Chemoenzymatic preparation of optically active 4-aryl-5-carboxy-6-methyl-3, 4-dihydro-2(1H)-pyridone derivatives

Article abstract of DOI:10.1016/j.tet.2014.05.012

A series of racemic 4-aryl-5-(tert-butoxycarbonyl)-6-methyl-3,4-dihydro- 2(1H)-pyridones have been prepared by means of a modified Hantzsch reaction using commercially available starting materials. An easy removal of the tert-butyl group of these pyridones and subsequent reaction with cesium carbonate and chloromethyl 2-methylpropanoate provided us suitable substrates (±)-5 to be used in lipase-catalyzed hydrolysis reactions. Lipase B from Candida antarctica (CAL-B) was the most adequate lipase in the hydrolysis of (±)-5. Despite the low enantioselectivity values obtained (E≤12), several optically active pyridone derivatives were finally isolated with high enantiomeric excesses (ee≥91%) and moderate yields.

Full text of DOI:10.1016/j.tet.2014.05.012

Accepted Manuscript
Chemoenzymatic preparation of optically active 4-aryl-5-carboxy-6-methyl-3,4-
dihydro-2(1H)-pyridone derivatives
Susana Y. Torres , Estael Ochoa , Yamila Verdecia , Francisca Rebolledo
PII:
S0040-4020(14)00672-3
10.1016/j.tet.2014.05.012
TET 25561
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 9 April 2014
Revised Date: 2 May 2014
Accepted Date: 3 May 2014
Please cite this article as: Torres SY, Ochoa E, Verdecia Y, Rebolledo F, Chemoenzymatic preparation
of optically active 4-aryl-5-carboxy-6-methyl-3,4-dihydro-2(1H)-pyridone derivatives, Tetrahedron (2014),
doi: 10.1016/j.tet.2014.05.012.
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Chemoenzymatic preparation of optically
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dihydro-2(1H)-pyridone derivatives
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Susana Y. Torres,a,b Estael Ochoa,b Yamila Verdecia,b and Francisca Rebolledoa,
aDepartamento de Química Orgánica e Inorgánica, Universidad de Oviedo, 33006 Oviedo, Spain. bLaboratorio
de Síntesis Orgánica, Facultad de Química, Universidad de La Habana, 10400 La Habana, Cuba.

1
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Tetrahedron
journal homepage: www.elsevier.com
Chemoenzymatic preparation of optically active 4-aryl-5-carboxy-6-methyl-3,4-
dihydro-2(1H)-pyridone derivatives.
Susana Y. Torres,a,b Estael Ochoa,b Yamila Verdecia,b and Francisca Rebolledoa,
a Departamento de Química Orgánica e Inorgánica, Universidad de Oviedo, 33006 Oviedo, Spain.
b Laboratorio de Síntesis Orgánica, Facultad de Química, Universidad de La Habana, 10400 La Habana, Cuba.
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
A series of racemic 4-aryl-5-(tert-butoxycarbonyl)-6-methyl-3,4-dihydro-2(1H)-pyridones have
been prepared by means of a modified Hantzsch reaction using commercially available starting
materials. An easy removal of the tert-butyl group of these pyridones and subsequent reaction
with cesium carbonate and chloromethyl 2-methylpropanoate provided us suitable substrates
(±)-5 to be used in lipase-catalyzed hydrolysis reactions. Lipase B from Candida antarctica
(CAL-B) was the most adequate lipase in the hydrolysis of (±)-5. Despite the low
enantioselectivity values obtained (E ≤ 12), several optically active pyridone derivatives were
finally isolated with high enantiomeric excesses (ee ≥ 91%) and moderate yields.
Available online
Keywords:
3,4-Dihydro-2(1H)-pyridone
Resolution
2009 Elsevier Ltd. All rights reserved
.
Hydrolysis
Lipase
2-Methylpropanoyloxymethyl esters
1. Introduction
Given that the biological activity of a compound is closely
related to the configuration of its chiral centres, the synthesis of
optically active 3,4-DHP-2-ones is necessary to investigate their
pharmacological activities. In this sense, methods for the
preparation of enantioenriched 3,4-DHP-2-ones are scarce.
Among them, the resolution by preparative enantioselective
HPLC of a racemic mixture allowed to obtain an optically active
precursor of 2.3b Recently, the asymmetric synthesis of a 4-(p-
fluorophenyl)-5-carboxy-3,4-DHP-2-one derivative by means of
an organocatalyst has been published,4 as well as the preparation
The 3,4-dihydro-2(1H)-pyridone (3,4-DHP-2-one) core is
present in natural products such as the homoclausenamide
alkaloids 1,1 and the structural analogy of these heterocycles with
1,4-dihydropyridines making them good candidates for the
development of calcium channel modulators.2 In addition,
different functionalized 3,4-DHP-2-ones have been used as
precursors in the synthesis of biologically active molecules3
(Figure 1). For instance, several carboxamide derivatives 2 have
shown to be potent and selective α1a receptor antagonists, and
they could be used for the treatment of benign prostatic
hyperplasia. In this case, the (R)-(−) enantiomer was significantly
more active than its (S)-(+)-counterpart.3b
of
a
wide variety of optically active 4-substituted-5-
(alkoxycarbonyl or cyano)-3,4-DHP-2-ones derivatives by N-
heterocyclic carbene catalyzed aza-Claisen reactions.5
Taking the importance of the development of green
procedures into account, we decided to investigate the utility of
hydrolytic enzymes for the resolution of 3,4-DHP-2-one
derivatives. Nowadays, enzymes are recognized as excellent tools
for preparing optically active compounds, either by kinetic
resolution (KR) of racemic mixtures or by desymmetrization of
meso compounds.6 We report herein an efficient method to
prepare a set of racemic 4-aryl-5-carboxy-3,4-DHP-2-ones,
which have been subsequently converted into adequate substrates
to carry out its resolution via lipase-catalyzed hydrolysis
reactions. The resulting optically active compounds could be
used as precursors of benzodiazepine-dihydropyridine hybrid
molecules with potential activity as neuroprotective agents.2c
Figure 1. Some representative 3,4-dihydro-2(1H)-pyridone derivatives.
———
Corresponding author. Tel.: +34 985 102 982; fax: +34 985 103 446; e-mail: frv@uniovi.es.

2
Tetrahedron
acetone mixture. Although in almost all cases the yields of this
ACCEPTED MANUSCRIPT
last step were high (>75%), the poor results of the first step cut
2. Results and discussion
down the overall yields of the products (±)-9 (15-59%).
2.1. Preparation of racemic 3,4-DHP-2-one derivatives.
The syntheses of some racemic 4-aryl-6-methyl-5-
(methoxycarbonyl)-3,4-DHP-2-ones (±)-3 and their ethyl ester
analogues (±)-4 have previously been published.7 Some initial
attempts to carry out the hydrolysis of these esters using different
lipases (Enz-OH) were unsuccessful (Figure 2). This absence of
reactivity may be due to both steric and electronic factors. The
steric congestion around the ester function could avoid the
adequate fitting of the substrate in the active site of the enzyme.
In addition, the high electronic density at the olefinic C-5
position7a could diminish the electrophilic character of the
adjacent carbonyl carbon. These facts agree with the difficulties
previously found in the enzymatic hydrolysis of related esters
derived from 4-aryl-1,4-dihydropyridines.8 Similarly to those
cases, the hydrolysis resistance could be circumvented by
Scheme 1. Synthesis of (±)-9a-h from 2-cyanoethyl esters (±)-8a-h. The Ar
groups for 8 and 9 are the same shown in Table 1.
With the idea to enhance the yields of acids (±)-9, we decided
to check other esters whose conversion into acids does not imply
a nucleophilic attack to the carbonyl group. Thus, we planned to
access to compounds (±)-9 through the tert-butyl esters
derivatives (±)-10 (Table 1).
introducing
a
spacer such as, for instance, the
isobutyryloxymethyl unit beyond the carboxyl group. With these
new substrates (±)-5 (Fig. 2), the enzyme could be able to
catalyze the reaction of the outer ester group, the pyridone ring
being accommodated in the nucleophile binding site, and not in
the acyl donor site. Moreover, this outer ester function is
activated because it contains an optimal leaving group, which
spontaneously loses formaldehyde with the concomitant
formation of the corresponding non-racemic 4-aryl-5-carboxy-6-
Table 1. Synthesis of (±)-9 and tert-butyl esters (±)-10.a
O
Ar
O
Ar
O
O
Ar
H
But
TFA
PhOMe
But
O
OH
NH4OAc
O
6
+
CH2Cl2
AcOH
80 ºC
O
N
H
O
N
H
O
methyl-3,4-DHP-2-one 9
.
7b
10
(+)-
9
(+)-
Entry Ar
(±)-10
Yield
(%)b
(±)-9 Yield (%)c
1
2
3
4
5
6
7
8
Ph
10a
10b
10c
10d
10e
10f
61
69
72
73
74
88
84
92
9a
9b
9c
9d
9e
9f
52
52
58
50
53
65
76
83
2-NO2-C6H4
3-NO2-C6H4
4-NO2-C6H4
Figure 2. 4-Aryl-5-carboxy-6-methyl-3,4-DHP-2-one esters derivatives.
2-Cl-5-NO2-C6H3
4-Br-C6H4
To obtain diesters (±)-5 the corresponding carboxylic acids
(±)-9 are required as starting materials. However, several
attempts to carry out the basic hydrolysis of the easily available
compounds (±)-3 and (±)-4 failed. Thus, the treatment at room
temperature with NaOH or LiOH in a water-methanol mixture
was ineffective and the starting materials were recovered
unaltered after 48 h of reaction. Higher reaction temperatures led
to a complex mixture of products. On the other hand, some of
these acids have been prepared on a small scale (0.50 mmol) by
means of a four-step solid-phase synthesis.9
3-CH3O-C6H4
1-Naphthyl
10g
10h
9g
9h
aReactions were carried out using an equimolecular amount of 6, 7b, and
the aldehyde, and 50% excess of ammonium acetate in AcOH as the solvent.
bIsolated yields for (±)-10 after recrystallization or flash-chromatography.
cOverall two-steps yields for (±)-9.
With the idea to develop a straightforward and expeditious
alternative to the synthesis of carboxylic acids (±)-9, we initially
decided to prepare the 2-cyanoethyl ester derivatives (±)-8
(Scheme 1). In the presence of a base this kind of esters
experiences a β-elimination10 (initiated by the removal of the
proton α to the cyano group) affording the corresponding
carboxylate. Synthesis of (±)-8 was carried out similarly to the
preparation of (±)-3 or (±)-4, by means of a modified Hantzsch
reaction using the Meldrum’s acid 6.11 The four-component
reaction among equimolar amounts of 6, 2-cyanoethyl
acetoacetate 7a, and the appropriate aromatic aldehyde, as well as
1.5 equivalents of ammonium acetate in acetic acid at 110 ºC,
gave the corresponding 2-cyanoethyl esters (±)-8 with low to
moderate yields (Scheme 1). The removal of the cyanoethyl
group, and thus the conversion of esters (±)-8 into acids (±)-9,
easily took place by treatment with NaOH (4 equiv) in a water-
The four component reaction among 6, tert-butyl acetoacetate
7b, the corresponding aldehyde, and ammonium acetate, all of
them commercially available, happened at lower reaction
temperature (80 ºC) than the reaction using 7a. Consequently, the
formation of side compounds decreased drastically, and the tert-
butyl esters (±)-10 were obtained in higher yields (Table 1) than
those of (±)-8 (Scheme 1). After testing several reaction
conditions for the removal of the tert-butyl group of (±)-10, the
best results were obtained at 0 ºC using trifluoroacetic acid and
anisole as a cation scavenger.12 Thus, acids (±)-9 were finally
isolated in 50-83% overall yields (Table 1).
Transformation of carboxylic acids (±)-9 into the diesters (±)-
5 was easily carried out in one pot process by treatment with
cesium carbonate (1.6 equiv) in DMF, and subsequent reaction of
the carboxylate with a slight excess of chloromethyl isobutyrate,

3
at room temperature (Scheme 2). Isolated yields after the
purification of (±)-5 by flash-chromatography were very high
(78-91%).
Next, the best enzymatic hydrolysis conditions were applied
ACCEPTED MANUSCRIPT
to all the diesters (±)-5a-h (for the nature of each Ar substituent,
see Table 1). Some of the results obtained are collected in Table
3. Reactions using (±)-5b,d,e,h as starting materials happened
with E ≤ 2 and therefore were not included in Table 3. The E
value obtained in the reaction with substrate (±)-5a (Ar = Ph)
was also very low (Table 3, entry 1), the ee for the remaining
substrate (S)-5a and the product (R)-9a being very low (32% and
47% ee, respectively). However, taking advantage of the different
solubility of the racemic diester (±)-5a and the enantiopure form,
it was possible to access to (S)-5a with very high ee after a
simple recrystallization of the moderately enantioenriched
sample (see below in the text).
Scheme 2. Synthesis of diesters (±)-5a-h.
2.2. Enzymatic hydrolysis of (±)-5.
In order to check the enzymatic kinetic resolution of diesters
(±)-5, the 3-nitrophenyl derivative (±)-5c was chosen as a
substrate model. To improve the dissolution of the diester, the
enzymatic hydrolysis reactions were carried out in organic
solvents. Several combinations of lipases (PSL, CRL, CAL-A,
and CAL-B) and organic solvents were tested and a selection of
the results obtained is included in Table 2, entries 1-5. The best
results were achieved with CAL-B in tert-butyl methyl ether
(TBME), at 28 ºC (entry 3). In these conditions, the enantiomeric
excess (ee) of either the remaining substrate and the product was
only moderate, the enantioselectivity value being low (E = 12).13
Results achieved in the CAL-B-catalyzed hydrolysis of (±)-5f
and (±)-5g (Table 3, entries 3 and 4) were similar to those
obtained with 5c, the ee for both substrate and product being
moderate at a degree of conversion near to 50% (E = 10).
Nevertheless, from these reactions happening with E values near
to 10, the remaining substrates can be reached with high ee at
degrees of conversion around 65%. Effectively, when enzymatic
hydrolysis of (±)-5c,f,g were conducted to longer reaction times
(Table 3, entries 5-7) substrates (S)-5c,f,g were isolated with high
ee (93-95%) and moderate yields (30-31%), considering the
maximal 50% yield of a kinetic resolution. It is worth noting that
isolation of optically active compounds (S)-5 and (R)-9 was
easily carried out by base-acid extraction.
Table 2. Enzymatic hydrolysis of diesters (±)-5c and (±)-11.a
NO2
NO2
NO2
On the other hand, we also try to enhance the ee of these
compounds by recrystallization of the enantioenriched samples,
seeing as the racemates usually crystallize better than the
corresponding optically active samples. Based on this fact,
substrate (S)-5c (ee = 73%), isolated from the enzymatic
hydrolysis reaction (Table 3, entry 2), was recrystallized in
diethyl ether. After a slow crystallization, the solid was filtered
and the enantiomeric excesses of both the solid and the product
recovered from the filtrate were measured by enantioselective
HPLC. Whereas the crystallized solid (22% of recovering) was
almost racemic (ee = 2%), the ee of the compound (S)-5c
obtained from the filtrate (71% of recovering) was very high
(95% ee). This highly enantioenriched (S)-5c was obtained as a
viscous oil, with an overall yield (taking the two steps, enzymatic
hydrolysis and crystallization, into account) of 32%. These
values (ee and yield) are similar to those obtained from the
kinetic resolution at c = 64% (Table 3, entry 5). In addition,
recrystallization of (S)-5a with 32% ee (Table 3, entry 1) allowed
us to access to a 13% overall yield of almost enantiopure (S)-5a
(99% ee). However, when this method was applied to
enantioenriched (S)-5f,g of 71 and 69% ee, respectively, the same
trends was observed in both cases, but the ee of the resulting
samples were lower than 90%. From these results we can
conclude that the most adequate method to obtain the
enantioenriched esters (S)-5c,f,g (ee >90%) is by means of
enzymatic hydrolysis to degrees of conversion around 65%.
O
O
O
O
O
O
O
H2O
CAL-B
O
R
O
R
OH
+
Solvent
O
N
H
O
N
H
O
N
H
(+)-5c (R = i-Pr)
(+)-11 (R = Me)
(S)-5c (R = i-Pr)
(S)-11 (R = Me)
(R)-9c
(+)-12 (R = t-Bu)
Entry
R
Solvent
t, h eeS (%)b
eeP (%)c
32
cd
Ee
3
1
2
3
4
5
6
7
i-Pr
i-Pr
i-Pr
i-Pr
i-Pr
Me
Me
1,4-Dioxane
DIPE
9
58
78
80
44
44
35
91
64
65
54
41
38
73
65
24
9
42
5
TBME
67
12
7
TBMEf
9
64
TBMEg
1,4-Dioxane
TBME
3
72
9
2
13
2
2
49
9
aReactions were carried out at 28 ºC and 200 rpm using 25 mg of substrate
and the following solvents: water-saturated diisopropyl ether (15 mL) or
TBME (2.5 mL); or a mixture of 1,4-dioxane (1.0 mL) and H2O (50 µL).
bThe ee of remaining substrates 5c or 11 (eeS) was determined by
c
enantioselective HPLC analysis. The ee of the produced acid 9c (eeP) was
determined after treatment with diazomethane, and then enantioselective
HPLC analysis of the resulting methyl ester derivative 3c. dThe degree of
conversion (%) was calculated from eeS and eeP: c = 100eeS / (eeS + eeP). eSee
Several attempts to enhance the ee of the carboxylic acids (R)-
9 isolated from the enzymatic reactions were also carried out.
Since the recrystallization attempts failed for compounds (R)-9,
we decided to convert the enantioenriched samples (R)-9c,f,g into
the esters (R)-5c,f,g and submit these substrates to enzymatic
hydrolysis again. Thus, after 2-3 h of reaction, the new acids (R)-
9c,f,g were isolated with very high ee (>90%) and moderate
yields (Table 3, entries 8-10). Considering the three steps (KR-
esterification-KR), the overall yields for these acids were in the
20-25% range. 14
Ref. 13. Reaction was carried out at 10 ºC. gA two-phase 3:1 v/v system
f
TBME-H2O was used.
Other diesters such as the acetyl (±)-11 and pivaloyl (±)-12
derivatives (Table 2) were also prepared and checked in the
enzymatic hydrolysis. Whereas pivaloyl derivative was not
transformed, the enzyme catalyzed the hydrolysis of the acetyl
derivative (±)-11 quicker than that of (±)-5c, but with a slightly
lower E value (Table 2, entry 7). Besides hydrolysis, several
enzymatic transesterifications and aminolysis reactions of (±)-5c
were essayed in the presence of CAL-B, but the results were
always poorer than those attained in the hydrolysis reaction.

4
Tetrahedron
ACCEPTED MANUSCRIPT
Table 3. Enzymatic hydrolysis of diesters (±)-5.a
Entry
Starting
diester
Ar
Time
(h)
Remaining substrate
Product
c (%)
E
Substrate
(S)-5a
(S)-5c
(S)-5f
Yield (%)
56
eeS (%)b
32d
73e
71
Product
(R)-9a
(R)-9c
(R)-9f
(R)-9g
(R)-9c
(R)-9f
(R)-9g
(R)-9c
(R)-9f
(R)-9g
Yield (%) eeP (%)c
1
(±)-5a
(±)-5c
(±)-5f
(±)-5g
(±)-5c
(±)-5f
(±)-5g
(R)-5cf
(R)-5fg
(R)-5gh
Ph
9
39
47
71
64
66
53
48
50
95
93
91
41
51
52
51
64
66
65
4
2
3-NO2-C6H4
4-Br-C6H4
3-CH3O-C6H4
3-NO2-C6H4
4-Br-C6H4
3-CH3O-C6H4
3-NO2-C6H4
4-Br-C6H4
3-CH3O-C6H4
9
47
46
48
31
31
30
48
12
10
10
11
9
3
11
7
46
4
(S)-5g
(S)-5c
(S)-5f
69
48
5
16
18
12
3
95
62
6
93
59
7
(S)-5g
95
61
10
8
52 (24)i
41 (20)i
49 (25)i
9
2.5
2
10
aReactions were carried out at 100-1000 mg scale amount (see Experimental Section) using TBME saturated with water as solvent, at 28 ºC and 200 rpm. The
degree of conversion (c, %) and the E values were calculated as in Table 2.
bDetermined by enantioselective HPLC analysis (see Sec. 4.14). High ee values are bolded.
cDetermined by enantioselective HPLC analysis after treatment of products 9 with diazomethane (see Sec. 4.14). High ee values are bolded.
d(S)-5a almost enantiopure (99% ee) was isolated after submitting this sample to crystallization (see Sec. 4.10.1).
eRecrystallization of this sample afforded to (S)-5c with 95% ee (see Sec. 4.11.1).
fAn enantioenriched sample of (R)-5c with eeo = 67% was used as substrate.
gAn enantioenriched sample of (R)-5f with eeo = 62% was used as substrate.
hAn enantioenriched sample (R)-5g with eeo = 57% was used as substrate.
iThe corresponding overall yield calculated from racemic 5 is given between brackets.
Finally, optically active esters (S)-5 were smoothly converted
into the acids (S)-9 by conventional basic hydrolysis (aq 3N
NaOH, acetone, RT). No racemization took place in these
reactions as proven by enantioselective HPLC.
acyloxymethyl esters, but the E values obtained for these
processes were low. The remoteness between the asymmetric
carbon and the reactive carbonyl function could be the reason for
this poor enantioselectivity.15 Nevertheless, compounds with high
enantiomeric excesses and moderate yields were prepared by
combining either the enzymatic resolution with a selective
crystallization process or two consecutive KRs. As a result, both
enantiomers of the pyridone-carboxylic acids have been
achieved. The biological activities of these optically active
pyridone-carboxylic acids as well as some of their derivatives are
currently under study.
In order to establish the enantiopreference of the lipase in
these process, product 9f (ee = 93%, see Table 3, entry 9) was
treated with cesium carbonate in DMF and then with methyl
iodide. The resulting methyl ester (−)-3f ([α]D20 = −100.6 (c 1.05,
CHCl3)) retained the ee as shown by the enantioselective HPLC
analysis. Comparison of the sign of the optical rotation of this
ester with the published value5 establishes the (R) configuration
for (−)-3f and thus, for the acid 9f proceeding from the enzymatic
reaction. That means that CAL-B preferently catalyzes the
hydrolysis of the (R) enantiomer of the diester 5f. Based on the
structural resemblance among all the substrates reported here, we
have tentatively assigned the (R) configuration to the other
produced acids 9. In addition the (S) configuration for the
remaining optically active substrates 5a,c,g was also assigned.
4. Experimental section
Lipase B from Candida antarctica (CAL-B, Novozyme 435,
available immobilized on polyacrylamide, 7300 PLU/g) was
gifted by Novo Nordisk Co. Immobilized lipase A from Candida
antarctica (CAL-A, NZL-101, 6.2 U/g) was purchased from
Codexis. Immobilized lipase from Burkholderia cepacia (PSL-
IM, 783 U/g), which previously was classified as Pseudomonas
cepacia, was purchased from Amano Pharmaceutical Co.
Melting points were taken on samples in open capillary tubes and
are uncorrected. For the enzymatic hydrolysis reaction tert-butyl
methyl (TBME) saturated with water was used. IR spectra were
3. Conclusion
We have developed an efficient method to obtain some
carboxylic acids derived from 3,4-DHP-2-ones, as well as
several acyloxymethyl ester derivatives which are suitable
substrates to be used in lipase-catalyzed hydrolysis reactions.
Lipase B from Candida antarctica catalyzed the hydrolysis of the
1
recorded using KBr pellets. H NMR and proton-decoupled 13C
NMR spectra (CDCl3 solutions) were obtained using AC-300 or

5
DPX300 (1H, 300.13 MHz and 13C, 75.5 MHz) spectrometers
148.6 (C-2’), 150.9 (C-6), 165.5 (CO), 168.9 (C-2); HRMS
(ESI+): MH+, found: 330.1084. C16H16N3O5 requires 330.1069.
ACCEPTED MANUSCRIPT
using the δ scale (ppm) for chemical shifts. Calibration was made
on the signal of the solvent (13C: CDCl3, 76.95; DMSO-d6, 39.52
ppm) or the residual solvent partially or non-deuterated (1H:
CHCl3, 7.26; DMSO-d5, 3.50 ppm).
4.2.3. (±)-5-[(2-Cyanoethyl)oxycarbonyl] -6-methyl-
4-(3-nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-
8c]
In Figure 3 we show the 4-aryl-3,4-DHP-2-one unit with the
numbering used in the assignation of the NMR signals.
Reaction time: 16 h. White solid; m.p.: 158-160 ºC; yield
51%; νmax(KBr) 3213, 1707, 1687, 1623, 1527, 1485, and 1350
cm−1; δH (300.13 MHz, CDCl3) 2.49 (s, 3H, CH3), 2.63 (m, 2H,
CH2CN), 2.73 (br d, 1H, |2J| 16.6 Hz, HH-3), 3.03 (dd, 1H, 3J 7.6,
3
|2J| 16.6 Hz, HH-3), 4.28 (m, 2H, O-CH2), 4.38 (br d, 1H, J 7.6
Hz, H-4), 7.60-7.43 (m, 2H, H-5’ and H-6’), 7.79 (br s, 1H, NH),
4
3
4
8.03 (t, 1H, J 1.7 Hz, H-2’), 8.11 (dt, 1H, J 7.7, J 1.4 Hz, H-
4’); δC (75.5 MHz, CDCl3) 18.2 (CH2-CN), 19.8 (CH3), 37.8 (C-
3), 37.9 (C-4), 58.8 (O-CH2), 105.0 (C-5), 116.9 (C≡N), 121.8
(C-4’), 122.6 (C-2’), 130.2 (C-5’), 133.2 (C-6’), 144.1 (C-1’),
148.8 (C-3’), 149.2 (C-6), 165.7 (CO), 169.6 (C-2); MS (EI), m/z
(%) = 329 (M•+, 83), 312 (100), 259 (64), 231 (93); HRMS
(ESI+): MH+, found: 330.1084. C16H16N3O5 requires 330.1094.
Figure 3. 4-Aryl-3,4-DHP-2-one derivatives.
4.1. 2-Cyanoethyl 2-oxobutanoate (7a).
4.2.4. (±)-5-[(2-Cyanoethyl)oxycarbonyl] -6-methyl-
4-(4-nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-
8d]
A solution of 1,3-dioxin-4-one (10.0 mL, 76.5 mmol) and 3-
hydroxypropanonitrile (5.5 mL, 80 mmol) in toluene (15 mL)
was heated at reflux during 24 h. Elimination of the solvents
under reduced pressure yielded 7a (11.3 g, 95%). Spectroscopic
data for 7a are in good agreement with those previously
published.16
Reaction time: 13 h. White solid; m.p.: 118-119 ºC; yield
27%; νmax(KBr) 3220, 1697, 1631, 1567, 1417, and 1349 cm−1;
δH (300.13 MHz, CDCl3) 2.46 (s, 3H, CH3), 2.75-2.50 [m + br d,
3H, CH2CN and doublet centered to 2.68 corresponding to HH-3
4.2. General procedure for the synthesis of 2-cyanoethyl
esters (±)-8.
3
(|2J| 16.7 Hz)], 3.02 (dd, 1H, J 8.3, |2J| 16.6 Hz, HH-3), 4.43-
4.15 [m + br d, 3H, O-CH2 and doublet centered to 4.37
corresponding to H-4 (3J 8.1 Hz)], 7.36 (d, 2H, 3J 8.3 Hz, H-2’
and H-6’), 8.15 (d, 2H, 3J 8.4 Hz, H-3’ and H-5’), 8.54 (br s, 1H,
NH); δC (75.5 MHz, CDCl3) 18.2 (CH2-CN), 19.6 (CH3), 37.7
(C-3), 38.0 (C-4), 58.8 (O-CH2), 104.9 (C-5), 117.0 (C≡N), 124.4
(CH), 127.8 (CH), 147.3 (C), 149.2 (C), 149.5 (C), 165.6 (CO),
170.2 (C-2); HRMS (ESI+): MH+, found: 330.1084. C16H16N3O5
requires 330.1071.
To a solution of 2,2-dimethyl-1,3-dioxane-4,6-dione (4.73 g,
32.8 mmol) in acetic acid (9.5 mL), the aromatic aldehyde (32.8
mmol), 2-cyanoethyl 3-oxobutanoate (5.09 g, 32.8 mmol), and
ammonium acetate (3.78 g, 49.1 mmol) were added. After
refluxing during 7-16 h, the reaction mixture was poured into ice-
water, and the precipitation of (±)-8 took place. The solid was
filtered and successively washed with water and cold diethyl
ether yielding the corresponding ester (±)-8, which was purified
by flash chromatography (hexane/ethyl acetate mixtures).
4.2.5. (±)-4-(2-Chloro-5-nitrophenyl)-5-[(2-
cyanoethyl)oxycarbonyl] -6-methyl-3,4-dihydro-
2(1H)-pyridone [(±)-8e]
4.2.1. (±)-5-[(2-Cyanoethyl)oxycarbonyl] -6-methyl-
Reaction time: 13 h. White solid; m.p.: 161-163 ºC; yield
66%; νmax(KBr) 3222, 1705, 1685, 1643, 1525, and 1350 cm−1;
δH (300.13 MHz, CDCl3) 2.79-2.49 [m + s, 5H, CH2CN and
singlet centered to 2.55 corresponding to CH3], 2.72 (br d, 1H,
4-phenyl-3,4-dihydro-2(1H)-pyridone [(±)-8a]
Reaction time: 14 h. White solid; mp 110-111 ºC; yield 17%;
νmax(KBr) 3226, 1705, 1690, 1635, and 1525 cm−1; δH (300.13
MHz, CDCl3) 2.43 (s, 3H, CH3), 2.77-2.47 (m, 3H, CH2CN and
3
|2J| 16.8 Hz, HH-3), 3.01 (dd, 1H, J 8.6, |2J| 16.8 Hz, HH-3),
3
HH-3), 2.96 (dd, 1H, J 8.2, |2J| 16.6 Hz, HH-3), 4.33-4.15 (m,
3
4.30-4.15 (m, 2H, O-CH2), 4.78 (br d, 1H, J 8.3 Hz, H-4), 7.60
3H, O-CH2 and H-4), 7.36-7.12 (m, 5H, Ph), 7.83 (br s, 1H, NH);
δC (75.5 MHz, CDCl3) 17.9 (CH2-CN), 19.0 (CH3), 37.8 (C-4),
38.1 (C-3), 58.4 (O-CH2), 105.8 (C-5), 117.0 (C≡N), 126.6 (CH),
127.1 (C-4’), 128.8 (CH), 141.9 (C), 148.4 (C), 166.0 (CO),
171.4 (C-2); HRMS (ESI+): MH+, found: 385.1234. C16H17N2O3
requires 385.1219.
3
4
(br d, 1H, J 8.7 Hz, H-3’), 7.86 (d, 1H, J 2.6 Hz, H-6’), 8.07
4
3
(dd, 1H, J 2.6, J 8.7 Hz, H-4’), 8.16 (br s, 1H, NH); δC (75.5
MHz, CDCl3) 18.1 (CH2-CN), 19.6 (CH3), 35.4 (C-4), 35.9 (C-3),
58.9 (O-CH2), 103.8 (C-5), 116.6 (C≡N), 122.4 (CH), 123.8
(CH), 131.5 (C-3’), 140.4 (C), 140.4 (C), 147.1 (C-2’), 150.6 (C-
5’), 165.3 (CO), 169.2 (C-2); HRMS (ESI+): MH+, found:
364.0695. C16H15ClN3O5 requires 364.0687.
4.2.2. (±)-5-[(2-Cyanoethyl)oxycarbonyl] -6-methyl-
4-(2-nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-
8b]
4.2.6. (±)-4-(4-Bromophenyl-5-[(2-
cyanoethyl)oxycarbonyl] -6-methyl-3,4-dihydro-
2(1H)-pyridone [(±)-8f]
Reaction time: 14 h. White solid; m.p.: 146-149 ºC; yield
26%; νmax(KBr) 3223, 1710, 1698, 1636, 1522, 1491, and 1342
cm−1; δH (300.13 MHz, CDCl3) 2.58-2.42 [m + s, 5H, CH2CN
and singlet centered to 2.49 corresponding to CH3], 2.84 (br d,
1H, |2J| 16.5 Hz, HH-3), 3.09 (dd, 1H, 3J 9.2, |2J| 17.2 Hz, HH-3),
Reaction time: 14 h. White solid; m.p.:144-146 ºC; yield 47%;
ν
max(KBr) 3209, 1705, 1690, 1630, and 1487 cm−1; δH (300.13
MHz, CDCl3) 2.44 (s, 3H, CH3), 2.72-2.50 (m, 3H, CH2CN and
3
HH-3), 2.95 (dd, 1H, J 8.2, |2J| 16.6 Hz, HH-3), 4.35-4.15 (m,
3
4.16 (m, 2H, O-CH2), 4.74 (br d, 1H, J 9.0 Hz, H-4), 7.33-7.19
3H, O-CH2 and H-4), 7.06 [d, 2H, 3J 8.4 Hz, H-2’ and H-6’], 7.41
3
4
(m, 1H, H-6’), 7.40 [td, 1H, J 7.7 (t), J 1.1 (d) Hz, H-4’], 7.52
[td, 1H, 3J 7.7 (t), 4J 0.9 (d) Hz, H-5’], 7.84 (dd, 1H, 3J 8.1, 4J 1.0
Hz, H-3’); δC (75.5 MHz, CDCl3) 17.2 (CH2-CN), 18.5 (CH3),
33.3 (C-4), 37.1 (C-3), 58.9 (O-CH2), 103.0 (C-5), 118.1 (C≡N),
124.7 (C-3’), 127.8 (CH), 128.3 (CH), 133.7 (CH), 136.6 (C-1’),
3
[d, 2H, J 8.5 Hz, H-3’ and H-5’], 8.00 (br s, 1H, NH); δC (75.5
MHz, CDCl3) 18.2 (CH2-CN), 19.6 (CH3), 37.6 (C-4), 38.1 (C-3),
58.7 (O-CH2), 105.7 (C-5), 117.0 (C≡N), 121.1 (C), 128.6 (C-2’
and C-6’), 132.2 (C-3’ and C-5’), 141.0 (C), 148.5 (C), 165.9
(CO), 170.6 (C-2); HRMS (ESI+): MH+, found: 363.0339.
C16H16BrN2O3 requires 363.0339.

6
Tetrahedron
4.2.7. (±)-5-[(2-Cyanoethyl)oxycarbonyl] -4-(3-
4.3.2. (±)-4-(4-Bromophenyl)-5-carboxy-6-methyl-
ACCEPTED MANUSCRIPT
methoxyphenyl)-6-methyl-3,4-dihydro-2(1H)-
pyridone [(±)-8g]
3,4-dihydro-2(1H)-pyridone [(±)-9f]
White solid, m.p.: 199-201 ºC; yield 94%; νmax(KBr) 3414;
1694; 1676; 1639; 1488 cm−1; δH (300.13 MHz, DMSO-d6) 2.31
Reaction time: 14 h. White solid; m.p.: 123-124 ºC; yield
34%; νmax(KBr) 3550, 3475, 3414, 3224, 1705, 1692, 1620,
1579, and 1489 cm−1; δH (300.13 MHz, CDCl3) 2.42 (s, 3H,
CH3), 2.74-2.47 (m, 3H, CH2CN and HH-3), 2.94 (dd, 1H, 3J 8.2,
|2J| 16.6 Hz, HH-3), 3.77 (s, 3H, O-CH3), 4.33-4.15 (m, 3H, O-
CH2 and H-4), 6.73-6.69 (m, 1H, Ph), 6.80-6.73 (m, 2H, Ph), 7.21
(s, 3H, CH3), 2.36 (d, 1H, |2J| 16.1 Hz, HH-3), 2.92 (dd, 1H, J
3
7.8, |2J| 16.2 Hz, HH-3), 4.08 (d, 1H, J 7.2 Hz, H-4), 7.10 (d,
3
3
3
2H, J 8.4 Hz, H-2’, H-6’), 7.48 (d, 2H, J 8.5 Hz, H-3’, H-5’),
9.84 (s, 1H, NH), 12.01 (s, 1H, COOH); δC (75.5 MHz, DMSO-
d6) 18.2 (CH3), 37.1 (C-4), 38.2 (C-3), 105.0 (C-5), 119.5 (C-4’),
129.0 (C-2’ and C-6’), 131.4 (C-3’ and C-5’), 142.2 (C-1’), 148.0
(C-6), 168.1 (CO), 169.6 (C-2); MS (APCI+), m/z (%) = 266
([(M+H) − CO2]+, 100). Anal. calc. for C13H12BrNO3: C, 50.34;
H, 3.90; N, 4.52. Found: C, 50.12; H, 3.99; N, 4.78.
3
(t, 1H, J 7.9 Hz, H-5’), 8.27 (br s, 1H, NH); δC (75.5 MHz,
CDCl3) 18.2 (CH2-CN), 19.5 (CH3), 38.1 (C-4), 38.3 (C-3), 55.3
(O-CH3), 58.6 (O-CH2), 105.9 (C-5), 112.0 (CH), 113.0 (CH),
117.1 (C≡N), 119.1 (CH), 130.1 (CH), 143.7 (C), 148.3 (C),
160.0 (C-3’), 166.1(CO), 170.8 (C-2); HRMS (ESI+): MNa+,
found: 337.1159. C17H18N2NaO4 requires 337.1151.
4.3.3. (±)-5-Carboxy-4-(3-methoxyphenyl)-6-
methyl-3,4-dihydro-2(1H)-pyridone [(±)-9g]
4.2.8. (±)-5-[(2-Cyanoethyl)oxycarbonyl] -6-methyl-
4-(1-naphthyl)-3,4-dihydro-2(1H)-pyridone [(±)-
8h]
White solid m.p.: 204-205 ºC; yield 76%; νmax(KBr) 3448,
3212, 1695, 1673, 1639, 1599, 1489, and 1367 cm−1; δH (300.13
MHz, DMSO-d6) 2.31 (s, 3H, CH3), 2.39 (d, 1H, |2J| 15.9 Hz,
3
HH-3), 2.90 (dd, 1H, J 7.8, |2J| 16.1 Hz, HH-3), 3.71 (s, 3H,
Reaction time: 7h. White solid; m.p.: 197-198 ºC; yield 52%;
νmax(KBr) 3227; 1707; 1695; 1633; 1527 cm−1; δH (300.13 MHz,
CDCl3) 2.58-2.20 [m + s, 5H, CH2CN and singlet centered to
2.52 corresponding to CH3], 2.80 (br d, 1H, |2J| 17.3 Hz, HH-3),
3
OCH3), 4.07 (d, 1H, J 7.3 Hz, H-4), 6.80-6.65 (m, 3H, H-2’, H-
3
4’, and H-6’), 7.20 (t, 1H, J 7.9 Hz, H-5’), 9.78 (s, 1H, NH),
11.96 (s, 1H, COOH); δC (75.5 MHz, DMSO-d6) 18.2 (CH3),
37.5 (C-4), 38.4 (C-3), 54.9 (OCH3), 105.3 (C-5), 111.3 (CH),
112.9 (CH), 118.7 (CH), 129.6 (CH), 144.4 (C), 147.6 (C), 159.4
(C-3’), 168.2 (CO), 169.7 (CO); HRMS (ESI+): MH+, found:
262.1074. C14H16NO4 requires 262.1060.
3
3.09 (dd, 1H, J 8.6, |2J| 17.3 Hz, HH-3), 4.14-3.97 (m, 1H, O-
3
CHH), 4.27-4.14 (m, 1H, O-CHH), 5.14 (br d, 1H, J 8.3 Hz, H-
3
3
4), 7.18 (d, 1H, J 7.7 Hz, H-2’), 7.35 (t, 1H, J 7.7 Hz, H-3’),
3
7.66-7.43 [m, 2H, H-6’ and H-7’], 7.75 (d, 1H, J 8.2 Hz, H-4’),
7.89 (d, 1H, 3J 7.8 Hz, H-5’), 7.96 (br s, 1H, NH), 8.08 (d, 1H, 3J
8.4 Hz, H-8’); δC (75.5 MHz, CDCl3) 17.9 (CH2-CN), 19.4
(CH3), 33.7 (C-4), 37.5 (C-3), 58.5 (O-CH2), 105.7 (C-5), 116.8
(C≡N), 122.7 (C-8’), 123.1 (C-2’), 125.5 (C-3’), 125.8 (C-6’),
126.6 (C-7’), 128.1 (C-4’), 129.4 (C-5’), 130.6 (C), 134.6 (C),
136.4 (C), 149.1 (C), 166.1 (CO), 170.7 (C-2); HRMS (ESI+):
MH+, found: 335.1390. C20H19N2O3 requires 335.1554.
4.3.4. (±)-5-Carboxy-6-methyl-4-(1-naphthyl)-3,4-
dihydro-2(1H)-pyridone [(±)-9h]
White solid, m.p.: 200-201 ºC; yield 90%; νmax(KBr) 3550,
3478, 3414, 3219, 1691, 1640, 1606, and 1486 cm−1; δH (300.13
MHz, DMSO-d6) 2.74-2.13 (m + s, 4H, HH-3 and singlet
3
centered to 2.43 corresponding to CH3), 3.10 (dd, 1H, J 8.4, |2J|
16.1 Hz, HH-3), 4.97 (d, 1H, 3J 8.0, H-4), 7.14 (d, 1H, 3J 7.0 Hz,
3
4.3. General procedure for the hydrolysis of 2-cyanoethyl
esters (±)-8.
H-2’), 7.41 (t, 1H, J 7.5 Hz, H-3’), 7.73-7.48 [m, 2H, H-6’ and
3
3
H-7’], 7.80 (d, 1H, J 8.2 Hz, H-4’), 7.96 (d, 1H, J 7.5 Hz, H-
5’), 8.17 (d, 1H, 3J 8.2 Hz, H-8’), 9.86 (s, 1H, NH), 11.92 (s, 1H,
COOH); δC (75.5 MHz, DMSO-d6) 18.3 (CH3), 33.6 (C-4), 37.8
(C-3), 105.0 (C-5), 122.7 (CH), 123.1 (CH), 125.5 (CH), 125.6
(CH), 126.3 (CH), 127.2 (CH), 128.9 (CH), 130.2 (C), 134.0 (C),
137.1 (C), 148.7 (C), 168.2 (C-2), 169.5 (CO); HRMS (ESI+):
MH+, found: 282.1125. C17H16NO3 requires 282.1121.
To a solution of the cyanoethyl ester (±)-8 (5.0 mmol) in
acetone (70 mL), aq 3 M NaOH (20 mmol) and water (140 mL)
were added. The solution was stirred at room temperature until
disappearance of the starting material (TLC control). Then,
acetone was eliminated under reduced pressure and the basic
aqueous phase extracted with dichloromethane (2 x 30 mL).
After, aq 3 M HCl was added to the aqueous phase until pH was
1-2. The resulting acid aqueous phase was extracted with ethyl
acetate (3 x 60 mL). The organic layers were combined, washed
with brine (50 mL), dried with Na2SO4, and evaporated under
reduced pressure to give the corresponding carboxylic acid (±)-9
as a pure product. Spectroscopic data for 9a-d (white solids) are
in good agreement with those previously published.17 Yields
obtained were: 9a (90%), 9b (88%), 9c (60%), and 9d (78%).
4.4. Synthesis of tert-butyl esters (±)-10.
tert-Butyl esters (±)-10 (up to a scale of 5.0 mmol) were
prepared from tert-butyl acetoacetate (7b) following the
procedure described for the synthesis of (±)-8, except that
reactions were conducted at 80 ºC.
4.4.1. (±)-5-(tert-Butoxycarbonyl)-6-methyl-4-
phenyl-3,4-dihydro-2(1H)-pyridone [(±)-10a]
Reaction time: 11 h. White solid; mp 155-156 ºC; yield 61%;
νmax(KBr) 3414, 3218, 1690, 1634, 1483, 1384, and 1366 cm−1;
δH (300.13 MHz, CDCl3) 1.35 (s, 9H, But); 2.36 (s, 3H, CH3),
2.68 (dd, 1H, 3J 2.7, |2J| 16.5 Hz, HH-3), 2.92 (dd, 1H, 3J 8.1, |2J|
4.3.1. (±)-5-Carboxy-4-(2-chloro-5-nitrophenyl)-6-
methyl-3,4-dihydro-2(1H)-pyridone [(±)-9e]
White solid, m.p.: 220-221 ºC; yield 90%; νmax(KBr) 3245;
1709; 1676; 1622; 1525 cm−1; δH (300.13 MHz, DMSO-d6) 2.33
3
3
(d, 1H, |2J| 16.3 Hz, HH-3), 2.42 (s, 3H, CH3), 3.10 (dd, 1H, J
16.5 Hz, HH-3), 4.18 (br d, 1H, J 7.8 Hz, H-4), 7.32-7.12 (m,
3
8.3, |2J| 16.5 Hz, HH-3), 4.54 (d, 1H, J 8.0 Hz, H-4), 7.76 (d,
5H, Ph), 7.56 (br s, 1H, NH); δC (75.5 MHz, CDCl3) 19.1 (CH3),
28.3 (3 × CH3), 38.3 (C-3), 38.6 (C-4), 80.6 (C), 109.1 (C-5),
126.8 (CH), 126.9 (C-4’), 128.9 (CH), 142.7 (C), 144.7 (C),
166.3 (CO), 171.0 (C-2); HRMS (ESI+): MH+, found: 288.1594.
C17H22NO3 requires 288.1595.
4
3
1H, J 2.7 Hz, H-6’), 7.82 (br d, 1H, J 8.7, H-4’), 8.13 (dd, 1H,
4J 2.7, J 8.7 Hz, H-3’), 10.10 (s, 1H, NH), 12.17 (br s, 1H,
3
COOH); δC (75.5 MHz, DMSO-d6) 18.1 (CH3), 35.4 (C-4), 35.8
(C-3), 103.6 (C-5), 121.7 (CH), 123.7 (CH), 131.6 (C-3’), 139.5
(C), 141.3 (C), 146.7 (C), 149.8 (C), 167.6 (CO), 168.8 (C-2);
HRMS (ESI+): MH+, found: 311.0429. C13H12ClN2O5 requires
311.0399.
4.4.2. (±)-5-(tert-Butoxycarbonyl)-6-methyl-4-(2-
nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-10b]
Reaction time: 7 h. White solid; mp: 193-194 ºC (dec.); yield
69%; νmax(KBr) 3480, 3416, 3220, 1699, 1671, 1638, 1524, and

7
1365 cm−1; δH (300.13 MHz, CDCl3) 1.20 (s, 9H, But); 2.44 (s,
(CO), 170.3 (C-2); HRMS (ESI+): MH+, found: 366.0699.
C17H21BrNO3 requires 366.0683.
ACCEPTED MANUSCRIPT
3H, CH3), 2.83 (dd, 1H, 3J 2.7, |2J| 17.1 Hz, HH-3), 3.09 (dd, 1H,
3
3J 9.2, |2J| 17.1 Hz, HH-3), 4.67 (br d, 1H, J 9.1 Hz, H-4), 7.28
4.4.7. (±)-5-(tert-Butoxycarbonyl)-4-(3-
methoxyphenyl)-6-methyl-3,4-dihydro-2(1H)-
pyridone [(±)-10g]
4
3
4
3
(dd, 1H, J 1.4, J 7.9 Hz, H-6’), 7.38 [dt, 1H, J 1.4 (d), J 8.1
(t), H-4’], 7.52 [dt, 1H, 4J 1.4 (d), 3J 7.9 (t), H-5’], 7.79 (br s, 1H,
NH), 7.89 (dd, 1H, J 1.4, 3J 8.1 Hz, H-3’); δC (75.5 MHz,
4
Reaction time: 10 h. White solid; mp: 145-146 ºC; yield 84%;
νmax(KBr) 3480, 3414, 3219, 1699, 1686, and 1634 cm−1; δH
(300.13 MHz, CDCl3) 1.36 (s, 9H, But); 2.35 (s, 3H, CH3), 2.67
(dd, 1H, 3J 2.6, |2J| 16.5 Hz, HH-3), 2.90 (dd, 1H, 3J 8.1, |2J| 16.5
Hz, HH-3), 3.77 (s, 3H, O-CH3), 4.16 (br d, 1H, 3J 8.1 Hz, H-4),
CDCl3) 18.9 (CH3), 28.1 (3 × CH3), 34.6 (C-4), 37.4 (C-3), 80.9
(C), 107.7 (C-5), 125.0 (CH), 128.0 (CH), 128.4 (CH), 133.5
(CH), 137.8 (C), 146.8 (C), 149.1 (C), 165.5 (CO), 170.4 (C-2);
HRMS (ESI+): MH+, found: 333.1445. C17H21N2O5 requires
333.1427.
3
6.81-6.69 (m, 3H, H-2’, H-4’, and H-6’), 7.19 (t, 1H, J 7.9 Hz,
4.4.3. (±)-5-(tert-Butoxycarbonyl)-6-methyl-4-(3-
nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-10c]
H-5’), 7.53 (br s, 1H, NH); δC (75.5 MHz, CDCl3) 19.2 (CH3),
28.3 (3 × CH3), 38.2 (C-3), 38.6 (C-4), 55.3 (O-CH3), 80.6 (C),
109.0 (C-5), 111.9 (CH), 113.0 (CH), 119.2 (CH), 129.8 (CH),
144.3 (C), 144.7 (C), 159.9 (C-3’) 166.3 (CO), 170.8 (C-2);
HRMS (ESI+): MH+, found: 318.1700. C18H24NO4 requires
318.1704.
Reaction time: 9 h. White solid; mp: 173-173.4 ºC; yield 72%;
νmax(KBr) 3554, 3470, 3414, 3227, 1701, 1640, 1618, 1526,
1366, and 1347 cm−1; δH (300.13 MHz, CDCl3) 1.38 (s, 9H, But);
3
2.40 (s, 3H, CH3), 2.69 (dd, 1H, J 1.7, |2J| 16.7 Hz, HH-3), 2.99
(dd, 1H, 3J 8.2, |2J| 16.7 Hz, HH-3), 4.29 (br d, 1H, 3J 8.2 Hz, H-
4), 7.57-7.40 (m, 2H, H-6’ and H-5’), 7.71 (br s, 1H, NH), 8.14-
8.02 (m, 2H, H-2’ and H-4’); δC (75.5 MHz, CDCl3) 19.3 (CH3),
28.3 (3 × CH3), 37.8 (C-3), 38.3 (C-4), 81.2 (C), 107.9 (C-5),
122.1 (CH), 122.2 (CH), 129.9 (CH), 133.0 (CH), 144.8 (C),
145.9 (C), 148.6 (C), 165.8 (CO), 170.2 (C-2); HRMS (ESI+):
MH+, found: 333.1445. C17H21N2O5 requires 333.1435.
4.4.8. (±)-5-(tert-Butoxycarbonyl)-6-methyl-4-(1-
naphthyl)-3,4-dihydro-2(1H)-pyridone [(±)-10h]
Reaction time: 6 h. White solid, mp: 235-236 ºC; yield 92%;
νmax(KBr) 3216, 1694, 1645, 1526, 1366, and 1347 cm−1; δH
(300.13 MHz, CDCl3) 1.15 (s, 9H, But); 2.46 (s, 3H, CH3), 2.76
(dd, 1H, 3J 2.6, |2J| 16.4 Hz, HH-3), 3.05 (dd, 1H, 3J 8.7, |2J| 16.4
3
3
Hz, HH-3), 5.06 (br d, 1H, J 8.7 Hz, H-4), 7.21 (d, 1H, J 6.9
Hz, H-2’), 7.35 (dd, 1H, 3J 7.3, 8.0 Hz, H-3’), 7.60-7.44 (m, 2H,
4.4.4. (±)-5-(tert-Butoxycarbonyl)-6-methyl-4-(4-
nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-10d]
3
H-6’ and H-7’), 7.62 (br s, 1H, NH), 7.73 (d, 1H, J 8.1 Hz, H-
4’), 7.86 (dd, 1H, 3J 7.8, 4J 1.5 Hz, H-5’), 8.06 (d, 1H, 3J 8.4 Hz,
H-8’); δC (75.5 MHz, CDCl3) 19.1 (CH3), 28.1 (3 × CH3), 34.3
(C-4), 37.6 (C-3), 80.5 (C), 108.7 (C-5), 122.8 (CH), 123.2 (CH),
125.6 (2 × CH), 126.3 (CH), 127.7 (CH), 129.3 (CH), 130.8 (C),
134.5 (C), 137.2 (C), 145.6 (C), 166.2 (CO), 170.6 (C-2); HRMS
(ESI+): MH+, found: 338.1751. C21H24NO3 requires 318.1745.
Reaction time: 9 h. White solid; mp 230-232 ºC; yield 73%;
νmax(KBr) 3413, 3223, 1698, 1637, 1521, and 1349 cm−1; δH
(300.13 MHz, CDCl3) 1.36 (s, 9H, But); 2.40 (s, 3H, CH3), 2.65
(dd, 1H, 3J 1.6, |2J| 16.6 Hz, HH-3), 2.98 (dd, 1H, 3J 8.3, |2J| 16.6
3
3
Hz, HH-3), 4.29 (br d, 1H, J 8.2 Hz, H-4), 7.35 (d, 2H, J 8.6
Hz, H-2’ and H-6’), 7.71 (br s, 1H, NH), 8.16 (d, 2H, 3J 8.6 Hz,
H-3’ and H-5’); δC (75.5 MHz, CDCl3) 19.3 (CH3), 28.3 (3 ×
CH3), 37.7 (C-3), 38.6 (C-4), 81.2 (C), 107.8 (C-5), 124.2 (CH),
127.8 (CH), 145.8 (C), 147.1 (C), 150.3 (C), 165.8 (CO), 170.1
(C-2); HRMS (ESI+): MH+, found: 333.1445. C17H21N2O5
requires 333.1449.
4.5. General procedure for tert-butyl group removing of (±)-
10.
A solution of the corresponding tert-butyl ester (±)-10 (1.0
mmol) in dichloromethane (2.0 mL) was cooled at 0 ºC, and
anisole (0.48 mL, 4.4 mmol) and trifluoroacetic acid (1.89 mL)
were added. The solution was stirred at 0 ºC until disappearance
of the starting material (TLC monitoring). Then, organic solvent
and trifluoroacetic acid were eliminated under reduced pressure.
The resulting residue was dissolved in dichloromethane (15 mL)
and extracted with aq saturated NaHCO3 (2 × 10 mL). The
organic phase was discarded. Conc. aq HCl was added to the
aqueous phase until pH was 1-2. The resulting acid aqueous
phase was extracted with dichloromethane (3 x 15 mL). The
organic layers were combined, washed with brine (20 mL), dried
with Na2SO4, and evaporated under reduced pressure to give the
corresponding carboxylic acid (±)-9 as a pure product. Yields
obtained were: 9a (85%), 9b (76%), 9c (81%), 9d (68%), 9e
(72%), 9f (74%), 9g (91%), and 9h (90%).
4.4.5. (±)-5-(tert-Butoxycarbonyl)-4-(2-chloro-5-
nitrophenyl)-6-methyl-3,4-dihydro-2(1H)-pyridone
[(±)-10e]
Reaction time: 5 h. White solid, mp 226-228 ºC; yield 74%;
νmax(KBr) 3551, 3473, 3413, 3224, 1700, 1630, 1523, and 1343
cm−1; δH (300.13 MHz, CDCl3) 1.31 (s, 9H, But); 2.49 (s, 3H,
CH3), 2.66 (br d, 1H, |2J| 16.8 Hz, HH-3), 2.96 (dd, 1H, 3J 8.6, |2J|
3
3
16.8 Hz, HH-3), 4.68 (br d, 1H, J 8.2 Hz, H-4), 7.57 (d, 1H, J
8.7 Hz, H-3’), 7.59 (br s, 1H, NH), 7.89 (d, 1H, 4J 2.6 Hz, H-6’),
8.06 (dd, 1H, 4J 2.6, 3J 8.7 Hz, H-4’); δC (75.5 MHz, CDCl3) 19.1
(CH3), 28.2 (3 × CH3), 35.97 (C-3), 36.02 (C-4), 81.1 (C), 106.6
(C-5), 122.5 (CH), 123.4 (CH), 131.1 (CH), 140.4 (C), 141.4 (C),
147.1 (C), 147.4 (C), 165.4 (CO), 169.6 (C-2); HRMS (ESI+):
MH+, found: 367.1055. C17H20ClN2O5 requires 367.1063.
4.6. General procedure for the synthesis of diesters (±)-5.
4.4.6. (±)-4-(4-Bromophenyl)-5-(tert-
butoxycarbonyl)-6-methyl-3,4-dihydro-2(1H)-
pyridone [(±)-10f]
Carboxylic acid (±)-9 (5.18 mmol) was dissolved in N,N-
dimethylformamide (15.5 mL). Cesium carbonate (4.14 mmol)
was added to the solution and the suspension was stirred during
2-3 min after which chloromethyl 2-methylpropanoate (6.73
mmol) was added. After stirring at room temperature during 18-
24 h, dichloromethane (30 mL) was added to the mixture and the
resulting organic phase was successively washed with water (3 x
20 mL) and brine (3 x 20 mL). Organic solvent was eliminated
and the residue was submitted to flash chromatography
(hexane/ethyl acetate mixtures) to yield the corresponding diester
(±)-5.
Reaction time: 4 h. White solid, mp 186-187 ºC; yield 88%;
ν
max(KBr) 3216, 1687, 1630, 1484, 1380, and 1365 cm−1; δH
(300.13 MHz, CDCl3) 1.36 (s, 9H, But); 2.36 (s, 3H, CH3), 2.62
(dd, 1H, 3J 1.9, |2J| 16.5 Hz, HH-3), 2.91 (dd, 1H, 3J 8.1, |2J| 16.5
3
3
Hz, HH-3), 4.14 (br d, 1H, J 8.1 Hz, H-4), 7.05 (d, 2H, J 8.4
Hz, H-2’ and H-6’), 7.39 (d, 2H, 3J 8.4 Hz, H-3’ and H-5’), 7.64
(br s, 1H, NH); δC (75.5 MHz, CDCl3) 19.3 (CH3), 28.3 (3 ×
CH3), 38.06 (C-3), 38.11 (C-4), 80.9 (C), 108.6 (C-5), 120.8 (C),
128.6 (2 × CH), 131.9 (2 × CH), 141.6 (C), 144.9 (C), 166.1

8
Tetrahedron
ACCEPTED MANUSCRIPT
3
system, |2J| 5.7 Hz, O-CH2-O), 7.34 (d, 2H, J 8.6 Hz, H-2’ and
H-6’), 8.12 (br s, 1H, NH), 8.14 (d, 2H, 3J 8.6 Hz, H-3’ and H-
5’); δC (75.5 MHz, CDCl3) 18.7 (CH3), 19.8 (CH3), 33.8 (CH),
37.5 (C-3), 37.8 (C-4), 79.1 (CH2), 104.9 (C-5), 124.3 (CH),
127.9 (CH), 147.3 (C), 149.46 (C), 149.50 (C), 164.9 (CO),
169.9 (C-2), 175.9 (CO); HRMS (ESI): MH+ found: 377.1343.
C18H21N2O7 requires 377.1335.
4.6.1. (±)-6-Methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] -4-phenyl-
3,4-dihydro-2(1H)-pyridone [(±)-5a]
Reaction time: 22 h. White solid, mp 157-160 ºC; yield 83%;
νmax(KBr) 3210; 1758; 1716; 1690; 1628; 1492 cm−1, δH (300.13
MHz, CDCl3) 1.13-1.02 [6H, two overlapped d centered to 1.09
(3J 7.0 Hz) and 1.06 (3J 7.0 Hz), CH-(CH3)2], 2.52-2.39 [m + s,
4H, CH(CH3)2 and singlet centered to 2.42 corresponding to
CH3], 2.71 (br d, 1H, |2J| 16.6 Hz, HH-3), 2.94 (dd, 1H, 3J 8.2, |2J|
16.6 Hz, HH-3), 4.26 (br d, 1H, 3J 7.9 Hz, H-4), 5.75 (AB
system, |2J| 5.5 Hz, O-CH2-O), 7.30-7.11 (m, 5H, Ph), 7.80 (br s,
1H, NH); δC (75.5 MHz, CDCl3) 18.7 (CH3), 19.5 (CH3), 33.8
(CH), 37.7 (C-4), 38.1 (C-3), 79.0 (CH2), 106.0 (C-5), 126.8
(CH), 127.2 (C-4’), 129.0 (CH), 141.9 (C), 148.7 (C-6), 165.3
(CO), 171.1 (C-2), 175.9 (CO); HRMS (ESI+): MH+, found:
332.1492. C18H22NO5 requires 332.1446.
4.6.5. (±)-4-(2-Chloro-5-nitrophenyl)-6-methyl-5-
[(2-methylpropanoyloxy)methyloxycarbonyl] - 3,4-
dihydro-2(1H)-pyridone [(±)-5e]
Reaction time: 23 h. White solid, 154-155 ºC, yield 81%;
νmax(KBr) 3420; 1759; 1709; 1681; 1636; 1524 cm−1; δH (300.13
MHz, DMSO-d6) 0.97-0.85 [6H, two overlapped d centered to
0.93 (3J 7.0 Hz) and 0.90 (3J 6.9 Hz), CH-(CH3)2], 2.51-2.31 [m
+ s, 5H, CH(CH3)2, HH-3, and singlet centered to 2.44
corresponding to CH3], 3.16 (dd, 1H, J 8.5, |2J| 16.7 Hz, HH-3),
3
4.55 (br d, 1H, J 8.1 Hz, H-4), 5.62 (AB system, |2J| 5.8 Hz, O-
3
4.6.2. (±)-6-Methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] -4-(2-
nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-5b]
CH2-O), 7.72 (d, 1H, 4J 2.7 Hz, H-6’), 7.81 (d, 1H, 3J 8.7 Hz, H-
3’), 8.13 (dd, 1H, 4J 2.7, J 8.7 Hz, H-4’), 10.35 (s, 1H, NH), δC
3
(75.5 MHz, DMSO-d6) 18.2 (CH3), 18.3 (CH3), 32.8 (CH), 34.9
(C-4), 35.6 (C-3), 78.8 (CH2), 101.4 (C-5), 121.5 (CH), 123.6
(CH), 131.6 (CH), 139.4 (C), 140.9 (C), 146.6 (C), 152.7 (C),
164.3 (CO), 168.6 (C-2), 174.7 (CO); HRMS (ESI+): MH+,
found: 411.0954. C18H20ClN2O7 requires 411.0931.
Reaction time: 18 h. White solid, mp 155-156 ºC; yield 78%;
νmax(KBr) 3436; 3312; 1745; 1714; 1703; 1624; 1527 cm−1, δH
(300.13 MHz, DMSO-d6) 1.00-0.88 [6H, two overlapped d
centered to 0.96 (3J 7.0 Hz) and 0.93 (3J 7.0 Hz), CH-(CH3)2],
2.50-2.33 [m + s, 5H, CH(CH3)2, HH-3, and singlet centered to
3
2.39 corresponding to CH3], 3.18 (dd, 1H, J 8.9, |2J| 16.5 Hz,
4.6.6. (±)-4-(4-Bromophenyl)-6-methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] - 3,4-
dihydro-2(1H)-pyridone [(±)-5f]
HH-3), 4.50 (br d, 1H, 3J 8.6 Hz, H-4), 5.57 (AB system, |2J| 5.9
4
3
Hz, O-CH2-O), 7.25 (dd, 1H, J 1.3, J 7.7 Hz, H-6’), 7.50 [dt,
4
1H, 4J 1.3 (d), 3J 8.0 (t), H-4’], 7.64 [dt, 1H, J 1.4 (d), 3J 7.7 (t),
Reaction time: 24 h. White solid, 134-135 ºC; yield 83%;
νmax(KBr) 3418; 1760; 1721; 1695; 1626; 1490 cm−1; δH (300.13
MHz, CDCl3) 1.14-1.00 [6H, two overlapped d centered to 1.09
(3J 7.0 Hz) and 1.07 (3J 7.0 Hz), CH-(CH3)2], 2.55-2.39 [m + s,
4H, CH(CH3)2 and singlet centered to 2.43 corresponding to
CH3], 2.66 (br d, 1H, |2J| 16.6 Hz, HH-3), 2.94 (dd, 1H, 3J 8.1, |2J|
16.6 Hz, HH-3), 4.21 (br d, 1H, 3J 7.8 Hz, H-4), 5.74 (AB
4
H-5’], 7.93 (dd, 1H, J 1.3, 3J 8.0 Hz, H-3’), 10.26 (br s, 1H,
NH); δC (75.5 MHz, DMSO-d6) 18.2 (CH3), 18.4 (CH3), 32.8
(CH), 33.2 (C-4), 37.0 (C-3), 78.8 (CH2), 102.3 (C-5), 124.8
(CH), 127.8 (CH), 128.3 (CH), 133.7 (CH), 136.5 (C), 148.4 (C),
152.3 (C), 164.4 (CO), 168.9 (C-2), 174.6 (CO); MS (EI), m/z
(%) = 259 ([M − OCH2OC(O)iPr]+, 97), 197 (100), 71 (98). Anal.
calc. for C18H20N2O7: C, 57.44; H, 5.36; N, 7.44. Found: C,
57.72; H, 5.27; N, 7.53.
3
system, |2J| 5.6 Hz, O-CH2-O), 7.03 (d, 2H, J 8.3 Hz, H-2’ and
3
H-6’), 7.38 (d, 2H, J 8.5 Hz, H-3’ and H-5’), 7.69 (br s, 1H,
4.6.3. (±)-6-Methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] -4-(3-
nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-5c]
NH); δC (75.5 MHz, CDCl3) 18.7 (CH3), 19.6 (CH3), 33.8 (CH),
37.3 (C-4), 37.9 (C-3), 79.0 (CH2), 105.5 (C-5), 121.1 (C), 128.6
(2 × CH), 132.0 (2 × CH), 140.9 (C), 149.0 (C), 165.1 (CO),
170.8 (C-2), 175.9 (CO); HRMS (ESI): MH+ found: 410.0598.
C18H21BrNO5 requires 410.0606.
Reaction time: 24 h. White solid; m.p. 126-127 ºC; yield 85%;
νmax(KBr) 3229; 1749; 1719; 1631; 1531 cm−1; δH (300.13 MHz,
CDCl3) 1.07 [m, 6H, (CH3)2CH], 2.59-2.39 [m + s, 4H,
CH(CH3)2 and singlet centered to 2.47 corresponding to CH3],
4.6.7. (±)-4-(3-Methoxyphenyl)-6-methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] - 3,4-
dihydro-2(1H)-pyridone [(±)-5g]
2
3
2.71 (br d, 1H, J 16.7 Hz, HH-3), 3.03 (dd, 1H, J 8.3, |2J| 16.6
Hz, HH-3), 4.38 (br d, 1H, 3J 7.9 Hz, H-4), 5.73 (AB system, |2J|
5.7 Hz, O-CH2-O), 7.57-7.47 (m, 2H, H-5’ and H-6’), 8.01 (s,
Reaction time: 18 h. White solid; mp 105-107 ºC; yield 91%;
νmax(KBr) 3412; 1752; 1714; 1694; 1629; 1585 cm−1; δH (300.13
MHz, CDCl3) 1.14-1.03 [6H, two overlapped d centered to 1.09
(3J 7.0 Hz) and 1.07 (3J 7.0 Hz), CH-(CH3)2], 2.42 (s, 3H, CH3),
3
4
1H, H-2’), 8.09 (dd, 1H, J 7.7, J 0.9 Hz, H-4’), 8.16 (br s, 1H,
NH); δC (75.5 MHz, CDCl3) 18.7 (CH3), 19.7 (CH3), 33.8 (CH),
37.6 (C-4), 37.8 (C-3), 79.2 (CH2), 104.9 (C-5), 121.8 (C-4’),
122.5 (C-2’), 130.1 (C-5’), 133.2 (C-6’), 144.1 (C-1’), 148.8 (C-
3’), 149.7 (C-6), 164.9 (CO), 170.0 (C-2), 175.8 (CO); MS (EI),
m/z (%) = 376 (M•+, 11) 259 ([M − OCH2OC(O)iPr]+, 100);
HRMS (EI): M•+ found: 376.1281. C18H20N2O7 requires
376.1271.
3
2.25 [septet, 1H, J 7.0 Hz, CH(CH3)2 ], 2.71 (br d, 1H, |2J| 16.6
Hz, HH-3), 2.93 (dd, 1H, 3J 8.1, |2J| 16.6 Hz, HH-3), 3.77 (s, 3H,
O-CH3), 4.24 (br d, 1H, 3J 8.1 Hz, H-4), 5.75 (AB system, |2J| 5.6
Hz, O-CH2-O), 6.80-6.65 (m, 3H, H-2’, H-4’, and H-6’), 7.18 (t,
1H, 3J 7.9 Hz, H-5’), 7.65 (br s, 1H, NH); δC (75.5 MHz, CDCl3)
18.7 (2 × CH3), 19.5 (CH3), 33.8 (CH), 37.7 (CH), 38.1 (C-3),
55.2 (O-CH3), 79.0 (CH2), 105.8 (C-5), 111.9 (CH), 113.1 (CH),
119.1 (CH), 129.9 (CH), 143.5 (C), 148.9 (C), 159.9 (C-3’) 165.3
(CO), 171.1 (C-2), 175.9 (CO); HRMS (ESI): [M+Na] + found:
384.1418. C19H23NNaO6 requires 384.1426.
4.6.4. (±)-6-Methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] -4-(4-
nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-5d]
Reaction time: 22 h. White solid, m.p. 155-156 ºC; yield 84%;
νmax(KBr) 3318; 1753; 1735; 1692; 1634; 1516 cm−1; δH (300.13
MHz, CDCl3) 1.12-1.00 [6H, two overlapped d centered to 1.08
(3J 7.0 Hz) and 1.05 (3J 7.0 Hz), CH-(CH3)2], 2.54-2.40 [m + s,
4H, CH(CH3)2 and singlet centered to 2.46 corresponding to
CH3], 2.70 (br d, 1H, |2J| 16.8 Hz, HH-3), 3.01 (dd, 1H, 3J 8.4, |2J|
16.8 Hz, HH-3), 4.36 (br d, 1H, 3J 8.3 Hz, H-4), 5.74 (AB
4.6.8. (±)-6-Methyl-5-[(2-
methylpropanoyloxy)methyloxycarbonyl] -4-(1-
naphthyl)-3,4-dihydro-2(1H)-pyridone [(±)-5h]
Reaction time: 24 h. White solid, mp 164-166 ºC; yield 79%;
νmax(KBr) 3413; 1750; 1710; 1703; 1675; 1631 cm−1; δH (300.13
MHz, CDCl3) 0.96-0.81 [6H, two d centered to 0.92 (3J 7.0 Hz)

9
3
and 0.87 (3J 7.0 Hz), CH-(CH3)2], 2.27 [septet, 1H, J 7.0 Hz,
the other hand, the basic aqueous phase containing the produced
ACCEPTED MANUSCRIPT
CH(CH3)2 ], 2.50 (s, 3H, CH3), 2.81 (br d, 1H, |2J| 16.4 Hz, HH-
carboxylic acid (R)-9 was acidified with aq 3 M HCl (pH = 1-2)
and subsequently extracted with ethyl acetate (three times). The
normal work-up of this new organic phase gave the
corresponding acid (R)-9.
3), 3.07 (dd, 1H, 3J 8.6, |2J| 16.4 Hz, HH-3), 5.13 (br d, 1H, 3J 8.6
3
Hz, H-4), 5.63 (s, 2H, O-CH2-O), 7.15 (d, 1H, J 6.6 Hz, H-2’),
7.33 (dd, 1H, 3J 7.2, 8.2 Hz, H-3’), 7.64-7.43 (m, 2H, H-6’ and
3
3
4
H-7’), 7.73 (d, 1H, J 8.2 Hz, H-4’), 7.87 (dd, 1H, J 7.9, J 1.4
Hz, H-5’), 8.04 (d, 1H, J 8.4 Hz, H-8’), 8.14 (br s, 1H, NH); δC
4.10. Enzymatic hydrolysis of (±)-5a (Table 3, entry 1).
3
(75.5 MHz, CDCl3) 18.5 (CH3), 19.6 (CH3), 33.56 (CH), 33.62
(CH), 37.4 (C-3), 79.3 (CH2), 105.5 (C-5), 122.8 (CH), 123.1
(CH), 125.5 (CH), 125.7 (CH), 126.6 (CH), 128.1 (CH), 129.3
(CH), 130.6 (C), 134.7 (C), 135.9 (C), 149.5 (C), 165.3 (CO),
170.5 (C-2), 175.6 (CO); HRMS (ESI): MH+ found: 382.1649.
C22H24NO5 requires 382.1650.
To a solution of (±)-5a (100 mg, 0.30 mmol) in TBME (10
mL), CAL-B (100 mg) was added and the mixture was shaken at
28 ºC and 200 rpm during 9 h. After, the general procedure was
followed. Thus, the remaining substrate (S)-5a was isolated with
56% yield (56 mg) and 32% ee. The product (R)-9a was isolated
with 39% yield (27 mg) and 47% ee.
4.7. (±)-5-(Acetoxymethyloxycarbonyl)-6-methyl-4-(3-
nitrophenyl)-3,4-dihydro-2-(1H)-pyridone [(±)-11]
4.10.1. Enantioenrichment of the enzymatically
produced (S)-5a and (R)-9a.
Substrate (S)-5a (55 mg, 32% ee) was dissolved in diethyl
ether (2.0 mL) at room temperature. After cooling at −15 ºC
during 30 min, the formed solid was filtered. The ee of this solid
(40 mg) was 5% (S). In addition, from the removal of the solvent
of the filtrate, 13 mg of enantioenriched (S)-5a (99% ee) was
recovered as a viscous oil: [α]D20 +64.9 (c 0.65, CHCl3).
It was obtained from (±)-9a (1.1 mmol) and chloromethyl
acetate (1.4 mmol) following the general procedure for the
synthesis of diesters (±)-5. Flash chromatography (hexane/ethyl
acetate 2:1) yielded pure (±)-11 (230 mg, 60%) as a white solid;
mp 131-132 ºC; νmax(KBr) 3229; 1752; 1720; 1628; 1517 cm−1;
δH (300.13 MHz, CDCl3) 2.01 (s, 3H, CH3), 2.48 (s, 3H, CH3),
2
3
2.72 (br d, 1H, J 16.7 Hz, HH-3), 3.02 (dd, 1H, J 8.2, |2J| 16.7
The product (R)-9a (26 mg, 47% ee) was dissolved in
methanol (3.0 mL). After cooling at −15 ºC during 4 h, the
formed solid was filtered. The ee of this solid (8 mg) was 31%
(R). In addition, from the removal of the solvent of the filtrate, 17
mg of slightly enantioenriched (R)-9a (55% ee) was recovered as
white solid, [α]D20 −17.4 (c 0.85, DMSO), ee 55%.
3
Hz, HH-3), 4.37 (br d, 1H, J 7.9 Hz, H-4), 5.73 (s, 2H, O-CH2-
O), 7.57-7.46 (m, 2H, H-5’ and H-6’), 7.86 (br s, 1H, NH), 8.01
3
(s, 1H, H-2’), 8.09 (br d, 1H, J 7.9 Hz, H-4’); δC (75.5 MHz,
CDCl3) 18.5 (CH3), 20.3 (CH3), 36.9 (C-4), 37.7 (C-3), 78.7
(CH2), 102.5 (C-5), 121.3 (C-4’), 121.8 (C-2’), 130.2 (C-5’),
133.4 (C-6’), 144.9 (C-1’), 148.0 (C-3’), 151.8 (C-6), 164.7
(CO), 169.2 (C), 169.3 (C); MS (EI), m/z (%) = 348 (M•+, 14),
259 ([M − OCH2OC(O)CH3]+, 100); HRMS (CI): MH+ found
349.1035. C16H17N2O7 requires 349.1036.
4.11. Enzymatic hydrolysis of (±)-5c (Table 3, entry 2)
To a solution of (±)-5c (1.00 g, 2.66 mmol) in TBME (100
mL), CAL-B (1.00 g) was added and the mixture was shaken at
28 ºC and 200 rpm during 9 h. After following the general
procedure, the remaining substrate (S)-5c was isolated with 47%
yield (470 mg) and 73% ee. The product (R)-9c was isolated with
48% yield (352 mg) and 71% ee.
4.8. (±)-5-[(2,2-Dimethylpropanoyloxy)methyloxycarbonyl]-
6-methyl-4-(3-nitrophenyl)-3,4-dihydro-2(1H)-pyridone [(±)-
12]
It was prepared as described for (±)-5c from (±)-9a (0.60
mmol) and chloromethyl 2,2-dimethylpropanoate (0.78 mmol).
Flash chromatography (hexane/ethyl acetate 2:1) yielded pure
(±)-12 (103 mg, 44%) as a white solid; mp 168-170 ºC;
νmax(KBr) 3211; 1749; 1725; 1631; 1510 cm−1; δH (300.13 MHz,
DMSO-d6) 0.97 (s, 9H, But), 2.38 (s, 3H, CH3), 2.55-2.44 (m,
4.11.1. Enantioenrichment of the enzymatically
produced (S)-5c and (R)-9c.
A solution of (S)-5c (469 mg, 73% ee) in diethyl ether (13
mL) was maintained at room temperature during 36 h. The
crystallized solid was filtered (90 mg, 2% ee). Removal of the
solvent of the filtrate gave 319 mg of compound (S)-5c with 95%
ee; [α]D20 = +101.5 (c 1.0, CHCl3).
3
1H, HH-3), 3.05 (dd, 1H, J 8.0, |2J| 16.6 Hz, HH-3), 4.28 (br d,
3
1H, J 7.7 Hz, H-4), 5.67 (AB system, |2J| 5.8 Hz, O-CH2-O),
7.65-7.55 (m, 2H, H-5’ and H-6’), 8.01 (br s, 1H, H-2’), 8.08 (m,
1H, H-4’), 10.20 (br s, 1H, NH),); δC (75.5 MHz, CDCl3) 18.4
(CH3), 26.3 (3 × CH3), 36.9 (C-4), 37.8 (C-3), 38.0 (C), 78.9
(CH2), 102.4 (C-5), 121.2 (C-4’), 121.8 (C-2’), 130.2 (C-5’),
133.4 (C-6’), 144.9 (C-1’), 147.9 (C-3’), 151.7 (C-6), 164.7
(CO), 169.2 (C-2), 176.2 (CO); MS (EI), m/z (%) = 390 (M•+, 8),
259 ([M − OCH2OC(O)tBu]+, 100), 57 (tBu+, 54); HRMS (CI):
MH+ found 391.1517. C19H23N2O7 requires 391.1505.
A partially enantioenriched sample of product (R)-9c (408 mg,
1.48 mmol, 67% ee) was converted into (R)-5c (484 mg, 87%
yield) as described for the racemic mixture. After incubation of a
solution of (R)-5c (482 mg, eeo = 67%) in TBME (48 mL) with
CAL-B (482 mg) during 3 h (Table 3, entry 8), the acid (R)-9c
20
(184 mg, 52% yield) was isolated with 95% ee, [α]D = −142.6
(c 0.50, MeOH).
4.12. Enzymatic hydrolysis of (±)-5f (Table 3, entries 3 and 6)
4.9. General procedure for the enzymatic hydrolysis
reactions.
To a solution of (±)-5f (200 mg, 0.488 mmol) in TBME (20
mL), CAL-B (200 mg) was added and the mixture was shaken at
28 ºC and 200 rpm during 11 h (Table 3, entry 3). After
following the general procedure, the remaining substrate (S)-5f
was isolated with 46% yield (92 mg) and 71% ee. The product
(R)-9f was isolated with 46% yield (70 mg) and 64% ee.
Compound (±)-5 was dissolved in tert-butyl methyl ether
(TBME) previously saturated with water. CAL-B was added to
this solution, and the mixture was shaken at 28 ºC and 200 rpm.
After the time collected in Table 3, the enzyme was filtered and
thoroughly washed with methanol. Solvents were eliminated
under reduced pressure and the residue was dissolved with ethyl
acetate. The organic solution was extracted three times with aq
saturated NaHCO3. The organic phase was successively washed
with water and brine, dried with Na2SO4 and concentrated in
vacuo to give the optically active remaining substrate (S)-5. On
When the enzymatic hydrolysis of (±)-5f (100 mg) was
maintained during 18 h (Table 3, entry 6), the remaining
substrate (S)-5f (31 mg, 31% yield) was isolated with 93% ee;
[α]D20 = +73.6 (c 1.0, CHCl3).

10
Tetrahedron
ACCEPTED MANUSCRIPT
J. Chem. Soc. Perkin Trans. 1, 1996, 947-951. (b) Ochoa, E.;
Suárez, M.; Verdecia, Y.; Pita, B.; Martín, N.; Quinteiro, M.;
4.12.1. Enantioenrichment of the enzymatically
produced (R)-9f.
Seoane, C.; Soto, J. L.; Duque, J.; Pomés R. Tetrahedron 1998, 54,
12409-12420. (c) Nuñez-Figueredo, Y.; Ramírez-Sánchez, J.;
Delgado-Hernández, R.; Porto-Verdecia, M.; Ochoa-Rodríguez, E.;
Verdecia-Reyes, Y.; Marin-Prida, J.; González-Durruthy, M.;
Uyemura, S. A.; Rodrigues, F. P.; Curti, C.; Souza, D. O.; Pardo-
Andreu, G. L. Eur. J. Pharmacol. 2014, 726, 57-65.
A partially enantioenriched sample of acid (R)-9f (99 mg, 0.32
mmol, 62% ee) was converted into (R)-5f (106 mg, 81% yield) as
described for the racemic mixture. After incubation of a solution
of (R)-5f (105 mg, eeo = 62%) in TBME (10 mL) with CAL-B
(105 mg) during 2.5 h (Table 3, entry 9), the acid (R)-9f (33 mg,
3. (a) Danieli, B.; Lesma, G.; Martinelli, M.; Passarella, D.; Silvani,
A. J. Org. Chem. 1997, 62, 6519-6523. (b) Nantermet, P. G.;
Barrow, J. C.; Selnick, H. G.; Homnick, C. F.; Freidinger, R. M.;
Chang, R. S. L.; O’Malley, S. S.; Reiss, D. R.; Broten, T. P.;
Ransom, R. W.; Pettibone, D. J.; Olah, T.; Forray, C. Bioorg. Med.
Chem. Lett. 2000, 10, 1625-1628. (c) Goodman, K. B.; Cui, H.;
Dowdell, S. E.; Gaitanopoulos, D. E.; Ivy, R. L.; Sehon, C. A.;
Stavenger, R. A.; Wang, G. Z.; Viet, A. Q.; Xu, W.; Ye, G.; Semus,
S. F.; Evans, C.; Fries, H. E.; Jolivette, L. J.; Kirkpatrick, R. B.;
Dul, E.; Khandekar, S. S.; Yi, T.; Jung, D. K.; Wright, L. L.; Smith,
G. K.; Behm, D. J.; Bentley, R.; Doe, C. P.; Hu, E.; Lee. D. J. Med.
Chem. 2007, 50, 6-9. (d) Huang, X.; Broadbent, S.; Dvorak, C.;
Zhao, S.-H. Org. Process Res. Dev. 2010, 14, 612-616.
4. Huang, X.; Zhu, J.; Broadbent, S. Tetrahedron Lett., 2010, 51,
1554-1557.
20
41% yield) was isolated with 93% ee, [α]D = −82.3 (c 0.40,
MeOH).
4.13. Enzymatic hydrolysis of (±)-5g (Table 3, entries 4 and
7)
To a solution of (±)-5g (100 mg, mmol) in TBME (10 mL),
CAL-B (100 mg) was added and the mixture was shaken at 28 ºC
and 200 rpm during 7 h (Table 3, entry 4). After following the
general procedure, the remaining substrate (S)-5g was isolated
with 48% yield (48 mg) and 69% ee. The product (R)-9c was
isolated with 48% yield (35 mg) and 66% ee.
When the enzymatic hydrolysis of (±)-5g (100 mg) was
allowed to react during 12 h (Table 3, entry 7), the substrate (S)-
5g (30 mg, 30% yield) was isolated with 95% ee; [α]D = +72.6
5. Wanner, B.; Mahatthananchai, J.; Bode, J. W. Org. Lett., 2011, 13,
5378–5381.
20
6. Enzymes Catalysis in Organic Synthesis, 3rd Ed. Drauz, K.;
Gröger, H.; May, O. (Eds.); Wiley-VCH, Weinheim, 2012.
7. (a) Morales, A.; Ochoa, E.; Suárez, M.; Verdecia, Y.; González, L.;
Martín, N.; Quinteiro, M.; Seoane, C.; Soto, J. L. J. Heterocyclic
Chem. 1996, 33, 103-107. (b) Rodríguez, H.; Suárez, M.; Pérez, R.;
Petit, A.; Loupy, A. Tetrahedron Lett. 2003, 44, 3709-3712. (c) Fu,
G.-Y.; Zhang, X.-L.; Sheng, S.-R.; Wei, M.-H.; Liu, X.-L. Synth.
Commun. 2008, 38, 1249-1258.
8. (a) Holdgrün, X. K.; Sih, C. J. Tetrahedron Lett. 1991, 32, 3465-
3468. (b) Ebiike, H.; Terao, Y.; Achiwa, K. Tetrahedron Lett. 1991,
32, 5805-5808. (c) Sobolev, A.; Franssen, M. C. R.; Vigante, B.;
Cekavicus, B.; Zhalubovskis, R.; Kooijman, H.; Spek, A. L.;
Duburs, G.; Groot, A. J. Org. Chem. 2002, 67, 401-410 and
references therein cited. (d) Sobolev, A.; Franssen, M. C. R.;
Poikans, J.; Duburs, G.; de Groot, A. Tetrahedron: Asymmetry
2002, 13, 2389-2397.
(c 0.50, CHCl3).
4.13.1. Enantioenrichment of the enzymatically
produced (R)-9g.
A partially enantioenriched sample of acid (R)-9g (120 mg,
0.460 mmol, 57% ee) was converted into (R)-5g (148 mg, 89%
yield) as described for the racemic mixture. After incubation of a
solution of (R)-5g (148 mg, eeo = 57%) in TBME (15 mL) with
CAL-B (148 mg) during 2 h (Table 3, entry 10), the acid (R)-9g
20
(52 mg, 49% yield) was isolated with 91% ee, [α]D = −64.2 (c
0.90, DMSO).
4.14. Determination of the enantiomeric excesses.
9. Rodríguez, H.; Reyes, O.; Suárez, M.; Garay, H. E.; Pérez, R.;
Cruz, L. J.; Verdecia, Y.; Martín, N.; Seoane, C. Tetrahedron Lett.
2002, 43, 439-441.
10. Yamamoto, T.; Niwa, S.; Ohno, S.; Onishi, T.; Matsueda, H.;
Koganei, H.; Uneyama, H.; Fujita, S.-i.; Takeda, T.; Kito, M.; Ono,
Y.; Saitou, Y.; Takahara, A.; Iwata, S.; Shoji, M. Bioorg. Med.
Chem. Lett. 2006, 16, 798-802.
11. Lipson, V. V.; Gorobets, N. Y. Mol. Divers. 2009, 13, 399-419 and
references cited therein.
12. Bryan, D. B.; Hall, R. F.; Holden, K. G.; Huffman, W. F.; Gleason,
J. G. J. Am. Chem. Soc. 1977, 99, 2353-2355.
Enantioselective HPLC was used for the determination of the
enantiomeric excesses of compounds 5 and 9. Previously to the
measurement, carboxylic acids 9 were transformed into the
corresponding methyl ester 3 by treatment with diazomethane.
HPLC-conditions and chromatograms are included in the
Supplementary Material.
4.15. Assignment of the configuration: synthesis of (R)-3f.
Optically active (R)-9f (ee = 93%; 29 mg, 0.094 mmol) was
dissolved in DMF (0.30 mL) and cesium carbonate (24 mg, 0.15
mmol) was added. After ten minutes, methyl iodide (0.023 mL,
0.37 mmol) was added and the solution was allowed to react
during 4h at RT. Then, dichloromethane (10 mL) was added. The
organic phase was successively washed with water (5 mL) and
brine (5 mL) to yield the methyl ester (R)-3f, which was purified
by flash chromatography using hexane/ethyl acetate 5:2 (23 mg,
76%); [α]D20 = −100.6 (c 1.05, CHCl3).
13. (a) Chen, C. S.; Fujimoto, Y.; Girdaukas, G.; Sih, C. J. J. Am.
Chem. Soc. 1982, 104, 7294–7299. (b) Program “Selectivity” by
Faber, K.; Hoenig, H. http://www.cis.TUGraz.at/orgc/
14. Recrystallization of esters (R)-5c,f,g also allows their
enantioenrichment in some extension, but either the overall yields
and enatiomeric excesses attained for the acids were lower than
those achieved following the three steps KR-derivatization-KR.
15. For a recent example about the influence of the remote stereogenic
centre on the enantioselectivity of the lipase-catalyzed reactions,
see: Schönstein, L.; Forró, E.; Fülop, F. Tetrahedron: Asymmetry
2013, 24, 202-206.
16. Suárez, M.; de Armas, M.; Ramírez, O.; Alvarez, A.; Martínez-
Alvarez, R.; Molero, D.; Seoane, C.; Liz, R.; Novoa de Armas, H.;
Blaton, N. M.; Peeters, O. M.; Martín, N. New J. Chem. 2005, 29,
1567-1576.
Acknowledgments
17. Rodríguez, H.; Martín, O.; Ochoa, E.; Suárez, M.; Reyes, O.;
Garay, H.; Albericio, F.; Martín, N. QSAR Comb. Sci. 2006, 25,
921-927.
Financial support from the Spanish MICINN (CTQ2011-
24237) is gratefully acknowledged. S. Y. Torres thanks to the
University of Oviedo for a grant.
Supplementary Material
References and notes
1. Yang, M.-H.; Chen, Y.-Y.; Huang, L. Phytochemistry 1988, 27,
445-450.
2. (a) Verdecia, Y.; Suárez, M.; Morales, A.; Rodríguez, E.; Ochoa,
E.; González, L.; Martín, N.; Quinteiro, M.; Seoane, C.; Soto, J. L.
Copy of the HPLC chromatograms of the racemic and
optically active compounds.

11
ACCEPTED MANUSCRIPT

Supplementary Material
ACCEPTED MANUSCRIPT
Chemoenzymatic preparation of optically active 4-aryl-5-carboxy-6-methyl-3,4-
dihydro-2(1H)-pyridone derivatives
Susana Y. Torres,a,b Estael Ochoa,b Yamila Verdecia,b and Francisca Rebolledoa,
a Departamento de Química Orgánica e Inorgánica, Universidad de Oviedo, 33006 Oviedo, Spain.
b Laboratorio de Síntesis Orgánica, Facultad de Química, Universidad de La Habana, 10400 La Habana, Cuba.
Table of Contents:
I. General procedure for the derivatization of the carboxylic acids with diazomethane (S1)
Scheme I (S1)
II. Enantioselective HPLC conditions and chromatograms:
1. Enzymatic hydrolysis of (±)-5a and recrystallization of the resulting (S)-5a and (R)-9a. (S2)
2. Enzymatic hydrolysis reactions of (
3. Enzymatic hydrolysis reactions of (
4. Enzymatic hydrolysis reactions of (
±
±
±
)-5c and (R)-5c (eeo = 67%) (S3 and S4)
)-5f and (R)-5f (eeo = 62%) (S5)
)-5g and (R)-5g (eeo = 57%). (S6)
I. General procedure for the derivatization of the carboxylic acids with diazomethane.
An excess of a solution of diazomethane in a mixture of diethyl ether/methanol (40 mM) was added to a
sample of 1.0 mg of the corresponding carboxylic acid. After 2-3 min. solvents were evaporated and the
resulting methyl ester was analysed by enantioselective HPLC. The optically active carboxylic acids
(isolated from the enzymatic reactions) and the methyl esters derivatives are collected in Scheme I.
Scheme I. Methyl esters derivatives
Corresponding author. Tel.: +34 985 102 982; fax: +34 985 103 446; e-mail: frv@uniovi.es.
S1

II. Enantioselective HPLC conditions and chromatograms:
ACCEPTED MANUSCRIPT
1. Enzymatic hydrolysis of (±)-5a and recrystallization of the resulting (S)-5a and (R)-9a.
HPLC conditions for remaining substrate 5a: Chiralpak IA; hexane/propan-2-ol 95:5, 0.8 mL/min, 254
nm, 30 °C; tR = 22.0 (R) and 29.3 (S) min; RS = 5.0.
(±)-5a
(S)-5a with ee = 32% (Table 3, entry 1)
(S)-5a with ee = 99% (isolated from the filtrate
of the crystallization step
Product 9a was transformed into the methyl ester 3a (Scheme 1)
HPLC conditions for 3a: Chiralpak IA; hexane/propan-2-ol 95:5, 0.8 mL/min, 215 nm, 30 °C; tR = 18.3
(R) and 19.7 (S) min; RS = 1.4.
(±)-3a
(R)-3a with ee = 47% (Table 3, entry 1)
(R)-3a with ee = 55% (isolated from the filtrate
of the crystallization step)
S2

2. Enzymatic hydrolysis reactions of (±)-5c and (R)-5c (ee = 67%).
ACCEPTED MANoUSCRIPT
HPLC conditions for remaining substrate 5c: Chiralcel OD; hexane/propan-2-ol 70:30, 0.8 mL/min, 254
nm, 20 °C; tR = 11.2 (S) and 18.3 (R) min; RS = 6.4.
(±)-5c
(S)-5c with ee = 73% (Table 3, entry 2)
(S)-5c with ee = 95% (Table 3, entry 5)
Product 9c was transformed into the methyl ester 3c (Scheme 1)
HPLC conditions for 3c: Chiralcel OD; hexane/propan-2-ol 70:30, 0.8 mL/min, 254 nm, 20 °C; tR = 9.4
(S) and 19.2 (R) min; RS = 9.5.
(R)-3c with ee = 71% (Table 3, entry 2)
(±)-3c
S3

ACCEPTED MANUSCRIPT
(R)-3c with ee = 95% (Table 3, entry 8)
S4

3. Enzymatic hydrolysis reactions of (±)-5f and (R)-5f (eeo = 62%).
ACCEPTED MANUSCRIPT
HPLC conditions for remaining substrate 5f: Chiralpak IC; hexane/propan-2-ol 90:10, 0.8 mL/min, 215
nm, 30 °C; tR = 19.4 (S) and 23.7 (R) min; RS = 4.0.
(±)-5f
(S)-5f with ee = 71% (Table 3, entry 3)
(S)-5f with ee = 93% (Table 3, entry 6)
Product 9f was transformed into the methyl ester 3f (Scheme 1)
HPLC conditions for 3f: Chiralpak IC; hexane/propan-2-ol 90:10, 0.8 mL/min, 215 nm, 30 °C; tR = 17.9
(S) and 21.1 (R) min; RS = 3.6.
(±)-3f
(R)-3f with ee = 64% (Table 3, entry 3)
(R)-3f with ee = 93% (Table 3, entry 9)
S5

4. Enzymatic hydrolysis reactions of (±)-5g and (R)-5g (eeo = 57%).
ACCEPTED MANUSCRIPT
HPLC conditions for remaining substrate 5g: Chiralpak IC; hexane/propan-2-ol 75:25, 0.8 mL/min, 254
nm, 20 °C; tR = 14.9 (S) and 17.3 (R) min; RS = 2.6.
(S)-5g with ee = 69% (Table 3, entry 4)
(±)-5g
(S)-5g with ee = 95% (Table 3, entry 7)
Product 9g was transformed into the methyl ester 3g (Scheme 1)
HPLC conditions for 3g: Chiralpak IC; hexane/propan-2-ol 75:25, 0.8 mL/min, 215 nm, 20 °C; tR = 15.1
(S) and 18.4 (R) min; RS = 3.9.
(±)-3g
(R)-3g with ee = 66% (Table 3, entry 4)
(R)-3g with ee = 91% (Table 3, entry 10)
S6