# Structural and Vibrational Study on Monomer and Dimer Forms and Water Clusters of Acetazolamide

Authors: Aysen E. Ozel; Serda Kecel Gunduz; Sefa Celik; Sevim Akyuz
Journal: Journal of Spectroscopy
 (2013)

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## Abstract

Experimental IR and Raman spectra of solid acetazolamide have been analysed by computing the molecular structures and vibrational spectra of monomer and dimer forms and water clusters of acetazolamide. The possible stable conformers of free acetazolamide molecule in  the ground state were obtained by scanning the potential energy surface through the dihedral angles, D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C). The final geometry parameters for the obtained stable conformers were determined by means of geometry optimization, carried out at DFT/B3LYP/6-31G++(d,p) theory level. Afterwards the possible dimer forms of the molecule and acetazolamide-H2O clusters were formed and their energetically preferred conformations were investigated using the same method and the same level of theory. The effect of BSSE on the structure and energy of acetazolamide dimer has been investigated. The assignment of the vibrational modes was performed based on the potential energy distribution of the vibrational modes, calculated by using GAR2PED program. The experimental vibrational wavenumbers of solid acetazolamide are found to be in better agreement with the calculated wavenumbers of dimer form of acetazolamide than those of its monomeric form. NBO analysis has been performed on both monomer and dimer geometries.


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## Body

## 1. Introduction

Acetazolamide, C4H6N4O3S2 (1,3,4-thiadiazole-2-sulfonamide,5-acetamido), is a sulfonamide derivative and carbonic anhydrase inhibitor used clinically to lower intraocular pressure in glaucomatous patients. It is also used as a diuretic agent for treating acute high-altitude sickness and used for the treatment of epilepsy, and most frequently it is used as a drug for the prophylaxis of high-altitude disorders [1–3]. Recently, it has played a major role as a remedy against respiratory diseases and it has been used to prevent adverse effects of drugs in the treatment of influenza [4] and epilepsy and as diuretic [5]. Traditionally, carbonic anhydrase inhibitors derived from acetazolamide were shown to inhibit the growth of several tumor cell lines in vitro and in vivo [6, 7].Acetazolamide is known to exist in two crystal forms (modifications I and II), which was first reported by Pala in 1956 [8]. Mathew and Palenik solved the crystal structure of triclinic acetazolamide (polymorphic form A, modification II) [9]. Griesser et al. [10] reported the crystal structure of acetazolamide polymorphic form B (modification I) and notified that crystal polymorphism of acetazolamide was based on changes in the spatial molecular arrangement and in the connectivities of hydrogen bonds. The experimental and calculated vibrational spectra [11] and crystal structure [12] of an acetazolamide derivative (5-amino-1,3,4-thiadiazole-2-sulfonamide; Hats) have been reported. The experimental vibrational spectra of the two polymorphic forms (A, B) of acetazolamide were reported by Baraldi et al. [13]. The structure and spectroscopic behaviours of some 1,3,4-thiadiazole ligands containing –SO2NH2 groups and their metal clusters were examined [14–23]. Recently, Brandan et al. [24] calculated the harmonic vibrational wavenumbers for the optimized geometry of acetazolamide, using DFT/B3LYP method with the 6-31G* and 6-311++G** basis sets. Chaturvedi et al. [25] reported calculated vibrational spectra of monomer and dimer forms of acetazolamide, but the calculations were performed on the most stable conformer, obtained by geometry optimization, and on three possible dimers. The basis set superposition error (BSSE) correction was not taken into account on the dimer forms. In the present work, we have extended the investigation of the possible stable conformers of title molecule by means of torsion potential energy surface scan studies through dihedral angles, D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C), and calculated the vibrational modes and wavenumbers of the most stable conformer. Moreover, the structures and vibrational wavenumbers of four energetically favorable dimers and ten H2O clusters of acetazolamide were investigated and the effect of basis set superposition error (BSSE) on the structure and energy of acetazolamide dimers has been evaluated. Acetazolamide is very slightly soluble in water. However, its solubility is significantly important and its slight changes affect the bioavailability of acetazolamide [10]. The aims of this study are to elucidate the conformational preferences of acetazolamide and the effect of hydrogen bonding in the formation of molecular dimers and upon interaction with water molecules.


## 2. Experimental and Computational Methods

### 2.1. Experimental Part

The acetazolamide molecule was purchased from Sigma-Aldrich (≥%99) with CAS number 59-66-5 and used as received. The FT-IR spectra of KBr disc (for solid sample) and aqueous solution (between ZnSe plates) of the molecule were recorded on a Jasco FT/IR-6300 spectrometer in the range 400–4000 cm−1 with a resolution of 2 cm−1 based on averaging 200 samples and 100 background scans. The Raman spectrum of the sample was taken with a Jasco NRS-3100 micro-Raman spectrometer (1800 lines/mm or 1200 lines/mm grating and high sensitivity cooled CCD). Sample was excited with a 531.96 nm diode laser. The Raman system was calibrated with a silicon semiconductor using the Raman peak at 520 cm−1. A 20x microscope objective (Olympus) was used to focus the laser and collect Raman scattering on the sample. Spectral resolution was 1.08 cm−1 and 100 spectra were accumulated.


### 2.2. Computational Part

All of the calculations were carried out by using the Gaussian03© program suite [26]. Due to success in calculating the electronic structure and energy, the calculations were carried out by using the hybrid density functional theory (DFT/B3LYP) method. For the calculations of monomer, dimer forms of acetazolamide and acetazolamide-water clusters 6-31++G(d,p) basis set was used. The X-ray crystallographic results [9] of acetazolamide molecule were used as initial input geometrical data. The minimum energy conformers of acetazolamide were identified by scanning the potential energy surface by varying D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C) dihedral angles. The final geometry parameters for the obtained stable conformers were determined by means of geometry optimization carried out at DFT/B3LYP/6-31G++(d,p) level of theory. Furthermore, acetazolamide dimers were constructed by bringing two identical acetazolamide monomers together in possible configurations, and energetically preferred conformations of dimers were investigated using the same method and the same level of theory. In order to correct overestimation between unscaled and observed wavenumbers dual scaling factors were used. We scaled all the computed harmonic wavenumbers under 1800 cm−1 with the scale factor 0.977 and wavenumbers greater than 1800 cm−1 with the scale factor 0.955 [27]. The potential energy distribution (PED) of the vibrational modes of the molecules was calculated with GAR2PED program [28], and the fundamental vibrational modes were characterized by their PED values.


## 2.1. Experimental Part

The acetazolamide molecule was purchased from Sigma-Aldrich (≥%99) with CAS number 59-66-5 and used as received. The FT-IR spectra of KBr disc (for solid sample) and aqueous solution (between ZnSe plates) of the molecule were recorded on a Jasco FT/IR-6300 spectrometer in the range 400–4000 cm−1 with a resolution of 2 cm−1 based on averaging 200 samples and 100 background scans. The Raman spectrum of the sample was taken with a Jasco NRS-3100 micro-Raman spectrometer (1800 lines/mm or 1200 lines/mm grating and high sensitivity cooled CCD). Sample was excited with a 531.96 nm diode laser. The Raman system was calibrated with a silicon semiconductor using the Raman peak at 520 cm−1. A 20x microscope objective (Olympus) was used to focus the laser and collect Raman scattering on the sample. Spectral resolution was 1.08 cm−1 and 100 spectra were accumulated.


## 2.2. Computational Part

All of the calculations were carried out by using the Gaussian03© program suite [26]. Due to success in calculating the electronic structure and energy, the calculations were carried out by using the hybrid density functional theory (DFT/B3LYP) method. For the calculations of monomer, dimer forms of acetazolamide and acetazolamide-water clusters 6-31++G(d,p) basis set was used. The X-ray crystallographic results [9] of acetazolamide molecule were used as initial input geometrical data. The minimum energy conformers of acetazolamide were identified by scanning the potential energy surface by varying D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C) dihedral angles. The final geometry parameters for the obtained stable conformers were determined by means of geometry optimization carried out at DFT/B3LYP/6-31G++(d,p) level of theory. Furthermore, acetazolamide dimers were constructed by bringing two identical acetazolamide monomers together in possible configurations, and energetically preferred conformations of dimers were investigated using the same method and the same level of theory. In order to correct overestimation between unscaled and observed wavenumbers dual scaling factors were used. We scaled all the computed harmonic wavenumbers under 1800 cm−1 with the scale factor 0.977 and wavenumbers greater than 1800 cm−1 with the scale factor 0.955 [27]. The potential energy distribution (PED) of the vibrational modes of the molecules was calculated with GAR2PED program [28], and the fundamental vibrational modes were characterized by their PED values.


## 3. Result and Discussion

### 3.1. Conformational Analysis and H-Bonding Interactions

The molecular model of acetazolamide with the atom numbering scheme is given in Figure1. Stable low energy conformers of free acetazolamide molecule were obtained firstly by potential energy surface scan studies by iteratively varying D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C) dihedral angles with step angle of 60°. PES scan for acetazolamide calculated with D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C) dihedral angles at the B3LYP/6-31G++(d,p) method is shown in Figure S1 (see Figure S1 in Supplementary Materials available online at http://dx.doi.org/10.1155/2013/538917).  The 45 conformers were identified between 0 and 10 kcal/mol relative energy intervals. Their dihedral angles and relative energies were given in Table S1. The geometry optimization was then performed on the lowest energy conformer. The estimated four most stable conformers of acetazolamide dimers are illustrated in Figure 2. All possible conformations of acetazolamide interacting with one water molecule were investigated, and  acetazolamide interacting with five water molecules was determined as the final possible conformation of acetazolamide-water cluster. Figure 3 demonstrates the geometries of the 10 stable acetazolamide-water clusters obtained using DFT method with B3LYP/6-31++G(d,p) basis set. The energies of the four most stable dimers (I–IV) and energetically preferred H2O-acetazolamide clusters are given in Table 1. It is known that the basis set superposition error (BSSE) effect is rather significant on the structure and energy of dimer forms, so removing this effect is very important. Therefore, optimization of the dimer I was also carried out along with the counterpoise correction scheme proposed by Boys and Bernardi [29]. The BSSE uncorrected and corrected distances between two acetazolamide units {R(5C–22C)(Å)} and binding energy (ΔE) of acetazolamide dimer {ΔE=2*Emonomer-Edimer} are given in Table 1(b). As seen in Table 1(b), the energy of dimer I is found to be 17.39 kcal/mol lower then total energy of the two monomer (2*Emonomer) units, indicating that intermolecular hydrogen bonding plays an important role in stabilization of the molecule.Table 1
(a) The calculated energies of monomeric, dimeric (I–IV), and H2O clusters (I–IX). (b) The counterpoise uncorrected and corrected interatomic distance (Å) and binding energy (kcal/mol) of dimer I.
(a)
Calculated energy (kcal/mol)
Energy differences (kcal/mol)
Monomer
−876645.7
—
Dimer I
−1753308.7
0
Dimer II
−1753299.4
9.3
Dimer III
−1753297.9
10.8
Dimer IV
−1753301.5
7.2
Cluster I
−924613.6
6.1
Cluster II
−924617.1
2.6
Cluster III
−924613.6
6.0
Cluster IV
−924614.6
5.1
Cluster V
−924619.1
0.6
Cluster VI
−924619.7
0
Cluster VII
−924619.1
0.6
Cluster VIII
−924612.9
6.7
Cluster IX
−924614.6
5.1
(b)
BSSE uncorrected
BSSE corrected
R(5C–22C) (Å)
ΔE (kcal/mol)
R(5C–22C) (Å)
Δ(E + BSSE) (kcal/mol)
Dimer I
4.23
17.4
4.25
16.1Figure 1
The most stable geometry, atom numbering, and searched dihedral angles of acetazolamide.Figure 2
The energy minimized structures of four low energy conformers (I–IV) of the dimer forms of the acetazolamide, (I–IV), respectively.Figure 3
The energy minimized structures of ten low energy conformers (I–X) of H2O clusters of the acetazolamide (I–X), respectively.The predicted geometrical parameters such as bond lengths and bond angles of the stable conformation of acetazolamide monomer, dimer (dimer I), and water cluster (VI and X) of acetazolamide, calculated at B3LYP method with 6–31G++(d,p), are presented in Table S2 in accordance with the atom numbering scheme as given in Figure1. By comparing theoretical and experimental bond lengths [9], it is easily seen that the computed bond lengths at B3LYP/6-31++G(d,p) method were slightly longer since the theoretical calculations were performed on isolated molecule in the gaseous state, whereas the experimental results are for the solid phase of the molecule [9, 10, 12]. Main differences between crystal structure [9] and monomer acetazolamide occur in H containing bond lengths and angles, and this is probably due to insufficient predictions for light atoms in the crystal structure study done in 1974 [9]. The theoretical results are mainly consistent with reports of other studies [11, 24]. Comparison of the geometry parameters of monomer form with those of dimer and water cluster of acetazolamide clearly shows the effects of intermolecular hydrogen bonding.The intra- and intermolecular hydrogen bonds of the dimer forms (I–IV) of the four low energy conformations together with those of ten energetically preferred water clusters are tabulated in Table2. The lowest energy conformer of dimer form (dimer I) makes stronger interhydrogen bonding interactions.Table 2
The intra- and interhydrogen bonds of the dimer conformers (I–IV) and H2O clusters (I–X) of acetazolamide.
(a)
Dimer I
Intramolecular H-bonds
Dimer IIIntramolecular H-bonds
Dimer IIIIntramolecular H-bonds
Dimer IVIntramolecular H-bonds
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
3N–10H
3.377
3N–10H
3.342
3N–10H
3.148
4N–13H
2.468
3N–11H
2.975
3N–11H
2.962
3N–11H
2.816
20H–25N
2.468
8O–10H
2.698
8O–10H
2.717
8O–10H
2.700
7O–11H
2.696
7O–11H
2.694
7O–11H
2.715
4N–13H
2.512
4N–13H
2.467
4N–13H
2.488
29N–37H
3.377
29N–37H
3.196
29N–37H
3.141
29N–38H
2.975
29N–38H
2.766
29N–38H
2.857
35O–37H
2.968
35O–37H
2.689
35O–37H
2.697
34O–38H
2.696
34O–38H
2.717
34O–38H
2.713
20H–25N
2.512
20H–25N
2.476
20H–25N
2.458
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
13H–25N
1.889
10H–34O
1.973
10H–35O
4.141
15O–37H
1.971
4N–20H
1.889
11H–35O
—
11H–34O
4.363
15O–38H
—
13H–26O
2.022
10H–26O
1.972
11H–26O
—
(b)
Cluster IIntramolecular H-bonds
Cluster IIIntramolecularH-bonds
Cluster IIIIntramolecularH-bonds
Cluster IVIntramolecularH-bonds
Cluster VIntramolecularH-bonds
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
3N–10H
3.103
3N–10H
2.838
3N–10H
3.092
3N–10H
3.068
3N–10H
3.041
3N–11H
2.786
3N–11H
3.340
3N–11H
2.791
3N–11H
2.846
3N–11H
3.593
8O–10H
2.696
8O–10H
2.767
8O–10H
2.695
8O–10H
2.696
8O–10H
2.733
7O–11H
2.715
7O–11H
2.630
7O–11H
2.716
7O–11H
2.715
7O–11H
2.659
4N–13H
2.474
4N–13H
2.472
4N–13H
2.475
4N–13H
2.472
4N–13H
2.472
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
7O–22H
2.004
7O–20H
2.068
7O–22H
1.983
15O–22H
1.954
3N–22H
2.006
10H–21O
1.893
Cluster VI IntramolecularH-bonds
Cluster VIIIntramolecularH-bond
Cluster VIIIIntramolecularH-bonds
Cluster IXIntramolecularH-bonds
Cluster X (5 H2O) IntramolecularH-bonds
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
3N–10H
3.175
3N–10H
3.591
3N–10H
2.999
3N–10H
3.068
3N–10H
2.837
3N–11H
2.850
3N–11H
3.041
3N–11H
2.859
3N–11H
2.845
3N–11H
3.229
8O–10H
2.694
8O–10H
2.659
8O–10H
2.711
8O–10H
2.695
8O–10H
2.781
7O–11H
2.712
7O–11H
2.732
7O–11H
2.693
7O–11H
2.716
7O–11H
2.636
4N–13H
2.487
4N–13H
2.472
4N–13H
2.478
4N–13H
2.472
4N–13H
2.463
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
4N–22H
2.023
3N–22H
2.007
7O–20H
1.978
15O–22H
1.954
7O–25H
2.193
13H–21O
1.907
11H–21O
1.892
8O–32H
1.926
3N–30H
2.273
10H–29O
1.708
4N–22H
2.036
13H–21O
1.834
15O–28H
1.985
8O–23H
2.474


### 3.2. Vibrational Analysis

The experimental FT-IR (a) and micro-Raman spectra (b) of acetazolamide are given in Figure4. The experimental FT-IR and Raman spectra of the solid acetazolamide are also given in comparison with those of calculated gas phase spectra in Figures 5 and 6, respectively.Figure 4
Experimental FT-IR (a) and micro-Raman spectra (b) of acetazolamide in the regions 3600–2800 cm−1 and 1800–200 cm−1, respectively.Experimental FT-IR (solid) and calculated (scaled) absorption intensity spectra of acetazolamide in the regions 3600–2600 cm−1 (a) and 1800–400 cm−1 (b), respectively.
(a)
(b)Experimental (solid) and calculated (scaled) Raman intensities spectra of acetazolamide in the regions 3600–2800 cm−1 (a) and 1800–200 cm−1 (b), respectively.
(a)
(b)The calculated wavenumbers, the calculated Raman intensities, and the potential energy distributions of the vibrational modes of monomer and dimer (I) forms of acetazolamide and acetazolamide-H2O cluster (VI and X) are given in Table 3, in comparison with the experimental vibrational spectra of the investigated molecule. The calculated wavenumbers of the four low energy dimers of acetazolamide are given in Table S3 comparatively. The assignment procedure for the dimers was done according to the potential energy distribution of dimer I.Table 3
Experimental FT-IR (solid and aqueous solution) and Raman and calculated wavenumbes (cm−1) of the global conformations of monomeric, dimeric, and H2O clusters (VI and X) forms of acetazolamide molecule.
Assignment
This work
This workcalculated by DFT/6-31++G(d,p)
PED (≥5%) Free acetazolamide
This work
[13]
Experimental
Experimental
Free
Cluster VI
Cluster X
Dimer I
I
R
solid
I
R
aq
Raman
I
R
solid
Raman
ν
sca
.
ν
uns
RamanInt.Mono
ν
sca
.
ν
sca
.
ν
sca
.
Raman Int. Dimer I
ν
as(N–H)
ν
asN–H
3339 vs
—
3337 vw
3331 ma; 3301 sb
—
3473
3637
631
3474
3433
3484; 3484
1169; 22
ν
NH2
(
asym
,
) (100)
ν(N–H)
νN–H
3234 s
3193
3225 m
3227 ma; 3179 mb
3230 wa; 3183 shb
3451
3614
1980
3221
3099
3035; 2995
28; 4931
ν
NH (100)
ν
s(N–H)
ν
SN–H
3151 s
—
3145 w
3148 ma; 3093 wb
3152 wa; 3089 sb
3361
3520
1690
3360
3334
3364; 3364
119; 3162
ν
NH2
(
sym
) (100)
ν
as(CH3)
νCH3
3004 s
—
—
3001 vwa; 3014 vwb
3014b
3007
3149
1564
3011
3018
3015; 3014
181; 26336
ν
CH
3
(
asym
,
) (100)
ν
as(CH3)
νCH3
2948 m
—
2979 sh
2916 wa; 2985 vwb
2941 vsa; 2985b
2996
3138
1759
2995
2982
2992; 2992
3199; 575
ν
CH
3
(
asym
,
) (99)
ν
s(CH3)
νCH3
2923 m
—
2933 vs
2932b
2932b
2924
3061
5191
2924
2914
2923; 2923
7119; 3177
ν
CH
3
(
sym
,
) (100)
ν(C=O)
C=O
1699 vs
1694
1707 s
1698 sa; 1680 sb
1708 ma; 1677 wb
1724
1765
3123
1720
1719
1721; 1720
45; 5313
ν
OC (77) + νCC (6) + δCNC (7)
δNH2(scis,)
δNHip
1558 sh
1569
—
1574 wb
1571b
1562
1599
173
1560
1619
1554; 1554
200; 145
δ
NH2
(
scis
,
) (96)
ν(C=N) + δCNH
ν
asC=Nring
1550 vs
1552 s
1555 m
1542 vsa; 1553 vsb
1558b
1519
1554
6783
1546
1575
1581; 1565
2005; 15
ν
CN (60) + δCNH (28)
δ
as(CH3)
—
1457 w
1457
—
1454 shb
1455 shb
1460
1494
749
1459
1479
1463; 1462
72; 1099
δ
CHH (74) + δCHC (13)
δ
as(CH3)
ν
sC=N + δaCH3
1437 s
1435
1442 vs
1436 ma; 1437 mb
1443 vsa; 1434 mb
1440
1474
932
1443
1451
1449; 1448
611; 204
δ
CHH (81) + δCHC (8) + νCN (8)
ν
as(C=N)ring
ν
sC=N δaCH3
1420 sh
1418
1428 sh
1426 shb
1422b
1442
1476
2472
1449
1443
1456; 1456
7540; 1329
ν
CN (77) + δCHH (6)
ν
s(C=N)ring
δ
sCH3
1384 sh
1387
1370 w
1376 shb
1386b
1409
1442
5198
1424
1428
1439; 1435
72; 16412
ν
CN (54) + δCNH (23) + δCNN (6)
δ
s(CH3)
ν
asSO2
1365 s
1363
—
1368 vsb
1373b
1376
1409
1144
1377
1388
1379; 1379
1312; 42
δ
CHH (92) + νCC (6)
ν
as(SO2)
ν
asC–N–C
1346 vs
1344
1347 m
1343 vsa; 1322 sb
1348a
1312
1343
750
1314
1296
1319; 1319
1226; 690
ν
SO
2
(
asym
,
) (90) + δSNH (6)
ν(C–N)
νC–Nring
1282 s
—
1308 w
1280 ma,1286 mb
1242 wa, 1289a
1291
1322
109
1299
1249
1300; 1298
11; 186
ν
CN (62) + νCS (13) + δNCN (7) + δCCN (5)
δ
CNH
—
1234 s
1234
1241 m
1227 sa
1197
1225
629
1223
1319
1232; 1226
2041; 15
ν
CN (29) + δCNH (19) + νCC (13) + δNCO (12) + δCCH (9)
ν
s(SO2) + νs(N–N)ring
ν
sSO2
1168 vs
1166
1165 s
1163 vsa; 1176 vsb
1167 sa, 1168b
1121
1147
1022
1133
1100
1134; 1132
186; 1905
ν
OS (59) + νNN (34)
ν
s(SO2) + νs(N–N)ring
δ
ipCNNC
1105 m
1106
1103 sh
1122 mb
1121b
1100
1126
5630
1104
1140
1105; 1104
10516; 439
ν
OS (42) + νNN (39)
ρ(NH2)
“Ring mode”
1096 s
1097
1094 s
1095 sa; 1087 mb
1097sa; 1085mb
1071
1096
323
1070
1171
1063; 1063
14; 778
ρ
NH
2(91)
δ
ring
+
ν
ring
“Ring mode”
1038 w
1038
—
1045 ma,1045 mb
1041b
1042
1066
538
1045
1048
1046; 1045
139; 1528
δ
SCN (44) + νCS (42)
ρ(CH3)
—
—
—
—
1011a,1014b
1016b
1031
1055
10
1033
1045
1036; 1036
9; 29
δ
CCH (72) + δCHH (10) + γCNCO (16)
ρ(CH3)
νN–N
996 m
995
998 w
975 mb
955 ma, 968b
993
1016
276
1000
1014
1009; 1007
198; 1
δ
CCH (55) + νCC (18)
ν(C–N) + ν(C–C)
νS–N
944 s
947
956 m
939 sa,913 mb
916b
949
971
854
954
963
953;  949
2; 2584
ν
CN (25) + νCC (18) + νNN (9) + δCCN (15) + δCCH (8) + δOCN (7) + δNCN (6)
ν(S–N)
“Ring mode”
813 m
807 vw
813 wa,844 wb
—
824
843
2623
821
829
817;  817
573; 6259
ν
SN (55) + γS–NH2 (15)
δ
ring
+
ν
ring
“Ring mode”
783 m
—
780 ma,817 mb
821b
786
805
24
792
793
798;  786
9; 201
ν
CS (17) + νCC (7) + δCCN (12) + δNNC (42)
ν
as(C–S)ring
ν
a(C–S)ring
705 m
719
709 s
705 wa; 710 wb
712 ma; 711b
672
688
1385
681
682
683;  681
3652; 11
ν
CS (58) + νCC (9) + δNCO (9) + δNCC (12)
w(NH2)
ν
s(C–S)ring
675 s
668
680 vs
674 sa,673 sb
684 vsa, 680b
668
684
408
667
988
626; 624
5395; 178
γ
S–NH
2 (36) + νSN (19) + νCS (16) + δNSO (13)
ν
s(C–S)ring + δring
—
644 m
643
647 sh
643a; 651 wb
645b
645
660
5062
648
661
661;  659
8007; 3
δ
CSC (23) + νCS (23) + γS–NH2 (16) +νNS (6) + δNSO (6)
Γ
as(ring)
δ
ipN–C=O
621 vs
618
637 w
620 sa, 622 sb
636 wa,624b
624
639
225
639
636
635;  635
225; 69
Γ
NNCS (44) + γNNCS (30) + γCCNO (11)
γ
OCNC
—
608 sh
610 vw
613b
607b
608
622
16
778
591
597;  595
89; 52
γ
CNCO (24) + γNNCS (16) + ΓNNCS (20)+ ΓCNCO (13) + ΓNCNH (11) + γCCNH (8) + δCHC (7)
Γ
s
(
ring
)
δSO2
582 m
580 w
584 mb
588b
588
602
65
593
575
578;  576
123; 36
Γ
NCSC (50) + γNNCS (10) + γSCSN (21) + δCSO (6)
δ(C–C=O)
573 m
575
—
574 mb
578 ma, 574b
573
586
392
576
611
573;  570
28; 565
δ
CCO (19) + γS–NH2 (10) + νCS (16) + νCC (8) + δOSC (7) + δNCN (5)
w(SO2) + w(NH2)
δO=C–C
—
—
554 mb
560b
535
547
279
535
524
532;  531
106; 670
δ
NSO (34) + δCCO (10) + δCSN (10) + γS–NH2 (12) + νNS (5)
γ(N–H)
—
508 s
881
507 w
520 sb
507b
531
544
97
778
878
881;  864
3; 47
γ
CNCH (25) + ΓOCNH (20) + γCCNO (16) + ΓNNCS (10) + ΓNCNH (8)
δ
sciss(SO2)
wag SO2
457 vw
545
453 w
458 mb
450 wa,453b
467
478
1731
469
514
470;  470
702; 1107
δ
OSO (40) + δNSO (25) + νNS (11) + νCS (5)
ρ(SO2) + t(NH2)
ρSO2
406 vw
413 w
411 wb
417 wa,406b
432
442
208
435
468
433;  433
559; 16
δ
NSO (48) + γS–NH2 (16) + ΓNCNH (7) + ΓCSCN (7) + δSNH (7)
t(NH2) + t(SO2)
—
—
—
398b
395 shb
393
403
245
406
440
400;  400
35; 1655
δ
NSO (51) + ΓOSNH (18) + ΓNSCN (9)
δ
CCO
+
δ
OSO
Latice vib.
—
373 w
—
374 ma, 368b
386
395
1489
388
404
386;  384
2455; 190
δ
CCO (21) + δNCN (18) + δOSO (16) + δSCS (7) + δCCN (13) + νCS (7) + νCN (6)
δ
CCN
—
—
326 w
—
328 wa,330b
346
354
298
353
364
360;  355
15; 364
δ
CCN (41) + δNCS (23) + νCS (5)
t(NH2)
—
—
267 m
—
270 sa,  270b
291
298
503
302
386
316;  315
34; 594
δ
CSN (19) + δOSN (19) + ΓHNSO (19) + ΓNNCS (6)
t(NH2)
—
—
253 sh
—
256 ma,258b
274
280
363
274
—
275;  275
12; 952
Γ
HNSC (23) + δCSN (17) + δCSO (9) + δNSO (8) + ΓCSCN (9)
ν
C–S
—
—
—
—
230b
235
241
4727
237
270
237;  233
5884; 325
ν
CS (42) + δCSN (25) + δNCC (13) + δCSO (6)
Γ
CSNH
—
—
195 w
—
192b
219
224
291
230
231
243;  241
4402; 112
δ
OSN (32) + ΓHNSC (32) + ΓNSCS (18)
δ
CSN
—
—
160 w
—
136b
163
166
425
176
197
165;  163
1835; 12
δ
CSN (55) + δCNC (29)
γ
C–S
—
—
116
—
118b
123
126
1587
127
133
138;  132
4227; 114
γ
CSNS (33) + δCSO (27) + γCCNH (15) + ΓOCNH (10)
δ
SCS
—
—
105
—
105b
90
92
887
87
130
102;  100
567; 764
δ
SCS (44) + δCNC (13) + ΓCNCN (14)
Γ
NCNH
—
—
—
—
—
88
90
295
98
104
118;  104
57; 25
Γ
NCNH (54) + ΓCNCO (9) + γNCSN (7) + δCSN (6)
t(CH3)
—
—
—
—
—
56
57
1122
50
116
78; 62
817; 1641
γ
S–NH2 (36) + ΓNCCH (18) + γNCCH (10) + γNCSN (6) + ΓNNCS (6)
t(CH3)
—
—
—
—
—
39
40
2415
43
106
51; 48
459; 3691
Γ
NCCH (69) + γNCCO (17) + δHCH (6)
Γ
SCSN
—
—
—
—
—
11
11
1622
21
13
31; 12
572; 4369
Γ
SCSN (65) + γSCSN (15)
aRef [13], bRef [24].When acetazolamide goes from the monomer form to the dimer or to the water cluster, alterations in the vibrational spectra, due to formation of H-bonds, are estimated. The NH2, NH, C=O, and SO2 vibrational modes provide useful information on the intermolecular H-bonding interaction for the acetazolamide. On the other hand, NH stretching vibrations have anharmonic behavior [30–32]. Thus, in harmonic approximation procedure, NH2 stretching wavenumbers are overestimated due to neglect of anharmonicity [33]. The calculated values of ν(NH) stretching wavenumbers were 3451, 3035–2995 and 3221 cm−1 and 3099 cm−1 for monomer and dimer (dimer I) forms and water cluster (VI and X), respectively. As seen from Table 3, due to the participation of the −NH group in hydrogen bonds, ν(NH) in dimer I (with two N⋯H bonds; Figure 2) and water clusters VI and X (Figure 3) showed negative shifts, 456–416 and 230 cm−1 and 352 cm−1, respectively. The highest shifts (456 and 416 cm−1) were obtained for the dimer I. The calculated values suggest the presence of relatively strong N⋯H and NH⋯N hydrogen bond interactions in dimer I. The differences in hydrogen bonding strengths are responsible for these different wavenumber shifts. On the other hand C=O, and SO2 groups do not involve hydrogen bonding interaction in both dimer I form and water cluster VI, and we do not observe a remarkable change in this wavenumber on going from monomer form to dimer or cluster form.The acetazolamide C–N ring stretching modes were recorded at 1540 cm−1, 1425 cm−1, and 1275 cm−1 as strong intense bands in the IR and at 1425 cm−1 and 1260 cm−1 in Raman spectra of solid acetazolamide by Chufan et al. [19]. However, in other studies, these modes were obtained at 1498 cm−1 and 1450 cm−1 [11] and at 1571 cm−1 and 1452 cm−1 [34]. In the present study, the 1550, 1420, 1384, and 1311 cm−1 in IR and 1555, 1428, 1370, and 1308 cm−1 in Raman spectrum of solid acetazolamide are assigned to C–N stretching vibrations. The 1550 cm−1 (IR) C–N stretching mode is estimated to shift to higher wavenumber in both dimer and water cluster forms due to contribution of δCNH mode.Theνas(SO2) stretching mode was observed at 1345 cm−1 and 1347 cm−1 in IR and Raman spectra, respectively, and was calculated at 1312 cm−1, 1314 cm−1, 1296 cm−1, and 1319 cm−1 for monomer, clusters (VI and X), and dimer structure, respectively. Cami et al. [11] observed this mode at 1342 cm−1 and Baraldi recorded this mode at 1343 cm−1 and at 1348 cm−1 in IR and Raman spectra, respectively [13].The mixing ofδCNH+ν(C–N) modes was observed at 1234 and 1226 cm−1 as a strong band in the IR spectrum and at 1241 cm−1 in Raman spectrum as a medium band. This mode is calculated at 1197, 1223, 1319, and 1232–1226 cm−1 for free, clusters (VI and X), and dimer forms of acetazolamide, respectively.The bands at 1174 cm−1 and 1140 cm−1 were assigned to ν(N–N)ring stretching and νSO2 symmetric stretching vibrations, respectively, by Cami et al. [11]. In this study we observed a strong band at 1167 cm−1 in IR and 1165 cm−1 in Raman spectra and assigned it to νSO2+υ(N–N) mode according to PED calculations.. The PED result of this mode shows us that SO2 stretching and (N–N) stretching contributions are 59% and 44%, respectively.The strong band at 1095 cm−1 in the IR and Raman spectra is assigned to NH2 rocking mode, in agreement with PED. The corresponding mode was assigned at 1097 cm−1 [11] and at 1090 cm−1 [34] in previous studies. But Baraldi et al. assigned this strong intense band at 1095 cm−1 to a “ring mode” (1,3,4 thiadiazolic ring) [13]. Our assignments are in agreement with those of Cami et al. [11, 34].Cami et al. [11] assigned the 1058 cm−1 and 795 cm−1 bands to ring bending vibrations with contribution from ν(C–S) [11]. In this study, ring bending and ring stretching modes were found to be mixed and assigned to 1038 cm−1 and 782 cm−1 bands observed in IR spectrum as weak bands. We do not observe any band in the Raman spectrum attributable to these modes.We assigned theν(S–N) mode to the medium and weak bands observed at 812 cm−1 and 807 cm−1 in the IR and Raman spectra, respectively, according to the calculated results. This mode was assigned to the medium band at 872 cm−1 in [34], whereas Cami et al. [11] assigned the ν(S–N)+wagging (NH2) complicated mode to 944 cm−1 [11] and Baraldi et al. assigned it to 939 cm−1 [13].Theνas(C–S)ring was observed at 704 cm−1 as a medium band in the IR and at 709 cm−1 as strong band in the Raman spectra. The νs(C–S)ring stretching was mixed with ring bending and observed at 644 cm−1 and 647 cm−1 in IR and Raman spectra, respectively. The bands at 782 cm−1 and 679 cm−1 were assigned to ν(C-S) asymmetric and symmetric stretchings, respectively, by Cami et al. [34].In this study, the band located at 675 cm−1 in the IR and at 680 cm−1 in the Raman spectrum was assigned to w(NH2) mode. This mode with a contribution of CH bending vibration was assigned to 636 cm−1 by Cami et al. [11]. There was no assignment corresponding to the w(NH2) mode in [13].The ring torsion modes were expected in the region 530–650 cm−1. We assigned the 621 cm−1 and 582 cm−1 bands in IR spectrum and 637 and 580 cm−1 bands in the Raman spectrum to the ring torsion modes. These modes were observed at 650 cm−1 and 534 cm−1 by Cami et al. [11]. Baraldi et al. did not assign ring torsion modes [13]. However, the bands at 621 cm−1 in the IR spectrum and at 637 cm−1 in Raman spectrum were identified as δip(N–C=O) mode [13], and the band at 584 cm−1 in IR spectrum and at 588 cm−1 in the Raman spectrum was assigned to δSO2 [24].The acetazolamide SO2 wagging mode, coupled with NH2 wagging mode, was calculated at 535 cm−1 for free form. We did not observe this mode in the experimental IR or Raman spectra, but it was observed at 588 cm−1 [34] and at 552 cm−1 [11] in previous studies. This mode was assigned to 458 cm−1 in IR and 453 cm−1 in Raman spectra [24]. Rocking and twisting SO2 vibrations were situated at 416 cm−1 and 315 cm−1 [11]. In this study, corresponding values of these modes were assigned at 406 cm−1 and at 413 cm−1.The assignment in the range of 500–250 cm−1 is difficult due to the presence of several overlapping weak bands. These bands are associated with vibrational and lattice modes and most of them are strongly overlapped.The experimental micro-Raman (solid) and calculated (scaled) Raman intensity spectra and the experimental FT-IR (solid) and calculated (scaled) absorption intensity spectra of acetazolamide dimer (I–IV) are given in Figures7 and 8.Figure 7
Experimental micro-Raman (solid) and calculated (scaled) Raman intensity spectra of acetazolamide dimer (I–IV) in the region 1800–200 cm−1, respectively.Figure 8
Experimental FT-IR (solid) and calculated (scaled) absorption intensity spectra of acetazolamide dimer (I–IV) in the region 1800–400 cm−1, respectively.ν
C=O, νC=N, νaSO2, and νsSO2 vibrations are observed at 1699, 1550, 1346, and 1168 cm−1, respectively, in the IR spectrum of solid acetazolamide, but they are observed at 1694 (Δ=νaq.-νsolid=-5 cm−1), 1552 (Δ=νaq.-νsolid=+2 cm−1), 1344 (Δ=νaq.-νsolid=-2 cm−1), and 1166 (Δ=νaq.-νsolid=-2 cm−1) respectively, in that of aqueous solution. The water cluster structure (X) of Figure 3 simulates best the aqueous environment for the molecule. The experimental FT-IR (solid and aqueous solution) and calculated (scaled) absorption intensity spectra of H2O clusters (I–X) of acetazolamide in the region of 1800–800 cm−1 are given in Figure S2.


### 3.3. HOMO-LUMO and NBO Analysis

The frontier molecular orbitals are important in determining the molecular reactivity. The highest occupied molecular orbital (HOMO) energy characterizes the ability of electron giving, the lowest unoccupied molecular orbital (LUMO) energy characterizes the ability of electron accepting, and the gap between HOMO and LUMO characterizes the molecular stability. The HOMO and LUMO energies of acetazolamide are calculated by DFT method at B3LYP/6-31G** level of theory, and the atomic orbital compositions of the frontier molecular orbitals for acetazolamide are shown in Figure S3. LUMO is located over the ring, the methyl group, and C–N bond which is located on the chain of the molecule. The HOMO is located over the ring, carbonyl group, and amine group, and consequently the homo-lumo transition implies an electron density transfer to ring, methyl group, and C–N bond from the carbonyl group and amine group. The computed low HOMO-LUMO energy gap (−5.497 eV) shows the charge transfer interactions, taking place within the molecule.The NBO analysis of acetazolamide molecule is calculated by DFT method at B3LYP/6-31G** level of theory. The nature and credibility of the intermolecular hydrogen bonding can be analysed by searching the changes in electron density in the environments of N⋯H hydrogen bonds. The NBO analysis of acetazolamide clearly explains the evidence of the formation of strong H-bonded interaction between the LP(N) and σ*(N–H) antibonding orbitals. The hyperconjugative interactions CR(N4)→σ*(H20–N21), LP1(N4)→σ*(H20–N21), and CR(N25)→σ*(N12–H13), LP1(N25)→σ*(N12–H13) are obtained as 0.37, 23.34, 0.37, and 23.34 kcal/mol, respectively, and are shown in Table S4. The differences in E2 energies are reasonably due to the fact that the accrual of electron density is in the N–H bonds.The comparison of NBO analysis between dimer and monomer forms of acetazolamide indicates the formation of two H-bonded interactions between nitrogen lone electron pairs andσ*(N–H) antibonding orbitals.The magnitudes of charge transfer from lone pairs ofn(N4) and n(N25) of the hydrogen-bonded N atoms into the antibonds σ*(N21–H20) and σ*(N12–H13) increased upon dimerization (0.05298e and 0.05299e) (see Table S5). Similar conclusion can be obtained while considering the energy of each orbital. The electron density in the N–H antibonding orbitals σ*(N4–C5) and π*(N4–C5) is increased significantly (0.00291e and 0.04037e, resp.) because of the dimerization. The elongation (0.02 Å) thereby weakens the bond. This is associated with the downshifts of stretching frequency (Table S2 and Table 3).The intramolecular hyperconjugative interactionσ(N4–C5) distribution to σ*(C5–N12) and σ*(N12–C14) caused less stabilization of 1.09 and 2.06 kcal/mol. This interaction of π(N4–C5) with π*(C2–N3) leads to strong delocalization of 14.11 kcal/mol. There occurs a strong intramolecular hyperconjugative interaction from n(N12) to σ*(N4–C5) and σ*(C14–O15) with 47.21 and 54.82 kcal/mol (Table S4). The increased electron density at the N12 atom leads to the elongation of N–H bond and a lowering of the N–H stretching wavenumber (see Table S2 and Table 3).


## 3.1. Conformational Analysis and H-Bonding Interactions

The molecular model of acetazolamide with the atom numbering scheme is given in Figure1. Stable low energy conformers of free acetazolamide molecule were obtained firstly by potential energy surface scan studies by iteratively varying D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C) dihedral angles with step angle of 60°. PES scan for acetazolamide calculated with D1 (1S-2C-6S-9N), D2 (4N-5C-12N-14C), and D3 (5C-12N-14C-16C) dihedral angles at the B3LYP/6-31G++(d,p) method is shown in Figure S1 (see Figure S1 in Supplementary Materials available online at http://dx.doi.org/10.1155/2013/538917).  The 45 conformers were identified between 0 and 10 kcal/mol relative energy intervals. Their dihedral angles and relative energies were given in Table S1. The geometry optimization was then performed on the lowest energy conformer. The estimated four most stable conformers of acetazolamide dimers are illustrated in Figure 2. All possible conformations of acetazolamide interacting with one water molecule were investigated, and  acetazolamide interacting with five water molecules was determined as the final possible conformation of acetazolamide-water cluster. Figure 3 demonstrates the geometries of the 10 stable acetazolamide-water clusters obtained using DFT method with B3LYP/6-31++G(d,p) basis set. The energies of the four most stable dimers (I–IV) and energetically preferred H2O-acetazolamide clusters are given in Table 1. It is known that the basis set superposition error (BSSE) effect is rather significant on the structure and energy of dimer forms, so removing this effect is very important. Therefore, optimization of the dimer I was also carried out along with the counterpoise correction scheme proposed by Boys and Bernardi [29]. The BSSE uncorrected and corrected distances between two acetazolamide units {R(5C–22C)(Å)} and binding energy (ΔE) of acetazolamide dimer {ΔE=2*Emonomer-Edimer} are given in Table 1(b). As seen in Table 1(b), the energy of dimer I is found to be 17.39 kcal/mol lower then total energy of the two monomer (2*Emonomer) units, indicating that intermolecular hydrogen bonding plays an important role in stabilization of the molecule.Table 1
(a) The calculated energies of monomeric, dimeric (I–IV), and H2O clusters (I–IX). (b) The counterpoise uncorrected and corrected interatomic distance (Å) and binding energy (kcal/mol) of dimer I.
(a)
Calculated energy (kcal/mol)
Energy differences (kcal/mol)
Monomer
−876645.7
—
Dimer I
−1753308.7
0
Dimer II
−1753299.4
9.3
Dimer III
−1753297.9
10.8
Dimer IV
−1753301.5
7.2
Cluster I
−924613.6
6.1
Cluster II
−924617.1
2.6
Cluster III
−924613.6
6.0
Cluster IV
−924614.6
5.1
Cluster V
−924619.1
0.6
Cluster VI
−924619.7
0
Cluster VII
−924619.1
0.6
Cluster VIII
−924612.9
6.7
Cluster IX
−924614.6
5.1
(b)
BSSE uncorrected
BSSE corrected
R(5C–22C) (Å)
ΔE (kcal/mol)
R(5C–22C) (Å)
Δ(E + BSSE) (kcal/mol)
Dimer I
4.23
17.4
4.25
16.1Figure 1
The most stable geometry, atom numbering, and searched dihedral angles of acetazolamide.Figure 2
The energy minimized structures of four low energy conformers (I–IV) of the dimer forms of the acetazolamide, (I–IV), respectively.Figure 3
The energy minimized structures of ten low energy conformers (I–X) of H2O clusters of the acetazolamide (I–X), respectively.The predicted geometrical parameters such as bond lengths and bond angles of the stable conformation of acetazolamide monomer, dimer (dimer I), and water cluster (VI and X) of acetazolamide, calculated at B3LYP method with 6–31G++(d,p), are presented in Table S2 in accordance with the atom numbering scheme as given in Figure1. By comparing theoretical and experimental bond lengths [9], it is easily seen that the computed bond lengths at B3LYP/6-31++G(d,p) method were slightly longer since the theoretical calculations were performed on isolated molecule in the gaseous state, whereas the experimental results are for the solid phase of the molecule [9, 10, 12]. Main differences between crystal structure [9] and monomer acetazolamide occur in H containing bond lengths and angles, and this is probably due to insufficient predictions for light atoms in the crystal structure study done in 1974 [9]. The theoretical results are mainly consistent with reports of other studies [11, 24]. Comparison of the geometry parameters of monomer form with those of dimer and water cluster of acetazolamide clearly shows the effects of intermolecular hydrogen bonding.The intra- and intermolecular hydrogen bonds of the dimer forms (I–IV) of the four low energy conformations together with those of ten energetically preferred water clusters are tabulated in Table2. The lowest energy conformer of dimer form (dimer I) makes stronger interhydrogen bonding interactions.Table 2
The intra- and interhydrogen bonds of the dimer conformers (I–IV) and H2O clusters (I–X) of acetazolamide.
(a)
Dimer I
Intramolecular H-bonds
Dimer IIIntramolecular H-bonds
Dimer IIIIntramolecular H-bonds
Dimer IVIntramolecular H-bonds
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
3N–10H
3.377
3N–10H
3.342
3N–10H
3.148
4N–13H
2.468
3N–11H
2.975
3N–11H
2.962
3N–11H
2.816
20H–25N
2.468
8O–10H
2.698
8O–10H
2.717
8O–10H
2.700
7O–11H
2.696
7O–11H
2.694
7O–11H
2.715
4N–13H
2.512
4N–13H
2.467
4N–13H
2.488
29N–37H
3.377
29N–37H
3.196
29N–37H
3.141
29N–38H
2.975
29N–38H
2.766
29N–38H
2.857
35O–37H
2.968
35O–37H
2.689
35O–37H
2.697
34O–38H
2.696
34O–38H
2.717
34O–38H
2.713
20H–25N
2.512
20H–25N
2.476
20H–25N
2.458
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
13H–25N
1.889
10H–34O
1.973
10H–35O
4.141
15O–37H
1.971
4N–20H
1.889
11H–35O
—
11H–34O
4.363
15O–38H
—
13H–26O
2.022
10H–26O
1.972
11H–26O
—
(b)
Cluster IIntramolecular H-bonds
Cluster IIIntramolecularH-bonds
Cluster IIIIntramolecularH-bonds
Cluster IVIntramolecularH-bonds
Cluster VIntramolecularH-bonds
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
3N–10H
3.103
3N–10H
2.838
3N–10H
3.092
3N–10H
3.068
3N–10H
3.041
3N–11H
2.786
3N–11H
3.340
3N–11H
2.791
3N–11H
2.846
3N–11H
3.593
8O–10H
2.696
8O–10H
2.767
8O–10H
2.695
8O–10H
2.696
8O–10H
2.733
7O–11H
2.715
7O–11H
2.630
7O–11H
2.716
7O–11H
2.715
7O–11H
2.659
4N–13H
2.474
4N–13H
2.472
4N–13H
2.475
4N–13H
2.472
4N–13H
2.472
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
7O–22H
2.004
7O–20H
2.068
7O–22H
1.983
15O–22H
1.954
3N–22H
2.006
10H–21O
1.893
Cluster VI IntramolecularH-bonds
Cluster VIIIntramolecularH-bond
Cluster VIIIIntramolecularH-bonds
Cluster IXIntramolecularH-bonds
Cluster X (5 H2O) IntramolecularH-bonds
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
Atoms
Bond (Å)
3N–10H
3.175
3N–10H
3.591
3N–10H
2.999
3N–10H
3.068
3N–10H
2.837
3N–11H
2.850
3N–11H
3.041
3N–11H
2.859
3N–11H
2.845
3N–11H
3.229
8O–10H
2.694
8O–10H
2.659
8O–10H
2.711
8O–10H
2.695
8O–10H
2.781
7O–11H
2.712
7O–11H
2.732
7O–11H
2.693
7O–11H
2.716
7O–11H
2.636
4N–13H
2.487
4N–13H
2.472
4N–13H
2.478
4N–13H
2.472
4N–13H
2.463
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
IntermolecularH-bonds
4N–22H
2.023
3N–22H
2.007
7O–20H
1.978
15O–22H
1.954
7O–25H
2.193
13H–21O
1.907
11H–21O
1.892
8O–32H
1.926
3N–30H
2.273
10H–29O
1.708
4N–22H
2.036
13H–21O
1.834
15O–28H
1.985
8O–23H
2.474


## 3.2. Vibrational Analysis

The experimental FT-IR (a) and micro-Raman spectra (b) of acetazolamide are given in Figure4. The experimental FT-IR and Raman spectra of the solid acetazolamide are also given in comparison with those of calculated gas phase spectra in Figures 5 and 6, respectively.Figure 4
Experimental FT-IR (a) and micro-Raman spectra (b) of acetazolamide in the regions 3600–2800 cm−1 and 1800–200 cm−1, respectively.Experimental FT-IR (solid) and calculated (scaled) absorption intensity spectra of acetazolamide in the regions 3600–2600 cm−1 (a) and 1800–400 cm−1 (b), respectively.
(a)
(b)Experimental (solid) and calculated (scaled) Raman intensities spectra of acetazolamide in the regions 3600–2800 cm−1 (a) and 1800–200 cm−1 (b), respectively.
(a)
(b)The calculated wavenumbers, the calculated Raman intensities, and the potential energy distributions of the vibrational modes of monomer and dimer (I) forms of acetazolamide and acetazolamide-H2O cluster (VI and X) are given in Table 3, in comparison with the experimental vibrational spectra of the investigated molecule. The calculated wavenumbers of the four low energy dimers of acetazolamide are given in Table S3 comparatively. The assignment procedure for the dimers was done according to the potential energy distribution of dimer I.Table 3
Experimental FT-IR (solid and aqueous solution) and Raman and calculated wavenumbes (cm−1) of the global conformations of monomeric, dimeric, and H2O clusters (VI and X) forms of acetazolamide molecule.
Assignment
This work
This workcalculated by DFT/6-31++G(d,p)
PED (≥5%) Free acetazolamide
This work
[13]
Experimental
Experimental
Free
Cluster VI
Cluster X
Dimer I
I
R
solid
I
R
aq
Raman
I
R
solid
Raman
ν
sca
.
ν
uns
RamanInt.Mono
ν
sca
.
ν
sca
.
ν
sca
.
Raman Int. Dimer I
ν
as(N–H)
ν
asN–H
3339 vs
—
3337 vw
3331 ma; 3301 sb
—
3473
3637
631
3474
3433
3484; 3484
1169; 22
ν
NH2
(
asym
,
) (100)
ν(N–H)
νN–H
3234 s
3193
3225 m
3227 ma; 3179 mb
3230 wa; 3183 shb
3451
3614
1980
3221
3099
3035; 2995
28; 4931
ν
NH (100)
ν
s(N–H)
ν
SN–H
3151 s
—
3145 w
3148 ma; 3093 wb
3152 wa; 3089 sb
3361
3520
1690
3360
3334
3364; 3364
119; 3162
ν
NH2
(
sym
) (100)
ν
as(CH3)
νCH3
3004 s
—
—
3001 vwa; 3014 vwb
3014b
3007
3149
1564
3011
3018
3015; 3014
181; 26336
ν
CH
3
(
asym
,
) (100)
ν
as(CH3)
νCH3
2948 m
—
2979 sh
2916 wa; 2985 vwb
2941 vsa; 2985b
2996
3138
1759
2995
2982
2992; 2992
3199; 575
ν
CH
3
(
asym
,
) (99)
ν
s(CH3)
νCH3
2923 m
—
2933 vs
2932b
2932b
2924
3061
5191
2924
2914
2923; 2923
7119; 3177
ν
CH
3
(
sym
,
) (100)
ν(C=O)
C=O
1699 vs
1694
1707 s
1698 sa; 1680 sb
1708 ma; 1677 wb
1724
1765
3123
1720
1719
1721; 1720
45; 5313
ν
OC (77) + νCC (6) + δCNC (7)
δNH2(scis,)
δNHip
1558 sh
1569
—
1574 wb
1571b
1562
1599
173
1560
1619
1554; 1554
200; 145
δ
NH2
(
scis
,
) (96)
ν(C=N) + δCNH
ν
asC=Nring
1550 vs
1552 s
1555 m
1542 vsa; 1553 vsb
1558b
1519
1554
6783
1546
1575
1581; 1565
2005; 15
ν
CN (60) + δCNH (28)
δ
as(CH3)
—
1457 w
1457
—
1454 shb
1455 shb
1460
1494
749
1459
1479
1463; 1462
72; 1099
δ
CHH (74) + δCHC (13)
δ
as(CH3)
ν
sC=N + δaCH3
1437 s
1435
1442 vs
1436 ma; 1437 mb
1443 vsa; 1434 mb
1440
1474
932
1443
1451
1449; 1448
611; 204
δ
CHH (81) + δCHC (8) + νCN (8)
ν
as(C=N)ring
ν
sC=N δaCH3
1420 sh
1418
1428 sh
1426 shb
1422b
1442
1476
2472
1449
1443
1456; 1456
7540; 1329
ν
CN (77) + δCHH (6)
ν
s(C=N)ring
δ
sCH3
1384 sh
1387
1370 w
1376 shb
1386b
1409
1442
5198
1424
1428
1439; 1435
72; 16412
ν
CN (54) + δCNH (23) + δCNN (6)
δ
s(CH3)
ν
asSO2
1365 s
1363
—
1368 vsb
1373b
1376
1409
1144
1377
1388
1379; 1379
1312; 42
δ
CHH (92) + νCC (6)
ν
as(SO2)
ν
asC–N–C
1346 vs
1344
1347 m
1343 vsa; 1322 sb
1348a
1312
1343
750
1314
1296
1319; 1319
1226; 690
ν
SO
2
(
asym
,
) (90) + δSNH (6)
ν(C–N)
νC–Nring
1282 s
—
1308 w
1280 ma,1286 mb
1242 wa, 1289a
1291
1322
109
1299
1249
1300; 1298
11; 186
ν
CN (62) + νCS (13) + δNCN (7) + δCCN (5)
δ
CNH
—
1234 s
1234
1241 m
1227 sa
1197
1225
629
1223
1319
1232; 1226
2041; 15
ν
CN (29) + δCNH (19) + νCC (13) + δNCO (12) + δCCH (9)
ν
s(SO2) + νs(N–N)ring
ν
sSO2
1168 vs
1166
1165 s
1163 vsa; 1176 vsb
1167 sa, 1168b
1121
1147
1022
1133
1100
1134; 1132
186; 1905
ν
OS (59) + νNN (34)
ν
s(SO2) + νs(N–N)ring
δ
ipCNNC
1105 m
1106
1103 sh
1122 mb
1121b
1100
1126
5630
1104
1140
1105; 1104
10516; 439
ν
OS (42) + νNN (39)
ρ(NH2)
“Ring mode”
1096 s
1097
1094 s
1095 sa; 1087 mb
1097sa; 1085mb
1071
1096
323
1070
1171
1063; 1063
14; 778
ρ
NH
2(91)
δ
ring
+
ν
ring
“Ring mode”
1038 w
1038
—
1045 ma,1045 mb
1041b
1042
1066
538
1045
1048
1046; 1045
139; 1528
δ
SCN (44) + νCS (42)
ρ(CH3)
—
—
—
—
1011a,1014b
1016b
1031
1055
10
1033
1045
1036; 1036
9; 29
δ
CCH (72) + δCHH (10) + γCNCO (16)
ρ(CH3)
νN–N
996 m
995
998 w
975 mb
955 ma, 968b
993
1016
276
1000
1014
1009; 1007
198; 1
δ
CCH (55) + νCC (18)
ν(C–N) + ν(C–C)
νS–N
944 s
947
956 m
939 sa,913 mb
916b
949
971
854
954
963
953;  949
2; 2584
ν
CN (25) + νCC (18) + νNN (9) + δCCN (15) + δCCH (8) + δOCN (7) + δNCN (6)
ν(S–N)
“Ring mode”
813 m
807 vw
813 wa,844 wb
—
824
843
2623
821
829
817;  817
573; 6259
ν
SN (55) + γS–NH2 (15)
δ
ring
+
ν
ring
“Ring mode”
783 m
—
780 ma,817 mb
821b
786
805
24
792
793
798;  786
9; 201
ν
CS (17) + νCC (7) + δCCN (12) + δNNC (42)
ν
as(C–S)ring
ν
a(C–S)ring
705 m
719
709 s
705 wa; 710 wb
712 ma; 711b
672
688
1385
681
682
683;  681
3652; 11
ν
CS (58) + νCC (9) + δNCO (9) + δNCC (12)
w(NH2)
ν
s(C–S)ring
675 s
668
680 vs
674 sa,673 sb
684 vsa, 680b
668
684
408
667
988
626; 624
5395; 178
γ
S–NH
2 (36) + νSN (19) + νCS (16) + δNSO (13)
ν
s(C–S)ring + δring
—
644 m
643
647 sh
643a; 651 wb
645b
645
660
5062
648
661
661;  659
8007; 3
δ
CSC (23) + νCS (23) + γS–NH2 (16) +νNS (6) + δNSO (6)
Γ
as(ring)
δ
ipN–C=O
621 vs
618
637 w
620 sa, 622 sb
636 wa,624b
624
639
225
639
636
635;  635
225; 69
Γ
NNCS (44) + γNNCS (30) + γCCNO (11)
γ
OCNC
—
608 sh
610 vw
613b
607b
608
622
16
778
591
597;  595
89; 52
γ
CNCO (24) + γNNCS (16) + ΓNNCS (20)+ ΓCNCO (13) + ΓNCNH (11) + γCCNH (8) + δCHC (7)
Γ
s
(
ring
)
δSO2
582 m
580 w
584 mb
588b
588
602
65
593
575
578;  576
123; 36
Γ
NCSC (50) + γNNCS (10) + γSCSN (21) + δCSO (6)
δ(C–C=O)
573 m
575
—
574 mb
578 ma, 574b
573
586
392
576
611
573;  570
28; 565
δ
CCO (19) + γS–NH2 (10) + νCS (16) + νCC (8) + δOSC (7) + δNCN (5)
w(SO2) + w(NH2)
δO=C–C
—
—
554 mb
560b
535
547
279
535
524
532;  531
106; 670
δ
NSO (34) + δCCO (10) + δCSN (10) + γS–NH2 (12) + νNS (5)
γ(N–H)
—
508 s
881
507 w
520 sb
507b
531
544
97
778
878
881;  864
3; 47
γ
CNCH (25) + ΓOCNH (20) + γCCNO (16) + ΓNNCS (10) + ΓNCNH (8)
δ
sciss(SO2)
wag SO2
457 vw
545
453 w
458 mb
450 wa,453b
467
478
1731
469
514
470;  470
702; 1107
δ
OSO (40) + δNSO (25) + νNS (11) + νCS (5)
ρ(SO2) + t(NH2)
ρSO2
406 vw
413 w
411 wb
417 wa,406b
432
442
208
435
468
433;  433
559; 16
δ
NSO (48) + γS–NH2 (16) + ΓNCNH (7) + ΓCSCN (7) + δSNH (7)
t(NH2) + t(SO2)
—
—
—
398b
395 shb
393
403
245
406
440
400;  400
35; 1655
δ
NSO (51) + ΓOSNH (18) + ΓNSCN (9)
δ
CCO
+
δ
OSO
Latice vib.
—
373 w
—
374 ma, 368b
386
395
1489
388
404
386;  384
2455; 190
δ
CCO (21) + δNCN (18) + δOSO (16) + δSCS (7) + δCCN (13) + νCS (7) + νCN (6)
δ
CCN
—
—
326 w
—
328 wa,330b
346
354
298
353
364
360;  355
15; 364
δ
CCN (41) + δNCS (23) + νCS (5)
t(NH2)
—
—
267 m
—
270 sa,  270b
291
298
503
302
386
316;  315
34; 594
δ
CSN (19) + δOSN (19) + ΓHNSO (19) + ΓNNCS (6)
t(NH2)
—
—
253 sh
—
256 ma,258b
274
280
363
274
—
275;  275
12; 952
Γ
HNSC (23) + δCSN (17) + δCSO (9) + δNSO (8) + ΓCSCN (9)
ν
C–S
—
—
—
—
230b
235
241
4727
237
270
237;  233
5884; 325
ν
CS (42) + δCSN (25) + δNCC (13) + δCSO (6)
Γ
CSNH
—
—
195 w
—
192b
219
224
291
230
231
243;  241
4402; 112
δ
OSN (32) + ΓHNSC (32) + ΓNSCS (18)
δ
CSN
—
—
160 w
—
136b
163
166
425
176
197
165;  163
1835; 12
δ
CSN (55) + δCNC (29)
γ
C–S
—
—
116
—
118b
123
126
1587
127
133
138;  132
4227; 114
γ
CSNS (33) + δCSO (27) + γCCNH (15) + ΓOCNH (10)
δ
SCS
—
—
105
—
105b
90
92
887
87
130
102;  100
567; 764
δ
SCS (44) + δCNC (13) + ΓCNCN (14)
Γ
NCNH
—
—
—
—
—
88
90
295
98
104
118;  104
57; 25
Γ
NCNH (54) + ΓCNCO (9) + γNCSN (7) + δCSN (6)
t(CH3)
—
—
—
—
—
56
57
1122
50
116
78; 62
817; 1641
γ
S–NH2 (36) + ΓNCCH (18) + γNCCH (10) + γNCSN (6) + ΓNNCS (6)
t(CH3)
—
—
—
—
—
39
40
2415
43
106
51; 48
459; 3691
Γ
NCCH (69) + γNCCO (17) + δHCH (6)
Γ
SCSN
—
—
—
—
—
11
11
1622
21
13
31; 12
572; 4369
Γ
SCSN (65) + γSCSN (15)
aRef [13], bRef [24].When acetazolamide goes from the monomer form to the dimer or to the water cluster, alterations in the vibrational spectra, due to formation of H-bonds, are estimated. The NH2, NH, C=O, and SO2 vibrational modes provide useful information on the intermolecular H-bonding interaction for the acetazolamide. On the other hand, NH stretching vibrations have anharmonic behavior [30–32]. Thus, in harmonic approximation procedure, NH2 stretching wavenumbers are overestimated due to neglect of anharmonicity [33]. The calculated values of ν(NH) stretching wavenumbers were 3451, 3035–2995 and 3221 cm−1 and 3099 cm−1 for monomer and dimer (dimer I) forms and water cluster (VI and X), respectively. As seen from Table 3, due to the participation of the −NH group in hydrogen bonds, ν(NH) in dimer I (with two N⋯H bonds; Figure 2) and water clusters VI and X (Figure 3) showed negative shifts, 456–416 and 230 cm−1 and 352 cm−1, respectively. The highest shifts (456 and 416 cm−1) were obtained for the dimer I. The calculated values suggest the presence of relatively strong N⋯H and NH⋯N hydrogen bond interactions in dimer I. The differences in hydrogen bonding strengths are responsible for these different wavenumber shifts. On the other hand C=O, and SO2 groups do not involve hydrogen bonding interaction in both dimer I form and water cluster VI, and we do not observe a remarkable change in this wavenumber on going from monomer form to dimer or cluster form.The acetazolamide C–N ring stretching modes were recorded at 1540 cm−1, 1425 cm−1, and 1275 cm−1 as strong intense bands in the IR and at 1425 cm−1 and 1260 cm−1 in Raman spectra of solid acetazolamide by Chufan et al. [19]. However, in other studies, these modes were obtained at 1498 cm−1 and 1450 cm−1 [11] and at 1571 cm−1 and 1452 cm−1 [34]. In the present study, the 1550, 1420, 1384, and 1311 cm−1 in IR and 1555, 1428, 1370, and 1308 cm−1 in Raman spectrum of solid acetazolamide are assigned to C–N stretching vibrations. The 1550 cm−1 (IR) C–N stretching mode is estimated to shift to higher wavenumber in both dimer and water cluster forms due to contribution of δCNH mode.Theνas(SO2) stretching mode was observed at 1345 cm−1 and 1347 cm−1 in IR and Raman spectra, respectively, and was calculated at 1312 cm−1, 1314 cm−1, 1296 cm−1, and 1319 cm−1 for monomer, clusters (VI and X), and dimer structure, respectively. Cami et al. [11] observed this mode at 1342 cm−1 and Baraldi recorded this mode at 1343 cm−1 and at 1348 cm−1 in IR and Raman spectra, respectively [13].The mixing ofδCNH+ν(C–N) modes was observed at 1234 and 1226 cm−1 as a strong band in the IR spectrum and at 1241 cm−1 in Raman spectrum as a medium band. This mode is calculated at 1197, 1223, 1319, and 1232–1226 cm−1 for free, clusters (VI and X), and dimer forms of acetazolamide, respectively.The bands at 1174 cm−1 and 1140 cm−1 were assigned to ν(N–N)ring stretching and νSO2 symmetric stretching vibrations, respectively, by Cami et al. [11]. In this study we observed a strong band at 1167 cm−1 in IR and 1165 cm−1 in Raman spectra and assigned it to νSO2+υ(N–N) mode according to PED calculations.. The PED result of this mode shows us that SO2 stretching and (N–N) stretching contributions are 59% and 44%, respectively.The strong band at 1095 cm−1 in the IR and Raman spectra is assigned to NH2 rocking mode, in agreement with PED. The corresponding mode was assigned at 1097 cm−1 [11] and at 1090 cm−1 [34] in previous studies. But Baraldi et al. assigned this strong intense band at 1095 cm−1 to a “ring mode” (1,3,4 thiadiazolic ring) [13]. Our assignments are in agreement with those of Cami et al. [11, 34].Cami et al. [11] assigned the 1058 cm−1 and 795 cm−1 bands to ring bending vibrations with contribution from ν(C–S) [11]. In this study, ring bending and ring stretching modes were found to be mixed and assigned to 1038 cm−1 and 782 cm−1 bands observed in IR spectrum as weak bands. We do not observe any band in the Raman spectrum attributable to these modes.We assigned theν(S–N) mode to the medium and weak bands observed at 812 cm−1 and 807 cm−1 in the IR and Raman spectra, respectively, according to the calculated results. This mode was assigned to the medium band at 872 cm−1 in [34], whereas Cami et al. [11] assigned the ν(S–N)+wagging (NH2) complicated mode to 944 cm−1 [11] and Baraldi et al. assigned it to 939 cm−1 [13].Theνas(C–S)ring was observed at 704 cm−1 as a medium band in the IR and at 709 cm−1 as strong band in the Raman spectra. The νs(C–S)ring stretching was mixed with ring bending and observed at 644 cm−1 and 647 cm−1 in IR and Raman spectra, respectively. The bands at 782 cm−1 and 679 cm−1 were assigned to ν(C-S) asymmetric and symmetric stretchings, respectively, by Cami et al. [34].In this study, the band located at 675 cm−1 in the IR and at 680 cm−1 in the Raman spectrum was assigned to w(NH2) mode. This mode with a contribution of CH bending vibration was assigned to 636 cm−1 by Cami et al. [11]. There was no assignment corresponding to the w(NH2) mode in [13].The ring torsion modes were expected in the region 530–650 cm−1. We assigned the 621 cm−1 and 582 cm−1 bands in IR spectrum and 637 and 580 cm−1 bands in the Raman spectrum to the ring torsion modes. These modes were observed at 650 cm−1 and 534 cm−1 by Cami et al. [11]. Baraldi et al. did not assign ring torsion modes [13]. However, the bands at 621 cm−1 in the IR spectrum and at 637 cm−1 in Raman spectrum were identified as δip(N–C=O) mode [13], and the band at 584 cm−1 in IR spectrum and at 588 cm−1 in the Raman spectrum was assigned to δSO2 [24].The acetazolamide SO2 wagging mode, coupled with NH2 wagging mode, was calculated at 535 cm−1 for free form. We did not observe this mode in the experimental IR or Raman spectra, but it was observed at 588 cm−1 [34] and at 552 cm−1 [11] in previous studies. This mode was assigned to 458 cm−1 in IR and 453 cm−1 in Raman spectra [24]. Rocking and twisting SO2 vibrations were situated at 416 cm−1 and 315 cm−1 [11]. In this study, corresponding values of these modes were assigned at 406 cm−1 and at 413 cm−1.The assignment in the range of 500–250 cm−1 is difficult due to the presence of several overlapping weak bands. These bands are associated with vibrational and lattice modes and most of them are strongly overlapped.The experimental micro-Raman (solid) and calculated (scaled) Raman intensity spectra and the experimental FT-IR (solid) and calculated (scaled) absorption intensity spectra of acetazolamide dimer (I–IV) are given in Figures7 and 8.Figure 7
Experimental micro-Raman (solid) and calculated (scaled) Raman intensity spectra of acetazolamide dimer (I–IV) in the region 1800–200 cm−1, respectively.Figure 8
Experimental FT-IR (solid) and calculated (scaled) absorption intensity spectra of acetazolamide dimer (I–IV) in the region 1800–400 cm−1, respectively.ν
C=O, νC=N, νaSO2, and νsSO2 vibrations are observed at 1699, 1550, 1346, and 1168 cm−1, respectively, in the IR spectrum of solid acetazolamide, but they are observed at 1694 (Δ=νaq.-νsolid=-5 cm−1), 1552 (Δ=νaq.-νsolid=+2 cm−1), 1344 (Δ=νaq.-νsolid=-2 cm−1), and 1166 (Δ=νaq.-νsolid=-2 cm−1) respectively, in that of aqueous solution. The water cluster structure (X) of Figure 3 simulates best the aqueous environment for the molecule. The experimental FT-IR (solid and aqueous solution) and calculated (scaled) absorption intensity spectra of H2O clusters (I–X) of acetazolamide in the region of 1800–800 cm−1 are given in Figure S2.


## 3.3. HOMO-LUMO and NBO Analysis

The frontier molecular orbitals are important in determining the molecular reactivity. The highest occupied molecular orbital (HOMO) energy characterizes the ability of electron giving, the lowest unoccupied molecular orbital (LUMO) energy characterizes the ability of electron accepting, and the gap between HOMO and LUMO characterizes the molecular stability. The HOMO and LUMO energies of acetazolamide are calculated by DFT method at B3LYP/6-31G** level of theory, and the atomic orbital compositions of the frontier molecular orbitals for acetazolamide are shown in Figure S3. LUMO is located over the ring, the methyl group, and C–N bond which is located on the chain of the molecule. The HOMO is located over the ring, carbonyl group, and amine group, and consequently the homo-lumo transition implies an electron density transfer to ring, methyl group, and C–N bond from the carbonyl group and amine group. The computed low HOMO-LUMO energy gap (−5.497 eV) shows the charge transfer interactions, taking place within the molecule.The NBO analysis of acetazolamide molecule is calculated by DFT method at B3LYP/6-31G** level of theory. The nature and credibility of the intermolecular hydrogen bonding can be analysed by searching the changes in electron density in the environments of N⋯H hydrogen bonds. The NBO analysis of acetazolamide clearly explains the evidence of the formation of strong H-bonded interaction between the LP(N) and σ*(N–H) antibonding orbitals. The hyperconjugative interactions CR(N4)→σ*(H20–N21), LP1(N4)→σ*(H20–N21), and CR(N25)→σ*(N12–H13), LP1(N25)→σ*(N12–H13) are obtained as 0.37, 23.34, 0.37, and 23.34 kcal/mol, respectively, and are shown in Table S4. The differences in E2 energies are reasonably due to the fact that the accrual of electron density is in the N–H bonds.The comparison of NBO analysis between dimer and monomer forms of acetazolamide indicates the formation of two H-bonded interactions between nitrogen lone electron pairs andσ*(N–H) antibonding orbitals.The magnitudes of charge transfer from lone pairs ofn(N4) and n(N25) of the hydrogen-bonded N atoms into the antibonds σ*(N21–H20) and σ*(N12–H13) increased upon dimerization (0.05298e and 0.05299e) (see Table S5). Similar conclusion can be obtained while considering the energy of each orbital. The electron density in the N–H antibonding orbitals σ*(N4–C5) and π*(N4–C5) is increased significantly (0.00291e and 0.04037e, resp.) because of the dimerization. The elongation (0.02 Å) thereby weakens the bond. This is associated with the downshifts of stretching frequency (Table S2 and Table 3).The intramolecular hyperconjugative interactionσ(N4–C5) distribution to σ*(C5–N12) and σ*(N12–C14) caused less stabilization of 1.09 and 2.06 kcal/mol. This interaction of π(N4–C5) with π*(C2–N3) leads to strong delocalization of 14.11 kcal/mol. There occurs a strong intramolecular hyperconjugative interaction from n(N12) to σ*(N4–C5) and σ*(C14–O15) with 47.21 and 54.82 kcal/mol (Table S4). The increased electron density at the N12 atom leads to the elongation of N–H bond and a lowering of the N–H stretching wavenumber (see Table S2 and Table 3).


## 4. Conclusion

In this study, the monomer and dimer forms and water clusters of acetazolamide were studied by using the DFT method at B3LYP/6-31G++(d,p) level of theory. The effect of basis set superposition error (BSSE) on the structure and energy of acetazolamide dimer has been investigated. The most stable structure of the dimer (dimer I) possesses interaction energy of 16.116 kcal/mol after the basis set superposition error (BSSE) correction. The difference between the BSSE corrected (16.116 kcal/mol) and uncorrected (17.390 kcal/mol) interaction energies indicates the magnitude of error caused due to the basis set superposition. The energy of dimer I is found to be lower than the total energy of the two monomer(2*Emonomer) units, indicating that intermolecular hydrogen bonding plays an important role in stabilization of the molecule. The wavenumbers associated with the related molecule are in a good agreement with [11, 34]. The comparison of NBO analysis between dimer and monomer forms of acetazolamide indicates the formation of two H-bonded interactions between nitrogen lone electron pairs and σ*(N–H) antibonding orbitals. Increasing of the electron density in the N–H antibonding orbitals upon dimerization can be associated with the downshifts of N–H stretching frequency.



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*Source: 538917-2013-12-21.xml*