Determination of 1-aryl-4-propylpiperazine pKa values: The substituent on aryl modulates basicity

Article abstract of DOI:10.1016/j.bmc.2008.12.015

In order to design a potential drug, it is important to know its pK_a because the protonation state of the molecule will be critical for ligand-receptor interaction and for the pharmacokinetic of the molecule. pK_a values of a series o

Full text of DOI:10.1016/j.bmc.2008.12.015

Bioorganic & Medicinal Chemistry 17 (2009) 1339–1344
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Determination of 1-aryl-4-propylpiperazine pKa values: The substituent
on aryl modulates basicity
*
Enza Lacivita , Marcello Leopoldo, Paola De Giorgio, Francesco Berardi, Roberto Perrone
Università degli Studi di Bari, Dipartimento Farmaco-Chimico, via Orabona, 4, 70125 Bari, Italy
a r t i c l e i n f o
a b s t r a c t
Article history:
In order to design a potential drug, it is important to know its pKa because the protonation state of the
molecule will be critical for ligand–receptor interaction and for the pharmacokinetic of the molecule.
pKa values of a series of 1-(substitutedphenyl)-4-propylpiperazines were measured to study how the
presence of a substituent on the phenyl ring modulates the basicity of N-4 nitrogen. pKa values indicated
that the position of the substituent was crucial. In general, the introduction of the substituent in ortho-
position of the phenyl ring increased the basicity of the molecule. This effect appeared to be related to
steric and conformational effects and not to the electronic properties of the substituent. On the other
hand, meta- and para-substituted derivatives showed a slight decrease of pKa that was qualitatively con-
sistent with the electronic properties of the substituent.
Received 1 August 2008
Revised 13 November 2008
Accepted 8 December 2008
Available online 24 December 2008
Keywords:
1-Arylpiperazine
Basicity
pKa
Ó 2008 Elsevier Ltd. All rights reserved.
Potentiometric titration
1. Introduction
1-Arylpiperazines represent one of the most versatile template
for obtaining molecules acting at several receptor system such as
It is been estimated that 60–70% of drugs are ionizable mole-
cules and, thus, one of the most important physicochemical prop-
erties of a drug molecule is the acid dissociation constant (pKa).
Several key physicochemical properties that regulates absorption
and distribution processes, such as lipophilicity and solubility,
are pKa dependent.1 The ionization state is a key parameter not
only in ADME profiling, but also when the drug interacts with
the biological target because the interaction occurs in aqueous
environment at physiological pH. Moreover, pKa can be important
in determining the rate and the site of drug metabolism.2 Finally, in
drug formulation the ionization constant is important for choosing
the correct excipients and counterions.3 Therefore, pKa should be
accurately evaluated during drug optimization.
Ionization constant can be measured by several methods,
including potentiometric and UV–vis spectrophotometric titrations
which are frequently used. These methods require a lot of skills for
both monitoring the ionization state of the molecule while chang-
ing pH and interpreting the experimental data taking full account
of chemical equilibria theory, sample solubility, and overlapping
pKa values.1 This explains the paucity of experimental data on
the ionization constant of drug-like molecules. Several pKa predic-
tion models have been proposed, based on different descriptors,
including atomic charges, topological distances, chemical reactiv-
ity models, and group philicity. However, the predicted values
sometimes could be much off and can only be used as estimates.4
serotoninergic, adrenergic, and dopaminergic receptors.5–7 A num-
ber of studies have described the synthesis, structure-activity rela-
tionships, and pharmacological evaluation of various classes of
drugs with an arylpiperazine moiety in their structure. These re-
search efforts have led to the identification of several arylpiper-
azine derivatives that reached late stage clinical trials for the
treatment of depression, psychosis, and anxiety (Fig. 1). The specif-
ity of arylpiperazine-based compounds for a given receptor system
and the pharmacokinetic properties can be modulated through: (i)
appropriate choice of the substituent on the phenyl ring linked to
the piperazine; (ii) optimization of the intermediate alkyl chain
length; (iii) selection of the different terminal fragment. During
the last decade, our research group have studied several classes
of 1-arylpiperazine derivatives with specificity for serotonin 5-
HT1A and 5-HT7 receptors and for dopamine D3 and D4 recep-
tors.8–11 Although 1-arylpiperazine-based compounds have been
studied in depth for their pharmacological properties, only a few
papers dealt with ionization constant. Caccia et al. determined
the ionization constants of several unsubstituted 1-arylpiperazines
which may be formed during the in vivo biotransformation of cen-
tral nervous system acting drugs.12 Van Steen et al. determined the
pKa values of a series of N-4 alkylsubstituted heterobicyclic phe-
nylpiperazines in order to evaluate if there was a correlation
between the ionization state of the ligands and their affinities at
5-HT1A receptors.13 Bojarski et al. studied the effect of N-alkylation
on the basicity of several cyclic amines, including some arylpiper-
azines, that are common fragments of several adrenergic, seroto-
ninergic and dopaminergic ligands.14
* Corresponding author. Tel.: +39 080 5442750; fax: +39 080 5442231.
E-mail address: lacivita@farmchim.uniba.it (E. Lacivita).
0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.12.015

1340
E. Lacivita et al. / Bioorg. Med. Chem. 17 (2009) 1339–1344
O
N
O
O
N
N
N
O
N
N
N
N
N
N
Cl
buspirone
nefazodone
O
H
N
O
O
N
Cl
N
N
N
Cl
S
NH
N
Cl
ziprasidone
aripiprazole
Figure 1. Structures of arylpiperazines entered in clinical trials.
Recently, we have determined by using a potentiometric meth-
od the physicochemical properties of a series of dopamine D3 li-
gands with N-[4-(4-arylpiperazin-1-yl)butyl]arylcarboxamide
structure. During that study, we observed that the ionization con-
stant of the piperazine nitrogen linked to the butyl chain was influ-
enced by the nature and the position of the substituent on the
phenyl attached to the piperazine.15 Starting from this observation,
we prepared a series of 1-(substitutedphenyl)-4-propylpiperazines
in order to study how the nature and position of the substituent on
the phenyl ring can modulate the basicity of the N-propyl-substi-
tuted nitrogen. For this purpose, we selected substituents covering
a wide range of electronic properties (CH3, OCH3, Cl, OH, CF3, CN,
NO2, CONH2). The N-propyl-1-arylpiperazine scaffold was chosen
because a large number of biologically active 1-arylpiperazine
derivatives bears an alkyl chain of variable length on the nitrogen.
Finally, to the best of our knowledge, this aspect has not been doc-
umented in the literature.
sponding cyano derivatives 16a–c. The 1-arylpiperazines were ob-
tained from commercial sources or were prepared according to the
literature as detailed in Experimental Section.
3. Results and discussion
The experimental and the calculated pKa values of compounds
10a–c–17a–c are reported in Table 1. To evaluate if variations in
pKa values were related to Hammet r-parameters of the substitu-
ents, the unsubstituted 4-phenyl-1-propylpiperazine 9 was pre-
pared. pKa data of ortho- (10a–17a) and meta-substituted (10b–
17b) derivatives appeared to be not in correlation with the elec-
tronic properties of the substituents. The introduction of a substi-
tuent in ortho-position determined an increase of pKa value of the
N-4 piperazine nitrogen with exception of derivatives 15a
(R = NO2) and 16a (R = CN), that showed pKa values close to that
of the unsubstituted 9. Moreover, the basicity-increasing effect
was not related to the electronic properties of the substituent. In
fact, 10a (R = CH3) and 14a (R = CF3) showed similar pKa values de-
spite the very different inductive effect of the substituents. On the
other hand, the introduction of OCH3 or OH (compounds 11a and
12a, respectively), which share nearly the same inductive effect,
determined different variation in pKa value (pKa = 8.29 and 8.05,
respectively). These data suggested that other factors, such as ste-
ric or conformational effects, are able to modulate the basicity of
the N-4 nitrogen of the piperazine in a larger extent than electronic
2. Chemistry
The target compounds 9, 10a–c–16a–c were synthesized by
reductive alkylation of the corresponding 1-arylpiperazines as de-
picted in Scheme 1. Propionaldehyde reacted with the appropriate
1-arylpiperazine to give the intermediate enamine which was re-
duced with NaBH4 to give the target compound. The carboxamide
derivatives 17a–c were prepared by acidic hydrolysis of the corre-
O
A
+
N
NH
N
N
N
N
H
R
R
R
a = ortho-; b = meta-; c = para-substituted
1R= H
9: R= H
2a-c: R = CH3
3a-c: R = OCH3
4a-c: R = OH
5a-c: R = Cl
6a-c: R = CF3
7a-c: R = NO2
8a-c: R = CN
10a-c: R = CH3
11a-c: R = OCH3
12a-c: R = OH
13a-c: R = Cl
14a-c: R = CF3
15a-c: R = NO2
16a-c: R = CN
17a-c: R = CONH2
B
Scheme 1. Reagents and conditions: (A) NaBH4, rt; (B) H2SO4, 70 °C.

E. Lacivita et al. / Bioorg. Med. Chem. 17 (2009) 1339–1344
1341
Table 1
the vicinity of the molecule and changes the prototropic proper-
ties of the medium in the local microenvironment of the mole-
cule. This translates into a decrease of the pKa for the aniline
nitrogen which depends on the thermodynamic of the solvation
process and not on the electronic properties of the substituent.
Considering that ortho-substituted phenylpiperazines 10a–17a
are structurally related to ortho-substituted anilines, it can be
speculated that the ‘hindrance effect’ modulates the electron
withdrawing effect of the ‘anilinic’ nitrogen of the piperazine
Calculated and experimental pKa values of compounds 9, 10a–c–17a–c
N
N
R
a
Cpd
R
Calculated pKa
Experimental pKa
D
pKa
which transmitted via
r-pathways to the N-4 nitrogen, thus
9
H
7.14
7.15
7.23
7.19
7.16
7.16
7.16
7.08
7.14
7.23
7.03
7.06
7.08
7.03
7.08
7.00
6.96
6.97
6.91
6.99
7.00
7.03
7.07
7.07
7.04
7.96 0.02
8.39 0.03
7.83 0.01
7.92 0.02
8.29 0.03
7.86 0.03
7.88 0.02
8.05 0.03
7.82 0.06
7.92 0.05
8.07 0.02
7.83 0.01
7.83 0.02
8.80 0.01
7.81 0.01
7.84 0.01
7.94 0.01
7.70 0.04
7.59 0.03
7.92 0.01
7.67 0.06
7.78 0.01
8.14 0.03
7.76 0.04
7.84 0.02
—
determining a basicity increase. Moreover, a conformational ef-
fect of the ortho-substituent reflecting in pKa variation cannot
be ruled out. From a survey of crystal structures in Cambridge
Structural Database it emerges that N-phenylpiperazines adopt
different conformations depending on the presence of a substitu-
ent on the phenyl ring. In particular, unsubstituted derivatives
10a
10b
10c
11a
11b
11c
12a
12b
12c
13a
13b
13c
14a
14b
14c
15a
15b
15c
16a
16b
16c
17a
17b
17c
2-CH3
3-CH3
4-CH3
2-OCH3
3-OCH3
4-OCH3
2-OH
3-OH
4-OH
2-Cl
3-Cl
+0.43
À0.13
À0.04
+0.33
À0.10
À0.08
+0.09
À0.14
À0.04
+0.11
À0.13
À0.13
+0.84
À0.15
À0.12
À0.02
À0.26
À0.37
À0.04
À0.29
À0.18
+0.18
À0.20
À0.12
can adopt a
p-conjugated arrangement with the N-lone pair axis
of the ‘anilinic’ nitrogen almost perpendicular to the adjacent
aryl p-orbitals. The introduction of an ortho-substituent on the
phenyl ring twists the N-lone pair axis by 45–75° from the plane
4-Cl
of p-system, causing the uncoupling of p-conjugation. This twist-
2-CF3
3-CF3
4-CF3
2-NO2
3-NO2
4-NO2
2ÀCN
3-CN
ing effect is less pronounced when the substituent is in meta-po-
sition, whereas it is nearly absent for para-substituted
derivatives.
All the above considerations might explain the significant dif-
ference between experimental and calculated pKa values of com-
pounds 9, 10a–c–17a–c, because currently available pKa
prediction tools operate considering only the electronic properties
of a substituent and its topological distance from reaction centre.
4-CN
2-CONH2
3-CONH2
4-CONH2
4. Conclusion
a
Calculated with ACD/Labs 7.0 (Advanced Chemistry Development, Inc., Toronto
ON, Canada).
A series of 1-(substitutedphenyl)-4-propylpiperazines was pre-
pared in order to study the effect of the substituent on the phenyl
ring on the basicity of the alkylated nitrogen. The pKa values indi-
cated that the position of the substituent was crucial for the mod-
ulation of this physicochemical parameter. In general, the
introduction of the substituent in ortho-position of the phenyl ring
increased the basicity of the molecule. This effect appeared to be
related to steric and conformational effects caused by the introduc-
tion of the substituent and not to its electronic properties. On the
other hand, the presence of a substituent in meta- and para-posi-
tion determined a slight decrease of pKa that was qualitatively con-
sistent with the electronic properties of the substituent. Clearly,
the increasing-basicity effect of the ortho-substituent may reflect
on lipophilicity and solubility of this class of compounds. Finally,
calculated pKa values were significantly different from the experi-
mental ones. This aspect is not surprising because computational
pKa prediction tools are not yet sufficiently sophisticated to be of
general practical value in pharmaceutical industry. However, this
situation is expected to change as more experimental data become
available and continued effort are spent to refine existing tools.
effects. The introduction of the substituent in meta-position of the
phenyl ring linked to the piperazine had an overall basicity-lower-
ing effect. Also in this case, this effect seemed to be not related to
the electronic properties of the substituent. In fact, all compounds
showed very similar pKa values, being 16b (R = CN) the less basic
compound among the meta-substituted derivatives. The introduc-
tion of the substituent in para position determined a slight de-
crease of the pKa value as compared to that of 9. This effect was
qualitatively consistent with the Hammet
substituents. In particular, derivative 15c (R = NO2) showed the
more marked decrease in pKa as compared to that of
pKa = À0.37) due to strong inductive and mesomeric effect of ni-
tro group in para-position.
r-parameters of the
9
(D
Taken together, these data clearly indicated that the elec-
tronic properties of the substituent on the phenyl ring are not
the only feature that modulates the basicity of the N-4 nitrogen
of the piperazine. It is interesting to note that the trend ob-
served in the variation of pKa in this serie of compounds paral-
leled the findings reported on the basicity of substituted
anilines.16–18 Those studies evidenced good correlation between
pKa and electronic properties of the substituent only among
para-substituted anilines, whereas no correlation was found
among the ortho-substituted isomers. Such results were rational-
ized considering that the introduction of a substituent in ortho-
position increases the hydrophobicity in the region adjacent to
the nitrogen atom, hindering the translational motion of water
molecules. As a result, the space in which the substituent and
the nitrogen are enclosed becomes inaccessible for water mole-
cules. This creates the so-called ‘hindrance effect’ and leads to
the formation of a layer of more structurized water around the
species. Hydrophobic hydration decreases medium polarity in
5. Experimental
5.1. Chemistry
Column chromatography was performed with 1:30 Merck silica
gel 60A (63–200 lm) as the stationary phase. Melting points were
determined in open capillaries on a Gallenkamp electrothermal
apparatus. Elemental analyses (C, H, N) were performed on Euro-
vector Euro EA 3000 analyzer; the analytical results were within
0.4% of the theoretical values for the formula given. 1H NMR spec-
tra were recorded at 300 MHz on a Varian Mercury-VX spectrom-
eter. All spectra were recorded on free bases. All chemical shift

1342
E. Lacivita et al. / Bioorg. Med. Chem. 17 (2009) 1339–1344
values are reported in ppm (d). Recording of mass spectra was done
on an HP6890-5973 MSD gas chromatograph/mass spectrometer;
only significant m/z peaks, with their percentage of relative inten-
sity in parentheses, are reported. The purity of new compounds
that were essential to the conclusions drawn in the text was deter-
mined by HPLC on a Perkin–Elmer series 200 LC instrument using a
5.8. 1-(3-Methoxyphenyl)-4-propylpiperazine (11b)
GC–MS m/z 235 (M++1, 15), 234 (M+, 92), 205 (100), 162 (28). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.49–1.62 (m, 2H), 2.36 (app t, 2H),
2.60 (app t, 4H), 3.21 (app t, 4H), 3.79 (s, 3H), 6.39–6.43 (m, 1H),
6.46–6.47 (m, 1H), 6.55 (dd, 1H, J = 2.2, 8.2 Hz), 7.16 (t, 1H,
J = 8.2 Hz). Anal. (C14H22N2OÁ2HCl) C, H, N.
Phenomenex Gemini RP-18 column, (250 Â 4.6 mm, 5
lm particle
size) and equipped with a Perkin–Elmer 785A UV–vis detector set-
ting k = 254 nm. Compounds were eluted with CH3OH/H2O/Et3N,
4:1:0.01, v/v at a flow rate of 0.8 mL/min. A standard procedure
was used to transform final compounds into their hydrochloride
salts. The following compounds were synthesized according to
published procedures: 1-(2-trifluoromethylphenyl)piperazine
5.9. 1-(4-Methoxyphenyl)-4-propylpiperazine (11c)
GC–MS m/z 235 (M++1, 17), 234 (M+, 100), 205 (78), 135 (25). 1H
NMR: d 0.92 (t, 3H, J = 7.4 Hz), 1.51–1.68 (m, 2H), 2.33–2.38 (m,
2H), 2.61 (app t, 4H), 3.11 (app t, 4H), 3.76 (s, 3H), 6.81–6.85 (m,
2H), 6.88–6.92 (m, 2H). Anal. (C14H22N2OÁ2HCl) C, H, N.
(6a),19
phenyl)piperazine (8b).21
1-(3-nitrophenyl)piperazine
(7b),20
1-(3-cyano-
5.10. 2-(4-Propyl-1-piperazinyl)phenol (12a)
5.2. General procedure for the preparation of compounds 9,
10a,c–16a,c
GC–MS m/z 221 (M++1, 11), 220 (M+, 70), 191 (94), 148 (64), 134
(100), 120 (99). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.50–1.62 (m,
2H), 2.39 (app. t, 2H), 2.64 (br s, 4H), 2.92 (app. t, 4H), 5.0 (br s,
1H, D2O exchanged), 6.85 (dt, 1H, J = 1.4, 7.7 Hz), 6.94 (dd, 1H,
J = 1.4, 7.9 Hz), 7.04–7.10 (m, 1H), 7.18 (dd, 1H, J = 1.4, 7.7 Hz).
Anal. (C13H20N2OÁ2HClÁH2O) C, H, N.
To a solution of the appropriate 1-arylpiperazine (3.0 mmol) in
MeOH (20 mL) propionaldehyde (3.6 mmol) was added dropwise
and the mixture was stirred at rt for 1 h. After cooling at 0 °C,
NaBH4 (4.5 mmol) was added in small portions. The mixture was
warmed at room temperature, stirred for 2 h and, then, quenched
with H2O. MeOH was removed under reduced pressure and the
aqueous solution was extracted with CH2Cl2 (3 Â 20 mL). The or-
ganic phases were collected, washed with brine, dried over Na2SO4
and concentrated in vacuo. The crude residue was purified by chro-
matography (CHCl3/AcOEt, 1:1 as eluent) to give pure compounds
as oils in 40–50% yield.
5.11. 3-(4-Propyl-1-piperazinyl)phenol (12b)
GC–MS m/z 221 (M++1, 14), 220 (M+, 88), 191 (100), 148 (28). 1H
NMR: d 0.92 (t, 3H, J = 7.4 Hz), 1.50–1.63 (m, 2H), 2.35–2.40 (m,
2H), 2.61 (app. t, 4H), 3.19 (app. t, 4H), 4.85 (br s, 1H, D2O ex-
changed), 6.28–6.31 (m, 1H), 6.36–6.38 (m, 1H), 6.47–6.50 (m,
1H), 7.09 (t, 1H, J = 8.0 Hz). Anal. (C13H20N2OÁ2HClÁ0.3H2O) C, H, N.
5.3. 1-Phenyl-4-propylpiperazine (9)
5.12. 4-(4-Propyl-1-piperazinyl)phenol (12c)
GC–MS m/z 205 (M++1, 11), 204 (M+, 74), 175 (100), 132 (26). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.49–1.60 (m, 2H), 2.33–2.39 (m,
2H), 2.61 (app. t, 4H), 3.21 (app. t, 4H), 6.81–6.88 (m, 1H), 6.91–
6.95 (m, 2H), 7.24–7.30 (m, 2H). Anal. (C13H20N2Á2HClÁ0.5H2O) C,
H, N.
GC–MS m/z 221 (M++1, 16), 220 (M+, 100), 191 (89), 120 (31). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.52–1.62 (m, 3H, 1H D2O ex-
changed), 2.40 (app. t, 2H), 2.65 (br s, 4H), 3.12 (app. t, 4H),
6.74–6.80 (m, 2H), 6.82–6.91 (m, 2H). Anal. (C13H20N2O) C, H, N.
5.4. 1-(2-Methylphenyl)-4-propylpiperazine (10a)
5.13. 1-(2-Chlorophenyl)-4-propylpiperazine (13a)
GC–MS m/z 219 (M++1, 9), 218 (M+, 63), 189 (100), 118 (43).1H
NMR: d 0.94 (t, 3H, J = 7.4 Hz), 1.51–1.61 (m, 2H), 2.30 (s, 3H), 2.39
(app t, 2H), 2.62 (br s, 4H), 2.96 (app. t, 4H), 6.95–7.05 (m, 2H),
7.14–7.19 (m, 2H). Anal. (C13H20N2Á2HCl) C, H, N.
GC–MS m/z 240 (M++2, 11), 238 (M+, 33), 211 (33), 209 (100),
166 (16). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.49–1.62 (m, 2H),
2.35–2.41 (m, 2H), 2.64 (br s, 4H), 3.09 (app. t, 4H), 6.93–6.99
(m, 1H), 7.05 (dd, 1H, J = 1.7, 8.0 Hz), 7.19 (m, 1H), 7.35 (dd, 1H,
J = 1.7, 8.0 Hz). Anal. (C13H19ClN2ÁHCl) C, H, N.
5.5. 1-(3-Methylphenyl)-4-propylpiperazine (10b)
5.14. 1-(3-Chlorophenyl)-4-propylpiperazine (13b)
GC–MS m/z 219 (M++1, 14), 218 (M+, 88), 189 (100), 70 (44).1H
NMR: d 0.94 (t, 3H, J = 7.4 Hz), 1.49–1.62 (m, 2H), 2.32 (s, 3H),
2.33–2.38 (m, 2H), 2.60 (app. t, 4H), 3.20 (app. t, 4H), 6.67–6.76
(m, 3H), 7.12–7.18 (m, 1H). Anal. (C13H20N2Á2HCl) C, H, N.
GC–MS m/z 240 (M++2, 16), 238 (M+, 49), 211 (33), 209 (100),
166 (21). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.61 (m, 2H),
2.32–2.38 (m, 2H), 2.58 (app. t, 4H), 3.20 (app. t, 4H), 6.76–6.81
(m, 2H), 6.86–6.88 (m, 1H), 7.15 (t, 1H, J = 8.0 Hz). Anal.
(C13H19ClN2ÁHCl) C, H, N.
5.6. 1-(4-Methylphenyl)-4-propylpiperazine (10c)
GC–MS m/z 219 (M++1, 15), 218 (M+, 90), 189 (100), 70 (44).1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.61 (m, 2H), 2.30 (s, 3H), 2.36
(app t, 2H), 2.61 (app t, 4H), 3.16 (app. t, 4H), 6.85 (d, 2H,
J = 8.5 Hz), 7.07 (d, 2H, J = 8.8 Hz). Anal. (C13H20N2Á2HCl) C, H, N.
5.15. 1-(4-Chlorophenyl)-4-propylpiperazine (13c)
GC–MS m/z 240 (M++2, 18), 238 (M+, 54), 211 (34), 209 (100),
166 (21). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.61 (m, 2H),
2.33–2.38 (m, 2H), 2.59 (app. t, 4H), 3.17 (app. t, 4H), 6.81–6.86
(m, 2H), 7.17–7.19 (m, 2H). Anal. (C13H19ClN2Á2HCl) C, H, N.
5.7. 1-(2-Methoxyphenyl)-4-propylpiperazine (11a)
GC–MS m/z 235 (M++1, 14), 234 (M+, 83), 205 (100), 190 (36),
177 (23). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.50–1.62 (m, 2H),
2.38 (app t, 2H), 2.66 (br s, 4H), 3.11 (br s, 4H), 3.86 (s, 3H),
6.84–7.02 (m, 4H). Anal. (C14H22N2OÁ2HCl) C, H, N.
5.16. 1-(2-Trifluoromethylphenyl)-4-propylpiperazine (14a)
GC–MS m/z 273 (M++1, 6), 272 (M+, 20), 243 (100), 172 (12), 70
(16). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.53–1.60 (m, 2H), 2.39 (app

E. Lacivita et al. / Bioorg. Med. Chem. 17 (2009) 1339–1344
1343
t, 2H), 2.61 (br s, 4H), 2.98 (app. t, 4H), 7.21 (t, 1H, J = 7.7 Hz), 7.38
(d, 1H, J = 7.9 Hz), 7.48–7.53 (m, 1H), 7.61 (dd, 1H, J = 1.1, 7.4 Hz).
Anal. (C14H19F3N2ÁHCl) C, H, N.
5.25. General procedure for the preparation of compounds
17a–c
Compounds 16a–c (2 mmol) were slowly added to cooled concd
H2SO4 (10 mL). The mixture was stirred at 60 °C for 3 h, then, after
cooling, was poured on ice. The aqueous solution was basified with
concd NH4OH and extracted with AcOEt (3 Â 20 mL). The organic
phases were collected, washed with brine, dried over Na2SO4 and
concentrated under reduced pressure. The crude residue was puri-
fied by chromatography (CHCl3/MeOH, 19:1 as eluent) to give pure
compounds as white semisolids in 50% yield.
5.17. 1-(3-Trifluoromethylphenyl)-4-propylpiperazine (14b)
GC–MS m/z 273 (M++1, 6), 272 (M+, 36), 243 (100), 200 (23), 172
(22). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.51–1.61 (m, 2H), 2.33–
2.38 (m, 2H), 2.59 (app t, 4H), 3.29 (app. t, 4H), 6.91–6.93 (m,
1H), 7.10–7.12 (m, 2H), 7.27–7.29 (m, 1H). Anal.
(C14H19F3N2ÁHClÁ0.5H2O) C, H, N.
5.18. 1-(4-Trifluoromethylphenyl)-4-propylpiperazine (14c)
5.26. 1-(2-Carboxamidophenyl)-4-propylpiperazine (17a)
GC–MS m/z 273 (M++1, 6), 272 (M+, 36), 243 (100), 200 (22),
172 (20). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.52–1.61 (m, 2H),
2.35 (app t, 2H), 2.59 (br s, 4H), 3.29 (app. t, 4H), 6.92 (d, 2H,
J = 9.0 Hz), 7.46 (d, 2H, J = 9.0 Hz). Anal. (C14H19F3N2ÁHClÁ0.5H2O)
C, H, N.
GC–MS m/z 248 (M++1, 2), 247 (M+, 4), 162 (100), 158 (68), 132
(68), 98 (39). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.60 (m, 2H),
2.34–2.39 (m, 2H), 2.62 (br s, 4H), 3.05 (app. t, 4H), 5.88 (br s, 1H,
D2O exchanged), 7.18–7.24 (m, 2H), 7.43–7.48 (m, 1H), 8.14–8.17
(m,
1H)
9.55
(br
s,
1H,
D2O
exchanged).
Anal.
(C14H21N3OÁHClÁ0.5H2O) C, H, N.
5.19. 1-(2-Nitrophenyl)-4-propylpiperazine (15a)
5.27. 1-(3-Carboxamidophenyl)-4-propylpiperazine (17b)
GC–MS m/z 250 (M++1, 6), 249 (M+, 31), 220 (87), 202 (57),
131 (100), 119 (72). 1H NMR: d 0.92 (t, 3H, J = 7.4 Hz), 1.47–
1.59 (m, 2H), 2.34–2.39 (m, 2H), 2.60 (app t, 4H), 3.08 (app. t,
4H), 6.99–7.08 (m, 1H), 7.14 (dd, 1H, J = 1.1, 8.3 Hz) 7.43–7.49
(m, 1H) 7.74 (dd, 1H, J = 1.4, 8.1 Hz). Anal. (C13H19N3O2ÁHCl) C,
H, N.
GC–MS m/z 248 (M++1, 8), 247 (M+, 46), 218 (100), 175 (17). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.49–1.61 (m, 2H), 2.33–2.38 (m,
2H), 2.60 (app t, 4H), 3.27 (app. t, 4H), 5.64 (br s, 1H, D2O ex-
changed), 6.05 (br s, 1H, D2O exchanged), 7.04–7.08 (m, 1H),
7.14–7.16 (m, 1H), 7.30 (t, 1H, J = 7.9 Hz), 7.43–7.44 (m, 1H). Anal.
(C14H21N3OÁ2HCl) C, H, N.
5.20. 1-(3-Nitrophenyl)-4-propylpiperazine (15b)
5.28. 1-(4-Carboxamidophenyl)-4-propylpiperazine (17c)
GC–MS m/z 250 (M++1, 5), 249 (M+, 29), 220 (100), 177 (14). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.49–1.62 (m, 2H), 2.34–2.39 (m,
2H), 2.61 (app t, 4H), 3.29 (app. t, 4H), 7.18 (dd, 1H, J = 2.4,
8.4 Hz), 7.36 (t, 1H, J = 8.0 Hz), 7.64 (dd, 1H, J = 2.2, 8.0 Hz) 7.70–
7.72 (m, 1H). Anal. (C13H19N3O2ÁHClÁ0.5H2O) C, H, N.
GC–MS m/z 248 (M++1, 8), 247 (M+, 47), 218 (100), 175 (18), 132
(20). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.49–1.61 (m, 2H), 2.33–
2.38 (m, 2H), 2.58 (app t, 4H), 3.30 (app. t, 4H), 5.65 (br s, 2H,
D2O exchanged), 6.87–6.91 (m, 2H), 7.69–7.74 (m, 2H). Anal.
(C14H21N3OÁHClÁ0.5H2O) C, H, N.
5.21. 1-(4-Nitrophenyl)-4-propylpiperazine (15c)
5.29. Determination of pKa by the pH-metric technique
GC–MS m/z 250 (M++1, 5), 249 (M+, 29), 220 (100), 177 (15). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.60 (m, 2H), 2.33–2.38 (m,
2H), 2.57 (app t, 4H), 3.43 (app. t, 4H), 6.78–6.84 (m, 2H), 8.08–
8.14 (m, 2H). Anal. (C13H19N3O2ÁHClÁ0.3H2O) C, H, N.
pKa values were determined by potentiometric titration using
the Sirius GLpKa (Sirius Analytical Instruments Ltd., Forest Row,
East Sussex, UK). All experiments were carried out at 25 0.5 °C
under a slow argon flow to avoid CO2 absorption at high pHs.
Details about instrument and procedures can be found else-
where.22–24 To measure pKa values, a right amount of each com-
pounds was dissolved in ionic strength adjusted water (0.15 M
5.22. 1-(2-Cyanophenyl)-4-propylpiperazine (16a)
GC–MS m/z 230 (M++1, 4), 229 (M+, 24), 200 (100), 157 (41). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.61 (m, 2H), 2.36–2.41 (m,
2H), 2.66 (app t, 4H), 3.26 (app. t, 4H), 6.96–7.02 (m, 2H), 7.44–
7.50 (m, 1H), 7.54–7.57 (m, 1H). Anal. (C14H19N3ÁHClÁ0.4H2O) C,
H, N.
KCl) to achieve final sample concentration in the range 5 Â 10À4
–
1Â10À4 M. The low aqueous solubility of the investigated com-
pounds required pKa measurements to be performed in the pres-
ence of methanol as cosolvent. Three separate 20 mL-
semiaqueous solutions, in 15–35% w/w of MeOH (50–30% w/w of
MeOH for compound 14a), were initially acidified with 0.5 M HCl
to pH 3.0. The solutions were then titrated with 0.5 M KOH to pH
11. The pH change per titrant addition was limited to 0.2 pH units
and pH value in each point was collected when the pH-drift was
lower than 0.002 pH per minute. Check of the pH-electrode with
measurement of voltage in Orion pH 7.00 buffer and electrode cal-
ibration (Four-PlusTM parameters) by a blank titration were every-
day performed. The initial estimates of the psKa values, which are
the apparent ionization constants in the mixed solvent, were ob-
tained by Bjerrum plots. These values were then refined by a
weighted non linear least-squares procedure (Refinement Pro 1.0
software) to create a multiset, where the refined values were
extrapolated to zero co-solvent concentration using the Yasuda–
Shedlovsky equation.25
5.23. 1-(3-Cyanophenyl)-4-propylpiperazine (16b)
GC–MS m/z 230 (M++1, 6), 229 (M+, 33), 200 (100), 157
(20). 1H NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.61 (m, 2H),
2.32–2.38 (m, 2H), 2.59 (app t, 4H), 3.23 (app. t, 4H), 7.06–
7.11 (m, 3H), 7.44–7.27–7.33 (m, 1H). Anal. (C14H19N3ÁHClÁ
0.4H2O) C, H, N.
5.24. 1-(4-Cyanophenyl)-4-propylpiperazine (16c)
GC–MS m/z 230 (M++1, 6), 229 (M+, 35), 200 (100), 157 (18). 1H
NMR: d 0.93 (t, 3H, J = 7.4 Hz), 1.48–1.68 (m, 2H), 2.35 (app t, 2H),
2.57 (app t, 4H), 3.33 (app. t, 4H), 6.82–6.87 (m, 2H), 7.46–7.51 (m,
2H). Anal. (C14H19N3ÁHClÁ0.4H2O) C, H, N.

1344
E. Lacivita et al. / Bioorg. Med. Chem. 17 (2009) 1339–1344
11. Perrone, R.; Berardi, F.; Colabufo, N. A.; Leopoldo, M.; Tortorella, V. J. Med. Chem.
Acknowledgement
2000, 43, 270.
12. Caccia, S.; Fong, M. H.; Urso, R. J. Pharm. Pharmacol. 1985, 37, 567.
13. Van Steen, B. J.; van Wijngaarden, I.; Tulp, M. T.; Soudijn, W. J. Med. Chem. 1993,
36, 2751.
14. Bojarski, A. J.; Mokroz, M. J.; Paluchowska, M. H. Pharmazie 1995, 50, 569.
15. Leopoldo, M.; Lacivita, E.; De Giorgio, P.; Contino, M.; Berardi, F.; Perrone, R.
Bioorg. Med. Chem. 2009, 17, 758.
The authors wish to thank Dr. Concetta Pacifico and Dr. Antonio
Carrieri (Universitá degli Studi di Bari) for supporting in Cambridge
Structural Database search.
16. Pankratov, A. N.; Uchaeva, I. M.; Doronin, S. Y.; Chernova, R. K. J. Struct. Chem.
2001, 42, 739.
References and notes
17. Adam, K. R. J. Phys. Chem. A 2002, 106, 11963.
18. Graña, A. M.; Hermida-Ramón, J. M.; Mosquera, R. A. Chem. Phys. Lett. 2005,
412, 106.
1. Avdeef, A. Curr. Top. Med. Chem. 2001, 1, 277.
2. Upthagrove, A. L.; Nelson, W. L. Drug Metab. Dispos. 2001, 29, 1377.
3. Stahl, P. H. In The Pratice of Medicinal Chemistry; Vermuth, C., Ed.; Academic
Press: London, 2003; p 601.
4. Morgenthaler, M.; Schweizer, E.; Hoffmann-Röder, A.; Benini, F.; Martin, R. E.;
Jaeschke, G.; Wagner, B.; Fischer, H.; Bendels, S.; Zimmerli, D.; Schneider, J.;
Diederich, F.; Kansy, M.; Müller, K. ChemMedChem 2007, 2, 1100.
5. Lopez-Rodriguez, M. L.; Ayala, D.; Benhamù, B.; Morbillo, M. J.; Viso, A. Curr.
Med. Chem. 2002, 9, 443.
19. Elworthy, T. R.; Ford, A. P. D. W.; Bantle, G. W.; Morgans, D. J.; Ozer, R. S.;
Palmer, W. S.; Repke, D. B.; Romero, M.; Sandoval, L.; Sjogren, E. B.; Talamas, F.
X.; Vazquez, A.; Wu, H.; Arredondo, N. F.; Blue, D. R.; De Sousa, A.; Gross, L. M.;
Kava, M. S.; Lesnick, J. D.; Vimont, R. L.; Williams, T. J.; Zhu, Q.; Pfister, J. R.;
Clarke, D. E. J. Med. Chem. 1997, 40, 2674–2687.
20. Becker, O. M.; Dhanoa, D. S.; Marantz, Y.; Chen, D.; Shacham, S.; Cheruku, S.;
Heifetz, A.; Mohanty, P.; Fichman, M.; Sharadendu, A.; Nudelman, R.;
Kauffman, M.; Noiman, S. J. Med. Chem. 2006, 49, 3116.
21. Koyama, M.; Kikuchi, C.; Ushiroda, O.; Ando, T.; Nagaso, H.; Fuji, K.; Okuno, M.;
Hiranuma, T. PTC Appl. WO 9800400, 1998; Chem. Abstr. 1998, 128, 114961.
22. Avdeef, A. Quant. Struct.-Act. Relat. 1992, 11, 510.
23. Avdeef, A. J. Pharm. Sci. 1993, 82, 183.
24. Avdeef, A.; Box, K. J.; Comer, J. E.; Gilges, M.; Hadley, M.; Hibbert, C.; Patterson,
W.; Tam, K. Y. J. Pharm. Biomed. Anal. 1999, 20, 631.
6. Rosini, M.; Bolognesi, M. L.; Giardinà, D.; Minarini, A.; Tumiatti, V.; Melchiorre,
C. Curr. Top. Med. Chem. 2007, 7, 147.
7. Soskic´, V.; Joksimovic´, J. Curr. Med. Chem. 1998, 5, 493.
8. Perrone, R.; Berardi, F.; Colabufo, N. A.; Leopoldo, M.; Lacivita, E.; Tortorella, V.;
Leonardi, A.; Poggesi, E.; Testa, R. J. Med. Chem. 2001, 44, 4431.
9. Leopoldo, M.; Lacivita, E.; Contino, M.; Colabufo, N. A.; Berardi, F.; Perrone, R. J.
Med. Chem. 2007, 50, 4214.
10. Leopoldo, M.; Lacivita, E.; De Giorgio, P.; Colabufo, N. A.; Niso, M.; Berardi, F.;
Perrone, R. J. Med. Chem. 2006, 49, 358.
25. Takács-Novák, K.; Box, K. J.; Avdeef, A. Int. J. Pharm. 1997, 151, 235.