Novel Bisphosphonates Derived from 1H-Indazole, 1H-Pyrazolo[3,4-b]Pyridine, and 1H-Pyrazolo[3,4-b]Quinoline

Article abstract of DOI:10.1002/hc.21282

Novel tetraethyl ethylene-1,1-bisphosphonate esters derived from 1H-indazole, 1H-pyrazolo[3,4-b]pyridine, and 1H-pyrazolo[3,4-b]quinoline were synthesized by a Michael addition reaction of tetraethyl ethylidene-1,1-bisphosphonate with the corresponding heterocycle, using conventional heating and microwave-assisted methods. The microwave-assisted method provides shorter reaction times and better yields. The hydrolysis of bisphosphonates afforded the corresponding bisphosphonic acids or salt, using concentrated hydrochloric acid or TMSBr/collidine, respectively. All new compounds were fully characterized, and their structures were assigned using ¹H, ³¹P, and ¹³C NMR and IR spectroscopies and mass spectrometry. The molecular structure of compound 6 was confirmed by X-ray diffraction studies.

Full text of DOI:10.1002/hc.21282

Heteroatom Chemistry
Volume 27, Number 1, 2016
Novel Bisphosphonates Derived from
1H-Indazole, 1H-Pyrazolo[3,4-b]Pyridine,
and 1H-Pyrazolo[3,4-b]Quinoline
Fa´tima C. Teixeira,1 Carla Lucas,1 M. Joa˜o M. Curto,1
Anto´nio P. S. Teixeira,2,3 M. Teresa Duarte,3 and Vaˆnia Andre´3
1Laborato´rio Nacional de Energia e Geologia, I.P., Estrada do Pac¸o do Lumiar, 22, 1649-038,
Lisboa, Portugal
2
´
´
Departamento de Qu´ımica, Escola de Cieˆncias e Tecnologia, Centro de Qu´ımica de Evora,
´
Instituto de Investigac¸a˜o e Formac¸a˜o Avanc¸ada, Universidade de Evora, 7000-671, Evora,
Portugal
3Centro de Qu´ımica Estrutural, Instituto Superior Te´cnico, Universidade de Lisboa, 1049-001,
Lisboa, Portugal
Received 6 April 2015; revised 4 July 2015
INTRODUCTION
ABSTRACT:
Novel
tetraethyl
ethylene-1,1-
bisphosphonate esters derived from 1H-indazole,
1H-pyrazolo[3,4-b]pyridine, and 1H-pyrazolo[3,4-
b]quinoline were synthesized by a Michael addition
reaction of tetraethyl ethylidene-1,1-bisphosphonate
with the corresponding heterocycle, using conven-
tional heating and microwave-assisted methods.
The microwave-assisted method provides shorter
reaction times and better yields. The hydrolysis
of bisphosphonates afforded the corresponding
bisphosphonic acids or salt, using concentrated
hydrochloric acid or TMSBr/collidine, respectively. All
new compounds were fully characterized, and their
structures were assigned using 1H, 31P, and 13C NMR
and IR spectroscopies and mass spectrometry. The
molecular structure of compound 6 was confirmed
Bisphosphonates are an important class of
organophosphorus drugs known by their
broad spectrum of therapeutic applications
in the treatment of diseases characterized by
abnormal calcium metabolism, due to their
high affinity for calcium and target the bone
mineral [1–5]. Bisphosphonates present other
therapeutic applications against pathogenic agents
due to their antiparasitic [6–8], herbicidal [9]
and antibacterial activities [10]. Bisphosphonates
were also studied as ligands for radioactive metal
complexes [11, 12] and as chelating agents for the
treatment of human metal intoxications [13].
The activity of bisphosphonates, in the treat-
ment of bone disorders, is improved by the pres-
ence of a nitrogen containing group bonded to an
alkyl moiety or to a heteroaromatic cycle [14–16].
Indazole, pyrazolo[3,4-b]pyridine, and pyrazolo[3,4-
b]quinoline are heteroaromatic compounds with
two or three nitrogen atoms in an aromatic moi-
ety, resembling the heteroaromatic part of the most
potent third-generation bisphosphonates, and are
promising moieties for drug discovery.
ꢀC
by X-ray diffraction studies.
2015 Wiley Period-
icals, Inc. Heteroatom Chem. 27:3–11, 2016; View
this article online at wileyonlinelibrary.com. DOI
10.1002/hc.21282
Correspondence to: Fa´tima C. Teixeira; e-mail: fatima.teixeira@
lneg.pt.
Indazole, pyrazolo[3,4-b]pyridine and pyrazolo
[3,4-b]quinoline have been studied, and a large
ꢀC
2015 Wiley Periodicals, Inc.
3

4
Teixeira et al.
number of remarkable biological applications have
been described, such as anticancer, antidepressant
or antianxiety, antiinflammatory, and antiviral [17–
20]. They also found applications as organic func-
tional materials, including in fluorescent and op-
toelectronics materials, such as chemosensors and
organic light emitting diodes [21–23].
3 and tetraethyl ethylene-1,1-bisphosphonate 4. The
Michael addition reaction was carried out at reflux,
in THF, during 16 h, to give the corresponding N-1
substituted bisphosphonate esters 5–7, in good to
very good yields (Table 1). This method presents
complete regioselectivity, with formation of only the
N-1 heterocyclic regioisomer.
Taking into account the potential of bisphospho-
nates and these heterocycles, it can be conceived
that molecules bearing both moieties can provide
compounds with high potential for applications in
medicinal chemistry and also in materials chemistry.
Microwave-assisted heating is an invaluable
methodology in organic synthesis, usually with some
advantages over the standard heating techniques,
such as the acceleration of the reaction, reduction
of its time, high yields, and better selectivities [24,
25]. Microwave-assisted reactions were already used
to prepare hydroxymethylene-1,1-bisphosphonates,
from carboxylic acids, phosphorus trichloride and
phosphorous acid [26], and aminomethylenebispho-
sphonates, from the reaction of amines with triethyl
orthoformate and diethyl phosphite [27].
Following previous studies on bisphosphonates
derived from indazole and condensed pyrazole [12,
28–30], the present investigation reports the syn-
thesis of new tetraethyl ethylene-1,1-bisphosphonate
esters derived from 1H-indazole, 1H-pyrazolo[3,4-
b]pyridine, and 1H-pyrazolo[3,4-b]quinoline, and
their hydrolysis to the corresponding bisphospho-
nic acids or salt, using conventional thermal or mi-
crowave irradiation conditions, to evaluate and com-
pare reaction times, yields, and regioselectivity be-
tween both methods.
To develop a new methodology for the synthe-
sis of bisphosphonate esters and to compare the be-
havior of these compounds in this reaction under
other conditions, a Michael addition reaction was
carried out using microwave-assisted conditions. In
this method, the heteroaromatic derivative 1–3 and
tetraethyl ethylidene-1,1-bisphosphonate 4 in THF
was heated to 95°C, for 30 min, in a laboratory
microwave reactor to afford, after purification, the
ethylene-1,1-bisphosphonate derivative 5–7 in very
good to excellent yields, with the yields decreasing
with the extension of the aromatic system or the in-
crease in the number of nitrogen atoms (Table 1).
This methodology showed to be superior to the con-
ventional thermal method, affording the same prod-
ucts in shorter times, with higher yields and fewer
side products. Both methodologies showed complete
regioselectivity, with the formation of only the same
N-1 regioisomer.
The bisphosphonate esters were submitted to
acid hydrolysis with concentrated hydrochloric acid.
The free bisphosphonic acids 8 and 9 were easily
prepared and isolated in good yields, but the 31P
NMR spectra of the crude of the hydrolysis of bis-
phosphonate ester 7 showed a mixture of the corre-
sponding bisphosphonic acid 10 and other uniden-
tified phosphonated species (Table 2). All attempts
to purify compound 10 by precipitation with ace-
tone/methanol/water failed.
RESULTS AND DISCUSSION
Synthesis
An alternative procedure to convert bisphospho-
nate esters to bisphosphonic acids is the McKenna
procedure, using trimethylsilyl bromide, followed
by methanolysis [33]. A first attempt to hydrolyze
bisphosphonate ester 7 under McKenna conditions
gave also a mixture of bisphosphonic acid 10 and
other phosphonated species as revealed by 31P NMR.
Therefore, the current procedure by Wiemer et al.
[34] was used, which involves the bisphosphonate es-
ter 7 hydrolysis by treatment with trimethylsilyl bro-
mide and 2,4,6-collidine, followed by a basic workup
to afford quantitatively the corresponding bisphos-
phonic acid salt 11 (Scheme 1).
The synthesis of 1H-pyrazolo[3,4-b]pyridine
and 1H-pyrazolo[3,4-b]quinoline 3 was performed
in high yields, by reaction of 2-chloro-3-
2
a
formylpyridine or 2-chloro-3-formylquinoline and
hydrazine with p-TsOH using a process reported
in the literature [30, 31]. Tetraethyl ethylidene-1,1-
bisphosphonate 4 was obtained from tetraethyl
methylenebisphosphonate in two steps, according to
a literature procedure [32].
The synthesis of new bisphosphonate esters de-
rived from indazole and pyrazole-fused heterocy-
cles was initially performed by conventional ther-
mal conditions using a preheated oil bath. The for-
mation of tetraethyl ethylidene-1,1-bisphosphonate
esters 5–7 was carried out by a Michael addition
reaction between the corresponding heterocycle 1–
Spectroscopic Characterization
The structures of synthesized compounds 5–9
and 11 were determined from 1H NMR, 31P
NMR, 13C NMR, IR and mass spectra. The
Heteroatom Chemistry DOI 10.1002/hc

Novel Bisphosphonates Derived from 1H-Indazole, 1H-Pyrazolo[3,4-b]Pyridine, and 1H-Pyrazolo[3,4-b]Quinoline
5
TABLE 1 Synthesis of Bisphosphonates 5–7 by a Michael addition reaction
Heating Condition
Reaction Time (h)
Product
Yield (%)
Conventional
16
5
71
99
Microwave
0.5
Conventional
Microwave
16
0.5
16
6
7
81
95
79
86
Conventional
Microwave
0.5
TABLE 2 Synthesis of Bisphosphonic Acids 8–10
SCHEME 1
Reaction Time (h)
Product
Yield (%)
containing phosphonate groups and to show the
molecular ion, confirming the proposed formula of
the new compounds [35].
The IR spectra of the new compounds 5–9 and
11 present large and strong bands, usually with sev-
eral maxima, characteristic of the bisphosphonate
groups [36–38].
2
8
73
4
2
9
76
The structures of bisphosphonate esters and bis-
phosphonic acids and salt (compounds 5–9 and
11) were readily identified by analysis of the NMR
data, including bidimensional techniques. Analysis
of these compounds by 1H NMR shows signals with
the expected and characteristic multiplicity due to
the coupling of the methine and methylene protons
of the side chain with the two phosphorus atoms
attached to the same carbon.
a
10
aA mixture of bisphosphonic acid 10 and unidentified phosphonated
species was obtained.
structure of compound 6 was also confirmed by sin-
gle crystal X-ray diffraction.
Several mass spectrometry (MS) techniques
were used to show the characteristic fragment
patterns of the lateral chain of the heterocycles
13C NMR spectroscopy confirms the presence
of the bisphosphonate ester, acid and salt groups,
with the presence of a carbon triplet at δ 36.9–37.3
ppm (JCP = 130–133 Hz), at δ 39.3–40.4 ppm (JCP
=
121–123 Hz), and at δ 39.8 ppm (JCP = 109 Hz),
Heteroatom Chemistry DOI 10.1002/hc

6
Teixeira et al.
bond lengths and angles are in agreement with
its aromatic character. The C atom bonded to the
ring (C7) is also within the same plane (Fig. 2).
The planarity, bond lengths and angles, and the
positioning of C7 atom are similar to the previously
reported 1-hydroxybisphosphonic acid derived from
pyrazolo[3,4-b]pyridine [30].
The C8 carbon atom and both phosphonate ester
groups are above the plane of the ring (as shown in
Fig. 2), with one phosphonate ester moiety in a syn-
clinal position relatively to the ring, displaying a tor-
sion angle of –74.2(2)º between P2–C8–C7–N2, and
the other phosphonate group in an antiperiplanar
position, with a torsion angle of 157.8(2)º between
P1–C8–C7–N2.
FIGURE 1 ORTEP view of bisphosphonate ester 6, show-
ing the atomic labelling scheme; ellipsoids are set at 50%
probability [41].
C8–C7 and C7–N2 bond distances correspond
to usual single bond lengths. The P–C8 bond
lengths are shorter than the corresponding 1-
hydroxybisphosphonic acid bonds [30] but still fall
within the range of other alkylphosphonic acids and
esters [30,38,40,41].
The P=O and P–O–C bond lengths are similar
in both phosphonate ester groups, but P=O bonds
are shorter and P–O–C bonds are longer than in the
corresponding bisphosphonic acid [30]. This is ob-
served due to the mesomeric structure of the phos-
phonic acid with no true double P=O or single P–O
bonds.
respectively, supporting the proposed structure with
two phosphonate or phosphonic groups attached to
the same carbon (P–CH–P).
The proton-decoupled 31P NMR spectra of bis-
phosphonate esters 5–7 and bisphosphonic acids 8–
9 and salt 11 showed a single signal at 20.4–20.5 and
18.6–20.5 ppm, respectively, which is in agreement
with the two phosphorus atoms magnetically equiv-
alent attached to the same carbon atom, confirming
the proposed structure.
C7 and C8 atoms present a slightly defor-
mation from the ideal tetrahedral geometry (C8–
C7–N2 113.3(1)°, P1–C8–C7 110.1(1)°, P2–C8–C7
112.0(1)°, and P1–C8–P2 114.0(1)°), due to the pres-
ence of the bulky phosphonate ester groups. This is
also the cause of the deformation from the tetra-
hedral shape of both phosphorus atoms, which
present angles ranging from 116.77(8)–101.63(8)°
for P1, and 116.61(8)–100.02(8)° for P2, as observed
for alkylphosphonic acids [30, 38, 40, 41]. In the
phosphonate ester group, the P–O–C angles range
between 120.1(1)° and 122.1(1)°, corresponding to
oxygen atoms with sp2 hybridization, and thus
Single Crystal X-Ray Diffraction
Characterization
Bisphosphonate ester 6 was crystallized from an
EtOAc/Et2O/petroleum ether solution, yielding sin-
gle crystals suitable for X-ray diffraction analysis
that confirmed the assignment of spectroscopic data.
The molecular structure of compound 6, which
crystallizes in the P21/c monoclinic space group, is
shown in Fig. 1.
The crystal structure shows that the
pyrazolo[3,4-b]pyridine ring is planar, and its
FIGURE 2 Conformation of phosphonate ester groups relatively to the aromatic ring (a) in the best view to depict the relative
positioning of all the groups and (b) in a view along the plane of the ring.
Heteroatom Chemistry DOI 10.1002/hc

Novel Bisphosphonates Derived from 1H-Indazole, 1H-Pyrazolo[3,4-b]Pyridine, and 1H-Pyrazolo[3,4-b]Quinoline
7
TABLE 3 Details of the main short interactions responsible for the supramolecular arrangement for compound 6 (ORTEP
numbering scheme)
˚
˚
˚
D–H···A
Sym. Op.
D–H (A)
H···A (A)
D···A (A)
D-H···A (°)
C3–H3···O4
x, –1 + y, z
x, –1 + y, z
x, y, z
0.93
0.88(3)
0.97
0.97
0.97
2.39
2.43(3)
2.60
2.59
2.52
3.315(2)
3.278(3)
3.006(3)
3.442(3)
2.992(3)
3.535(3)
174
161(2)
106
C5–H5···O1
C9–H9A···O1
C9–H9B···N1
C13–H13A···O4
C16–H16B···O4
1 + x, y, z
147
110
168
x, y, z
1 – x, –1/2 + y, 3/2 – z
0.96
2.59
FIGURE 3 (a) Supramolecular arrangement of compound 6 in a view along the b axis, showing the formation of lines of
molecules along a, with the pyrazolo rings oriented in a head-to-tail fashion within the same line, but with inverted orientation in
consecutive lines. (b) Supramolecular arrangement of compound 6 in a view along the a axis, showing the formation of lines of
molecules along b, with the pyrazolo rings oriented in a head-to-tail fashion within the same line, but with inverted orientation
in consecutive lines (blue vs. purple). (c) Angle formed between the pyrazolo rings of consecutive lines.
reflecting their ability to lose the single P–O bond
character to an intermediate stage with electronic
delocalization between the P–O bonds of phospho-
nate groups.
Even though some C−H···O short interactions
are present in the supramolecular arrangement of
this compound (Table 3), no classic hydrogen bonds
are observed.
In a view along the b axis, it is clear that
the pyrazolo[3,4-b]pyridine rings are parallel among
them and align along a with the molecules ori-
ented in a head-to-tail fashion in the same line while
assuming opposite directions in consecutive lines
(Fig. 3a). Each line is based on C–H···O (3.315(2),
˚
˚
and 3.278(3) A) and C–H···N 3.442(3) A) short con-
tacts established between the pyrazolo ring and two
other molecules and reinforced by another C–H···O
˚
(3.006(3) and 2.992(3) A). Consecutive lines are con-
˚
nected by C–H···O (3.535(3) A) interactions involving
the phosphonate moiety. Also in a view along the a
axis, a similar packing is observed with lines grow-
ing along the b axis. (Fig. 3b). The pyrazolo rings
in consecutive lines form a 34.80(20)° angle among
them (Fig 3c).
Heteroatom Chemistry DOI 10.1002/hc

8
Teixeira et al.
CONCLUSIONS
distilled from sodium benzophenone ketyl. CH2Cl2
was distilled from calcium hydride. Column chro-
matography was performed on silica gel (230-400
mesh) under a positive pressure of nitrogen.
Microwave-assisted reactions were carried out
using a Milestone’s MicroSYNTH–Microwave Lab-
station for the Synthesis with power supply 1600 W.
Novel tetraethylethylene-1,1-bisphosphonate es-
ters derived from 1H-indazole, 1H-pyrazolo[3,4-
b]pyridine, and 1H-pyrazolo[3,4-b]quinoline were
synthesized using conventional heating and
microwave-assisted methods by a Michael addition
reaction of tetraethyl ethylidene-1,1-bisphosphonate
with the corresponding heterocycle, with good to
excellent yields. Both methods present complete
regioselectivity, with formation of only the N-1 hete-
rocyclic derivative. The microwave-assisted method
has the advantages of short reaction times and better
yields, while achieving the same regioselectivity.
The bisphosphonate esters 5–7 were hydrolyzed
to afford the corresponding bisphosphonic acids 8–9
or salt 11.
All new compounds were fully characterized us-
ing their mass, infrared, and 1H NMR, 31P NMR, and
13C NMR data. Compound 6 gave crystals suitable
for X-ray diffraction studies, and its molecular and
crystalline structure was presented and confirmed
the proposed structure based on the assignment of
spectroscopic data.
1H-Indazole
1
is commercially available
(Aldrich, Sintra, Portugal) and was used as received.
1H-Pyrazolo[3,4-b]pyridine 2 [30], 1H-pyrazolo[3,4-
b]quinoline 3 [31] and tetraethyl ethenylidene-1,1-
bisphosphonate 4 [32] were prepared according to
literature procedures.
General Procedure 1 (Thermal Conditions)
A mixture of azole 1–3 (1 mmol) and tetraethyl
ethenylidene-1,1-bisphosphonate 4 (1 mmol) in THF
was stirred under reflux for 16 h. The solvent
was evaporated in vacuum, and the crude mate-
rial was purified by column chromatography (silica,
1:1 ethyl acetate/acetone) to give the corresponding
ethylidene-1,1-bisphosphonate derivative.
General Procedure 2 (Microwave-Assisted
Conditions)
EXPERIMENTAL
NMR spectra were recorded on a Bruker AMX 300
and on a Bruker Avance II 300 (1H 300 MHz, 13C
75 MHz, 31P 121 MHz) and on a Bruker Avance
II 400 (1H 400 MHz, 13C 100 MHz, 31P 162 MHz)
spectrometers. Chemical shifts (δ) are reported in
ppm and coupling constants (J) in Hz. 1H and
13C NMR chemical shifts were assigned using Dis-
tortionless Enhancement by Polarization Transfer
(DEPT) and Attached Proton Test (APT) sequences,
and bidimensional 2D Correlation Spectroscopy
(COSY), Heteronuclear Single Quantum Correlation
(HSQC), and Heteronuclear Multiple-Bond Corre-
lation (HMBC) techniques. Infrared spectra were
recorded on a Perkin Elmer FT-IR 1725xIR Fourier
transform spectrophotometer using KBr disks or
film. The bands are quoted in cm−1. Low-resolution
and high-resolution (HRMS) mass spectra analyses
were obtained by electron impact ionization (EI),
fast atom bombardment (FAB), or electrospray ion-
ization (ESI). Mass spectrometry (MS) techniques
were performed at the ‘C.A.C.T.I. - Unidad de Espec-
trometria de Masas’ at the University of Vigo, Spain,
on a VG AutoSpect M, MicroTOF (Bruker Daltonics)
or APEX-Q (Bruker Daltonics) instrument. Melting
points were determined on a Reichert Thermovar
melting point apparatus and are not corrected.
In the microwave reactor, a mixture of azole
1–3 (1 mmol) and tetraethyl ethenylidene-1,1-
bisphosphonate 4 (1 mmol) in THF was heated for
30 min at 95°C under microwave irradiation, with a
power of 300 W. Upon removal of the solvent, the
crude material was purified by column chromatog-
raphy (silica, 1:1 ethyl acetate/acetone) to give the
corresponding
derivative.
ethylidene-1,1-bisphosphonate
Tetraethyl
(2-(1H-indazol-1-yl)ethane-1,1-diyl)
bisphosphonate (5). Compound 5 was prepared
from 1H-indazole 1 (0.500 g, 4.23 mmol), during
16 h, following general procedure 1 (1.25 g, 71%),
as a colorless oil.
Compound 5 was prepared from 1H-indazole 1
(100 mg, 0.85 mmol), following general procedure 2
(350 mg, 99%) as a colorless oil.
υmax (film) (cm−1): 2984, 2933, 2908 (C–H), 1617,
1501, 1467, 1444 (C=N, C=C), 1392, 1369, 1252
(P=O), 1164, 1023, 975 (P–OC). 1H NMR (300 MHz,
CDCl3): δ (ppm) = 1.18 (12H, m, OCH2CH3), 3.44
(1H, tt, JHP 23.0 and J 6.6, CH(PO3Et2)2), 4.06 (8H, m,
OCH2CH3), 4.93 (2H, td, JHP 13.7 and J 6.6, NCH2),
7.13 (1H, t, J 7.5, ArH, 5-H), 7.38 (1H, t, J 7.7, ArH,
6-H), 7.55 (1H, d, J 8.4, ArH, 7-H), 7.69 (1H, d, J 8.1,
ArH, 4-H), 8.02 (1H, s, ArH, 3-H). 13C NMR (75 MHz,
CDCl3): δ (ppm) = 15.7 (d, JCP = 6.4, CH3), 37.3 (t,
All solvents were distilled under a nitrogen at-
mosphere and were degassed before use. THF was
Heteroatom Chemistry DOI 10.1002/hc

Novel Bisphosphonates Derived from 1H-Indazole, 1H-Pyrazolo[3,4-b]Pyridine, and 1H-Pyrazolo[3,4-b]Quinoline
9
JCP = 130.5, CH(PO3Et2)2), 44.3 (t, JCP = 3.1, NCH2),
62.4 (dd, JCP = 26.6 and 6.6, OCH2), 109.3 (Ar:
C7), 120.2 (Ar: C5), 120.4 (Ar: C4), 123.3 (Ar: C3a),
125.9 (Ar: C6), 133.3 (Ar: C3), 139.7 (Ar: C7a).31P
NMR (121 MHz, H3PO4/ CDCl3): δ (ppm) = 20.5. MS
(FAB): m/z = 419 (MH+, 90%), 373 (M+ – OEt , 9%),
301 (M+ – Indz, 100%), 281 (M+ – PO3Et2, 16%).
HRMS (FAB) m/z calcd for C17H29N2O6P2 419.1501
[MH]+, found 419.1500.
3.77 (1H, tt, JHP 23.0 and J 7.0, CH(PO3Et2)2), 4.11
(8H, m, OCH2CH3), 5.22 (2H, td, JHP 13.6 and J
7.3, NCH2), 7.45 (1H, dt, J 7.6 and 0.8, ArH, 6-H),
7.76 (1H, dt, J 7.8 and 1.6, ArH, 7-H), 7.97 (1H,
d, J 8.0, ArH, 5-H), 8.17 (1H, d, J 8.4, ArH, 8-H),
8.25 (1H, s, ArH, 3-H), 8.63 (1H, s, ArH, 4-H). 13C
NMR (100 MHz, CDCl3): δ (ppm) = 16.47 (d, JCP
5.1, CH2CH3), 16.53 (d, JCP 5.2, CH2CH3), 37.1 (t, JCP
130.7, CH(PO3Et2)2), 44.2 (NCH2), 63.1 (t, JCP 6.4,
OCH2), 117.5 (Ar: C3a), 124.2 (Ar: C6), 124.5 (Ar:
C4a), 128.0 (Ar: C8), 129.6 (Ar: C5), 131.3 (Ar: C4
and C7), 133.4 (Ar: C3), 147.5 (Ar: C8a), 149.7 (Ar:
C9a). 31P NMR (121 MHz, H3PO4/ CDCl3): δ (ppm)
Tetraethyl
(2-(1H-pyrazolo[3,4-b]pyridin-1-yl)
ethane-1,1-diyl)bisphosphonate (6). Compound 6
was prepared from 1H-pyrazolo[3,4-b]pyridine 2
(500 mg, 4.17 mmol), during 16 h, following general
procedure 1 (1.42 g, 81%), as a pale yellow solid.
= 20.5. MS (EI): m/z = 469 (M+, 3%), 332 (M+
–
–
PO3Et2, 10%), 195 (M+ – (PO3Et2)2, 3%), 182 (M+
Compound
6
was prepared from 1H-
CH(PO3Et2)2, 9%), 169 (C10H7N3+, 100%), 168 (M+
– CH2CH(PO3Et2)2, 6%). HRMS (EI) m/z calcd for
C20H29N3O6P2 469.1532 [M]+, found 469.1527.
pyrazolo[3,4-b]pyridine 2 (100 mg, 0.84 mmol),
following general procedure 2 (335 mg, 95%) as a
pale yellow solid.
mp 50–51°C. υmax (film) (cm−1): 3095, 3053 (C–
H Ar), 2984, 2905 (C–H), 1597, 1574, 1502, 1461,
1445 (C=N, C=C), 1249 (P=O), 1017, 972 (P–OC).
1H NMR (300 MHz, CDCl3): δ (ppm) = 1.13 (6H,
t, J 7.0, OCH2CH3), 1.99 (6H, t, J 7.1, OCH2CH3),
3.66 (1H, tt, JHP 23.0 and J 7.8, CH(PO3Et2)2), 4.06
(8H, m, CH2CH3), 5.05 (2H, td, JHP 13.4 and J 6.9,
NCH2), 7.10 (1H, dd, J 7.8 and 4.6, ArH, 5-H), 8.00
(1H, s, ArH, 3-H), 8.03 (1H, dd, J 8.1 and 1.4, ArH,
4-H), 8.51 (1H, dd, J 4.5 and 1.4, ArH, 6-H). 13C
NMR (75 MHz, CDCl3): δ (ppm) = 16.1 (d, JCP
4.0, OCH2CH3), 16.2 (d, JCP 4.0, OCH2CH3), 36.9 (t,
JCP 132.5, CH(PO3Et2)2), 43.2 (m, NCH2), 62.7 (m,
OCH2), 115.6 (Ar: C3a), 116.8 (Ar: C5), 129.8 (Ar:
C4), 132.1 (Ar: C3), 148.5 (Ar: C6), 150.4 (Ar: C7a).31P
NMR (121 MHz, H3PO4/ CDCl3): δ (ppm) = 20.4. MS
(EI): m/z = 419 (M+, 12%), 282 (M+ – PO3Et2, 100%),
145 (M+ – (PO3Et2)2,14%), 132 (M+ – CH(PO3Et2)2,
27%), 118 (M+ – CH2CH(PO3Et2)2, 15%). HRMS (EI)
m/z calcd for C16H27N3O6P2 419.1375 [M]+, found
419.1377.
(2-(1H-Indazol-1-yl)ethane-1,1-diyl)bisphos-
phonic Acid (8). A solution of compound
5
(100 mg, 0.24 mmol) in concentrated HCl (1 mL)
was refluxed for 2 h. After solvent removal under
reduced pressure, the residue was precipitated
with acetone and methanol. Compound 8 (53 mg,
73%) was isolated as a white solid. mp 245°C. υmax
(KBr) (cm−1): 3500–2200 (O–H), 3123, 3018 (C–H
Ar), 2987, 2891 (C–H), 2632 (PO–H), 1628, 1575,
1520, 1457 (C=N, C=C), 1234 (P=O), 1195, 1164,
1
1125, 1056, 998, 988 (P–OC). H NMR (400 MHz,
DMSO-d6): δ (ppm) = 2.99 (1H, tt, JHP 22.2 and J
6.2, CH(PO3H2)2), 4.80 (2H, td, JHP 13.8 and J 6.4,
NCH2), 7.14 (1H, t, J 7.4, ArH, 5-H), 7.40 (1H, t, J
7.6, ArH, 6-H), 7.64 (1H, d, J 8.8, ArH, 7-H), 7.76
(1H, d, J 8.0, ArH, 4-H), 8.10 (1H, s, ArH, 3-H).
13C NMR (75 MHz, DMSO-d6): δ (ppm) = 40.4 (t,
JCP 121.6, CH(PO3H2)2), 45.8 (NCH2), 111.1 (Ar:
C7), 121.1 (Ar: C5), 121.6 (Ar: C4), 124.4 (Ar: C3a),
126.7 (Ar: C6), 133.8 (Ar: C3), 140.5 (Ar: C7a).31P
NMR (121 MHz, H3PO4/ DMSO-d6): δ (ppm) = 18.8.
MS (ESI): m/z = 307 (MH+, 100%). HRMS (ESI)
m/z calcd for C9H13N2O6P2 307.0243 [MH]+, found
307.0250.
Tetraethyl (2-(1H-pyrazolo[3,4-b]quinolin-1-yl)
ethane-1,1-diyl)bisphosphonate (7). Compound 7
was prepared from 1H-pyrazolo[3,4-b]quinoline 3
(500 mg, 2.96 mmol), during 16 h, following general
procedure 1 (1.10 g, 79%), as a pale yellow solid.
(2-(1H-Pyrazolo[3,4-b]pyridin-1-yl)ethane-1,1-
diyl)bisphosphonic Acid (9). A solution of com-
pound 7 (100 mg, 0.24 mmol) in concentrated HCl
(1 mL) was refluxed for 4 h. After solvent removal
under reduced pressure, the residue was precipi-
tated with acetone and methanol. Bisphosphonate
9 (56 mg, 76%) was isolated as a white solid. mp
245°C (decomp.). υmax (KBr) (cm−1): 3500–2300
(O–H), 3102, 3096 (C–H Ar), 2927 (C–H), 2718
(PO–H), 1649, 1544, 1459 (C=N, C=C), 1241 (P=O),
1150, 1114, 1066, 1051, 998, 973, 954 (P-OC). 1H
Compound
7
was prepared from 1H-
pyrazolo[3,4-b]quinoline 3 (145 mg, 0.86 mmol),
following general procedure 2 (347 mg, 86%) as a
pale yellow solid.
mp 81–82°C. υmax (KBr) (cm−1): 3099, 3035 (C–H
Ar), 2984, 2905 (C–H), 1618, 1570, 1498, 1455, 1438
(C=N, C=C), 1257, 1246 (P=O), 1046, 1028, 971, 946
(P–OC). 1H NMR (400 MHz, CDCl3): δ (ppm) = 1.17
(6H, t, J 7.0, OCH2CH3), 1.23 (6H, t, J 7.0, OCH2CH3),
Heteroatom Chemistry DOI 10.1002/hc

10 Teixeira et al.
NMR (400 MHz, DMSO-d6): δ (ppm) = 3.27 (1H, tt,
JHP 22.0 and J 6.4, CH(PO3H2)2), 4.92 (2H, td, JHP
13.6 and J 6.8, NCH2), 7.23 (1H, dd, J 8.0 and 4.4,
ArH, 5-H), 8.17 (1H, s, ArH, 3-H), 8.25 (1H, dd, J
8.0 and 1.2, ArH, 4-H), 8.57 (1H, d, J 3.2, ArH, 6-H).
13C NMR (100 MHz, DMSO-d6): δ (ppm) = 39.3 (t,
JCP 123.4, CH(PO3H2)2), 44.1 (NCH2), 116.3 (Ar:
C3a), 117.7 (Ar: C5), 131.3 (Ar: C4), 132.9 (Ar: C3),
149.2 (Ar: C6), 150.8 (Ar: C7a).31P NMR (121 MHz,
H3PO4/ DMSO-d6): δ (ppm) = 18.6. MS (ESI): m/z =
308 ([M + H]+, 100%). HRMS (ESI) m/z calcd for
C9H12N3O6P2 308.0195 [MH]+, found 308.0210.
collection was monitored by the SMART program
(Bruker, 2003) [42]. All the data were corrected
for Lorentzian, polarization, and absorption effects
using the SAINT and SADABS programs (Bruker,
2003) [42]. All non-hydrogen atoms were refined
by full matrix least squares on F2 with anisotropic
thermal motion parameters, whereas H-atoms were
placed in idealized positions and allowed to refine
isotropically riding on the parent C atom. The struc-
ture was solved by direct methods with SIR97 [43]
and refined by full matrix least squares on F2
with SHELXL97 [44], both included in the pack-
age of programs WINGX-VERSION 1.70.01 [45].
Graphical representations were prepared using
ORTEP3.5 [39] and Mercury 3.3 programs [46].
CCDC 1050362 contains all crystallographic de-
tails of this publication and is available free of charge
at www.ccdc.cam.ac.uk/conts/retrieving.html or can
be ordered from the following address: Cambridge
Crystallographic Data Centre, 12 Union Road, Cam-
bridge CB2 1EZ; UK, fax: +44-(0)1223-336033; or
deposit@ccdc.cam.ac.uk.
(2-(1H-Pyrazolo[3,4-b]quinolin-1-yl)ethane-1,1-
diyl)bisphosphonic Acid Tetrasodium Salt (11). To
a solution of bisphosphonate ester 9 (50 mg, 0.107
mmol) in dry CH2Cl2 at 0°C, collidine (0.15 mL,
1.07 mmol) and TMSBr (0.15 mL, 1.07 mmol)
were added. After 2 h at 0ºC, the reaction mixture
was allowed to warm to room temperature and
was stirred for 18 h. To the mixture, toluene was
added and the solvents were removed in vacuum to
obtain a white solid. To the solid, an aqueous NaOH
solution (1 M) (5 mL) was added. After 24 h, the
mixture was poured into acetone and stored at 0ºC
for 24 h. The mixture was decanted, and the residue
was washed with cold hexane and dried in vacuum
to afford bisphosphonate 11 (48 mg, quantitative)
as a white solid.
ACKNOWLEDGMENTS
We thank Fundac¸a˜o para a Cieˆncia e a Tec-
nologia (POCTI, POCI and FEDER programs)
for provision of funding (Projects POCI/QUI/
55508/2004, PPCDT/QUI/55508/2004, RECI/QEQ-
QIN/0189/2012, PEst-OE/QUI/UI0100/2013, and
SFRH/BPD/78854/2011), and Portuguese NMR
Network (IST-UTL Center - FCT project RECI/QEQ-
QIN/0189/2012) for providing access to the NMR
facilities.
mp >350°C. υmax (KBr) (cm−1): 3600–2900 (O–
H), 1688, 1624, 1458 (C=N, C=C), 1219 (P=O), 1083,
1
954 (P–OC). H NMR (300 MHz, D2O): δ (ppm) =
2.85 (1H, tt, JHP 20.0 and J 7.6, CH(PO3H2)2), 4.76
(2H, m, NCH2), 7.39 (1H, t, J 7.4, ArH, 6-H), 7.73
(1H, t, J 7.6, ArH, 7-H), 7.83 (1H, d, J 8.8, ArH, 5-
H), 7.87 (1H, d, J 8.4, ArH, 8-H), 8.21 (1H, s, ArH,
3-H), 8.58 (1H, s, ArH, 4-H). 13C NMR (100 MHz,
D2O): δ (ppm) = 39.8 (t, JCP 109.2, CH(PO3Na2)2),
45.1 (NCH2), 117.2 (Ar: C3a), 123.6 (Ar: C6), 125.8
(Ar: C4a), 129.6 (Ar: C8), 131.5 (Ar: C5), 133.0 (Ar:
C4 and C7), 133.6 (Ar: C3), 146.9 (Ar: C8a), 148.7 (Ar:
C9a). 31P NMR (121 MHz, D2O): δ (ppm) = 20.5. MS
(ESI): m/z = 424 ([M – Na + 2H]+). MS (negative ion
- ESI): m/z = 356 ([M – 4Na + 3H]). HRMS (ESI)
m/z calcd for C12H11N3O6P2Na3 423.9811 [M – Na +
2H]+, found 423.9812.
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