Synthesis and characterisation of new 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides as potential adenosine receptor ligands

Article abstract of DOI:10.1016/j.molstruc.2015.02.009

Abstract Chromones are 4H-benzopyran-4-one heterocycles that have been thoroughly studied due to their interesting biological activities. Thiazole based compounds have been used in therapeutics as antimicrobial, antiviral and as antifungal agents for a long time but, in the past decades, they have been identified as potent and selective ligands for adenosine receptor. In continuation of our project related to the syntheses of pharmacologically important heterocycles, a new series of chromone-thiazole hybrids have been designed as potential ligands for human adenosine receptors. In this context, new 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides were synthesized from chromone-2-carboxylic acid by two different amidation methods. The development of dissimilar synthetic approaches provided the possibility of working with diverse reaction conditions, namely with conventional heating and/or microwave irradiation. The structure of the compounds has been established on the basis of NMR and MS spectroscopy and X-ray crystallography. Relevant data related to the molecular geometry and conformation of the chromone-thiazole hybrids has been acquired which can be of the utmost importance to understand ligand-receptor binding.

Full text of DOI:10.1016/j.molstruc.2015.02.009

Accepted Manuscript
Synthesis and characterisation of new 4-oxo-N-(substituted-thiazol-2-yl)-4H-
chromene-2-carboxamides as potential adenosine receptor ligands
Fernando Cagide, Fernanda Borges, Ligia R. Gomes, John Nicolson Low
PII:
S0022-2860(15)00094-0
DOI:
http://dx.doi.org/10.1016/j.molstruc.2015.02.009
MOLSTR 21307
Reference:
Journal of Molecular Structure
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18 October 2014
17 January 2015
5 February 2015
Please cite this article as: F. Cagide, F. Borges, L.R. Gomes, J.N. Low, Synthesis and characterisation of new 4-
oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides as potential adenosine receptor ligands, Journal of
Molecular Structure (2015), doi: http://dx.doi.org/10.1016/j.molstruc.2015.02.009
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Synthesis and characterisation of new 4-oxo-N-(substituted-thiazol-
2-yl)-4H-chromene-2-carboxamides as potential adenosine receptor
ligands
Fernando Cagide,a Fernanda Borges,a* Ligia R. Gomesb and John Nicolson Lowc
a CIQ/Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade do Porto,
4169-007 Porto, Portugal
b REQUIMTE, FP-ENAS-Faculdade de Ciências de Saúde, Escola Superior de Saúde da UFP,
Universidade Fernando Pessoa, P-4200-150 Porto, Portugal
c Department of Chemistry, University of Aberdeen, Meston Walk, Old Aberdeen, AB24 3UE,
Scotland
Abstract
Chromones are 4H-benzopyran-4-one heterocycles that have been thoroughly studied due to
their interesting biological activities. Thiazole based compounds have been used in therapeutics
as antimicrobial, antiviral and as antifungal agents for a long time but, in the past decades, they
have been identified as potent and selective ligands for adenosine receptor. Thus as a continual
ion of a project related to the syntheses of pharmacologically important heterocycles, a new
series of chromone-thiazole hybrids have been designed as potential ligands for human
adenosine receptors. In this context, new 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-
carboxamides were synthesized from chromone-2-carboxylic acid by two different amidation
methods. The development of dissimilar synthetic approaches provided the possibility of
working with diverse reaction conditions, namely with conventional heating and/or microwave
irradiation. The structure of the compounds has been established on the basis of NMR and MS
spectroscopy and X-ray crystallography. Relevant data related to the molecular geometry and
conformation of the chromone-thiazole hybrids has been acquired which can be of the utmost
importance to understand ligand-receptor binding.
Keywords: chromone-thiazole hybrids; conventional heating; microwave
irradiation

Synthesis and characterisation of new 4-oxo-N-(substituted-thiazol-
2-yl)-4H-chromene-2-carboxamides as potential adenosine receptor
ligands
Fernando Cagide,a Fernanda Borges,a Ligia R. Gomesb and John Nicolson Lowc
a CIQ/Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade do Porto,
4169-007 Porto, Portugal
b REQUIMTE, FP-ENAS-Faculdade de Ciências de Saúde, Escola Superior de Saúde da UFP,
Universidade Fernando Pessoa, P-4200-150 Porto, Portugal
c Department of Chemistry, University of Aberdeen, Meston Walk, Old Aberdeen, AB24 3UE,
Scotland
1. Introduction
Chromones are compounds based on 4H-benzopyran-4-one and they form a large class of natural
and synthetic heterocyclic compounds that have been studied due to their interesting biological
activities [1]. Numerous biological effects (e.g. enzyme inhibition properties, anti-inflammatory,
anticancer, antimicrobial and antioxidant activities) [1, 2] have been ascribed to this benzo-γ-
pyrone nucleus in the popular medicine leading to an interest in their therapeutic application.
Recently, the chromone scaffold has been used for development of new MAO-B inhibitors [3-6]
as well as a model ligand for adenosine receptors [7,8]. Thiazoles also act as ligands towards a
great variety of biological substrates, and this being so they are very interesting functional
groups for application in medicinal chemistry. Thiazoles are used in therapeutics as
antimicrobial, antiviral and as antifungal agents but, in the past decades, they have been
investigated for other pharmacological activities e.g. anti-allergic [9], anti-hypertension [10],
pain treatment [11] and for controlling schizophrenia [12] symptoms. A comprehensive review
of the biological properties and uses of thiazoles derivatives, before 2005, has been recently
published [13,14]. Since that time thiazoles have been assayed as agents for controlling
inflammation [15] and they have been, like chromones, identified as potent and selective ligands
for adenosine receptors [16]. Thus, in continuation of our syntheses of pharmacologically
important heterocycles, a new series of chromone-thiazole hybrids have been designed as ligands
for human adenosine receptors. This work reports the synthesis and structural characterization of
some chromone-thiazole hybrids that are depicted in Figure 1.

2. Experimental
2.1. Materials, methods and apparatus
Chromone-2-carboxylic acid, aniline derivatives, phosphorus (V) oxychloride, benzotriazol-1-
yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyBOP), N,N-diisopropylethylamine
(DIPEA), dimethylformamide (DMF) and its derivatives were purchased from Sigma-Aldrich
Química S.A. (Sintra, Portugal). All other reagents and solvents were pro analysis grade and
were acquired from Merck (Lisbon, Portugal) and used without additional purification.
Thin-layer chromatography (TLC) was carried out on precoated silica gel 60 F254 (Merck) with
layer thickness of 0.2 mm. For analytical control the following systems were used: ethyl
acetate/petroleum ether, ethyl acetate/methanol, chloroform/methanol in several proportions. The
spots were visualized under UV detection (254 and 366 nm) and iodine vapour. Column
chromatography was performed using silica gel 60 0.2-0.5 or 0.040-0.063 mm (Merck).
Microwave-assisted synthesis was performed in a Biotage® Initiator Microwave Synthesizer.
2.2. Synthesis
Synthesis of 4-oxo-N-(thiazol-2-yl)-4H-chromene-2-carboxamide (1).
To a solution of chromone-2-carboxylic acid (1 g, 5.26 mmol) in DMF (12 mL) at 4ºC was
added N,N-diethylpropan-2-amine (0.92 ml, 5.26 mmol) and a solution of PyBOP (2.73 mg, 5.26
mmol) in CH2Cl2 (12 mL). The mixture was kept in an ice bath and stirred for half hour. After
this period thiazol-2-amine (526 mg, 5.26 mmol) was added and the mixture was allowed to then
warm up to room temperature. The reaction was kept with stirring by 4 hours. The solution was
diluted with water and the formed precipitate was filtered and washed with water and MeOH. 4-
Oxo-N-(thiazol-2-yl)-4H-chromene-2-carboxamide was obtained as a white powder (Yield:
55%). 1H NMR (DMSO): δ= 7.05 (1H, s, H(3)), 7.38 (1H, d, J = 3.7 Hz, H(3’), 7.55 (1H, ddd, J
= 8.1, 7.0, 1.2 Hz, H(6)), 7.64 (1H, d, J = 3.7 Hz, H(4’)), 7.84 (1H, dd, J = 8.5, 0.8 Hz,H(8)),
7.90 (1H, ddd, J = 8.6, 7.0, 1.7 Hz, H(7)), 8.06 (1H, dd, J = 8.0, 1.3 Hz, H(5)), 13.32 (1H, bs,
NH). 13C NMR (DMSO): δ= 111.8 (C3), 114.6 (C3’), 119.1 (C8), 123.8 (C4a), 124.9 (C5),
126.1 (C6), 135.1 (C7, C4’), 155.3 (C2, C8a, CONH), 159.5 (C1’) 177.4 (C4). MS/EI m/z: 272.1
(M+, 58), 244 (18), 216 (13), 173 (49), 145 (30), 89 (100).
Synthesis of N-(5-methylthiazol-2-yl)-4-oxo-4H-chromene-2-carboxamide (2)

To a solution of the chromone-2-carboxylic acid (1 g, 5.26 mm) in DMF (12 mL) at 4ºC was
added a solution of PyBOP (2.73 mg, 5.26 mmol) in CH2Cl2 (12 mL). The mixture was kept in
an ice bath and stirred for half hour. After this period 5-methylthiazol-2-amine (599 mg, 5.26
mmol) was added and the mixture was allowed to warm up to room temperature. The reaction
was kept with stirring by 4 hours. The solution was diluted with water and the formed precipitate
formed was filtered and washed with water and MeOH. N-(5-Methylthiazol-2-yl)-4-oxo-4H-
1
chromene-2-carboxamide was obtained as a white powder (Yield: 48%). H NMR (CDCl3): δ=
2.39 (3H, d, J = 1.1 Hz, CH3), 7.04 (1H, s, H(3)), 7.32 (1H, q, J = 1.1 Hz, H(4’)), 7.56 (1H, ddd,
J = 8.1, 7.2, 1.1 Hz, H(6)), 7.83 (1H, d, J = 7.9 Hz, H(8)), 7.91 (1H, ddd, J = 8.6, 7.1, 1.7 Hz,
H(7)), 8.07 (1H, dd, J = 8.0, 1.4 Hz, H(5)), 13.18 (1H, bs, NH).13C NMR (CDCl3): δ= 11.4
(CH3), 111.7 (C3), 119.1 (C8, C3’), 123.8 (C4a), 124.9 (C5), 126.1 (C6), 135.1 (C7, C4’), 155.4
(C1’, C2, C8a, CONH), 177.4 (C4). MS/EI m/z: 288 (M+, 53), 286 (86), 258 (72), 230 (62), 230
(62), 173 (95), 145 (82), 68 (100).
Synthesis of N-(4,5-dimethylthiazol-2-yl)-4-oxo-4H-chromene-2-carboxamide (3)
To a solution of chromone-2-carboxylic acid (0.5 g, 2.63 mmol) in DMF (4 mL), POCl3 (241 l,
2.6 mmol) was added. The mixture was stirred at room temperature for 30 min, with in situ
formation of the corresponding acyl chloride. Then N,N-diethylpropan-2-amine (459 l, 2.63
mmol) and 4,5-dimethylthiazol-2-amine hydrochloride (433 mg, 2.63 mmol) were added. The
system was heated 120 °C for 5 min in a microwave apparatus. After, the mixture was diluted
with dichloromethane (20 mL), washed with H2O (2X10 mL) and with saturated NaHCO3
solution (2X10 mL). The organic phase was dried with Na2SO4, filtered and concentrated under
reduced pressure. The residue was purified by flash chromatography (50% AcOEt/n-hexane). N-
(4,5-Dimethylthiazol-2-yl)-4-oxo-4H-chromene-2-carboxamide as a yellow powder (Yield 40%).
1H NMR (CDCl3): δ= 2.26 (3H, s, CH3). 2.35 (3H, s, CH3), 7.27 (1H, s, H(3)), 7.44 (1H, d, J =
8.4 Hz, H(8)), 7.49 (1H, ddd, J = 8.0, 7.2, 1.0 Hz, H(6)), 7.76 (1H, ddd, J = 8.7, 7.2, 1.7 Hz,
H(7)), 8.23 (1H, dd, J = 8.0, 1.5 Hz, H(5)).13C NMR (CDCl3): δ= 11.0 (CH3), 14.3 (CH3), 113.3
(C3), 118.0 (C8), 122.0 (C3’), 124.4 (C4a), 126.2 (C5), 126.3 (C6), 134.9 (C7), 142.0 (C4’),
153.3 (C8a), 156.2 (C1’), 157.0 (C2, CONH) , 177.6 (C4). MS/EI m/z: 300 (M+, 100), 272 (17),
199 (27), 163 (31), 139 (76), 101 (41).

Synthesis of Ethyl 4-methyl-2-(4-oxo-4H-chromene-2-carboxamido)thiazol-5-carboxylate
(4)
To a solution of chromone-2-carboxylic acid (0.5 g, 2.6 mmol) in DMF (4 mL), POCl3 (241 l,
2.6 mmol) was added. The mixture was stirred at room temperature for 30 min, with in situ
formation the corresponding acyl chloride. Then ethyl 2-amino-4-methylthiazol-5-carboxylate
(490 mg, 2.6 mmol) was added. The system was heated 120 °C for 5 min in a microwave
apparatus. After, the mixture was diluted with dichloromethane (20 mL), washed with H2O
(2X10 mL) and with saturated NaHCO3 solution (2X10 mL). The organic phase was dried with
Na2SO4, filtered and concentrated under reduced pressure. The residue was purified by
crystallization
in
methanol/n-hexane.
The
ethyl-methyl-2-(4-oxo-4H-chromene-2-
1
carboxamido)thiazol-5-carboxylate was obtained as a yellow powder (Yield 43%). H NMR
(CDCl3): 1.39 (3H, t, J = 7.1 Hz, CH2CH3), 2.70 (3H,s,CH3), 4.35 (2H, q, J = 7.1 Hz,CH2CH3),
7.32 (1H, s, H(3)), 7.58 – 7.46 (2H, m, H(8), H(6)), 7.80 (1H, ddd, J = 8.7, 7.2, 1.7 Hz, H(7)),
8.25 (1H, dd, J = 8.0, 1.6 Hz, H(5)), 10.22 (1H, bs, NH). 13C NMR (CDCl3): δ= 14.3 (CH2CH3),
17.0 (CH3), 61.2 (CH2CH3), 113.9 (C3), 117.7 (C3’), 118.0 (C8), 124.4 (C4a), 126.3 (C5), 126.6
(C6), 135.2 (C7), 152.2 (C4’), 155.1 (C8a), 156.2 (C1’), 157.3 (C2), 157.9 (CONH), 162.2
(COOCH2CH3), 177.4 (C4). MS/EI m/z: 357.9 (M+, 100), 173 (98), 150 (49), 89 (96).
2.4. Measurements and characterization
2.4.1. Spectroscopy
The IR spectra were acquired in a Nicolet 6700 FT-IR spectrometer using potassium bromide
disk. NMR was recorded on a Bruker AMX 400 NMR spectrometer. NMR spectra were
recorded at room temperature in 5mm outside diameter (o.d.) tubes. Tetramethylsilane (TMS)
was used as internal standard, chemical shifts are expressed in ppm (δ) and J in Hz. Electron
impact mass spectra (EIMS) were carried out on a VG AutoSpec, (Fisons, Ipswich, United
Kingdom) instrument; the data are reported as m/z (percentage of relative intensity of the most
important fragments).
2.4.2. Single crystal X-ray data collection, structure solution and refinement
The crystals for diffractometry of compounds 1-3 were grown at room temperature by slow
vaporisation of CH2Cl2/n-hexane (1:1) solutions of the compounds, while 4 was crystallised from
a methanol/n-hexane (1:1) solution.

Crystallographic data were collected on a Rigaku Saturn724+, AFC12 Kappa 3 circle
diffractometer. The structure was solved using the following computer programs: CrystalClear
SM Expert 2.0 r13, Rigaku Corporation, Tokyo, Japan, SHELXS, SHELXL [17], OSCAIL [18],
PLATON [19], MERCURY [20], ShelxLe [21]. Detailed crystal data and structural refinement
parameters are summarized in table 1 of supplementary information. Compounds 1, 2 and 4
crystallised in spacegroup P21/c while 3 crystallised in the monoclinic C2/c space group.
Hydrogen atoms were refined as riding atoms with C—H(aromatic), 0.95Å N–H, 0.88Å with
Uiso = 1.2Ueq(C) and C—H(methyl), 0.98Å, with Uiso = 1.5Ueq(C). The H atoms attached to
the N2 atom and the methyl group were checked on a final difference Fourier map.
In 3 the highest difference map peaks are associated with the S atom. These crystals were very
thin plates and although the Rint value was 0.057 the refinement gave high R and weighted R
values, Table 1 supplementary information, as a result of the measurement a large number of
very weak reflections. The crystals for 4 were very small; accordingly, many high angle
reflections were weak. This resulted in a data set with a high Rint values.
2.4.3. Gaseous phase quantum chemical calculations
Ab initio geometry optimisations for four gas-phase conformers of compound 1 (–anti rotamer
conformations A1 and A2, as set in figure 10, with the nitrogen atoms of the amide and thiazole
ring –trans related and –cis related) were performed using density functional theory. The
quantum chemical calculations were performed at the B3LYP exchange correlation functional,
which combines the hybrid exchange functional of Becke [22] with the gradient-correlation
functional of Lee et al. [23] and the split-valence polarised 6- 311+G(d,p) basis set [24] level of
theory. The GaussView 3.0 [25] program was used to get visual animation and the normal
vibrational modes descriptions. At the same theoretical level, analytical frequency calculations
were performed to ensure true minima (Nimg = 0). All theoretical calculations were performed
with the GAUSSIAN 03 program package [26]. The bonding characteristics of the compounds
studied were investigated using natural population (NPA) analysis of Reed and Weinhold
[27,28]. Values for the atomic natural total charges have been retrieved.
3. Results and discussion
3.1. Chemistry

4-Oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides were synthesized by two
different processes (Scheme 1). In the first method, a one-pot condensation reaction, chromone-
2-carboxylic acid is activated by reaction with benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate (PyBOP) and the in situ formed intermediate react with the aminothiazole
or its derivatives giving rise to the intended chromone. In the second method the chromone
carboxamides have been obtained by a condensation reaction that requires the previous
activation of the chromone-2-carboxylic with phosphorus oxychloride and subsequent addition
of the aminothiazole derivative to the formed acyl chloride in a process assisted by microwave.
Moderate yields have been obtained by both methods. However, it must be stressed that PyBOP
reactions were more time-consuming and that the coupling reagent is quite expensive.
3.2. FTIR analysis
From the infrared spectra of the chromone-thiazole hybrids a characteristic band correspondent
to the N-H str (stretching) at 3349-3435 cm -1 was observed. The carbonyl group is confirmed by
a C=O str band at 1674 cm -1 for compounds 1 and 2 and 1683 and 1686 cm -1 for compounds 3
and 4, respectively. In addition, an N-H bending vibration of the amide group is also detected
-1
(1651-1637 cm ) in the infrared spectra of all compounds. The C=O bond of ester function
present in compound 4 is observed at 1708 cm -1. The FTIR data confirmed the formation of the
amide functional group generated in the amidation reaction of chromone carboxylic acid.
3.3. NMR analysis
The structural characterization of compounds 1-4 was achieved in all cases from NMR, ESI-
MS data, and by a single crystal structure determination. The numbering of chromone
carboxamide core used along the analysis is depicted in Figure 2.
The assignment of the protons of compound 1 was done using 1D and 2D 1H-NMR. The protons
at position H3 (δ= 7.05 as singlet), H5 (δ= 8.06 as double doublet), H3’ (δ= 7.38 as doublet) and
the amide proton (δ= 13.32 as broad singlet) were easily assigned by their chemical shifts (δ) and
1
multiplicities (Table 1). The other protons of chromone 1 were assigned by a H–1H COSY
(COrrelation SpectroscopY) experiment as it clearly indicate correlation with coupled that is
determined by cross peaks (correlation peaks). In the present case cross peaks between H5 and
H6 (δ= 7.55, ddd), H6 and H7 (δ= 7.90, ddd), H7 and H8 (δ= 7.84, dd) and H3’ and H4' (δ=
7.55, ddd) where observed (Figure 3). The assignment of the protons of the other chromones

were done by 1D NMR based on a comparison of chemical shifts (δ), multiplicities and the
structural similarity with compound 1 (Table 1): The protons of the chromone rings were
assigned as H3 (δ= 7.04-7.32), H5 (δ= 7.04-7.32), H6 (δ= 7.46-7.58), H8 (δ= 7.83-7.44) and H7
(δ= 7.90-7.76). The protons of the heterocyclic ring and substituents (R1 and R2) were also
allocated: the hydrogen at position 4’ of compound 2 was observed to δ= 7.64 ppm and the
methyl substituents of compounds 2-4 were observed at δ= 2.70- 2.26 ppm as a singlet. The ethyl
group of the compound 4 was detected as a triplet (δ= 1.39) and quadruplet (δ= 4.35). The
protons of the amide group were observed as a broad singlet (δ= 13.32-10.22), except the proton
of the compound 4 that was not detected in the present experimental conditions. The carbons
1
linked to the protons of compound 1 were assigned by heteronuclear NMR, namely H–13C
HSQC (Heteronuclear Single Quantum Coherence) spectroscopy experiments (Figure 4). Cross
peaks between the following protons and carbons were observed: H3 and C3 (δ= 111.8), H5 and
C5 (δ= 124.9), H6 and C6 (δ= 126.2), H7 and C7 (δ= 135.1), H8 and C8 (δ= 119.1), H3 ' and
C3’ (δ= 114.7) and H4 ' and C4' (δ= 135.1). The quaternary carbons at position C4a (δ= 123.8)
and C4 (δ= 177.4) were easily assigned by their chemical shifts. The other quaternary carbons of
the chromone were assigned by HMBC (Heteronuclear Multiple Bond Correlation) spectroscopy
experiment (Figure 5) where cross peaks between C1’ (δ= 159.5) and H3, H3’ and H4’ and
between C2, C8a, CONH (δ= 155.3) and H3, H5, H6, H7 and H8 were observed. The carbons
linked to the protons and the missing quaternary carbons were assigned by 13C NMR and DEPT
(Distortionless Enhancement by Polarisation Transfer) (Table 2): C2 (δ= 157.3-155.4), C3 (δ=
118.8-111.7), C4 (δ= 177.6-177.4), C4a (δ= 129.4-124.4), C6 (δ= 126.3-126.6), C5 (δ= 126.3-
124.9), C7 (δ= 135.2- 134.9), C8 (δ= 119.1-118.0), C8a (δ= 155.4-155.1), C1’ (δ= 156.2-155.3),
C3’(δ= 123.8-114.7), C4’ (δ= 152.2-135.1) CONH (δ= 157.9-155.3). The methyl substituents
were observed at a high field between δ= 11.0 and δ= 17.0 ppm. The methylenic and carbonyl
groups of the ester substituent present in compound 4 were detected at δ= 61.2 and 162.2 ppm,
respectively.
3.4. Molecular conformation
Thermal ellipsoid diagrams with the adopted atomic labelling are shown in Figures 6–9.
Compound 4 crystallised with two molecules in the asymmetric unit that are identified as
molecule #1 and #2.

The conformation around the C–N rotamer for chromone carboxamides can be either –anti or –
syn. The former conformation appears to be more probable in these compounds since it lowers
the steric hindrance between the two aromatic rings as compared to the –syn rotamer
conformation. Structural characterizations made previously in other 4H-chromene-2-
carboxamides [29] show that, when the amide oxygen atom is trans related to the pyran oxygen
atom the –anti conformation predominates since it permits the establishment of a short
intramolecular N---H...O(pyran) contact, as depicted in the A1 representation in Figure 10.
Compounds 1-4 follow this pattern as can be seen from figures 3-6: they present an hydrogen
bond
that
originates
a
S(5)
ring
[29,
30]. The values for parameters characterizing those intramolecular interactions are given in table
3. In all compounds the S atom of the thiazole ring is in a –cis position with respect to the
carbonyl O atom of the amide.
Relevant data for the discussion of molecular geometry and conformation is presented in table 4;
θChr-Thz refers to the dihedral angle between the mean planes of the chromone and thiazole rings,
θChr-amide to the dihedral angle between the best plane of the chromone and the plane defined by
atoms O2-C21-N2 of the amide moiety whereas θThz-amide refers to the dihedral angle between
this plane and the best plane of the thiazole ring. Values for dC2 and dC22 are the deviations of the
C2 and C22 atoms from the plane defined by the O2-C21-N2 atoms. The relative conformational
position of the heteroaromatic rings with respect to the O2-C21-N2 plane of the amide may be
evaluated according to the dC2 and dC22 values: when the vectors dC2 and dC22 have the same
directions as in 3 and in molecule #1 of compound 4 the molecule has a butterfly conformation
as seen in Figure 11. In 1 and 2 C2 is co-planar with the O2-C21-N2 plane. As said before,
dihedral θ angles may also be used as quantitative parameters for evaluation of the overall
conformation of those molecules. Since the heteroaromatic rings are practically planar, the
dihedral θChr-Thz quantifies the degree of bend and/or twist between them and can be used for
evaluation of the distortion of the molecule from planarity when one of the dihedrals, θChr-amide or
θThz-amide, is relatively small. This is the case for compound 4 where the deviation to planarity for
both molecules is mainly due to the torsion of the chromone residue. In 1 and 2 and since both
θChr-amide and θThz-anide assume values significantly different from zero, the conformation of the
molecule may be better analysed looking at those values. Since C2 is co-planar with the O2-C21-
N2 plane in both compounds, they assume a conformation where the chromone rings are
predominantly twisted relatively to the amide plane while in 3 the heteroaromatic rings are
predominantly bent with respect to that plane giving the butterfly conformation, as can be seen in

Figure 8. Those observations suggest that these molecules may have several energetically
accessible conformations that can be present in the biological environment indicating a degree of
flexibility for the docking with substrates.
3.5. Supramolecular structures.
Values for intermolecular H bonds and contacts are presented in Table 4. In all compounds the
main feature is the formation of an intermolecular N2—H2···O4 hydrogen bond which links the
molecules into C(7) chains [30] that run parallel to the c-axis an example, compound 1, is shown
in Figure 12. These bonds are supplemented by weak C–H···O interactions as shown in Figure
13. The symmetry codes are inserted in Table 4. Detailed discussion of the supramolecular
structures for all compounds is given in S2 Supplementary Information.
3.6. Gaseous phase quantum chemical calculations
Adiabatic quantum chemical calculations were performed in order to get some inside concerning
the configurations and conformations for those componds as given by X-ray crystallography
results. Geometric optimisations of conformers of compound 1 were made with different starting
geometries that were identified as A1_0, A1_1, A2_0 and A2_1, Figure 10. A1 and A2 refer to
the –anti rotamer configurations; depending on the rotation of the thiazole ring, the oxygen atom
of the amide and the nitrogen atom of the thiazole may be -trans related (A1_0 and A2_0) or -cis
related (A1_1 and A2_1).
The total electronic energies, at the B3LYP/6-311+G(d,p) level of theory, are given in
Supporting information S3.The zero-point energy correction was scaled using a 0.9887 scaling
factor [32]. The cartesian coordinates as obtained from quantum chemical gas phase geometry
optimisations, at the B3LYP/6-311+G(d) level of theory, are given in Table Supporting
information S3. The geometries obtained after optimisations are depicted in Figure 10.
Conformers A1 optimized to the same final geometry as can be seen in figure 10. This conformer
as obtained from geometry optimisation is the most stable in gas phase taking the B3LYP/6-
311+G (d,p) level of theory. The geometry obtained is similar to that presented in the solid state
given by single crystal diffractometry. The molecule is mostly planar with dihedral angles
between the amide moiety and the chromone/thiazole rings being close to 180º. Opposed to this,
the geometrical optimisation made for the conformers A2 show two distinct local minima that
present torsions of the heterorings around the amide rotamer. In conformer A2_0 the dihedral

angle between the chromone ring and the amide moiety, θChr-amide, is of 28º and the dihedral angle
between the thiazole and the amide spacer, θThz-amide, is of 1º showing that loss of planarity is due
to the twist of the chromone ring with respect to the amide. In conformer A2_1 those dihedrals
are of 30º and 61º respectively pointing out for significant torsions of both rings.
Furthermore, in order to obtain the natural atomic charges, a population analysis was performed,
using the natural bond orbital (NBO) analysis [27,28]. Natural atomic charges represent nuclear
charges minus the summed natural populations of the natural atomic orbitals on the atoms and
characterize the ground electronic state of the molecule. The calculated charges potentially give
some insight into the electronic distribution around the molecule and, thus, contribute to an
understanding of the preferred geometries for compound 1. The charge distributions for all the
optimisations are shown in figures in Supporting Information S4 and are presented within a
relative charge range of -1.000 to +1.000. Figure 14 shows some charge values for the most
stable configuration, A1; negative charges are located at the two N atoms and at the three
oxygen atoms while the positive charges reside at the amide carbon as well as at the sulphur
atom of the thiazole ring. The same qualitative pattern is observed in the other conformers A2.
The geometry obtained for conformation A1 allows the minimisation of steric hindrance between
the oxygen atoms of the amide and the pyrone in the planar geometry while conformers A2 does
not. In addition, conformation A1 is probably further stabilized by the cis related nitrogen where
the sulfur is pointing to the carboxyl oxygen atom of the amide. In A2 the torsion of the
chromone ring with respect to the amide permits a minimisation of the steric hindrance between
the carboxylic amide oxygen atom and the pyrone oxygen atom and in A2_1 the torsion of the
thiazole ring is probably related with the minimisation of the of the hindrance between the two
nitrogen atoms since the stabilisation of the geometry in which the thiazole ring is planar with
the amide is not present due to the position of the positive charged sulphur atom that is far from
the range field of the negative charged atoms.
4. Conclusion
New 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides were synthesized from
chromone-2-carboxylic acid by two different amidation methods. The development of dissimilar
synthetic approaches provided the possibility of working with diverse reaction conditions,
namely with conventional heating and/or microwave irradiation. Relevant data related to the
molecular geometry and conformation of the chromone-thiazole hybrids has been acquired
which can be of the utmost importance to understand ligand-receptor binding.

Acknowledgements: The authors thank the Foundation for Science and Technology (FCT) of
Portugal (PEst-C/QUI/UI0081/2013). F. Cagide (SFRH/BPD/74491/2010) is supported by FCT
grant. Thanks are also due to the staff at the National Crystallographic Service, University of
Southampton for the data collection, help and advice [31].

Scheme 1. Synthesis of 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides.

Figures and captions
R1
O
S
R2
O
O
N
N
1 R1= H; R2=H
H
2 R1= CH3; R2=H
3 R1= CH3; R2=CH3
4 R1= COOEt; R2=CH3
Figure 1 – Chemical structure of the 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-
carboxamides.
Figure 2 – Numbering of 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamide core.
H3’
H4’
H3
H6
H7 H8
H5
H3
H3’
H6
H4’
H8
H7
H5
Figure 3 – 1H–1H COSY spectrum of compound 1

H3
H4’
H3’
H7 H8
H6
H5
C3
C3’
C8
C5
C6
C7, C4’
Figure 4 – 1H–13C HSQC spectrum of compound 1
H3
H4’
H3’
H6
H7 H8
H5
C4a
C2, C8a, CONH
C1’
C4
Figure 5 – 1H–13C HMBC spectrum of compound 1

Figure 6 - Thermal ellipsoid (70%) plot of 4-Oxo-N-(thiazol-2-yl)-4H-chromene-2-carboxamide
(1)
Figure 7 - Thermal ellipsoid (70%) plot of N-(5-methylthiazol-2-yl)-4-oxo-4H-chromene-2-
carboxamide (2)
Figure 8 - Thermal ellipsoid (70%) plot of N-(4,5-dimethylthiazol-2-yl)-4-oxo-4H-chromene-2-
carboxamide (3).

Figure 9 - Thermal ellipsoid (70%) plot of ethyl-4-methyl-2-(4-oxo-4H-chromene-2-
carboxamido)thiazole-5-carboxylate (4).

Figure 10 – Possible –anti rotamer conformations for the studied compounds. A1- with the
amide oxygen atom –trans related with the pyrone oxygen atom; A2 – with the amide oxygen
atom –cis related with the pyrone oxygen atom; depending on the rotation of the thiazole ring,
the oxygen atom of the amide and the nitrogen atomof the thiazole may be -trans related (A1_0
and A2_0) or -cis related (A1_1 and A2_1). The presented compounds present the A1_0 –anti
conformation allowing for the establishment of a short intramolecular N---H...O(pyran) contact.
Molecular representations are referred to the conformations obtained from quantum chemical gas
phase geometry optimisations at the B3LYP/6-311+G(d) level of theory.

Figure 11– A perspective view of the molecules 1–3 (from top to bottom) through the amide
plane depicting the differences in conformation of the molecules that are given by the twist
and/or bent of the heteroaromatic rings in relation to the amide plane. In molecules 1 and 2 the
rings are predominantly twisted while molecule 3 they are predominantly bent to the same
direction conferring a “butterfly” conformation to the molecule.
Figure 12 – Representation of the C(7) chain running along the c-axis for 1. Compounds 2 and 3
make similar chains. The hydrogen atoms not involved in the connectivity were omitted for
clarity.

Figure 13 – Representation of the C(8) chain parallel to the b-axis made by the weak C5—
H5···O2 hydrogen contact for compound 1. Compounds 2 and 3 exhibit similar patterns. The
hydrogen atoms not involved in the connectivity were omitted for clarity.
Figure 14 - Natural atomic charges from population analysis (NBO), at the B3LYP/6-311+G(d)
level of theory for the most stable configuration, A1. The charge distributions are presented
within a relative charge range of -1.000 to +1.000.

Tables and captions
Table 1: 1H-MNR data of the 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides
Compounds
H3
H5
H6
H7
H8
R1
R2
NH
7.05
8.06
7.55
7.90
7.84
7.38
7.64
13.32
1
(s)
(dd)
(ddd)
(ddd)
(dd)
(d) H
(d) H
(bs)
7.04
(s)
8.07
(dd)
8.23
(dd)
8.25
(dd)
7.56
(ddd)
7.49
7.91
(ddd)
7.76
7.83
(dd)
2.39
(d) CH3
7.32
(q) H
2.26
13.18
(bs)
-
2
3
4
7.27
(s)
7.44
2.35
(ddd)
7.58-7.46
(m)
(ddd)
7.76
(dd)
(s) CH3
(s) CH3
2.70
7.32
(s)
7.58-7.46
(m)
1.39
10.22
(ddd)
(t) COOCH2CH3
4.35
(s) CH3
(bs)
(q)COOCH2CH3

Table 2: 13C-MNR data of the 4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides
Compounds C2
C3
C4
C4a
C5
C6
C7
C8
C8a
C1’
C3’
C4’
R1
-
R2
-
CONH
155.3
1
2
155.3 111.8 177.4 123.8 124.9 126.2 135.1 119.1 155.3 159.5 114.7 135.1
155.4 111.7 177.4 123.8 124.9 126.5 135.1 119.1 155.4 155.4 119.1 135.1 11.4 CH3
-
155.4
14.3
CH3
3
4
157.0 113.3 177.6 124.4 126.2 126.3 134.9 118.0 153.3 156.2 122.0 142.0 11.0 CH3
157.3 113.9 177.4 124.4 126.3 126.6 135.2 118.0 155.1 156.2 117.7 152.2 14.3
157.0
157.9
17.0
CH3
COOCH2CH3
61.2
COOCH2CH3
162.2
COOCH2CH3

Table 3: Selected hydrogen-bond parameters
D—H···A
D—H (Å)
H···A (Å)
D···A (Å)
D—H···A (°)
1
N2—H2···O1
N2—H2···O4i
C5—H5···O2ii
C7—H7···N23iii
C7—H7···S21iv
2
0.89 (3)
0.89 (3)
0.95
2.30 (3)
2.01 (3)
2.33
2.652 (4)
2.887 (3)
3.244 (4)
3.266 (4)
3.628 (3)
103 (2)
165 (3)
161.2
0.95
2.59
128.8
0.95
2.81
144.6
N2—H2···O1
N2—H2···O4v
C5—H5···O2vi
3
0.866 (19)
0.87 (2)
0.95
2.362 (19)
2.03 (2)
2.34
2.6751 (18)
2.8835 (17)
3.276 (2)
101.7 (15)
169.2 (18)
166.6
N2—H2···O1
N2—H2···O4vii
C8—H8···O4vii
4
0.82 (9)
0.82 (9)
0.95
2.29 (8)
2.20 (9)
2.57
2.627 (5)
3.012 (5)
3.348 (6)
105 (6)
171 (8)
138.9
N12—H12···O11
N12—H12···O24
0.82 (5)
0.82 (5)
2.39 (5)
2.12 (6)
1.97 (5)
2.32 (4)
2.48
2.687 (6)
2.934 (7)
2.923 (6)
2.686 (6)
3.432 (7)
3.405 (8)
103 (4)
174 (5)
169 (4)
102 (3)
175.6
N22—H22···O14viii 0.97 (5)
N22—H22···O21 0.97 (5)
C15—H15···N223ix 0.95
C25—H25···N123 0.95
2.52
154.4
Symmetry code(s): (i) x, -y+1/2, z-1/2; (ii) -x+1, y-1/2, -z+3/2; (iii) -x+1, y-1/2, -z+1/2; (iv) x-1, -y+1/2, z-1/2; (v) x, -y+3/2,
z+1/2; (vi) -x+1, y+1/2, -z+1/2; (vii) x, -y+1, z+1/2; (viii) x, y+1, z; (ix) x, y-1, z;

Table 4: Selected dihedral angles and distances for compounds.
θºChr-Thz
θºChr-amide
θºThz-amide
dC2 (Å)
dC22 (Å)
8.64(15)
12.30(8)
14.4(2)
13.4(5)
17.4(2)
7.1(4)
7.9(5)
6.2(2)
11.5(4)
3.9(8)
2.9(8)
-0.001(2)
0.0046(16)
0.053(5)
-0.073(2)
-0.0716(16)
0.084(5)
1
2
3
4 #1
4 #2
17.75(16)
15.6(2)
14.4(6)
18.3(6)
0.026(5)
0.062(5)
-0.060(5)
0.023(5)

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Graphical abstract
Highlights
 Cromones as potential adenosine receptor ligands
 Novel
synthesized
Compounds were characterized by NMR spectroscopy and X-ray crystallography.
4-oxo-N-(substituted-thiazol-2-yl)-4H-chromene-2-carboxamides
were
