First total syntheses of (±)-isopiline, (±)-preocoteine, (±)-oureguattidine and (±)-3-methoxynordomesticine and the biological activities of (±)-3-methoxynordomesticine

Article abstract of DOI:10.1248/cpb.57.368

A convenient and economical synthesis of 4-hydroxy-2,3- dimethoxybenzaldehyde has been developed. This was used as the starting material for the first total syntheses of (±)-isopiline, (±)-preocoteine, (±)-oureguattidine and (±)-3-methoxynordomesticine in

Full text of DOI:10.1248/cpb.57.368

368
Chem. Pharm. Bull. 57(4) 368—376 (2009)
Vol. 57, No. 4
First Total Syntheses of (ꢀ)-Isopiline, (ꢀ)-Preocoteine,
(ꢀ)-Oureguattidine and (ꢀ)-3-Methoxynordomesticine and the
Biological Activities of (ꢀ)-3-Methoxynordomesticine
Surachai NIMGIRAWATH, ,a Phansuang UDOMPUTTIMEKAKUL,a Thongchai TAECHOWISAN,b
*
c
Asawin WANBANJOB,a and Yuemao SHEN
a Department of Chemistry, Faculty of Science, Silpakorn University; b Department of Microbiology, Faculty of Science,
Silpakorn University; Nakorn Pathom 73000, Thailand: and c Kunming Institute of Botany, The Chinese Academy of
Sciences; Kunming 650204, P. R. China.
Received November 28, 2008; accepted January 29, 2009; published online January 30, 2009
A convenient and economical synthesis of 4-hydroxy-2,3-dimethoxybenzaldehyde has been developed. This
was used as the starting material for the first total syntheses of (ꢀ)-isopiline, (ꢀ)-preocoteine, (ꢀ)-oureguatti-
dine and (ꢀ)-3-methoxynordomesticine in which the key step involved formation of ring C of the aporphines by a
radical-initiated cyclisation. Although (ꢀ)-3-methoxynordomesticine possesses weak antimicrobial activity, it
inhibits the production of nitric oxide (NO), prostaglandin (PG)E2, tumor necrosis factor (TNF)-a, interleukin
(IL)-1b and IL-6 and the expression of inducible nitric oxide synthase (iNOS) and cycloxygenase (COX)-2 in
macrophages stimulated with LPS in vitro.
Key words alkaloid; aporphine; isoquinoline; synthesis; antimicrobial; anti-inflammatory activity
The aporphine alkaloids constitute a large group in the work, only one synthesis of aldehyde (2e) had been reported.
family of isoquinoline alkaloids and occur in a large number The synthesis, involving a Reimer–Tiemann formylation on
of plant families. They also exhibit a large variety of oxy- 2,3-dimethoxyphenol, was extremely tedious and ineffi-
genation patterns and biological activities. Aporphines with a cient,2) making laboratory scale preparation of this aldehyde
1-hydroxy-2,3-dimethoxy-substituted ring A are rather rare by this route not an attractive undertaking. We therefore
and occur in a restricted number of species.1) The structures sought to develop a more attractive and economical synthesis
of all these alkaloids were proposed based on spectroscopic of 2e. Thus, nitration of vanillin acetate with fuming nitric
analysis and conversion to known aporphines. No total syn- acid followed by hydrolysis gave 2b3) which was reduced
thesis of any member of the aporphines in this group has with iron powder and iron(II) sulfate to 2c. This was con-
ever been reported and the minute quantities of the natural verted to 2d under standard diazotization and iodination con-
alkaloids isolated from plant sources make it impossible to ditions. Treatment of 2d with sodium methoxide in the pres-
carry out any biological activity evaluation of these alka- ence of copper(II) chloride gave 2e in good yield. Benzyla-
loids. In view of this limitation, it was therefore highly desir- tion then afforded the desired 2f. This was converted into ni-
able to carry out the total syntheses of a number of these al- trostyrene (2g) which was reduced with lithium aluminium
kaloids to confirm their structures and to evaluate their bio- hydride to the desired amine (2h). All steps in the above syn-
logical activities.
thesis can be carried out safely on a laboratory scale. With
the desired amine (2h) available in ample quantity, it was de-
cided to carry out total syntheses of the racemic forms of
Results and Discussion
Total Syntheses of Aporphine Alkaloids The lack of (ꢀ)-isopiline (1a),4) (ꢁ)-preocoteine (1b),5) (ꢀ)-oureguatti-
synthesis of this group of aporphine alkaloids may be part- dine (1c)6) and (ꢁ)-3-methoxynordomesticine (1d).7)
ly due to the difficult access to 4-hydroxy-2,3-dimethoxy-
The strategy employed was based on the construction of
benzaldehyde (2e), the key starting compound necessary for the ring C by a radical-initiated cylisation.8) Such a cyclisa-
the conversion to the key intermediate, 4-benzyloxy-2,3- tion on a 7-benzyloxyisoquinoline precursor has not hitherto
dimethoxyphenethyl amine (2h). At the beginning of this been reported. It was anticipated from the outset that due to
Reaction conditions: (A) (i) fuming HNO3, dry ice/acetone; (ii) 10% NaOH; (B) Fe(II)SO4·7H2O, iron power/ethanol–water; (C) (i) NaNO2/20% HCl; (ii) KI; (D) CuCl2/DMF,
NaOCH3/methanol; (E) BnCl, anh. K2CO3/ethanol; (F) CH3NO2, NH4OAc/acetic acid; (G) LAH/THF.
Chart 1. Synthesis of 4-Benzyloxy-2,3-dimethoxyphenethylamine (2h)
∗ To whom correspondence should be addressed. e-mail: surachai@su.ac.th
© 2009 Pharmaceutical Society of Japan

April 2009
369
steric hindrance of the benzyloxy group at that position, the (5a—d) were converted respectively to dihydroisoquinolines
yields of the cyclisation step would be quite low. Condensa- (6a—d) which were substances of extremely low stability.
tion of amine (2h) with acid chlorides (3d—f) afforded in These were quickly reduced with sodium borohydride to give
good yields amides (5a—c) respectively. Due to its poor sol- tetrahydroisoquinolines (7a—d). (7a), (7c) and (7d) were
ubility, acid (4) cannot be converted into the corresponding converted into the trifluoroacetyl derivatives (8a), (8c) and
acid chloride. Condensation of amine (2h) and acid (4) was (8d) while 7b was converted into urethane (8b). Treatment of
carried out in boiling xylene with water removal to give 8a, 8b, 8c and 8d with tributyltin hydride in the presence of
amide (5d). By a Bischler–Napieralski reaction, amides azobis(isobutyronitrile) gave noraporphines (9a), (9b), (9c)
and (9d) respectively in 8.3—10.9% yields. In all cases, cy-
clisation took place with concurrent loss of the benzyl pro-
tecting group at C-1 of the aporphine nucleus and the forma-
tion of the corresponding hydrogenolysis products (8e—h) in
13.1—41.6% yields. The structure of 9a was supported by
the presence of a doublet at dH 8.39, characteristic of the pro-
ton present at C-11 of the aporphine nucleus. Similarly, the
1H-NMR spectra of 9b, 9c and 9d also exhibited such low-
field proton signals. Removal of the trifluoroacetyl groups
from 9a and 9d gave (ꢂ)-isopiline (1a) and (ꢂ)-3-
methoxynordomesticine (1d) respectively. On the other hand,
Reaction condition: (H) SOCl2/benzene.
Chart 2. Syntheses of Acid Chlorides 3d—3f
reduction of 9b with lithium aluminium hydride gave (ꢂ)-
Reaction conditions: (I) 10% NaHCO3/chloroform; (J) reflux/xylene.
Chart 3. Syntheses of Amides 5a—5d
Reaction conditions: (K) POCl3/acetonitrile; (L) NaBH4/ethanol; (M) (CF3CO)2O, Et3N/chloroform (7a, 7c, 7d→8a, 8c, 8d); (N) ClCOOCH3, Et3N/chloroform (7b→8b); (O)
Bu3SnH, AIBN/dry toluene; (P) K2CO3/methanol–water; (Q) LAH/THF; (R) H2, Pd/C/ethanol.
Chart 4. Syntheses of Aporphine Alkaloids 1a—1d

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Vol. 57, No. 4
Fig. 3. Effect of (ꢂ)-3-Methoxynordomesticine on PEG2 Production in
LPS-Induced RAW 264.7 Macrophage for 24 h
Fig. 1. Structures of (ꢂ)-Isopiline (1a), (ꢂ)-Preocoteine (1b), (ꢂ)-
Oureguattidine (1c) and (ꢂ)-3-Methoxynordomesticine (1d)
The values are the means of at least 3 determinationsꢂS.D. Probability level (Stu-
dent’s t-test): ∗ pꢃ0.05 vs. LPS-treated group.
Fig. 2. Evaluation of Nitrite Production by RAW 264.7 Cells Stimulated
for 24 h with LPS Alone or Combination with Increasing Concentrations
(1—5 mg/ml) of (ꢂ)-3-Methoxynordomesticine
Fig. 4. Effect of (ꢂ)-3-Methoxynordomesticine on iNOS Protein Produc-
tion by LPS-Induced RAW 264.7 Macrophage for 24 h
The values are the means of at least 3 determinationsꢂS.D. Probability levels (Stu-
dent’s t-test): ∗ pꢃ0.05 vs. LPS-treated group.
preocoteine (1b) while catalytic removal of the benzyl group
of (9c) at C-9 followed by removal of the trifluoroacetyl
1
group gave (ꢂ)-oureguattidine (1c). The H- and 13C-NMR
spectral data of synthetic (ꢂ)-isopiline (1a), (ꢂ)-preocoteine
(1b), (ꢂ)-oureguattidine (1c) and (ꢂ)-3-methoxynordomes-
ticine (1d) were identical in all respects with those of the re-
spective natural alkaloids.
ꢀ
Biological Activities of ( )-3-Methoxynordomesticine
(ꢂ)-3-Methoxynordomesticine was more effective against
Gram-negative Escherichia coli ATCC10536 than Gram-pos-
itive Staphylococcus aureus ATCC25932 and budding yeast
Candida albicans ATCC90028. The MIC value of 256 mg/ml
was obtained for E. coli ATCC10536, whereas the MIC val-
ues lower or equal to 512 mg/ml were obtained for S. aureus
Fig. 5. Effect of (ꢂ)-3-Methoxynordomesticine and LPS-Induced COX-2
Protein Expression in RAW 264.7 Cells
ATCC25932 and C. albicans ATCC90028. The MMC value domesticine on NO and PGE2 production, we investigated
lower or equal to 512 mg/ml were obtained in all tested mi- their effects on inducible nitric oxide synthase (iNOS) and
croorganisms. In the course of our studies on anti-inflam- cyclooxygenase (COX)-2 expression levels, respectively. In
matory, we have found that (ꢂ)-3-methoxynordomesticine response to LPS, the iNOS and COX-2 inductions were
inhibits nitric oxide (NO) production in murine macro- markedly increased, and (ꢂ)-3-methoxynordomesticine sig-
phage RAW 264.7 cells stimulated with lipopolysaccharide nificantly decreased the iNOS and COX-2 protein expression
(LPS) (Fig. 2), we then investigated the effect of (ꢂ)-3- in a concentration-dependent manner (Figs. 4, 5). In contrast
methoxynordomesticine on the release of prostaglandin E2 to iNOS and COX-2, (ꢂ)-3-methoxynordomesticine had no
(PGE2). Compared with the untreated control, LPS (1 mg/ml) effect on the expression of b-actin and COX-1 (data not
induced a great production of PGE2 in RAW 264.7 cells. shown). This finding indicates that (ꢂ)-3-methoxynordomes-
(ꢂ)-3-Methoxynordomesticine (1—5 mg/ml) inhibited the ticine could suppress NO and PGE2 production in LPS-stim-
production of PEG2 in RAW 264.7 cells stimulated with LPS ulated RAW 264.7 cells by inhibiting iNOS and COX-2 pro-
in a concentration-dependent manner (Fig. 3). To elucidate tein expression, respectively. It has been reported that cy-
the mechanism of the inhibitory effect of (ꢂ)-3-methoxynor- tokines such as tumor necrosis factor (TNF)-a, interleukin

April 2009
371
3-methoxynordomesticine at all concentrations used (1—
5 mg/ml) in the absence or presence of LPS.
In conclusion, we found that (ꢂ)-3-methoxynordomes-
ticine not only possesses antibacterial activity agaianst E.
coli but also possesses anti-inflammatory activity. It inhibits
the production of NO, PGE2, TNF-a, IL-1b and IL-6 and the
expression of iNOS and COX-2 in macrophages stimulated
with LPS in vitro. From these results it is expected that (ꢂ)-
3-methoxynordomesticine could be potentially useful for the
treatment of inflammatory diseases.
Experimental
Melting points were determined on a Stuart Scientific SMP 2 melting
point apparatus and are uncorrected. Infrared spectra were recorded on
CH2Cl2-films with a Perkin Elmer Spectrum GX FT-IR spectrophotometer.
Ultraviolet spectra were recorded on methanol solutions with a Perkin-Elmer
Lambda 35 UV–VIS spectrophotometer. 1H- and 13C-NMR spectra were
recorded on (D) chloroform solutions at 300 MHz for 1H and 75 MHz for
13C with a Bruker AVANCE 300 spectrometer. Tetramethylsilane was used
as the internal standard. Mass spectra were recorded on a POLARIS Q or
HEWLETT PACKARD 5973 mass spectrometer. Elemental analyses were
performed on a Perkin Elmer 2400 Elemental Analyser.
Fig. 6. Effect of (ꢂ)-3-Methoxynordomesticine on LPS-Induced TNF-a
Production by RAW 264.7 Cells
The values are the means of at least 3 determinationsꢂS.D. Probability level (Stu-
dent’s t-test): ∗ pꢃ0.05 vs. LPS-treated group.
4-Hydroxy-2-nitro-3-methoxybenzaldehyde (2b) Vanillin acetate (2a)
(50.0 g, 257.7 mmol) was slowly added with stirring to fuming nitric acid
(200 ml) at ꢀ10—5 °C and stirring continued for 10 min. The mixture was
poured into ice-water (800.0 g). The yellow precipitate was filtered, washed
with some water and heated with stirring to boiling in 10% NaOH (400 ml).
After cooling to room temperature, the mixture was acidified with conc. HCl
and the crude product was filtered and recrystallized from water to give 2b
as a pale yellow solid (44.9 g, 88.6%), mp 136—138 °C [lit.3) mp 136 °C].
1H-NMR d: 3.92 (3H, s, O–CH3), 7.19 (1H, d, Jꢄ8.5 Hz, Ar–H), 7.52 (1H,
d, Jꢄ8.5 Hz, Ar–H), 9.72 (1H, s, CHO). 13C-NMR d: 66.00 (q), 122.29 (d),
123.08 (s), 133.74 (d), 144.04 (s), 148.55 (s), 162.07 (s), 190.82 (d).
2-Amino-4-hydroxy-3-methoxybenzaldehyde (2c) A mixture of 2b
(16.0 g, 81.2 mmol), iron(II) sulfate heptahydrate (4.0 g), iron powder
(40.0 g), ethanol (150 ml) and water (50 ml) was refluxed for 4 h. The iron
was filtered and the solvent removed under vacuum and the residue recrys-
tallized from water (about 100 ml) to give 2c as a pale brown solid (11.5 g,
84.8%), mp 137—138 °C [lit.10) mp 138—139 °C]. 1H-NMR d: 3.81 (3H, s,
O–CH3), 6.39 (1H, d, Jꢄ8.6 Hz, Ar–H), 7.20 (1H, d, Jꢄ8.6 Hz, Ar–H), 9.71
(1H, s, CHO). 13C-NMR d: 58.81 (q), 105.72 (d), 113.01 (s), 131.77 (d),
132.17 (s), 144.96 (s), 154.87 (s), 191.05 (d).
Fig. 7. Effect of (ꢂ)-3-Methoxynordomesticine on IL1-b Production by
RAW 264.7 Cells
The values are the means of at least 3 determinationsꢂS.D. Probability level (Stu-
dent’s t-test): ∗ pꢃ0.05 vs. LPS-treated group.
2-Iodo-4-hydroxy-3-methoxybenzaldehyde (2d) A solution of sodium
nitrite (9.5 g) in water (80 ml) was added to a stirred solution of 2c (21.0 g,
125.7 mmol) in 20% HCl (250 ml) at 0—5 °C and stirring continued for
15 min. A solution of potassium iodide (100 g) in water (300 ml) was slowly
added to the diazonium solution and stirring continued overnight. Chloro-
form (600 ml) and excess sodium thiosulfate were added and the chloroform
layer was separated, washed with brine, then dried over anh. Na2SO4. Re-
moval of the solvent under vacuum followed by recrystallization from
ethanol–water gave 2d as yellow needles (30.6 g, 87.5%), mp 154—156 °C
[lit.11) mp 155—156 °C]. 1H-NMR d: 3.92 (3H, s, O–CH3), 7.03 (1H, d,
Jꢄ8.5 Hz, Ar–H), 7.70 (1H, d, Jꢄ8.5 Hz, Ar–H), 9.97 (1H, s, CHO). 13C-
NMR d: 64.40 (q), 105.13 (s), 121.32 (d), 131.69 (d), 132.03 (s), 151.84 (s),
160.86 (s), 198.87 (d).
Fig. 8. Effect of (ꢂ)-3-Methoxynordomesticine on LPS Induced IL-6 Pro-
duction by RAW 264.7 Cells
4-Hydroxy-2,3-dimethoxybenzaldehyde (2e) A mixture of 2d (4.2 g,
15.1 mmol), CuCl2 (1.0 g) in N,N-dimethylformamide (60 ml) was added to
sodium methoxide (9.7 g) in methanol (60 ml) and the mixture heated in an
oil bath (ꢀ160 °C) for 2 h. Water (100 ml) was added and the mixture acidi-
fied with 6 N HCl. The mixture was filtered and the filtrate extracted with
ethyl acetate (1ꢅ100 ml, 2ꢅ50 ml). The extract was washed with water
(5ꢅ100 ml) then dried over anh. Na2SO4. Removal of the solvent under vac-
uum gave a yellow oil which was chromatograped over silica gel using ben-
zene as eluent to give aldehyde (2e) as pale yellow prisms (2.2 g, 78.0%),
mp 72—73 °C [lit.12) mp 75 °C]. 1H-NMR d: 3.96 (3H, s, O–CH3), 4.01 (3H,
s, O–CH3), 6.81 (1H, d, Jꢄ8.7 Hz, Ar–H), 7.57 (1H, d, Jꢄ8.7 Hz, Ar–H),
10.20 (1H, s, CHO). 13C-NMR d: 61.05 (q), 62.05 (q), 111.54 (d), 122.95
(s), 124.97 (d), 139.25 (s), 156.05 (s), 156.34 (s), 188.67 (d).
The values are the means of at least 3 determinationsꢂS.D. Probability level (Stu-
dent’s t-test): ∗ pꢃ0.05 vs. LPS-treated group.
(IL)-1b and IL-6 are pro-inflammatory in vitro as well as in
vivo.9) The present study also demonstrated that (ꢂ)-3-
methoxynordomesticine has inhibitory effects on the produc-
tion of TNF-a, IL-1b and IL-6 in LPS-stimulated RAW
264.7 cells. As shown in Figs. 6—8, LPS-induced produc-
tions of TNF-a, IL-1b and IL-6 were significantly inhibited
by (ꢂ)-3-methoxynordomesticine in a concentration-depend-
ent manner. In addition, the cytotoxic effect of (ꢂ)-3-
methoxynordomesticine was evaluated in the absence or
presence of LPS (more than 95% cell viability). There is no
significant difference on cell viability when treated with (ꢂ)-
4-Benzyloxy-2,3-dimethoxybenzaldehyde (2f) A mixture of 2e (5.0 g,
27.5 mmol), benzyl chloride (3.8 g), anhydrous potassium carbonate (4.2 g)
in ethanol (30 ml) was refluxed for 5 h. The potassium carbonate was fil-
tered, and the filtrate cooled to room temperature to give 2f as pale yellow

372
Vol. 57, No. 4
prisms (6.9 g, 92.4%), mp 63—64 °C [lit.12) liquid]. 1H-NMR d: 3.90 (3H, s,
O–CH3), 4.02 (3H, s, O–CH3), 5.17 (2H, s, Ph–CH2), 6.79 (1H, d, Jꢄ8.8 Hz,
Ar–H), 7.23—7.50 (5H, m, Ph–H), 7.55 (1H, d, Jꢄ8.8 Hz, Ar–H), 10.24
(1H, s, CHO). 13C-NMR d: 60.98 (q), 62.35 (q), 70.83 (t), 108.93 (d),
123.48 (s), 124.00 (d), 127.28 (d), 128.25 (d), 128.70 (d), 136.02 (s), 141.98
(s), 157.07 (s), 158.43 (s), 188.77 (d).
(d), 125.02 (s), 126.58 (s), 127.25 (d), 127.93 (d), 128.57 (d), 137.05 (s),
142.77 (s), 148.70 (s), 148.93 (s), 151.78 (s), 151.84 (s), 170.14 (s). EI-MS
m/z (%): 544 [M]ꢁ (3), 270 (8), 179 (17), 91 (100). Anal. Calcd for
C27H30BrNO6: C, 59.56; H, 5.55; N, 2.57, Found: C, 59.31; H, 5.72; N, 2.68.
2-(5-Benzyloxy-2-bromophenyl)-N-(4-benzyloxy-2,3-dimethoxy-
phenethyl)acetamide (5c) In a similar manner, 5c was obtained as a pale
yellow solid from ethanol in 72.9% yield, mp 127—128 °C. UV lmax
(MeOH) nm (log e): 206 (4.87), 227 (4.33), 280 (3.22). IR (CH2Cl2-film)
cmꢀ1: 3300, 3065, 3033, 2934, 2873, 2827, 1652, 1595, 1571, 1533, 1492,
1467, 1417, 1380, 1280, 1241, 1194, 1171, 1096, 1057, 1016, 910. 1H-NMR
d: 2.69 (2H, t, Jꢄ6.6 Hz, Ar–CH2), 3.41 (2H, apparent q, Jꢄ6.6 Hz,
N–CH2), 3.61 (2H, s, CO–CH2), 3.81 (3H, s, O–CH3), 3.86 (3H, s, O–CH3),
5.02 (2H, s, Ph–CH2), 5.06 (2H, s, Ph–CH2), 5.81 (1H, br s, NH), 6.57 (1H,
d, Jꢄ8.5 Hz, Ar–H), 6.66 (1H, d, Jꢄ8.5 Hz, Ar–H), 6.78 (1H, dd, Jꢄ3.0,
8.8 Hz, Ar–H), 6.95 (1H, d, Jꢄ3.0 Hz, Ar–H), 7.30—7.45 (11H, m,
Ph–Hꢅ10 and Ar–H). 13C-NMR d: 29.52 (t), 40.84 (t), 44.25 (t), 60.83 (q),
61.01 (q), 70.24 (t), 70.98 (t), 109.46 (d), 115.62 (s), 115.84 (d), 117.94 (d),
124.40 (d), 125.12 (s), 127.25 (d), 127.49 (d), 127.91 (d), 128.17 (d), 128.56
(d), 128.65 (d), 133.61 (d), 135.80 (s), 136.34 (s), 137.08 (s), 142.78 (s),
151.74 (s), 151.86 (s), 158.32 (s), 169.44 (s). EI-MS m/z (%): 498 (9), 179
(25), 91 (70). Anal. Calcd for C32H32BrNO5: C, 65.09; H, 5.46; N, 2.37,
Found: C, 64.91; H, 5.62; N, 2.48.
4-Benzyloxy-2,3-dimethoxy-b-nitrostyrene (2g)
A mixture of 2f
(39.7 g, 145.9 mmol), ammonium acetate (29.2 g) and nitromethane (29.2 g)
in acetic acid (440 ml) was refluxed for 2 h, then cooled to room tempera-
ture. Ice-water (2 l) was added to the mixture and the yellow precipitate was
filtered. Drying of the solid in air gave 2g as yellow needles (36.4 g, 79.2%),
mp 80—81 °C [lit.12) mp 83—84 °C]. 1H-NMR d: 3.90 (3H, s, O–CH3), 4.00
(3H, s, O–CH3), 5.18 (2H, s, Ph–CH2), 6.76 (1H, d, Jꢄ8.8 Hz, Ar–H), 7.15
(1H, d, Jꢄ8.8 Hz, Ar–H), 7.30—7.49 (5H, m, Ph–H), 7.75 (1H, d,
Jꢄ13.6 Hz, CH), 8.08 (1H, d, Jꢄ13.6 Hz, CH). 13C-NMR d: 60.99 (q),
61.26 (q), 70.93 (t), 109.28 (d), 117.26 (s), 126.40 (d), 127.26 (d), 128.29
(d), 128.74 (d), 135.28 (d), 136.04 (s), 136.63 (d), 142.81 (s), 154.42 (s),
156.44 (s).
4-Benzyloxy-2,3-dimethoxyphenethylamine (2h)
A solution of 2g
(28.0 g, 88.9 mmol) in dry tetrahydrofuran (350 ml) was slowly added to a
stirred and cooled suspension of lithium aluminium hydride (11.2 g) in dry
tetrahydrofuran (140 ml) and stirring continued for 3 h. Diethyl ether
(280 ml) was added and with continued cooling and vigorous stirring, water
(15 ml), 15% NaOH (15 ml) and water (45 ml) were added in succession.
The pale yellow granular residue was filtered and washed with ether. The
combined organic phase was extracted with 12% HCl (1ꢅ100 ml, 2ꢅ60 ml).
The extract was basified with conc. ammonia to litmus. The oil was sepa-
rated and the aqueous layer was extracted with chloroform (2ꢅ100 ml). The
oil and chloroform layer were combined, then dried over anh. Na2CO3. Re-
moval of the solvent under vacuum gave 2h as a yellow viscous oil (22.1 g,
86.6%) [lit.12) oil]. 1H-NMR d: 2.69 (2H, t, Jꢄ6.9 Hz, CH2), 2.89 (2H, t,
Jꢄ6.9 Hz, CH2), 3.90 (6H, s, O–CH3ꢅ2), 5.09 (2H, s, Ph–CH2), 6.65 (1H, d,
Jꢄ8.5 Hz, Ar–H), 6.79 (1H, d, Jꢄ8.5 Hz, Ar–H), 7.28—7.45 (5H, m,
Ph–H). 13C-NMR d: 33.89 (t), 42.80 (t), 60.79 (q), 61.02 (q), 70.97 (t),
109.29 (d), 124.32 (d), 125.91 (s), 127.25 (d), 127.85 (d), 128.53 (d), 137.15
(s), 142.91 (s), 151.48 (s), 152.20 (s).
2-(2-Bromo-4,5-methylenedioxyphenyl)-N-(4-benzyloxy-2,3-di-
methoxyphenethyl)acetamide (5d)
A solution of acid (4) (11.3 g,
43.6 mmol) and amine (2h) (12.5 g, 43.6 mmol) in xylene (150 ml) was re-
fluxed for 24 h with water removal with a Dean-Stark trap. The xylene was
then removed under vacuum to yield a brown sticky residue which was dis-
solved in chloroform (100 ml). The chloroform layer was washed with 5%
HCl (3ꢅ100 ml), water (150 ml), 10% NaCO3 (3ꢅ150 ml) and then dried
over anh. Na2SO4. Removal of the solvent under vacuum gave a residue
which was recrystallized with ethanol to give 5d as white prisms (8.3 g,
36.1%), mp 139—141 °C. UV lmax (MeOH) nm (log e): 212 (4.48), 228 sh
(4.23), 247 sh (3.99), 282 sh (3.97), 295 (4.01). IR (CH2Cl2-film) cmꢀ1
:
3296, 2921, 1638, 1603, 1500, 1480, 1417, 1380, 1256, 1234, 1196, 1163,
1096, 1056, 1037, 971, 938, 926. 1H-NMR d: 2.70 (2H, t, Jꢄ6.6 Hz,
Ar–CH2), 3.42 (2H, apparent q, Jꢄ6.4 Hz, N–CH2), 3.55 (2H, s, CO–CH2),
3.83 (3H, s, O–CH3), 3.87 (3H, s, O–CH3), 5.08 (2H, s, Ph–CH2), 5.85 (1H,
br s, NH), 5.95 (2H, s, O–CH2–O), 6.61 (1H, d, Jꢄ8.5 Hz, Ar–H), 6.69 (1H,
d, Jꢄ8.5 Hz, Ar–H), 6.75 (1H, s, Ar–H), 6.98 (1H, s, Ar–H), 7.29—7.50
(5H, m, Ph–H). 13C-NMR d: 29.49 (t), 40.81 (t), 43.77 (t), 60.81 (q), 61.02
(q), 70.97 (t), 101.91 (t), 109.45 (d), 111.08 (d), 112.84 (d), 115.40 (s),
124.46 (d), 125.09 (s), 127.25 (d), 127.67 (s), 127.93 (d), 128.58 (d), 137.07
(s), 142.78 (s), 147.64 (s), 147.80 (s), 151.76 (s), 151.85 (s), 169.77 (s). EI-
MS m/z (%): 527 [M]ꢁ (1), 257 (9), 179 (30), 91 (6). Anal. Calcd for
C26H26BrNO6: C, 59.10; H, 4.96; N, 2.65, Found: C, 59.28; H, 4.81; N, 2.40.
1-(2-Bromobenzyl)-7-benzyloxy-5,6-dimethoxy-3,4-dihydroisoquino-
line (6a) A solution of 5a (3.0 g, 6.2 mmol) and phosphorus oxychloride
(10.0 g) in acetonitrile (80 ml) was refluxed for 3 h. The excess reagent and
solvent were removed under vacuum. The residue was shaken with chloro-
form (50 ml) and dilute ammonium hydroxide (50 ml). The chloroform layer
was washed with water (50 ml), then dried over anh. Na2CO3. Removal of
the solvent under vacuum gave 6a as a pale brown viscous oil (2.8 g) which
was unstable and decomposed on standing. It was immediately used in
2-(2-Bromophenyl)-N-(4-benzyloxy-2,3-dimethoxyphenethyl)ac-
etamide (5a) A mixture of 2-bromophenylacetic acid (3.8 g, 17.6 mmol)
and thionyl chloride (6.2 g) in benzene (100 ml) was refluxed for 1 h, then
the solvent and excess thionyl chloride were removed under vacuum. The re-
sulting crude acid chloride (3a) was dissolved in ethanol-free chloroform
(50 ml) and added portionwise to
a mixture of amine (2h) (5.0 g,
17.42 mmol) in chloroform (50 ml) and 10% NaHCO3 (100 ml). The mixture
was stirred at room temperature for 3 h. The chloroform layer was washed
with 10% NaHCO3 (3ꢅ50 ml), water (2ꢅ100 ml), 5% HCl (3ꢅ50 ml),
water (100 ml), then dried over anh. Na2SO4. Removal of the solvent under
vacuum gave
a pale yellow solid which was recrystallized from
benzene–hexane to give amide (5a) as a pale yellow solid (3.6 g, 42.8%), mp
101—103 °C. UV lmax (MeOH) nm (log e): 209 (4.72), 223 sh (4.35), 271
(3.16). IR (CH2Cl2-film) cmꢀ1: 3301, 2926, 1648, 1601, 1542, 1535, 1491,
1
1467, 1439, 1417, 1379, 1279, 1258, 1200, 1096, 1058, 1026. H-NMR d:
2.69 (2H, t, Jꢄ6.6 Hz, Ar–CH2), 3.42 (2H, apparent q, Jꢄ6.5 Hz, N–CH2),
3.67 (2H, s, CO–CH2), 3.80 (3H, s, O–CH3), 3.87 (3H, s, O–CH3), 5.09 (2H,
s, Ph–CH2), 5.79 (1H, br s, NH), 6.58 (1H, d, Jꢄ8.5 Hz, Ar–H), 6.66 (1H, d,
Jꢄ8.5 Hz, Ar–H), 7.10—7.50 (8H, m, Ar–H), 7.54 (1H, d, Jꢄ7.8 Hz,
Ar–H). 13C-NMR d: 29.49 (t), 40.86 (t), 44.02 (t), 60.84 (q), 61.00 (q),
70.98 (t), 109.46 (d), 124.40 (d), 125.10 (s), 127.24 (d), 127.89 (d), 127.93
(d), 128.58 (d), 128.98 (d), 131.78 (d), 133.05 (d), 134.85 (s), 137.09 (s),
142.76 (s), 143.64 (s), 151.73 (s), 151.8 4 (s), 169.64 (s). EI-MS m/z (%):
483 [M]ꢁ (1), 270 (34), 179 (9), 91 (100). Anal. Calcd for C25H26BrNO4: C,
61.99; H, 5.41; N, 2.89, Found: C, 61.75; H, 5.60; N, 3.02.
1
the next step without further purification. H-NMR d: 2.69 and 2.73 (total
2H, 2t, Jꢄ7.6 Hz, CH2 of both conformers), 3.71and 3.82 (total 2H, 2t,
Jꢄ7.6 Hz, CH2 of both conformers), 3.85, 3.88, 3.90 and 3.96 (total 6.7H,
4 s, O–CH3ꢅ2 of both conformers and CH2 of one conformer), 4.15 (1.30H,
s, CH2 of another conformer), 5.01 and 5.12 (total 2H, 2 s, Ph–CH2 of both
conformers), 6.85 (0.6H, s, Ar–H of one conformer), 7.04—7.65 (9.4H, m,
Ph–Hꢅ5 of both conformers and Ar–Hꢅ4.4 of both conformer). 13C-NMR
d: (both conformers) 18.53 (t), 19.04 (t), 24.85 (t), 42.38 (t), 46.79 (t), 48.04
(t), 60.97 (q), 61.00 (q), 61.04 (q), 61.08 (q), 71.12 (t), 71.20 (t), 107.94 (d),
109.21 (d), 120.66 (d), 121.53 (s), 123.90 (s), 124.54 (s), 124.85 (s), 124.92
(s), 127.38 (d), 127.50 (d), 127.60 (d), 127.64 (d), 128.00 (d), 128.02 (d),
128.15 (d), 128.24 (d), 128.57 (d), 129.69 (d), 129.88 (d), 130.20 (d), 131.02
(d), 132.49 (d), 132.85 (d), 133.07 (d), 136.63 (s), 137.45 (s), 140.19 (s),
145.09 (s), 150.04 (s), 150.09 (s), 150.79 (s), 150.88 (s), 163.64 (s), 165.08
(s).
2-(2-Bromo-4,5-dimethoxyphenyl)-N-(4-benzyloxy-2,3-dime-
thoxyphenethyl)acetamide (5b) In a similar manner, 5b was obtained
as a pale yellow solid from benzene–hexane in 65.8% yield, mp 114—
116 °C. UV lmax (MeOH) nm (log e): 207 (4.89), 230 sh (4.26), 282 (3.51),
294 sh (3.30). IR (CH2Cl2-film) cmꢀ1: 3307, 2935, 2843, 1652, 1603, 1506,
1466, 1439, 1417, 1386, 1260, 1219, 1193, 1166, 1096, 1058, 1029, 969.
1H-NMR d: 2.70 (2H, t, Jꢄ6.6 Hz, Ar–CH2), 3.42 (2H, apparent q, Jꢄ
6.5 Hz, N–CH2), 3.60 (2H, s, CO–CH2), 3.82 (3H, s, O–CH3), 3.83 (3H, s,
O–CH3), 3.85 (3H, s, O–CH3), 3.87 (3H, s, O–CH3), 5.09 (2H, s, Ph–CH2),
5.85 (1H, br s, NH), 6.59 (1H, d, Jꢄ8.5 Hz, Ar–H), 6.67 (1H, d, Jꢄ8.5 Hz,
Ar–H), 6.81 (1H, s, Ar–H), 7.01 (1H, s, Ar–H), 7.32—7.43 (5H, m, Ph–H).
13C-NMR d: 29.48 (t), 40.89 (t), 43.50 (t), 56.09 (q), 56.20 (q), 60.80 (q),
61.02 (q), 70.97 (t), 109.40 (d), 113.89 (d), 114.87 (s), 115.57 (d), 124.39
1-(2-Bromo-4,5-dimethoxybenzyl)-7-benzyloxy-5,6-dimethoxy-3,4-di-
hydroisoquinoline (6b) In a similar manner, the unstable 6b was obtained
in 42.1% yield from benzene–hexane as a pale yellow solid, mp 148—
150 °C. 1H-NMR d: 2.67 (2H, t, Jꢄ7.3 Hz, CH2), 3.70 (2H, t, Jꢄ7.3 Hz,
CH2), 3.73 (3H, s, O–CH3), 3.79 (3H, s, O–CH3), 3.83 (3H, s, O–CH3), 3.89
(3H, s, O–CH3), 4.13 (2H, s, CH2), 5.07 (2H, s, Ph–CH2), 6.80 (1H, s,

April 2009
373
Ar–H), 6.93 (1H, s, Ar–H), 7.01 (1H, s, Ar–H), 7.24—7.45 (5H, m, Ph–H).
13C-NMR d: 19.13 (t), 42.16 (t), 47.04 (t), 55.81 (q), 56.00 (q), 60.86 (q),
60.90 (q), 71.09 (t), 107.71 (d), 112.34 (d), 114.07 (s), 115.30 (d), 123.96
(s), 124.72 (s), 127.40 (d), 127.93 (d), 128.47 (d), 129.36 (s), 136.67 (s),
144.81 (s), 148.29 (s), 148.48 (s), 149.96 (s), 150.77 (s), 165.08 (s).
Ar–H), 6.87 (1H, d, Jꢄ3.0 Hz, Ar–H), 7.26—7.49 (11H, m, Ph–Hꢅ10 and
Ar–H). 13C-NMR d: 23.78 (t), 39.53 (t), 42.93 (t), 54.97 (d), 60.46 (q),
60.93 (q), 70.19 (t), 71.11 (t), 108.03 (d), 114.76 (d), 115.58 (s), 118.51 (d),
122.02 (s), 127.31 (d), 127.45 (d), 127.84 (d), 128.10 (d), 128.53 (d), 128.63
(d), 133.57 (d), 133.88 (s), 136.49 (s), 137.22 (s), 139.64 (s), 141.09 (s),
150.45 (s), 151.22 (s), 157.93 (s). Anal. Calcd for C32H32BrNO4: C, 66.90;
H, 5.61; N, 2.44, Found: C, 66.74; H, 5.80; N, 2.58.
1-(2-Bromo-4,5-methylenedioxybenzyl)-7-benzyloxy-5,6-dimethoxy-
1,2,3,4-tetrahydroisoquinoline (7d) In a similar manner, 7d was obtained
in 64.0% yield from ethanol as a pale yellow solid, mp 105—106 °C. UV
lmax (MeOH) nm (log e): 213 (4.34), 230 sh (4.09), 276 sh (3.83), 293
(3.88). IR (CH2Cl2-film) cmꢀ1: 3336, 2935, 1602, 1585, 1500, 1478, 1456,
1436, 1412, 1378, 1343, 1322, 1273, 1231, 1201, 1163, 1112, 1037, 964,
932. 1H-NMR d: 2.62—3.02 (4H, m, CH2ꢅ2), 3.12—3.26 (2H, m, CH2),
3.88 (3H, s, O–CH3), 3.90 (3H, s, O–CH3), 4.13 (1H, dd, Jꢄ3.6, 10.1 Hz, H-
1), 5.09 (2H, s, Ph–CH2), 5.95 (2H, s, O–CH2–O), 6.62 (1H, s, Ar–H), 6.76
(1H, s, Ar–H), 7.03 (1H, s, Ar–H), 7.27—7.48 (5H, m, Ph–H). 13C-NMR d:
23.59 (t), 39.57 (t), 42.52 (t), 55.22 (d), 60.50 (q), 60.95 (q), 71.16 (t),
101.72 (t), 108.02 (d), 111.38(d), 112.91 (d), 114.95 (s), 121.89 (s), 127.29
(d), 127.87 (d), 128.56 (d), 131.45 (s), 133.58 (s), 137.21 (s), 141.15 (s),
147.22 (s), 147.31 (s), 150.52 (s), 151.22 (s). Anal. Calcd for C26H26BrNO5:
C, 60.95; H, 5.11; N, 2.73, Found: C, 60.80; H, 5.28; N, 2.90.
1-(5-Benzyloxy-2-bromobenzyl)-7-benzyloxy-5,6-dimethoxy-3,4-dihy-
droisoquinoline (6c) In a similar manner, 6c was obtained in almost quan-
titative yield as an unstable pale brown viscous oil which decomposed
quickly on standing. It was immediately used in the next step without fur-
ther purification. 1H-NMR d: 2.63 (2H, t, Jꢄ7.5 Hz, CH2), 3.68 (2H, t,
Jꢄ7.5 Hz, CH2), 3.84 (3H, s, O–CH3), 3.90 (3H, s, O–CH3), 4.07 (2H, s,
CH2), 4.94 (2H, s, Ph–CH2), 5.01 (2H, s, Ph–CH2), 6.71 (1H, dd, Jꢄ8.8,
2.8 Hz, Ar–H), 6.82 (1H, s, Ar–H), 6.88 (1H, d, Jꢄ2.8 Hz, Ar–H), 7.27—
7.40 (10H, m, Ph–H), 7.44 (1H, d, Jꢄ8.8 Hz, Ar–H). 13C-NMR d: 19.03 (t),
42.80 (t), 47.09 (t), 60.96 (q), 61.00 (q), 70.13 (t), 71.14 (t), 107.78 (d),
115.20 (d), 116.45 (d), 124.00 (s), 124.81 (s), 127.40 (d), 127.42 (d), 127.97
(d), 128.05 (d), 128.56 (d), 133.35 (d), 136.50 (s), 136.72 (s), 138.65 (s),
144.85 (s), 150.01 (s), 150.77 (s), 158.17 (s), 164.69 (s).
1-(2-Bromo-4,5-methylenedioxybenzyl)-7-benzyloxy-5,6-dimethoxy-
3,4-dihydro isoquinoline (6d) In a similar manner, 6d was obtained in al-
most quantitative yield as an unstable pale brown viscous oil which decom-
posed quickly on standing. It was immediately used in the next step without
1
further purification. H-NMR d: 2.67 (2H, t, Jꢄ7.5 Hz, CH2), 3.70 (2H, t,
2-Trifluoroacetyl-1-(2-bromobenzyl)-7-benzyloxy-5,6-dimethoxy-
1,2,3,4-tetrahydroisoquinoline (8a) Trifluoroacetic anhydride (7.8 g) was
added dropwise to a stirred mixture of 7a (2.8 g, 6.0 mmol) and triethyl-
amine (5.0 g) in chloroform (50 ml) at 0—10 °C. Stirring was continued at
room temperature for 3 h. Chloroform (50 ml) was added and the chloroform
layer was washed with 10% NaHCO3 (5ꢅ60 ml), water (50 ml), 5% HCl
(5ꢅ60 ml) and brine (50 ml), then dried over anh. Na2SO4. Removal of the
solvent under vacuum gave a yellow viscous oil which was chromatographed
over alumina using benzene as eluent to gave 8a as a pale yellow solid
(2.7 g, 78.3%), mp 106—108 °C. UV lmax (MeOH) nm (log e): 206 (4.87),
224 (4.49), 275 (3.52), 283 (3.50). IR (CH2Cl2-film) cmꢀ1: 2938, 1694,
1603, 1587, 1492, 1459, 1438, 1417, 1375, 1350, 1325, 1265, 1244, 1198,
Jꢄ7.5 Hz, CH2), 3.85 (3H, s, O–CH3), 3.91 (3H, s, O–CH3), 4.02 (2H, s,
CH2), 5.06 (2H, s, Ph–CH2), 5.93 (2H, s, O–CH2–O), 6.73 (1H, s, Ar–H),
6.85 (1H, s, Ar–H), 7.01 (1H, s, Ar–H), 7.30—7.42 (5H, m, Ph–H). 13C-
NMR d: 19.05 (t), 42.34 (t), 46.94 (t), 60.94 (q), 60.96 (q), 71.11 (t), 101.71
(t), 107.70 (d), 109.55 (d), 112.56 (d), 114.41 (s), 123.89 (s), 124.84 (s),
127.32 (d), 127.95 (d), 128.55 (d), 130.45 (s), 136.61 (s), 144.93 (s), 147.03
(s), 147.56 (s), 150.06 (s), 150.80 (s), 164.89 (s).
1-(2-Bromobenzyl)-7-benzyloxy-5,6-dimethoxy-1,2,3,4-tetrahydroiso-
quinoline (7a) Sodium borohydride (0.4 g, 10.0 mmol) was added portion-
wise to a stirred solution of 6a (2.8 g, 6.0 mmol) in ethanol (80 ml) and the
mixture was refluxed for 1 h. Removal of the ethanol under vacuum gave a
residue which was shaken with water (50 ml) and chloroform (50 ml). The
chloroform layer was dried. Removal of the solvent gave a crude yellow vis-
cous oil which was chromatograped over alumina using benzene as eluent to
give tetrahydroisoquinoline (7a) as a pale yellow viscous oil (2.4 g, 85.4%).
UV lmax (MeOH) nm (log e): 207 (4.44), 224 sh (4.05), 276 (2.94). IR
(CH2Cl2-film) cmꢀ1: 3332, 2935, 2829, 1601, 1491, 1456, 1437, 1413, 1377,
1342, 1322, 1274, 1240, 1200, 1110, 1026, 916, 751, 697. 1H-NMR d:
2.58—2.78 (2H, m, CH2), 2.84—2.94 (2H, m, CH2), 3.10—3.29 (2H, m,
CH2), 3.86 (3H, s, O–CH3), 3.89 (3H, s, O–CH3), 4.16 (1H, dd, Jꢄ3.3,
10.2 Hz, H-1), 5.06 (2H, s, Ph–CH2), 6.64 (1H, s, Ar–H), 7.00—7.45 (8H,
m, Ph–Hꢅ5 and Ar–Hꢅ3), 7.55 (1H, d, Jꢄ7.8 Hz, Ar–H). 13C-NMR d:
23.79 (t), 39.53 (t), 42.81 (t), 54.94 (d), 60.39 (q), 60.86 (q), 71.02 (t),
107.97 (d), 121.99 (s), 124.88 (s), 127.25 (d), 127.36 (d), 127.79 (d), 128.21
(d), 128.49 (d), 131.95 (d), 132.99 (d), 133.98 (s), 137.21 (s), 138.67 (s),
141.04 (s), 150.40 (s), 151.19 (s). Anal. Calcd for C25H26BrNO3: C, 64.11;
H, 5.59; N, 2.99, Found: C, 64.29; H, 5.42; N, 2.81.
1
1171, 1142, 1119, 1094, 1046, 1027, 905. H-NMR d: 2.70—3.36 (5H, m,
H-3b, CH2ꢅ2), 3.89 (6H, s, O–CH3), 4.00—4.13 (1H, m, H-3a), 5.00 (2H,
s, Ph–CH2), 5.69—5.74 (1H, m, H-1), 6.43 (1H, s, Ar–H), 7.06—7.50 (8H,
m, Ph–Hꢅ5 and Ar–Hꢅ3), 7.54 (1H, d, Jꢄ7.5 Hz, Ar–H). 13C-NMR d:
(both conformers) 21.87 (t), 23.47 (t), 37.10 (t), 39.61 (t), 41.50 (t), 53.66
(d), 60.67 (q), 60.98 (q), 71.02 (t), 107.84 (d), 108.22 (d), 114.47 (s), 118.29
(s), 119.67 (s), 125.41 (s), 127.25 (d), 127.31 (d), 127.71 (d), 128.04 (d),
128.61 (d), 128.64 (d), 128.74 (d), 128.90 (d), 129.88 (s), 130.09 (s), 131.28
(d), 131.46 (d), 132.85 (d), 136.39 (s), 136.81 (s), 141.61 (s), 150.91 (s),
151.30 (s), 155.68 (s). Anal. Calcd for C27H25BrF3NO4: C, 57.46; H, 4.46;
N, 2.48, Found: C, 57.30; H, 4.60; N, 2.62.
1-(2-Bromo-4,5-dimethoxybenzyl)-7-benzyloxy-2-carbomethoxy-5,6-
dimethoxy-1,2,3,4-tetrahydroisoquinoline (8b) Methyl chloroformate
(4.1 g) was slowly added dropwise to a stirred mixture of 7b (3.8 g,
7.2 mmol) and triethylamine (4.4 g) in chloroform (30 ml) at 0—10 °C. Stir-
ring was continued at room temperature for 3 h. Chloroform (50 ml) and
water (50 ml) were added and the chloroform layer was washed with 10%
HCl (6ꢅ100 ml), water (50 ml), then dried over anh. Na2SO4. Removal of
the solvent under vacuum gave a pale yellow viscous oil. The viscous oil
was chromatograped over alumina using benzene as eluent to give 8b as a
white solid (3.2 g, 76.2%), mp 113—114 °C. UV lmax (MeOH) nm (log e):
207 (4.60), 233 sh (3.94), 284 (3.28). IR (CH2Cl2-film) cmꢀ1: 2936, 2843,
1698, 1603, 1509, 1452, 1409, 1382, 1342, 1258, 1220, 1165, 1106, 1088,
1-(2-Bromo-4,5-dimethoxybenzyl)-7-benzyloxy-5,6-dimethoxy-1,2,3,4-
tetrahydroisoquinoline (7b) In a similar manner, 7b was obtained in
98.0% yield as a pale yellow viscous oil. UV lmax (MeOH) nm (log e): 210
(4.80), 229 sh (4.42), 284 sh (3.79), 295 sh (3.57). IR (CH2Cl2-film) cmꢀ1
:
3429, 2935, 2839, 1602, 1506, 1456, 1438, 1413, 1380, 1322, 1259, 1219,
1164, 1109, 1029, 959. 1H-NMR d: 2.60—2.95 (4H, m, CH2ꢅ2), 3.15—
3.27 (2H, m, CH2), 3.82 (3H, s, O–CH3), 3.83 (3H, s, O–CH3), 3.87 (3H, s,
O–CH3), 3.89 (3H, s, O–CH3), 4.15 (1H, dd, Jꢄ3.2, 9.8 Hz, H-1), 5.07 (2H,
s, Ph–CH2), 6.62 (1H, s, Ar–H), 6.74 (1H, s, Ar–H), 7.04 (1H, s, Ar–H),
7.24—7.47 (5H, m, Ph-H). 13C-NMR d: 23.86 (t), 40.12 (t), 42.45 (t), 55.40
(d), 56.12 (q), 56.16 (q), 60.47 (q), 60.93 (q), 71.15 (t), 107.99 (d), 114.40
(d), 114.71 (s), 115.75 (d), 122.11 (s), 127.34 (d), 127.87 (d), 128.55 (d),
130.45 (s), 133.85 (s), 137.24 (s), 141.10 (s), 148.28 (s), 148.37 (s), 150.47
(s), 151.26 (s). Anal. Calcd for C27H30BrNO5: C, 61.37; H, 5.72; N, 2.65,
Found: C, 61.15; H, 5.88; N, 2.78.
1
1029, 986. H-NMR d: 2.51—3.50 (5H, m, H-3b, CH2ꢅ2), 3.43 and 3.65
(total 3H, 2 s, COO–CH3 of both conformers), 3.78, 3.83, 3.85, 3.87, 3.88
and 3.89 (total 12H, 6 s, O–CH3ꢅ4 of both conformers), 3.86—3.98 and
4.22—4.35 (total 1H, m, H-3a), 4.93 and 5.05 (total 2H, 2 s, Ph–CH2 of
both conformers), 5.18—5.35 (1H, m, H-1), 6.26, 6.46, 6.50, 6.59, 7.02 and
7.04 (total 3H, 6 s, Ar–H of both conformers), 7.30—7.50 (5H, m, Ph–H).
13C-NMR d: (both conformers) 22.24 (t), 22.42 (t), 37.12 (t), 38.44 (t),
41.17 (t), 42.02 (t), 52.27 (q), 52.61 (q), 53.32 (d), 54.90 (d), 56.03 (q),
56.12 (q), 56.23 (q), 60.62 (q), 60.69 (q), 60.97 (q), 71.04 (t), 71.14 (t),
108.27 (d), 108.59 (d), 113.76 (d), 114.10 (d), 115.14 (d), 115.29 (d), 121.07
(s), 121.27 (s), 127.15 (d), 127.26 (d), 127.82 (d), 127.96 (d), 128.40 (d),
128.60 (d), 129.59 (s), 129.76 (s), 131.55 (s), 131.66 (s), 137.02 (s), 141.42
(s), 148.10 (s), 148.16 (s), 148.20 (s), 148.33 (s),150.72 (s), 150.85 (s),
151.14 (s), 155.84 (s). 155.98 (s). Anal. Calcd for C29H32BrNO7: C, 59.39;
H, 5.50; N, 2.39, Found: C, 59.54; H, 5.37; N, 2.10.
1-(5-Benzyloxy-2-bromobenzyl)-7-benzyloxy-5,6-dimethoxy-1,2,3,4-
tetrahydroisoquinoline (7c) In a similar manner, 7c was obtained in
92.1% yield as a pale yellow solid, mp 93—96 °C. UV lmax (MeOH) nm
(log e): 206 (4.77), 229 (4.31), 282 (3.43). IR (CH2Cl2-film) cmꢀ1: 3335,
3089, 3064, 3032, 2935, 2829, 1592, 1570, 1492, 1456, 1435, 1413, 1342,
1
1321, 1277, 1240, 1201, 1169, 1111, 1028, 1015, 913. H-NMR d: 2.65—
2.94 (4H, m, CH2ꢅ2), 3.10—3.25 (2H, m, CH2), 3.87 (3H, s, O–CH3), 3.89
(3H, s, O–CH3), 4.14 (1H, dd, Jꢄ3.3, 9.8 Hz, H-1), 5.02 (2H, s, Ph–CH2),
5.08 (2H, s, Ph–CH2), 6.65 (1H, s, Ar–H), 6.76 (1H, dd, Jꢄ3.0, 8.8 Hz,
2-Trifluoroacetyl-1-(5-benzyloxy-2-bromobenzyl)-7-benzyloxy-5,6-
dimethoxy-1,2,3,4-tetrahydroisoquinoline (8c) In a similar manner to

374
Vol. 57, No. 4
both conformers), 6.11 and 6.28 (total 1H, 2 s, Ar–H of both conformers),
6.52—6.65 (2H, m, Ar–H of both conformers), 6.76 (1H, apparent t,
Jꢄ8.5 Hz, Ar–H of both conformers), 7.26—7.44 (5H, m, Ph–H of both
conformers). 13C-NMR d: (both conformers) 22.21 (t), 37.53 (t), 38.69 (t),
41.98 (t), 42.43 (t), 52.45 (q), 52.58 (q), 55.75 (q), 55.84 (q), 55.90 (q),
56.02 (d), 56.12 (d), 60.62 (q), 60.69 (q), 60.92 (q), 70.79 (t), 71.04 (t),
108.44 (d), 108.72 (d), 110.85 (d), 111.01 (d), 112.73 (d), 120.89 (s), 121.32
(s), 121.79 (d), 121.9 1(d), 127.11 (d), 127.21 (d), 127.86 (d), 127.91 (d),
128.53 (d), 130.63 (s), 131.71 (s), 136.96 (s), 141.13 (s), 141.32 (s), 147.65
(s), 147.73 (s), 148.61 (s), 150.34 (s), 150.58 (s), 150.77 (s), 151.11 (s),
155.89 (s), 156.00 (s). Noraporphine (9b) was obtained in 8.3% yield as a
pale yellow solid, mp 212—213 °C. UV lmax (MeOH) nm (log e): 222
(4.54), 271 sh (3.91), 280 (4.00), 304 (4.12), 315 (4.12). IR (CH2Cl2-film)
cmꢀ1: 3383, 2936, 2849, 1686, 1609, 1592, 1513, 1459, 1448, 1396, 1342,
1250, 1197, 1167, 1126, 1111, 1078, 1049, 1021, 974. 1H-NMR d: 2.47—
2.98 (5H, m, H-5b, CH2ꢅ2), 3.77, 3.86, 3.91, 3.93 and 3.97 (total 15H, 5 s,
O–CH3ꢅ5), 4.38—4.50 (total 1H, m, H-5a), 4.72—4.82 (total 1H, m, H-
6a), 6.39 (1H, br s, OH), 6.78 (1H, s, H-8), 8.05 (1H, s, H-11). 13C-NMR d:
23.57 (t), 34.25 (t), 38.78 (t), 52.08 (d), 52.68 (q), 55.84 (q), 55.98 (q), 60.45
(q), 60.98 (q), 111.25 (d), 111.92 (d), 116.43 (s), 119.57 (s), 124.35 (s),
128.75 (s), 128.95 (s), 138.53 (s), 144.80 (s), 147.26 (s), 147.70 (s), 147.90
(s), 155.09 (s). EI-MS m/z (%): 415 [M]ꢁ (34), 383 (36), 327 (100), 88 (4).
Anal. Calcd for C22H25NO7: C, 63.60; H, 6.07; N, 3.37, Found: C, 63.42; H,
6.21; N, 3.50.
8a, 8c was obtained in 62.1% yield from ethanol as a white solid, mp 108—
110 °C. UV lmax (MeOH) nm (log e): 215 (4.66), 227 sh (4.52), 283 (4.12).
IR (CH2Cl2-film) cmꢀ1: 2936, 1691, 1592, 1571, 1492, 1459, 1437, 1416,
1376, 1350, 1324, 1283, 1265, 1243, 1196, 1169, 1141, 1117, 1093, 1019,
1
905. H-NMR d: 2.70—3.08 (4H, m, CH2ꢅ2), 3.57—3.69 (1H, m, H-3b),
3.88 (6H, s, O–CH3ꢅ2), 3.99—4.09 (1H, m, H-3a), 4.97 (2H, s, Ph–CH2),
4.98 (2H, s, Ph–CH2), 5.69 (1H, dd, Jꢄ5.4, 8.9 Hz, H-1), 6.41 (1H, s,
Ar–H), 6.70—6.78 (2H, m, Ar–H), 7.28—7.45 (11H, m, Ph–Hꢅ10 and
Ar–H). 13C-NMR d: 23.42 (t), 39.68 (t), 41.66 (t), 53.74 (d), 60.68 (q),
60.99 (q), 70.23 (t), 71.03 (t), 108.19 (d), 115.22 (d), 116.12 (s), 117.88 (d),
118.24 (s), 119.68 (s), 127.24 (d), 127.47 (d), 128.03 (d), 128.13 (d), 128.65
(d), 130.01 (s), 133.33 (d), 136.47 (s), 136.81 (s), 137.38 (s), 141.63 (s),
150.90 (s), 151.32 (s), 155.51 (s), 157.97 (s). Anal. Calcd for
C34H31BrF3NO5: C, 60.90; H, 4.66; N, 2.09, Found: C, 60.80; H, 4.8; N,
2.17.
2-Trifluoroacetyl-1-(2-bromo-4,5-methylenedioxybenzyl)-7-benzyloxy-
5,6-dimethoxy-1,2,3,4-tetrahydroisoquinoline (8d) In a similar manner
to 8a, 8d was obtained in 65.7% yield from ethanol as a pale yellow solid,
mp 152—154 °C. UV lmax (MeOH) nm (log e): 212 (4.70), 226 sh (4.43),
285 (4.08), 294 (4.09). IR (CH2Cl2-film) cmꢀ1: 2938, 1690, 1604, 1587,
1502, 1478, 1460, 1438, 1416, 1374, 1350, 1325, 1267, 1232, 1199, 1169,
1142, 1115, 1093, 1037, 933, 905. 1H-NMR d: 2.70—3.26 (4H, m, CH2ꢅ2),
3.57—3.70 (1H, m, H-3b), 3.88 (3H, s, O–CH3), 3.89 (3H, s, O–CH3),
4.00—4.11 (1H, m, H-3a), 5.04 (2H, s, Ph–CH2), 5.64 (1H, dd, Jꢄ5.4,
9.0 Hz, H-1), 5.93—5.97 (2H, m, O–CH2–O), 6.45 (1H, s, Ar–H), 6.56 (1H,
s, Ar–H), 6.98 (1H, s, Ar–H), 7.28—7.45 (5H, m, Ph–H). 13C-NMR d: (both
conformers) 21.86 (t), 23.48 (t), 39.67 (t), 39.72 (t), 41.40 (t), 42.29 (t),
53.93 (d), 60.69 (q), 61.00 (q), 71.15 (t), 101.77 (t), 108.29 (d), 110.53 (d),
112.75 (d), 115.56 (s), 118.34 (s), 119.74 (s), 127.20 (d), 128.06 (d), 128.67
(d), 129.33 (s), 130.05 (s), 136.83 (s), 141.65 (s), 147.33 (s), 147.47 (s),
150.92 (s), 151.35 (s), 155.79 (s). Anal. Calcd for C28H25Br F3NO6: C,
55.27; H, 4.14; N, 2.30, Found: C, 55.02; H, 4.31; N, 2.47.
1-Hydroxy-2,3-dimethoxy-6-trifluoroacetylnoraporphine (9a) A so-
lution of azobis (isobutyronitrile) (0.6 g, 3.6 mmol) and tributyltin hydride
(4.1 g, 14.0 mmol) in toluene (40 ml) was added dropwise in four equal por-
tions over 3 h to a refluxing solution of 8a (2.0 g, 3.6 mmol) in toluene
(40 ml) and the resulting mixture was then refluxed for another 24 h. The
solvent was then removed under vacuum and the residue was dissolved in
acetonitrile (50 ml) and washed with hexane (3ꢅ50 ml), then dried over anh.
Na2SO4. Removal of the solvent gave the crude noraporphine (1.1 g) as a
brown viscous oil which was chromatograped over silica gel using
hexane–ethyl acetate as eluent. The earlier fractions gave the hydrogenolysis
product (8e) as a white solid (317.7 mg, 18.5%), mp 115—116 °C. 1H-NMR
d: 2.71—2.78 (2H, m, H-4 of both conformers), 3.08 (2H, apparent d,
Jꢄ6.8 Hz, Ar–CH2), 3.38—3.50 (1H, m, H-3b of both conformers), 3.86,
3.87, 3.88 and 3.89 (total 6H, 4 s, O–CH3ꢅ2 of both conformers), 3.90—
3.98 (1H, m, H-3a), 4.87 (2H, s, Ph–CH2), 5.55 (1H, apparent t, Jꢄ6.8 Hz,
H-1), 6.17 (1H, s, Ar–H), 7.04—7.12 (2H, m, Ar–H of both conformers),
7.22—7.41 (8H, m, Ph–Hꢅ5 and Ar–Hꢅ3 of both conformers). 13C-NMR
d: (both conformers) 21.83 (t), 23.22 (t), 39.98 (t), 40.02 (t), 41.78 (t), 55.30
(d), 60.71 (q), 60.97 (q), 70.78 (t), 108.33 (d), 114.58 (s), 118.40 (s), 119.69
(s), 122.21 (s), 126.95 (d), 127.25 (d), 128.02 (d), 128.46 (d), 128.61 (d),
129.74 (d), 136.78 (s), 136.87 (s), 141.46 (s), 150.88 (s), 150.98 (s), 155.74
(s). The latter fractions gave pure noraporphine (9a) as a pale brown solid
(151.6 mg, 10.9%), mp 205—206 °C. UV lmax (MeOH) nm (log e): 215
(4.51), 274 (4.22), 292 sh (4.07), 309 (4.02). IR (CH2Cl2-film) cmꢀ1: 3393,
2921, 2850, 1682, 1589, 1456, 1440, 1420, 1377, 1300, 1277, 1186, 1173,
1147, 1127, 1079, 1046, 1019, 952, 926. 1H-NMR d: 2.57—3.34 (5H, m, H-
5b, CH2ꢅ2), 3.89 (3H, s, O–CH3), 3.97 (3H, s, O–CH3), 4.19—4.28 (1H, m,
H-5a), 5.08 (1H, dd, Jꢄ4.2, 13.7 Hz, H-6a), 6.41 (1H, s, OH), 7.19—7.38
(3H, m, Ar–H), 8.39 (1H, d, Jꢄ7.9 Hz, H-11). 13C-NMR d: 23.93 (t), 33.19
(t), 41.09 (t), 52.33 (d), 60.55 (q), 61.03 (q), 114.48 (s), 116.43 (s), 118.26
(s), 122.11 (s), 127.06 (d), 127.29 (d), 128.12 (d), 128.49 (d), 131.36 (s),
134.86 (s), 138.80 (s), 146.13 (s), 148.32 (s), 155.81 (s). EI-MS m/z (%):
393 [M]ꢁ (43), 361 (6), 267 (100), 126 (2). Anal. Calcd for C20H18F3NO4: C,
61.07; H, 4.61; N, 3.56, Found: C, 61.24; H, 4.48; N, 3.40.
9-Benzyloxy-1-hydroxy-2,3-dimethoxy-6-trifluoroacetylnoraporphine
(9c) In a similar manner, 8g was obtained in 38.6% yield as a yellow oil.
1H-NMR d: 2.70—2.77 (2H, m, H-4 of both conformers), 3.04 (2H, appar-
ent d, Jꢄ6.8 Hz, Ar–CH2), 3.34—3.47 (1H, m, H-3b), 3.84, 3.85 and 3.86
(total 6H, 3 s, O–CH3ꢅ2 of both conformers), 3.88—3.98 (1H, m, H-3a),
4.65 (0.2H, ABq, Jꢄ11.7 Hz, Ph–CH2 of one conformer), 4.85 (1.8H, ABq,
Jꢄ12.1 Hz, Ph–CH2 of another conformer), 4.96 and 4.98 (total 2H, 2 s,
Ph–CH2 of both conformers), 5.56 (1H, apparent t, Jꢄ6.7 Hz, H-1), 6.17
(1H, s, Ar–H), 6.64—6.75 (2H, m, Ar–H of both conformers), 6.81—6.91
(1H, m, Ar–H of both conformers), 7.12—7.21 (1H, m, Ar–H of both con-
formers), 7.23—7.41 (total 10H, m, Ph–H of both conformers). 13C-NMR d:
(both conformers) 23.18 (t), 39.99 (t), 40.03 (t), 41.80 (t), 55.23 (d), 60.65
(q), 60.92 (q), 69.87 (t), 70.76 (t), 108.31 (d), 113.50 (d), 114.62 (s), 116.10
(d), 118.44 (s), 119.66 (d), 122.40 (d), 127.25 (d), 127.47 (d), 127.92 (d),
127.98 (d), 128.54 (d), 128.58 (d), 129.45 (d), 129.73 (d), 129.97 (s), 136.77
(s), 136.91 (s), 138.48 (s), 141.45 (s), 150.85 (s), 150.99 (s), 155.71 (s),
158.86 (s). Noraporphine (9c) was obtained in 9.7% yield as a pale yellow
solid, mp 183—184 °C. UV lmax (MeOH) nm (log e): 213 (4.67), 231 sh
(4.43), 274 (4.28), 283 (4.31), 299 (4.19), 312 (4.11). IR (CH2Cl2-film)
cmꢀ1: 3583, 3411, 2942, 1686, 1609, 1501, 1459, 1414, 1378, 1342, 1310,
1
1281, 1233, 1201, 1175, 1153, 1083, 1049, 1016, 974. H-NMR d: 2.57—
3.10 (5H, m, CH, CH2ꢅ2), 3.21—3.33 (1H, m, H-5b), 3.87 (3H, s, O–CH3),
3.97 (3H, s, O–CH3), 4.18—4.28 (1H, m, H-5a), 5.10 (2H, s, Ph–CH2), 6.35
(1H, s, OH), 6.90 (1H, d, Jꢄ2.4 Hz, H-8), 6.95 (1H, dd, Jꢄ2.4, 8.7 Hz, H-
10), 7.30—7.50 (5H, m, Ph–H), 8.32 (1H, d, Jꢄ8.7 Hz, H-11). 13C-NMR d:
23.92 (t), 33.55 (t), 41.12 (t), 52.31 (d), 60.54 (q), 60.99 (q), 69.95 (t),
113.28 (d), 114.76 (d), 116.45 (s), 118.20 (s), 118.29 (s), 124.43 (s), 127.28
(s), 127.51 (d), 127.97 (d), 128.61 (d), 129.48 (d), 136.68 (s), 136.99 (s),
138.80 (s), 145.49 (s), 147.69 (s), 155.80 (s), 157.75 (s). EI-MS m/z (%):
499 [M]ꢁ (10), 373 (16), 91 (23), 32 (100). Anal. Calcd for C27H24F3NO5: C,
64.93; H, 4.84; N, 2.80, Found: C, 64.76; H, 4.98; N, 2.97.
1-Hydroxy-2,3-dimethoxy-9,10-methylenedioxy-6-trifluoroacetylnora-
porphine (9d) In a similar manner, 8h was obtained in 13.1% yield as a
pale yellow solid, mp 129—130 °C. 1H-NMR d: 2.71—2.78 (2H, m, H-4 of
both conformers), 2.99 (2H, apparent d, Jꢄ6.7 Hz, Ar–CH2), 3.36—3.48
(1H, m, H-3b), 3.87 (3H, s, O–CH3), 3.89 (3H, s, O–CH3), 3.91—4.00 (1H,
m, H-3a), 4.95 (2H, s, Ph–CH2), 5.49 (1H, apparent t, Jꢄ6.7 Hz, H-1),
5.89—5.95 (2H, m, O–CH2–O of both conformers), 6.24 (1H, s, Ar–H),
6.51 (1H, dd, Jꢄ1.4, 7.8 Hz, Ar–H), 6.54 and 6.56 (total 1H, 2d, Jꢄ1.4 Hz,
Ar–H of both conformers), 6.70 and 6.75 (total 1H, 2d, Jꢄ7.8 Hz, Ar–H of
both conformers), 7.28—7.42 (5H, m, Ph–H of both conformers). 13C-NMR
d: (both conformers) 23.23 (t), 39.98 (t), 40.02 (t), 41.41 (t), 55.33 (d),
58.28 (d), 60.69 (q), 60.96 (q), 70.95 (t), 100.95 (t), 108.17 (d), 108.38 (d),
109.97 (d), 114.57 (s), 118.39 (s), 119.77 (s), 122.69 (d), 127.19 (d), 128.02
(d), 128.62 (d), 129.97 (s), 130.52 (s), 136.76 (s), 141.50 (s), 146.52 (s),
147.63 (s), 150.87 (s), 151.04 (s), 155.75 (s). Noraporphine (9d) was ob-
tained in 10.4% as a pale yellow solid, mp 251—252 °C. UV lmax (MeOH)
nm (log e): 220 (4.60), 235 sh (4.40), 274 sh (4.03), 283 (4.09), 309 (4.27),
320 (4.28). IR (CH2Cl2-film) cmꢀ1: 3402, 2919, 2850, 1686, 1506, 1488,
1459, 1431, 1414, 1373, 1354, 1305, 1281, 1239, 1225, 1186, 1146, 1094,
1-Hydroxy-6-carbomethoxy-2,3,9,10-tetramethoxynoraporphine (9b)
In a similar manner, 8f was obtained in 41.6% yield as a pale yellow solid,
mp 121—122 °C. 1H-NMR d: 2.53—2.84 (2H, m, H-4 of both conformers),
2.84—3.10 (2H, m, Ar–CH2 of both conformers), 3.10—3.30 (1H, m, H-
3b), 3.52, 3.68, 3.71, 3.81, 3.84, 3.85, 3.87 and 3.88 (total 15H, 8 s,
O–CH3ꢅ4 of both conformers), 4.05—4.22 (1H, m, H-3a), 4.85 (1.06H,
ABq, Jꢄ12.1 Hz, Ph–CH2 of one conformer), 4.96 (0.94H, s, Ph–CH2 of an-
other conformer), 5.05—5.15 and 5.15—5.25 (total 1H, apparent 2m, H-1 of

April 2009
375
1
118.28 (s), 125.23 (s), 130.69 (d), 132.05 (s), 138.02 (s), 140.77 (s), 147.25
(s), 149.74 (s), 157.52 (s). EI-MS m/z (%): 313 [M]ꢁ (88), 312 (100), 282
(23). Anal. Calcd for C18H19NO4: C, 68.99; H, 6.11; N, 4.47, Found: C,
68.78; H, 6.30; N, 4.58.
1045, 971, 940, 924. H-NMR d: 2.54—3.09 (4H, m, CH2ꢅ2), 3.19—3.20
(1H, m, H-5b), 3.87 (3H, s, O–CH3), 3.97 (3H, s, O–CH3), 4.17—4.27 (1H,
m, H-5a), 5.02 (1H, dd, Jꢄ4.2, 13.8 Hz, H-6a), 5.97 (2H, s, O–CH2–O),
6.40 (1H, s, OH), 6.74 (1H, s, H-8), 7.95 (1H, s, H-11). 13C-NMR d: 23.88
(t), 33.18 (t), 41.09 (t), 52.48 (d), 60.54 (q), 61.01 (q), 101.01 (t), 108.80 (d),
108.82 (d), 114.47 (s), 116.54 (s), 118.17 (s), 124.93 (s), 127.38 (s), 129.17
(s), 138.77 (s), 145.42 (s), 146.26 (s), 146.56 (s), 147.80 (s), 155.83 (s). EI-
MS m/z (%): 437 [M]ꢁ (38), 405 (3), 311 (100), 126 (9). Anal. Calcd for
C21H18F3NO6: C, 57.67; H, 4.15; N, 3.20, Found: C, 57.48; H, 4.30; N, 3.37.
(ꢀ)-Isopiline (1a) A mixture of 9a (60.0 mg, 15.3 mmol), potassium
carbonate (1.0 g), methanol (20 ml) and water (5 ml) was refluxed for 3 h.
The solvent was then removed under vacuum and water (20 ml) and 10%
sodium bicarbonate (20 ml) were added to the residue followed by extraction
with chloroform (2ꢅ20 ml). Removal of the solvent gave a brown solid
which was triturated with ethanol to give (ꢂ)-isopiline (1a) as a pale grey
solid (29.3 mg, 64.7%), mp 170—171 °C. UV lmax (MeOH) nm (log e): 214
(4.45), 220 sh (4.42), 232 sh (4.19), 274 (4.20), 291 sh (4.00), 309 (3.89). IR
(CH2Cl2-film) cmꢀ1: 3583, 3305, 2931, 2849, 1587, 1490, 1463, 1445, 1418,
1380, 1338, 1305, 1267, 1226, 1196, 1152, 1110, 1086, 1052, 1025, 991,
960. 1H-NMR d: 2.72—3.25 (5H, m, H-5b, CH2ꢅ2), 3.39—3.49 (1H, m,
H-5a), 3.83—3.94 (1H, m, H-6a), 3.88 (3H, s, O–CH3), 3.95 (3H, s,
O–CH3), 7.14—7.34 (3H, m, Ar–H), 8.32 (1H, d, Jꢄ7.2 Hz, H-11). 13C-
NMR d: 23.12 (t), 36.82 (t), 42.82 (t), 53.80 (d), 59.95 (q), 60.83 (q), 115.39
(s), 118.32 (s), 126.69 (d), 126.82 (d), 127.73 (d), 127.84 (d), 131.59 (s),
132.18 (s), 134.90 (s), 138.75 (s), 145.31 (s), 149.09 (s). EI-MS m/z (%):
297 [M]ꢁ (73), 296 (100), 266 (18). Anal. Calcd for C18H19NO3: C, 72.71;
H, 6.44; N, 4.71, Found: C, 72.60; H, 6.54; N, 4.86.
(ꢀ)-3-Methoxynordomesticine (1d) In a similar manner to 1a, (ꢂ)-3-
methoxynordomesticine (1d) was obtained in 78.6% yield as a pale brown
solid, mp 206—208 °C. UV lmax (MeOH) nm (log e): 213 (4.73), 234 sh
(4.40), 271 sh (4.41), 281 (4.45), 295 (4.29), 310sh (4.15). IR (CH2Cl2-film)
cmꢀ1: 3437, 2919, 2850, 1620, 1542, 1502, 1463, 1430, 1413, 1381, 1357,
1
1303, 1292, 1255, 1228, 1196, 1148, 1126, 1096, 1039, 982, 932. H-NMR
d: 2.58—3.00 (6H, m, CH2ꢅ3), 3.35—3.44 (1H, m, H-6a), 3.87 (3H, s,
O–CH3), 3.94 (3H, s, O–CH3), 5.95 (2H, ABq, Jꢄ1.4 Hz, O–CH2–O), 6.72
(1H, s, H-8), 7.90 (1H, s, H-11). 13C-NMR d: 23.51 (t), 37.29 (t), 42.99 (t),
54.02 (d), 59.93 (q), 60.81 (q), 100.78 (t), 108.14 (d), 108.66 (d), 115.53 (s),
118.63 (s), 125.74 (s), 129.49 (s), 131.87 (s), 138.58 (s), 144.48 (s), 145.69
(s), 146.21 (s), 148.61 (s). EI-MS m/z (%): 341 [M]ꢁ (87), 340 (100), 310
(22). Anal. Calcd for C19H19NO5: C, 66.85; H, 5.61; N, 4.10, Found: C,
66.68; H, 5.84; N, 4.03.
Minimum Inhibitory Concentration (MIC) MIC of (ꢂ)-3-
methoxynordomesticine was determined by NCCLS microbroth dilution
methods.13) (ꢂ)-3-Methoxynordomesticine was weighed and dissolved in
DMSO to make a solution of concentration 2.56 mg/ml. From this stock so-
lution two-fold serial dilution has been carried out to give a series of solu-
tions from 256 to 0.50 mg/ml with culture medium in 96-well microplates
(100 ml of total volume). Three different microorganisms were selected viz.
Staphytolcoccus aureus ATCC25932, Escherichia coli ATCC10536 and
Candida albicans ATCC90028. They were subcultured on nutrient broth
supplemented with 10% glucose (NBG) (for bacteria) or Sabouraud glucose
broth (for yeast) and incubated at 37 °C for 24 h. A final concentration of
1ꢅ105 cfu/ml of test bacteria or yeast was added to each dilution. The plates
were incubated at 37 °C for 48 h. MIC was defined as the lowest concentra-
tion of the test agent that inhibited bacterial or yeast growth, as indicated by
the absence of turbidity. Test agent-free broth containing 5% DMSO was in-
cubated as the growth control. Minimum microbicidal concentration (MMC)
was determined by inoculating onto nutrient agar plates (for bacteria) or
Sabouraud agar plates (for yeast) a 100 ml of medium from each of the wells
from the MIC test which showed no turbidity. MMCs were defined as the
lowest concentration of the test agent where there was no microbial growth
on the plates.
(ꢀ)-Preocoteine (1b) A mixture of 9b (100.0 mg, 0.24 mmol) and
lithium aluminium hydride (2.0 g) in dry tetrahydrofuran (40 ml) was re-
fluxed for 3 h. Water (10 ml) was added dropwise followed by dilute ammo-
nium hydroxide (25 ml). The pale yellow granular residue was filtered and
washed with chloroform. The organic phase was separated and then dried
over anh. Na2SO4. Removal of the solvent under vacuum gave a pale yellow-
white solid which was recrystallized from benzene to give (ꢂ)-preocoteine
(1b) as a pale yellow solid (66.6 mg, 74.5%), mp 179—180 °C. UV lmax
(MeOH) nm (log e): 219 (4.38), 271sh (3.71), 281 (3.82), 308 (4.00), 319
(3.99). IR (CH2Cl2-film) cmꢀ1: 3400, 2937, 2846, 1609, 1593, 1515, 1463,
1430, 1397, 1375, 1347, 1310, 1284, 1253, 1226, 1215, 1195, 1115, 1082,
1
1058, 1036, 1014, 990, 971. H-NMR d: 2.70 (3H, s, N–CH3), 2.68—3.36
Anti-inflammatory Activity Murine macrophage RAW 264.7 cell line
obtained from American Type Culture Collection (ATCC, Maryland,
U.S.A.), was maintained in DMEM supplemented with 10% heat inactivated
FBS, penicillin G (100 IU/ml), streptomycin (100 mg/ml), and L-glutamine
(2 mM) and incubated at 37 °C in a humidified atmosphere containing 5%
CO2. Cells (1ꢅ106/ml) were pre-incubated 2 h with (ꢂ)-3-methoxynordo-
mesticine (1, 2.5, 5 mg/ml) and further cultured 24 h with LPS (1 mg/ml) in
24-well plates. Supernatants were removed at the allotted times and NO,
PGE2, TNF-a, IL-1b and IL-6 levels were quantified by immunoassay kits
(7H, m, CH, CH2ꢅ3), 3.87, 3.91 and 3.96 (12H, 3 s, O–CH3ꢅ4), 6.77 (1H,
s, H-8), 7.96 (1H, s, H-11). 13C-NMR d: 21.72 (t), 30.34 (q), 33.06 (t), 52.60
(t), 55.91 (q), 56.03 (q), 60.08 (q), 60.87 (q), 62.24 (d), 111.03 (d), 111.73
(d), 115.92 (s), 117.28 (s), 124.31 (s), 126.93 (s), 135.80 (s), 139.06 (s),
145.01 (s), 147.52 (s), 147.69 (s), 148.07 (s). EI-MS m/z (%): 371 [M]ꢁ
(100), 339 (35), 326 (17). Anal. Calcd for C21H25NO5: C, 67.91; H, 6.78; N,
3.77, Found: C, 67.80; H, 6.85; N, 3.90.
1,9-Dihydroxy-2,3-dimethoxy-6-trifluoroacetylnoraporphine (9e)
A
solution of 9c (115.0 mg, 0.23 mmol) in ethanol (30 ml) was hydrogenolysed
over 10% Pd/C (10.0 mg) at atmospheric pressure for 48 h. The catalyst was
filtered and the solvent removed under vacuum. The resulting white residue
was recrystallized from ethanol to give 9e as a pale brown solid (84.3 mg,
89.4%), mp 274—275 °C. UV lmax (MeOH) nm (log e): 214 (4.64), 232 sh
(4.39), 274 sh (4.25), 282 (4.28), 299 (4.16), 312 (4.08). IR (CH2Cl2-film)
cmꢀ1: 3502, 3347, 2920, 2850, 1671, 1619, 1587, 1506, 1459, 1378, 1343,
1302, 1280, 1264, 1234, 1201, 1190, 1176, 1144, 1081, 1049, 1020, 973.
1H-NMR (acetone-d6) d: 2.57—3.13 (4H, m, CH2ꢅ2), 3.26—3.38 (1H, m,
H-5b), 3.88 (3H, s, O–CH3), 3.94 (3H, s, O–CH3), 4.17—4.28 (1H, m, H-
5a), 5.02 (1H, dd, Jꢄ5.1, 12.9 Hz, H-6a), 6.76—6.85 (2H, m, Ar–H), 8.31
(1H, d, Jꢄ8.4 Hz, H-11). 13C-NMR (acetone-d6) d: 24.10 (t), 33.70 (t),
41.32 (t), 52.69 (d), 60.53 (q), 60.92 (q), 113.95 (d), 115.24 (d), 117.02 (s),
118.00 (s), 118.60 (s), 123.72 (s), 127.20 (s), 130.06 (d), 136.95 (s), 139.46
(s), 146.20 (s), 148.15 (s), 155.73 (s), 156.16 (s). EI-MS m/z (%): 409 [M]ꢁ
(29), 377 (7), 283 (100), 126 (3). Anal. Calcd for C20H18F3NO5: C, 58.68; H,
4.43; N, 3.42, Found: C, 58.45; H, 4.60; N, 3.55.
(ꢀ)-Oureguattidine (1c) In a similar manner to 1a, (ꢂ)-oureguattidine
(1c) was obtained in 86.5% yield as a brown solid, mp 228—229 °C. UV
lmax (MeOH) nm (log e): 209 sh (4.40), 221 (4.53), 269 sh (3.96), 303
(4.14), 314 (4.13). IR (CH2Cl2-film) cmꢀ1: 3412, 2922, 2850, 1610, 1463,
1420, 1380, 1341, 1298, 1227, 1195, 1154, 1112, 1085, 1048, 1026, 992.
1H-NMR (pyridine-d5) d: 2.81—3.06 (5H, m, H-5b, CH2ꢅ2), 3.39—3.49
(1H, m, H-5a), 3.86 (3H, s, O–CH3), 3.88 (3H, s, O–CH3), 4.00—4.15 (1H,
m, H-6a), 7.21 (1H, d, Jꢄ2.6 Hz, H-8), 7.27 (1H, dd, Jꢄ2.6, 8.6 Hz, H-10),
9.03 (1H, d, Jꢄ8.6 Hz, H-11). 13C-NMR (pyridine-d5) d: 23.97 (t), 38.03 (t),
43.42 (t), 54.85 (d), 60.01 (q), 60.56 (q), 114.37 (d), 115.67 (d), 117.70 (s),
according to the manufacture’s protocols (Assay Designs’ Correlate-EIATM
Stressgen, U.S.A.), respectively.
,
Western blot Cellular proteins were extracted from the control and (ꢂ)-3-
methoxynordomesticine-treated RAW 264.7 cells. The washed cell pellets
were resuspended in lysis buffer buffer (50 mM HEPES pH 7.0, 250 mM
NaCl, 5 mM EDTA, 0.1% Nonidet P-40, 1 mM phenylmethylsulfonyl fluo-
ride, 0.5 mM dithiothreitol, 5 mM NaF, 0.5 mM Na orthovanadate) containing
5 mg/ml each of leupeptin and aprotinin and incubated for 15 min at 4 °C.
Cell debris was removed by microcentrifugation, followed by quick freezing
of the supernatants. Protein concentration was determined by BioRad pro-
tein assay reagent according to the manufactures instruction, 40—50 mg of
cellular proteins from treated and untreated cell extracts were electroblotted
onto nitrocellulose membrane following separation on a 10% SDS-polyacryl-
amide gel electrophoresis. The immunoblot was incubated overnight with
the blocking solution (5% skim milk) at 4 °C, followed by incubation for 4 h
with a 1 : 500 dilution of monoclonal anti-iNOS and COX-2 antibody (San-
tacruz, CA, U.S.A.). Blots were washed 2 times with PBS and incubated
with a 1 : 1000 dilution of horseradish peroxidase-conjugated goat anti-
mouse IgG secondary antibody (Santacruz, CA, U.S.A.) for 1 h at room tem-
perature. Blots were again washed three times in Tween 20/Tris-buffered
saline (TTBS) and then developed by enhanced chemiluminescence (Amer-
sham Life Science, Arlington Heights, IL, U.S.A.). Cytotoxicity assay. 3-
(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cyto-
toxicity assay was performed according to the method previously de-
scribed.14) MTT solution was added at a concentration of 50 mg/ml into each
well, which also contain 1, 2.5 and 5 mg/ml of (ꢂ)-3-methoxynordomes-

376
Vol. 57, No. 4
ticine. After 4 h of incubation at 37 °C, the medium was discarded and the
formazan blue, which formed in the cells, was dissolved in 50 ml DMSO.
The optical density at 540 nm was determined with a microplate reader. The
optical density of formazan formed in control (untreated) cells was taken as
100% of viability.
6) Leboeuf M., Cortes D., Hocquemiller R., Cave A., Planta Med., 48,
234—245 (1983).
7) Castro O., Hasbun C., Calderon M., Fitoterapia, 62, 72—73 (1991).
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50, 2107—2114 (1994).
9) Mannel D. N., Echtenacher B., Chem. Immunol., 74, 141—161 (2000).
10) Burger A. P. N., Brandt E. V., Roux D. G., Phytochemistry, 22, 2813—
2817 (1983).
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