Synthesis and Antibacterial Activity of New Spiro[thiadiazoline-(pyrazolo[3,4-d]pyrimidine)] Derivatives

El Fal, Mohammed; Ramli, Youssef; Zerzouf, Abdelfettah; Talbaoui, Ahmed; Bakri, Youssef; Essassi, El Mokhtar

doi:https://doi.org/10.1155/2015/982404

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Research Article | Open Access

Volume 2015 | Article ID 982404 | https://doi.org/10.1155/2015/982404

Synthesis and Antibacterial Activity of New Spiro[thiadiazoline-(pyrazolo[3,4-d]pyrimidine)] Derivatives

Mohammed El Fal,¹Youssef Ramli,²Abdelfettah Zerzouf,³Ahmed Talbaoui,⁴Youssef Bakri,⁴and El Mokhtar Essassi¹

Academic Editor: Marco Radi

Received24 Feb 2015

Revised16 Apr 2015

Accepted20 Apr 2015

Published13 May 2015

Abstract

New heterocyclic compounds spiroderivatives of allopurinol of biological interest were prepared from allopurinol via thionation and 1,3-dipolar cycloaddition and were produced in high to excellent yields. These compounds were characterized on the basis of spectral and spectroscopic data (¹H NMR, ¹³C, IR, and MS). The antibacterial activity of the synthesized products was studied using bacterial strains: Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, and Pseudomonas aeruginosa. Compounds having an ethyl group showed the best activity with MIC value of 31.25 µg/mL against Staphylococcus aureus and Streptococcus fasciens.

1. Introduction

Heterocycles are widely distributed in nature and play a key role in the metabolism of all living cells. Among the many heterocyclic compounds containing nitrogen, the pyrazolopyrimidine ring is very interesting and versatile scaffold for the synthesis of potential drugs or molecular tools (Figure 1). Among their many applications, pyrazolo,4-d]pyrimidines (Figure 2) were used as inhibitor kinases [1, 2] antiviral agents [3, 4], adenosine antagonists [5–7], glutamate modulators [8], antituberculosis agents [3], and as antibiotics inhibit bacterial growth [9, 10].

In light of the great importance of spirocyclic systems containing one carbon atom common to two rings [11–13], in recent years efforts have been made in developing methodologies for the synthesis of these compounds especially by cycloaddition reactions [14–22].

1,3-Dipolar cycloaddition is a subject of intense research during the last decade due to its great synthetic value. The cycloaddition is a method of synthesis of five membered heterocycles, which are difficult to prepare by other means.

Generally, reactions of 1,3-dipoles with true heterocyclic thiones having the thione form A proceed via 1,3-dipolar cycloaddition to the C=S double bond to form the spirocycloadducts, namely, spirothiadiazoles. The reaction of heterocyclic thiones A with nitrilimines, generated in situ by base-catalyzed dehydrohalogenation of hydrazonoyl halides, has been described for synthesis of various derivatives of spiro[heterocycle-n,2′-3H-1,3,4-thiadiazole] B (Figure 3).

In continuation of our work on the synthesis of the excess of allopurinol [23–28], compounds 2 and 2a-b have been synthesized by using reported methods [29–31]. Herein we report simple efficient synthesis of spiro thiadiazoline-(pyrazolo[3,4-d]pyrimidine) derivatives 4a-b by 1,3-dipolar cycloaddition of diphenyl hydrazonoyl chloride 3 with an equimolecular amount of pyrazolo[3,4-d]pyrimidin-4(5H)-thione derivatives 2a-b. All the synthesized compounds were evaluated for their antibacterial activity.

2. Materials and Methods

Generally the melting points were taken on an electrothermal capillary melting point apparatus. Infrared spectra (ν-cm⁻¹) were recorded on a Perkin Elmer 577, using KBr disks. ¹H NMR and ¹³C NMR spectra were recorded on Bruker Avance 300 NMR Spectrometer in DMSO-. Spectra were internally referenced to TMS. Peaks are reported in ppm downfield of TMS. Mass spectra are recorded in a SYNAPT G2 HDMS (Waters) Spectrometer in electrospray ionization (ESI).

2.1. Thionation

3.67 mmol of pyrazolo[3,4-d]pyridine is refluxed in pyridine with 3.67 mmol of phosphorus pentasulfide for 4 hours. Then the solvent is evaporated under reduced pressure, and the precipitate formed is washed with hot water to remove residual dimerized P₂S₅ until there is colorless filtrate.

1H-Pyrazolo[3,4-d]pyrimidine-4(5H)-thione (2). Yield = 90%; mp: 151°C. IR: cm⁻¹¹H NMR (DMSO-): δppm: 8.11 (1H, s, CH); 8.22 (1H, s, CH); 13.47 (1H, s, NH); 13.92 (1H, s, NH). ¹³C NMR (DMSO-) δppm: 105.05; 134.05 (Cq); 150.77 (CH); 151.20 (CH); 156.79 (Cq C=S). HRMS (ESI) [M + H]: m/z = 153.17.

1,5-Diethyl-1H-pyrazolo[3,4-d]pyrimidine-4(5H)-thione (2a). Yield = 75%; mp: 150°C. ¹H NMR (DMSO-) δ ppm: 1.3 (3H, t, Hz. CH₃); 1.36 (3H, t, , CH₃); 4.30 (2H, q, Hz, CH₂); 4.51 (2H, q, Hz, CH₂); 8.15 (1H, s, CH); 8.72 (1H, s, CH). ¹³C NMR (DMSO-) δppm: 14.56 (CH₃); 15.15 (CH₃); 42.38 (CH₂); 46.34 (CH₂); 118.17; 137.09 (Cq); 145.09 (CH); 149.99 (CH); 179.11 (Cq, C=S). HRMS (ESI) [M + H]: m/z = 209.07.

1,5-Dibenzyl-1H-pyrazolo[3,4-d]pyrimidine-4(5H)-thione (2b). Yield = 70%; mp: 160°C. ¹H NMR (DMSO-) δ ppm: 3.25 (2H, s, CH₂); 3.78 (2H, s, CH₂); 6.85–7.12 (q, ); 8.21 (1H, s, CH), 8.89 (1H, s, CH). ¹³C NMR (DMSO-) δppm: 51.83 (CH₂); 54.31 (CH₂); 118.24; 127.94 (Cq); 128.11–137.89 (); 145.54 (CH); 150.95 (CH); 179.82 (Cq, C=S). HRMS (ESI) [M + H]: m/z = 33.10.

2.2. 1,3-Dipolar Cycloaddition

To a solution of 1,5-diethyl-1H-pyrazolo[3,4-d]pyrimidine-4(5H)-thione (10 mmol) and diphenylnitrilimine (1.3 × 10 mmol) in THF (30 mL) triethylamine (2 mL) was added. The mixture was refluxed for 24 hours. The precipitate was collected by filtration and was separated by silica gel chromatography (hexane/ethyl acetate: 8/2).

2,5-Diethyl-3′,5′-diphenyl-2,5-dihydro-3′H-spiro[pyrazolo[3,4-d]pyrimidine-1,2′-[1,3,4]thiadiazole] (4a). Yield = 60%; mp: 165°C. ¹H NMR (DMSO-) δ ppm 1.12 (3H, t, Hz, CH₃); 1.28 (3H, t, Hz, CH₃); 3.37 (2H, q, Hz. CH₂); 4.11 (2H, q, Hz, CH₂); 6.83–7.67 (q, Hz, ); 7.69 (1H, s, CH); 7.69 (1H, s, CH). ¹³C NMR (DMSO-) δppm: 15.61 (CH₃); 16.03 (CH₃); 41.88 (CH₂); 42.28 (CH₂); 99.70; 100.79; 117.03; 126.38 (Cq); 129.26–142.20 (); 143.87 (CH); 148.01 (CH). HRMS (ESI) [M + H]: m/z = 403.16.

2,5-Dibenzyl-3′,5′-diphenyl-2,5-dihydro-3′H-spiro[pyrazolo[3,4-d]pyrimidine-1,2′-[1,3,4]thiadiazole] (4b). Yield = 60%; mp: 185°C. ¹H NMR (DMSO-) δ ppm: 3.65 (2H, s, CH₂); 4.28 (2H, s, CH₂); 7.25–7.32 (q, Hz, ); 8.81 (1H, s, CH); 8.99 (1H, s, CH). ¹³C NMR (DMSO-) δppm: 50.83 (CH₂); 53.21 (CH₂); 77.2; 118.24; 121.02; 125.13; 127.94; 128.04; 128.15; 128.46; 128.79 (Cq); 129.12–137.9 (CH_Ar); 145.54 (CH); 150.95 (CH). HRM (ESI) [M + H]: m/z = 527.39.

3. Results and Discussion

We first prepared 1,5-diethyl-1H-pyrazolo[3,4-d]pyrimidine-4(5H)-thiones 2a-b from 1a-b by refluxing phosphorus pentasulfide in pyridine. The identification of the product was determined by ¹H NMR, ¹³C NMR, IR, and mass spectra.

IR-spectra of compounds 2a-d did not contain C=O-group signals, the signal of C=S group (1500cm⁻¹) was present. The spectra of ¹³C NMR corroborated the results of IR. In addition, its mass spectrum shows a molecular ion peak at m/z 152 (M+) corresponding to the molecular formula C₅H₄N₄S compound 2, molecular ion at m/z 208.07 (M+) corresponding to the molecular formula C₉H₁₂N₄S compound 2a and molecular ion at m /z 332.10 (M+) corresponding to the molecular formula C₁₉H₁₆N₄S compound 2b (Scheme 1, Figure 4) [23].

We investigated the reaction of 1,3-dipolar cycloaddition of diphenyl hydrazonoyl chloride 3 with equimolecular amount of 1,5-diethyl-1H-pyrazolo[3,4-d]pyrimidin-4(5H)-thione 2a in dry tetrahydrofuran in presence of triethylamine (Scheme 2); one cycloadduct was obtained as a result of 1,3-dipolar cycloaddition of diphenyl nitrile imine ylide generated in situ from diphenyl hydrazonoyl chloride and triethylamine, on the dipolarophilic group C=S.

The structure of these compounds was confirmed on the basis of their spectroscopic characteristics. The ¹H NMR spectra of 4a (in DMSO-) showed an aromatic multiplet in the region of 6.83 to 7.67 ppm corresponding to the aromatic protons. Two downfield singlets were observed in the region of 7691–7968 ppm representing the protons for CH in pyrimidine ring and CH in pyrazole ring because of the high incidence of the aromatic ring system deshielding protons CH pyrimidine CH pyrazole. 1H NMR spectra of 4a also showed the CH₂ and CH₃ signals as triplets and multiplets and between 1.12 and 1.27 ppm and between 3.37 and 4.11 ppm, respectively. ¹³C NMR spectra of 4a exhibit in signal spirocarbon to 99.70 ppm, aromatic carbons 100.79 to 129.38 ppm, and the imine carbon to 142.2 and 139.93 ppm (HC=N). The mass spectrum shows a peak at m/z 403.16 corresponding to [M + H].

In order to examine the N-substitution effect of alkyl group on the 1,3-dipolar cycloaddition, 1,5-dibenzyl-1H-pyrazolo[3,4-d]pyrimidin-4(5H)-thione 2a was chosen to be employed to react with DPNI. So in this case it was found that C=S group underwent the 1,3-dipolar cycloaddition reaction and formed of novel spiro[thiadiazoline-(pyrazolo[3,4-d]pyrimidine)] (Scheme 3).

The structure of compound 4b was established by IR, ¹H NMR, ¹³C NMR, and mass spectrum. Its IR spectrum showed a characteristic absorption band at 1647 cm⁻¹ for the C=N- indicating the spirocarbon formation. Its ¹H NMR spectrum exhibited peaks at 7.25–7.32 ppm (20H) indicating the presence of aromatic protons. The two singlets at 3.65 ppm and 4.28 ppm relating to two protons of the two CH₂ groups were also observed. ¹³C NMR spectra of 4b exhibited, in particular, a spiro carbon signal at 77.2 ppm. The mass spectrum shows a peak at m/z 527.39 corresponding to [M + H].

3.1. In Vitro Antibacterial Activity

We studied spiropyrazolo[3,4-d]pyrimidines newly synthesized for their antibacterial activity against Escherichia coli (ATTC-25922), Staphylococcus aureus (ATCC-25923), Pseudomonas aeruginosa (ATCC 27853), and Enterococcus faecalis (ATCC-29212) strains of bacteria by the diffusion method disk [32]. The MIC values of the compound against bacteria are presented in Table 1.

We synthesized new spirocompounds with a high yield by cycloaddition reaction. They have showed moderate antibacterial activity against selected bacteria. All these compounds showed at an average concentration low antibacterial activity against the bacteria, except that compounds 1a and 4a showed higher activity, even at higher concentrations. Compound 4b showed higher activity against S. aureus and E. faecalis compared to bacteria E. coli and P. aeruginosa.

4. Conclusion

In conclusion, a new class of heterocyclic spiro[thiadiazoline pyrazolopyrimidine] compounds was synthesized and their structure was determined and also tested for their antibacterial activity in vitro. This study is expected to take the tests of anti-inflammatory drugs, antifungal, and anticancer activity because the literature gives some very interesting results on these topics.

Conflict of Interests

The authors declare that there is no conflict of interests regarding the publication of this paper.

Acknowledgment

The authors thank Pharmaceutical Laboratories PHARMA 5 for supporting this study.

References

P. Dinér, J. P. Alao, J. Söderlund, P. Sunnerhagen, and M. Grøtli, “Preparation of 3-substituted-1-isopropyl-1 H-pyrazolo[3,4-d]pyrimidin-4- amines as RET kinase inhibitors,” Journal of Medicinal Chemistry, vol. 55, no. 10, pp. 4872–4876, 2012.
View at: Publisher Site | Google Scholar
L. L. Yang, G. B. Li, H. X. Yan et al., “Discovery of N6-phenyl-1H-pyrazolo[3,4-d]pyrimidine-3,6-diamine derivatives as novel CK1 inhibitors using common-feature pharmacophore model based virtual screening and hit-to-lead optimization,” European Journal of Medicinal Chemistry, vol. 56, pp. 30–38, 2012.
View at: Publisher Site | Google Scholar
J.-H. Chern, K.-S. Shia, T.-A. Hsu et al., “Design, synthesis, and structure-activity relationships of pyrazolo[3,4-d]pyrimidines: a novel class of potent enterovirus inhibitors,” Bioorganic and Medicinal Chemistry Letters, vol. 14, no. 10, pp. 2519–2525, 2004.
View at: Publisher Site | Google Scholar
T. A. Bektemirov, E. V. Chekunova, I. A. Korbukh, N. Bulychev Yu, N. G. Yakunina, and M. N. Preobrazhenskaya, “Antiviral activity of substituted 6-methylmercaptopyrazolo(3,4-d)pyrimidines and their ribosides,” Acta Virologica, vol. 25, no. 5, pp. 326–329, 1981.
View at: Google Scholar
S. Taliani, C. La Motta, L. Mugnaini et al., “Novel N²-Substituted pyrazolo[3,4-d]pyrimidine adenosine A₃ receptor antagonists: inhibition of A₃-Mediated human glioblastoma cell proliferation,” Journal of Medicinal Chemistry, vol. 53, no. 10, pp. 3954–3963, 2010.
View at: Publisher Site | Google Scholar
D. Briel, R. Aurich, U. Egerland, and K. Unverferth, “Synthese substituierter 6-Phenylpyrazolo[3,4-d]pyrimidine mit potenziell Adenosin-A_2A-antagonistischer Wirkung,” Die Pharmazie, vol. 60, no. 10, pp. 732–735, 2005.
View at: Google Scholar
S.-A. Poulsen and R. J. Quinn, “Synthesis and structure-activity relationship of pyrazolo[3,4- d]pyrimidines: potent and selective adenosine A₁ receptor antagonists,” Journal of Medicinal Chemistry, vol. 39, no. 21, pp. 4156–4161, 1996.
View at: Publisher Site | Google Scholar
C. M. Niswender, E. P. Lebois, Q. Luo et al., “Synthesis and anti-tubercular evaluation of some novel pyrazolo[3,4-d]pyrimidine derivatives,” Medicinal Chemistry Research, vol. 21, pp. 1887–1891, 2012.
View at: Publisher Site | Google Scholar
B. S. Holla, M. Mahalinga, M. S. Karthikeyan, P. M. Akberali, and N. S. Shetty, “Synthesis of some novel pyrazolo[3,4-d]pyrimidine derivatives as potential antimicrobial agents,” Bioorganic and Medicinal Chemistry, vol. 14, no. 6, pp. 2040–2047, 2006.
View at: Publisher Site | Google Scholar
D. Bhambi, V. K. Salvi, J. L. Jat, S. Ojha, and G. L. Talesara, “Synthesis and antimicrobial activity of some new indole containing isoxazolines and phthalimidoxy derivatives of thiazolidinone and thiohydantoin,” Journal of Sulfur Chemistry, vol. 28, no. 2, pp. 155–163, 2007.
View at: Publisher Site | Google Scholar
R. A. M. Fatty, A. M. Gaafar, and A. M. S. Youssef, “A simple and efficient synthesis of new substituted benzothienopyridazine spiro- derivatives of pyrazolone, pyrimidinone and diazepinone,” Journal of Chemical and Pharmaceutical Research, vol. 3, pp. 222–228, 2011.
View at: Google Scholar
N. N. Gajera and M. C. Patel, “Synthesis, characterization and biological screening of new spirochromanones,” Journal of Chemical and Pharmaceutical Research, vol. 4, no. 7, pp. 3377–3382, 2012.
View at: Google Scholar
B. S. Rane, S. V. Deshmukh, M. G. Ghagare, R. V. Rote, and M. N. Jachak, “Synthesis of spiro-pyrimido [4,5-b]quinoline and study of their antimicrobial activities,” Journal of Chemical and Pharmaceutical Research, vol. 4, no. 7, pp. 3562–3567, 2012.
View at: Google Scholar
J. R. Yerrabelly, V. Chakravarthula, H. Yerrabelly et al., “Unusual tandem ring closing metathesis—Claisen rearrangement enroute to spiro annulation of O-allyl biscoumarins,” Tetrahedron Letters, vol. 56, no. 17, pp. 2180–2182, 2015.
View at: Publisher Site | Google Scholar
K. De, A. Bhaumik, B. Banerjee, and C. Mukhopadhyay, “An expeditious and efficient synthesis of spiro-pyrazolo[3,4-b]pyridines catalysed by recyclable mesoporous aluminosilicate nanoparticles in aqueous-ethanol,” Tetrahedron Letters, vol. 56, no. 13, pp. 1614–1618, 2015.
View at: Publisher Site | Google Scholar
S. Abouricha, E. M. Rakib, N. Benchat, M. Alaoui, H. Allouchi, and B. El Bali, “Facile synthesis of new spirothiadiazolopyridazines by 1,3-dipolar cycloaddition,” Synthetic Communications, vol. 35, no. 16, pp. 2213–2221, 2005.
View at: Publisher Site | Google Scholar
E. V. Budarina, N. N. Labeish, V. K. Bel'Skii, and V. A. Galishev, “Hetrocyclic thiones and their analogs in reactions of 1,3-dipolar addition: v. Reactions of 2,3,3-triphenyl-1-thioxophthalimidine with nitrile imines,” Russian Journal of Organic Chemistry, vol. 41, no. 5, pp. 758–761, 2005.
View at: Publisher Site | Google Scholar
E. M. Essassi, C. Ahoya, R. Bouhfid et al., “Synthesis and antibacterial activity of new spiro[thiadiazoline-quinoxaline] derivatives,” Arkivoc, vol. 2011, no. 2, pp. 217–226, 2011.
View at: Publisher Site | Google Scholar
K. Al Mamari, H. Ennajih, H. Zouihri, R. Bouhfid, S. W. Ng, and E. M. Essassi, “Synthesis of novel dispiro-oxindoles via 1,3-dipolar cycloaddition reactions of azomethine ylides,” Tetrahedron Letters, vol. 53, no. 18, pp. 2328–2331, 2012.
View at: Publisher Site | Google Scholar
Y.-X. Zhang, L. Ding, X.-Y. Liu et al., “Spiro-fused N-phenylcarbazole-based host materials for blue phosphorescent organic light-emitting diodes,” Organic Electronics, vol. 20, pp. 112–118, 2015.
View at: Publisher Site | Google Scholar
P. Das, A. O. Omollo, L. J. Sitole et al., “Synthesis and investigation of novel spiro-isoxazolines as anti-cancer agents,” Tetrahedron Letters, vol. 56, no. 14, pp. 1794–1797, 2015.
View at: Publisher Site | Google Scholar
J. Naga Siva Rao and R. Raghunathan, “A facile synthesis of glyco 3-nitrochromane hybrid pyrrolidinyl spiro heterocycles via [3+2] cycloaddition of azomethine ylides,” Tetrahedron Letters, vol. 56, no. 17, pp. 2276–2279, 2015.
View at: Publisher Site | Google Scholar
M. El Fal, Y. Ramli, E. M. Essassi, M. Saadi, and L. El Ammari, “The crystal structure of 1,5-dibenzyl-1H-pyrazolo[3,4-d]pyrimidine-4(5H)-thione,” Acta Crystallographica E, vol. 71, pp. o95–o96, 2015.
View at: Publisher Site | Google Scholar
M. El Fal, Y. Ramli, E. M. Essassi, M. Saadi, and L. El Ammari, “Crystal structure of 4-allylsulfanyl-1H-pyrazolo[3,4-d]pyrimidine,” Acta Crystallographica E, vol. 70, p. o1038, 2014.
View at: Publisher Site | Google Scholar
M. El Fal, Y. Ramli, E. M. Essassi, M. Saadi, and L. El Ammari, “Crystal structure of 1-ethylpyrazolo[3,4-d]pyrimidine-4(5H)-thione,” Acta Crystallographica Section E: Structure Reports Online, vol. 70, no. 9, pp. o1005–o1006, 2014.
View at: Publisher Site | Google Scholar
A. Alsubari, Y. Ramli, E. M. Essassi, and H. Zouihri, “5-(Pyridin-4-ylmeth-yl)-1Hpyrazolo-[3,4-d]pyrimidin-4(5H)-one,” Acta Crystallographica Section E: Structure Reports Online, vol. 67, part 8, Article ID o1926, 2011.
View at: Publisher Site | Google Scholar
M. El Fal, Y. Ramli, E. M. Essassi, M. Saadi, and L. El Ammari, “4-benzylsulfanyl-1H-pyrazolo[3,4-d]pyrimidine,” Acta Crystallographica Section E, vol. 69, no. 11, p. o1650, 2013.
View at: Publisher Site | Google Scholar
M. El Fal, Y. Ramli, E. M. Essassi, M. Saadi, and L. El Ammari, “Crystal structure of 1-methyl-4-methylsulfanyl-1H-pyrazolo[3,4-d]pyrimidine,” Acta Crystallographica, vol. E70, Article ID o1281, 2014.
View at: Publisher Site | Google Scholar
I. El Ouali, B. Hammouti, A. Aouniti et al., “Thermodynamic characterisation of steel corrosion in HCl in the presence of 2-phenylthieno (3, 2-b) quinoxaline,” Journal of Materials and Environmental Science, vol. 1, no. 1, pp. 1–8, 2010.
View at: Google Scholar
M. Z. A. Badr, G. M. El-Naggar, H. A. H. El-Sherief, A. E. S. Abdel-Rahman, and M. F. Aly, “Reaction of quinoxaline derivatives with nucleophilic reagents,” Bulletin of the Chemical Society of Japan, vol. 56, no. 1, pp. 326–330, 1983.
View at: Publisher Site | Google Scholar
O. S. Moustafa, “Synthesis and some reactions of Quinoxalinecarboazides,” Journal of the Chinese Chemical Society, vol. 47, pp. 351–357, 2000.
View at: Google Scholar
J. Sirot, Bactériologie Médicale, Flammarion, Paris, France, 2nd edition, 1990.

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