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Heterocyclic Chemistry is a thrust area for Chemists. Biologically active substituted benzenethiols in recent years have aroused our interest in the synthesis and biological activity of these heterocycles. This article reports synthesis of prospective bioactive substituted benzenethiol that have shown significant antimicrobial activity1-2 against some selected strains of fungi, bacteria and lead us to develop a possibly potent class of biologically active agents. Heterocyclic compounds such as 10H-Phenothiazines, 4H,1-4 Benzothiazines and their derivatives sulfones, ribofuranosides etc. used as tranquilizers, antihistamines, diuretics, analgesics, neurolepitcs, sedatives, antipsychotics, CNS depressants, anticancer, antidepressants, antipyretics, antiparkinson drugs3-7 etc. Substituted benzenethiol were prepared from substituted aryl amines.

In last few decades a considerable amount of attention has been focused on synthesis of nitrogen and sulfur containing heterocycles The investigation of substituted substituted aminobenzenethiols has steadily flourished because they exhibit a large scope of applications. Due to the pharmaceutical properties, substituted aminobenzenethiols are used for clinical purposes, like sedative, antihelmintics, antiinflammatory, antimalarials, antibacterial, anticonvulsants, pesticides etc. Slight change in substitution pattern in substituted aminobenzenethiols nucleus causes distinguishable difference in their biological activities.

Arun Goyal, Shikha Agarwal and D.C.Gautam, Indian Journal of Chemistry, 2022 in the article " different novel 10H-phenothiazines, their sulfones and ribofuranosides were synthesized using commercially available starting materials and identified by spectral analysis and have shown moderate to significant antimicrobial properties. The different bacterial and fungal strains showed different susceptibility towards the tested compounds, which may be possibly explained by different resistance mechanism exhibited by each strain. The MIC values of antibacterial and antifungal screening revealed that the excellent antibacterial and antifungal activities against all the three selected strains of bacteria and fungi respectively".

Nishidha Khandelwal, Abhilasha Yadav et al., Nucleosides Nucleotides Nucleic Acids, 2012 in the article "Synthesized phenothiazines on oxidation with 30% hydrogen peroxide in glacial acetic acid yield sulfones, and when treated with sugar give ribofuranosides. These compounds are evaluated for their anthelmintic and antimicrobial activities.

A mixture of concentrated hydrochloric acid (9 ml) and water (25 ml) with substituted aniline (I) (0.1 mole), were taken in a 250 ml R.B. flask, and heated for half an hour, and then 0.1 mole ammonium thiocyanate was added and refluxed. Solid phenyl thiourea (II) separated out was filtered. washed with water, dried and crystallized from ethanol. 0.1 Mole synthesized substituted phenylthiourea (II) and Bromine (0.1 mole) in chloroform (100 ml) were taken in a two necked R.B. flask (500 ml) well equipped with a mechanical stirred and the temperature was maintained below 5°C. The reaction mixture was refluxed until the evolution of hydrogen bromide vapors ceased (about 4 hrs). Solid substituted 2-aminobenzothiaozole ( 2-Amino-3,5-dichloro-6-fluoro-benzenethiol) was treated with sulfur dioxide water and filtered.

                                                 

Table- Biological Activity of Substituted 2-aminobenzenethiol:

Compound Name	Minimum Inhibitory Concentrations MICs of bacterial strains in mg/ml
	Escherichia coli MTCC 2939	Bacillus subtilis MTCC 441	Streptomyces griseus MTCC 1998
2-Amino-3,5-dichloro-6-fluoro-benzenethiol	128	116	118
Streptomycin	23	18	21

Spectral assessment and physical data of the synthesized substituted 2-aminobenzothiaozole given below.

2-Amino-3,5-dichloro-6-fluoro-benzenethiol

Yield 68%, m.p. 92°C, color: brown; IR (KBr, v): 3450, 3290, 2590, 780 cm^–1. ¹H NMR (300.40 MHz, DMSO-d₆): d 7.79 (s, 1H, Ar-H), d 4.35 (s, 2H, -NH₂) d 1.76 (s, 1H, -SH). ¹³C NMR (75.45 MHz, CDCl₃,): d 120.7 (C-1), 129.5 (C-2), 128.4 (C-3), 144.2 (C-4), 128.3 (C-5), 124.8(C-6). "Anal. Calcd for C₆H₄Cl₂FNS: C, 33.96; H, 1.88; N, 6.60; Found : C, 33.45; H, 6.25; N, 6.84".

Biological Activity

The synthesized substituted 2-aminobenzenethiol found active in primary screening were further tested in a second set of dilution against microorganisms. This substituted 2-aminobenzenethiol were diluted. The highest dilution showing at least 99% inhibition was taken as MIC. Antibacterial activities¹¹ of the bacterial strains were carried out in Luria broth (Himedia) medium and all fungi were cultivated in Sabouraud Dextrose Agar (Himedia) at pH 6.9 by the spectrophotometric method and serial dilution followed by incubated on a rotary shaker at 37°C. At the end of incubation period, MIC values were recorded.

In the present study, a novel substituted 2-aminobenzenethiols was synthesized using commercially available starting materials and identified by spectral analysis and have shown moderate to significant biological properties. The different bacterial and fungal strains showed different susceptibility towards the tested compound, which may be possibly explained by different resistance mechanism exhibited by each strain. The MIC values of antibacterial screening (2-Amino-3,5-dichloro-6-fluoro-benzenethiol show 128 mg/ml against Escherichia coli MTCC 2939, 116 mg/ml against Bacillus subtilis MTCC 441 and 118 mg/ml against Streptomyces griseus MTCC 1998) revealed that the moderate antibacterial activities against all the three selected strains of bacteria was exhibited by compound. The motive of our research is to extend the area of research by synthesizing new and better templates of 2-aminobenzenethiols and screening them as potential antibacterial drugs but further biomedical research is required.

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