Synthesis and pharmacological activity of alkaloids from embryo of lotus, Nelumbo nucifera

Article abstract of DOI:10.1248/cpb.c12-00820

Bisbenzylisoquinoline alkaloid, nelumboferine which was recently isolated from the embryo of Nelumbo nucifera, and stereoisomers of neferine, which is a major alkaloid of the embryo of N. nucifera, were stereoselectively synthesized. Pharmacological activity of nelumboferine, stereoisomers of neferine, liensinine, isoliensinine, and O-methylneferine were evaluated.

Full text of DOI:10.1248/cpb.c12-00820

Regular Article
January 2013
Chem. Pharm. Bull. 61(1) 59–68 (2013)
59
Synthesis and Pharmacological Activity of Alkaloids from Embryo of
Lotus, Nelumbo nucifera
,a
Katsumi Nishimura,* Shinji Horii,a Takao Tanahashi,a Yumi Sugimoto,b and Jun Yamadab
b
a Kobe Pharmaceutical University; Higashinada-ku, Kobe 658–8558, Japan: and Yokohama College of Pharmacy;
Totsuka-ku, Yokohama 245–0066, Japan.
Received September 19, 2012; accepted October 11, 2012; advance publication released online October 26, 2012
Bisbenzylisoquinoline alkaloid, nelumboferine which was recently isolated from the embryo of Nelumbo
nucifera, and stereoisomers of neferine, which is a major alkaloid of the embryo of N. nucifera, were stereo-
selectively synthesized. Pharmacological activity of nelumboferine, stereoisomers of neferine, liensinine, iso-
liensinine, and O-methylneferine were evaluated.
Key words nelumboferine; neferine; liensinine; isoliensinine; O-methylneferine
Lotus (Nelumbo nucifera Gaertner, Nelumbonaceae) has (THF) at −78°C, followed by treating with benzyl chloride 7a
a wide distribution, such as in Japan, China, India, and Aus- at −98°C to give benzylisoquinoline (l)-(R)-8a in 59% yield
tralia. All parts of this plant have been used from ancient (Chart 2). Chiral auxiliary in 8a was removed by treating
times in traditional medicine.1) Recently in Japan, lotus seeds with hydrazine and toluenesulfonic acid. The resulting amino
have been recorded in the Japanese Pharmacopoeia. Chemical group was reductively methylated with formalin and sodium
constituents of the embryo of N. nucifera were studied in the borohydride in methanol to give N-methylated isoquinoline
1960s to isolate neferine,2) liensinine,3–5) and isoliensinine6) as (R)-9a in 70% yield. Chiral stationary phase HPLC analysis of
major alkaloids (Fig. 1).
(R)-9a showed enantiomeric excess of >99%. Removal of the
Biological studies on the lotus embryo reported biological benzyl protective group by catalytic hydrogenation gave ben-
activities such as antihypertensive activity7) and anti-pulmo- zyltetrahydroisoquinoline (R)-5a, the top half of nelumbofer-
nary fibrosis8); however, most of these activities were exam- ine, in quantitative yield. The bottom half of nelumboferine
ined by using a crude extract of the lotus embryo, and there ((R)-5b) was synthesized using the same procedure (Chart 3);
have been few experiments using pure constituents.9) Recently, asymmetric alkylation of (l)-6 with benzyl chloride 7b gave
we investigated the chemical constituents of the lotus embryo isoquinoline (l)-(R)-8b in 57% yield. Removal of the chiral
to isolate and structurally elucidate a new alkaloid, nelum- auxiliary, followed by reductive N-methylation, gave the bot-
boferine, as a minor component (Fig. 1).10) We also found that tom half (R)-5b in 70% yield with >99% optical purity.
neferine decreases locomotor activities in mice.11,12) These re-
sults prompted us to synthesize nelumboferine and to evaluate of tetrahydroisoquinoline,2) we attempted to couple the top and
its pharmacological activity. bottom halves. Reaction of phenol (R)-5a with bromide (R)-5b
According to the reported procedure for Ullmann coupling
Neferine, a major alkaloid, has two chiral centers in the in the presence of copper(II) oxide, potassium carbonate and
molecule, and there are structural differences among nefer- potassium iodide in refluxing pyridine gave the desired iso-
ine, liensinine, and isoliensine as to whether is O-methylated. quinoline dimer 10 only in 10% yield (Chart 4). The yield was
From the viewpoint of the structure–activity relationship, we improved by treating (R)-5a and (R)-5b with copper(I) bro-
synthesized three stereoisomers of neferine and O-methyl- mide-dimethyl sulfide complex15) and cesium carbonate when
neferine,5,13,14) and examined these pharmacological activities.
refluxing pyridine for 20h to give bisbenzylisoquinoline 10
in 34% yield along with 42% recovery of unchanged (R)-5a.
Deprotection of both benzyl and isopropyl groups with boron
Results and Discussion
Neferine, liensinine, and isoliensinine were isolated from trichloride in dichloromethane at −15°C proceeded smoothly
the embryo of N. nucifera. O-Methylneferine was prepared by
methylation of neferine with trimethylsilyldizaomethane.10)
Synthesis of Nelumboferine Our synthesis plan is shown
in Chart 1. Because nelumboferine is a dimer of benzyliso-
quinoline, it is synthesized by Ullmann coupling15,16) of two
chiral isoquinoline monomers (R)-5a (top half) and (R)-5b
(bottom half). Chiral 5a and 5b are synthesized by Gawley’s
asymmetric alkylation of lithiated isoquinoline (l)-6-Li, which
has an oxazolidine derived from (l)-valine as a chiral auxilia-
ry, with substituted benzyl chlorides 7a and 7b, respectively.17)
Tetrahydroisoquionoline (l)-6, which is a substrate for
asymmetric alkylation, was prepared according to the re-
ported procedure.17,18) Asymmetric alkylation was carried out
by lithiation of (l)-6 with n-butyllithium in tetrahydrofuran
The authors declare no conflict of interest.
Fig. 1. Structures of Alkaloids of the Embryo of Nelumbo nucifera
© 2013 The Pharmaceutical Society of Japan
*To whom correspondence should be addressed. e-mail: nishi@kobepharma-u.ac.jp

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Vol. 61, No. 1
Chart 1. Retrosynthesis of Nelumboferine
Reagents and conditions: (a) n-BuLi, 7b, THF, −98 to 0°C, 2h, 57%; (b) hydra-
zine, p-TsOH, EtOH, reflux, 2h; (c) formalin, NaBH4, MeOH, rt, 1h, 70%.
Reagents and conditions: (a) n-BuLi, 7a, THF, −98 to 0°C, 2h, 59%; (b) hydra-
zine, p-TsOH, EtOH, reflux, 2h; (c) formalin, NaBH4, MeOH, rt, 1h, 70%; (d) H2,
Pd/C, MeOH, rt, 7h, 99%.
Chart 3. Asymmetric Alkylation of Bottom Half
Chart 2. Asymmetric Alkylation of Top Half
to give nelumboferine in 70% yield. Spectroscopic and ana- dichloromethane at −15°C gave 1-epi-neferine in 68% yield.
lytical data of synthetic nelumboferine were agreed with those Ullmann coupling of (R)-5c and (S)-13, followed by deprotec-
of natural product.
tion gave 1′-epi-neferine. In the same way, Ullmann coupling
Synthesis of Stereoisomers of Neferine Three stereo- of (S)-5c and (S)-13, followed by deprotection, gave ent-
isomers of neferine (1-epi-neferine, 1′-epi-neferine and ent- neferine.
neferine) were synthesized by the same methodology used for
Pharmacological Activity Previously we demonstrated
the synthesis of nelumboferine. Thus, top and bottom halves that neferine reduces locomotor activity in mice dose depen-
were prepared by asymmetric alkylation, and two isoquinoline dently, showing that neferine has sedative effects;11) however,
units were connected by Ullmann coupling. Reaction of (l)-6 it is not yet clear whether liensinine and isoliensinine, and
with butyllithium followed by 4-methoxybenzyl chloride in nelumboferine isolated from the embryo of N. nucifera, have
THF at −98°C gave benzylisoquinoline (l)-(R)-8c in 54% sedative effects or not. In the present study, therefore, we
yield (Chart 5). Removal of the chiral auxiliary with hydra- examined the effects of these alkaloids, including a newly
zine, followed by reductive N-methylation with formalin and synthesized nelumboferine, on locomotor activity in mice. The
sodium borohydride gave (R)-9c in 60% yield. Deprotection of effects of O-methylneferine were also studied.
the benzyl group by catalytic hydrogenation gave the top half
Effects of neferine, liensinine, isoliensinne, nelumbofer-
(R)-5c in quantitative yield. Enantiomer of the top half ((S)-5c) ine and O-methylneferine on locomotor activity are shown
was also synthesized by asymmetric alkylation of (d)-6, which in Fig. 2. As shown in Results, all compounds decreased
has a chiral auxiliary derived from d-valine. Both enantiomers locomotor activity in mice. These results suggest that the
of the bottom half ((R)- and (S)-13) were also synthesized bisbenzylisoquinoline alkaloids examined in the present study
through asymmetric alkylation of (l)- or (d)-11, removal of the possess sedative effects. This is the first study demonstrating
chiral auxiliary, and reductive N-methylation (Chart 6).
that alkaloids from the embryo of N. nucifera liensinine, iso-
Ullmann coupling of (S)-5c and (R)-13 with copper(I) liensinine, and nelumboferine have sedative effects similar to
bromide-dimethyl sulfide and cesium carbonate in refluxing neferine. Neferine and O-methylneferine at 25mg/kg had no
pyridine gave isoquinoline dimer 14 in 35% yield (Chart 7). effect but had significant effects at a dosage of 50mg/kg. In
Deprotection of the benzyl group with boron trichloride in contrast, liensinine, isoliensinine, and nelumboferine elicited

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61
Reagents and conditions: (a) CuBr·SMe2, Cs2CO3, pyridine, reflux, 20h, 34%; (b) BCl3, CH2Cl2, −15°C, 1h, 70%.
Chart 4. Synthesis of Nelumboferine
Reagents and conditions: (a) n-BuLi, 7b, THF, −98 to 0°C, 2h, 75%; (b) hydra-
zine, p-TsOH, EtOH, reflux, 2h; (c) formalin, NaBH4, MeOH, rt, 1h, 64%.
Reagents and conditions: (a) n-BuLi, p-methoxybenzyl chloride, THF, −98 to
0°C, 2h, 54%; (b) hydrazine, p-TsOH, EtOH, reflux, 2h; (c) formalin, NaBH4,
MeOH, rt, 1h, 60%; (d) H2, Pd/C, MeOH, rt, 16h, 99%.
Chart 6. Asymmetric Alkylation of Bottom Half
Chart 5. Asymmetric Alkylation of Top Half
apparent effects on locomotor activity at 25 and 50mg/kg. nucifera are not clear at present; however, we reported that
Thus, the efficacy of liensinine, isoliensinine and nelum- neferine has antidepressant effects in mice mediated by the
boferine is more potent than neferine and O-methylneferine. serotonin 5-HT1A receptor12); therefore, the sedative effects of
Liensinine, isoliensinine, and nelumboferine have a hydroxy these compounds may be related to serotonergic transmission.
group at C-12 and/or C-7′ respectively, different from neferine
and O-methylneferine. O-methylation of the hydroxy group at
C-12′ in neferine could not increase pharmacological activity
Conclusions
We synthesized nelumboferine, an alkaloid of the embryo
and rather shortened the sedative effects, since the effects of of Nelumbo nucifera by using asymmetric alkylation and Ull-
O-methylneferine disappeared 50min after treatment. These mann coupling as key reactions. This is the first synthesis of
results suggest that the presence of a hydroxy group at C-12 or nelumboferine. We also synthesized stereoisomers of neferine,
C-7′, in place of a methoxy group, may contribute to enhanc- a major alkaloid of the embryo of N. nucifera and its deriva-
ing the sedative effects of bisbenzylisoquinolines.
tive O-methylneferine.
From the viewpoint of the structure–activity relationship,
Alkaloids from the embryo of N. nucifera, liensinine, iso-
three stereoisomers of neferine were synthesized and the ef- liensinine, and nelumboferine, have sedative effects similar to
fects of stereochemistry on locomotor activity were examined neferine. Synthesized nelumboferine showed a stronger effect
(Fig. 3). All compounds decreased locomotor activity in mice. than neferine. The newly synthesized O-methylneferine also
Neferine and ent-neferine elicited significant sedative effects elicited significant sedative effects. From the structure–activ-
at 50mg/kg, while at doses of 10 and 25mg/kg they did not ity relationship, the presence of a hydroxyl group at C-12 or
alter locomotor activity. 1-epi-Neferine and 1′-epi-neferine C-7′ may contribute to enhancing sedative activity. Among
decreased locomotor activity above 25 and 10mg/kg, respec- stereoisomers of neferine, activities of 1-epi- and 1′-epi-
tively. 1′-epi-Neferine elicited the most potent sedative effects neferine were higher than those of neferine. These interesting
among stereoisomers and 1′-epi-neferine at 50mg/kg elicited findings implied that the configuration change at C-1 or C-1′
strong sedative effects, with the result that 30% of mice died might enhance the activity of other alkaloids such as liensi-
during experiments.
nine.
These results reveal that the activity of the enantiomer
was not improved; however, 1-epimer and 1′-epimer both had
greater activity than neferine.
Experimental
All melting points were recorded on Yanagimoto hot plate
The detailed mechanisms of the sedative effects of bis- melting points apparatus and are uncorrected. IR spectra were
benzylisoquinoline alkaloids from the embryo of Nelumbo taken by Shimadzu FTIR-8200 spectrophotometer. NMR

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Vol. 61, No. 1
Reagents and conditions: (a) CuBr·SMe2, Cs2CO3, pyridine, reflux, 20h; (b) BCl3, CH2Cl2, −15°C, 1h.
Chart 7. Synthesis of 1-epi- and 1′-epi- and ent-Neferine
1
spectra were taken by Varian Mercury 300 spectrometer at (c=1.00, benzene). H-NMR (300MHz, CDCl3) δ: 0.84 (3H, d,
300MHz for 1H- and 75MHz for 13C-NMR or by Varian J=6.9Hz), 0.95 (3H, d, J=6.9Hz), 1.69 (1H, septet of d, J=6.9,
1
VXR-500 spectrometer at 500MHz for H- and 125MHz for 6.6Hz), 2.78 (2H, dd, J=5.8, 5.8Hz), 3.56 (1H, dt, J=12.6,
13C-NMR. MS and HRMS spectra were taken by Hitachi 5.8Hz), 3.63 (1H, dt, J=12.6, 5.8Hz), 3.81 (1H, ddd, J=8.8,
M-4000 spectrometer.
6.8, 6.6Hz), 3.86 (3H, s), 4.00 (1H, dd, J=8.0, 6.8Hz), 4.26
7-(Benzyloxy)-2-[(4S)-4,5-dihydro-4-(1-methylethyl)-2- (1H, dd, J=8.8, 8.0Hz), 4.39 (1H, d, J=17.3Hz), 4.45 (1H, d,
oxazolyl]-6-methoxy-1,2,3,4-tetrahydroisoquinoline ((l)-6) J=17.3Hz), 5.11 (2H, s), 6.60 (1H, s), 6.63 (1H, s), 7.28–7.44
To a solution of 7-(benzyloxy)-6-methoxy-1,2,3,4-tetrahydro- (5H, m). 13C-NMR (75MHz, CDCl3) δ: 17.5, 18.8, 28.0, 33.1,
isoquinoline18) (7.30g, 27mmol) in benzene (100mL) were 42.8, 46.8, 55.9, 70.0, 70.4, 71.0, 111.8, 111.9, 125.0, 126.7,
added (S)-2-ethoxy-4-isopropyl-4,5-dihydrooxazole17) (4.30g, 127.0, 127.6, 128.4, 137.0, 146.6, 148.1, 160.7.
27mmol) and p-toluenesulfonic acid (92mg, 0.54mmol), and
the mixture was refluxed for 2h. After being cooled to room
(d)-6. mp 76–77°C. [α]D23 +25.0 (c=1.00, benzene).
(1R)-7-Benzyloxy-2-[(4S)-4,5-dihydro-4-(1-methylethyl)-
temperature (rt), the solution was washed with saturated so- 2-oxazolyl]-1-(4-isopropylbenzyl)-6-methoxy-1,2,3,4-tetra-
dium bicarbonate, dried over sodium sulfate, filtered, and hydroisoquinoline ((l)-(R)-8a) To a solution of isoquinoline
evaporated. Silica gel column chromatography (ethyl acetate/ (l)-6 (3.82g, 10mmol) in THF (80mL) was added n-butyl-
ethanol=4/1) gave (l)-6 (9.0g, 88%) as white solid of mp lithium (1.22m in hexane, 8.6mL, 11mmol) at −80°C, and
72–74°C. MS (electron ionization (EI)) m/z: 380 (M+), 337, the mixture was stirred for 10min. A solution of the benzyl
289, 91. HR-MS (EI) m/z: 380.2113 (Calcd for C23H28N2O3 chloride 7a (2.27g, 12mmol) in THF (20mL) was added at
(M+): 380.2100). IR (Nujol) cm−1: 1654, 1609. [α]D28 −29.8 −98°C, and the mixture was stirred for 2h from −98 to 0°C.

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Fig. 2. Effects of Benzylisoquinoline Alkaloids on Locomotor Activity in Mice
Results are shown as the means S.E.M. of 5–9 mice. (A) Neferine; (B) liensinine; (C) isoliensinine; (D) nelumboferine; (E) O-methylneferine. *p<0.05, **p<0.01,
significantly different from the respective saline-treated group.
Saturated ammonium chloride solution was added, and the 2.48 (1H, ddd, J=15.9, 4.4, 4.0Hz), 2.78 (1H, m), 2.82 (1H,
whole was extracted with ethyl acetate. Organic layers were ddd, J=15.9, 9.8, 6.3Hz), 2.88 (1H, dd, J=13.5, 6.5Hz), 2.99
washed with brine, dried over sodium sulfate, filtered, and (1H, dd, J=13.5, 6.5Hz), 3.33 (1H, ddd, J=13.6, 9.8, 4.4Hz),
evaporated. Silica gel column chromatography (ethyl acetate/ 3.80 (1H, m), 3.85 (3H, s), 3.95 (1H, dd, J=8.5, 8.5Hz), 4.08
ethanol=9/1) gave (l)-(R)-8a (3.10g, 59%) as an amorphous (1H, dd, J=8.5, 8.5Hz), 4.48 (1H, septet, J=6.1Hz), 4.89 (1H,
solid of mp 85–88°C. MS (EI) m/z: 529 (MH+), 379, 91. d, J=12.4Hz), 4.95 (1H, d, J=12.4Hz), 5.01 (1H, t, J=6.5Hz),
HR-MS (EI) m/z: 529.3056 (Calcd for C33H41N2O4 (MH+): 6.30 (1H, s), 6.58 (1H, s), 6.75 (2H, d, J=8.5Hz), 6.93 (2H, d,
529.3066). IR (Nujol) cm−1: 1644. [α]D27 −135.4 (c=1.00, ben- J=8.5Hz), 7.27–7.37 (5H, m). 13C-NMR (75MHz, CDCl3) δ:
1
zene). H-NMR (300MHz, CDCl3) δ: 0.81 (3H, d, J=6.6Hz), 17.6, 18.4, 21.7, 21.8, 27.7, 33.1, 39.5, 41.1, 55.6, 57.1, 69.4, 69.6,
0.89 (3H, d, J=6.6Hz), 1.30 (6H, d, J=6.1Hz), 1.66 (1H, m), 70.1, 70.6, 111.4, 113.0, 115.2, 126.6, 126.9, 127.5, 128.1, 128.2,

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Vol. 61, No. 1
Fig. 3. Effects of Neferine and Its Stereoisomers on Locomotor Activity in Mice
Results are shown as the means S.E.M. of 5–9 mice. (A) Neferine; (B) 1-epi-neferine; (C) 1′-epi-neferine; (D) ent-neferine; *p<0.05, **p<0.01, significantly different
from the respective saline-treated group.
130.2, 130.6, 136.9, 145.6, 147.9, 156.1, 160.1.
(1H, dd, J=13.5, 5.0Hz), 3.15 (1H, m), 3.60 (1H, dd, J=7.7,
(R)-7-(Benzyloxy)-1-(4-isopropoxybenzyl)-6-methoxy- 5.0Hz), 3.83 (3H, s), 4.48 (1H, septet, J=6.1Hz), 4.75 (1H, d,
2-methyl-1,2,3,4-tetrahydroisoquinoline ((R)-9a) To a so- J=12.1Hz), 4.82 (1H, d, J=12.1Hz), 6.08 (1H, s), 6.57 (1H, s),
lution of (l)-(R)-8a (1.00g, 1.89mmol) in ethanol (19mL) 6.78 (2H, d, J=8.5Hz), 6.94 (2H, d, J=8.5Hz), 7.24–7.33 (5H,
were added hydrazine hydrate (1.8mL, 27.8mmol) and p- m). 13C-NMR (75MHz, CDCl3) δ: 22.0, 25.6, 40.0, 42.6, 46.9,
toluenesulfonic acid monohydrate (718mg, 3.78mmol), and the 55.8, 64.7, 69.6, 70.6, 111.6, 113.7, 115.4, 126.4, 127.2, 127.5,
mixture was refluxed for 2h. Water was added, and the whole 128.3, 129.2, 130.6, 131.7, 137.2, 145.3, 147.7, 156.1. HPLC:
was extracted with ethyl acetate. Organic layers were washed Daicel Chiralpak IB, hexane–2-propanol–ethylenediamine=
with brine, dried over sodium sulfate, filtered, and evaporated 80:20:0.1, 1.0mL/min, UV 270nm, R: 5.2min (dl-9a: 4.7,
to give crude amine. To a solution of this amine in metha- 5.3 min).
nol (29mL) were successively added formalin (37%, 1.4mL,
(R)-1-(4-Isopropoxybenzyl)-6-methoxy-2-methyl-1,2,3,4-
18.9mmol) and sodium borohydride (472mg, 12.6mmol) at tetrahydroisoquinolin-7-ol ((R)-5a) To a solution of (R)-9a
0°C, and the mixture was stirred for 1h at rt. Ten percent (757mg, 1.75mmol) in methanol (150mL) were added 10%
acetic acid and 10% ammonia solution were successively palladium on carbon (208mg) and acetic acid (0.42mL), and
added to the reaction mixture at 0°C, and the whole was ex- the mixture was stirred under hydrogen atmosphere for 7h at
tracted with ether. Organic layers were washed with brine, rt. Reaction mixture was filtered, and the filtrate was evapo-
dried over potassium carbonate, filtered, and evaporated. rated. The residue was dissolved in dichloromethane, and the
Silica gel column chromatography (ethyl acetate/ethanol= solution was washed with saturated sodium bicarbonate and
4/1) gave (R)-9a (570mg, 70%) as an amorphous solid of mp brine, dried over sodium sulfate, filtered, and evaporated.
76–78°C. MS (EI) m/z: 432 (MH+), 282, 191. HR-MS (EI) m/z: Silica gel column chromatography (chloroform/methanol=9/1)
432.2536 (Calcd for C28H34NO3 (MH+): 432.2539). IR (Nujol) gave (R)-5a (583mg, 99%) as a colorless oil. MS (EI) m/z: 342
1
cm−1: 1606. [α]D28 −8.0 (c=1.00, benzene). H-NMR (300MHz, (MH+), 192. HR-MS (EI) m/z: 342.2072 (Calcd for C21H28NO3
CDCl3) δ: 1.29 (3H, d, J=6.1Hz),1.30 (3H, d, J=6.1Hz), 2.50 (MH+): 342.2069). IR (neat) cm−1: 3543, 3408, 1610. [α]D27
(3H, s), 2.54–2.86 (3H, m), 2.69 (1H, dd, J=13.5, 7.7Hz), 3.06 +27.6 (c=1.00, benzene). 1H-NMR (300MHz, CDCl3) δ:

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1.32 (6H, d J=6.0Hz), 2.45 (3H, s), 2.53 (1H, m), 2.67–2.77 J=13.7, 5.2Hz), 3.05–3.15 (2H, m), 3.51–3.64 (2H, m), 3.75
(2H, m), 2.82 (1H, dd, J=14.0, 6.0Hz), 3.02 (1H, dd, J=14.0, (3H, s), 3.80 (3H, s), 4.38 (1H, septet, J=6.0Hz), 4.84 (2H, s),
6.0Hz), 3.16 (1H, m), 3.65 (1H, t, J=6.0Hz), 3.84 (3H, s), 4.50 5.03 (2H, s), 6.18 (1H, s), 6.39 (1H, s), 6.51 (1H, s), 6.61 (1H,
(1H, septet, J=6.0Hz), 6.39 (1H, s), 6.52 (1H, s), 6.77 (2H, d, s), 6.62 (2H, d, J=8.5Hz), 6.65 (1H, dd, J=8.3, 2.2Hz), 6.80
J=8.5Hz), 7.02 (2H, d, J=8.5Hz). 13C-NMR (75MHz, CDCl3) (1H, d, J=8.3Hz), 6.87 (2H, d, J=8.5Hz), 7.22–7.37 (11H,
δ: 22.0, 22.0, 25.0, 40.5, 42.4, 46.8, 55.6, 64.5, 69.7, 110.5, m). 13C-NMR (75MHz, CDCl3) δ: 22.0, 25.5, 25.8, 40.2, 40.4,
113.9, 115.5, 125.1, 130.0, 130.3, 131.8, 143.4, 145.2, 156.0.
42.6, 42.6, 46.9, 47.3, 55.8, 55.8, 64.3, 64.5, 69.6, 70.8, 70.9,
(1R)-7-(Benzyloxy)-1-(4-(benzyloxy)-3-bromobenzyl)-2- 111.6, 112.3, 113.6, 114.8, 115.4, 117.9, 120.3, 120.6, 124.7,
[(4S)-4,5-dihydro-4-(1-methylethyl)-2-oxazolyl]-6-me- 126.6, 127.0, 127.3, 127.4, 127.6, 128.2, 128.3, 129.1, 129.5,
thoxy-1,2,3,4-tetrahydroisoquinoline
((l)-(R)-8b) This 130.3, 131.5, 133.3, 137.2, 137.3, 144.0, 145.6, 146.4, 147.6,
compound was prepared from the isoquinoline (l)-6 and the 147.8, 148.6, 155.9.
benzyl chloride 7b as described above in 57% yield. MS (EI)
Nelumboferine (4) To a solution of 10 (410mg, 0.5mmol)
m/z: 654 (M+), 379, 91. HR-MS (EI) m/z: 654.2103 (Calcd for in dichloromethane (35mL) was added a solution of boron tri-
C37H39BrN2O4 (M+): 654.2093). IR (neat) cm−1: 1647. [α]D24 chloride (1m in dichloromethane, 2.5mL, 2.5mmol) at −15°C,
1
−107.6 (c=1.00, benzene). H-NMR (300MHz, CDCl3) δ: 0.80 and the mixture was stirred for 1h. Water and 27% ammonia
(3H, d, J=6.6Hz), 0.88 (3H, d, J=6.6Hz), 1.62 (1H, m), 2.46 solution were added, and the whole was extracted with dichlo-
(1H, ddd, J=15.9, 4.4, 4.4Hz), 2.73–2.84 (2H, m), 2.85 (1H, romethane. Organic layers were washed with brine, dried over
dd, J=13.5, 6.6Hz), 2.94 (1H, dd, J=13.5, 6.6Hz), 3.34 (1H, sodium sulfate, filtered, and evaporated. Silica gel column
ddd, J=9.6, 4.4, 4.4Hz), 3.76 (1H, m), 3.85 (3H, s), 3.93 (1H, chromatography (chloroform/methanol/water=80/20/1) gave
dd, J=8.0, 6.0Hz), 4.05 (1H, dd, J=8.5, 8.0Hz), 4.91 (1H, d, nelumboferine (210mg, 70%) as a pale yellow amorphous solid
J=12.4Hz), 4.98 (1H, d, J=12.4Hz), 5.00 (1H, t, J=6.6Hz), of mp 117–120°C. UV λmax (methanol) nm (logε): 225 (4.51),
5.11 (2H, s), 6.32 (1H, s), 6.58 (1H, s), 6.79 (1H, d, J=8.4Hz), 284 (4.02). MS (EI) m/z: 596 (M+), 489, 192. HR-MS (EI) m/z:
6.90 (1H, dd, J=8.4, 2.0Hz), 7.27–7.46 (11H, m). 13C-NMR 596.2876 (Calcd for C36H40N2O6 (M+): 596.2886). IR (KBr)
(75MHz, CDCl3) δ: 17.7, 18.6, 27.8, 33.2, 39.7, 41.1, 55.9, 57.1, cm−1: 3385, 1612, 1510. [α]D26 −66.0 (c=1.00, MeOH). Lit.10)
69.8, 70.3, 70.7, 71.0, 111.7, 111.8, 113.1, 113.4, 126.8, 126.9, [α]D −64.0 (c=1.0, MeOH). 1H-NMR (500MHz, CDCl3) δ:
127.1, 127.7, 127.8, 127.9, 128.4, 129.7, 132.6, 134.4, 136.5, 2.52 (3H, s,), 2.54 (3H, s), 2.62 (1H, m), 2.66 (1H, dd, J=13.5,
137.0, 145.9, 148.2, 153.4, 160.2.
10.0Hz) 2.69 (1H, dd, J=13.5, 10.5Hz), 2.75 (1H, ddd, J=12.0,
(R)-7-(Benzyloxy)-1-(4-(benzyloxy)-3-bromobenzyl)- 6.0, 6.0Hz), 2.79–2.87 (2H, m), 2.91 (1H, m), 2.97 (1H, m),
6-methoxy-2-methyl-1,2,3,4-tetrahydroisoquinoline 3.10 (1H, dd, J=13.5, 3.0Hz), 3.20 (1H, dd, J=13.5, 2.5Hz),
((R)-5b) This compound was prepared from (l)-(R)-8b as 3.21 (1H, m), 3.42 (1H, m) 3.51 (1H, dd, J=10.5, 2.5Hz) 3.67
described above in 58% yield. MS (EI) m/z: 557 (M+), 282. (1H, dd, J=10.0, 3.0Hz) 3.85 (3H, s), 3.90 (3H, s), 5.97 (1H,
HR-MS (EI) m/z: 557.1580 (Calcd for C32H32BrNO3 (M+): s), 6.34 (1H, s), 6.46 (1H, dd, J=8.0, 2.0Hz), 6.56 (1H, s),
557.1566). IR (neat) cm−1: 1607. [α]D25 +24.4 (c=1.00, benzene). 6.68 (1H, s), 6.75 (1H, d, J=8.0Hz), 6.75 (1H, d, J=2.0Hz),
1H-NMR (300MHz, CDCl3) δ: 2.48 (3H, s), 2.49 (1H, m), 6.76 (2H, d, J=8.5Hz), 6.94 (2H, d, J=8.5Hz). 13C-NMR
2.73–2.82 (3H, m,), 2.97 (1H, dd, J=13.7, 5.0Hz), 3.11 (1H, m), (125MHz, CDCl3) δ: 22.1, 26.3, 39.8, 40.5, 42.3, 42.6, 44.5,
3.57 (1H, t, J=5.0Hz), 3.84 (3H, s), 4.81 (1H, d, J=12.3Hz), 47.6, 55.8, 56.0, 64.2, 65.3, 110.8, 112.2, 114.4, 115.5, 116.6,
4.89 (1H, d, J=12.3Hz), 5.10 (2H, s), 6.14 (1H, s), 6.56 (1H, s), 118.2, 121.3, 123.2, 127.0, 127.8, 129.5, 130.5, 130.6, 130.9,
6.78 (1H, d, J=8.5Hz), 6.84 (1H, dd, J=8.5, 1.9Hz), 7.23–7.46 143.2, 143.3, 143.9, 145.6, 146.7, 148.3, 155.6. Hydrochloride:
(11H, m). 13C-NMR (75MHz, CDCl3) δ: 25.6, 39.7, 42.6, 47.1, To a solution of 4 (230mg, 0.39mmol) in methanol (5mL) was
55.8, 64.5, 70.7, 70.8, 111.7, 111.8, 113.3, 113.6, 126.8, 127.1, added hydrochloric acid (12m, 0.2mL) at 0°C. Evaporation of
127.6, 127.7, 128.3, 128.4, 128.8, 129.6, 133.9, 134.3, 136.5, the solvent gave hydrochloride (250mg). mp 207–210°C. Anal.
137.2, 145.6, 147.9, 153.1. HPLC: Daicel Chiralpak IB, hex- Calcd for C36H42Cl2N2O6·2H2O: C, 61.27; H, 6.57; N, 3.97.
ane–2-propanol–ethylenediamine=80:20:0.1, 1.0mL/min, UV Found: C, 61.26; H, 6.63; N, 3.89.
270nm, R: 8.3min (dl-5b: 7.2, 8.4min).
(1 R) -7- ( Ben z yloxy) -2 -[(4S ) - 4, 5 - dihydro - 4 - (1-
(R)-7-(Benzyloxy)-1-[(4-(benzyloxy)-3-[(R)-1-(4- methylethyl)-2-oxazolyl]-6-methoxy-1-(4-methoxy-benzyl)-
isopropoxybenz yl) - 6 -methoxy-2-methyl-1, 2, 3, 4 - 1,2,3,4-tetrahydroisoquinoline ((l)-(R)-8c) This compound
tetrahydroisoquinolin-7-yloxy]benzyl)-6-methoxy-2-methyl- was prepared from (l)-6 and p-methoxybenzyl chloride as
1,2,3,4-tetrahydroisoquinoline (10) To a solution of (R)-5a described above in 54% yield. MS (EI) m/z: 501 (MH+), 379.
(191mg, 0.56mmol) and (R)-5b (310mg, 0.56mmol) in pyri- HR-MS (EI) m/z: 501.2741 (Calcd for C31H37N2O4 (MH+):
dine (3.9mL) were added copper(I) bromide–dimethyl sulfide 501.2753). IR (neat) cm−1: 1644. [α]D24 −67.8 (c=1.00, benzene).
complex (230mg, 1.12mmol) and cesium carbonate (1.09g, 1H-NMR (300MHz, CDCl3) δ: 0.81 (3H, d, J=6.6Hz), 0.89
3.36mmol), and the mixture was refluxed for 20h. Reaction (3H, d, J=6.6Hz), 1.64 (1H, m), 2.46 (1H, ddd, J=15.9, 4.4,
mixture was filtered, and the filtrate was evaporated. Silica gel 4.4Hz), 2.71 (1H, m), 2.79 (1H, ddd, J=15.9, 9.9, 5.8Hz), 2.90
column chromatography (chloroform/methanol=9/1) gave 10 (1H, dd, J=13.5, 6.6Hz), 3.00 (1H, dd, J=13.5, 6.3Hz), 3.32
(156mg, 34%) as a brown oil. MS (EI) m/z: 819 (MH+), 282, (1H, ddd, J=12.9, 9.9, 4.4Hz), 3.75 (1H, m), 3.76 (3H, s), 3.84
192. HR-MS (EI) m/z: 819.4368 (Calcd for C53H59N2O6 (MH+): (3H, s), 3.95 (1H, dd, J=8.0, 6.0Hz,), 4.09 (1H, dd, J=8.5,
819.4371). IR (neat) cm−1: 1508. [α]D26 −76.8 (c=1.00, MeOH). 8.0Hz), 4.89 (1H, d, J=12.6Hz), 4.95 (1H, d, J=12.6Hz),
1H-NMR (300MHz, CDCl3) δ: 1.23 (3H, d, J=6.0Hz), 1.25 5.02 (1H, dd, J=6.6, 6.3Hz), 6.30 (1H, s), 6.58 (1H, s), 6.77
(3H, d, J=6.0Hz), 2.43 (3H, s), 2.44 (3H, s), 2.55 (1H, m), (2H, d, J=8.8Hz), 6.94 (2H, d, J=8.8Hz), 7.24–7.37 (5H, m).
2.62 (1H, dd, J=13.7, 5.8Hz) 2.64–2.83 (5H, m), 2.75 (1H, dd, 13C-NMR (75MHz, CDCl3) δ: 17.6, 18.5, 27.8, 33.2, 39.7, 41.2,
J=14.6, 5.5Hz), 2.90 (1H, dd, J=14.6, 5.8Hz), 2.97 (1H, dd, 55.0, 55.8, 57.1, 69.7, 70.2, 70.8, 111.5, 113.1, 113.3, 126.8,

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Vol. 61, No. 1
127.0, 127.6, 128.3, 130.4, 130.5, 130.7, 137.0, 145.7, 148.0, 1.55–1.66 (1H, m), 2.52 (1H, ddd, J=15.9, 4.4, 4.4Hz),
157.9, 160.2.
2.78–2.87 (2H, m), 2.92 (1H, dd, J=13.5, 6.6Hz), 3.04 (1H,
dd, J=13.5, 6.9Hz), 3.40 (1H, ddd, J=12.9, 9.6, 4.4Hz), 3.66
(d)-(S)-8c. [α]D24 +80.4 (c=1.00, benzene).
(R)-7-(Benzyloxy)-6-methoxy-1-(4-methoxybenzyl)-2- (3H, s), 3.72 (1H, ddd, J=8.8, 6.0, 6.0Hz), 3.84 (3H, s), 3.93
methyl-1,2,3,4-tetrahydroisoquinoline ((R)-9c) This com- (1H, dd, J=8.0, 6.0Hz), 4.05 (1H, dd, J=8.8, 8.0Hz), 5.09 (1H,
pound was prepared from (l)-(R)-8c as described above in dd, J=6.9, 6.6Hz), 5.13 (2H, s), 6.29 (1H, s), 6.58 (1H, s), 6.81
60% yield. MS (EI) m/z: 404 (MH+), 282. HR-MS (EI) m/z: (1H, d, J=8.3Hz), 6.98 (1H, dd, J=8.3, 2.2Hz), 7.28–7.48 (6H,
404.2201 (Calcd for C26H30NO3 (MH+): 404.2226). IR (neat) m). 13C-NMR (75MHz, CDCl3) δ: 17.7, 18.5, 27.7, 33.2, 39.6,
cm−1: 1610, 1511. [α]D27 +15.2 (c=1.00, benzene). 1H-NMR 41.2, 55.6, 55.7, 57.1, 69.8, 70.4, 70.7, 110.2, 111.2, 111.9, 113.4,
(300MHz, CDCl3) δ: 2.50 (3H, s), 2.57 (1H, m), 2.69–2.85 126.1, 126.8, 127.8, 128.1, 128.4, 129.6, 132.6, 134.4, 136.5,
(2H, m), 2.71 (1H, dd, J=13.7, 7.4Hz), 3.06 (1H, dd, J=13.7, 146.8, 147.6, 153.3, 160.2.
5.2Hz), 3.16 (1H, m), 3.60 (1H, dd, J=7.4, 5.2Hz) 3.77 (3H, s),
3.84 (3H, s), 4.77 (1H, d, J=12.4Hz), 4.84 (1H, d, J=12.4Hz),
(d)-(S)-12. [α]D28 +91.0 (c=1.00, benzene).
(R)-1-(4-(Benzyloxy)-3-bromobenzyl)-6,7-dimethoxy-2-
6.01 (1H, s), 6.57 (1H, s), 6.79 (2H, d, J=8.8Hz), 6.95 (2H, d, methyl-1,2,3,4-tetrahydroisoquinoline ((R)-13) This com-
J=8.8Hz), 7.27–7.34 (5H, m). 13C-NMR (75MHz, CDCl3) δ: pound was prepared from (l)-(R)-12 as described above
25.6, 39.9, 42.6, 46.9, 55.0, 55.7, 64.7, 70.6, 111.5, 113.4, 113.7, in 64% yield. MS (EI) m/z: 482 (MH+), 206. HR-MS (EI)
126.5, 127.0, 127.5, 128.2, 129.2, 130.6, 131.9, 137.2, 145.3, m/z: 482.1323 (Calcd for C26H29BrNO3 (MH+): 482.1331). IR
147.7, 157.7. HPLC: Daicel Chiralpak IB, hexane–2-propanol– (neat) cm−1: 1607. [α]D24 −73.0 (c=1.00, MeOH). 1H-NMR
ethylenediamine=80:20:0.1, 1.0mL/min, UV 270nm, R: (300MHz, CDCl3) δ: 2.50 (3H, s), 2.57 (1H, m), 2.72–2.86
6.8min (dl-9c: 5.8, 6.8min).
(2H, m), 2.73 (1H, dd, J=13.7, 7.1Hz), 3.07 (1H, dd, J=13.7,
5.5Hz), 3.16 (1H, ddd, J=12.4, 8.8, 8.8Hz), 3.58 (3H, s), 3.65
(S)-9c. [α]D24 −11.2 (c=1.00, benzene).
(R)-6-Methoxy-1-(4-methoxybenzyl)-2-methyl-1,2,3,4- (1H, dd, J=7.1, 5.5Hz), 3.84 (3H, s), 5.13 (2H, s), 6.08 (1H,
tetrahydroisoquinolin-7-ol ((R)-5c) This compound was s), 6.55 (1H, s), 6.81 (1H, d, J=8.2Hz), 6.91 (1H, dd, J=8.2,
prepared from (R)-9c as described above in 99% yield. MS 2.2Hz), 7.27–7.48 (6H, m). 13C-NMR (75MHz, CDCl3) δ:
(EI) m/z: 314 (MH+), 192. HR-MS (EI) m/z: 314.1751 (Calcd 25.1, 39.7, 42.3, 46.6, 55.3, 55.4, 64.4, 70.4, 110.6, 111.0, 111.7,
for C19H24NO3 (MH+): 314.1756). IR (neat) cm−1: 1611, 1509. 113.2, 125.8, 126.6, 127.5, 128.2, 128.7, 129.3, 133.8, 134.1,
[α]D27 +40.0 (c=1.00, benzene). 1H-NMR (300MHz, CDCl3) 136.3, 146.2, 147.0, 152.9. HPLC: Daicel Chiralpak IB, hex-
δ: 2.54 (3H, s), 2.55 (1H, m), 2.68–2.78 (2H, m), 2.84 (1H, dd, ane–2-propanol–ethylenediamine=80:20:0.1, 1.0mL/min, UV
J=14.1, 6.0Hz), 3.06 (1H, dd, J=14.1, 6.0Hz), 3.17 (1H, m), 270nm, R: 12.5min (dl-13: 7.9, 12.1min).
3.67 (1H, t, J=6.0Hz) 3.78 (3H, s), 3.84 (3H, s), 6.39 (1H, s),
6.53 (1H, s), 6.79 (2H, d, J=8.5Hz), 7.04 (2H, d, J=8.5Hz).
(S)-13. [α]D26 +75.2 (c=1.00, MeOH).
(R) -1-[4 - ( B en z ylox y) -3 -[(S ) - 6 - methox y-1- (4 -
13C-NMR (75MHz, CDCl3) δ: 24.9, 40.4, 42.3, 46.8, 55.0, methoxybenzyl)-2-methyl-1,2,3,4-tetrahydro-isoquinolin-7-
55.6, 64.5, 110.5, 113.3, 113.8, 124.9, 129.6, 130.3, 131.6, 143.4, yloxy]benzyl]-6,7-dimethoxy-2-methyl-1,2,3,4-tetrahy-
145.3, 157.7.
droisoquinoline (14) This compound was prepared from
(S)-5c and (R)-13 as described above in 35% yield. MS (EI)
(S)-5c. [α]D24 −39.4 (c=1.00, benzene).
2-[(4S)-4,5-Dihydro-4-(1-methylethyl)-2-oxazolyl]-6,7- m/z: 715 (MH+), 593, 296, 206. HR-MS (EI) m/z: 715.3723
dimethoxy-1,2,3,4-tetrahydroisoquinoline
((l)-11) This (Calcd for C45H51N2O6 (MH+): 715.3747). IR (Nujol) cm−1:
compound was prepared from commercially available 6,7-di- 1611, 1509. [α]D23 −46.6 (c=1.00, benzene). 1H-NMR (300MHz,
methoxy-1,2,3,4-tetrahydroisoquinoline and (S)-2-ethoxy-4- CDCl3) δ: 2.46 (6H, s), 2.52–2.63 (2H, m), 2.66–2.85 (5H, m)
isopropyl-4,5-dihydrooxazole17) as described above in 82% 2.70 (1H, dd, J=13.7, 8.0Hz), 2.94 (1H, dd, J=14.0, 5.8Hz),
yield. mp 67–69°C. MS (EI) m/z: 304 (M+), 261. HR-MS 3.07 (1H, dd, J=13.7, 5.2Hz), 3.12–3.15 (2H, m), 3.52 (3H, s),
(EI) m/z: 304.1803 (Calcd for C17H24N2O3 (M+): 304.1787). 3.57–3.64 (2H, m), 3.67 (3H, s), 3.78 (3H, s), 3.82 (3H, s), 5.06
IR (Nujol) cm−1: 1660, 1609. [α]D27 −40.8 (c=1.00, benzene). (2H, s), 6.03 (1H, s), 6.33 (1H, s), 6.52 (1H, s), 6.62 (1H, s),
1H-NMR (300MHz, CDCl3) δ: 0.85 (3H, d, J=6.6Hz), 0.95 6.63 (2H, d, J=8.8Hz), 6.65 (1H, dd, J=8.2, 1.9Hz), 6.72 (1H,
(3H, d, J=6.6Hz), 1.69 (1H, septet of d, J=6.6, 6.6Hz), 2.79 d, J=1.9Hz), 6.82 (1H, d, J=8.2Hz) 6.89 (2H, d, J=8.8Hz),
(2H, t, J=6.0Hz), 3.59 (1H, dt, J=11.5, 6.0Hz), 3.65 (1H, dt, 7.22–7.32 (5H, m). 13C-NMR (75MHz, CDCl3) δ: 25.2, 25.8,
J=11.5, 6.0Hz), 3.82 (1H, ddd, J=8.8, 6.6, 6.6Hz), 3.84 (3H, s), 40.0, 40.4, 42.4, 42.6, 46.7, 47.2, 54.8, 55.3, 55.5, 55.7, 64.2,
3.85 (3H, s), 4.01 (1H, dd, J=8.0, 6.6Hz), 4.27 (1H, dd, J=8.8, 64.5, 70.8, 110.7, 111.0, 112.1, 113.2, 114.8, 117.6, 120.6, 124.7,
8.0Hz), 4.47 (1H, d, J=17.0Hz), 4.53 (1H, d, J=17.0Hz), 6.56 125.9, 126.8, 127.4, 128.1, 129.0, 129.3, 129.8, 130.1, 131.4,
(1H, s), 6.61 (1H, s). 13C-NMR (75MHz, CDCl3) δ: 17.3, 18.5, 133.3, 137.2, 143.9, 146.2, 146.2, 147.0, 147.4, 148.4, 157.5.
27.7, 32.9, 42.6, 46.7, 55.5, 69.7, 70.2, 108.7, 111.2, 124.8, 125.8,
147.2, 147.2, 160.6.
1-epi-Neferine This compound was prepared from 14
as described above in 68% yield. mp 69–72°C. UV λmax
(methanol) nm (logε): 227 (4.48), 284 (3.94). MS (EI) m/z:
(d)-11. mp 64–65°C. [α]D26 +37.8 (c=1.00, benzene).
(1R)-1-[4-(Benzyloxy)-3-bromobenzyl]-2-[(4S)-4,5-di- 625 (MH+), 503, 206. HR-MS (EI) m/z: 625.3279 (Calcd for
hydro-4-(1-methylethyl)-2-oxazolyl]-6,7-dimethoxy-1,2,3,4- C38H45N2O6 (MH+): 625.3278). IR (KBr) cm−1: 3422, 1611,
tetrahydroisoquinoline ((l)-(R)-12) This compound was 1510. [α]D22 −27.8 (c=1.00, CHCl3). 1H-NMR (500MHz,
prepared from (l)-11 and 7b as describe above in 75% yield. CDCl3) δ: 2.46 (3H, s, NCH3-2′), 2.49 (3H, s, NCH3-2), 2.55
MS (EI) m/z: 579 (MH+), 303. HR-MS (EI) m/z: 579.1853 (1H, ddd, J=16.0, 4.5, 4.5Hz, H-4′), 2.62 (1H, ddd, J=16.5,
(Calcd for C31H36BrN2O4 (MH+): 579.1858). IR (neat) cm−1: 5.0, 5.0Hz, H-4) 2.67 (1H, dd, J=14.0, 8.0Hz, H-α′), 2.71–2.75
1650. [α]D22 −104.4 (c=1.00, benzene). 1H-NMR (300MHz, (2H, m, H-3 or 3′), 2.75 (1H, dd, J=14.0, 6.5Hz, H-α), 2.81
CDCl3) δ: 0.79 (3H, d, J=6.9Hz), 0.88 (3H, d, J=6.9Hz), (1H, ddd, J=16.0, 8.0, 6.0Hz, H-4′), 2.83 (1H, ddd, J=16.5,

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8.0, 6.0Hz, H-4), 3.02 (1H, dd, J=14.0, 5.5Hz, H-α), 3.08 (1H, OCH3-6), 3.81 (3H, s, OCH3-7′), 5.99 (1H, s, H-8′), 6.38 (1H,
dd, J=14.0, 5.0Hz, H-α′), 3.11–3.17 (2H, m, H-3 or 3′), 3.55 s, H-8), 6.51 (1H, s, H-5′), 6.55 (1H, d, J=2.0Hz, H-10′), 6.63
(3H, s, OCH3-6′) 3.61 (1H, dd, J=8.0, 5.0Hz, H-1′) 3.63 (1H, (1H, s, H-5), 6.69 (2H, d, J=8.5Hz, H-11, 13), 6.70 (1H, dd,
dd, J=6.5, 5.5Hz, H-1) 3.72 (3H, s, OCH3-12), 3.80 (3H, s, J=8.0, 2.0Hz, H-14′), 6.85 (1H, d, J=8.0Hz, H-13′), 6.90
OCH3-6), 3.82 (3H, s, OCH3-7′), 6.01 (1H, s, H-8′), 6.39 (1H, (2H, d, J=8.5Hz, H-10, 14). 13C-NMR (125MHz, CDCl3) δ:
s, H-8), 6.53 (1H, s, H-5′), 6.58 (1H, d, J=2.0Hz, H-10′), 6.64 25.2, 26.0, 39.9, 40.6, 42.5, 42.7, 46.7, 47.0, 55.1, 55.5, 55.7,
(1H, s, H-5), 6.68 (1H, dd, J=8.0, 2.0Hz, H-14′), 6.70 (2H, 55.8, 64.4, 64.8, 110.9, 111.1, 112.3, 113.4, 115.4, 119.1, 120.0,
d, J=8.5Hz, H-11, 13), 6.85 (1H, d, J=8.0Hz, H-13′), 6.93 125.3, 130.4, 131.0, 131.3, 131.9, 142.8, 144.6, 145.4, 146.4,
(2H, d, J=8.5Hz, H-10, 14). 13C-NMR (125MHz, CDCl3) δ: 147.3, 148.9, 157.8. Hydrochloride: mp 187–190°C. Anal. Calcd
25.2, 26.0, 39.9, 40.6, 42.5, 42.7, 46.7, 47.0, 55.1, 55.5, 55.7, for C38H46Cl2N2O6·H2O: C, 63.77; H, 6.76; N, 3.91. Found: C,
55.8, 64.4, 64.8, 110.9, 111.1, 112.3, 113.4, 115.4, 119.1, 120.0, 63.92; H, 6.64; N, 3.94.
125.3, 125.6, 129.0, 130.4, 130.5, 131.0, 131.4, 131.9, 142.8,
Determination of Locomotor Activity Experiments
144.6, 145.3, 146.3, 147.2, 148.9, 157.8. Hydrochloride: mp were undertaken in accordance with the Guiding Principles
181–183°C. Anal. Calcd for C38H46Cl2N2O6·1.5H2O: C, 62.98; for Care and Use of Laboratory Animals as approved by The
H, 6.82; N, 3.87. Found: C, 63.51; H, 6.83; N, 3.97.
Japanese Pharmacological Society. The study protocol was ap-
(S ) -1- (4 - ( B en z ylox y) -3 - ((R) - 6 -methox y-1- (4 - proved by the Ethics Committee of the Yokohama College of
methoxybenzyl)-2-methyl-1,2,3,4-tetrahydroisoquinolin-7- Pharmacy (Yokohama, Japan). Male ICR mice (age, 5 weeks)
yloxy)benzyl)-6,7-dimethoxy-2-methyl-1,2,3,4-tetrahy- were purchased from SLC Japan Inc. (Shizuoka, Japan). Mice
droisoquinoline (15) This compound was prepared from were housed in groups of five under a controlled 12-h–12-h
(R)-5c and (S)-13 as described above in 30% yield. [α]D27 light–dark cycle (light from 7 a.m. to 7 p.m.) with a room tem-
+43.4 (c=1.00, benzene).
perature of 23 1°C and humidity of 55 5%. Mice had free
1′-epi-Neferine This compound was prepared from access to food and water. Each mouse was only used once. All
15 in 84% yield. mp 77–80°C. [α]D25 +24.4 (c=1.00, drugs were injected via the intraperitoneal (i.p.) route. Mice in
CHCl3). Hydrochloride: mp 185–189°C. Anal. Calcd for the control group received saline. The locomotor activity of
C38H46Cl2N2O6·1.5H2O: C, 62.98; H, 6.82; N, 3.87. Found: C, mice was measured using a digital counter with an infrared
63.32; H, 6.93; N, 3.92.
sensor (NS-AS01, Neuroscience Inc., Tokyo, Japan) follow-
(S ) -1- (4 - ( B en z ylox y) - 3 - ((S ) - 6 - methox y-1- (4 - ing the method described by previous reports.12) An infrared
methoxybenzyl)-2-methyl-1,2,3,4-tetrahydroisoquinolin-7- sensor was set over an open-top clear polycarbonate cage
yloxy)benzyl)-6,7-dimethoxy-2-methyl-1,2,3,4-tetrahy- (22.5×33.8×14.0cm) into which each mouse was placed. Lo-
droisoquinoline (16) This compound was prepared from comotor activity was determined over 60min. The apparatus
(S)-5c and (S)-13 as described above in 45% yield. [α]D24 was used to detect and record a digital count of the horizonal
−46.4 (c=1.00, benzene). MS (EI) m/z: 715 (MH+), 593, 296, movements of animals. Results are shown as means S.E.M.
206. HR-MS (EI) m/z: 715.3723 (Calcd for C45H51N2O6 (MH+): of 5–9 mice in behavioral studies. Results were analyzed by
1
715.3725). IR (Nujol) cm−1: 1611, 1509. H-NMR (300MHz, one-way analysis of variance (ANOVA) followed by Dunnet’s
CDCl3) δ: 2.45 (3H, s), 2.46 (3H, s), 2.50–2.62 (2H, m), multiple comparison post-hoc test.
2.66–2.84 (5H, m) 2.76 (1H, dd, J=14.3, 6.5Hz), 2.92 (1H,
dd, J=14.3, 5.8Hz), 3.02–3.15 (3H, m), 3.54 (3H, s), 3.57–3.64
Acknowledgments We thank Dr. M. Sugiura and Dr. A.
(2H, m), 3.68 (3H, s), 3.78 (3H, s), 3.80 (3H, s), 5.06 (2H, Takeuchi (Kobe Pharmaceutical University) for NMR and MS
s), 6.06 (1H, s), 6.34 (1H, s), 6.50 (1H, s), 6.62 (1H, s), 6.63 spectra. This work was supported by MEXT/JSPS KAKENHI
(2H, d, J=8.8Hz), 6.66 (1H, dd, J=8.2, 2.2Hz), 6.72 (1H, Grant Number 19710194.
d, J=2.2Hz), 6.81 (1H, d, J=8.2Hz) 6.88 (2H, d, J=8.8Hz),
7.22–7.32 (5H, m). 13C-NMR (75MHz, CDCl3) δ: 25.2, 25.9,
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