Synthesis and the crystal structure of (E)-2-(7-(3-(Thiophen-2-yl) acrylamido)-2,3-dihydro-5-oxobenzo[e][1,4]oxazepin-1(5H)-yl)ethyl acetate

Article abstract of DOI:10.1007/s10870-009-9588-y

Details of the synthesis and crystal structure determination of (E)-2-(7-(3-(thiophen-2-yl)acrylamido)-2,3-dihydro-5-oxobenzo[e][1,4] oxazepin-1(5H)-yl)ethyl acetate are presented. The compound crystallizes in the triclinic P-1 space group (a = 8.3377(17)

Full text of DOI:10.1007/s10870-009-9588-y

J Chem Crystallogr (2009) 39:902–907
DOI 10.1007/s10870-009-9588-y
ORIGINAL PAPER
Synthesis and the Crystal Structure of (E)-2-(7-(3-(Thiophen-2-yl)
acrylamido)-2,3-dihydro-5-oxobenzo[e][1,4]oxazepin-1(5H)-yl)
ethyl acetate
Yun J. Lee Æ Jason R. King Æ Bala Chandra Chenna Æ Samuel B. Owens Jr. Æ
Jason L. Freeman Æ Gary M. Gray Æ Sadanandan E. Velu
Received: 31 December 2008 / Accepted: 7 June 2009 / Published online: 1 July 2009
Ó Springer Science+Business Media, LLC 2009
Abstract Details of the synthesis and crystal structure
determination of (E)-2-(7-(3-(thiophen-2-yl)acrylamido)-
2,3-dihydro-5-oxobenzo[e][1,4]oxazepin-1(5H)-yl)ethyl
acetate are presented. The compound crystallizes in the
triclinic P-1 space group (a = 8.3377(17), b = 9.792(2),
identified from these studies has an IC50 value of 75 lM
[1] (Fig. 1).
Inhibitor 1 has a three-ring structure consisting of a thi-
ophene ring, phenyl ring and a morpholine ring. Thiophene
and phenyl rings are connected by an acrylamido linkage in
which the double bond has a trans stereochemistry. Middle
phenyl ring bears a carboxylic acid group. The morpholine
ring is connected to the middle phenyl ring at the position
ortho to the carboxylic acid group through the morpholine
N atom. The FlexX docking model of inhibitor 1 revealed
several critical interactions of the inhibitor within the active
site residues. The model indicated that the morpholine ring O
has a H bonding interaction with a backbone NH in SrtAD59
active site along with several other critical interactions. In
order to improve the H bonding interaction of the ring O it is
proposed to synthesize the analog 2 which has a bis(2-
hydroxyethyl)amino group in the place of morpholine ring.
The FlexX docking model of the proposed analog 2 has
revealed possible additional H bonding interactions with NH
of Trp194, CO of the Ala92 backbone and CO of Gly192
backbone in the enzyme active site. We have synthesized
this new analog 2 using a six step synthesis starting from 2-
fluoro-5-nitrobenzoic acid (3). The synthesis involves a 7
membered dihydrooxazepinone as a key intermediate (10)
in the penultimate step. The structure of this intermediate
has been confirmed by a high resolution X-ray crystallo-
graphic structure. This manuscript describes the synthesis
of inhibitor 2 and the X-ray structure determination of the
7-membered ring intermediate dihydrooxazepinone, 10.
˚
c = 12.469(3) A, a = 96.39(3)°, b = 108.50(3)°, c =
3
˚
97.68(3)°, V = 943.9(3) A , Z = 2). Interesting features
of the structure include intermolecular hydrogen bond-
ing between the amide proton on one molecule and the
carbonyl oxygen of the dihydrooxazepinone ring on an
adjacent molecule, the boat conformation of the dihydro-
oxazepinone ring, and p-p stacking between thiophene and
phenyl rings on adjacent molecules with a distance
˚
between centroids of 3.79(2) A.
Keywords E isomer ꢀ Arylacrylamide ꢀ Oxazepinone ꢀ
X-ray ꢀ SrtA inhibitor ꢀ Hydrogen bonding
Introduction
We have identified inhibitors of the bacterial surface
enzyme Sortase A by conducting in silico virtual screening
of commercial compound libraries against the Sortase A
active site using FlexX software package. These small
molecules have inhibited the catalytic activity of the
enzyme with IC50 values ranging from low to high
micromolar values [1]. The most active inhibitor (1)
Y. J. Lee ꢀ J. R. King ꢀ B. C. Chenna ꢀ S. B. Owens Jr. ꢀ
J. L. Freeman ꢀ G. M. Gray ꢀ S. E. Velu (&)
Department of Chemistry, University of Alabama
at Birmingham, 901, 14th Street South, Birmingham,
AL 35294-1240, USA
Experimental Section
Synthesis: 5-[(E)-3-(thiophen-2-yl)acrylamido)-2-(bis
(2-hydroxyethyl)amino]benzoic acid (2) is prepared in six
e-mail: svelu@uab.edu
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J Chem Crystallogr (2009) 39:902–907
903
HO
complete. Solvent was completely removed and the residue
obtained was diluted wit EtOAc (50 mL). The EtOAc
extract was washed with water (3 9 25 mL), brine (25 mL)
and dried over Na2SO4. The drying agent was filtered off,
and the solvent was completely removed in vacuo to obtain
the crude product. This was purified by chromatography
over Si gel (20 9 2 cm) using 10% EtOAc in hexanes as
eluent to afford the pure product 4 (0.53 g, 99%); 1H NMR
(CDCl3) d 3.95 (s, 3H), 7.34(t, 1H, J = 9 Hz), 8.38–8.47
O
OH
N
N
S
S
COOH
COOH
N
N
1
2
O
H
O
H
Fig. 1 Sortase A inhibitors
(m, 1H) and 8.85 (dd, 1H, J1 = 6.3 Hz, J2 = 3.0 Hz); 13
C
steps from commercially available methyl 2-fluoro-5-
nitrobenzoate (3) as outlined in Scheme 1. The experi-
mental details follow.
NMR (CDCl3) d 53.0, 118.3 and 118.6 (C–F coupling),
119.7 and 119.9 (C–F coupling), 128.2 and 128.3, 129.4 and
129.6 (C–F coupling), 143.9, 162.6 and 163.3 (C–F cou-
pling) and 166.9; MS (ES?) m/z 169 (M-OCH3).
The melting point was determined using a Mel-Temp II
1
melting point instrument apparatus and is uncorrected. H
and 13C NMR spectra were recorded on a Bruker 300 MHz
spectrometer using TMS as internal standard. The values of
chemical shifts (d) are given in ppm and coupling constants
(J) in Hz. Reactions were monitored by TLC (Whatmann,
Si gel, UV 254, 25 lM plates). The solvents used for
reactions were purchased as anhydrous in Sure-SealTM
bottles from Aldrich chemical company.
2,3-Dihydro-1-(2-hydroxyethyl)-7-
nitrobenzo[e][1,4]oxazepin-5(1H)-one (6)
To a solution of compound 4 (0.59 g, 2.9 mmol) in anhy-
drous THF (12 mL), bis(2-hydroxyethyl)amine (0.4 g,
3.8 mmol) and triethylamine (0.67 mL, 4.8 mmol) were
added and stirred at room temperature for 17 h. TLC
analysis (25% EtOAc in hexane) revealed that the reaction
was complete. The solvent was removed, and the residue
was diluted with EtOAc (100 mL). The EtOAc extract was
washed with water (3 9 35 mL) and brine (35 mL) and
dried over Na2SO4. The drying agent was filtered off, and
the solvent was removed in vacuo to obtain the crude
product. The crude product was purified by crystallization
from CHCl3/EtOAc/MeOH (70:25:5) to afford the pure
product 6 (0.67 g, 90%); 1H NMR (acetone-d6) d 3.75
Methyl 2-fluoro-5-nitrobenzoate (4)
To a 2-fluoro-5-nitrobenzoic acid (0.5 g, 2.7 mmol) in
anhydrous DMF (8 mL), K2CO3 (0.86 g, 6.23 mmol) was
added and stirred for 20 min at room temperature under N2
atmosphere. CH3I (0.5 mL, 8.0 mmol) was added to the
mixture and stirred for additional 30 min. TLC examination
(50% EtOAc in hexanes) showed that the reaction is
Scheme 1 Synthesis
of compound 2
HO
OH
O
O2N
COOH
F
O2N
COOCH3
F
O2N
O
N
CH3I
5
H
K2CO3, DMF
Et3N, THF
N
4
6
3
OH
O
O
O
9
S
COOH
O2N
H2N
O
H2, Pd/C
EtOAc
EDAC, DMAP
CH2Cl2
Ac2O
N
N
DMAP
8
7
OAc
OAc
HO
OAc
OH
N
N
1N. NaOH
S
S
O
THF:MeOH
COOH
O
N
H
N
H
10
O
2
O
123

904
J Chem Crystallogr (2009) 39:902–907
(t, 2H, J = 5.4 Hz), 3.89 (t, 2H, J = 5.4 Hz), 4.03(t, 2H,
J = 3.9 Hz), 4.60 (t, 2H, J = 3.9 Hz), 7.20 (d, 1H,
J = 9.6 Hz), 8.13 (dd, 1H, J1 = 9.6 Hz, J2 = 3.0 Hz) and
8.61(d, 1H, J = 3.0 Hz); 13C NMR (acetone-d6) d
56.4(2C), 59.4, 66.1, 116.8, 117.5, 128.7, 132.7, 138.10,
152.0 and 170.2; MS (ES?) m/z 253 (M ? H).
reaction. The mixture was diluted with CH2Cl2 (20 mL) and
washed with 1 M NaHCO3 (3 9 20 mL), water
(2 9 20 mL), brine (20 mL) and then dried over Na2SO4.
The drying agent was filtered off, and solvent was com-
pletely removed to yield the crude product. The crude
product was purified by column chromatography over Si gel
(20 9 2 cm) using CHCl3 as the eluent to afford the pure
product 10 (0.162 g, 64%); 1H NMR (DMSO-d6) d 1.96 (s,
3H), 3.35–3.58 (m, 4H), 4.18 (t, 2H, J = 6.0 Hz), 4.36 (t,
2H, J = 4.5 Hz), 6.51(d, 1H, J = 15.3 Hz), 7.03(d, 1H,
J = 9.0 Hz), 7.14 (dd, 1H, J1 = 9.0 Hz, J2 = 4.6 Hz), 7.44
(d, 1H, J = 3.3 Hz), 7.65 (d, 1H, J = 5.1 Hz), 7.71(d, 1H,
J = 15.6 Hz), 7.80 (dd, 1H, J1 = 8.7 Hz, J2 = 2.7 Hz) and
7.86 (d, 1H, J = 2.4 Hz); 13C NMR (DMSO-d6) d 20.6,
50.1, 54.2, 60.6, 65.1, 118.6, 120.6, 122.3, 122.5, 124.7,
128.4(2C), 131.2, 132.2, 132.9, 139.7, 141.8, 162.9, 170.3
and 170.8; MS (ES-) m/z 399 (M - H); Anal Calcd for
C20H20N2O5S: C, 59.99; H, 5.03; N, 7.00. Found: C, 59.33;
H, 4.98; N, 6.87.
2-(2,3-Dihydro-7-nitro-5-oxobenzo[e][1,4]oxazepin-
1(5H)-yl)ethyl Acetate (7)
A solution of compound 6 (0.103 g, 0.41 mmol) and
DMAP (0.011 g, 0.09 mmol) in acetic anhydride (2 mL)
was stirred at room temperature for 12 h. TLC (50%
EtOAc in CHCl3) examination indicated the completion of
the reaction. Acetic anhydride was completely removed
under high vacuum, and the residue was diluted with
EtOAc (25 mL). The EtOAc extract was washed with
water (3 9 15 mL) and brine (15 mL) and then dried over
Na2SO4. The drying agent was filtered off, and the filtrate
was concentrated in vacuo to yield the pure product 7
1
(0.091 g, 75%); H NMR (CDCl3) d 2.05 (s, 3H), 3.72 (t,
5-((E)-3-(Thiophen-2-yl)acrylamido)-2-(bis
(2-hydroxyethyl)amino)benzoic acid (2)
2H, J = 6.0 Hz), 3.82–3.91(m, 2H), 4.35 (t, 2H,
J = 6.0 Hz), 4.49–4.59 (m, 2H), 6.92 (d, 1H, J = 9.6 Hz),
8.19 (dd, 1H, J1 = 9.6 Hz, J2 = 3.0 Hz) and 8.72 (d, 1H,
J = 3.0 Hz); 13C NMR (CDCl3) d 21.0, 52.0, 55.1, 60.3,
65.1, 116.4, 117.3, 129.0, 132.3, 138.9, 150.5, 169.6 and
170.9; MS (ES?) m/z 295 (M ? H).
To a solution of compound 10 (0.087 g, 0.22 mmol) in
MeOH/THF (6 mL, 1:1), 1 N NaOH (0.5 mL) was added,
and the mixture was stirred at room temperature for 4 h.
TLC examination (25% EtOAc/hexanes) showed the
completion of the reaction. Solvents were completely
evaporated in vacuo, and the residue was dissolved in
water (20 mL). The aqueous solution was extracted with
CHCl3 (2 9 10 mL) and acidified with 6 N HCl to pH 2 to
precipitate the crude product. The crude product was fil-
tered off and purified by column chromatography over Si
gel (10 9 1 cm) using 20% MeOH/CHCl3 as the eluent to
furnish the pure product 2 (0.21 g, 95%) as a white solid;
(E)-2-(7-Amino-2,3-dihydro-5-
oxobenzo[e][1,4]oxazepin-1(5H)-yl)ethyl acetate (8)
To a solution of compound 7 (0.165 g, 0.56 mmol) in
EtOAc (40 mL), 10% Pd/C (0.034 g) was added and then
this mixture was stirred at room temperature for 1 h under
H2 gas. TLC examination (25% EtOAc in CHCl3) showed
that the reaction was complete. The catalyst was removed
by filtration through CeliteÒ. The filtrate was concentrated
in vacuo to obtain the product 8 (0.148 g, 100%); 1H NMR
(CDCl3) d 2.05 (s, 3H), 3.33–3.39 (m, 4H), 4.21 (t, 2H,
J = 6.0 Hz), 4.29 (t, 2H, J = 5.6 Hz), 6.77–6.84 (m, 2H)
and 6.93–6.98 (m, 1H); 13C NMR (CDCl3) d 20.7, 50.5,
54.6, 61.1, 65.2, 117.2, 120.0, 120.3, 126.2, 137.3, 141.8,
170.9 and 171.9; MS (ES?) m/z 265 (M ? H).
1
mp: 116 °C; H-NMR (DMSO-d6): d 3.21–3.53 (m, 8H),
4.91(bs, 2H), 6.69 (d, 1H, J = 15.3 Hz), 7.13 (s, 1H), 7.46
(s, 1H), 7.58–7.90 (m, 3H), 8.07 (d, 1H, J = 7.8 Hz), 8.43
(s, 1H) and 10.88 (s, 1H); 13C NMR (DMSO-d6) d 57.1,
58.7, 120.9, 121.0, 121.1, 121.2, 124.6, 124.7, 125.2,
129.0, 129.2, 132.0, 134.1, 140.1, 164.1 and 167.4; MS
(ES-) m/z 375 (M - H).
X-ray Data Collection and Structure Solution
(E)-2-(7-(3-(Thiophen-2-yl)acrylamido)-2,3-dihydro-5-
oxobenzo[e][1,4]oxazepin-1(5H)-yl)ethyl acetate (10)
Compound 10 was crystallized by slow diffusion of hex-
anes into a THF solution of 10 to give yellow plate-like
crystals. A suitable single crystal of compound 10 was
glued on a glass fiber with epoxy and aligned upon an
Enraf Nonius CAD4 single crystal diffractometer under
aerobic conditions. Standard peak search and automatic
indexing routines followed by least squares fits of 25
accurately centered reflections resulted in accurate unit cell
To a solution of compound, 8 (0.166 g, 0.63 mmol) and
(E)-3-(thiophen-2-yl)acrylic acid 9 (0.116 g, 0.75 mmol) in
CH2Cl2 (10 mL), EDAC (0.246 g, 1.28 mmol) and DMAP
(0.01 g, 0.082 mmol) were added and the mixture was
stirred under N2 atmosphere for 12 h. TLC examination
(25% EtOAc in CHCl3) indicated the completion of the
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J Chem Crystallogr (2009) 39:902–907
905
parameters. The space group was assigned on the basis of
systematic absences and intensity statistics. All data col-
lection was carried out using the CAD4-PC software [2],
and details of the data collection are given in Table 1. The
analytical scattering factors of the complex were corrected
for both Df0 and iDf00components of anomalous dispersion.
All data were corrected for the effects of absorption using a
psi scan with four reflections with v C 80° and for Lorentz
and polarization effects.
Table 2 Selected torsion angles (°) for compound 10
C(2)–C(3)–C(4)–S
0.2(6)
-177.0(4)
178.7(4)
C(1)–C(2)–C(3)–C(4)
N(1)–C(1)–C(2)–C(3)
C(2)–C(1)–N(1)–C(8)
C(9)–C(8)–N(1)–C(1)
-174.5(4)
161.4(4)
anisotropic thermal parameters for all these atoms versus
F2 was carried out. All other hydrogen atoms were placed
in calculated positions with the appropriate molecular
All crystallographic calculations were performed with
the Siemens SHELXTL-PC program package [3]. All
heavy atom positions were located using Direct Methods
while the amide hydrogen atom was located in difference
Fourier maps. Full matrix refinement of the positional and
˚
geometry and the d(C–H) = 0.96A. The isotropic thermal
parameter associated with each hydrogen atom was fixed
equal to 1.2 times the Ueq of the atom to which it was
bound.
Selected torsion angles for compound 10 are given in
Table 2. Crystallographic data for the complex has been
deposited with the Cambridge Crystallographic Database
(10: CCDC 712737).
Table 1 Crystal and structure data for 10
Empirical formula
CCDC deposit no.
Formula weight
Temperature
C20H20N2O5S
712737
400.44
293(2) K
˚
Wavelength
0.71073 A
Triclinic
P-1
Results and Discussion
Crystal system
Space group
X-ray Crystal Structure
˚
a = 8.3377(17) A
Unit cell dimensions
˚
b = 9.792(2) A
The crystal structure of compound 10 shows that the
molecule has a rigid backbone with a conformationally
flexible dihydrooxazepinone ring and side chain as seen in
Fig. 2. Table 2 gives selected torsion angles of the back-
bone portion of the molecule. The data show that the thi-
ophene, alkene and amide are essentially coplanar as
indicated by the small deviation ( 5°) in torsion angles
from 180°. This conformation of the backbone suggests
˚
c = 12.469(3) A
a = 96.39(3)°
b = 108.50(3)°
c = 97.68(3)°
3
˚
Volume
943.9(3) A
Z
2
Density (calculated)
Absorption coefficient
1.409 mg/m3
0.207 mm-1
420
F
(000)
Crystal size
0.4 9 0.5 9 0.8 mm3
2.13°–22.47°
-8 B h B 8
-10 B k B 1
-13 B l B 13
2969
h range for data collection
Index ranges
Reflections collected
Independent reflections
Completeness to h = 22.47°
Absorption correction
Maximum transmission
Minimum transmission
Refinement method
2450 [R(int) = 0.0575]
100.0%
Empirical
0.6643
0.4442
Full-matrix least-squares on F2
Data/restraints/parameters
Goodness-of-fit on F2
Final R indices [I [ 2r (I)]
R indices (all data)
2450/0/258
1.113
R1 = 0.0694, wR2 = 0.1720
R1 = 0.0982, wR2 = 0.1935
Fig. 2 ORTEP [6] drawing of the molecular structure of compound
10. Thermal ellipsoids are drawn at 50% and hydrogen labels are
omitted for clarity
-3
˚
0.670 and -0.441 e A
Largest diff. peak and hole
123

906
J Chem Crystallogr (2009) 39:902–907
that conjugation is preserved throughout the backbone. In
contrast, the amide and phenyl ring are twisted by
approximately 18.6(4)° relative to each other, which breaks
the conjugation of the p-system.
dihydrooxazepinone ring adopts a boat conformation as
opposed to the half-chair conformation seen in other dihy-
drooxazepinone rings [4, 5]. This conformation is further
supported by the lack of axial-axial interactions between
H(15) and C(10) or C(11) due their lack of hydrogens.
One final point of interest is the manner in which the
molecules stack in the crystal lattice. Figure 3 shows that a
portion of the molecules stack about a center of symmetry,
which allows for intermolecular hydrogen bonding between
the amide hydrogen and the carbonyl of the dihydrooxaz-
Another important feature of the structure of compound
10 is the conformation of the dihydrooxazepinone ring.
Besides the side chain, the dihydrooxazepinone ring is the
most conformationally flexible portion of the molecule. The
dihydrooxazepinone ring adopts a boat conformation with
C(15) occupying the bow and C(10)–C(11) occupying the
stern. A least squares plane through C(14), O(3), N(2), and
C(16) shows that these four atoms are only slightly out of
plane. This creates a mirror plane of symmetry through the
C(10)–C(11) bond and bisecting C(15), which is indicative
of the boat conformation. Fillers et al. also found that the
˚
epinone ring. The O(2)–HN1 distance is 2.08(4) A and the
˚
N(1)–HN(1) distance is 0.87(4) A. The angle between N(1)–
HN(1)–O(2) is almost linear with a value of 174(4)°. The
orientation of the rings relative to each other in the crystal
lattice shows that p-p stacking is present. The thiophene
Fig. 3 ORTEP [6] Packing Diagram of the molecular structure of compound 10 showing centrosymmetric intermolecular hydrogen bonding.
Thermal ellipsoids are drawn at 50% and hydrogens are omitted for clarity
123

J Chem Crystallogr (2009) 39:902–907
907
References
ring stacks directly over an adjacent phenyl ring with a
˚
distance between centroids of 3.79(2) A. However, the
1. Chenna BC, Shinkre BA, King JR, Lucius AL, Narayana SV, Velu
SE (2008) Bioorg Med Chem Lett 18:380
2. CAD4-PC Ver. 1.2;Enraf Nonius :Delft TN, 1988
3. Sheldrick GM (2001) In SHELXTL NT Ver. 6.12; Bruker AXS
:Madison, WI
rings are not coplanar due to the free rotation about the
N(1)–C(8) bond causing the phenyl group to twist relative
to the rigid backbone.
4. Antel J, Sheldrick GM, Pfeiffer T, Tietze L-F (1988) Acta
Crystallogr C44:1860
5. Fillers JP, Hawkinson SW (1982) Acta Crystallogr B38:3041
6. Burnett MN, Johnson CK (1996) ORTEP-III: Oak Ridge Thermal
Ellipsoid Plot Program for Crystal Structure Illustrations, Oak
Ridge National Laboratory Report ORNL-6895
Supplementary Material
CCDC 712737 contains the supplementary crystallographic
data for this paper. These data can be obtained free of
charge via (please use the link below) by e-mailing
data_request@ccdc.cam.ac.uk, or by contacting The
Cambridge Crystallographic Data Centre, 12 Union Road,
Cambridge CB2 1EZ, UK, Fax: ? 44(0)1223-336033.
123