Synthesis and antifungal activity of novel polyheterocyclic compounds containing fused 1,2,4-triazine moiety

3-Amino-4-(4-chlorophenyl)-7-hydrazino-8 H -pyrazolo[4,3-e ][1,2,4]triazolo[1`,5`- a ]pyridine-5-carbonitrile ( 4 ) was synthesized from 4-(4-chlorophenyl)-1,6-diamino-2-oxo-1,2-dihydro-pyridine-3,5-dicarbonitrile ( 1 ). Reaction of 4 with α , β -bifunctional compounds gave pyrazolotriazinotriazolopyridines ( 8-14 ). The behavior of 4 towards condensation reactions with indole-2,3-dione in different media gave different products 15-16 . Acetylation of 16 led to different products depending on the reaction conditions. Structures of the products have been deduced from analytical and spectral data (UV, IR, 1 H NMR, 13 C NMR and mass spectra). Some of the products were screened for antifungal activity.

C NMR of compounds 9 and 10.
Finally, the behavior of compound 16 towards acetylation reactions has been studied under different conditions.Thus, refluxing 16 with glacial acetic acid afforded the monoacetyl derivative 18.However, when the reaction was carried out in boiling glacial acetic acid containing a few drops of acetic anhydride, the diacetyl derivative 19 was isolated (Scheme 4).Structures of mono-and diacetyl derivatives 18 and 19 were established form their elemental analysis and spectral data.IR spectrum of 18 showed an absorption band at 1764 cm -1 assigned to one C=O, while 19 showed two absorption bands at 1740 and 1702 cm -1 assigned to two C=O functions. 1H NMR of 18 showed a signal at δ 1.91 ppm, characteristic for one COCH 3 group, while that of 19 showed two signals at δ 1.56 and 1.92 ppm, characteristic for two COCH 3 groups.

Fungicidal activity
4] The tested compounds were dissolved in DMF [which has no inhibition activity] to get 1 mg/ml solution.The antibiotic flucanazole was used as standard antifungal reference.The inhibition zones of the microbial growth surrounding the filter paper disc (2.5 mm) were measured in millimeters at the end of an incubation period at 30 ο C for 3 days (Table 1).
All the tested compounds showed variable activities toward the two species in comparison to the standard flucanazole which revealed that these compounds are biologically active due to the presence of different heterocycles and functional groups.
From the results obtained, it is clear that most of the tested compounds showed moderate activity toward the tested fungi except compound 13 showed higher activity towards Alternaria alternata fungi which mainly due to the expected dihydroxy structure (Table 1)