Design, synthesis, and biological evaluation of combretabenzodiazepines: A novel class of anti-tubulin agents

Article abstract of DOI:10.1021/jm5016389

In the present manuscript, starting from the 1,4-benzodiazepin-2-one nucleus, a privileged structure in medicinal chemistry, we have synthesized a novel class of cis-locked combretastatins named combreatabenzodiazepines. They show similar cytotoxic and antitubulin activity compared to combretastatin A-4 in neuroblastoma cells, showing a better pharmacokinetic profile. This class of compounds has therefore the potential for further development as antitubulin agents.

Full text of DOI:10.1021/jm5016389

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Article
Design, Synthesis and Biological Evaluation of
Combretabenzodiazepines: a Novel Class of Antitubulin Agents
Ubaldina Galli, Cristina Travelli, Silvio Aprile, Elena Arrigoni, Simone Torretta, Giorgio Grosa, Alberto
Massarotti, Giovanni Sorba, Pier Luigi Canonico, Armando A. Genazzani, and Gian Cesare Tron
J. Med. Chem., Just Accepted Manuscript • DOI: 10.1021/jm5016389 • Publication Date (Web): 13 Jan 2015
Downloaded from http://pubs.acs.org on January 21, 2015
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Design, Synthesis and Biological Evaluation of
Combretabenzodiazepines: a Novel Class of
Antitubulin Agents
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#
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Ubaldina Galli, Cristina Travelli, Silvio Aprile, Elena Arrigoni, Simone Torretta, Giorgio
Grosa, Alberto Massarotti, Giovanni Sorba, Pier Luigi Canonico, Armando A. Genazzani,* and
Gian Cesare Tron*
Dipartimento di Scienze del Farmaco, Università degli Studi del Piemonte Orientale “A.
Avogadro”, Largo Donegani 2, 28100 Novara, Italy.
ABSTRACT: In the present manuscript, starting from the 1,4ꢀbenzodiazepinꢀ2ꢀone nucleus, a
privileged structure in medicinal chemistry, we have synthesized a novel class of cisꢀlocked
combretastatins named combreatabenzodiazepines. They show similar cytotoxic and antitubulin
activity compared to combretastatin Aꢀ4 in neuroblastoma cells, showing a better
pharmacokinetic profile. This class of compounds has therefore the potential for further
development as antitubulin agents.
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INTRODUCTION
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The naturallyꢀoccurring combretastatin Aꢀ4 (CAꢀ4, 1) and its analogue phenstatin (2; Figure
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), serendipitously discovered in 1998, block the polymerization of tubulin, strongly interacting
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with the colchicine binding site. These have displayed a high degree of cytotoxicity on several
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cancer cell lines, including those that exhibit multidrug resistance, and have antiꢀtumoural
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properties in vivo models. Furthermore, both agents induce a rapid vascular shutdown in
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tumors, occluding the blood flow, with the net result of starving the proliferating cancer cells.
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CAꢀ4, in its water soluble phosphate proꢀdrug, both as a single agent and in combination
therapy, has entered clinical trials for the treatment of anaplastic thyroid cancer and ageꢀrelated
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macular degeneration.
Figure 1. CAꢀ4 (1) and phenstatin (2).
Over the last decades, these two relative simple molecules have been the starting point of many
medicinal chemistry programs directed at the rational design of novel antitubulin agents. For
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CAꢀ4, the replacement of the unstable olefinic bond with several fiveꢀmembered heterocyclic
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rings has been the main strategy pursued. In the case of phenstatin, the replacement of the
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ketone with an ethylene bridge (3), a methyl (4), a Nꢀmethyl group (5) or the formation of
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rigid analogues between the ethylene group and the Bꢀring (6, 7, 8, 9, 10 ) or the Aꢀring
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(
11) has unveiled novel agents endowed with cytotoxic activity (Figure 2).
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Figure 2. Analogues of phenstatin endowed with strong cytotoxic activity.
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The important features that maintain the tubulin interaction for CAꢀ4, phenstatin and all the
related compounds shown in Figure 2 are: i) the presence of the trimethoxyphenyl Aꢀring and ii)
the lack of planarity for the two phenyl rings which must be inclined towards each other with a
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dihedral angle between 50° and 80°.
In search of novel potential rigid analogues of phenstatin, we hypothesised that the
replacement of the benzosuberene ring of 9 with a 1,4ꢀbenzodiazepineꢀ2ꢀone nucleus (12)
(Figure 3) could be a successful strategy to unravel a novel class of antitubulin agents. Indeed,
the 1,4ꢀbenzodiazepineꢀ2ꢀone nucleus is one of the most widely used molecular scaffold in
medicinal chemistry due to its favourable pharmacokinetic features and its ability to act as a
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privileged structure when properly decorated. In this manuscript, we report the design, synthesis,
and biological evaluation of a novel class of antitubulin agents named combretabenzodiazepines.
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Figure 3. The combretabenzodiazepine scaffold.
RESULTS AND DISCUSSION
The synthesis of the combretabenzodiazepines starts with a Grignard reaction between the 5ꢀ
bromoꢀ1,2,3ꢀtrimethoxybenzene (13) and the appropriate aldehydes (14ꢀ18) to afford the
corresponding 2ꢀnitrobenzydrols (19ꢀ23). The latter were oxidized with pyridinium dichromate
(PDC) to benzophenones (24ꢀ28). The nitro group was then reduced using iron/acetic
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acid/hydrochloric acid under reflux conditions to give the 2ꢀaminobenzophenones (29ꢀ33). The
amino group was acetylated in the presence of chloroacetyl chloride to form the
chloroacetamidobenzophenones (34ꢀ38). Finally, the 1,4ꢀbenzodiazepineꢀ2ꢀone nucleus was
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obtained using an improved BlaževićꢀKajfež procedure via ammonium acetate and
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hexamethylenetetramine (HMTA) to afford final compounds 39ꢀ43. These
combretabenzodiazepines were then N1ꢀmethylated to give compounds 44ꢀ48 using sodium
hydride and methyl iodide in DMF (Scheme 1).
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Scheme 1. Scheme for the synthesis of combretabenzodiazepines 39-48.
In order to investigate the cytotoxicity and tubulinꢀpolymerization aspects of these compounds,
we first performed an MTT assay on the SHꢀSY5Y neuroblastoma cell line, as we have evidence
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that this model is highly responsive to CAꢀ4 and phenstatin. The initial screening at 1 µM
showed that most compounds (except 41, 46) were active at this concentration.
To investigate the activity further, we chose the compounds that induced at least 50% cell
death and performed complete concentrationꢀresponse curves (Figure 4).
A
B
%
of
Inhibition of
tubulin
polymerization
IC50 (µM)
1.3 ± 0.1
9.0 ± 1.3
n.d.
viability
SH-SY5Y
(1 µM)
IC50 (nM)
MTT assay
Comp.
1
33.4 ± 3.0
24.7 ± 3.0
33.9 ± 4.3
102 ± 6.7
46.8 ± 3.0
35.1 ± 6.5
32.9 ± 7.1
44.4 ± 2.9
105.2 ± 5.4
76.4 ± 6.5
46.4 ± 5.8
27.5 ± 2.9
95.2 ± 2.9
64.8 ± 4.6
2.7 ± 2.1
25.1 ± 14.7
255.6 ± 60.4
n.d.
90.3 ± 14.3
71.0 ± 14.1
279.5 ± 115.9
862.0 ± 338.7
n.d.
n.d
n.d.
3.9 ± 2.6
n.d.
414.8 ± 345.0
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n.d.
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n.d.
n.d.
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Figure 4. Cytotoxicity in SH-SY5Y cells of the synthesized compounds. (A) Values in the
second column represent the % of viable cells (determined by MTT assay) after 48 hours of
treatment with 1 µM of the indicated compounds. Values in the third column represent the
calculated IC values (using Kaleidagraph software). Values in last column represent the IC
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value calculated for inhibition of tubulin polymerization in vitro. n.d.: not determined since full
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cytotoxicity was not reached at concentration at 1 µM and up to 1 µM. Values are the mean ± SD
of 8ꢀ12 determinations from 3 different experiments. (B) Concentration response curves for
cytotoxicity (MTT assay) in SHꢀSY5Y treated for 48 hours with compounds 39, 61, 62 and 64.
Values are the mean ± SD of 8 determinations from 2 different experiments. n.d.: not
determined.
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Remarkably, the Nꢀmethylated combretabenzodiazepines 44-48 were far less active compared to
to the corresponding combretabenzodiazepines 39-43.
Compounds 42 and 43 displayed IC s of 90.3 ± 14.3 nM and 71.0 ± 14.1 nM, respectively.
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The most potent compound of this series was compound 39, displaying an apparent IC of 25.1
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14.7 nM. To investigate whether the cytotoxic effect of the two most potent compounds could
be reꢀconducted to tubulin polymerization, we performed flow cytometry analysis of the cell
cycle of cells treated with 39 or 43. The typical signature of tubulin depolymerizing agents, i.e. a
G2/M arrest, was evident and quantitatively similar to CAꢀ4, for both compounds (Figure 5).
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Figure 5. CA-4 and the most potent compounds synthesized induce G2/M arrest. Cell
cycle analysis (using BD Accuri C6 flow cytometry) of SHꢀSY5Y cells treated for 16 hours with
vehicle (CTRL) or with the indicated compounds at the IC or 3 x IC values. The Yꢀaxis
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represents the cell number, and the Xꢀaxis represents propidium iodide (PI) fluorescence on a
linear scale. FL2ꢀA: PI staining. Data are representative of 3 different experiments.
To confirm this finding, we also performed an in vivo tubulin polymerization assay in SHꢀ
SY5Y cells. Briefly, in this assay, cells are grown for 16 h in the presence of pharmacological
agents after which cells are lysed in the presence of paclitaxel to maintain tubulin status.
Polymerized tubulin is then pelleted by centrifugation while monomers remain in the
supernatant. As it can be observed in Figure 6, 1, 39, and 43 significantly reduced the amount of
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pelletable (polymerized) tubulin, strongly suggesting that these agents act on tubulin by
inhibiting polymerization.
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Figure 6. CA-4 and compounds 43, 61 and 39 synthesized affect tubulin polymerization.
Western blot of αꢀtubulin extracted in the presence of paclitaxel from SHꢀSY5Y cells treated
with vehicle (CTRL) or the indicated compounds at 3 X IC value for 16 hours. Results are
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representative of 3 separate experiments. P = pelletable fraction, S = soluble fraction.
Given that the two methods above may be seen as indirect evidences of an effect on tubulin
polymerization, we also visualized tubulin in intact cells via immunocytochemistry and
investigated directly the effect of compounds 43 and 39 on tubulin polymerization in vitro. As
predicted, the ordered tubulin network seen in control cells by immunocytochemistry was
deranged when cells were treated with combretastatin, 39 or 43 at the IC value (see supporting
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information). Furthermore, tubulin polymerization was inhibited by both 39 and 43 in a doseꢀ
dependent manner (Figure 7) with IC50s of 9.0 ± 1.3 ꢁM and 13.6 ± 1.1 ꢁM, respectively
(compared to 1.3 ± 0.1 ꢁM of 1).
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Figure 7. Representative traces of the tubulin in vitro polymerization assay using a commercial
kit (see materials and methods). Experiments depicted are from a single 96ꢀwell plate and
therefore the control and paclitaxel traces are identical for all four panels are presented each time
for comparison.
From previous SAR studies of 1 and 2, an isovanillin moiety in the Bꢀring significantly
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increases potency, but due to the rigidity of the 1,4ꢀbenzodiazepineꢀ2ꢀone scaffold two
positions were available to graft the phenyl. To understand the spatial orientation of
combretabenzodiazepines in the tubulin pocket we pursued two strategies: (i) we synthesized the
two isovanillinꢀcontaining combretabenzodiazepines; and (ii) we performed docking studies.
For the synthesis, starting from the aldehydes 49 and 50, the same procedure developed for the
combretabenzodiazepines 39ꢀ43 was used. In order to successfully carry out the Grignard
reaction, the hydroxyl groups of the aldehydes were protected with tertꢀbutyldimethylsilyl
chloride to afford the protected aldehydes 51 and 52. The reduction of the nitro group to amino
was not carried out with iron/acetic acid/hydrochloric acid as in the presence of these strong acid
conditions the silyl protecting group was cleaved (the silyl deprotection was even observed when
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the nitro reduction was attempted using sodium dithionite in acetone at 50 °C). In this case
hydrogenation in presence of Pd/C 10% successfully afforded the silyl protected 2ꢀ
aminobenzophenones 57 and 58. We point out that during the cyclization procedure, besides the
desired combretabenzodiazepine 61, obtained in 36% yield, we isolated a second
combretabenzodiazepine 62 in 21% yield. The extra ring was formed by the reaction between the
formaldehyde deriving from thermal decomposition of HMTA with the NH and OH group of the
Bꢀring. Remarkably the silyl group was lost during this stage, while to obtain 64 it was necessary
performed a deprotection TBAF mediated (Scheme 2).
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Scheme 2. Scheme for the synthesis of combretabenzodiazepines 61 and 64.
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When testing the three new compounds, 62 was devoid of activity up to 1 µM, compound 64
was poorly active while 61 gained a 10ꢀfold potency over the parent compound 39, and showed
similar potency in respect to CAꢀ4 displaying an IC of 3.9 ± 2.6 nM (Figure 4).
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As expected, compound 61 induced a G2/M cell cycle arrest (Figure 5), inhibited tubulin
polymerization in the in vivo (Figure 6) and in the in vitro assays (Figure 7), and deranged
tubulin organization in intact cells as visualized by immunocytochemistry (see Supporting
Information Figure S5). This confirmed that the mechanism of action of 61 is to inhibit tubulin
polymerization.
Given that we wanted to explore the docking mode of 61 to tubulin, we first needed to provide
formal proof that, similarly to 1 and 2, it bound to the colchicine binding site. To investigate this,
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we performed [ H]colchicine radioligand binding assays. As shown in Table 1, 61 inhibited
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[ H]colchicine binding at similar concentrations compared to nonꢀradioactive colchicine, which
was used as control.
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Table 1. Compound 61 competes with [ H]colchicine for the same binding site on tubulin.
Values represent mean ± SEM of at least 7 replicates from three separate experiments.
Compound Inhibition of colchicine binding
%
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1
1
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1
1
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1
48 ± 6
80 ± 4
0
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0
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2
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4
5
6
7
8
9
0
Molecular modeling studies were then performed to investigate the potential pose of 61 to the
colchicine binding site of α,βꢀtubulin. Docking studies showed that 61, the most active
compound, occupies the colchicine binding site of tubulin in agreement with the Xꢀray structure
2
6
complex of DAMAꢀcolchicineꢀα,βꢀtubulin (PDB id: 1SA0). The trimethoxyphenyl moiety in
ring A of 61 was positioned in the binding cavity buried in the βꢀsubunit. The thiol group of
Cysβ241 formed a hydrogen bond with the oxygen atom of one of the methoxy groups, and
several amino acids of βꢀtubulin formed hydrophobic interactions with the trimethoxyphenyl
moiety. The hydroxyl group of ring B formed a hydrogen bond with the main chain nitrogen
atom of Valα181 (Figure 8). In the case of compound 64 the docked pose is slight shifted
compared to the colchicine binding mode, the trimethoxyphenyl group occupies the same cavity
around Cysβ241, without forming an hydrogen bond. The different position of the hydroxyl
group on ring B implies a missing hydrogen bonding interaction with Valα181 and an
unfavorable interaction with the hydrophobic region around Metβ259.
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49
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53
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60
Figure 8. Superimposition of the docked conformation of compounds 61 (a) and 64 (b) as cyan
stick models on top of the Xꢀray structure of DAMAꢀcolchicine (pink wire model). The
backbone of tubulin is shown as ribbon representation (αꢀtubulin: yellow, βꢀtubulin: green). The
amino acids of tubulin within 4.0 Å from colchicine are shown as wire models. Hydrogen bonds
(distance < 3 Å) are shown as dotted yellow lines.
CONCLUSIONS
In conclusion, we now report a novel class of compounds named combretabenzodiazepines
with similar efficacy and potency compared to CAꢀ4. Although CAꢀ4 has entered clinical trials,
doubleꢀbond isomerisation, a short halfꢀlife and metabolic stability could represent an Achille’s
heel and therefore a point of improvement. We therefore also compared the metabolic profile
(phase I/II) of CAꢀ4 and 61. When incubated in the presence of human liver microsomes, CAꢀ4
underwent a relevant phase I oxidative transformation, with a residual substrate after 1 h ranging
between 75% to 67% depending on the presence or absence of GSH respectively (Table 2); in
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the phase II incubations the residual substrate was about 39%. Conversely, a greater metabolic
stability was observed for 61, undergoing poor substrate depletion compared to CAꢀ4, being the
residual substrate >95% and 92% for phase I/II respectively (Table 2). The structural
characterization of the metabolites and the corresponding metabolic pathways involved in the
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
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7
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9
0
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9
0
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9
0
2
metabolism of 61 were achieved by tandem mass spectrometry (LCꢀESIꢀMS ) analysis and are
present in the supporting information.
Table 2. Metabolic profile for 1 and 61. Data are expressed as the peak area percent relative to
controls (without microsomes) obtained from LCꢀUV analyses of incubation extracts (t = 60 min,
phase I or 30 min, phase II incubations).
Phase I
HLM+NADPH
Phase II
HLM+UDPGA
39%
Compound
HLM
100%
95%
HLM+NADPH+GSH
1
75%
96%
67%
97%
61
92%
Overall, these data show that 61 is characterized by a greater in vitro phase I/II metabolic
stability with respect to the reference compound CAꢀ4. The 1,4ꢀbenzodiazepinꢀ2ꢀone nucleus
appears, alongside avoiding the isomerization problem associated with the double bond, to
9
mitigate the occurrence of both the Oꢀdemethylation and aromatic hydroxylation pathways and
2
7
the extensive glucuronidation of the phenolic function.
Table 3. Summary of metabolic profile for 1 and 61.
Metabolic pathways
1
61
Aromatic hydroxylation
+
ꢀ
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Oꢀdemethylation
+
+
+
ꢀ
Reactive metabolites
Isomerization
+
ꢀ
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53
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60
Glucuronidation
+++
ꢀ
+
ꢀ
Hydrolysis (in vitro 1h)
Finally, as a consequence of the lack of aromatic hydroxylation, the formation of glutathione
adducts was not observed indicating the absence of reactive metabolites in the metabolism of 61
(
Table 3). On the contrary the metabolic fate of CAꢀ4 is characterized by the formation of
electrophilic quinone species that reacted with nucleophilic thiols.
These compounds, in our opinion deserve consideration for the development of novel
antitubulin agents.
EXPERIMENTAL SECTION
Chemistry General Procedures. Commercially available reagents and solvents were
purchased from Aldrich or Alfa Aesar and were used without further purification.
Tetrahydrofuran (THF) was distilled immediately before use from Na/benzophenone under a
slight positive atmosphere of dry nitrogen. N,N’ꢀDimethylformamide (DMF) was distilled under
vacuum from KOH and stored on activated molecular sieves (4 Å). Dichloromethane was dried
by distillation from P O and stored on activated molecular sieves (4 Å). When needed the
2
5
reactions were performed in flameꢀ or ovenꢀdried glassware under a positive pressure of dry
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nitrogen. Melting points were determined in open glass capillary with a Stuart scientific SMP3
apparatus and are uncorrected. All compounds were checked by IR (FTꢀIR THERMOꢀNICOLET
1
13
AVATAR), H and C APT (JEOL ECP 300 MHz spectrometer), and mass spectrometry
0
1
2
3
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9
0
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7
8
9
0
(Thermo Finningan LCQꢀdeca XPꢀplus) equipped with an ESI source and an ion trap detector.
Chemical shifts are reported in parts per million (ppm). Flash column chromatography was
performed on silica gel (Merck Kieselgel 60, 230–400 mesh ASTM). Thin layer chromatography
(TLC) was carried out on 5x20 cm plates with a layer thickness of 0.25 mm (Merck Silica gel 60
F ). When necessary they were developed with KMnO reagent. Purity of tested compounds
2
54
4
was established by elemental analysis. Elemental analysis (C, H, N) of the target compounds are
within ±0.4% of the calculated values.
Preparation of aldehydes 14ꢀ18, 49 and 50. The 4ꢀmethoxyꢀ2ꢀnitrobenzaldehyde (14) was
28
synthesized according to the literature. The 5ꢀmethoxyꢀ2ꢀnitrobenzaldehyde (15) was
2
9
synthesized according to the literature. The 4,5ꢀdimethoxyꢀ2ꢀnitrobenzaldehyde (16) and 6ꢀ
nitrobenzo[1,3]dioxoleꢀ5ꢀcarbaldehyde (18) were commercially available. The 3,4ꢀdimethoxyꢀ2ꢀ
30
nitrobenzaldehyde (17) was synthesized according to the literature. The 3ꢀhydroxyꢀ4ꢀmethoxyꢀ
2ꢀnitrobenzaldehyde (49) and 5ꢀhydroxyꢀ4ꢀmethoxyꢀ2ꢀnitrobenzaldehyde (50) were synthesized
3
1
according to the literature.
Preparation of OTBDMSꢀprotected aldeydes 51 and 52. The 3ꢀ(tertꢀbutyldimethylsilyloxy)ꢀ4ꢀ
methoxyꢀ2ꢀnitrobenzaldehyde (51) and the 5ꢀ(tertꢀbutyldimethylsilyloxy)ꢀ4ꢀmethoxyꢀ2ꢀ
nitrobenzaldehyde (52) were synthesized starting from aldehydes 49 and 50 respectively,
3
2
according to the literature.
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General procedure for the preparation of 2ꢀnitrobenzhydrols 19ꢀ23, 53ꢀ54. To a dried threeꢀ
necked flask equipped with a dropping funnel, reflux condenser, and magnetic stirrer was added
magnesium turnings (10 mmol, 1.4 eq), a small piece of iodine and 3.0 mL of dry THF. The
mixture was gentle heated while approximately 1/4 of 3,4,5ꢀtrimethoxybromobenzene solution
(13) (10 mmol, 1.4 eq) in dry THF (10.0 mL) was added dropwise. As soon as the solution
became colorless the remaining 3,4,5ꢀtrimethoxylbromobenzene solution was added dropwise
under mild reflux and stirring was continued for 1 h at room temperature. Afterward the 3,4,5ꢀ
trimethoxyphenylmagnesium bromide solution prepared was slowly added to the corresponding
substituted 2ꢀnitrobenzaldehyde (7 mmol, 1 eq) in 10.0 mL of dry THF at 0 °C. After complete
addition, the solution was allowed to stir at room temperature for 12 h. A saturated aqueous
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53
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59
60
NH Cl solution was slowly added to hydrolyze the adduct at 0 °C, and the mixture was stirred
4
for 10 min. The phases were separated, and the aqueous layer was extracted with EtOAc (x3).
The combined organic layers were washed with brine (x1), dried over sodium sulfate, and
filtered. The filtrate was concentrated in vacuo, and the residue was purified by column
chromatography to afford the desired 2ꢀnitrobenzhydrol.
(4-methoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanol (19). The title compound was
prepared from 14. The crude material was purified by column chromatography using PE/EtOAc
8
:2 and PE/EtOAc 7:3 as eluants and was crystallized with MeOH to give a pale yellow solid,
ꢀ
yield 37%, m.p. 130ꢀ132 °C. IR (KBr) 3471, 2956, 2828, 1595, 1537, 1324, 1128, 1003, 851 cm
1
1
.
HꢀNMR (300 MHz, CDCl ) δ 7.49 (d, J = 8.5 Hz, 1ꢀH), 7.39 (d, J = 1.5 Hz, 1ꢀH), 7.01 (dd, J
3
1
3
=
8.5/1.8 Hz, 1ꢀH), 6.55 (s, 2ꢀH), 6.24 (s, 1ꢀH), 3.84 (s, 3ꢀH), 3.80ꢀ3.78 (m, 9ꢀH) ppm; CꢀNMR
(
75 MHz, CDCl ) δ 159.3, 153.3, 149.1, 137.7, 137.4, 119.7, 109.4, 103.9, 70.9, 60.9, 56.2, 56.0
3
+
ppm; MS (ESI) m/z 332 [M+HꢀH O] .
2
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(
5-methoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanol (20). The title compound was
prepared from 15. The crude material was purified by column chromatography using PE/EtOAc
8
3
:2 and PE/EtOAc 7:3 as eluants and was crystallized with MeOH to give a brown solid, yield
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
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7
8
9
0
1
2
3
4
5
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7
8
9
0
ꢀ1 1
9%, m.p. 110ꢀ112 °C. IR (KBr) 3505, 2941, 2841, 1592, 1506, 1237, 1126, 835 cm . HꢀNMR
(300 MHz, CDCl ) δ 8.08 (d, J = 9.1 Hz, 1ꢀH), 7.19 (d, J = 2.7 Hz, 1ꢀH), 6.90 (dd, J = 9.1/2.7
3
1
3
Hz, 1ꢀH), 6.57 (s, 2ꢀH), 6.44 (s, 1ꢀH), 3.88 (s, 3ꢀH), 3.82 (s, 3ꢀH), 3.81 (s, 6ꢀH) ppm; CꢀNMR
(
75 MHz, CDCl ) δ 163.8, 153.3, 142.0, 141.2, 137.6, 137.3, 128.0, 114.5, 113.0, 104.2, 71.6,
3
+
60.9, 56.2, 56.0 ppm; MS (ESI) m/z 332 [M+HꢀH O] .
2
(4,5-dimethoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanol (21). The title compound
was prepared from 16. The crude material was purified by column chromatography using
PE/EtOAc 8:2 and PE/EtOAc 5:5 as eluants and was crystallized with MeOH to give a pale
yellow solid, yield 30%, m.p. 148ꢀ149 °C. IR (KBr) 3438, 2935, 2836, 1586, 1526, 1336, 1262,
ꢀ1 1
1
2
211, 1126, 1067, 742 cm . HꢀNMR (300 MHz, CDCl ) δ 7.60 (s, 1ꢀH), 7.16 (s, 1ꢀH), 6.58 (s,
3
1
3
ꢀH), 6.44 (s, 1ꢀH), 3.95 (s, 3ꢀH), 3.92 (s, 3ꢀH), 3.82 (s, 3ꢀH), 3.81 (s, 6ꢀH) ppm; CꢀNMR (75
MHz, CDCl ) δ 153.4, 153.1, 148.0, 140.3, 137.8, 137.4, 134.0, 110.5, 108.0, 104.1, 71.2, 60.8,
3
+
56.4, 56.3, 56.1 ppm; MS (ESI) m/z 362 [M+HꢀH O] .
2
(3,4-dimethoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanol (22). The title compound
was prepared from 17. The crude material was purified by column chromatography using
PE/EtOAc 8:2 and PE/EtOAc 7:3 as eluants and was crystallized with MeOH to give a yellowꢀ
orange solid, yield 33%, m.p. 126ꢀ127 °C. IR (KBr) 3420, 2941, 2831, 1650, 1596, 1539, 1504,
ꢀ1 1
1
282, 1131, 1054 cm . HꢀNMR (300 MHz, CDCl ) δ 7.03 (d, J = 8.7, Hz, 1ꢀH), 6.96 (d, J = 8.7
3
1
3
Hz, 1ꢀH), 6.59 (s, 2ꢀH), 5.79 (s, 1ꢀH), 3.92 (s, 3ꢀH), 3.88 (s, 3ꢀH), 3.82 (brs, 9ꢀH) ppm; CꢀNMR
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(
75 MHz, CDCl ) δ 153.1, 152.5, 145.0, 140.4, 137.8, 137.1, 127.9, 123.2, 114.0, 103.3, 70.6,
3
+
6
2.1, 60.8, 56.2, 56.0 ppm; MS (ESI) m/z 362 [M+HꢀH O] .
2
(6-nitrobenzo[d][1,3]dioxol-5-yl)(3,4,5-trimethoxyphenyl)methanol (23). The title
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
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39
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48
49
50
51
52
53
54
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56
57
58
59
60
compound was prepared from 18. The crude material was purified by column chromatography
using PE/EtOAc 8:2 and PE/EtOAc 7:3 as eluants and was crystallized with MeOH to give a
yellow solid, yield 71%, m.p. 195ꢀ197 °C. IR (KBr) 3064, 2950, 2842, 1591, 1520, 1501, 1330,
ꢀ1 1
1
263, 1132, 1029, 920 cm . HꢀNMR (300 MHz, CDCl ) δ 7.47 (s, 1ꢀH), 7.04 (s, 1ꢀH), 6.59 (s,
3
13
2ꢀH), 6.35 (s, 1ꢀH), 6.11 (s, 1ꢀH), 3.82 (brs, 9ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 153.3,
3
1
52.2, 147.3, 142.3, 137.6, 137.5, 136.4, 108.3, 105.5, 103.9, 103.2, 70.9, 60.9, 56.2 ppm; MS
+
(
ESI) m/z 346 [M+HꢀH O] .
2
(3-(tert-butyldimethylsilyloxy)-4-methoxy-2-nitrophenyl)(3,4,5-
trimethoxyphenyl)methanol (53). The title compound was prepared from 51. The crude
material was purified by column chromatography using PE/EtOAc 8:2 and PE/EtOAc 7:3 as
eluants and was crystallized with MeOH to give a yellow solid, yield 61%, m.p. 99ꢀ101 °C. IR
ꢀ1 1
(
KBr) 3291, 2937, 2858, 1533, 1506, 1292, 1129, 830 cm . HꢀNMR (300 MHz, CDCl ) δ 6.89
3
(d, J = 8.6 Hz, 1ꢀH), 6.84 (d, J = 8.6 Hz, 1ꢀH), 6.56 (s, 2ꢀH), 5.75 (s, 1ꢀH), 3.80–3.76 (m, 12ꢀH),
13
0
.94–0.91 (m, 9ꢀH), 0.18–0.16 (m, 6ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 153.3, 150.3, 143.5,
3
1
37.5, 137.3, 137.2, 128.0, 120.0, 112.8, 103.4, 71.2, 60.9, 56.2, 55.6, 25.7, 18.6, ꢀ3.96 ppm; MS
+
(ESI) m/z 462 [M+HꢀH O] .
2
(5-(tert-butyldimethylsilyloxy)-4-methoxy-2-nitrophenyl)(3,4,5-
trimethoxyphenyl)methanol (54). The title compound was prepared from 52. The crude
material was purified by column chromatography using PE/EtOAc 8:2 and PE/EtOAc 7:3 as
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1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
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4
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5
5
5
5
5
5
5
5
6
eluants and was crystallized with MeOH to give a yellow solid, yield 32%, m.p. 118ꢀ119 °C. IR
ꢀ1 1
(
KBr) 3448, 2937, 1595, 1516, 1459, 1330, 1282, 1131, 1060, 836, 796 cm . HꢀNMR (300
MHz, CDCl ) δ 7.55 (s, 1ꢀH), 7.00 (s, 1ꢀH), 6.54 (s, 2ꢀH), 6.32 (s, 1ꢀH), 3.85–3.77 (m, 12ꢀH),
3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
13
3
35 (brs, 1ꢀH), 0.92 (brs, 9ꢀH), 0.11 (brs, 6ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 153.2, 150.3,
3
150.1, 141.1, 137.7, 137.2, 133.7, 120.7, 108.9, 103.9, 71.1, 60.8, 56.0, 56.0, 25.6, 18.5, ꢀ4.4
+
ppm; MS (ESI) m/z 462 [M+HꢀH O] .
2
General procedure for the preparation of 2ꢀnitrobenzophenones 24ꢀ28, 55ꢀ56. To a stirred
solution of the corresponding 2ꢀnitrobenzhydrol (4 mmol, 1 eq) and 4 Å molecular sieves (0.40
g) in dry dichloromethane (30.0 mL) was added pyridinium dichromate (PDC, 6 mmol, 1,5 eq)
portionꢀwise at 0 °C. The reaction mixture was stirred under nitrogen atmosphere at room
temperature for 12 h. Afterward the mixture was diluted with dichloromethane (30.0 mL) and
filtered through a pad of Celite. The filtrate was concentrated in vacuo and the residue was
purified by column chromatography to obtain the desired 2ꢀnitrobenzophenone.
(4-methoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanone (24). The title compound
was prepared from 19. The crude material was purified by column chromatography using
PE/EtOAc 7:3 and PE/EtOAc 5:5 as eluants and was crystallized with MeOH to give an off
white solid, yield 82%, m.p. 164ꢀ165 °C. IR (KBr) 3103, 2945, 2843, 1678, 1584, 1531, 1332,
ꢀ1 1
1
233, 1129, 995, 864 cm . HꢀNMR (300 MHz, CDCl ) δ 7.64 (d, J = 1.8 Hz, 1ꢀH), 7.44 (d, J =
3
8.5 Hz, 1ꢀH), 7.26 (dd, J = 8.2/3.4 Hz, 1ꢀH), 6.98 (s, 1ꢀH), 3.96 (s, 3ꢀH), 3.91 (s, 3ꢀH), 3.82 (s, 6ꢀ
13
H) ppm; CꢀNMR (75 MHz, CDCl ) δ 192.1, 161.2, 153.3, 148.8, 143.3, 131.6, 130.6, 127.9,
3
+
1
19.8, 109.4, 107.0, 61.0, 56.4, 56.3 ppm; MS (ESI) m/z 348 [M+H] .
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(
5-methoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanone (25). The title compound
was prepared from 20. The crude material was purified by column chromatography using
PE/EtOAc 8:2 and PE/EtOAc 5:5 as eluants to give a pale yellow solid, yield 82%, m.p. 156ꢀ157
10
11
12
13
14
15
16
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19
20
21
22
23
24
25
26
27
28
29
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31
32
33
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46
47
48
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50
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53
54
55
56
57
58
59
60
ꢀ1 1
°
C. IR (KBr) 3101, 2972, 2943, 2845, 1671, 1579, 1333, 1233, 1124, 755 cm . HꢀNMR (300
MHz, CDCl ) δ 8.22 (d, J = 9.2 Hz, 1ꢀH), 7.07 (dd, J = 9.2/2.8 Hz, 1ꢀH), 6.99 (s, 2ꢀH), 6.88 (d, J
3
1
3
=
2.8 Hz, 1ꢀH), 3.92 (s, 3ꢀH), 3.90 (s, 3ꢀH), 3.81 (s, 6ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ
3
1
92.1, 164.1, 153.3, 143.4, 139.6, 138.7, 131.0, 127.1, 115.4, 113.6, 106.8, 61.0, 56.4, 56.4 ppm;
+
MS (ESI) m/z 348 [M+H] .
(4,5-dimethoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanone
(26).
The
title
compound was prepared from 21. The crude material was purified by column chromatography
using PE/EtOAc 7:3 and PE/EtOAc 4:6 as eluants to give an off white solid, yield 82%, m.p. 151
ꢀ1
1
°
C. IR (KBr) 3101, 2937, 2832, 1661, 1582, 1524, 1415, 1334, 1284, 1219, 1130, 757 cm . Hꢀ
NMR (300 MHz, CDCl ) δ 7.71 (s, 1ꢀH), 6.99 (s, 2ꢀH), 6.86 (s, 1ꢀH), 4.03 (s, 3ꢀH), 3.98 (s, 3ꢀH),
3
1
3
3
1
.92 (s, 3ꢀH), 3.82 (s, 6ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 192.3, 153.9, 153.4, 149.8,
3
43.4, 139.8, 131.4, 130.2, 110.2, 106.9, 106.8, 61.1, 56.8, 56.7, 56.4 ppm; MS (ESI) m/z 378
+
[M+H] .
(
3,4-dimethoxy-2-nitrophenyl)(3,4,5-trimethoxyphenyl)methanone (27). The title
compound was prepared from 22. The crude material was purified by column chromatography
using PE/EtOAc 6:4 and PE/EtOAc 5:5 as eluants to give a brown solid, yield 66%, m.p. 161ꢀ
ꢀ1
1
1
63 °C. IR (KBr) 3002, 2944, 2141, 1659, 1536, 1335, 1280, 1132, 990, 754 cm . HꢀNMR (300
MHz, CDCl ) δ 7.03 (d, J = 8.7 Hz, 1ꢀH), 6.96 (d, J = 8.7 Hz, 1ꢀH), 6.59 (s, 2ꢀH), 5.79 (s, 1ꢀH),
3
1
3
3
.92 (s, 3ꢀH), 3.88 (s, 3ꢀH), 3.82 (brs, 9ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 153.1, 152.5,
3
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5
5
5
6
1
45.0, 140.4, 137.8, 137.1, 127.9, 123.2, 114.0, 103.3, 70.6, 62.1, 60.8, 56.2 ppm; MS (ESI) m/z
+
3
78 [M+H] .
(6-nitrobenzo[d][1,3]dioxol-5-yl)(3,4,5-trimethoxyphenyl)methanone (28). The title
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
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2
3
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7
8
9
0
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2
3
4
5
6
7
8
9
0
1
2
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4
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6
7
8
9
0
compound was prepared from 23. The crude material was purified by column chromatography
using PE/EtOAc 5:5 and PE/EtOAc 2:8 as eluants to give a pale orange solid, yield 91%, m.p.
ꢀ
1
95ꢀ197 °C. IR (KBr) 3051, 2939, 2 l’842, 1661, 1584, 1536, 1504, 1421, 1333, 1128, 1030 cm
1
1
.
HꢀNMR (300 MHz, CDCl ) δ 7.62 (s, 1ꢀH), 6.97 (s, 2ꢀH), 6.81 (s, 1ꢀH), 6.21 (s, 2ꢀH), 3.89 (s,
3
13
3ꢀH), 3.81 (brs, 6ꢀH), ppm; CꢀNMR (75 MHz, CDCl ) δ 191.6, 153.3, 152.7, 149.1, 143.3,
3
ꢀ
1
41.4, 132.6, 131.0, 107.8, 106.7, 105.0, 103.8, 61.0, 56.4 ppm; MS (ESI) m/z 360 [MꢀH] .
(3-(tert-butyldimethylsilyloxy)-4-methoxy-2-nitrophenyl)(3,4,5-
trimethoxyphenyl)methanone (55). The title compound was prepared from 53. The crude
material was purified by column chromatography using PE/EtOAc 6:4 as eluant to give a white
solid, yield 84%, m.p. 138ꢀ139 °C. IR (KBr) 3009, 2937, 2358, 1652, 1543, 1500, 1416, 1331,
ꢀ1 1
1
127, 840 cm . HꢀNMR (300 MHz, CDCl ) δ 7.14 (d, J = 8.5 Hz, 1ꢀH), 7.04 (s, 2ꢀH), 7.98 (d, J
3
1
3
=
8.5 Hz, 1ꢀH), 3.92–3.85 (m, 12ꢀH), 0,94 (brs, 9ꢀH), 0.21 (brs, 6ꢀH) ppm; CꢀNMR (75 MHz,
CDCl ) δ 191.3, 153.8, 153.0, 143.5, 142.8, 138.6, 131.6, 125.0, 123.3, 111.5, 107.5, 61.1, 56.4,
3
+
5
5.9, 25.6, 18.6, ꢀ4.0 ppm; MS (ESI) m/z 478 [M+H] .
(5-(tert-butyldimethylsilyloxy)-4-methoxy-2-nitrophenyl)(3,4,5-
trimethoxyphenyl)methanone (56). The title compound was prepared from 54. The crude
material was purified by column chromatography using PE/EtOAc 8:2 as eluant to give a pale
yellow solid, yield 81%, m.p. 150ꢀ152 °C. IR (KBr) 3040, 2941, 2856, 1675, 1567, 1518, 1415,
ꢀ1 1
1
331, 1294, 1134, 838 cm . HꢀNMR (300 MHz, CDCl ) 7.70 (s, 1ꢀH), 6.97 (s, 2ꢀH), 6.83 (s, 1ꢀ
3
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H), 3.94 (s, 3ꢀH), 3.89 (s, 3ꢀH), 3.79 (s, 6ꢀH), 0.96 (brs, 9ꢀH), 0.18 (brs, 6ꢀH) ppm; CꢀNMR (75
MHz, CDCl ) δ 192.1, 153.3, 151.7, 150.8, 143.3, 140.3, 131.5, 130.2, 120.0, 107.9, 106.8, 61.0,
3
+
5
6.4, 56.1, 25.6, 18.6, ꢀ4.4 ppm; MS (ESI) m/z 478 [M+H] .
10
11
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18
19
20
21
22
23
24
25
26
27
28
29
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31
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49
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54
55
56
57
58
59
60
General procedure for the preparation of 2ꢀaminobenzophenones 29ꢀ33. A mixture of the
corresponding 2ꢀnitrobenzophenone (2.8 mmol, 1 eq) and iron powder (7.7 eq), in ethanol (10.0
mL), acetic acid (10 mL), water (5.0 mL), and 35% hydrochloric acid (2 drops) was vigorously
stirred and refluxed for 2 h. The suspension was cooled and filtered through a pad of Celite. The
filtrate was diluted with water and extracted with dichloromethane (x3). The combined organic
layers were washed with saturated aqueous NaHCO solution (x1), water (x1), dried over sodium
3
sulfate and concentrated in vacuo. Finally the residue was purified by column chromatography to
afford the desired 2ꢀaminobenzophenone.
(2-amino-4-methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (29). The title compound
was prepared from 24. The crude material was purified by column chromatography using
PE/EtOAc 7:3 as eluant to give a yellow solid, yield 98%, m.p. 94ꢀ95 °C. IR (KBr) 3441, 3338,
ꢀ1 1
2
935, 2832, 1616, 1582, 1343, 1228, 1120, 770 cm . HꢀNMR (300 MHz, CD OD) δ 7.34 (d, J
3
= 9.2 Hz, 1ꢀH), 6.82 (s, 2ꢀH), 6.31 (d, J = 2.5 Hz, 1ꢀH), 6.15 (dd, J = 9.2/2.4 Hz, 1ꢀH), 3.84 (s, 6ꢀ
13
H), 3.83 (s, 3ꢀH), 3.80 (s, 3ꢀH) ppm; CꢀNMR (75 MHz, CD OD) δ 198.3, 166.2, 156.0, 154.1,
3
+
1
41.3, 137.6, 137.5, 112.6, 107.6, 105.0, 99.9, 61.2, 56.7, 55.7 ppm; MS (ESI) m/z 318 [M+H] .
(2-amino-5-methoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (30). The title compound
was prepared from 25. The crude material was purified by column chromatography using
PE/EtOAc 7:3 as eluant to give a yellow solid, yield 76%, m.p. 130ꢀ132 °C. IR (KBr) 3420,
ꢀ1 1
3
308, 2944, 2832, 1639, 1583, 1411, 1333, 1213, 1134, 777 cm . HꢀNMR (300 MHz, CDCl ) δ
3
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2
2
2
2
2
2
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3
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5
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6
7
.01ꢀ6.94 (m, 4ꢀH), 6.77 (d, J = 8.6 Hz, 1ꢀH), 3.92 (s, 3ꢀH), 3.87 (br s, 6ꢀH), 3.68 (s, 3ꢀH) ppm;
1
3
CꢀNMR (75 MHz, CDCl ) δ 197.5, 152.9, 149.9, 145.3, 141.1, 134.9, 122.8, 118.6, 116.7,
3
+
1
07.0, 61.0, 56.4, 56.1 ppm; MS (ESI) m/z 318 [M+H] .
0
1
2
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8
9
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8
9
0
(2-amino-4,5-dimethoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (31). The title
compound was prepared from 26. The crude material was purified by column chromatography
using PE/EtOAc 7:3 as eluant to give a yellow solid, yield 71%, m.p. 129ꢀ131 °C. IR (KBr)
ꢀ1 1
3452, 3323, 2944, 2833, 1629, 1582, 1414, 1337, 1207, 1134, 775 cm . HꢀNMR (300 MHz,
CDCl ) δ 7.00 (s, 1ꢀH), 6.88 (s, 2ꢀH), 6.35 (s, 1ꢀH), 3.91 (s, 6ꢀH), 3.86 (s, 6ꢀH), 3.69 (s, 3ꢀH)
3
1
3
ppm; CꢀNMR (75 MHz, CDCl ) δ 196.1, 155.4, 152.8, 148.2, 140.4, 139.8, 135.8, 116.3,
3
+
1
10.0, 106.5, 99.6, 61.0, 56.6, 56.3, 55.9 ppm; MS (ESI) m/z 348 [M+H] .
(2-amino-3,4-dimethoxyphenyl)(3,4,5-trimethoxyphenyl)methanone (32). The title
compound was prepared from 27. The crude material was purified by column chromatography
using PE/EtOAc 7:3 as eluant to give a yellow solid, yield 89%, m.p. 140ꢀ141 °C. IR (KBr)
ꢀ1 1
3
481, 3346, 2968, 2942, 2833, 1617, 1578, 1529, 1241, 1129, 997, 743 cm . HꢀNMR (300
MHz, CD OD) δ 7.27 (d, J = 9.3 Hz, 1ꢀH), 6.84 (s, 2ꢀH), 6.23 (d, J = 9.3 Hz, 1ꢀH), 3.90–3.85
3
13
(m, 15ꢀH) ppm; CꢀNMR (75 MHz, CD OD) δ 197.1, 156.0, 152.8, 146.2, 140.5, 135.8, 134.7,
3
+
1
31.3, 113.6, 106.6, 99.8, 61.0, 59.8, 56.3, 55.9 ppm; MS (ESI) m/z 348 [M+H] .
(6-aminobenzo[d][1,3]dioxol-5-yl)(3,4,5-trimethoxyphenyl)methanone (33). The title
compound was prepared from 28. The crude material was purified by column chromatography
using PE/EtOAc 7:3 and PE/EtOAc 5:5 as eluants to give a yellow solid, yield 86%, m.p. 150ꢀ
ꢀ1
1
52 °C. IR (KBr) 3450, 3332, 3003, 2904, 1583, 1560, 1496, 1413, 1333, 1233, 1133, 783 cm .
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HꢀNMR (300 MHz, CDCl ) δ 6.85 (s, 1ꢀH), 6.77 (s, 2ꢀH), 6.42 (brs, 2ꢀH), 6.18 (s, 1ꢀH), 5.84 (s,
3
1
3
2
ꢀH), 3.85–3.82 (brs, 9ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 196.1, 153.2, 152.8, 150.5, 140.0
3
+
1
38.3, 136.1, 111.3, 110.0, 106.1, 101.3, 96.8, 60.9, 56.2 ppm; MS (ESI) m/z 332 [M+H] .
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
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51
52
53
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57
58
59
60
General procedure for the preparation of OTBDMSꢀprotected 2ꢀaminobenzophenones 57ꢀ58.
To a solution of the corresponding OTBDMS protected 2ꢀnitrobenzophenone (0.122 g, 0.25
mmol, 1 eq) in dichloromethane (7.0 mL), was added Pd/C 10% (0.036 g). The resulting mixture
was vigorously stirred under a H atmosphere (1 atm) at room temperature. After 10 h the
2
suspension was filtered through a pad of Celite and the solvent was removed in vacuo The crude
material was purified by column chromatography using PE/EtOAc 8:2 and PE/EtOAc 7:3 as
eluants.
(2-amino-3-(tert-butyldimethylsilyloxy)-4-methoxyphenyl)(3,4,5-
trimethoxyphenyl)methanone (57). The title compound was prepared from 55. Yellow oil,
ꢀ1 1
yield 78%. IR (KBr) 3498, 3359, 2934, 2857, 1609, 1579, 1502, 1275, 1127, 834 cm . HꢀNMR
(
300 MHz, CDCl ) δ 7.15 (d, J = 9.1 Hz, 1ꢀH), 6.86 (s, 2ꢀH), 6.22 (d, J = 9.1 Hz, 1ꢀH), 3.90–
3
13
3
.86 (m, 12ꢀH), 1.04 (brs, 9ꢀH), 0.22 (brs, 6ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 197.2, 152.8,
3
144.9, 140.3, 136.0, 130.7, 128.5, 113.5, 106.6, 99.5, 61.0, 56.3, 55.0, 26.2, 18.8, ꢀ3.9 ppm; MS
+
(
ESI) m/z 448 [M+H] .
(2-amino-5-(tert-butyldimethylsilyloxy)-4-methoxyphenyl)(3,4,5-
trimethoxyphenyl)methanone (58). The title compound was prepared from 56. Yellow solid,
yield 75%,%, m.p. 124ꢀ127 °C. IR (KBr) 3470, 3359, 2934, 2856, 1623, 1578, 1533, 1251, 1116,
ꢀ1 1
8
3
32, 783 cm . HꢀNMR (300 MHz, CDCl ) δ 6.94 (s, 1ꢀH), 6.82 (s, 2ꢀH), 6.17 (s, 1ꢀH), 3.90–
3
13
.83 (m, 12ꢀH), 0.91 (brs, 9ꢀH), 0.07 (brs, 6ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 196.6, 157.2,
3
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2
2
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2
2
2
2
2
2
2
3
3
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3
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52.9, 148.4, 140.2, 136.1, 135.2, 124.7, 110.8, 106.3, 99.3, 61.1, 56.3, 55.5, 25.8, 18.4,ꢀ4.5
+
ppm; MS (ESI) m/z 448 [M+H] .
General procedure for the preparation of 2ꢀchloroacetamides 34ꢀ38, 59ꢀ60. To a cooled (0 °C)
solution of the corresponding 2ꢀaminobenzophenone (1 eq) in dry dichloromethane (0.2 M), was
added chloroacetyl chloride (1.1 eq) dropwise, and the reaction mixture was stirred at room
0
1
2
3
4
5
6
7
8
9
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1
2
3
4
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8
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8
9
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7
8
9
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2
3
4
5
6
7
8
9
0
temperature for 1h. Afterward a saturated aqueous NaHCO solution was added at 0 °C and the
3
mixture was vigorously stirred for 10 min. The organic layer was separated, washed with
aqueous saturated NaHCO solution (x1), dried over sodium sulfate and concentrated under
3
reduced pressure. Finally the crude material was purified by column chromatography to afford
the desired 2ꢀchloroacetamide.
2
-chloro-N-(5-methoxy-2-(3,4,5-trimethoxybenzoyl)phenyl)acetamide (34). The title
compound was prepared from 29. The crude material was purified by column chromatography
using PE/EtOAc 8:2 and PE/EtOAc 4:6 as eluants to give a white solid, yield 86%, m.p. 183ꢀ185
ꢀ1 1
°
C. IR (KBr) 3140, 2969, 2839, 1669, 1579, 1408, 1341, 1248, 1127, 617 cm . HꢀNMR (300
MHz, CDCl ) δ 12.05 (br s, 1ꢀH), 8.32 (s, 1ꢀH), 7.59 (d, J = 8.8 Hz, 1ꢀH), 6.91 (s, 2ꢀH), 6.65 (d,
3
13
J = 8.8 Hz, 1ꢀH), 4.20 (s, 2ꢀH), 3.92 (s, 3ꢀH), 3.91 (s, 3ꢀH), 3.87 (s, 6ꢀH) ppm; CꢀNMR (75
MHz, CDCl ) δ 197.5, 165.8, 164.3, 152.9, 142.2, 141.7, 135.8, 134.2, 116.8, 109.5, 107.4,
3
+
1
05.8, 61.0, 56.4, 55.8, 43.3 ppm; MS (ESI) m/z 394 [M+H] .
2-chloro-N-(4-methoxy-2-(3,4,5-trimethoxybenzoyl)phenyl)acetamide (35). The title
compound was prepared from 30. The crude material was purified by column chromatography
using PE/EtOAc 7:3 and PE/EtOAc 6:4 as eluants to give a pale yellow solid, yield 88%, m.p.
ꢀ1 1
1
39ꢀ140 °C. IR (KBr) 3208, 2954, 2835, 1674, 1585, 1540, 1337, 1137, 775 cm . HꢀNMR (300
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MHz, CDCl ) δ 10.90 (br s, 1ꢀH), 8.39 (d, J = 8.9 Hz, 1ꢀH), 7.16ꢀ7.10 (m, 2ꢀH), 7.02 (s, 2ꢀH),
3
1
3
4
1
5
.16 (s, 2.H), 3.94 (s, 3ꢀH), 3.87 (br s, 6ꢀH), 3.78 (s, 3ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ
3
97.1, 165.0, 155.1, 153.0, 142.6, 132.8, 131.6, 126.8, 123.9, 118.8, 117.7, 107.9, 61.1, 56.4,
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
+
5.8, 43.1 ppm; MS (ESI) m/z 394 [M+H]
2-chloro-N-(4,5-dimethoxy-2-(3,4,5-trimethoxybenzoyl)phenyl)acetamide (36). The title
compound was prepared from 31. The crude material was purified by column chromatography
using PE/EtOAc 5:5 and EtOAc as eluants to give an off white solid, yield 98%, m.p. 185ꢀ186
ꢀ1 1
°C dec. IR (KBr) 3119, 2958, 2832, 1671, 1583, 1524, 1328, 1208, 1136, 988, 776 cm . Hꢀ
NMR (300 MHz, CDCl ) δ 11.9 (br s, 1ꢀH), 8.37 (s, 1ꢀH), 7.13 (s, 1ꢀH), 6.96 (s, 2ꢀH), 4.19 (s, 2ꢀ
3
1
3
H), 4.00 (s, 3ꢀH), 3.94 (s, 3ꢀH), 3.87 (br s, 6ꢀH), 3.77 (s, 3ꢀH) ppm; CꢀNMR (75 MHz, CDCl )
3
δ 197.0, 165.5, 153.8, 153.0, 144.0, 141.8, 135.8, 134.0, 116.2, 115.7, 107.4, 104.7, 61.1, 56.4,
+
5
6.3, 43.3 ppm; MS (ESI) m/z in 424 [M+H] .
2-chloro-N-(2,3-dimethoxy-6-(3,4,5-trimethoxybenzoyl)phenyl)acetamide (37). The title
compound was prepared from 32. The crude material was purified by column chromatography
using PE/EtOAc 7:3 and PE/EtOAc 6:4 as eluants to give a pale yellow solid, yield 84%, m.p.
ꢀ1 1
137ꢀ139 °C. IR (KBr) 3389, 3023, 2944, 2840, 1698, 1654, 1413, 1323, 1130, 999, 765 cm . Hꢀ
NMR (300 MHz, CDCl ) δ 9.13 (brs, 1ꢀH), 7.19 (d, J = 8.4 Hz, 1ꢀH), 7.09 (s, 2ꢀH), 6.80 (d, J =
3
13
8
.7 Hz, 1ꢀH), 4.08 (s, 2ꢀH), 3.93–3.85 (m, 15ꢀH) ppm; CꢀNMR (75 MHz, CDCl ) δ 194.1,
3
164.4, 155.5, 152.9, 142.9, 142.3, 132.6, 129.3, 126.7, 126.0, 108.5, 107.9, 61.0, 60.8, 56.4, 56.1,
+
4
3.0 ppm; MS (ESI) m/z 446 [M+Na] .
2
-chloro-N-(6-(3,4,5-trimethoxybenzoyl)benzo[d][1,3]dioxol-5-yl)acetamide (38). The title
compound was prepared from 33. The crude material was purified by column chromatography
29
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