Mechanism-based design of parasite-targeted artemisinin derivatives: Synthesis and antimalarial activity of benzylamino and alkylamino ether analogues of artemisinin

Article abstract of DOI:10.1021/jm9604944

Several artemisinin derivatives linked to benzylamino and alkylamino groups were synthesized in order to enhance accumulation within the malaria parasite. The in vitro antimalarial activity was assessed against the chloroquine sensitive HB3 strain and the

Full text of DOI:10.1021/jm9604944

J . Med. Chem. 1996, 39, 4511-4514
4511
Mech a n ism -Ba sed Design of P a r a site-Ta r geted Ar tem isin in Der iva tives:
Syn th esis a n d An tim a la r ia l Activity of Ben zyla m in o a n d Alk yla m in o Eth er
An a logu es of Ar tem isin in
Paul M. O’Neill,* Laurence P. Bishop, Richard C. Storr,† Shaun R. Hawley, J ames L. Maggs,
Stephen A. Ward, and B. Kevin Park
Departments of Chemistry and of Pharmacology and Therapeutics, The University of Liverpool,
P.O. Box 147, Liverpool L69 3BX, U.K.
Received J uly 8, 1996X
Several artemisinin derivatives linked to benzylamino and alkylamino groups were synthesized
in order to enhance accumulation within the malaria parasite. The in vitro antimalarial activity
was assessed against the chloroquine sensitive HB3 strain and the chloroquine resistant K1
strain of Plasmodium falciparum. In general the incorporation of amino functionality enhances
the activity relative to artemisinin. The most potent analogue in the series was compound 6
which was severalfold more active than artemisinin against both strains of P. falciparum used
in the study.
Ch a r t 1. Structures of Artemisinin, Dihydroartemisinin,
and Radical Intermediates 3 and 4
In tr od u ction
Malaria is one of the world's most deadly diseases and
is becoming an increasingly serious problem as malaria
parasites develop resistance to drugs such as chloro-
quine and mefloquine. There is, therefore, considerable
urgency to develop new classes of antimalarials. Arte-
misinin (1) (qinghaosu) is an unusual 1,2,4-trioxane
which has been used clinically in China for the treat-
ment of multidrug resistant Plasmodium falciparum
malaria.1 However the clinical application of artemisi-
nin has been limited by the drug's pharmacokinetic
properties. This has provided the impetus for the
investigation of derivatives of this compound, some of
which include esters and ethers of the the corresponding
lactol, dihydroartemisinin (DHA) (2). Analogues of this
type are currently being developed as potent and rapidly
acting antimalarials.2
Studies on the mode of action of artemisinin and
related 1,2,4-trioxanes have indicated that the peroxide
bridge is cleaved homolytically by heme (ferriprotopor-
phyrin IX), a ubiquitous cellular component of P. falci-
parum, to give a reactive intermediate that alkylates
vital parasite protein molecules.3 There is evidence that
this key intermediate is the radical at C-4 (4) formed
by an intramolecular abstraction of the 4RH by the
initially formed alkoxy radical 3.4
to enhanced cleavage and generation of the required
“cytotoxic radical species”.7 However, neither of these
approaches resulted in compounds with significantly
greater antimalarial potency.
Existing 4-aminoquinoline antimalarials such as chlo-
roquine and amodiaquine incorporate a basic alky-
lamino side chain, the role of which is probably 2-fold.
Firstly, protonation of the drug within the acidic vacuole
of the parasite assists accumulation since the resultant
cation cannot pass out of the parasite.8 Secondly, the
protonated alkylamino groups might also assist binding
to the proposed cellular receptor, heme,9 within the
acidic food vacuole, i.e., the site of action of antimalarial
peroxides.
We have linked artemisinin to benzylamino and
alkylamino groups (Chart 2) with the aim of increasing
the activity of artemisinin by enhancing the intracel-
lular accumulation of drug within the parasite food
vacuole by “ion trapping”. This should increase the
amount of drug available for interaction with heme and
hence generation of the required alkylating species.
Several “mechanism-based approaches” have been
investigated for improving the antimalarial activity of
artemisinin derivatives. In a recent study Posner has
investigated the effect of radical-stabilizing groups at
the C-4 position in the artemisinin framework.5,6 It was
proposed that incorporation of such groups would
increase the formation of the carbon-centred radical and
that this would be in turn reflected by increased
antimalarial activity. In another approach Yuthavong
synthesized a series of analogues covalently linked to
iron chelators. The rationale behind this series of
compounds was that the chelation of available “free
iron” in the food vacuole of the parasite would locate
available Fe(III)/Fe(II) species close to the peroxide
bridge of the artemisinin derivative leading ultimately
Ch em istr y
The preparation of the benzylamino ether derivatives
of artemisinin is shown in Scheme 1. Artemisinin (1)
was reduced with sodium borohydride to give DHA (2)
† Department of Chemistry.
X Abstract published in Advance ACS Abstracts, September 1, 1996.
S0022-2623(96)00494-3 CCC: $12.00 © 1996 American Chemical Society

4512 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22
Notes
Ch a r t 2. Structures of Amine-Substituted Artemisinin
Ch a r t 3. Amine Peroxide Antimalarials
Derivatives
coupled with 2-bromoethanol in anhydrous ether using
BF3 etherate catalysis to give a mixture of R and â
stereoisomers.11 The â isomer (10, R ) Br) was sepa-
rated by column chromatography to give the product as
a yellow solid. The bromo compound was then allowed
to react with the appropriate amine in dioxane at 50
°C for 1 h, and the product was finally purified by
column chromatography.
0
Br
An tim a la r ia l Activity
The antimalarial activity of chloroquine and the
artemisinin analogues was determined12,13 against two
strains of P. falciparum: the uncloned K1 strain (chlo-
roquine resistant) and the HB3 strain (chloroquine
sensitive) using standard techniques.
Sch em e 1. Synthesis of Benzylamino and Alkylamino
Ether Analogues of Artemisinin
Resu lts a n d Discu ssion
As formulated by De Duve,14,15 the explanation of the
weak base effect is based upon two main assumptions:
(i) the neutral (uncharged) forms of the weak bases
readily cross both plasma and vesicle membranes and
(ii) these membranes are impermeable (or much less
permeable) to the protonated forms of the bases. The
logical consequences of these assumptions are consistent
with the known effects of weak bases on both mam-
malian cells and plasmodia,16 i.e., weak bases are
concentrated by protonation in their nondiffusable form
within acidic intracellular vesicles.
In addition to the wide variety of artemisinin ana-
logues that have been examined to date,17 simple
peroxides such as tert-butyl hydroperoxide and hydrogen
peroxide have been screened and found to have limited
antimalarial potency.18 Since both of these agents
result in unwanted side effects, such as hemolysis of
uninfected erythrocytes at parasiticidal concentrations,
Vennerstrom synthesized a series of amine peroxides
(e.g., 13 and 14) with the idea that a selective drug
delivery would circumvent toxicity to the host.18 The
rationale for the intended specificity of action was the
selective concentration of these weak base derivatives
in the acidic digestive vacuoles of the parasite (pH g
5.0) where the oxidant sensitivity of the malaria para-
site could be exploited to maximum effect. The study
revealed that linking tert-butyl peroxide to various
amine groups enhanced antimalarial activity by 1 order
of magnitude.
The purpose of the present study was to explore the
effect on antimalarial activity of linking artemisinin to
different amine functionalities. It was anticipated that
the incorporation of a moiety that can undergo proto-
nation in the acidic food vacuole of the parasite would
enhance cellular accumulation by ion trapping and
thereby provide larger quantities of drug available for
interaction with heme, the “cellular activator” of anti-
malarial peroxide drugs. It can be seen from Table 1
that all of the analogues tested have significant activity
against both strains of P. falciparum used in the study.
Of the benzylamino compounds, the derivatives 6 and
9 are the most active against the chloroquine sensitive
according to the literature procedure.10 Treatment of
(2) with 4-(hydroxymethyl)benzyl alcohol, in the pres-
ence of BF3 etherate catalyst, gave the required benzyl
alcohol 5, (Scheme 1) in 80% yield. NMR analysis of
the product revealed a small coupling (J ) 3.30 Hz,
C9H-C10H) indicating that the stereochemistry of the
product was â at the C-10 carbon atom. Treatment of
the benzylic alcohol in dichloromethane with mesyl
chloride in the presence of triethylamine at low tem-
perature gave the mesylate which was used without
further purification in the next step of the synthesis.
The solvent was removed, dry dioxane was added, and
the mesylate was allowed to react with 2 equiv of the
appropriate amine for 1 h at 40 °C. The solvent was
removed, and the required product was purified by
preparative thin layer chromatography.
Two alkylamino ether derivatives were also synthe-
sized for comparison according to Scheme 1. DHA was

Notes
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22 4513
Ta ble 1. Antimalarial Activity against the HB3 and K1
Strains of P. falciparum
Assays were performed in sterile 96-well microliter plates, each
well containing 100 µL of medium which was seeded with 10
µL of a parasitized red blood cell mixture to give a resulting
initial parasitemia of 1% with a 5% hematocrit. Control wells
(which constituted 100% parasite growth) consisted of the
above, with the omission of the drug.
HB3 IC50
(nM)
K1 IC50
(nM)
drug
(SE
(SE
chloroquine
artemisinin (1)
5
6
7
8
20
1.1
0.6
0.8
0.9
1.9
9
250
6.4
4.4
1.4
9.8
35
7.1
0.5
0.9
0.2
1.7
17
0.3
0.4
0.04
7.3
4.2
3.1
12.2
50
After 24 h incubation at 37 °C, 0.5 µCi of [3H]hypoxanthine
was added to each well. After a further 24 h incubation, the
cells were harvested onto filter mats, dried overnight, placed
in scintillation vials with 4 mL of scintillation fluid, and
counted on a liquid scintillation counter.
IC50 values were calculated by interpolation of the probit
transformation of the log dose-response curve. Each com-
pound was tested in triplicate against both strains to ensure
reproducibility of the results.
9
11
12
2.3
2.7
4.1
0.3
0.4
0.04
2.3
2.4
4.1
Ch em istr y. Merck Kieselgel 60 F 254 precoated silica
plates for TLC were obtained from BDH, Poole, Dorset, U.K.
Column chromatography was carried out on Merck 938S silica
gel. Infra red (IR) spectra were recorded in the range 4000-
600 cm-1 using a Perkin Elmer 298 infrared spectrometer.
Spectra of liquids were taken as films. Sodium chloride plates
(Nujol mull) and solution cells (dichloromethane) were used
as indicated.
Proton NMR spectra were recorded using Perkin Elmer R34
(220 MHz) and Bruker (400 and 200 MHz) NMR spectrom-
eters. Solvents are indicated in the text, and tetramethylsi-
lane was used as an internal reference. Mass spectra were
recorded at 70 eV using a VG7070E mass spectrometer. The
samples were introduced using a direct insertion probe. In
the text the parent ion (M+) is given followed by peaks
corresponding to major fragment losses with intensities in
parentheses.
HB3 strain, having approximately 2-fold greater activity
than artemisinin and being several times more active
than chloroquine. Analogue 6 was also the most active
compound tested against the chloroquine resistant K1
strain. However, although the derivatives 6-9 show
good activity, it is clear that there is only slight
improvement over compound 5 which was included in
the study as a control. This analogue is also more
potent than artemisinin but does not contain an amino
group. Thus the observed increases in activity for
analogues 6 and 9 may be on account of not only
increased cellular accumulation but also increased
lipophilicity brought about by the presence of the
substituted benzene ring.
In general the rank order of drug activity within each
amine series was similiar for both strains of plasmodia.
Notably the morpholino compound 8 had reduced activ-
ity. The two alkylamino derivatives 11 and 12 had
comparable activity in both strains with slightly im-
proved activity relative to artemisinin. Interestingly
none of the derivatives was subject to resistance; all of
the analogues have equipotent activity against chloro-
quine resistant and sensitive isolates.
1 0 â-[ [ p -( H y d r o x y m e t h y l ) b e n z y l ] o x y ] d i h y d r o -
a r tem isin in (5). Dihydroartemisinin (0.5 g, 1.77 mmol) was
dissolved in anhydrous ether (70 mL) under nitrogen. BF3‚-
Et2O (0.25 mL) was then added by syringe followed by
4-(hydroxymethyl)benzyl alcohol (0.37 g, 2.65 mmol). The
reaction mixture was left to stir for 20 h, after which water
was added. The ether layer was washed with sodium sulfate
solution (30% w/v) and dried with magnesium sulfate and the
solvent removed under reduced pressure to give the crude
product as an oil. Column chromatography (silica, 40% ethyl
acetate/60% petroleum ether) gave the product as a colorless
viscous syrup (0.56 g, 78%): 1H NMR (CDCl3, 200 MHz) δ
7.08-7.20 (m, 4H, Ar-H), 5.30 (s, 1H, C-H at C12), 4.75 (d,
1H, J ) 3.30 Hz, C-H at C10), 4.73 (d, 1H, 4.52, J ) 12.10 Hz,
-OCH2Ar), 4.52 (s, 2H, CH2OH), 4.35 (d, 1H, J ) 12.10 Hz,
-OCH2Ar), 2.50 (m, 1H), 2.25 (dt, 1H), 1.10-1.95 (m, 10H),
1.30 (s, 3H, CH3 at C3), 0.90 (d, 3H, CH3 at C6), 0.85 (d, 3H,
CH3 at C9); 13CNMR (CDCl3, 400 MHz) 140, 127, 126, 104,
101, 87, 81, 69, 65, 52, 44, 37, 36, 34, 30, 26, 24.5, 20, 12; LCMS
Su m m a r y a n d Con clu sion
The aim of the present study was to develop simple
chemical routes to a series of amine-functionalized
artemisinin derivatives and to investigate the effect of
amine substitution on antimalarial activity. The fact
that all the compounds tested have good activity il-
lustrates the potential of selectively targeting the
malarial parasite, and further studies are in progress
to examine whether linking artemisinin to a range of
diprotic side chains can enhance antimalarial activity
further.
(75 V, MeOH-AcOH) 437 (M + MeOH + 1, 100), 422 (M+
+
NH4, 65), 405 (M + 1, 1), 359 (M + 1 - CO2H2, 9), 267 (M +
1 - CHPhCH2OH - H2O, 75), 221 (14); CIMS 422 (M+ + NH4,
8), 376 (M+ - HCO2H, 12), 359 (M+ - HCO2H - NH3, 100),
317 (8), 284 (39), 267 (36), 251 (16), 221 (96), 138 (33); IR (CH2-
Cl2) 3510 (OH), 1420, 1260, 1200, 890 (O-O), 815 (O-O). Anal.
C,H.
Exp er im en ta l Section
An tim a la r ia l Testin g. Two strains of P. falciparum from
Thailand were used in this study: (a) the uncloned K1 strain,
which is known to be chloroquine resistant, and (b) the HB3
strain, which is sensitive to all antimalarials.
Parasites were maintained in continuous culture using a
method derived from that of J ensen and Trager.12 Cultures
were maintained in culture flasks containing human eryth-
rocytes (2-5%) with parasitemia ranging from 0.1% to 10%.
The parasites were suspended in RPMI 1640 medium supple-
mented with 25 mM HEPES buffer, 32 mM NaHCO3, and 10%
human serum (complete medium). Cultures were gassed with
a mixture of 3% O2, 4% CO2, and 93% N2.
Antimalarial activity was assessed by an adaptation of the
48 h sensitivity assay of Desjardins et al.13 which uses [3H]-
hypoxanthine incorporation as an assessment of parasite
growth. Stock drug solutions were dissolved in 100% ethanol
and diluted to an appropriate concentration with complete
medium (final concentrations contained less than 1% ethanol).
1 0 â-[[p -[(D i e t h y l a m i n o )m e t h y l ]b e n z y l ]o x y ]d i -
h yd r oa r tem isin in (6). The benzyl alcohol derivative 5 (0.10
g, 0.246 mmol) was dissolved in dry dichloromethane (10 mL)
under nitrogen, and the solution was cooled to 0 °C. Mesyl
chloride (0.42 g, 0.36 mmol) was added, and the solution was
allowed to stir for 45 min (TLC). The solvent was removed
under reduced pressure, and the residue was dissolved in
dioxane (5 mL). The reaction vessel was flushed with nitrogen,
and diethylamine (0.035 g, 0.50 mmol, 0.05 mL) was added
via syringe. The reaction mixture was heated at 50 °C for 1
h, and the solvent was removed under reduced pressure. The
residue was applied to a preparative TLC plate (1% MeOH/
99% DCM), and the product was obtained as a colorless oil,
0.52 g (49%): 1H NMR (CDCl3, 200 MHz) δ 7.11-7.31 (m, 4H,
Ar-H), 5.40 (s, 1H, C-H at C12), 4.90 (d, 1H, J ) 3.10 Hz, CH
at C10), 4.84 (d, 1H, J ) 12.10 Hz, -OCH2Ar), 4.45 (d, 1H, J
) 12.10 Hz, -OCH2Ar), 3.50 (s, 2H, CH2N), 2.60 (m, 1H), 2.45
(q, 4H, NCH2CH3), 2.32 (dt, 1H), 1.10-2.12 (10H, m), 1.40 (s,

4514 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22
Notes
3H, CH3 at C3), 0.96 (t, 6H, NCH2CH3), 0.89 (d, 3H, CH3 at
C6), 0.85 (d, 3H, CH3 at C9);13CNMR (CDCl3, 400 MHz) 136,
128, 127, 104, 101, 87, 81, 69, 57, 52, 46, 44, 37, 36, 34, 30, 26,
24, 20, 12, 11; LCMS (75 V, 50-75% MeOH-AcOH) 460 (M+
+ MeOH + 1, 100), 401(M+ - HCO2H), 372 (23), 195 (14), 176
(M + 1 - DHA, 15); IR (CH2Cl2) 3056, 2987, 1421, 1258, 1193,
1175, 1158, 1139, 1099, 895 (O-O), 820 (O-O). Anal. C,H,N.
10â-[[p -(P i p e r i d i n o m e t h y l)b e n z y l]o x y ]d i h y d r o -
a r tem isin in (7). This compound was prepared as described
for 6. The yield was 40%.: 1H NMR (CDCl3, 200 MHz) δ 7.10-
7.30 (m, 4H, Ar-H), 5.45 (s, 1H, C-H at C12), 4.90 (d, 1H, CH
at C10), 4.86 (d, 1H, J ) 12.65 Hz, -OCH2Ar), 4.50 (d, 1H, J
) 12.65 Hz, -OCH2Ar), 3.53 (s, 2H, CH2N), 2.65 (m, 1H), 2.40
(m, 5H), 1.62 (m, 4H, NCH2CH2-), 1.44 (s, 3H, CH3 at C3),
1.15-2.10 (m, 11H), 0.96 (d, 3H, CH3 at C6), 0.92 (d, 3H, CH3
at C9); 13C NMR (CDCl3, 400 MHz) 137, 136, 129, 104, 101,
87, 81, 69, 67, 63, 54, 52, 44, 37, 34, 30, 26, 25, 24.64, 24.48,
24.03, 20, 13, 9; LCMS (85 V, 75% MeOH-AcOH) 494 (M +
Na, 100), 472 (M+ + 1, 100); IR (CH2Cl2) 3050, 2900, 1430,
1260, 1175, 895 (O-O), 820 (O-O). Anal. C,H,N.
(2H, m, -OCH2), 2.80-3.20 (4H, m, CH2NCH2), 2.40 (dt, 1H),
1.10-1.95 (m, 10H), 1.30 (s, 3H, CH3 at C3), 1.12 (t, 3H,
NCH2CH3), 0.90 (s, 3H, CH3 at C6), 0.89 (s, 3H, CH3 at C9);
IR (KBr) 1700, 1630, 1580, 1470, 1360, 825 (O-O). Anal.
C,H,N.
Ack n ow led gm en t. We thank the Wellcome Trust
(B.K.P., S.A.W.), MRC (S.R.H.), Roche (Welwyn) (P.M.O.)
and the University of Liverpool for financial support.
The authors also thank the WHO for supplying arte-
misinin.
Refer en ces
(1) Zaman, S. S.; Shrma, R. P. Some Aspects of the Chemistry and
Biological Activity of Artemisinin and Related Antimalarials.
Heterocycles 1991, 32, 1593-1638.
(2) Klayman, D. L. Quinghaosu (Artemisinin):an antimalarial drug
from China. Science 1985, 228, 1049-1055.
10â-[[p -(Mo r p h o lin o m e t h y l)b e n zy l]o x y ]d ih y d r o -
a r tem isin in (8). This compound was prepared as described
for 6. 8: 1H NMR (CDCl3, 200 MHz) δ 7.18-7.30 (m, 4H, Ar-
H), 5.45 (s, 1H, C-H at C12), 4.91 (d, 1H, J ) 4.10 Hz, CH at
C10), 4.89 (d, 1H, J ) 12.65 Hz, -OCH2Ar), 4.50 (d, 1H, J )
12.65 Hz, -OCH2Ar), 3.70 (m, 4H, NCH2CH2O), 3.50 (s, 2H,
CH2N), 2.67 (m, 1H), 2.45 (m, 4H, NCH2CH2O-), 1.45 (s, 3H,
CH3 at C3), 1.25-2.10 (m, 11H), 0.97 (d, 3H, CH3 at C6), 0.93
(d, 3H, CH3 at C9); LCMS (85 V, 75% MeOH-AcOH) 474 (M+
+ 1, 100), 190 (10); IR (CH2Cl2) 3050, 895 (O-O), 820 (O-O).
Anal. C,H,N.
10â-[[p -(P y r r o lid in o m e t h y l)b e n zy l]o x y ]d ih y d r o -
a r tem isin in (9). This compound was prepared as described
for 6. The yield after preparative TLC was 40%: 1H NMR
(CDCl3, 200 MHz) δ 7.43 (m, 4H, Ar-H), 5.31 (s, 1H, C-H at
C12), 4.95 (d, 1H, J ) 12.65 Hz, -OCH2Ar), 4.91 (d, 1H, J )
3.50 Hz, C-H at C10), 4.55 (d, 1H, J ) 12.65 Hz, -OCH2Ar),
4.06 (s, 2H, CH2N), 3.06 (m, 4H,NCH2CH2), 2.67 (m, 1H), 2.40
(dt, 1H), 2.05 (m, 4H, NCH2CH2), 1.34 (s, 3H, CH3 at C3),
1.22-1.87 (m, 10H), 0.96 (d, 3H, CH3 at C6), 0.94 (d, 3H, CH3
at C9); LCMS (75 V, 50-75% MeOH-AcOH) 458 (M+ + 1,
100), 444 (7), 430 (23), 412 (7), 174 (28); IR (CH2Cl2) 3060,
2951, 1421, 1272, 895 (O-O), 820 (O-O). Anal. C,H,N.
10â-(2-Br om oeth oxy)d ih yd r oa r tem isin in (10). Dihy-
droartemisinin (0.25 g, 0.88 mmol) was dissolved in dry ether,
and the reaction vessel was flushed with nitrogen. 2-Bromo-
ethanol (2 equiv) was added via syringe, and the reaction
mixture was allowed to stir at room temperature overnight.
The solvent was removed under reduced pressure, and the
mixture of R and â isomers was separated by column chroma-
tography (hexane/ethyl acetate, 9:1): 1H NMR (CDCl3, 200
MHz) δ 5.38 (s, 1H, C-H at C12), 4.71 (d, 1H, J ) 3.60 Hz,
C-H at C10), 3.78-4.11 (m, 2H), 3.48 (m, 2H, CH2Br), 2.65
(m, 1H), 2.40 (dt, 1H), 1.10-1.95 (m, 10H), 1.30 (s, 3H, CH3
at C3), 0.90 (s, 3H, CH3 at C6), 0.89 (s, 3H, CH3 at C9); MS
374, 376 (M+, 5), 314, 316; IR (CH2Cl2) 895 (O-O), 818 (O-O).
Anal. C,H.
10â-[2-(N-Meth ylam in o)eth oxy]dih ydr oar tem isin in (11).
The bromo compound 11 (0.20 g, 5.33 mmol) was dissolved in
dioxane; 2 equiv of aqueous methylamine was added, and the
solution was heated at 50 °C for 1.2 h. The product was
purified by preparative TLC to give 11 as an oil: 1H NMR
(CDCl3, 200 MHz) δ 5.36 (s, 1H, C-H at C12), 4.76 (d, 1H, J )
3.30 Hz, C-H at C10), 3.50-3.93 (m, 2H), 2.73 (m, 2H, CH2N),
2.42 (s, 3H, N-CH3), 2.40 (dt, 1H), 2.11 (m, 1H, NH), 1.10-
1.95 (m, 11H), 1.30 (s, 3H, CH3 at C3), 0.90 (s, 3H, CH3 at
C6), 0.89 (s, 3H, CH3 at C9); MS 374, 376 (M+, 5), 314, 316;
IR (CH2Cl2) 895 (O-O), 818 (O-O). Anal. C,H,N.
10â-[2-(Eth yla m in o)eth oxy]d ih yd r oa r tem isin in (12).
The bromo compound 11 (0.20 g, 5.33 mmol) was dissolved in
dioxane; 2 equiv of aqueous ethylamine (70%) was added, and
the solution was heated at 50 °C for 1.2 h as for 11. The
product was purified by preparative TLC to give 12 as an oil
which was converted to the corresponding maleate salt: 1H
NMR (CDCl3, 200 MHz) 5.94 (s, 2H, CH:CH), δ 5.36 (s, 1H,
C-H at C12), 4.62 (d, 1H, J ) 3.30 Hz, C-H at C10), 3.32-4.01
(3) Asawamahasakda, W.; Ittarat, I.; Pu, Y.-M.; Ziffer, H.; Meshnick,
S. R. Alkylation of Parasite Specific Proteins by Endoperoxide
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(4) Posner, G. H.; Oh, C. H. A Regiospecifically Oxygen-18 Labelled
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Mechanism(s) for the Antimalarial Activity of Artemisinin
(Quinghaosu). J . Am. Chem. Soc., 1992, 114, 8328-8329.
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Meshnick, S. R.; Asawamahasakda, W. Mechanism-Based De-
sign, Synthesis and In Vitro Antimalarial Testing of New
4-Methylated Trioxanes Structurally Related to Artemisinin:The
Importance of a Carbon Centred Radical for Antimalarial
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