Separation of planar rotamers through intramolecular hydrogen bonding in polysubstituted 5-nitrosopyrimidines

Article abstract of DOI:10.1039/c4cc06978a

While purifying new polysubstituted 5-nitrosopyrimidines, the unique separation of pairs of rotamers as chemical species, stabilized by a single intramolecular hydrogen bond and differing only in nitroso group orientation, was achieved. This interesting stereochemical phenomenon is compared to the well-known atropisomerism. This journal is

Full text of DOI:10.1039/c4cc06978a

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Separation of planar rotamers through
intramolecular hydrogen bonding in
polysubstituted 5-nitrosopyrimidines†‡
Cite this: Chem. Commun., 2014,
50, 14892
Received 4th September 2014,
Accepted 8th October 2014
L. Cechova,§ E. Prochazkova,§ac I. Cısarova,c M. Dracınsky*a and Z. Janeba*a
ab
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DOI: 10.1039/c4cc06978a
www.rsc.org/chemcomm
While purifying new polysubstituted 5-nitrosopyrimidines, the unique atoms are sp2-hybridized (as in amides, enols of b-diketones and
separation of pairs of rotamers as chemical species, stabilized by a b-enaminones, heteroaromatic rings, etc.).1 Some researchers
single intramolecular hydrogen bond and differing only in nitroso suggest that the stability of such systems is due to stabilization
group orientation, was achieved. This interesting stereochemical by a resonance-assisted hydrogen bond (RAHB).3 Others believe
phenomenon is compared to the well-known atropisomerism.
that RAHB is not the primary reason behind the strength of the
IMHBs in a series of enols of b-diketones and b-enaminones,
The ability to form strong hydrogen bonds is one of the most but rather that the molecular structure (s-skeleton) keeps the
important properties of biologically active compounds. Intermole- hydrogen donor and hydrogen acceptor coplanar and, therefore,
cular hydrogen bonding is crucial for encoding genetic informa- closer to each other.4
tion (Watson–Crick base pairs); intramolecular hydrogen bonding
An effect of the internal hydrogen bond on the conformational
is pivotal to the formation of biologically relevant three- isomerism of 2-amino-2-methylpropanol was theoretically studied
dimensional structures of macromolecules, such as nucleic acids by Andrade et al.5 Recently, conformers of C-nitroso derivatives of
and proteins. Nevertheless, the ability to form intramolecular indolizines and azaindolizines with restricted rotation about the
hydrogen bonds (IMHBs) also has a great impact on the mole- C–NO bond were studied by the Katritzky group using solution
cular structure and physico-chemical properties of small organic NMR methods.6 Conjugated nitrosamines can evidently form
molecules, making the closed form (with an IMHB) more lipo- planar systems with enhanced p-delocalization and, thus, belong
philic and the open form more water-soluble.1 Furthermore, the to a class of compounds able to form strong IMHBs.7 During
cyclic system formed due to the presence of an IMHB can mimic recent studies of 5-nitrosopyrimidines bearing distinct amino
the structural motifs of other biologically relevant molecules. substituents in the C4 and C6 positions, two possible rotamers,
Thus, replacing the real ring with a ‘‘pseudo’’ ring in drug-like stabilized by strong IMHBs, were identified as two sets of signals
molecules has become an important medicinal chemistry strategy in 1H NMR spectra.8 In our previous paper,8d the ratio of the two
in the search for new lead compounds.1,2
rotamers was found to be significantly substituent-dependent
The high stability of IMHBs has often been observed in planar with a broad range of conformation ratios. NMR spectroscopy
systems where a six-membered ring is formed and the linker showed that the interconversion barriers of these rotamers were
as high (420 kcal molꢀ1) as those reported for atropisomers,9
which are conformational isomers isolable through sterically
hindered rotation. The high interconversion barriers of these
rotamers led us to wonder: could the use of low temperatures
with appropriately substituted amino groups in positions C4
and/or C6 potentially lead to the separation of a pair of IMHB-
stabilized rotamers? As non-covalent interactions (e.g. hydrogen
bonds) are considerably less stable compared to covalent bonds, it
has been suggested by Szumna10 that separation of isomers based
on a single hydrogen bond is not possible, unless additional
auxiliaries are introduced. Indeed, the isolation of such isomers
has so far been successful either in relatively complex molecules
with an ability to form cooperative robust systems of many hydrogen
bonds,11 or in the case of relatively stable formamide rotamers.12
a Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech
´
Republic, v.v.i., Flemingovo nam. 2, CZ-166 10 Prague 6, Czech Republic.
E-mail: janeba@uochb.cas.cz, dracinsky@uochb.cas.cz
b Department of Chemistry of Natural Compounds, Institute of Chemical Technology
´
Prague, Technicka 5, CZ-166 28 Prague 6, Czech Republic
c Faculty of Science, Charles University in Prague, Hlavova 2030,
CZ-128 40 Prague 2, Czech Republic
† This paper is dedicated to the memory of Professor Alan R. Katritzky, who died
on February 10, 2014.
‡ Electronic supplementary information (ESI) available: Supplementary figures
and Table, X-ray diffraction data, kinetics, DFT calculations, synthesis and
characterization of the prepared compounds. CCDC 1001296 and 1001297. For
ESI and crystallographic data in CIF or other electronic format see DOI: 10.1039/
c4cc06978a
§ L.C. and E.P. contributed equally to this work.
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Fig. 1 Two possible geometries of the nitroso group in position C5 for
compounds 1–4. Rotamer A has a torsion angle C6–C5–N–O close to 01;
rotamer B has a torsion angle C6–C5–N–O close to 1801.
Fig. 2 General characteristics of atropisomerism and the herein reported
Although spatial separation using an electric field has recently stereochemical phenomenon. A a B and A0 a B0, A/A0 has priority over B/B0.
been developed to enable the study of small molecule confor-
mers,13 the isolation and full characterization of small mole-
both sides of the axis; in our case, there are differently substituted
amino groups in the C4 and C6 positions, and an oxygen and
free electron pair on the nitrosyl nitrogen. However, unlike
atropisomers,9 which contain an element of chirality (usually
through the axis), our planar rotamers (shortly planamers),
precisely because they are planar, lack any element of chirality.
The solid-state 13C cross polarization-magic angle spinning
(CP-MAS) NMR spectra of rotamers 1A and 1B obtained after
gentle evaporation of the solvents from the chromatographic
fractions are depicted in Fig. 3 and Fig. S1 (ESI‡). According to these
spectra, both rotamers were pure; the amount of the second one in
the pair was lower than the solid-state NMR detection limit (ca. 5%).
The 13C NMR signals of rotamer 1A are sharp, indicating long-range
order in the solid, while the spectrum of rotamer 1B exhibits the
features typical of amorphous structures; namely, broad signals with
multiple overlapped resonances. Interestingly, the solid-state
13C NMR spectra show that while the crystalline rotamer 1A was
perfectly stable at room temperature, the solid amorphous
rotamer 1B slowly re-crystallized, over a period of months, into
the crystalline rotamer 1A (Fig. 3 and Fig. S1 in ESI‡).
cule rotamers of aromatic compounds as separate chemical
species stabilized by a single resonance-assisted IMHB has not
yet been reported.
In this paper, we present unique examples of the separation
and full characterization of such planar rotamers isolable
through a single intramolecular hydrogen bond.
Our previously described methodology8d (see ESI‡) was used to
prepare a new set of polysubstituted 5-nitrosopyrimidines bearing
two different hydrogen-bond donors in positions C4 and C6 of the
pyrimidine moiety. While purifying compounds 1–4 (Fig. 1), two
products were observed as partially separated UPLC peaks and as
well-separated spots on thin layer chromatography (TLC) plates.
These products were isolated by routine silica gel chromatography
and, to our surprise, found to have identical mass and sub-
sequently identified as A and B rotamer pairs (Fig. 1), differing
only in their 5-nitroso group orientation. The purification of
compounds 1 and 2 led to the isolation of pure rotamers 1A/1B
and 2A/2B. In the case of compounds 3 and 4, it was only possible
to isolate 3A and 4A as pure isomers; 3B and 4B were always, to
some extent, contaminated by 3A and 4A, respectively.
The solid-state IR spectra of rotamers 1A and 1B were acquired on
samples dispersed in KBr pellets. Similar to the NMR spectra, there
were significant differences between the rotamers. For example,
a difference of up to 40 cmꢀ1 was observed between their anti-
symmetric N–H vibrations. Furthermore, as with the NMR
spectra, the signals of the amorphous isomer 1B were slightly
We were quite astonished by the fact that we were able to
separate rotamers A and B (Fig. 1) at room temperature and
characterize them using just standard spectroscopic methods.
Initially, we considered this phenomenon to be a special case of
atropisomerism because atropisomers are, by definition,9 stereo-
isomers with restricted rotation around a single bond where the
rotational barrier is high enough to permit isolation of the isomeric
species. However, we quickly realized that there were significant
differences between typical atropisomers and our rotamers, the
most obvious being their mode of stabilization. Steric hindrance
stabilizes atropisomers, but our planar rotamers were stabilized by
intramolecular hydrogen bonding (Fig. 2).
Having made this distinction, we explored further similarities
and differences between atropisomers and our planar rotamers.
We found that there are two necessary preconditions for both the
phenomena. The first precondition is a rotationally stable axis,
which, in our case, lies along the C5–NO bond. Analogous to the
axis of the most common class of atropisomers represented
by biaryl compounds (the sp2–sp2 family),9 the herein reported
rotamers are characterized by a rotationally stable bond between
sp2 carbon (C5) and sp2 nitrogen (nitroso group) atoms. The second
precondition involves the presence of different substituents on
Fig. 3 The aromatic region of 13C CP-MAS solid-state NMR spectra of
planar rotamers 1A and 1B. Solid amorphous 1B slowly transforms into
solid crystalline 1A. The full spectrum is available in Fig. S1 in ESI.‡
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Fig. 4 X-ray crystallographic analysis of pure rotamer 1A (A) and of the
monohydrate of compound 1 (B). Carbon is shown in grey, nitrogen in
blue, oxygen in red and phosphorus in yellow. The less occupied position
of the nitroso group of rotamer 1B is drawn with dashed lines (B). The
displacement ellipsoids are drawn at a 30% probability level. The disor-
dered atoms of the phosphonate moiety and solvating water molecule are
omitted for clarity.
Fig. 5 Interconversion of rotamers 1A and 1B. Interconversion was monitored
by 1H NMR spectroscopy in a DMSO-d6 solution. R1 = CH2–CH2–O–CH2–
P(O)(OEt)2.
broader than those of the crystalline isomer 1A, with no visible
contamination by each other (Fig. S2 in ESI‡).
After the re-crystallization of compound 1 (a mixture of 1A and
1B) from dry acetone, a monocrystal suitable for X-ray diffraction
(XRD) was obtained (Fig. 4A). XRD confirmed that the orientation
of the nitroso group corresponded to that of pure rotamer 1A. The
solid-state 13C NMR spectrum of this recrystallized material was
identical to the spectrum of 1A obtained by column chromato-
graphy. However, following re-crystallization from an acetone–
water mixture, a monohydrate of compound 1 was obtained. In
this monohydrate (Fig. 4B), both rotamers were present, 1A being
predominant (ca. 80%). Clearly, in a solid state, the conformational
stability of a rotamer depends on the polymorphic/crystalline form
of the parent compound. In this way, our planamers meet Oki’s
criterion for atropisomers:14 physically separable species at a given
temperature, having a half-life (t) of at least 1000 s (B16.7 min),
which is the time considered to be the minimum lifetime for a
molecule to be isolable.
NH hydrogen atoms and the pyrimidine carbon atoms. These
correlations were observed when the atoms involved were
arranged in the form of a W-like coupling path (Fig. S4 in
ESI‡). Furthermore, the orientation of the nitroso group had
a significant influence on the chemical shifts of C4 and C6: by
ca. 12 ppm, the C6 carbon was more shielded in rotamer A and
the C4 carbon more shielded in rotamer B.
Fig. 6 shows the progress of the isotope exchange of hydrogen
C6–NH for compound 1 in a DMSO-d6–CD3OD mixture. As can
be seen from the rates of the hydrogen-to-deuterium exchange
reaction, the formation of intramolecular hydrogen bonds
reduced the accessibility of the solvent to the hydrogen bond
donors. In rotamer 1A, where C6–NH is involved in the hydrogen
bond, the exchange reaction was much slower than in rotamer
1B. As is normal for free amino groups, the hydrogen atoms of
the amino group in position C2 were completely replaced by
deuterium before the first spectrum could be acquired. Interest-
ingly, both amino hydrogens in position C4 in isomer 1B were
also fully replaced before the first data acquisition. We assume
that this was caused by the fast isotope exchange of the unbound
amino hydrogen followed by fast rotation around the C4–NH2
Immediately after dissolution of the separated rotamers A or B
1
in DMSO-d6 or CD3OD, their H and 13C NMR spectra contained
one major set of signals together with the low intensity signals of its
corresponding second rotamer, B or A, respectively. Gradually, the
A/B signal ratio changed until, after several hours, equilibrium was
reached. The kinetics of these transformations were determined
by solution-state 1H NMR spectroscopy (Fig. 5 and Fig. S3 in ESI‡).
The NH hydrogen signals of the two rotamers were usually
well separated and, thus, their integration enabled the mixture
composition to be determined at various times after dissolution.
Table S1 (see ESI‡) summarizes the rate constants (calculated
according to the procedure in ESI‡) and the concentrations of
rotamers 1A–4A in equilibrium in various solvents. It is evident that
the rotamer equilibrium ratio was not only strongly substituent
dependent, but also solvent dependent.
The signals of the NH hydrogens involved in the hydrogen
bond were shifted downfield (10–13 ppm) with respect to the
signals of the free NH hydrogens (7–9 ppm). The position and
orientation of the hydrogen-bond donor atom was confirmed
both by the splitting of the signal, which resulted from indirect
spin–spin interactions with neighbouring hydrogen atoms, and
by the two- and three-bond correlations observed in the HMBC
spectra. We also observed four-bond correlations between the
Fig. 6 The progress of the H–D exchange of hydrogen atoms of the
amino group in position C6 in the equilibrium mixture of rotamers 1A and
1B in a DMSO-d6–CD3OD mixture (9 : 1).
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bond. This hypothesis is supported by the density functional Notes and references
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Chem. Commun., 2014, 50, 14892--14895 | 14895