An expeditious synthesis of quercetin 3-O-β-d-glucuronide from rutin

Article abstract of DOI:10.1016/j.tetlet.2011.06.108

We report the synthesis of the major human metabolite of quercetin, quercetin 3-O-β-d-glucuronide, from rutin (quercetin-3-rutinoside), which is commercially available at low cost. This straightforward synthesis is based on the key intermediate 3′,4′,5,7-tetra-O-benzyl-quercetin which is obtained in only two steps by the total benzylation of rutin followed by acid hydrolysis of the rutinoside residue. Glycosylation of the free 3 hydroxyl group by 1-bromo-3,4,6-tetra-O-acetyl-α-d-glucopyranoside yields the protected glucoside. TEMPO-mediated oxidation of primary alcohol on the deprotected glucoside gives access to the benzylated glucuronide. Removal of the benzyl groups which protect the quercetin hydroxyl groups by H₂ (10% Pd/C) yields quercetin 3-O-β-d-glucuronide.

Full text of DOI:10.1016/j.tetlet.2011.06.108

Tetrahedron Letters 52 (2011) 4738–4740
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
An expeditious synthesis of quercetin 3-O-b-D-glucuronide from rutin
⇑
Mohammed Kajjout, Rajae Zemmouri, Christian Rolando
Université de Lille 1, Sciences et Technologies, USR CNRS 3290 Miniaturisation pour la synthèse, l’Analyse & la Protéomique, 59655 Villeneuve d’Ascq Cedex, France
a r t i c l e i n f o
a b s t r a c t
Article history:
We report the synthesis of the major human metabolite of quercetin, quercetin 3-O-b-D-glucuronide,
Received 20 May 2011
Revised 25 June 2011
Accepted 27 June 2011
Available online 2 July 2011
from rutin (quercetin-3-rutinoside), which is commercially available at low cost. This straightforward
synthesis is based on the key intermediate 30,40,5,7-tetra-O-benzyl-quercetin which is obtained in only
two steps by the total benzylation of rutin followed by acid hydrolysis of the rutinoside residue. Glyco-
sylation of the free 3 hydroxyl group by 1-bromo-3,4,6-tetra-O-acetyl-a-D-glucopyranoside yields the
protected glucoside. TEMPO-mediated oxidation of primary alcohol on the deprotected glucoside gives
access to the benzylated glucuronide. Removal of the benzyl groups which protect the quercetin hydroxyl
groups by H2 (10% Pd/C) yields quercetin 3-O-b-D-glucuronide.
Keywords:
Rutin (quercetin-3-rutinoside)
Quercetin 3-O-b-
TEMPO oxidation
D-glucuronide
Ó 2011 Elsevier Ltd. All rights reserved.
Flavonoids are the most important family of plant polyphenolic
compounds and are diverse both in chemical structures and charac-
teristics. Flavonoids are categorized into flavonols, flavones, flava-
nones, isoflavones, catechins, anthocyanidins, dihydroflavonols,
and chalcones. Quercetin is the major individual flavonol in fruits,
vegetables, and beverages and the human daily consumption ranges
between 10 and 100 mg according to eating and drinking habits.1–4
Quercetin is an antioxidant and appears to be active in many dis-
eases related to aging like cardiovascular,5 cancer6,7 and neurode-
generative diseases.8 Quercetin glucuronides and sulfates are the
main circulating metabolites of quercetin along with methylated
quercetins and their glucuronide and sulfate conjugates.9 In humans
bromide in the presence of silver oxide.13 This synthesis has been
improved by Needs et al. using a desiccant (either calcium sulfate
or molecular sieves) in pyridine at 0 °C, which prevents scrambling
of the acetate group protecting the glucuronate residue during the
alkylation step. After debenzylation and hydrolysis of the acetate
protecting the sugar secondary alcohols and of the methyl ester
protecting the carboxylic acid, quercetin 3-O-b-
been obtained in 11% yield from quercetin.14 The preparation of
quercetin-3-O-b- -glucuronide has also been previously described
D-glucuronide has
D
in our group in modest yield (25% from protected quercetin) by
selective oxidation of the glucoside by NaOCl/TEMPO in the pres-
ence of potassium bromide and tetrabutylammonium bro-
quercetin 3-O-b-
most abundant quercetin metabolites and have even more potent
biological activities than quercetin itself.10 Quercetin 3-O-b-
-glu-
D
-glucuronide and quercetin 30-O-sulfate are the
mide.15,16 The required 3-O-b-
D glucoside quercetin protected at
all phenolic positions was obtained in five steps. First the quercetin
B catechol ring was protected by neat heating at 180 °C with dic-
hlorodiphenylmethane in low yield. The intermediate was glycos-
ylated under phase transfer conditions at the more reactive 3
position. Before TEMPO oxidation the 5 and 7 positions were pro-
tected by benzylation and the glucose moiety was deprotected
using sodium methylate. The deprotection of the ketal was more
difficult than expected and the diphenylmethylene group required
a strong catalyst such as 30% palladium on charcoal.
D
curonide, which is not commercially available for biological studies,
has been isolated from green beans11 and from Reynoutria sachialin-
ensis.12 We developed a novel synthesis of quercetin 3-O-b-
D-glucu-
ronide from rutin. Our approach is based on the protection of all
hydroxyl groups and mild hydrolysis of the 3-O-rutinoside natural
protecting group, leading to 30,40,5,7-tetrabenzylquercetin. Glyco-
sylation of the free hydroxyl at the 3 position, oxidation of the
primary alcohol of the glucoside catalyzed by TEMPO (2,2,6,6-tetra-
methyl-1-piperidinyloxy) and the deprotection of hydroxyl groups
Our present method provides an efficient and easy approach to
access of quercetin 3-O-b-D-glucuronide 5 in just four steps from ru-
gave quercetin 3-O-b-
straightforward access to quercetin functionalized at the position
3 via the key intermediate 2.
D
-glucuronide. This synthetic pathway gives
tin as depicted in Scheme 1, taking advantage of the natural protec-
tion of the 3 position in rutin which is readily available and cheap.
Huang et al.17 obtained 30,40,7 tribenzyl-quercetin after treatment
of rutin 1 using 3.35 equiv of benzyl bromide in N,N0-dimethylform-
amide (DMF) at 60 °C and hydrolysis by HCl/ethanol (15:85, v/v)
during 2 h at 70 °C. In our approach all hydroxyl groups must be pro-
tected during the oxidation step. We previously described that to-
tally benzylated quercetin is obtained by alkylating quercetin with
6 equiv of benzylbromide.18 So, we expected that rutin would be
In the literature quercetin 3-O-b-
D-glucuronide has been syn-
thesized first by direct glucuronidation of 40,7-di-O-benzylquerce-
tin using methyl 2,3,4-tri-O-acetyl-a-D-glucopyranosyluronate
⇑
Corresponding author. Tel.: +33 320434977; fax: +33 320336136.
E-mail address: Christian.rolando@univ-lille1.fr (C. Rolando).
0040-4039/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2011.06.108

M. Kajjout et al. / Tetrahedron Letters 52 (2011) 4738–4740
4739
OBn
In conclusion, we developed an efficient methodology for the
synthesis of quercetin 3-O-b- -glucuronide 5 based on the readily
OH
3'
OBn
OH
D
BnBr (exces)
K2CO3 / DMF
4'
1)
2)
available rutin. The reported strategy based on easy access to the
key intermediate 30,40,5,7-tetrabenzylated quercetin 2 which can
be very easily deprotected and which is not prone to protecting
group scrambling could be applied to the synthesis of 3-O-meth-
ylquercetin18 or labile derivatives of quercetin like sulfate which
are compatible with the hydrogenolysis of the O-benzyl protecting
group.14
OBn
O
7
HO
O
OH
MeOH / HCl
98/2
OH
OH
OH
3
O
5
OH
OBn O
O
O
2
1
O
O
OAc
OAc
OAc
Br
OH
HO
OBn
OH
OBn
O
Acknowledgments
BnO
O
MeONa / MeOH
OAc
K2CO3 / DMF
OAc
The NMR and Mass Spectrometry facilities used in this study
are funded by the European community (FEDER), the Région
Nord-Pas de Calais (France), the CNRS, and the Université de Lille
1, Sciences et Technologies. Part of this work was funded by the
European Contract QLK1-CT-1999-00505 POLYBIND. The authors
thank Dr. Maria van Agthoven for her help with editing this
manuscript.
OAc
O
OBn O
O
OAc
OAc
3
OBn
OH
OBn
OH
1) NaOCl,
TEMPO
BnO
O
HO
O
OH
OH
OH
OH
O
OH
O
2) H2 / Pd/C
References and notes
OBn O
OH
O
O
O
OH
OH
4
5
1. Hertog, M. G. L.; Hollman, P. C. H.; Katan, M. B. J. Agric. Food Chem. 1992, 40,
2379–2383.
2. Hertog, M. G. L.; Hollman, P. C. H.; van de Putte, B. J. Agric. Food Chem. 1993, 41,
1242–1246.
3. Hollman, P. C. H.; Katan, M. B. Food Chem. Toxicol. 1999, 37, 937–942.
4. Manach, C.; Scalbert, A.; Morand, C.; Remesy, C.; Jimenez, L. Am. J. Clin. Nutr.
2004, 79, 727–747.
OH
O
Scheme 1. Synthesis of quercetin 3-O-b-D-glucuronide.
5. Hollman, P. C. H.; Geelen, A.; Kromhout, D. J. Nutr. 2010, 140, 600–604.
6. Gibellini, L.; Pinti, M.; Nasi, M.; De Biasi, S.; Roat, E.; Bertoncelli, L.; Cossarizza,
A. Cancers 2010, 2, 1288–1311.
7. Wang, L.; Lee, I. M.; Zhang, S. M.; Blumberg, J. B.; Buring, J. E.; Sesso, H. D. Am. J.
Clin. Nutr. 2009, 89, 905–912.
8. Spencer, J. P. E.; Vauzour, D.; Rendeiro, C. Arch. Biochem. Biophys. 2009, 492, 1–9.
9. Mullen, W.; Edwards, C. A.; Crozier, A. Br. J. Nutr. 2006, 96, 107–116.
10. Chao, C. L.; Hou, Y. C.; Lee Chao, P. D. L.; Weng, C. S.; Ho, F. M. Br. J. Nutr. 2009,
101, 1165–1170.
11. Price, K. R.; Colquhoun, I. J.; Barnes, K. A.; Rhodes, M. J. C. J. Agric. Food Chem.
1998, 46, 4898–4903.
12. Zhang, X.; Thuong, P. T.; Jin, W.; Su, N. D.; Sok, D. E.; Bae, K.; Kang, S. S. Arch.
Pharm. Res. 2005, 28, 22–27.
13. Wagner, H.; Danninger, H.; Seligmann, O.; Farkas, L. Chem. Ber. 1970, 103,
3674–3677.
14. Needs, P. W.; Kroon, P. A. Tetrahedron 2006, 62, 6862–6868.
15. Bouktaib, M.; Atmani, A.; Rolando, C. Tetrahedron Lett. 2002, 43, 6263–6266.
16. Kajjout, M.; Rolando, C. Tetrahedron 2011, 67, 4731–4741.
17. Huang, H.; Jia, Q.; Ma, J.; Qin, G.; Chen, Y.; Xi, Y.; Lin, L.; Zhu, W.; Ding, J.; Jiang,
H.; Liu, H. Eur. J. Med. Chem. 2009, 44, 1982–1988.
tetrabenzylated under the same conditions. We observed also in our
previous work16,18 that the cleavage of the benzyl group at position
5, during the attempted selective deprotection of catechol ring of
3,5,7-tribenzylated quercetin whose B cycle was protected by the
diphenylmethane ketal under acidic conditions using a mixture of
acetic acid/water (80:20) at reflux. So in order to avoid an additional
step of rebenzylation of the 5-hydroxy group, we decided to work
under diluted acidic conditions to cleave the rutinoside group. The
four free hydroxyl groups of rutin were benzylated using an excess
quantity (8 equiv) of benzyl bromide and potassium carbonate in
DMF during 10 h at room temperature. Then the hydrolysis of tet-
rabenzylated rutin was performed using a mixture of HCl/methanol
(2/98, v/v) at reflux (ca 65 °C) which led to the desired product
30,40,5,7-tetrabenzylated quercetin
2 in 60% yield in gram
quantities.19
18. Bouktaib, M.; Lebrun, S.; Atmani, A.; Rolando, C. Tetrahedron 2002, 58, 10001–
10009.
19. Procedure for the synthesis of the key intermediate 5,7-bisbenzyloxy-2-(3,4-
bisbenzyloxyphenyl)-3-hydroxyl-4H-chromen-4-one 2. All commercially
available products were purchased from Aldrich (Saint-Quentin Fallavier,
The O-glucosylation was achieved by condensing 1-bromo-3,4,6-
tetra-O-acetyl- -glucopyranoside (acetobromoglucose) on pro-
a-D
tected quercetin 2 in DMF at room temperature using simply potas-
sium carbonate as a base. Then the glycosyl moiety was deprotected
by the transesterification of the four acetate groups with sodium
methylate.20 Finally oxidation of the protected quercetin-glucoside
to the corresponding glucuronic acid was accomplished by sodium
hypochlorite (NaOCl) catalyzed by TEMPO (2,2,6,6-tetramethyl-1-
piperidinyloxy) in the presence of potassium bromide and tetrabu-
tylammonium bromide. This step, which is sensitive to the precise
structure of the molecule to be oxidized,16,21 proceeded smoothly
and was not affected by the change of the protecting group on the
B catechol cycle from diphenylmethyleneketal to dibenzylethers.
The synthesis ended by the cleavage of the benzyl groups by a mild
hydrogenolysis using palladium hydroxide at 10% on charcoal in a
THF/EtOH mixture. It must be pointed out that a higher content of
palladium (30%) was required to deprotect the diphenylmethyl-
France) and used as received. To
a solution of rutin hydrate 1 (5.00 g,
approximately 8.0 mmol) in DMF (60 ml), potassium carbonate (9.04 g,
65.52 mmol) and benzyl bromide (7.79 ml, 65.52 mmol) were added under
argon. The reaction mixture was stirred vigorously at room temperature for
10 h. The resulting mixture was diluted with 200 ml of ethyl acetate and was
washed with water (2 ꢀ 150 ml). The residue obtained after evaporation of the
solvent was solubilized in a mixture of MeOH/HCl (98:2). The solution was
refluxed until a precipitate was formed. After cooling the precipitate was
filtered off and was washed with 50 mL of cold MeOH to afford 2 (3.20 g, 60%
yield). 1H NMR (CDCl3): 5.08 (s, 2H, OCH2Ph), 5.20 (s, 2H, OCH2Ph), 5.22 (s, 2H,
OCH2Ph), 5.23 (s, 2H, OCH2Ph), 6.45 (d, J = 1.9 Hz, 1H, aromatic H), 6.55 (d,
J = 1.9 Hz, 1H, aromatic H), 7.01 (d, J = 8.5 Hz, 1H, aromatic H), 7.19–7.47 (m,
20H, aromatic H), 7.59 (dd, J = 8.5, 1.9 Hz, 1H, aromatic H), 7.75 (d, J = 1.9 Hz,
1H, aromatic H). 13C NMR (CDCl3): 70.4 (OCH2Ph), 70.6 (OCH2Ph), 70.9
(OCH2Ph), 72.5 (OCH2Ph), 93.6, 97.5, 106.7, 114.0, 114.1, 121.2, 124.3, 126.7,
127.2, 127.5, 127.7, 127.8, 127.9, 128.5, 128.6, 128.7, 128.9, 135.6, 136.2, 136.8,
137.1, 137.7, 141.9, 148.6, 150.1, 158.7, 159.3, 163.2, 171.7 (C@O). Elemental
Anal. Calcd for C43H34O7: C, 77.94; H, 5.14; O, 16.92. Observed C, 77.96; H, 5.16;
O, 16.94.
eneketal group we used previously. The desired quercetin 3-O-b-D-
glucuronide 5 was obtained after a final purification on a C18 reverse
phase, solid phase extraction (SPE) cartridge using a mixture of
methanol/water as the eluent, and was identified by its ESI mass
spectrum and its 1H and 13C NMR spectra.22 Glucuronide 5 was ob-
tained in 28% yield from rutin 1.
20. Procedure for the synthesis of 5,7-bisbenzyloxy-2-(3,4-bisbenzyloxyphenyl)-3-O-b-
D
-glucopyranosyloxy-4H-chromen-4-one 4: (1.00 g, 1.51 mmol) of compound 2,
(1.24 g, 3.02 mmol) of acetobromoglucose and (417 mg, 3.02 mmol) of
potassium carbonate were dissolved in 20 ml of DMF under argon. The
mixture was agitated for 6 h. The reaction mixture was diluted with 150 ml of
ethyl acetate and was washed with water (2 ꢀ 75 ml). The organic phase was

4740
M. Kajjout et al. / Tetrahedron Letters 52 (2011) 4738–4740
dried over MgSO4 and the solvent was evaporated. The residue obtained was
bromide. Then 10 ml of a solution of 5.5% NaOCl were added dropwise. After
1 h the resulting mixture was diluted with 30 ml of ethyl acetate and 30 ml of
water. The precipitate thus formed was filtered off. The solid obtained was
dissolved in a mixture of EtOH (20 ml) and THF (20 ml), and was treated with
palladium hydroxide (10%, 30 mg) under a hydrogen flow for 12 h. The reaction
mixture was then filtered on Celite and rinsed with EtOH (30 ml). The residue
obtained after evaporation of the solvent was purified by chromatography on a
C18 reverse phase SPE (solid phase extraction) cartridge using a mixture of
MeOH/water (40/60) as the eluent pushed through the cartridge by a slight
purified by flash column chromatography using a mixture of CH2Cl2/EtOAc (85/
15) as the eluent to give 1.19 g of 3 (79% yield). Then (500 mg, 0.50 mmol) of
the obtained compound 3 was dissolved in 30 ml of a mixture of MeOH/THF
(50/50). A solution sodium methylate, prepared from 10 mg of sodium metal in
methanol (10 ml) was added at room temperature. When the deprotection was
completed, the solution was neutralized by adding 2.0 g of an ion-exchange
resin (H+ form). The agitation was maintained for 30 min, and then the resin
was filtered off. Methanol was eliminated by evaporation under vacuum, at
room temperature to give 4, which was used without further purification in the
next step, TEMPO mediated oxidation (383 mg, 93% yield).
argon pressure to afford 5 (18 mg, 63% yield). Data for quercetin 3-O-b-D-
glucuronide 5: 1H NMR (CD3OD) 3.45–3.83 (m, 4H), 5.28 (d, J = 7.2 Hz, 1H), 6.18
(d, J = 2.2 Hz, 1H), 6.42 (d, J = 2.2 Hz, 1H), 6.85 (d, J = 8.5 Hz, 1H), 7.62 (dd,
J = 8.5 Hz, 1H), 7.69 (d, J = 2.0 Hz, 1H). 13C NMR (CD3OD) 73.1, 75.5, 77.5, 78.4,
95.0, 100.4, 104.1, 106.0, 116.1, 118.3, 122.0, 123.1, 135.7, 146.0, 149.7, 157.9,
160.0, 162.9, 165.7, 171.8, 178.6. HR-FAB-MS (m/z): observed, 479.0799; calcd
for C21H19O13, 479.0826 [M+H]+.
21. Wang, P.; Zhang, Z.; Yu, B. J. Org. Chem. 2005, 70, 8884–8889.
22. Procedure for the preparation of 3-O-b-
D-glucuronide 5: to a solution of
compound (50 mg, 0.06 mmol) in CH2Cl2 were added 1 ml of solution
4
saturated in potassium carbonate, 1.5 mg of TEMPO (2,2,6,6-tetramethyl-1-
piperidinyloxy), 5 mg of potassium bromide, and 2.0 mg tetrabutylammonium