL × 3). The extract was
washed with water (30 mL × 2) and brine (30 mL × 2) and
dried with MgSO4. Concentration in vacuo gave crude prod-
ucts, which were purified by a silica gel column (hexane/ethyl
acetate ) 5:2 as eluent) (Rf ) 0.53) to give 12-(p-vinyl)-
homobenzyldeoxoartemisinin 12 (47 mg) in 35% yield as a
colorless foam: [R]25 +18.2° (c 0.1, CHCl3); 1H NMR (CDCl3,
D
250 MHz) δ 7.34 (2H, d, J ) 8.1 Hz), 7.20 (2H, d, J ) 8.0 Hz),
6.74 (1H, dd, J ) 10.9, 17.6 Hz), 5.73 (1H, d, J ) 17.6 Hz),
5.36 (1H, s), 5.21 (1H, d, J ) 10.9 Hz), 4.24-4.17 (1H, m),
3.04-2.89 (1H, m), 2.79-2.53 (2H, m), 2.33 (1H, ddd, J ) 3.8,
3.0, 3.8 Hz), 2.09-2.01 (2H, m), 1.94-1.78 (3H, m), 1.74-1.64
(2H, m), 1.57-1.46 (1H, m), 1.44 (3H, s), 1.42-1.20 (3H, m),
1.17-1.04 (1H, m), 0.96 (3H, d, J ) 5.9 Hz), 0.85 (3H, d, J )
7.6 Hz); 13C NMR (CDCl3, 63 MHz) δ 142.4, 136.8, 135.4, 128.8,
126.4, 113.1, 103.4, 89.2, 81.3, 75.5, 52.6, 44.6, 37.6, 36.8, 34.6,
33.8, 31.9, 30.4, 26.3, 26.1, 24.9, 20.4, 13.2; IR νmax 3373, 2925,
2864, 1627, 1510, 1448, 1382, 1179, 1128, 1056, 1011, 832.,
756 cm-1; HRMS (EI, 70 eV) m/z 398.2463 (obsd), 398.2457
(calcd for C25H34O4).
Biology. In Vivo An tim a la r ia l Stu d ies. Female mice
(BALB/c, specific pathogen free), 18-20 g, were infected
intravenously (iv) with 1 × 107 P. chabaudi ANKA infected
erythrocytes from donor mice on day 1 of the experiment. Blood
was taken from donor mice, in serum, and diluted in RPMI
(Sigma, U.K.) to a parasitemia of 1% (the equivalent of 1 ×
107 infected erythrocytes), and 0.2 mL was administered to
each mouse iv. Mice were randomly sorted into groups of five.
Then 2 h of postinoculation dosing commenced. The control
drug, chloroquine, was given ip every day for 4 days. The route
of administration of experimental compounds was ip. Drugs
were administered at 0.2 mL every day for 4 days in 10%
DMSO in phosphate-buffered saline (Sigma, U.K.). On day 5,
postinfection blood smears of all animals were prepared and
stained with 10% Giemsa. Parasitemia was determined mi-
croscopically by counting a minimum of 1000 red cells. Results
are reported as the percentage of infected erythrocytes and
compared to the chloroquine control group and the uninfected
control group.39
P r ep a r a tion of Deoxoa r telin ic Acid (13) fr om 12-(p-
Vin yl)h om oben zyld eoxoa r tem isin in (12). A solution of 12-
(4’-p-vinylhomobenzyldeoxoartemisinin 12 (40 mg, 0.10 mmol)
in HPLC grade acetone was slowly mixed with NaHCO3 (4.2
mg, 0.05 mmol) and KMnO4 (47 mg, 0.30 mmol). The reaction
mixture was stirred at room temperature for 1 h and then
treated with 10% aqueous HCl (3 mL) and stirred at room
temperature for 5 h. The mixture was extracted with diethyl
ether (10 mL × 3) and was washed with brine (10 mL × 2).
The extract was dried over MgSO4 and was concentrated in
vacuo to give crude products, which were purified by a silica
gel column (hexane/ethyl acetate ) 1:2 as eluent) (Rf ) 0.46)
to give deoxoartelinic acid 13 (35 mg) in 83% yield as a white
1
solid: mp 135-137 °C; [R]20 +58.3° (c 0.1, CHCl3); H NMR
D
(CDCl3, 250 MHz) δ 8.03 (2H, d, J ) 8.2 Hz), 7.34 (2H, d, J )
8.1 Hz), 5.36 (1H, s), 4.26-4.20 (1H, m), 3.12-3.00 (1H, m),
2.75-2.65 (2H, m), 2.34 (1H, ddd, J ) 3.8, 2.9, 3.8 Hz), 2.13-
2.01 (1H, m), 1.96-1.89 (3H, m), 1.84-1.73 (3H, m), 1.68-
1.48 (4H, m), 1.44 (3H, s), 1.38-1.17 (2H, m), 0.96 (3H, d, J )
5.6 Hz), 0.86 (3H, d, J ) 7.5 Hz); 13C NMR (CDCl3, 63 MHz)
δ 171.6, 149.2, 130.6, 128.9, 127.1, 103.5, 89.3, 81.3, 75.0, 52.5,
Ack n ow led gm en t. This work was supported by
Korea Research Foundation (Grant KRF-2000-A1004-

4944 J ournal of Medicinal Chemistry, 2002, Vol. 45, No. 22
J ung et al.
(21) J ung, M.; Yu, D.; Bustos, D. A.; ElSohly, H. N.; McChesney, J .
DP0267). H.K., B.R., and S.L.C. are supported by grants
from the UNDP/World Bank/WHO Special Programme
for Research and Training in Tropical Diseases (TDR).
D.
A Concise Synthesis of 12-(3′-Hydroxy-n-propyl)-deoxo-
artemisinin. Bioorg. Med. Chem. Lett. 1991, 1, 741-744.
(22) Haynes, R. K.; Vonwiller, S. C. Efficient Preparation of Novel
Qinghaosu (Artemisinin) Derivatives. Synlett 1992, 481-483.
(23) Pu, Y. M.; Ziffer, H. Synthesis and Antimalarial Activities of
12-â-Allyldeoxoartemisinin and Its Derivatives. J . Med. Chem.
1995, 38, 613-616.
(24) Abouabdellah, A.; Begue, J .-P.; Bonnet-Delpon, D.; Gantier, J .-
C.; Nga, T. T. T.; Thac, T. D. Synthesis and in Vivo Antimalarial
Activity of 12R-Trifluoromethyl-Hydroartemisinin. Bioorg. Med.
Chem. Lett. 1996, 6, 2717-2720.
(25) Woo, S. H.; Parker, M. H.; Ploypradith, P.; Northrop, J .; Posner,
G. H. Direct Conversion of Pyranose Anomeric OH f F f R in
the Artemisinin Family of Antimalarial Trioxanes. Tetrahedron
Lett. 1998, 39, 1533-1536.
(26) Nga, T. T. T.; Menage, C.; Begue, J . P.; Bonnet-Delpon, D.;
Gantier, J .-C. Synthesis and Antimalarial Activities of Fluoro-
alkyl Derivatives of Dihydroartemisinin. J . Med. Chem. 1998,
41, 4101-4108.
(27) Posner, G. H.; Parker, M. H.; Northrop, J .; Elias, J . S.; Ploypra-
dith, P.; Xie, S.; Shapiro, T. A. Orally Active, Hydrolytically
Stable, Semisynthetic Antimalarial Trioxanes in the Artemisinin
Family. J . Med. Chem. 1999, 42, 300-304.
(28) O’Neill, P. M.; Searle, N. L.; Kan, K.-W.; Storr, R. C.; Maggs, J .
L.; Ward, S. A.; Raynes, K.; Park, B. K. Novel, Potent, Semi-
synthetic Antimalarial Carba Analogues of the First-Generation
1,2,4-Trioxane Artemether. J . Med. Chem. 1999, 42, 5487-5493.
(29) Chorki, F.; Crousse, B.; Bonnet-Delpon, D.; Begue, J . P.; Brigaud,
T.; Portella, C. C-10 Fluorinated Derivatives of Dihydro-
artemisinin: Difluoromethylene Ketones. Tetrahedron Lett.
2001, 42, 1487-1489.
(30) J ung, M.; Lee, K.; J ung, H. First synthesis of (+)-Deoxo-
artemisitene and Its Novel C-11 Derivatives. Tetrahedron Lett.
2001, 42, 3997-4000.
(31) J ung, M.; Freitas, A. C. C.; McChesney, J . D.; ElSohly, H. N. A
Practical and General Synthesis of (+)-Carboxyalkyldeoxo-
artemisinins. Heterocycles 1994, 39, 23-29.
(32) Luo, X. D.; Shen, C. C. The Chemistry, Pharmacology, and
Clinical Applications of Qinghaosu (Artemisinin) and Its Deriva-
tives. Med. Res. Rev. 1987, 7, 29-52.
(33) Baker, H. K.; McChesney, J . D.; Chi, H. T. Decomposition of
Arteether in Simulated Stomach Acid Yielding Compounds
Retaining Antimalarial Activity. Pharm. Res. 1993, 10, 662-
666.
(34) J ung, M.; Schinazi, R. F. Synthesis and in Vitro Anti-Human
Immunodeficiency Virus Activity of Artemisinin (Qinghaosu)-
Related Trioxanes. Bioorg. Med. Chem. Lett. 1994, 4, 931-934.
(35) Ponnudurai, T.; Leeuwenberg, A. D.; Meuwissen, J . H. Chloro-
quine Sensitivity of Isolates of Plasmodium falciparum Adapted
to in Vitro Culture. Trop. Geogr. Med. 1981, 33, 50-54.
(36) Thaithong, S.; Beale, G. H. Resistance of ten Thai isolates of
Plasmodium falciparum to chloroquine and pyrimethamine by
in vitro tests. Trans. R. Soc. Trop. Med. Hyg. 1981, 75, 271-
273.
(37) Desjardins, R. E.; Canfield, J .; Haynes, D.; Chuly, D. J . Quan-
titative Assessment of Antimalarial Activity in Vitro by a Semi-
automated Microdilution Technique. Antimicrob. Agents Chemo-
ther. 1979, 16, 710-718.
Refer en ces
(1) Klayman, D. L. Qinghaosu (Artemisinin): An Antimalarial Drug
from China. Science 1985, 228, 1049-1055.
(2) Zaman, S. S.; Sharma, R. P. Some Aspects of the Chemistry and
Biological Activity of Artemisinin and Related Antimalarials.
Heterocycles 1991, 32 (8), 1593-1638.
(3) J ung, M. Current Developments in the Chemistry of Artemisinin
and Related Compounds. Curr. Med. Chem. 1994, 1, 35-49.
(4) (a) Zhou, W. S.; Xu, X. X. Total Synthesis of the Antimalarial
Sesquiterpene Peroxide Qinghaosu and Yingzhaosu A. Acc.
Chem. Res. 1994, 27, 211-216. (b) Robert, A.; Dechy-Cabaret,
O.; Cazelles, J .; Meunier, B. From Mechanistic Studies on
Artemisinin Derivatives to New Modular Antimalarial Drugs.
Acc. Chem. Res. 2002, 35, 167-174.
(5) J efford, C. W. Peroxidic Antimalarials. In Advances in Drug
Research; Academic Press: London, 1997; Vol. 29, pp 271-325.
(6) Haynes, R. K.; Vonwiller, S. C. From Qinghao, Marvelous Herb
of Antiquity, to the Antimalarial Trioxane Qinghaosu and Some
Remarkable New Chemistry. Acc. Chem. Res. 1997, 30, 73-79.
(7) Avery, M. A. Current Progress in the Chemistry, Medicinal
Chemistry and Drug Design of Artemisinin Based Antimalarials.
Curr. Pharm. Des. 1999, 5 (2), 101-138.
(8) Bhattacharya, A. K.; Sharma, R. P. Recent Developments on the
Chemistry and Biological Activity of Artemisinin and Related
AntimalarialssAn Update. Heterocycles 1999, 51 (7), 1681-
2000.
(9) Dhingra, V.; Rao, K. V.; Narasu, M. L. Current Status of
Artemisinin and Its Derivatives as Antimalarial Drugs. Life Sci.
2000, 66, 279-300.
(10) McCullough, K. J .; Nojima, M. Recent Advances in the Chemistry
of Cyclic Peroxides. Curr. Org. Chem. 2001, 5, 601-636.
(11) (a) Woerdenbag, H. J .; Moskal, T. A.; Pras, N.; Maringle, T. M.;
ElFeraly, F. S.; Kampinga, H. H.; Konings, A. W. T. Cytotoxicity
of Artemisinin-Related Endoperoxides to Ehrlich Ascites Tumor
Cells. J . Nat. Prod. 1993, 56, 849-856. (b) Beekman, A. C.;
Woerdenbag, H. J .; Kampinga, H. H.; Konings, A. W. T.
Cytotoxicity of Artemisinin, a Dimer of Dihydroartemisinin,
Artemisitene and Eupatoriopicrin as evaluated by the MTT and
Clonogenic assay. Phytother. Res. 1996, 10, 140-144.
(12) Zheng, G.-Q. Cytotoxic Terpenoids and Flavonoids from Arte-
misia annua. Planta Med. 1994, 60, 54-57.
(13) Posner, G. H.; Ploypradith, P.; Parker, M. H.; O’Dowd, H.; Woo,
S.-H.; Northrop, J .; Krasavin, M.; Dolan, P.; Kensler, T. W.; Xie,
S.; Shapiro, T. A. Antimalarial, Antiproliferative, and Antitumor
Activities of Artemisinin-Derived, Chemically Robust, Trioxane
Dimers. J . Med. Chem. 1999, 42, 4275-4280.
(14) J ung, M. Synthesis and Cytotoxicity of Novel Artemisinin
Analogs. Bioorg. Med. Chem. Lett. 1997, 7, 1091-1094.
(15) Tropical Disease Research Progress 1995-96, Thirteenth Pro-
gramme Report of UNDP/ World Bank/ WHO Special Pro-
gramme for Research & Training in Tropical Disease (TDR);
World Health Organization: Geneva, 1997; p 51.
(16) Lin, A. J .; Klayman, D. L.; Milhous, W. K. Antimalarial Activity
of New Water-Soluble Dihydroartemisinin Derivatives. J . Med.
Chem. 1987, 30, 2147-2150.
(38) O’Neill, M. J .; Bray, D. H.; Boardman, P.; Phillipson, J . D.;
Warhurst, D. C. Plants as Sources of Antimalarial Drugs. Part
1. In vitro test method for the Evaluation of Crude Extracts from
Plants. Planta Med. 1985, 5, 394-398.
(17) Lin, A. J .; Lee, M.; Klayman, D. L. Antimalarial Activity of New
Water-Soluble Dihydroartemisinin Derivatives. 2. Stereospeci-
ficity of the Ether Side Chain. J . Med. Chem. 1989, 32, 1249-
1252.
(39) Peters, W.; Portus, J . H.; Robinson, B. L. The Chemotherapy of
Rodent Malaria XXII. The Value of Drug Resistant Strains of
Plasmodium berghei in Screening for Blood Schizontocidal
Activity. Ann. Trop. Med. Parasitol. 1975, 69, 155-171.
(40) Posner, G. H.; J eon, H. B.; Parker, M. H.; Krasavin, M.; Paik,
I.-H.; Shapiro, T. A. Antimalarial Simplified 3-Aryltrioxanes:
Synthesis and Preclinical Efficacy/Toxicity Testing in Rodents.
J . Med. Chem. 2001, 44, 3054-3058.
(18) J ung, M.; Li, X.; Bustos, D. A.; ElSohly, H. N.; McChesney, J .
D. A Short and Stereospecific Synthesis of (+)-Deoxoartemisinin
and (-)-Deoxodesoxyartemisinin. Tetrahedron Lett. 1989, 30,
5973-5976.
(19) J ung, M.; Li, X.; Bustos, D. A.; ElSohly, H. N.; McChesney, J .
D.; Milhous, W. K. Synthesis and Antimalarial Activity of (+)-
Deoxoartemisinin. J . Med. Chem. 1990, 33, 1516-1518.
(20) J ung, M.; Bustos, D. A.; ElSohly, H. N.; McChesney, J . D. A
Concise and Stereoselective Synthesis of (+)-12-n-Butyldeoxo-
artemisinin. Synlett 1990, 743-744.
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