Synthesis, characterization, X-ray diffraction studies and biological evaluation of tert-butyl 4-(2-ethoxy-2-oxoethyl)-piperazine-1-carboxylate and tert-butyl 4-(2-hydrazino-2-oxoethyl)piperazine-1-carboxylate

Article abstract of DOI:10.1007/s11164-016-2542-7

Two derivatives of N-Boc piperazine, an ester derivative, i.e., tert-butyl 4-(2-ethoxy-2-oxoethyl)-piperazine-1-carboxylate (1), and, a hydrazide derivative tert-butyl 4-(2-hydrazino-2-oxoethyl)piperazine-1-carboxylate (2) were synthesized and were characterized by FT-IR, ¹H & ¹³C NMR and LCMS spectroscopic studies. The structures of both 1 and 2 were further confirmed by single crystal X-ray diffraction analysis. The molecule of 1 is linear in shape with the ethyl acetate moiety adopting fully extended conformation, while the molecule of 2 is L-shaped with the molecule being twisted at the C10 atom. The crystal structure of 1 adopts a two-dimensional zig-zag architecture featuring C–H…O intermolecular interactions, while that of 2 features strong N–H…O hydrogen bonds and intermolecular interactions of the type N–H…N and C–H…N, resulting in a two-dimensional structure. Furthermore, a detailed analysis of the intermolecular interactions and crystal packing of 1 and 2 via Hirshfeld surface analysis and fingerprint plots was performed. The antibacterial and antifungal activities of both the compounds have been studied against several microorganisms, and were found to be moderately active.

Full text of DOI:10.1007/s11164-016-2542-7

Res Chem Intermed
DOI 10.1007/s11164-016-2542-7
Synthesis, characterization, X-ray diffraction studies
and biological evaluation of tert-butyl 4-(2-ethoxy-2-
oxoethyl)-piperazine-1-carboxylate and tert-butyl 4-(2-
hydrazino-2-oxoethyl)piperazine-1-carboxylate
Bhuvan Kulkarni1 Sharada Thimmappaiah2 Harisha Padigar2
•
•
•
Suchetan Parameshwar Adimule3 Naveen Shivalingegowda4
Lokanath Krishnappagowda Neratur5 Manjunatha Kumsi1
•
•
•
Received: 1 December 2015 / Accepted: 7 April 2016
Ó Springer Science+Business Media Dordrecht 2016
Abstract Two derivatives of N-Boc piperazine, an ester derivative, i.e., tert-butyl
4-(2-ethoxy-2-oxoethyl)-piperazine-1-carboxylate (1), and, a hydrazide derivative
tert-butyl 4-(2-hydrazino-2-oxoethyl)piperazine-1-carboxylate (2) were synthesized
and were characterized by FT-IR, 1H & 13C NMR and LCMS spectroscopic studies.
The structures of both 1 and 2 were further confirmed by single crystal X-ray
diffraction analysis. The molecule of 1 is linear in shape with the ethyl acetate moiety
adopting fully extended conformation, while the molecule of 2 is L-shaped with the
molecule being twisted at the C10 atom. The crystal structure of 1 adopts a two-
dimensional zig-zag architecture featuring C–H…O intermolecular interactions,
while that of 2 features strong N–H…O hydrogen bonds and intermolecular inter-
actions of the type N–H…N and C–H…N, resulting in a two-dimensional structure.
Furthermore, a detailed analysis of the intermolecular interactions and crystal
packing of 1 and 2 via Hirshfeld surface analysis and fingerprint plots was performed.
The antibacterial and antifungal activities of both the compounds have been studied
against several microorganisms, and were found to be moderately active.
&
Manjunatha Kumsi
manju.kumsi@gmail.com
1
2
3
4
5
Department of Chemistry, Nagarjuna College of Engineering and Technology, Devanahalli,
Bangalore 562164, India
Department of Physics, Nagarjuna College of Engineering and Technology, Devanahalli,
Bangalore 562164, India
Department of Chemistry, University College of Science, Tumkur University, Tumkur 572103,
India
Institution of Excellence, University of Mysore, VijnanaBhavan, Manasagangotri, Mysore,
India
Department of Studies in Physics, University of Mysore, Manasagangotri, Mysore, India
123

B. Kulkarni et al.
Keywords Piperazine Á Crystal structure Á Hirshfeld surface Á Antibacterial
activity Á Antifungal activity
Introduction
Piperazine and N-Boc piperazine and their simple derivatives such as tert-butyl
4-(2-ethoxy-2-oxoethyl)piperazine-1-carboxylate (1) and tert-butyl 4-(2-hydrazino-
2-oxoethyl)piperazine-1-carboxylate (2) serve as useful building blocks/intermedi-
ates in the synthesis of several novel organic compounds such as amides,
sulphonamides, Mannich bases, Schiff’s bases, thiazolidinones, azetidinones, and
imidazolinones [1–4]. These derived compounds have shown a wide spectrum of
biological activities such as the antibacterial [5–7], antifungal [8], anticancer [9,
10], antiparasitic [11, 12] antihistamine [13] and antidepressive activities [14]. Such
diverse biological activities of the compounds containing piperazine rings can be
attributed to the conformational flexibility of the piperazine ring and the polar
nitrogen atoms in the piperazine ring and, thereby, enhancing favorable interaction
with macromolecules. Further, due to its easy modificability, proper alkality, water
solubility, and the capacity for the formation of hydrogen bonds and adjustment of
molecular physicochemical properties, the incorporation of the piperazine ring is
considered as an important synthetic strategy in the field of drug discovery [3, 4].
Hirshfeld surface analysis is an effective tool for exploring packing modes and
intermolecular interactions in molecular crystals [15–19]. It offers a visual picture of
intermolecular interactions and of molecular shape in a crystalline environment.
Surface features characteristic of different types of intermolecular interactions can be
identified, and these features can be revealed by colour coding distances from the
surface to the nearest atom exterior (de plots) or interior (di plots) to the surface. This
gives a visual picture of different types of interactions present, and also reflects their
relative contributions from molecule to molecule. Furthermore, 2-D fingerprint plots,
in particular the breakdown of 2-D fingerprint plots into specific atom…atom contacts
in a crystal, provide a quantitative idea of the types of intermolecular contacts
experienced by molecules in the bulk and presents this information in a convenient
colour plot. Thus, based on the above facts, compounds 1 and 2 were synthesized, and
their molecular and crystal structures were established by single crystal X-ray
diffraction analysis. A detailed analysis of the intermolecular interactions and crystal
packing of 1 and 2 via Hirshfeld surface analysis and fingerprint plots was performed.
Furthermore, 1 and 2 were tested for their antibacterial and antifungal activities.
Experimental
Materials
All reagents were purchased from Spectrochem Pvt. Ltd., India, and were used
without further purifications. Melting points of 1 and 2 were determined in open
capillary tubes.
123

Synthesis, characterization, X-ray diffraction studies…
Preparation of tert-butyl 4-(2-ethoxy-2-oxoethyl)piperazine-1-carboxylate (1)
A mixture of N-Bocpiperzine (0.01 mol) and triethyl amine (0.02 mol) were placed
in acetonitrile (250 ml) and stirred for 4 h, and the mixture was cooled to 0 °C
(Scheme 1). To this solution, ethylbromoacetate (0.01 mol) was added dropwise
over a period of 15 min. The resulting reaction mixture was stirred for 1 h at 0 °C
and then at room temperature (27 °C) for 15 h. The completion of the reaction was
monitored by Thin Layer Chromatography (TLC). The excess solvent was then
distilled off under reduced pressure and then ethyl acetate was extracted by water.
The resulting concentrated solution was poured into ice cold water. The separated
solid was filtered, washed with water, and dried.The crude product thus obtained
was recrystallized from ethanol. (Yield. 85 %; M.p. 393 K).
Preparation of tert-butyl 4-(2-hydrazino-2-oxoethyl)piperazine-1-carboxylate (2)
A
mixture of tert-butyl 4-(2-ethoxy-2-oxoethyl)piperazine-1-carboxylate (1)
(2.723 g, 0.01 mol) and hydrazine hydrate (0.02 mol) was refluxed in absolute
ethanol (50 ml) for 8 h (Scheme 2). The completion of the reaction was monitored
by TLC. The excess solvent was then distilled off under reduced pressure, and the
concentrated solution was poured into ice cold water. The separated solid was
filtered, washed with water, and dried. The crude product thus obtained was
recrystallized from ethanol. (Yield. 80 %; M.p. 428 K).
Spectral measurements
Infrared spectra were recorded on a Shimadzu FT-IR 157 spectrophotometer in the
range of 400–4000 cm-1 in the solid state using KBr pellets. The 1H and 13C NMR
spectra were recorded on a Bruker Avance II-400 (400 MHz) and a Bruker Avance
(100 MHz) spectrometer using TMS as an internal standard and CDCl3 as the
solvent. LCMS were recorded on an Agilent Mass spectrometer.
Scheme 1 Synthesis of 1
123

B. Kulkarni et al.
Scheme 2 Synthesis of 2
Preparation of crystals of 1 and 2
Single crystals of both 1 and 2 suitable for single crystal X-ray studies were
obtained from the slow solvent evaporation technique at room temperature.
Compound 1 (50 mg) was dissolved in aqueous ethanol (50 ml, with a few drops of
water) and the solution was stirred for 15 min with moderate heating to get a clear
solution. The hot solution was filtered, and the solvent was allowed to evaporate at
room temperature (27 °C). Colourless prism-like crystals of 1 were obtained after
7 days. The crystals obtained were separated from the supersaturated solution by
filtration and dried. The colourless prism-like crystals of 2 were obtained from a
similar procedure after 5 days.
X-ray crystallographic study
The X-ray intensity data were collected at temperatures of 173 K (for 1) and 296 K
(for 2) on a Bruker Proteum2 CCD diffractometer equipped with an X-ray generator
˚
operating at 45 kV and 10 mA, using Cu-Ka radiation of wavelength 1.54178 A.
Data were collected for 24 frames per set with different settings of u (0° and 90°),
keeping the scan width of 0.5°, exposure time of 5 s, the sample-to-detector distance
of 45.10 mm, and 2h value at 46.6°. Image processing and data reduction were done
using SAINT-Plus and XPREP [20]. The structure was solved by direct methods
using SHELXS-97 [21]. All the non-hydrogen atoms were revealed in the first-
difference Fourier map itself and were refined anisotropically. All the hydrogen
atoms were positioned geometrically. In 1, all H atoms were positioned geomet-
˚
˚
rically, with C–H = 0.99 A for methylene H, and C–H = 0.98 A for methyl H, and
refined using a riding model with Uiso(H) = 1.2Ueq(C) for methylene H, and
U
iso(H) = 1.5Ueq(C) for methyl H atoms. The amino H atoms were located in a
123

Synthesis, characterization, X-ray diffraction studies…
difference map and were refined isotropically with the bond length restraint N–
˚
H = 0.90(1) A. In 2, all H atoms were positioned geometrically, with C–H = 0.97
˚
˚
A for methylene H, and C–H = 0.96 A for methyl H, and refined using a riding
model with Uiso(H) = 1.2 Ueq(C) for methylene H, and Uiso(H) = 1.5 Ueq(C) for
methyl H atoms. All the geometrical calculations for 1 and 2 were carried out using
the program PLATON [22] within the WinGX suite [23]. The molecular and packing
diagrams were generated using the software MERCURY [24]. The crystallographic
data and refinement parameters for 1 and 2 are summarized in Table 1.
Hirshfeld surface calculations
Hirshfeld surface analyses were carried out and finger print plots were plotted using
the software CrystalExplorer 3.0 [25]. The dnorm plots were mapped with colour scale
in between -0.18 au (blue) and 1.4 au (red). The 2-D fingerprint plots [15, 16] were
˚
displayed by using the expanded 0.6–2.8 A view with the de and di distance scales
displayed on the graph axes. When the cif files of 1 and 2 were uploaded into the
Table 1 Crystal data and structure refinement for 1 and 2
Compound code
CCDC deposit No.
Empirical formula
Formula weight
Crystal system
Space group
1
2
1435775
1435776
C
13N2O4H24
C11H22N4O3
258.33
272.34
Monoclinic
Triclinic
P21/c
P-1
˚
a/A
8.125(3)
5.5647(5)
6.5288(6)
19.7375(19)
99.367(4)
90.527(5)
103.326(3)
687.65(11)
2
˚
b/A
15.871(6)
˚
c/A
12.323(4)
a/°
b/°
c/°
90.00
98.47(2)
90.00
3
˚
Volume/A
1571.7(10)
Z
4
D
calc/g/cm3
1.151
1.248
l/mm-1
0.699
0.758
F(000)
Crystal size/mm3
592.0
280.0
0.29 9 0.22 9 0.18
9.14 to 128.44
9177
0.27 9 0.26 9 0.24
9.1 to 129.38
6360
2h range for data collection/°
Reflections collected
Independent reflections
Data/restraints/parameters
Goodness-of-fit on F2
Final R indexes (I C 2r (I))
Final R indexes (all data)
2495
2248
2495/0/176
2248/3/178
1.083
1.046
R1 = 0.0992, wR2 = 0.2380
R1 = 0.1224, wR2 = 0.2648
0.41/-0.73
R1 = 0.0639, wR2 = 0.1833
R1 = 0.0659, wR2 = 0.1852
0.45/-0.34
-3
˚
Largest diff. peak/hole/e A
123

B. Kulkarni et al.
CrystalExplorer software, all bond lengths to hydrogen were automatically modified
˚
to typical standard neutron values, i.e., C–H = 1.083 and N–H = 1.009 A.
Antimicrobial studies
The antimicrobial activities were determined using disc diffusion method [26] by
measuring the zone of inhibition in mm. Both 1 and 2 were screened in vitro at
concentrations of 10 lg/disc for their antibacterial activities against two Gram-
positive strains (Staphylococcus aureus and Bacillus subtilis) and two Gram-
negative strains (Escherichia coli and Pseudomonas aeruginosa). Antifungal
evaluations of 1 and 2 were carried out against Candida albicans and Aspergillus
niger at concentrations of 10 lg/disc. Standard antibacterial drug ciprofloxacin
(10 lg/disc) and antifungal drug fluconazole (10 lg/disc) were also tested under
similar conditions against these organisms.
Results and discussion
FT-IR spectral analysis
The FT-IR spectra of compounds 1 and 2 are respectively shown in Figs. 1 and 2. In
the IR spectrum of 1 (Fig. 1), there are two sharp characteristic absorption bands at
1748.2 and 1678.7 cm-1, respectively, which are assigned to the two carbonyl
groups of the –N–CO–O– and –CH2–CO–O– segments, while the IR spectrum of 2
(Fig. 2), shows two sharp absorption bands at 1686.4 and 1666.2 cm-1 corre-
sponding to the two carbonyl groups, and a strong band at 3290.9 cm-1, which is
assigned to the N–H stretching. Thus, the IR spectra clearly show the formation of
these compounds from their respective starting materials.
1H and 13C NMR spectral analysis
1
The H NMR spectra of 1 and 2 are shown in Figs. 3 and 4, respectively.
Fig. 1 FT-IR spectrum of 1
123

Synthesis, characterization, X-ray diffraction studies…
Fig. 2 FT-IR spectrum of 2
Fig. 3 1H NMR spectrum of 1
1
In the H NMR spectrum of 1, two sharp singlet’s at d 1.46 and 3.22 ppm
integrated to nine and two protons, respectively, due to –C(CH3)3 and –N–CH2–CO
protons. A triplet centered at d 1.28 ppm corresponding to three methyl protons of –
O–CH2–CH3 with J = 6.7 Hz is observed. Two characteristic triplets at d 2.53 ppm
with J = 5.2 Hz and d 3.48 with J = 5.0 Hz are integrated to four protons due to
the methylene protons of the piperazine ring system. The spectrum also showed a
quartet at d 4.19 ppm and integrates to two protons of the ester group due to methyl
protons with J = 7.3 Hz. The 1H NMR spectrum of 2 showed no signal for methyl
and methylene protons of the ester group, but showed two new broad singlets at d
123

B. Kulkarni et al.
Fig. 4 1H NMR spectrum of 2
3.87 and d 8.09 ppm, which are assigned to the NH2 and NH protons, respectively.
Thus, the conversion of compound 1 to compound 2 is confirmed by this. Further,
prominent singlet’s at d 1.46 and 3.22 ppm are as a result of –C(CH3)3 and –N–
CH2–CO protons, respectively. Two characteristic triplets at d 2.47 ppm with
J = 5.2 Hz and d 3.09 ppm with J = 5.0 Hz integrates to four protons that are due
to the methylene protons of the piperazine ring.
The 13C NMR spectra of 1 and 2 are shown in Figs. 5 and 6, respectively.
The 13C NMR spectrum of compound 1 showed characteristic signals at d 14.19
(C13), 28.36 (C7, C8 & C9), 52.64 (C1 & C4), 59.39 (C2 & C3), 60.65 (C11),
79.65 ppm (C6) for various carbon atoms, and also showed the presence of two
carbonyl carbon atoms C5 and C11 at 154.63 and 170.07 ppm, respectively. In 2,
13C NMR spectrum showed characteristic signals at 28.33 (C7, C8 & C9), 53.38 (C1
& C4), 60.69 (C2 & C3), 79.91 ppm (C6) for various carbon atoms, and signals at
154.51 and 170.06 ppm, respectively, for the two carbonyl carbon atoms C5 and
C11.
LCMS analysis
The mass spectrum of compound 1 showed an intense molecular ion (M??1) peak
at m/z 273 corresponding to its molecular formula, C13H24N2O4 (Mol. Wt. 272)
(Fig. 7). Similarly, the mass spectrum of compound 2 (Fig. 8) showed an intense
molecular ion (M??1) peak at m/z 259 corresponding to its molecular formula,
C11H22N4O3 (Mol. Wt. 258).
123

Synthesis, characterization, X-ray diffraction studies…
Fig. 5 13C NMR spectrum of 1
Fig. 6 13C NMR spectrum of 2
Single-crystal X-ray diffraction
A
colourless prism-like single crystal of
1
of dimension (in mm3)
0.29 9 0.22 9 0.18 was mounted on the X-ray diffractometer. The crystallographic
123

B. Kulkarni et al.
Fig. 7 LCMS of 1
Fig. 8 LCMS of 2
data and refinement parameters for 1 are summarized in Table 1. The details of the
programs/software’s employed for solving the crystal structure and generating
molecular and crystal packing diagrams are already described in the experimental
section and, hence, not mentioned here again. Similarly, a colourless prism-like
single crystal of 2 of dimension (in mm3) 0.27 9 0.26 9 0.24 was mounted on the
X-ray diffractometer for recording the X-ray intensity data. The crystallographic
data and refinement parameters for 2 are summarized in Table 1. The list of
important bond lengths, bond angles and torsional angles in 1 and 2 are given in
Tables 2 and 3, respectively.
123

Synthesis, characterization, X-ray diffraction studies…
Table 2 Selected bond lengths, angles and torsions in 1
˚
Length/A
Bond
Bond
Angle/°
Bond
Angle/°
O2-C6
O2-C5
O4-C11
O4-C12
O1-C5
N2-C3
N2-C10
N2-C2
N1-C4
N1-C5
N1-C1
C11-O3
1.486(3)
1.340(3)
1.335(3)
1.444(4)
1.218(3)
1.470(3)
1.462(3)
1.456(3)
1.460(3)
1.348(3)
1.469(3)
1.185(3)
C5-O2-C6
121.37(16)
117.8(2)
O2-C6-C9
C7-C6-C9
C8-C6-C7
C8-C6-C9
O2-C5-N1
O1-C5-O2
O1-C5-N1
O4-C12-C13
O2-C6-C7
O2-C6-C8
111.3(2)
109.6(2)
110.2(2)
113.3(2)
111.47(19)
124.3(2)
124.2(2)
108.9(3)
101.82(18)
110.1(2)
C11-O4-C12
C10-N2-C3
C2-N2-C10
C5-N1-C4
112.10(17)
110.66(19)
120.43(19)
124.7(2)
C5-N1-C1
O4-C11-C10
O3-C11-O4
O3-C11-C10
N2-C10-C11
110.4(2)
123.6(2)
126.0(3)
114.4(2)
Torsion
Angle/°
177.73(18)
Torsion
Angle/°
O4-C11-C10-N2
C3-N2-C10-C11
C11-O4-C12-C13
O3-C11-C10-N2
C10-N2-C3-C4
C10-N2-C2-C1
C4-N1-C5-O2
C2-N2-C10-C11
C6-O2-C5-O1
C6-O2-C5-N1
C5-N1-C4-C3
C5-N1-C1-C2
C1-N1-C4-C3
C1-N1-C5-O2
C12-O4-C11-O3
C12-O4-C11-C10
-161.2(2)
7.9(4)
78.1(3)
-171.4(4)
-3.9(4)
-171.2(2)
-142.5(2)
142.5(3)
51.2(3)
-176.9(2)
174.9(2)
-172.5(2)
8.4(4)
-7.9(4)
-1.4(4)
177.0(3)
C4-N1-C5-O1
C1-N1-C5-O1
173.1(3)
Molecular and crystal structure of 1
Figure 9 represents the ORTEP diagram of molecule 1 with thermal ellipsoids
drawn at 50 % probability.
The molecule of 1 is linear in shape with the ester moiety (ethyl acetate moiety)
adopting fully extended conformation. The ethyl acetate moiety in 1 is almost in a
single plane with the root mean squared deviation (rmsd) of non-hydrogen atoms
˚
being 0.053(1) A. The piperazine ring in the structure adopts a chair conformation,
˚
which is confirmed by the puckering amplitude Q = 0.568 (3) A, and angles
h = 7.8(2)° and U = 170(2)°. Furthermore, the bonds N1-C5 and N2-C10 make an
angle 72.35(16)° and 75.87(15)°, respectively, with the Cremer and Pople (C & P)
plane of the piperazine ring [27], and, thus, are in the equatorial plane of the
piperazine ring. The dihedral angle between the mean plane of the piperazine ring
(A) (excluding hydrogen atoms) and the mean plane defined by the ester chain C10-
C11(O3)-O4-C12-C13 (B) is 48.9(6)°, while that between A and the mean plane of
the ester chain O1-C5-O2-C6 (C) is 23.7(8)°, and that between B and C is
123

B. Kulkarni et al.
Table 3 Selected bond lengths, angles and torsions in 2
˚
Length/A
Bond
Bond
Angle/°
Bond
Angle/°
O3-C11
O1-C5
O2-C5
O2-C6
N4-N3
N3-C11
N1-C5
C10-C11
C6-C8
C6-C7
C6-C9
N2-C10
1.244(2)
1.212(3)
1.352(3)
1.467(3)
1.422(2)
1.330(3)
1.362(3)
1.518(3)
1.511(3)
1.505(4)
1.513(4)
1.459(2)
C5-O2-C6
C11-N3-N4
C10-N2-C3
C2-N2-C10
C5-N1-C4
C5-N1-C1
O1-C5-O2
O1-C5-N1
O2-C5-N1
O3-C11-N3
121.55(17)
121.54(16)
115.07(16)
111.90(16)
118.21(17)
123.42(17)
125.06(19)
124.62(19)
110.31(18)
123.28(19)
O3-C11-C10
N3-C11-C10
O2-C6-C8
O2-C6-C7
O2-C6-C9
C8-C6-C9
C7-C6-C8
C7-C6-C9
N2-C10-C11
121.56(17)
115.16(17)
111.14(19)
110.06(19)
101.97(19)
110.1(2)
111.6(2)
111.6(3)
112.48(16)
Torsion
Angle/°
Torsion
Angle/°
N4-N3-C11-O3
N4-N3-C11-C10
N2-C10-C11-O3
N2-C10-C11-N3
C5-O2-C6-C8
C5-O2-C6-C7
C5-O2-C6-C9
C5-N1-C4-C3
C5-N1-C1-C2
C10-N2-C3-C4
-10.5(3)
168.72(17)
-170.43(18)
10.4(2)
C3-N2-C10-C11
C2-N2-C10-C11
C4-N1-C5-O1
C4-N1-C5-O2
C1-N1-C5-O1
C1-N1-C5-O2
C6-O2-C5-O1
C6-O2-C5-N1
C10-N2-C2-C1
-139.53(17)
94.5(2)
-13.3(3)
167.96(18)
-170.5(2)
10.8(3)
58.3(3)
-65.9(3)
175.5(2)
-1.8(3)
148.65(18)
-150.91(19)
170.26(16)
176.94(18)
-168.17(16)
Fig. 9 ORTEP view of the molecule 1 with thermal ellipsoids drawn at 50 % probability
76.42(13)°. The N1 and N2 nitrogen atoms of the piperazine ring are sp3 hybridized
with pyramidal geometries. The important bond lengths, bond angles, and torsions
of the side chains are listed in Table 2. All the bond lengths and angles in 1 are
within the expected ranges.
123

Synthesis, characterization, X-ray diffraction studies…
The crystal structure of 1 features C10-H10B…O1 intermolecular interactions
(Table 4), which connect the symmetrically identical molecules into C(8) chains
[28] parallel to the crystallographic a axis (Fig. 10). The molecules in the adjacent
chains are related to each other by twofold rotation symmetry, and these chains are
nearly perpendicular to one another. These adjacent chains are interconnected by
C7-H7B…O3 intermolecular interactions (Table 4) leading into zigzag sheets
(Fig. 11), thus, the overall supramolecular architecture is two.
Molecular and crystal structure of 2
Figure 12 represents the ORTEP diagram of molecule 2 with thermal ellipsoids
drawn at 50 % probability.
The molecule of 2 (i.e., the hydrazide derivative of N-Boc piperazine) is
L-shaped (Fig. 12) with the molecule being twisted at the C10 atom, in contrast to
the linear shape of the ester derivative, i.e., molecule 1. Similar to 1, the piperazine
ring in the molecule of 2 adopts a chair conformation, which is confirmed by the
˚
puckering amplitude Q = 0.571 (1) A, and angles h = 171.9(5)° and
U = 5.028(8)° [27].
Further, the bonds N1-C5 and N2-C10 make an angle 78.50(1)° and 82.9(4)°,
respectively, with the Cremer and Pople (C & P) plane of the piperazine ring [27],
and, thus, are in the equatorial plane of the piperazine ring. All the bond lengths
and angles in 2 are within the expected ranges. In particular, the length of the
C = O bond and the trigonal planar geometry around C11 and N3 atoms are
consistent with the expected resonance forms of the hydrazide group, the rmsd of
2
˚
C10-C11(O3)-N3-N4 plane being 0.058(1) A. The N3 (-NH group) atom is sp
hybridized, while the geometry around the N4 atom (-NH2 group) is pyramidal
(sp3 hybridized), the sum of valence angles around N4 being 323°. N1 and N2
atoms of the piperazine ring are also sp3 hybridized with pyramidal geometries.
The important bond lengths, bond angles, and torsions of the side chains are listed
in Table 3. The dihedral angle between the mean plane of the piperazine ring
(A) (excluding hydrogen atoms) and the plane defined by the side chain C10-
C11(O3)-N3-N4 (B) is 77.7(7)°, which is significantly larger than that observed
between the piperazine ring and the ester group B in 1. The dihedral angle
between A and the ester plane O1-C5-O2-C6 (C) is 12.8(8)°, which is quite
smaller than that observed in 1. The dihedral angle between the two side chains B
and C is 79.7(3)°, similar to that in molecule 1.
The molecule 2 possesses three potential H-bond donors: one in the –NH and two
in the –NH2 moieties of the hydrazide chain, and possesses three H-bond acceptors:
N atom of the –NH2 group and the O atoms of the two carbonyl groups (one in
hydrazide and one in the ester moiety). In the initial stage of packing, the molecules
related by the inversion centre are interconnected via strong N4-H2N4…O3
(Table 4) hydrogen bonds forming R22(10) dimers (Fig. 13). The C=O group (of the
hydrazide chain) is in fact involved as a bifurcated acceptor in H-bonding resulting
in another strong N4-H1N4…O3 (Table 4) hydrogen bonds giving rise to a
tetrameric pattern across inversion centres, whose graph set descriptor is R42(8)
(Fig. 13). These alternating ring patterns form ribbons parallel to a axis (Fig. 13). In
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B. Kulkarni et al.
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Synthesis, characterization, X-ray diffraction studies…
Fig. 10 Packing of molecules of 1 in the crystal structure forming a two-dimensional architecture.
Intermolecular interactions are shown as light blue lines. (Color figure online)
Fig. 11 Zigzag sheets observed in the crystal structure of 1 when viewed down the a axis. The molecules
shown in white and green are related by twofold rotation symmetry. (Color figure online)
addition to these, the nitrogen atom of the –NH2 group is involved as a bifurcated
acceptor in hydrogen bonding, resulting in N3-HN3…N4 and C10-H10B…N4
intermolecular interactions (Table 4) that form R22(6) and R22(10) dimers, respec-
tively (Fig. 14). The ester carbonyl group is not involved in any hydrogen bonds,
and overall, the crystal structure of 2 is a two-dimensional supramolecular
architecture.
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B. Kulkarni et al.
Fig. 12 ORTEP view of the molecule 2 with thermal ellipsoids drawn at 50 % probability
Fig. 13 Packing of molecules of 2 in the crystal structure resulting in ribbons parallel to a axis.
Hydrogen bonds are shown as dotted lines
123

Synthesis, characterization, X-ray diffraction studies…
Hirshfeld surface analysis of 1 and 2
Hirshfeld surface analysis is an effective tool for exploring packing modes and
intermolecular interactions in molecular crystals [15–18], as they provide a visual
picture of intermolecular interactions and of molecular shapes in a crystalline
environment. Surface features characteristic of different types of intermolecular
interactions can be identified, and these features can be revealed by colour coding
distances from the surface to the nearest atom exterior (de plots) or interior (di plots)
to the surface. This gives a visual picture of different types of interactions present,
and also reflect their relative contributions from molecule to molecule. Further, 2-D
fingerprint plots (FP), in particular the breakdown of FP into specific atom…atom
contacts in a crystal, provide a quantitative idea of the types of intermolecular
contacts experienced by molecules in the bulk and presents this information in a
convenient colour plot. Hirshfeld surfaces comprising dnorm surface plots and FP
were generated and analysed for the crystals of 1 and 2 in order to explore their
packing modes and intermolecular interactions, and to demonstrate the efficiency of
this tool in demonstrating the crystal structures of the two compounds.
The dnorm surfaces of molecules 1 and 2 are shown in Fig. 15, and the FP for the
overall contacts and individual atom…atom contacts in 1 and 2 are shown in
Fig. 16.
The dark-red spots on the dnorm surface arise as a result of the short interatomic
contacts, i.e., strong hydrogen bonds, while the intermolecular interactions appear as
light-red spots. The analysis of the dnorm surface and FP gives a pictorial
conformation (both qualitatively as well as quantitatively) to the nature and
Fig. 14 Display of two dimensional architecture in 2. Hydrogen bonds are shown as dotted lines
123

B. Kulkarni et al.
Fig. 15 dnorm mapped on a Hirshfeld surface for visualizing the intermolecular contacts in 1 and 2
geometries of the hydrogen bonds and intermolecular interactions described in the
crystal structures of both 1 and 2.
The dnorm and FP of 1 are consistent with the observed intermolecular
interactions. The light red spots on the dnorm surface of 1 located near the O1 and
O3 carbonyl oxygen atoms, and close to the H7B and H10B atoms (Fig. 15) can be
attributed to the H10B…O1 and H7B…O3 contacts displayed in 1. The analysis of
the FP of 1 (Fig. 16-Left) shows that the major contribution to the Hirshfeld surface
is from H…H contact, with a 75.4 % contribution (Fig. 16), the closest H…H
˚
contact occurs at di ? de = 2.2 A. The second largest contribution comes from the
˚
O…H/H…O contacts (19.7 %), the closest one observed at di ? de = 2.35 A,
which is slightly less than the interatomic distance of 2.44 A (Table 4) observed for
˚
the H10B…O1 contact in the crystal structure of 1.
The two red spots in the dnorm surface of 2 near the apex of the pyramidal
geometry around the N4 atom (Fig. 15) gives conclusive evidence for the
participation of the lone pair of electrons of N4 atom as bifurcated hydrogen bond
acceptors in the crystal structure of 2. Similarly, the two red spots around the
carbonyl oxygen atom O3 is in agreement with its participation as bifurcated
acceptors in hydrogen bond formation in 2. Further, the participation of the hydrogen
atoms of –NH2, –NH and –CH2 groups in hydrogen bonding can be identified in the
form of red spots around the respective H-atoms in the dnorm surfaces (Fig. 15). Also,
it can be seen that there are no red spots around the carbonyl oxygen of the ester
group, and, thus, is not involved in any intermolecular interactions. The FP of 2 is
shown in Fig. 16 (Right). The analysis of FP of individual atom…atom contacts was
made. The major contribution to the dnorm surface of 2 is from H…H contact
˚
(71.9 %) with the shortest distance at di ? de = 2.2 A, similar to that observed for 1.
The O…H/H…O contacts contributes 18.2 %, with the shortest distance occurring at
123

Synthesis, characterization, X-ray diffraction studies…
Fig. 16 Fingerprint plots of 1 and 2 for the overall contacts (Top) and for individual atom…atom contacts
123

B. Kulkarni et al.
Table 5 Antimicrobial activity data of the compounds 1 and 2
Compound
Zone of inhibition/mm
Antibacterial activity
Antifungal activity
S. aureus
B. subtilis
E. coli
P. aeruginosa
C. albicans
A. niger
1
11
15
28
–
10
16
27
–
13
14
26
–
15
17
24
–
10
16
–
14
20
–
2
Ciprofloxacin
Flucanazole
27
26
˚
di ? de = 2.1 A. This is in close agreement with the contact distances of 2.17 A and
˚
˚
2.21 A observed for N4-H1N4…O3 and N4-H2N4…O3 hydrogen bonds (Table 4)
in the crystal structure of 2. The H…N/N…H contacts contribute 6.5 % to the
Hirshfeld surface, with shortest contact distance appearing at di ? de = 2.3 A. This
˚
distance is almost same as that for the intermolecular interaction N3-HN3…N4
observed at 2.35 A in the crystal structure of 2. Thus, the Hirshfeld surface and FP
˚
analysis of both 1 and 2 give conclusive evidence, both qualitatively and
quantitatively, of the various hydrogen bonds/intermolecular interactions displayed
in the crystal structure of 1 and 2.
Biological studies
Both the compounds were screened in vitro at a concentration of 10 lg/disc for their
antibacterial activity against two Gram-positive strains (Staphylococcus aureus and
Bacillus subtilis) and two Gram-negative strains (Escherichia coli and Pseu-
domonas aeruginosa). Furthermore, antifungal evaluation was also carried out
against Candida albicans and Aspergillus niger at a concentration of 10 lg/disc.
Standard antibacterial drug ciprofloxacin (10 lg/disc) and antifungal drug flucona-
zole (10 lg/disc) were also tested under similar conditions against these organisms.
Each experiments were done in triplicate and the average readings were taken. Both
1 and 2 exhibited moderate antibacterial and antifungal activities. The molecules
were classified as highly active (if the zone of inhibition C26 mm), moderately
active (if the zone of inhibition = 11–25 mm), and least active (if the zone of
inhibition \11 mm), based on their biological activities. Comparison of the results
for 1 and 2 with the standards showed that both the compounds 1 and 2 exhibited
moderate antibacterial as well as antifungal activities. The results of antibacterial
and antifungal activities of compounds 1, 2 and the standards are expressed in terms
of zone of inhibition in Table 5.
Conclusion
Two derivatives of N-Boc piperazine, an ester derivative, i.e., tert-butyl 4-(2-
ethoxy-2-oxoethyl)-piperazine-1-carboxylate (1), and, a hydrazide derivative tert-
butyl 4-(2-hydrazino-2-oxoethyl)piperazine-1-carboxylate (2) were synthesized and
123

Synthesis, characterization, X-ray diffraction studies…
were characterized by FT-IR, 1H & 13C NMR and LCMS spectroscopic studies and
single crystal XRD data. The molecule structures of 1 and 2 are quite different: 1
being a linear-shaped molecule and the molecule of 2 is L-shaped. The crystal
structures of 1 and 2 prefer different packing patterns. The crystal structure of 1
features C–H…O intermolecular interactions, while that of 2 features strong N–
H…O hydrogen bonds and intermolecular interactions of the type N–H…N and C–
H…N, which were further confirmed by analysis of the Hirshfeld surfaces and the
fingerprint plots. Both 1 and 2 displayed moderate antibacterial and antifungal
activities against several microorganisms.
Supplementary materials
CCDC 1435775 & 1435776 contains the supplementary crystallographic data for
this paper. These data can be obtained free of charge from the Cambridge
Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax:
(?44) 1223 336 033; or e-mail: deposit@ccdc.cam.ac.uk.
Acknowledgments The authors are thankful to the Institution of Excellence, Vijnana Bhavana,
University of Mysore, Mysuru, for providing the single-crystal X-ray diffraction facility. The authors are
also thankful to Sapala Organics Pvt. Ltd. Hyderabad, India for recording the FT-IR, 1H & 13C NMR and
LCMS spectra. BK, TS, PH, and KM are thankful to the Management and to Dr. S. G. Gopala Krishna,
Principal, Nagarjuna college of Engg. & Tech., Devanahalli, Bangalore, for providing laboratory
facilities.
Compliance with ethical standards
Conflict of interest The authors declare that they have no conflict of interest.
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