Assignment of the absolute configuration of (-)-linarinic acid by theoretical calculation and asymmetric total synthesis

Article abstract of DOI:10.1016/j.tetasy.2005.11.029

The specific rotation of (-)-linarinic acid calculated using Hartree-Fock and density functional theory on 14 representative conformers of this molecule were all negative. The most stable conformer was found to be almost identical to the crystallographic

Full text of DOI:10.1016/j.tetasy.2005.11.029

Tetrahedron:
Asymmetry
Tetrahedron: Asymmetry 17 (2006) 179–183
Assignment of the absolute configuration of (ꢀ)-linarinic acid
by theoretical calculation and asymmetric total synthesis
Maosheng Cheng,a,* Qiang Li,a Bin Lin,a Yu Sha,a Jinhong Ren,a Yan He,a
Qinghe Wang,a Huiming Huab and Kenneth Ruudc
aSchool of Pharmaceutical Engineering, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenhe District,
Shenyang 110016, PR China
bSchool of Traditional Chinese Materia Medica, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenhe District,
Shenyang 110016, PR China
cDepartment of Chemistry, University of Tromsø, N-9037 Tromsø, Norway
Received 29 July 2005; revised 23 October 2005; accepted 29 November 2005
Abstract—The specific rotation of (ꢀ)-linarinic acid calculated using Hartree-Fock and density functional theory on 14 representa-
tive conformers of this molecule were all negative. The most stable conformer was found to be almost identical to the crystallo-
graphic structure. These results are consistent with the assumption that (ꢀ)-linarinic acid has a (1S)-configuration. An
asymmetric total synthesis with starting products chosen on the basis of the theoretical calculations was found to corroborate
the theoretical assignment. On the basis of the theoretical calculations of specific rotations and a total asymmetric synthesis, the
absolute configuration of the natural product (ꢀ)-linarinic acid was determined to be 1S.
ꢀ 2005 Elsevier Ltd. All rights reserved.
1. Introduction
lute configuration of the stereogenic carbon could not
be assigned. We decided to combine recent advances in
the calculation of specific rotations with a total asym-
metric synthesis in order to determine the absolute con-
figuration of (ꢀ)-linarinic acid.
Linaria vulgaris Mill. (Scrophulariaceae), distributed
in the northeast and Inner Mongolia of China, is
known as a folk medicine for antitussive expectorant,
antiasthmatic, diuretic, laxative and curing headache,
jaundice and dermatosis.1 (ꢀ)-Linarinic acid, [1,2,3,9-
tetrahydro-pyrrolo(2,1-b)quinazolin-1-carboxylic acid] is
a new compound isolated from the ethanol extract of
the whole herb of L. vulgaris Mill. by Hua et al.2
According to the biosynthetic regular pattern of alkal-
oids, (ꢀ)-linarinic acid was biosynthesized from a cer-
tain amino acid. It is well known that almost all
naturally existing amino acids are of the (S)-configura-
tion. Thus, it has been assumed that (ꢀ)-linarinic acid
also has an (S)-configuration. Herein, we calculated
the specific rotation of (S)-linarinic acid using a recently
developed theoretical approach implemented in Dalton
2.0 for accurate calculations of specific rotations.5 Based
on the agreement of the calculated sign of the specific
rotation with the experimentally observed specific
rotation of the natural product, an asymmetric total
synthesis of (S)-linarinic acid was carried out using
o-nitrobenzaldehyde and L-glutamic acid as the starting
material. The absolute configuration of (ꢀ)-linarinic
acid was unambiguously assigned as having an (S)-con-
figuration by both theoretical calculations and asymmet-
ric total synthesis.
Using UV, IR, NMR, MS, 2D NMR, and X-ray data
analysis, the structure was identified as a new tricyclic
quinazoline alkaloid.2 A notable characteristic of the
compound is the existence of a 1-carboxylic and func-
tional group, which results in a stereogenic carbon. A
single-crystal X-ray diffraction analysis of this com-
pound was performed and its result deposited with the
Cambridge Structural Database.3,4 However, the abso-
*
Corresponding author. Tel.: +86 24 2398 6101; fax: +86 24 2399
5043; e-mail: mscheng@263.net
0957-4166/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2005.11.029

180
M. Cheng et al. / Tetrahedron: Asymmetry 17 (2006) 179–183
Table 1. Theoretical specific rotations at the sodium D-line of 14
2. Results and discussion
representative conformers of (S)-linarinic acid using the aug-cc-pVDZ
basis set
2.1. Theoretical calculation
Conformer
HF
ꢀ58.98
DFT/B3LYP
The first ab initio calculations of specific rotations were
reported in 1997 by Polavarapu.6 Since then, a number
of investigations have been published.7,8 Recently, theo-
retical calculations of specific rotations were used suc-
cessfully to assist synthetic chemists in elucidating the
absolute configuration of some chiral molecules.9 The
theoretical calculations proved particularly successful
for molecules with no more than two stereogenic centers
when combined with other theories such as vanÕt HoffÕs
rule, which is applicable to rather complex natural prod-
ucts.9 Here, we use theoretical calculations to assign the
absolute configuration for (ꢀ)-linarinic acid.
1
2
3
4
5
6
7
8
ꢀ57.64
ꢀ40.12
ꢀ71.41
ꢀ271.08
ꢀ243.34
ꢀ213.87
ꢀ142.90
ꢀ115.38
ꢀ88.78
ꢀ79.35
ꢀ75.14
ꢀ115.15
ꢀ83.34
ꢀ119.88
ꢀ491.94
ꢀ435.72
ꢀ355.86
ꢀ269.63
ꢀ189.02
ꢀ124.10
ꢀ83.25
9
10
11
12
13
14
ꢀ73.48
ꢀ313.05
ꢀ250.13
(ꢀ)-Linarinic acid has only one stereogenic center.
There is therefore no need to apply vanÕt HoffÕs rule
and a direct calculation of its specific rotations will suf-
fice to determine the absolute configuration of the mole-
cule. In addition to being strongly basis set dependent,
the specific rotation is also known to depend strongly
on the molecular geometry and in particular on the
molecular conformation.10 Based on a conformational
search and cluster analysis, 14 stable and representative
conformations were identified (Chart 1).
Chart 2. Specific rotations of (S)-linarinic acid calculated at the HF
level with different basis sets.
As shown in Table 1, all conformers give rise to a nega-
tive specific rotation at both the Hartree-Fock and
DFT/B3LYP levels of theory (the only exceptions to
this observation were conformers 1 and 3 using the
(inadequate) 6-31G basis set at the Hartree-Fock level
of theory). This sign is in agreement with the experimen-
tal observations for (ꢀ)-linarinic acid. London atomic
orbitals13 have been used in our calculation to ensure
that the Gab tensor determining the optical rotation is
independent of the gauge origin.14–16
Chart 1. 14 Representative conformers of (S)-linarinic acid.
These conformations were optimized using the Hartree-
Fock method with a 6-31G** basis set. The specific
rotations of these conformations were calculated using
several basis sets, of which the largest was the aug-
cc-pVDZ set, in accordance with the recommendations
of Stephens et al.11 The specific rotations of the opti-
mized conformers were evaluated using the Dalton
quantum chemistry package12 with Hartree-Fock and
density functional theory with the B3LYP hybrid func-
tional.11 The results obtained at the Hartree-Fock and
DFT levels of theory using the aug-cc-pVDZ basis are
collected in Table 1. In Chart 2, we have illustrated
the specific rotation obtained for the different basis sets
at the Hartree-Fock level of theory.
The crystal structure of (ꢀ)-linarinic acid is available in
the Cambridge Structural Database with deposition
number CCDC 242576. This crystal structure is almost
identical to the most stable conformer given by the con-
formational search with an rmsd of 0.603 (Chart 3). A
Hartree-Fock calculation of the specific rotation was
also carried out for this structure, giving a specific
rotation of ꢀ263. This indicates that the conforma-
tional search gave a correct and reasonable set of

M. Cheng et al. / Tetrahedron: Asymmetry 17 (2006) 179–183
181
Table 2. Comparison of the theoretical specific rotations at four
different wavelengths of 14 representative conformers of (S)-linarinic
acid using the aug-cc-pVDZ basis set with experimental data
Wavelength
589
578
546
436
Experimental
Calculations
ꢀ290
ꢀ356
ꢀ300
ꢀ375
ꢀ340
ꢀ443
ꢀ750
ꢀ993
2.2. Chemical synthesis
Different synthetic routes were attempted to synthe-
size the target compound. In order to synthesize the
stereoisomers asymmetrically and on the basis of the
theoretical calculations described in the preceding
section, L-glutamic acid was used as the chiral block
and o-nitrobenzaldehyde as the starting material.
Chart 3. The alignment of crystal structure and most stable structure
given by conformation search.
First, o-nitrobenzaldehyde and L-glutamic acid were
transformed into compound 4 via a reductive aminated
reaction in 65.9% yield.18 Compound 4 was then
refluxed in ethanol for 3 h and dehydration gave com-
pound 5 in 97.1% yield.18 The two above-mentioned
products were used in the following steps without fur-
ther purification. Reductive approaches using catalytic
hydrogenation19 and SnCl2/HCl20 failed due to poor
solubility in most solvents. However hydrazine hydrate,
ferric chloride, and active carbon were used to reduce
the nitro group. The product obtained was confirmed
to be our target compound (S)-linarinic acid in 59.6%
yield (Scheme 1).
conformations, and that the most stable conformer
found was indeed responsible for the largest contribu-
tion to the overall specific rotation.
Carboxylic acids are known to dimerize in low concen-
tration; and this may affect our calculated optical
rotations, as shown by Polavarapu.17 However, consid-
ering the good agreement obtained between theory and
experiment for (ꢀ)-linarinic acid, we do not believe this
to be a serious concern for this molecule.
Considering that predictions of rotation at a single
wavelength do not reveal the full story, we also calcul-
ated the specific rotations at different wavelengths and
compared them with the experimental data. This is
shown in Table 2. This comparison again confirmed that
our calculations are correct.
In this synthetic scheme, (S)-linarinic acid was obtained
in an unexpected way. The mechanism of the last step is
still unknown and further investigations of this mecha-
nism are in progress.
The specific rotations of the intermediates and (S)-linar-
inic acid were measured. The minus sign of the specific
18
In summary, the theoretical calculations of most con-
formers at the HF and DFT levels gave a negative sign
for the optical rotation, which is consistent with the nat-
ural product. These results strongly suggest that the
absolute configuration at carbon-1 for natural (ꢀ)-linar-
inic acid is S.
rotation of (S)-linarinic acid {½aꢁD ¼ ꢀ290:0 (c 0.01,
MeOH)} was consistent with that of the natural product
18
{½aꢁD ¼ ꢀ217:0 (c 0.01, MeOH)}.2 Based on the above
comparison, we concluded that the absolute configura-
tion at the C-1 position of natural (ꢀ)-linarinic acid is
an (S)-configuration (Chart 4).
O
O
OH
OH
1) NaOH
2) NaBH4
CHO
H N
2
N
H
+
3) HCl/H2O
OH
NO
NO
2
2
OH
OH
O
O
3
2
4
O
O
OH
CH3CH2OH
reflux
NH2NH2·H2O/FeCl3/active carbon
CH3CH2OH reflux
N
N
N
O
NO
2
1
5
Scheme 1. Synthetic route to (ꢀ)-linarinic acid.

182
M. Cheng et al. / Tetrahedron: Asymmetry 17 (2006) 179–183
COOH
where xn is the excitation energy from the electronic
ground state w0 to the excited state wn, and la and mb
are components of the electric and magnetic dipole oper-
ator, respectively.
N
N
Chart 4. The structure of (S)-linarinic acid.
Four basis sets were used for the calculation of the spe-
cific rotation: 6-31G, 6-31G*, 6-31G**, and aug-cc-
pVDZ. The final specific rotation was determined by
Boltzmann weighting of specific rotations for all
adopted conformers.
3. Conclusions
Using both theoretical calculations and an asymmetric
total synthesis of (S)-linarinic acid, the determination
of the absolute configuration for natural (ꢀ)-linarinic
acid has been accomplished herein. The specific rotation
of (S)-linarinic acid calculated by ab initio methods were
all negative. Based on this result, the absolute configura-
tion of natural (ꢀ)-linarinic acid was predicted to be
(1S). On the basis of theoretical calculations, the asym-
metric synthesis of (S)-(ꢀ)-linarinic acid was achieved in
three steps in an overall yield of 38.1%. A comparison
performed between the natural product and the synthe-
sized product demonstrated that the sign of the specific
rotation of the synthesized product was consistent
with the natural product. We therefore concluded that
the absolute configuration at C-1 in naturally existing
(ꢀ)-linarinic acid is (1S).
4.2. Organic general
All melting points were determined on a Buchi-540 melt-
¨
ing point apparatus and are uncorrected. The IR spectra
were recorded on a Bruker IR S-55 instrument. 1H
NMR and 13C NMR spectra were measured on a Bru-
ker AC (E)-300 instrument using tetramethylsilane as
an internal standard. EI-MS was obtained on Finnigan
LCQ–MS spectrometer. ESI-MS was obtained on Agi-
lent 1100 series LC/MS Trap. Specific rotation was mea-
sured on Perkin–Elmer 241MC Polarimeter. Analytical
thin-layer chromatography (TLC) was performed on
precoated plates of silica gel HF254 (0.5 mm, Qingdao,
China). Flash column chromatography was performed
on silica gel H (60 lm, Qingdao, China).
4.2.1. Preparation of (S)-(ꢀ)-2-(2-nitro-benzylamino)-
pentanedioic acid 4. L-Glutamic acid (3) (9.80 g,
66.7 mmol) was added at room temperature to a solu-
tion of sodium hydroxide (2 M, 50 mL). Then an etha-
4. Experimental
4.1. Conformational search and calculations of optical
rotations
nol solution (150 mL) of o-nitrobenzaldehyde
2
(8.30 g, 54.9 mmol) was added dropwise. The mixture
was stirred at room temperature for 30 min and cooled
to 0 ꢁC. Sodium borohydride (1.20 g, 32.4 mmol) was
added in small portions at 0–5 ꢁC. The mixture was stir-
red for 90 min at room temperature, and another por-
tion of o-nitrobenzaldehyde (1.70 g, 11.2 mmol) added.
After 20 min, a second portion of sodium borohydride
(0.31 g, 8.1 mmol) was added as before, and the mixture
stirred for 45 min at room temperature. The resulting
solution was extracted with ether (150 mL · 3). The
aqueous layer was acidified to pH 2–3 at 0–5 ꢁC with
concentrated hydrochloric acid. The resulting suspen-
The conformational search of (S)-linarinic acid was per-
formed using Systematic Search by MMFFs force field
in SYBYL21 by rotating two bonds of the carboxyl group.
Water was used as the ÔsolventÕ in the conformational
search. A cluster analysis was then accomplished based
on those conformations to give 14 representative
conformers.
These 14 representative conformers obtained from the
cluster analysis were optimized at the HF/6-31G** level
using the Dalton program, and the corresponding
specific rotations were calculated at the frequency of
the sodium D-line (589.3 nm), also using Dalton12
at the HF and DFT levels of theory, respectively. The
specific optical rotation is calculated as follows:5
sion was filtered to give a white dusty solid 12.40 g, yield
18
65.9%. Mp 186.1–190.0 ꢁC. ½aꢁD ¼ þ47:7 (c 0.51,
DMSO). IR mmax (KBr) cmꢀ1, 3422, 3040, 1708, 1601,
1
1530, 1438, 1343, 1260, 1178, 860, 813, 792, 740. H
NMR (DMSO-d6, 300 MHz) d 1.76 (2H, m), 2.30 (2H,
m), 3.07 (1H, t, J = 5.4 Hz), 3.85 (1H, d, J = 14.8 Hz),
4.07 (1H, d, J = 14.9 Hz), 7.52 (1H, m), 7.70 (2H, m),
7.93 (1H, d, J = 9.8 Hz), 9.25 (2H, s). EI-MS m/z: 264
(0.79), 237 (11.48), 218 (14.39), 136 (100.00), 130
(24.68), 119 (10.15), 92 (12.42), 78 (84.02), 65 (27.40),
51 (23.85), 39 (29.31).
½aꢁD ¼ 0:1343 ꢂ 10ꢀ3bm2=M
ꢀ
with b in units of bohr.4 M is the molar mass in g/mol,
and m is the frequency of the incident light in cmꢀ1. b is
ꢀ
the electric dipole-magnetic dipole polarizability, calcu-
lated by
X
1
b ¼
ðG0aaÞ
4.2.2. Preparation of (S)-(+)-1-(2-nitro-benzyl)-5-oxo-
pyrrolidine-2-carboxylic acid 5. The suspension of 2-
3x
a
where x is the frequency of the incident light, and G0 is
(2-nitro-benzylamino)-pentanedioic acid
4 (12.01 g,
the response function
42.5 mmol) in ethanol (420 mL) was heated at reflux
for 3 h. The resulting solution was filtered and concen-
X
x
G0ab ¼ ꢀ2
Im½hw0jlajwnihwnjmbjw0iꢁ
trated in vacuum to give an ivory-white dusty solid
x2n ꢀ x2
18
n¼0
10.91 g, yield 97.1%. Mp 193.9–196.6. ½aꢁD ¼ þ10:2 (c

M. Cheng et al. / Tetrahedron: Asymmetry 17 (2006) 179–183
183
0.50, DMSO). IR mmax (KBr) cmꢀ1, 3411, 2805, 2515,
depositing the crystal structure and providing the CIF
and mol2 file of (ꢀ)-linarinic acid.
1743, 1645, 1578, 1521, 1457, 1407, 1335, 1260, 1217,
1
975, 791, 728, 677. H NMR (DMSO-d6, 300 MHz) d
2.03 (1H, m), 2.41 (2H, m), 4.08 (1H, d, J = 8.9 Hz),
4.38 (1H, d, J = 16.8 Hz), 5.01 (1H, d, J = 16.8 Hz),
7.43 (1H, d, J = 7.7 Hz), 7.55 (1H, t, J = 7.6, 7.8 Hz),
7.72 (1H, t, J = 7.5, 7.6 Hz), 8.04 (1H, d, J = 8.1 Hz),
13.00 (1H, s). ESI-MS m/z: 265.1 (M++1).
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4.2.3. Preparation of (S)-(ꢀ)-linarinic acid 1. To a solu-
tion of 1-(2-nitro-benzyl)-5-oxo-pyrrolidine-2-carbox-
ylic acid 5 (10.00 g, 37.6 mmol) in ethanol (360 mL)
was added active carbon (1.00 g) and a catalytic amount
of ferric chloride (0.95 g, 3.5 mmol). Then the suspen-
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(5.52 mL, 113.5 mmol) was added dropwise. The mix-
ture was heated at reflux for about 64 h. Then the mix-
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vacuum evaporation. The residue was purified by flash
chromatography on silica gel using methanol–ethyl acet-
ate (5:4, v/v) to give a white solid 4.87 g, yield 59.6%;
18
mp 229.9–232.5 ꢁC dec. ½aꢁD ¼ ꢀ290:0 (c 0.01, MeOH).
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1
1376, 1292, 771. H NMR (CD3OD, 300 MHz) d 2.22–
2.31 (1H, m), 2.54–2.62 (1H, m), 2.98–3.10 (2H, m), 4.30
(1H, dd, J = 9.3, 4.1 Hz), 4.75 (1H, d, J = 15.4 Hz), 4.96
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7.28–7.33 (1H, m). 13C NMR (CD3OD, 75 MHz) d
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128.29, 130.28, 132.34, 165.00, 174.92. ESI-MS m/z:
217.0 (M++1), 215.0 (M+ꢀ1). EI-MS m/z: 216 (59.47),
215 (62.09), 171 (41.22), 169 (46.73), 145 (11.58), 144
(100.00), 129 (7.91), 118 (26.71), 91 (9.34), 77 (19.57),
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Acknowledgments
We thank Peter R. Taylor at Centre for Scientific Com-
puting, University of Warwick, United Kingdom for
discussion and help with the theoretical calculation of
the specific rotation. We also thank Gill Heale at Cam-
bridge Crystallographic Data Centre for her help of
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MO 63144, USA.