Access to 1,2,3,4-Tetraoxygenated Benzenes via a Double Baeyer-Villiger Reaction of Quinizarin Dimethyl Ether: Application to the Synthesis of Bioactive Natural Products from Antrodia camphorata

Article abstract of DOI:10.1021/acs.joc.5b02861

The first systematic investigation into the Baeyer-Villiger reaction of an anthraquinone is presented. The double Baeyer-Villiger reaction of quinizarin dimethyl ether is viable, directly providing the dibenzo[b,f][1,4]-dioxocin-6,11-dione ring-system, which is otherwise difficult to prepare. This methodology provides rapid access to 1,2,3,4-tetraoxygenated benzenes, and has been exploited by application to the total synthesis of a natural occurring benzodioxole and its biphenyl dimer, which both display noteworthy biological activity. Interestingly, the axially chiral biphenyl was found to be configurationally stable, but the resolved enantiomers exhibit no optical activity at the αD-line.

Full text of DOI:10.1021/acs.joc.5b02861

Subscriber access provided by ORTA DOGU TEKNIK UNIVERSITESI KUTUPHANESI
Article
Access to 1,2,3,4-tetraoxygenated benzenes via a double Baeyer–
Villiger reaction of quinizarin dimethyl ether: application to the
synthesis of bioactive natural products from Antrodia camphorata
Harriet L Newson, Duncan Andrew Wild, Sing Yee Yeung, Brian W.
Skelton, Gavin Ray Flematti, Jane E Allan, and Matthew John Piggott
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.5b02861 • Publication Date (Web): 22 Mar 2016
Downloaded from http://pubs.acs.org on March 29, 2016
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a free service to the research community to expedite the
dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts
appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been
fully peer reviewed, but should not be considered the official version of record. They are accessible to all
readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered
to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published
in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just
Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor
changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers
and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors
or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
The Journal of Organic Chemistry is published by the American Chemical Society.
1155 Sixteenth Street N.W., Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Access to 1,2,3,4ꢀtetraoxygenated benzenes via a double Baeyer–Villiger reaction of
quinizarin dimethyl ether: application to the synthesis of bioactive natural products from
Antrodia camphorata
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
Harriet L. Newson,† Duncan A. Wild,† Sing Yee Yeung,† Brian W. Skelton,‡ Gavin R. Flematti,†
Jane E. Allan§ and Matthew J. Piggott†*
The University of Western Australia, Perth, WA, Australia
† School of Chemistry and Biochemistry
‡ Centre for Microscopy, Characterisation and Analysis
§ School of Medicine and Pharmacology, Fiona Stanley Hospital Unit
* Corresponding author: Eꢀmail: matthew.piggott@uwa.edu.au
Dedicated to the memory of Professor Emilio Ghisalberti
Table of Contents/Abstract Graphic:
O
O
O
O
O
O
O
O
O
O
O
O
O
O
Baeyer-Villiger
O
O
O
O
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 2 of 32
1
2
3
4
ABSTRACT
5
6
7
8
The first systematic investigation into the Baeyer–Villiger reaction of an anthraquinone is
presented. The double Baeyer–Villiger reaction of quinizarin dimethyl ether is viable, directly
providing the dibenzo[b,f][1,4]ꢀdioxocinꢀ6,11ꢀdione ringꢀsystem, which is otherwise difficult to
prepare. This methodology provides rapid access to 1,2,3,4ꢀtetraoxygenated benzenes, and has
been exploited by application to the total synthesis of a natural occurring benzodioxole and its
biphenyl dimer, which both display noteworthy biological activity. Interestingly, the axially
chiral biphenyl was found to be configurationally stable, but the resolved enantiomers exhibit no
optical activity at the αDꢀline.
9
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
INTRODUCTION
The 1,2,3,4ꢀtetraoxygenated benzene moiety is abundant in biologically active natural products
spanning several chemotypes, including many food constituents valued for their antioxidant
properties. Common and representative examples include apiol (Chart 1), a principal component
of the essential oil of parsley;1 ubiquinol, the reduced form of coenzyme Q10; the cytotoxic
coumarin, isofraxidin;2 the sea urchinꢀderived, red naphthoquinone pigment, echinochrome;3 the
flavone auranetin, a constituent of orange peel;4 the bisbenzylisoquinoline alkaloid
hernandezine;5 and tannins such as the oenotheins.6
ACS Paragon Plus Environment

Page 3 of 32
The Journal of Organic Chemistry
1
2
3
4
Chart 1. Common 1,2,3,4ꢀtetraoxygenated benzeneꢀcontaining natural products.
5
6
7
8
O
OH
H
O
O
O
O
10
9
O
OH
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
apiol
ubiquinol
O
OH
O
HO
O
O
O
HO
HO
OH
OH
O
isofraxidin
echinochrome
O
O
O
N
O
O
O
O
O
O
O
O
O
O
O
N
auranetin
hernandezine
Previous syntheses of 1,2,3,4ꢀtetroxygenated benzenes (Scheme 1) include directed lithiation of
7
1,2,4ꢀtrimethoxybenzene, followed by a lowꢀyielding quench with O2 (eq 1); a directed
lithiation/borylation/oxidation sequence, as part of a total synthesis of ecteinascidin 7438 (eq 2);
a threeꢀstep biocatalytic synthesis from Dꢀglucose, involving four enzymeꢀcatalysed reactions
9
and an acidꢀcatalysed dehydration/aromatisation, applied to a synthesis of coenzyme Q3 (eq 3); a
threeꢀstep procedure from diisopropyl squarate, used in the total synthesis of enchinochrome10
(eq 4); a peroxyacetic acidꢀinduced Baeyer–Villiger oxidation of 2,6ꢀdimethoxyacetophenone
accompanied by electrophilic aromatic hydroxylation11 (eq 5); and a lowꢀyielding and
incompletely
regioselective
palladiumꢀcatalysed
C–H
acetoxylation
of
1,2,3ꢀ
trimethoxybenzene12 (eq 6). The most common approach has been a regioselective Vilsmeier–
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 4 of 32
1
2
3
4
5
6
Haack formylation or Friedel–Crafts acylation of a 1,2,3ꢀtetraalkoxybenzene, followed by
Baeyer–Villiger oxidation13 (eq 7).
7
8
9
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
Scheme 1. Examples of previous syntheses of 1,2,3,4ꢀtetraoxygenated benzenes.
O
O
O
O
1. BuLi, TMEDA
(eq 1)
2. O2
3. H2 O
HO
O
O
OMOM
OMOM
HO
O
1. BuLi
(eq 2)
2. B(OMe)3
3. H2 O2
O
O
O
OH
OH
OH
HO
HO
HO
HO
OH
HO
OH
OH
enz.
H2SO4
(eq 3)
O
HO
OH
O
OH
O
iPrO
iPrO
iPrO
iPrO
iPrO
iPrO
O
O
O
°
1. 160 C
BrMg
(eq 4)
2. MeI
KOH
OH
O
O
O
O
AcO
H2O2, TsOH
AcOH
(eq 5)
O
O
OH
O
O
O
O
O
O
Pd(cat), MesI(OAc)2
AcOH, Ac2O
(eq 6)
(eq 7)
OAc
OR
OR
RO
OR
RO
RO
RO
RO
RO
OR'
O
R'
ACS Paragon Plus Environment

Page 5 of 32
The Journal of Organic Chemistry
1
2
Building on our interest in the total synthesis of highlyꢀoxygenated benzenoid natural products,14
we now report a novel and rapid approach to the 1,2,3,4ꢀtetraoxygenated benzene nucleus,
particularly suited to symmetrical compounds, involving a double Baeyer–Villiger oxidation of
the anthraquinone, quinizarin dimethyl ether (1) (Scheme 2). The methodology has been applied
to the total synthesis of two natural products, 4 and 5 (Scheme 2), which demonstrate biological
activity worthy of further investigation.
3
4
5
6
7
8
9
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
Scheme 2. Overview of the work described herein.
O
O
O
O
O
O
O
O
O
O
1
2
O
O
O
O
O
O
O
O
O
O
O
O
O
O
O
O
5
3
4
RESULTS AND DISCUSSION
To the best of our knowledge, and somewhat surprisingly, there are only two reported examples
of Baeyer–Villiger reactions of anthraquinones. Microbial transformation of chrysazin (6) gave
the oxepine lactone 7 (Scheme 3),15 a reaction that would be very difficult to replicate through
chemical (as opposed to enzymatic) means, since oxygen insertion occurs at the least electron
rich site.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 6 of 32
1
2
3
4
5
6
7
8
9
Treatment of anthraquinone (8) with peroxytrifluoroacetic acid was reported to give a “very low
yield” of dibenzo[b,f][1,4]dioxocineꢀ6,11ꢀdione (10).16 The inefficiency of this reaction was
attributed to strain and rigidity in the sevenꢀmembered ring of the intermediate oxepine 9. The
ring system present in 10 is quite rare, with the only other reported syntheses involving very lowꢀ
yielding (3–10%) condensation of phthaloyl chlorides with catechol.16ꢀ17 Our studies suggest that
the previous low yields of 10 are, at least in part, due to its instability (see below).
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
Scheme 3. Precedents for the microbial15 and chemical19 Baeyer–Villiger oxidation of
anthraquinones.
OH
O
OH
OH
O
OH
Str eptomyces aureofaciens
O
O
O
6
7
O
O
O
O
F3CCO3H
DCM
O
O
O
O
O
8
9
10
In the Baeyer–Villiger oxidation of quinizarin dimethyl ether (1), based on migratory aptitudes,18
we expected oxygen insertion to occur adjacent to the more electronꢀrich dimethoxybenzene.
Indeed, treatment of 1 with excess mꢀchloroperbenzoic acid (mCPBA), alone or with TFA,19
gave a mixture of the oxepine 11 and dioxocin 2 (Table 1); however, the reaction stalled and
could not be pushed to favour the double Baeyer–Villiger product 2. Thus, alternative oxidants
were explored; the more promising results are summarised in Table 1. The reaction of quinizarin
dimethyl ether with hydrogen peroxide and sulfuric acid gave only very polar products, perhaps
arising from sulfonation. Urea–hydrogen peroxide complex/trifluoroacetic anhydride20 gave the
ACS Paragon Plus Environment

Page 7 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
desired dioxocin in low yield, which was improved with sodium percarbonate in trifluoroacetic
acid (TFA).21 The best yields were obtained with sodium perborate tetrahydrate.22
7
8
9
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
Although a rapid and inexpensive way to access 1,2,3,4ꢀtetraoxygenated benzenes, the Baeyer–
Villiger reaction of quinizarin dimethyl ether is somewhat capricious. Yields are very sensitive to
reaction time, presumably due to competing transesterification by trifluoroacetic acid (TFA),
and/or hydrolysis, and subsequent further oxidation of the electron rich 1,2,3,4ꢀtetraoxygenated
benzene. Attempts to minimise the decomposition of 2 by using acetic acid in place of TFA, or
DCM with a smaller excess of TFA, were unsuccessful. The reaction with sodium perborate
monohydrate was significantly slower than that with the tetrahydrate, with only a small amount
of the oxepine 8 present by TLC after 90 min. Similarly, in situ removal of the water by use of
TFA/trifluoroacetic anhydride (TFAA) completely shut the Baeyer–Villiger reaction down and
resulted in complex mixtures. These results suggest that water facilitates the reaction to some
extent. However, deliberate addition of a small amount of water (0.1 mL in 5 mL TFA) led to a
decrease in reaction rate. Ultimately, reasonable yields and purities of dioxocin 2 were obtained
by increasing the number of equivalents of oxidant, and keeping reaction times short.
It should be noted that sodium perborate tetrahydrate is very cheap (<US$100/kg), safe (used in
some teeth bleaching products) and environmentally friendly (hydrolyses to H2O2 and boric
acid), so its use in stoichiometric excess is not problematic.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 8 of 32
1
2
3
4
Table 1. Optimisation of the double Baeyer–Villiger oxidation of quinizarin dimethyl ether (2).
5
6
7
8
O
O
O
O
O
O
O
O
NMR Yield*
O
9
O
O
O
O
O
O
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
1
11
2
(isolated yield) %
Oxidant
Urea.H2O2, TFAA¥
Na2CO3.1.5 H2O2
NaBO3.4H2O
NaBO3.4H2O
NaBO3.4H2O
NaBO3.4H2O
NaBO3.4H2O
NaBO3.4H2O
NaBO3.4H2O
Eqv. (scale)# Time (min)
2
14
11
53
phthalic anhydride
8, 2.5
135
120
120
105
120
90
13
nd
12
23
5
4
3
<50
nd
18
67 (28)
trace
–
5
58
5
45
6
66
–
21
nd
nd
31
6 (2)
6 (10)
10
90
<66ǂ (37)c
<70ǂ (39)x
63 (22)x
nd
75w
30
nd
9
#All reactions were conducted in TFA on a 1 mmol scale unless indicated. Solvent was DCM;
¥
*Calculated from the mass and 1H NMR spectra of crude dioxocin 2 following aqueous workup;
ǂMaximum yield based on crude mass; After chromatography; After recrystallization from
c
x
EtOAc; wWater bath used to dissipate initial exotherm; nd = not determined.
Dioxocin 2 is unstable on silica gel, which results in significant losses upon chromatography.
The instability of 2 is presumably due to strain in the 8ꢀmembered ring, which forces the
carbonyl groups out of conjugation with the ringꢀoxygen lone pair electrons, and the adjacent
aromatic ring, as is apparent in the Xꢀray crystal structure (Figure 1). The latter was obtained
after crystallization from EtOAc, the best method of purification if pure dioxocin is required.
ACS Paragon Plus Environment

Page 9 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
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
Figure 1. Representation of the crystal structure of dioxocin 2 showing the pronounced buckling
of the central ring, which forces the carbonyl groups out of conjugation with the adjacent
aromatic ring, and presumably contributes to its hydrolytic instability. Ellipsoids in this and
subsequent figures are shown at 50% probability amplitudes with hydrogen atoms assigned
arbitrary radii.
An attempt was also made to optimize the yield of the lactone 11, and while the crude yield
could indeed be increased by reducing the number of equivalents of oxidant (up to a ratio of 4:1
1
of oxepine 11 to dioxocin 2, based on the H NMR spectrum of the crude product), purification
proved difficult; the lactone underwent partial hydrolysis during chromatography. Again, a pure
sample of 11 was obtained by crystallization, although the recovery was quite poor (28%).
The dioxocin 2 was then used in the synthesis of two natural products isolated from the Antrodia
camphorata23 (also called Antrodia cinnamomea, Taiwanofungus camphoratus, niuꢀchangꢀchih
or jangꢀjy), a fungus that grows only on the stout camphor tree, tree Cinnamomum kanehirae, in
Taiwan, where it is a traditional medicine of great value. The simple benzodioxole 4 exhibits
promising in vitro24 and in vivo25 antiꢀcolon cancer activity, while its dimer, biphenyl 5, is
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 10 of 32
1
2
3
4
5
6
reported to exhibit antiviral effects against wildꢀtype and lamivudineꢀresistant hepatitis B virus
(HBV).26
7
8
9
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
Initial attempts to cleanly hydrolyse the dioxocin 2 under basic or acidic conditions gave several
byꢀproducts in addition to low yields of the desired catechol 12 (Scheme 4). Therefore, we
attempted a oneꢀpot, twoꢀstep reaction on the predication that a mildly nucleophilic base might
cleave the rather unstable dioxocin, under conditions that would allow the catechol to be trapped
in situ. In practice this worked well, providing the known benzodioxole 314a in good yield –
when undistilled molecular sieveꢀdried DMF was used as solvent. However, the yield dropped
dramatically when freshly distilled, dry DMF was used. We reasoned that adventitious water, or
perhaps dimethylamine from partial decomposition of DMF, facilitates the cleavage of the
dioxocin. Indeed, when a little water was added to the freshly distilled, dry DMF, yields of the
dioxocin improved.
ACS Paragon Plus Environment

Page 11 of 32
The Journal of Organic Chemistry
1
2
3
4
Scheme 4. Synthesis of the anticancer natural product 4. B–V = Baeyer–Villiger, NMF = Nꢀ
5
6
methylformanilide.
7
8
9
O
O
O
O
O
O
O
HO
HO
H3O+ or HO−
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
O
2
12
85%
O
O
O
O
O
O
−
1. B V reaction
2. CH2Br2, Cs2CO3, DMF
O
O
32%
1
3
POCl3, NMF 97%
O
O
CHO
O
O
O
O
H2, Pd/C
AcOH
O
O
84%
4
13
Given the chromatographic instability of dioxocin 2, we also attempted the methylenation of the
crude product. Although this did provide the easily purified benzodioxole 3, yields were quite
poor (32% from quinizarin dimethyl ether, i.e., over two steps), suggesting that byꢀproducts were
interfering with the desired methylenation step. On one occasion, clearly when the double
Baeyer–Villiger reaction had not gone to completion, the methylenation of the crude Baeyer–
Villiger product gave the unusual spirodioxine 14 as a major product, as confirmed with an Xꢀ
ray crystal structure. A mechanism explaining its formation is shown in Scheme 5. The ring
system present in 14 is unprecedented; discovery of related acetal–lactones lacking the fused
benzenes was recently reported,27 with some derivatives shown to have potent mammalian
cytoxicity.28
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 12 of 32
1
2
3
4
5
6
7
Scheme 5. Representation of the crystal structure of 14 and proposed mechanism leading to its
formation.
8
9
O
O
O
O
O
O
O
O
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
O
O
HO
O
HO
O
O
HO
O
11
CH2Br2, Cs2 CO3
O
O
O
O
O
O
14
The synthesis of the anticancer natural product 4 (Scheme 4) was completed by Vilsmeier–
Haack formylation of 3 to give the benzaldehyde 13,29 followed by reductive deoxygenation.
Interestingly, the first attempted hydrogenation/hydrogenolysis produced a small amount of the
decarbonylation product 3. There is precedent for decarbonylation of benzaldehydes with Pd/C,
but only at high temperatures.30 On the assumption that the decarbonylation requires ‘naked’
metal sites, the reaction was repeated with Pd/C that was preꢀcharged with H2, and indeed the
decarbonylation product was not observed. Synthetic 4 was isolated as a colourless, amorphous
solid, while the natural product was reported to be a yellow liquid; however, the NMR spectra
for synthetic 4 match the data reported for the natural product.
During the course of our work a synthesis of 4 from 2,3,4,5ꢀteramethoxytoluene was reported.31
ACS Paragon Plus Environment

Page 13 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
Our attention then turned to the dimeric antiviral biphenyl 5 (Scheme 6). A single attempt at a
PIFA [phenyliodine bis(trifluoroacetate)]ꢀmediated oxidative coupling32 of 4 gave a complex
7
8
9
1
mixture of products, perhaps containing a trace of the desired biphenyl as judged from the 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
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
NMR spectrum of the crude material. Therefore, a more conventional approach was investigated.
Specifically, we were attracted by the capacity of the Ullmann coupling to deliver sterically
congested biaryls.33
Bromination of 4 proceeded smoothly, but the Ullmann coupling of 15 under standard conditions
failed (Scheme 6). Similar reactions of 2ꢀhaloꢀ3ꢀalkylanisoles and related compounds are
known,34 but the yields are variable and very high temperatures are often required. Copperꢀ
mediated Ullmann biaryl coupling is favoured by coordinating substituents ortho to the
halogen,33 and there are several examples of Ullmann couplings of 2ꢀhaloꢀ3ꢀ
methoxybenzaldehyde,35 so we chose to explore the reaction of bromobenzaldehyde 16.
Bromination of 13 with Br2 was very sluggish, and did not go to completion, even with excess
Br2 and heating. In contrast, NBS in TFA36 at room temperature led to rapid bromination, but
also extensive formation of the dibromide 17.37 Fortunately, the bromodecarbonylation was
minimal at –10°C, and 17 was easily separated from the bromobenzaldehyde 16, which was
isolated in excellent yield.
As predicted, the Ullmann coupling of 16 gave an excellent yield of bi(benzaldehyde) 18,
accompanied by a small amount (9%) of the dehalogenation product 13. In contrast to the
hydrogenation/hydrogenolysis of 13, the analogous deoxygenation of 18 was not a clean
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 14 of 32
1
2
3
4
5
6
7
8
9
reaction. Fortunately, deoxygenation with acidified triethylsilane was rapid and efficient,
providing an almost quantitative yield of pure 5, after a simple aqueous workup. The NMR
spectra for synthetic 5 matched the data reported for the natural product.23
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
Scheme 6. Synthesis of the biphenyl natural product 5.
O
O
O
O
O
O
Br2
DCM
Br
O
O
quant.
4
15
Cu
°
DMF, 160 C
trace
O
O
CHO
O
O
O
O
O
O
Et3SiH
TFA
O
O
O
O
O
OHC
O
97%
O
O
18
5
Cu
°
85%
DMF, 160 C
O
O
CHO
Br
CHO
O
O
O
NBS
−
°
TFA, 10 C
O
O
O
85%
16
13
O
Br
Br
O
O
O
17
The natural product 5 was isolated as a racemate, as supported by the lack of optical activity, and
confirmed by the centrosymmetric space group of the reported crystal structure.23 We were
interested to ascertain whether this is due to racemic biosynthesis or because the atropisomers are
configurationally unstable. Accordingly, the synthetic racemate 5 was subjected to semiꢀ
ACS Paragon Plus Environment

Page 15 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
preparative normalꢀphase enantioselective chromatography, affording the two atropisomers in
greater than 96% e.r. An Xꢀray crystal structure of the less mobile enantiomer was obtained,
which showed it to be the Sꢀatropisomer Sꢀ5 (Figure 2). To our surprise, the atropisomers exhibit
no optical activity at the αꢀD line in CHCl3 solution. They do, however, give rise to equal and
opposite circular dichroism spectra (Figure 3).
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
Figure 2. Representation of the crystal structure of the S atropisomer of 5.
Figure 3. Circular dichroism spectra of Rꢀ5 and Sꢀ5.
The configurational stability of the atropisomers was assessed by heating a solution of the Rꢀ
enantiomer in refluxing toluene and monitoring racemisation by analytical enantioselective
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 16 of 32
1
2
3
4
5
6
chromatography over several days. Although some degradation was observed during this period,
none of the Sꢀatropisomer was detected (see supporting information).
7
8
9
To gain an estimate of the energy barrier to rotation about the biaryl bond, a conformational
analysis was undertaken. Using density functional theory, the geometry of the complex was fully
optimised and subsequently determined to be a minimum via vibrational frequency analysis.
Following on, a relaxed potential energy scan was undertaken along the biaryl dihedral angle
coordinate, with the results represented as the torsional potential in Figure 6. All calculations
were undertaken at B3LYP/6ꢀ31G* using the Gaussian 09 program suite.38
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
As depicted in Figure 4, the lowest energy pathway, in which the methyls pass by the methoxy
groups, involves a highlyꢀstrained conformation that lies ~250 kJ.mol–1 above the ground state
energy minimum (biaryl dihedral = 80°). Of course, these calculations only give approximate
energies for the barriers to rotation as they do not consider the influence of vibrational excitation
of specific modes, which could facilitate passage over the barrier. Although the structure of the
transition state was not determined, the calculated barriers shown in Figure 4 are on par with
those reported in similar work, wherein the halfꢀlife was estimated to be greater than 10 years.39
ACS Paragon Plus Environment

Page 17 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
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
Figure 4. Torsional potential energy scan about the biaryl dihedral angle, produced at the
B3LYP/6ꢀ31G* level of theory.The structures shown correspond to the tip of the first peak in the
plot. The orientation on the left emphasises the extreme distortion from coplanarity of the biaryl
bond/benzene, which results from steric repulsion between the methyl and methoxy groups.
The results above suggest that the biosynthesis of 5 is, indeed, racemic. It seems likely that
oxidative coupling of a phenolic precursor gives a biphenol, which is not configurationally stable
about the biaryl axis, and subsequent Oꢀmethylation gives racꢀ5.
AntiꢀHCV activity
The mode of action of biaryl natural product 5 against the DNA encoded wildꢀtype, and
lamuvidineꢀresistant, HBV is not known, but it may be a polymerase inhibitor. Some drugs
inhibit both reverse transcriptase and DNA polymerases. For example, lamivudine is approved
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 18 of 32
1
2
for the treatment of HIV and chronic hepatitis B.40 Alternatively, 5 may work through some
hepatocyteꢀspecific pathways. Therefore, we were interested to see if the in vitro activity
reported for natural product 5 against HBV extended to the RNA encoded hepatitis C virus
(HCV), which also has tropism for hepatocytes.
3
4
5
6
7
8
9
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
Attempts were made to evaluate the natural product 5 and the dialdehyde precursor 18 in the
human hepatoma (Huh7ꢀJ20) cell line, which secretes alkaline phosphatase upon HCV
infection.41 Concomitantly, cell viability (cytotoxicity) was assessed by standard MTT assay.
However, the natural product 5 was found to have poor aqueous solubility. At low concentrations
5 was inactive against HCV replication, and the concentrations of DMSO necessary to achieve
higher concentrations of 5 (comparable to those reported previously for HBV assays26) were
cytotoxic. The aldehyde 18 was more waterꢀsoluble and had some antiviral activity, with an
approximate EC50 of 1.7 µM; however, at 10 µM 18 was also cytotoxic, complicating the EC50
determination. Thus, at this time, the potential of 5 as an antiviral lead is not particularly
encouraging. It remains to be seen if similar activity can be demonstrated in more polar/water
soluble analogues that are less cytotoxic than dialdehyde 18.
CONCLUSION
The first systematic investigation into the Baeyer–Villiger reaction of an anthraquinone has been
presented. The double Baeyer–Villiger reaction of quinizarin dimethyl ether with the inexpensive
and environmentally benign oxidant sodium perborate tetrahydrate is viable, and provides rapid
access to symmetrical 1,2,3,4ꢀtetraoxygenated benzenes. This work has been applied to the first
ACS Paragon Plus Environment

Page 19 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
total synthesis of the antiviral biphenyl natural product 5, which was prepared in five steps and
27% overall yield (3 steps and 70% overall yield from known aldehyde 13).
7
8
9
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
EXPERIMENTAL SECTION
Materials and Methods
All solvents were distilled prior to use. Anhydrous DMF was obtained by drying over activated
3A molecular sieves for 24 h, followed by distillation under reduced pressure onto activated 3A
sieves. ‘Hexanes’ refers to the hydrocarbon fraction distilling from 64–67 °C. All other reagents
and materials were purchased from commercial suppliers and used as received.
Temperatures reported for heated reactions refer to bath temperatures unless the reactions were
heated under reflux. Organic extracts were dried over anhydrous MgSO4. Solvents were
evaporated under reduced pressure at approximately 45°C, and then traces of solvent were
removed under a flow of nitrogen.
Reaction progress was monitored by analytical thin layer chromatography (TLC) using Merck
aluminiumꢀbacked TLCF254 plates. Spots were visualised with a UV lamp (254 nm) and/or by
staining with acidified ceric sulfate, or dinitrophenylhydrazine (DNP). Chromatography was
performed with either Merck, Silicycle or Davisil silica gel (average particle size: 40–63 ꢁm;
average pore size: 60 Å). RSF = rapid silica filtration.42
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 20 of 32
1
2
1H and 13C NMR spectra were obtained using 300 , 400 MHz, 500 or 600 MHz spectrometers, as
indicated. Deuterochloroform (CDCl3) was used as the solvent, with residual CHCl3 (1H, δ =
7.26 ppm) or CDCl3 (13C, δ = 77.16 ppm) being used for calibration.
3
4
5
6
7
8
9
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
Infrared (IR) spectra were acquired using an ATR attachment. Mass spectra were recorded using
fast atom bombardment (FAB+) or electron impact ionisation (EI) (both with magnetic sector
mass analyser), or electrospray ionization (ESI+) and quadrupole mass analyser, as indicated.
CD spectra were recorded on acetonitrile solutions (1 mg.mL–1). Melting points were determined
on a hot stage melting point apparatus.
Synthesis
1,4ꢀDimethoxyanthraquinone (quinizarin dimethyl ether) (1). A mixture of quinizarin (12.01
g, 50.00 mmol), anhydrous K2CO3 (27.64 g, 200.0 mmol), methyl iodide (7.5 mL, 120 mmol)
and DMF (50 mL) was sealed with a septum and stirred at 40°C overnight. TLC after this time
indicated that the reaction was incomplete, so additional methyl iodide (3.75 mL, 60 mmol) was
added and the sealed reaction mixture was stirred for a further 24 h at 40°C. The reaction mixture
was diluted with water (500 mL) and 1 M NaOH (100 mL) and extracted with DCM (4 × 100
mL). The extract was washed with 0.2 M NaOH (4 × 100 mL) and the washes were backꢀ
extracted with DCM (3 × 100 mL). The combined organic phase was gravity filtered to remove
insoluble impurities, then washed with 0.2 M NaOH (4 × 200 mL), brine (200 mL), dried and
evaporated to give 1 as an orange solid (12.72 g, 95%), mp 170–174°C [lit.43 170.5–171.5°C]. Rf
1
= 0.15 (1:1 EtOAc/hexanes); H NMR (500 MHz):
δ 8.17 (m [AA' part of AA'XX'], 2H), 7.71
ACS Paragon Plus Environment

Page 21 of 32
The Journal of Organic Chemistry
1
2
1
3
4
(m [XX' part of AA'XX'], 2H), 7.35 (s, 2H), 4.00 (s, 6H). The H NMR data were identical to
5
6
those reported.44
7
8
9
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
1,4ꢀDimethoxydibenzo[b,f][1,4]ꢀdioxocinꢀ6,11ꢀdione (2). Sodium perborate tetrahydrate (9.23
g, 60.0 mmol) was added to a stirred solution of quinizarin dimethyl ether (1) (2.683 g, 10.00
mmol) in TFA (50 mL) under N2, in a water bath to dissipate the heat generated. After 75 min
the reaction mixture was poured onto iceꢀwater (300 mL) and extracted with DCM (3 × 100 mL).
The extract was washed with water (100 mL) and brine (100 mL), dried and evaporated to give
crude 2 as an orange solid (2.100 g), which crystallized from EtOAc as colourless needles (1.178
1
g, 39%), mp 211–215°C. Rf = 0.25 (1:1 EtOAc/hexanes); IR ν (cm–1): 1761 (C=O); H NMR
(300 MHz):
166.7, 145.7, 135.5, 135.5, 132.4, 127.5, 109.7, 56.7. MS (EI): m/z 300 (M•+, 95 %), 229 (51),
δ
7.52 (AA'BB' [apparent s], 4H); 6.65 (s, 2H); 3.79 (s, 6H); 13C NMR (75.5 MHz):
δ
104 (100), 76 (64), 69 (31). C16H12O6 requires C, 64.0; H, 4.0; found: C, 64.2; H, 4.1 %.
1,4ꢀDimethoxydibenzo[b,e]oxepineꢀ6,11ꢀdione (11). Sodium perborate tetrahydrate (2.311 g,
15.02 mmol) was added to a stirred solution of 1 (1.341 g, 4.999 mmol) in TFA (20 mL) at 0°C
under argon. After 90 min the ice bath was removed and stirring was continued for 30 min, then
poured onto iceꢀwater (200 mL) and extracted with DCM (4 × 50 mL). The extract was washed
with water (2 × 50 mL), and brine (50 mL), dried and evaporated to give a brown solid (1.312 g),
which was subjected to flash chromatography. Elution with 1:5 EtOAc/hexanes gave 11 as a
1
yellow solid (1.110 g, 73%, containing 5% dioxocin 2 based on the H NMR spectrum;
Percentages are yields, not proportions). The pure oxepine 11 crystallized from MeOH as pale
yellow needles (0.394 g, 28%), mp 162–166°C. Rf = 0.35 (1:1 EtOAc/hexanes); IR (ATR) ν (cm–
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 22 of 32
1
2
1
1): 1736 (OC=O), 1691 (C=O). H NMR (500 MHz): δ 8.17 (ddd, J = 7.7, 1.4, 0.4 Hz, 1H),
3
4
5
6
7.65–7.76 (m, 3H), 7.01 (d, J = 9.0 Hz, 1H), 6.77 (d, J = 9.0 Hz, 1H), 3.90 (s, 3H), 3.83 (s, 3H).
7
8
9
13C NMR (500 MHz): δ 191.2, 163.1, 150.1, 145.0, 142.6, 139.3, 134.5, 133.3, 132.5, 127.4,
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
+
124.8, 124.1, 116.1, 109.5, 57.0, 56.9. MS (ESI+): m/z observed: 285.0748 ([M+H+]; C16H13O5
requires 285.0763.
4,7ꢀDimethoxyꢀ1,3ꢀbenzodioxole (3). Method A: A mixture of dibromomethane (0.60 mL, 8.1
mmol), 2 (1.201 g, 4.000 mmol), Cs2CO3 (5.22 g, 16 mmol) and 3A molcular sieveꢀdried (but
not distilled) DMF (40 mL) was stirred at 80°C under N2 overnight. The reaction mixture was
allowed to cool then diluted with 0.1 M NaOH (400 mL) and exttracted with DCM (3 × 100
mL). The extract was washed with water (2 × 100 mL) and brine (100 mL), dried and evaporated
to give 3 as a pale yellow crystalline solid (618 mg, 85%), mp 77–79°C [lit.29 76.5–77.5°C]. H
1
1
NMR (300 MHz)
those reported.14a
δ
6.45 (s, 2H); 5.99 (s, 2H); 3.86 (s, 6H). The H NMR data are identical to
Method B: Sodium perborate tetrahydrate (7.69 g, 50.0 mmol) was added to a stirred solution of
quinizarin dimethyl ether (1.344 g, 5.010 mmol) in TFA (25 mL) under argon, in a water bath to
dissipate any heat generated. After 45 min the reaction mixture was poured into iceꢀwater (350
mL) and extracted with DCM (3 × 150 mL). The extract was washed with water (100 mL) and
brine (100 mL), dried and evaporated to give crude an orange solid (1.064 g). The crude
dioxocin was dissolved in in DMF (20 mL), treated with dibromomethane (0.60 mL, 8.1 mmol)
and Cs2CO3 (3.58 g, 11.0 mmol), and stirred at 80°C under argon for 22 h. Water (5 drops) was
added the reaction mixture was stirred at 80ºC for a further 2 h, then cooled, diluted with 0.1 M
ACS Paragon Plus Environment

Page 23 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
NaOH (250 mL) and extracted with Et2O (4 × 100 mL). The extract was washed with water (2 ×
100 mL) and brine (100 mL), dried and evaporated to give the an orange solid (0.558 g), which
was subjected to flash chromatography. Elution with 1:9 EtOAc/hexanes gave 3 as white needles
(0.294 g, 32% over 2 steps), identical with the material described above.
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
On one occasion, the attempted methylenation of a crude mixture of 2 and 8 (0.738 g), as
described above, gave, after flash chromatography, eluting with 1:5 EtOAc/hexanes, 5,8ꢀ
dimethoxyꢀ3'Hꢀspiro[benzo[d][1,3]dioxineꢀ4,1'ꢀisobenzofuran]ꢀ3'ꢀone (14) as a white solid
(0.377 g), which crystallized from DCM as small white rhomboids, mp 188–198; 200–202°C
(there were two crystal types). Rf = 0.50 (1:1 EtOAc/hexanes); IR (ATR) ν (cm–1): 1771 (C=O).
1H NMR (500 MHz) = δ 7.91 (pseudo d, 1H), 7.60 (ddd [app. dt], J1 = J2 = 7.5, J3 = 1.5 Hz, 1H),
7.56 (ddd [app. dt], J1 = J2 = 7.5, J3 = 1.5 Hz, 1H), 7.27 (pseudo d, 1H), 6.88 (d, J = 9.0 Hz, 1H),
6.33 (d, J = 9.0 Hz, 1H), 5.52 (d, J = 5.5 Hz, 1H), 5.49 (d, J = 5.5 Hz, 1H), 3.89 (s, 3H), 3.26 (s,
13
3H). C NMR (500 MHz) = δ 168.3, 150.8, 148.3, 144.3, 142.3, 134.1, 130.3, 128.1, 124.8,
121.9, 113.1, 109.7, 103.6, 101.3, 87.9, 56.6, 55.7. MS (ESI+): m/z observed: 315.0855
+
([M+H+]; C17H15O6 requires 315.0869.
2,5ꢀDimethoxyꢀ3,4ꢀmethylendioxybenzaldehyde (13).29 POCl3 (1.4 mL, 15 mmol) was added
slowly to a stirred mixture of 3 (1.822 g, 10.00 mmol) and Nꢀmethylformanilide (1.49 g, 11.0
mmol) at 0°C. The suspension was stirred at room temperature for 3 h, then allowed to stand
overnight, before being partitioned between EtOAc (50 mL) and cold, saturated NaHCO3 (50
mL). The phases were separated and the aqueous phase was extracted with EtOAc (2 × 20 mL).
The combined organic phase was washed with water (30 mL) and brine (30 mL), dried and
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 24 of 32
1
2
3
4
evaporated to give 13 as a crystalline tan solid (2.04 g, 97%), which crystallized from hexanes to
5
6
1
give 13 as tan prisms, mp 99–101°C [lit.29 101.5–102.5°C]. Rf = 0.65 (1:1 EtOAc/hexanes); H
7
8
9
NMR (500 MHz): δ 10.23 (s, 1H), 7.08 (s, 1H), 6.09 (s, 2H), 4.05 (s, 3H), 3.89 (s, 3H). The
NMR data are slightly different from those reported.45
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
4,7ꢀDimethoxyꢀ5ꢀmethylꢀ1,3ꢀbenzodioxole (4). A degassed solution of 13 (211 mg, 1.00
mmol) in AcOH (5 mL) was added to a stirred suspension of 10% Pd/C (0.21 g) in AcOH (5 mL)
under a balloon of H2. After 24 h, TLC showed the reaction to be complete. The suspension was
diluted with Et2O (50 mL), vacuumꢀfiltered through a pad of Celite, and washed through with
Et2O (50 mL). The filtrate was washed with saturated NaHCO3 (4 × 30 mL) [FOAMING], water
(30 mL), and brine (30 mL), dried and evaporated to give pink oil, which was subjected to RSF.
Elution with 1:99 EtOAc/hexanes gave 4 as a colourless oil that solidified on standing (165 mg,
84%), which precipitated from hexanes as an amorphous solid, mp = 39–40°C (lit.23 yellow
liquid). Rf = 0.35 (1:9 EtOAc/hexanes); 1H NMR (400 MHz):
δ
6.30 (br q, J = 0.7 Hz, 1H), 5.93
13
(s, 2H,), 3.87 (s, 3H), 3.84 (s, 3H), 2.17 (d, J = 0.6 Hz, 3H); C NMR (100 MHz):
δ 138.8,
138.6, 136.5, 134.6, 123.6, 108.8, 101.4, 59.8, 56.8, 15.8. The NMR data are identical to those
reported,23 except for the fine benzylic coupling in the 1H NMR spectrum.
5ꢀBromoꢀ4,7ꢀdimethoxyꢀ6ꢀmethylꢀ1,3ꢀbenzodioxole (15). A solution of Br2 (25 ꢁL, 0.49
mmol) in DCM (1 mL) was added dropwise to a stirred solution of 4 (87 mg, 0.44 mmol) in
DCM (2 mL) at 0°C. Stirring was continued for 15 min then the resulting solution was poured
into 1 M sodium thiosulfate (15 mL). The mixture was extracted with Et2O (2 × 15 mL) and the
extract was washed with water (10 mL) and brine (10 mL), dried and evaporated to give 15 as a
ACS Paragon Plus Environment

Page 25 of 32
The Journal of Organic Chemistry
1
2
3
4
crystalline white solid (122 mg, quant.), which crystallized from hexanes as white needles. Rf =
5
6
7
1
0.35 (1:9 EtOAc/hexanes); H NMR (400 MHz):
δ 5.94 (s, 2H), 3.93 (s, 3H), 3.88 (s, 3H), 2.25
13
8
9
(s, 3H); C NMR (100 MHz):
δ 138.3 , 137.5 , 137.2 , 136.3 , 124.3, 110.2, 101.6, 60.3, 60.1,
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
16.0; MS (FAB+): m/z 276 [M(81Br)+H]+ (97%), 274 [M(79Br)+H]+ (100%), 196 [M–Br+H]+,
(16%). C10H11BrO4 requires: C, 43.7; H, 4.0; found: C, 43.5; H, 4.0 %.
6ꢀBromoꢀ2,5ꢀdimethyoxyꢀ3,4ꢀmethylendioxybenzaldehyde (16). NBS (0.43 g, 2.4 mmol) was
added portionwise to a stirred solution of 13 (420 mg, 2.00 mmol) in TFA (10 mL) at –10°C
(ice/MeOH with occasional liquid N2). Stirring was continued for 90 min, after which time an 1H
NMR spectrum of a sample showed the reaction to be complete. The reaction mixture was
diluted with ice/water (200 mL) and basified with solid NaHCO3, whereupon a precipitate
formed. The suspension was extracted with Et2O (4 × 50 mL). The extract was washed with 1 M
sodium thiosulfate (2 × 50 mL), water (50 mL) and brine (50 mL), dried and evaporated to give a
crystalline, pale yellow solid, which was subjected to RSF. Elution with 1:9 EtOAc/hexanes gave
1
5,6ꢀdibromoꢀ4,7ꢀdimethoxyꢀ1,3ꢀbenzodioxole (17) as a colourless solid (51 mg, 8%). H NMR
13
(300 MHz): 6.01 (s, 2H), 3.94 (s, 6H); C NMR (75 MHz): 139.0 , 137.4 , 111.5, 102.3 , 60.5 .
The 1H NMR data are similar to those reported.37a
Further elution with 1:4 EtOAc/hexanes gave 16 as a crystalline yellow solid (489 mg, 85%),
which crystallized from DCM/hexanes as pale yellow needles. Rf = 0.20 (1:5 (EtOAc/hexanes);
IR ν (cm–1): 1682 (C=O); 1H NMR (300 MHz):
3H); 13C NMR (75 MHz):
δ 10.20 (s, 1H, 6.07 (s, 2H), 3.97 (s, 3H), 3.91 (s,
δ
189.4, 144.0 , 141.5 , 138.4 , 136.6 , 121.1, 112.6, 102.8 , 61.0 , 60.5
; MS (FAB+): m/z 289 [M(79Br)+H]+ (100%), 291 [M(81Br)+H]+ (88%). C10H9BrO5 requires: C,
41.6; H, 3.1; found: C, 41.4; H, 2.9. The 1H NMR data are similar to those reported.37a
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 26 of 32
1
2
3
4
5
6
7
8
Biꢀ(2,5ꢀdimethoxyꢀ3,4ꢀmethylenedioxybenzaldehyde) (18). A degassed suspension of
activated46 copper bronze (0.29 g, 4.6 mmol) in a solution of 16 (289 mg, 1.00 mmol) in dry
DMF (1 mL) was stirred at 100°C under a positive pressure of argon for 1.5 d. The reaction
mixture was allowed to cool, then diluted with water (100 mL) and extracted with EtOAc (3 × 30
mL). The extract was filtered, washed with water (3 × 30 mL) and brine (30 mL), dried and
evaporated to give a yellow solid (200 mg), which was subjected to RSF. Elution with 1:4
EtOAc/hexanes gave 13 (17 mg, 9%). Further elution with 1:1 EtOAc/hexanes gave 18 as a
9
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
yellow solid (177 mg, 85%), which crystallized from DCM/hexanes as yellow rhomboids, mp
1
185–188°C. Rf = 0.34 (1:1 EtOAc/hexanes); IR ν (cm–1): 1681 (C=O); H NMR (500 MHz):
δ
13
10.02 (s, 2H), 6.10 (AA' [app. q, ‘J’ = 1.5 Hz], 4H), 4.06 (s, 6H), 3.69 (s, 3H); C NMR (75
MHz):
δ 188.8, 143.9 , 142.3 , 137.9 , 136.7 , 125.9, 121.0, 102.3 , 60.7 , 60.0 ; MS (FAB+): m/z
419 ([M+H]+, 100%), 389 (36%), 373 (64%). C20H18O10 requires: C, 57.4; H, 4.3; O, 38.2;
found: C, 57.6; H, 4.1.
Biꢀ(2,5ꢀdimethoxyꢀ6ꢀmethylꢀ3,4ꢀmethylenedioxybenzene) (5). Triethylsilane (0.31 mL, 2.0
mmol) was added dropwise to a stirred solution of 18 (21 mg, 0.050 mmol) in TFA (0.5 mL) at
0°C, whereupon the bright orange solution immediately turned colourless. After 30 min the
reaction mixture was poured onto ice and diluted with saturated NaHCO3 (80 mL), then
extracted with Et2O (3 × 20 mL). The extract was washed with brine, dried and evaporated to
give 5 as a colourless oil that solidified on standing (19 mg, 97%), which crystallized from
MeOH as colourless rhomboids. Rf = 0.35 (1:4 EtOAc/hexanes); 1H NMR (500 MHz):
δ
5.95 (s,
4H), 3.92 (s, 6H, 3.73 (s, 3H, 1.78 (s, 6H); 13C NMR (150 MHz):
δ 138.0 , 137.4 , 137.0 , 136.6 ,
ACS Paragon Plus Environment

Page 27 of 32
The Journal of Organic Chemistry
1
2
123.3*, 101.2 , 60.04 , 60.00 , 12.8 . The NMR data were identical with those reported.23 *Nonꢀ
identical carbons are isochronous.
3
4
5
6
7
8
9
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
Enantioselective chromatography
Racemic 5 was resolved into its individual atropisomers using semiꢀpreparative HPLC
performed on a Hewlettꢀ Packard 1050 system fitted with an Astec Cellulose DMP (3,5ꢀ
dimethylphenyl carbamateꢀderivatized) HPLC column (250 mm long, 10 mm i.d., 5 ꢁm particle
size, Supelco, Bellafonte, PA, USA). Separation was achieved using a flow rate of 2 mL/min
with 5% isopropanol/hexanes. Under these conditions the R enantiomer eluted first. Analytical
enantioselective HPLC was conducted using an Astec Cellulose DMP HPLC column (250 mm
long, 4.6 mm i.d., 5 ꢁm particle size, Supelco, Bellafonte, PA, USA). Separation was achieved
using a flowrate of 0.5 mL/min with 5% isopropanol/hexanes.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 28 of 32
1
2
3
4
5
6
7
8
9
Kinetic stability of Rꢀbi(2,5ꢀdimethoxyꢀ6ꢀmethylꢀ3,4ꢀmethylenedioxybenzene) (Rꢀ5).
A solution of the single atropisomer Rꢀ5 (0.5 mg) in toluene (5 mL) was heated under reflux
under argon. Samples were taken at regular intervals over the course of 3 d and analysed by
analytical enantioselective HPLC as described above.
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
Crystallography
Crystallographic data were acquired at 100(2) K Cu Kα (for Sꢀ5) or Mo Kα (for 2, 14) radiation.
Following multiꢀscan absorption corrections and solution by direct methods, the structures were
refined against F2 with fullꢀmatrix leastꢀsquares using the program SHELXLꢀ97.47 All hydrogen
atoms were added at calculated positions and refined by use of riding models with isotropic
displacement parameters based on those of the parent atoms. Anisotropic displacement
parameters were employed throughout for the nonꢀhydrogen atoms.
Biological assays
Cell culture. The human hepatoma reporter cell line, Huh7ꢀJ20,58 kindly provided by Dr A.
Patel (University of Glasgow), was grown in RPMI supplemented with 10% foetal bovine serum
(FBS) with gentamicin (0.16 mg/mL) and puromycin (8 ꢁg/mL) at 37°C with 5% CO2 in a
humidified atmosphere. Monolayers were dispersed with trypsin–EDTA to provide single cell
suspensions for assays.
ACS Paragon Plus Environment

Page 29 of 32
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Addition of test compounds to tissue culture. Culture medium (100 ꢁL/well) was removed
from 96 well plates and replaced with the test compounds diluted in RPMI/10% FBS to give
triplicate wells with the final concentrations: 0.08, 0.16, 0.31, 0.63, 1.30, 2.50, 5.00 µM for
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
natural product 5; and 0.08, 0.16, 0.25, 0.31, 0.50, 0.63, 0.75, 1.00, 1.25, 1.40, 1.90, 2.50, 3.80,
5.00 µM for 18 and 0.44 % (v/v) DMSO. Negative control cultures consisted of medium with
0.44 % (v/v) DMSO (final concentration).
MTT Assay for cell viability. Cells were incubated overnight at 6 × 104 cells/well in 96 well
plates and compounds 5 or 18 were added as described above, and incubated for 3 d. MTT dye
(0.5 mg/mL, 20 ꢁL/well) was added and after 4 h, 100 µL/well media was replaced with lysis
buffer consisting of 20% sodium dodecyl sulphate in 50% aqueous DMF. Absorbance was read
at 540 nm.
Testing antiꢀHCV activity. Cells to be infected were dispensed at 100 µL/well at 6 × 103
cells/well into 96ꢀwell plates 5 h prior to infection. HCV strain JFHꢀ1, originally kindly provided
by Dr M. Watson (Institute for Immunology and Infectious Diseases, Murdoch University), was
added in 100 µL per well and the plates incubated at 37°C for at least 2 h prior to addition of test
compounds. IFNαꢀ2b (10 units/100 ꢁL) was used as an antiviral compound of known activity.
The EC50 was calculated using GraphPad Prism 6 software with log concentrations and a 3
parameter dose response curve.
ACS Paragon Plus Environment