The new generation dihydropyridine type calcium blockers, bearing 4-phenyl oxypropanolamine, display α-/β-adrenoceptor antagonist and long-acting antihypertensive activities

Article abstract of DOI:10.1016/S0968-0896(01)00318-2

A new series of dihydropyridine derivatives, bearing oxypropanolamine moiety on phenyl ring at the 4-position of the dihydropyridine base, were prepared. Oxypropanolamine was synthesized by replacing the phenolic OH of vanillin or other compounds, having a phenyl aldehyde group, with epichlorohydrin, followed by cleavaging the obtained epoxide compounds with tert-butylamine, n-butylamine or 2-methoxy-1-oxyethylamino benzene (guaiacoxyethylamine), respectively. Obtained various oxypropanolamine compounds, still remaining a phenyl aldehyde moiety, were then performed by Hantzsch condensation reaction with methylacetoacetate or ethylacetoacetate, respectively, to give our new series of dihydropyridine linked with the 4-phenyl ring. These compounds were evaluated for inotropic, chronotropic, and aorta contractility that associated with calcium channel and adrenoceptor antagonist activities. Dihydropyridine derivatives that with oxypropanolamine side chain on their 4-phenyl ring associated α-/β-adrenoceptor blocking activities created a new family of calcium entry and the third generation β-adrenoceptor blockers. Optimizing this research to obtain more potent α-/β-adrenoceptor blocking and long-acting antihypertensive oxypropanolamine on the 4-phenyl ring of dihydropyridine series compounds was thus accomplished and classified as third generation dihydropyridine type calcium channel blockers, in comparison with previous short-acting type nifedipine and long-acting type amlodipine. We concluded that compounds 1a, 1b and 1g showed not only markedly high calcium-antagonistic activity but also the highest antihypertensive effect; compounds 1b, 1c, 1f, 1g, 1i and 1j induced sustained antihypertensive effects are major and attributed to their calcium entry and α-adrenoceptor blocking activities in the blood vessel due to their introduction of 2-methoxy, 1-oxyethylamino benzene moiety in the side chain on the 4-phenyl ring of dihydropyridine. Bradycardiac effects of all the compounds 1a-1j resulted from calcium entry and β-adrenoceptor blocking, which attenuate the sympathetic activation-associated reflex tachycardia in the heart. We selected compound 1b as candidate compound for further pharmacological and pre-clinical evaluation studies.

Full text of DOI:10.1016/S0968-0896(01)00318-2

Bioorganic & Medicinal Chemistry 10 (2002) 719–730
The New Generation Dihydropyridine Type Calcium Blockers,
Bearing 4-Phenyl Oxypropanolamine, Display ꢀ-/ꢁ-Adrenoceptor
Antagonist and Long-Acting Antihypertensive Activities
Jhy-Chong Liang, Jwu-Lai Yeh, Chia-Sui Wang, Shwu-Fen Liou,
Chieh-Ho Tsai and Ing-Jun Chen*
Department of Pharmacology, Kaohsiung Medical University, 100 Shih-Chuan 1st Road, Kaohsiung 807, Taiwan, ROC
Received 11 May 2001; accepted 10 September 2001
Abstract—A new series of dihydropyridine derivatives, bearing oxypropanolamine moiety on phenyl ring at the 4-position of the
dihydropyridine base, were prepared. Oxypropanolamine was synthesized by replacing the phenolic OH of vanillin or other com-
pounds, having a phenyl aldehyde group, with epichlorohydrin, followed by cleavaging the obtained epoxide compounds with tert-
butylamine, n-butylamine or 2-methoxy-1-oxyethylamino benzene (guaiacoxyethylamine), respectively. Obtained various oxypro-
panolamine compounds, still remaining a phenyl aldehyde moiety, were then performed by Hantzsch condensation reaction with
methylacetoacetate or ethylacetoacetate, respectively, to give our new series of dihydropyridine linked with the 4-phenyl ring. These
compounds were evaluated for inotropic, chronotropic, and aorta contractility that associated with calcium channel and adreno-
ceptor antagonist activities. Dihydropyridine derivatives that with oxypropanolamine side chain on their 4-phenyl ring associated
a-/b-adrenoceptor blocking activities created a new family of calcium entry and the third generation b-adrenoceptor blockers.
Optimizing this research to obtain more potent a-/b-adrenoceptor blocking and long-acting antihypertensive oxypropanolamine on
the 4-phenyl ring of dihydropyridine series compounds was thus accomplished and classified as third generation dihydropyridine
type calcium channel blockers, in comparison with previous short-acting type nifedipine and long-acting type amlodipine. We
concluded that compounds 1a, 1b and 1g showed not only markedly high calcium-antagonistic activity but also the highest anti-
hypertensive effect; compounds 1b, 1c, 1f, 1g, 1i and 1j induced sustained antihypertensive effects are major and attributed to their
calcium entry and a-adrenoceptor blocking activities in the blood vessel due to their introduction of 2-methoxy, 1-oxyethylamino
benzene moiety in the side chain on the 4-phenyl ring of dihydropyridine. Bradycardiac effects of all the compounds 1a–1j resulted
from calcium entry and b-adrenoceptor blocking, which attenuate the sympathetic activation-associated reflex tachycardia in the
heart. We selected compound 1b as candidate compound for further pharmacological and pre-clinical evaluation studies. # 2002
Elsevier Science Ltd. All rights reserved.
Introduction
able effects is the reflex tachycardia produced by dihy-
dropyridines.7 This trouble is also found in the use of an
a-adrenoceptor blocker for enhancing calcium channel
blocking activity.8 Obviously, a dihydropyridine type
calcium channel and a-adrenoceptor blockers without
revealing reflex tachycardia activity is an important tar-
get in the research of new drugs in this field. To resolve
this problem, b-adrenoceptor blockade is suggested to
combine with calcium channel or a-adrenoceptor
blockade to reduce their vasodilatory effects associated
tachycardia reflex.9ꢀ12 Chemical incorporation of a
dihydropyridine type calcium channel blocker and a-/b-
adrenoceptor blocker molecule to reveal these three
blockades is suggested to be reasonable in this study.
Our series of compounds in this study are the hybrid
products of these three blockades.
Dihydropyridine drugs, such as nifedipine, nicardipine,
amlodipine and others, are clinically effective cardio-
vascular agents for the treatment of hypertension and
have been intensively studied to elucidate the molecular
and conformational requirements for their attractive
calcium antagonist activities.1,2
Short-acting dihydropyridine drugs must be used with
caution in patients with coronary heart disease because
of the increased mortality observed in patients treated
with these drugs.3ꢀ6 One of the most discussed undesir-
*Corresponding author. Tel.: +886-7-323-4686; fax: +886-7-323-
4686; e-mail: ingjun@kmu.edu.tw
0968-0896/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(01)00318-2

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J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
Scheme 1.
Scheme 2.
To date, the structure–activity relationship of the dihy-
dropyridines indicated that desired structural char-
acteristic of the substituents at the 4-position of the
dihydropyridine nucleus had been thought to be the
phenyl ring. Modification of the phenyl ring of dihy-
dropyridine compounds demonstrated that the intro-
duction of electron donating substituents, such as the 3-
hydroxy and 2-methoxy group, results in a loss of cor-
onary blood flow and electron-withdrawing sub-
stitutents appear to contribute to increasing or
maintaining the activities.13 An electron-withdrawing
meta substituent in the 4-phenyl ring of dihydropyridine
enhanced relaxing activity but lengthy (not bulky) meta
substituent and any para substituent was described to
reduce this activity.14 In order to obtain additional b- or
a-adrenoceptor blocking activity in one molecule, we
intended to discover appropriate substituents on the
phenyl ring at the 4-position of the dihydropyridines
nucleus. In the beginning of this study, we speculated
that substitution of the hydroxy group on phenyl ring
could be one of the candidates to introduce any moiety
with adrenergic blocking function. However, by repla-
cing the phenolic hydroxyl group of vanilloids with
oxypropanolamine, we have recently succeeded in syn-
thesizing a series of vanilloid based b-adrenoceptor
blockers that with distinguish properties.15ꢀ18 Previous
success prompted us to extend the scope in this research
so we decided to synthesize a new family of dihy-
dropyridines by introducing electron-withdrawing sub-
stituents with a basic property on their 4-phenyl ring,
given that a third generation b-adrenoceptor blocker
carvedilol with additional vasodilatory activity, derived
from a-adrenoceptor and calcium channel entry block-
ing activities, was reported to be useful in the treatment
of hypertension and heart failure.19ꢀ21 In the present
study, we tried to introduce an oxypropanolamine moi-
ety that associated with b-adrenoceptor blocking activ-
ity, on the phenyl ring of dihydropyridines, expecting to
have extra adrenergic antagonist activities. In addition,
new type long-acting antihypertensive dihydropyridines
with adrenoceptor blocking activities were also expected
to resist hydrolysis or metabolism in vivo. Taking into
consideration that nearly all the dihydropyridine drugs
were essentially neutral ester molecule with low aqueous
solubility and marked first-pass effect in the liver,14
we decided to incorporate oxypropanolamine on the
4-position with the phenyl ring that is linked with
dihydropyridine. Our compounds are of basic property,
however, which was not the case as alkoxymethy amine,
substituted on the 2-position of the dihydropyridine
base,22 such as amlodipine. Actually, this decision of
synthesis was also adapted to a previous claim about
basic substitutents, such as found in amlodipine. Our
basic 4-phenyl oxypropanolamine, linked with dihy-
dropyridine, was thus also supposed to display long-
acting antihypertensive activity.23,24
Since guaiacoxyethylamino moiety has been taken as
a1-adrenoceptor blockade-associated functional group
previously,15ꢀ18 in the present study, as shown in
Schemes 1 and 2, an a-blockade-associated guaiacoxy-
ethylamino moiety was also designed to be involved in
our dihydropyridine series to enhance original dihy-
dropyridine-associated calcium channel blocking activ-
ity in vascular smooth muscle. Propanolamine moiety
(related to b-blockade) located, by chemical reaction,
between the phenyl ring and guaiacoxyethylamino
group (related to a-blockade), shown in Schemes 1 and
2, was suggested to have a proper a/b-adrenoceptor
blockade.25ꢀ33 The aim of this study in developing long-

J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
721
acting antihypertensive dihydropyridines was to com-
bine the advantages of both a- and b-adrenoceptor
blockades, calcium channel blocking effect, without the
presence of a tachycardia reflex, and thus to compensate
for the untoward circulatory effect in classical short-
acting dihydropyridine derivatives and a-adrenoceptor
blockers.
replacing the phenolic OH of vanillin with epi-
chlorohydrin, followed by cleaving the obtained epoxide
compound with 2-methoxy, 1-oxyethylamino benzene
(guaiacoxyethylamine), tert-butylamine or n-butyla-
mine, respectively, to yield middle products. Ortho-sub-
stituted oxypropanolamine derivatives were synthesized
by replacing the phenolic OH of hydroxy benzaladhyde
and then repeated as above (2a–2j) (Fig. 1). All obtained
compounds were then reacted with methylacetoacetate
or ethylacetoacetate, followed by Hantzsch condensa-
tion reaction, to obtain various dihydropyridines (com-
pounds 1b–1j) (Fig. 2). In contrast, vanillin was also
directly followed by Hantzsch condensation reaction,
without oxypropanolamine moiety, to present the pos-
sible calcium channel blocking activity as control, in
comparison with compounds with oxypropanolamine
moiety.2
Results
Chemistry
Obtained compounds were represented by the general
Scheme 1 for that with para-substituted oxypropanola-
mine moiety, and Scheme 2 for that with ortho-sub-
stituted oxypropanolamine moiety on the 4-phenyl ring.
The synthesis of compound 1 represents a typical
example of the general synthesis of para-oxypropanola-
mine derivatives of vanillin (Scheme 1) and compound
1f represents an example of the general synthesis of
ortho-substituted oxypropanolamine derivatives of
hydroxy benzaldehyde (Scheme 2), respectively. Para-
substituted oxypropanolamines were synthesized by
Pharmacology
Intravenous compounds 1a–1j (1.0mg/kg) produced
blood pressure lowering and heart rate decreasing
effects in pentobarbital-anesthetized normotensive Wis-
tar rats. Blood pressure lowering effects of compounds
Figure 1. Structures of compounds 1a–1j.
Figure 2. Structures of compounds 2a–2j.

722
J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
1a–1j reached their maximum about 1 min after dosing
and only compounds 1a, and 1b lasted for above 1 h
(Figs 3 and 4). Nifedipine at a dose of 1.0mg/kg iv
produced a temporary fall in blood pressure with a
marked increase in heart rate (Figs 3 and 4).
a potent and long-lasting antihypertensive effect. Com-
pound 1b-induced sustained depressor effect in SHR
might be major and attributed to its calcium entry and
a-adrenoceptor blocking activities in the blood vessel
and partly attributed to b-adrenoceptor/calcium channel
blockade-associated decreased cardiac output in the heart.
Compound 1a is devoid of a-adrenoceptor blocking and
is less potent than compound 1b in anti-hypertensive
activity and a/b ratio, but is more potent than com-
pound 1b in decreasing of mechanical cardiac function.
Two of the most effective compounds 1a and 1b were
selected and their antihypertensive activities were
examined in pentobarbital-anesthetized normotensive
Wistar rats and unanesthetized spontaneous hypertensive
rats (SHR). Compound 1a is reported as ‘‘vanidipinedilol’
and compound 1b is reported as ‘‘labedipinedilol-A’.23,24
These two compounds showed high activity and longer-
lasting actions under these experimental conditions.
Oral administration of compounds 1a and 1b (10–50
mg/kg) in SHR reduced the blood pressure and heart
rate for 24 h. More promisingly, heart rate remained
relatively constant in comparison with blood pressure
changes after administration of compound 1b and
returned to basal value at 24 h. The hypotensive effects
of compounds 1a and 1b (25–50mg/kg) reached max-
imum about 1 h after dosing and lasted over 10h.
Compound 1b was shown to have more potent vasodi-
In the isolated Wistar rat atrium, compounds 1a–1j
(10ꢀ6 M) competitively antagonized (ꢀ)isoprenaline
(10ꢀ10–10ꢀ5 M)-induced positive chronotropic and ino-
tropic effects in a concentration-dependent manner.
Compounds 1a–1j (10ꢀ6 M) also caused a shift to the
right of the (ꢀ)isoprenaline concentration–response
curve. The following order of activity was found for
>
these compounds: 1a>1b>1g>1i>1h>1j>1f 1d>
¼
1e>1c. In electrically driven rat left atrial strips, com-
pounds 1a–1j also antagonized (ꢀ)isoprenaline-induced
positive inotropic responses and produced rightward
shifts of the cumulative concentration–response curves
to (ꢀ)isoprenaline. The following order of activity was
ꢀ1
latory effect due to calcium antagonism with the pKCa
value of 8.46 (nifedipine: pKCa 9.02). Despite its
ꢀ1
>
found for these compounds: 1b>1a>1j 1g>1h>1i>
¼
weaker vasodilatory action than that of nifedipine in
vitro, the oral administration of compound 1b produced
1e>1c>1d>1f. The pA2 values and slope of regression
lines were indicated in Table 1.
Figure 3. Effects of intravenous injection of compounds 1a–1j and
nifedipine (1.0mg/kg) on blood pressure in normotensive Wistar rats,
anesthetized with pentobarbital. Saline was used as a control. Vertical
bars represent SE change from basal line value, which was 114ꢁ5 mm
Hg for mean arterial blood pressure (MBP). Each value represents
meanꢁSEM; n=6.
Figure 4. Effects of intravenous injection of compounds 1a–1j and
nifedipine (1.0mg/kg) on heart rate in normotensive Wistar rats,
anesthetized with pentobarbital. Saline was used as a control. Vertical
bars represent SE change from basal line value, which was 381ꢁ25
beats/min for heart rate. Each value represents meanꢁSEM; n=6.

J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
723
Compounds 1a–1j competitively antagonized the
(ꢀ)isoprenaline-induced relaxation from the sponta-
neous tone of guinea pig tracheal strips. Likewise, com-
pounds 1a–1j produced parallel shifted to the right of
the agonist concentration–response curves. The follow-
ing order of activity was found for these compounds:
>
>
>
1a>1g>1h 1b>1f 1i>1j 1e>1d>1c. The apparent
¼
¼
¼
pA2 values for compounds 1a–1j on tracheal strips were
indicated in Table 1. Compounds 1a, 1b and 1g with
para-substituted tert-butylamino or 2-methoxy, 1-oxy-
ethylamino benzene (guaiacoxyethylamino) terminal
moiety in the side chain on 4-phenyl ring showed the
greater activity in the isolated Wistar rat atrium and
guinea pig tracheal strips than others in these experi-
ments so they had more potent competitive b1- and b2-
blockade activities.
In isolated Wistar rat aorta rings, compounds 1a–1j
(10ꢀ6 M) competitively inhibited norepinephrine-
induced contraction effects, causing a shift to the right
of the norepinephrine concentration–response curves.
The following order of activity was found for these
>
compounds: 1b>1g 1j>1i>1f>1h=1e=1c>1d>1a.
¼
The pA2 values and slope of regression lines were shown
in Table 1. Compounds 1b, 1f, 1g, 1i and 1j with sub-
stitutive guaiacoxyethylamino terminal moiety showed
the greater activity in the isolated Wistar rat aorta rings
than others in these experiments confirming that the
substitution with guaiacoxyethylamino terminal moiety
was found to produce a-adrenoceptor blocking
activity.15ꢀ18
In isolated rat thoracic aortic rings, compounds 1a–1j
produced inhibition or calcium channel blocking of
CaCl2-induced contraction with a pKCa value showed
ꢀ1
in Table 1. The slopes for compounds 1a–1j on the
experiments were not significantly different from unity.
The following order of activity was found for these
compounds: 1i>1h>1b>1j>1d>1g>1f>1a>1e>1c.
Compounds 1h and 1i with substitutive 5-NO2 group on
phenyl ring and compounds 1b and 1j with substitutive
guaiacoxyethylamino terminal moiety in the side chain
on 4-phenyl ring showed the greater activity in the iso-
lated Wistar rat aorta rings than others. On the other
hand, compounds 1a–1j also inhibited high K+-induced
voltage-dependent calcium influx-associated contrac-
tion. The maximum percent relaxation and IC50 values
of compounds 1a–1j on KCl (75 mM)-contracted thor-
acic aortic rings are shown in Table 2. The following
order of activity was found for these compounds:
1g>1b>1h>1i>1a>1e>1j>1f>1d>1c. Substitution
with guaiacoxyethylamino terminal or with NO2 group
on 4-phenyl ring seemed to potentiate the voltage-
dependent relaxing activities in this experiment.
Discussion
Our present series of compounds, bearing oxypropano-
lamine on the 4-phenyl ring, belong to a new family of
dihydropyridine type calcium channel blockers with
additional adrenoceptor blocking properties. The con-
tribution of the channel and receptor blockades of these

724
J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
compounds were suggested to provide a novel balance
among them. In this study, we have investigated the
effects of structural modifications, bearing oxypropano-
lamine on 4-phenyl ring of dihydropyridine. Although
several previous studies suggested that the hydrogen
atom was preferable at the para position of the 4-phenyl
ring of dihydropyridine,14 however, on which, we first
test to introduce 2,5-, 2,4- or 3,4-disubstitutents on 4-
phenyl ring of dihydropyridine. Substituents on the 4-
phenyl ring of dihydropyridine were selected from NO2,
chloro, methoxyl, hydroxy and oxypropanolamine moi-
eties. On the 4-phenyl ring of dihydropyridine, the
replacement of its 4-hydroxyl, 3-hydroxyl and 2-
hydroxyl group with a oxy- propanolamine moiety
might be added with a- and b- adrenoceptor blocking
activities to enhance calcium channel blocking activity
that derived from their dihydropyridine base. Because of
the bulkiness, basic characteristics, and hydrophobicity
of this branched oxypropanolamine side chain, these
compounds were expected to resist hydrolysis and had
more potent and longer antihypertensive activity.
sive rats, and with long-acting antihypertensive action
in SHR. Compound 1a was the most potent for the
bradycardia effect both in normotensive rats and SHR.
It was shown that all the compounds could either
strongly or weakly inhibit Ca2+ dependent chrono-
tropic, inotropic and aortic contractile effects; therefore,
some of the compounds (1a, 1b and 1g) appeared to be
potent and competitive calcium entry blocking agents.
The IC50 values indicated that the relaxation of aorta
rings by compound 1g was more effective than other
compounds. Compounds 1a, 1b and 1g showed not only
markedly high calcium-antagonistic activity but also the
highest antihypertensive effect. In all cases, the 3,5-
dicarboethoxy substituent derivatives were less active
than those of the 3,5-dicarbomethoxy substituent on
dihydropyridine base. Ester substituents at the 3,5-
position of the dihydropyridine derivatives weakened
the activity with increasing substituent size. From the
results of the compounds, we concluded that com-
pounds 1b, 1c, 1f, 1g, 1i and 1j induced sustained anti-
hypertensive effect is major and attributed to their
calcium entry and a-adrenoceptor blocking activities in
the blood vessel due to their introduction of guaia-
coethyl terminal moiety in the side chain on the 4-
phenyl ring of dihydropyridine. Bradycardiac effects of
all the compounds 1a–1j are resulted from calcium entry
and b-adrenoceptor blocking, which attenuate the sym-
pathetic activation-associated reflex tachycardia in the
heart.
The results, expressed as blood pressure decreasing
activity, calcium blocking activity and pA2 values of the
negative inotropic, chronotropic and calcium antagonist
potencies, together with the respective percentage
decreases of the maximum effects, are shown in Figures
3 and 4, Tables 1 and 2. These data show that all the
synthetic manipulations were detrimental for calcium
channel modulation and added with adrenoceptor
blocking activities as far as vascular smooth muscle and
heart were concerned.
A favorable substituent on the 4-phenyl ring of dihy-
dropyridine derivatives was suggested to be an electron-
withdrawing group such as a nitro group, a cyano
group, a trifluoromethyl group, or a chlorine atom.34
Dihydropyridine derivatives with nitroso moiety, an
electron-withdrawing substitutent on the phenyl ring,
was described as having coronary blood flow increasing
activity.8 Introduction of electron-donating substituents
on the phenyl ring such as the hydroxy and methoxy
groups found in dihydropyridines resulted in a loss of
hypotensive effect and coronary blood flow,35 however,
the modification of the hydroxy group on the phenyl
ring of the dihydropyridine derivative with the methoxy
group and oxypropanolamine such as compound 1a or
compound 1b still revealed potent vasorelaxing activity,
in contrast to previous results.8 In addition, 2-chloro, 5-
chloro and 5-NO2 phenyl derivatives exhibit almost
decreased activities for voltage-dependent calcium
channel blocking and for the hypertensive effects; this
was an interesting result that had never been seen in the
case of 4-substituted phenyl derivatives. Conforma-
tional and electronic change caused by the substitution
at different sites of the phenyl ring might affect interac-
tions of the concerned molecules with drug receptors
and thereby influence their potencies.36 The introduc-
tion of electron withdrawing substituents on the phenyl
ring, such as found in 4-hydroxy derivative,24 results in
a loss of these activities in comparison with compounds
1a and 1b. The finding implied that the substitution on
the para-hydroxy group of the 4-phenyl ring with oxy-
propanolamine may extend to have an additional adre-
noceptor blockade and may be more important than the
Significant differences in the structure–activity relation-
ship exist between these compounds. Intravenous
administration of these compounds produced both
decrease in mean blood pressure and heart beat. Of the
compounds, shown in Figure 1, with a 2-chloro sub-
stituent (R3), guaiacoxyethyl terminal (R1) (1g) and 3-
methoxy substituent (R3), t-butyl terminal (R1) (1a) on
nitrogen in the side chain on the 4-phenyl ring were
strongly active, and of the compound with a 3-methoxy
substituent (R3), guaiacoxyethyl terminal (R1) on nitro-
gen (1b) in the side chain on the 4-phenyl ring showed
the most potent antihypertensive activities in this series.
Compound 1b, with less tachycardiac reflex effect, was
the most potent for the hypotensive effect in normoten-
Table 2. Maximum relaxation (%) and IC50 values (M) of new
dihydropyridine calcium antagonists in isolated rat thoracic aorta
precontracted with 75 mM [K+
]
0
Compd
Maximum relaxation
(%) at 10ꢀ4 M
IC50 values
(M)
1a
1b
1c
1d
1e
1f
1g
1h
1i
78.5ꢁ5.2
89.8ꢁ2.4
56.4ꢁ4.4
70.5ꢁ4.8
72.8ꢁ6.3
75.3ꢁ2.5
96.2ꢁ3.7
97.6ꢁ4.8
100.6ꢁ5.1
90.6ꢁ3.9
1.26ꢂ10ꢀ5
3.69ꢂ10ꢀ6
9.24ꢂ10ꢀ5
2.94ꢂ10ꢀ5
1.66ꢂ10ꢀ5
2.07ꢂ10ꢀ5
2.99ꢂ10ꢀ6
4.43ꢂ10ꢀ6
5.02ꢂ10ꢀ6
1.75ꢂ10ꢀ5
1j

J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
725
chloro or NO2 groups on the phenyl ring for their cal-
cium channel blocking and their hypotensive activities.
In contrast, the oxypropanolamine derivative (1j) with-
out an electron donating 3-methoxy group decreases the
hypotensive and adrenoceptor/calcium channel block-
ing effects37 in comparison with compound 1b. Our 5-
nitro group substituted compounds did not show any
better influence upon the antihypertensive action, but
showed the most potent calcium antagonist activity
(shown in compounds 1h and 1i, in comparison with
compounds 1a and 1b). The 5-chloro-substituted com-
pounds 1d, 1e and 1f lost their antihypertensive activ-
ities, though the 2-chloro-substituted compound 1g was
a little less potent than the two most potent compounds,
1a and 1b. Compounds with 3-methoxy substituent (1a
and 1b) in our series seems significantly to increase the
hypotensive and adrenoceptor/calcium channel block-
ing activities, in comparison with that without the 3-
methoxy group (1j).37 For the substitutions on the 3,5-
position, compounds with dicarbomethyl substitutent
reveal more antihypertensive activity than those with
dicarboethyl substituent. This fact indicates that it is not
necessary to increase the bulk of the ester substituent in
our series of compounds. So far, the 3-methoxy group,
para substituted oxypropanolamine on the phenyl ring
and the 3,5-dicarcomethyl group of dihydropyridine
have been found to be the most favorable for our series
of compounds.
Scheme 3.
their effects in the cardiovascular system: (1) short-act-
ing calcium channel blockers, such as nifedipine; (2)
long-acting calcium channel blockers, such as amlodi-
pine; and (3) long-acting calcium channel blockers with
adrenoceptor blocking activity.37 Our new compounds
and YM-430and YM-15430-1 with b- or a-/b-adreno-
ceptor blocking activities,12,38 are suggested to belong to
the third generation dihydropyridine type calcium
channel blockers.
Compounds 1h and 1i, with nitro substituent, showed
more Ca/b1 selectivity (Table 1). This fact indicated that
they have more potent calcium channel blocking activity
on the vascular smooth muscle than b1-adrenoceptor
blocking activity on the cardiac muscle. However, tissue
selectivity is also dependent on other structural compo-
nents. Compounds 1b, 1c, 1f, 1g and 1j, containing
guaiacoxyethylamino terminal moiety that associated
with a-adrenoceptor blocking activity, are suggested to
be more highly tissue-selective (a>b1) on the vascular
smooth muscle (Table 1). Our compounds with structu-
rally different substituents on the 4-phenyl ring of dihy-
dropyridine type compounds exhibit selectivity for
vascular over cardiac tissue, except compound 1a with
more b1-adrenoceptor blocking activity in comparison
with its own a-adrenoceptor and calcium blocking
activities.
Experimental
General information
All melting points were measured with a Yanaco MP-J3
micromelting point apparatus and are uncorrected.
Infrared spectra were recorded through a KBr disk (n in
cmꢀ1) on a Hitachi 270-30 IR spectrophotometer. H
1
nuclear magnetic resonance spectra were recorded on a
Varian Gemini-200 FT-NMR spectrometer, using
DMSO-d6 as solvent and TMS as internal standard
(chemical shift in d, ppm). Mass spectra were recorded
with a JEOR-D100 GC-mass spectrometer. Column
chromatography was performed on silica gel (Merck
Kieselgel 60, 230–400 mesh).
In conclusion, this result suggested that our newly syn-
thetic compounds belong to hypotensive and anti-
hypertensive agents. Most of these compounds have
higher vascular calcium entry and then a-adrenoceptor
blocking activities. Some of them have predominantly
with b-adrenoceptor blocking activity. Since these com-
pounds have higher molecular weight and ester sub-
stituents, the lipophilities of theses compounds may be
easily partitioned into the cell membrane of vascular
smooth muscle where the binding events occur. From
these profiles, compound 1b was suggested to be a use-
ful compound for the treatment of hypertension and
associated cardiac hypertrophy.23 This compound is
now selected as candidate for further preclinical investi-
gations. Dihydropyridine type calcium channel blockers
have been functionally divided into three classes for
Vanillin and phenyl aldehyde such as 5-chloro-
salicylaldehyde, 2-chloro-4-hydroxy-benzaldehyde, 5-
nitrosalicylaldehyde were all obtained from Tokyo Che-
mical Industry Co. (TCI, Tokyo, Japan). (ꢀ)Isoprenaline
bitartrate and norepinephrine HCl were purchased from
Sigma Chemical Co. (St. Louis, MO, USA). Other
reagents used in this study were EP-grade products of E.
Merck (Darmstadt, Germany). Synthetic compounds
were prepared in stock solution in isotonic saline, with
the addition of 10% DMSO if necessary. Animals were
obtained from the National Laboratory Animal Breed-
ing and Research Center, Taipei, Taiwan.

726
J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
Synthesis
4-N{4-[2-Hydroxy-3-(tert-butylamino)propoxy]-3-meth-
oxy}benzyl-2,6-dimethyl-3,5-dicarbomethoxy-1,4-dihy-
2-methoxy-1-oxyethylamino benzene (guaiacoethylamine)
(Scheme 3). Guaiacol (0.2 mol) and 1,2-dibromoethane
(0.4 mol) were heated to 100 ꢃC with vigorous stirring,
and 125 mL of 1.6 N NaOH was added to the mixture
during 30min. Vigorous stirring and heating were con-
tinued until the pH approached 7. Then the cold reac-
tion mixture, consisting of two layers, was taken up in
ether or chloroform, the aqueous layer separated (up to
80% of unreacted quaicol can be removed from the
aqueous phase), and the organic phase was washed
thoroughly with 2 N NaOH (NaOH solution was
necessary to remove unreacted guaiacol) and once with
saturated NaCl and dried (MgSO4), and the solvent was
evaporated. The 3a (2-methoxy-1-oxyethylbromide
benzene) was obtained by distillation.
dropyridine (1a). Epichlorohydrin (5 mol) was added to
a
solution of 4-hydroxy-3-methoxy-1-benzaldehyde
(vanillin) (1 mol) in an aqueous ethanol solution (100 mL)
containing 8 g of sodium hydroxide. The resulting solution
was heated at 70 ꢃC for 3h with stirring. The hot reaction
mixture was filtered to remove the white precipitate, and
the filtrate was then concentrated under reduced pressure.
The residue was purified by column chromatography on
silica gel with an eluent (n-hexane: EtOAc=1:9). The
objective fragments were combined and evaporated and
the residue triturated with n-hexane. The resulting solid
was collected, washed with n-hexane and dried to afford
N-[4-(2,3-epoxypropoxy)-3-methoxy] benzaldehyde (4a)
1
as a pale yellow powder. H NMR (CDCl3) d 2.77–2.97
(m, 2H,–CH(O)CH2), 3.39–3.47 (m, 1H,–CH(O)CH2),
3.94 (s, 3H, Ar–O–CH3), 4.06–4.43 (m, 2H, Ar–O–
CH2), 7.01–7.47 (m, 3H, Ar-H), 9.87 (s, 1H, CHO); MS
m/s: 208 (Scan FAB+). Anal. (C11H12O4) C, H, N.
3a (2-methoxy-1-oxyethylbromide benzene). 1H NMR
(CDCl3) d 3.62–3.69 (m, 2H, CH2–Br), 3.87 (s, 3H, Ar–
O–CH3), 4.30–4.37 (m, 2H, Ar–O–CH2), 6.89–6.99 (m,
4H, Ar–H); MS m/s: 230(Scan FAB+). Anal.
(C9H11O2Br) C, H, N.
To 1 mol of tert-butylamine were added 1 mol of com-
pound 4a and 100 mL of absolute ethanol. The mixture
was heated at 50–55 ꢃC for 4 h, with stirring. The reac-
tion mixture was clarified by filtration and the filtrate
concentrated under reduced pressure. The product was
filtered and washed with absolute ethanol and dried,
yielding a pale yellow powder. Recrystallization from
EtOH afforded N-{4-[2-hydroxy-3-(tert-butylamino)
Potassium phthalimide (0.2 mol) was added to a solu-
tion of 3a (0.2 mol) in 200 mL of dimethylformamide.
The reaction was slightly exothermic, the temperature
rising to 55 ꢃC in 5 min. Stirring was continued for 30
min, and the temperature dropped slowly to 25 ꢃC.
After the addition of 300 mL of chloroform, the mixture
was poured into 500 mL of water. The aqueous phase
was separated and extracted with two 100 mL portions
of chloroform. The combined chloroform extract was
washed with 200 mL of 0.2 N NaOH (to remove
unreacted phthalimide) and 200 mL of water. After
drying (MgSO4) the chloroform was removed. The
crystalline residue was triturated with 40mL of ether,
and then 3b (2-methoxy-1-oxyethylphthalimide ben-
zene) was collected by filtration.
propoxy]-3-methoxy}benzaldehyde
(2a).1H
NMR
(CDCl3) d 1.51 (s, 9H, 2ꢂCH3), 3.17–3.51 (m, 2H, CH2–
N), 3.90(s, 3H, Ar–O–C H3), 4.17–4.25 (m, 2H, Ar–O–
CH2), 4.60–4.80 (s, 1H, CH–OH), 7.02–7.43 (m, 3H, Ar-
H), 9.86 (s, 1H, CHO); MS m/s: 282 (Scan FAB+).
Anal. (C15H23O4N) C, H, N.
A solution of N-{4-[2-hydroxy-3-(tert-butylamino)pro-
poxy]-3-methoxy} benzaldehyde (0.01 mol), methyl
acetoacetate (2.4 mL, 0.02 mol) and ammonia (10 mL)
in EtOH (15 mL) was refluxed for 15 h. The mixture was
evaporated and treated with 50mL aqueous Na 2CO3
solution. The solution was extracted with CHCl3 and
the extract was concentrated, followed by acidification
with 2 N EtOH–HCl. The residue was purified by col-
umn chromatography on silica gel with an eluent
(MeOH/EtOAc=3:7). The objective fragments were
combined and evaporated and the residue triturated
with EtOAc. The resulting solid was collected, washed
with EtOAc and recrystallization from a mixture of
MeOH and EtOAc (1:9). Compound 1a was obtained as
a yellow powder. UV (MeOH) lmax nm (log e) 229.0
3b (2 - methoxy - 1 - oxyethylphthalimide benzene). 1H
NMR (CDCl3) d 3.75 (s, 3H, Ar–O–CH3), 4.10–4.15 (t,
2H, CH2–phthalimide), 4.25–4.32 (t, 2H, Ar–O–CH2),
6.83–7.88 (m, 8H, Ar–H); MS m/s: 297 (Scan FAB+).
Anal. (C17H15O4N) C, H, N.
Product 2 (0.07 mol) and hydrazine hydrate (0.07 mol)
in 70mL of absolute ethanol were heated on the steam
bath for 45 min. A thick white precipitate appeared, to
which 20mL of 18% HCl was added. Heating was con-
tinued for 1 h and solid material was filtered off and
washed with ethanol. The solvent was evaporated from
filtrate, and the residue was basified with a 20% NaOH
solution. The mixture was extracted with chloroform
and dried (MgSO4), and the solvent was evaporated.
The residual oil was distilled to obtain 2-methoxy-1-
oxyethylamino benzene (guaiacoethylamine).
1
(3.89); H NMR (CDCl3) d 1.46 (s, 9H, 3ꢂCH3), 2.33
(s, 6H, 2ꢂCH3), 3.00–3.45 (m, 2H, CH2–NH), 3.65 (s,
6H, 2 ꢂCO–OCH3 ꢂ2), 3.77 (s, 3H, Ar–OCH3), 3.93–
4.14 (m, 2H, Ar–OCH2), 4.53 (m, 1H, CH–OH), 4.94 (s,
1H, Ar–CH<), 5.45 (br s, 1H, exchangeable,–NH–),
6.33–6.87 (m, 3H, Ar–H), 8.33 (br s, 1H, exchangeable,
CH2–NH–C), 9.28 (br s, 1H, exchangeable, OH); IR
(KBr): 3500, 3350, 3000 and 2850, 1710, 800 cmꢀ1; MS
m/s: 476 (Scan FAB+). Anal. (C25H36O7N2) C, H, N.
2 - methoxy - 1 - oxyethylamino benzene (guaiacoethyla-
mine). 1H NMR (CDCl3) d 3.08–3.13 (t, 2H, CH2–
NH2), 3.87 (s, 3H, Ar–O–CH3), 4.02–4.08 (t, 2H, Ar–O–
CH2), 6.88–6.95 (m, 4H, Ar-H); MS m/s: 167 (Scan
FAB+). Anal. (C9H13O2N) C, H, N.
The other compounds, including epoxides (compounds
4a–4j, data not shown), compounds 2b–2j and 1b–1j,

J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
727
1
were synthesized in a similar manner to that of com-
pound 1a, during the course of synthesis. The general
method for the amination using 3–10mL of each amine
such as n-butylamine, tert-butylamine, and 2-methoxy-1-
oxyethylamino benzene (guaiacoxyethylamine) was added
with 5 g (14.3 mol) of each epoxide and 50mL of metha-
nol, respectively. Each mixture was heated under nitro-
gen at 50–55ꢃC for 4 h, with stirring. The reaction mixture
was evaporated to dryness under reduced pressure. The
residual product was passed through SiO2 gel column
chromatography and eluted with EtOAc and n-hexane
mixture to produce various oxypropanolamines (2b–2j).
(4.33); H NMR (CDCl3) d 1.15–1.27 (s, 9H, 3 CH3),
1.36 (m, 6H, 2 ꢂCO–OCH2CH3), 2.29–2.30(d, 6H, 2
ꢂCH3), 2.69–2.98 (m, 2H, CH2–NH), 3.46–3.76 (s, 4H,
2 ꢂCO–OCH2CH3), 3.88–4.23 (m, 2H, Ar–OCH2), 4.27
(s, 1H, CH–OH), 5.21 (s, 1H, Ar–CH<), 5.74 (br s, 1H,
exchangeable,–NH–), 6.67–7.54 (m, 3H, Ar–H); IR
(KBr): 3460, 3261, 3009 and 2960, 1677, 1222 and 1127,
829 cmꢀ1
(C26H37O6N2 Cl) C, H, N.
; MS m/s: 508.5 (Scan FAB+). Anal.
N-{2-[2-Hydroxy-3-(n-butylamino)propoxy]-5-choloro}-
benzaldehyde (2e). 1H NMR (CDCl3): 1.13–1.24 (m,
7H,–N–CH2–CH2–CH2–CH3), 2.19–2.75 (m, 2H, CH2–
N–CH2), 3.97–4.17 (m, 2H, Ar–O–CH2), 4.32–4.52 (s,
1H, CH–OH), 6.65–7.28 (m, 3H, Ar–H), 9.25 ( s, 1H,
CHO); MS m/s: 285.5 (Scan FAB+). Anal.
(C14H20O3NCl) C, H, N.
The synthesis of various dihydropyridine derivatives
were prepared by 1 mol of aldehyde groups of oxypro-
panolamines and 2 mols of acetoacetic acid ester
(methylacetoacetate or ethylacetoacetate) and 1 mol of
ammonia according to the Hantzsch condensation
reaction (1b–1j).
4-{{N-{2-[2-Hydroxy-3-(n-butylamino)propoxy]-5-cholor-
o}benzyl}}-2,6-dimethyl-3,5-dicarboethoxy-1,4-dihydro-
pyridine (1e). UV (MeOH) lmax nm (log e) 226.0(4.35);
N-{4-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminobenze-
ne)propoxy] - 3 - methoxy}benzaldehyde (2b). 1H NMR
(CDCl3) d 2.50–2.89 (m, 4H, CH2–N–CH2), 3.71–3.79
(d, 6H, 2ꢂAr–O–CH3), 3.86–4.03 (m, 4H, 2ꢂAr–O–
CH2), 4.19–4.24 (t, 1H, CH–OH), 6.84–7.37 (m, 7H,
Ar–H), 8.31 (s, 1H, CHO); MS m/s: 375 (Scan FAB+).
Anal. (C20H25O6N) C, H, N.
1H NMR (CDCl3)
d
1.11–1.22 (m, 7H, NH–
CH2CH2CH2CH3), 1.22–1.29 (m, 6H,
2
ꢂCO–
OCH2CH3), 2.04 (s, 6H, 2 ꢂCH3), 2.17–2.71 (m, 4H,
CH2–NH–CH2), 3.4 (m, 4H, 2 ꢂCO–OCH2CH3), 3.97–
4.14 (m, 2H, Ar–OCH2), 4.3–4.5 (m, 1H, CH–OH), 5.33
(s, 1H, Ar–CH<), 6.65–7.27 (m, 3H, Ar–H); IR (KBr):
3475, 3365, 2967 and 2900, 1686, 1107 and 1043,
808 cmꢀ1
; MS m/s: 508.5 (Scan FAB+). Anal.
(C26H37O6N2 Cl) C, H, N.
4-{{N-{4-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminoben-
zene)propoxy]-3-methoxy}benzyl}}-2,6-dimethyl-3,5-di-
carbomethoxy - 1,4 - dihydropyridine (1b). UV (MeOH)
lmax nm (log e) 219.0(4.48); 1H NMR (CDCl3) d: 2.26
(s, 6H, 2 ꢂCH3), 2.50–2.89 (m, 4H, CH2–NH–CH2),
3.56 (s, 6H, 2ꢂCO–OCH3), 3.68–3.74 (d, 6H, 2 ꢂAr–
OCH3), 3.86–4.05 (m, 4H, 2 ꢂAr–OCH2), 4.4 (s, 1H,
CH--OH), 4.83 (s, 1H, Ar–CH<), 6.84–7.37 (m, 7H,
Ar–H), 8.33 (s, 1H,–NH–); IR (KBr): 3535, 3276, 2778,
1651, 1127 and 1023, 810 and 748 cmꢀ1; MS m/s: 570
(Scan FAB+). Anal. (C30H38O9N2) C, H, N.
N-{2-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminobenze-
ne)propoxy] - 5 - choloro}benzaldehyde (2f). 1H NMR
(CDCl3) d 2.60–3.21 (m, 4H, CH2–N–CH2), 3.73–3.84
(d, 3H, Ar–O–CH3), 4.07–4.18 (m, 4H, 2ꢂAr–O–CH2),
4.51–4.62 (t, 1H, CH–OH), 6.79–7.27 (m, 7H, Ar–H),
7.70(s, 1H, C HO); MS m/s: 379.5 (Scan FAB+). Anal.
(C19H22O5NCl) C, H, N.
4-{{N-{2-[2-Hydroxy-3-(2-methoxy-1-oxyethylamino
benzene) propoxy]-5-choloro} benzyl}}-2,6-dimethyl-3,5-
dicarboethoxy - 1,4 - dihydropyridine (1f). UV (MeOH)
lmax nm (log e) 226.0(4.26); 1H NMR (CDCl3) d 1.17–
1.28 (m, 6H, 2 ꢂCO–OCH2CH3), 2.27–2.29 (m, 6H, 2
ꢂCH3), 2.50–2.89 (m, 4H, CH2–NH–CH2), 3.67–3.77
(m, 4H, 2 ꢂCO–OCH2CH3), 3.84 (s, 3H, Ar–OCH3),
3.85–4.14 (m, 4H, 2 ꢂAr–OCH2), 4.26–4.33 (m, 1H,
CH–OH), 5.3 (s, 1H, Ar–CH<), 5.6 (br s, 1H,
exchangeable,–NH–), 6.71–7.16 (m, 7H, Ar–H); IR
(KBr): 3467, 3332, 2918 and 2850, 1707, 1621, 1101 and
1031, 810 & 753 cmꢀ1; MS m/s: 602.5 (Scan FAB+).
Anal. (C31H39O8N2 Cl) C, H, N.
4-{{N-{4-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminoben-
zene)propoxy]-3-methoxy} benzyl}}-2,6-dimethyl-3,5-di-
carboethoxy-1,4-dihydropyridine (1c). UV (MeOH) lmax
nm (log e) 226.0(4.42); 1H NMR (CDCl3) d: 1.20–1.27
(t, 6H, 2 ꢂCO–OCH2CH3), 2.33 (s, 6H, 2 ꢂCH3), 2.50–
2.89 (m, 4H, CH2–NH–CH2), 3.45 (m, 4H, 2 ꢂCO–
OCH2CH3), 3.78–3.84 (d, 6H, 2 ꢂOCH3), 3.88–3.92 (m,
4H, 2 ꢂAr–OCH2), 4.09–4.12 (m, 1H, CH–OH), 4.95 (s,
1H, Ar–CH<), 5.73 (br s, 1H, exchangeable,–NH--),
6.76–6.91 (m, 7H, Ar–H); IR (KBr) 3420, 3332, 2968 &
2850, 1687, 1217 and 1127, 753 cmꢀ1; MS m/s: 598
(Scan FAB+). Anal. (C32H42O9N2) C, H, N.
N-{2-[2-hydroxy-3-(tert-butylamino)propoxy]-5-choloro}-
benzaldehyde (2d). 1H NMR (CDCl3) d 1.29–1.34 (s,
9H, 3ꢂCH3), 2.86–2.96 (m, 2H, CH2–N–), 4.07–4.17
(m, 2H, Ar–O–CH2), 4.27–4.41 (s, 1H, CH–OH), 6.81–
7.89 (m, 3H, Ar–H), 10.34 (s, 1H, CHO); MS m/s: 285.5
(Scan FAB+). Anal. (C14H20O3NCl) C, H, N.
N-{4-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminobenze-
ne)propoxy] - 3 - choloro}benzaldehyde (2g). 1H NMR
(CDCl3) d 2.81–3.12 (m, 4H, CH2–N–CH2), 3.92–3.86
(t, 3H, Ar–O–CH3), 4.02–4.04 (m, 4H, 2ꢂAr–O–CH2),
4.31–4.42 (t, 1H, CH–OH), 6.88–7.98 (m, 7H, Ar–H),
8.71 (s, 1H, CHO); MS m/s: 379.5 (Scan FAB+). Anal.
(C19H22O5NCl) C, H, N.
4-{{N-{2-[2-Hydroxy-3-(tert-butylamino)propoxy]-5-cho-
loro}benzyl}}-2,6-dimethyl-3,5-dicarboethoxy-1,4-dihy-
dropyridine (1d). UV (MeOH) lmax nm (log e) 227.0
4-{{N-{4-[2-Hydroxy-3-(2-methoxy-1-oxyethylamino
benzene) propoxy]-3-choloro} benzyl}}-2,6-dimethyl-3,5-

728
J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
dicarboethoxy - 1,4 - dihydropyridine (1g). UV (MeOH)
lmax nm (log e) 221.0(4.39); 1H NMR (CDCl3) d 1.21–
1.29 (m, 6H, 2 ꢂCO–OCH2CH3), 2.04 (m, 6H, 2
ꢂCH3), 2.29–2.50(m, 4H, C H2–NH–CH2), 3.82–3.86
(m, 4H, 2 ꢂCO–OCH2CH3), 3.87–3.98 (m, 3H, Ar–
OCH3), 4.06–4.24 (m, 4H, 2 ꢂAr–OCH2), 4.6 (m, 1H,
CH–OH), 5.31 (s, 1H, Ar–CH<), 5.75 (br s, 1H,
exchangeable,–NH–), 6.73–6.98 (m, 7H, Ar–H); IR
(KBr): 3463, 3383, 2938, 1742, 1656, 1120, 810 and 748
2.78–3.08 (m, 4H, CH2–NH–CH2), 3.63 (s, 6H, 2 ꢂCO–
OCH3), 3.82 (s, 3H, Ar–OCH3), 3.92 (s, 1H, CH–OH),
4.10–4.14 (m, 4H, 2 ꢂAr–OCH2), 4.93 (s, 1H, Ar–
CH<), 6.73–7.17 (m, 7H, Ar–H), 8.39 (s, 1H,–NH–);
IR (KBr): 3438, 3346, 3019 and 2840, 1694, 1124 and
1124, 1029 and 769 cmꢀ1; MS m/s: 540(Scan FAB+).
Anal. (C29H35O8N2) C, H, N.
Pharmacology
cmꢀ1
; MS m/s: 602.5 (Scan FAB+). Anal.
(C31H39O8N2 Cl) C, H, N.
Measurement of hypotensive effect. All the compounds
were examined for antihypertensive activity. The
experiments were accomplished as described pre-
viously.23,24 In brief, male Wistar rats, weighing 250–
300 g were anesthetized with pentobarbital (40 mg/kg,
i.p.). Following each tracheal cannulation, systemic
arterial blood pressure and heart rate were taken from
the femoral arterial with a pressure transducer (Model
P10EZ; Spectramed, Oxnard, USA) connected to an
amplifier (Model 13-4615-52, Gould, Valley View, OH,
USA) and displayed on a recorder (Model 8188-4402,
Gould, Valley View, OH, USA). The body temperatures
were maintained at 37 ꢃC with an electric heating pad. A
femoral vein was cannulated for iv injection. All drugs
were administered in a volume of 0.4 ml/kg. Equivolu-
metric injections of the vehicle were administered to
control animals. The magnitudes of effects elicited after
injections were evaluated by measuring the changes in
arterial blood pressure and heart rate and comparing
them to basal values.
N-{2-[2-Hydroxy-3-(n-butylamino)propoxy]-5-nitro}ben-
1
zaldehyde (2h). H NMR (CDCl3) d 1.16–1.30( m, 9H,
3ꢂ CH3), 2.71–2.95 (m, 2H, CH2–N–), 4.03–4.07 (m,
2H, Ar–O–CH2), 4.17–4.26 (s, 1H, CH–OH), 6.95–7.26
(m, 3H, Ar–H), 10.45 (s, 1H, CHO); MS m/s: 296 (Scan
FAB+). Anal. (C14H20O5N2) C, H, N.
4-{{N-{2-[2-Hydroxy-3-(n-butylamino)propoxy]-5-nitro}-
benzyl}}-2,6-dimethyl-3,5-dicarboethoxy-1,4-dihydropyri-
dine (1h). UV (MeOH) lmax nm (log e) 233.0(4.27); 1H
NMR (CDCl3) d 1.14–1.16 (s, 9H, 3 ꢂCH3), 1.17–1.24
(m, 6H, 2 ꢂCO–OCH2CH3), 2.05 (d, 6H, 2 ꢂCH3),
2.33–2.34 (m, 4H, CH2–NH), 2.85–2.98 (m, 4H, 2
ꢂCO–OCH2CH3), 3.98–4.17 (m, 2H, Ar–OCH2), 4.24–
4.26 (s, 1H, CH–OH), 5.39 (s, 1H, Ar–CH<), 6.82–8.14
(m, 3H, Ar–H); IR (KBr): 3485, 3296, 2967 and 2880,
1681, 1197 and 1077, 848 and 768 cmꢀ1; MS m/s: 519
(Scan FAB+). Anal. (C26H37O8N3) C, H, N.
N-{2-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminobenze-
ne)propoxy] - 5 - nitro}benzaldehyde (2i). 1H NMR
(CDCl3) d 2.86–3.15 (m, 4H, CH2–N–CH2), 3.81–3.89
(t, 3H, Ar–O–CH3), 4.05–4.17 (m, 4H, 2ꢂAr–O–CH2),
4.33–4.39 (t, 1H, CH–OH), 6.84–7.03 (m, 7H, Ar–H),
8.72 (s, 1H, CHO); MS m/s: 390(Scan FAB+). Anal.
(C19H22O7N2) C, H, N.
Measurement of antihypertensive effect. Some of the
compounds (1a and 1b) with potent hypotensive activity
in normotensive rats were tested for antihypertensive
activity SHR. Male SHR of SHR/N strain weighing
200–300 g were used. Systolic blood pressure and heart
rate were measured by the indirect tail cuff method
using a rat tail manometer-tachometer (Natsume KN-
210, Natsume Seisakusho Co. Ltd., Japan). The rats
were restrained in a Plexiglas holder where temperature
was maintained at 37 ꢃC for 15–20min to raise their
body temperature, leading to dilation of the caudal
artery and thereby allowing the pressure pulse to be
easily detected. In all cases, at least three consecutive
measurements were obtained, and the average was
reported as the systolic blood pressure. Only rats with a
systolic pressure of 180mm Hg or higher were used.
The measurement was performed just before and 0.5, 1,
2, 3, 4, 5, 6, 12 and 24 h even 30h after single oral dose
of compounds 1a and 1b (5, 10, 25, 50 mg/kg), which
were dissolved in ethanol prior to diluting in acidic sal-
ine solution (pH 5.3). The solutions were protected from
light and administered orally via a stomach tube in a
volume of 5 mL/kg.
4-{{2-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminobenze-
ne)propoxy]-5-nitro}phenyl}}-2,6-dimethyl-3,5-dicarbo-
ethoxy-1,4-dihydropyridine (1i). UV (MeOH) lmax nm
(log e) 224.0(4.28); 1H NMR (CDCl3) d 1.14–1.28 (m,
6H, 2 ꢂCO–OCH2CH3), 2.04 (s, 6H, 2 ꢂCH3), 2.22–
2.33 (m, 4H, CH2–NH–CH2), 3.40–3.60 (m, 4H, 2
ꢂCO–OCH2CH3), 3.83–3.85 (s, 3H, Ar–OCH3), 3.99–
4.13 (m, 4H, 2 ꢂAr–OCH2), 4.3 (m, 1H, CH–OH), 5.41
(s, 1H, Ar–CH<), 6.5 (br s, 1H, exchangeable,–NH–),
6.82–8.12 (m, 7H, Ar–H); IR (KBr): 3483, 3342, 2989
and 2875, 1636, 1132 and 1046, 815 and 748 cmꢀ1; MS
m/s: 609 (Scan FAB+). Anal. (C31H35O10N3) C, H, N.
N-{4-[2-hydroxy-3-(tert-butylamino)propoxy]}benzalde-
1
hyde (2j). H NMR (CDCl3) d: 2.86–3.12 (m, 4H, CH2–
N–CH2), 3.81–3.85 (s, 3H, Ar–O–CH3), 3.98–4.15 (m,
4H, 2ꢂAr–O–CH2), 4.34 (s, 1H, CH–OH), 6.86–6.97
(m, 7H, Ar–H), 9.89 (s, 1H, CHO); MS m/s: 345 (Scan
FAB+). Anal. (C19H23O5N) C, H, N.
Measurement of ꢁ1-, ꢁ2-adrenergic antagonist effects.
The experiments were carried out at 37 ꢃC in a Krebs
solution of the following composition (in mM): NaCl
113; KCl 4.8; CaCl2 2.2; KH2PO4 1.2; MgCl2 1.2;
NaHCO3 25; Dextrose 11.0; bubbled with a 95% O2/
5% CO2 mixture. The right and left atrial strips were
prestretched to a baseline tension of 1.0g. The atria
were equilibrated for 90min in aerated Krebs solution
4-{{N-{4-[2-Hydroxy-3-(2-methoxy-1-oxyethylaminoben-
zene)propoxy]}benzyl}}-2,6-dimethyl-3,5-dicarbomethoxy
-1,4-dihydropyridine (1j). UV (MeOH) lmax nm (log e)
219.0(4.48); 1H NMR (CDCl3) d 2.31 (s, 6H, 2 CH3),

J.-C. Liang et al. / Bioorg. Med. Chem. 10 (2002) 719–730
729
before the experimental protocols were initiated. Iso-
lated atria were used to evaluate the possible involve-
ment of all compounds effects in b-adrenergic
mechanism.
compounds for 10min, using four segments per phar-
macological subset. Normal Krebs solution was then
exchanged for a calcium-free and depolarizing solution
(containing 40mM KCl, 0.5 mM EGTA, with or with-
out test compounds, pH 7.5). Cumulative contraction–
response curves for CaCl2 were then generated by add-
ing CaCl2. Responses were recorded in the presence and
absence of three concentrations of the test compounds.
For the assessment of b1-adrenergic blocking activity, a
control cumulative concentration–response curve to the
chronotropic effect of (ꢀ)isoprenaline was established.
The right atria were allowed 60min to restabilize, after
which time various concentrations of the test agent were
incubated with the atria 30min before the cumulative
concentration of the (ꢀ)isoprenaline (10ꢀ10–10ꢀ4 M)
were added. Responses were calculated as a percentage
of the maximum control response to (ꢀ)isoprenaline.
On the other hand, quiescent left atria were dissected
free of connective tissue and mounted in organ cham-
bers under a resting tension of 1.0g. The tissues were
bathed in an aerated Krebs solution (37 ꢃC) and were
driven at 2-s intervals via two platinum electrodes
placed at either side of the atrium. b1-Adrenergic
antagonist activity was determined as follows. Cumula-
tive concentration–response curves to the positive ino-
tropic effects of (ꢀ)isoprenaline were obtained in the
absence and presence of various concentrations of a test
compound. An incubation time of 30min was allowed
for the test compound. Data were calculated as a per-
centage of the increase in force induced by (ꢀ)iso-
)isoprenaline.
Voltage-dependent vasorelaxing activity. The other
method of measurement of vasorelaxing activity was a
modification of one we published previously to evaluate
the aorta relaxing activity of these compounds.23 The
tissues were allowed to equilibrate for 1 h in physi-
ological solution. To induce stable contractions of
thoracic aorta rings mounted in an organ bath, high
potassium solution (75 mM KCl) was added to the bath.
When response was considered 100%, compounds were
added. The 75 mM KCl solution was prepared by sub-
stituting 70mM NaCl for an equimolar amount of KCl.
Statistical evaluation of data. In the in vitro study, the
dose ratio was obtained from the ratio of the EC50
values of the agonist with and without the test com-
pound. Dissociation constant (KB) in a-, b1- and b2-
adrenoceptor antagonism of the test compounds were
calculated from the method described by our previous
study,23,24 using the equation: KB=[antagonist]/(dose
ratio-1). The pA2 values were then expressed as the
negative logarithm of KB and the slope of the regression
line were calculated by the method described by our
previous study.23,24 Since compounds reduced the max-
imum responses to CaCl2 in the rat aorta, double-reci-
procal regression plots according to our previous report
were performed.15 In the radioligand binding assay, the
Kd (the dissociation constant of radioligand), Bmax (the
maximum binding capacity) and Ki (inhibition constant)
values were determined by Scatchard analysis using the
non-linear curve fitting LIGAND program. All values
were presented in terms of mean ꢁ SEM. Analysis of
the data and plotting of the figures were done with the
aid of software (SigmaStat and SigmaPlot, Version 5.0,
San Rafael, CA, USA; GraphPad PRISMTM, Version
2.0, San Diego, CA, USA) run on an IBM compatible
computer and a Power Macintosh.
Tracheal b2-adrenoceptor blocking activity was eval-
uated according to the method of our previous
report.23,24 Tracheal strips were suspended in Krebs
solution at 37 ꢃC under resting tension of 1.5 g. After the
preparation were contracted by 3ꢂ10ꢀ6 M carbacol,
cumulative concentration-relaxant response curve for
(ꢀ)isoprenaline were constructed by increasing bath
concentration of the agonist approximately 3-fold. The
response to (ꢀ)isoprenaline was examined after 30min
incubation with the test compound. Relaxations were
expressed as percentage of the maximum relaxation
obtained by the first challenge of the tissues.
ꢀ1-Adrenoceptor antagonism. Vascular a-adrenoceptor
blocking activity was evaluated in the Wistar rat aorta
preparation. Isolated rat aorta preparations were
mounted in a 10mL organ bath and suspended in Krebs
solution at 37 ꢃC under a resting tension of 1.0g. Com-
pounds were added to the bath medium after a control
Acknowledgements
concentration-response curve to norepinephrine (10ꢀ9
–
10ꢀ4 M) has been obtained. The tissue was exposed to
compounds for 30min before re-challenging with nor-
epinephrine. Inhibitions were expressed as percentage of
the maximum contraction obtained by the challenge of
the tissues with norepinephrine.
This work in chemical synthesis was supported by
research grants from the Lotus Medical Supply Inc.
(Taipei, Taiwan).
Calcium antagonism activity. Calcium antagonistic
activity was evaluted according to the method of our
previous study.23,24 Isolated rat aorta was sectioned into
2.5-mm rings segments. An aortic ring segment was
mounted between two triangular pins and was sus-
pended in Krebs solution at 37 ꢃC under a resting ten-
sion of 1.0g and was equilibrated for 60min. The
segments were then incubated with or without test
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