4-Ylmethylethylation Lab Report

Results and Discussion: A reaction involving two bond formation was employed for the preparation of 3,5-disubstituted 1,2,4-triazole as nucleobase analog. The hydrazides 2a,b were allowed to react with the heteroaryl substituted acetonitrile derivatives 1a-c using catalytic amount of K2CO3 and n-butanol as a solvent to afford the disubstituted 1,2,4-triazoles 3-7 bearing benzothiazole, benzotriazole, quinoloxymethyl and benziimidazole ring systems in moderate yields. The structures of compounds 3-7 were confirmed by their spectral data. Their IR spectra revealed the disappearance of the characteristic hydrazide carbonyl and cyano absorption bands. The 1H NMR spectra showed signals corresponding to the two methylene protons in addition to the aryl and NH proton of the 1,2,4-triazole ring.
Glycosylation of the 1,2,4-triazole substituted with benzotriazol-1-ylmethyl, quinoloxy-8-ylmethyl,benzothiazolyl-2-ylmethyl 3,4 and 7, respectively by reaction with acetylated gluco- and xylopyranosyl bromide produced the corresponding heteroaryl-1,2,4-N-glycosides 8-13 in good yields.
The Infra-Red spectra showed the carbonyl of the acetyl groups in addition to the disappearance of the NH band. Their 1H NMR spectra showed the acetyl methyl signals, the sugar protons signals and the aromatic signals. The coupling constant value of the anomeric protons 9.8-10.2 Hz indicated that attachment of the sugar moiety is in the beta conformation leading to the formation of the
1,2,4-triazole-β-glycoside.
Compounds 8-13 were deacetylated by methanolic ammonia to give the deprotected glycosides 14-19 with free hydroxyls. The NMR and IR spectra of the latter free hydroxyl glycosides are in agreement with the assigned structure. Reaction of the 1,2,4-triazole substituted with benzoimidazol-2-ylmethyl- moiety incorporating two -NH centers 5 and 6 with tri-O-acetyl-D-xylopyranosyl bromide resulted in the formation of the acetylated glycoside derivatives 20 and 21, respectively in good yields. Their corresponding spectral data revealed the attachment of two xylopyranosyl moieties at the two NH positions in the compound. The mode of attachment as β-type was also obvious from the coupling J value of H-1 in the sugar moiety.
Deacetylation of the latter benzoimidazolyl-1,2,4-triazolxylopyranoside derivatives 20 and 21 in saturated methanolic ammonia solution resulted in the formation of the free hydroxyl glycoside derivatives 22 and 23, respectively. Their IR data showed the presence of the hydroxyl bands and the NMR spectra agreed with the assigned structure.

Antiviral activity:
A number of the synthesized compounds were studied for their antiviral activity against H5N1 influenza virus strain A/Egypt/M7217B/2013 using MTT cytotoxicity assay (TC50) and Plaque reduction assay investigating the inhibition % and cytotoxicity% values. The results of inhibition activities and cytotoxicity were formulated in tables 1 and 2 in addition to fig. 1.
The results revealed that the tested compounds displayed a range from no inhibition to week and moderate inhibition. Compound 8 showed no cytotoxicity at all concentrations in this investigation with TC50 more than 200 μg/μl. On the other hand Compounds 8, 6, 9 and 3 showed no inhibition activity. The results of antiviral activity indicated that compounds 21, 7 and 4 were the most active with 34%, 30.5% and 29% inhibition. Compound 7 showed higher inhibition activity at 25 μg/μl concentration than 50 μg/μl. Compounds 5 and 10 were found to be weak in inhibition activity at only 50 μg/μl. It was also found that compounds 21 and 4 showed little cytotoxicity values at most of the concentrations with TC50 values 1250 and 173 μg/μl.
In correlation of the obtained results with the structures of tested compounds, the results indicated that the 1,2,4-triazole compound linked to benzoimidazole and benzo-1,2,3-triazole ring systems and incorporating two glycosyl moieties was the most active in such investigation against H5N1. This indication reveals the importance of the xylosyl units as the activity was raised by the attachment of such glycosyl constituents to the free triazole compound linked to both ring systems.
On the other hand, in this investigation, it was also found that the unsymmetrical 1,2,4-triazole compound with both benzotriazole and benzothiazole in addition to the 1,2,4-triazole having both quinolin-8-yloxy 1,2,3-benzotriazole structures were higher in activity than symmetrical compounds or other triazoles with free NH groups.