6-Amino-2,4,5-trimethylpyridin-3-ols: A new general synthetic route and antiangiogenic activity

Article abstract of DOI:10.1016/j.ejmech.2014.03.045

A new synthetic strategy for preparation of a wide range of 6-amino-2,4,5-trimethylpyridin-3-ols from pyridoxine·HCl via a six-step sequence has been developed. This approach features an introduction of various amino groups to C(6)-position of 3-benzyloxy-6-bromo-2,4,5-trimethylpyridine (13), a key intermediate, by a Buchwald-Hartwig amination reaction using palladium(0) transition metal, which certainly renders an expanded scope of amino substituents. Some analogs prepared using the methods described here showed high level of antiangiogenic and antitumor activities in chick chorioallantoic membrane (CAM) assay, demonstrating the potential of these new aminopyridinols as antiangiogenic agents.

Full text of DOI:10.1016/j.ejmech.2014.03.045

European Journal of Medicinal Chemistry 78 (2014) 126e139
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
6-Amino-2,4,5-trimethylpyridin-3-ols: A new general synthetic route
and antiangiogenic activity
,
,
,
Dong-Guk Kim a 1, Youra Kang a 1, Hyunji Lee a, Eun Kyung Lee a, Tae-gyu Nam b
*
,
Jung-Ae Kim a , Byeong-Seon Jeong
,
a,
*
*
a College of Pharmacy and Institute for Drug Research, Yeungnam University, Gyeongsan 712-749, Republic of Korea
b Department of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan 426-791, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
A new synthetic strategy for preparation of a wide range of 6-amino-2,4,5-trimethylpyridin-3-ols from
pyridoxine$HCl via a six-step sequence has been developed. This approach features an introduction of
various amino groups to C(6)-position of 3-benzyloxy-6-bromo-2,4,5-trimethylpyridine (13), a key in-
termediate, by a BuchwaldeHartwig amination reaction using palladium(0) transition metal, which
certainly renders an expanded scope of amino substituents. Some analogs prepared using the methods
described here showed high level of antiangiogenic and antitumor activities in chick chorioallantoic
membrane (CAM) assay, demonstrating the potential of these new aminopyridinols as antiangiogenic
agents.
Received 26 June 2013
Received in revised form
28 February 2014
Accepted 14 March 2014
Available online 17 March 2014
Keywords:
Aminopyridinol
Angiogenesis
Ó 2014 Elsevier Masson SAS. All rights reserved.
Antiangiogenic
Antitumor
Chick chorioallantoic membrane assay
1. Introduction
as a novel class of monocyclic 6-aminopyridin-3-ol antioxidants
[8,11]. In addition to a highly practical synthetic method starting
6-Aminopyridin-3-ol (1) was first proposed by Porter and col-
leagues in 2003 as a novel scaffold for antioxidants with improved
air-stability [1]. Since then, several types of analogs, including 2 and
3, have been designed and prepared as potent and air-stable anti-
oxidants [2e16]. Some lipophilic analogs were found to have better
from pyridoxine HCl (5), a component of vitamin B6, some de-
rivatives in this class also have potent radical-scavenging activities.
A lipophilic analog with N-palmityl chain (i.e., R1 ¼ R2 ¼ R3 ¼ Me,
R4 ¼ n-C16H33, R5 ¼ H in 2) showed greater capacity for prevention
of lipid peroxidation in a model membrane system using 1-
palmitoyl-2-linoleoyl-sn-glycero-3-phosphatidylcholine (PLPC) li-
posomes, and higher efficacy for protection of cells from oxidative
antioxidant properties, in many respects, than a-tocopherol (4), the
most effective natural lipophilic antioxidant. Not only did they
show the better antioxidant activities measured by peroxyl radical
trapping rate constant (kinh) but also much higher binding affinity
injury than a-tocopherol [8,11]. On the other hand, a hydrophilic,
water-soluble analog, 6-amino-2,4,5-trimethylpyridin-3-ol (i.e.,
R1 ¼ R2 ¼ R3 ¼ Me, R4 ¼ R5 ¼ H in 2), was found to act as a co-
antioxidant in heterogeneous system, which may serve to pro-
to tocopherol transfer protein (TPP) than
a-tocopherol, protection
of endogenous -tocopherol from oxidative consumption, and no
a
participation in tocopherol-mediated peroxidation [4,5]. Therefore,
interest in 6-aminopyridin-3-ols is growing, however, only limited
areas in analog space were explored.
long the half-life of a-tocopherol [11].
Antioxidant activities against reactive oxygen species (ROS)
have mainly been studied as the primary effects of aminopyridinols
on biological systems while only a few studies on their pharma-
cological actions against disease-related systems have been re-
ported [14e16] (Fig. 1).
6-Amino-2,4,5-trimethylpyridin-3-ols (2, R1 ¼ R2 ¼ R3 ¼ Me),
first reported by some of us and colleagues in 2010, were developed
Angiogenesis is a biological process of generating new blood
vessels from preexisting vasculature [17e21]. This process involves
the growth, migration, and differentiation of endothelial cells,
which line the inside wall of blood vessels. It is one of the normal
physiological functions playing important roles in reproduction,
development, and wound repair [20,21]. However, at an
* Corresponding authors.
E-mail addresses: tnam@hanyang.ac.kr (T.-g. Nam), jakim@yu.ac.kr (J.-A. Kim),
jeongb@ynu.ac.kr (B.-S. Jeong).
1
These authors contributed equally to this work and should be considered co-
first authors.
http://dx.doi.org/10.1016/j.ejmech.2014.03.045
0223-5234/Ó 2014 Elsevier Masson SAS. All rights reserved.

D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
127
Fig. 1. Aminopyridinol antioxidants.
abnormally high rate, it is deeply implicated in the pathogenesis of
many diseases, including cancer, age-related macular degeneration,
diabetic retinopathy, and so on [22,23]. Solid tumors, in particular,
desperately require new blood vessels for a sufficient supply of
nutrients and oxygen, which are essential for growth over a few
mm3 in size. In addition, new blood vessels enable cancer cells to
invade adjacent tissue and to migrate throughout the body, which
is known as metastasis [24]. Therefore, suppression of such path-
ological angiogenesis has been one of the most promising ap-
proaches in prevention and treatment of cancer [22,23].
Considerable effort has been made in development of angiogenesis
inhibitors that block tumor angiogenesis. Synthetic antiangiogenic
drugs such as sorafenib and sunitinib were approved by FDA for
treatment of cancer [25,26] and various types of angiogenesis in-
hibitors are currently being tested in clinical trials [27].
Fig. 2. The previous synthetic strategy for 6-aminopyridin-3-ols from pyridoxine$HCl
(5) and its limitations on the scope of derivatization.
consisting of 1) N,O-diacylation of 6, 2) selective hydrolysis of an O-
acyl group to afford 8, and 3) borane reduction of the amide
carbonyl group was used.
Although this synthetic strategy is highly practical and efficient
in preparation of 6-amino-2,4,5-trimethylpyridin-3-ols in quantity,
it has a limitation on the scope of alkyl groups to be attached to
C(6)-nitrogen. This route involves the formation of either an imi-
nium intermediate for N,N-dialkyl derivatives 7 or an amide in-
termediate
8 for N-monoalkyl analogs 9, in both cases,
intermediates are to be reduced in order to afford the final product.
As a result, the newly introduced alkyl moieties in the final prod-
ucts (7 and 9) always tether two hydrogen atoms at a-position to
Based on the notion that ROS are related to angiogenesis, many
natural and synthetic antioxidants including vitamin E and propolis
constituents were examined as antiangiogenic agents. For example,
the C(6)-nitrogen. Therefore, secondary and tertiary alkyl groups,
aryl groups which hold one or no hydrogen, and heterocyclic
groups which include C(6)-nitrogen cannot be introduced to the
C(6)-nitrogen under these conditions (Fig. 2). Here, we report a new
synthetic route by which those refractory substituents are suc-
cessfully attached.
For synthesis of a series of 6-amino-2,4,5-trimethylpyridin-3-
ols that cannot be prepared via the aforementioned route shown
in Fig. 2, direct introduction of various amines to the C(6)-position
of 3-alkoxy-6-halopyridine was attempted (Fig. 3).
vitamin E constituents such as a-tocopherol and d-tocotrienol and
components from propolis including artepillin C and quercetin
were found to possess a certain level of antiangiogenic activity
[28e32].
We set forth to expand the scope of the analogs of this attractive
aminopyridinol scaffold and to explore their biological activities in
more physiologically and pathologically relevant systems. Here, we
report our data on the feasibility of novel series of 6-amino-2,4,5-
trimethylpyridin-3-ol analogs as antiangiogenic agents. A general
synthetic strategy for preparation of the analogs was developed
and their antiangiogenic and antitumoreformation activities were
examined using chick chorioallantoic membrane (CAM) assay as an
in vivo model [33,34].
Among a number of synthetic methods for formation of a
s-
bond between aryl carbon and a nitrogen atom, we chose the
BuchwaldeHartwig amination, which features the palladium-
catalyzed cross-coupling of amines with aryl halides [35e43].
Accordingly, a new key intermediate for this strategy, which is
equipped with a halide leaving group at the C(6)-position and a
proper protective group on the C(3)-OH was required.
Scheme 1 shows the synthesis of the key intermediate, 3-
benzyloxy-6-bromo-2,4,5-trimethylpyridine (13). Two benzylic
hydroxy groups of pyridoxine$HCl (5) were chlorinated with excess
amounts of thionyl chloride and catalytic amount of DMF to give
40,50-dichloropyridoxine 10 as an HCl salt in 93% yield. A solvent
2. Results and discussion
2.1. New general route for synthesis of 6-amino-2,4,5-
trimethylpyridin-3-ol analogs
A general description of the synthesis of 6-dialkylamino- and 6-
monoalkylamino-2,4,5-trimethylpyridin-3-ol derivatives (7 and 9)
starting from pyridoxine hydrochloride (5), which was previously
reported by us and colleagues is shown in Fig. 2 [11]. The key in-
termediate, 6-amino-2,4,5-trimethylpyridin-3-ol (6), was prepared
in high yield from 5 through a four-step sequence. Then, the
attachment of alkyl group(s) to the C(6)-NH2 was achieved using
two different methods. The dialkylamino moiety in 7 was suc-
cessfully attached by a reductive alkylation with various aldehydes.
For synthesis of 6-monoalkylaminopyridin-3-ols (9), for which the
reductive alkylation was not very effective, a three-step strategy
Fig. 3. A new synthetic strategy for aminopyridinols.

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D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
such as 1,2-dichloroethane was often used for better stirring of the
reaction mixture. In this case, the color of the product became
darker; however, the purity was intact. Two benzylic chloride
groups of 10 were then removed with three equivalents of zinc dust
in refluxing acetic acid to give the 2,4,5-trimethyl compound 11.
Then, in order to introduce a leaving group at C(6)-position, an
electrophilic aromatic bromination was performed using 1,3-
dibromo-5,5-dimethylhydantoin (DBDMH) to afford the C(6)-
bromo compound 12 from 10 at an overall yield of approximately
70%. Finally, the phenolic OH of 12 was protected with a benzyl
group using benzyl chloride to give the key intermediate 13 in 97%
yield.
Scheme 1. Synthesis of the key intermediate 13.
Coupling of the key intermediate 13 with various amines was
performed under one of the typical BuchwaldeHartwig amination
reaction conditions with tris(dibenzylideneacetone)dipalladium(0)
Scheme 2. Synthesis of 6-amino-2,4,5-trimethylpyridin-3-ols by the CeN bond formation.

D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
129
(Pd2(dba)3)
and
2,20-bis(diphenylphosphino)-1,10-binaphthyl
the most effective of the derivatives (80.9 ꢀ 2.8% inhibition) and
compound 15m being the least effective.
(BINAP) as a ligand in the presence of sodium tert-butoxide in
refluxing toluene (Scheme 2) [35e43].
Because VEGF stimulates tyrosine kinase activity within its re-
ceptor [46], and ROS production in downstream signal transduction
pathway [47], we also compared the inhibitory effects of the 6-
aminopyridin-3-ol derivatives to SU4312, a known receptor tyro-
sine kinase inhibitor [48]. While all of the test compounds showed
considerable activities against angiogenesis, in particular, com-
pounds 6 and 15h showed the most significant inhibitory activities
compared to SU4312. We then examined the 50% inhibitory dose
(ID50) of the three compounds (SU4312, 6 and 15h) for VEGF-
induced angiogenesis (Fig. 2). Each compound was prepared in
A variety of primary and secondary amines were introduced to
the C(6)-position under these reaction conditions. In particular,
secondary alkyl amino- (14d) and aryl amino-moieties (14fei) were
successfully introduced in the pyridine ring, which was not possible
using the previously reported methods [11]. In the case of intro-
duction of 3-aminopropan-1-ol to the pyridine ring, a dimeric by-
product, 14ce2 (36% yield) was obtained along with the desired
product, 14ce1 (30% yield), in which a hydroxy group of 3-
aminopropan-1-ol also participated in the reaction with the bro-
mopyridine 13 under the reaction conditions. The benzyl protecting
group of 3-benzyloxy-6-aminopyridin-3-ols 14aej was then
removed by catalytic hydrogenolysis to afford 15’s. In the case of
compounds such as 14h and 14i, which contain functional group(s)
that are vulnerable under catalytic hydrogenolysis conditions, BCl3
was employed as an alternative reducing agent [44]. In fact, reac-
tion of 14i under catalytic hydrogenolysis conditions afforded the
debrominated compound (i.e., 15f as an HBr salt) in 95% yield.
Secondary amines, including diphenyl amine, cyclic amines, pyr-
role, and aminopyridine, were also introduced in moderate to high
yields. Debenzylation of 14kep was performed by catalytic
hydrogenolysis to give 15kep in high yields.
A slightly different approach was required for installation of an
ammonia (eNH2 group). Because ammonia can poison palladium
metal during the palladium-catalyzed CeN bond forming reaction,
we used a commercially available benzophenone imine as an
ammonia equivalent [45]. This strategy was applied successfully to
our system; a benzophenone imino-pyridinol 16 was afforded in
83% yield and the subsequent hydrolysis of the imine moiety under
acidic condition (17, 83%) followed by debenzylation gave the final
product 6 (Scheme 3).
stock solution and diluted with PBS at a concentration of
was taken from each of the five solutions of different concentra-
tions (0.001, 0.01, 0.1, 1.0 and 10 M) and each was added directly to
mM; 10 mL
m
the disk on top of CAM. Because we measured the blood vessels in
the area under the disk, the drug concentration inhibiting angio-
genesis was calculated as 10
m
L/CAM ꢁ compound concentration.
The inhibitory effects of the three compounds were dose-
dependent, and ID50’s of SU4312, 6, and 15h for VEGF-induced
angiogenesis were 10.5 pg/CAM, 3.2 pg/CAM, and 4.4 pg/CAM,
respectively.
2.3. Antiangiogenic antitumor effects of 6-amino-2,4,5-
trimethylpyridin-3-ols on A549 human lung cancer cell-inoculated
CAMs
The potential of 6 and 15h as antiangiogenic antitumor agents
was examined using cancer cell-inoculated CAM assay. As shown in
Table 2, implanted A549 cancer cells in a mixture of Matrigel onto
CAM dramatically induced angiogenesis and corresponding tumor
growth. In contrast, after exposure to compounds (once at the time
of implantation), dose-dependent inhibition of cancer cell-induced
angiogenesis was observed for compounds 6 and 15h. Similarly, the
compounds induced a significant reduction in the size and weight
of excised tumors. Results showing proportionate suppression of
tumor growth along with the inhibition of angiogenesis suggest
that antitumor activity of 6 and 15h was due to suppression of
angiogenesis.
Angiogenesis is a very complex biological process involving a
variety of events regarding vascular endothelial cell growth,
migration, and differentiation. In this study, we have presented a
novel series of 6-amino-2,4,5-trimethylpyridin-3-ols which have
significant antiangiogenic activities. Some of them showed greater
antiangiogenic activities (2.4- to 3.3-fold) than SU4312, a well-
known VEGFR tyrosine kinase inhibitor.
Although some of the phenolic antioxidants such as rosmarinic
acid, exhibited antiangiogenic activities [49], structural de-
terminants for antiangiogenic activity have not been clearly
defined. In this study, aminopyridinols with a hydrogen and a p-
nitrophenyl group on the C(6)-amino position (6 and 15h) showed
the best antiangiogenic activity (75e81% inhibition) while rela-
tively electron-rich aminoalkyl derivatives, such as 15b, 15ce1,
15d, 15m, and 15e, showed rather weak antiangiogenic activities
(30e57% inhibition). In addition, aminopyridinols with aliphatic
substituents (15a, 15b, 15ce1, 15d, 15m) were generally weaker
than those with aromatic substituents. Initially, we expected sig-
nificant variation in antiangiogenic activity among various 6-
amino-2,4,5-trimethylpyridin-3-ol structures shown in this
study, however, the window of their antiangiogenic activities was
narrower than expected. Compared to the previously known
antioxidant scaffold, it might be possible that aminopyridinols in
this study possess similar and high level of antioxidant activities
[11], which resulted in a narrow range of antioxidant and anti-
angiogenic activity.
2.2. Inhibitory effects of 6-amino-2,4,5-trimethylpyridin-3-ols on
VEGF-induced angiogenesis in vivo
Antiangiogenic activity of the compounds was determined us-
ing the quantitative chick chorioallantoic membrane (CAM) assay
[33,34], one of the most useful in vivo assay model of angiogenesis.
CAM and rabbit cornea assay are the assay used most widely as an
in vivo model for angiogenesis study. Of these, CAM provides ad-
vantages of being simpler to use, less expensive, reproducible and
reliable. CAMs were treated with vascular endothelial growth factor
(VEGF), the best characterized pro-angiogenic factor, prior to the
compound treatment. As shown in Table 1, treatment of VEGF
resulted in significant enhancement of the number of newly
formed blood vessel branch points compared to the phosphate
buffered saline (PBS)-treated control group. At a fixed dose of 0.01
nmol/CAM, the 6-aminopyridin-3-ol derivatives induced signifi-
cant suppression of VEGF-induced angiogenesis; compound 6 was
Scheme 3. Introduction of eNH2 group.

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D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
Table 1
Inhibitory effects of 6-amino-2,4,5-trimethylpyridin-3-ols (6 and 15aep) on the VEGF-induced angiogenesis in vivo.a
Treatment
Compound
Inhibition (%)b
VEGF (20 ng/CAM)þ
SU4312c
6
15a
15b
15ce1
15ce2
15d
15e
15f
15g
15h
15i
15j
15k
15l
15m
15n
15o
15p
70.5 ꢀ 6.5#
80.9 ꢀ 2.8#
53.7 ꢀ 10.3#
30.1 ꢀ 20.0
46.3 ꢀ 12.0#
67.3 ꢀ 13.9#
36.8 ꢀ 10.7
56.7 ꢀ 16.8#
69.4 ꢀ 5.7#
55.3 ꢀ 20.3
75.0 ꢀ 15.8#
61.7 ꢀ 7.7#
62.4 ꢀ 11.6
65.4 ꢀ 14.6#
69.5 ꢀ 12.7#
24.7 ꢀ 21.3
59.0 ꢀ 11.0#
44.0 ꢀ 17.2
42.2 ꢀ 21.7
#P < 0.05 compared to the VEGF-treated CAM sample.
a
Quantitation of new branches formed from existing blood vessels was performed. Photographs were imported into an Image software program, to measure the new
branch points. The data is expressed as the mean ꢀ S.E.M. of at least six chick embryos. Dose was 0.01 nmol/CAM for both test compounds and SU4312, which is equivalent to
2e3 ng/CAM depending on the molecular weights.
b
Inhibition (%) ¼ [(the number of vessel branch points induced by VEGF)ꢂ(the number of vessel branch points induced by test compound and VEGF)/(the number of vessel
branch points induced by VEGF)] ꢁ 100.
c
SU4312 is known as a potent and selective inhibitor of VEGF receptor.
.

D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
131
Table 2
Antiangiogenic antitumor effects of the compounds 6 and 15h on A549 human lung cancer cell-inoculated CAMs.a.
Treatment (pmol/CAM)
Angiogenesis inhibition (%)
Tumor inhibition (%)
SU4312
0.01
0.1
1.0
0.01
0.1
1.0
0.01
0.1
1.0
20.8 ꢀ 1.4#
25.1 ꢀ 2.8#
36.2 ꢀ 1.9#
16.6 ꢀ 3.2#
18.0 ꢀ 7.8#
20.7 ꢀ 3.6#
19.9 ꢀ 1.9
16.2 ꢀ 20.4
23.4 ꢀ 9.3
26.8 ꢀ 4.9
20.5 ꢀ 5.1
25.2 ꢀ 8.0
27.4 ꢀ 7.8#
25.3 ꢀ 5.7#
44.4 ꢀ 6.0#
50.1 ꢀ 5.6#
6
15h
25.6 ꢀ 4.5#
34.5 ꢀ 2.8#
#P < 0.05 compared to the vehicle-treated group.
a
A549 human lung cancer cells (2 ꢁ 106 cells/CAM) were inoculated on top of CAM, and different concentrations of SU4312, 6, and 15h were introduced. The number of new
vessel branches was quantitated the same method as described in Table 1. Tumor mass grown on top of CAM was isolated and weighed. The data represent the means ꢀ S.E.M.
of at least six chick embryos.

132
D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
There are several important stimulators that are known for
mixture was stirred for 3 h at 80 ꢃC. Ethyl ether (70 mL) was added,
followed by stirring for 1 h in an iced bath. The mixture was filtered
and the filter cake was washed with ethyl ether. The filter cake was
dried to give 10 (5.5 g, 93%) as a white solid.
angiogenesis, including fibroblast growth factor (FGF), VEGF, angio-
poietin (Ang) I and II, and matrix metalloproteinase (MMP) etc. Of
these, VEGF has received special attention in the context of tumori-
genic condition. Cancer cells utilize angiogenic cues, including VEGF,
to promote sprouting of new blood vessels out of preexisting vascular
system. In this study, we report another novel finding that the ami-
nopyridinols, 6 and 15h, which exhibited the best antiangiogenic
activities in VEGF-stimulated system, strongly inhibited angiogenesis
which was stimulated by inoculated A549 human lung cancer cells
(Table 2). The inhibition was dose-dependent, in line with their
antiangiogenic and antitumor activities.
Without identifying a specific target protein or enzyme with
which our aminopyridinols interact to exert the antiangiogenic
activity, it still remains quite difficult to conduct a target-based
optimization or rational drug design. However, we believe that
the novel antiangiogenic activity of the aminopyridinols deserves
future study, including extensive medicinal chemistry work, in
order to identify structural determinants for the activity and a lead
structure for further optimization.
4.2.2. 2,4,5-Trimethylpyridin-3-ol (11) [CAS no. 5622-78-6]
Zinc dust (8.08 g, 123.69 mmol) was added in small portions to a
suspension of 10 (10 g, 41.23 mmol) in acetic acid (50 mL). The
solution was refluxed for 3 h, and then cooled to room temperature.
The solution was filtered and washed with acetic acid. The pH of the
filtrate was adjusted to 6 with 10 M NaOH and saturated with NaCl.
The aqueous layer was extracted with EtOAc (100 mL ꢁ 6). The
combined organic solution was dried over MgSO4 and concen-
trated. The residue was purified by silica gel column chromatog-
raphy (CHCl3:MeOH ¼ 20:1) to give 11 (5.2 g, 92%) as a white solid.
4.2.3. 6-Bromo-2,4,5-trimethylpyridin-3-ol (12)
1,3-Dibromo-5,5-dimethylhydantoin (DBDMH, 2.5 g, 9.11 mmol)
was added to a suspension of 11 (2.5 g, 18.22 mmol) in THF (30 mL)
and the resulting mixture was stirred for 3 h at room temperature.
The mixture was concentrated and the residue was diluted with
EtOAc and water. The aqueous layer was extracted with EtOAc. The
combined organic solution was dried over MgSO4 and concen-
trated. The residue was purified by silica gel column chromatog-
raphy (EtOAc:Hexanes ¼ 1:4) to give 12 (3.22 g, 80%) as a yellow
3. Conclusion
A new synthetic strategy for preparation of a wider range of 6-
amino-2,4,5-trimethylpyridin-3-ols from pyridoxine$HCl was
developed. This route features the introduction of various amino
groups to the C(6)-position of 3-benzyloxy-6-bromo-2,4,5-
trimethylpyridine using the BuchwaldeHartwig amination reac-
tion using palladium(0) transition metal. It was shown that 6-
solid. mp 140 ꢃC; 1H NMR (CHCl3-d)
d
5.56 (br s, 1H), 2.42 (s, 3H),
2.31 (s, 3H), 2.25 (s, 3H) ppm.; 13C NMR (CHCl3-d)
148.7, 142.7,
135.3, 133.1, 131.8, 18.7, 18.3, 13.3 ppm.; IR (KBr) 3419, 2926, 2709,
d
n
2595, 2515, 1707, 1582, 1550, 1444, 1404, 1335, 1267, 1246, 1218,
1099, 1011, 954, 920, 768, 729, 689, 613, 541, 519, 475 cmꢂ1; MS(ES-
API) [M þ H] 216 (79Br).
amino-2,4,5-trimethylpyridin-3-ols
have
significant
anti-
angiogenic activity. In particular, 6 and 15h showed greater anti-
angiogenic activities than SU4312, a well-known VEGFR tyrosine
kinase inhibitor. In addition, 6 and 15h exhibited strong anti-
angiogenic antitumor activities, and the inhibitory level of 15h
against tumor growth was even higher than that of SU4312. Study
of other pharmacological activities, including antioxidant activities,
of the novel compounds prepared through this strategy is
underway.
4.2.4. 3-Benzyloxy-6-bromo-2,4,5-trimethylpyridine (13)
K2CO3 (20.78 g, 150.04 mmol), benzyl chloride (5.2 mL,
45.12 mmol) was added to a solution of 12 (6.5 g, 30.08 mmol) in
DMF (15 mL) and the resulting mixture was stirred for 12 h at room
temperature. The mixture was diluted with EtOAc (700 mL) and
water. The organic layer was washed with water (20 mL ꢁ 10). The
organic solution was dried over MgSO4 and concentrated. The
residue was purified by silica gel column chromatography
(EtOAc:Hexanes ¼ 1:20) to give 13 (8.9 g, 97%) as a white solid. m.p.
4. Experimental section
4.1. General
50 ꢃC; 1H NMR (CHCl3-d)
d
7.38e7.43 (m, 5H), 4.77 (s, 2H), 2.46 (s,
3H), 2.32 (s, 3H), 2.24 (s, 3H) ppm.; 13C NMR (CHCl3-d)
151.8,150.7,
142.0, 138.4, 136.8, 132.3, 129.0, 128.8, 128.3, 75.4, 19.5, 19.2,
14.2 ppm.; IR (KBr) 3444, 3067, 3036, 2999, 2954, 2913, 2869,
d
Materials were purchased from commercial supplier and used
without further purification. Progress of the reaction was moni-
tored by thin-layer-chromatography (TLC) using silica gel F254
plates. Purification of the products was performed using a Biotage
‘Isolera One’ with indicated solvents. Melting points were deter-
mined using a FischereJones melting point apparatus and are not
corrected. NMR spectra were obtained using a Bruker-250 spec-
trometer 250 MHz for 1H NMR and 62.5 MHz for 13C NMR and are
reported as ppm from the internal standard tetramethylsilane
(TMS). IR spectra were recorded on a Perkin Elmer Spectrum GX FT-
IR spectrometer. Low-resolution mass spectra (LRMS) were recor-
ded on an Agilent Technologies Quadruple 6130 LC/MS. High-
resolution mass spectra (HRMS) were obtained using a Thermo
Scientific LTQ Orbitrap XL mass spectrometer, and recorded in a
positive ion mode using an electrospray (ESI) source.
n
1945, 1885, 1808, 1748,1578, 1541, 1497, 1454, 1436, 1417, 1399, 1357,
1260, 1240, 1225, 1212, 1096, 1028, 993, 981, 942, 905, 841, 766, 748,
703, 687, 626, 585, 547, 524, 487, 419 cmꢂ1; MS (ES-API) [M þ H]þ
306 (79Br).
4.2.5. 5-Benzyloxy-N-hexyl-3,4,6-trimethylpyridin-2-amine (14a)
Hexylamine (0.97 mL, 7.35 mmol) was added to a mixture of 13
(1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), tris(dibenzylide-
neacetone)dipalladium(0) (Pd2(dba)3, 101 mg, 0.10 mmol), 2,20-
bis(diphenylphosphino)-1,10-binaphthyl
(BINAP,
125
mg,
0.20 mmol) in toluene (25 mL) and the resulting mixture was
refluxed for 2 h. The mixture was cooled to room temperature, and
then diluted with EtOAc (500 mL) and water. The organic layer was
washed with brine (20 mL ꢁ 5). The organic solution was dried over
MgSO4 and concentrated. The residue was purified by silica gel
column chromatography (EtOAc:Hexanes ¼ 1:15) to give 14a
4.2. Synthesis of 6-amino-2,4,5-trimethylpyridin-3-ols
4.2.1. 4,5-Bis(chloromethyl)-2-methylpyridin-3-ol hydrochloride
(10) [CAS no. 39984-50-4]
DMF (0.2 mL, 2.583 mmol) was added to a suspension pyr-
idoxine$HCl (5) (5 g, 24.31 mmol) in thionyl chloride (30 mL). The
(1.58 g, 97%) as a yellow solid. m.p. 45 ꢃC; 1H NMR (CHCl3-d)
d 7.34e
7.48 (m, 5H), 4.69 (s, 2H), 3.47 (t, J ¼ 6.8 Hz, 2H), 2.43 (s, 3H), 2.19 (s,
3H), 1.99 (s, 3H), 1.57e1.65 (m, 2H), 1.25e1.35 (m, 6H), 0.87e0.92
(m, 3H) ppm; 13C NMR (CHCl3-d)
d 152.9, 144.2, 137.8, 128.9, 128.4,

D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
133
128.3, 75.5, 42.7, 32.0, 30.4, 30.1, 27.2, 23.0, 14.4, 13.2, 13.0 ppm; IR
cooled to room temperature, and then diluted with EtOAc (500 mL)
and water. The organic layer was washed with brine (20 mL ꢁ 5). The
organic solution was dried over MgSO4 and concentrated. The res-
idue was purified by silica gel column chromatography
(EtOAc:Hexanes ¼ 1:4) to give 14ce1 (550 mg, 30%) as a yellow solid,
(KBr)
n 3404, 3061, 3031, 2922, 2855, 1590, 1488, 1456, 1402, 1362,
1335, 1212, 1154, 1096, 985, 920, 754, 714, 694, 544, 465 cmꢂ1; MS
(ES-API) [M þ H]þ 327.
1
4.2.6. 6-Hexylamino-2,4,5-trimethylpyridin-3-ol (15a)
and 14ce2 (630 mg, 36%) as a yellow solid. 14ce1: m.p. 130 ꢃC; H
To a solution of 14a (100 mg, 0.306 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15a (72 mg, 99%) as
NMR (DMSO-d6)
3H), 3.41e3.49 (m, 2H), 3.30e3.40 (m, 2H), 2.24 (s, 3H), 2.11 (s, 3H),
1.93 (s, 3H), 1.64e1.74 (m, 2H) ppm; 13C NMR (DMSO-d6)
152.7,
143.9, 142.7, 137.9, 137.5, 128.2, 127.9, 127.7, 112.7, 74.3, 58.8, 38.4,
32.6, 18.9, 12.3, 12.3 ppm; IR (KBr) 3389, 2929, 1603, 1520, 1455,
d
7.31e7.48 (m, 5H), 5.47 (t, J ¼ 5.5 Hz, 1H), 4.64 (s,
d
n
1397, 1371, 1252, 1229, 1160, 1096, 1068, 990, 935, 862, 752, 712, 695,
a yellow oil. 1H NMR (DMSO-d6)
d
7.47 (s, 1H), 5.10 (s, 1H), 3.21e3.26
545, 492, 418 cm-1; MS (ES-API) [M þ H]þ 301. 14ce2: m.p. 90 ꢃC; 1H
(t, J ¼ 6.8 Hz, 2H), 2.21 (s, 3H), 2.05 (s, 3H),1.92 (s, 3H),1.50e1.52 (m,
NMR (DMSO-d6)
d
7.30e7.47 (m, 10H), 5.49 (t, J ¼ 5.4 Hz, 1H), 4.70 (s,
2H), 1.27 (m, 6H), 0.83e0.86 (m, 3H) ppm; 13C NMR (DMSO-d6)
2H), 4.63 (s, 2H), 4.29 (t, J ¼ 6.2 Hz, 2H), 3.44e3.51 (m, 2H), 2.29 (s,
3H), 2.24 (s, 3H), 2.14 (s, 3H), 2.10 (s, 3H), 2.06 (s, 3H), 2.00e2.05 (m,
d
150.1, 140.1, 138.4, 134.8, 112.7, 41.4, 31.2, 29.2, 26.4, 22.1, 19.1, 13.9,
12.5, 12.4 ppm; IR (KBr) 2926, 2851, 1747, 1731, 1714, 1696, 1681,
n
2H),1.94 (s, 3H) ppm; 13C NMR (DMSO-d6)
d 156.4, 152.5, 145.9, 144.1,
1668, 1650, 1633, 1614, 1538, 1518, 1505, 1487, 1471, 1455, 1434,
1416, 1394, 1218, 1094, 667, 417 cm-1; MS (ES-API) [M þ H]þ 237;
HRMS calcd for C14H25N2O [M þ H]þ 237.1967, found 237.1961.
143.9, 142.8, 140.7, 137.7, 137.5, 137.1, 128.3, 128.2, 127.9, 127.8, 127.7,
116.1, 112.7, 79.1, 74.3, 63.5, 59.6, 38.4, 29.1, 19.0, 18.7, 12.4, 12.2,
11.4 ppm; IR (KBr)
n 3429, 2948, 1733, 1716, 1698, 1592, 1558, 1540,
1507, 1488, 1455, 1364, 1332, 1228, 1130, 1091, 991, 745, 717, 697,
4.2.7. 5-Benzyloxy-N-hexadecyl-3,4,6-trimethylpyridin-2-amine
(14b)
418 cmꢂ1; MS (ES-API) [M þ H]þ 526.
1-Hexadecylamine (1.96 g, 7.35 mmol) was added to a mixture
of 13 (1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), Pd2(dba)3
(101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 2 h. The mixture
was cooled to room temperature, and then diluted with EtOAc
(500 mL) and water. The organic layer was washed with brine
(20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (CHCl3:MeOH ¼ 9:1) to give 14b (1.42 g, 63%) as a
4.2.10. 6-[(3-Hydroxypropyl)amino]-2,4,5-trimethylpyridin-3-ol
(15ce1)
To a solution of 14ce1 (100 mg, 0.333 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15ce1 (70 mg,
100%) as a yellow oil. 1H NMR (DMSO-d6)
d 7.42 (s, 1H), 5.16 (s, 1H),
yellow solid. m.p. 59 ꢃC; 1H NMR (CHCl3-d)
d
7.24e7.49 (m, 5H),
3.45 (t, J ¼ 6.0 Hz, 2H), 3.31 (t, J ¼ 6.4 Hz, 2H), 2.19 (s, 3H), 2.05 (s,
4.69 (s, 2H), 3.44 (t, J ¼ 7.0 Hz, 2H), 2.41 (s, 3H), 2.24 (s, 3H), 2.18 (s,
3H), 1.91 (s, 3H), 1.60e1.70 (m, 2H) ppm; 13C NMR (DMSO-d6)
3H), 1.56e1.65 (m, 2H), 1.13e1.36 (m, 26H), 0.85e0.91 (m, 3H) ppm;
d
150.6, 140.2, 138.8, 134.6, 112.6, 58.8, 38.5, 32.8, 19.3, 12.5 ppm; IR
13C NMR (CHCl3-d)
d
153.2, 146.0, 144.3, 138.9, 138.0, 128.9, 128.3,
(KBr) 3913, 3897, 3878, 3860, 3849, 3833, 3812, 3798, 3775, 3764,
n
128.3, 113.1, 75.4, 42.5, 32.3, 30.5, 30.1, 30.0, 29.9, 29.7, 27.6, 23.1,
3741, 3730, 3719, 3707, 3685, 3666, 3644, 3625, 3605, 3584, 3389,
2938, 1865, 1841, 1789, 1746, 1730, 1714, 1694, 1681, 1667, 1650,
1644, 1633, 1614, 1573, 1556, 1537, 1514, 1504, 1470, 1455, 1372,
1232, 1072, 1032, 941, 666 cmꢂ1; MS (ES-API) [M þ H]þ 211; HRMS
calcd for C11H19N2O2 [M þ H]þ 211.1447, found 211.1443.
19.6, 14.5, 13.1, 12.9 ppm; IR (KBr)
n 3405, 2915, 2848, 1592, 1489,
1470, 1370, 1219, 1154, 1091, 984, 751, 715, 693, 506, 455 cmꢂ1; MS
(ES-API) [M þ H]þ 467.
4.2.8. 6-Hexadecylamino-2,4,5-trimethylpyridin-3-ol (15b)
To a solution of 14b (100 mg, 0.214 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15b (73 mg, 90%) as
4.2.11. 6-[3-{(5-Hydroxy-3,4,6-trimethylpyridin-2-yl)amino}
propoxy]-2,4,5-trimethylpyridin-3-ol (15ce2)
To a solution of 14ce2 (100 mg, 0.190 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15ce2 (64 mg, 97%)
a brown solid. m.p. 80 ꢃC; 1H NMR (DMSO-d6)
1H), 3.20e3.37 (m, 2H), 2.18 (s, 3H), 2.03 (s, 3H), 1.88 (s, 3H), 1.21e
1.49 (m, 28H), 0.83 (s, 3H) ppm; 13C NMR (DMSO-d6)
150.2, 140.1,
139.2, 133.9, 112.3, 41.4, 31.3, 29.3, 29.0, 28.7, 26.7, 22.1, 19.5, 13.2,
12.4 ppm; IR (KBr) 3902, 3838, 3734, 3689, 3421, 2917, 2848, 1716,
d 7.32 (s, 1H), 4.87 (s,
d
as a yellow oil. 1H NMR (DMSO-d6)
d 7.87 (br s, 1H), 7.43 (br s, 1H),
4.23 (t, J ¼ 6.3 Hz, 2H), 3.40 (t, J ¼ 6.3 Hz, 2H), 2.25 (s, 3H), 2.21 (s,
n
3H), 2.13 (s, 3H), 2.09 (s, 3H), 2.06 (s, 3H), 1.85 (s, 3H) ppm; 13C NMR
1698, 1683, 1646, 1636, 1558, 1540, 1507, 1456, 1219, 1092, 669,
418 cm-1; MS (ES-API) [M þ H]þ 377; HRMS calcd for C24H44N2O
[M þ H]þ 377.3532, found 377.3539.
(CHCl3-d)
d
154.0, 150.0, 143.4, 140.3, 138.2, 136.1, 115.3, 112.8, 68.1,
3897, 3860,
63.3, 48.5, 29.2, 21.6, 19.1, 12.4, 12.4, 11.4 ppm; IR (KBr)
n
3849, 3834, 3812, 3798, 3764, 3741, 3730, 3707, 3685, 3666, 3644,
3625, 3443, 1865, 1841, 1789, 1730, 1714, 1694, 1681, 1659, 1650,
1644, 1633, 1573, 1556, 1537, 1514, 1504, 1486, 1470, 1455, 1416,
1372, 1228, 1093, 1054, 1032, 666 cmꢂ1; MS (ES-API) [M þ H]þ 346;
HRMS calcd for C19H28N3O3 [M þ H]þ 346.2131, found 346.2129.
4.2.9. 3-[(5-Benzyloxy-3,4,6-trimethylpyridin-2-yl)amino]propan-
1-ol (14ce1) & 5-Benzyloxy-N-[3-{(5-benzyloxy-3,4,6-
trimethylpyridin-2-yl)oxy}propyl]-3,4,6-trimethylpyridin-2-amine
(14ce2)
3-Amino-1-propanol (0.6 mL, 7.83 mmol) was added to a mixture
of 13 (2 g, 6.53 mmol), NaOtBu (906 mg, 9.14 mmol), Pd2(dba)3
(135 mg, 0.13 mmol), BINAP (166 mg, 0.26 mmol) in toluene (30 mL)
and the resulting mixture was refluxed for 4 h. The mixture was
4.2.12. 5-Benzyloxy-N-cyclohexyl-3,4,6-trimethylpyridin-2-amine
(14d)
Cyclohexylamine (0.81 g, 7.10 mmol) was added to a mixture of
13 (1.8 g, 5.91 mmol), NaOtBu (820 mg, 8.27 mmol), Pd2(dba)3

134
D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
(122 mg, 0.12 mmol), BINAP (150 mg, 0.23 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 2 h. The mixture
was cooled to room temperature, and then diluted with EtOAc
(500 mL) and water. The organic layer was washed with brine
(20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (EtOAc:Hexanes ¼ 1:19) to give 14d (1.73 g, 90%) as a
1096, 1032, 699 cmꢂ1; MS (ES-API) [M þ H]þ 243; HRMS calcd for
C
15H19N2O [M þ H]þ 243.1497, found 243.1501.
4.2.16. 5-Benzyloxy-3,4,6-trimethyl-N-phenylpyridin-2-amine
(14f)
Aniline (0.71 mL, 7.838 mmol) was added to a mixture of 13 (2 g,
6.53 mmol), NaOtBu (906 mg, 9.14 mmol), Pd2(dba)3 (135 mg,
0.13 mmol), BINAP (166 mg, 0.26 mmol) in toluene (30 mL) and the
resulting mixture was refluxed for 2 h. The mixture was cooled to
room temperature, and then diluted with EtOAc (500 mL) and
water. The organic layer was washed with brine (20 mL ꢁ 5). The
organic solution was dried over MgSO4 and concentrated. The
residue was purified by silica gel column chromatography
(EtOAc:Hexanes ¼ 1:19) to give 14f (1.94 g, 93%) as a yellow solid.
yellow solid. m.p. 78 ꢃC; 1H NMR (CHCl3-d)
d 7.29e7.48 (m, 5H),
4.68 (s, 2H), 3.92e4.01 (m, 1H), 2.38 (s, 3H), 2.16 (s, 3H), 2.04e2.11
(m, 2H), 1.95 (s, 3H), 1.09e1.77 (m, 8H) ppm; 13C NMR (CHCl3-d)
d
152.3, 145.9, 144.0, 138.6, 137.9, 128.6, 128.1, 112.7, 75.2, 49.7, 34.1,
26.3, 25.3, 19.6, 12.9, 12.7 ppm; IR (KBr) 3421, 2928, 2844, 1592,
n
1558, 1540, 1507, 1489, 1456, 1396, 1362, 1340, 1212, 1151, 1100,
1014, 884, 739, 713, 694, 669, 418 cmꢂ1; MS (ES-API) [M þ H]þ 325.
m.p. 95 ꢃC; 1H NMR (CHCl3-d)
d
7.25e7.51 (m, 9H), 6.93e6.99 (m,
1H), 6.38 (s, 1H), 4.79 (s, 2H), 2.49 (s, 3H), 2.27 (s, 3H), 2.11 (s, 3H)
ppm; 13C NMR (CHCl3-d)
149.0, 147.0, 146.3, 142.5, 141.6, 137.5,
129.2, 129.0, 128.6, 128.3, 121.5, 118.5, 118.1, 75.5, 19.2, 14.1,
13.5 ppm; IR (KBr) 3399, 3025, 3005, 2918, 2884, 2844, 1924,
4.2.13. 6-Cyclohexylamino-2,4,5-trimethylpyridin-3-ol (15d)
d
To a solution of 14d (100 mg, 0.308 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15d (71 mg, 98%) as
n
1869, 1809, 1749, 1597, 1583, 1517, 1497, 1441, 1413, 1392, 1364, 1299,
1256, 1213,1180,1116,1088, 1028, 1014, 996, 925, 889, 877, 828, 743,
715, 695, 665, 620, 552, 494, 473 cmꢂ1; MS (ES-API) [M þ H]þ 319.
a yellow oil. 1H NMR (DMSO-d6)
1H), 2.19 (s, 3H), 2.04 (s, 3H), 1.90 (s, 3H), 1.10e1.66 (m, 10H) ppm;
13C NMR (DMSO-d6)
149.5, 140.1, 139.2, 134.1, 112.4, 49.0, 33.1,
25.8, 25.1, 19.7, 12.5, 12.5 ppm; IR (KBr) 2926, 2844, 1735, 1596,
d 7.41 (s, 1H), 4.51 (s, 1H), 3.77 (s,
4.2.17. 2,4,5-Trimethyl-6-(phenylamino)pyridin-3-ol (15f)
To a solution of 14f (100 mg, 0.314 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15f (70 mg, 98%) as
d
n
1494, 1450, 1358, 1298, 1239, 1210, 1137, 1098, 1051, 1032, 921, 889,
757, 655, 546 cmꢂ1; MS (ES-API) [M þ H]þ 235; HRMS calcd for
C
14H23N2O [M þ H]þ 235.1810, found 235.1815.
a yellow solid. m.p. 172 ꢃC; 1H NMR (DMSO-d6)
d 8.07 (br s, 1H), 7.53
4.2.14. N-Benzyl-5-benzyloxy-3,4,6-trimethylpyridin-2-amine
(14e)
(s, 1H), 7.25e7.28 (m, 2H), 7.10e7.16 (m, 2H), 6.68e6.73 (m, 1H),
2.27 (s, 3H), 2.13 (s, 3H), 2.08 (s, 3H) ppm; 13C NMR (DMSO-d6)
Benzylamine (0.8 mL, 7.35 mmol) was added to a mixture of 13
(1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), Pd2(dba)3
(101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 4 h. The mixture
was cooled to room temperature, and then diluted with EtOAc
(500 mL) and water. The organic layer was washed with brine
(20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (EtOAc:Hexanes ¼ 1:15) to give 14e (1.6 g, 97%) as a
d
145.5, 144.2, 143.9, 139.9, 134.5, 128.2, 118.4, 118.4, 116.4, 19.4, 13.5,
12.6 ppm; IR (KBr) n 3385, 2890, 2495, 1603, 1558, 1540, 1499, 1436,
1375, 1296, 1211, 1090, 744, 266, 418 cmꢂ1; MS (ES-API) [M þ H]þ
229; HRMS calcd for C14H17N2O [M þ H]þ 229.1341, found 229.1345.
4.2.18. 5-Benzyloxy-N-(2-isopropylphenyl)-3,4,6-trimethylpyridin-
2-amine (14g)
2-Isopropylaniline (1.03 mL, 7.35 mmol) was added to a mixture
of 13 (1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), Pd2(dba)3
(101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 2 h. The mixture
was cooled to room temperature, and then diluted with EtOAc
(500 mL) and water. The organic layer was washed with brine
(20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (CHCl3:MeOH ¼ 99:1) to give 14g (1.78 g, 99%) as a
yellow solid. m.p. 85 ꢃC; 1H NMR (CHCl3-d)
d
7.24e7.52 (m, 10H),
4.74 (s, 2H), 4.70 (s, 2H), 4.18 (s, 1H), 2.46 (s, 3H), 2.22 (s, 3H), 2.00
(s, 3H) ppm; 13C NMR (CHCl3-d)
152.7, 146.1, 144.8, 141.2, 139.1,
138.0, 129.0, 128.9, 128.8, 128.4, 128.3, 128.3, 127.4, 113.3, 75.4, 46.7,
19.7, 13.1, 12.9 ppm; IR (KBr) 3450, 3028, 2905, 2866, 1591, 1505,
d
n
1452, 1396, 1356, 1272, 1210, 1152, 1096, 1027, 993, 911, 855, 755,
730, 710, 695, 604, 451, 418 cmꢂ1; MS (ES-API) [M þ H]þ 333.
brown solid. m.p. 70 ꢃC; 1H NMR (CHCl3-d)
d 7.25e7.49 (m, 7H),
4.2.15. 6-Benzylamino-2,4,5-trimethylpyridin-3-ol (15e)
7.06e7.13 (m, 1H), 6.94e7.01 (m, 1H), 5.92 (s, 1H), 4.77 (s, 2H), 3.13e
To a solution of 14e (100 mg, 0.301 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15e (66 mg, 90%) as
3.24 (m, 1H), 2.42 (s, 3H), 2.23 (s, 3H), 2.03 (s, 3H), 1.32 (s, 3H), 1.29
(s, 3H) ppm; 13C NMR (CHCl3-d)
d
149.9, 147.1, 147.0, 140.0, 139.9,
137.6, 137.5, 128.7, 128.3, 128.1, 126.2, 125.6, 122.1, 119.6, 118.3, 75.2,
27.9, 22.9, 19.3, 14.1, 13.2 ppm; IR (KBr) 3483, 3031, 2963, 1749,
n
1716, 1698, 1683, 1602, 1521, 1472, 1455, 1418, 1395, 1363, 1294,
1218, 1091, 1019, 750, 730, 693, 444, 418 cmꢂ1; MS (ES-API)
[M þ H]þ 361.
a yellow oil. 1H NMR (DMSO-d6)
J ¼ 5.6 Hz, 1H), 4.50 (d, J ¼ 5.3 Hz, 2H), 2.17 (s, 3H), 2.06 (s, 3H), 1.97
(s, 3H) ppm; 13C NMR (DMSO-d6)
149.8, 142.2, 140.5, 139.1, 134.3,
127.8, 127.3, 126.0, 112.6, 44.5, 19.6, 12.5, 12.4 ppm; IR (KBr) 3913,
d 7.15e7.34 (m, 5H), 5.69 (t,
d
4.2.19. 6-[(2-Isopropylphenyl)amino]-2,4,5-trimethylpyridin-3-ol
(15g)
n
3897, 3878, 3860, 3849, 3833, 3812, 3798, 3775, 3764, 3741, 3730,
3707, 3685, 3666, 3644, 3625, 3605, 3584, 3562, 3361, 1865, 1841,
1789,1746,1730,1714,1694,1681,1667,1650,1644,1633,1614,1573,
1556, 1537, 1514, 1504, 1494, 1486, 1470, 1454, 1433, 1416, 1228,
To a solution of 14g (100 mg, 0.277 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite

D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
135
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15g (72 mg, 96%) as
a yellow solid. m.p. 140 ꢃC; 1H NMR (DMSO-d6)
4.2.23. 6-[(4-Bromophenyl)amino]-2,4,5-trimethylpyridin-3-ol
(15i)
d
7.19 (d, J ¼ 7.4 Hz,
To a solution of 14i (100 mg, 0.251 mmol) in CH2Cl2 (3 mL) was
added 1 M BCl3 (0.55 mL, 0.55 mmol) in an iced bath. The mixture
was stirred for 30 min in an iced bath. A solution of chloroform and
methanol (volume ratio of 9:1, 1 mL) was added to the reaction
solution and stirred for 1 h at room temperature. The reaction so-
lution was concentrated and the residue was purified by silica gel
column chromatography (CHCl3:MeOH ¼ 9:1) to give 15i (75 mg,
1H), 6.77e7.01 (m, 4H), 3.11e3.22 (m, 1H), 2.20 (s, 3H), 2.13 (s, 3H),
2.02 (s, 3H), 1.17 (s, 3H), 1.14 (s, 3H) ppm; 13C NMR (DMSO-d6)
d
147.1, 144.0, 141.2, 140.4, 138.6, 134.5, 125.5, 125.1, 121.1, 119.8,
118.6, 26.7, 22.9, 22.3, 19.2, 13.6, 12.6 ppm; IR (KBr) 3734, 3401,
n
2960, 2877, 1698, 1602, 1558, 1540, 1507, 1473, 1456, 1418, 1338,
1243, 1205, 1094, 1022, 754, 457, 418 cmꢂ1; MS (ES-API) [M þ H]þ
271; HRMS calcd for C17H23N2O [M þ H]þ 271.1810, found 271.1819.
97%) as a yellow oil. 1H NMR (DMSO-d6)
d
8.44 (br s,1H), 7.99 (s,1H),
7.30 (d, J ¼ 8.9 Hz, 2H), 7.19 (d, J ¼ 8.9 Hz, 2H), 2.30 (s, 3H), 2.16 (s,
3H), 2.09 (s, 3H) ppm; 13C NMR (CHCl3-d)
144.6, 143.2, 139.1, 131.0,
129.2, 118.4, 115.1, 109.8, 48.5, 18.3, 13.4, 12.8 ppm; IR (KBr) 1592,
d
4.2.20. 5-Benzyloxy-3,4,6-trimethyl-N-(4-nitrophenyl)pyridin-2-
amine (14h)
n
1573, 1556, 1537, 1513, 1504, 1487, 1462, 1454, 1415, 1352, 1224,
1088, 1072, 1004, 815, 666, 416 cmꢂ1; MS (ES-API) [M þ H]þ 307;
HRMS calcd for C14H16BrN2O [M þ H]þ 307.0446, found 307.0456.
4-Nitroaniline (1.03 mL, 7.35 mmol) was added to a mixture of
13 (2.18 g, 7.12 mmol), NaOtBu (987 mg, 9.96 mmol), Pd2(dba)3
(147 mg, 0.14 mmol), BINAP (177 mg, 0.28 mmol) in toluene (25 mL)
and the resulting mixture was refluxed for 6 h. The mixture was
cooled to room temperature, and then diluted with EtOAc (500 mL)
and water. The organic layer was washed with brine (20 mL ꢁ 5).
The organic solution was dried over MgSO4 and concentrated. The
residue was purified by silica gel column chromatography (CHCl3
only) to give 14h (2.36 g, 91%) as a yellow solid. m.p.171 ꢃC; 1H NMR
4.2.24. 2,4,5-Trimethyl-6-(phenylamino)pyridin-3-ol hydrobromide
(15feHBr)
To a solution of 14i (100 mg, 0.251 mmol) in methanoleTHF (1:1,
2 mL) was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15feHBr (76 mg,
(CHCl3-d)
1H), 4.77 (s, 2H), 2.46 (s, 3H), 2.24 (s, 3H), 2.15 (s, 3H) ppm; 13C NMR
(CHCl3-d) 148.7, 148.3, 147.6, 146.8, 141.1, 140.6, 137.1, 128.8, 128.5,
128.1, 125.7, 119.3, 116.0, 75.3, 19.5, 13.7, 13.3 ppm; IR (KBr) 3396,
d
8.12 (dd, J ¼ 7.2, 1.9 Hz, 2H), 7.35e7.46 (m, 7H), 6.50 (s,
d
98%) as a yellow solid. m.p. 226 ꢃC; 1H NMR (DMSO-d6)
1H), 8.82 (br s, 1H), 7.25e7.31 (m, 2H), 6.92e7.02 (m, 3H), 2.44 (s,
3H), 2.32 (s, 3H), 2.17 (s, 3H) ppm; 13C NMR (DMSO-d6)
146.2,
145.6, 142.3, 141.2, 135.7, 129.4, 125.4, 121.6, 117.1, 14.8, 14.1,
13.8 ppm; IR (KBr) 3734, 3419, 3254, 2980, 2791, 1749, 1683, 1646,
d 9.63 (br s,
n
1593, 1525, 1504, 1475, 1455, 1405, 1374, 1319, 1300, 1268, 1240,
1215, 1184, 1109, 1086, 990, 845, 748, 708, 696, 575, 418 cmꢂ1; MS
(ES-API) [M þ H]þ 364.
d
n
1635, 1594, 1558, 1540, 1507, 1473, 1456, 1375, 1350, 1198, 1096,
1026, 903, 745, 693, 418 cmꢂ1; MS (ES-API) [M þ H]þ 229.
4.2.21. 2,4,5-Trimethyl-6-[(4-nitrophenyl)amino]pyridin-3-ol (15h)
To a solution of 14h (100 mg, 0.277 mmol) in CH2Cl2 (3 mL) was
added 1 M BCl3 (0.55 mL, 0.55 mmol) in an iced bath. The mixture
was stirred for 30 min in an iced bath. A solution of chloroform and
methanol (volume ratio of 9:1, 1 mL) was added to the reaction
solution and stirred for 1 h at room temperature. The reaction so-
lution was concentrated and the residue was purified by silica gel
column chromatography (CHCl3:MeOH ¼ 9:1) to give 15h (56 mg,
4.2.25. N-(Benzo[d][1,3]dioxol-5-yl)-5-benzyloxy-3,4,6-
trimethylpyridin-2-amine (14j)
3,4-(Methylenedioxy)aniline (1.01 g, 7.35 mmol) was added to a
mixture of 13 (1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol),
Pd2(dba)3 (101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in
toluene (25 mL) and the resulting mixture was refluxed for 2 h. The
mixture was cooled to room temperature, and then diluted with
EtOAc (500 mL) and water. The organic layer was washed with
brine (20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (EtOAc:Hexanes ¼ 1:9) to give 14j (1.57 g, 88%) as a
74%) as a yellow solid. m.p. 223 ꢃC; 1H NMR (DMSO-d6)
d
8.86 (s,
1H), 8.44 (s, 1H), 8.03 (d, J ¼ 9.1 Hz, 2H), 7.21 (d, J ¼ 9.1 Hz, 2H), 2.31
(s, 3H), 2.15 (s, 3H), 2.10 (s, 3H) ppm; 13C NMR (CHCl3-d)
151.2,
d
146.1, 142.7, 140.9, 137.5, 134.7, 125.3, 121.6, 114.1, 19.2, 13.2,
12.5 ppm; IR 3392, 2923, 1597, 1498, 1479, 1319, 1277, 1197, 1113,
841, 810, 751, 467 cmꢂ1; MS (ES-API) [M þ H]þ 274; HRMS calcd for
brown solid. m.p. 130 ꢃC; 1H NMR (CHCl3-d)
d 7.33e7.47 (m, 5H),
C
14H16N3O3 [M þ H]þ 274.1192, found 274.1196.
7.17 (d, J ¼ 2.0 Hz, 1H), 6.61e6.72 (m, 2H), 5.89 (s, 3H), 4.73 (s, 2H),
2.41 (s, 3H), 2.20 (s, 3H), 2.07 (s, 3H) ppm; 13C NMR (CHCl3-d)
4.2.22. 5-Benzyloxy-N-(4-bromophenyl)-3,4,6-trimethylpyridin-2-
amine (14i)
d
149.4, 147.9, 146.6, 146.5, 142.1, 140.1, 137.5, 137.3, 128.7, 128.3,
128.1, 116.4, 111.2, 108.2, 101.7, 101.0, 75.2, 19.4, 13.5, 13.1 ppm; IR
(KBr) 3413, 2874, 1635, 1592, 1558, 1503, 1489, 1455, 1416, 1395,
4-Bromoaniline (869 mg, 4.90 mmol) was added to a mixture of
13 (1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), Pd2(dba)3
(101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 6 h. The mixture
was cooled to room temperature, and then diluted with EtOAc
(500 mL) and water. The organic layer was washed with brine
(20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (EtOAc:Hexanes ¼ 1:9) to give 14i (1.23 g, 63%) as a
n
1365, 1328, 1281, 1258, 1216, 1192, 1142, 1109, 1098, 1080, 1042,
1021, 935, 844, 815, 805, 789, 754, 736, 720, 695, 667, 606, 568, 458,
418 cmꢂ1; MS (ES-API) [M þ H]þ 363.
4.2.26. 6-(Benzo[d][1,3]dioxol-5-ylamino)-2,4,5-trimethylpyridin-
3-ol (15j)
To a solution of 14j (100 mg, 0.276 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 5 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15j (75 mg, 100%) as
yellow solid. m.p. 150 ꢃC; 1H NMR (CHCl3-d)
d
7.25e7.48 (m, 9H),
6.02 (s, 1H), 4.74 (s, 2H), 2.42 (s, 3H), 2.22 (s, 3H), 2.09 (s, 3H) ppm;
13C NMR (CHCl3-d)
148.4, 146.9, 146.8, 141.5, 140.4, 137.3, 131.8,
128.8, 128.3, 128.1, 119.6, 117.2, 112.8, 75.2, 19.4, 13.6, 13.2 ppm; IR
(KBr) 3444, 3032, 2912, 1596, 1570, 1540, 1508, 1486, 1455, 1402,
d
n
a brown solid. m.p. 168 ꢃC; 1H NMR (DMSO-d6)
1H), 6.71 (s, 2H), 5.87 (s, 2H), 2.24(s, 3H), 2.10 (s, 3H), 2.05 (s, 3H)
ppm; 13C NMR (DMSO-d6)
146.8, 146.1, 143.3, 139.8, 139.6, 138.9,
d 7.33 (s, 1H), 7.11 (s,
1372, 1299, 1254, 1213, 1177, 1082, 1071, 1031, 989, 911, 822, 808,
749, 713, 703, 694, 591, 498, 444 cmꢂ1; MS (ES-API) [M þ H]þ 397.
d

136
D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
134.6, 117.0, 109.3, 107.7, 100.2, 99.8, 19.4, 13.3, 12.6 ppm; IR (KBr)
n
4.2.30. 2,4,5-Trimethyl-6-(pyrrolidin-1-yl)pyridin-3-ol (15l)
3838, 3731, 3648, 3568, 3565, 3393, 2890, 1749, 1683, 1646, 1635,
To a solution of 14l (100 mg, 0.337 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 10 h. The solid in the reaction mixture was filtered through
Celite pad and the filtrate was filtered again with syringe filter
(AdvantecÒ JP050AN). The filtrate was concentrated to give 15l
1558, 1540, 1507, 1488, 1456, 1201, 1038, 928, 786, 669, 418 cmꢂ1
;
MS (ES-API) [M þ H]þ 273; HRMS calcd for C15H17N2O3 [M þ H]þ
273.1239, found 273.1246.
4.2.27. 5-Benzyloxy-3,4,6-trimethyl-N,N-diphenylpyridin-2-amine
(14k)
(70 mg,100%) as a pink oil. 1H NMR (DMSO-d6)
(t, J ¼ 6.5 Hz, 4H), 2.27 (s, 3H), 2.10 (s, 6H), 1.78e1.84 (m, 4H) ppm;
13C NMR (DMSO-d6)
151.4, 143.9, 138.4, 136.0, 119.4, 50.2, 24.4,
18.8, 14.8, 12.7 ppm; IR (KBr) 3913, 3897, 3878, 3860, 3850, 3834,
d 8.04 (br s,1H), 3.23
Diphenylamine (829 mg, 4.90 mmol) was added to a mixture of
13 (1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), Pd2(dba)3
(101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 2 h. The mixture
was cooled to room temperature, and then diluted with EtOAc
(500 mL) and water. The organic layer was washed with brine
(20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (EtOAc:Hexanes ¼ 1:9) to give 14k (695 mg, 36%) as a
d
n
3812, 3798, 3764, 3741, 3730, 3707, 3685, 3666, 3644, 3625, 3442,
1865, 1841, 1789, 1746, 1730, 1714, 1694, 1681, 1659, 1650, 1644,
1633, 1573, 1556, 1537, 1518, 1504, 1494, 1486, 1470, 1454, 1415,
1346, 1216, 1087, 1032, 666 cmꢂ1; MS (ES-API) [[M þ H]þ 207;
HRMS calcd for C12H19N2O [M þ H]þ 207.1497, found 207.1494.
brown oil. 1H NMR (CHCl3-d)
d
7.35e7.49 (m, 5H), 7.17e7.23 (m,
5H), 6.89e6.96 (m, 5H), 4.83 (s, 2H), 2.42 (s, 3H), 2.21 (s, 3H),1.94 (s,
3H) ppm; 13C NMR (CHCl3-d)
151.8, 150.4, 149.7, 146.8, 141.9, 137.1,
129.1, 128.8, 128.7, 128.4, 128.0, 121.8, 121.7, 77.4, 74.9, 19.6, 14.6,
13.3 ppm; IR (KBr) 3913, 3897, 3878, 3860, 3849, 3833, 3812,
4.2.31. 1-(5-Benzyloxy-3,4,6-trimethylpyridin-2-yl)-4-
methylpiperazine (14m)
N-Methylpiperazine (0.73 mL, 6.53 mmol) was added to a
mixture of 13 (2 g, 6.53 mmol), NaOtBu (906 mg, 9.14 mmol),
Pd2(dba)3 (135 mg, 0.13 mmol), BINAP (166 mg, 0.26 mmol) in
toluene (30 mL) and the resulting mixture was refluxed for 6 h. The
mixture was cooled to room temperature, and then diluted with
EtOAc (500 mL) and water. The organic layer was washed with
brine (20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (CHCl3:MeOH ¼ 20:1) to give 14m (918 mg, 43%) as a
d
n
3798, 3775, 3764, 3741, 3730, 3719, 3707, 3697, 3685, 3666, 3645,
3625, 3605, 3584, 3562, 3541, 3028, 1865, 1841, 1789, 1746, 1730,
1714,1694,1681,1667,1650,1644,1633,1587,1573,1556,1537,1515,
1504, 1494, 1486, 1470, 1454, 1433, 1416, 1367, 1216, 1089, 750,
694 cmꢂ1; MS (ES-API) [M þ H]þ 395.
yellow solid. m.p. 98 ꢃC; 1H NMR (CHCl3-d)
d
7.32e7.47 (m, 5H), 4.71
(s, 2H), 3.07 (t, J ¼ 4.7 Hz, 4H), 2.55 (br s, 4H), 2.41 (s, 3H), 2.33 (s,
3H), 2.16 (s, 3H), 2.14 (s, 3H) ppm; 13C NMR (CHCl3-d)
157.3, 147.0,
146.4, 140.5, 137.5, 128.7, 128.2, 128.0, 122.2, 74.9, 55.7, 50.4, 46.5,
19.5, 14.8, 13.2 ppm; IR (KBr) 3838, 3734, 3446, 2965, 2932, 2851,
4.2.28. 6-(Diphenylamino)-2,4,5-trimethylpyridin-3-ol (15k)
To a solution of 14k (100 mg, 0.253 mmol) in methanoleTHF
(1:1, 2 mL) was added palladium (10% on activated carbon, 20 mg).
The mixture was stirred with hydrogen balloon at room tempera-
ture for 5 h. The solid in the reaction mixture was filtered through
Celite pad and the filtrate was filtered again with syringe filter
(AdvantecÒ JP050AN). The filtrate was concentrated to give 15k
(68 mg, 87%) as a yellow solid. m.p. 227 ꢃC; 1H NMR (DMSO-d6)
d
n
2789, 1576, 1507, 1497, 1455, 1404, 1376, 1364, 1284, 1265, 1229,
1212, 1161, 1146, 1088, 1009, 972, 914, 845, 796, 750, 718, 691, 619,
591, 508, 418 cmꢂ1; MS (ES-API) [M þ H]þ 326.
d
7.16e7.22 (m, 4H), 6.79e6.98 (m, 6H), 2.28 (s, 3H), 2.13 (s, 3H),1.91
(s, 3H) ppm; 13C NMR (DMSO-d6)
148.1, 146.9, 146.3, 143.2, 134.6,
128.9, 127.7, 120.9, 120.3, 19.4, 13.8, 12.5 ppm; IR (KBr) 3418, 2922,
4.2.32. 2,4,5-Trimethyl-6-(4-methylpiperazin-1-yl)pyridin-3-ol
(15m)
d
n
To a solution of 14m (100 mg, 0.307 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 2 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15m (72 mg, 99%) as
2619,1739,1590,1493,1416,1380,1293,1273,1260,1217,1177,1093,
1029, 933, 748, 693, 586, 527, 510 cmꢂ1; MS (ES-API) [M þ H]þ 305;
HRMS calcd for C20H21N2O [M þ H]þ 305.1654, found 305.1660.
4.2.29. 3-Benzyloxy-2,4,5-trimethyl-6-(pyrrolidin-1-yl)pyridine
(14l)
a yellow solid. m.p. 149 ꢃC; 1H NMR (DMSO-d6)
4H), 2.50 (t, J ¼ 1.9 Hz, 4H), 2.27 (s, 3H), 2.25 (s, 3H), 2.08 (s, 6H)
ppm; 13C NMR (DMSO-d6)
153.3, 145.2, 139.9, 134.4, 121.6, 54.8,
49.9, 45.5, 19.4, 13.7, 12.5 ppm; IR (KBr) 3735, 3348, 2967, 2922,
d
2.87 (t, J ¼ 4.7 Hz,
Pyrrolidine (0.40 mL, 4.90 mmol) was added to a mixture of 13
(1.5 g, 4.90 mmol), NaOtBu (680 mg, 6.86 mmol), Pd2(dba)3
(101 mg, 0.10 mmol), BINAP (125 mg, 0.20 mmol) in toluene
(25 mL) and the resulting mixture was refluxed for 4 h. The
mixture was cooled to room temperature, and then diluted with
EtOAc (500 mL) and water. The organic layer was washed with
brine (20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column
chromatography (EtOAc:Hexanes ¼ 1:9) to give 14l (1.04 g, 71%) as
d
n
2860, 1735, 1587, 1455, 1413, 1368, 1307, 1270, 1229, 1105, 1065,
1032, 918, 901, 843, 697, 614 cmꢂ1; MS (ES-API) [M þ H]þ 236;
HRMS calcd for C13H22N3O [M þ H]þ 236.1763, found 236.1769.
4.2.33. 4-(5-Benzyloxy-3,4,6-trimethylpyridin-2-yl)morpholine
(14n)
a yellow oil. 1H NMR (CHCl3-d)
d
7.33e7.48 (m, 5H), 4.71 (s, 2H),
3.36 (t, J ¼ 6.7 Hz, 4H), 2.41 (s, 3H), 2.18 (s, 3H), 2.14 (s, 3H), 1.85e
1.93 (m, 4H) ppm; 13C NMR (CHCl3-d)
156.3, 146.3, 145.2, 140.2,
137.7, 128.7, 128.1, 128.0, 118.9, 75.0, 50.4, 25.5, 19.5, 15.7, 13.2 ppm;
IR (KBr) 3913, 3897, 3878, 3860, 3849, 3833, 3812, 3797, 3764,
Morpholine (0.59 mL, 6.53 mmol) was added to a mixture of 13
(2 g, 6.53 mmol), NaOtBu (906 mg, 9.14 mmol), Pd2(dba)3 (135 mg,
0.13 mmol), BINAP (166 mg, 0.26 mmol) in toluene (30 mL) and the
resulting mixture was refluxed for 5 h. The mixture was cooled to
room temperature, and then diluted with EtOAc (500 mL) and
water. The organic layer was washed with brine (20 mL ꢁ 5). The
organic solution was dried over MgSO4 and concentrated. The
residue was purified by silica gel column chromatography
(EtOAc:Hexanes ¼ 1:9) to give 14n (1.38 g, 67%) as a yellow solid.
d
n
3741, 3730, 3707, 3685, 3666, 3644, 3625, 3605, 3584, 3562, 2959,
2867, 1865, 1841, 1789, 1768, 1746, 1730, 1714, 1694, 1681, 1667,
1650, 1644, 1633, 1574, 1556, 1537, 1514, 1495, 1454, 1406, 1368,
1345, 1218, 1146, 1087, 1013, 914, 734, 716, 696, 622 cmꢂ1; MS (ES-
API) [M þ H]þ 297.
m.p. 97 ꢃC; 1H NMR (CHCl3-d)
d 7.33e7.47 (m, 5H), 4.72 (s, 2H), 3.83

D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
137
(t, J ¼ 4.6 Hz, 4H), 3.03 (t, J ¼ 4.6 Hz, 4H), 2.42 (s, 3H), 2.17 (s, 3H),
2.16 (s, 3H) ppm; 13C NMR (CHCl3-d)
157.0, 148.3, 146.6, 140.7,
137.4, 128.7, 128.3, 128.0, 122.5, 74.9, 67.5, 51.0, 29.8, 19.8, 14.6,
13.2 ppm; IR (KBr) 3902, 3838, 3734, 3648, 3566, 3419, 2970,
2915, 2851, 1716, 1698, 1683, 1636, 1575, 1558, 1540, 1521, 1507,
1488, 1456, 1418, 1362, 1266, 1223, 1112, 1092, 981, 920, 836, 748,
720, 699, 617, 418 cmꢂ1; MS (ES-API) [M þ H]þ 313.
The mixture was cooled to room temperature, and then diluted
with EtOAc (500 mL) and water. The organic layer was washed with
brine (20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (CHCl3:MeOH ¼ 50:1) to give 14p (1.78 g, 81%) as a
d
n
yellow solid. m.p. 49 ꢃC; 1H NMR (CHCl3-d)
d 8.17e8.20 (m, 1H),
7.30e7.48 (m, 6H), 6.55e6.58 (m, 1H), 6.11 (d, J ¼ 8.5 Hz, 1H), 4.82
(s, 2H), 3.40 (s, 3H), 2.47 (s, 3H), 2.23 (s, 3H), 1.98 (s, 3H) ppm; 13C
4.2.34. 2,4,5-Trimethyl-6-morpholinopyridin-3-ol (15n)
NMR (CHCl3-d)
128.8, 128.6, 128.4, 128.0, 112.9, 108.1, 77.4, 74.9, 36.5, 19.5, 14.4,
13.4 ppm; IR (KBr) 2915, 1598, 1560, 1483, 1422, 1397, 1365, 1293,
d 158.3, 151.5, 150.7, 149.7, 148.1, 141.8, 137.0, 136.9,
To a solution of 14n (100 mg, 0.320 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 2 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15n (71 mg, 99%) as
n
1270, 1229, 1191, 1159, 1102, 1054, 985, 914, 839, 764, 754, 730, 720,
697, 613, 589, 522 cmꢂ1; MS (ES-API) [M þ H]þ 334.
4.2.38. 2,4,5-Trimethyl-6-[methyl(pyridin-2-yl)amino]pyridin-3-ol
(15p)
a yellow solid. m.p. 148 ꢃC; 1H NMR (DMSO-d6)
4H), 2.83 (t, J ¼ 4.6 Hz, 4H), 2.27 (s, 3H), 2.10 (s, 3H), 2.08 (s, 3H)
ppm; 13C NMR (DMSO-d6)
153.1, 145.4, 140.0, 136.3, 121.7, 66.4,
50.8, 48.2, 19.4, 13.7, 12.5 ppm; IR (KBr) 3855, 3839, 3736, 3677,
d
3.69 (t, J ¼ 4.5 Hz,
To a solution of 14p (100 mg, 0.299 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 10 h. The solid in the reaction mixture was filtered through
Celite pad and the filtrate was filtered again with syringe filter
(AdvantecÒ JP050AN). The filtrate was concentrated to give 15p
d
n
3650, 3630, 2949, 2828, 2473, 1735, 1637, 1585, 1455, 1365, 1274,
1236, 1215, 1150, 1092, 1051, 1032, 1005, 929, 802, 777, 669, 452,
415 cmꢂ1; MS (ES-API) [M þ H]þ 223; HRMS calcd for C12H19N2O2
[M þ H]þ 223.1447, found 223.1451.
(72 mg, 98%) as a yellow oil. 1H NMR (DMSO-d6)
d 9.48 (br s, 1H),
8.07 (d, J ¼ 4.2 Hz, 1H), 7.56e7.66 (m, 1H), 6.80 (t, J ¼ 6.0 Hz, 1H),
4.2.35. 3-Benzyloxy-2,4,5-trimethyl-6-(1H-pyrrol-1-yl)pyridine
(14o)
6.40 (d, J ¼ 8.6 Hz, 2H), 3.33 (s, 3H), 2.40 (s, 3H), 2.23 (s, 3H), 1.98 (s,
3H) ppm; 13C NMR (DMSO-d6)
d 155.7, 149.4, 144.1, 142.8, 139.6,
Pyrrole (0.45 mL, 6.53 mmol) was added to a mixture of 13 (2 g,
6.53 mmol), NaOtBu (906 mg, 9.14 mmol), Pd2(dba)3 (135 mg,
0.13 mmol), BINAP (166 mg, 0.26 mmol) in toluene (30 mL) and the
resulting mixture was refluxed for 12 h. The mixture was cooled to
room temperature, and then diluted with EtOAc (500 mL) and
water. The organic layer was washed with brine (20 mL ꢁ 5). The
organic solution was dried over MgSO4 and concentrated. The
residue was purified by silica gel column chromatography
(EtOAc:Hexanes ¼ 1:9) to give 14o (1.43 g, 75%) as a yellow solid.
128.8, 113.3, 109.1, 48.5, 37.0, 18.3, 13.5, 13.2 ppm; IR (KBr) n 3913,
3897, 3878, 3860, 3849, 3834, 3812, 3798, 3775, 3764, 3741, 3730,
3707, 3685, 3666, 3645, 3625, 365, 3584, 3562, 3225, 1865, 1841,
1789, 1746, 1730, 1714, 1694, 1681, 1667, 1644, 1633, 1600, 1566,
1556,1537,1515,1504,1486,1470,1454,1415,1216,1103,1054,1032,
770 cmꢂ1; MS (ES-API) [M þ H]þ 244; HRMS calcd for C14H18N3O
[M þ H]þ 244.1450, found 244.1455.
4.2.39. 5-Benzyloxy-N-diphenylmethylene-3,4,6-trimethylpyridin-
2-amine (16)
m.p. 118 ꢃC; 1H NMR (CHCl3-d)
d
7.36e7.50 (m, 5H), 6.96 (t,
J ¼ 2.1 Hz, 2H), 6.30 (t, J ¼ 2.1 Hz, 2H), 4.82 (s, 2H), 2.49 (s, 3H), 2.27
(s, 3H), 2.14 (s, 3H) ppm; 13C NMR (CHCl3-d)
151.0, 148.9, 147.0,
141.8, 136.9, 128.8, 128.5, 128.1, 124.8, 121.4, 109.2, 75.1, 19.4, 14.9,
13.4 ppm; IR (KBr) 3902, 3838, 3734, 3432, 3083, 2954, 2898,
Benzophenone imine (1.73 mL, 9.80 mmol) was added to a
mixture of 13 (3 g, 9.80 mmol), NaOtBu (1.36 g, 13.71 mmol),
Pd2(dba)3 (203 mg, 0.20 mmol), BINAP (249 mg, 0.39 mmol) in
toluene (30 mL) and the resulting mixture was refluxed for 12 h.
The mixture was cooled to room temperature, and then diluted
with EtOAc (700 mL) and water. The organic layer was washed with
brine (20 mL ꢁ 5). The organic solution was dried over MgSO4 and
concentrated. The residue was purified by silica gel column chro-
matography (EtOAc:Hexanes ¼ 1:4) to give 16 (3.28 g, 83%) as a
d
n
2855, 1698, 1588, 1540, 1520, 1507, 1497, 1474, 1455, 1430, 1405,
1367, 1312, 1250, 1220, 1105, 1082, 1016, 1001, 926, 903, 835, 746,
728, 697, 670, 647, 590, 458, 418 cmꢂ1; MS (ES-API) [M þ H]þ 293.
4.2.36. 2,4,5-Trimethyl-6-(1H-pyrrol-1-yl)pyridin-3-ol (15o)
To a solution of 14o (100 mg, 0.342 mmol) in methanol (2 mL)
was added palladium (10% on activated carbon, 20 mg). The
mixture was stirred with hydrogen balloon at room temperature
for 2 h. The solid in the reaction mixture was filtered through Celite
pad and the filtrate was filtered again with syringe filter (AdvantecÒ
JP050AN). The filtrate was concentrated to give 15o (69 mg, 99%) as
yellow solid. m.p.133 ꢃC; 1H NMR (CHCl3-d)
d
7.80 (d, J ¼ 7.1 Hz, 2H),
7.17e7.48 (m, 13H), 4.69 (s, 2H), 2.29 (s, 3H), 2.03 (s, 3H), 1.91 (s, 3H)
ppm.; 13C NMR (CHCl3-d)
169.6, 157.2, 147.9, 147.2, 140.1, 139.3,
137.2,137.0,131.0,129.7,129.0,128.7,128.6,128.4,128.3,128.2,128.1,
127.6, 119.7, 75.1, 29.8, 19.1, 14.0, 12.8 ppm.; IR (KBr) 2921, 1737,
d
n
1637, 1448, 1398, 1363, 1314, 1225, 1055, 1032, 976, 951, 915, 783,
a yellow solid. m.p.181 ꢃC; 1H NMR (DMSO-d6)
J ¼ 2.0 Hz, 2H), 6.15 (t, J ¼ 2.0 Hz, 2H), 2.34 (s, 3H), 2.18 (s, 3H), 2.00
(s, 3H) ppm; 13C NMR (DMSO-d6)
148.4, 142.7, 141.9, 134.4, 124.1,
121.3, 107.9, 19.2, 14.1, 12.6 ppm; IR (KBr) 3734, 2869, 1736, 1595,
d
8.79 (s,1H), 6.90 (t,
753, 695, 611, 419 cmꢂ1; MS (ES-API) [M þ H]þ 407.
d
4.2.40. 5-Benzyloxy-3,4,6-trimethylpyridin-2-amine (17)
n
16 (2 g, 4.920 mmol) was dissolved in a mixed solvent of MeOH
(50 mL) and THF (5 mL), and 2% methanolic hydrogen chloride
(20 mL) was added to the solution. The resulting mixture was
stirred for 12 h at room temperature, and then concentrated. The
residue was diluted with EtOAc (300 mL) and washed with sat.
NaHCO3 solution (20 mL ꢁ 4). The organic solution was dried over
MgSO4 and concentrated. The residue was purified by silica gel
column chromatography (CHCl3:MeOH ¼ 20:1) to give 17 (992 mg,
1486, 1455, 1373, 1217, 1086, 1056, 1032, 1015, 778, 729, 669, 527,
419 cmꢂ1; MS (ES-API) [M þ H]þ 203; HRMS calcd for C12H15N2O
[M þ H]þ 203.1184, found 203.1187.
4.2.37. 5-Benzyloxy-N-3,4,6-tetramethyl-N-(pyridin-2-yl)pyridin-
2-amine (14p)
2-(Methylamino)pyridine (0.68 mL, 6.53 mmol) was added to a
mixture of 13 (2 g, 6.53 mmol), NaOtBu (906 mg, 9.14 mmol),
Pd2(dba)3 (135 mg, 0.13 mmol), BINAP (166 mg, 0.26 mmol) in
toluene (30 mL) and the resulting mixture was refluxed for 12 h.
83%) as a yellow solid. m.p. 127 ꢃC; 1H NMR (CHCl3-d)
d
7.31e7.45
(m, 5H), 4.68 (s, 2H), 4.25 (br s, 1H), 2.34 (s, 3H), 2.16 (s, 3H), 1.99 (s,
3H) ppm; 13C NMR (CHCl3-d)
152.5,146.1,145.4,140.2,137.4, 128.7,
d

138
D.-G. Kim et al. / European Journal of Medicinal Chemistry 78 (2014) 126e139
128.2, 128.1, 113.7, 75.2, 18.8, 13.2, 12.5 ppm; IR (KBr)
n
3676, 3458,
Acknowledgment
3306, 3169, 3027, 2906, 2859, 1634, 1588, 1447, 1417, 1367, 1273,
1230, 1144, 1083, 1029, 985, 911, 862, 754, 716, 696, 612, 509,
431 cmꢂ1; MS (ES-API) [M þ H]þ 243.
This research was supported by Basic Science Research Program
through the National Research Foundation of Korea (NRF) funded
by the Ministry of Education, Science and Technology
(2012R1A1A2039174).
4.2.41. 6-Amino-2,4,5-trimethylpyridin-3-ol (6) [CAS no. 1245315-
08-5]
To a solution of 17 (50 mg, 0.206 mmol) in methanol (2 mL) was
added palladium (10% on activated carbon,10 mg). The mixture was
stirred with hydrogen balloon at room temperature for 3 h. The
solid in the reaction mixture was filtered through Celite pad and the
filtrate was filtered again with syringe filter (AdvantecÒ JP050AN).
The filtrate was concentrated to give 6 (31 mg, 100%) as an orange
solid.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.03.045.
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