Synthesis and biological evaluation of febrifugine analogues as potential antimalarial agents

Article abstract of DOI:10.1016/j.bmc.2009.05.011

Febrifugine is an alkaloid isolated from Dichroa febrifuga Lour as the active component against Plasmodium falciparum. Adverse side effects have precluded febrifugine as a potential clinical drug. In this study novel febrifugine analogues were designed an

Full text of DOI:10.1016/j.bmc.2009.05.011

Bioorganic & Medicinal Chemistry 17 (2009) 4496–4502
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and biological evaluation of febrifugine analogues as potential
antimalarial agents
a
b
a
a
c
Shuren Zhu a, , Quan Zhang , Chandrashekar Gudise , Lai Wei , Erika Smith , Yuling Zeng
*
a Radix Pharmaceuticals, Inc., 13303 Sunny Brooke PL, Rm 101, Potomac, MD 20854, USA
b Indian Institute of Chemical Technology, Uppal Road, Hyderabad 500 007, India
c Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Shanghai 201203, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 18 February 2009
Revised 2 May 2009
Accepted 5 May 2009
Available online 9 May 2009
Febrifugine is an alkaloid isolated from Dichroa febrifuga Lour as the active component against Plasmo-
dium falciparum. Adverse side effects have precluded febrifugine as a potential clinical drug. In this study
novel febrifugine analogues were designed and synthesized. Lower toxicity was achieved by reducing or
eliminating the tendency of forming chemically reactive and toxic intermediates and metabolites. Syn-
thesized compounds were evaluated for acute toxicity and in vitro and in vivo antimalarial efficacy. Some
compounds are much less toxic than the natural product febrifugine and existing antimalarial drug chlo-
roquine and are expected to possess wide therapeutic windows. These compounds, as well as the under-
lying design rationale, may find usefulness in the discovery and development of new antimalarial drugs.
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Antimalarial
Febrifugine
1. Introduction
its availability. Subsequent pre-clinical researches have found that
febrifugine possesses adverse side effects. Strong liver toxicity has
Malaria is one of the most common infectious diseases in over
100 countries in Africa, Southeast Asia, and South America. It kills
more people than any other communicable disease except for
tuberculosis. Worldwide, malaria affects approximately 300 mil-
lion patients, leading to more than 2 million death a year.1 The
increasing prevalence of multiple drug resistant strains of Plasmo-
dium falciparum in most malaria endemic areas has significantly re-
duced the efficacy of current antimalarial drugs for prophylaxis
and treatment of this disease. For instance, resistance to the inex-
pensive antimalarial mainstays, such as chloroquine, is worldwide.
Similarly, resistance to mefloquine, which was proposed as the
drug of choice for chloroquine-resistant malaria, has been reported
from Africa and Southeast Asia.2,3 Therefore, medicinal agents
based on novel mode of action are required to overcome the emer-
gence of resistance and to control an ever-increasing number of
epidemics caused by the malaria parasite.
Febrifugine (1) and isofebrifugine (2) were isolated as the active
components against malaria in the Chinese herb Chang Shan (Dich-
roa febrifuga Lour),4,5 which has been employed by the local people
as medicine against fevers caused by malaria parasites for a long
time. Febrifugine acts by impairing haemazoin formation required
for maturation of the parasite at the trophozoite stage.6 The use of
febrifugine as antimalarial agent is initially appealing not only be-
cause of its rapid effect and no drug resistance, but also because of
precluded febrifugine as a clinical drug.7–9 Major drug toxicities
can be grouped into five categories in terms of the underlying tox-
icity mechanism: (I) on-target, or mechanism-relayed, toxicity; (II)
hypersensitivity and related immunological reactions; (III) off-tar-
get pharmacology; (IV) biological activation to toxic metabolites;
(V) idiosyncratic toxicities. One concept in the study of toxicity is
the so-called ‘metabolic activation’ of drugs and other chemicals.
One explanation is that drugs can be transformed into reactive
electrophiles, which bind covalently with proteins, DNA and low-
molecular-mass nucleophiles such as glutathione (GSH).
Total synthesis and structural modifications of febrifugine have
been made.10–16 It has been elucidated the essential role played by
the 4-quinazolinone ring and 100-amino group in the appearance of
activity. It is also known that febrifugine 1 and isofebrifugine 2 are
in equilibrium when they are in solution (Scheme 1). This equilib-
rium most likely follows a three-step sequence: (i) retro-Michael
reaction (1?3); (ii) Michael addition (3?4); (iii) hemi-ketal cycli-
zation (4?2). Intermediate 3, as a Michael acceptor, is highly elec-
trophilic. If half-life of 3 is sufficiently long, then it is very possible
that this electrophile will non-selectively alkylate biomolecules
such as proteins, peptides, DNA, or RNA inside the host cell, result-
ing in the observed toxicity.
Febrifugine could be metabolized to the corresponding arene
oxide 5 by cytochrome P-450 enzymes (Scheme 2). When arene
oxide 5 escapes deactivation process by certain enzymes such as
epoxide hydrase or glutathione S-transferase, toxicity can result
because this reactive electrophile will form covalent adducts with
* Corresponding author. Tel.: +1 301 675 3714; fax: +1 301 424 1927.
E-mail address: shuren.zhu@gmail.com (S. Zhu).
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.05.011

S. Zhu et al. / Bioorg. Med. Chem. 17 (2009) 4496–4502
4497
As a comparative study, compounds 21 and 22 were also de-
signed. These compounds each bear an electron-donating methoxy
group on the C-7 or C-8 position of the aromatic ring. These com-
pounds have comparable or even greater tendency to undergo oxi-
dation. Biological data from these compounds should further
validate, or nullify, the hypothesis that some oxidized febrifugine
metabolites have contributed to the observed toxicity.
2. Chemistry
Scheme 3 illustrated the synthesis of compound 7. Other com-
pounds (8–22) were synthesized by similar procedures. The known
alcohol 23, available from borane reduction of commercially avail-
able trans-4-benzyloxy-N-Cbz-L-proline, was converted to the cor-
responding alkyl iodide 24. Coupling between 24 and vinyl
magnesium bromide, in the presence of copper(I) iodide, furnished
alkene 25 in good yield.18 Although initial oxidation of alkene 25
by mCPBA to give oxirane 26 was not successful, the transforma-
tion was smooth and high-yielding via a three-step sequence: (i)
dihydroxylation; (ii) tosylation of the primary alcohol; and (iii)
epoxide ring formation facilitated by potassium carbonate in
methanol. 3H-Quinazolin-4-one condensed with oxirane 26, to fur-
nish corresponding alcohol 27, as a pair of diastereomers, this fol-
lows the methods of Ogasawara et al.10 Compound 27 then
underwent TPAP oxidation19 and hydrogenolysis, to afford com-
pound 7. The synthesis of compounds 8–22 follows similar route.
The respective aromatic moieties are available from previously
published procedures.20–22
Scheme 1.
DNA, RNA, proteins, or other biomolecules of the host. Such bind-
ing can cause mutations and result in cell damage. In a recent met-
abolic study of febrifugine, Oshima and co-workers isolated
metabolite 6.14 This study indicates that arene oxide 5 is most
probably a short-lived reactive metabolic intermediate because
metabolite 6 would have been derived from intermediate 5
through the rearrangement known as NIH shift.17
There are two possible ways to make this process unfavorable:
(I) Blocking the C-5 or C-6 position of the quinoline ring; and (II)
Increasing the oxidation potential of the molecule. As part of an
ongoing malaria chemotherapy project in our laboratory, we
undertook the synthesis and antimalarial activity evaluation of
some febrifugine analogues (compounds 7–22, shown in Fig. 1).
Lower toxicity of these newly designed compounds will be
achieved by reducing or eliminating the tendency to form chemi-
cally reactive and toxic intermediates. Formation of five-mem-
bered rings are kinetically more favored than that of six-
membered rings due to entropy effect. In the case of compounds
7–22 where the original piperidine ring was replaced by a pyrrol-
idine ring, retro-Michael reaction would less likely occur. Also, the
half-life of the corresponding Michael acceptor, once formed,
would be much shorter. Thus, the host is less exposed to toxic elec-
trophiles resulted from the retro-Michael reaction of the adminis-
tered drug. For compounds 8–16, an extra nitrogen atom or a
substitution group was also introduced on the aromatic ring to
block the C-5 or C-6 position of the quinazolinone ring or to in-
crease the oxidation potential of the molecule. Compounds 17
and 18 each bear a bulky group, on C-5 or C-6 position of the qui-
nazolinone ring. Compounds 19 and 20 each bear a benzyloxy
group on C-300 position of the quinazolinone ring. These com-
pounds are designed to block the aforementioned unwarranted
metabolic pathway. Overall, these compounds closely resemble
febrifugine itself by possessing a planar aromatic ring, a 100-amino
group and C-20, C-300 O-functionality and are therefore expected to
possess same or similar mode of action. Meanwhile, they would be
much less likely to produce toxic intermediates because of the
replacement of the original piperidine ring by a pyrrolidine moiety,
the blocking of C-5 and C-6 position or the increase of the oxida-
tion potential of the molecule.
3. Results and discussion
Synthesized compounds (7–22) were tested against P. falcipa-
rum clones W-2, a chloroquine resistant cell line for in vitro effi-
cacy. All compounds were also evaluated for antimalarial efficacy
and toxicity in mouse models. The data are summarized in Table
1. Most compounds (7–16, 19–22) have exhibited comparable or
superior in vitro and in vivo antimalarial activity, as compared to
parent natural product febrifugine. This confirms earlier observa-
tion that it is crucial to possess a planar aromatic ring, a 100-amino
group and C-20, C-300 O-functionality in order to preserve the excel-
lent antimalarial activity of febrifugine. Compounds 17 and 18,
bearing a bulky tert-butyl group at the C-5 or C-6 position of the
aromatic ring, are less active. The acute toxicity study of new com-
pounds was conducted and the MTDs (maximum tolerated dose)
were determined. Compound 7, bearing a pyrrolidine ring instead
of the original piperidine, becomes 4 times less toxic as compared
to febrifugine itself. Compounds 8–18, where the aromatic ring has
either an extra nitrogen atom, an electron-withdrawing group, or a
bulky group, have shown drastically reduced toxicity.
Compounds 19 and 20 each bears a benzyloxy group on C-300
position of the aromatic ring. They possess excellent in vitro anti-
malarial activity. The IC50’s are comparable to compounds 8 and
15, which bear a free hydroxyl group on C-300 position of the aro-
matic ring. It is interesting to note that the in vivo efficacy of com-
pounds 19 and 20 is superior to that of 8 and 15. The toxicity level
is also elevated. Compounds 19 and 20, with a benzyloxy group on
Scheme 2.

4498
S. Zhu et al. / Bioorg. Med. Chem. 17 (2009) 4496–4502
Figure 1.
Scheme 3.
C-300 position of the aromatic ring, are more lipophilic. Conse-
quently, they may possess improved bioavailability.
In conclusion, new febrifugine analogues were designed and
synthesized. Some of these compounds (8, 9, 19, 20) possess a ther-
apeutic index over ten times superior to that of the commonly used
antimalarial drug chloroquine. These compounds, as well as the
underlying design rationale, may find usefulness in the discovery
and development of new antimalarial drugs.
Although the antimalarial efficacy of compounds 21 and 22 is
about the same as that of compound 7, these compounds are con-
siderably more toxic. Compounds 21 and 22 each possess an elec-
tron-donating methoxy group attached to the C-7 or C-8 of the
aromatic ring and possess even greater tendency to undergo oxida-
tion than that of compound 7. These biological data further vali-
date the hypothesis that some oxidized febrifugine metabolites
have contributed to the observed toxicity.
4. Experimental
4.1. Chemistry
Antimalarial potency was also measured by determination of
the MCD (minimum clearance dose), a commonly used efficacy
measurement. The therapeutic indices, which were obtained by
using the MCDs as the effective parameters, were calculated. Based
on the therapeutic-index data, compounds 8–16, 19, and 20 are
much less toxic than the natural product febrifugine and existing
antimalarial drug chloroquine and are expected to possess wide
therapeutic windows. Four compounds (17, 18, 21, and 22) possess
narrow therapeutic windows. They are either less active (com-
pounds 17, 18) or quite toxic (compounds 21, 22).
Melting points were determined on a Mettler FP62 melting
point apparatus and are uncorrected. Unless otherwise noted, all
nonaqueous reactions were performed under an oxygen-free
atmosphere of nitrogen with rigid exclusion of moisture from re-
agents and glassware. Analytical thin layer chromatography (TLC)
was performed using EM Reagents 0.25 mm silica gel 60-F plates.
Visualization of the developed chromatogram was performed by
UV absorbance, aqueous potassium permanganate, or ethanolic
anisaldehyde. Liquid chromatography was performed using a force

S. Zhu et al. / Bioorg. Med. Chem. 17 (2009) 4496–4502
4499
TIb
Table 1
Antimalarial efficacy and toxicity evaluationa
Test compounds
In vitro W2-strain (IC50, ng/mL)
In vivo P. berghei (ED50, mg/kg)
MCD (mg/kg)
MTD (mg/kg)
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
0.56
0.12
0.14
0.31
0.34
0.29
0.26
0.40
0.23
0.28
2.9
1.6
8.4
3.1
2.9
6.6
5.9
7.1
6.8
8.9
4.7
7.9
39
144
345
310
338
439
399
425
379
335
440
429
504
102
88
17
111
107
51
74
56
63
43
71
56
11
12
132
128
10
10
3
0.44
0.49
1.2
1.3
1.0
1.1
1.7
0.74
1.5
8.7
3.4
10
42
0.14
0.19
0.62
0.57
0.76
224
0.14
0.12
1.7
1.5
2.3
0.77
0.69
5.6
4.9
12
55
51
35
240
Febrifugine
Chloroquine
4.5
40.0
6
a
Antimalarial activities were determined after i.p. administration of the compounds once daily for 4 days to infected mice.
Therapeutic indices were calculated as MTD divided by MCD.
b
flow (flash chromatography) of the indicated solvent system on EM
product. Silica gel flash chromatography (15% ethyl acetate in hex-
anes) furnishes 24 as viscous oil. Yield: 85% (two steps). 1H NMR:
7.31–7.27 (m, 10H), 5.34 (m, 2H), 4.61 (m, 2H), 4.09 (m, 1H),
3.85 (m, 1H), 3.57 (m, 2H), 3.38 (dd, J = 11 and 5.4 Hz, 1H), 3.24
(dd, J = 11 and 4.2 Hz, 1H), 1.94 (m, 2H); 13C NMR: 159.4, 140.9,
137.9, 128.7, 128.5, 127.7, 127.6, 127.4, 127.3, 80.2, 76.2, 70.8,
59.0, 39.1, 28.5, 13.5. HRESIMS m/z 474.0531 [M+Na]+ (calcd for
C20H22INNaO3þ, 474.0542) (100%). Anal. Calcd for C20H22INO3: C,
53.23; H, 4.91; I, 28.12; N, 3.10. Found: C, 53.29; H, 4.88; I,
28.21; N, 3.04.
Reagents Silica Gel 60 (70–230 mesh). Preparative TLC was per-
formed using Whatman Silica Gel C8 TLC plates (PLK5F). 1H NMR
spectra were recorded in deuteriochloroform, unless otherwise
noted, on a Bruker Avance 600 spectrometer at the frequency of
600.1 MHz. Chemical shifts are reported in parts per million on
the d scale from an internal standard of tetramethylsilane. Data
are reported as follows: chemical shifts, multiplicity (s = singlet,
d = doublet, t = triplet, q = quartet, qn = quintet, m = multiplet,
and br = broad), coupling constant in Hertz, integration, and
assignment. 13C NMR spectra were recorded in deuteriochloro-
form, unless otherwise noted, on a Bruker Avance 600 spectrome-
ter at the frequency of 150.9 MHz. Chemical shifts are reported in
parts per million on the d scale from an internal standard of tetra-
methylsilane. Combustion analyses were performed by Atlantic
Microlab, Inc. (Norcross, Georgia). All purified compounds possess
a purity of at least 95%. When necessary, solvents and reagents
were dried as follows: ether, tetrahydrofuran, benzene, and tolu-
ene were stored and distilled from sodium benzophenone ketyl;
dichloromethane, triethylamine, pyridine, and hexane were dis-
tilled over calcium hydride. Unless otherwise stated, the reagents
were purchased from Fisher Scientific, Aldrich Chemical Company,
Lancaster, or Fluka, and used as received.
4.1.2. 2-Allyl-3-benzyloxy-pyrrolidine-1-carboxylic acid benzyl
ester (25)
Vinyl magnesium bromide [1.0 M in tetrahydrofuran (THF),
53.0 mL] was added into a stirred solution of alkyl iodide 24
(11.0 g) and copper(I) iodide (CuI, 2.5 g) in 100 mL of THF at
ꢀ78 °C under nitrogen atmosphere. The reaction mixture was kept
at ꢀ78 °C for 1 h, then slowly warmed to room temperature over a
period of 2 h. The reaction mixture was quenched with aqueous so-
dium bicarbonate (60 mL). After partition between ethyl acetate
(120 mL) and water (120 mL), the separated organic layer was
washed with brine (55 mL), dried over anhydrous sodium sulfate,
and evaporated in a rotary evaporator under reduced pressure to
furnish the crude product. Silica gel flash chromatography (15%
ethyl acetate in hexanes) furnished 25 as viscous oil. Yield: 94%.
1H NMR: 7.31–7.27 (m, 10H), 5.73 (m, 1H), 5.34 (m, 2H), 5.07 (m,
1H), 4.99 (m, 1H), 4.61 (m, 2H), 4.09 (m, 1H), 3.85 (m, 1H), 3.57
(m, 2H), 2.38 (m, 2H), 1.94 (m, 2H); 13C NMR: 158.6, 142.1,
140.5, 138.2, 128.6, 128.4, 127.8, 127.7, 127.5, 127.2, 115.5, 79.2,
4.1.1. 3-Benzyloxy-2-iodomethyl-pyrrolidine-1-carboxylic acid
benzyl ester (24)
Mesyl chloride (MsCl, 8.0 mL) was added into a stirred solution
of alcohol 23 (25.3 g), triethylamine (Et3N, 30 mL), and 4-dimethyl-
aminopyridine (DMAP, 1.0 g) in 200 mL of methylene chloride
(CH2Cl2) in an ice-water bath. The reaction mixture was then
warmed to room temperature and stirred for additional 2 h. Sol-
vent was evaporated. The resulting slurry was dissolved in ethyl
acetate (180 mL), washed with water (150 mL), then aqueous so-
dium bicarbonate (80 mL), and brine (70 mL), dried over anhy-
drous sodium sulfate, and evaporated in a rotary evaporator
under reduced pressure to furnish the essentially pure mesylate.
This mesylate (29.5 g) and sodium iodide (NaI, 23.0 g) were dis-
solved in 180 mL of acetone. The resulting solution was then
heated at reflux for 12 h. Solvent was evaporated. The resulting
slurry was dissolved in ethyl acetate (150 mL), washed with water
(130 mL), then aqueous sodium bicarbonate (60 mL), and brine
(50 mL), dried over anhydrous sodium sulfate, and evaporated in
a rotary evaporator under reduced pressure to furnish the crude
75.8, 71.2, 57.6, 38.7, 31.4, 27.5. HRESIMS m/z 374.1739 [M+Na]+
þ
(calcd for C22H25NNaO3
, 374.1732) (100%). Anal. Calcd for
C22H25NO3: C, 75.19; H, 7.17; N, 3.99. Found: C, 75.24; H, 7.15;
N, 3.92.
4.1.3. 3-Benzyloxy-2-oxiranylmethyl-pyrrolidine-1-carboxylic
acid benzyl ester (26)
Alkene 25 (9.0 g), osmium tetraoxide (OsO4, 4% in water,
1.0 mL), and N-methyl morpholine N-oxide (NMO, 10.5 g) were
dissolved in 80 mL of water and 80 mL of tetrahydrofuran (THF).
The resulting heterogeneous solution is stirred vigorously for
4 days at room temperature. After partition between ethyl acetate
(150 mL) and water (100 mL), the separated organic layer was
washed with brine (70 mL), dried over anhydrous sodium sulfate,

4500
S. Zhu et al. / Bioorg. Med. Chem. 17 (2009) 4496–4502
and evaporated in a rotary evaporator under reduced pressure to
furnish the essentially pure diol. Tosyl chloride (TsCl, 4.6 g) was
added into a stirred solution of this diol (8.9 g), triethylamine
(Et3N, 7.0 mL), and 4-dimethylaminopyridine (DMAP, 300 mg) in
100 mL of methylene chloride (CH2Cl2) in an ice-water bath. The
reaction mixture was then warmed to room temperature and stir-
red for additional 2 h. Solvent was evaporated. The resulting slurry
was dissolved in ethyl acetate (100 mL), washed with water
(80 mL), then aqueous sodium bicarbonate (80 mL), and brine
(50 mL), dried over anhydrous sodium sulfate, and evaporated in
a rotary evaporator under reduced pressure to furnish the essen-
tially pure mono-tosylate. Potassium carbonate (K2CO3, 1.0 g)
was added into a stirred solution of this mono-tosylate (11.4 g)
in 25 mL of methanol (MeOH) at room temperature. After 4 h, reac-
tion mixture was partitioned between ethyl acetate (80 mL) and
water (80 mL), the separated organic layer was then washed with
brine (40 mL), dried over anhydrous sodium sulfate, and evapo-
rated in a rotary evaporator under reduced pressure to furnish
the crude product. Silica gel flash chromatography (25% ethyl ace-
tate in hexanes) furnishes 26 as viscous oil. Yield: 76% (three
steps). 1H NMR: 7.30–7.22 (m, 10H), 5.37 (m, 2H), 4.65 (m, 2H),
3.85 (m, 1H), 3.57 (m, 2H), 3.34 (m, 1H), 3.05 (m, 1H), 2.79 (m,
1H), 2.52 (m, 1H), 1.94 (m, 2H), 1.87–1.82 (m, 2H); 13C NMR:
158.8, 141.7, 139.3, 128.5, 128.2, 127.9, 127.6, 127.4, 126.8, 76.2,
73.8, 71.9, 51.4, 47.5, 43.3, 35.8, 34.8, 28.4. HRESIMS m/z
390.1677 [M+Na]+ (calcd for C22H25NNaO4þ, 390.1681) (100%).
Anal. Calcd for C22H25NO4: C, 71.91; H, 6.86; N, 3.81. Found: C,
71.84; H, 6.91; N, 3.85.
in a Parr apparatus for 12 h. Solid was filtered off and the solution
was evaporated under vacuum to dryness. Recrystallization from
ethanol–water (with addition of dilute aqueous HCl solution, 4–
5 equiv of HCl) furnishes compound 7 (HCl salt) as off-white crys-
tals. Yield: 901 mg, 76% (two steps). Mp: 217–218 °C. ½a D25
ꢃ
+24.9 (c
0.51, EtOH). 1H NMR (CD3OD): 8.27 (dd, J = 7.9, 1.0 Hz, 1H), 7.92 (s,
1H), 7.77 (ddd, J = 8.1, 7.9, 1.0 Hz, 1H), 7.71 (dd, J = 8.1, 0.9 Hz, 1H),
7.50 (dt, J = 7.9, 0.9 Hz, 1H), 5.11 (d, J = 17.1 Hz, 1H), 4.96 (d,
J = 17.1 Hz, 1H), 4.41 (m, 1H), 4.14 (m, 1H), 3.49–3.42 (m, 2H),
2.89 (dd, J = 15.9, 4.2 Hz, 1H), 2.81 (dd, J = 15.9, 6.1 Hz, 1H), 2.24
(m, 1H), 1.90 (m, 1H); 13C NMR (CD3OD): 204.1, 169.2, 165.2,
147.9, 133.2, 128.1, 127.4, 124.3, 121.9, 75.1, 57.9, 56.4, 39.2,
33.4, 31.7. HRESIMS m/z 310.1159 [M+Na]+ (calcd for
C15H17N3NaO3þ, 310.1168) (100%). Anal. Calcd for C15H17N3O3ꢂHCl:
C, 55.64; H, 5.60; Cl, 10.95; N, 12.98. Found: C, 55.60; H, 5.63; Cl,
10.99; N, 12.89.
4.1.6. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
3H-pyrido[3,2-d]pyrimidin-4-one (8)
Yellow crystal. Mp: 261 °C. ½a D25
ꢃ
+21.8 (c 0.5, EtOH). 1H NMR
(CD3OD): 9.71 (d, J = 7.2 Hz, 1H), 9.24 (d, J = 7.2 Hz, 1H), 8.80 (s,
1H), 8.21 (t, J = 7.2 Hz, 1H), 5.08 (d, J = 17.3 Hz, 1H), 4.92 (d,
J = 17.3 Hz, 1H), 4.41 (m, 1H), 4.14 (m, 1H), 3.49–3.42 (m, 2H),
2.89 (m, 1H), 2.81 (dd, J = 15.1 and 6.1 Hz, 1H), 2.24 (m, 1H), 1.90
(m, 1H); 13C NMR (CD3OD): 202.1, 163.7, 159.2, 151.5, 149.6,
146.4, 132.9, 131.2, 81.0, 57.9, 54.1, 51.9, 44.7, 36.7. HRESIMS m/
z 311.1116 [M+Na]+ (calcd for C14H16N4NaO3þ, 311.1120) (100%).
Anal. Calcd for C14H16N4O3ꢂHCl: C, 51.78; H, 5.28; Cl, 10.92; N,
17.25. Found: C, 51.82; H, 5.26; Cl, 10.88; N, 17.28.
4.1.4. 3-Benzyloxy-2-[2-hydroxy-3-(4-oxo-4H-quinazolin-3-yl)-
propyl]-pyrrolidine-1-carboxylic acid benzyl ester (27)
4.1.7. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
3H-pyrido[4,3-d]pyrimidin-4-one (9)
Potassium hydride (KH, 30% in mineral oil, 3.6 g) was suspended
in 50 mL of dimethylformamide (DMF). It was cooled in an ice-
water bath, and solid 3H-quinazolin-4-one (4.0 g) was added in.
After 30 min, a solution of oxirane 26 (2.2 g) in 10 mL of DMF was
added in. The reaction mixture was then heated at 80 °C for 12 h
under nitrogen atmosphere. It was partitioned between ethyl ace-
tate (80 mL) and water (80 mL), separated organic layer was
washed with water (3 ꢁ 60 mL), then brine (40 mL), dried over
anhydrous sodium sulfate, and evaporated in a rotary evaporator
under reduced pressure to furnish the crude product. Silica gel flash
chromatography (80% ethyl acetate in hexanes) furnished 27 as off
white solid. Yield: 77%. Mp: 187 °C. 1H NMR: 8.28 (d, J = 7.5 Hz, 1H),
8.24 (s, 1H), 7.73 (m, 2H), 7.48 (m, 1H), 7.31–7.27 (m, 10H), 5.45 (m,
2H), 5.19 (br, 1H), 4.81 (m, 2H), 4.24 (m, 1H), 4.16 (m, 1H), 3.85 (m,
2H), 3.62 (m, 2H), 3.23 (m, 1H), 1.89 (m, 2H), 1.57 (m, 2H); 13C NMR:
168.4, 163.1, 158.7, 145.3, 141.2, 138.2, 134.1, 129.3, 128.9, 128.6,
127.8, 127.6, 127.4, 127.1, 127.0, 126.7, 123.8, 80.8, 77.4, 71.3,
Pale-yellow crystal. Mp: 251 °C. ½a D25
ꢃ
ꢀ31.4 (c 0.5, EtOH). 1H
NMR (CD3OD): 9.54 (s, 1H), 9.19 (d, J = 7.5 Hz, 1H), 8.71 (s, 1H),
8.13 (d, J = 7.5 Hz, 1H), 5.07 (d, J = 17.4 Hz, 1H), 4.90 (d,
J = 17.4 Hz, 1H), 4.39 (m, 1H), 4.18 (m, 1H), 3.46–3.41 (m, 2H),
2.94 (m, 1H), 2.85 (dd, J = 15.1 and 6.1 Hz, 1H), 2.23 (m, 1H), 1.87
(m, 1H); 13C NMR (CD3OD): 202.6, 166.6, 163.6, 159.1, 154.5,
149.0, 147.1, 135.2, 80.3, 56.9, 55.3, 50.9, 44.3, 35.6. HRESIMS m/
z 311.1128 [M+Na]+ (calcd for C14H16N4NaO3þ, 311.1120) (100%).
Anal. Calcd for C14H16N4O3ꢂHCl: C, 51.78; H, 5.28; Cl, 10.92; N,
17.25. Found: C, 51.74; H, 5.31; Cl, 10.90; N, 17.22.
4.1.8. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
3H-pyrido[3,4-d]pyrimidin-4-one (10)
Yellow crystal. Mp: 221–222 °C. ½a D25
ꢃ
+27.4 (c 0.43, EtOH). 1H
NMR (CD3OD): 9.24 (s, 1H), 8.74 (d, J = 7.3 Hz, 1H), 8.49 (d,
J = 7.3 Hz, 1H), 8.25 (s, 1H), 5.16 (d, J = 17.1 Hz, 1H), 5.03 (d,
J = 17.1 Hz, 1H), 4.37 (m, 1H), 4.23 (m, 1H), 3.44–3.38 (m, 2H),
2.91 (m, 1H), 2.89 (dd, J = 15.4 and 6.2 Hz, 1H), 2.27 (m, 1H), 1.92
(m, 1H); 13C NMR (CD3OD): 202.6, 167.8, 164.6, 159.2, 155.2,
154.7, 139.9, 134.8, 79.2, 56.1, 54.9, 51.2, 45.1, 34.8. HRESIMS m/
z 311.1114 [M+Na]+ (calcd for C14H16N4NaO3þ, 311.1120) (100%).
Anal. Calcd for C14H16N4O3ꢂHCl: C, 51.78; H, 5.28; Cl, 10.92; N,
17.25. Found: C, 51.84; H, 5.24; Cl, 10.99; N, 17.31.
66.4, 55.1, 51.9, 39.4, 32.4, 26.6. HRESIMS m/z 536.2170 [M+Na]+
þ
(calcd for C30H31N3NaO5
, 536.2161) (100%). Anal. Calcd for
C30H31N3O5ꢂHCl: C, 65.51; H, 5.86; Cl, 6.45; N, 7.64. Found: C,
65.56; H, 5.83; Cl, 6.51; N, 7.62.
4.1.5. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
3H-quinazolin-4-one (7)
A solution of alcohol 27 (2.1 g) in 5 mL of CH2Cl2 was added into
a stirred slurry of tetrapropylammonium perruthenate (TPAP,
70 mg), N-methyl morpholine N-oxide (NMO, 936 mg), and
grounded molecular sieve (800 mg) in 10 mL of CH2Cl2 at room
temperature. After 1 h, the reaction mixture was loaded directly
into a short column of silica gel and eluted with 5% MeOH/EtOAc.
Concentration of the eluant under reduced pressure afforded the
corresponding ketone. This ketone (1.89 g) was dissolved in
15 mL of 95% EtOH/H2O. Three hundred milligrams of 10% Pd on
carbon was added in. It was then treated with hydrogen (60 psi)
4.1.9. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
3H-pyrido[2,3-d]pyrimidin-4-one (11)
Yellow crystal. Mp: 213–214 °C. ½a D25
ꢃ
ꢀ19.6 (c 0.42, EtOH). 1H
NMR (CD3OD): 8.88 (d, J = 6.8 Hz, 1H), 8.75 (d, J = 6.8 Hz, 1H),
8.28 (s, 1H), 8.19 (t, J = 6.8 Hz, 1H), 5.03 (d, J = 16.8 Hz, 1H), 4.92
(d, J = 16.8 Hz, 1H), 4.37 (m, 1H), 4.24 (m, 1H), 3.45–3.40 (m, 2H),
2.82 (dd, J = 15.5, 4.6 Hz, 1H), 2.76 (dd, J = 15.5, 6.2 Hz, 1H), 2.28
(m, 1H), 1.84 (m, 1H); 13C NMR (CD3OD): 204.1, 171.8, 167.3,
162.8, 152.1, 142.6, 133.1, 130.9, 77.4, 58.5, 56.8, 38.7, 33.9, 32.1.
HRESIMS m/z 311.1122 [M+Na]+ (calcd for C14H16N4NaO3
,
þ

S. Zhu et al. / Bioorg. Med. Chem. 17 (2009) 4496–4502
4501
311.1120) (100%). Anal. Calcd for C14H16N4O3ꢂHCl: C, 51.78; H,
5.28; Cl, 10.92; N, 17.25. Found: C, 51.83; H, 5.25; Cl, 10.81; N,
17.34.
1H), 4.39 (m, 1H), 4.24 (m, 1H), 3.47–3.41 (m, 2H), 2.85 (dd,
J = 15.7, 4.4 Hz, 1H), 2.77 (dd, J = 15.7, 5.9 Hz, 1H), 2.20 (m, 1H),
1.92 (m, 1H); 13C NMR (CD3OD): 203.3, 167.8, 162.6, 147.3, 137.1,
133.5, 129.8, 128.1, 127.4, 119.2, 76.9, 58.2, 57.1, 38.8, 33.1, 30.9.
HRESIMS m/z 378.1035 [M+Na]+ (calcd for C16H16F3N3NaO3
,
þ
4.1.10. (2R,3S)-5-Fluoro-3-[3-(3-hydroxy-pyrrolidin-2-yl)-2-
oxo-propyl]-3H-quinazolin-4-one (12)
378.1041) (100%). Anal. Calcd for C16H16F3N3O3ꢂHCl: C, 49.05; H,
4.37; Cl, 9.05; F, 14.55; N, 10.73. Found: C, 48.98; H, 4.41; Cl, 9.11;
F, 14.49; N, 10.67.
Pale-yellow crystals. Mp: 242 °C. ½a D25
ꢃ
ꢀ33.8 (c 0.5, EtOH). 1H
NMR (CD3OD): 8.32 (s, 1H), 8.20 (d, J = 7.1 Hz, 1H), 7.73 (d,
J = 7.2 Hz, 1H), 7.55 (t, J = 7.1 Hz, 1H), 5.10 (d, J = 17.4 Hz, 1H),
4.93 (d, J = 17.4 Hz, 1H), 4.43 (m, 1H), 4.10 (m, 1H), 3.48–3.40 (m,
2H), 2.82 (m, 1H), 2.77 (dd, J = 15.2 and 5.8 Hz, 1H), 2.28 (m, 1H),
1.93 (m, 1H); 13C NMR (CD3OD): 203.1, 164.1, 158.5, 152.7,
4.1.15. (2R,3S)-5-tert-Butyl-3-[3-(3-hydroxy-pyrrolidin-2-yl)-2-
oxo-propyl]-3H-quinazolin-4-one (17)
Off-white crystal. Mp: 193–194 °C. ½a D25
ꢃ
+12.4 (c 0.51, EtOH). 1H
NMR (CD3OD): 8.25 (d, J = 7.2 Hz, 1H), 8.17 (s, 1H), 7.74 (d,
J = 7.2 Hz, 1H), 7.63 (t, J = 7.2 Hz, 1H), 5.12 (d, J = 16.9 Hz, 1H),
5.00 (d, J = 16.9 Hz, 1H), 4.34 (m, 1H), 4.12 (m, 1H), 3.49–3.42 (m,
2H), 2.91 (dd, J = 15.5, 4.5 Hz, 1H), 2.82 (dd, J = 15.5, 5.8 Hz, 1H),
2.22 (m, 1H), 1.89 (m, 1H), 1.31 (s, 9H); 13C NMR (CD3OD): 202.3,
169.5, 165.1, 152.9, 147.2, 143.1, 133.9, 130.1, 129.9, 77.3, 58.6,
149.3, 147.5, 132.8, 132.2, 129.6, 80.2, 58.6, 53.2, 50.9, 43.8, 35.9.
HRESIMS m/z 328.1082 [M+Na]+ (calcd for C15H16FN3NaO3
,
þ
328.1073) (100%). Anal. Calcd for C15H16FN3O3ꢂHCl: C, 52.71; H,
5.01; Cl, 10.37; F, 5.56; N, 12.30. Found: C, 52.80; H, 5.00; Cl,
10.26; F, 5.51; N, 12.34.
56.4, 39.4, 35.6, 33.6, 31.9, 30.5. HRESIMS m/z 366.1782 [M+Na]+
, 366.1794) (100%). Anal. Calcd for
C19H25N3O3ꢂHCl: C, 60.07; H, 6.90; Cl, 9.33; N, 11.06. Found: C,
4.1.11. (2R,3S)-6-Fluoro-3-[3-(3-hydroxy-pyrrolidin-2-yl)-2-
oxo-propyl]-3H-quinazolin-4-one (13)
þ
(calcd for C19H25N3NaO3
Pale-yellow crystal. Mp: 196–197 °C. ½a D25
ꢃ
+26.3 (c 0.5, EtOH).
60.13; H, 6.87; Cl, 9.29; N, 11.12.
1H NMR (CD3OD): 8.24 (s, 1H), 7.84 (d, J = 7.1 Hz, 1H), 7.53 (s,
1H), 7.46 (d, J = 7.1 Hz, 1H), 5.08 (d, J = 17.3 Hz, 1H), 4.95 (d,
J = 17.3 Hz, 1H), 4.48 (m, 1H), 4.15 (m, 1H), 3.47–3.39 (m, 2H),
2.86 (m, 1H), 2.79 (dd, J = 15.1 and 5.6 Hz, 1H), 2.32 (m, 1H), 1.89
(m, 1H); 13C NMR (CD3OD): 202.9, 167.9, 163.6, 147.1, 133.2,
4.1.16. (2R,3S)-6-tert-Butyl-3-[3-(3-hydroxy-pyrrolidin-2-yl)-2-
oxo-propyl]-3H-quinazolin-4-one (18)
Off-white crystal. Mp: 190–191 °C. ½a D25
ꢃ
ꢀ10.3 (c 0.45, EtOH). 1H
NMR (CD3OD): 8.20 (s, 1H), 7.94 (d, J = 7.1 Hz, 1H), 7.69 (d,
J = 7.1 Hz, 1H), 7.56 (s, 1H), 5.09 (d, J = 17.1 Hz, 1H), 4.97 (d,
J = 17.1 Hz, 1H), 4.30 (m, 1H), 4.18 (m, 1H), 3.48–3.40 (m, 2H),
2.93 (dd, J = 15.9, 4.6 Hz, 1H), 2.84 (dd, J = 15.9, 5.9 Hz, 1H), 2.29
(m, 1H), 1.92 (m, 1H), 1.34 (s, 9H); 13C NMR (CD3OD): 202.9,
167.9, 164.6, 154.1, 143.2, 134.7, 129.9, 128.8, 128.4, 79.2, 59.1,
129.7, 128.9, 128.0, 127.7, 78.4, 60.4, 54.0, 51.1, 42.5, 34.5. HRE-
SIMS m/z 328.1066 [M+Na]+ (calcd for C15H16FN3NaO3
,
þ
328.1073) (100%). Anal. Calcd for C15H16FN3O3ꢂHCl: C, 52.71; H,
5.01; Cl, 10.37; F, 5.56; N, 12.30. Found: C, 52.64; H, 5.05; Cl,
10.42; F, 5.52; N, 12.24.
56.9, 38.8, 36.2, 34.2, 31.7, 30.2. HRESIMS m/z 366.1803 [M+Na]+
, 366.1794) (100%). Anal. Calcd for
C19H25N3O3ꢂHCl: C, 60.07; H, 6.90; Cl, 9.33; N, 11.06. Found: C,
4.1.12. (2R,3S)-5,6-Difluoro-3-[3-(3-hydroxy-pyrrolidin-2-yl)-2-
oxo-propyl]-3H-quinazolin-4-one (14)
þ
(calcd for C19H25N3NaO3
Pale-yellow crystal. Mp: 241 °C. ½a D25
ꢃ
ꢀ11.8 (c 0.5, EtOH). 1H
60.01; H, 6.93; Cl, 9.37; N, 10.98.
NMR (CD3OD): 8.31 (s, 1H), 7.80 (d, J = 6.9 Hz, 1H), 7.74 (d,
J = 6.9 Hz, 1H), 5.06 (d, J = 17.4 Hz, 1H), 4.92 (d, J = 17.4 Hz, 1H),
4.39 (m, 1H), 4.12 (m, 1H), 3.48–3.40 (m, 2H), 2.92 (m, 1H), 2.78
(dd, J = 14.9 and 5.8 Hz, 1H), 2.28 (m, 1H), 1.87 (m, 1H); 13C NMR
(CD3OD): 202.8, 162.8, 161.6, 158.5, 152.1, 149.4, 147.2, 132.4,
130.5, 80.7, 58.5, 53.8, 51.4, 43.6, 37.2. HRESIMS m/z 346.0984
[M+Na]+ (calcd for C15H15F2N3NaO3þ, 346.0979) (100%). Anal. Calcd
for C15H15F2N3O3ꢂHCl: C, 50.08; H, 4.48; Cl, 9.85; F, 10.56; N, 11.68.
Found: C, 50.14; H, 4.44; Cl, 9.91; F, 10.49; N, 11.75.
4.1.17. (2R,3S)-3-[3-(3-Benzyloxy-pyrrolidin-2-yl)-2-oxo-
propyl]-3H-pyrido[3,2-d]pyrimidin-4-one (19)
Yellow crystal. Mp: 232–233 °C. ½a D25
ꢃ
+25.7 (c 0.5, EtOH). 1H NMR
(CD3OD): 9.61 (d, J = 7.2 Hz, 1H), 9.19 (d, J = 7.2 Hz, 1H), 8.76 (s, 1H),
8.31 (t, J = 7.2 Hz, 1H), 7.22–7.18 (m, 5H), 5.08 (d, J = 17.2 Hz, 1H),
4.89 (d, J = 17.2 Hz, 1H), 4.69 (m, 2H), 4.40 (m, 1H), 4.12 (m, 1H),
3.48–3.42 (m, 2H), 2.88 (m, 1H), 2.78 (dd, J = 15.1 and 6.0 Hz, 1H),
2.26 (m, 1H), 1.92 (m, 1H); 13C NMR (CD3OD): 204.3, 163.2, 159.9,
151.1, 149.3, 146.1, 137.9, 133.8, 131.6, 128.9, 127.8, 127.5, 81.1,
75.1, 573, 54.8, 51.3, 44.2, 36.2. HRESIMS m/z 401.1586 [M+Na]+
4.1.13. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
5-trifluoromethyl-3H-quinazolin-4-one (15)
þ
(calcd for C21H22N4NaO3
, 401.1590) (100%). Anal. Calcd for
Off-white crystals. Mp: 262 °C. ½a D25
ꢃ
+27.5 (c 0.5, EtOH). 1H NMR
C21H22N4O3ꢂHCl: C, 60.79; H, 5.59; Cl, 8.55; N, 13.50. Found: C,
(CD3OD): 8.28 (s, 1H), 7.87 (d, J = 7.2 Hz, 1H), 7.59 (d, J = 7.2 Hz,
1H), 7.50 (t, J = 7.2 Hz, 1H), 5.05 (d, J = 16.8 Hz, 1H), 4.84 (d,
J = 16.8 Hz, 1H), 4.40 (m, 1H), 4.19 (m, 1H), 3.47–3.41 (m, 2H),
2.85 (m, 1H), 2.78 (dd, J = 15.3 and 6.2 Hz, 1H), 2.27 (m, 1H), 1.93
(m, 1H); 13C NMR (CD3OD): 202.8, 162.7, 158.2, 150.5, 148.3,
60.83; H, 5.55; Cl, 8.62; N, 13.44.
4.1.18. (2R,3S)-3-[3-(3-Benzyloxy-pyrrolidin-2-yl)-2-oxo-
propyl]-5-trifluoromethyl-3H-quinazolin-4-one (20)
Pale yellow crystals. Mp: 229–230 °C. ½a D25
ꢀ17.9 (c 0.5, EtOH).
ꢃ
1H NMR (CD3OD): 8.22 (s, 1H), 7.84 (d, J = 7.1 Hz, 1H), 7.62 (d,
J = 7.1 Hz, 1H), 7.55 (t, J = 7.1 Hz, 1H), 7.23–7.19 (m, 5H), 5.09 (d,
J = 16.9 Hz, 1H), 4.82 (d, J = 16.9 Hz, 1H), 4.71 (m, 2H), 4.39 (m,
1H), 4.13 (m, 1H), 3.46–3.40 (m, 2H), 2.89 (m, 1H), 2.79 (dd,
J = 15.4 and 6.0 Hz, 1H), 2.29 (m, 1H), 1.91 (m, 1H); 13C NMR
(CD3OD): 203.7, 162.1, 158.9, 151.2, 149.1, 140.7, 139.2, 133.5,
132.4, 131.9, 128.9, 127.8, 127.7, 116.8, 80.1, 76.2, 57.9, 52.9,
141.4, 133.3, 132.7, 131.7, 116.8, 80.5, 57.6, 53.6, 50.5, 42.4, 36.5.
HRESIMS m/z 378.1050 [M+Na]+ (calcd for C16H16F3N3NaO3
,
þ
378.1041) (100%). Anal. Calcd for C16H16F3N3O3ꢂHCl: C, 49.05; H,
4.37; Cl, 9.05; F, 14.55; N, 10.73. Found: C, 49.12; H, 4.33; Cl,
9.01; F, 14.62; N, 10.82.
4.1.14. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
6-trifluoromethyl-3H-quinazolin-4-one (16)
51.5, 42.1, 33.8. HRESIMS m/z 468.1519 [M+Na]+ (calcd for
Off-white crystal. Mp: 206–207 °C. ½a D25
ꢃ
+16.7 (c 0.5, EtOH). 1H
NMR (CD3OD): 8.26 (s, 1H), 7.81 (d, J = 7.2 Hz, 1H), 7.75 (s, 1H),
7.58 (d, J = 7.2 Hz, 1H), 5.09 (d, J = 17.4 Hz, 1H), 4.98 (d, J = 17.4 Hz,
þ
C23H22F3N3NaO3
,
468.1511)
(100%).
Anal.
Calcd
for
C23H22F3N3O3ꢂHCl: C, 57.32; H, 4.81; Cl, 7.36; F, 11.83; N, 8.72.
Found: C, 57.39; H, 4.76; Cl, 7.42; F, 11.74; N, 8.80.

4502
S. Zhu et al. / Bioorg. Med. Chem. 17 (2009) 4496–4502
4.1.19. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
7-methoxy-3H-quinazolin-4-one (21)
the donor mice was then diluted with normal mouse serum to a
concentration of 1 ꢁ 106 P. berghei-infected erythrocytes per inoc-
ulum (0.1 mL). The groups of experimental and control mice were
inoculated with this parasitized blood on day 0. The tested mice
were treated with either candidate antimalarial drugs or with vehi-
cle alone to serve as negative controls from day 3 to day 10 orally
(PO) or subcutaneously (SC) once a day for eight days. Each exper-
imental group received a different dose level, with up to five differ-
ent dose groups per compound. Blood films and body weights were
taken on the third and sixth days post-infection, then at weekly
intervals through day 60. Films were Giemsa stained and examined
microscopically to determine parasitemia. All mice were observed
twice a day to assess their clinical signs that were recorded. All
treated mice with negative smear on day 60 were considered
cured.
Off-white crystal. Mp: 181–182 °C. ½a D25
ꢃ
+18.3 (c 0.5, EtOH). 1H
NMR (CD3OD): 8.13 (s, 1H), 7.72 (s, 1H), 7.43 (d, J = 7.2 Hz, 1H),
7.33 (d, J = 7.2 Hz, 1H), 5.08 (d, J = 17.3 Hz, 1H), 4.92 (d,
J = 17.3 Hz, 1H), 4.38 (m, 1H), 4.11 (m, 1H), 3.81 (s, 3H), 3.43–
3.36 (m, 2H), 2.91 (dd, J = 15.4, 4.3 Hz, 1H), 2.79 (dd, J = 15.4,
6.2 Hz, 1H), 2.29 (m, 1H), 1.81 (m, 1H); 13C NMR (CD3OD): 202.5,
168.3, 165.5, 161.3, 188.5, 138.4, 134.5, 130.6, 128.9, 76.4, 58.7,
57.2, 55.9, 38.8, 34.1, 31.4. HRESIMS m/z 340.1285 [M+Na]+ (calcd
þ
for C16H19N3NaO4
,
340.1273) (100%). Anal. Calcd for
C16H19N3O4ꢂHCl: C, 54.32; H, 5.70; Cl, 10.02; N, 11.88. Found: C,
54.41; H, 5.67; Cl, 10.11; N, 11.79.
4.1.20. (2R,3S)-3-[3-(3-Hydroxy-pyrrolidin-2-yl)-2-oxo-propyl]-
8-methoxy-3H-quinazolin-4-one (22)
Off-white crystal. Mp: 184–185 °C. ½a D25
ꢃ
ꢀ15.6 (c 0.5, EtOH). 1H
Acknowledgments
NMR (CD3OD): 8.18 (s, 1H), 7.55 (t, J = 7.3 Hz, 1H), 7.36 (d,
J = 7.3 Hz, 1H), 7.19 (d, J = 7.3 Hz, 1H), 5.09 (d, J = 17.5 Hz, 1H),
5.01 (d, J = 17.5 Hz, 1H), 4.43 (m, 1H), 4.08 (m, 1H), 3.77 (s, 3H),
3.51–3.44 (m, 2H), 2.81 (dd, J = 16.1, 4.3 Hz, 1H), 2.75 (dd,
J = 16.1, 6.0 Hz, 1H), 2.27 (m, 1H), 1.92 (m, 1H); 13C NMR (CD3OD):
203.1, 166.2, 162.3, 160.5, 147.3, 137.4, 132.9, 129.9, 128.7, 80.1,
This research was partially supported by a grant from the Na-
tional Institutes of Health (R43 AI077109 to S.Z.). The content is so-
lely the responsibility of the authors and does not necessarily
represent the official views of the funding agency.
58.9, 57.3, 56.9, 39.7, 34.8, 30.8. HRESIMS m/z 340.1266 [M+Na]+
References and notes
þ
(calcd for C16H19N3NaO4
, 340.1273) (100%). Anal. Calcd for
C16H19N3O4ꢂHCl: C, 54.32; H, 5.70; Cl, 10.02; N, 11.88. Found: C,
1. WHO, The World Health Report, 2004.
2. Winstanley, P. A. Parasitol. Today 2000, 16, 146.
54.25; H, 5.74; Cl, 9.98; N, 11.96.
3. Oduola, A. M.; Milhous, W. K.; Salako, L. A.; Walker, O.; Desjardins, R. E. Lancet
1987, 2, 1304.
4. Koepfly, J. B.; Mead, J. F.; Brockman, J. A., Jr. J. Am. Chem. Soc. 1947, 69, 1837.
5. Koepfly, J. B.; Mead, J. F.; Brockman, J. A., Jr. J. Am. Chem. Soc. 1949, 71, 1048–
1054.
6. WHO Report, Meeting on Antimalarial Drug Development, Shanghai, China, 16–
17 November 2001.
7. Ablondi, F.; Gordon, S.; Morton, J., II; Williams, J. H. J. Org. Chem. 1952, 17, 14.
8. Hewitt, R. I.; Wallace, W. S.; Gill, E. R.; Williams, J. H. Am. J. Trop. Med. Hyg. 1952,
1, 768.
4.2. Parasite culture
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Febrifugine and analogs were tested in a cell-based in vitro drug
susceptibility assay to determine if they were capable of interrupt-
ing Plasmodium metabolism and growth. The semi-automated mi-
cro-dilution technique was used to assess the sensitivity of the
parasites to the selected compounds. The incorporation of
[3H]hypoxanthine into the parasites was measured as a function
of compound concentration to determine IC50 values.
4.4. In vivo efficacy test25
In conducting the research described in this report, the investi-
gators adhered to the Guide for the Care and Use of Laboratory Ani-
mals by the Institute of Laboratory Animal Resources, National
Research Council. Typically, 4–5 weeks old IRC mice were housed
in plastic cages with free access to water and food. Plasmodium
berghei parasite-infected erythrocytes were obtained from donor
mice. On experiment day 0, the donor mice were anesthetized
and exsanguinated via cardiac puncture. The pooled blood from