Synthesis of (-)-lobeline via enzymatic desymmetrization of lobelanidine

Article abstract of DOI:10.1016/j.bmc.2009.01.055

The bioactive alkaloid (-)-lobeline was synthesized via the stereoselective acylation (desymmetrization) of meso-lobelanidine by vinyl acetate in the presence of Candida antarctica lipase B.

Full text of DOI:10.1016/j.bmc.2009.01.055

Bioorganic & Medicinal Chemistry 17 (2009) 1837–1839
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Bioorganic & Medicinal Chemistry
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Synthesis of (ꢀ)-lobeline via enzymatic desymmetrization of lobelanidine
*
Robert Chênevert , Pierre Morin
Département de Chimie, PROTEO, Faculté des Sciences et de Génie, Université Laval, Québec (QC), Canada G1V 0A6
a r t i c l e i n f o
a b s t r a c t
Article history:
The bioactive alkaloid (ꢀ)-lobeline was synthesized via the stereoselective acylation (desymmetrization)
of meso-lobelanidine by vinyl acetate in the presence of Candida antarctica lipase B.
Ó 2009 Elsevier Ltd. All rights reserved.
Received 9 December 2008
Revised 20 January 2009
Accepted 24 January 2009
Available online 31 January 2009
Keywords:
Lobeline
Lobelanidine
Candida antarctica lipase B
Desymmetrization
OH
O
OH
OH
1. Introduction
Ph
N
Ph
Ph
N
Ph
S
S
R
Lobeline 1 (Fig. 1) is the main alkaloid constituent of Lobelia inf-
lata, a plant indigenous to North America. The plant is also known
as Indian tobacco because native Americans smoked the dried
leaves as a substitute for tobacco. Lobeline has a long history of
therapeutic usage and, during the 19th century, was prescribed
as an emetic or a respiratory stimulant for the treatment of various
ailments.1
Me
Me
(-)-Lobeline 1
Lobelanidine 2
Figure 1. Structure of (ꢀ)-lobeline 1 and lobelanidine 2.
2. Results and discussion
Several recent studies have shown that lobeline has multiple
mechanisms of action,1 acting as a ligand for nicotinic acetylcho-
Lobelanidine 2 was prepared in two steps according to litera-
ture procedures.15,18,19 The Mannich condensation of glutaralde-
hyde, benzoylacetic acid and methylamine hydrochloride
provided the diketone lobelanine which was reduced with sodium
borohydride to yield a mixture of cis–trans diastereomeric diols.
The major cis isomer, lobelanidine 2, was purified by
recrystallization.
We then completed some screening experiments in order to
find lipases with the ability to distinguish the enantiotopic groups
of meso-lobelanidine 2 (Table 1). The enantiomeric excess (ee) of
monoacetate 3 was determined by chiral HPLC. Acylation of 2 with
vinyl acetate as the acylation reagent and solvent in the presence
of Candida antarctica lipase B (CAL-B) provided monoester 3 in fair
yield and high enantiomeric excess (ee = 97%) but the reaction was
very slow (320 h) at room temperature (entry 1). CAL-B is a ther-
mostable enzyme, allowing operation at temperature up to
90 °C.20 At 50 °C, the reaction gave a better yield and the same ee
in a shorter reaction time (18 h) (entry 2). Substrate 2 is insoluble
in solvents of low polarity (hexane, toluene, ether) commonly used
in lipase-catalyzed esterification and acetylation in acetonitrile/vi-
nyl acetate (9:1) led to lower enantioselectivity. Acylation of 2 with
line receptors,2–4
l
-opioid receptors,5 and monoamine transport-
ers (dopamine, serotonine, norepinephrine, vesicular monoamine
transporter-2).6,7 Lobeline has been considered as a useful lead
for the development of medications to treat psychostimulants
addictions (smoking, cocaine, amphetamines)8–11 and neurological
disorders such as Alzheimer’s and Parkinson’s diseases.1,12
To date, a few syntheses of racemic or enantioenriched lobeline
have been reported.1,13,14 Recently, both enantiomers of lobeline
were prepared via the enantioselective acylation of lobelanidine
2 with propionic anhydride in the presence of a chiral organocata-
lyst15 and Stoltz et al. developed another approach via PdCl2(spar-
teine)-catalyzed aerobic oxidation of 2.16 Herein, we report the
synthesis of natural (ꢀ)-lobeline 1 via the enzymatic desymmetri-
zation17 of lobelanidine 2.
* Corresponding author. Tel.: +1 418 656 3283; fax: +1 418 656 7916.
E-mail address: robert.chenevert@chm.ulaval.ca (R. Chênevert).
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.01.055

1838
R. Chênevert, P. Morin / Bioorg. Med. Chem. 17 (2009) 1837–1839
Table 1
Enzymatic acylation (desymmetrization) of lobelanidine 2
OH
OH
OAc
OH
OAc
OAc
Ph
Lipase
+
Ph
N
Ph
Ph
N
Ph
Ph
N
Vinyl acetate
R
S
Me
Me
Me
2
3
4
Entry
Lipase
T (°C)
Time (h)
Monoacetate 3
Eed (%)
Diacetate 4
Yieldc (%)
Absolute configuratione
Yieldc (%)
1
2
3
CAL-Ba
CAL-Ba
PFLb
25
50
25
320
18
183
65
70
71
97
>98
49
S
S
S
6
3
27
a
CAL-B: Candida antarctica lipase B.
PFL: Pseudomonas fluorescens lipase.
Isolated yield.
Measured by HPLC on a chiral phase.
At the CHOAc center.
b
c
d
e
OAc
OAc
Ph
OH
OH
Ph
Ac2O
pyridine
Ph
N
Me
Ph
N
Me
4
2
OAc
OH
OH
OH
Ph
lipase
buffer
+
Ph
N
Ph
Ph
N
Me
Me
3
2
Scheme 1. Preparation and enzymatic hydrolysis of diacetate 4.
OAc
OH
Ph
OAc
O
OH
O
CrO3-H2SO4
Acetone
MeOH
HCl
Ph
N
Ph
N
Ph
Ph
N
Ph
Me
3
Me
Me
1
Scheme 2. Preparation of (ꢀ)-lobeline 1 from monoacetate 3.
Pseudomonas fluorescens lipase (PFL) resulted in a lower ee (49%)
(entry 3), and several lipases including Aspergillus niger lipase
(ANL), Mucor sp. lipase (MSL), Candida rugosa lipase (CRL), and
Pseudomonas sp. lipase (PSL) were not active.
transformation into (ꢀ)-lobeline 1 (Scheme 2). The Jones oxidation
of the alcohol function followed by acidic alcoholysis of the ester
function provided (ꢀ)-lobeline 1. Comparison of the optical rota-
tion of this product with literature data revealed its absolute con-
figuration to be identical with that of the natural product. As a
result, the absolute configuration of monoacetate 3 was deduced
to be S at the CHOAc center. Spectroscopic and physical data of
synthetic (ꢀ)-lobeline were in agreement with literature values;
as already noted by others, we observed a cis–trans epimerisation
in solution.1,14,16
Diacetate 4 was prepared by the acetylation of 2 with acetic
anhydride in pyridine (Scheme 1). Hydrolysis of diacetate 4 by
CAL-B in a phosphate buffer at 50 °C was very slow (several days)
and provided enantiomerically pure (ee P 95%) monoacetate 3 in
low yield (23%) and the corresponding achiral diol 2 (74%) indicat-
ing that the monoacetate is also a substrate for the lipase (overhy-
drolysis). Hydrolysis in water-saturated diisopropylether did not
improve the rate of conversion or the yield of monoester 3. Unex-
pectedly, the enzymatic hydrolysis provided the same monoester
as the acylation. In general, the esterification of meso-diols and
the hydrolysis of the corresponding meso-diesters are complemen-
tary and give opposite enantiomers but exceptions have been
reported.21,22
3. Experimental
3.1. General
NMR spectra were recorded on a Varian Inova AS 400 spectrom-
eter (400 MHz). Infrared spectra were recorded on a Bomem MB-
100 spectrometer. Optical rotations were measured using a JASCO
DIP-360 polarimeter (c as gram of compound per 100 mL). High
resolution mass spectra (HRMS) were recorded on an Agilent
The reactions were monitored by TLC analysis and terminated
when all the starting material was consumed (conversion = 100%).
The absolute configuration of monoester 3 was determined by

R. Chênevert, P. Morin / Bioorg. Med. Chem. 17 (2009) 1837–1839
1839
MSTOF-6210 (electrospray) spectrometer. Flash column chroma-
tography was performed on aluminum oxide (activated, neutral,
58 Å, from Aldrich). Thin layer chromatography (TLC) was carried
out using aluminum oxide 60 F254 (neutral, type E, from Merck).
The enantiomeric excess of compound 3 was determined by HPLC
with a chiral stationary phase column (Regis Technologies Inc.
(S,S)-Whelk-O1, 4.6 mm ꢁ 250 mm), using n-hexane/ethanol (4:1)
as the eluent, at a flow rate of 1.5 mL/min with a detection wave-
length of 220 and 254 nm, t(S) = 8.25 min, t(R) = 5.55 min. Enzymes
were from Sigma-Fluka (PFL, CRL, PSL), Amano International En-
zyme (ANL, MSL), and Roche Molecular Biochemicals (CAL-B,
Chirazyme L-2, carrier-fixed, C2, from C. antarctica fraction B,
4.5 kU/g dry carrier, with tributyrin as the substrate). (ꢀ)-Lobeline
was purchased from Sigma.
quenched by filtration of the enzyme when all the starting material
was consumed. The solvents were evaporated (water–MeOH azeo-
trope) and the crude product was purified by flash chromatogra-
phy (neutral alumina, CH2Cl2/1% EtOH) to give monoacetate 3
(10.6 mg, 23%, ee P 95%) and lobelanidine 2 (29.9 mg, 74%). Spec-
tral data as above.
3.5. Synthesis of (ꢀ)-lobeline 1 from lobelanidine monoacetate
3
To
0.204 mmol) in acetone (5 mL) cooled to 0 °C, was added Jones re-
agent (25 mg of CrO3, 20 L of H2SO4 and 70 L of H2O). The mix-
a
solution of lobelanidine monoacetate
3
(78 mg,
l
l
ture was stirred at room temperature for 1 h. The mixture was
diluted with chloroform (30 mL) and saturated aqueous NaHCO3
was slowly added until the organic phase became colorless. The or-
ganic phase was separated, dried, and evaporated to yield the
crude intermediate keto-ester as an oil which was dissolved in
methanol (15 mL) and 3 N HCl (8 mL). The mixture was stirred at
reflux overnight. The solvent was then evaporated and the crude
product was purified by recrystallization (43 mg, 57% for two
steps). All characterization data were identical to those of a com-
mercial sample of (ꢀ)-lobeline hydrochloride.
3.2. Enzymatic acylation of lobelanidine 2
To a solution of lobelanidine 2 (100 mg, 0.27 mmol) in vinyl
acetate (10 mL) was added C. antarctica lipase B (3700 units). The
mixture was stirred at 50 °C for 18 h. The enzyme was removed
by filtration, and the filtrate was evaporated to dryness. The crude
product was purified by flash chromatography (neutral alumina,
CH2Cl2 containing 1% EtOH) to provide lobelanidine monoacetate
3 (72 mg, 70%) as a colorless oil and lobelanidine diacetate 4
(3.6 mg, 3%) as a white solid. Lobelanidine monoacetate 3: ½a D23
ꢂ
Acknowledgments
ꢀ2.7 (c 0.57, CHCl3); IR (NaCl) 3380, 2940, 1730, 1237,
1026 cmꢀ1
;
1H NMR (400 MHz, CDCl3) d 1.39–1.80 (m, 8H), 2.04
We acknowledge the financial support of this work by the Nat-
ural Sciences and Engineering Research Council of Canada (NSERC).
(s, 3H), 2.17–2.24 (m, 1H), 2.51 (s, 3H), 2.54–2.61 (m, 1H), 3.27–
3.35 (m, 1H), 3.76 (br s, 1H), 4.83 (dd, J = 10.8 and 1.6 Hz, 1H),
5.80 (dd, J = 9.8 and 4.0 Hz, 1H), 7.22–7.40 (m, 10H), 8.2 (br s,
OH); 13C NMR (100 MHz, CDCl3) d 21.6, 23.2, 23.7, 39.8, 41.8,
60.1, 63.1, 72.9, 125.7, 126.8, 127.4, 128.6, 128.9, 139.9, 145.1,
170.8; HRMS (ES) m/z calcd for C24H32NO3 (MH)+: 382.2377. Found
382.2389.
References and notes
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1997, 282, 410.
3.3. Lobelanidine diacetate 4
5. Miller, D. K.; Lever, J. R.; Rodvelt, K. R.; Baskett, J. A.; Will, M. J.; Kracke, G. R.
Drug Alcohol Depend. 2007, 89, 282.
6. Zheng, F.; Zheng, G.; Deaciuc, A. G.; Zhan, C. G.; Dwoskin, L. P.; Crooks, P. A.
Bioorg. Med. Chem. 2007, 15, 2975.
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Chem. Lett. 2006, 16, 5018.
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2008, 75, 1411.
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2008, 18, 6509.
To a solution of lobelanidine 2 (200 mg, 0.53 mmol) in pyridine
(5 mL) was added acetic anhydride (1 mL, 10.5 mmol) and the mix-
ture was stirred overnight at room temperature. The solvent was
evaporated and the crude product was purified by flash chroma-
tography (neutral alumina, CH2Cl2) to give lobelanidine diacetate
4 (173 mg, 77%) as a white solid: mp: 205–210 °C (dec.); IR (KBr)
3038, 2934, 1737, 1521, 1457, 1243 cmꢀ1 1H NMR (400 MHz,
;
10. Neugebauer, N. M.; Harrod, S. B.; Stairs, D. J.; Crooks, P. A.; Dwoskin, L. P.;
Bardo, M. T. Eur. J. Pharmacol. 2007, 571, 33.
11. Thayer, A. Chem. Eng. News 2006, 84, 21.
CD3OD) d 1.54–1.69 (m, 3H), 1.85–2.10 (m, 5H), 2.12 (s, 6H),
2.28–2.36 (m, 2H), 2.70 (br s, 3H), 3.47–3.54 (m, 2H), 5.79 (dd,
J = 13.4 and 4.9 Hz, 2H), 7.31–7.40 (m, 10H); 13C NMR (100 MHz,
CDCl3) d 21.4, 21.6, 22.7, 38.9, 53.7, 72.3, 126.7, 128.7, 129.1,
139.5, 170.8, 175.8; HRMS (ES) m/z calcd for C26H34NO4 (MH)+:
424.2482. Found 424.2487.
12. Zheng, G.; Dwoskin, L. P.; Crooks, P. A. The AAPS J. 2006, 8, E682.
13. Felpin, F. X.; Lebreton, J. J. Org. Chem. 2002, 67, 9192.
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B. M. J. Am. Chem. Soc. 2008, 130, 13745.
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18. Klingler, F. D.; Sobotta, R. (Boehringer Ingleheim). Patent US2006/0014791,
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3.4. Enzymatic hydrolysis of lobelanidine diacetate 4
19. Schöpf, C.; Lehmann, G. Annalen 1935, 518, 1.
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E.; Parmar, V. S. Tetrahedron Lett. 2005, 46, 4511.
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19, 1333.
To a solution of lobelanidine diacetate 4 (50 mg, 0.118 mmol) in
diethyl ether (1 mL) were added a phosphate buffer (pH 7.1, 9 mL)
and CAL-B (1000 units). The mixture was stirred at 50 °C. The reac-
tion was monitored by TLC (neutral alumina, CH2Cl2/2% EtOH) and
22. Chênevert, R.; Jacques, F. Tetrahedron: Asymmetry 2006, 17, 1017.