The synthesis of radiolabeled Irbesartan using N,N-dimethyl[ 14C]formamide as a source of carbon-14 isotope

Article abstract of DOI:10.1002/jlcr.1846

Irbesartan (Avapro) is in clinical use as an antihypertensive drug. It is a nonpeptide angiotensin II receptor antagonist. A radiolabeled version of this drug, intended for use in environmental fate studies, was prepared from N,N-dimethyl[¹⁴C]f

Full text of DOI:10.1002/jlcr.1846

Research Article
Received 20 August 2010,
Revised 12 October 2010,
Accepted 13 October 2010
Published online 29 December 2010 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.1846
The synthesis of radiolabeled Irbesartan using
N,N-dimethyl[14C]formamide as a source of
carbon-14 isotope
Ã
I. Victor Ekhato, and Samuel Bonacorsi Jr
Irbesartan (AvaproTM) is in clinical use as an antihypertensive drug. It is a nonpeptide angiotensin II receptor anta-
gonist. A radiolabeled version of this drug, intended for use in environmental fate studies, was prepared from
N,N-dimethyl[14C]formamide. Our method of synthesis gave 18% overall yield and a radiochemical purity of 98.22%.
Keywords: angiotensin receptor antagonist; antihypertensive; sartan; Irbesartan
Introduction
Experimental
Irbesartan is a member of the sartan class of antihypertensive All reactions were carried out under an atmosphere of argon
drugs.1 These nonpeptides interact with the renin–angiotensin unless otherwise specified. Solvents were of commercial grade
system (RAS), a central regulator of blood pressure and and used without purification or drying. Column chromatogra-
homeostasis.2 Angiotensin I is a decapeptide formed from the phy was carried out on Merck Kiesegel 60 (230 m) silica gel. Flash
action of renin on angiotensinogen. Angiotensin I is further chromatographic separations were performed on a Biotage
converted by angiotensin converting enzyme (ACE) into Flash System using pre-packed silica gel cartridges. TLC
angiotensin II, an octapeptide. Binding of angiotensin II to its visualization reagents included (10% iodine plus 10% AcOH) in
receptor (AT1) produces a powerful vasoconstrictor.3 Since 40% aqueous KI and, Cerium sulfate in 10% sulfuric acid. 1H NMR
angiotensin II can be formed in vivo by enzymes other4 than spectra were recorded at 300, 400, or 500 MHz Spectrometer.
ACE, a small molecule-mediated modulation of the RAS system Chemical shifts are given in ppm relative to tetramethylsilane.
that blocks the action of angiotensin II at receptor level made a Agilent 1100 HPLC System including solvent degasser, pump,
better therapeutic target than the inhibition of angiotensin I automated injector, and a variable UV detector connected to IN/
conversion to angiotensin II. It is the selective antagonism of the US BetaRam model 4 Flow Detector with a 0.25-mL detector cell
biological activity of angiotensin II that underlies the mechanism was used in the analyses of compounds. The HPLC column was
of action of the sartans, and several compounds in this group YMC-Park Pro C18, 3 m, 4.6 Â 150 mm. Elution Solvent System
including Irbesartan (BMS), Losartan (Merck), Telmisartan (Boeh- A = water:acetonitrile 80:20 (0.1% TFA), and B = water:acetonitrile
ringer Ingelheim), etc., are currently in clinical use.5 Essential 20:80 (0.1% TFA) under Gradient Condition of 0–8 min, 0% B;
hypertension is a major risk factor in cardiovascular disease and 20 min, 100% B; 21 min, 0% B; 27 min, 0% B, and a flow rate of
it accounts for a third of global deaths.6 Irbesartan and the 1.0 mL/min, with detection by UV at 254 nm. LC/MS analysis was
related sartans are useful in the treatment of hypertension and performed on a Finnigan LXQ Mass Spectrometer System, LC/MS
other diseases, such as heart failure, renal insufficiency, method: Generic-B20 (1p ESI full mass). N,N-Dimethyl [14C]for-
glaucoma, and diabetic retinopathy, which are linked with this mamide (Lot ] B37, 100 mCi, Sp Act. 56 mCi/mmol) was
condition.7
purchased from GE Healthcare, UK Limited, Amersham Place,
Our interest in the synthesis of a radiolabeled analog Little Chalfont, Buckinghamshire, PH7 9NA, UK.
was prompted by studies of the environmental fate of
Irbesartan, the ecological impact of its clinical use as a drug 4-Bromo[14C]benzyl bromide 4
and the derivatives therefrom. An earlier labeled synthesis of
To a stirred solution of 1,4-dibromobenzene (1.18 g, 5 mmol) in
dry THF (40 mL) at À781C was slowly added n-BuLi (2.5 M
Irbesartan (unpublished) utilized [14C]cyclopentanone to install
the label. We opted instead for a comparatively less expensive
starting labeled reagent and a short synthetic route that takes
advantage of intermediates in the known syntheses. Dimethyl-
formamide-(carbonyl-14C), 1,4-dibromobenzene, 5-phenyl-1H-
tetrazole, and SR48001A met these requirements for a cost
effective and expeditious preparation, and the details are as
follows.
Department of Chemical Synthesis, Bristol-Myer Squibb Company, Rt. 206/
Provinceline Road, Princeton, NJ 08543, USA
*Correspondence to: I. Victor Ekhato, Department of Chemical Synthesis, Bristol-
Myer Squibb Company, Rt. 206/Provinceline Road, Princeton, NJ 08543, USA.
E-mail: ihoezo.ekhato@bms.com
J. Label Compd. Radiopharm 2011, 54 202–205
Copyright r 2011 John Wiley & Sons, Ltd.

I. V. Ekhato and S. Bonacorsi Jr
solution, 2 mL, 5 mmol) over 45 min. After 1 h, a solution of hexanes, 46.16 mL, 115.4 mmol) was added slowly and the
DMF-14C (purchased as 100 mCi, Sp. Act. 56 mCi/mmol and reaction was stirred at À20 to À101C under argon atmosphere
diluted to 374.5 mg, 5.0 mmol, Sp. Act. 20 mCi/mmol, 100 mCi) in for 1 h. The reaction mixture was again cooled to À301C,
THF (5 mL) was added. It was stirred at À781C for 1 h, 1 N HCl triisopropylborate (30.6 mL, 130 mmol) was added and the
(15 mL) was added and the reaction mixture was allowed to reaction mixture was stirred and slowly warmed to 101C over
slowly warm to room temperature. The reaction was extracted 1 h. After the volatiles were removed by rotary evaporation,
with dichloromethane (3 Â 50 mL), and the combined organic 160 mL of THF was added followed by 60 mL of isopropyl
phases were washed with water (50 mL), brine (50 mL), and alcohol. The mixture was cooled to 01C and saturated NH4Cl
dried. Solvent was removed under reduced pressure and the (40 mL) was added ensuring that temperature stayed below
product was dissolved in THF:MeOH (60 mL, 7:3 v/v). It was 101C. The slurry was warmed to room temperature and stirred
cooled in ice-water bath, sodium borohydride (274.3 mg, for 30 min. Water (100 mL) was added over 20 min and the
7.25 mmol) was added, and the reaction was stirred at room mixture was kept for 2 h during which white solid separated. It
temperature for 30 min. Excess borohydride was destroyed by was collected by filtration, washed with isopropyl alcohol/water/
the careful addition of dil. HCl and the reaction was extracted triethylamine (50:50:2, 100 mL), and dried under high vacuum
with dichloromethane (3 Â 30 mL). The combined organic overnight to give 2-(1-trityl-1H-tetrazol-5-yl)phenylboronic acid
portion was washed with water, brine, and dried. It was purified (43.2 g, 98 %). 1H-NMR (DMSO-d6) d 7.94 À7.84 (m), 7.60–7.39
by column on silica gel (40% ether in hexane) to afford white (m), 7.10 À7.08 (m), 1.74 (brS).
solid, 4-bromobenzyl alcohol-14C 3 (690 mg, 3.68 mmol, 73.7%).
1H NMR (400 MHz, CDCl3), d1.96 (brs, 1H), 4.61 (s, 2H), 7.21 (d, 2H,
Irbesartan-14C 6
J = 8.3 Hz), 7.60 (d, 2H, J = 8.3 Hz). 13C-NMR (125.7 MHz, CDCl3),
139.79, 131.62, 128.59, 121.43, 64.52. To a solution of 3 and
triethylamine (1.02 mL, 7.36 mmol) in 20 mL dry dichloro-
methane at 01C was added methanesulfonyl chloride (401 mL,
5.16 mmol) and the mixture was stirred to warm to room
temperature in 1 h. It was diluted with diethyl ether (40 mL) and
washed with saturated NaHCO3 (30 mL), water (30 mL), brine
(40 mL), and dried over MgSO4. The solution was filtered and
evaporated to give the crude mesylate (992 mg). The mesylate
and LiBr (800 mg, 9.2 mmol) in dry acetone (120 mL) were
refluxed for 2 h and concentrated to a small volume under
reduced pressure. It was partitioned between ether and water,
the aqueous was further extracted with ether (3 Â 30 mL), and
the organic portions were combined and dried. Evaporation
gave 4-bromo-[14C]benzyl bromide 4 (922 mg, 3.6 mmol, 72%,
To a solution of 5 (1.3 g, 3.56 mmol) in methanol (13 mL) was
added potassium hydroxide (400 mg, 7.01 mmol). After the
solution was flushed with argon for 10 min, it was refluxed for
45 min under argon atmosphere. A solution of 2-(1-trityl-1H-
tetrazol-5-yl)phenylboronic acid (2.7 g, 6.23 mmol) and potas-
sium hydroxide (400 mg, 7.01 mmol) in methanol (13 mL) was
added. The reaction mixture was heated under reflux for 10 h
before it was cooled to room temperature. It was diluted
with water (15 mL) and toluene (15 mL); the organic was
separated and the aqueous portion was further extracted with
toluene (12 mL). The organic portions were discarded; the
aqueous was acidified with acetic acid and it was evaporated to
a residue. The residue was purified by using flash column
chromatography (C18 column) using step-gradient Twenty to
sixty percent of MeOH in water to give the phenylboronic
acid impurity followed by pure fractions of labeled compound.
The fractions were combined and while it was being concen-
trated under reduced pressure a white solid separated. It
was collected by filtration and dried under high vacuum to
afford Irbesartan-14C 6, (380 mg, 16.43 mCi, 43.26 mCi/mg or
18.53 mCi/mmol, 25%). Radiochemical purity was determined
to be 98.22% at Retention time of 15.1 min (authentic
Reference Lot ] 6C17210), Flow rate of 1 mL/min and at UV
detection wavelength of 254 nm. MS [M11]1: 429.33 and
431.25. 1H-NMR (300 MHz, CDCl3), 7.90 (d, J = 7.6 Hz, 1H), 7.58
(tr, J = 7.5 Hz, 1H), 7.50 (tr, J = 7.5 Hz, 1H), 7.40 (d, J = 7.58, 1H),
7.14 (d, J = 8.0 Hz, 2H), 7.04 (d, J = 8.0 Hz, 2H), 4.62 (s, 2H), 2.1
(tr, J = 8.5 Hz, 2H), 1.83–1.60 (br m, 8H), 1.4 (m, 2H), 1.25 (m, 2H),
0.79 (tr, 3H).
1
from DMF). H-NMR (400 MHz, CDCl3) d 4.32 (s, 2H), 7.17 (d, 2H,
J = 8.52 Hz), 7.34 (d, 2H, J = 8.3 Hz).
3-(4-Bromo-[14C]benzyl)-2-butyl-1,3-diazaspiro[4,4]non-1-
en-4-one 5
Potassium carbonate (1.53 g, 11.06 mmol) was added to
SR48001A (850 mg, 3.68 mmol) in dry DMF (20 mL) and stirred
for 45 min at room temperature. 4-Bromobenzyl bromide-14C 4
(922.0 mg, 3.68 mmol) in dry DMF (5 mL) was added and the
mixture was stirred at room temperature overnight. The solvent
was removed under high vacuum and the residue was partition
between water/n-heptane/ether (50 mL, 20:20:10 v/v/v). The
aqueous was further extracted with heptane/ether (3 Â 30 mL,
20:10v/v), and the combined organic was dried and evaporated
to give 5 (1.3 g, 3.56 mmol, 96%). 1H-NMR (400 MHz, CDCl3), d
7.39 (d, J = 8.3 Hz, 2 H), 6.97 (d, J = 8.3 Hz, 2H), 4.55 (s, 2H), 2.21
(overlapping q, J = 7.8 Hz, 2H), 1.9 (brm, 6H), 1.7 (brm, 2H), 1.49
(m, 2H), 1.24 (m, 2H), 0.80 (tr, 3H).
Results and discussion
In an earlier labeled synthesis, [14C]-2-n-butyl-1,3-diazo-spiro-
[4,4]-non-1-ene hydrochloride (SR48001A) was generated from
[14C]cyclopentanone and elaborated into labeled Irbesartan 6.8
2-(1-Trityl-1H-tetrazol-5-yl)phenylboronic acid
To a stirred solution of 5-phenyltetrazole (14.9 g, 100 mmol) and Besides that it started from the relatively more expensive
triethylamine (14.8 mL, 105 mmol) in dry THF (260 mL) at 401C [14C]cyclopentanone, we considered the reaction sequence to
was added a solution of trityl chloride (29.9 g, 105 mmol) in dry the -spiro-[4,4]non-1-ene intermediate to be cumbersome. The
THF (120 mL). After 30 min, the reaction was cooled to 01C and additional steps required downstream to convert the labeled
filtered to remove the salt. The filtrate was placed under argon intermediate SR48001A-14C to Irbesartan-14C were likely to
atmosphere and cooled to À251C. n-BuLi (2.5 M solution in further depress the overall radiochemical yield.
J. Label Compd. Radiopharm 2011, 54 202–205
Copyright r 2011 John Wiley & Sons, Ltd.
www.jlcr.org

I. V. Ekhato and S. Bonacorsi Jr
Scheme 1. Synthesis of Irbesartan 14C.
It had been established in earlier work that 4-bromobenzyl 4-bromobenzyl alcohol 3. A mesylate prepared from 3 was
bromide-[14C] 4, easily made from dimethyl[14C]formamide and reacted with LiBr in refluxing acetone to make 4 in 72% yields
1,4-dibromobenzene 1, may be used to insert the bridging from labeled DMF. Reaction of SR48001A with 4 gave 3-[4-
methylene to the heterocycle to make these sartans.9 With bromobenzyl]-2-butyl-1,3-diazoSpiro[4,4]-non-1-en-4-one
labeled DMF instead of [14C]cyclopentanone, we expected to 96% yield.
5 in
realize a significant cost saving.
A phenylboronic acid derivative for cross-coupling with 5 was
In addition, we were encouraged by the precedence in prepared from commercially obtained 5-phenyl-1H-tetrazole.
synthetic literature on Irbesartan indicating that N-alkylation of Our best procedure for making the boronic acid required the
SR48001A with 4-bromobenzyl bromide10 proceeded in good protection of the tetrazole as a triphenylmethyl homolog,
yield. Therefore, our critical task became the reaction of followed by tetrazole moiety directed ortho-lithiation of the
SR48001A with 4-bromobenzyl bromide to produce 5 for the phenyl ring.11 Triisopropyl borate was the superior reagent for
Suzuki cross-coupling reaction that would follow. Such a cross- incorporating boron onto the phenyltetrazole after lithiation at
coupling reaction with 2-(1-trityl-1H-tetrazole-5-yl)phenylboro- À301C. By this protocol, the requisite boronic acid was made in
nic acids has been used extensively in making these sartans,11 98% yield after deprotection with dilute acid. The cross-coupling
and it should provide labeled Irbesartan after the removal reaction was carried out with Pd(PPh3)4 and KOH in methanol
of protecting groups. Accordingly in Scheme 1, the labeled and afforded a low yield of 25%. It could be that the low yield
4-bromobenzyl bromide 4 was prepared as follows. A lithium– was due to the quality of the palladium catalyst. Acidification
halogen exchange reaction was conducted on 1 using 1.0 equiv. with acetic acid to a pH 4 provided Irbesartan-14C in 18% overall
of n-BuLi at À781C. After 45 min, the reaction mixture was Radiochemical yield.
treated with DMF-[114C-carbony] and stirred for a further 1 h
reaction time. Dil HCl was added and the reaction mixture was
stirred to warm to room temperature. The resulting 4-bromo-
benzaldehyde 2 was isolated by extraction method and reduced
immediately with sodium borohydride to give the labeled
Conclusions
From dimethylformamide-(carbonyl-14C), Irbesartan-14C was
prepared in 18% overall yield. The synthetic sequence entailed
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J. Label Compd. Radiopharm 2011, 54 202–205

I. V. Ekhato and S. Bonacorsi Jr
generating labeled benzaldehyde from dimethylformamide-
(carbonyl-14C) and 1,4-dibromobenzene, its reduction to the
alcohol, followed by conversion of the latter compound to the
benzyl bromide. Thereon in the sequence, known intermediates,
other established reaction steps like N-alkylation of SR48001A
and Suzuki cross-coupling were employed to make Irbesar-
tan-14C. The present sequence is a cost effective and an efficient
route to labeled Irbesartan.
Pharmacological Basis of Therapeutics, 7th edn (Eds.: A. G. Gilman,
L. S. Goodman, T. W. Rall, F. Murad), Macmillan Pub. Co.,
New York, 1975, pp. 639–659.
[3] M. A. Ondetti, D. W. Cushman, J. Med. Chem. 1981, 24,
355–361.
[4] H. Okunishi, M. Miyazaki, T. Okamura, N. Toda, Biochem. Biophys.
Res. Commun. 1987, 149, 1186.
[5] P. C. Wong, T. B. Barnes, A. T. Chiu, D. D. Christ, J. V. Duncia,
W. F. Herblin, P. B. M. W. M. Timmermans, Cardiovasc. Drug Rev.
1991, 9, 317–399. b) D. J. Carini, J. V. Duncia, EPA EP0253310A2,
1998.
[6] World Health Organization; International Society of Hypertension
Writing Group, J. Hypertens. 2003, 21, 1983–1992.
[7] M. McIntyre, R. J. MacFadyen, P. A. Meredith, R. Brouard, J. L. Reid,
J. Cardiovasc. Pharmacol. 1996, 28, 101–106
Acknowledgements
I. V. E. gratefully acknowledges the Department of Chemical
Synthesis for support, John Chen for the supply of SR48001A,
Reference Irbesartan, and Sharon X Gong for the analysis of final
product.
[8] J. R. Durrwachter, US Patent 5, 910,595,
8 June 1999;
b) C. Bernhart, J. C. Brelier, J. Clement, D. Nisato, P. Perreault,
C. Muneaux, Y. Muneaux, US Patent 5,270,317, December
1993.
[9] I. V. Ekhato, C. C. Huang, J. Label. Compd. Radiopharm 1994, 34,
213–220.
References
[10] B. S. Y. Sumalatha, S. Venkatraman, G. M. Reddy, P. P. Reddy,
Synthetic Commun. 2005, 35, 1979–1982 and references cited
therein.
[1] C. Bernhart, J.-C. Brelier, J. Clement, D. Nisato, P. Perreaut, EPA
1991; 454511. EPA PCT 1991; 91/14679.
[2] G. Berellini, G. Cruciani, R. Mannhold, J. Med. Chem. 2005, 48,
4389–4399; b) W.W. Douglas, Goodman and Gilman’s The
[11] S. L. Young, T. R. Lucius, D. J. Meloni, J. R. Moore, Y.-C. Lee, J. F.
Arnett, J. Heterocyclic. Chem. 1995, 32, 355–357.
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