NMR analysis of a series of imidazobenzoxazines

Article abstract of DOI:10.1002/mrc.2603

The complete ¹H and ¹³C NMR assignment of a series of imidazobenzoxazines by a combination of one- and two-dimensional experiments (COSY, HSQC and HMBC) is studied. Moreover, 2D NOESY and 1D selective NOESY are reported. This procedure allows the identification of the regioisomers obtained. Copyright

Full text of DOI:10.1002/mrc.2603

MRC Letters
Received: 11 February 2010
Revised: 2 March 2010
Accepted: 4 March 2010
Published online in Wiley Interscience: 13 April 2010
(www.interscience.com) DOI 10.1002/mrc.2603
NMR analysis of a series
of imidazobenzoxazines
Mirko Rivara,a Marco Fantini,a∗ Domenico Acquottib
and Valentina Zuliania
The complete 1H and 13C NMR assignment of a series of imidazobenzoxazines by a combination of one- and two-dimensional
experiments (COSY, HSQC and HMBC) is studied. Moreover, 2D NOESY and 1D selective NOESY are reported. This procedure
c
allows the identification of the regioisomers obtained. Copyright ꢀ 2010 John Wiley & Sons, Ltd.
Keywords: NMR; 2D NMR; HSQC; HMBC; COSY; NOESY; imidazobenzoxazines
Introduction
the already assigned C4. It is interesting to observe that for
this rigid scaffold, a number of 4J correlation peaks, such as
H11-C8 and H13-C4, are present in the spectrum. Moreover, a
similar 1H and 13C chemical shift behavior was observed in a
series of benzoxazine derivatives previously published.[5] Once
H10 is assigned and considering the correlations obtained by 2D
COSY, H11, H12 and H13 can be endowed easily; the respective
carbons were allotted from HSQC. In the HMBC spectrum, H10
with δ = 8.01 ppm showed a 2J coupling constant with the
quaternary carbon signal at δ = 117.02 ppm, identified as C9.
The protons of the second aromatic ring (H15–H17) are easily
assigned on the basis of multiplicity and 2D COSY relations; the
only doublet not assigned in 1D 1H NMR spectrum (δ = 7.79 ppm)
belongs to H15 and, consequently, H16 (δ = 7.36 ppm) and
H17 (δ = 7.23 ppm) were identified. The corresponding carbons
are directly assigned through a HSQC experiment. The same
methodology described was applied to the imidazobenzoxazine
2. The correct conformation of each regioisomer was obtained
with 2D NOESY and 1D selective NOESY. The 1D selective NOESY
spectrum of structure 1 shows two spatial contacts between H7
and both H2 and H15, whereas for the structure 2 only one
cross-peak, between H2 and H15, is detected. Figure 2 shows the
differences between the spatial interactions of the two isomers,
clearlyidentifiedbothinthe2DNOESYaswellasinthe1Dselective
NOESY spectra.
As part of our research on the design and synthesis of new sodium
channel blockers potentially useful as anticonvulsants,[1,2] we have
been involved in the preparation of a series of imidazole-based
privileged structures (Fig. 1), obtained both under conventional
as well as microwave heating conditions in high yields.[3] The
synthetic route employed to obtain the imidazobenzoxazines
1–6 allowed to the formation of two regioisomers, separated and
isolated. These new polycyclic heterocycles have been described
by Mahesh only in 1985[4] and detailed information about 1H and
13C NMR signals of this class of compounds is still lacking. Thus,
the present study deals with the complete 1H and 13C assignments
of these structures, also focusing on the identification of the
regioisomers obtained.
Results and Discussion
The complete proton and carbon chemical shifts’ assignments
were achieved by a combination of one- and two-dimensional
NMR experiments. Considering the structure of 1 (Fig. 2), the 1D
1H NMR spectrum shows two singlets at δ = 5.74 and 7.12 ppm,
integrating for 1 and 2 protons, respectively: these data suggest
that the signal at δ = 5.74 belongs to H2 and that at δ = 7.12 ppm
toH7. Theheteronuclearsinglequantumcoherence(HSQC)shows
that H2 correlates with the carbon at δ = 75.0 ppm and the
proton H7 with the carbon at δ = 111.1 ppm (Table 1). The long-
range heteronuclear multiple bond coherence (HMBC), showing
2J and 3J CH coupling constants, allows to assign a number of
quaternary carbons. In particular, the proton H2 correlates with
the previously assigned C7 and with two quaternary carbons at
δ = 141.1 and 152.1 ppm; the proton H7 shows a long-distance
correlation with C2 and with two quaternary carbons at δ = 141.1
and 143.2 ppm. Their only possible common correlation must
be with C4, assigned to the carbon signal at δ = 141.1 ppm.
Consequently, the remaining two quaternary carbons considered
can be identified as C6 (δ = 143.2 ppm) and C8 (δ = 152.1 ppm)
correlated to H7 and H2, respectively.
All 1H and 13C data regarding the two isomers 2-phenyl-5H-
imidazo[1,2-c][1,3] benzoxazine (1) and 3-phenyl-5H-imidazo[1,2-
c][1,3]benzoxazine (2) are given in Table 1.
For compounds 3, 4, 5 and 6, the protons and carbons were
assigned using the same strategy and the data obtained are
reported in Tables 2 and 3.
∗
`
Correspondence to: Marco Fantini, Dipartimento Farmaceutico, Universita
degli Studi di Parma, Viale G.P. Usberti 27/A, I-43124 Parma, Italy.
E-mail: marco-fantini@libero.it
`
a
Dipartimento Farmaceutico, Universita degli Studi di Parma, V.le G.P. Usberti,
27/A, I-43124 Parma, Italy
The HMBC analysis allows to discriminate H10 and H13:
considering the structure 1, only H10 can correlate (3J) with
b
Centro Interdipartimentale Misure ‘Giuseppe Casnati’, V.le G.P. Usberti, 23/A,
I-43124 Parma, Italy
c
Magn. Reson. Chem. 2010, 48, 500–503
Copyright ꢀ 2010 John Wiley & Sons, Ltd.

NMR analysis of imidazobenzoxazines
NMR measurements
B
OCH3
O
OH
N
1
H
(i)
The H NMR experiments were performed in CDCl3 on a Varian
B
A
+
A
N
Inova 600 spectrometer operating at 599.74 MHz using a Standard
Bore 5 mm inverse triple resonance Z gradient probe. 13C NMR
experiments were performed on the same instrument operating
at 150.82 MHz.
(ii)
H
O
O
1H and 13C NMR, gCOSY, NOESY, gHSQC, gHMBC spectra were
acquired from samples in CDCl3 (0.6 ml) solutions (50 mg ml−1).
1H and 13C NMR spectra were measured at 25 ◦C in standard
5-mm o.d. tubes (Norell, S600). All experiments were performed at
298◦K without spinning. Chemical shifts are reported as δ (ppm);
coupling constants (J) are expressed in Hz. 1H NMR spectra were
referenced to residual CDCl3 at δ = 7.24 ppm; 13C assignments
were referenced to CDCl3 at δ = 77.00 ppm. In 1D 1H NMR data,
16 transients were collected with a 30◦ pulse flip angle and 1.5-s
relaxation delay using 32 K data points. 13C NMR spectra were run
operating typically with a 45◦ pulse flip angle, a relaxation delay
time of 3–5 s and a spectral width of 25 000 Hz with 64 K data
points and zero-filled to a digital resolution of 0.4 Hz.
(iii)
O
N
N
B
A
1-6
A, B = H, CH3, n-CH2CH2CH3, Ph
Figure 1. Synthesis of the imidazobenzoxazines 1–6.
Reagentsandconditions:(i) CH3COONH4,CH3OH,rt,2 h.(ii) 48%
HBr, MW, 150 ◦C, 200 W, 10 min. (iii) CH2I2, Cs2CO3, CH3CN, MW,
80 ◦C, 150 W, 10 min.
The NOESY spectra allow to identify the two isomers of each
couple; they show two spatial contacts H2–H18 and H18–H15 for
3 while for 4 H2 presents a NOE cross-peak with H15 and H18,
indicating that the phenyl ring is linked in position 6 in 3 and in
position 7 in 4. For 5 only the H2–H14 contact is detected while
the spectrum of 6 shows cross peaks of H2 with the chain protons
H15, H16 and H17.
The data for the gCOSY, gHSQC and gHMBC spectra were
collected with 1 K data points for F2 and 128–256 increments for
F1 with pulse sequences that allowed gradient selection; spectral
1
windows were 5500–6000 and 27 000–37 000 Hz in the H and
13C dimensions, respectively. In the homonuclear 1H, 2D COSY and
NOESY experiments, the relaxation delay was set to 1.5–2 s. The
resulting FIDs were zero-filled to a 2 K × 1 K data matrix, apodized
withasinefunctionforCOSY, andashiftedsinefunctionforNOESY
in both the ω 1 and ω 2 dimensions prior to Fourier transformation.
In NOESY spectra mixing time was 0.7 s.
The 1D NOESY was achieved using the standard Varian DPFGSE
NOE pulse sequence using the pulse sequence using a q3 shape
pulse (Gaussian cascade 180◦ inversion); the selective 180◦ pulse
was arrayed between 25.5 and 49.4 ms. The 1D NOESY spectra
were acquired with a mixing time of 700 ms, a relaxation delay of
1 s and acquisition time of 1.7 s.
Experimental
Synthesis
The imidazobenzoxazines reported in this study were prepared in
our laboratories as previously described.[4]
1
13
O
8
2
12
11
1
N3
13
4
O
8
9
7
2
12
11
10
16
15
6
5N
17
N3
4
15
7
14
15
16
9
10
14
16
17
16
5N
15
6
1
2
2
7
2
15
15
Figure 2. 1D selective NOESY spectra of compounds 1 and 2.
c
Magn. Reson. Chem. 2010, 48, 500–503
Copyright ꢀ 2010 John Wiley & Sons, Ltd.
www.interscience.wiley.com/journal/mrc

M. Rivara et al.
Table 1. 1H and 13C chemical shifts (in ppm) and HMBC correlations of 1 and 2a
1
O
1
O
13
13
10
8
8
9
2
12
11
2
12
11
N3
4
N3
4
7
9
7
10
6
5N
5N
6
15
14
15
16
16
17
1
2
Position
1H
13C
HMBC
Position
1H
13C
HMBC
2
5.74 (s)
–
75.0
141.1
143.2
111.1
152.1
117.1
123.7
123.7
130.1
116.9
133.7
125.1
128.6
127.1
4, 7, 8
2
4
5.82 (s)
–
73.9
141.6
128.8
131.0
152.2
117.5
123.6
123.8
130.1
116.9
128.7
127.8
129.1
128.1
4, 7, 8
4
–
–
6
–
–
2, 4, 6
–
6
7.24 (s)
–
2, 4, 7
7
7.12 (s)
–
7
–
–
8
8
–
9
–
–
9
–
–
10
11
12
13
14
15
16
17
8.01 (d)
7.13 (t)
7.27 (t)
7.02 (d)
–
4, 8, 9, 11, 12
8, 9, 10, 12, 13
8, 10, 11, 13
8, 9, 11, 12
–
10
11
12
13
14
15
16
17
7.96 (d)
7.14 (t)
7.28 (t)
7.04 (d)
–
4, 8, 11, 12, 13
8, 9, 10, 12, 13
8, 9, 10, 11, 13
8, 9, 11, 12
–
7.79 (d)
7.36 (t)
7.23 (t)
6, 16, 17
7.33 (d)
7.43 (t)
7.35 (t)
7, 16, 17
14, 15, 17
15, 16
14, 15, 17
15, 16
Multiplicities: s, singlet; d, doublet, t, triplet.
a Recorded in CDCl3.
Table 2. 1H and 13C chemical shifts (in ppm) and HMBC correlations of 3 and 4a
1
O
1
O
13
13
8
9
8
9
2
12
11
2
12
11
N3
4
N3
4
18
7
10
10
7
6
5N
N
6
5
15
14
15
16
18
16
17
3
4
Position
1H
13C
HMBC
Position
1H
13C
HMBC
2
5.69 (s)
–
73.3
139.5
139.2
120.7
151.8
117.2
123.5
123.7
129.8
116.8
134.6
127.3
128.4
126.6
9.4
4, 7, 8
2
4
5.66 (s)
–
74.0
139.8
136.8
126.0
152.3
117.3
123.4
123.6
129.8
116.8
129.0
129.1
128.9
127.9
13.2
4, 7, 8
4
–
–
–
–
–
–
6
–
6
–
–
7
–
7
–
–
–
8
–
8
–
9
–
9
–
–
10
11
12
13
14
15
16
17
18
7.99 (d)
7.11 (t)
7.23 (t)
7.01 (d)
–
4, 8, 9, 11, 12
8, 9, 10, 12, 13
8, 10, 11, 13
8, 9, 11, 12
–
10
11
12
13
14
15
16
17
18
7.95 (d)
7.12 (t)
7.25 (t)
7.01 (d)
–
4, 8, 9, 11, 12
8, 9, 10, 12, 13
8, 10, 11, 13
8, 9, 10, 11, 12
–
7.65 (d)
7.39 (t)
7.25 (t)
2.38 (s)
6, 14, 16, 17
14, 15, 17
15, 16
7.27 (d)
7.45 (t)
7.36 (t)
2.33 (s)
7, 14, 16, 17
14, 15, 17
15, 16
6, 7
6, 7
Multiplicities: s, singlet; d, doublet, t, triplet.
a Recorded in CDCl3.
c
www.interscience.wiley.com/journal/mrc
Copyright ꢀ 2010 John Wiley & Sons, Ltd.
Magn. Reson. Chem. 2010, 48, 500–503

NMR analysis of imidazobenzoxazines
Table 3. 1H and 13C chemical shifts (in ppm) and HMBC correlations of 5 and 6a
1
O
1
13
13
8
8 O
2
12
11
2
12
11
N3
4
N3
4
9
15
7
14
9
17
10
10
7
6
16
5N
5N
6
16
15
14
17
5
6
Position
1H
13C
HMBC
Position
1H
13C
HMBC
2
5.57 (s)
–
73.3
138.7
139.7
119.6
151.6
117.4
123.1
123.5
129.3
116.7
8.0
4, 7, 8, 9
2
4
5.61 (s)
–
73.4
138.7
135.4
124.4
151.6
117.4
123.0
123.5
129.4
116.7
12.6
4, 7, 8, 9
4
–
–
–
–
–
–
6
–
6
–
–
7
–
7
–
–
–
8
–
8
–
9
–
9
–
–
10
11
12
13
14
15
16
17
7.89 (d)
7.08 (t)
7.21 (t)
6.99 (d)
2.17 (s)
2.51 (t)
1.66 (m)
0.93 (t)
4, 8, 9, 11, 12, 13
4, 8, 9. 10, 12, 13
8, 10, 11, 13
8, 9, 11, 12
6, 7, 15
10
11
12
13
14
15
16
17
7.87 (d)
7.08 (t)
7.21 (t)
6.98 (d)
2.22 (s)
2.51 (t)
1.50 (m)
0.92 (t)
4, 8, 9, 11, 12, 13
4, 8, 9, 10, 12, 13 1311313
4, 8, 10, 11, 13
8, 9, 11, 12
6, 7, 15
29.1
6, 7, 16, 17
6, 15, 17
24.7
6, 7, 16, 17
7, 15, 17
23.4
23.4
13.9
15, 16
13.4
15, 16
Multiplicities: s, singlet; d, doublet, t, triplet.
a Recorded in CDCl3.
Heteronuclear spectra were recorded with 1 K × 256 data References
points, zero-filled only in F1 to a 1 K × 512 data matrix, and
[1] M. Rivara, A. R. Baheti, M. Fantini, G. Cocconcelli, C. Ghiron,
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[2] M. Fantini, M. Rivara, V. Zuliani, C. L. Kalmar, F. Vacondio, C. Silva,
A. R. Baheti, N. Singh, E. C. Merrick, R. S. Katari, G. Cocconcelli,
C. Ghiron, M. K. Patel, Bioorg. Med. Chem. 2009, 17, 3642.
[3] M. Fantini, V. Zuliani, M. A. Spotti, M. Rivara, J. Comb. Chem. 2010, 12,
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[4] Y. K. Mahesh, M. Maheswari, R. Sharma, R. Sharma, Can. J. Chem.
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apodized in both dimensions with a shifted sine function. HSQC
experiments were acquired using adiabatic pulses for inversion
◦
of 13C, optimized for J(CH) = 145 Hz. The 90 13C pulse length
1
was 13.8 µs. The gHMQC was acquired with 13C sweep width of
32 000 Hz and 256 t1 increments. Each increment was acquired
with 32 transients. Sinebell weighting was applied to the F2
dimension before zero-filled to 2 K points, and a sinebell was
applied to the F1 and zero-filled to 2 K points before Fourier
transformation. A one-bond coupling constant delay was set using
140 HzandWURSTdecouplingwasappliedduringacquisition.The
gHMBCwasacquiredusing64transientsperincrementwith256F1
increments. The one-bond coupling constant of 140 Hz and long-
range coupling constant varying between 5 and 12 Hz were used
to set the delays in the pulse sequence. A sinebell and Gaussian
weighting functions were applied to 13C and 1H dimensions and
zero-filled to 1 and 4 K, respectively.
Acknowledgements
FinancialsupportfromItalianMIURisgratefullyacknowledged.We
are grateful to the Centro Interdipartimentale Misure ‘Giuseppe
Casnati’ of the University of Parma for providing the NMR
instrumentation.
c
Magn. Reson. Chem. 2010, 48, 500–503
Copyright ꢀ 2010 John Wiley & Sons, Ltd.
www.interscience.wiley.com/journal/mrc