Synthesis of [2-13C, 4-13C]-(2R,3S)-catechin and [2-13C, 4-13C]-(2R,3R)-epicatechin

Article abstract of DOI:10.1002/jlcr.1791

The first synthesis of doubly labeled, [2-¹³C, 4- ¹³C]-(2R,3S)-catechin 15 and [2-¹³C, 4- ¹³C]-(2R,3R)-epicatechin 18 starting from labeled 2-hydroxy-4, 6-bis(benzyloxy)acetophenone 3 and labeled 3, 4-bis(benzyloxy)-benzaldehyde 7 are described. Copyright

Full text of DOI:10.1002/jlcr.1791

Research Article
Received 24 February 2010,
Revised 19 April 2010,
Accepted 20 April 2010
Published online 30 July 2010 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.1791
Synthesis of [2-13C, 4-13C]-(2R,3S)-catechin
and [2-13C, 4-13C]-(2R,3R)-epicatechin
Ã
Pradeep K. Sharma,a Min He,a Leo J. Romanczyk Jr,b and Hagen Schroeterb
The first synthesis of doubly labeled, [2-13C, 4-13C]-(2R,3S)-catechin 15 and [2-13C, 4-13C]-(2R,3R)-epicatechin 18 starting
from labeled 2-hydroxy-4, 6-bis(benzyloxy)acetophenone 3 and labeled 3, 4-bis(benzyloxy)-benzaldehyde 7 are described.
Keywords: flavan-3-ol; [2-13C, 4-13C]-(2R, 3S)-catechin; [2-13C, 4-13C]-(2R, 3R)-epicatechin; CH3 13CN; Me2N13CHO
An intriguing possibility has been raised by the recent plant
literature. Several reports have indicated that plant nuclei and
Introduction
histones are the target for flavanols and proanthocyanidins.23 In
There is great interest in the role played by the polyphenolic
one instance23b flavanols have been detected in calf small
class of compounds in health, nutrition and treatment of
disease. In particular, dietary interventions using flavanol-
containing foods and beverages have substantiated epidemio-
logical data regarding the improvement of vasodilation1 and
blood pressure.2 Moreover, numerous investigations have been
conducted in vitro aimed at elucidating flavanol-mediated
bioactivities. These activities include, but are not limited to
inhibiting HIV-1 replication in vitro,3 reducing the risk of heart
disease,4 suppressing ulcer formation,5 possessing antimuta-
genic,6 neuroprotective,7 anti-inflammatory,8 antibacterial,9
hypertensive10 properties, and can inhibit the growth of cancer
cells.11 Moreover, flavanols can affect a host of cellular functions
in vitro such as signaling pathways,12 alter cell membrane
characteristics13 and receptor function14 to imply physiological
effects in vivo. It should be noted that these in vitro studies use
the natural form of the flavanols as they appear in nature,
whereas the biological effects elicited in vivo are the con-
sequences of adsorption (bioavailability), their metabolism and
distribution within body organs. Substantial data exists to
support the in vivo biotransformation and metabolism of
flavanols within the gastrointestinal tract by resident micro-
flora15 as well as their hepatic metabolism.16
intestinal cell nuclei upon addition to cells in vitro raising the
possibility that flavanols and accompanying proanthocyanidins
may be ‘hidden’ in other mammalian cell nuclei. Verification of
this observation and its extension to other cell lines in vitro has
not yet been established.
To complement and extend this validation process, we have
developed the synthesis for doubly 13C-labeled (2R,3S)-catechin
and (2R,3R)-epicatechin for more rigorous in vitro and in vivo
studies. These labeled monomers have been developed for use
‘as is’ and for the synthesis24 of B-type dimers and trimers up to
the multigram level to address the fates of these procyanidins in
vivo. A doubly labeled target was chosen to accommodate the
various types of chemistry active at the C-2 and C-4 positions of
the pyran ring to account for the known products of
biotransformation and metabolic processes, as well as account
for the possible biotransformation sites of higher oligomers. This
would include the possible oxidation of B-types to A-types,25
where doubly linked A-type dimers and trimers could be
formed. In this case, all of the cited synthesis steps could be
adapted for the 14C-labeled synthesis of the monomers and their
resultant B-type dimers and trimers.
To validate flavanol activities in vivo, isotopic labeling has
proved to be an invaluable tool. Multiple approaches have been
reported in the literature to include deuterium labeling,17 14C-
labeling in plants via incorporation of 14CO2, 14C-acetate and
14C-phenylalanine,18 and direct synthesis of 13C and 14C-labeled
flavanols.19 These approaches have been applied to small,
monomeric flavanols, but little information is available to
describe the adsorption, distribution and metabolic fates of
higher molecular weight oligomers (procyanidins)20 which are
abundantly present in many foods and beverages. This is in spite
of their reported stability during gastric transit,21 where it was
originally thought that the acidic conditions within the stomach
would hydrolyze the larger oligomers into smaller species down
to the monomers. Additionally, in vitro studies with Caco-2 cells
have shown dimers and trimers to cross multiple layers of
cells.22
As a first step in this process, the first doubly (at C-2 and C-4)
13C-labeled synthesis of (2R,3S)-catechin 15 and (2R,3R)-
epicatechin 18 (Figure 1) were established. The synthesis
developed here for 15 and 18 could then be applied to obtain
the other isomers, which are not as abundant in nature as
(2R, 3S)-catechin.
For these applications, we decided to synthesize [2-13C,
4-13C]-(2R,3S)-catechin 15 and [2-13C, 4-13C]-(2R,3R)-epicatechin
aChemical Process R&D, Johnson Matthey Pharmaceutical Materials, 25 Patton
Road, Devens, MA 01434, USA
bMARS Incorporated, 6885 Elm Street, McLean, VA 22101, USA
*Correspondence to: Pradeep K. Sharma, Chemical Process R&D and CMC,
ARIAD Pharmaceuticals Inc., 26 Landsdowne Street, Cambridge, MA 02139, USA.
E-mail: sharmap76@yahoo.com
J. Label Compd. Radiopharm 2010, 53 605–612
Copyright r 2010 John Wiley & Sons, Ltd.

P. K. Sharma et al.
18 starting from labeled 2-hydroxy-4, 6-bis(benzyloxy)acetophe- corresponding doubly labeled chalcone 8 via a Claisen-Schmidt
none 3 and labeled 3,4-bis(benzyloxy)benzaldehyde 7.
reaction in 86% yield.19d,e Attempts to reduce the conjugated
ketone 8 to alkene 9 using ethyl chloroformate and NaBH4 as
reported by Kijima27 either at low temperature or elevated
temperature did not produce alkene 9 and the starting material
was recovered. Alternatively, the selective reduction of the
conjugated ketone 8 with NaBH4 in the presence of cerium
chloride (CeCl3 ꢀ 7H2O) in a mixture of ethanol and THF at 0–51C
provided the alkene 9 after chromatography in 84% yield.
Performing the reaction at low temperature was essential to
obtain the high yield of 9 since elevated temperatures resulted
in the formation of by-products. This newly developed method
to obtain 9 was an improvement over the reported procedure
by Li.26 The phenolic hydroxyl group of 9 was protected
with TBDMSC1 and imidazole in DMF to yield compound
10 in quantitative yield. The asymmetric dihydroxylation of
compound 10 under Sharpless dihydroxylation conditions
using AD-mix-a and methanesulfonamide in a mixture of
t-butanol, H2O and THF yielded compound 11 in good
yield.19b The use of co-solvent THF, resulted in a short reaction
time and high enantio-selectivity. Use of other co-solvents
such as CH2Cl2, EtOAc or toluene resulted in longer reaction
times, low yield and purity of the desired compound. The
reaction of compound 11 with nBu4NF in THF resulted in
the deprotection of the TBMDS group to produce 12.26
However, chiral HPLC indicated the presence of isomers due
to the isomerization of 12, possibly at the C-2 position.
Alternatively, when a cold mixture of equimolar amounts of
nBn4NF and glacial acetic acid in THF was reacted at 0–51C with
11, the triol 12 was produced in 88% yield after chromato-
graphy. Chiral HPLC indicatedo2% isomerization of 12 under
these newly developed conditions. Thus, these deprotection
conditions of the TBDMS group avoided the isomerization at
the C-2 position.
Results and discussions
Our synthetic approach for the construction of 15 and
18 is depicted in Scheme 3. The requisite starting materials, 3
and 7 are synthesized from commercially available starting
materials. The synthesis of 3 was accomplished as outlined in
Scheme 1.
The reaction of phloroglucinol 1 with labeled-acetonitrile
(CH313CN) in the presence of ZnCl2 in Et2O resulted in the
formation of 2,4,6-trihydroxyacetophen-[2-13C]-one
2 as a
yellow solid in 39% yield.16 The reaction of 2 with BnCl in the
presence of K2CO3 in DMF at 75–801C yielded 2-hydroxy-4,
6-bis(benzyloxy)acetophen-[13C]-one 3 in 33% yield.
The synthesis of compound 7 was achieved as outlined in
Scheme 2.
The reaction of 4-bromo veratrole 4 with BBr3 in CH2Cl2
resulted in 4-bromo-catechol 5 in quantitative yield. The
reaction of 5 with BnBr in the presence of K2CO3 in DMF
resulted in the formation of 3, 4-bis(benzyloxy)bromobenzene 6
in 27% yield after chromatography. The treatment of 6 with Mg
metal in refluxing THF and in the presence of a catalytic amount
of 2, 4-dibromoethane followed by reaction with N, N-dimethyl-
[
hyde 7 as an off-white solid.
13C]-formamide yielded 3, 4-bis(benzyloxy)benz-[2-13C]-alde-
Once the synthesis of key starting materials was achieved, our
efforts were directed to the target compounds. The syntheses of
15 and 18 are based upon our and other research.19,26 The
synthesis of these compounds was achieved as depicted in
Scheme 3. The syntheses of 15 and 18 were accomplished from
a common intermediate 12 in high optical purity. This key
intermediate was obtained from commercially available starting
materials 1 and 4 in gram quantities.
The reaction of triol 12 with triethylorthoformate in PPTS
in 1, 2-dichloromethane at 601C resulted in 13a in 68% yield and
84% ee. Alternatively, when triol 12 was reacted with
triethylorthopropionate [EtC(OEt)3] in place of triethylorthofor-
mate, and PPTS in 1,2-dichloroethane at 601C for 2–3 h, these
conditions afforded [2-13C, 4-13C]-3-O-propyl ester 5,7,30,40-
tetra-O-benzyl-(2R,3S)-catechin 13 in quantitative yield with
98% ee.26 The use of triethylorthopropionate in place of
triethylorthoformate avoided the isomerization of 13 under
As depicted in Scheme 3, the base-catalyzed condensation
between labeled 2-hydroxy-4, 6-bis(benzyloxy)acetophenone 3
and labeled 3,4-bis(benzyloxy)benzaldehyde 7 furnished the
OH
OH
OH
OH
HO
O
HO
O
*
*
1
these conditions as determined by H NMR and chiral HPLC.26
OH
OH
*
*
The ester 13 was then hydrolyzed using K2CO3 in methanol at
ambient temperature to produce 14 in 82% yield. The optical
purity of 14 was further improved upon crystallization from
toluene. The reaction of 14 with palladium hydroxide on carbon
in EtOAc at room temperature under hydrogen atmosphere
resulted in the formation of [2-13C, 4-13C]-(2R,3S)-catechin 15 as
an off-white solid in quantitative yield.
OH
OH
[2-13C, 4-13C]-(2R,3R)-
Epicatechin [* = 13C]
18
[2-13C, 4-13C]-(2R,3S)-
Catechin [* = 13C]
15
Figure 1. Structures of (2R, 3S)-catechin 15 and (2R, 3R)-epicatechin 18
CH313CN
ZnCl2
Et2O
BnCl
K2CO3
DMF
BnO
OH
HO
OH
HO
OH
33%
*
*
39%
OBn O
OH
O
OH
1
2
3
[* = 13C]
Scheme 1. Synthesis of 2-hydroxy-4,6-bis(benzyloxy)acetophen-[13C]-one (3)
www.jlcr.org
Copyright r 2010 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2010, 53 605–612

P. K. Sharma et al.
BnBr
BBr3
CH2Cl2
OCH3
OCH3
OH
OH
OBn
OBn
K2CO3
27%
100%
Br
Br
Br
5
6
4
Mg, THF
Me2N
OBn
OBn
13C
HO
H
*
77%
O
7
Scheme 2. Synthesis of 3,4-bis(benzyloxy)benz-[13C]-aldehyde (7)
2, 4, 6-Trihydroxyacetophen-[13C]-one (2)
The oxidation of the C-3 hydroxyl group of 14 with Dess-
Martin periodinane (DMP)28 in wet CH2Cl2 resulted in 16 in 90%
yield. The presence of water in the reaction was required to
accelerate the reaction rate.24,28 The reaction of 16 with Al(OiPr)3
and 2-propanol in refluxing toluene under Meerwein–
Ponndorf–Verley reduction conditions resulted in the formation
of 17 in 74% yield after crystalization.24 The treatment of 17
with palladium hydroxide on carbon in ethyl acetate at room
temperature under hydrogen atmosphere using a balloon
produced 18 in 77% yield.
Phloroglucinol dihydrate (9.98 g, 61.6mmol, 1 eq.) was dried
under high vacuum at 125731C to a constant weight (Note: It
tookꢃ72h at this scale to achieve a constant weight. After this
time, onlyꢃ1.3% water content remained in the compound).
Under N2 atmosphere, a 2 M solution of ZnCl2 in Et2O (80mL) was
added and the mixture was stirred at room temperature for 0.5 h.
A clear solution was obtained. Then, CH3 13CN (5g, 121.8 mmol,
1.98eq.) was added and the resulting mixture was stirred for
188 h at RT. The solids were suction filtered. The solids were
dissolved in H2O (80 mL) and refluxed for 4 h. The reaction
mixture was cooled to RT. The solids were suction filtered and
dried under high vacuum at RT for 20h to give 5.16g (39% yield)
of 2 as a yellow solid. HPLC analysis using Method A showed the
product to be 100% pure with a retention time of 7.2 min.1H NMR
(300MHz, CDCl3) d = 2.53 (d, J = 5.8 Hz, 3H), 5.8 (s, 2H), 10.26 (br s,
1H), 11.2 (br s, 1H). 13C NMR (75 MHz, CDCl3) d = 202.4.
The optical rotation of 15 and 18 were consistent with those
reported in the literature.29
Experimental
General
All the solvents were purchased from Aldrich Chemical
Company in Sure/SealTM bottles and were used as received.
The labeled N, N-dimethyl-[13C]-formamide (99 atom % 13C) and
CH313CN (99 atom % 13C) were purchased from Cambridge
Isotope Laboratory and were used as received. Phloroglucinol
dihydrate and 4-bromo veratrole were purchased from Alfa
Aesar. 1H NMR spectra were recorded on a 300 MHz Bruker,
whereas 13C NMR spectra were recorded on a 75 MHz Bruker
NMR and TMS was used as an internal standard. In 13C NMR, only
the chemical shift of the 13C-labeled position-values in the
molecule were reported. Specific rotations were determined for
solutions by irradiating with the sodium D line ( = 589 nm) using
a Perkin Elmer 341 polarimeter: specific rotation, [a]D values are
given in units 10ꢁ1deg ꢀ cm2gꢁ1 where the concentration c is
given in g/100 mL. The chemical purity (Method A) was
2-Hydroxy-4, 6-bis(benzyloxy)acetophen-[13C]-one (3)
To preheated (internal temperatureꢃ351C) DMF (130mL) was
added 2,4,5-trihydroxyacetophenone (13.6 g, 72.84 mmol, 1 eq.)
and the solution was degassed for 15 min by bubbling with N2.
K2CO3 (22.1g, 160.2 mmol, 2.2eq.) was added and the internal
temperature rose toꢃ551C. To this was added, BnCl (20.28g,
160.2 mmol, 2.2 eq.) in one portion and the mixture was stirred at
55721C with stirring for 3–3.5h. The reaction mixture was cooled
to room temperature and the solids were suction filtered. The
filtrate was diluted with EtOAc (600mL) and washed with H2O
(2 ꢂ 250 mL). The combined aqueous layer was extracted with
EtOAc (1 ꢂ 100 mL). The organic layers were combined and
washed with 10% aqueous brine (4ꢂ 100 mL), dried (Na2SO4),
filtered and the solvent removed in vacuo to give the crude
product. The crude product was purified by silica gel chromato-
graphy using CH2Cl2/heptane (1/1, v/v) as the eluent to give 9.3 g
(33% yield) of 3 as an off-white solid. HPLC analysis using Method
A showed the product to be 99% pure with a retention time of
10.3 min. 1H NMR (300 MHz, CDCl3) d = 2.53 (d, J13C–H = 5.8 Hz,
3H), 5.02 (s, 4H), 6.1 (d, 1H, J = 1.4Hz), 6.22 (d, 1H, J = 2.3Hz),
7.2–7.55 (m, 10H), 14.02 (s, 1H). 13C NMR (75MHz, CDCl3) d = 203.2.
determined by standard HPLC (containing a PDA detector)
˚
A
using
a
Phenomenex Synergi 4 m Fusion-RP 80
(150 mm ꢂ 4.6 mm) column at wavelength of 280 nm, using a
gradient of 5–90% of acetonitrile (containing 0.01% TFA) with
water (containing 0.01% TFA) up to 20 min, the column
temperature was 251C, and the flow rate was 1 mL/min. The
enantiomeric purity (Method B) was determined by chiral HPLC
equipped with a PDA detector using a Chiralpak AD-RH 5 m
(150 mm ꢂ 4.6 mm) column. The solvent for isocratic programs
were acetonitrile/water (65/35, v/v) containing 0.01% TFA, run
time 40 min, detection wavelength was 210 nm, column
temperature was 601C, and the flow rate was 1 mL/min. All
the compounds were monitored against reference materials
including the starting materials.
4-Bromocatechol (5)
BBr3 (1 M solution in CH2Cl2, 130.3 mL, 130.3 mmol, 1.45 eq.) was
added slowly to 4-bromo veratrole (19.5 g, 89.86 mmol, 1 eq.)
with stirring at 01C under N2. The resulting mixture was heated
J. Label Compd. Radiopharm 2010, 53 605–612
Copyright r 2010 John Wiley & Sons, Ltd.
www.jlcr.org

P. K. Sharma et al.
OBn
OBn
OBn
OBn
H
O
H
O
CeCl3.7H2O
NaBH4
BnO
BnO
BnO
OH
7
, NaH,
*
*
THF/EtOH
DMF
*
*
*
OBn O
8
86%
84%
OBn
OBn O
9
3
OBn
OBn
OBn
OBn
TBDMS
TBDMS
O
TBDMSCl
Imidazole BnO
DMF
α
AD-mix-
BnO
nBu4NF
AcOH
O
MeSO2NH2
*
*
OH
88%
100%
88%
*
OH
*
OBn
OBn
11
10
OBn
OBn
OBn
OBn
H
BnO
O
RC(OEt)3
BnO
O
K2CO3
*
*
*
PPTS
100%
MeOH
OH
OH
82%
*
*
O
R
OBn
OBn
O
12
13a: R = H
13: R = Et
OBn
OBn
OBn
OBn
Dess-Martin
Periodinane
wet CH2Cl2
BnO
O
BnO
O
*
90%
*
Al(OiPr)3
2-Propanol
*
OH
O
74%
OBn
OBn
14
16
OBn
OBn
100%
(Overall 46%)
Pd(OH)2/C
EtOAC
BnO
O
*
*
OH
OBn
OH
17
HO
O
*
OH
77%
(Overall 24%)
Pd(OH)2/C
EtOAC
*
OH
OH
15
OH
HO
O
*
OH
*
OH
OH
18
Scheme 3. Synthesis of [2-13C, 4-13C]-(2R, 3S)-catechin 15 and [2-13C, 4-13C]-(2R, 3R)-epicatechin 18
at reflux for 18–20 h. The reaction mixture was cooled to (o51C) showed the product to be 98.3% pure with a retention time of
1
and H2O (15 mL) was added and stirred for 15 min. To this was 8.1 min. H NMR (300 MHz, CDCl3) d = 5.88 (br s, 1H), 6.5 (br s,
added, 1 N aqueous sodium hydroxide solution (220 mL). The 1H), 6.72 (d, 1H, J = 8.5 Hz), 6.9 (dd, 1H, J = 2.3, 8.5 Hz), 7.0 (d, 1H,
mixture was stirred for 1 h at room temperature (pH of the J = 2.3 Hz). Note: The crude material was used in the next step
solution was ꢃ9.0 as judged by pH paper). The reaction mixture without further purification, as it was unstable at room
was then acidified with 1 N hydrochloric acid (120 mL, pHꢃ0 to temperature. However, it was stable at–201C.
1). The reaction mixture was then extracted with tert-butyl
methyl ether (3 ꢂ 300 mL). The organic layers were combined
and washed with H2O (3 ꢂ 250 mL), brine (1 ꢂ 250 mL), dried
3, 4-Bis(benzyloxy)bromobenzene (6)
To an ice cold solution (o51C) of 5 (25.3 g, 89.86 mmol, 1 eq.) in
(Na2SO4), filtered and the solvent removed in vacuo to give
DMF (250 mL) was added K2CO3 (31 g, 224.64 mmol, 2.5 eq.) and
25.3 g (100% yield) of 5 as an oil. HPLC analysis using Method A
www.jlcr.org
Copyright r 2010 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2010, 53 605–612

P. K. Sharma et al.
stirred for 15 min. To this was added, BnBr (30.8 g, 180 mmol, were filtered and washed with CH3OH (2 ꢂ 50 mL) and dried
2 eq.) slowly and the reaction mixture was stirred at room under high vacuum at room temperature to give 9.71 g (86%
temperature for 1820 h. HPLC indicated the completion of the yield) of 8 as a yellow solid. HPLC analysis using Method A
reaction. The reaction –mixture was diluted with EtOAc (600 mL) showed the product to be 98% pure with a retention time of
and the solids were suction filtered. The filtrate was washed with 8.9 min. 1H NMR (300 MHz, CDCl3) d = 4.9 (s, 2H), 5.01 (s, 2H), 5.02
H2O (3 ꢂ 500 mL), brine (1 ꢂ 500 mL), dried (Na2SO4), filtered and (s, 2H), 5.2 (s, 2H), 6.1 (br s, 1H), 6.2 (d, 1H, J = 2.1 Hz), 6.6–6.7 (m,
the solvent removed in vacuo to furnish the crude product. The 1H), 6.72 (d, 1H, J = 8.4 Hz), 6.81 (dd, 1H, J = 1.6, 4.6 Hz), 7.1–7.5
crude product was triturated with EtOH (175 mL) at room (m, 20H), 7.6–8.0 (m, 1H), 14.4 (s, 1H). 13C NMR (75 MHz, CDCl3)
temperature with stirring for 2 h. The solids were filtered. The d = 142.7, 192.6.
filtered solids were again re-dissolved in hot EtOAc (25 mL) and
ethanol (75 mL). The solution was then cooled to room (E)-[1-13C, 3-13C]-2,4-Bis(benzyloxy)-2-(3-(3’, 4’-bis(benzylox-
y)phenyl)allyl)phenol (9)
temperature with stirring. The solids were suction filtered and
washed with EtOH (2 ꢂ 25 mL) and dried under high vacuum at
room temperature for 20 h to give 13.11 g (27% yield) of 6 as an
off-white solid. HPLC analysis using Method A showed the
product to be 98.8% pure with a retention time of 13.6 min. H
NMR (300 MHz, CDCl3) d = 5.1 (s, 4H), 6.78 (d, 1H, J = 5.8 Hz), 6.98
(dd, 1H, J = 2.3, 8.6 Hz), 7.05 (d, 1H, J = 8.6 Hz), 7.26–7.47 (m, 10H).
A solution of 8 (9.7g, 14.97 mmol, 1 eq). in THF (225mL) and EtOH
(45 mL) was cooled to 0–21C. Then, CeCl3 ꢀ 7H2O (13.94 g,
37.42 mmol, 2.5 eq). was added to the reaction mixture and
stirring continued until a clear solution was obtained. Solid NaBH4
(1.42 g, 37.42 mol, 2.5 eq.) was added in portions over a period of
1 h while keeping the internal temperaturer41C for an additional
1.5–2 h. (Note: The color of the reaction mixture turned to light
brown from light yellow and the evolution of hydrogen gas was
observed). HPLC indicated the consumption of the starting
material and the formation of the desired product (84% AUC)
and a by-product (14% AUC). The reaction mixture was quenched
with 5% aqueous citric acid solution (50 mL) while keeping the
internal temperatureo51C for 15min. (Note: The reaction mixture
solution turned turbid). The reaction mixture was diluted with
EtOAc (250mL) and the solids were removed by suction filtration.
The filtrate was washed with H2O (1 ꢂ 75mL), brine (1 ꢂ 100 mL),
dried (Na2SO4), filtered and the solvent removed in vacuo to
furnish the crude product. The crude product was triturated with
CH2Cl2/heptane (1/1, v/v) at 501C (bath temperature) for
10–15 min. The mixture was cooled to room temperature and
the solids were suction filtered to give crude 9. The combined
filtrate was concentrated in vacuo and purified by silica gel
chromatography using CH2Cl2/heptane (1/1–4/1, v/v) as the
eluent to give 8.01g (84% yield) of 9. HPLC analysis using Method
A showed the product to be 98.8% pure with a retention time of
1
3. 4-Bis(Benzyloxy)benz-[13C]-aldehyde (7)
In a flame dried flask under positive nitrogen atmosphere, was
added Mg (234 mg, 9.76 mmol, 1.24 eq.) in dry THF (50 mL) and
the resulting mixture was heated at 50731C (bath temperature).
To this, was slowly added a solution of 3, 4-bis(benzyloxy)-
bromobenzene (2.9 g, 7.86 mmol, 1 eq.) and 1,2-dibromoethane
(0.2 mL) in dry THF (10 mL). The bath temperature was raised to
6573 1C and maintained for 3–4h until all of the magnesium
was dissolved. The reaction mixture was cooled (o5 1C) and
then Me2N13CHO (1 g, 5.78 mmol, 0.74 eq.) was added and the
resulting reaction mixture stirred at this temperature for 1 h. The
reaction mixture was quenched with a cold solution (o5 1C) of
10% aqueous hydrochloric acid (15 mL) and extracted with
EtOAc (2 ꢂ 50 mL). The combined organic layers were washed
with brine (1 50 mL), dried (Na2SO4), filtered and the solvent
removed in vacuo to give the crude product. The crude product
was purified by silica gel chromatography using CH2Cl2/heptane
(1/1, v/v) as the eluent to furnish 1.3 g (77% yield) of 7 as an off-
white solid. HPLC analysis using Method A showed the product
to be 99.2% pure with a retention time of 14.4 min. 1H NMR
(300 MHz, CDCl3) d = 5.2 (s, 2H), 5.22 (s, 2H), 6.98 (d, 1H,
J = 8.2 Hz), 7.26–7.54 (m, 12H), 9.82 (d, J13C–H = 144 Hz, 1H). 13C
NMR (75 MHz, CDCl3) d = 190.8.
1
13.9min. H NMR (300MHz, CDCl3) d = 3.28–3.36 (m, 1H), 3.7–3.8
(m, 1H), 4.95 (s, 2H), 4.98 (s, 2H), 5.06 (s, 1H), 5.08 (s, 2H), 5.12 (s,
2H), 6.1 (d, 1H, J = 6 Hz), 6.14 (d, 1H, J = 2.1 Hz), 6.26 (d, 1H,
J = 2.1 Hz), 6.75–6.78 (m, 2H), 6.94 (d, 1H, J = 3.3 Hz), 7.26–7.5 (m,
20H). 13C NMR (75 MHz, CDCl3) d = 26.7, 128.6.
(E)-[1-13C, 3-13C)-(3, 5-Bis(benzyloxy)-2-(3-(30, 40-bis(benzy-
loxy)phenyl)allyl)phenoxy)(tert-butyldimethyl-silane (10)
(E)-1-(2, 4-Bis(benzyloxy)-6-hydroxyphenyl)-3-(3’, 4’-bis(ben-
zyloxy)phenyl)prop-2-[13C]en-1-[13C]-one (8)
To a solution of 9 (7.91 g, 12.48 mmol, 1 eq). and imidazole
(2.55 g, 37.44 mmol, 1 eq.) in DMF (60 mL) at room temperature
was added TBDMSCl (3.76 g, 24.95 mmol, 2 eq). under N2. The
resulting solution was stirred at room temperature for 20 h.
Water (150 mL) was added to the reaction mixture. The reaction
mixture was extracted with EtOAc (2 ꢂ 200 mL). The organic
layers were combined and washed with H2O (2 ꢂ 100 mL), brine
(1 ꢂ 100 mL), dried (Na2SO4), filtered and the solvent removed in
vacuo to give the crude product. The crude product was purified
by silica gel chromatography using CH2Cl2/heptane (1/1–4/1,
v/v) as the eluent to give 9.88 g (100% yield) of 10 as a viscous
oil. 1H NMR (300 MHz, CDCl3) d = 0.22 (s, 6H), 1.02 (s, 9H), 3.32 (t,
1H, 6 Hz), 3.78 (t, 1H, J = 6 Hz), 5.04 (s, 2H), 5.06 (s, 2H), 5.16 (s,
2H), 5.18 (s, 2H), 6.5 (m, 1H), 6.21 (m, 2H), 6.35 (d, 1H, J = 2.1 Hz),
6.78–6.9 (m, 2H), 6.94–7.0 (m, 1H), 7.28–7.5 (m, 20H). 13C NMR
(75 MHz, CDCl3) d = 26.8, 128.6.
A suspension of NaH (60% dispersion in oil, 0.845 g, 21.13 mmol,
1.2 eq.) in DMF (20 mL) was cooled in an ice bath (ꢃ51C) under
N2. To this was added a solution of 3 (6.13 g, 17.61 mmol, 1 eq.)
in DMF (20 mL). The reaction mixture was stirred for 0.5 h at this
temperature. Then, a solution of 7 (5.6 g, 17.61 mmol, 1 eq.) in
DMF (25 mL) was slowly added. The cooling bath was removed
and the resulting reaction mixture was allowed to warm to room
temperature and stirred for 3.5 4 h. The reaction mixture was
poured into a mixture of ice (100 g) and 1N HCl (50 mL) and
stirred for 15 min. The solids were suction filtered and washed
with CH3OH (2 ꢂ 50 mL). The solids were dissolved in EtOAc
(250 mL) and washed with H2O (1 ꢂ 100 mL), brine (1 ꢂ 100 mL),
dried (Na2SO4), filtered and the solvent removed in vacuo to give
the crude product. The crude product was triturated with
CH3OH (100 mL) at room temperature for 15 min. The solids
J. Label Compd. Radiopharm 2010, 53 605–612
Copyright r 2010 John Wiley & Sons, Ltd.
www.jlcr.org

P. K. Sharma et al.
(1S, 2S)-[1-13C, 3-13C]-3-(2, 4-bis(benzyloxy)-6-(tert-butyldi-
methylsilyloxy)phenyl)-1-(30,40-bis(benzyloxy)-phenyl)pro-
pane-1, 2-diol (11)
temperature) for 5 h. The reaction mixture was cooled to room
temperature and passed through a silica gel plug. The silica gel
plug was eluted with CH2Cl2 (4 ꢂ 50 mL). The combined filtrate
was concentrated in vacuo to give 6.97 g (100% yield) of 13 as
an oil. HPLC analysis using Method A showed the product to be
92.3% pure with a retention time of 14.2 min. The ee was
determined using chiral HPLC analysis (Method B) and was
found to be 98% ee. [a]D25 = 129.5 [c 1, Acetone]. 1H NMR
A cold solution (0–21C) of tert-butanol (130 mL), H2O (130 mL)
and AD-mix-a (48.9 g, 5 eq. by weight) was added to a solution
of 10 (9.78 g, 13.07 mmol, 1 eq.) in THF (150 mL) under N2. The
internal temperature rose to 101C. The reaction mixture was
then cooled to 01C and methanesulfonamide (1.62 g, 17 mmol,
1.3 eq.) was added; followed by stirring forꢃ42 h at 0–21C. 10%
Aqueous sodium bisulfite solution (w/v, 150 mL) was added
slowly with stirring and the reaction mixture was allowed to
warm to room temperature. The reaction mixture was diluted
with EtOAc (200 mL) and H2O (200 mL). The organic layer was
separated. The aqueous layer was extracted with EtOAc
(1 ꢂ 200 mL). The organic layers were combined and washed
with H2O (1 ꢂ 150 mL), brine (1 ꢂ 150 mL), dried (Na2SO4),
filtered and the solvent removed in vacuo to give the crude
product. The crude product was purified by silica gel
chromatography using EtOAc/heptane (2/8, v/v) as the eluent
to give 9 g (88% yield) of 11 as an off-white solid. HPLC analysis
using Method A showed the product to be 100% pure with a
retention time of 17.9 min. 1H NMR (300 MHz, CDCl3) d = 0.18
(s, 6H), 0.78 (s, 9H), 1.98 (br s, 1H, OH), 2.4 (d, 2H, J = 3.5 Hz), 2.82
(d, 1H, J = 5.6 Hz), 3.03 (t, 1H, J = 2.6 Hz), 3.6–3.76 (m, 1H), 4.8
(s, 2H), 4.82 (s, 2H), 4.93 (s, 2H), 4.97 (s, 2H), 5.95 (d, 1H, J = 2 Hz),
6.14 (d, 1H, J = 2 Hz), 6.68 (s, 2H), 6.83 (d, 1H, J = 3.2 Hz), 7.08–7.3
(m, 20H). 13C NMR (75 MHz, CDCl3) d = 27.2, 76.8.
(300 MHz, CDCl3) d = 0.95 (t, 3H, J = 7.6 Hz), 2.1 (q, 2H, J = 7.6 Hz),
C–H
13
4.69 (d, J13C–H = 15 Hz), 5.2 (d, J = 30 Hz, 1H), 5.0 (s, 2H), 5.08
(s, 2H), 5.12 (s, 2H), 5.26–5.38 (m, 1H), 6.25 (dd, 1H, J = 2 8 Hz),
6.88 (br s, 2H), 6.95 (d,1H, J = 3.7 Hz), 7.2–7.5 (m, 22H), 7.7–7.83
(m, 1H).13C NMR (75 MHz, CDCl3) d = 24.2, 78.4.
5,7,30,40-Tetra-O-benzyl-[2-13C, 4-13C]-(2R,3S)-catechin (14)
To a solution of 13 (8.9 g, 12.08 mol, 1 eq.) in CH2Cl2 (100 mL)
and CH3OH (50 mL) was added K2CO3 (1.63 g, 16.63 mol, 1.5 eq.)
and the resulting suspension was stirred at room temperature
for 6–8 h. The reaction mixture was concentrated in vacuo
keeping the bath temperaturer261C. The residue was dissolved
in CH2Cl2 (100 mL) and washed with H2O (1 ꢂ 50 mL). The
organic layer was passed through a silica gel plug (150 g). The
silica gel plug was washed with CH2Cl2 (2 ꢂ 50 mL). The filtrates
were combined and the solvent was removed in vacuo to give
an off-white solid (ꢃ14 g). The solid was dissolved in hot
toluene (250 mL) atꢃ451C (bath temperature). The solution was
allowed to cool to room temperature and kept at room
temperature for 65–72 h. The solid obtained was suction filtered.
The solids were dried under high vacuum at room temperature
for 20 h to give 7.3 g (82% yield) of 14 as an off-white solid. HPLC
analysis using Method A showed the product to be 99.8% pure
with a retention time of 12.9 min. The ee was determined using
chiral HPLC analysis (Method B) and was found to be 97% ee.
[a]2D5 = ꢁ59.8 [c 1, Acetone]. 1H NMR (300 MHz, CDCl3) d = 1.6
(t, 1H, J = 4.5 Hz), 2.33–2.48 and 3.22–3.4 (each m, 2H,
J13C–H = 54 MHz), 2.76–2.95 (m, 1H), 3.97 (br s, 1H), 4.32 and
4.86 (each dd, 1H, J = 2.6, 8.2 Hz, J13C–H = 300 MHz), 5.02 (s, 2H),
5.08 (s, 2H), 5.12 (s, 4H), 6.22 (d, 1H, J = 2 Hz), 6.28 (d,1H, J = 2 Hz),
6.95 (s, 2H), 7.04 (d, 1H, J = 3.6 Hz), 7.1–7.5 (m, 18H). 13C NMR
(75 MHz, CDCl3) d = 28.3, 81.6.
(1R, 2R)-[1-13C, 3-13C]-3-(2, 4-bis(benzyloxy)-6-hydroxy phe-
nyl)-1-(30, 40-bis(benzyloxy)-phenyl)propane-1, 2-diol (12)
To an ice cold solution (0–51C) of 11 (8.9 g, 11.35 mmol, 1 eq.) in
THF (50 mL) was slowly added a premixed solution of nBu4NF
(1M solution in THF, 22.7 mL, 22.7 mol, 2 eq.) and glacial AcOH
(1.36 g, 22.7 mol, 2 eq.) under N2. The resulting reaction mixture
was stirred at this temperature for 0.5 h as HPLC indicated the
completion of the reaction. The solvent was removed in vacuo.
The residue was dissolved in CH2Cl2 (100 mL) and washed with
H2O (1 ꢂ 100 mL), brine (1 ꢂ 100 mL), dried (Na2SO4), filtered and
the solvent removed in vacuo to give the crude product. The
crude product was triturated with EtOAc/heptane (1/9, v/v,
100 mL) at room temperature for 0.5 h. The solids were suction
filtered, washed with cold EtOAc/heptane (ꢃ51C, 1/9, v/v,
2 ꢂ 25 mL), and dried under high vacuum at room temperature
for 18 h to give 6.7 g (88% yield) of 12 as an off-white solid. HPLC
analysis using Method A showed the product to be 99.9% pure
with a retention time of 8.7 min. The ee was determined using
chiral HPLC analysis (Method B) and was found to be 98.6% ee.
[a]2D5 = ꢁ21.6 [c 1, Acetone]. 1H NMR (300 MHz, CDCl3) d = 3.74
(br s, 2H), 4.08 (br s, 1H), 4.52 (br s, 1H), 4.86 and 5.1 (each br s,
1H), 4.95 (s, 2H), 4.98 (s, 2H), 5.03 (s, 2H), 5.07 (s, 2H), 6.1 (d, 1H,
J = 1.7 Hz), 6.21 (d, 1H, J = 1.7 Hz), 6.78 (d, 1H, J = 7.3 Hz), 6.96 (d,
1H, J = 8.3 Hz), 7.04 (br s, 1H), 7.2–7.5 (m, 20H), 9.3 (s, 1H). 13C
NMR (75 MHz, CDCl3) d = 26.9, 75.5 (d, J = 7.5 Hz).
[2-13C, 4-13C]-(2R,3S)-catechin (15)
To a solution of 14 (350 mg, 0.54 mmol) in EtOAc (5 mL) was
added 20% Pd(OH)2/C (50% wet, 70 mg, 20 weight %). The
resulting suspension was stirred at RT under H2 atmosphere
using a balloon forꢃ2 h. The reaction mixture was filtered
through a 0.45-micron filter. The filter was washed with EtOAc
(5 mL). The filtrates were combined and the solvent was
removed in vacuo. The residue was dissolved in EtOAc (2 mL)
and diluted with H2O (HPLC grade, 15 mL) and lyophilized to
give 160 mg (100% yield) of 15 as an off-white solid. HPLC
analysis using Method A showed the product to be 100% pure
with a retention time of 8.7 min. The ee was determined using
chiral HPLC analysis (Method B) and was found to be 99% ee.
[a]2D5 = 136.6 [c 1, Acetone]. 1H NMR (300 MHz, Acetone-d6)
d = 2.62–2.8 (m, 2H), 3.06–3.2 (m, 1H), 3.85–3.95 (m, 1H), 3.96–4.1
[2-13C, 4-13C]-3-O-propiolate ester-5,7,30,40-tetra-O-benzyl-
(2S,3S)-catechin (13)
To
a suspension of 12 (6.6 g, 9.85 mmol, 1 eq.) in 1,2- (m, 1H), 4.28–4.37 and 4.75–4.85 (each m, 1H), 5.84 (d, 1H,
dichloroethane (130 mL) was added EtC(OEt)3 (3.13 g, J = 2.4 Hz), 6.02 (d, 1H, J = 2.4 Hz), 6.7–6.83 (m, 1H), 6.85–6.92 (m,
17.73 mmol, 1.8 eq.) and PPTS (1.34 g, 5.32 mmol, 0.54 eq.) and 1H), 7.84 (br s, 1H), 7.95 (br s, 1H), 8.13 (br s, 1H). 13C NMR
the resulting reaction mixture was heated atꢃ651C (bath (75 MHz, CDCl3) d = 29.3, 82.8. MS = 293.1 [M11H, 99%].
www.jlcr.org
Copyright r 2010 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2010, 53 605–612

P. K. Sharma et al.
(2R)-[2-13C, 4-13C]-5, 7-bis(benzyloxy)-2-(30, 40-bis(benzyl-
oxy)chroman-3-one (16)
white fluffy solid. HPLC analysis using Method A showed the
product to be 98.5% pure with a retention time of 9.2 min. The
ee was determined using chiral HPLC analysis (Method B) and
was found to be 98.7% ee. [a]2D5 = -33.9 [c 1, Acetone]. H NMR
1
To a solution of 14 (2.3 g, 3.53 mmol, 1 eq.) in CH2Cl2 (50 mL) was
added DMP reagent (1.75 g, 4.13 mmol, 1.17 eq.) under N2 at
room temperature. Water (0.1 mL) was added to the reaction
mixture which was continuously stirred at room temperature for
2 h. The reaction mixture was quenched with saturated aqueous
NaHCO3 and the pH was adjusted to ꢃ8 (pH paper). The layers
were separated. The aqueous layer was extracted with CH2Cl2
(2 ꢂ 50 mL). The organic layers were combined and washed with
H2O (1 ꢂ 50 mL), and brine (1 ꢂ 50 mL). The organic layer was
passed through a silica gel plug (1.5’’ diameter) and the plug
was washed with CH2Cl2 (4 ꢂ 50 mL). The combined filtrates
were concentrated in vacuo to give 2.06 g (90% yield) of 16 as
an off-white solid. HPLC analysis using Method A showed the
product to be 97% pure with a retention time of 9.9 min. 1H
NMR (300 MHz, CDCl3) d = 3.3 (dd, 1H, J = 21.5, 48 Hz), 3.74 (dd,
1H, J = 21.5, 48 Hz), 4.96–5.05 (m, 5H), 5.08 (s, 2H), 5.11 (s, 2H),
6.33 (d, 1H, J = 2.5 Hz), 6.82 (br s, 2H), 6.94 (d, 1H, J = 2.5 Hz),
7.27–7.52 (m, 21H). 13C NMR (75 MHz, CDCl3) d = 33.6
(d, J = 7.5 Hz), 83.0 (d, J = 7.5 Hz).
(300 MHz, Acetone-d6) d = 2.42–2.74 (m, 1H), 2.88–3.16 (m, 1H),
3.47–3.68 (m, 1H), 4.2 (br s, 1H), 4.66 (br s, 1H), 5.12 (br s, 1H), 5.9
(d, 1H, J = 2.1 Hz), 6.02 (br s, 1H, J = 2.1 Hz), 6.75–6.88 (m, 2H),
7.0–7.08 (m, 1H), 7.72 (br s, 1H), 7.9 (br s, 1H), 8.06 (br s, 1H). 13
C
NMR (75 MHz, Acetone-d6) d = 28.8, 79.7 (d, J = 7.5 Hz).
MS = 293.1 [M111, 99%].
Conclusions
The synthesis of doubly labeled [2-13C, 4-13C]-(2R,3S)-catechin
15 and [2-13C, 4-13C]-(2R,3R)-epicatechin 18 were achieved for
the first time in 46 and 24% overall yield, respectively, with high
chemical purity and ee in milligram quantities. The incorporation
of labeled 13C at the C-2 and C-4 positions was accomplished
using commercially available acetonitrile (CH313CN) and N, N-
dimethylformaide (Me2N13CHO). The reduction of chalcone 8 to
alkene 9 was accomplished under Luche conditions resulting in
the desired alkene 9 in excellent yield. Use of glacial AcOH with
nBu4NF for the deprotection of the TBDMS group was
accomplished and it avoided isomerization at the C-2 position.
Replacement of triethylorthoformate with triethylorthopropio-
nate for the synthesis of 13 from 12 resulted in shorter reaction
time along with minimal isomerization (o2 %) either at C-2 and/
or C-3 positions as determined by 1H NMR and chiral HPLC.26
The oxidation of the C-3 hydroxyl group of 14 was performed
with wet DMP reagent. The presence of a small amount of water
(ꢃ0.1–0.2 mL) in the reaction mixture allowed the reaction to
proceed with ease of operation and complete conversion. The
synthetic methodology developed here could not only be
applicable for the synthesis of the corresponding radiolabeled
analogs, but also applied to obtain gram quantities of other
epimers of 15 and 18, as well as higher oligomers.
5,7,30,40-tetra-O-benzyl-[2-13C, 4-13C]-(2R,3R)-epicatechin (17)
A suspension of 16 (2.3 g, 3.54mmol, 1 eq.), Al(OiPr)3 (1.45 g,
7.08 mmol, 2 eq.) and IPA (18mL) in toluene (60mL) was heated at
110–1151C (bath temperature) using a Dean–Stark unit (to remove
the acetone formed during the reaction in order to complete the
reaction) under N2 with stirring forꢃ1.5–2 h. The reaction mixture
was cooled to room temperature, poured into ice (100g) and the
pH of the mixture was adjusted toꢃ1 (pH paper) with 1 N H2SO4.
The layers were separated. The aqueous layer was extracted with
EtOAc (1 ꢂ 50mL). The organic layers were combined and washed
with H2O (1ꢂ 100mL), dried (Na2SO4), filtered and the solvent
removed in vacuo to give the crude product. The crude product
was purified by silica gel chromatography (75g) using CH2Cl2/
heptane/EtOAc (7/7/1, v/v/v) as the eluent. The fractions contain-
ing the desired compound were combined and the solvent was
removed in vacuo to give the desired compound (17, 95% ee as
judged by chiral HPLC Method B).
The product was further purified by dissolving the compound
in CH2Cl2 (10 mL) followed by the addition of CH3OH (20 mL).
The solution was allowed to stand at ambient temperature
forꢃ0.5 h. The solids were suction filtered and washed with cold
(o51C) CH3OH (2 ꢂ 3 mL) and dried under high vacuum at room
temperature to give 1.7 g (74% yield) of 17 as an off-white solid.
HPLC analysis using Method A showed the product to be 100%
pure with a retention time of 12.8 min. The ee was determined
Acknowledgements
The authors thank Drs Heather Taft, Richard Lombardy and
Daniel J. Coughlin of JMPM and Harold H. Schmitz of MARS
Incorporated for their constant encouragement and Ms Yanni
Gou for analytical method development. The authors are
grateful to the reviewers for their helpful suggestions.
using chiral HPLC analysis (Method B) and was found to be References
98.7% ee. [a]2D5 = 19.8 [c 1, Acetone]. H NMR (300 MHz, CDCl3)
1
d = 1.68 (br s, 1H), 2.66–2.8 (m, 1H), 3.08–3.27 (m, 1H), 4.2 (br s,
1H), 5.01 (s, 4H), 5.1–5.23 (m, 5H), 6.28 (s, 2H), 6.92–7.08 (m, 2H),
7.15 (br s, 1H), 7.24–7.53 (m, 20H). 13C NMR (75 MHz, CDCl3)
d = 28.3, 78.4.
[1] N. D. Fisher, M. Hughes, M. Gerhard-Herman, N. K. Hollenberg,
J. Hypertens. 2003, 21, 2281–2286.
[2] a) D. Taubert, R. Berkels, R. Roesen, W. Klaus, J. Am. Med. Assoc.
2003, 290, 1029–1030; b) H. Schroeter, C. Heiss, J. Balzer,
P. Kleinbongard, C. L. Keen, N. K. Hollenberg, H. Sies, Kwick-K.
Uribe, H. H. Schmitz, M. Kelm, PNAS. 2006, 103, 1024–1029.
[3] A. A. Shahat, S. I. Ismail, F. M. Hammouda, S. A. Azzam, G. Lemiere,
T. De Bruyne, S. De Swaet, I. Piteters, A. Vlietinck, Phytomedicine
1998, 5, 133–136.
[4] a) M. Aviram, B. Fuhrman, Artherosclerosis 1998, 137, S45–S50;
b) J. Yamakoshi, S. Kataoka, T. Koga, T. Ariga, Artherosclerosis
1999, 142, 139–149
[2-13C, 4-13C]-(2R,3R)-epicatechin (18)
A suspension of 17 (1.65 g, 2.53 mol) and 20% palladium
hydroxide (50% wet, 0.33 g, 20% by weight) in ethyl acetate
(HPLC grade, 25 mL) was stirred under hydrogen atmosphere
using a balloon at room temperature for 1 h. Following the
workup similar to 15 provided 573 mg (77% yield) of 18 as a
[5] M. Saito, H. Hosoyama, T. Ariga, S. Kataoka, N. Yamaji, J. Agric.
Food Chem. 1998, 46, 1460–1467.
J. Label Compd. Radiopharm 2010, 53 605–612
Copyright r 2010 John Wiley & Sons, Ltd.
www.jlcr.org

P. K. Sharma et al.
[6] a) L. Liviero, P. P. Puglisis, P. Marazzoni, E. Bombardelli, Fitoterapia
1994, 65, 203–209; b) A. Constable, N. Varga, J. Richoz,
R. H. Stadler, Mutagenesis 1996, 11, 189–194; c) O. Inanmi,
Y. Watanabe, B. Syuto, M. Nakano, M. Kuwabara, Free Radic. Res.
1998, 29, 359–365.
Dispos. 2002, 30, 897–903; e) G. Van’t Slot, H. U. Humpf, J. Agric.
Food Chem. 2009, 57, 8041–8048.
`
[17] a) M-C Pierre, C. Cheze, J. Vercauteren, Tetrahedron. Lett. 1997, 32,
5639–5642; b) S. Rasku, K. Wa¨ha¨la¨, Tetrahedron 2000, 56,
913–916.
´
[7] O. Inanami, Y. Watanabe, B. Syuto, M. Nakano, M. Kuwabara, Free
Radic. Res. 1998, 29, 359–365.
[8] C. Hartisch, H. Kolodziej, F. von Bruchhausen, Planta Med. 1977,
63, 106–110.
[9] A. Scalbert, Phytochemistry 1991, 30, 3875–3883.
[18] a) S. Deprez, I. Mila, A. Scalbert, J. Agric. Food Chem. 1999, 47,
4219–4230; b) J. Laparra, J. Michaud, M. F. Lesca, P. Blanquest,
J. Masquelier, Bull. Soc. Pharm. Bordeaux 1977, 116, 17–23;
c) S. Krisa, P. Waffo Teguo, A. Decendit, G. Deffieux,
J. Vercauteren, J-M. Merillon, Phytochemistry 1999, 51, 651–656.
[19] a) B. Nay, V. Arnaudinaud, J-F. Peyrat, A. Nuhrich, G. Deffieux,
J-M. Merillon, J. Vercauteren, Eur. J. Org. Chem. 2000, 1279–1283; b)
H. van Rensburg, P. S. van Heerden, B. C. B. Bezuidenhoudt,
D. Ferreira, Tetrahedron Letts 1997, 38, 3089–3092; c) H. van
Rensburg, P. S. van Heerden, D. Ferreira, J. Chem. Soc. Perkin Trans I.
1997, 3415–3421; d) B. Nay, V. Arnaudinaud, J. Vercauteren, Chimie
2002, 5, 517–523; e) L. Claisen, A. Claparede, Chem. Ber. 1881, 14,
2460–2468; f) J. G. Schmidt, Chem. Ber. 1881, 14, 1459–1465.
[20] C. Tsang, C. Auger, B. Mullen, A. Bornet, J. M. Rouanet, A. Crozier,
4th Tannin Conference, 228th ACS National Meeting, Philadel-
phia, PA, August 22–26, 2004.
[21] L. Y. Rios, R. N. Bennet, S. A. Lazarus, C. Remsesy, A. Scalbert,
G. Williamson, Am. J. Clin. Nutr. 2002, 76, 1106–1110.
[22] S. Deprez, I. Mila, J. F. Huneau, D. Tome, A. Scalbert, Antiox. Redox
Signal. 2001, 6, 957–967.
[23] a) J. Polster, H. Dithmar, R. Burgemeister, G. Friedman, W. Feucht,
Physiol. Plant. 2006, 128, 163–174; b) J. Polster, H. Dithmar,
F. Watter, Biol. Chem. 2003, 384, 997–1008; c) W. Feucht,
D. Treatter, H. Dithmar, J. Polster, Tree Physiol. 2008, 28,
1783–1791; d) W. Feucht, D. Treutter, J. Polster, Plant Cell Rep.
2004, 22, 430–436.
[10] K. Kameda, T. Takau, H. Okuda, Y. Kimura, T. Okuda, T. Hatano,
I. Agara, S. Arichi, J. Nat. Prod. 1987, 50, 680–683.
[11] a) F. V. So, N. Guthrie, A. F. Chambers, K. K. Carroll, Cancer
Lett. 1997, 112, 127–133; b) H. Kolodziej, C. Haberland,
H. Woerdenbag, A. W. T. Konigs, Phytother. Res. 1995, 9,
410–415; c) D. Ramljak, L. J. Romanczyk Jr, L. J. Metheny-Barlow,
N. Thompson, V. Knezevic, M. Galperin, A. Ramesh, R. B. Dickson,
Mol. Cancer Ther. 2005, 4, 537–546.
[12] a) G. M. Polya, B. H. Wang, L. Y. Foo, Phytochemistry 1995, 38,
307–315; b) T. K. Mao, J. van der Water, C. L. Keen, H. H. Schmitz,
M. E. Gerschwin, Dev. Immunol 2002, 9, 135–141; c) Y. C. Park,
G. Rimbach, C. Saliou, G. Valucchi, L. Parker, FEBS Lett 2000, 465,
93–97; d) T. P. Kenny, C. L. Keen, P. Jones, H-J. Kung,
H. H. Schmitz, M. E. Gerschwin, Exp. Biol. Med 2004, 229,
255–263; e) T. K. Mao, J. J. Powell, J. van der Water, C. L. Keen,
H. H. Schmitz, M. E. Gerschwin, Int. J. Immunol 1999, 15, 23–29.
[13] a) C. Agarwal, R. P. Singh, R. Agarwal, Carcinogenesis 2002, 23,
1869–1876; b) S. V. Verstraeten, J. F. Hammerstone, C. L. Keen,
C. G. Fraga, P. I. Oteiza, J. Agric. Food Chem. 2005, 53, 5041–5048;
c) F. Ollila, K. Halling, P. Vuorela, J. P. Slotte, Arch. Biochem.
Biophys 2002, 399, 103–108; d) S. V. Veratraeten, C. L. Keen,
H. H. Schmitz, C. G. Fraga, P. I. Oteiza, Free Rad. Biol. Med. 2003,
34, 84–92.
´
´
[24] a) W. Tu¨ckmantel, A. P. Kozikowski, L. J. Romanczyk Jr, J. Am.
Chem. Soc. 1999, 121, 12073–12081; b) A. P. Kozikowski,
¨
[14] a) H. P. Ciolino, P. J. Daschner, G. C. Yeh, Biochem. J. 1999, 340,
715–722; b) M. Zhu, J. D. Phillipson, P. M. Greengrass,
N. E. Bowery, Y. Cai, Phytochemistry 1997, 44, 441–447.
[15] a) P. C. Hollman, M. B. Katan, Biomed. Pharmacother. 1997, 51,
305–310; b) C. Rice-Evans, J. P. E. Spencer, H. Schroeter,
A. R. Rechner, Drug Metab. Drug Interact. 2000, 17, 291–298;
c) P. C. Hollman, M. B. Katan, Free Rad. Res. 1999, 31, S75–S86;
d) P. C. Hollman, M. B. Katan, Arch. Toxicol. 1998, 20, 237–248;
e) J. P. E. Spencer, H. Schroeter, A. Rechner, C. Rice-Evans, Antiox.
Redox, Signal. 2001, 3, 1023–1039; f) T. P. Mulder, A. G. Rietveld,
J. M. van Amelsvoort, Am. J. Clin. Nutr. 2005, 81, S565–S575.
[16] a) A. Scalbert, G. Williamson, J. Nutr. 2000, 130, 20735–20855;
b) K. Okushio, M. Suzuki, N. Matsumoto, F. Nanjo, Y. Hara, Biosci.
Biotech. Biochem. 1999, 63, 430–432; c) K. Okushio, M. Suzuki,
N. Matsumoto, F. Nanjo, Y. Hara, Drug Metab. Drug Dispos. 1999,
27, 309–316; d) J. B. Vaidyanathan, T. Walle, Drug Metab. Drug
W. Tu¨ckmantel, G. Bottcher, L. J. Romanczyk Jr, J. Org. Chem.
2003, 68, 1641–1658; c) P. K. Sharma, A. Kolchinski, H. A. Shea,
J. J. Nair, Y. Gou, L. J. Romanczyk Jr, H. H. Schmitz, Org. Proc. Res.
Dev. 2007, 11, 422–430; d) L. J. Romanczyk Jr, P. K. Sharma,
D. Gou, Y. Gou, PCT Patent Application WO2007002877 A1
(Jan 4, 2007).
[25] a) D-Y. Xie, R. A. Dixon, Phytochemistry 2005, 66, 2127–2144;
b) D. Ferreira, D. Slade, Nat. Prod. Rep. 2002, 19, 517–541.
[26] L. Li, T. H. Chan, Org. Lett. 2001, 3, 739–741.
[27] N. Minami, S. Kijima, Chem. Pharm. Bull. 1979, 27, 1490–1494.
[28] S. D. Meyer, S. L. Schreiber, J. Org. Chem. 1994, 59,
7549–7552.
[29] a) J. Zhang, K. Takahashi, Y. Kono, Y. Suzuki, S. Takeuchi,
T. Shimizu, I. Yamaguchi, M. Chijimatsu, A. Sakurai, Y. Sato,
H. Kitamura, J. Pesticide Sci. 1990, 15, 585–591; b) H. Wang,
K. Helliwell, Food Chem. 2000, 70, 337–344.
www.jlcr.org
Copyright r 2010 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2010, 53 605–612