Enantioselective synthesis of (R)-(+)- and (S)-(-)-higenamine and their analogues with effects on platelet aggregation and experimental animal model of disseminated intravascular coagulation

Article abstract of DOI:10.1016/j.bmcl.2008.05.094

Optically active tetrahydroisoquinoline alkaloids, (R)-(+)-higenamine (1R) and (S)-(-)-higenamine (1 S), and their optically active 1-naphthylmethyl analogues (2 and 3), were synthesized by enantioselective hydrogenation of the corresponding dihydroisoqui

Full text of DOI:10.1016/j.bmcl.2008.05.094

Bioorganic & Medicinal Chemistry Letters 18 (2008) 4110–4114
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Enantioselective synthesis of (R)-(+)- and (S)-(ꢀ)-higenamine and their
analogues with effects on platelet aggregation and experimental animal
model of disseminated intravascular coagulation
b
b
b
c
Mi Kyung Pyo a, , Duck-Hyung Lee b, , Doo-Hyun Kim , Ji-Hye Lee , Jong-Cheon Moon , Ki Churl Chang ,
*
Hye Sook Yun-Choi a,
*
a Natural Product Research Institute, College of Pharmacy, Seoul National University, Yeongeon-dong 28, Jongro-gu, Seoul 110-460, Republic of Korea
b Department of Chemistry, Sogang University, Shinsoo-dong 1, Mapo-gu, Seoul 121-742, Republic of Korea
c Department of Pharmacology, School of Medicine, Institute of Health Sciences and Gyeongsang National University, 92 Chilamdong, Jinju 660-751, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 28 February 2008
Revised 3 April 2008
Accepted 23 May 2008
Available online 29 May 2008
Optically active tetrahydroisoquinoline alkaloids, (R)-(+)-higenamine (1R) and (S)-(ꢀ)-higenamine (1 S),
and their optically active 1-naphthylmethyl analogues (2 and 3), were synthesized by enantioselective
hydrogenation of the corresponding dihydroisoquinoline intermediates 7 as a key step. The evaluation
of the platelet anti-aggregation effect demonstrated clearly that the (S)-(ꢀ)-enantiomers, 1S, 2S, and
3S, had higher inhibitory potency than the corresponding (R)-(+)-antipodes, 1R, 2R, and 3R, respectively,
to platelet aggregation induced by epinephrine. 1S enantiomer was superior to the corresponding 1R
enantiomer in attenuating all of the disseminated intravascular coagulation (DIC) and multiple organ fail-
ure (MOF) parameters tested, while the S enantiomers 2S and 3S ameliorated some of the DIC and MOF
parameters more effectively than the corresponding antipodes 2R and 3R.
Keywords:
Enantioselective synthesis
(R)-(+)- and (S)-(ꢀ)-higenamine
Platelet aggregation
Disseminated intravascular coagulation
(DIC)
Ó 2008 Published by Elsevier Ltd.
Multiple organ failure (MOF)
Higenamine (1, CAS 5843-65-2), a benzyltetrahydroisoquino-
line alkaloid, was isolated as an optically inert racemic mixture
from several plant materials, including Aconite root, Asiasaari ra-
dix, Annona squamosa, and Gnetum parvifolium,1–5 and as a (R)-
(+)-enantiomer from the embryo of Nelumbo nucifera.6 Racemic
higenamine was reported to possess cardiac b-adrenoceptor stim-
ulating activity, vasodilating and platelet anti-aggregating activi-
volve one asymmetric center in their structure and exist as
enantiomers. Although two optically active enantiomers have the
same achiral physical properties in general, discrepancies in phys-
iological activities are quite often observed between the two enan-
tiomers. Therefore, the development of a single enantiomeric
compound instead of racemic mixtures is strongly encouraged in
new drug development these days.
Racemic higenamine (1) could be either partially synthesized by
demethylation of racemic coclaurine (6-O-methylhigenamine) ob-
tained from the natural sources6 or by total syntheses.19 Prepara-
tion of optically active (R)-(+)-higenamine (1R) or (S)-(ꢀ)-
higenamine (1S) was also successful via resolution of the racemic
norarmepavine (6,7-di-O-methylhigenamine) followed by demeth-
ylation.20 However, despite the enormous potential as useful ther-
apeutic candidates and crucial needs to secure the optically active
higenamine and its derivatives, general and efficient methods to
obtain sufficient amount of enantiomerically pure compounds of
1,2, and 3 have not been reported yet. We report herein the first
enantioselective total syntheses of both (R)-(+)- and (S)-(ꢀ)-enan-
tiomers of higenamine (1S and 1R) and its 1-naphthylmethyl
derivatives, YS-49 (2S and 2R) and YS-51 (3S and 3R) as their
HBr salts in multi-gram quantities, together with their platelet
anti-aggregation effects and the effects on experimental DIC and/
or accompanying MOF.
ties through
a
-adrenoceptor,7–9 and to inhibit inducible nitric
oxide synthase (iNOS) expression and NO production in RAW
264.7, vascular smooth muscle and other cells.10–12 Two of the 1-
naphthylmethyl analogues of higenamine, 1-(a-naphthylmethyl)-
6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (2, YS-49, CAS
213179-96-5) and 1-(b-naphthylmethyl)-6,7-dihydroxy-1,2,3,4-
tetrahydroisoquinoline (3, YS-51, CAS 213179-96-5) were also re-
ported to possess various biological activities either equivalent or
superior to higenamine.7,13–15 In addition, these three tetrahydro-
isoquinolines1–3 were reported to have therapeutic potential for
disseminated intravascular coagulation (DIC) and/or accompany-
ing multiple organ failure (MOF).16–18 Tetrahydroisoquinolines in-
* Corresponding authors.
E-mail address: dhlee@sogang.ac.kr (D.-H. Lee).
Present address: Department of Physiology, College of Veterinary Medicine,
Konkuk University, Hwayang-dong 1, GwangJin-gu, Seoul 143-701, Republic of Korea.
0960-894X/$ - see front matter Ó 2008 Published by Elsevier Ltd.
doi:10.1016/j.bmcl.2008.05.094

M. K. Pyo et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4110–4114
4111
Synthesis of (R)-(+)-higenamine (1R) is summarized in Scheme
1. A mixture of commercially available 3,4-dimethoxyphenethyl-
amine (4) and 4-methoxyphenylacetic acid (5) was heated at
200 °C for 4 h to provide the amide 6 in 94% yield after recrystalli-
zation in ethyl acetate.21 The amide 6 was subjected to Bischler–
Napieralski cyclization (POCl3, CHCl3, reflux, 12 h),22 and the crude
iminium salt was subsequently converted to free imine 7 in 98%
overall yield by neutralization reaction using saturated sodium
bicarbonate solution at 0 °C. The imine 7 should be used immedi-
ately because it turned out to be relatively unstable at room
temperature.
Catalytic asymmetric reduction of 7, a key step in this route,
was carried out in DMF–HCO2H–TEA at rt for 12 h, using Noyori’s
RuCl[(S,S)-TsDPEN](p-cymene)] (8S) (1 mol%) as a catalyst.23 The
reaction proceeded smoothly to give a product 9 along with a sig-
nificant amount of greenish by-product, which interfered the puri-
fication of the desired secondary amine 9 in large scale reaction.
Fortunately, purification of the free amine 9 was successfully car-
ried out by a combination of acid–base reaction during the work-
up procedure, precipitation as its hydrobromide salt, and neutral-
ization reaction in 62% yield with >99% ee by HPLC analysis.24
The purified amine 9 turned out to be rather unstable at room tem-
perature unless it was very pure and therefore converted into its
hydrobromide salt 10 immediately by treatment with 48% HBr–
AcOH solution at rt in 84% yield.24 The salt 10 was then subjected
to BBr3 in methylene chloride at 0 °C for 3 h to afford the optically
active (R)-(+)-higenamine (1R) (78%) in excellent purity (>98%) and
enantioselectivity (>99% ee).25
Noyori catalyst (8R) was employed in place of (S,S)-catalyst (8S) in
reaction (d) of Scheme 1 (Scheme 2).
Other optically active higenamine derivatives such as (R)-(+)-
YS-49 (2R), (S)-(ꢀ)-YS-49 (2S), (R)-(+)-YS-51 (3R) and (S)-(ꢀ)-YS-
51 (3S) were also synthesized in a similar manner with >98% ee
in all cases (Scheme 3), starting from the commercially available
1-naphthylacetic acid (11) and 2-naphthylacetic acids (12),
respectively.26,27
The yields in each step of the synthesis of (R)-higenamine (1R),
(R)-YS-49 (2R), and (R)-YS-51 (3R) are summarized in Table 1. In
general, amide condensation, Bischler–Napieralski cyclization,
and neutralization reaction proceeded smoothly in all cases (at
least >85% yield in entries 1–3). And chiral reduction, salt forma-
tion, and final demethylation reaction also occurred in moderate
yields of 61–89% (entries 4–6).
Platelet anti-aggregating activities of six enantiomers were
then measured as follows. Blood, collected from the rat heart
after surgery using syringe containing 0.1 volume of 2.2% sodium
citrate, was centrifuged at 200g for 10 min to obtain platelet rich
plasma (PRP). The supernatant PRP was diluted with saline to
adjust the number of platelets (400–450 ꢁ 106 platelets/ml) with
the aid of platelet counter. Platelet poor plasma (PPP) was pre-
pared from the residue by centrifugation at 900g for 30 more
minutes. After 3-min pre-incubation of the adjusted PRP, sample
or vehicle was added and an aggregation inducing agent [ADP
(2–5
the sample addition on platelet aggregometer.28 Epinephrine
(1–4 M), arachidonic acid (AA, 10–40 M), and U46619 (1–
M) induced rat platelet aggregation were measured in the
presence of the threshold concentration of collagen (0.5–1.0 g/
lM) or collagen (2–5 lg/ml)] was added at 1 min after
l
l
(S)-(ꢀ)-Higenamine (1S), the antipode of 1R, was prepared fol-
5 l
lowing the same protocol as the synthesis of 1R, except that (R,R)-
l
ml), since rat platelets were observed not to aggregate in re-
sponse to epinephrine, AA, or U46619 (PGH2/TXA2 receptor ago-
nist) alone in the concentration dependent manner.29 The
concentrations of the compounds causing 50% inhibitory effects
(IC50) were determined from the Regression Wizard from the
SigmaPlot equation library.
O
MeO
OH
MeO
MeO
NH
a
MeO
O
+
NH2
Effects of these enantiomers on LPS-induced DIC model in the
rat were evaluated as follows. Six enantiomers suspended in dis-
OMe
OMe
4
5
6
MeO
MeO
MeO
MeO
HO
b, c
d
N
NH
NH2Br
(S,S
)-catalyst
(8
HO
S
)
9
MeO
MeO
OMe
OMe
OH
7
NBr
(
R)-Higenamine (1R)
5
HO
HO
MeO
MeO
HO
HO
7
OH
f
e
NH2Br
NH2Br
NH2Br
(
R,R)-catalyst
(8R)
10
OH
OMe
OH
(
R)-Higenamine (1R)
(
S
)-Higenamine (1
S)
Scheme 2. Concepts in the synthesis of 1R, and 1S.
Ts
N
Ts
N
Ru
Ru
O
O
O
N
H
N
H
Cl
Cl
OH
OH
OH
8
S
8
R
(RuCl[(
S,S
)-TsDPEN(p-cymene)] )
(RuCl[(R,
R
)-TsDPEN(p-cymene)] )
OMe
Scheme 1. Synthesis of (R)-(+)-higenamine (1R). Reagents and conditions: (a) 20-
0 °C, neat, 4 h, 94%; (b) POCl3, CHCl3, reflux, 12 h; (c) saturated NaHCO3, 0 °C, 98%
overall yield; (d) (i) RuCl[(S,S)-TsDPEN(p-cymene)] (8S) (1 mol%), HCO2H/TEA = 5:2,
DMF, rt, 12 h; (ii) purification, 62%; (e) 48% HBr, AcOH, rt, 84%; (f) CH2Cl2, BBr3, ꢀ78
to 0 °C, 3 h, 78%.
5
11
(sm for 2)
12
(sm for 1)
(sm for 3)
Scheme 3. Starting materials for the synthesis of 1, 2 and 3.

4112
M. K. Pyo et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4110–4114
Table 1
Table 2
Yields (%) in each step for the route to (R)-higenamine (1R), (R)-YS-49 (2R), and (R)-
YS-51 (3R)
IC50
(lM) values for platelet anti-aggregating activities of each enantiomer of
higenamine (1), YS-49 (2) and YS-51 (3) in rat platelets
Entry
Reaction types
Higenamine
YS-49
YS-51
Sample
ADPa
Collagenb
Epinephrinec,f
NaAAd,f
U46619e,f
(1)
(2)
(3)
ASAg
1R
1S
2R
2S
>1000
>1000
>1000
540
270
540
420
300
450
140
69
53
14
1.6
2.4
0.38
15
66
11
340
15
25
20
16
49
61
1
2
3
4
5
6
Amide condensation (a)
Bischler–Napieralski cyclization (b)
Neutralization (c)
97
98
85
65
70
75
92
94
96
70
70
61
92
99
88
71
70
89
9.1
11
5.6
7.9
6.2
Chiral reductiona (d)
Salt formation (e)
3R
3S
150
150
Deprotectiona (f)
460
0.9
a
a
The purities of the isolated compounds were >98% pure and >99% ee in all cases
as determined by chiral HPLC analysis.
ADP 2–5 ꢁ 10ꢀ6 M.
b
c
d
e
f
Collagen 2–5 ꢁ 10ꢀ6 g/ml.
Epinephrine 1–4 ꢁ 10ꢀ6 M.
NaAA 1–4 ꢁ 10ꢀ5 M.
U46619 1–5 ꢁ 10ꢀ6 M.
With the threshold.
tilled water were administered orally to rats (Sprague–Dawley,
male, 230–270 g) once a day for two consecutive days at the dose
of 10 mg or 25 mg/10 ml/day after fasting 2 h. At 1 h after the
second oral administration of vehicle (for normal and LPS groups)
or test sample, rats were anesthetized with ketamine (250 mg/kg,
im). After 30 min, physiological saline (10 ml/kg for normal
group) or LPS (20 mg/10 ml of saline/kg) was continuously in-
fused into the tail vein for 3 h through the scalp vein set (needle
size; 25 G ꢁ 19 mm) with the aid of a syringe pump (KASP 005/
150MT, Keun-A Mechatronics, Korea). Blood from the abdominal
aorta was collected either in a glass tube containing soy bean
trypsin inhibitor and Bothrops atrox venom for the measurement
of fibrinogen/fibrin degradation product (FDP, Murex Diagnostics,
England) or in a plastic tube containing 2.2% sodium citrate (1/10,
v/v). After centrifugation of the coagulated blood at 1200g for
5 min twice, the supernatant serum was separated and stored
in the freezer for at least 12 h before FDP testing. The citrated
blood was centrifuged at 2000g for 30 min and the supernatant
plasma was used for the measurement of prothrombin time
(PT), activated partial thrombin time (aPTT), and fibrinogen level
(Sigma Chem. Co., USA). A part (0.4 ml) of the plasma was centri-
fuged with 100 mM CaCl2 (0.1 ml) to obtain serum for the mea-
surement of aspartate aminotransferase (AST), alanine
aminotransferase (ALT), urea nitrogen (BUN), and creatinine lev-
els. Platelet count was measured from the citrated whole blood
using an automatic platelet counter (PLT-4, Texas International
Lab). The PT and aPTT time were measured on a Beckton–Dickin-
son BBL fibrosystem (Coctkeysville, USA) according to the specifi-
cations set by the manufacturer. Thrombo-wellcotest kit was used
for FDP assay. The assay was performed semi-quantitatively, with
the up and down dilution method according to the procedure in
the kit. AST, ALT, BUN, and creatinine levels were determined
according to the specifications set by the manufacturer with the
use of an Autobiochemical analyzer (HITACH 747, Japan) in the
Green Cross Reference Laboratory. The experimental data were
expressed as means standard error of the mean (SEM). The re-
sults obtained from the experiments were analyzed with Turkey’s
honesty significant difference test for multiple comparisons to
make the paired comparisons between groups after the analysis
of one-way ANOVA test. P values less than 0.05 were considered
as significant.
enantiomers, at least partly, differentially interfere with the adren-
ergic -receptors.
a
Lipopolysaccharide (LPS, endotoxin), an integral component of
outer membrane of Gram-negative bacteria, has been considered
one of the major pathogenic factors contributing to symptoms of
septicemia and sepsis-related DIC and MOF. The iv infusion of
LPS to animals induces human DIC-like symptoms and has been
established as an experimental animal DIC model.30–32 The acti-
vation of blood coagulation system results in lowering of blood
platelet count and fibrinogen level, enhancing FDP concentration
and the prolongation of coagulation time. In the LPS-treated
group of rats (LPS control), most of parameters related to DIC
deviated significantly from those of the normal group of rats
as shown in Figure 1. The oral administration of each enantio-
mer (1R, 1S, 2R, 2S, 3R, and 3S) significantly ameliorated the in-
crease of FDP and BUN level, and prolongation of PT and aPTT
time induced by the iv infusion of LPS with showing more favor-
able effects with S enantiomer than with corresponding R enan-
tiomer. The decrease of platelet counts was also suppressed with
1S, 2R, 3R, and 3S. The prevention of LPS-induced decrease of
fibrinogen level was observed only with 1S and 3S. The LPS-in-
duced AST increase was suppressed by 1R, 1S, 3R, and 3S. No
significant differences were observed in ALT and creatinine levels
among normal, LPS control, and all of the compounds treated
groups of rats. The overall effects showed that 1S enantiomer
was superior to the corresponding 1R enantiomer while the
enantiomers of 2 and 3 did not show significant differences,
although the S enantiomers 2S and 3S attenuated some DIC
and MOF parameters more effectively than the corresponding
antipodes 2R and 3R (Fig. 2).
In summary, enantioselective synthesis of optically active 1R,
1S, 2R, 2S, 3R, and 3S were completed in multi-gram quantities
using Noyori’s asymmetric hydrogenation protocol as a key step
in 27–37% overall yields, respectively. The (S)-(ꢀ)-enantiomers,
1S, 2S and 3S, showed selectively higher inhibitory potency than
the (R)-(+)-enantiomers, 1R, 2R, and 3R in platelet anti-aggregating
activities induced by epinephrine. 1S enantiomer was superior to
The anti-platelet activity of each enantiomer (1R, 1S, 2R, 2S, 3R,
and 3S) was summarized in Table 2. All the compounds showed
dose-dependent inhibitory activities to ADP, collagen, epinephrine,
AA, and U46619 induced platelet aggregation. Both R-(+)- and S-
(ꢀ)-enantiomers of 1, 2, and 3 showed the same order of inhibitory
effects on ADP, collagen, AA, and U46619 induced aggregation.
However, 1S and 2S isomers were approximately 10-fold stronger
inhibitors than the corresponding 1R and 2R isomers, respectively,
and 3S isomer was about 20 times more active than the 3R anti-
pode on epinephrine-induced aggregation, suggesting that these
HO
HO
HO
HO
HO
HO
NH
NH
NH
OH
(
(
R
S
)-Higenamine (1
)-Higenamine (1
R
)
(
R
)-YS-49 (2
)-YS-49 (2
R
)
(
R
)-YS-51 (3
R)
S
)
(
S
S
)
(
S
)-YS-51 (3S)
Figure 1. Higenamine (1) and its derivatives (2, 3).

M. K. Pyo et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4110–4114
4113
Figure 2. Effects of each enantiomer of tetrahydroisoquinoline alkaloids (po) on LPS (20 mg/kg, 3 h iv infusion) induced DIC model in the rat. Each column represents the
relative mean SEM. ꢂp < 0.05, ꢂꢂp < 0.01, ꢂꢂꢂp < 0.001, significantly different from the control group (LPS group), #p < 0.05, ##p < 0.01, ###p < 0.001, significantly different from
the normal group, ^p < 0.05, ^^p < 0.01, ^^^p < 0.001, significantly different from the same sample, different dose treated group from Tukey’s HSD test after the one-way
ANOVA test. (A) Normal (n = 8), (B) LPS control (n = 13), (C) 1R 10 mg/kg (n = 6), 25 mg/kg (n = 9), (D) 1S 10 mg/kg (n = 8), 25 mg/kg (n = 7), (E) 2R 10 mg/kg (n = 9), 25 mg/kg
(n = 7), (F) 2S 10 mg/kg (n = 11), 25 mg/kg (n = 9), (G) 3R 10 mg/kg (n = 10), 25 mg/kg (n = 11), H: 3S 10 mg/kg (n = 11), 25 mg/kg (n = 9).
the corresponding 1R enantiomer in attenuating DIC and MOF
parameters, while the S enantiomers 2S and 3S ameliorated some
of the DIC and MOF parameters more effectively than the corre-
sponding antipodes 2R and 3R.
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award to M.K. Pyo, from the BK 21 project supported from Ministry
of Education, Republic of Korea.

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5.12; N, 4.02; tR (CHIREX 3020G-EO column by Phenomenex Co.
4.6 mm ꢁ 25 cm, 53:35:12 hexane/dichloromethane/(trifluoroacetic acid/
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26. Spectral data for (R)-YS-49 (2R)-HBr. TLC Rf 0.50 (benzene/acetone/
MeOH = 5:4:2, then three drops of 28% ammonium hydroxide solution); mp
28
255 °C; ½a D
ꢃ
¼ ꢀ68:7 (c 0.048, MeOH); 1H NMR (500 MHz, DMSO-d6) d 9.12
(s, 1H), 8.80 (s, 1H), 8.20 (d, 1H, J = 8.5 Hz), 8.02 (d, 1H, J = 8.0 Hz), 7.92 (d, 1H,
J = 8.0 Hz), 7.57–7.65 (m, 2H), 7.49 (t, 1H, J = 8.0 Hz), 7.42 (d, 1H, J = 8.5 Hz),
6.61 (s, 1H), 6.39 (s, 1H), 4.68 (t, 1H, J = 6.5 Hz), 3.79 (m, 1H), 3.52 (m, 2H), 3.20
(m, 1H), 2.98 (m, 1H), 2.83 (m, 1H); IR (KBr pellet) 3419, 2935, 2788, 2559,
1632, 1535 cmꢀ1; Anal. Calcd for C20H20BrNO2: C, 62.19; H, 5.22; N, 3.63.
Found: C, 62.21; H, 5.26; N, 3.66; tR (CHIREX 3020G-EO column by
Phenomenex Co. 4.6 mm ꢁ 25 cm, 53:35:12 hexane/dichloromethane/
(trifluoroacetic acid/EtOH = 1:20), flow rate 0.9 ml/min) 9.9 min for (R)-YS-49
(2R)-HBr, 11.9 min for (S)-YS-49 (2S)-HBr.
19. Chang, K. C.; Yun-Choi, H. S.; Lim, J. K.; Park, C. W. Arch. Pharm. Res. 1984, 7, 133.
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27. Spectral data for (R)-YS-51 (3R)-HBr. TLC Rf 0.50 (benzene/acetone/
21. Rice, K. C.; Brossi, A. J. Org. Chem. 1980, 45, 592.
22. Ott, H.; Hardtmann, G.; Denzer, M.; Frey, A. J.; Gogert, J. H.; Leslie, G. H.;
Trapold, J. H. J. Med. Chem. 1968, 11, 777.
MeOH = 5:4:2, then three drops of 28% ammonium hydroxide solution); mp
¼ þ10:4 (c 0.051, MeOH); 1H NMR (500 MHz, DMSO-d6) d 9.11
28
244 °C; ½a D
ꢃ
(s, 1H), 8.84 (s, 1H), 7.89–7.96 (m, 2H), 7.82–7.89 (m, 2H), 7.46–7.56 (m, 3H),
23. Uematsu, N.; Fujii, A.; Hashiguchi, S.; Ikariya, T.; Noyori, R. J. Am. Chem. Soc.
6.59 (s,1H), 6.57 (s, 1H), 4.72 (t, 1H, J = 6.5 Hz), 3.47 (m, 1H), 3.35 (m, 1H), 3.17
1996, 118, 4916.
(m, 2H), 2.88 (m, 1H); IR (KBr pellet) 3162, 2966, 2800, 1628, 1541, 1441 cmꢀ1
;
28
24. Spectral data for 9. TLC Rf 0.32 (EtOH/hexane = 3:1); ½a D
ꢃ
¼ þ25:0 (c 0.05,
Anal. Calcd for C20H20BrNO2: C, 62.19; H, 5.22; N, 3.63. Found: C, 62.19; H, 5.22;
N, 3.67; tR (CHIREX 3020G-EO column by Phenomenex Co. 4.6 mm ꢁ 25 cm,
53:35:12 hexane/dichloromethane/(trifluoroacetic acid/EtOH = 1:20), flow rate
0.9 ml/min) 9.7 min for (R)-higenamine (3R)-HBr, 10.9 min for (S)-higenamine
(3S)-HBr.
CHCl3); 1H NMR (500 MHz, CDCl3) d 7.17 (d, 2H, J = 8.5 Hz), 6.87 (d, 2H,
J = 8.5 Hz), 6.64 (s, 1H), 6.59 (s, 1H), 4.10 (m, 1H), 3.86 (s, 3H), 3.82 (s, 3H), 3.80
(s, 3H), 3.10–3.26 (m, 2H), 2.81–2.97 (m, 2H), 2.62–2.81 (m, 2H); MS (m/z)
calcd for
C
19H23NO2 (M+) 313.39, found 313.55; tR (Daicel Chiralcel,
4.6 mm ꢁ 25 cm, 40:10:0.05 hexane/2-propanol/diethylamine, flow rate:
28. McNicol, A. Platelets preparation and estimation of functional responses. In
Platelets-A practical approach; Watson, S. P., Authi, K. S., Eds.; Oirl Press at
Oxford University Press: New York, 1996; p 1.
0.5 ml/min) 25.4 min for (R)-isomer (9R), 20.6 min for (S)-isomer (9S).
25. Spectral data for (R)-higenamine (1R)-HBr. TLC Rf 0.50 (benzene/acetone/
MeOH = 5:4:2, then three drops of 28% ammonium hydroxide solution); mp
29. Yun-Choi, H. S.; Park, K. M.; Pyo, M. K. Throm. Res. 2000, 100, 511.
30. Onaya, J.; Kyogashima, M.; Sunose, A.; Miyauchi, S.; Horie, K. Jpn. J. Pharmacol.
1998, 76, 397.
¼ þ25:0 (c 0.05, MeOH); 1H NMR (500 MHz, DMSO-d6) d 9.35 (s,
28
249 °C; ½a D
ꢃ
1H), 9.06 (s, 1H), 8.84 (s, 1H), 7.10 (d, 2H, J = 8.5 Hz), 6.70 (d, 2H, J = 8.5 Hz),
6.56 (s, 1H), 6.52 (s, 1H), 4.5 (br s, 1H), 3.32 (m, 1H), 3.15 (m, 2H), 2.97 (m, 1H),
31. Aoki, Y.; Ota, M.; Katsuura, Y.; Komoriya, K.; Nakagaki, T. Arzneim.-Forsch./Drug
Res. 2000, 50, 809.
2.86 (m, 1H), 2.77 (m, 1H); IR (KBr pellet) 3231, 2798, 1627, 1520, 1454 cmꢀ1
;
Anal. Calcd for C16H18BrNO3: C, 54.56; H, 5.15; N, 3.98. Found: C, 54.55; H,
32. Fujita, M.; Izutani, W.; Komurasaki, Y. Thromb. Haemost 2000, 84, 54.