Microwave-assisted synthesis of new imide-and formamide-derivatives of 2(3 H )- benzoxazolinones and 2(3 H )-benzothiazolinones

With the aim to obtain 2(3 H )-benzoxazolinones and 2(3 H )-benzothiazolinone derivatives, we synthesized new imide compounds 5a-c , 6c-d and 7a-d by treating their 6-amino precursors with succinic, maleic or phthalic anhydride. The microwave (MW)-assisted reaction provided them more efficiently than through the corresponding isolated amic acids. The same nucleophile reagents were used in a MW-assisted formylation to obtain new formamide products 8-11 , that were detected as cis/trans forms by NMR analysis. The ratio was in favor of the cis isomer and affected by the solvent, as also supported by DFT calculations.


Introduction
Molecules bearing a 2(3H)-benzoxazolinone or 2(3H)-benzothiazolinone moiety are considered privileged scaffolds in drug design according to their chemical properties and reactivity. 1 They have shown a high flexibility in chemical modifications, allowing changes on characteristic side-chains anchored to the rigid scaffold, able to provide molecules with interesting therapeutic applications.Several potentially useful compounds based on these pharmacophores were developed, producing molecules with a broad spectrum of bioactivities which include analgesic and anti-inflammatory, [2][3][4] antioxidant, 5 cytotoxic [6][7] and antinociceptive 8 properties.
Substituted derivatives are reported as scaffolds of non-steroidal anti-inflammatory drugs (NSAID), based on an observed analgesic activity.For example, tiaramide (1-[2-(5-chloro-2-benzothiazolinone3-yl) acetyl]-4hydroxyethyl piperazine) is a drug that prevents the synthesis of prostaglandins and therefore has a strong anti-inflammatory activity. 9Quantum chemical calculations were applied to a small library of 6acylbenzothiazolone derivatives, with the aim to establish a structure-activity relationship useful for the design of new analgesic drugs. 10,11ecently, a series of urea and thiourea derivatives of 3-methyl-2(3H)-benzothiazolone have been evaluated for antibacterial and antifungal activities. 12Besides 2(3H)-benzoxazolinones and 2(3H)-benzothiazolinones with a N-methyl group, the corresponding N-H heterocycles find a particular interest in medicinal chemistry as precursors of differently N-functionalized analogues. 13rocedure where the conventional heating is replaced by microwave (MW) irradiation have been widely applied in organic synthesis, due to its general high efficiency in terms of reduced reaction time and increased product yield, eco-friendly nature by possible solvent-free conditions, and experimental ease of manipulation. 14,15][18] The study on formylation of the amine group has attracted attention as a reaction with a biological relevance.It occurs on amino acids and was identified to be involved in the initiation of protein synthesis in bacteria 19 and recently in a histone modification which may modulate gene expression. 20The isomerism of the amide group is of some interest due to its partial double bond character and has been studied in order to understand protein structure. 21e report here on the MW-assisted synthesis of new 2-benzoxazolinone and 2-benzozathiazolinone derivatives, bearing either a HN(3) or a methyl-N(3) group, and functionalized at the 6-amino group as imide or as formamide.In the latter case, insights into the molecular structures of the isomeric products by a formylation reaction have been taken into account by NMR analysis and density functional theory (DFT) calculation.

Results and Discussion
In the context of bioisosterism, based on the hypothesis that molecules with similar electronic structures tend to exert similar biologic activities, 6-aminobenzoxazolinone and 6-amino-benzothiazolinone are promising precursors for the production of new O/S isosters with potential improved bioactivities.Both 6-amino-2(3H)benzoxazolinone (1a) and 6-amino-2(3H)-benzothiazolone (1c) are commercially available, whereas the Nmethyl analogues 1b and 1d can be obtained starting from 3-methyl-2(3H)-benzoxazolone or from the corresponding thia-compound after aromatic nitration with HNO3/ Ac2O at 0-5 °C and reduction with SnCl2•2H2O under reflux in ethanol according to the reported procedure. 22he series of amic acids 2a-d, 3c-d and 4a-d were obtained treating each of the corresponding 6aminobenzoxazolinones 1a, 1b or 6-aminothiazolinones 1c, 1d with succinic, maleic or phthalic anhydride in THF at room temperature for 1-2 hours (Scheme 1).After partial solvent concentration, the products were isolated in good yields by precipitation from the reaction mixture and with a good purity as established by NMR and MS analysis.In details, ESI-MS/MS experiments on [M-H] -ions gave fragments corresponding to the loss of carbon dioxide and water, whereas electron impact MS(EI) spectra showed intense signals due to the loss of water from the corresponding undetectable molecular peak.This fact can be attributed to the production of imide derivatives (reported below) under the high temperature conditions applied in the measurement.
The presence of a carboxylic group on these molecules offers the opportunity to further functionalize them.The benzoxazolinones 2a has been reported, 23 and both 2b and 3b are now commercially available.By heating the amic acids 2a-d, 3c-d and 4a-d at 80 °C for one hour in acetic anhydride with an equimolecular amount of sodium acetate, 24 we have obtained a series of N-arylimide derivatives ( 5a-d, 6c-d and 7a-d).
MW irradiation of the 6-amino aryl reagent 1a-d with the suitable anhydride in TFA at 73 °C for 15 minutes provided the corresponding imide products in higher yields if compared with the two-step conventional method.In detail, by a single step and shorter reaction time, starting from 1c-d, the imides 5c-d were obtained by reaction with succinic anhydride, 6c-d by reaction with maleic anhydride and 7b-d by reaction with phthalic anhydride (Scheme 1).The products were isolated in solid form and were structurally characterized by NMR analysis and mass spectrometry (Experimental Section).
This one-pot production of N-arylimide derivatives is less time-consuming and more efficient since it gives higher yields (86-91%) compared to the two-step method where the amic acids obtained at room temperature are subsequently cyclized upon conventional heating, with global yields in the range 57-69% starting from 1ad (Scheme 1).
1 HNMR spectrum of compound 8 in CDCl3 shows the presence of two related molecules, in particular we found pairs of signals as doublets centered at δ 8.57 (J 11.6 Hz) and δ 8.37 (J 1.6 Hz) for the formyl proton, broad singlets at δ 7.70 and δ 7.21 for the NH amide proton and singlets at δ 3.45 and δ 3.43, all these in a 1: 2 ratio, respectively.The phenomenon is due to the partial single-double bond feature of the HN-CHO amide bond that restricts the rotation around this bond so that the cis and trans forms are both possible when referring to the relative position of formyl and secondary amide hydrogen atoms,which correspond to Z and E isomers, respectively.
The coupling constant values 11.6 and 1.6 Hz for the formyl protons allow the assignments of the signals to the trans and cis isomers, respectively.This result is in agreement with reported data for N-methylformamide and N-phenylformamide, 25 as well as for N-formyl-o-toluidine and a series of strictly related Nmonoalkylsubstituted formamides. 26Further support to these assignments comes from the resonance of the trans-C(O)H proton at lower fields than for the cis form, attributable to the closer proximity of the trans -C(O)H proton to the phenyl ring that produces a larger ring-current. 23The downfield shifts for H-5 (δ 7.37) and H-7 (δ 7.84) protons observed in trans form compared with the signals (δ 7.06 and 7.20, respectively) for the cis isomer are also in agreement with the behavior of the corresponding aromatic proton in ortho-position reported for cis/trans forms of Nformyl-o-toluidine. 26 It was also observed that the amide NH signals shifted depending on the concentration of 8 in CDCl3 solution, so that the detectable signals at δ 7.70 for the cis (br d, J 11.6 Hz) and at δ 7.31 (br s) for the trans isomers moved to δ 7.76 and 7.83 respectively in an about four times more concentrated solution.This solvent effect is compatible with the increase of solute-solute hydrogen bonding in more concentrated solutions. 25Neither shifts for HC(O) proton nor isomer ratio change was instead observed upon dilution or concentration of the solution.
The isomeric ratio resulted sensitive to the type of solvent used for the NMR analysis, in fact, replacing CDCl3 with CD3OD, the trans/cis ratio moved to 1:5, with the cis form predominant also in this case(Table 1).We could attribute the signalsat δ 8.25 and δ 8.64 to the trans and cis forms respectively, assuming also here the aforementioned downfield shift of the trans-formyl proton due to the closer proximity to the phenyl group.This assignment finds a support in DFT-minimized structures, where the 2.23 Å distance was evaluated between H-5 and amide oxygen.In addition, as expected, the signal for HC(O) protons became singlets due to the H/D exchange on NH position.The presence of trans/cis forms is also confirmed by the doubling of the signals for all the carbon atoms observed in 13 C NMR spectra.In details, the values for the trans isomer are at lowerfields if compared to those corresponding to the cis isomer, which are in a higher proportion, in agreement with reported data. 26Similar results were found for the 3-methyl benzoxazolylformamide 9 (Exper.Section).
Due to the poor CDCl3 solubility observed for the more polar products 10 and 11, their NMR spectra were recorded in CD3OD.Also for these compounds the presence of two forms was shown, with the CHO signals as singlets at δ 8.26 and 8.62 in 5:2 ratio for 10 and at δ 8.24 and 8.61 in 4:1 ratio for 11, attributable to each cis/ trans pair respectively, based on the downfield position of doublet for the minor trans isomer in CDCl3 spectra of 8 and 9.In addition, the cis form resulted the major one also in DMSO-d6, where the 3:1 cis/trans ratio evaluated for 11 was based on the relative intensities of the corresponding CHO signals at δ 8.23 as a broad singlet and at δ 8.63 as a doublet (J 11.5 Hz), (Table 1).A confirmation to these data came from energy-minimized structures obtained by DFT calculation.Phenyl and substituted-phenylformamides have been studied as simple models for peptide-like configurations 27,28 with different basis sets tested in the investigation of small model amide, e.g.N-methylamide.The efficacy of the methods was reported comparing the calculated amide frequencies with those measured in solution by IR spectroscopy.This allowed to select BPW91 hybrid functional with a small basis set as the best method to predict the amide frequencies both in gas-phase and in solution with a good approximation. 29,30In our hands, DFT (BPW91) method also resulted the best approach when applied to arylformamides 8-11, with 6-31G (d,p) as the basis set of choice for all atoms, except for sulfur atom in molecules 8 and 11, where 6-311+(3df,p) basis set was used.Energy-minimized structures for the isomeric forms of compound 8 display lengths of 1.37 Å for N-CO and 1.23 Å for C=O bonds.The cis isomer presents a full planar disposition, whereas the trans isomer presents a dihedral angle of 31.9° between the plane containing the aryl group and the one containing the formyl unit (Figure 2).A similar behavior has been also observed for the cis/trans forms of 9-11, where the cis isomer always displays a planar structure.The calculated energy values obtained for the minimized structure of formamide 8 in chloroform solution confirmed that the cis form is more stable than the trans, in agreement with the 2:1 ratio observed in CDCl3 by 1 H NMR spectrum.Besides this, the highest amount of the cis over the trans form observed in 1 HNMR spectra recorded for each formamide 8-11 in CD3OD can be correlated to their higher stability, as established by DFT calculation in methanol solutions(Table 2).The N(3)-demethyl products 10 and 11 present the possibility of different tautomeric forms, among which the most probable are the 2-hydroxybenzoxazolyl and 2-hydroxybenzthiazolyl ones (Figure 1).For the four isomers taken into account in DFT calculation for molecules 10 and 11, the cis form of the 2-oxo tautomer resulted the most stable (Table 2).In addition the high energy values obtained for the cis and trans 2hydroxytautomers are in line with the not appreciable presence of the corresponding signals in NMR spectra.

Conclusions
6-Amino functionalized 2(3H)-benzoxazolinones and 2(3H)-benzothiazolinones and their N(3)-methyl derivatives have been converted into the corresponding imide derivatives upon treatment with succinic, maleic or phthalic anhydrides by one-pot MW-assisted reactions.This procedure is more efficient compared with the two-step method where the amic acids are produced at room temperature and subsequently cyclized upon conventional heating.The same aryl amines were also employed as nucleophilic reagents in a MWassisted formylation to give the corresponding formamide derivatives.We investigated the cis/trans ratio of these products by both NMR analysis (under different concentration and solvent conditions) and by considering their minimized energy structures obtained by DFT calculations.

Experimental Section
General.Melting points were determined using a Reichert Thermovar apparatus.Infrared spectra were recorded by using a FT-IR Tensor 27 Bruker spectrometer equipped with an Attenuated Transmitter Reflection (ATR) device at 1 cm −1 resolution in the absorption region Δν 4000-1000 cm −1 .A thin solid layer was obtained by evaporation of the methanol solution of the sample.The instrument was purged with a constant dry nitrogen flow.Spectral processing was made using the Opus software package.NMR spectra were recorded by an Avance 400 Bruker spectrometer: 1 H at 400 MHz and 13 C at 100 MHz in CDCl3 or CD3OD; δ values are reported in ppm relative to the solvent residual signals = 7.25 and δC = 77.00ppm for CDCl3, δH = 3.31 and δC = 49.00 for CD3OD, δH = 2.49 and δC = 39.50 ppm for DMSO-d6 relative to SiMe4 (=0 ppm); J values in Hertz.
Assignments are supported by heteronuclear single quantum correlation (HSQC) and heteronuclear multiple bond correlation (HMBC) experiments.Electron impact (EI) mass spectra (m/z; rel.%) and high-resolution EI data performed on a Kratos-MS80 mass spectrometer with a home-built computerized acquisition software.Electrospray ionization (ESI)-MS mass spectra and tandem fragmentation spectra (MS/MS) were taken with a Bruker Esquire-LC spectrometer equipped with an electrospray ion source used in positive or negative ion mode by direct infusion of a methanolic solution of the sample, under the following conditions: source temperature 300 °C, drying gas N2, 4 Lmin -1 , scan range m/z 100-1,000.High-resolution ESI MS measurements for compounds 2-4 were obtained by direct infusion using an Orbitrap Fusion Tribrid ® mass spectrometer.Solvents and reagents were purchased from Sigma Aldrich Europe or Alfa Aesar and were used without purification.Thin-layer chromatography (TLC) was carried out on Merck Kieselgel Si60 PF254 and spots visualized at 254 nm.Preparative TLC (PLC) was realized on 20 × 20 cm Merck Kieselgel 60 F254 0.5 mm plates.MW-assisted reactions were carried out using a mono-mode CEM Discover reactor in a sealed vessel, monitoring the temperature of reaction mixtures by an external infrared control system.All evaporations were carried out at reduced pressure.Products 2a-2d, 3c-3d, 4a-4d were recrystallized from methanol and imides 5a-5d, 6c,6d and 7a-7d from hexane/ethyl acetate before carrying out analysis for structural characterization.
Computational details.Starting structures were generated by molecular mechanics minimization, using PC Model (Serena Software, version 7.00).Quantum chemical calculations were performed using the Gaussian 03W revision E.01 package program set. 31 The 6-31G (d,p) was the standard basis set of choice for all atoms of the first and second row, whereas 6-311+(3df,p) basis set was used for sulfur atom.The gradient-corrected DFT was BPW91 method which uses the GGA (Generalized Gradient Approximation) exchange functional of Becke 32 and the PW91 GGA correlation functional of Perdew and Wang. 33The functional was parameterized using the Monte Carlo results of Ceperley and Alder. 34Perdew and Wang considered all the Ceperley and Alder data, including the uncertainty in each data point.They fit the functional to match the correct low-density behavior for the correlation. 33The simulations were made in the presence of methanol or chloroform using the polarizable continuum model (PCM).The optimized structural parameters were employed in the vibrational energy calculations at the DFT levels to characterize all stationary points as minima.For each optimized structure, no imaginary wavenumber modes were obtained, proving that a local minimum on the potential energy surface was actually found.stirred at room temperature for 1-2 hours.The progress of the reaction was monitored by TLC (ethyl acetate/hexane 6:4 v/v).Once the reaction was completed, the solvent was evaporated under reduced pressure to obtain the corresponding solid amic acids (53.9 mg, yield: 90%).

Figure 2 .
Figure 2.Energy-minimized structures of cis (left) and trans(right) forms for formamide 8, calculated at the BPW91/6-31G (d,p) level of theory in methanol solution.

Table 2 .
Data by DFT calculation at the level of theory BPW91/6-31G (d,p) for formamides 8-11, added of BPW91/6-311+(3df,p) for sulfur atom in 8 and 11: relative energy values for the trans-minimized structures with respect to the corresponding cis-form (taken as reference =0 KJ/mol) and dihedral angles (in degrees) between the aromatic and formamide units, in the indicated solvents.