Synthesis and characterization of in vitro and in vivo profiles of hydroxybupropion analogues: Aids to smoking cessation

Article abstract of DOI:10.1021/jm1003232

To create potentially superior aids to smoking cessation and/or antidepressants and to elucidate bupropions possible mechanisms of action(s), 23 analogues based on its active hydroxymetabolite (2S,3S)-4a were synthesized and tested for their abilities to inhibit monoamine uptake and nAChR subtype activities in vitro and acute effects of nicotine in vivo. The 3′,4′-dichlorophenyl [(±)-4n], naphthyl (4r), and 3-chlorophenyl or 3-propyl analogues 4s and 4t, respectively, had higher inhibitory potency and/or absolute selectivity than (2S,3S)-4a for inhibition of DA, NE, or 5HT uptake. The 3′-fluorophenyl, 3′-bromophenyl, and 4-biphenyl analogues 4c, 4d, and 4l, respectively, had higher potency for antagonism of α4β2-nAChR than (2S,3S)-4a. Several analogues also had higher potency than (2S,3S)-4a as antagonists of nicotine-mediated antinociception in the tail-flick assay. The results suggest that compounds acting via some combination of DA, NE, or 5HT inhibition and/or antagonism of α4β2-nAChR can potentially be new pharmacotherapeutics for treatment of nicotine dependence.

Full text of DOI:10.1021/jm1003232

J. Med. Chem. 2010, 53, 4731–4748 4731
DOI: 10.1021/jm1003232
Synthesis and Characterization of in Vitro and in Vivo Profiles of Hydroxybupropion
Analogues: Aids to Smoking Cessation
†
‡
§
‡
‡
‡
ꢀ
Ronald J. Lukas, Ana Z. Muresan, M. Imad Damaj, Bruce E. Blough, Xiaodong Huang, Hernan A. Navarro,
S. Wayne Mascarella,‡ J. Brek Eaton,† Syndia K. Marxer-Miller,† and F. Ivy Carroll*,‡
†Division of Neurobiology, Barrow Neurological Institute, 350 West Thomas Road, Phoenix, Arizona 85013, ‡Center for Organic and Medicinal
Chemistry, Research Triangle Institute, P.O. Box 12194, Research Triangle Park, North Carolina 27709, and Department of Pharmacology and
Toxicology, Virginia Commonwealth University, Richmond, Virginia 23298
§
Received March 10, 2010
To create potentially superior aids to smoking cessation and/or antidepressants and to elucidate
bupropion’s possible mechanisms of action(s), 23 analogues based on its active hydroxymetabolite
(2S,3S)-4a were synthesized and tested for their abilities to inhibit monoamine uptake and nAChR
subtype activities in vitro and acute effects of nicotine in vivo. The 30,40-dichlorophenyl [(()-4n],
naphthyl (4r), and 3-chlorophenyl or 3-propyl analogues 4s and 4t, respectively, had higher inhibitory
potency and/or absolute selectivity than (2S,3S)-4a for inhibition of DA, NE, or 5HT uptake. The
30-fluorophenyl, 30-bromophenyl, and 4-biphenyl analogues 4c, 4d, and 4l, respectively, had higher
potency for antagonism of R4β2-nAChR than (2S,3S)-4a. Several analogues also had higher potency
than (2S,3S)-4a as antagonists of nicotine-mediated antinociception in the tail-flick assay. The results
suggest that compounds acting via some combination of DA, NE, or 5HT inhibition and/or antagonism
of R4β2-nAChR can potentially be new pharmacotherapeutics for treatment of nicotine dependence.
Introduction
of different combinations of genetically distinct subunits, (R1,
R2-R10, β1-β4, γ, δ, ε).5,6 The predominant nAChR sub-
types found in the brain are thought to be heteromeric R4β2-
nAChR or homomeric R7-nAChR.7 However, appreciable
amounts of R3β4*- and R6β2*-nAChRs (where the * indi-
cates that other subunits are known or possible assembly
partners with those specified) also are in brain regions impli-
cated in reward and drug dependence.8-13
Tobacco use is the leading preventable cause of disease,
disability, and death in the United States (U.S.). According to
the Centers for Disease Control (CDCa) 2008 Smoking and
Tobacco Use;Fact Sheet,1 cigarette smoking results in more
than 400000premature deaths inthe U.S. eachyear, about1 in
every 5 deaths. On average, adults who smoke die 14 years
earlier than nonsmokers.2 Cigarette smoking accounts for
about one-third of all cancers, including 90% of lung cancer
cases. Smoking also causes lung diseases such as chronic
bronchitis and emphysema and increases the risk of stroke,
heart attack, vascular disease, and aneurysm.2 In spite of these
documented connections between tobacco use and disease, an
unacceptable number of people continue to use tobacco
products. In 2008, 28.6% of the US population 12 years of
age and older (70.9 million people) used a tobacco product at
least once in the prior month to being interviewed. This figure
includes 3.1 million young people aged 12-17 (12.4% of this
age group).3
Nicotine exposure can stimulate activity of somatodendri-
tic nAChRs to alter neuronal electrical activity and neuro-
transmitter release as a consequence of neuronal activation.
However, by actingatnAChRspositioned on nerveterminals,
nicotine also can increase neurotransmitter release as a con-
sequence of local depolarization of the nerve terminal mem-
brane potential and/or calcium ion mobilization in terminals.
The integration of these effects are likely to contribute to
nicotine’s actions, including those that are presumably in-
volved in its reinforcement of tobacco product use such as
effects in monoaminergic reward pathways.14,15
Nicotine (1) is considered to be the main psychoactive
component in tobacco smoke that causes and maintains
tobacco use.4 Nicotine’s pharmacological and behavioral
effects result from the activation of different nicotinic acetyl-
choline receptor (nAChR) subtypes. The subtypes are
either homo- or heteropentameric ion channels, consisting
*To whom correspondence should be addressed. Phone: 919-541-
6679. Fax: 919-541-8868. E-mail: fic@rti.org.
aAbbreviations: CDC, Centers for Disease Control; DA, dopamine;
5HT, serotonin; NE, norepinephrine; SR, sustained release; HEK,
human embryonic kidney; DAT, dopamine transporter; SERT, seroto-
nin transporter; NET, norepinephrine transporter; nAChR, nicotine
acetylcholine receptor; VTA, ventral tegmental area; NRT, nicotine
replacement therapy; MPE, maximum possible effect.
Even though nicotine dependence has a huge impact on
global health, pharmacotherapies for treating tobacco use are
limited. They include nicotine-replacement therapies (NRTs),
r
2010 American Chemical Society
Published on Web 05/28/2010
pubs.acs.org/jmc

4732 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
Scheme 1a
a Reagents: (a) RCH2MgBr; (b) TBSOTf, CH2Cl2, Et3N; (c) AD-mix-β, t-BuOH/H2O; (d) Tf2O, proton sponge, 2-amino-2-methyl-1-propanol, CH3CN.
bupropion (2), and varenicline (3). Because only about one-
fifth of smokers are able to maintain long-term (12 months)
abstinence with any of the present pharmacotherapies,16,17
new and improved drugs are needed.
Chemistry
Analogues 4a-g and 4q-t were synthesized in a fashion
similar to that reported in the literature for optically active 4a
starting with an aryl ketone (Scheme 1).24 The commercially
unavailable ketones (8) were synthesized by Grignard addi-
tionstocommercially available arylnitriles(7). (Z)-tert-Butyl-
dimethylsilylenol ether formation from these propiophe-
nones, 8a-k, using t-butyldimethylsilyl triflate in methylene
chloride, gave high yields of the (Z)-enol ethers 9a-k. The key
transformation in this sequence is a chiral Sharpless hydro-
xylation reaction of these enol ethers, which when using
AD-mix-β, provided the (R)-R-hydroxy ketones 10a-k. The
products of these reactions were not checked for optical purity
but were found to be optically active, so optical induction was
successful at some level. Because of possible epimerization
throughout the process, it was decided to aminate the ketone
before establishing optical purity. Initial efforts to reproduce
the literature preparation of (2S,3S)-4a by converting (R)-1-
(3-chlorophenyl)-2-hydroxypropan-1-one (10a) to the desired
product [(2S,3S)-4a] using the literature conditions with
2-amino-2-methyl-1-propanol and 2,6-lutidine failed or were
low yielding. The presence of 2,6-lutidine overwhelmed silica
gel chromatography, making purification difficult. A mod-
ified approach was developed using proton sponge and
provided (2S,3S)-4a in good yields. This modified procedure
was used to synthesize compounds 4a-g and 4q-t.
Analogues 4n, 4p, 5, and 6 were synthesized as racemic
mixtures by following the standard synthesis of bupropion
analogues25 except substituting 2-amino-2-methyl-1-propanol
for t-butylamine, shown in Scheme 2. As illustrated in
Scheme 2, the appropriate ketones (8l-o) were first synthe-
sized by the addition of ethylmagnesium bromide to the nitriles
(7), or in the case of the 3-pyridyl analogue was synthesized by
lithium halogen exchange starting with 3-bromopyridine (11)
and adding proprionitrile. Simple bromination to form the
R-bromo ketones (12a-d) followed by amination with 2-amino-
2-methyl-1-propanol provided the desired analogues in good
yield. It should be noted that the optically active syntheses of 5
and 6 were attempted using the approach in Scheme 1, but the
Sharpless reaction failed to provide the desired product.
Bupropion [(()-2-tert-butylamino-30-chloropropiophenone]
is used clinically as a racemic mixture of its (R)- and(S)-isomers,
(R)-bupropion [(R)-2] and (S)-bupropion [(S)-2], respectively.
Bupropion is extensively metabolized, with less than 1%
recovered intact in urine.16 Major metabolites result from
hydroxylation of the N-tert-butyl group by P450-(CYP)2B6
isoenzyme.17-19 The resulting hydroxylated metabolites cyclizes
to give (2R,3R)- and (2S,3S)-hydroxybupropion [(2R,3R)-4a
and (2S,3S)-4a, respectively]. Several studies suggest that
(2S,3S)-hydroxybupropion [(2S,3S)-4a] contributes to the anti-
depressant and smoking cessation efficacy of 2.20,21 Peak plasma
and cerebrospinal fluid concentrations of (2S,3S)-4a exceed
those of 2by 4- to 7-fold, and (2S,3S)-4a has a longer elimination
half-life than the parent drug.22,23
In a previous study, we reported that (2S,3S)-4a was an
inhibitor of both dopamine (DA) and norepinephrine (NE)
uptake.21 Importantly, we found that (2S,3S)-4a was a non-
competitive functional antagonist at R4β2-nAChRs, with an
IC50 value of 3.3 μM, a concentration that is comparable
to those needed to inhibit DA and NE uptake. In addition,
(2S,3S)-4a was 3-10 times more potent than 2 after acute
administration in mice in antagonizing nicotine-induced
hypomobility and hypothermia and nicotine-induced analgesia
in tail-flick and hot-plate tests. Compound (2S,3S)-4a also
was equally potent with 2 in the mouse forced-swim test of
depression. Because (2S,3S)-4a has higher potency at relevant
targets compared to 2, it may be a better drug candidate for
smoking cessation pharmacotherapy. Moreover, (2S,3S)-4a can
serve as a lead structure to design analogues with greater potency
at the relevant targets and to possess better drug-like properties.
In this study, we report the synthesis and biological evalua-
tion of 23 hydroxybupropion analogues 4b-4v, 5, and 6 (all
with the (2S,3S)-stereochemistry except for 4n, 4p, 5, and 6,
which are racemic). Some of the analogues have higher
potencies than 2 for DA and NE uptake inhibition as well
as for antagonism of the R4β2-nAChR. In addition, some of
the compounds antagonize the antinociceptive, hypolocomo-
tion, and hypothermic effects of acutely administered nicotine
in mice with potencies greater than that of 2 or (2S,3S)-4a.
2-(30-Bromophenyl)-3,5,5-trimethylmorpholin-2-ol (4d), which
was 19-fold more potent in inhibition of DA uptake and 6-fold
more potent as an R4β2*-nAChR antagonist than (2S,3S)-4a,
has one of the more interesting in vitro properties.
The synthesis of 4 h-m and 4o was accomplished by a novel
convergent synthetic approach for the preparation of 2-sub-
stituted morpholinols as outlined in Scheme 3. This new
approach utilized a nucleophilic addition of Grignard reagents
to (3S)-3,5,5-trimethylmorpholin-2-one (15). Treatment of methyl
(R)-(þ)-lactate (13) with trifluoromethanesulfonic anhydride

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4733
Scheme 2a
a Reagents: (a) EtMgBr; (b) nBuLi, CH3CH2CN; (c) Br2; (d) 2-amino-2-methyl-1-propanol, CH3CN.
Scheme 3a
a Reagents: (a) Tf2O, 2,6-lutidine; (b) 2-amino-2-methyl-1-propanol, CH2Cl2, -40 °C to RT; (c) arylmagnesium bromide.
Scheme 4a
and 2,6-lutidine at 0 °C gave methyl (2R)-2-{[(trifluoromethyl)-
sulfonyl]oxy}propionate (14) in 77% yield. The alkylation of
2-amino-2-methyl-1-propanol with triflate 14 at -40 °C for 2 h
and overnight at room temperature, and subsequent cyclization
afforded 15 in 63% yield. To test the approach, reaction of
lactone 15 with 3-chlorophenylmagnesium bromide resulted
in the formation of (2S,3S)-trimethyl-2-(30-chlorophenyl)mor-
pholin-2-ol [(2S,3S)-4a] in 32% yield (98% ee), 16% overall
from methyl-(R)-(þ)-lactate (13). The addition of the appro-
priate arylmagnesium bromide to 15 provided 4 h-m and 4o.
The C-3 stereocenter of these compounds was derived from
the lactate, not created by a synthetic transformation such as
the Sharpless hydroxylation used in Scheme 1. This center was
then leveraged to create the second C-2 stereocenter. The
resulting stereochemistry at C-2 was a result of either facial
selectivity during the Grignard addition anti to the C-3 methyl
group and/or a thermodynamic equilibrium of the final pro-
duct to the S,S-configuration because the resulting product can
ring open and close. The ring opened form loses its C-2
stereochemistry, forming a ketone. This route was more con-
vergent than the Sharpless hydroxylation route, and was more
reliable, requiring far less analytical work. All of the com-
pounds could have been synthesized by the new route, but this
was deemed inefficient because those compounds were already
available for pharmacological testing.
N-Methylated compounds 4u and 4v were synthesized from
their nonalkylated analogues (4a and 4i, respectively) by reac-
tion with methyl iodide in the presence of potassium carbonate
(Scheme 4).
In Vitro Assays. The (2S,3S)-4a analogues 4b-4v and 5
and 6 were evaluated for their ability to block reuptake of
[3H]dopamine ([3H]DA), [3H]serotonin ([3H]5HT), and [3H]-
norepinephrine ([3H]NE), respectively, into HEK293 cells
stably expressing human DA transporters [(h)DAT], 5HT
transporters [(h)SERT], or NE transporters [h(NET)] using
methods similar to those previously reported.21,26 The results
are given in Table 1.
a Reagents: (a) CH3I, K2CO3.
Compound (2S,3S)-4a and analogues 4b-4v and 5 and 6
also were evaluated for their ability to antagonize functional
responses of R3β4*-, R4β2-, R4β4-, and R1*-nAChR using
previously reported methods21 modified as described in
Experimental Section. Results are given in Table 1 and in
Figures 1-2.
In Vivo Assays. Compound (2S,3S)-4a analogues 4b-4v
and 5 and 6 also were evaluated for their ability to antagonize
behavioral responses to acute nicotine administration as
previously described.21 Results are given in Table 2.
Results
Effects on Monoamine Uptake. Compound 2 inhibits DA
reuptake (IC50 = 660 nM), which would increase synaptic
levels of DA and presumed reward. Compound (2S,3S)-4a
(IC50 = 630 nM), but not (2R,3R)-4a (IC50 > 10 μM), is as
effective as 2 in inhibiting DA uptake inhibition (Table 1).
Compound 2 also inhibits NE reuptake (IC50 = 1850 nM),
which increases synaptic levels of NE. Interestingly, (2S,3S)-4a
(IC50=241 nM), but not (2R,3R)-4a (IC50=9900 nM), is 7.7
times more potent than 2 in inhibiting NE uptake (Table 1).
Neither 2 nor its hydroxymetabolites are active (IC50 > 10 μM;
Table 1) as inhibitors of 5HT uptake.
Among the new hydroxybupropion analogues tested, the
propyl extended chain form 4t and the (()-30,40-dichlorophenyl

4734 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
Table 1. Inhibition of Monoamine Uptake and nAChR Function for Hydroxybupropion Analogues
monoamine uptake inhibitiona IC50 (nM)
nAChR inhibitionb IC50 (μM)
compdc
R1
R2
X
Y
Z
[3H]DA
[3H]NE
[3H]5HT
R3β4*
R4β2
R4β4
R1*β1
2
(2R,3R)-4a CH3
660 ( 178
IA
1850 ( 300
9900 ( 1400
241 ( 60
IA
IA
1.8 (1.15)
6.5 (1.20)
11 (1.48)
8.9 (1.23)
15 (1.12)
3.2 (1.12)
8.6 (1.12)
11 (1.07)
14 (1.10)
20 (1.15)
5.1 (1.07)
8.6 (1.07)
11 (1.10)
1.3 (1.12)
12 (1.15)
31 (1.12)
3.3 (1.07)
6.4 (1.23)
1.3 (1.17)
12 (1.07)
41 (1.07)
30 (1.10)
92 (1.29)
IA
7.9 (1.12)
7.6 (1.12)
28 (1.45)
IA
H
Cl
Cl
H
F
H
H
H
H
H
H
H
H
F
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
F
(2S,3S)-4a
4b
4c
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
C2H5
C3H7
CH3
CH3
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
630 ( 50
1065 ( 30
1380 ( 360
3340 ( 680
2600 ( 400
16,000 ( 2000 3000 ( 900
12,000 ( 4000 1210 ( 340
4200 ( 700
285 ( 70
832 ( 260
IA
IA
IA
550 ( 90
740 ( 150
920 ( 300
1130 ( 20
IA
IA
IA
4d
4e
Br
CH3
0.55 (1.23) 30 (1.07)
18 (1.07)
33 (1.07)
49 (1.07)
96 (1.10)
69 (1.17)
19 (1.15)
20 (1.15)
25 (1.10)
IA
IA
6.0 (1.20)
10 (1.26)
4.8 (1.26)
12 (1.10)
9.2 (1.17)
12 (1.12)
27 (1.23)
1.8 (1.12)
64 (1.20)
IA
4f
4g
CH3O
NO2
IA
IA
80 (1.10)
IA
4h
4i
H
3800 ( 600
830 ( 90
1680 ( 330
IA
H
Cl
4600 ( 900
IA
IA
33 (0.05)
62 (1.10)
72 (1.12)
4j
H
CH3
CH3O
C6H5
F
4k
4l
H
H
IA
10,300 ( 1500 IA
8.1 (1.07) 5.9 (1.15)
4m
4nc
4o
4pc
F
Cl
2140 ( 180
70 ( 20
1020 ( 190
740 ( 110
114 ( 30
151 ( 43
2440 ( 730
IA
11.9 (1.15) 12 (1.07)
IA
36 (1.2)
7.2 (1.12)
23 (1.12)
7.2 (1.07)
6.6 (1.10)
11 (1.12)
14 (1.10)
10 (1.07)
57 (1.05)
43 (1.05)
IA
Cl
H
360 ( 40
IA
IA
2.6 (1.10)
11 (1.15)
3.9 (1.07)
5.2 (1.15)
2.0 (1.05)
4.3 (1.12)
4.8 (1.10)
6.5 (1.05)
20 (1.07)
6.3 (1.51)
11 (1.05)
9.0 (1.07)
6.5 (1.07)
2.9 (1.10)
7.5 (1.05)
7.1 (1.07)
42 (1.17)
IA
14 (1.17)
62 (1.07)
18 (1.17)
13 (1.10)
11 (1.10)
16 (1.05)
18 (1.07)
43 (1.20)
91 (1.12)
IA
F
Cl
H
Cl 8250 ( 720
4q
4r
1-napthyl
2-napthyl
Cl
;
10,000 ( 4000 411 ( 53
1565 ( 215
334 ( 42
2500 ( 540
4130 ( 770
IA
453 ( 4
204 ( 23
30 ( 4
3400 ( 600
2870 ( 820
IA
1570 ( 430
43.4 ( 72
31 ( 10
415 ( 9
527 ( 104
IA
4s
H
H
H
Cl
H
H
H
H
4t
4u
Cl
CH3 Cl
CH3
4v
5c
H
6480 ( 1280 4.6 (1.15)
IA
IA
IA
IA
6c
IA
7950 ( 1800
IA
IA
IA
a Values for mean ( standard error of three independent experiments, each conducted with triplicate determination. b Mean micromolar IC50 values
(to two significant digits) for bupropion and the indicated analogues from three independent experiments for inhibition of functional responses to an
EC80-EC90 concentration of carbamylcholine mediated by nAChR subtypes composed of the indicated subunits (where * indicates that additional
subunits are or may be additional assembly partners with the subunits specified; see Experimental Section). Numbers in parentheses indicate SEM as a
multiplication/division factor of the mean micromolar IC50 values shown [i.e., the value 1.8 (1.15) reflects a mean IC50 value of 1.8 μM with an SEM
range of 1.8 ꢀ 1.15 μM to 1.8/1.15 μM or 1.6-2.1 μM]. The value 11 (1.48) reflects a mean IC50 value of 11 μM with an SEM range of 11 ꢀ 1.40 μM to 11/
1.48 μM or 7.4-16 μM. IA: IC50 > 100 μM. c Compounds 4b-4m, and 4o-4v are all (2S,3S)-isomers. Compounds 4n, 4p, 5, and 6 are racemic materials.
derivative (()-4n of (2S,3S)-4a with IC50 values of 30 and
70 nM, respectively, the ethyl extended chain form 4s (IC50
204 nM) and the 4-chlorophenyl analogue 4i (IC50=285 nM)
have higher potency than (2S,3S)-4a (IC50=630 nM) as inhi-
bitors of DA uptake.
In terms of activity for NE uptake inhibition, the ethyl and
propyl extended chain forms, 4s and 4t (IC50 values of 43 and
31 nM, respectively), and the (()-30-,40-dichlorophenyl deri-
vative (()-4n (IC50=114 nM), are more potent than (2S,3S)-
4a (IC50=241 nM).
The only analogues that had submicromolar IC50 values
for inhibition of 5HT uptake were the (()-30,40-dichlorophe-
nyl analogue (()-4n (IC50 = 360 nM) and the 2-napthyl
analogue 4r (IC50 =334 nM). (2S,3S)-4a, (2R,3R)-4a, and
16 of the analogues were inactive at the SERT. The remain-
ing analogues had IC50 values greater than 1560 nM.
Compound 2 has ∼3-fold selectivity for inhibition at DA
over NE uptake and is inactive at inhibition of 5HT uptake.
Neither (2R,3R)-4a nor (2S,3S)-4a shows activity for 5HT
uptake. Compound (2R,3R)-4a’s poor activity overall pre-
cludes comments about its transporter selectivity. On the
other hand, (2S,3S)-4a has the opposite DA/NE inhibition of
uptake selectivity relative to 2, exhibiting ∼3-fold selectivity for
inhibition of uptake of NE over DA. Analogues 4b-4g, 4l-4m,
4o-4q, 4s, and 4u-4v share with (2S,3S)-4a selectivity for
inhibition of NE over DA uptake inhibition. Relative to
(2S,3S)-4a, 4s and 4o have the best combination of higher
potency in NE uptake inhibition and selectivity for NE over
DA uptake inhibition. The propyl-extended chain analogue, 4t,
has much higher potency than (2S,3S)-or(2R,3R)-4a at each of
the monoamine transporter targets, but it is essentially equipo-
tent for NE and DA uptake inhibition (IC50 = 31 and 30 nM).
The only analogues tested with selectivity for inhibition of DA
over NE uptake [discounting the small preference for DA
over NE and 5HT inhibition shown by (()-30,40-dichloro-
phenyl analogue (()-4n] are the 40-chlorophenyl analogue 4i or
40-methylphenyl analogues 4j and 2-napthyl analogue (4r)
(3-5-fold). Only the 2-napthyl analogue 4r shows slight selectiv-
ity for the SERT (IC50 = 334 nM) over NET (IC50 = 1570 nM).
However, by contrast to 4r, the structurally related 1-napthyl
analogue (4q) has 24-fold selectivity for inhibition of NE over DA
uptake. Thus, alkyl extension as well as phenyl substitution can
impact inhibitory potency and selectivity of the hydroxybupro-
pion analogues for monoamine transporters.
=
In Vitro Profiles for Drug Action at nAChR. Effects of
hydroxybupropion analogues 4b-4v and 5 and 6 on function

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4735
Figure 1. Specific 86Rbþ efflux (ordinate; percentage of control) was determined for functional, human muscle-type R1β1γδ-nAChR (b),
ganglionic R3β4*-nAChR (O), R4β2-nAChR (2), or R4β4-nAChR (3) naturally or heterologously expressed in human cell lines in the presence
of a receptor subtype-specific, EC80-EC90 concentration of the full agonist, carbamylcholine, either alone or in the presence of the indicated
concentrations (abscissa, log molar) of (2S,3S)-hydroxybupropion [(2S,3S)-4a] or its analogues (compounds 4d, 4c, and 4g) as indicated. Mean
micromolar IC50 values and SEM as a multiplication/division factor of the mean micromolar IC50 value are provided in Table 1.
Figure 2. Specific 86Rbþ efflux (ordinate; percentage of control) was determined for functional, human muscle-type R1β1γδ-nAChR (b),
ganglionic R3β4*-nAChR (O), R4β2-nAChR (2), or R4β4-nAChR (3) naturally or heterologously expressed in human cell lines in the presence
of a receptor subtype-specific, EC80-EC90 concentration of the full agonist, carbamylcholine, either alone or in the presence of the indicated
concentrations (abscissa, log molar) of (2R,3R)-hydroxybupropion [(2R,3R)-4a] or analogues (compounds 4s, 4w, and 4t) as indicated. Mean
micromolar IC50 values and SEM as a multiplication/division factor of the mean micromolar IC50 value are provided in Table 1.
of diverse, human nAChR subtypes naturally or heterologously
expressed by human cell lines were assessed using 86Rbþ efflux
assays that are specific only for nAChR function in the cells
used. None of the analogues has activity as an agonist at R1*-,
R3β4*-, R4β2-, or R4β4-nAChR because 86Rbþ efflux in the
presence of these ligands alone at concentrations from ∼5nMto
100 μM (data not shown here) was indistinguishable from
responses in cells exposed only to efflux buffer.

4736 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
Table 2. Pharmacological Evaluation of Hydroxybupropion Analogues As Noncompetitive Nicotinic Antagonistsa
AD50 (mg/kg)
compdb
tail-flickc
1.2 (1-1.8)
0.2 (0.1-1.1)
2.5 (1.7-3.5)
IA
hot-platec
locomotionc
hypothermiac
2
(2S,3S)-4a
(2R,3R)-4a
15 (6-19)
1.0 (0.5-4.5)
10.3 (8.9-15)
IA
4.9 (0.9-46)
0.9 (0.38-3.7)
IA
9.2 (4-23)
1.5 (0.95-2.6)
IA
4b
4c
4d
4e
4f
4g
4h
4i
IA
IA
IA
4.4 (1.3-14.5)
1.7 (0.5-6.8)
2.3 (0.4-11)
IA
0.012 (0.002-0.16)
0.16 (0.05-0.6)
0.054 (0.04-0.066)
5.85 (3.8-8.8)
4.9 (1.6-15)
0.013 (0.005-0.03)
0.019 (0.063-0.1)
0.004 (0.002-0.012)
0.019 (0.06-0.064)
0.021 (0.005-0.1)
0.006 (0.004-0.01)
8.8 (4.3-18)
8.6 (0.7-10.5)
4.3 (1.8-9.8)
7.6 (2-29)
IA
2.6 (0.7-10.1)
IA
IA
IA
IA
IA
4.7 (3.5-6.2)
5.67 (2.5-12.8)
IA
IA
IA
IA
IA
4j
IA
IA
IA
IA
4k
4l
IA
IA
IA
IA
IA
IA
4m
4n
4o
4p
4q
4r
4s
4t
4u
4v
5
IA
IA
IA
IA
IA
IA
0.0056 (0.004-0.009)
IA
IA
IA
1.9
IA
IA
IA
0.034 (0.001-0.1)
0.04 (0.001-0.6)
0.004 (0.001-0.03)
0.004 (0.001-0.03)
0.016 (0.004-0.06)
0.32 (0.04-2.5)
n/a
IA
IA
IA
10.3 (1.4-75)
4.7
IA
IA
n/a
n/a
IA
7 (4.5-10.8)
IA
IA
IA
n/a
n/a
3.7 (0.8-17)
IA
IA
IA
n/a
n/a
6
n/a
a Results were expressed as AD50 (mg/kg) ( confidence limits (CL) or % effect at the highest dose tested. Dose-response curves were determined
using a minimum of four different doses of test compound, and at least eight mice were used per dose group. b Compounds 4b-4m, and 4o-4v are all
(2S,3S)-isomers. Compounds 4n, 4p, 5, and 6 are racemic materials 1. c n/a: not assayed; IA: AD50 > 15 mg/kg; NFT = no further testing.
86Rbþ efflux assays also were used to assess whether ligands
had activity as antagonists at human nAChR. Representative
concentration-response curves for selected ligands (2S,3S)-
4a, 4d, 4c, and 4g (Figure 1) and (2R,3R)-4a, 4s, 4u, and 4t
(Figure 2) illustrate nAChR in vitro inhibitory profiles (see also
Table 1). As we have shown previously,21 (2S,3S)-4a is a full
antagonist at all of the nAChR subtypes tested and is selective
( p<0.05) for R4β2- over R3β4*-, R4β4-, and R1*-nAChR
(upper left panel, Figure 1). Compounds 4c, 4d, and 4g also
block all nAChR function at an adequately high concentration,
4c and 4d have higher potency (p< 0.05) than (2S,3S)-4a at
R4β2-nAChR, and all three compounds are more selective than
(2S,3S)-4a for R4β2-nAChR over the other nAChR subtypes
(Figure 1). By contrast, (2R,3R)-4a, which also is a full antago-
nist at all of the nAChR subtypes tested, is selective (p<0.05)
for R3β4*- and R1*-nAChR over R4β2- and R4β4-nAChR
(upper left panel, Figure 2). Compounds 4s and 4u have very
similar potency as inhibitors of R4β2- and R3β4*-nAChR
function and are selective for those over R1*- and R4β4-nAChR
(Figure 2). Compound 4t is another full antagonist at all nAChR
subtypes but with diminished selectivity for R3β4*-nAChR
(Figure 2). Note, more generally, that all of the ligands tested
were full antagonists in that they blocked all nAChR function at
the highest concentration tested or tended to do so if there still
was submaximal block at 100 μM. Not shown here are other
studies demonstrating that antagonism in all cases was mediated
noncompetitively in that agonist concentration-ion flux response
curves in the presence of ∼IC50 concentrations of analogues
showed diminished efficacy of agonist relative to response curves
obtained in the absence of analogues and that agonist apparent
EC50 values were unaffected by the presence of analogues.
As was previously noted,21 (2S,3S)-4a has an altered nAChR
functional inhibitory profile relative to 2, showing higher
potency at R4β2-nAChR (IC50 = 3.3 μM compared to 12 μM
for 2) and higher selectivity for R4β2- over other nAChR
(∼3-fold) (Table 1).
Relative to (2R,3R)-4a and (2S,3S)-4a (IC50 values of 6.5 and
11 μM, respectively), the 30-deschlorophenyl-analogue 4b and
the 30-methylphenyl and 40-methylphenyl-analogues 4e and
4j have comparable inhibitory potencies at R3β4*-nAChR
(IC50 = 8.5-8.9 μM; Table 1). By contrast, there is slightly
higher potency at R3β4*-nAChR for the 30-bromophenyl 4d,
30-chlorophenyl-4-(N-methyl) 4u, ethyl and propyl chain ex-
tended analogues 4s and 4t, 40-chlorophenyl 4i, 30,50-dichloro-
phenyl 4p, 1-naphthyl 4q, and 40-chlorophenyl-4-(N-methyl) 4v
analogues (IC50 = 3.2-6.5 μM; p<0.05 for IC50 value
differences between (2R,3R)-4a and 4d or 4p). Also having
higher antagonist potencies (p < 0.05) than (2R,3R)-4a at
R3β4*-nAChR are the (()-30,40-dichlorophenyl analogue
(()-4n and the 2-naphthyl analogue 4r (IC50 values of 2.6
and 2.0 μM, respectively (Table 1)). For these analogues,
IC50 values for inhibition of R1*-nAChR function are
higher (>33 μM for 4b, 4e, 4u, and 4v) or are in the range
(5.9-19 μM for 4d, 4i-4j, 4l, (()-4n, 4p-4t) of IC50 values
for inhibition of R1*-nAChR by (2S,3S)-4a or (2R,3R)-4a
(28 or 7.6 μM, respectively).
In absolute terms, the 30-bromophenyl- (4d), 30-fluorophenyl-
(4c), biphenyl 4l, and ethyl extended chain analogue 4s have
higher inhibitory potency at R4β2-nAChR than (2S,3S)-4a
[IC50 values of 0.55, 1.3, 1.8 (all p<0.05) and 2.9 relative to
3.3 μM, respectively].
Of the analogues tested, 4v has the largest selectivity for
R3β4*-nAChR over the other nAChR subtypes (Table 1).
(2R,3R)-4a is barely selective for R3β4*-nAChR over R1*-
nAChR, but about 5-fold selective for R3β4*- over R4β2-
nAChR (Table 1). Of all the analogues tested, 4b-4e, 4 g-4h,

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4737
4o, and 4s have reasonable selectivity for R4β2-nAChR over
other nAChR subtypes. Compounds 4f, 4m, and 4u are about
equipotent at R4β2- and R3β4*-nAChR. From another per-
spective, only the racemic 30,40-dicholoro (()-4n (∼8-fold)
and 40-chloro-4-(N-methyl) 4v (∼9-fold) have higher selectivity
for R3β4*- over R4β2-nAChR than (2R,3R)-4a (∼5-fold) or 2
(∼6.7-fold). The 30-fluorophenyl 4c (∼12-fold) and 30-bromo-
phenyl 4d (∼6-fold) analogues have selectivity for R4β2-nAChR
over other nAChR subtypes better than or comparable to that
for (2S,3S)-4a (∼3-fold).
is blocked by 2 with an AD50 of 1.2 mg/kg. Compound (2S,3S)-
4a with an AD50 = 0.2 mg/kg (Table 2) has a 6-fold increase in
effectiveness in this assay. Fifteen of the new analogues have
even higher potency. AD50 values (in mg/kg) are 0.004 for 4j, 4s,
and 4t, ∼0.006 for 4m and 4o, 0.012-0.013 for 4c and 4h, 0.016
for 4u; 0.019 for 4i and 4k, 0.021 for 4l, between 0.034 and 0.054
for 4q, 4r, and 4e, and 0.16 for 4d (Table 2).
At the doses tested, none of the compounds showed
sedative, convulsive, or other obvious toxic effects in mice.
Thus, in four assays of the ability of analogues to block
acute actions of nicotine, 30-bromophenyl analogue 4d was
more effective than 2 in each and rivaled effects of (2S,3S)-4a
in tail-flick and hypothermia assessments. The 30-fluorophe-
nyl analogue 4c, 30-methylphenyl analogue 4e, and ethyl
extended chain analogue 4s exceeded 2’s potency in three
assays and had higher potency than (2S,3S)-4a in the tail-
flick assay. Compounds 4c, 4d, and 4s also are remarkable
for their higher inhibitory effectiveness at R4β2-nAChR than
(2S,3S)-4a. Eleven of the other ligands had better effective-
ness than 2 in one of the acute assays, with potency in the tail-
flick assay correlating with IC50 values <10 μM for inhibi-
tion of R4β2-nAChR function for 4c-4e, 4l, 4o, and 4q-4u
but not for 4 h-4k, 4m and 4v, which were potent in the tail-
flick assay but not as R4β2-nAChR antagonists, or for 4b, 4f,
and 4g, which lacked effectiveness relative to (2S,3S)-4a in
the tail-flick assay but have sub-10 μM IC50 values at R4β2-
nAChR. Lacking potency in tail-flick or inhibition of R4β2-
nAChR function, but having activity as antagonists of
R3β4*-nAChR, are 4n and 4p. Increased in vitro inhibition
of DA and NE uptake did not necessarily correlate with
increased in vivo inhibitory potency in nicotine-sensitive
assays if there also was no increase in effectiveness at
R4β2-nAChR or selectivity for that target (e.g., 4n). On the
other hand, the ligands with the highest inhibitory potency
for NE uptake inhibition (and highest or higher effectiveness
than (2S,3S)-4a at DA uptake inhibition) were among the
most potent inhibitors of nicotine-induced analgesia in the
tail-flick assay (4s and 4t). Improvement over (2S,3S)-4a in
the ability to block nicotine-induced analgesia in the tail-
flick assay as seen for 4j, 4m, and 4o has no obvious basis as
judged from in vitro assays.
Multiple Target Actions of Hydroxybupropion Analogues.
We compared inhibitory potencies across transporters and
nAChR relative to (2S,3S)-4a, which has ∼3-fold selectivity
for inhibition of NE over DA uptake inhibition and ∼14-fold
selectivity for inhibition of NE uptake inhibition over
R4β2-nAChR function. Interestingly, there is a change to
absolute selectivity for inhibition of R4β2-nAChR over DA
uptake inhibition (∼6-fold) and over NE uptake inhibition
(∼1.7-fold), and there is ∼3.6-fold selectivity for inhibition
of NE over DA uptake for 30-bromophenyl 4d. There is an
even more striking increase in selectivity for inhibition of
R4β2-nAChR function over transporter inhibition for bi-
phenyl analogue 4l (>5-fold over DA uptake inhibition and
6-fold over NE uptake inhibition). Selectivity for inhibition
of NE uptake inhibition over R4β2-nAChR function also is
reduced relative to that for (2S,3S)-4a for analogues 4c
(∼1.7-fold), 4h (∼3-fold), 4f (∼3.3-fold), 4g and 4r (∼4-fold),
4e (∼5.3-fold), and 4b (∼12-fold), and for analogue 4p (∼4.5-
fold). Conversely, there is an increase in selectivity for
inhibition of NE uptake over R4β2-nAChR function for 4t
(∼240-fold), (()-4n (∼175-fold), 4v (80-fold), 4s (∼67-fold),
and 4o (42-fold), although selectivity for inhibition of NE
uptake over inhibition of R3β4*- instead of R4β2-nAChR
function is less for 4t, (()-4n, and 4v (∼154-, 23-, and 9-fold,
respectively; recall that 4t has comparable activity for DA
and NE uptake inhibition). Although its selectivity for
inhibition of DA over NE uptake is marginal, (()-4n has
∼37-fold selectivity for inhibition of DA uptake over R3β4*-
nAChR and >285-fold selectivity for inhibition of DA
uptake over R4β2-nAChR. The biphenyl analogue 4l has 7.9-
fold selectivity for inhibition of R3β4*-nAChR over inhibition
of NE uptake, surpassing selectivity seen for (2R,3R)-4a
(∼1.5-fold).
Discussion
In Vivo Effects of Analogues. Compound 2 blocks nicotine-
induced increases in locomotor activity with an AD50 value
of 4.9 mg/kg, while (2S,3S)-4a has a lower AD50 value of
0.9 mg/kg in the same assay, but none of the analogues is better
than (2S,3S)-4a, although 4d and 4p (2.6 and 1.9 mg/kg AD50,
respectively) have slightly higher potency than 2 (Table 2).
The ability of (2S,3S)-4a to block the nicotine-induced
decrease in body temperature is better than that for 2
[AD50 = 1.5 and 9.2 mg/kg, respectively (Table 2)]. Com-
pound 4d (AD50 = 1.7 mg/kg) also rivals (2S,3S)-4a in this
assay, and analogues 4c, 4e, 4g, 4h, and 4s have intermediate
potencies (2.3-7 mg/kg AD50 values).
In the hot-plate assay, 2 blocks nicotine-induced, suprasp-
inally mediated analgesia with an AD50 value of 15 mg/kg.
Compound (2S,3S)-4a is 15-fold more potent (AD50 value of
1 mg/kg), but blockade of nicotine-induced hot-plate analge-
sia is no better for any of the new analogues tested. However,
4c, 4d, 4e, and 4s (3.7-8.6 mg/kg AD50 values) are more
potent than 2 (Table 2).
New analogues of the hydroxybupropion isomer (2S,3S)-
4a were generated and assessed for their ability to affect
monoamine uptake, the function of four different nAChR
subtypes, and the acute effects of nicotine on nociception,
locomotor activity, and hypothermia.
We succeeded in generating analogues with reasonably
higher inhibitory potency than 2 or either of its hydroxy-
metabolite isomers (2R,3R)-4a and (2S,3S)-4a for DA uptake
inhibition (4i, 4n, 4s, and 4t), NE uptake inhibition [(()-4n, 4s,
and 4t], or 5HT uptake inhibition [(()-4n and 4r], or for
functional inhibition of R3β4*-nAChR (4l) or R4β2-nAChR
(4c, 4d, 4l, and4s). Selectivityfor inhibitionof DAuptakeover
nAChR or NE uptake inhibition better than that of 2 was
achieved with retention of reasonable potency at DAT for 4i.
As predicted based on our previous studies of hydroxymeta-
bolites, many of the compounds evaluated have some selec-
tivity for inhibition of NE over DA uptake. This was
improved relative to (2S,3S)-4a for 4f-4g, 4s, 4v, 4o, 4u,
and 4q, but only 4s and 4o have improved potency at NE
uptake inhibition. Compounds (()-4n, 4t, and 4s are more
Nicotine-induced analgesia in the tail flick assay, which
assesses spinal processes involved in nicotine antinociception,27

4738 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
selective than (2S,3S)-4a for inhibition of NE uptake inhibi-
tion over nAChR function. Only (()-4n and 4v have increased
selectivity for R3β4*-nAChR over other nAChR subtypes
relative to 2 or (2R,3R)-4a. Compounds 4l and 4k have
improved selectivity for R3β4*-nAChR functional inhibition
over DA and NE uptake inhibition relative to that of (2R,3R)-
4a. Compound 4d has absolute selectivity for inhibition of
R4β2-nAChR over inhibition of R3β4*-nAChR as well as
over NE, 5HT, or DA uptake inhibition, and 4l’s similar
potency at R4β2- and R3β4*-nAChR also means that it is
selective for inhibition of R4β2-nAChR over monoamine
transporters. Selectivity for R4β2-nAChR over R3β4*- and
other nAChR subtypes was increased for 4c and 4d relative to
that for 2 and (2S,3S)-4a. Selectivity for inhibition of R4β2-
nAChR over NE uptake inhibition was increased relative to
that of (2S,3S)-4a for 4b, 4c, 4e, 4f, 4g, 4p, and 4r, with 4d
actually being absolutely selective for inhibition of R4β2-
nAChR function over other nAChR subtypes and over
inhibition of DA and NE uptake.
From a chemical structure perspective, changes in the
30-chlorophenyl group in (2S,3S)-4a to 30-bromophenyl 4d
or 30-fluorophenyl 4c afforded ligands with improved affinity
and selectivity for R4β2-nAChRs. Selectivity for monoamine
transporter uptake over R4β2-nAChRs functional block was
decreased while selectivity for R4β2-nAChRs over other
nAChR subtypes was preserved by other changes in the
phenyl substitution (nitro 4g, methyl 4e) or lack of chloro
group (4b) but not for 30-methoxyphenyl substitution (4f).
The dichlorophenyl analogues (()-4n and 4p have increased
selectivity for R3β4*-nAChR over other nAChR subtypes. In
addition, the (()-30,40-dichlorophenyl analogue [(()-4n] also
has a marked increase in affinity for inhibition of DA uptake.
Replacement of the 3-methyl group on the morpholinol ring
with an ethyl or propyl group (4s and 4t) affords ligands with
notable improvements in selectivity for inhibition of NE
uptake over nAChR and increases in potencies for inhibition
of DA and NE uptake. Changing the 3-chlorophenyl ring to a
pyridine ring leads to ligands 5 and 6, which are without
activity. Naphthyl analogues 4q and 4r have modestly altered
activities compared to (2S,3S)-4a. Interestingly, moving the
3-substituent of the phenyl group of (2S,3S)-4a and 4e to the
40 position affords ligands (4i-4j) that are selective for DA over
NE uptake and nAChR functional inhibition and for inhibi-
tion of R3β4*- over R4β2-nAChR. However, the biphenyl
analogue (4l) has almost no activity at monoamine transpor-
ters yet is very potent as an inhibitor of R4β2 and R3β4*-
nAChR. Moving the 30-substituent of the phenyl group of the
N-methyl analogue 4u to the 40 position (4v) has little effect
on inhibition or DA or NE uptake but increases selectivity
for functional blockage of R3β4*- over R4β2-nAChRs from
negligible to ∼9-fold.
selectivity for inhibition of NE over DA uptake and for
inhibition of DA over NE uptake, respectively, but having
comparable effects on nAChR function, both had ∼5-fold
higher activity in the tail-flick assay than (2S,3S)-4a. How-
ever, the other in vivo assays mostly did not reveal striking
inhibition by analogues of acute nicotine action. Moreover,
the tail-flick assay could reflect CNS actions of ligands at
a higher level than the presumed spinal level of nicotine-
mediated antinociception in the test.
The current studies dovetail with those based on modifica-
tions to the bupropion template28 to provide insights into
further target-directed drug development. The new analogues
also illuminate roles of specific molecular targets in nicotine-
mediated behavioral effects. Extensions of the R1 methyl
group in (2S,3S)-4a to an ethyl or propyl group (4s and 4t)
preferentially improve activities for inhibition of DA and NE
uptake. The naphthyl analogues (4q and 4r) afford ligands
with better potency for 5HT uptake inhibition, and changing
the 30-chlorophenyl moiety of (2S,3S)-4a to a 30-fluoro or
30-bromophenyl group leads to ligands with better activity at
R4β2-nAChRs (4c and 4d).
Greater challenges come in correlating the in vivo effects to
each other and to the in vitro fingerprints. Formation of
metabolites, differences in pharmacokinetics, or difficulties in
brain penetration following systemic administration could
confound interpretations. However, even for assays that are
done soon after delivery, the chemical modifications that
selectively increase in vitro activity at transporters or at
R4β2-nAChR afford ligands with potential as nicotine an-
tagonists in vivo. That is, increased activity at either R4β2-
nAChR or for inhibition of DA or NE uptake (or for one
ligand, at 5HT uptake inhibition) correlates well with im-
provement in ligand antagonist potency in the tail-flick assay.
Thus, the behavioral results suggest that effects of nicotine
dependence and/or depression could be countered by ligands
acting at DAT, NET, or R4β2-nAChR or any combination of
the three.
Conclusions
The current findings provide part of a preclinical founda-
tion for development of compounds related to (2S,3S)-4a.
Recently, Silverstone et al.29 reported that the racemic hydroxy-
bupropion induced convulsions in mice after ip administration.
However, the potency of (2S,3S)-4a in our mouse testing is
close to 50-fold lower than the racemic hydroxy bupropion
metabolite active doses reported by Silverstone et al.29 Further-
more, we found no evidence for seizure or convulsive activities
at any of the doses tested in our study. Thus, we will continue
exploring use of hydroxybupropion analogues as potentially
superior aids to smoking cessation.
In silico predictions that all of the compounds synthesized
would have drug-like character and activity in the central
nervous system were consistent with the results of behavioral
studies. We obtained several analogues with higher potency
than (2S,3S)-4a or 2 as antagonists of nicotine-mediated
antinociception in the tail-flick assay. The ethyl and propyl
extended chain ligands 4s and 4t with high affinity for inhibi-
tion of DA and NE uptake, the 30-fluoro and 30-bromo phenyl
substituted analogues 4c and 4d with high affinity and good
selectivity for R4β2-nAChR, and to a lesser extent the
30-methylphenyl analogue 4e also with good activity at R4β2-
nAChR, also had better potency in the tail-flick assay than
2 or (2S,3S)-4a. The naphthyl analogues 4q and 4r with
Experimental Section
Nuclear magnetic resonance (1H NMR and 13C NMR) spectra
were recorded on a 300 MHz (Bruker AVANCE 300) unless
otherwise noted. Chemical shift data for the proton resonances
were reported in parts per million (δ) relative to internal (CH3)4Si
(δ 0.0). Optical rotations were measured on an AutoPol III
polarimeter, purchased from Rudolf Research. Elemental ana-
lyses were performedbyAtlantic Microlab, Norcross, GA. Purity
of compounds (>95%) was established by elemental analyses.
Analytical thin-layer chromatography (TLC) was carried out on
plates precoated with silica gel GHLF (250 μM thickness). TLC
visualization was accomplished with a UV lamp or in an iodine
chamber or ninhydrin staining. All moisture-sensitive reactions

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4739
were performed under a positive pressure of nitrogen maintained
by a direct line from a nitrogen source. Anhydrous solvents were
purchased from Aldrich Chemical Co. 3-Methoxypropiophe-
none (8f), 3,4-dichloropropiophenone (8l), 3,5-dichloropropio-
phenone (8m), 1-(pyridin-2-yl)propan-1-one (8n), and 1-(pyridin-
3-yl)propan-1-one (8o), were synthesized, but are now commer-
cially available. 3-Chlorobutyrophenone (8j) and 3-chloropenta-
phenone (8k) were described in an earlier paper on bupropion
analogues.30
Synthesis of (2S,3S)-4a Using Optically Active Sharpless Hydro-
xylation Chemistry. Step 1: (Z)-tert-Butyl-[1-(3-chlorophenyl)prop-
1-enyloxy]dimethylsilane (9a). In a 250 mL flask, 30-chloropropio-
phenone (8a, 10 g, 0.059 mol) was dissolved in 100 mL in CH2Cl2
and cooled with an ice water bath. Et3N (13 mL, 95 mmol) was
added to the solution, followed by slow addition of TBDMSOTf
(15 mL, 65 mmol). After stirring overnight at room temperature,
the reaction mixture was diluted with CH2Cl2 and washed with
NaHCO3. The organic layer was separated, dried (Na2SO4), and
concentrated. The oily residue was purified by column chromato-
graphy on neutral alumina using hexanes (a few drops of Et3N were
added) as the eluent to give 16.4 g (98%) of title product as a
colorless oil. 1H NMR (CDCl3) δ 7.46-7.44 (m, 1H), 7.35-7.32
(m, 1H), 7.21-7.20 (m, 2H), 5.23 (q, 1H, J = 6.9 Hz), 1.73 (d, 3H,
J = 6.9 Hz), 0.99 (s, 9H), -0.03 (s, 6H). C15H23ClOSi.
(2R,3R)-2-(3-Chlorophenyl)-3,5,5-trimethylmorpholin-2-ol
((2R,3R)-4a) Hemi-L-tartrate. Following the procedure described
for (2S,3S)-4a, asampleof(S)-10a (4.5 g, 0.024 mol), was dissolved
in CH2Cl2 (75 mL) and treated with Proton sponge (6.3 g) and
cooled to -50 °C. Next, triflic anhydride (4.5 mL, 32 mmol) was
added slowly, and the reaction mixture was stirred at 0 °C for an
additional hour. The resulting orange slurry was transferred by
syringe to another flask containing a solution of 2-amino-2-
methyl-1-propanol, (4.7 g, 0.052 mol) in CH3CN. After purifica-
tion, 3.3 g (78%) of the (2R,3R)-4a was isolated and converted to
2.7 g of the Hemi-L-tartrate salt (>99%ee): mp 128-131 °C;
[R]23 -13.0° (c 0.79, CH3OH). Characterization data is similar
D
with data reported in the literature.24
(2S,3S)-2-Phenyl-3,5,5-trimethylmorpholin-2-ol (4b) Hemi-
D-tartrate. Compound 4b was synthesized by a procedure similar to
that described for (2S,3S)-4a using (R)-1-phenyl-2-hydroxypro-
pan-1-one (10b, 3.49 g, 0.0233 mol), Proton sponge (5.9 g), triflic
anhydride (4.2 mL, 26 mmol), and 2-amino-2-methyl-1-propanol
(4.6 g, 0.052 mol) in CH2Cl2 (50 mL). After purification by
chromatography on silica gel, 3.52 g (68%) of the free base 4b
was isolated and converted to 1.19 g of the hemi-D-tartrate
salt, which had >99%ee: mp 112-113 °C; [R]20D þ15.8° (c 1.1,
1
CH3OH). H NMR (methanol-d4) δ 7.61-7.59 (m, 2H), 7.44-
7.36 (m, 3H), 4.32 (s, 1H), 4.24 (d, 1H, J= 12.3 Hz), 3.58-3.48 (m,
2H), 1.64 (s, 3H), 1.39 (s, 3H), 1.09 (d, 3H, J = 6.6 Hz). 13C NMR
(methanol-d4) δ 179.1, 142.4, 130.3, 129.6 (2C), 127.9 (2C), 97.3,
75.4, 67.4, 55.47, 55.27, 24.1, 21.3, 14.3. LCMS (ESI) m/z 222.4
Step 2: (R)-1-(3-Chlorophenyl)-2-hydroxypropan-1-one ((R)-10a).
A mixture of AD-mix-β (50.6 g) and CH3SO2NH2 (3.5 g,
0.037 mol) in tert-butyl alcohol-water (120 mL/120 mL) was
cooled at 0 °C and treated with 9a (10 g, 0.036 mol). The reaction
mixture was stirred for 16 h at 0 °C. Sodium sulfite (36 g) was
added, and the mixture was stirred for another hour. The mixture
was filtered through a celite pad and washed with ether. The filtrate
was transferred to a separation funnel and the lower dark-colored
layer was discarded. The upper yellowish phase was separated,
dried (Na2SO4), filtered, and concentrated. The crude product was
purified by column chromatography on silica gel using hexanes-
EtOAc (10:1 to 3:1) as the eluent to give 5.8 g (87%) of title product
(M - tartrate)þ. Anal. (C15H22NO5 0.5H2O) C, H, N.
3
(2S,3S)-2-(3-Fluorophenyl)-3,5,5-trimethylmorpholin-2-ol (4c)
Hemi-D-tartrate. Compound 4c was synthesized by a procedure
similar to that described for (2S,3S)-4a using (R)-1-(3-fluoro-
phenyl)-2-hydroxypropan-1-one (10c, 3.94 g, 0.024 mol), Proton
sponge (6.0 g), triflic anhydride (4.6 mL, 25.8 mmol), and 2-amino-
2-methyl-1-propanol (4.6 g, 0.052 mol) in acetonitrile (50 mL).
After purification, 2.2 g (39%) of the free base 4c was isolated and
converted to the hemi-D-tartrate salt, which had >99%ee: mp
131-132 °C; [R]20D þ20.4° (c 1.0, CH3OH). 1H NMR (methanol-
d4) δ 7.45-7.41 (m, 2H), 7.35-7.31 (m, 1H), 7.11-7.09 (m, 1H),
4.33 (s, 1H), 4.26 (d, 1H, J = 12.0 Hz), 3.56-3.45 (m, 2H), 1.59 (s,
3H), 1.34 (s, 3H), 1.06 (d, 3H, J = 6.6 Hz). 13C NMR (methanol-
d4) δ 178.1, 165.4, 162.1, 144.8, 130.8, 123.2, 116.3 (d), 114.3 (d),
96.2, 74.5, 67.0, 54.3, 23.6, 20.8, 13.7. LCMS (ESI) m/z240.0 [(M -
as light-greenish oil: [R]20 þ64.2° (c 1.2, CHCl3). 1H NMR
D
(CDCl3) δ 7.91 (s, 1H), 7.80 (d, 1H, J = 7.8 Hz), 7.62-7.57
(m, 1H), 7.46 (t, 1H, J=7.8 Hz), 5.15-5.08 (m, 1H), 3.68-3.65
(m, 1H), 1.45 (d, 3H, J=7.1 Hz). C9H9ClO2. Characterization
data is similar with that reported in the literature.24
Step 3: (2S,3S)-2-(3-Chlorophenyl)-3,5,5-trimethylmorpholin-
2-ol [(2S,3S)-4a] Hemi-D-tartrate. A sample of (R)-10a (2.47 g,
0.0134 mol) was added to a 250 mL flask and dissolved in
CH2Cl2 (40 mL). Proton sponge (3.5 g, 0.016 mol) was added
to the reaction flask, and the reaction mixture was cooled to
-50 °C. Triflic anhydride (2.47 mL, 14.7 mmol) was slowly
added to the reaction flask, the temperature was allowed to rise
to 0 °C, and the reaction mixture was stirred for an additional
hour. The resulting orange slurry was transferred by syringe to
another flask containing a solution of 2-amino-2-methyl-1-propa-
nol (2.6 g, 0.029 mol) in CH3CN (40 mL) at -10 °C. After stirring
for 4 h at 0 °C, the precipitate was removed by filtration and the
filtrate was concentrated. The residue was extracted with ether and
solid was removed by filtration and discarded. The filtrate was
concentrated to an oil. Purification of the residue by chromatog-
raphy on silica gel using EtOAc with 1% NH4OH as the eluent
gave 1.39 g (41%) of (2S,3S)-4a as a white solid. Characterization
data is similar with data reported in the literature.24
tartrate)þ, M = C15H21FNO5]. Anal. (C15H21FNO5 0.5H2O)
3
C, H, N.
(2S,3S)-2-(3-Bromophenyl)-3,5,5-trimethylmorpholin-2-ol (4d)
Hemi-D-tartrate. Compound 4d was synthesized by a procedure
similar to that described for (2S,3S)-4a using (R)-1-(3-bromo-
phenyl)-2-hydroxypropan-1-one (10d, 4.0 g, 0.018 mol), Proton
sponge (4.5 g, 0.021 mol), triflic anhydride (3.2 mL, 192 mmol),
and 2-amino-2-methyl-1-propanol (3.4 g, 0.038 mol) in acetonitrile
(50 mL). After purification, 2.04 g (39%) of the free base 4d was
isolated and converted to 1.6 g of the hemi-D-tartrate salt, which
had >99%ee: mp 129-130 °C; [R]20D þ9.6° (c 1.0, CH3OH). 1H
NMR (methanol-d4) δ 7.77-7.76 (m, 1H), 7.63-7.53 (m, 2H),
7.38-7.32 (m, 1H), 4.37 (s, 1H), 4.14 (d, 1H, J = 12.0 Hz),
3.58-3.39 (m, 2H), 1.57 (s, 3H), 1.32 (s, 3H), 1.07-1.02 (m, 3H).
13C NMR (methanol-d4) δ 178.8, 145.1, 133.3, 131.1, 126.9, 123.6,
96.7, 75.2, 67.7, 55.0, 24.4, 21.5, 14.4; LCMS (ESI) m/z 300.6
[(M-tartrate)þ, M = C15H21BrNO5]. Anal. (C15H21BrNO5
0.25H2O) C, H, N.
3
The product freebase (1.2 g, 0.0047 mol) was dissolved in 20 mL
of MeOH and treated with a solution of D-tartaric acid (350 mg,
2.30 mmol) in MeOH (3 mL). After stirring for 5 min at room
temperature, the reaction mixture was concentrated, the sample
dissolved in CH2Cl2 (30 mL) and MeOH was added until the
solution was clear. Next, ether was added slowly until it became
cloudy or small crystals started to form. After keeping the mixture
at 0 °C for 1 h, the white solid was collected by filtration and
(2S,3S)-2-(m-Tolyl)-3,5,5-trimethylmorpholin-2-ol (4e) Hemi-
D-tartrate. Compound 4e was synthesized by a procedure similar
to that described for (2S,3S)-4a using (R)-1-(3-methylphenyl)-2-
hydroxypropan-1-one (10e, 4.2 g, 0.026 mol), Proton sponge (6.5 g,
0.030 mol), triflic anhydride (4.60 mL, 282 mmol), and 2-amino-2-
methyl-1-propanol (5.0 g, 0.056 mol) in acetonitrile (55 mL). After
purification, 4.0 g (66%) of the free base 4e was isolated and con-
verted to 3.6 g of the hemi-D-tartrate salt, which had 94%ee: mp
104-105 °C; [R]20D þ11.9° (c 0.85, CH3OH). 1H NMR (methanol-
d4) δ 7.43-7.38 (m, 2H), 7.28 (t, 1H, J = 7.8 Hz), 7.21-7.18
recrystallized to give 0.7 g of (2S,3S)-4a hemi-D-tartrate as a white
3
solid (ee 98.4%): mp 128-131 °C; [R]23D þ13.7° (c 0.76, CH3OH).
Anal. (C15H21ClNO5 0.5H2O) C, H, N.
3

4740 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
(m, 1H), 4.33 (s, 1H), 4.20 (d, 1H, J = 12.0 Hz), 3.52-3.45 (m,
2H), 2.38 (s, 3H), 1.61 (s, 3H), 1.36 (s, 3H), 1.06 (d, 3H, J = 6.6
Hz). 13C NMR (methanol-d4) δ 179.0, 142.3, 139.4, 130.9, 129.5,
128.4, 125.0, 97.2, 75.3, 67.4, 55.2, 24.2, 21.9, 21.4, 14.4. LCMS
(ESI) m/z 236.2 [(M - tartrate)þ, M = C16H24NO5]. Anal.
(ESI) m/z240.3 [(M - fumaric)þ, M=C13H18FNO2 0.5C4H4O4].
3
Anal. (C15H20FNO4 H2O) C, H, N.
3
(2S,3S)-2-(4-Chlorophenyl)-3,5,5-trimethylmorpholin-2-ol (4i)
Hemi-Fumarate. A solution of 15 (357 mg, 2.50 mmol) in dry
THF (2.5 mL, 1M) under an N2 atmosphere was cooled to
-78 °C and treated with 4-chlorophenylmagnesium bromide
(2 equiv, 5.00mmol, 5.00 mL, 1 M solutioninether). Thereaction
mixture was stirred at -78 °C for 2 h. A saturated aqueous
solution of NH4Cl was added to the reaction vessel, and the
mixture was allowed to warm to room temperature. EtOAc was
added, and the organic layer was separated and the aqueous
phase was extracted with EtOAc (trice). The combined organic
extracts were washed (water, brine), dried (Na2SO4), and con-
centrated. The residue was purified by column chromatography
on silica gel using CH2Cl2 to CH2Cl2-MeOH (90:10) as the
eluenttoafford 180 mg(29%) of4ias a white solid: [R]23D þ33° (c
0.4, CHC13). 1H NMR (CDCl3) δ 7.54 (d, 2H, J = 8.5 Hz), 7.32
(d, 2H, J = 8.5 Hz), 3.83 (d, 1H, J = 11.3 Hz), 3.40 (d, 1H, J =
11.3 Hz), 3.18 (q, 1H, J = 6.5 Hz), 1.38 (s, 3H), 1.08 (s, 3H), 0.78
(d, 3H, J = 6.5 Hz). 13C NMR (CDCl3) δ 128.5, 128.1, 127.6,
127.2, 101.6, 95.9, 69.5, 53.4, 49.7, 27.3, 22.8, 16.4. LCMS (ESI)
m/z 256.3 [(M þ H)þ, M = C13H18ClNO2].
(C16H24NO5 0.75H2O) C, H, N.
3
(2S,3S)-2-(3-Methoxyphenyl)-3,5,5-trimethylmorpholin-2-ol (4f)
Hemi-D-tartrate. Compound 4f was synthesized by a procedure
similar to that described for (2S,3S)-4a using (R)-1-(3-methoxy-
phenyl)-2-hydroxypropan-1-one (10f, 4.2 g, 0.023 mol), Proton
sponge (5.9 g, 0.028 mol), triflic anhydride (4.2 mL, 257 mmol),
and 2-amino-2-methyl-1-propanol (4.5 g, 0.051 mol) in CH2Cl2
(50 mL). After purification, 4.16 g (71%) of the free base 4f was
isolated and converted to 1.24 g the hemi-D-tartrate salt, which had
91%ee: mp 99-100 °C; [R]20D þ7.9° (c 1.1, CH3OH). 1H NMR
(methanol-d4) δ 7.32 (t, 1H, J = 7.8 Hz), 7.19-7.14 (m, 2H),
6.96-6.92 (m, 1H), 4.33 (s, 1H), 4.18 (d, 1H, J = 12.3 Hz), 3.81 (s,
3H), 3.52 (d, 1H, J = 12.3 Hz), 3.48-3.45 (m, 1H), 1.59 (s, 3H),
1.34 (s, 3H), 1.06 (d, 3H, J = 6.6 Hz). 13C NMR (methanol-d4) δ
178.8, 161.4, 144.1, 130.7, 120.1, 115.4, 113.8, 97.1, 75.2, 67.6, 56.1,
55.2, 24.4, 21.5, 14.5. LCMS (ESI) mlz 252.3 [(M - tartrate)þ,
M=C16H24NO6]. Anal. (C16H24NO6 0.5H2O) C, H, N.
3
A sample of 4i (161 mg, 0.629 mmol) in ether (3 mL) was treated
with a solution of fumaric acid (73.0 mg, 0.629 mmol) in MeOH
(1.2 mL). The mixture was stirred at room temperature overnight.
Filtration and washing of the filter cake with ether, followed
by recrystallization from MeOH-ether gave 123 mg (53%) of
(2S,3S)-2-(3-Nitrophenyl)-3,5,5-trimethylmorpholin-2-ol (4g)
Hemi-D-tartrate. Compound 4g was synthesized by a procedure
similar to that described for (2S,3S)-4a using (R)-1-(3-nitrophenyl)-
2-hydroxypropan-1-one (10g, 4.0 g, 0.021 mol), Proton sponge
(5.2 g, 0.0246 mol), triflic anhydride (3.7 mL, 0.023 mol), and
2-amino-2-methyl-1-propanol (4.0 g, 0.045 mol) in acetonitrile
(45 mL). After purification, 1.0 g (18%) of the free base 4g was
isolated and converted to the hemi-D-tartrate salt, which had 94%
4i 0.5fumarate as a white solid; mp 187-190 °C; [R]22 þ22°
D
3
(c 0.75, MeOH). 1H NMR (methanol-d4) δ 7.58 (d, 2H, J = 8.6
Hz), 7.41 (d, 1H, J = 8.6 Hz), 6.67 (s, 1H), 4.11 (d, 1H, J = 12.0
Hz), 3.51 (d, 1H, J = 12.0 Hz), 3.38-3.36 (m, 1H), 1.53 (s, 3H),
1.28 (s, 3H), 1.00 (d, 3H, J = 6.6 Hz). 13C NMR (methanol-d4) δ
172.9, 141.1, 136.7, 135.8, 129.3, 96.6, 67.4, 54.68, 54.37, 24.1, 21.2,
14.1. LCMS (ESI) m/z 256.6 [(M - fumaric)þ, M = C13H18-
ClNO2 0.5C4H4O4]. Anal. (C15H20ClNO4 0.5H2O) C, H, N.
ee: mp 192-193 °C; [R]20 þ6.5° (c 1.0, CH3OH). 1H NMR
D
(methanol-d4)
δ 8.47-8.45 (m, 1H), 8.31-8.26 (m, 1H),
8.05-8.01 (m, 1H), 7.73-7.66 (m, 1H), 4.34 (s, 1H), 4.18 (d, 1H,
J = 12.1 Hz), 3.59 (d, 1H, J = 6.6 Hz), 3.50 (q, 1H, J = 6.6 Hz),
1.60 (s, 3H), 1.35 (s, 3H), 1.07 (d, 3H, J = 6.6 Hz). 13C NMR
(methanol-d4) δ 178.3, 149.9, 145.3, 134.4, 131.0, 125.0, 123.0,
96.8, 75.0, 68.1, 54.8, 24.7, 21.7, 14.6. LCMS (ESI) mlz 267.3
3
3
(2S,3S)-3,5,5-Trimethyl-2-(4-methylphenyl)morpholin-2-ol (4j)
Hemi-Fumarate. A solution of morpholin-2-one 15 (270 mg,
1.88 mmol) in anhydrous THF (1.9 mL) was cooled to -78 °C
and treated with p-tolylmagnesium bromide (1.2 equiv, 2.26 mmol,
2.26 mL, 1 M solution in THF) under an N2 atmosphere. After
stirring the reaction mixture at -78 °C for 1.5 h, saturated aqueous
solution of NH4Cl (30 mL) was added to the reaction vessel, and
the mixture was allowed to warm to room temperature. Ether
(30 mL) was added to the reaction flask, and the organic layer was
separated. The aqueous phase was extracted with ether (twice). The
combined organic extracts were washed (water, brine), dried
(Na2SO4), and concentrated. The residue was purified by column
chromatography on silica gel using CH2Cl2 to CH2Cl2-MeOH
(90:10) as the eluent to give 271 mg (41%) of 4j as a yellow foam:
[R]22D þ21.2° (c 0.9, CHCl3). 1H NMR (CDCl3) δ7.47 (d, 2H, J =
8.2 Hz), 7.16 (d, 2H, J = 8.0 Hz), 3.85 (d, 1H, J = 11.3 Hz), 3.40
(d, 1H, J = 11.2 Hz), 3.20-3.07 (m, 1H), 2.35 (s, 3H), 1.39 (s, 3H),
1.08 (s, 3H), 0.81 (d, 3H, J = 6.5 Hz). 13C NMR (CDCl3) δ 129.6,
128.60, 128.49, 125.9, 103.2, 96.2, 69.4, 53.9, 53.4, 49.6, 27.3, 22.8,
16.5. LCMS (ESI) m/z 236.3 [(M þ H)þ, M = C14H21NO2].
A sample of 4j (240 mg, 0.683 mmol) in ether (3 mL) was treated
with a solution of fumaric acid (87.0 mg, 0.751 mmol) in MeOH
(2 mL) and stirred at room temperature overnight. Ether was
added to the reaction mixture. The solid was recrystallized from
[(M-tartrate)þ, M=C15H21N2O7]. Anal. (C15H21N2O7 0.25H2O)
3
C, H, N.
(2S,3S)-2-(4-Fluorophenyl)-3,5,5-trimethylmorpholin-2-ol (4h)
Hemi-Fumarate. A solution of 15 (166 mg, 1.16 mmol) in dry THF
(1.2 mL, 1M) under an N2 atmosphere was cooled to -78 °C and
treated with 4-fluorophenylmagnesium bromide (1.3 equiv,
1.5 mmol, 1.9 mL, 0.8 M solution in THF). The reaction mixture
was stirred at -78 °C for 3 h. A saturated aqueous solution of
NH4Cl was added to the reaction vessel, and the mixture was
allowed to warm to room temperature. EtOAc (5 mL) was added
to the reaction vessel, and the organic layer was separated. The
aqueous phase was extracted with EtOAc (three times). The
combined organic extracts were washed (water, brine), dried
(Na2SO4), and concentrated. The residue waspurified by column
chromatography on silica gel using CH2Cl2 to CH2Cl2-MeOH
(90:10) as the eluent to afford 80 mg of 4h as a white solid: [R]22
D
þ31.2° (c 0.5, CHC13). 1H NMR (CDCl3) δ 7.60-7.55 (m, 2H),
δ 7.08-7.00 (m, 2H), 3.83 (d, 1H, J = 11.3 Hz), 3.40 (d, 1H, J =
11.3 Hz), 3.17 (q, 1H, J = 6.4 Hz), 1.38 (s, 3H), 1.08 (s, 3H), 0.78
(d, 3H, J = 6.4 Hz). 13C NMR (CDCl3) δ 128.0, 127.9, 114.87,
114.58, 103.2, 98.4, 96.8, 69.5, 53.5, 27.3, 22.8, 16.4. LCMS (ESI)
m/z 240.0 [(M þ H)þ, M = C13H18FNO2].
A sample of 4h (56.0 mg, 0.234 mmol) in ether (2 mL) was
treated with a solution of fumaric acid (30.0 mg, 0.258 mmol) in
MeOH (0.6 mL). The mixture was stirred at room temperature
overnight. Filtration and washing of the filter cake with ether,
followed by recrystallization of the solid from MeOH-ether
MeOH-ether to give 140 mg (67%) of 4j 0.5fumarate as a white
3
1
solid: mp 178-182 °C; [R]22 þ19° (c 0.6, MeOH). H NMR
D
(methanol-d4) δ 7.47 (d, 2H, J = 8.2 Hz), 7.21 (d, 2H, J = 8.0 Hz),
6.65 (s, 1H), 4.18 (d, 1H, J = 12.2 Hz), 3.51 (d, 1H, J = 12.2 Hz),
3.46 (q, 1H, J=6.6Hz), 2.35(s, 3H), 1.58(s, 3H), 1.34(s, 3H), 1.04
(d, 1H, J = 6.6 Hz). 13C NMR (methanol-d4) δ 139.9, 139.2, 136.9,
129.8, 127.4, 96.8, 67.2, 54.9, 54.6, 24.1, 21.1, 14.1. LCMS (ESI)
gave 45 mg (54%) of 4h 0.5fumarate: mp 178-182 °C; [R]22D þ29°
3
1
(c 0.6, MeOH). H NMR (methanol-d4) δ 7.65-7.59 (m, 2H),
7.15-7.08 (m, 2H), 6.66 (s, 1H), 4.15 (d, 1H, J = 12.2 Hz), 3.52 (d,
1H, J= 12.2 Hz), 3.41-3.33 (m, 1H), 1.56 (s, 3H), 1.32 (s, 3H), 1.03
(d, 3H, J = 6.6 Hz). 13C NMR (methanol-d4) δ 136.7, 129.75,
129.64, 115.92, 115.63, 67.5, 54.8, 54.1, 24.3, 21.3, 14.3. LCMS
m/z 236.2 [(M - fumaric)þ C14H21NO2 0.5C4H4O4]. Anal.
3
(C16H23NO4 0.25H2O) C, H, N.
(2S,3S)-2-(4-Methoxyphenyl)-3,5,5-trimethylmorpholin-2-ol (4k)
Hydrochloride. A solution of 15 (434 mg, 3.03 mmol) in anhydrous
3

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4741
THF (3 mL, 1M) was cooled to -78 °C and treated with 4-methoxy-
phenylmagnesium bromide (1.3 equiv, 3.93 mmol, 3.9 mL of 1 M
solution in THF). The reaction mixture was stirred at -78 °C and
under an N2 atmosphere for 1.5 h. A saturated aqueous solution of
NH4Cl (30 mL) was added to the reaction vessel, and the mixture
was allowed to warm to room temperature. The mixture was
extracted with ether (three times). The combined ether extracts were
washed (water, brine), dried (Na2SO4), and concentrated. The
residue was purified by column chromatography on silica gel using
CH2Cl2 to CH2Cl2-MeOH-NH4OH (90:9:1) as the eluent to give
380 mg of a yellow solid.
to CH2Cl2-MeOH-NH4OH (90:9:1) as the eluent gave 190 mg
(25%) of 4m as a yellow solid: [R]22D þ18.2° (c 1.0, CHCl3). 1H
NMR (CDCl3) δ 7.45-7.39 (m, 1H), 7.34-7.32 (m, 1H),
7.18-7.10 (m, 1H), 3.82 (d, 1H, J = 11.3 Hz), 3.40 (d, 1H, J =
11.3 Hz), 3.18 (q, 1H, J = 6.5 Hz), 1.38 (s, 3H), 1.08 (s, 3H), 0.78
(d, 3H, J = 6.5 Hz). 13C NMR (CDCl3) δ 122.40, 122.31 116.74,
116.51, 115.8, 115.6, 96.5, 69.6, 53.4, 49.8, 27.3, 22.8, 16.5. LCMS
(ESI) m/z 258.6 [(M þ H)þ, M = C13H17F2NO2].
A sample of 4m (180 mg, 0.699 mmol) in ether (3 mL) was
treated with a solution of fumaric acid (80.0 mg, 0.689 mmol) in
MeOH (2.5 mL). The mixture was stirred at room temperature
overnight. Ether was added to the reaction mixture. The suspen-
sion was sonicated, centrifuged, and decanted to afford a solid
pellet that was recrystallization from MeOH-ether. The suspen-
sion was sonicated, centrifuged, and decanted three times to
The sample (379 mg, 0.783 mmol) in ether (5 mL) was treated
with HCl (1.00 mmol, 0.250 mL, 4 M solution in dioxane). The
mixture was stirred at room temperature overnight. Ether was
added to the reaction mixture. The suspension was sonicated,
centrifuged, and decanted three times to afford a solid pellet.
The solid material was recrystallized from MeOH-ether to give
afford 119 mg (53%) of 4m 0.5fumarate as a white solid: mp
3
187-189 °C; [R]22D þ29.6° (c 0.5, MeOH). 1H NMR (methanol-
d4) δ 7.52-7.41 (m, 2H), 7.38-7.22 (m, 1H), 6.65 (s, 1H), 4.34 (s,
1H), 4.15 (d, 1H, J = 12.2 Hz), 3.54 (d, 1H, J = 12.2 Hz), 3.46 (q,
1H, J = 6.6 Hz), 1.57 (s, 3H), 1.33 (s, 3H), 1.06 (d, 3H, J = 6.6
Hz). 13C NMR (methanol-d4) δ 136.7, 124.4, 118.1, 117.8, 117.1,
116.9, 98.2, 67.5, 54.58, 54.36, 24.1, 21.2, 14.1. LCMS (ESI) m/z
215 mg (95%) of 4k HCl as a pale-yellow solid: mp 170-172 °C;
3
[R]20D þ19.6° (c 1.0, MeOH). 1H NMR (methanol-d4) δ 7.52 (d,
2H, J = 8.9 Hz), 6.95 (d, 2H, J = 8.9 Hz), 4.22 (d, 1H, J = 12.4
Hz), 3.81 (s, 3H), 3.58-3.45 (m, 2H), 1.62 (s, 3H), 1.39 (s, 3H),
1.10 (d, 3H, J = 6.6 Hz). 13C NMR (methanol-d4) δ 161.8, 133.5,
132.8, 128.8, 114.6, 96.7, 66.6, 55.80, 55.7, 55.2, 23.4, 20.8, 13.7.
258.6, [(M - fumaric)þ M = C13H17F2NO2 0.5C4H4O4]. Anal.
3
LCMS (ESI) m/z 286.4 (M - H)þ, M = C14H21NO3 HCl].
(C15H19F2NO4 0.5H2O) C, H, N.
3
3
Anal. (C14H22ClNO3) C, H, N.
2-(3,4-Dichlorophenyl)-3,5,5-trimethylmorpholin-2-ol [(()-4n]
Hemi-Fumarate. To a solution of 30,40-dichloropropiophenone
(8l, 5.02 g, 0.247 mol) in CH2Cl2 (100 mL) was added 10 drops of
bromine. After stirring at room temperature under nitrogen
for several min, the characteristic red color of bromine disap-
peared, indicating initiation of the reaction. The remainder of
the bromine (1.27 mL, 24.7 mmol) was added dropwise, and the
reaction solution was allowed to stir at room temperature under
nitrogen atmosphere for 1.75 h. Analysis by TLC (silica, 2:1
hexane:CH2Cl2) indicated consumption of starting material.
The reaction solution was quenched and brought to a pH of 9
with a saturated aqueous solution of NaHCO3 and concentrated
NH4OH. The solution was extracted with CH2Cl2, dried
(Na2SO4), filtered, concentrated, and dried under vacuum to
give 7.14 g (100%) of 2-bromo-(30,40-dichlorophenyl)propan-
1-one as a white solid. Characterization data is similar with data
reported in the literature.31
(2S,3S)-2-Biphenyl-4-yl-3,5,5-trimethylmorpholin-2-ol (4l) Hemi-
Fumarate. A solution of 15 (394 mg, 2.75 mmol) in anhydrous
THF (2.75 mL, 1M) was cooled to -78 °C and treated with
4-biphenylmagnesium bromide (1.2 equiv, 3.30 mmol, 6.60 mL,
0.5 M solution in THF) under an N2 atmosphere. After stirring at
-78 °C for 1.5 h, the reaction mixture was treated with saturated
aqueous solution of NH4Cl (40 mL) and EtOAc was added
(30 mL). The organic layer was separated. The aqueous phase
was extracted with EtOAc trice. The combined organic extracts
were washed (water, brine), dried (Na2SO4), and concentrated.
Purification of the residue by column chromatography on silica
gel using CH2Cl2 to CH2Cl2-MeOH (90:10) as the eluent
afforded 300 mg (37%) of 4l as a white solid: [R]23 þ15.2° (c
D
1
0.3, CHC13). H NMR (CDCl3) δ 7.70-7.33 (m, 9H), 3.88 (d,
1H, J = 11.3 Hz), 3.44 (d, 1H, J = 11.3 Hz), 3.29-3.25 (m, 1H),
1.42 (s, 3H), 1.10 (s, 3H), 0.86 (d, 3H, J = 6.5 Hz). 13C NMR
(CDCl3) δ 140.9, 129.0, 128.97, 128.74, 128.4, 127.54, 127.30,
127.13, 126.7, 126.5, 96.2, 69.5, 53.4, 51.4, 49.6, 27.4, 22.8, 16.6.
LCMS (ESI) m/z 298.4, 280.3 [(M þ H)þ, M = C19H23NO2].
A sample of 4l (379 mg, 1.27 mmol) in CH2Cl2 (4 mL) was
treated with a solution of fumaric acid (148 mg, 1.27 mmol) in
MeOH (4 mL). The mixture was stirred at room temperature
overnight. Filtration and washing of the filter cake with ether,
followed by recrystallization from MeOH-ether gave 130 mg
2-Bromo-(30,40-dichlorophenyl)propan-1-one (6.97 g, 0.025 mol)
in a minimal amount of CH2Cl2 was transferred to a sealable
reaction tube. Most of the CH2Cl2 was removed via positive
nitrogen flow. 2-Amino-2-methyl-1-propanol (23.6 mL, 247 mmol)
was added in one portion, and the tube was sealed and placed in an
oil bath heated to 75 °C. After stirring at 75 °C overnight, analysis
by TLC (silica, 9:1:20 ether-Et3N-hexane) showed only a trace
amount of starting material remaining and the reaction was allowed
to cool to room temperature. The reaction mixture was quenched
and brought to a pH of 10 with a saturated aqueous solution of
NaHCO3, and the product was extracted with CH2Cl2. The organic
layer was separated, dried (Na2SO4), filtered, concentrated, and
dried under vacuum to give 11.29 g of a yellow oil. The residue was
purified by column chromatography on silica gel using ether-
Et3N-hexane (9:1:50) as eluent to give 3.50 g (49%) of 4n as a white
solid. 1H NMR (250 MHz, DMSO-d6) δ 7.70-7.67 (m, 1H),
7.47-7.39 (m, 2H), 3.81 (d, 1H), 3.40 (d, 1H), 3.22-3.14 (m,
1H), 1.38 (s, 3H), 1.08 (s, 3H), 0.78 (d, 3H).
(25%) of 4l 0.5fumarate as a white solid: mp 197-202 °C; [R]22
D
3
þ18° (c 1.6, MeOH). 1H NMR (methanol-d4) δ 7.70-7.63 (m,
6H), 7.49-7.35 (m, 3H), 6.69 (s, 1H), 4.20 (d, 1H, J = 12.1 Hz),
3.55 (d, 1H, J = 12.1 Hz), 3.47 (q, 1H, J = 6.7 Hz), 1.60 (s, 3H),
1.34 (s, 3H), 1.08 (d, 3H, J = 6.6 Hz). 13C NMR (methanol-d4) δ
142.9, 141.8, 141.4, 136.7, 129.9, 128.6, 128.04, 128.02, 127.7,
96.9, 67.5, 54.8, 54.1, 30.7, 24.4, 21.3, 14.4. LCMS (ESI) m/z
298.6, 280.3 [(M - fumaric)þ, M = C19H23NO2 0.5C4H4O4].
3
Anal. (C21H25NO4 0.75H2O) C, H, N.
3
(2S,3S)-2-(3,4-Difluorophenyl)-3,5,5-trimethylmorpholin-2-ol
(4m) Hemi-Fumarate. A solution of morpholin-2-one 15 (410 mg,
2.86 mmol) in anhydrous THF (2.9 mL, 1M) was cooled to
-78 °C and treated with 3,4-difluorophenylmagnesium bromide
(1.2 equiv, 3.43 mmol, 6.90 mL, 0.5 M solution in THF) under an
N2 atmosphere. After stirring the reaction mixture at -78 °C
under an inert atmosphere for 1.5 h, saturated aqueous solution
of NH4Cl was added to the reaction vessel. The reaction mixture
was allowed to warm to room temperature and extracted with
ether (three times). The combined organic extracts were washed
(water, brine), dried (Na2SO4), and concentrated. Purification of
the residuebycolumn chromatography on silicagel usingCH2Cl2
A solution of (()-4n (3.34 g, 0.012 mol) in methanol was
treated with fumaric acid (1.34 g, 0.012 mol). The mixture was
allowed to stir for 15 min and a white solid precipitated out of
solution, which was collected by vacuum filtration to afford 2.23
g (53%) of 4n 0.5fumarate as a white solid: mp 188-189 °C. 1H
3
NMR (250 MHz, DMSO-d6) δ 7.68-7.62 (m, 2H), 7.52-7.48
(m, 1H), 3.77 (d, 1H), 3.36 (d, 1H), 3.13-3.07 (q, 1H), 1.33 (s,
3H), 1.07 (s, 3H), 0.76 (d, 3H). Anal. (C15H19Cl2NO4) C, H, N.
(2S,3S)-2-(3,5-Difluorophenyl)-3,5,5-trimethylmorpholin-2-ol
(4o) Hydrochloride. A solution of 15 (448 mg, 3.13 mmol) in
anhydrous THF (3 mL, 1M) was cooled to -78 °C and treated

4742 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
with 3,5-dipfluorophenylmagnesium bromide (1.2 equiv, 3.75
mmol, 7.5 mL, 0.5 M solution in THF). The reaction mixture
was stirred at -78 °C and under an N2 atmosphere for 1.5 h.
Saturated aqueous solution of NH4Cl was added to the reaction
vessel, and the mixture was allowed to warm to room tempera-
ture. The reaction mixture was extracted with ether (three
times). The combined ether extracts were washed (water, brine),
dried (Na2SO4), and concentrated. Purification of the residue by
column chromatography on silica gel using CH2Cl2 to CH2Cl2-
MeOH-NH4OH (90:9:1) as the eluent gave 105 mg (13%) of 4o
δ 136.5, 132.3, 131.9, 130.5, 130.2, 128.1, 127.4, 127.0, 126.1, 99.0,
75.1, 68.1, 67.3, 53.9, 24.8, 22.9, 15.8, 15.2. LCMS (ESI) m/z 272.3
[(M- tartrate)þ, M=C19H24NO5]. Anal. (C19H24NO5 0.75H2O)
C, H, N.
3
(2S,3S)-2-(Naphthalen-2-yl)-3,5,5-trimethylmorpholin-2-ol (4r)
Hemi-D-tartrate. Compound 4r was synthesized by a procedure
similar to that described for (2S,3S)-4a using (R)-2-hydroxy-
1-(naphthalen-2-yl)propan-1-one (10i, 2.5 g, 0.013 mol), Proton
sponge (3.24 g, 0.0151 mol), triflic anhydride (2.4 mL, 137 mmol),
and 2-amino-2-methyl-1-propanol (2.4 g, 0.027 mol) in aceto-
nitrile (40 mL). After purification, 2.2 g (65%) of the free base 4r
was isolated and converted to the hemi-D-tartrate salt: mp 179-
180 °C; [R]20D -1.5° (c 0.55, CH3OH). 1H NMR (methanol-d4) δ
8.13-8.12 (m, 1H), 7.95-7.88 (m, 3H), 7.75-7.71 (m, 1H),
7.57-7.50 (m, 2H), 4.33 (s, 1H), 4.25 (d, 1H, J = 11.9 Hz),
3.66-3.57 (m, 2H), 1.65 (s, 3H), 1.37 (s, 3H), 1.09 (d, 3H, J = 6.6
Hz). 13C NMR (methanol-d4) δ 178.7, 139.9, 135.3, 134.6, 129.8,
129.33, 129.00, 128.0, 127.8, 127.4, 125.4, 97.4, 75.2, 67.7, 55.1,
24.5, 21.6, 14.6. LCMS (ESI) m/z 272.5 [(M - tartrate)þ, M =
C19H24NO5]. Anal. (C19H24NO5) C, H, N.
as a yellow solid: [R]22 þ19.5° (c 0.8, CHCl3). 1H NMR
D
(CDCl3) δ 7.16-7.10 (m, 2H), 6.82-6.67 (m, 1H), 3.81 (d, 1H,
J = 12.0 Hz), 3.40 (d, 1H, J = 12.0 Hz), 3.18 (q, 1H, J = 6.0 Hz),
1.39 (s, 3H), 1.12 (s, 3H), 0.83 (d, 3H, J = 6.5 Hz). 13C NMR
(CDCl3) δ 109.63, 109.40, 103.9, 103.5, 103.2, 95.8, 69.2, 53.3,
50.1, 49.6, 26.9, 22.5, 15.2. LCMS (ESI) m/z 258.8 [(M þ H)þ,
M = C13H17F2NO2].
A sample of 4o (71.0 mg, 0.275 mmol) in CH2Cl2 (1.5 mL) was
treated with HCl (0.386 mmol, 0.100 mL solution 4 M in dioxane).
The mixture was stirred at room temperature overnight, and ether
was added to the reaction mixture. The suspension was sonicated,
centrifuged, and decanted to afford a solid pellet; this procedure
was repeated three times. The solid material was recrystallized
from methanol-ether. The suspension was sonicated, centrifuged,
(2S,3S)-2-(3-Chlorophenyl)-3-ethyl-5,5-dimethylmorpholin-2-ol
(4s) Hemi-D-tartrate. Compound 4s was synthesized by a proce-
dure similar to that described for (2S,3S)-4a using (R)-1-(3-
chlorophenyl)-2-hydroxybutan-1-one (10j, 1.13 g, 0.0568 mol),
Proton sponge (1.43 g, 0.0667 mol), triflic anhydride (1.0 g, 0.063
mol), and 2-amino-2-methyl-1-propanol (1.09 g, 0.0122 mol) in
CH2Cl2 (13 mL). After purification, the free base 4r was converted
to 0.230 g of its D-tartrate salt: mp 160-161 °C;[R]20D þ5.6° (c 0.8,
decanted, and dried to give 60 mg (74%) of 4o HCl as a pale-
3
1
yellow solid: mp 199-204 °C; [R]22 þ21.5° (c 1.0, MeOH). H
D
NMR (methanol-d4) δ 7.23-7.18 (m, 2H), 7.03-6.92 (m, 1H),
4.19 (d, 1H, J = 12.0 Hz), 3.65-3.54 (m, 2H), 1.62 (s, 3H), 1.39 (s,
3H), 1.12 (d, 3H, J = 6.0 Hz). 13C NMR (methanol-d4) δ 166.1,
162.8, 111.1, 110.9, 105.6, 105,2, 104.9, 66.9, 55.6, 54.5, 23.5, 20.9,
13.7. LCMS (ESI) m/z 258.5 [(M - HCl)þ, M = C13H17F2-
1
CH3OH). H NMR (methanol-d4) δ 7.49-7.46 (m, 1H), 7.41-
7.35 (m, 3H), 4.37-4.34 (m, 1H), 4.30-4.24 (m, 1H), 3.81-3.62
(m, 2H), 1.57 (s, 3H), 1.47-1.36 (m, 2H), 1.33 (s, 3H), 0.81-0.71
(m, 3H). 13C NMR (methanol-d4) δ 178.5, 145.2, 135.6, 131.2,
130.3, 128.2, 126.5, 97.1, 75.0, 67.8, 61.1, 55.0, 24.3, 23.1, 21.6,
11.2. LCMS (ESI) m/z 270.4 [(M - tartrate)þ, M = C16H23-
NO2 HCl]. Anal. (C13H18ClF2NO2) C, H, N.
3
2-(3,5-Dichlorophenyl)-3,5,5-trimethylmorpholin-2-ol [(()-4p]
Hemi-Fumarate. To a stirred solution of 3,5-dichloropropio-
phenone (8m, 3.81 g, 0.0188 mol) in CH2Cl2 (28 mL) was added
bromine (0.986 mL, 19.1 mmol) dropwise. The bromine was
immediately consumed upon addition of each drop until the end
of addition when it had a consistent brownish color, indicative
of excess bromine. The reaction was immediately quenched with
saturated aqueous NaHCO3, extracted three times with CH2Cl2.
The organic layer was separated, combined, and concentrated
under reduced pressure without drying to afford a yellow oil.
The oil was dissolved in anhydrous diethyl ether (50 mL) and
2-amino-2-methyl-1-propanol (6.7 g, 0.075 mol) was added. The
reaction mixture was stirred overnight and quenched with
saturated aqueous NaHCO3. The aqueous layer was extracted
three times with CH2Cl2, and the organic layers were separated,
dried, and concentrated to afford another yellow oil. Purification by
column chromatography on silica gel afforded 2.51 g (46%) of the
title compound. The hemifumarate salt was prepared by dissolving
the free base in methanol and adding 1.0 g of fumaric acid to form
the title compound: mp 188-190 °C. 1H NMR (DMSO-d6)
δ7.58-7.57 (m, 1H), 7.47-7.46 (m, 2H), 6.67 (s, 0.5H), 4.09 (d, 1H,
J = 11.9 Hz), 3.53 (d, 1H, J = 12.0 Hz), 3.41-3.37 (m, 1H), 1.53
(s, 3H), 1.28 (s, 3H), 1.02 (d, 3H, 6.6 Hz). 13C NMR (DMSO-d6)
δ 134.9, 133.6, 127.7, 125.2, 94.7, 67.2, 52.5, 50.1, 25.0, 21.6, 14.6.
ClNO5). Anal. (C16H23ClNO5 0.25H2O) C, H, N.
3
(2S,3S)-2-(3-Chlorophenyl)-5,5-dimethyl-3-propyl-morpholin-
2-ol (4t) Hemi-D-tartrate. Compound 4t was synthesized by a
procedure similar to that described for (2S,3S)-4a using (R)-1-(3-
chlorophenyl)-2-hydroxypent-1-one (10k, 1.5 g, 0.0704 mol), Pro-
ton sponge (1.8 g, 0.0840 mol), triflic anhydride (1.29 g, 0.081 mol),
and 2-amino-2-methyl-1-propanol (1.42 g, 0.0159 mol) in CH2Cl2
(7 mL). After purification, 790 mg (40%) of the free base 4t was
isolated and converted to the hemi-D-tartrate salt, which had 99%
1
ee: mp 151-152 °C, [R]20 -10.1° (c 0.77, CH3OH). H NMR
D
(methanol-d4) δ 7.50-7.47 (m, 1H), 7.43-7.36 (m, 3H), 4.38 (s,
1H), 4.32 (d, 1H, J = 10.0 Hz), 3.81-3.66 (m, 1H), 1.58 (s, 3H),
1.49-1.38 (m, 2H), 1.35 (s, 3H), 1.34-1.27 (m, 1H), 1.02-0.92 (m,
1H), 0.77 (t, 3H, J = 7.0 Hz). 13C NMR (methanol-d4) δ 176.1,
142.9, 133.3, 128.9, 128.0, 125.9, 124.2, 94.8, 72.7, 65.4, 57.0, 52.7,
29.9, 22.0, 19.3, 18.2, 12.1. Anal. (C17H25ClNO5) C, H, N.
(2S,3S)-2-(3-Chlorophenyl)-3,4,5,5-tetramethylmorpholin-2-ol (4u)
Di-p-tolyl-L-tartrate. A sample of (2S,3S)-2-(3-Chlorophenyl)-
3,5,5-trimethylmorpholin-2-ol (4i, 107 mg, 0.42 mmol) in DMF
(2.0 mL) was treated with K2CO3 (174 mg, 1.26 mmol). After
stirring the reaction mixture at room temperature under an inert
atmosphere for 1.5 h, CH3I (19 μL, 0.30 mmol) was added to the
reaction flask and the reaction mixture was stirred at 70 °C for 24 h.
The reaction mixture was cooled to 0 °C water added, followed by
extraction with ether (thee times). The combined organic extracts
were washed (water, brine), dried (Na2SO4), and concentrated to a
pale-yellow oil. Purification of the residue by column chromato-
graphy gave a 72 mg (63%) of 4u as a white solid. The compound
4u was converted to the corresponding di-p-tolyl-L-tartrate salt: mp
128-129 °C; [R]20D -50.0° (c 0.83, CH3OH). 1H NMR (DMSO-
d6) δ 7.85 (d, 4H, J = 7.8 Hz), 7.50-7.40 (m, 4H), 7.32 (d, 4H, J =
7.8 Hz), 5.6 (s, 2H), 4.07 (d, 1H, J = 12.4 Hz), 3.50-3.40 (m, 1H),
2.56 (s, 3H), 2.36 (s, 6H), 1.40 (s, 3H), 1.21 (s, 3H), 0.90 (d, 3H, J =
6.2 Hz). 13C NMR (DMSO-d6) δ 168.2, 164.9, 143.9, 132.7, 129.9,
129.3 (d), 128.6, 126.67, 126.31, 125.3, 96.4, 72.2, 66.2, 60.4, 59.2,
Anal. (C15H19Cl2NO4 0.25H2O) C, H, N.
3
(2S,3S)-2-(Naphthalen-1-yl)-3,5,5-trimethylmorpholin-2-ol (4q)
Hemi-D-tartrate. Compound 4q was synthesized by a procedure
similar to that described for (2S,3S)-4a employing (R)-2-hydroxy-
1-(naphthalen-1-yl)propan-1-one (10h, 1.15 g, 0.058 mol), Proton
sponge (1.5 g, 0.070 mol), triflic anhydride (1.1 mL, 64 mmol),
and 2-amino-2-methyl-1-propanol (1.1 g, 0.012 mol) in CH3CN
(45 mL). After purification, 1.35 g (86%) of the free base 4q was
isolated and converted to the D-tartrate salt, which was recrystal-
lized from H2O-MeOH-Et2O solvent system: mp 115-116 °C;
[R]20D -9.5° (c 0.74, CH3OH). 1H NMR (methanol-d4) δ 8.89 (d,
1H, J = 8.4 Hz), 7.97-7.89 (m, 3H), 7.54-7.50 (m, 3H), 4.33 (s,
1H), 3.70 (d, 1H, J = 12.2 Hz), 3.48 (q, 1H, J = 7.0 Hz), 1.77 (s,
3H), 1.41 (s, 3H), 0.96 (d, 3H, J=6.6Hz).13C NMR (methanol-d4)
33.3, 21.1, 15.9, 11.5. Anal. (C34H38ClNO10 2H2O) C, H, N.
3

Article
(2S,3S)-2-(4-Chlorophenyl)-3,4,5,5-tetramethylmorpholin-2-ol (4v)
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4743
7.34-7.26 (m, 1H), 6.00 (s, 1H), 3.97-3.91 (m, 1H), 3.47-3.37
Fumarate. A sample of (4i) (144 mg, 0.563 mmol) in anhydrous
THF (1.9 mL) was treated with K2CO3 (4 folds, 311 mg,
2.25 mmol). After stirring the reaction mixture at room tempera-
ture under inert atmosphere for 1 h, CH3I (1.3 equiv, 46.0 μL,
0.731 mmol) was added to the reaction flask. The reaction mixture
was stirred at room temperature for 24 h, cooled to 0 °C, and water
added, followed by extraction with ether (thee times). The com-
bined organic extracts were washed (water, brine), dried (Na2SO4),
and concentrated to a pale-yellow oil. Purification of the residue by
column chromatography on silica gel and CH2Cl2-MeOH-
NH4OH (90:9:1) as the eluent gave 93.0 mg (61%) of a white
solid: mp = 75-78 °C; [R]22D þ28.0° (c 1.0, CHCl3). 1H NMR δ
7.58-7.53 (m, 2H), 7.34-7.31 (m, 2H), 4.52-4.49 (br, 1H), 3.90
(d, 1H, J = 11.6 Hz), 3.32 (d, 1H, J = 11.6 Hz), 2.85 (q, 1H, J =
6.5 Hz), 2.20 (s, 3H), 1.19 (s, 3H), 1.07 (s, 3H), 0.76 (d, 3H, J = 6.5
Hz). 13C NMR δ 133.9, 128.59, 128.36, 128.05, 127.7, 97.5, 70.4,
59.3, 53.8, 32.3, 25.5, 14.4, 13.1. MS (ESI) m/z 270.4 [(M þ H)þ
M = C14H20ClNO2].
(m, 2H), 1.42 (s, 3H), 1.10 (s, 3H), 0.74 (d, 3H, J = 6.5 Hz). 13
C
NMR (CDCl3) δ 158.6, 147.2, 137.5, 123.6, 120.5, 94.2, 70.0, 52.1,
48.9, 27.1, 22.9, 16.2. LCMS (ESI) m/z 223.4 [(M - tartrate)þ,
M = C16H24N2O8]. Anal. (C16H24N2O8 0.5H2O) C, H, N.
3
1-Naphthalen-1-ylpropan-1-one (8q). A sample of 1-cyano-
naphthylene (7a, 5.0 g, 0.0327 mol) in dry diethyl ether (200 mL)
was treated with ethyl magnesium bromide (49.2 mol, 16.4 mL
3 M solution in ether) under nitrogen atmosphere at room
temperature. The solution was stirred overnight at 25 °C and
then cooled to 0 °C. The reaction was quenched slowly with 1 N
aqueous HCl and allowed to warm to room temperature over 1 h
with stirring. The aqueous layer was extracted with ether. The
combined organic layers were washed with aqueous NaHCO3,
water, and brine, and dried (Na2SO4). The organic layers were
concentrated, and the resulting yellow oil was purified by
column chromatography on silica gel using hexane-EtOAc
(20:1) to (10:1) as the eluent afforded 3.55 g (60%) of the title
product. 1H NMR (CDCl3) δ 8.59-8.52 (m, 1H), 8.00-7.94 (m,
1H), 7.90-7.81 (m, 2H), 7.62-7.45 (m, 3H), 3.08 (q, 2H, J = 7.3
Hz), 1.29 (t, 3H, J = 7.3 Hz). C13H12O.
A sample of 4v (90 mg, 0.33 mmol) in ether (1.5 mL) was treated
with a solution of fumaric acid (38 mg, 0.33 mmol) in MeOH
(1 mL). The mixture was stirred at room temperature overnight.
Ether was added to the reaction mixture. The suspension was
sonicated, centrifuged, and decanted to afford a solid pellet; this
procedure was repeated three times. Recrystallization from
MeOH-ether afforded the title product as white solid 101 mg
1-Naphthalen-2-ylpropan-1-one (8r). Compound 8r was synt-
hesized by a procedure similar to that described for a procedure
similar to 8q using identical amounts, except commercially
available 2-cyanonaphthylene (7b) was used in place of 1-cyano-
naphthylene. Purification by column chromatography on silica
gel using hexane-EtOAc (20:1) to (10:1) afforded 4 g (66%)
(77%): mp =167-169 °C; [R]22 þ44.3° (c 1.0, MeOH). 1H
D
1
NMR (methanol-d4) δ 7.60 (d, 2H, J = 8.6 Hz), 7.43 (d, 2H, J =
8.6 Hz), 6.68 (s, 2H), 4.35 (d, 1H, J = 12.8 Hz), 3.64-3.54 (m,
2H), 2.79 (s, 3H), 1.60 (s, 3H), 1.41 (s, 3H), 1.14 (d, 3H, J = 6.5
Hz). 13C NMR (methanol-d4) δ 171.4, 140.9, 136.3, 129.4, 98.1,
67.6, 63.4, 62.5, 34.3, 21.8, 20.8, 16.9, 12.3. MS (ESI) m/z 270.4
of the title product as a white solid. H NMR (CDCl3) δ 8.48
(s, 1H), 8.05 (d, 1H, J=7.5 Hz), 8.00-7.94 (m, 1H), 7.93-7.85
(m, 2H), 7.64-7.51 (m, 2H). 3.15 (q, 2H, J = 7.2 Hz), 1.29
(t, 3H, J = 7.2 Hz). 13C NMR (CDCl3) δ 129.9, 128.8 (d), 128.2,
127.1, 124.3, 32.3, 8.8. C13H12O.
(Z)-tert-Butyl(1-phenylprop-1-enyloxy)dimethylsilane (9b). Com-
pound 9b was synthesized by a procedure similar to 9a using
commercially available propiophenone (8b, 5.0 g, 0.037 mol),
TBDMSOTf (9.4 mL, 41.0 mmol), and of Et3N (8.3 mL) in CH2Cl2
(50 mL). After purification, 7.32 g (79%) of 9b was isolated as
colorless oil. 1H NMR (CDCl3) δ 7.50-7.40 (m, 2H), 7.35-7.22
(m, 3H), 5.23 (q, 1H, J = 6.9 Hz), 1.77 (d, 3H, J = 6.9 Hz), 1.03 (s,
9H), -0.04 (s, 6H). 13C NMR (CDCI3) δ 150.4, 140.0, 128.0, 127.4,
125.9, 105.9, 25.9, 18.5, 11.9, -3.9. C15H24OSi.
[(M - fumaric)þ, M = C14H20ClNO2 C4H4O4]. Anal. (C18H24-
3
ClNO6 0.5H2O) C, H, N.
3
3,5,5-Trimethyl-2-(pyridin-3-yl)morpholin-2-ol (5) D-Tartrate.
A sample of 1-(pyridin-3-yl)propan-1-one (8o, 685 mg, 5.00 mmol)
was dissolved in CCl4 (20 mL). Bromine (0.26 mL, 5 mmol) was
added to the reaction flask, and the reaction mixture was gently
refluxed for 1 h. The solvent was decanted. The deep-red solid at
the bottom of flask was washed with ether, dried, suspended in
CH3CN (40 mL), and treated with 2-amino-2-methyl-1-propanol
(0.89 g, 0.01 mol). The reaction mixture was stirred for 8 h. The
precipitate was removed by filtration, and the filtrate was extrac-
ted with EtOAc. The organic layer was washed with aqueous
NaHCO3, dried (Na2SO4), and concentrated to a deep-red oil
residue. The oily residue was purified by column chromatography
on silica gel and CH2Cl2-MeOH (20:1 to 5:1) with 1% NH4OH) to
(Z)-tert-Butyl(1-(3-fluorophenyl)prop-1-enyloxy)dimethylsilane
(9c). Compound 9c was synthesized by a procedure similar to that
described for 9a using commercially available 3-fluoropropio-
phenone (8c, 5.0 g, 0.033 mol), TBDMSOTf (8.3 mL, 36.1 mmol),
and Et3N (7.3 mL) in CH2Cl2 (50 mL). After purification, 8.1 g
1
(93%) of 9c was isolated as colorless oil. H NMR (CDCl3) δ
give 56 mg free base that was converted to 5 tartrate as a yellow
7.38-7.10 (m, 2H), 7.00-6.87 (m, 1H), 5.26 (q, 1H, J = 6.9 Hz),
1.75 (d, 3H, J = 6.9 Hz), 1.00 (s, 9H), -0.02 (s, 6H). 13C NMR
(CDCl3) δ 164.5, 161.2, 149.2, 142.4, 129.4, 121.4, 114.3, 114.0,
112.8, 112.5, 107.1, 105.9, 26.0, 18.5, 11.9, -3.9. C15H23FOSi.
(Z)-tert-Butyl(1-(3-bromophenyl)prop-1-enyloxy)dimethylsilane
(9d). Compound 9d was synthesized by a procedure similar to that
described for 9a using commercially available 3-bromopropio-
phenone (8d, 1.0 g, 0.0047 mol), TBDMSOTf (1.4 mL, 5.1 mmol),
and Et3N (1.1 mL) in CH2Cl2 (10 mL). After workup, 1.15 g
(75%) of crude 9d was isolated as colorless oil with a Z-E ratio of
94:6. 1H NMR (CDCl3) δ 7.64-7.58 (m, 1H), 7.41-7.34 (m, 2H),
7.17 (t, 1H, J = 8.04 Hz), 5.25 (q, 1H, J = 6.9 Hz), 1.75 (d, 3H,
J = 6.9 Hz), 1.02 (s, 9H), -0.03 (s, 6H). 13C NMR (CDCl3) δ
148.8, 141.9, 130.2, 129.5, 128.7, 124.1, 122.1, 25.8, 18.3, 11.8, -4.0.
C15H23BrOSi.
3
solid: mp 115-116 °C. 1HNMR(CDCl3) δ8.85-8.83(m,1H),8.54
(dd, 1H, J = 4.7, 1.7 Hz), 7.91 (tt, 1H, J = 8.0, 2.0 Hz), 7.31-7.25
(m, 1H), 3.85-3.82 (m, 1H), 3.42 (d, 1H, J = 11.2 Hz), 3.21 (q, 1H,
J = 6.5 Hz), 1.40 (s, 3H), 1.10 (s, 3H), 0.81 (d, 3H, J = 6.5 Hz). 13
C
NMR (CDCl3) δ 149.4, 148.1, 137.2, 134.1, 122.8, 95.4, 69.4, 53.55,
49.8, 27.4, 22.8, 16.4. LCMS (ESI) m/z223.3 [(M - tartrate)þ, M=
C16H24N2O8]. Anal. (C16H24N2O8 0.75H2O) C, H, N.
3
3,5,5-Trimethyl-2-(pyridin-2-yl)morpholin-2-ol (6) D-Tartrate.
1-(Pyridin-2-yl)propan-1-one (8n, 1.71 g, 0.013 mol) was dissolved
in CCl4 (50 mL). Bromine (0.66 mL, 12.7 mmol) was added and
gently refluxed for 1 h. The solvent was decanted. The orange solid
at the bottom of flask was washed with ether, vacuum-dried, and
then suspend in CH3CN (40 mL), followed by the addition of
2-amino-2-methyl-1-propanol (2.30 g, 0.0254 mol). The reaction
mixture was stirred for 24 h. The precipitate was separated by
filtration and was extracted with EtOAc. The organic layer was
washed with aqueous NaHCO3 and dried (Na2SO4) and concen-
trated to give a orange oil. The residue was purified by column
chromatography on silica gel using, CH2Cl2-MeOH (50:1 to 20:1)
with 1% NH4OH) to give 0.94 g (33%) of free base that was
converted to the corresponding tartrate salt. 1H NMR (CDCl3) δ
8.55-8.51 (m, 1H), 7.84-7.59 (m, 1H), 7.63-7.57 (m, 1H),
(Z)-tert-Butyl(1-(m-tolyl)prop-1-enyloxy)dimethylsilane (9e).
Compound 9e was synthesized by a procedure similar to that
described for 9a using commercially available 3-methylpropio-
phenone (8e, 5.0 g, 0.033 mol), TBDMSOTf (8.5 mL, 36.1
mmol), and Et3N (7.5 mL) in CH2Cl2 (50 mL). After purifica-
tion by column chromatography on silica gel, 7.45 g (84%) of 9e
1
was isolated as colorless oil. H NMR (CDCl3) δ 7.31-7.24
(m, 2H), 7.23-7.17 (m, 1H), 7.10-7.05 (m, 1H), 5.22 (q, 1H,

4744 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
J = 6.9 Hz), 2.37 (s, 3H), 1.76 (d, 3H, J = 6.9 Hz), 1.03 (s, 9H),
-0.03 (s, 6H). 13C NMR (CDCl3) δ 150.7, 140.1, 137.7, 128.4,
127.9, 126.8, 123.2, 105.9, 26.3, 21.8, 18.7, 12.1, -3.6. C16H26OSi.
(Z)-tert-Butyl-(1-(3-methoxyphenyl)prop-1-enyloxy)dimethyl-
silane (9f). Compound 9f was synthesized by a procedure similar
to that described for 9a using 3-methoxypropiophenone (8f,
6.00 g, 0.0366 mol), TBDMSOTf (9.2 mL, 40.2 mmol), and Et3N
(8.1 mL) in CH2Cl2 (60 mL). After purification on alumina,
8.14 g (80%) of 9f was isolated as colorless oil. 1H NMR (CDCl3)
δ 7.24-7.16 (m, 1H), 7.07-6.98 (m, 2H), 6.84-6.76 (m, 1H),
5.23 (q, 1H, J = 6.9 Hz), 3.81 (s, 3H), 1.75 (d, 3H, J = 6.9 Hz),
1.01 (s, 9H), -0.01 (s, 6H). 13C NMR (CDCl3) δ 159.4, 150.1,
141.5, 129.0, 118.4, 113.3, 111.2, 106.1, 55.3, 26.0, 18.5, 11.9,
-3.9. C16H26O2Si.
(43.5 g), and CH3SO2NH2 (3a, 3.0 g, 0.032 mol) in tert-butyl
alcohol-water (120 mL:120 mL). The reaction mixture was
quenched with sodium sulfite (31.1 g). After purification, 4.51 g
(78%) of the (S)-10a was isolated. Characterization data is similar
with data reported in the literature.24
(R)-1-Phenyl-2-hydroxypropan-1-one (10b). Compound 10b was
synthesized by a procedure similar to that described for (R)-10a
using (Z)-tert-butyl(1-phenylprop-1-enyloxy)dimethylsilane (9b,
7.2 g, 0.029 mol), AD-mix-β (40.7 g), and CH3SO2NH2 (2.8 g,
0.0294 mol) in tert-butyl alcohol-water (110 mL:110 mL). The
reaction was quenched with sodium sulfite (29.1 g). After purifica-
tion, 2.49 g (80%) of the title product was isolated: [R]20D þ84.9° (c
1.6, CHCl3). 1H NMR (CDCI3) δ 7.96-7.90 (m, 2H), 7.66-7.58
(m, 1H), 7.54-7.47 (m, 2H), 5.23-5.12 (m, 1H), 3.84 (d, 1H, J =
6.3 Hz), 1.45 (d, 3H, J = 7.05 Hz). 13C NMR (CDCI3) δ 202.4,
134.0, 133.4, 128.9, 128.7, 69.3, 22.3. C9H10O2.
(R)-1-(3-Fluorophenyl)-2-hydroxypropan-1-one (10c). Compound
10c was synthesized by a procedure similar to that described for
(R)-10a using (Z)-tert-butyl(1-(3-fluorophenyl)prop-1-enyloxy)-
dimethylsilane, (9c, 8.0 g, 0.030 mol), AD-mix-β (42.1 g), and
CH3SO2NH2 (2.90 g, 0.0301 mol) in tert-butyl alcohol-water
(120 mL:120 mL). The reaction was quenched with sodium sulfite
(30.1 g). After purification, 4.4 g (87%) of the desired product 10c
was isolated: [R]20D þ58.1° (c 3.2, CHCl3). 1H NMR (CDCl3) δ
7.74-7.59 (m, 2H), 7.54-7.45 (m, 1H), 7.37-7.28 (m, 1H), 5.19-
5.07 (m, 1H), 1.76 (d, 1H, J = 6.3 Hz), 1.46 (d, 3H, J = 6.9 Hz).
13C NMR (CDCl3) δ 201.6, 164.9, 131.0, (d), 124.7, 121.3 (d),
115.8 (d), 69.9, 22.4. C9H9FO2.
(R)-1-(3-Bromophenyl)-2-hydroxypropan-1-one (10d). Compound
10d was synthesized by a procedure similar to that described for
(R)-10a using (Z)-tert-butyl(1-(3-bromophenyl)prop-1-enyloxy)-
dimethylsilane, (9d, 1.15 g, 0.0035 mol), AD-mix-β (4.9 g), and
CH3SO2NH2 (334 mg, 3.5 mmol) in tert-butyl alcohol-water
(17.5 mL:17.5 mL). The reaction was quenched with sodium
sulfite (3.5 g). After purification by column chromatography
(Z)-tert-Butyl-(1-(3-nitrophenyl)prop-1-enyloxy)dimethylsilane
(9g). Compound 9g was synthesized by a procedure similar to that
described for 9a using commercially available 3-nitropropiophe-
none (8g, 5.0 g, 0.028 mol), TBDMSOTf (7.1 mL, 30.7 mmol), and
Et3N (6.2 mL) in CH2Cl2 (50 mL). After purification on alumina,
8.0 g (98%) of 9g was isolated as colorless oil. 1H NMR (CDCl3) δ
8.34-8.30 (m, 1H), 8.12-8.06 (m, 1H), 7.80-7.75 (m, 1H), 7.52-
7.42 (m, 1H), 5.40 (q, 1H, J = 6.9 Hz), 1.78 (d, 3H, J= 6.9 Hz), 1.02
(s, 9H), -0.02 (s, 6H). 13C NMR (CDCI3) δ 148.8, 141.6, 131.3,
129.0, 122.1, 120.5, 108.6, 106.8, 25.9, 18.4, 12.0, -3.8. C15H23NO3Si.
(Z)-tert-Butyldimethyl-(1-(naphthalen-1-yl)prop-1-enyloxy)silane
(9q). Compound 9q was synthesized by a procedure similar to that
described for 9a using 1-naphthalen-1-yl-propan-1-one (8h, 3.5 g,
0.019 mol), TBDMSOTf (4.8 mL, 0.021 mol), and Et3N(4.3mL) in
CH2Cl2 (26 mL). After purification, 4.58 g (81%) of title product
1
was isolated as colorless oil. H NMR (CDCl3) δ 8.27-8.24 (m,
1H), 7.83-7.75 (m, 2H), 7.48-7.38 (m, 4H), 5.04 (q, 1H, J =
6.0 Hz), 1.83 (d, 3H, J = 6.0 Hz), 0.87 (s, 9H), -0.29 (s, 6H). 13
C
NMR (CDCl3) δ 150.0, 138.3, 133.6, 131.6, 128.1, 128.0, 126.8,
126.2, 125.7, 125.1, 108.4, 26.3, 25.7, 18.2, -4.6. C19H26OSi.
(Z)-tert-Butyldimethyl-(1-(naphthalen-2-yl)prop-1-enyloxy)silane
(9r). Compound 9r was synthesized by a procedure similar to that
described for 9a using 1-naphthalen-2-ylpropan-1-one (8i, 4.0 g,
0.0217 mol, TBDMSOTf (5.5 mL, 23.9 mmol), and Et3N (4.9 mL)
in CH2Cl2 (26 mL). After purification, 4.7 g (73%) of the title
product was isolated as colorless oil. 1H NMR (CDCl3) δ
7.86-7.74 (m, 1H), 7.86-7.74 (m, 3H), 7.59 (dd, 1H, J = 8.6,
1.7 Hz), 7.51-7.41 (m, 2H), 5.38 (q, 1H, J = 6.9 Hz), 1.81 (d, 3H,
J = 6.9 Hz), 1.04 (s, 9H), 0.02 (s, 6H). 13C NMR (CDCl3) δ 150.5,
137.5, 133.5, 133.2, 128.5, 127.9, 127.8, 126.4, 126.0, 107.0, 105.9,
26.3, 26.1, 18.8, -3.6. C19H26OSi.
on silica gel, 0.700 g (88%) of the desired product was isolated:
1
[R]20 þ61.1° (c 1.3, CHCl3). H NMR (CDCl3) δ 8.09-8.04
D
(m, 1H), 7.87-7.81 (m, 1H), 7.78-7.72 (m, 1H), 7.39 (t, 1H, J =
8.0 Hz), 5.18-5.05 (m, 1H), 3.66 (d, 1H, J = 6.4 Hz), 1.45 (d, 3H,
J = 7.2 Hz). 13C NMR (CDCl3) δ 201.2, 136.8, 135.2, 131.6,
130.4, 127.1, 123.3, 69.5, 22.1. C9H9BrO2.
(R)-1-(m-Tolyl)-2-hydroxypropan-1-one (10e). Compound 10e
was synthesized by a procedure similar to that described for
(R)-10a using (Z)-tert-butyl(1-(3-methylphenyl)prop-1-enyloxy)-
dimethylsilane, (9e, 7.4 g, 0.028 mol), AD-mix-β (39.5 g),
and CH3SO2NH2 (2.73 g, 0.029 mol) in tert-butyl alcohol-water
(110 mL:110 mL). The reaction was quenched with sodium
sulfite (28.3 g). After purification, 4.2 g (85%) of the desired
10e was isolated: [R]20D þ83.9° (c 2.0, CHCl3). 1H NMR (CDCl3)
δ 7.78-7.67 (m, 2H), 7.46-7.33 (m, 2H), 5.20-5.09 (m, 1H), 3.86
(Z)-tert-Butyl(1-(3-chlorophenyl)but-1-enyloxy)dimethylsilane
(9s). Compound 9s wassynthesized by a procedure similarto that
described for 9a using 3-chlorobutyrophenone (8j, 3.10 g, 0.0169
mol), TBDMSOTf (4.3 mL, 18.6 mmol), and Et3N (3.8 mL) in
CH2Cl2 (30 mL). After purification, 3.7 g (74%) of the title product
1
(d, 1H, J = 6.3 Hz), 2.42 (s, 3H), 1.44 (d, 3H, J = 7.0 Hz). 13
C
was isolated as colorless oil. H NMR (CDCl3) δ 7.48-7.44 (m,
1H), 7.38-7.32 (m, 1H), 7.31-7.29 (m, 2H), 5.16 (t, 1H, J =
7.1 Hz), 2.30-2.19 (m, 2H), 1.09- 1.03 (m, 3H), 1.01 (s, 9H), -0.04
(s, 6H). 13C NMR (CDCl3) δ 147.5, 141.7, 133.9, 129.2, 127.3,
125.9, 123.8, 114.9, 25.8, 19.5, 14.1, 0.0, -4.1. C16H25ClOSi.
(Z)-tert-Butyl(1-(3-chlorophenyl)pent-1-enyloxy)dimethylsilane
(9t). Compound 9t was synthesized by a procedure similar to that
described for 9a using 3-chloropentaphenone (8k, 2.7 g, 0.014 mol),
TBDMSOTf (3.5 mL, 15 mmol), and Et3N (3.1 mL) in CH2Cl2
(25 mL). After purification, 4.18 g (96%) of the title product was
isolated as colorless oil. 1H NMR (CDCl3) δ 7.47-7.44 (m, 1H),
7.38-7.32 (m, 1H), 7.30-7.28 (m, 1H), 7.25-7.22 (m, 1H), 5.18 (t,
1H, J = 7.2 Hz), 2.20 (q, 2H, J = 7.5 Hz), 1.54-1.38 (m, 2H), 1.02
(s, 9H), 1.00-0.90 (m, 3H), -0.05 (s, 6H). 13C NMR (CDCl3) δ
148.0, 141.8, 133.9, 129.2, 127.6, 125.9, 123.9, 113.0, 28.3, 25.8,
22.8, 14.0, 0.0, -4.01. C17H27ClOSi.
NMR (CDCl3) δ 202.6, 138.8, 134.8, 133.5, 129.1, 128.7, 125.9,
69.4, 22.3, 21.4. C10H12O2.
(R)-1-(3-Methoxyphenyl)-2-hydroxypropan-1-one (10f). Com-
pound 10f was synthesized by a procedure similar to that
described for (R)-10a using (Z)-tert-butyl(1-(3-methoxyphenyl)-
prop-1-enyloxy)dimethylsilane, (9f, 8.1 g, 0.029 mol), AD-mix-β
(40.7 g), and CH3SO2NH2 (2.8 g, 0.0294 mol) in tert-butyl
alcohol-water (110 mL:110 mL). The reaction was quenched
with sodium sulfite (29.1 g). After purification, 4.2 g (80%) of the
desired 10f was isolated: [R]20D þ71.1° (c 1.1, CHCl3). 1H NMR
(CDCl3) δ 7.51-7.45 (m, 2H), 7.44-7.37 (m, 1H), 7.19-7.13 (m,
1H), 5.19-5.09 (m, 1H), 3.87 (s, 3H), 3.76 (d, 1H, J = 6.5 Hz),
1.45(d, 3H, J = 7.1 Hz). 13C NMR (CDCl3) δ 202.3, 160.0, 134.7,
129.9, 121.1, 120.3, 113.1, 69.4, 55.5, 22.4. C10H12O3.
(R)-1-(3-Nitrophenyl)-2-hydroxypropan-1-one (10g). Compound
10g was synthesized by a procedure similar to that described for
(R)-10a using (Z)-tert-butyl(1-(3-nitrophenyl)prop-1-enyloxy)-
dimethylsilane, (9g, 8.0 g, 0.027 mol), of AD-mix-β (38 g), and
CH3SO2NH2 (2.64 g, 0.0277 mol) in tert-butyl alcohol-water
(S)-1-(3-Chlorophenyl)-2-hydroxypropan-1-one [(S)-10a]. Com-
pound (S)-10a was synthesized by a procedure similar to that
described for (R)-10a using (Z)-tert-butyl(1-(3-chlorophenyl)-
prop-1-enyloxy)dimethylsilane (9a, 8.8 g, 0.031 mol), AD-mix-R

Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4745
(110 mL:110 mL). The reaction was quenched with sodium sulfite
(27.4 g). After purification by column chromatography on silica
gel, 4.0 g (75%) of the desired product was isolated: [R]20D þ63.8°
(c 1.3, CHCl3). 1H NMR (CDCI3) δ 8.80-8.74 (m, 1H),
8.52-8.44 (m, 1H), 8.31-8.24 (m, 1H), 7.83-7.68 (m, 1H),
5.27-5.13 (m, 1H), 3.59 (d, 1H, J = 6.5 Hz), 1.49 (d, 3H, J =
7.08 Hz). 13C NMR (CDCI3) δ 200.4, 148.6, 134.9, 134.1, 130.2,
128.1, 121.6, 69.8, 21.9. C9H9NO4.
on silica gel using CH2Cl2 as the eluent afforded 9.15 g (77%) of
14 as a light-pink oil with characterization data as previously
reported:32 [R]25D þ40.5° (c 1.0, CHC13). 1H NMR (CDCl3) δ
5.27 (q, 1H, J = 6.9 Hz), 3.85 (s, 3H), 1.71 (d, 3H, J = 6.9 Hz).
13C NMR (CDCl3) δ 167.8, 79.9, 53.3, 18.0. LCMS (ESI) m/z
240.1 [(M þ 4H)þ, M = C5H7F3O5S].
(3S)-3,5,5-Trimethylmorpholin-2-one (15). A solution of tri-
flate 14 (5.00 g, 0.021 mol) in anhydrous CH2Cl2 (80 mL) under
an N2 atmosphere, was cooled to -40 °C and was treated with a
solution of 2-amino-2-methyl-1-propanol (2.5 folds, 4.68 g,
0.0525 mol) in anhydrous CH2Cl2 (10 mL). After stirring for
2 h at -40 °C, the reaction mixture was warmed slowly to 0 °C
then to room temperature and stirred overnight. The reaction
mixture was treated with saturated aqueous NaHCO3 solution
(100 mL). The organic phase was washed (water, brine), sepa-
rated, and dried (Na2SO4). The aqueous layer was extracted
with EtOAc (twice). The organic layer was separated, washed
(water, brine), and dried (Na2SO4). The organic extracts were
combined and concentrated to give a yellow oil. Column
chromatography on silica gel using hexanes-EtOAc (1:2) to
EtOAc gave 1.88 g (63%) of 15 as a light-yellow oil: [R]23D -75°
(c 1.0, CHC13). 1H NMR (CDCl3) δ 4.11 (s, 2H), 3.71 (q, 1H,
(R)-2-Hydroxy-1-(naphthalen-1-yl)propan-1-one (10h). Com-
pound 10h was synthesized by a procedure similar to that described
for (R)-10a using (Z)-tert-butyl-(1-(naphthalen-1-yl)prop-1-eny-
loxy)dimethylsilane, (9h, 4.58 g, 0.015 mol), AD-mix-β (21.4 g),
and CH3SO2NH2 (1.5 g, 0.0158 mol) in tert-butyl alcohol-water
(60 mL:60 mL). The reaction was quenched with sodium sulfite
(15.3 g). After purification by column chromatography on silica
gel, 1.15 g (38%) of the title product was isolated plus 2.1 g of
starting olefin was recovered, raising the effective yield to 70%:
[R]20D þ140.2° (c 3.2, CHCl3), 1H NMR (CDCl3) δ 8.51-8.46 (m,
1H), 8.04 (d, 1H, J = 8.3 Hz), 7.93-7.87 (m, 1H), 7.80-7.75 (m,
1H), 7.66-7.48 (m, 3H), 5.30-5.17 (m, 1H), 3.96 (d, 1H, J = 5.8
Hz), 1.36 (d, 3H, J = 7.1 Hz), 13C NMR (CDCl3) δ 205.7, 134.1,
133.5, 132.5, 130.7, 128.7, 128.4, 127.7, 126.9, 125.5, 124.4, 71.2,
21.3. LCMS (ESI) m/z 201.2 [(M þ H)þ, M = C13H12O2].
(R)-2-Hydroxy-1-(naphthalen-2-yl)propan-1-one (10i). Com-
pound 10i was synthesized by a procedure similar to that des-
cribed for (R)-10a using (Z)-tert-butyl(1-(naphthalen-2-yl)-
prop-1-enyloxy)dimethylsilane, (9i, 4.7 g, 0.016 mol), AD-mix-
β (22.0 g), and CH3SO2NH2 (1.55 g, 0.016 mol) in tert-butyl
alcohol-water (60 mL:60 mL). The reaction was quenched with
sodium sulfite (15.7 g). After purification by column chroma-
tography on silica gel, 2.5 g (80%) of the desired product was
isolated: [R]20D þ115° (c 0.7, CHCl3). 1H NMR (CDCl3) δ 8.44
(s, 1H), 8.01-7.86 (m, 4H), 7.69-7.54 (m, 2H), 5.37-5.27 (m,
1H), 3.86 (d, 1H, J = 6.5 Hz), 1.52 (d, 3H, J = 7.0 Hz). 13C
NMR (CDCl3) δ 202.3, 136.0, 132.4, 130.7, 130.5, 129.7, 129.0,
128.8, 127.9, 127.1, 124.0, 69.4, 22.5. C13H12O2.
J = 6.8), 1.39 (d, 3H, J = 6.8 Hz), 1.26 (s, 3H), 1.18 (s, 3H). 13
C
NMR (CDCl3) δ 229.2, 77.6, 49.5, 49.3, 27.2, 24.2, 18.4. LCMS
(APCI) m/z 144.3 [(M þ H)þ, M = C7H13NO2].
Note: 1H NMR data is similar with the data reported in the
literature for the racemic compound.33
Cell Lines and Culture. Human embryonic kidney (HEK-293)
cells stably expressing human DAT, NET or SERT were main-
tained as previously described.26
Use was made of several human cell lines that naturally or
heterologously express specific, functional, human nAChR sub-
types.34 Cells of the TE671/RD line naturally expresses muscle-
type nAChR (R1β1γδ- or R1*-nAChR), and SH-SY5Y neuro-
blastoma cells naturally express autonomic R3β4*-nAChRs
(containing R3, β4, probably R5, and sometimes β2 subunits).
Different clones of SH-EP1 epithelial cell lines have been
engineered to heterologously express either R4β2-nAChR, which
are thought to be the most abundant, high affinity nicotine-
binding nAChR in mammalian brain, or R4β4-nAChR, another
possible brain nAChR subtype (SH-EP1-hR4β2 or R4β4 cells, res-
pectively).35,36 These cells were maintained as low passage number
(1-26 from our frozen stocks) cultures to ensure stable expression
of native or heterologously expressed nAChR as previously
described.34 Cells were passaged once weekly by splitting just-
confluent cultures 1/300 (TE671/RD), 1/5 (SH-SY5Y), or 1/20
(transfected SH-EP1) in serum-supplemented medium to maintain
log-phase growth.
(R)-1-(3-Chlorophenyl)-2-hydroxybutan-1-one (10j). Compound
10j was synthesized by a procedure similar to that described for
(R)-10a using (Z)-tert-butyl(1-(3-chlorophenyl)but-1-enyloxy)-
dimethylsilane (9j, 3.7 g, 0.013 mol), AD-mix-β (17.5 g), and
CH3SO2NH2 (1.2 g, 0.0126 mol) in tert-butyl alcohol-water
(45 mL:45 mL). The reaction was quenched with sodium sulfite
(12.5 g). After purification by column chromatography on silica
gel, 2.2 g (79%) of the title product was isolated: [R]20D þ31.4° (c
1.0, CHCl3). 1H NMR (CDCl3) δ 7.91-7.88 (m, 1H), 7.81-7.75
(m, 1H), 7.62-7.56 (m, 1H), 7.45 (t, 1H, J = 7.8 Hz), 5.06-4.98
(m, 1H), 3.60 (d, 1H, J = 6.5 Hz), 2.04-1.87 (m, 1H), 1.70-1.51
(m, 1H), 0.94 (t, 3H, J = 7.4 Hz). 13C NMR (CDCl3) δ 201.4,
135.8, 135.7, 134.2, 130.6, 128.9, 126.9, 74.5, 29.1, 9.2. C10H11ClO2.
(R)-1-(3-Chlorophenyl)-2-hydroxypentan-1-one (10k). Compound
10k was synthesized by a procedure similar to that described for
(R)-10a using (Z)-tert-butyl(1-(3-chlorophenyl)pent-1-enyloxy)-
dimethylsilane (9k, 4.1 g, 0.013 mol), AD-mix-β (18.5 g), and
CH3SO2NH2 (1.3 g, 0.014 mol) in tert-butyl alcohol-water
(50 mL:50 mL). The reaction was quenched with sodium sulfite
(13.2g). After purification, 2.4g (77%) of the desiredproduct was
isolated: [R]20D þ33.3° (c 1.1, CHCl3). 1H NMR (CDCl3) δ 7.91-
7.87 (m, 1H), 7.80-7.75 (m, 1H), 7.63-7.17 (m, 1H), 7.45 (t, 1H,
J = 7.6 Hz), 5.08-5.00 (m, 1H), 3.58 (d, 1H, J = 6.5 Hz), 1.89-
Transporter Assays. The abilities of 2 and its analogues to
inhibit uptake of [3H]dopamine ([3H]DA), [3H]serotonin ([3H]5-
HT), or [3H]norepinephrine ([3H]NE) by the respective, human
transporters were evaluated using the appropriate HEK-293 cell
line as previously reported.26
nAChR Functional Assays. Cells were harvested at confluence
from 100 mm plates by mild trypsinization (Irvine Scientific,
Santa Ana, CA) and trituration or (for SH-SY5Y cells) by
trituration alone before being suspended in complete medium
and evenly seeded at a density of 1.25-2 confluent 100 mm
plates per 24-well plate (Falcon; ∼100-125 μg of total cell
protein per well in a 500 μL volume). After cells had adhered
(generally overnight, but no sooner than 4 h later), the medium
was removed and replaced with 250 μL per well of complete
medium supplemented with ∼350000 cpm of 86Rbþ (Perkin-
Elmer Life and Analytical Sciences, Boston, MA) and counted
at 40% efficiency using Cerenkov counting (TriCarb 1900 liquid
scintillation analyzer, 59% efficiency; PerkinElmer Life Sciences).
After at least 4 h and typically overnight, 86Rbþ efflux was
measured using the “flip-plate” technique.34 Briefly, after aspira-
tion of the bulk of 86Rbþ loading medium from each well of the
“cell plate,” each well containing cells was rinsed 3ꢀ with 2 mL of
1.75 (m, 1H), 1.61-1.35 (m, 3H), 0.93 (t, 3H, J = 7.2 Hz). 13
C
NMR (CDCl3) δ 201.1, 135.4, 133.8, 130.2, 128.6, 126.5, 73.2,
37.8, 18.2, 13.8. C11H13ClO2.
Methyl (2R)-2-[(trifluoromethyl)sulfonyl]oxypropionate) (14).
Following a reported procedure27 with modification, a solution
of methyl-(R)-(þ)-lactate (13) (5.20 g, 0.05 mol) in anhydrous
CH2Cl2 (200 mL, 0.25 M) was cooled to 0 °C and was treated
with trifluoromethane sulfonic anhydride (8.8 mL, 52.5 mmol)
and 2,6-lutidine (6.10 mL, 52.5 mmol) under an N2 atmosphere.
After stirring for 20 min at 0 °C, the reaction mixture was
concentrated to a pink oil residue. Column chromatography

4746 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12
Lukas et al.
fresh 86Rbþ efflux buffer (130 mM NaCl, 5.4 mM KCl, 2 mM
CaCl2, 5 mM glucose, 50 mM HEPES, pH 7.4) to remove
extracellular 86Rbþ. Following removal of residual rinse buffer
by aspiration, the flip-plate technique was used again to simulta-
neously introduce 1.5 mL of fresh efflux buffer containing drugs of
choice at indicated final concentrations from a 24-well “efflux/
drugplate”intothe wellsofthecellplate. Aftera5minincubation,
the solution was “flipped” back into the efflux/drug plate and any
remaining buffer in the cell plate was removed by aspiration. Cells
remaining in the cell plate were lysed and suspended by addition of
1.5 mL of 0.1 M NaOH, 0.1% sodium dodecyl sulfate to each well.
Suspensions in each well were then subjected to Cerenkov count-
ing (Wallac Micobeta Trilux 1450; 25% efficiency) after place-
ment of inserts (Wallac 1450-109) into each well to minimize
cross-talk between wells.
For quality control and normalization purposes, the sum of
86Rbþ in cell plates and efflux/drug plates was defined to
confirm material balance (i.e., that the sum of 86Rbþ released
into the efflux/drug plates and 86Rbþ remaining in the cell plate
were the same for each well). This assured that 86Rbþ efflux was
the same whether measured in absolute terms or as a percentage
of loaded 86Rbþ. Similarly, the sum of 86Rbþ in cell plates and
efflux/drug plates also determined the efficiency of 86Rbþ load-
ing (the percentage of applied 86Rbþ actually loaded into cells).
Control, total 86Rbþ efflux was assessed in the presence of
only a fully efficacious concentration of carbamylcholine (1 mM
for SH-EP1-hR4β2, SH-EP1-hR4β4 cells or TE671/RD cells; 3
mM for SH-SY5Y cells). Control, nonspecific 86Rbþ efflux was
measured either in the presence of the fully efficacious concen-
tration of carbamylcholine plus 100 μM mecamylamine, which
gave full block of agonist-induced and spontaneous nAChR-
mediated ion flux, or in the presence of efflux buffer alone.
Either determination of nonspecific efflux was equivalent. Spe-
cific efflux was then taken as the difference in control samples
between total and nonspecific 86Rbþ efflux. Any intrinsic ago-
nist activity of test drugs was ascertained using samples contain-
ing test drug only at different concentrations and was
normalized, after subtraction of nonspecific efflux, to specific
efflux in test drug-free, control samples. Antagonism of carba-
mylcholine-evoked 86Rbþ efflux was assessed in samples con-
taining the full agonist at a concentration where it stimulates
80-90% of maximal function (i.e., its EC80-EC90 value) when
exposed alone to a given nAChR subtype (i.e, 460 μM for
TE671/RD cells, 2 mM for SH-SY5Y cells; 200 μM for SH-
EP1-hR4β2 or -R4β4 cells) and test drugs at the concentrations
shown. After subtraction of nonspecific efflux, results were
normalized to specific ion flux in control samples. For studies
of mechanism of antagonism, concentration-response curves
were obtained using samples containing the full agonist, carba-
mylcholine, at the indicated concentrations alone or in the
presence of a concentration of the test ligand close to its IC50
value for inhibition of nAChR function.
effects makes it difficult to measure a single representative “nico-
tinic” acute response in animals. We opted instead in our study for
a battery of tests where these different effects of the drug are
measured. Factors such as agonist potency, time-course, site of
action, and nicotinic receptor subtypes mediating these effects
were taken into consideration. For obvious reasons, these nico-
tinic effects measured are centrally mediated. Although little is
known about the different nicotinic receptor subtypes activated
for some of these different responses, the antinociceptive effects of
nicotine were among the best described. Recent report showed
that neuronal R4β2 nAChRs subtypes are involved in nicotine-
induced antinociception in the tail-flick and hot-plate tests. How-
ever, the primary sites of action are different for the two tests (the
tail-flick assay involves a spinal reflex but in contrast, supraspinal
sites aremore likelyto be involved in the hot-plate test). Moreover,
spinal sites seems to involve both R4β2 and non-R4β2 receptors,
whereas supraspinal sites are more likely to involve R4β2 neuronal
subtypes as major component.37 We therefore decided to measure
nicotinic responses in both pain tests. All animal experiments were
conducted in accordance with the NIH Guide for the Care and
Use of Laboratory Animals and Institutional Animal Care and
Use Committee guidelines.
Animals. Male Institute of Cancer Research (ICR) mice
(weighing 20-25 g) obtained from Harlan (Indianapolis, IN)
were used throughout the study. Animals were housed in an
Association for Assessment and Accreditation of Laboratory
Animal Care-approved facility, were placed in groups of six, and
had free access to food and water. Studies were approved by the
Institutional Animal Care and Use Committee of Virginia
Commonwealth University.
Tail-Flick Test. Antinociception for pain mediated at the
spinal level was assessed by the tail-flick method of D’Amour
and Smith.38 In brief, mice were lightly restrained while a
radiant heat source was shone onto the upper portion of the
tail. To minimize tissue damage, a maximum latency of 10 s was
imposed. Latency to remove the tail from the heat source was
recorded for each animal. A control response (2-4 s) was
determined for each mouse before treatment, and a test latency
was determined after drug administration (nicotine as an an-
algesic 5 min after subcutaneous administration at 2.5 mg/kg;
nicotine administration 15 min after exposure to saline of
bupropion analogue to assess the latter drug’s ability to block
nicotine-mediated antinociception). Antinociceptive response
was calculated as the percentage of maximum possible effect
(%MPE), where %MPE = [(test control)/(10 control)] ꢀ 100.
Hot-Plate Test. Mice were placed into a 10 cm wide glass
cylinder on a hot plate (Thermojust Apparatus) maintained at
55 °C for assessment of pain responses mediated at supraspinal
levels. To minimize tissue damage, a maximum exposure to the
hot plate 40 s was imposed. Measures of control latencies (time
until the animal jumped or licked its paws; typically 8-12 s) were
done twice for stimuli applied at least 10 min apart for each
mouse. Antinociceptive responses after test drug administra-
tions were determined and calculated as the %MPE, where %
MPE = [(test latency in s - control latency in s)/(40 s - control
latency in s) ꢀ 100]. Groups of 8-12 animals were used for each
drug condition. Antagonism studies were carried in mice pre-
treated with either saline or bupropion metabolites 15 min
before nicotine administration. The animals were then tested 5
min after administration of a subcutaneous dose of 2.5 mg/kg
nicotine.
Ion flux assay results were fit using Prism (GraphPad) to the
Hill equation, F = Fmax/(1 þ (X/Z)n), where F is the test sample
specific ion flux as a percentage of control, Fmax is specific ion
flux in the absence of test drug (i.e., for control samples), X is the
test ligand concentration, Z is the EC50 (n > 0 for agonists) or
IC50 (n < 0 for antagonists), and n is the Hill coefficient. All
concentration-ion flux response curves were simple and fit well,
allowing maximum and minimum ion flux values to be deter-
mined by curve fitting, but in cases where antagonists had weak
functional potency, minimum ion flux was set at 0% of control.
Note that because agonist concentrations used for test ligand
antagonism assessments were EC80-EC90 values, not all of the
data, even at the lowest concentrations of test antagonist,
approaches 100% of specific efflux as separately determined in
sister samples exposed to fully efficacious concentrations of
agonist.
Locomotor Activity. Mice were placed into individual Omni-
tech photocell activity cages (28 cm ꢀ 16.5 cm; Omnitech
Electronics, Columbus, OH) 5 min after subcutaneous admin-
istration of either 0.9% saline or nicotine (1.5 mg/kg). Interrup-
tions of the photocell beams (two banks of eight cells each) were
then recorded for the next 10 min. Data were expressed as the
number of photocell interruptions. Antagonism studies were
carried out by pretreating the mice with either saline or bupro-
pion metabolites 15 min before nicotine administration.
Behavior. Mice were tested for nicotine-induced antinocicep-
tion, hypothermia, and hypomotility. The complex set of nicotinic

Article
Body Temperature. Rectal temperature was measured by a
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 12 4747
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Acknowledgment. This work was supported by National
Institutes of Health National Cooperative Drug Discovery
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