Synthesis and muscarinic activity of 1,2,3,4-tetrahydropyrimidine derivatives

Article abstract of DOI:10.1002/1521-4184(200103)334:3<79::AID-ARDP79>3.0.CO;2-7

3-Methyl-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde O-substituted oximes 4 and 1-(3-methyl-1,2,3,4-tetrahydropyrimidin-5-yl)ethanone O-substituted oximes 9 have been prepared as bioisosteric congeners of arecoline which is a muscarinic agonist for treatment of Alzheimer's disease. Starting from pyrimidine-5-carbaldehyde 1, formation of the 3-methylpyrimidinium salt and subsequent reduction afforded 1,2,3,4-tetrahydropyrimidine derivatives which were converted into oxalate salts in the interest of purity and stability. Binding affinities of prepared compounds for the cloned human muscarinic M₁ receptor (h-M₁) were determined by radioligand binding assay using [³H]-N-methylscopolamine (NMS).

Full text of DOI:10.1002/1521-4184(200103)334:3<79::AID-ARDP79>3.0.CO;2-7

1,2,3,4-Tetrahydropyrimidine Derivatives
79
Synthesis and Muscarinic Activity of 1,2,3,4-Tetrahydropyrimidine
Derivatives✩
Myung Hee Jung*, Jewn-Giew Park, Kong Jae Yang, and Mi-Jeoung Lee
Korea Research Institute of Chemical Technology, P.O. Box 107, Yusong, Taejon 305-600, Korea
Key Words: 1,2,3,4-Tetrahydropyrimidine; muscarinic agonist; pyrimidine-5-carbaldehyde oxime;
pyrimidine-5-ethanone oxime
Summary
Synthesis
As shown in Scheme 1, the starting pyrimidine-5-carbalde-
3-Methyl-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde O-substi-
tuted oximes 4 and 1-(3-methyl-1,2,3,4-tetrahydropyrimidin-5-
yl)ethanone O-substituted oximes 9 have been prepared as
bioisosteric congeners of arecoline which is a muscarinic agonist
for treatment of Alzheimer’s disease. Starting from pyrimidine-5-
carbaldehyde 1, formation of the 3-methylpyrimidinium salt and
subsequent reduction afforded 1,2,3,4-tetrahydropyrimidine de-
rivatives which were converted into oxalate salts in the interest of
purity and stability. Binding affinities of prepared compounds for
the cloned human muscarinic M1 receptor (h-M1) were determined
by radioligand binding assay using [3H]-N-methylscopolamine
(NMS).
hyde 1 was first synthesized from 4,6-dihydroxypyrimidine
[6]
in accordance with the procedure desribed in reference .
Compounds 2 and 2a were easily prepared by condensation
of pyrimidine-5-carbaldehyde 1 with hydroxylamine and
methoxylamine in 40% and 62% yield, respectively. The
reaction afforded 2 and 2a as the sterically more favored
[7]
E-isomers as the major products along with small amounts
of less favored Z-isomers. Both isomers were readily sepa-
1
rated by column chromatography on silica gel. In the H
NMR spectra, the aldimine protons of the E-isomers always
showed downfield shifts (8.13, 8.03 ppm) relative to those of
[8,9]
Z-isomers (7.46, 7.26 ppm)
. The oxime 2 obtained was
reacted with various alkyl halides in the presence of a base to
give compounds 2b–2g. Subsequent treatment of 2a–2g with
methyl iodide in acetone for 48–72 hours afforded the quar-
ternized compounds 3a–3g in almost quantitative yields.
Although several products were expected in the hydride
Introduction
Recent research effort has focused on the development of
centrally acting muscarinic agonists as a potential remedy for
Alzheimer’s disease . The classical muscarinic agonist
arecoline (Chart 1) led to little improvement in Alzheimer-
type dementia patients . However, arecoline is compro-
mised by its poor metabolic stability and peripheral side
effects. With a view to developing arecoline relating entities
in which the ester group has been replaced by an oxime ether
group. One of them is milameline (Chart 1), whose pharma-
cological effects are 2–3 times more potent than those of
arecoline; it also has longer duration of action . While both
arecoline and milameline have a tetrahydropyridine ring, in
our effort to develop a possible M -selective muscarinic
agonist, we have introduced a 1,2,3,4-tetrahydropyrimidine
ring employing the bioisosteric replacement concept. Tetra-
hydropyrimidine rings are structures exerting interesting bio-
logical activity in medicines and should be more stable
against metabolic degradation: Moreover, further introduc-
[1]
reduction of 5-substituted pyrimidines, NaBH reduction of
4
3a–3g in MeOH yielded the 1,2,3,4-tetrahydropyrimidine
derivatives as the sole products, after purification by column
chromatography. The structure of reductive compounds,
1,2,3,4-tetrahydropyrimidine derivatives, can be deduced
[2]
[10]
fromX-raycrystallography . They are structural congeners
of milameline, and were further converted into the oxalate
salts 4a–4g to improve the stability and purity of the com-
[3]
1
13
pounds ( H NMR data and C NMR data are summarized in
Table 1). All the compounds 4a–4g were the E-isomers, and
1
1
in the H NMR spectra, the oxime protons exhibited their
small variational chemical shifts of 7.65–7.76 ppm. All the
compounds (4a–4g) had a typical singlet signal at 3.61–
3.70 ppm corresponding to the C4-protons without excep-
[4]
[5]
tion .
Scheme 2 shows the synthesis of O-substituted 1-(5-
pyrimidinyl)-1-ethanone oximes 9 starting from pyrimidine-
5-carbaldehyde 1. The Grignard reaction of the starting
aldehyde 1 and subsequent pyridinium dichromate oxidation
yielded the key intermediate 1-(5-pyrimidinyl)-1-ethanone 6.
Similarly to the previous experiment, the resultant ethanone
6 was reacted with hydroxylamine and methoxylamine to
give 7 and 7a, respectively. Again, the reaction yielded
sterically more favored E-isomers as the major products.
Interestingly, in the case of 7, it afforded a small amount of
3-methyl-isoxazolo[5,4-d]pyrimidine instead of the Z-iso-
mer. The methyl proton of 7 showed a chemical shift of
2.28 ppm in NMR spectrum, whereas that of 3-methylisoxaz-
olo[5,4-d]pyrimidine was 2.42 ppm. The chemical shift of
tion of various substituents on the 5-position of the tetrahy-
dropyrimidine ring should give a meaningful result in
Structure-Activity Relationship (SAR) studies
[5]
.
Chart 1. Muscarinic agonists.
Arch. Pharm. Pharm. Med. Chem.
© WILEY-VCH Verlag GmbH, D-69451 Weinheim, 2001
0365-6233/01/0303/0079 $17.50 +.50/0

80
Jung, Park, Yang, and Lee
Table 1. 1H NMR data (DMSO) and 13C NMR data (DMSO) of the compound 4.
—————————————————————————————————————————————————————————
Proton
δ [ppm]
4-H
Carbon
δ [ppm]
C-5
Compd
2-H
6-H
CH=N
C-2
C-4
C-6
C=N
—————————————————————————————————————————————————————————
4a
4b
4c
4d
4e
4f
4.17
4.17
4.16
4.18
4.18
4.25
4.25
3.62
3.63
3.61
3.62
3.63
3.70
3.68
6.82
6.80
6.82
6.87
6.84
6.85
6.86
7.65
7.66
7.70
7.71
7.74
7.74
7.76
61.3
61.4
61.4
61.3
61.4
61.4
61.4
49.1
49.2
49.2
49.1
49.2
49.2
49.1
97.1
97.5
97.2
96.7
97.2
97.2
97.1
137.0
136.7
137.1
137.9
137.3
137.2
137.4
148.7
148.3
149.0
149.9
149.3
149.1
149.5
4g
—————————————————————————————————————————————————————————
.
.
Scheme 1. Reagents: i) HONH2 HCl; ii) CH3ONH2 HCl; iii) RX; iv) CH3I ; v) a. NaBH4/MeOH b. (CO2H)2.
ketoxime methyl of E-isomer of 7a was observed at Experimental Section, the methyl protons of ketoximes are
observed in the range of 1.85–1.90 ppm in DMSO-d .
1.99 ppm, whereas that of the Z-isomer was at 2.06 ppm. This
once again reveals the structural outcomes. The E-isomers
were separated by column chromatography, then reacted with
alkyl halides. Subsequent quarternization to 8a–8g and re-
duction under similar conditions for the aldimines afforded
the only isolable products, 1,2,3,4-tetrahydropyrimidine de-
6
Results and Discussion
In an attempt to develop more selective, active, and biologi-
cally stable M muscarinic agonists, we have synthesized
1
[10]
rivatives , which were treated with oxalic acid to give the
1,2,3,4-tetrahydropyrimidine derivatives 4 and 9 as arecoline
final oxalate salts 9a–9g (Table 2). As can be seen in the and milameline congeners. The rationales for the design of
Arch. Pharm. Pharm. Med. Chem. 334, 79–85 (2001)

1,2,3,4-Tetrahydropyrimidine Derivatives
81
Table 2. 1H NMR data (DMSO) and 13C NMR data (DMSO) of the compound 9.
—————————————————————————————————————————————————————————
Proton
δ [ppm]
4-H
Carbon
δ [ppm]
C-6
Compd
2-H
6-H
-N=CCH3
C-2
C-4
C-5
C=N
—————————————————————————————————————————————————————————
9a
9b
9c
9d
9e
9f
4.23
4.23
4.24
4.24
4.24
4.24
4.24
3.76
3.76
3.76
3.75
3.76
3.76
3.74
6.92
6.90
6.93
6.97
6.94
6.93
6.95
1.85
1.85
1.88
1.87
1.90
1.88
1.90
61.2
61.2
61.1
61.1
61.1
61.1
61.1
49.8
49.8
49.8
49.7
49.8
49.8
49.7
99.7
100.1
99.8
99.2
99.7
99.8
99.6
132.8
132.5
132.8
133.5
132.9
132.9
133.1
153.1
152.7
153.3
153.4
153.6
153.4
153.8
9g
—————————————————————————————————————————————————————————
.
.
Scheme 2. Reagents: i) MeMgBr; ii) PDC/HOAc; iii) HONH2 HCl; iv) CH3ONH2 HCl; v) RX; vi) CH3I; vii) a. NaBH4 b. (CO2H)2.
these compounds are as follows: 1) naturally occurring determined indirectly by assaying displacement affinity of
3
1,2,3,4-tetrahydropyrimidine could contribute to formation [ H]-NMS in the suspension of cloned human M expressed
1
of biologically stable compounds and fits a bioisosteric re- in CHO cells. Table 3 and 4 show that milameline has higher
placement concept; 2) various substituents on the 5-position affinity than arecoline at both 10 and 100 µM concentrations
of tetrahydropyrimidine ring may result better agonists than and these previously characterized agonists are used as con-
current arecoline and milameline. The binding affinities were trols for the newly synthesized compounds. Alkyl deriva-
Arch. Pharm. Pharm. Med. Chem. 334, 79–85 (2001)

82
Jung, Park, Yang, and Lee
Table 4. Inhibition of [3H]-NMS to h-M1 by 1,2,3,4-tetrahydropyrimidine-
5-ethanone oximes.
Table 3. Inhibition of [3H]-NMS to h-M1 by 1,2,3,4-tetrahydropyrimidine-
5-carbaldehyde oximes.
——————————————————————————————
——————————————————————————————
% inhibition of [3H]NMS binding IC50 (µM)
Compounds
% inhibition of [3H]NMS binding IC50 (µM)
Compounds
100 µM
10 µM
Mean ± S.E
100 µM
10 µM
Mean ± S.E
——————————————————————————————
——————————————————————————————
9a
9b
38.1
70.5
77.6
75.1
77.6
78.5
86.4
44.7
73.7
8.5
N.D.
41.3
4a
2.3
0.7
N.D.
15.9
26.0
16.7
19.2
23.8
24.2
7.1
4b
4c
49.7
44.6
68.7
69.5
71.4
84.0
44.7
73.7
12.9
5.1
99.54 ± 21.87
N.D.
9c
27.66 ± 9.17
37.23 ± 5.13
33.29 ± 4.67
29.77 ± 5.27
23.66 ± 3.69
N.D.
9d
4d
16.6
29.1
29.0
29.0
7.1
49.16 ± 10.57
33.73 ± 3.40
32.00 ± 1.37
23.19 ± 2.90
N.D.
9e
4e
9f
4f
9g
4g
arecoline
milameline
arecoline
milameline
24.4
52.36 ± 8.55
24.4
52.36 ± 8.55
——————————————————————————————
* N.D: Not determined.
——————————————————————————————
* N.D: Not determined.
* Data represent the mean ± S.E from three assays each performed in
duplicate.
* Data represent the mean ± S.E from three assays each performed in
duplicate.
Pyrimidine-5-carbaldehyde Oxime (2)
tives, 4a, 4b, 9a, and 9b, regardless of their (methyl or pentyl)
chain length, show no better affinity than arecoline and
milameline. The benzyl derivatives (4e, 4f, 4g, 9e, 9f, and 9g)
displayed high binding affinities. Among these, ligand-recep-
To a stirred solution of pyrimidine-5-carbaldehyde 1 (5.36 g, 49.58 mmol)
in toluene (50 ml) was added hydroxylamine hydrochloride (3.79 g, 54.54
mmol). The mixture was heated under reflux for 1 h, then cooled to room
temperature, and washed with 200 ml water. The aqueous layer was neutral-
ized with solid K2CO3, concentrated to dryness, and then combined with the
organic concentrate. The combined residue was subjected to silica gel
chromatography (EtOAc:Hex = 1:1) to give 2.43 g (40%) of a pale yellow
solid. mp 141–142 °C. IR (KBr) ν: 3300–2800, 1560, 1400.– 1H-NMR (200
MHz, CD3OD) δ: 9.09 (1H, s, C2-H), 8.96 (2H, s, C4-H, C6-H), 8.13 (1H,
s, CH=N).– 13C-NMR (75 MHz, CD3OD) δ: 159.4 (C2), 156.8 (C4, C6),
144.5 (C=N), 129.9 (C5).
tor interaction of 4-chlorobenzyl derivatives 4g (IC = 23.19
50
± 2.90) and 9g (IC = 23.66 ± 3.69) was more than twice as
50
efficient as that of milameline (IC = 52.36 ± 8.55). Unlike
50
4c, the affinity of 9c was unexpectedly comparable to that of
the highest ones 4g and 9g. In summary, the data indicate that
these 5-position variants of 1,2,3,4-tetrahydropyrimidine are
efficient agonists towards M muscarinic receptors. In par-
1
ticular, the 4g, 9f, and 9g compounds justify further studies
Pyrimidine-5-carbaldehyde O-Methyloxime (2a)
aiming at the development of M muscarinic agonists more
1
selective and efficacious than currently available ones for the
treatment of Alzheimer’s disease. Functional selectivity of
To a stirred solution of pyrimidine-5-carbaldehyde 1 (3.72 g, 34.42 mmol)
in benzene (60 ml) was added methoxylamine hydrochloride (3.16 g,
37.84 mmol). The mixture was heated under reflux for 2 h using a Dean-Stark
column, cooled to room temperature, and washed with aqueous NaHCO3
solution. The aqueous layer was extracted with CH2Cl2 and then combined
to the organic layer, concentrated in vacuo. The residue was subjected to
silica gel flash chromatography (EtOAc:Hex = 4:1) to give 2a as a yellow
solid. Yield 62%. mp 49–50 °C. IR (KBr) ν: 3600, 2900, 1700, 1650.–
1H-NMR (200 MHz, CDCl3) δ: 9.19 (1H, s, C2-H), 8.92 (2H, s, C4-H,
C6-H), 8.03 (1H, s, CH=N), 4.03 (3H, s, CH3).– 13C-NMR (50 MHz, CDCl3)
δ: 158.3 (C2), 154.5 (C4, C6), 142.4 (C=N), 126.6 (C5), 62.6 (CH3).– MS
m/z: 137.0589 (Calcd for C6H7N3O: 137.0590).
these compounds for M versus M , M , M , and M mus-
1
2
3
4
5
carinic receptors has to be assessed urgently. Biochemical
and molecular modeling will help to determine interaction
between key amino acid residues and our synthetic ligands
and to understand steric and electronic requirements for the
future design of new compounds. The in vivo stability of
tetrahydropyrimidine also needs to be verified. Taken to-
gether, the simple compounds presented showed relatively
high affinity to the M receptor. The data here, therefore, are
1
a basis for the further useful studies toward the development
General Procedure for the Preparation of Pyrimidine-5-carbaldehyde
O-Substituted Oximes (2b–2g)
of efficacious M receptor agonists and ultimately the treat-
1
ment of Alzheimer’s disease.
To a stirred solution of pyrimidine-5-carbaldehyde oxime 2 in acetonitrile
was added alkyl halide (1.5 eq) and K2CO3 (2.0 eq). The resulting mixture
was then heated under reflux for 5–6 h and cooled to room temperature,
insolubles were filtered off, and the filtrate was concentrated. The residue
was subjected to silica gel flash chromatography (EtOAc:Hex = 4:1) to give
the titled compounds.
Acknowledgments
We wish to thank the Korea Ministry of Science and Technology (KN
9925-01) and (KK 0003-KO) for financial grants.
(2b): Yield 85%. IR (NaCl, neat) ν: 2950, 1550, 1410.– 1H-NMR
(200 MHz, CDCl3) δ: 9.18 (1H, s, C2-H), 8.92 (2H, s, C4-H, C6-H), 8.04
(1H, s, CH=N), 4.22 (2H, t, J = 6.7 Hz, OCH2), 1.8–1.7 (2H, m, CH2), 1.35
(4H, m, 2 × CH2), 0.93 (3H, m, CH3).– 13C-NMR (75 MHz, CDCl3) δ: 158.9
(C2), 154.7 (C4, C6), 142.2 (C=N), 126.9 (C5), 75.2 (OCH2), 28.8, 28.0, 22.5
(3 × CH2), 14.0 (CH3).– MS m/z: 193.1217 (Calcd for C10H15N3O:
193.1215).
Experimental
Melting points were determined on a Thomas-Hoover apparatus and are
uncorrected. Infrared spectra were recorded on a Shimadzu 435 spectrometer
and Mattson Genesis II FTIR. Nuclear magnetic resonance spectra were
measured on a Brucker AM-300 spectrometer and a Varian Gemini-2000.
Mass spectra were determined on JEOL JMS-DX 303 Mass Spectrometer
JEOL JMA-DA 5000 mass data system focusing high resolution mass
spectrometers.
(2c): Yield 70%. IR (NaCl, neat) ν: 3400, 2900, 1580.– 1H-NMR
(300 MHz, CDCl3) δ: 9.19 (1H, s, CH=N), 8.93 (2H, s, C4-H, C6-H), 8.09
(1H, s, C2-H), 6.04 (1H, ddt, J = 17.0, 10.0, 6.0 Hz, CH=CH2), 5.37 (1H,
Arch. Pharm. Pharm. Med. Chem. 334, 79–85 (2001)

1,2,3,4-Tetrahydropyrimidine Derivatives
83
ddd, J = 17.0, 3.0, 1.5 Hz, C=CHtrans), 5.29 (1H, ddd, J = 10.0, 3.0, 1.1 Hz,
C=CHcis), 4.73 (2H, ddd, J = 6.0, 1.5, 1.1 Hz, OCH2).– 13C-NMR (75 MHz,
CDCl3) δ: 158.9 (C2), 154.7 (C4, C6), 142.8 (C=N), 133.3 (CH=CH2), 125.8
(C5), 118.5 (CH=CH2), 75.8 (OCH2).– MS m/z: 163.0746 (Calcd for
C8H9N3O: 163.0746).
(2d): Yield 83%. mp 76–79 °C. IR (KBr) ν: 3300, 2100, 1580, 940.–
1H-NMR (300 MHz, CDCl3) δ: 9.22 (1H, s, C2-H), 8.96 (2H, s, C4-H, C6-H),
8.13 (1H, s, CH=N), 4.83 (2H, d, J = 2.4 Hz, OCH2), 2.57 (1H, t, J = 2.4 Hz,
C≡CH).– 13C-NMR (75 MHz, CDCl3) δ: 159.6 (C2), 155.3 (C4, C6), 144.6
(C=N), 126.5 (C5), 79.2 (OCH2), 75.7 (C≡CH), 62.8 (C≡CH).– Anal. Calcd.
For C8H7N3O: C, 59.62; H, 4.38; N, 26.07. Found: C, 59.34; H, 4.35; N,
26.07.
brs, NH), 6.82 (1H, d, C6-H), 5.91 (1H, ddt, J = 17.0, 10.0, 6.0 Hz, CH=CH2),
5.23 (1H, ddd, J = 17.0, 3.0, 1.5 Hz, C=CHtrans), 5.14 (1H, ddd, J = 10.0,
3.0, 1.1 Hz, C=CHcis), 4.50 (brs, COOH), 4.39 (2H, ddd, J = 6.0, 1.5, 1.1
Hz, OCH2), 4.16 (2H, s, C2-H), 3.61 (2H, s, C4-H), 2.62 (3H, s, NCH3).–
13C-NMR (75 MHz, DMSO-d6) δ: 164.7 (C=O), 149.0 (C=N), 137.1 (C6),
135.1 (CH=CH2), 117.3 (CH=CH2), 97.2 (C5), 73.8 (OCH2), 61.4 (C2), 49.2
(C4), 38.9 (NCH3).– MS m/z: 181.1213 (Calcd for C9H15N3O: 181.1215).
(4d): Yield 31%. mp >135 °C (decomp). IR (KBr) ν: 3400, 3300, 2920,
1
1640.– H-NMR (200 MHz, DMSO-d6) δ: 7.71 (1H, s, CH=N), 7.09 (1H,
brs, NH), 6.87 (1H, s, C6-H), 5.16 (brs, COOH), 4.50 (2H, d, J = 2.4 Hz,
OCH2), 4.18 (2H, s, C2-H), 3.62 (2H, s, C4-H), 3.38 (1H, t, J = 2.4 Hz,
C≡CH), 2.63 (3H, s, NCH3).– 13C-NMR (75 MHz, DMSO-d6) δ: 164.7
(C=O), 149.9 (C=N), 137.9 (C6), 96.7 (C5), 80.9 (C≡CH ), 77.2 (OCH2),
61.3 (C2), 60.7 (C≡CH), 49.1 (C4), 38.9 (NCH3).– MS m/z: 179.1055 (Calcd
for C9H13N3O: 179.1059).
(2e): Yield 60%. mp 42–44 °C. IR (neat) ν: 3000, 1580, 1410.– 1H-NMR
(300 MHz, CDCl3) δ: 9.17 (1H, s, CH=N), 8.89 (2H, s, C4-H, C6-H), 8.07
(1H, s, C2-H), 7.35–7.40 (5H, m, Ph-H), 5.24 (2H, s, OCH2).– 13C-NMR (75
MHz, CDCl3) δ: 158.8 (C2), 154.6 (C4, C6), 142.9 (C=N), 136.7 (C1′), 128.4
(C2′, C3′), 128.1 (C4′), 126.4 (C5), 76.9 (OCH2).– Anal. Calcd. For
C12H11N3O: C, 67.59; H, 5.20; N, 19.71. Found: C, 67.47; H, 5.23; N, 19.48.
(2f): Yield 69%. mp 85–88 °C. IR (KBr) ν: 2900, 1610, 1580, 1000.–
1H-NMR (300 MHz, CDCl3) δ: 9.17 (1H, s, CH=N), 8.89 (2H, s, C4-H,
C6-H), 8.06 (1H, s, C2-H), 7.20 (4H, 2×d, J = 8.0 Hz, Ph-H), 5.20 (2H, s,
OCH2), 2.35 (3H, s, CH3).– 13C-NMR (75 MHz, CDCl3) δ: 158.8 (C2), 154.6
(C4, C6), 142.9 (C=N), 138.1 (C1′), 133.6 (C4′), 129.1 (C3′), 128.6 (C2′),
126.5 (C5), 76.9 (OCH2), 21.1 (CH3).– Anal. Calcd. For C13H13N3O: C,
68.70; H, 5.77; N, 18.49. Found: C, 68.26; H, 5.93; N, 18.50.
(4e): Yield 24%. mp >148 °C (decomp). IR (KBr) ν: 3400, 3280, 3030,
1
1640.– H-NMR (300 MHz, DMSO-d6) δ: 7.74 (1H, s, CH=N), 7.32 (5H,
m, Ph-H), 7.05 (1H, brs, NH), 6.84 (1H, s, C6-H), 5.3 (brs, COOH), 4.93
(2H, s, OCH2), 4.18 (2H, s, C2-H), 3.63 (2H, s, C4-H), 2.63 (3H, s, NCH3).–
13C-NMR (75 MHz, DMSO-d6) δ: 164.6 (C=O), 149.3 (C=N), 138.4 (C1′),
137.3 (C6), 128.5, 128.2, 127.8 (C2′, C3′, C4′), 97.2 (C5), 74.9 (OCH2), 61.4
(C2), 49.2 (C4), 38.9 (NCH3).– MS m/z: 231.1369 (Calcd for C13H17N3O:
231.1372).
(4f): Yield 19%. mp >146 °C (decomp). IR (KBr) ν: 3420, 3300, 2930,
1640.– 1H-NMR (300 MHz, DMSO-d6) δ: 9.34 (brs, COOH), 7.74 (1H, s,
CH=N), 7.27 (5H, 2×d, brs, Ph-H, NH), 6.85 (1H, s, C6-H), 4.88 (2H, s,
OCH2), 4.25 (2H, s, C2-H), 3.70 (2H, s, C4-H), 2.68 (3H, s, NCH3), 2.27
(3H, s, CH3).–13C-NMR (75 MHz, DMSO-d6) δ: 164.7 (C=O), 149.1 (C=N),
137.2 (C6), 136.9 (C1′), 135.3 (C4′), 129.0, (C2′), 128.4, (C3′), 97.2 (C5),
74.8 (OCH2), 61.4 (C2), 49.2 (C4), 38.9 (NCH3), 21.0 (CH3).– MS m/z:
245.1530 (Calcd for C14H19N3O: 245.1528).
(4g): Yield 20%. mp >149 °C (decomp). IR (KBr) ν: 3410, 3280, 2930,
1640.– 1H-NMR (300 MHz, DMSO-d6) δ: 8.53 (brs, COOH), 7.76 (1H, s,
CH=N), 7.36 (4H, 2×d, Ph-H), 7.21 (1H, brs, NH), 6.86 (1H, s, C6-H), 4.92
(2H, s, OCH2), 4.25 (2H, s, C2-H), 3.68 (2H, s, C4-H), 2.67 (3H, s, NCH3).–
13C-NMR (75 MHz, DMSO-d6) δ: 164.6 (C=O), 149.5 (C=N), 137.5 (C1′),
137.4 (C6), 132.3 (C4′), 129.9, (C2′), 128.5, (C3′), 97.1 (C5), 73.9 (OCH2),
61.4 (C2), 49.1 (C4), 38.9 (NCH3).– MS m/z: 265.0981 (Calcd for
C13H16N3OCl: 265.0982).
(2g): Yield 75%. mp 75–77 °C. IR (KBr) ν: 2950, 1600, 1550.– 1H-NMR
(300 MHz, CDCl3) δ: 9.18 (1 H, s, CH=N), 8.89 (2H, s, C4-H, C6-H), 8.08
(1H, s, C2-H), 7.34 (4H, s, Ph-H), 5.20 (2H, s, OCH2).– 13C-NMR (75 MHz,
CDCl3) δ: 158.9 (C2), 154.7 (C4, C6), 143.3 (C=N), 135.2 (C1′), 133.9 (C4′),
129.7 (C3′), 128.6 (C2′), 126.3 (C5), 76.0 (OCH2).– Anal. Calcd. For
C12H10N3OCl: C, 58.19; H, 4.07; N, 16.97. Found: C, 57.95; H, 3.91; N,
16.64.
General Procedure for the Preparation of 3-Methyl-1,2,3,4-tetrahy-
dropyrimidine-5-carbaldehyde O-Substituted Oxime Oxalate (4a–4g)
To a stirred solution of pyrimidine-5-carbaldehyde O-alkyl-oxime 2 in
acetone (5 ml/mmol) was added MeI (12.0 eq), then the mixture was heated
under reflux for 48–72 h. Yellow precipitates were collected by filtration and
dried under vacuum. Attempted purification by recrystallization from MeOH
resulted in deterioration of the material, thus the crude products were used
in the next step without further purification. The methiodide 3a–3g was
dissolved in MeOH (5ml/mmol) then cooled to –30 °C. To this was added
portions of NaBH4 (1.2 eq) maintaining the temperature below –20 °C. The
reaction was quenched with a small amount of water then the solvent was
evaporated in vacuo. The residue was dissolved in 50 ml CH2Cl2 then washed
with water, dried over MgSO4 then concentrated. The residue was subjected
to silica gel flash chromatography (MeOH:EtOAc = 1:9) to give a deep
red-brown syrup. To a stirred solution of above syrup in 2 ml MeOH was
added a solution of oxalic acid (1.0 eq) in 1.5 ml MeOH. After 1 h stirring
at room temperature, the precipitates were filtered off to afford a pale yellow
powder upon drying under vacuum.
(4a): Yield 18%. mp >135 °C(decomp). IR (KBr) ν: 3340, 2900, 1640.–
1H-NMR (200 MHz, DMSO-d6) δ: 7.65 (1H, s, CH=N), 6.97 (1H, brs, NH),
6.82 (1H, s, C6-H), 5.40 (COOH), 4.17 (2H, s, C2-H), 3.66 (3H, s, OCH3),
3.62 (2H, s, C4-H), 2.62 (3H, s, NCH3).– 13C-NMR (50 MHz, DMSO-d6) δ:
164.7 (C=O), 148.7 (C=N), 137.0 (C6), 97.1 (C5), 61.3 (C2), 61.0 (OCH3),
49.1 (C4), 38.9 (NCH3).– MS m/z: 155.1056 (Calcd for C7H13N3O:
155.1059).
1-(5-Pyrimidinyl)-1-ethanol (5)
To a stirred solution of pyrimidine-5-carbaldehyde 1 (1.50 g, 13.87 mmol)
in dry ether (10 ml) at 0 °C was added dropwise CH3MgBr (5.09 ml, 3.0 M
in ether). The resulting suspension was stirred for another 1 h at room
temperature then cooled to 0 °C. A saturated aqueous solution of NH4Cl was
added until the cloudy mixture became clear. The solvent was evaporated,
and the residue was mixed well with silica gel then subjected to flash
chromatography on silica gel (EtOAc) to afford 5 (1.15 g, 67%) as a clear
light yellow oil. IR (NaCl, neat) ν: 3300 (OH), 2950, 1560.– 1H-NMR (300
MHz, CDCl3) δ: 9.05 (1H, s, C2-H), 8.74 (2H, s, C4-H, C6-H), 5.00 (1H, q,
J = 6.8 Hz, CH(OH)CH3), 4.38 (1H, brs, OH), 1.57 (3H, d, J = 6.8 Hz,
CH(OH)CH3).– 13C-NMR (75 MHz, CDCl3) δ: 157.2 (C2), 154.3 (C4, C6),
138.8 (C5), 65.7 (CHOH), 24.9 (CH3).– MS m/z: 124.0635 (Calcd for
C6H8N2O: 124.0637).
1-(5-Pyrimidinyl)-1-ethanone (6)
1-(5-Pyrimidinyl)-1-ethanol 5 (0.31 g, 2.50 mmol) was dissolved in 15 ml
CH2Cl2 then to this was added 4Å Molecular sieves powder (2.00 g,
0.80 g/mmol), activated overnight at 80 °C), pyridinium dichromate (0.94 g,
2.50 mmol), and acetic acid (0.25 ml, 4.37 mmol) in the order. After 20 min
stirring at room temperature, filtered through a Celite pad, the filter cake was
washed well with CH2Cl2 then the filtrate was washed with aq. NaHCO3
solution. The organic layer was dried over MgSO4 then concentrated. The
residue was subjected to flash chromatography on silica gel (EtOAc:Hex =
1:1) to afford 6 (0.20 g, 67%) as a white solid. mp 80–81 °C. IR (KBr) ν:
3000, 1680, 1280.– 1H-NMR (300 MHz, CDCl3) δ: 9.39 (1H, s, C2-H), 9.25
(2H, s, C4-H, C6-H), 2.69 (3H, s, CH3).– 13C-NMR (75 MHz, CDCl3) δ:
(4b): Yield 10%. mp>125 (decomp). IR (KBr) ν: 3370, 2940, 1640.–
1H-NMR (200 MHz, DMSO-d6) δ: 7.66 (1H, s, CH=N), 6.95 (1H, brs, NH),
6.80 (1H, s, C6-H), 5.50 (brs, COOH), 4.17 (2H, s, C2-H), 3.85 (2H, t,
OCH2), 3.63 (2H, s, C4-H), 2.63 (3H, s, NCH3), 1.54 (2H, m, CH2), 1.26
(4H, m, 2×CH2), 0.85(3H, t, CH3).– 13C-NMR (50 MHz, DMSO-d6) δ: 164.5
(C=O), 148.3 (C=N), 136.7 (C6), 97.5 (C5), 72.9 (OCH2), 61.4 (C2), 49.2
(C4), 38.9 (NCH3), 28.5, 28.0, 22.2(3×CH2), 14.2 (CH3).– MS m/z: 211.1683
(Calcd for C11H21N3O: 211.1685).
(4c): Yield 20%. mp >140 °C(decomp). IR (KBr) ν: 3400, 3290, 2930,
1
1640.– H-NMR (300 MHz, DMSO-d6) δ: 7.70 (1H, s, CH=N), 6.98 (1H,
Arch. Pharm. Pharm. Med. Chem. 334, 79–85 (2001)

84
Jung, Park, Yang, and Lee
(7f): Yield 92%. mp 62–64 °C. IR (KBr) ν: 2920, 1600, 1410.– 1
H-NMR
(300 MHz, CDCl3) δ: 9.16 (1H, s, C2-H), 8.96 (2H, s, C4-H, C6-H), 7.24
(4H, ABq, Ph-H), 5.21 (2H, s, OCH2), 2.34 (3H, s, CH3), 2.23 (3H, s,
=CCH3).– 13C-NMR (75 MHz, CDCl3) δ: 158.3 (C2), 153.9 (C4, C6), 149.6
(C=N), 137.7 (C1′), 134.1 (C4′), 129.9 (C5), 129.0 (C3′, C5′), 128.4 (C2′,
C6′), 76.6 (OCH2), 21.1 (Ph-CH3), 11.8 (N=CCH3).– Anal.: C14H15N3O.C,
69.69; H, 6.27; N, 17.41. Found: C, 69.54; H, 6.38; N, 17.36
194.9 (C=O), 161.3 (C2), 156.6 (C4, C6), 129.4 (C5), 26.6 (CH3).– MS m/z:
122.0480 (Calcd for C6H6N2O: 122.0481).
1-Pyrimidin-5-yl-ethanone Oxime (7)
Hydroxylmine hydrochloride (1.24 g, 17.81 mmol) was neutralized with
saturated aqueous NaHCO3 solution and then added to a stirred solution of
1-(5-pyrimidinyl)-1-ethanone 6 (1.45 g, 11.87 mmol) in 50 ml benzene. The
resulting mixture was heated under reflux for 2 h using a Dean-Stark column.
The solvent was evaporated in vacuo, the residue was mixed well with silica
gel then was subjected to flash chromatography on silica gel (EtOAc:Hex =
1:1) to give 7 (1.28 g, 78%) as a white solid. mp 140–141 °C. IR (KBr) ν:
3150 (OH), 2850, 1550.– 1H-NMR (300 MHz, CD3OD) δ: 9.12 (1H, s,
C2-H), 9.06 (2H, s, C4-H, C6-H), 2.28 (3H, s, CH3).– 13C-NMR (75 MHz,
CD3OD) δ: 158.9 (C2), 156.6 (C4, C6), 150. 8 (C=N), 132.9 (C5), 11.2
(N=CCH3).
(7g): Yield 77%. mp 61–63 °C. IR (KBr) ν: 3000, 1610, 1410.– 1H-NMR
(200 MHz, CDCl3) δ: 9.17 (1H, s, C2-H), 8.96 (2H, s, C4-H, C6-H), 7.32
(4H, s, Ph-H), 5.21 (2H, s, OCH2), 2.26 (3H, s, CH3).– 13C-NMR (50 MHz,
CDCl3) δ: 158.4 (C2), 153.8 (C4, C6), 150.1 (C=N), 135.7 (C1′), 133.7 (C4′),
129.7 (C5), 129.5 (C3′), 128.4 (C2′), 75.7 (OCH2), 11.8 (N=CCH3).– Anal.:
C13H12N3OCl, C, 59.66; H, 4.62; N, 16.06. Found: C, 59.66; H, 4.73; N,
16.10.
General Procedure for the Preparation of 1-(3-Methyl-1,2,3,4-tetrahydro-
pyrimidin-5-yl)ethanone O-Substituted Oxime Oxalate (9a–9g)
1-Pyrimidin-5-yl-ethanone O-Methyloxime (7a)
To a stirred solution of 1-pyrimidin-5-yl-ethanone O-substituted oxime
7a–7g in acetone (5 ml/mmol) was added CH3I (12.0 eq), and the mixture
was then heated under reflux for 48–72 h. Yellow precipitates were collected
by filtration and dried under vacuum. Attempted purification by recrystalli-
zation from CH3OH resulted in deterioration of the material, thus used for
the next step without further treatment. The resulting methiodide 8a–8g was
dissolved in CH3OH (5ml/mmol) then cooled to –30 °C. To this was added
portions of NaBH4 (1.2 eq) maintaining the temperature below –30 °C. The
reaction was allowed to come to room temperature, then the solvent was
evaporated in vacuo. The residue was dissolved in 50 ml CH2Cl2 then washed
with water, organic layer was dried over MgSO4 then concentrated. The
residue was subjected to purification by HPLC (ODS-S-50B, CH3OH:H2O
= 19:1) to give a deep red-brown syrup. The product is easily decomposed
during evaporation of the solvents, thus the bath temperature should be no
higher than 20 °C. The oily product was used immediately after drying. To
a stirred solution of above syrup in CH3OH (1–2 ml) was added a solution
of oxalic acid (1.0 eq) in CH3OH (0.5–1 ml). After 1 h stirring at room
temperature the precipitates were collected by filtration to afford a pale
yellow powder upon drying under vacuum.
To a stirred solution of methoxylamine hydrochloride (0.75 g, 8.98 mmol)
in 30 ml methanol was added 0.83 g (9.83 mmol) of solid NaHCO3 then the
mixture was stirred for 30 min at room temperature. To this was added
1-pyrimidin-5-yl-ethanone 6 (0.50 g, 4.09 mmol) then stirring was continued
for 1 h at the same temperature. The reaction mixture was dissolved in water,
extracted with CH2Cl2, the organic layer was dried over MgSO4, then
concentrated. The residue was subjected to flash chromatography on silica
gel (EtOAc:Hex = 1:4) to give 7a (0.39 g, 63%) as a white solid. mp
38–39 °C. IR (KBr) ν: 2950, 1610, 1420.– 1H-NMR (200 MHz, CDCl3) δ:
8.94 (1H, s, C2-H), 8.75 (2H, s, C4-H, C6-H), 3.79 (3H, s, OCH3), 1.99 (3H,
s, N=CCH3).– 13C-NMR (50 MHz, CDCl3) δ:158.1 (C2), 153.6 (C4, C6),
149.1 (C=N), 129.6 (C5), 61.9 (OCH3), 11.3 (N=CCH3).– MS m/z: 151.0746
(Calcd for C7H9N3O: 151.0745).
General Procedure for the Preparation of 1-Pyrimidin-5-yl-ethanone
O-Substituted Oxime (7b–7g)
A mixture of 1-pyrimidin-5-yl-ethanone oxime 7 (2.00 mmol), K2CO3
(4.00 mmol), and alkyl halide (3.00 mmol) in 10 ml acetonitrile was heated
under reflux for 5–6 h. Insolubles were filtered off, the filtrate was concen-
trated. The residue was subjected to flash chromatography on silica gel
(EtOAc:Hex = 1:1) to give the title compounds.
(9a): Yield 10%. mp 141–142 °C. IR (KBr) ν: 3310, 2930, 2610, 1640.–
1H-NMR (300 MHz, DMSO-d6) δ: 8.56 (brs, COOH), 6.92 (2H, s, C6-H,
NH), 4.23 (2H, s, C2-H), 3.76 (2H, s, C4-H), 3.71 (3H, s, OCH3), 2.69 (3H,
s, NCH3), 1.85 (3H, s, CH3).– 13C-NMR (75 MHz, DMSO-d6) δ: 164.9
(C=O), 153.1 (C=N), 132.8 (C6), 99.7 (C5), 61.23 (OCH3), 61.16 (C2), 49.8
(C4), 38.7 (NCH3), 9.9 (N=CCH3).– MS m/z: 169.1215 (Calcd for
C8H15N3O: 169.1215).
1
(7b): Yield 68%. IR (NaCl, neat) ν: 2950, 1610, 1570.– H-NMR (300
MHz, CDCl3) δ: 9.18 (1H, s, C-2 H), 8.99 (2H, s, C4-H, C6-H), 4.23 (2H, t,
J = 6.8 Hz, OCH2), 2.25 (3H, s, CH3), 1.74 (2H, m, CH2), 1.39 (4H, m, CH2),
0.93 (3H, t, J = 6.8 Hz, CH2CH3).– 13C-NMR (75 MHz, CDCl3) δ: 158.3
(C2), 153.9 (C4, C6), 148.9 (C=N), 130.2 (C5), 74.9 (OCH2), 28.8 (C3′),
28.0 (C2′), 22.4 (C4′), 13.9 (CH3), 11.7 (N=CCH3).
1
(9b): Yield 13%. mp 127–129 °C. IR (KBr) ν: 3295, 2930, 1640.– H-
NMR (300 MHz, DMSO-d6) δ: 8.96 (brs, COOH), 6.90 (2H, s, C6-H, NH),
4.23 (2H, s, C2-H), 3.91 (2H, t, OCH2), 3.76 (2H, s, C4-H), 2.69 (3H, s,
NCH3), 1.85 (3H, s, CH3), 1.56 (2H, m, CH2), 1.27 (4H, m, 2×CH2), 0.85
(3H, t, CH3).– 13C-NMR (75 MHz, DMSO-d6) δ: 164.9 (C=O), 152.7 (C=N),
132.5 (C6), 100.1 (C5), 73.1 (OCH2), 61.2 (C2), 49.8 (C4), 38.7 (NCH3),
28.6, 27.9, 22.2 (C2′, C3′, C4′), 14.2 (CH3), 9.9 (N=CCH3).– MS m/z:
225.1839 (Calcd for C12H23N3O: 225.1841).
(9c): Yield 26%. mp 139–140 °C. IR (KBr) ν: 3250, 2990, 2615, 1640.–
1H-NMR (300 MHz, DMSO-d6) δ: 8.80 (brs, COOH), 6.93 (2H, s, C6-H,
NH), 5.95 (1H, m, CH=CH2), 5.20 (2H, m, CH=CH2), 4.45 (2H, d, OCH2),
4.24 (2H, s, C2-H), 3.76 (2H, s, C4-H), 2.69 (3H, s, NCH3), 1.88 (3H, s,
CH3).– 13C-NMR (75 MHz, DMSO-d6) δ: 164.8 (C=O), 153.3 (C=N), 135.1
(CH=CH2), 132.8 (C6), 117.2 CH=CH2), 99.8 (C5), 74.0 (OCH2), 61.1 (C2),
49.8 (C4), 38.7 (NCH3), 10.1 (N=CCH3).– MS m/z: 195.1371 (Calcd for
1
(7c): Yield 83%. IR (NaCl, neat) ν: 2900, 1610, 1550.– H-NMR (300
MHz, CDCl3) δ: 9.19 (1H, s, C2-H), 8.99 (2H, s, C4-H, C6-H), 6.05 (1H, 2
× dt, J = 17.0, 10.0, 6.0 Hz, CH=CH2), 5.35 (1H, ddd, J = 17.0, 3.0, 1.5 Hz,
C=CHtrans), 5.27 (1H, dd, J = 10.0, 3.0 Hz, C=CHcis), 4.74 (2H, dt, J = 6.0,
1.5 Hz, OCH2), 2.28 (3H, s, CH3).– 13C-NMR (75 MHz, CDCl3) δ: 158.5
(C2), 154.0 (C4, C6), 149.7 (C=N), 133.8 (CH=CH2), 130.1 (C5), 117.9
(CH=CH2), 75.6 (OCH2), 11.9 (N=CCH3).– MS m/z: 177.0903 (Calcd for
C9H11N3O: 177.0902).
(7d): Yield 94%. mp 46–47 °C. IR (KBr) ν: 3250, 3030, 1630.– 1H-NMR
(300 MHz, CDCl3) δ: 9.20 (1H, s, C2-H), 9.00 (2H, s, C4-H, 6-H), 4.84 (2H,
d, J = 2.2 Hz, CH2C≡CH), 2.54 (1H, t, J = 2.2 Hz, CH2C≡CH), 2.29 (3H, s,
CH3).– 13C-NMR (75 MHz, CDCl3) δ: 158.7 (C2), 154.1 (C4, C6), 151.1
(C=N), 129.7 (C5), 79.3 (C≡CH), 74.8 (C≡CH), 62.1 (OCH2), 12.1
(N=CCH3).– Anal.: C9H9N3O, C, 61.70; H, 5.18; N, 23.99. Found: C, 61.68;
H, 5.14; N, 23.91.
C
10H17N3O: 195.1372).
(9d): Yield 28%. mp 133–134 °C. IR (KBr) ν: 3335, 2920, 2680, 1640.–
1H-NMR (300 MHz, DMSO-d6) δ: 9.83 (brs, COOH), 7.07 (1H, brs, NH),
6.97 (1H, s, C6-H), 4.56 (2H, d, J = 2.0 Hz, OCH2), 4.24 (2H, s, C2-H), 3.75
(2H, s, C4-H), 3.39 (1H, t, J = 2.0 Hz, C≡CH), 2.70 (3H, s, NCH3), 1.87 (3H,
s, CH3).– 13C-NMR (75 MHz, DMSO-d6) δ: 164.9 (C=O), 153.4 (C=N),
133.5 (C6), 99.2 (C5), 81.0 (C≡CH), 77.2 (C≡CH), 61.1 (C2), 60.9 (OCH2),
49.7 (C4), 38.8 (NCH3), 10.1 (N=CCH3).– MS m/z: 193.1214 (Calcd for
(7e): Yield 56%. mp 28–31 °C. IR (KBr) ν: 3010, 1650, 1420.– 1H-NMR
(300 MHz, CDCl3) δ: 9.18 (1H, s, C2-H), 8.97 (2H, s, C4-H, C6-H), 7.37
(5H, m, Ph-H), 5.26 (2H, s, OCH2), 2.26 (3H, s, CH3).– 13C-NMR (75 MHz,
CDCl3) δ: 158.4 (C2), 156.1 (C4 , C6), 149.8 (C=N), 137.2 (C1′), 129.9 (C5),
128.4 (C3′, C5′), 128.1 (C4′), 127.9 (C2′, C6′), 76.6 (OCH2), 11.9
(N=CCH3).
C
10H15N3O: 193.1215).
Arch. Pharm. Pharm. Med. Chem. 334, 79–85 (2001)

1,2,3,4-Tetrahydropyrimidine Derivatives
85
(9e): Yield 8%. mp 140–142 °C. IR (KBr) ν: 3300, 3030, 2860, 1645.–
1H-NMR (300 MHz, DMSO-d6) δ: 8.74 (brs, COOH), 7.33 (5H, m, Ph-H),
6.94 (2H, s, C6-H, NH), 5.00 (2H, s, OCH2), 4.24 (2H, s, C2-H), 3.76 (2H,
s, C4-H), 2.69 (3H, s, NCH3), 1.90 (3H, s, CH3).– 13C-NMR (75 MHz,
DMSO-d6) δ: 164.7 (C=O), 153.6 (C=N), 138.4, 128.5, 128.1, 127.8 (Ph-C),
132.9 (C6), 99.7 (C5), 75.0 (OCH2), 61.1 (C2), 49.8 (C4), 38.7 (NCH3), 10.2
(N=CCH3).– MS m/z: 245.1528 (Calcd for C14H19N3O: 245.1528).
(9f): Yield 17%. mp 140–142 °C. IR (KBr) ν: 3230, 2965, 2550, 1640.–
1H-NMR (300 MHz, DMSO-d6) δ: 7.92 (COOH), 7.17 (4H, ABq, Ph-H),
7.00 (1H, brs, NH), 6.93 (1H, s, C6-H), 4.94 (2H, s, OCH2), 4.24 (2H, s,
C2-H), 3.76 (2H, s, C4-H), 2.70 (3H, s, NCH3), 2.27 (3H, s, Ph-CH3), 1.88
(3H, s, CH3).–13C-NMR (75MHz, DMSO-d6) δ: 164.9 (C=O), 153.4 (C=N),
132.9 (C6), 137.0, 135.4, 129.1, 128.3 (Ph-C), 99.8 (C5), 74.9 (OCH2), 61.1
(C2), 49.8 (C4), 38.7 (NCH3), 21.0 (Ph-CH3), 10.2 (N=CCH3).– MS m/z:
259.1680 (Calcd for C15H21N3O: 259.1685).
(9g): Yield 10%. mp 133–135 °C. IR (KBr) ν: 3290, 3015, 2965, 1640.–
1H-NMR (300 MHz, DMSO-d6) δ: 7.36 (6H, brs, m, COOH, Ph-H), 7.01
(1H, brs, NH), 6.95 (1H, s, C6-H), 4.98 (2H, s, OCH2), 4.24 (2H, s, C2-H),
3.74 (2H, s, C4-H), 2.69 (3H, s, NCH3), 1.90 (3H, s, CH3).– 13C-NMR (75
MHz, DMSO-d6) δ: 164.8 (C=O), 153.8 (C=N), 133.1(C6), 137.6, 132.3,
129.9, 128.5 (Ph-C), 99.6 (C5), 74.1 (OCH2), 61.1 (C2), 49.7 (C4), 38.7
(NCH3), 10.2 (N=CCH3).– MS m/z: 279.1136 (Calcd for C14H18N3OCl:
279.1138).
References
✩
Dedicated to the memory of Professor Dr. Klaus Hartke.
[1] a) E. K. Moltzen, B. Bjorholm, Drugs Future 1995, 20, 37–54.
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Biological Methods
Receptor Binding Assay. Binding of compounds to muscarinic M1 recep-
tor (M1) was performed by the radioligand binding assay as described
previously[11,12]. Binding affinity was determined indirectly by the ability of
compound to compete with 1 nM [3H]-N-methylscopolamine (NMS) in the
suspension of cloned human M1 (h-1M) expressed in CHO cells (Biosignal).
Nonspecific binding was evaluated by the inclusion of 1 µM atropine in a
separate set of samples. Each sample contained approximately 16 µg of
protein (h-M1) in 50 mM Tris buffer (pH 7.2) and varying concentrations of
each compound in a final volume of 250 µl. Samples were incubated for 1 h
at 27 °C in shaking incubator and then filtered through Wallac glass fiber
filtermat (GF/C) using Inotech cell harvester of 96-well format. The radio-
activity bound to filter was counted by Micro-β counter (Wallac).
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Received: September 1, 2000 [FP520]
Arch. Pharm. Pharm. Med. Chem. 334, 79–85 (2001)