1. INTRODUCTION:

Scientists have been interested in the coordination chemistry of heterocyclic thione. The primary reason thione rings have attracted such chemical interest is that they are bifunctional ligands that bond to atoms by donating electrons from sulfur or nitrogen atoms¹ and effective biological importance². References have shown that the presence of the azomethine group (C=N) is a prerequisite for biological activity³ result from the condensation of primary amines with ketones or aldehydes⁴^,5. Heterocyclic compounds offer a high degree of structural diversity and have proven to be broadly and economically useful as therapeutic agents^6,7 .It is the backbone of a wide range of synthetic materials and therapeutic drugs used as antifungals, antibacterials, anti-inflammatory agents, antivirals, antitumors, and others⁸. 1,2,4-Thiotriazines and 1,2,4-Thiotriazoles are cyclic thionates derived from thiocarbohydrazides. Their derivatives have demonstrated biological activity, such as antibacterial agents, antibiotics, and insecticides⁹. Triazole derivatives are synthesized from amine and thione substituents. The presence of the exocyclic thione group on the heterocyclic group is of great importance, because the presence of both amine and thione groups can form compounds with effective and synergistic coordination ability¹⁰. The biological activity of metal complexes with thiosemicarbazone and thiocarbohydrazide derivatives has also been studied and found to be among the most potent anti-inflammatory, anti-viral, and anti-tumor agents. Anti-tumor activity is one of the most important biological findings of research on thiosemicarbazone compounds and their metal complexes¹¹. The bonding between the amine and thione groups usually results in the formation of complexes with metals that have important five-membered rings. ¹²The general formula for the bonding between the mercapto-triazole system is R3N3SC2 ¹³ .

The 1,2,4-triazole nucleus has been used in the synthesis of many drug candidates of therapeutic importance, including sedatives and anxiolytics. Several 1,3,4-thiadiazoline derivatives have been used as antibacterial and antitumor agents, insecticides, dyes, analytical reagents, and lubricants¹⁴.

In this study, the structures of the compounds were investigated using infrared spectroscopy, mass spectrometry (MS), and single-nuclear magnetic resonance (1H-NMR) data analysis. These compounds were then studied in vitro to reveal their antifungal against Candida albicans and antioxidant activity

2. MATERIALS AND METHODS:

All chemicals were purchased from Sigma Aldrich Chemical Co. and Merck Chemical Co. Melting points of the compounds were determined by using an electrothermal digital melting point apparatus. The infrared (IR) spectra of the compounds were recorded in the region of 4000-400 cm-1 on a) FT-IR Perkin-Elmer) spectrophotometer, Mass spectra were recorded on a (Shimadzu Agilent Technologies 5975C at 70(. 1H-NMR spectra were recorded with a model) Bruker AMX400 MHZ (spectrometer using DMSO as a solvent.

The antimicrobial screening was carried out at with Muller Hinton agar and the isolates were Candida albicans fungi. Nystatin was used as control. The susceptibility test, the minimum inhibitory concentration, and minimum fungicidal concentration were carried out.

The antioxidant activities is dependent on free radical scavenging activity of the1,2,4-triazole derivatives as the concentration of the test compounds increases, the radical scavenging activity increases and lower IC50 value reflects better protective action.

2.1. Chemistry:

The compounds were variedly color solids (data in Table 1). They were generally insoluble in common organic solvents but soluble in coordinating solvents such as DMSO and DMF. They were stable in air and exist in crystalline form. They have high melting points indicating strong bonding network within the compound.

The prepared compound was described by chemical analyses such the infrared (IR) spectra in the region of 4000-400 cm-1 on a) FT-IR (spectrophotometer, Mass spectra ,1H- NMR spectra spectrometer using DMSO as a solvent.

Table 1. Physical data of the compounds

Physical data of the compounds	Chemical formular	M.wt g/mole	Color	Melting point m.p. °C	solvent
L₁	C₁₄H₁₄N₈S₂	358.44	white	280	DMSO-DMF
L₂	C₁₄H₁₄N₈S₂	358.44	Yellow	285	DMSO-DMF

2.2 Antimicrobial Study:

The newly synthesized Schiff base L1–L2 were test against antifungal activity was carried out with isolates Candida albican fungi (E. coli) by the disk diffusion method^3,1⁵. The test compounds (ligand L1) were dissolved in DMSO to get 10mg/mL solution. A known volume (10 lL) of the solution was applied with the help of a micropipette onto the sterilized filter paper disks. The disks were dried at room temperature overnight and stored in sterilized dry containers. Disks soaked with 10L of DMSO and dried in air at room temperature were used as the negative control. The standard antibiotic disks used as positive control were prepared as mentioned above in the laboratory by applying a known concentration of the standard antibiotic solution. Nystatin was used as standard antibiotic. Bacterial culture was grown in nutrient broth medium at 22°C overnight and spread onto solidified nutrient agar medium in Petri plates using sterilized cotton swabs. Test and control disks were then applied to the medium surface with the help of sterilized forceps. The plates were incubated at 22°C for 24h. The results were recorded by measuring the zone of inhibition in mm against each compound. The experiments were carried out in triplicate and the values obtained were statistically analyzed.

2.3 Antioxidant Activities:

The radical scavenging activity of the synthesized compounds against stable free radical 2.2- diphenyl-2-picrylhydrazyl hydrate (DPPH, Sigma-Aldrich Chemie, Steinheim, Germany) was determined spectrophotometric ally. When DPPH reacts with antioxidant compounds, which can donate hydrogen, it is reduced. Following the reduction, its deep violet color in methanol bleached to yellow, showing a significant absorption decrease at 517nm.

Then 1000μL of various concentrations (60,120,180,240,300,360,450, and 600μg/ml) of the compounds(L1-L2) dissolved in methanol were added to 1000 μL of ethanol solution of DPPH (0.6μM). After a 30min incubation period at room temperature, the absorbance was read against a blank at 515 nm (ATI-UnicamUV-2 UV-Vis spectrophotometer, Cambridge, UK) Ascorbic acid was used as the reference compound. All tests and analyses were done in three replicates and the results were averaged.

Free radical DPPH inhibition in percentage (AA %) was calculated as follows:

DPPH scavenging effect (%) = (A control – A sample/A control) × 100

Where control is the absorbance of the control reaction (containing all reagents except the test compound) and sample is the absorbance of the test compound.

3. RESULTS AND DISCUSSION:

3.1 Chemistry:

The compounds were identified by MS, FT-IR spectrum,¹HNMR spectrum and The results were discussed using this articles¹⁶^–20

N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,3-methylidene) benzene (L1):

Schiff base of N, N bis (3-methyl-1,2,4-triazole-5- thione)-1,3-methylidene) benzene (L1) figure 1, IR showed absorption bands at 3068 cm-1 due to C-H aromatic ,2938cm-1 due to C-H aliphatic, 2742 cm-1 for S-H group and absorption band at 1593 cm-1 for C=N group figure 2.

¹HNMR spectrum shows appearances of singlet signal at 13.97 ppm for 2H of S-H, singlet signal at 10.13 ppm due to 2H of azomethine group (CH=N), and signals (8.41-8.15-7.75) for (s, 1 H), (d, 2 H), (t,1 H) of phenyl group respectively, and singlet signal at 2.38 ppm for 6H of methyl group figure 3.

MS (AEI) (M+): the mass spectra figure 4 showed the base peak M+ at M/Z+= (358.2) corresponds to the original molecular weight of ligand molecular ion, where the mass spectrum was given to ligand a number of fragmentation and these fragmentation represented in the mass spectra in term of relative abundance compared to M/Z+, and the main peak show via mass spectrum is relatively to molecular weight of ligand [C₁₄H₁₄N₈S₂]⁺.

Figure 1: L1

Figure 2: FT-IR Spectrum for Compound (L1)

Figure 3: 1HNMR Spectrum for Compound (L1)

Figure 4: Mass Spectrum for Compound(L1)

N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,4 -methylidene )benzene (L2) .

Schiff base of N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,4 -methylidene )benzene (L₂ ) figure 5, FTIR spectrum of compound (L₂) showed absorption bands at 3102 cm-1 due to C-H aromatic and absorption bands at 2948 cm^-1 for C-H aliphatic, 2776 cm^-1 for S-H group, 1635 cm^-1 due to C=N group figure 6.

MS (AEI) (M⁺): the mass spectra showed the base peak M⁺ at M/Z⁺= (358.2).

Figure 5 : N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,4 -methylidene )benzene (L2)

Figure 6: FT-IR Spectrum for Compound (L2)

3.2Antimicrobial analysis:

To determine the of fungi activity of the synthesized compounds, the prepared compounds were examined at a concentration of (250, 500, 1000 ppm) in (DMSO), against of fungi (Candida albicans). The zone of inhibition was compared after 24h of incubation at 22°C against suitable antibiotics.

The results indicated that tow compounds exhibited good inhibitory activity against tested pathogenic microorganism.

The following table2 shows the effect of against of fungi (Candida albicans).

Table 2: The effect of against of fungi (Candida albicans)

Ppm	250		500		1000
N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,3-methylidene ) benzene (L₁).
Damping diameter (mm)	24.5	25	27	27	28	28
Average	24.7		27		28
N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,4 -methylidene )benzene (L₂).
Damping diameter(mm)	25.2	25	26.8	27	27	28
Average	25.1		26.9		27.5
Standard antibiotic nystatin
Damping diameter (mm)	27.4		29.8		33.8

Both N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,3-methylidene ) benzene (L₁) and N, N bis()3-methyl-1,2,4-triazole-5- thione (-1,4 -methylidene )benzene (L₂) showed good activity against Candida albicans, especially at 250 ppm, and poor activity at 1000 ppm compared to the standard antibiotic Nystatin as the figures 7,8and 9 shows.

Figure 7: Biological activity of compound L₁ against Candida albicans.

Figure 8: Biological activity of compound L₂ against Candida albicans.

Figure 9: The biological activity of the standard antibiotic nystatin against Candida albicans.

3.3 Antioxidant Activities:

Free radical scavenging activity of the1,2,4-triazole derivatives is concentration dependent, as the concentration of the test compounds increases, the radical scavenging activity increases and lower IC50 value reflects better protective action. From results, it may be postulated that compounds (L₁- L₂) were able to reduce the stable free radical DPPH to the yellow-colored diphenyl picrylhydrazine exhibiting better free radical scavenging activity than the standard antioxidant Ascorbic acid.

The effect of the different synthetic compounds on DPPH radical scavenging was compared to control. Results showed in Table 3.

Structure activity relationship study showed that the antioxidant activity of these 1,2,4-triazole derivatives could be due to that consist of atoms with low electronegativity and species with relatively small ionization energiess²¹.

Table 3: Percentage of Scavenging Effect of synthesized compounds

Comp/deplete %	60μg/ml	120μg/ml	180μg/ml	240μg/ml	300μg/ml	360μg/ml	450μg/ml	600μg/ml
L₁	21.9	25	27.6	32.1	36.9	41.8	48.8	57.29
L₂	32.85	37.5	41.4	48.15	55.35	62.70	73.20	85.94

As shown in table 3 the most active compound was L₂ that showed highest antioxidant activity, which reached 85.94% at concentration 600μg/ml that may be due to 1,2,4-triazole- ring which possess a high biological activity.

4. CONCLUSION:

The prepared compounds were characterized by known spectroscopies (IR, MS, and 1H-NMR).

The activity of the prepared thiazolidine derivatives against Candida albicans was tested, both two compounds studied demonstrated good antifungal activity against Candida albicans at a concentration of 250 ppm. This is due to the presence of the thione group (C=S), which is known for its antibiotic activity.

The results of the antioxidant activity study showed that the most active compound was L2, which showed the highest antioxidant activity, reaching a free radical scavenging rate of 85.94% at a concentration of 600 µg/ml, which may be due to the 1,2,4-triazole ring, which has high biological activity.

5. ACKNOWLEDGMENT:

The cooperation of the Department of Chemistry, Faculty of Science, Damascus University, Syria.

6. AUTHORS' DECLARATION:

Conflicts of Interest: None.

We hereby confirm that all the Figures and Tables in the manuscript are mine ours.

Ethical Clearance: The project was approved by Faculty of Science, Damascus University, Syria.

7. AUTHORS' CONTRIBUTIONS STATEMENT.

M A designed the study from preparation compounds and performed the experiments and analysis of the results. And M A wrote the paper, a contributed to the design and implementation of the research, to the analysis of the results and to the writing of the manuscript.

REFERENCES:

1. Shirinkam B. Tabatabaee M. Kukovec B. M. Oliver C. L. Ghassemzadeh, M. Preparation, spectroscopic characterization, and crystal structure of a mixed-ligand silver (I) complex with 1, 2, 4-triazole-based Schiff base and triphenylphosphine. Monatshefte für Chemie-Chemical Monthly. 2014; 145(11): 1753-1757.‏

2. Iniama G. E. Eno E. A. Pt (II) and Ru (III) Complexes Of 5-Substituted-4-Amino-3-Mercapto-1, 2, 4-Triazole Schiff Base: Synthesis, Spectral Characterization And Antimicrobial Studies. International Journal of Engineering, Science and Mathematics.2018; 7(9): 21-27.‏

3. Hanif M. Chohan Z. H. Design, spectral characterization and biological studies of transition metal (II) complexes with triazole Schiff bases. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy.2013; 104: 468-476.‏

4. Kumar R. Sharma S. Synthesis, spectroscopic, thermal studies and biological activity of a new schiff base and its copper complexes. Res J Sci Technol. 2020; 12(4): 251-256. doi:10.5958/2349-2988.2020.00033.9

5. Ashour M. Kandil F. Alazrak A. Synthesis, and Spectroscopic Characterization study of some Schiff Bases N, N bis (3-methyl-1, 2, 4-triazole-5-thione (1, 3-methylidene) benzene and N, N bis (3-methyl-1, 2, 4-triazole-5-thione (1, 4-methylidene) benzene. Research Journal of Pharmacy and Technology.2014; 17(4): 1486-1490.

6. Hanan A. Abdel Fattah. Synthesis, Cytotoxic And Antimicrobial Activities Of Novel 1,2,4- Triazole Derivatives Incorporating Aryl Sulfonamide Moiety. Asian Journal of Pharmaceutical Analysis and Medicinal Chemistry. 2014 ; 2(3): 145 - 162

7. Bhalerao R. Rishipathak D. Udavant P. Kshirsagar S. Synthesis and Evaluation of Antidiabetic Activity of Few 5-(4-(5-Substitutedphenyl)-1, 3, 4-Oxadiazlole-2-Yl) Methoxy)Benzylidene) Thiazolidine 2, 4-Dione Derivatives. Asian J Pharm Res. 2017; 7(4): 261. doi:10.5958/2231-

8. Muktham R. Bhargava S K. Bankupalli S. and Ball A S. A Review on 1, 2, 4 - Triazoles. J. Sustain. Bioenergy Syst. 2016,. 6(3): 72-92 Available: http://www.scirp.org/journal/doi.aspx?DOI=10.4236/jsbs.2016.63008.

9. Ghaneian M T. Synthesis of Ag(I) and Cu(I) Complexes with 4-Amino-5-Methyl-2h-1,2,4-Triazole-3(4h)-Thione Ligand as Thiocarbohydrazide Derivatives and their Antimicrobial Activity. Pharm. Chem. J.2015; 49(3):210-212. doi: 10.1007/s11094-015-1258-0.

10. Shah H J. and Chaudhari J A. Synthesis, Characterization and biological studies of some novel benzimidazole derivatives. J. Chem. Pharm. Res.2014; 6(7) 1890–1894.

11. Kumar G. Kumar D. Singh C. P. Kumar A. and Rana V. B. Synthesis, physical characterization and antimicrobial activity of trivalent metal Schiff base complexes. J. Serbian Chem. Soc.2010; 75)5(: 629–637doi: 10.2298/JSC090704037K.

12. Barnali Das S C D. Sajal Srivastava Y R P. Synthesis, characterization and biological studies of some novel piperidinothiophenes. Springerplus. 2013; 1–6.

13. Hameed A A. Synthesis, Characterization and Antioxidant Activity of Some 4-Amino-5-Phenyl- 4h-1,2, 4-Triazole-3-Thiol Derivatives. Indian J. Heterocycl. Chem.2014; 4(2): 397–400. doi: 10.13140/RG.2.2.18501.96484

14. Al-amin M. and Islam M. R. Synthesis of some bis-triazole derivatives as probes for cytotoxicity study. A J. Bangladesh Pharmacol. Soc.2006; 21–26. doi: 10.3329/bjp.v1i1.483.

15. Kumar K. Singh B. K. Synthesis, characterization and anti-microbial activity of some 4-thiazolidinone conjugatives. Asian journal of pharmaceutical analysis.2020; 10(4).195-200.‏

16. Jubie S. Gowramma B. Kalirajan R. Gomathy S. Elango K. Synthesis and Biological Evaluation of Some 3 - Derivatives. Asian J Res Chem. 2009; 1(1):32-38.

17. Vaijayanthimala P. Perumal P. Synthesis, Characterization and Anti-microbial Evaluation of Substituted Benzo [ g ] indol-2-one Derivatives. Res J Sci Tech. 2011; 3(6):313-317.

18. Gupta V. Singh S. Kgupta Y. Synthesis and Antimicrobial Activities of Zn (II) Complex of 2,5-diamino-1,3,4-thiadiazole. Res J Sci Tech. 2013; 5(4):452-465.

19. Kalkotwar RS. Saudagar RB. Design, Synthesis and anti-microbial, anti-inflammatory, Antitubercular activities of some 2, 4, 5-trisubstituted imidazole derivatives. Asian J Pharm Res. 2013; 3(4):159-165.

20. Shendge AS. Tayade DT. Kale PR. Synthesis of 2-(3-Substitutedamino-2, 4-Dithiobiuretoformamidino-4-isobutoxyphenyl)-4-Methyl-5-Carboxy-1, 3-Thiazoles. Asian J Pharm Anal. 2017; 7(4):214. doi:10.5958/2231-5675.2017.00034.5

21. Hameed A. A. Hassan F. Synthesis, characterization, and antioxidant activity of some 4-Amino-5-Phenyl-4H-1,2,4-Triazole-3-Thiol Derivatives. Int. J. Appl. Sci.Technol.2014; 4(2): 202–211.doi: 10.13140/RG.2.2.18501.96484.

Received on 10.11.2025 Revised on 20.12.2025

Accepted on 29.01.2026 Published on 25.04.2026

Available online from April 28, 2026

Research J. Science and Tech. 2026; 18(2):138-144.

DOI: 10.52711/2349-2988.2026.00019

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