Total synthesis of robustaflavone, a potential anti-hepatitis B agent

Article abstract of DOI:10.1021/jo981186b

Robustaflavone, a naturally occurring compound, is an inhibitor of hepatitis B virus replication in vitro. Robustaflavone is a biflavanoid composed of two units of apigenin (5,7,4'-trihydroxyflavone) joined via a biaryl linkage between the 6-position of one unit and the 3'-position of the other (I6,II3'-biapigenin). The natural material was isolated from the seed- kernels of Rhus succedanea. To provide ready access to sufficient quantities of material for continued biological studies, as well as to provide a general route for the preparation of structural analogues, a total synthesis of robustaflavone was pursued. The total synthesis was approached by constructing apigenin ethers containing functionalities at the 6- and 3'- positions which could be cross-coupled using transition metal catalysis. Key steps of the synthesis included development of a regioselective iodination of an apigenin derivative at the 6-position. Also key was the formation of an apigenin 3'-boronate using a palladium-catalyzed exchange of the corresponding 3'-iodide with a diboron reagent. Finally, identification of appropriate reaction conditions for Suzuki coupling to form the sterically congested 6-3''' biaryl bond of robustaflavone provided access to the desired biflavanoid system. This work represents the first total synthesis of robustaflavone.

Full text of DOI:10.1021/jo981186b

9300
J . Org. Chem. 1998, 63, 9300-9305
Tota l Syn th esis of Robu sta fla von e, a P oten tia l An ti-Hep a titis B
Agen t
David E. Zembower* and Heping Zhang
MediChem Research, Inc., 12305 South New Avenue, Lemont, Illinois, 60439
Received J une 18, 1998
Robustaflavone, a naturally occurring compound, is an inhibitor of hepatitis B virus replication in
vitro. Robustaflavone is a biflavanoid composed of two units of apigenin (5,7,4′-trihydroxyflavone)
joined via a biaryl linkage between the 6-position of one unit and the 3′-position of the other (I6,II3′-
biapigenin). The natural material was isolated from the seed-kernels of Rhus succedanea. To provide
ready access to sufficient quantities of material for continued biological studies, as well as to provide
a general route for the preparation of structural analogues, a total synthesis of robustaflavone
was pursued. The total synthesis was approached by constructing apigenin ethers containing
functionalities at the 6- and 3′-positions which could be cross-coupled using transition metal
catalysis. Key steps of the synthesis included development of a regioselective iodination of an
apigenin derivative at the 6-position. Also key was the formation of an apigenin 3′-boronate using
a palladium-catalyzed exchange of the corresponding 3′-iodide with a diboron reagent. Finally,
identification of appropriate reaction conditions for Suzuki coupling to form the sterically congested
6-3′′′ biaryl bond of robustaflavone provided access to the desired biflavanoid system. This work
represents the first total synthesis of robustaflavone.
In tr od u ction
We recently reported that robustaflavone, a naturally
occurring biflavanoid, is a potent nonnucleoside inhibitor
of hepatitis B virus (HBV) replication.1 HBV is one of
the most serious health problems in the world today and
is listed as the ninth leading cause of death by the World
Health Organization.2 Approximately 300 million persons
are chronically infected with HBV worldwide, with over
one million of those in the United States. The Centers
for Disease Control estimates that over 300000 new cases
of acute HBV infection occurs in the United States each
year, resulting in 4000 deaths due to cirrhosis and 1000
due to hepatocellular carcinoma.3 The highest incidence
of HBV infection occurs in the Far East and sub-Saharan
Africa, where approximately 20% of the population are
chronically infected.4 Infection can be prevented through
the use of several extremely effective recombinant vac-
cines.5 Despite the availability of these vaccines, HBV
infection remains the most significant viral pathogen
infecting man, particularly in underdeveloped countries.
succedanea. Robustaflavone is a biflavanoid composed of
two apigenin (5,7,4′-trihydroxyflavone) units connected
via a biaryl linkage between their 6- and 3′-positions. No
synthesis of robustaflavone has been reported. A recent
synthesis of a related biflavanoid, amentoflavone (I8,II3′-
biapigenin), which consists of two apigenin units con-
nected via a biaryl linkage between respective 8- and 3′-
positions, was achieved using Suzuki coupling8 of an
apigenin 8-boronic acid derivative with an appropriate
3′-iodoapigenin analogue.9 We adopted this general meth-
odology in our approach to the total synthesis of robust-
aflavone.
Robustaflavone was first6 isolated in 1973, as its hexa-
O-methyl ether, from leaf extracts of Agathis robusta, and
later7 in larger quantities from the seed-kernels of Rhus
* To whom correspondence should be addressed. Telephone:
(630)-257-1500. FAX:
mcr.medichem.com.
(630)-257-1505. E-mail:
dzembower@
(1) Lin, Y.-M.; Zembower, D. E.; Flavin, M. T.; Schure, R. M.;
Anderson, H. M.; Korba, B. E.; Chen, F.-C. Bioorg. Med. Chem. Lett.
1997, 7, 2325-2328.
(2) Hoofnagle, J . H. N. Engl. J . Med. 1990, 323, 337-339.
(3) Aach, R. D. Ann. Intern. Med. 1988, 109, 89-91.
(4) Martin, P.; Friedman, L. S. In Innovations in Antiviral Develop-
ment and the Detection of Virus Infections; Block, T. M., J unkind, D.,
Crowell, R. L., Denison, M., Walsh, L. R., Eds.; Plenum Press: New
York, 1992; pp 111-120.
Ch em istr y
The total synthesis of robustaflavone was approached
via construction of two apigenin derivatives, one substi-
(5) Maynard, J . E.; Kane, M. A.; Hadler, S. C. Rev. Infect. Dis. 1989,
11, S574-S578.
(6) Varshney, A. K.; Rahman, W.; Okigawa, M.; Kawano, N. Ex-
perimentia 1973, 29, 784-786.
(8) Miyaura, N.; Suzuki, A. Chem. Rev. 1995, 95, 2457-2483.
(9) Muller, D.; Fleury, J .-P. Tetrahedron Lett. 1991, 32, 2229-2232.
(7) Lin, Y.-M.; Chen, F.-C. Phytochemistry 1974, 13, 1617-1619.
10.1021/jo981186b CCC: $15.00 © 1998 American Chemical Society
Published on Web 11/14/1998

Total Synthesis of Robustaflavone
J . Org. Chem., Vol. 63, No. 25, 1998 9301
Sch em e 1
Sch em e 2
salts in the iodination of phenols16 (Scheme 2). Selective
demethylation of apigenin trimethyl ether (5b) in the
5-position was accomplished with 1.1 equiv of boron
tribromide, to afford apigenin 7,4′-dimethyl ether (6).
Iodination of 6 with 1.0 equiv of I2 in the presence of 1.2
equiv of thallium(I) acetate in CH2Cl2 provided 6-iodoapi-
genin 7,4′-dimethyl ether (7), in excellent yield, contain-
tuted in the 6-position and one substituted in the
3′-position, with groups that could be coupled using
transition metal-catalyzed cross-coupling methodology.
The synthesis of apigenin derivatives substituted in the
3′-position is straightforward (Scheme 1), involving es-
terification of phloroacetophenone dimethyl ether (1) with
3-substituted p-anisoyl chlorides, such as 3-iodo-p-anisoyl
chloride (2a ). Baker10-Venkataraman11 rearrangement
of the resulting ester (3a ) to the â-diketone 4a was
achieved by heating in pyridine at 100 °C in the presence
of powdered KOH. Cyclization of the diketone 4a under
acidic conditions provided 3′-iodoapigenin trimethyl ether
(5a ).12 This route was also utilized to prepare apigenin
trimethyl ether (5b), starting with 1 and p-anisoyl
chloride.
1
ing only trace amounts (less than 1% determined by H
NMR) of the 8-iodinated species. Methylation of 7 with
dimethyl sulfate afforded the desired 6-iodoapigenin
trimethyl ether (8).
Formation of biaryl systems is efficiently achieved via
the palladium-catalyzed cross-coupling of aryl halides
and aryl boronic acids (Suzuki coupling)8 or aryl halides
and aryl stannanes (Stille coupling),17 either of which
could be applied to the synthesis of robustaflavone from
the iodinated species 5a or 8. Conceivably, the iodide of
either of these derivatives could be converted to a boronic
acid or a stannane and then cross-coupled with the other
iodide to afford robustaflavone hexamethyl ether.
In a related example, derivatives of the biflavanoid
amentoflavone were synthesized using the palladium-
catalyzed cross-coupling of apigenin ethers bearing a
boronic acid in the 8-position with apigenin ethers having
an iodide in the 3′-position, in very good yields.9 The
apigenin-8-boronic acids were synthesized from the 8-io-
dinated derivatives via halogen-lithium exchange, fol-
lowed by quenching with trimethyl borate and aqueous
workup. Attempted conversion of the 3′-iodide 5a via
halogen-lithium exchange followed by trimethyl borate
quench were unsuccessful in our hands, as for others.9
Additionally, our attempt to prepare the corresponding
apigenin 6-boronic acid derivative from 8 using this
technique were also unsuccessful, in contrast to the
reported9 simple conversion of the 8-iodinated isomer to
its corresponding boronic acid.
The preparation of apigenin derivatives substituted in
the 6-position presented a more difficult challenge, as
direct electrophilic substitution of apigenin ethers occurs
preferentially in the 8-position. An extensive search of
the chemical literature yielded only two examples of
6-halogenated apigenin derivatives; the first, described
in 1939, reported 6-bromoapigenin trimethyl ether as an
intermediate in a total synthesis of apigenin.13 However,
it was later determined that the position of the ring
bromination had been incorrectly assigned in the original
report, and that the actual intermediate was 8-bromoapi-
genin trimethyl ether.14 A second route described the
iodination of apigenin 7,4′-dimethyl ether with iodine in
a solution of iodic acid and ethanol, which provided a
mixture of the 6-iodo and 8-iodo derivatives, in a 1:4
ratio.15 The desired 6-iodinated derivative was reportedly
purified by fractional recrystallization, but in very poor
yield.
Milder general methods for the preparation of both
stannanes18 and boronic acids19 have been described,
using the palladium-catalyzed exchange of aryl halides
with nucleophilic distannane and diboron reagents,
respectively. These methods were applied to compound
5a to afford both the corresponding stannane derivative
9a and boronate 9b, illustrated in Scheme 3. Treatment
The desired 6-iodinated species was prepared in excel-
lent yield with almost exclusive regioselectivity by ex-
ploiting the ortho-directing capabilities of thallium(I)
(10) Baker, W. J . Chem. Soc. 1933, 1381-1389.
(11) Mahal, H. S.; Venkataraman, K. J . Chem. Soc. 1934, 1767-
1769.
(16) Cambie, R. C.; Rutledge, P. S.; Smith-Palmer, T.; Woodgate,
P. D. J . Chem. Soc., Perkin Trans. 1 1976, 1161-1164.
(17) Stille, J . K. Angew. Chem., Int. Ed. Engl. 1986, 25, 508-524.
(18) Azizian, H.; Eaborn, C.; Pidcock, A. J . Organomet. Chem. 1981,
215, 49-58.
(19) Ishiyama, T.; Murata, M.; Miyaura, N. J . Org. Chem. 1995, 60,
7508-7510.
(12) Nakazawa, K. Chem. Pharm. Bull. 1962, 10, 1032-1038.
(13) Hutchins, W. A.; Wheeler, T. S. J . Chem. Soc. 1939, 91-94.
(14) Chang, C. T.; Chen, T. S.; Chen, F. C. J . Org. Chem. 1961, 26,
3142-3143.
(15) Ahmad, S.; Razaq, S. Pak. J . Sci. Ind. Res. 1972, 15, 361-362.

9302 J . Org. Chem., Vol. 63, No. 25, 1998
Zembower and Zhang
Sch em e 3
Sch em e 4
being formed, we were comforted to learn that others22
had successfully prepared biaryl systems containing
similar degrees of steric crowding utilizing Suzuki meth-
odology.
of 5a with commercially available hexamethylditin in the
presence of catalytic (Ph3P)4Pd in refluxing toluene
afforded the trimethylstannane 9a . Similarly, treatment
of 5a with bis(pinacolato)diboron in the presence of
catalytic PdCl2(dppf) and K2CO3 in DMF at 85 °C
provided boronate ester 9b in 64% yield. Bis(pinacolato)-
diboron was prepared via treatment of tetrakis(trimethyl-
amino)diboron20 with pinacol as previously described21
and recently became commercially available.
Evaluation of a variety of reaction conditions, con-
ducted by changing the solvent, palladium catalyst, and
base, identified conditions that afforded the desired
robustaflavone hexamethyl ether (10). Conducting the
reaction in DMF containing 10% H2O, 4.0 equiv of NaOH,
and 10 mol % Pd(PPh3)4 as catalyst afforded 10 in 30%
yield, with only small amounts of 5b formed as a
byproduct, and trace amounts of unreacted iodide 8
(Scheme 4). Utilization of NaOH as base resulted in
greatly accelerated reaction rates relative to those ob-
served when using K2CO3, and the Suzuki coupling was
generally complete within 1 h. Increasing the equivalence
of 9b to 1.2 increased the yield of 10 to 35%, and further
increasing the stoichiometry of 9b to 2.0 equiv increased
the yield of 10 to 50%. The desired material, robustafla-
vone hexamethyl ether (10), was readily purified using
silica gel column chromatography.
Attempted Stille coupling of 9a with iodide 8 in a
variety of solvents (DMF, toluene, dioxane, THF) using
several palladium catalysts [(Ph3P)4Pd, Pd(OAc)2, PdCl2,
(Ph3P)2PdCl2] failed to provide any significant formation
of robustaflavone hexamethyl ether (10, data not shown).
When using toluene as solvent, a very small amount of
10 could be detected by TLC, by comparison with an
authentic standard prepared by methylation of natural
robustaflavone with dimethyl sulfate,7 but the major
product formed in the reaction was apigenin trimethyl
ether (5b). It is commonly accepted that transmetalation
from tin to palladium represents the rate-limiting step
in Stille couplings.17 Thus, because iodide 8 is particularly
reactive toward oxidative addition, accelerated by the
presence of the two electron-donating o-methoxyl groups,
reduction of the aryl iodide apparently occurred much
faster than transmetalation. However, it is unclear
whether 5b was formed exclusively via reduction of the
iodide 8, or also by proteodesilylation of 9a . Following
the reaction progress by TLC appeared to indicate that
the concentration of 8 decreased over time, while that of
9a was unchanged, suggesting that reduction of 8 was
the likely source of 5b.
Deprotection of 10 was initially attempted using
standard mineral acid conditions, such as HBr and HI.
In all cases, the use of mineral acids resulted in Wessely-
Moser rearrangement,23 and amentoflavone was the
major product isolated. Similar results were obtained
using aqueous (HBr and HI) or anhydrous (HBr in HOAc)
conditions.
Complete demethylation of 10 was achieved by treat-
ment with 12 equiv of BBr3 in refluxing CHCl3. Attempts
to deprotect using lesser amounts of BBr3, or at lower
temperatures, did not achieve complete demethylation,
and the resulting products contained significant quanti-
ties of partially demethylated materials. Following de-
methylation, crude robustaflavone was obtained in 88.9%
yield. Column chromatography through silica gel (toluene/
pyridine/formic acid, 20:10:1) afforded robustaflavone in
30% yield.
In contrast to Stille coupling, transmetalation from
boron to palladium in Suzuki couplings is rapid, and
oxidative addition is generally the rate-limiting step.8 We
thus turned to coupling of boronate 9b with aryl iodide
8. Using standard conditions described19 for coupling of
pinacol boronate esters with aryl halides (PdCl2(dppf),
DMF, KOAc, 80 °C), we observed that, again, the major
product formed was apigenin trimethyl ether (5b). Though
we anticipated that coupling between 9b and 8 may be
sluggish, due to the steric congestion of the biaryl bond
Su m m a r y
Robustaflavone represents an important lead com-
pound in the search for potential anti-hepatitis B agents.
Our preliminary in vitro evaluations determined that
robustaflavone possesses anti-hepatitis B activity com-
(20) Brotherton, R. J .; McCloskey, A. L.; Petterson, L. L.; Steinberg,
H. J . Am. Chem. Soc. 1960, 82, 6242-6245.
(21) No¨th, H. Z. Naturforsch. 1984, 39b, 1463-1466.
(22) J ohnson, M. G.; Foglesong, R. J . Tetrahedron Lett. 1997, 38,
7001-7002.
(23) Wessely, F.; Moser, G. H. Monatsh. Chem. 1930, 56, 97-105.

Total Synthesis of Robustaflavone
J . Org. Chem., Vol. 63, No. 25, 1998 9303
d6) δ 3.91 (s, 3 H), 7.00 (d, 1 H, J ) 8.7 Hz), 7.95 (dd, 1 H, J
) 8.7, 1.8 Hz), 8.27 (d, 1 H, J ) 1.8 Hz), 12.89 (br s, 1 H).
3-Iod o-p-a n isoyl Ch lor id e (2a ). To a suspension of PCl5
(33.0 g, 158 mmol) in 30 mL of CHCl3 was added, with
magnetic stirring, 3-iodo-p-anisic acid (40.0 g, 144 mmol) in
small portions. The solution was heated at reflux for 1 h under
a gentle nitrogen sweep, during which time the solution
became homogeneous. The solvent was evaporated under
reduced pressure, and then the residue was distilled under
high vacuum. The fraction distilling at 152-156 °C (5 mmHg;
lit.12 bp 183-185 °C, 12-13 mmHg) was collected, which
rapidly solidified as a pink solid. Recrystallization from 200
mL of hexane/CH2Cl2 afforded the desired product as white
needles (36.4 g, 85.5%): mp 57-58 °C; 1H NMR (CDCl3) δ 3.99
(s, 3 H), 6.88 (d, 1 H, J ) 8.7 Hz), 8.13 (dd, 1 H, J ) 8.7, 2.4
Hz), 8.52 (d, 1 H, J ) 2.4 Hz); EI-MS m/z 296 (M+, 42), 261
(100).
4,6-Dim et h oxy-2-(3′-iod o-4′-m et h oxyb en zoyloxy)a ce-
top h en on e (3a ). To a solution of 1 (5.00 g, 25.5 mmol) in 15
mL of pyridine was added 2a (9.08 g, 30.6 mmol), and the
solution was heated to 100 °C in an oil bath with magnetic
stirring for 10 min. The solution was cooled to room temper-
ature and then diluted with 20 mL of MeOH. The solution was
cooled in an ice bath and, following scratching with a glass
rod, colorless needles formed. The crystals were collected on a
Bu¨chner funnel, rinsed with cold MeOH and air-dried to
provide 10.1 g (86.4%) of the desired product (mp 146 °C),
which was used without further purification. An analytical
sample was obtained via recrystallization from acetone: mp
156-157 °C (lit.12 mp 158 °C; 1H NMR (CDCl3) δ 2.47 (s, 3 H),
3.83 (s, 3 H), 3.87 (s, 3 H), 3.96 (s, 3 H), 6.34 (d, 1 H, J ) 2.1
Hz), 6.41 (d, 1 H, J ) 2.1 Hz), 6.87 (d, 1 H, J ) 8.7 Hz), 8.12
(dd, 1 H, J ) 8.7, 2.1 Hz), 8.55 (d, 1 H, J ) 2.1 Hz); EI-MS m/z
456 (M+, 41), 261 (100).
parable to several nucleoside analogues currently in
clinical trials1 as well as acts synergistically with these
agents.24
We have described the first total synthesis of robust-
aflavone. The key steps included a thallium-assisted
regioselective iodination of apigenin 7,4′-dimethyl ether
in the 6-position, allowing efficient access to 6-iodoapi-
genin trimethyl ether (8). This step was critical, because
an efficient route to 6-halogenated flavones has not been
described previously. Another key step was conversion
of 3′-iodoapigenin trimethyl ether (5b) to its correspond-
ing 3′-pinacol boronate derivative 9b, via the palladium-
catalyzed cross-coupling of 5b with bis(pinacolato)-
diboron. The corresponding 3′-stannane 9a failed to
couple with iodide 8 under Stille conditions, and attempts
to convert either of iodides 5b or 8 to their corresponding
boronic acids using standard methods (halogen-lithium
exchange/trialkylborate quench) failed. Identification of
reaction conditions which allowed Suzuki coupling be-
tween boronate 9b with iodide 8 furnished the critical
6-3′′′ biaryl linkage, to afford the desired robustaflavone
skeleton in the form of its hexamethyl ether (10). Finally,
demethylation under nonaqueous conditions using BBr3
provided access to synthetic robustaflavone, which was
identical in all respects to the natural product. This total
synthesis will allow access to needed quantities of ro-
bustaflavone for further biological studies as well as
provide an efficient method for the synthesis of structural
analogues.
1-(3′-Io d o-4′-m e t h ox yp h e n y l)-3-(2′′-h y d r o x y -3′′,6′′-
d im eth oxyp h en yl)-1,3-p r op a n ed ion e (4a ). To a suspension
of 3a (9.12 g, 20.0 mmol) in 20 mL of pyridine was added 2.80
g (50 mmol) of powdered KOH, and the solution was heated
to 100 °C with magnetic stirring in an oil bath for 10 min.
The solution was cooled to room temperature and treated with
10 mL of HOAc, which produced a yellow paste. Addition of
20 mL of MeOH and cooling in an ice bath afforded a yellow
powder, which was collected on a Bu¨chner funnel, rinsed with
100 mL of MeOH, and air-dried to provide 5.68 g (62.3%) of
the desired product: mp 170-171 °C (lit.12 mp 168 °C); 1H
NMR (CDCl3) δ 3.51 (s, 3 H), 3.82 (s, 3 H), 3.97 (s, 3 H), 4.48
(s, 2 H), 5.85 (d, 1 H, J ) 2.1 Hz), 6.09 (d, 1 H, J ) 2.1 Hz),
6.89 (d, 1 H, J ) 8.7 Hz), 7.96 (dd, 1 H, J ) 8.7, 2.1 Hz), 8.40
(d, 1 H, J ) 2.1 Hz), 13.68 (s, 1 H); EI-MS m/z 456 (M+, 70),
425 (40), 261 (83), 181 (100).
3′-Iod o-5,7,4′-tr im eth oxyfla von e (5a ). To a magnetically
stirred solution of 4a (5.00 g, 11.0 mmol) in 60 mL of HOAc at
100 °C (water bath) was added 10 mL of 20% H2SO4/HOAc.
After stirring at 100 °C for 10 min, the solution was poured
into 250 mL of H2O, which produced a white precipitate. The
solid was collected on a Bu¨chner funnel, rinsed with 500 mL
of H2O, and air-dried. Recrystallization from 150 mL of dioxane
produced white needles (2.83 g, 58.9%). Evaporation of the
mother liquor and recrystallization afforded an additional 1.35
g (total yield 4.18 g, 87.0%): mp 209-210 °C (lit.12 mp 223
°C); 1H NMR (DMSO-d6) δ 3.83 (s, 3 H), 3.91 (s, 3 H), 3.92 (s,
3 H), 6.50 (d, 1 H, J ) 2.1 Hz), 6.73 (s, 1 H), 6.91 (d, 1 H, J )
2.1 Hz), 7.14 (d, 1 H, J ) 8.7 Hz), 8.06 (dd, 1 H, J ) 8.7, 2.4
Hz), 8.41 (d, 1 H, J ) 2.4 Hz); EI-MS m/z 438 (M+, 100), 407
(16). Anal. Calcd for C18H15IO5: C, 49.29; H, 3.42. Found: C,
48.98; H, 3.39.
Exp er im en ta l Section
Gen er a l Exp er im en ta l. Column chromatography was
conducted with EM Science silica gel 60 (70-230 mesh) with
indicated eluents. Analytical thin-layer chromatography was
performed with silica gel 60 F254 precoated glass-backed plates
(250 µm) with indicated eluents. Bis(pinacolato)diboron was
prepared from tetrakis(dimethylamino)diboron20 as previously
described.21 All other reagents and solvents were purchased
from commercial sources and used without further purification.
2,4-Dim eth oxy-6-h yd r oxya cetop h en on e (1). To a solu-
tion of phloroacetophenone hydrate (20.5 g, 110 mmol) and
K2CO3 (22.1 g, 160 mmol) in 150 mL of acetone was added
dimethyl sulfate (27.7 g, 220 mmol) slowly over 30 min with
mechanical stirring. The solution was heated at reflux over-
night and then poured into 500 mL of H2O, which produced a
white solid. The material was collected on a Bu¨chner funnel,
rinsed with 1 L of H2O and air-dried. Recrystallization from
80 mL of 90% MeOH afforded white needles (16.1 g, 74.4%):
mp 79-80 °C (lit.12 mp 82-83 °C); 1H NMR (CDCl3) δ 2.61 (s,
3 H), 3.82 (s, 3 H), 3.86 (s, 3 H), 5.92 (d, 1 H, J ) 2.4 Hz), 6.06
(d, 1 H, J ) 2.4 Hz), 14.03 (s, 1 H).
3-Iod o-p-a n isic Acid . To a mechanically stirred suspension
of p-anisic acid (59.90 g, 394 mmol) and iodine (100.0 g, 394
mmol) in a mixture of 325 mL of glacial acetic acid and 60 g
of concd H2SO4, heated to 45 °C with a water bath, was added
dropwise a solution of 40 g of concd HNO3 in 60 mL of HOAc
at such a rate that the temperature was maintained between
40 and 50 °C (ca. 90 min). After addition, the mixture was
stirred for 30 min at 50 °C and then diluted with 400 mL of
H2O, which produced a pink solid. The material was collected
on a Bu¨chner funnel, rinsed with 1 L of 10% Na2S2O4 and 1 L
of H2O, and then air-dried. Recrystallization from 700 mL of
pyridine/MeOH (1:1) provided colorless plates. The crystals
were collected on a Bu¨chner funnel, rinsed with 500 mL of
MeOH, and then allowed to air-dry. After drying under high
vacuum overnight, 71.4 g (65.2%) of the desired product was
obtained; mp 243-244 °C (lit.12 mp 238 °C); 1H NMR (DMSO-
4,6-Dim e t h oxy-2-(4′-m e t h oxyb e n zoyloxy)a ce t op h e -
n on e (3b). To a solution of 1 (20.0 g, 102 mmol) in 60 mL of
pyridine was added 20.9 g (122 mmol) p-anisoyl chloride (2b),
and the solution was heated at 100 °C for 10 min with
magnetic stirring in an oil bath. The solution was cooled to
room temperature and then diluted first with 50 mL of EtOH
followed by 50 mL of H2O. The mixture was cooled to 0 °C in
an ice bath which, after scratching with a glass rod, produced
shiny white plates. The crystals were collected on a Bu¨chner
(24) Zembower, D. E.; Lin, Y.-M.; Flavin, M. T.; Chen, F.-C.; Korba,
B. E. Antiviral Res. 1998, 39, 81-88.

9304 J . Org. Chem., Vol. 63, No. 25, 1998
Zembower and Zhang
funnel, rinsed with cold 50% EtOH, and air-dried to afford 27.4
g (81.4%) of desired product: mp 97-98 °C (lit.25 mp 115-
116 °C); 1H NMR (CDCl3) δ 2.46 (s, 3 H), 3.82 (s, 3 H), 3.86 (s,
3 H), 3.88 (s, 3 H), 6.37 (d, 1 H, J AB ) 2.1 Hz), 6.39 (d, 1 H,
J AB ) 2.1 Hz), 6.96 (d, 2 H, J ) 8.9 Hz), 8.09 (d, 2 H, J ) 8.9
Hz); EI-MS m/z 330 (M+, 23), 135 (100). Anal. Calcd for
C18H18O6: C, 65.45; H, 5.49. Found: C, 65.55; H, 5.57.
suspension of thallium salts precipitated. The solution was
stirred at room temperature overnight and then filtered
through a bed of Celite to remove the precipitated salts. The
filtrate was extracted sequentially with 500 mL each of 5%
NaHCO3, 10% Na2S2O4, and saturated brine and then dried
over magnesium sulfate and filtered. Evaporation in vacuo
provided an orange solid. Recrystallization from 300 mL of
CHCl3/EtOH (1:2) afforded fine yellow needles (3.10 g, 73.1%);
mp 227-228 °C (lit.15 mp 205-207 °C); 1H NMR (CDCl3) δ
3.90 (s, 3 H), 4.00 (s, 3 H), 6.54 (s, 1 H), 6.64 (s, 1 H), 7.02 (d,
1-(4′-Meth oxyp h en yl)-3-(2′′-h yd r oxy-3′′,6′′-d im eth oxy-
p h en yl)-1,3-p r op a n ed ion e (4b). To a solution of 3b (30.0 g,
90.8 mmol) in 120 mL of pyridine was added 12.7 g (153 mmol)
powdered KOH, and the solution was heated to 100 °C in an
oil bath with magnetic stirring for 10 min. The mixture was
cooled to room temperature and then treated with 50 mL of
glacial acetic acid, which produced a thick yellow paste. The
mixture was diluted with 100 mL of EtOH, which afforded a
homogeneous solution, followed by 100 mL of H2O. After being
cooled to 0 °C, the product crystallized as yellow prisms. The
crystals were collected on a Bu¨chner funnel, rinsed with cold
50% EtOH, and air-dried to provide 20.72 g (69.1%) of desired
product: mp 132-133 °C (lit.25 mp 147-149 °C); 1H NMR
(CDCl3) δ 3.48 (s, 3 H), 3.81 (s, 3 H), 3.89 (s, 3 H), 4.51 (s, 2
H), 5.83 (d, 1 H, J ) 2.4 Hz), 6.09 (s, 1 H, J ) 2.4 Hz), 6.97 (d,
2 H, J ) 9.1 Hz), 7.94 (d, 2 H, J ) 9.1 Hz), 13.74 (s, 1 H);
FTIR (KBr) 3071, 1724, 1584, 1182, 1138 cm-1; EI-MS m/z 330
(M+, 46), 135 (100). Anal. Calcd for C18H18O6: C, 65.45; H, 5.49.
2 H, J ) 9.0 Hz), 7.85 (d, 2 H, J ) 9.0 Hz), 13.84 (s, 1 H); 13
C
NMR (75 MHz, CDCl3) δ 181.7, 164.4, 163.5, 162.9, 161.3,
158.3, 128.2, 123.3, 114.6, 104.4, 104.3, 90.5, 90.4, 56.8, 55.5;
EI-MS m/z 424 (M+, 100). Anal. Calcd for C17H13IO5: C, 48.14;
H, 3.09. Found: C, 48.30; H, 3.11.
6-Iod o-5,7,4′-t r im et h oxyfla von e (8). To a solution of 7
(3.00 g, 7.08 mmol) and K2CO3 (1.47 g, 10.6 mmol) in 150 mL
of THF was added dimethyl sulfate (1.07 g, 8.49 mmol), and
the solution was heated at reflux overnight. The solvent was
evaporated and the residue partitioned between 50 mL each
of CHCl3 and H2O. The organic phase was separated and
washed with saturated brine, dried over magnesium sulfate,
filtered, and evaporated to provide a yellow solid. Recrystal-
lization from CHCl3/EtOH (1:2, 150 mL) afforded pale-yellow
needles (1.49 g). Column chromatography of the evaporated
mother liquor (100 g of silica gel, 2% MeOH/CH2Cl2) afforded
an additional 720 mg of desired product (total yield 2.21 g,
71.3%); mp 202-204 °C (lit.15 mp 191-194 °C); 1H NMR
(CDCl3) δ 3.89 (s, 3 H), 3.94 (s, 3 H), 4.01 (s, 3 H), 6.62 (s, 1
H), 6.78 (s, 1 H), 7.01 (d, 2 H, J ) 9.1 Hz), 7.83 (d, 2 H, J )
9.1 Hz); EI-MS m/z 438 (M+, 65), 311 (100). Anal. Calcd for
1
Found: C, 65.38; H, 5.54. The H NMR spectra also indicated
the presence of enol tautomers.
5,7,4′-Tr im et h oxyfla von e (a p igen in t r im et h yl et h er ,
5b). A suspension of 4b (18.6 g, 56.4 mmol) in 200 mL of glacial
acetic acid was heated to 100 °C with magnetic stirring in an
oil bath. To this suspension was added 40 mL of 20% H2SO4
in acetic acid, and the mixture was stirred at 100 °C for 10
min. The mixture was poured into 1 L of H2O, which produced
a pale-yellow gelatinous solid. The solid was collected on a
Bu¨chner funnel, allowed to partially dry by drawing air
through the funnel, and then partitioned between 600 mL each
of CHCl3 and H2O. The organic layer was separated and
washed with 600 mL each of 5% NaHCO3 and saturated brine,
dried over magnesium sulfate, filtered, and evaporated to
afford a light yellow solid. Recrystallization from 300 mL of
acetone provided white needles (12.45 g, 70.8%): mp 159-
160 °C (lit.26 mp 156 °C); 1H NMR (DMSO-d6) δ 3.83 (s, 3 H),
3.85 (s, 3 H), 3.90 (s, 3 H), 6.50 (d, 1 H, J ) 2.1 Hz), 6.67 (s, 1
H), 6.84 (d, 1 H, J ) 2.1 Hz), 7.09 (d, 2 H, J ) 8.9 Hz), 7.99 (d,
C
18H15IO5: C, 49.34; H, 3.45. Found: C, 48.58; H, 3.21.
5,7,4′-Tr im eth oxy-3′-(tr im eth ylsta n n yl)fla von e (9a ). To
a solution of 5a (438 mg, 1.00 mmol) in 30 mL of toluene were
added Pd(Ph3P)4 (150 mg, 0.13 mmol) and hexamethylditin
(655 mg, 2.00 mmol), and the solution was heated to reflux
under N2 for 16 h. The solution was filtered and the solvent
evaporated in vacuo. The residue was dissolved in 75 mL of
CHCl3, washed with 75 mL of saturated brine, dried over
magnesium sulfate, filtered, and evaporated to provide a white
crystalline solid. The solid was triturated with 10 mL of EtOH,
collected on a Buchner funnel, rinsed with fresh EtOH, and
air-dried (270 mg, 56.8%). An analytical sample was obtained
via recrystallization from EtOH, which produced fine colorless
needles: mp 154-155 °C; 1H NMR (DMSO-d6) δ 0.31 (s, 9 H),
3.83 (s, 3 H), 3.85 (s, 3 H), 3.91 (s, 3 H), 6.51 (d, 1 H, J ) 2.4
Hz), 6.65 (s, 1 H), 6.82 (d, 1 H, J ) 2.4 Hz), 7.08 (d, 1 H, J )
8.7 Hz), 7.86 (d, 1 H, J ) 2.4 Hz), 8.01 (dd, 1 H, J ) 8.7, 2.4
Hz); 13C NMR (75 MHz, CDCl3) δ 177.9, 166.5, 164.0, 161.4,
161.0, 160.1, 134.3, 131.7, 128.6, 124.1, 109.1, 107.7, 96.0, 92.9,
56.4, 55.7, 55.5; EI-MS m/z 476 (M+, 64), 461 (100). Anal. Calcd
for C21H24O5Sn: C, 53.09; H, 5.09. Found: C, 52.74; H, 5.01.
P in a col 5,7,4′-tr im eth oxyfla von e-3′-bor on a te (9b). A
solution of 5a (2.00 g, 4.56 mmol), bis(pinacolato)diboron (1.50
g, 5.93 mmol), KOAc (1.79 g, 18.24 mmol), and PdCl2(dppf)
(372 mg, 0.456 mmol) in 60 mL of DMF was stirred at 80 °C
overnight. The reaction mixture was filtered, diluted with 200
mL of EtOAc, and then washed with H2O (3×) and brine. After
drying over sodium sulfate, the solvent was evaporated. The
product was chromatographed over silica gel (EtOAc/MeOH,
96:4) to afford 1.21 g (60.5%) of the desired product as a gray
solid. The material could be recrystallized from CH2Cl2/EtOAc,
as pale gray needles: mp 218-220 °C; 1H NMR (CDCl3) δ 1.39
(s, 12 H), 3.91 (s, 3 H), 3.92 (s, 3 H), 3.96 (s, 3 H), 6.38 (d, 1 H,
J ) 2.2 Hz), 6.60 (d, 1 H, J ) 2.2 Hz), 6.66 (s, 1 H), 6.96 (d, 1
H, J ) 8.9 Hz), 7.96 (dd, 1 H, J ) 8.9, 2.4 Hz), 8.18 (d, 1 H, J
) 2.4 Hz); 13C NMR (CDCl3) δ 177.9, 166.6, 164.0, 161.0, 160.9,
160.0, 134.9, 130.4, 123.3, 114.4, 110.6, 109.3, 107.8, 96.0, 93.0,
83.9, 56.4, 55.9, 55.7, 24.7; FTIR (KBr) 1645, 1602, 1330, 1148
2 H, J ) 8.9 Hz); FTIR (KBr) 1644, 1348, 1256, 1121, 831 cm-1
;
EI-MS m/z 312 (M+, 100). Anal. Calcd for C18H16O5: C, 69.22;
H, 5.16. Found: C, 69.35; H, 5.32.
7,4′-Dim eth oxy-5-h yd r oxyfla von e (6). To a solution of 5b
(10.0 g, 32.1 mmol) in 200 mL of anhydrous CH2Cl2 was added
a 1 M solution of BBr3 (35.3 mL, 35.3 mmol) dropwise over 15
min at room temperature with magnetic stirring. A thick
yellow precipitate formed rapidly during the addition. After
stirring 5 h, the reaction was quenched by adding 200 mL of
EtOH, and the solvent was then evaporated in vacuo. The
yellow residue was triturated with 300 mL of boiling 50%
EtOH. After cooling to room temperature, the yellow solid was
collected on a Bu¨chner funnel, rinsed with 500 mL of 50%
EtOH, and air-dried. Recrystallization from 1.5 L of EtOH
provided 8.16 g (85.4%) of the title product as fine, pale-yellow
needles: mp 176-177 °C (lit.27 mp 174-175 °C); 1H NMR
(CDCl3) δ 3.88 (s, 3 H), 3.89 (s, 3 H), 6.36 (d, 1 H, J ) 2.1 Hz),
6.48 (d, 1 H, J ) 2.1 Hz), 6.57 (s, 1 H), 7.01 (d, 2 H, J AB ) 9.0
Hz), 7.84 (d, 2 H, J AB ) 9.0 Hz), 12.81 (s, 1 H); EI-MS m/z 298
(M+, 100). Anal. Calcd for C17H14O5: C, 68.45; H, 4.73.
Found: C, 68.25; H, 4.77.
7,4′-Dim eth oxy-5-h yd r oxy-6-iod ofla von e (7). To a solu-
tion of 6 (2.98 g, 10.0 mmol) in 300 mL of CH2Cl2 was added
thallium(I) acetate (3.16 g, 10.2 mmol). With magnetic stirring,
a solution of iodine (2.54 g, 10.0 mmol) in 200 mL of CH2Cl2
was added dropwise over 1 h. During the addition, a fine
cm-1; APCI-MS m/z 439 (MH+, 100). Anal. Calcd for C24H27
-
BO7‚1/2H2O: C, 64.45; H, 6.31. Found: C, 64.17; H, 6.05.
Robu sta fla von e Hexa m eth yl Eth er (10). To a solution
of 8 (25.0 mg, 0.057 mmol) and 9a (50.0 mg, 0.114 mmol) in
DMF/H2O (9:1), which had been deoxygenated for 15 min by
bubbling N2, were added NaOH (9.1 mg, 0.23 mmol) and Pd-
(25) Roy, D.; Khanna, R. N. Indian J . Chem. 1980, 19B, 583-586.
(26) Schmid, L.; Waschkau, A. Monatsh. Chem. 1928, 49, 83-91.
(27) Dawson, R. M.; Henrick, C. A.; J efferies, P. R.; Middleton, E.
J . Aust. J . Chem. 1965, 18, 1871-1875.

Total Synthesis of Robustaflavone
J . Org. Chem., Vol. 63, No. 25, 1998 9305
(PPh3)4 (6.6 mg, 0.0057 mmol), and the reaction was stirred
at 80 °C for 2 h. The solution was diluted with CH2Cl2 and
then extracted with H2O and saturated brine. The organic
layer was dried over magnesium sulfate, filtered, and evapo-
rated in vacuo. Column chromatography (silica gel, CH2Cl2/
MeOH, 96:4) afforded the desired product 10 (17.7 mg, 50%),
identical with an authentic sample prepared via methylation
with MeOH, the solvent again evaporated in vacuo, and the
solid partitioned between EtOAc and 1 M NaOH. The organic
layer was discarded, and the aqueous layer was extracted with
EtOAc. After cooling to 0 °C, the aqueous layer was carefully
acidified to pH 3.0 by the dropwise addition of 3 M HCl. The
resulting yellow precipitate was collected by vacuum filtration,
rinsed with water, and air-dried (38.7 mg). The crude material
1
of robustaflavone:7 mp 296-297 °C (lit.7 mp 303-305 °C) H
was chromatographed through silica gel, eluting with
a
NMR (CDCl3) δ 3.62 (s, 3 H), 3.85 (s, 3 H), 3.87 (s, 3 H), 3.91
(s, 6 H), 3.96 (s, 3 H), 6.37 (d, 1 H, J ) 2.1 Hz), 6.60 (d, 1 H,
J ) 2.4 Hz), 6.69 (s, 1 H), 6.74 (s, 1 H), 6.89 (s, 1 H), 7.04 (d,
2 H, J ) 9.0 Hz), 7.10 (d, 1 H, J ) 8.7 Hz), 7.81 (d, 1 H, J )
2.4 Hz), 7.89 (d, 2 H, J ) 9.0 Hz), 7.89 (dt, 1 H, J ) 2.1, 8.7
Hz); HR-EIMS m/z 623.1917 (MH+, requires 623.1917). Anal.
Calcd for C36H30O10‚0.2CH2Cl2: C, 68.34; H, 4.81. Found: C,
68.28; H, 5.19. (Note: dichloromethane was extremely difficult
to remove from this material, consistent with our observations
of the hexamethyl ether prepared from natural robustafla-
vone).
Robu sta fla von e. To a solution of 10 (75.0 mg, 0.12 mmol)
in 10 mL of dry CHCl3 was added BBr3 (1.0 M in CH2Cl2, 1.45
mL, 1.45 mmol), and the resulting yellow slurry was stirred
at reflux overnight. The reaction mixture was cooled to room
temperature, quenched by the careful addition of MeOH, and
evaporated in vacuo. The resulting orange solid was triturated
mixture of toluene/pyridine/formic acid (20:10:1). Appropriate
fractions were combined and evaporated to afford 19.4 mg
(30.0%) robustaflavone. An analytical sample was obtained via
recrystallization from pyridine/H2O (1:1): mp 370-372 °C, dec
(lit.1 mp 350-352 °C). Spectral data of synthetic robustafla-
vone was identical to that recently reported1 for the natural
product isolated from Rhus succedanea. HR-FABMS m/z
538.0912 (M+, requires 538.0900). Anal. Calcd for C30H18O10
H2O: C, 64.75; H, 3.60. Found: C, 64.95; H, 3.91.
‚
Ackn owledgm en t. Portions of this work were funded
under Small Business Innovation Research grant 1R43
AI40745-01, awarded to D.E.Z. by the National Insti-
tutes of Health.
J O981186B