Orally active esters of dihydroartemisinin: Synthesis and antimalarial activity against multidrug-resistant Plasmodium yoelii in mice

Article abstract of DOI:10.1016/j.bmcl.2007.12.074

A series of artemisinin derived esters 7a-j, incorporating pharmacologically privileged substructure, such as biphenyl, adamantane and fluorene, have been prepared and evaluated for antimalarial activity against multidrug-resistant (MDR) Plasmodium yoelii

Full text of DOI:10.1016/j.bmcl.2007.12.074

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 18 (2008) 1436–1441
Orally active esters of dihydroartemisinin:
Synthesis and antimalarial activity against multidrug-resistant
Plasmodium yoelii in miceq
Chandan Singh,a,* Sandeep Chaudharya and Sunil K. Purib
aDivision of Medicinal and Process Chemistry, Central Drug Research Institute, Lucknow 226001, India
bDivision of Parasitology, Central Drug Research Institute, Lucknow 226001, India
Received 27 November 2007; revised 25 December 2007; accepted 29 December 2007
Available online 5 January 2008
Abstract—A series of artemisinin derived esters 7a–j, incorporating pharmacologically privileged substructure, such as biphenyl,
adamantane and fluorene, have been prepared and evaluated for antimalarial activity against multidrug-resistant (MDR) Plasmo-
dium yoelii nigeriensis by oral route. Several of these compounds were found to be more active than the antimalarial drugs b-artee-
ther 4 and artesunic acid 5. Ester 7i, the most active compound of the series, provided 100% and 80% protection to the infected mice
at 24 mg/kg · 4 days and 12 mg/kg · 4 days, respectively. In this model b-arteether provided 100% and 20% protection at 48 mg/
kg · 4 days and 24 mg/kg · 4 days, respectively.
ꢀ 2008 Elsevier Ltd. All rights reserved.
15
The discovery of artemisinin 1, as the active principle of
H
H
H
5
H
4
6
the Chinese traditional drug against malaria, Artemisia
annua, is a major breakthrough in malaria chemother-
apy. The derivatives of artemisinin, for example, dihyd-
roartemisinin 2, artemether 3, arteether 4 and artesunic
acid 5 (Fig. 1), are more active than the parent com-
pound, and are currently the drugs of choice for the
treatment of malaria caused by multidrug-resistant
P. falciparum.2,3
2
7
8
O
O
3
O
O
5a
8a
14
1
O
O
O
O
13
12a
10
O
O
O
O
12
11
H
H
H
H
O
O
O
O
9
O
16
OR
O
O
OH
OH
O
5
3, R = Me
4, R = Et
1
2
Figure 1. Artemisinin and its clinically useful derivatives.
While these compounds show high efficacy when admin-
istered by systemic routes, they are comparatively
less active when given by oral route. We have recently
reported the synthesis of a series of lipophilic ether
derivatives of dihydroartemisinin, incorporating phar-
macologically privileged substructures such as biphenyl,
adamantane and fluorene, some of which showed high
order of antimalarial activity against multidrug-resistant
P. yoelii in mice by oral route.4 A noticeable feature of
these derivatives was that a-isomers, the minor products
of acid catalysed etherification of dihydroartemisinin,
were significantly more active than the corresponding
b-isomers. Since base-catalysed esterification of dihyd-
roartemisinin with acid chlorides/anhydrides furnishes
exclusively a-isomers,5 it made commercial sense to us
to prepare and evaluate for activity the corresponding
ester derivatives, incorporating these pharmacologically
privileged substructures. In the event, several of these
derivatives, showed very promising antimalarial activity
against multidrug-resistant malaria in mice by oral
route. Herein, we report the details of this study.6
Dihydroartemisinin 2 was prepared from artemisinin 1
using the known procedure.7 The acid chlorides RCOCl
6a–j were prepared from the corresponding carboxylic
acids by heating with thionyl chloride at 50–60 ꢁC for
2–3 h and reacted with dihydroartemisinin 2 in the pres-
ence of triethylamine in dry dichloromethane at 0 ꢁC for
Keywords: Dihydroartemisinin; Arteether; Antimalarial; Multidrug-
resistant; 1,2,4-Trioxanes.
q
Ref. 1.
Corresponding author. Tel.: +91 0522 2612411 18x4220; fax: +91 522
2623405; e-mail: chandancdri@yahoo.com
*
0960-894X/$ - see front matter ꢀ 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.12.074

C. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1436–1441
1437
H
O
O
O
H
SOCl2
50-60 C, 2-3h
2 / Et3N
O
RCOCl
RCOOH
º
Dry CH2Cl2, 0 ºC,2h
O
R
6a-j
O
7a-j
a
R =
R =
e
f
R =
R =
i
R =
H2C
H2C
b
H2C
CH2
O
g
R =
R =
c
R =
R =
H2C
j
R =
H2C
O
d
h
H2C H2C
Scheme 1. Synthesis of ester derivatives 7a–j.
2 h to furnish ester derivatives 7a–j in 49–58% yields
(Scheme 1, Table 1).8
Table 1. Ester derivatives 7a–j
Compound
Mp
% Yield
H
O
Antimalarial drugs b-arteether and artesunic acid, when
given orally at 48 mg/kg · 4 days, provide 100% protec-
tion to the mice infected with multidrug-resistant P. yoelii
nigeriensis. At 24 mg/kg · 4 days, while artesunic acid
does not provide any protection, b-arteether provides
only 20% protection. Since the objective of the present
study was to select compounds having activity profile
better than that of b-arteether and artesunic acid, all
the newly prepared ester derivatives 7a–j were initially
screened against multidrug-resistant P. yoelii nigeriensis
in Swiss mice at 48 mg/kg · 4 days by oral route.9 All
these ester derivatives provided 100% protection at this
dose and therefore all these active compounds were fur-
ther screened at 24 mg/kg · 4 days. Compounds 7f, 7i
and 7j which showed 100% protection at 24 mg/kg · 4
days were further tested at 12 mg/kg · 4 days. The re-
sults are summarized in Table 2.
O
O
H
R
O
O
O
R =
7a
7b
59–61 ꢁC
63–65 ꢁC
57
57
H2C
H2C
O
7c
57–59 ꢁC
56
7d
7e
7f
H2C
86–88 ꢁC
85–87 ꢁC
115–117 ꢁC
53
49
58
O
The semi-synthetic derivatives of artemisinin such as
artemether 3, arteether 4 and artesunic acid 5 are highly
effective against both chloroquine-sensitive and resistant
malaria. They are fast acting and are currently the drugs
of choice for the treatment of complicated cases of ma-
laria caused by multidrug-resistant P. falciparum. While
these drugs show high efficacy when administered by
systemic routes, they are comparatively less active when
given orally. In our search for artemisinin derivatives
with high efficacy by oral route, we had recently re-
ported a series of ether derivatives of dihydroartemisi-
nin, incorporating adamantane, biphenyl and fluorene
moieties. Several of these lipophilic derivatives were
found to be 2–4 times more active than arteether by oral
route.5,10,11 We also observed that the a-isomers, that is,
8a–d of these ether derivatives were significantly more
active than the corresponding b-isomers 9a–d (Fig. 2).
H2C
H2C
7g
7h
73–75 ꢁC
57
55
115–116 ꢁC
H2C·H2C
7i
7j
75–77 ꢁC
73–75 ꢁC
52
57
H2C
This was a serious limitation because the a-isomers were
the minor products (20–25% of the a- and b-mixture)
and the isolation of the pure isomers required extensive
chromatographic separation. The observation, however,
suggested that a bulkier group on the a-face of the mol-
ecule has beneficial effect on antimalarial activity and

1438
C. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1436–1441
Table 2. Blood schizontocidal activity of esters 7a–j against multidrug-resistant (MDR) strain P. yoelii nigeriensis in Swiss mice via oral route9
Compound
LogP
Dose (mg/kg · 4 days)a
% Suppression of parasitaemia on day 4b,c
Cured/ Treated
7a
6.95
48
24
48
24
48
24
12
48
24
48
24
48
24
12
48
24
12
48
24
12
48
24
12
48
24
12
48
24
48
24
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
5/5
3/5
5/5
3/5
5/5
2/5
0/5
5/5
1/5
5/5
0/5
5/5
5/5
1/5
5/5
4/5
0/5
5/5
3/5
0/5
5/5
5/5
4/5
5/5
5/5
0/5
5/5
1/5
5/5
0/5
7b
7c
6.89
6.53
7d
7e
7f
6.53
6.05
5.99
7g
7h
7i
5.85
6.41
6.61
6.79
7j
4
5
3.84
3.04
a The drug dilutions of compounds were prepared in groundnut oil and administered to a group of mice at each dose, from day 0 to 3, once daily.
b Percent suppression = [(C–T)/C] · 100; where C = parasitaemia in control group and T = parasitaemia in treated group.
c 100% suppression of parasitaemia means no parasites were detected in 50 oil immersion fields during microscopic observation.12
H
H
rable with that of the active ether derivatives 8a–d
(LogP 5.51–6.91) earlier reported by us.
O
O
O
O
O
O
H
H
O
O
As can be seen from Table 2, all these compounds
provided 100% protection at 48 mg/kg · 4 days and
therefore all these derivatives are at least as effective as
b-arteether which provided 100% and 20% protection
at 48 mg/kg · 4 days and 24 mg/kg · 4 days; respec-
tively. Among the biphenyl derivatives only 7a and
7b provided significant protection (60% protection)
at 24 mg/kg · 4 days. The adamantane-based esters
showed better activity profile; while compound 7f pro-
vided 100% protection at 24 mg/kg, compounds 7g and
7h provided 80% and 60% protection, respectively, at
this dose. Even at 12 mg/kg · 4 days compound 7f
showed 100% clearance of parasitaemia12 on day 4
and 20% of the treated mice survived beyond day 28.
In this series adamantane moiety linked through C-1
and separated by one CH2 from the ester group proved
to be the optimum for activity; absence of CH2 or
replacement of CH2 with CH2CH2 proved detrimental
to biological activity. The fluorene derivative 7i, the
most active compound of the series, provided 100% pro-
tection at 24 mg/kg · 4 days. At 12 mg/kg · 4 days, it
provided 80% protection. Its homologue 7j showed
100% protection at 24 mg/kg. At 12 mg/kg · 4 days, it
showed 100% clearance of parasitaemia on day 4 but
none of the treated mice survived beyond day 28.
OR
OR
9a-d
8a-d
a
R =
R =
H2C
H2C
c
R =
R =
H2C H2C
O
H2C
d
b
Figure 2. Structure of active ether derivatives of dihydroartemisinin
8a–d.
prompted us to prepare ester derivatives of dihydroarte-
misinin incorporating similar substructure. The ester
derivatives of dihydroartemisinin have the advantage
over the ether derivatives as they are obtained exclu-
sively as a-isomers and therefore their purification does
not require elaborate chromatographic separation. In
the event we prepared and screened against multidrug-
resistant P. yoelii 10 ester derivatives 7a–j incorporating
adamantane, biphenyl and fluorene moieties. The lipo-
philicity of these derivatives (LogP 5.85–6.95) is compa-

C. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1436–1441
1439
Shen, C. C. Med. Res. Rev. 1987, 7, 29–52; (c) Cumming,
J. N.; Ploypradith, P.; Posner, G. H. Adv. Pharmacol.
1997, 37, 253; (d) Bhattacharya, A. K.; Sharma, R. P.
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Res. Rev. 2004, 24, 425; (i) Jefford, C. W. Curr. Opin.
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3. (a) Asthana, O. P.; Srivastava, J. S.; Valecha, N.
J. Parasitic Diseases 1997, 211, 1–12; (b) Jambou, R.;
Legrand, E.; Niang, M.; Khim, N.; Lim, P.; Volney, B.;
Therese Ekala, M.; Bouchier, C.; Esterre, P.; Fandeur, T.;
Mercereau-Puijalon, O. Res. Lett. 2005, 366, 1960–1963.
4. Singh, C.; Chaudhary, S.; Puri, S. K. J. Med. Chem. 2006,
49(24), 7227–7233.
5. Haynes, R. K.; Chan, H.-W.; Cheung, M.-K.; Lam,
W.-L.; Soo, M.-K.; Tsang, H.-W.; Voerste, A.;
Williams, I. D. Eur. J. Org. Chem. 2002, 113.
6. This work has been covered in an Indian patent: Singh, C.;
Chaudhary, S.; Puri, S. K. Indian Patent Appl. No. 0391
DEL 2006, Filing Date 13-02-2006.
7. Brossi, A.; Venugopalan, B.; Dominquez, G. L.; Yeh, H.
J. C.; Flippen, A. J. L.; Buchs, P.; Wo, X. D.; Milhous,
W.; Peters, W. J. Med. Chem. 1988, 31, 645.
Figure 3. Three-dimensional structure of compounds 7i, 8d and 9d.
Since the present work on ester derivatives was inspired
by the excellent results shown by the structurally similar
ether derivatives,4 it is worthwhile to compare the bio-
logical activity of these two series. The ester derivatives
incorporating the biphenyl moiety on the whole are
much less active than their ether counterparts, two of
which (8a and 8b) had shown 100% protection at
12 mg/kg · 4 days. In the ether series, in fact, biphe-
nyl-based compounds were more active than the ada-
mantane- and fluorene-based derivatives. In the
adamantane-based compounds, both the series have
similar level of activity; in both these series the most ac-
tive compounds are twice as active as b-arteether. In the
fluorene-based series, the esters are marginally more ac-
tive than the ethers.
8. The preparation and antimalarial activity of esters 7a and
7d have been reported earlier: (a) Li, Y.; Yu, P.-L.; Chen,
I.-H.; Chi, J.-Y. Yao Hsueh Tung Pao 1980, 15, 38, Chem.
Abstr. 1980, 96, 6883u; (b) Li, Y.; Yu, P.; Chen, Y.; Ji, R.
HuaXue XueBao 1982, 40, 557–561; Chem. Abstr. 1982, 98,
4420h. (b) General procedure for esterification of dihyd-
roartemisinin (compound 7a as representative). To a
solution of dihydroartemisinin (0.50 g, 1.75 mmol) and
biphenyl-4-carbonyl chloride (1.14 g, 3 equiv, 5.25 mmol)
dissolved in dry dichloromethane (30 mL) was added
triethylamine (0.73 ml, 3 equiv, 5.29 mmol) dropwise at
0 ꢁC. The mixture was stirred at the same temperature for
2 h. The reaction mixture was then quenched with
saturated sodium bicarbonate solution (25 mL) and
extracted with dichloromethane (3· 25 mL). The organic
layer was washed with 10% aqueous HCl solution (2·
20 mL), then with water, dried over anhyd Na2SO4 and
concentrated under reduced pressure. The crude product
on column chromatography over silica gel using ethyl
acetate/hexane (1:25) as eluant gave pure 7a (465 mg, 57%)
as a white solid. (c) Selected spectral data. Compound 7a:
White solid; FT-IR (KBr, cmÀ1): 2932.3, 2876.9, 1729.0,
1608.1, 1452.4, 1378.2, 1269.4, 1201.8, 1130.6, 1087.8,
1016.9, 754.2; 1H NMR (200 MHz, CDCl3) d 0.94 (d, 3H,
J = 7.1 Hz, CH3), 0.98 (d, 3 H, J = 5.8 Hz, CH3), 1.25–
2.10 (m, 10H), 1.43 (s, 3H, CH3), 2.32–2.46 (m, 1H), 2.74–
2.80 (m, 1H), 5.54 (s, 1H, C12–H), 6.03 (d, 1H, J = 9.8 Hz,
C10–H), 7.38–7.51 (m, 3H), 7.60–7.68 (m, 4H), 8.18 (d, 2H,
J = 8.4 Hz); FABMS (m/z): 465 [M+H]+; ESMS (m/z): 482
[M+NH4]+, 487 [M+Na]+; Anal. Calcd for (C28H32O6Æ0.1-
H2O): C 72.38 H 6.94; Found C 72.11 H 6.96. Compound
7b: White solid; FT-IR (KBr, cmÀ1): 2931.5, 2875.8,
1748.8, 1487.4, 1452.0, 1379.1, 1243.1, 1141.1, 1099.7,
1019.8, 756.3; 1H NMR (200 MHz, CDCl3) d 0.74 (d, 3H,
J = 7.0 Hz, CH3), 0.95 (d, 3H, J = 5.1 Hz, CH3), 1.17–2.07
(m, 10H), 1.45 (s, 3H, CH3), 2.30–2.44 (m, 1H), 2.52–2.63
(m, 1H), 3.75 (s, 2H), 5.44 (s, 1H, C12–H), 5.80 (d, 1H,
J = 9.8 Hz, C10–H), 7.29–7.59 (m, 9H); 13C NMR
(50 MHz, CDCl3) d 12.37 (CH3), 20.60 (CH3), 22.38
(CH2), 24.99 (CH2), 26.39 (CH3), 32.30 (CH), 34.49
(CH2), 36.64 (CH2), 37.66 (CH), 41.31(CH2), 45.68
(CH), 51.98 (CH), 80.55 (C), 91.97 (CH), 92.71 (CH),
We also compared the three-dimensional structures of
ester 7i, the most active compound of the series, with
the corresponding ethers 8d (a-isomers) and 9d (b-iso-
mers) (Fig. 3).13 Clearly, there is a similarity in the
three-dimensional structures of 7i and 8d. In both these
compounds the C10–O moieties are placed away from
the trioxane group while in 9d it is very close. This prob-
ably accounts for the better activity profiles shown by a-
isomers of ethers and esters.14
In conclusion, we have prepared a series of lipophilic es-
ter derivatives of dihydroartemisinin, several of which
show better activity profile than b-arteether and artesu-
nic acid. Compound 7i, the most active compound of
the series, is more than twice as active as b-arteether
and artesunic acid.
Acknowledgments
Sandeep Chaudhary is thankful to the Council of Scien-
tific and Industrial Research (CSIR), New Delhi, for the
award of Senior Research Fellowship. We are thankful
to Mr. G.P. Yadav for his help in generating 3D
structures.
References and notes
1. CDRI Communication No.: 6926.
2. For reviews on artemisinin and its analogues, see: (a)
Klayman, D. L. Science 1985, 228, 1049; (b) Luo, X. D.;

1440
C. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1436–1441
104.89 (C), 127.46 (CH), 127.66 (CH), 129.15 (CH), 130.23
(C), 91.84 (CH), 92.00 (CH), 104.78 (C), 172.58 (C);
FABMS (m/z): 461 [M+H]+; ESMS (m/z): 478 [M+NH4]+,
483 [M+Na]+; Anal. Calcd for (C27H40O6): C 70.40 H
8.75; Found C 70.12 H 9.10. Compound 7h: White solid;
FT-IR (KBr, cmÀ1): 2920.0, 2853.7, 1751.5, 1591.3,
(CH), 133.03 (C), 140.49 (C), 141.20 (C), 170.76 (C);
ESMS (m/z): 501 [M+Na]+; Anal. Calcd for (C29H34O6): C
72.77 H 7.16; Found C 72.47 H 7.53. Compound 7c: White
solid; FT-IR (KBr, cmÀ1): 1774.7, 1637.8, 1480.8, 1438.5,
1379.1, 1218.0, 1131.9, 1103.4, 1013.8, 770.4; 1H NMR
(200 MHz, CDCl3) d 0.77 (d, 3H, J = 7.1 Hz, CH3), 0.96
(d, 3H, J = 5.3 Hz, CH3), 1.25–2.07 (m, 10H), 1.44 (s, 3H,
CH3), 2.30–2.44 (m, 1H), 2.50–2.57 (m, 1H), 4.67 (s, 2H),
5.44 (s, 1H, C12–H), 5.80 (d, 1H, J = 9.8 Hz, C10–H), 6.87–
7.62 (m, 9H); 13C NMR (50 MHz, CDCl3) d 12.38 (CH3),
20.60 (CH3), 22.37 (CH2), 24.98 (CH2), 26.32 (CH3), 32.18
(CH), 34.47 (CH2), 36.61 (CH2), 37.68 (CH), 45.64 (CH),
51.93 (CH), 66.08 (CH2), 80.49 (C), 91.99 (CH), 93.07
(CH), 104.96 (C), 113.22 (CH), 122.59 (CH), 127.50 (CH),
128.45 (CH), 128.93 (CH), 130.04 (CH), 131.67 (CH),
138.43 (C), 155.08 (C), 168.32 (C); ESMS (m/z): 512
[M+Na]+; Anal. Calcd for (C29H34O7): C 70.43 H 6.93;
Found C 70.28 H 7.15. Compound 7d: White solid; FT-IR
(KBr, cmÀ1): 2928.9, 2874.4, 1774.3, 1607.4, 1519.4,
1488.0, 1452.0, 1379.4, 1354.4, 1273.2, 1178.5, 1133.1,
1103.1, 1081.8, 1013.2, 764.3; 1H NMR (200 MHz,
CDCl3) d 0.83 (d, 3H, J = 7.0 Hz, CH3), 0.96 (d, 3H,
J = 5.1 Hz, CH3), 1.17–2.07 (m, 10H), 1.43 (s, 3H, CH3),
2.31–2.44 (m, 1H), 2.55–2.62 (m, 1H), 4.70 and 4.80 (2 · d,
2H, J = 16.1 Hz, COCH2O), 5.46 (s, 1H, C12–H), 5.91 (d,
1H, J = 9.8 Hz, C10–H), 6.96–7.55 (m, 9H); ESMS (m/z):
517 [M+Na]+; HR-EIMS for (C29H34O7): Measured
494.2326, Calculated 494.2305; Anal. Calcd for
(C29H34O7Æ0.1H2O): C 70.43 H 6.93; Found C 70.21 H
6.94. Compound 7e: White solid; mp 85–87 ꢁC; FT-IR
(KBr, cmÀ1): 3020.4, 2910.3, 1742.5, 1451.9, 1368.5,
1219.4, 1133.5, 1098.3, 1022.0, 769.0; 1H NMR
(200 MHz, CDCl3) d 0.82 (d, 3H, J = 7.1 Hz, CH3), 0.96
(d, 3H, J = 5.6 Hz, CH3), 1.25–2.01 (m, 25H), 1.42 (s, 3H,
CH3), 2.29–2.43 (m, 1H), 2.53–2.59 (m, 1H), 5.42 (s, 1H,
C12–H), 5.75 (d, 1H, J = 9.7 Hz, C10–H); 13C NMR
(50 MHz, CDCl3) d 12.58 (CH3), 20.63 (CH3), 22.42
(CH2), 24.97 (CH2), 26.29 (CH3), 28.28 (3 · CH), 32.37
(CH2), 34.53(CH2), 36.87 (3 · CH2), 37.64 (CH), 38.98
(3 · CH2), 41.20 (CH), 45.73 (CH), 52.07 (CH), 80.58 (C),
91.94 (CH), 104.71 (C), 176.63 (C); ESMS (m/z): 469
[M+Na]+; Anal. Calcd for (C26H38O6): C 69.92 H 8.57;
Found: C 69.48 H 8.46. Compound 7f: White solid; mp
115–117 ꢁC; FT-IR (KBr, cmÀ1): 2914.3, 1751.0, 1452.2,
1
1457.0, 1378.5, 1352.1, 1161.9, 1096.4, 1022.0; H NMR
(200 MHz, CDCl3) d 0.86 (d, 3H, J = 7.0 Hz, CH3), 0.95
(d, 3H, J = 5.6 Hz, CH3), 1.25–2.10 (m, 27H), 1.43 (s, 3H,
CH3), 2.15–2.24 (m, 2H), 2.36–2.45 (m, 1H), 2.53–2.59 (m,
1H), 5.43 (s, 1H, C12–H), 5.77 (d, 1H, J = 9.8 Hz, C10–H);
13C NMR (50 MHz, CDCl3) d 12.85 (CH3), 20.63 (CH3),
22.39 (CH2), 24.99 (CH2), 26.35 (CH3), 29.01 (3 · CH),
31.99 (CH), 33.22 (C), 34.53 (CH2), 35.15 (CH2), 36.64
(CH2), 37.13 (3 · CH2), 37.69 (CH), 42.74 (3 · CH2),
45.78 (CH), 49.19 (CH2), 52.00 (CH), 80.52 (C), 91.77
(CH), 91.97 (CH), 104.76 (C), 170.93 (C); FABMS (m/z):
475 [M+H]+; ESMS (m/z): 497 [M+Na]+; Anal. for
(C28H42O6): Calcd C 70.86 H 8.92; Found C 70.57 H
8.99. Compound 7i: White solid; FT-IR (KBr, cmÀ1
)
2943.8, 1750.1, 1596.9, 1451.5, 1353.0, 1276.0, 1190.4,
1139.1, 1100.8, 1015.9, 744.8; 1H NMR (200 MHz,
CDCl3) d 0.80 (d, 3H, J = 7.1 Hz, CH3), 0.96 (d, 3H,
J = 5.6 Hz, CH3), 1.25–2.09 (m, 10H), 1.47 (s, 3H, CH3),
2.27–2.48 (m, 1H), 2.68–2.74 (m, 1H), 4.99 (s, 1H, benzylic
H), 5.46 (s, 1H, C12–H), 5.87 (d, 1H, J = 9.8 Hz, C10–H),
7.32–7.45 (m, 4H), 7.65–7.82 (m, 4H); 13C NMR (50 MHz,
CDCl3) d 12.69 (CH3), 20.61 (CH3), 22.34 (CH2), 25.01
(CH2), 26.39 (CH3), 32.35 (CH), 34.48 (CH2), 36.67
(CH2), 37.63 (CH), 45.78 (CH), 51.96 (CH), 53.61 (CH),
80.52 (C), 91.99 (CH), 93.31 (CH), 104.87 (C), 120.33
(CH), 120.48 (CH) 126.21 (CH), 126.79 (CH), 127.66
(CH), 127.82 (CH), 128.60 (CH), 140.49 (C), 140.78 (C),
141.90 (C), 169.87 (C); FABMS (m/z): 476 [M]+,
477[M+H]+; ESMS (m/z): 499 [M+Na]+; Anal. Calcd for
(C29H32O6): C 73.09 H 6.77; Found C 73.38 H 6.73.
Compound 7j: White solid; FT-IR (KBr, cmÀ1) 2929.9,
2877.2, 1747.2, 1653.9, 1529.6, 1450.4, 1350.9, 1275.6,
1216.2, 1145.2, 1097.4, 1014.1, 755.9; 1H NMR (200 MHz,
CDCl3) d 0.84 (d, 3H, J = 7.1 Hz, CH3), 0.98 (d, 3H,
J = 5.3 Hz, CH3), 1.25–2.08 (m, 10H), 1.45 (s, 3H, CH3),
2.31–2.38 (m, 1H), 2.53–2.63 (m, 1H), 2.79 (dd, 1H,
J = 16.12 Hz, 6.55 Hz), 2.96 (dd, 1H, J = 16.4 Hz,
7.56 Hz), 4.44 (br t, 1H, benzylic H), 5.50 (s, 1H, C12–
H), 5.92 (d, 1H, J = 9.8 Hz, C10–H), 7.28–7.41 (m, 4H),
7.51–7.58 (m, 2H), 7.75 (d, 2H, J = 7.4 Hz); 13C NMR
(50 MHz, CDCl3) d 12.61 (CH3), 20.65 (CH3), 22.46
(CH2), 25.03 (CH2), 26.38 (CH3), 32.11 (CH), 34.53
(CH2), 36.63 (CH2), 37.72 (CH), 39.10 (CH2), 43.77
(CH), 45.68 (CH), 51.98 (CH), 80.55 (C), 91.90 (CH),
92.77 (CH), 104.90 (C), 120.30 (CH), 120.42 (CH), 124.77
(CH), 124.94 (CH), 127.68 (CH), 127.88 (CH), 128.01
(CH), 141.20 (C), 146.41 (C), 146.66 (C), 171.88 (C);
ESMS (m/z): 508 [M+NH4]+, 513 [M+Na]+; Anal. Calcd
for (C30H34O6): C 73.45 H 6.99; Found C 73.30 H 6.95.
9. (a) Peters, W. In Chemotherapy and Drug Resistance in
Malaria; Academic Press: London, 1970; pp 64–136; (b) In
vivo antimalarial efficacy test. The blood schizontocidal
activity of the test compounds was evaluated in rodent
model using multidrug-resistant strain of Plasmodium
yoelii nigeriensis. Multidrug-resistant Plasmodium yoelii
nigeriensis used in this study is resistant to chloroquine,
mefloquine and halofantrine. The colony bred Swiss mice
of either sex (20 2 g) were inoculated intraperitoneally
with 1 · 105 P. yoelii (MDR) parasites on day zero, and
treatment was administered to group of five mice at each
dose, from day 0 to 3, once daily. The drug dilutions of all
compounds were prepared in groundnut oil so as to
contain the required amount of the drug (0.6 mg/kg for a
dose of 48 mg/kg, 0.3 mg for a dose of 24 mg/kg, 0.15 mg
1
1401.9, 1375.0, 1267.5, 1161.1, 1096.0, 1020.3; H NMR
(200 MHz, CDCl3) d 0.87 (d, 3H, J = 7.0 Hz, CH3), 0.96
(d, 3H, J = 5.6 Hz, CH3), 1.25–2.08 (m, 25H), 1.42 (s, 3H,
CH3), 2.15–2.23 (m, 2H), 2.29–2.45 (m, 1H), 2.50–2.61 (m,
1H), 5.43 (s, 1H, C12–H), 5.77 (d, 1H, J = 9.8 Hz, C10–H);
13C NMR (50 MHz, CDCl3) d 12.84 (CH3), 20.62 (CH3),
22.38 (CH2), 25.01 (CH2), 26.34 (CH3), 29.00 (3 · CH),
31.98 (CH), 33.21 (C), 34.54 (CH2), 36.63 (CH2), 37.12
(3 · CH2), 37.68 (CH), 42.73 (3 · CH2), 45.77 (CH), 49.18
(CH2), 51.99 (CH), 80.51 (C), 91.76 (CH), 91.96 (CH),
104.75 (C), 170.92 (C); FABMS (m/z): 461 [M+H]+; ESMS
(m/z): 483 [M+Na]+; Anal. Calcd for (C27H40O6): C 70.40
H 8.75; Found C 70.87 H 9.09. Compound 7g: White
solid; mp 73–75 ꢁC; FT-IR (KBr, cmÀ1) 2909.7, 2855.5,
1742.2, 1452.4, 1411.7, 1377.5, 1251.0, 1150.0, 1099.7,
1021.8, 757.0; 1H NMR (200 MHz, CDCl3) d 0.83 (d, 3H,
J = 6.9 Hz, CH3), 0.96 (d, 3H, J = 5.0 Hz, CH3), 1.25–2.06
(m, 25H), 1.43 (s, 3H, CH3), 2.24–2.37 (m, 1H), 2.46–2.57
(m, 3H), 5.44 (s, 1H, C12–H), 5.78 (d, 1H, J = 9.8 Hz, C10–
H); 13C NMR (50 MHz, CDCl3) d 12.46 (CH3), 20.60
(CH3), 22.37 (CH2), 24.99 (CH2), 26.33 (CH3), 28.24
(2 · CH), 31.90 (2 · CH2), 32.17 (3 · CH), 34.51 (CH2),
36.62 (CH2), 37.64 (CH), 38.56 (2 · CH2), 39.24
(2 · CH2), 41.36 (CH), 45.70 (CH), 51.97 (CH), 80.50

C. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1436–1441
1441
for a dose of 12 mg/kg) in 0.1 mL and administered orally
for each dose. Parasitaemia level was recorded from thin
blood smears on day 4 and subsequently twice a week until
day 28. The animals which did not develop patent
infection until day 28 were recorded as cured.15 Mice
treated with b-arteether served as positive control.
Posner, G. H. J. Med. Chem. 2006, 49(9), 2731–2734; (f)
Posner, G. H.; Paik, I.-H.; Chang, W.; Borstnik, K.;
Sinishtaj, S.; Rosenthal, A. S.; Shapiro, T. A. J. Med.
Chem. 2007, 50(10), 2516–2519.
12. (a) 100% suppression of parasitamia means no parasites
were detected in 50 oil immersion microscopic fields
(parasites if at all present were below the detection limit).
The parasites present below the detection limit can
multiply and eventually can be detected during observa-
tion on subsequent days. In such cases though the drug is
providing near 100% suppression of the parasitaemia on
day 4 it will not provide full protection to the treated mice
in the 28 day survival assay; (b) 100% protection means
none of the treated mice developed patent infection during
the 28 days observation period and hence were recorded as
cured. Similarly 60% protection means only 3 out of 5
mice were cured.
10. (a) Also in our programme on synthetic 1,2,4-trioxanes,
we had observed that trioxane built around these moieties
showed promising antimalarial activity: Singh, C.; Kan-
chan, R.; Puri, S. K. Indian Patent Appl. No. 1554 DEL
99, 1999; (b) Singh, C.; Tiwari, P.; Puri, S. K. U.S. Patent
6,737,438 B2, 2004; (c) Singh, C.; Kanchan, R.; Sharma,
U.; Puri, S. K. J. Med. Chem. 2007, 50(3), 521–527.
11. (a) Hindley, S.; Ward, S. A.; Storr, R. C.; Searle, N. L.;
Bray, P. G.; Park, B. K.; Davies, J.; O’Neill, P. M. J. Med.
Chem. 2002, 45, 1052–1063; (b) Avery, M. A.; Alvim-
Gaston, M.; Vroman, J. A.; Wu, B.; Ager, A.; Peters, W.;
Robinson, B. L.; Charman, W. J. Med. Chem. 2002, 45,
4321–4335; (c) Posner, G. H.; Paik, I.-H.; Sur, S.;
McRiner, A. J.; Borstnik, K.; Xie, S.; Shapiro, T. A.
J. Med. Chem. 2003, 46, 1060–1065; (d) Grellepois, F.;
Chorki, F.; Ourevitch, M.; Charneau, S.; Grellier, P.;
McIntosh, K. A.; Charman, W. N.; Pradines, B.; Crousse,
B.; Bonnet-delpon, D.; Begue, J. P. J. Med. Chem. 2004,
47, 1423–1433; (e) Paik, I.-H.; Xie, S.; Shapiro, T. A.;
Labonte, T.; Narducci Sarjeant, A. A.; Baege, A. C.;
13. Energy minimized three-dimensional structures were gen-
erated using Builder (Insight II) module.
14. Following a suggestion from one of the reviewers, we
compared the stabilities of artemether 3 and ester 7i under
acidic conditions (1:1 mixture of THF and 10% aqueous
HCl). While artemether was completely degraded within
96 h, 80% of the ester 7i was recovered unchanged under
the same conditions.
15. Puri, S. K.; Singh, N. Expl. Parasitol. 2000, 94, 8–14.