Synthesis and Antimicrobial Screening of Some Novel 2-amino Benzothiazole Derivatives

Heterocyclic compounds, also called heterocycle, it is major class of an organic chemical components identify by the fact some or all the molecules in that atoms are joined in rings containing at least one or more molecule an atom other than carbon. Some hetero atoms such as nitrogen, oxygen and sulphur¹. Heterocyclic system is used in building blocks for new materials for chemical and biological properties. There are number of compounds such as vitamins, hormones, essential amino acids, alkaloids, dyes, and various types of drugs that have heterocyclic ring systems². There was various number of basic heterocyclic compounds such as pyrrole, pyrrolidine, pyridine, pyrimidine, furan, having important application in drug design³.

Medicinal chemistry plays a vital role in drug design and discovery. Medicinal chemistry mainly focus on new drug discovery, design and development. Medicinal chemists also focus on finding the basis of an disease with the help of biochemistry. After the identification of the chemical structure of the targets, it becomes easy to design new drugs for the treatment of disease⁴. 2-aminobenzothiazoles is an important heterocyclic compound shows the biological activity.

There is the major problem that we are facing in the content of infectious disease is the spreading of the antimicrobial resistance. The studies of new series of 2-aminobenzothiazole derivatives for infectious disease such as the antimicrobial activities. This the heterocyclic nuclei such as 2-aminobenzothiazole hence they are the basically two main rings are present like benzene ring and thiazole ring⁵. Thiazole are the five membered ring and one benzene ring also be attached with amine group named as 2-amino benzothiazole because amine group as 2- position in thiazole ring. In the present study on 2-amino benzothiazole derivatives and they are fused heterocyclic compound. 2-aminobenzothiazole derivatives are the reported for some diversified activity to seen such as antimalarial, anthelmintic, antitumor, anticonvulsant activities show these biological activities⁶.

These are highly reactive compound and its utilized-on reaction intermediate to synthesized different derivatives. They are studied as a central muscle relaxant and they have been further studied to found a diverse reactive and broad spectrum of biological activities.

Table 1: Characteristics of 2-aminobenzothiazole

MATERIAL AND METHOD

Reagents and Solvents:

Chemicals and solvents used for the research work commercially procured from Sigma Aldrich, S.D. Fine Merck, Qualigens, Yarrow chemical and GLR Innovation. Aminobenzothiazole, Triethylamine, Chloroacetylchloride, 1-Bromo-3-Chloro propane, 2-Nitrobenzene sulfonamide, Sulfanilamide, sulfanilic acid, 4-chlorobenzene sulfonamide, 2-chlorobenzene sulfonamide, Acetone, Benzene, Ethanol ⁷etc.

The silica gel G used for analytical chromatography (TLC) was obtained from Qualigens. Solvent system used were Benzene: Methanol (8:2) cyclohexane: ethylacetate (8:2) Benzene: Acetone (8:2).

Iodine chambers and U.V. lamps were used for visualization of TLC spots.

Apparatus & glassware used:

Reflux condenser, round bottom flask, beaker, separating funnel, iodine flask, glass rod, magnetic stirrer, pipettes, heating mantle and TLC plates, magnetic stirrer with hot plate, melting point apparatus, hot air oven, volumetric flask, magnetic beads, UV chamber and weighing balance etc.

Analytical Work:

The melting points were found in an open glass capillary using a melting point equipment (Veego, Bombay, India). Using a variety of solvent systems, iodine vapours, and a UV chamber as visualizing agents, reactions were tracked using thin layer chromatography (TLC) on silica gel G plates.

Material used for antimicrobial assay:

Autoclave (Sciencetech), Incubator (Scientific Laboratory India), Nutrient Agar (Himedia, E. coli (Escherichia coli) (Gram-negative, Hi-media), B. subtilis (Bacillus subtilis) (Gram- positive, Hi-media), Sulfathiazole (GLR Innovation).

Scheme1: Synthesis of 2-aminobenzothiazole derivatives ^{8, 9}. (Compound A1-A2)

Table 2: Physical data of synthesized compounds (A1-A2)

Sr. No	1	2
Compounds	A1	A2
Molecular formula	C15H14O3N4S2	C15H13O4N3S2
Molecular weight	369 gm/mol	363 gm/mol
Appearance	Pale yellow ppt.	Brown ppt.
Yield	33.3%	42%
Rfvalue	0.73	0.36
Solubility	Soluble in methanol and DMSO	Soluble in methanol and DMSO
Melting point(Co)	97oC	141oC

Acetamide (0.01mol,1.5gm) and Sulphanilamide (0.01mol,1.7gm) in presence of potassium carbonate or triethylamine and the solvent used as benzene was stirred about 6-7 hours at room temperature¹⁰.The solid was filtered. The solvent system was used as Benzene: Methanol (8:2), Cyclohexane: Ethyl acetate (8:2).

Spectral data:

Solid, mp-96-98, IR (VmaxKBr): 3363.8 (-NH Aromatic, stretching vibration), 2924.03 (-CH Aliphatic, Stretching), 3739.1 (-OH, Stretching), 1384.6 (-S=O, Stretching), 1607 (C=O, Stretching) MS:(ESI+)m/z=369 (M+)

Synthesis of 4-(2-(Benzothiazole-2-ylamino)-2-oxoethyl) amino) benzene sulfonic acid. (Compound A2)

Same procedure was followed as mentioned in the synthesis of (compound A2) only sulphanilamide replaced by sulfanillic acid. System was used as benzene: methanol (8:2). Cyclohexane: Ethyl acetate (8:2).

Spectral data:

Solid, mp- 140-142, IR (VmaxKBr): 3384 (-NH Aromatic, stretching vibration), 3060 (-CH Aliphatic, Stretching),1315 (-S=O, Stretching), 1637 (C=O, Stretching), 2360(S-H, Streching)

MS:(ESI+)m/z=364(M+1)

Scheme 2: Synthesis of 2-aminobenzothiazolederivatives^{11, 12}. (Compound B1-B2)

Table 3:Physical data of synthesized compound (B1 and B2)

Sr. No	1	2
Compounds	B1	B2
Molecular formula	C16H18O2N4S2	C16H17O3N3S2
Molecular weight	362 gm/mol	363.45 gm/mol
Appearance	White ppt.	White powder
Yield	38.6%	48.2%
Rf value	0.71	0.51
Solubility	Soluble in methanol and DMSO	Soluble in methanol and DMSO
Melting point(Co)	80-82	92-94

Synthesis of N-3-Chloropropyl-benzothiazole-2-amine¹³ (IntermediateY1)

1-bromo-3-chloropropane (1.07 mol, ml)was added dropwise to astirred solution of2-amino benzothiazole (0.01mol, 1.5 gm) in the presence of Triethyl amine (0.01mol, 0.78 ml) under an ice-cold environment, with benzene as the solvent. At room temperature, the reaction mixturewas stirred for 6 to 7 hours N-3-Chloropropyl-benzothiazole-2-amine was produced when the solids were filtered, washed with water, and dried and synthesized.

Synthesis of 4-(3-(Benzothiazol-2-ylamino) propyl) amino) benzene sulfonamide^{14, 15}. (Compound B1)

Equimolar solution of N-3-Chloro propyl-benzothiazole-2-amine (0.01mol, 1.5 gm) and Sulphanilamide (0.01mol, 1.7 gm) in presence of potassium carbonate or triethyl amine and the solvent used as dichloromethane was stirred about 6-7 hours at room temperature. The solid was filtered. The solvent system was used as Benzene: Methanol (8:2), Cyclohexane: Ethyl acetate (8:2).

Spectral data:

Solid, mp-80-82, IR (VmaxKBr): 3371.8 (-NH Aromatic, stretching vibration), 2924.03 (-CH Aliphatic, Stretching), 3739.1 (-OH, Stretching), 1384.6 (-S=O, Stretching)

MS:(ESI+)m/z=363 (M+1)

Synthesis of 4-(3-(Benzothiazol-2-ylamino) propyl) amino) benzene sulfonic acid. (Compound B2)

Same procedure was followed as mentioned in the synthesis of (compound B2) only sulphanilamide replaced by sulfanillic acid.

Spectral data:

Solid, m p- 92-94, IR (VmaxKBr): 3254 (-NH Aromatic, stretching vibration), 2935.4 (-CH Aliphatic, Stretching), 1309.5 (S=O, Stretching)

Evaluation of Anti-microbial activity

The cup plate method was employed for the purpose of antimicrobial testing. Nutrient Agar was carefully dispensed into the sterilized petri dishes, with each dish receiving 25-30 mL of the medium. The poured material was allowed to solidify for 30 minutes, after which cups with a diameter of 3 mm created¹⁶.

The test compound solution (0.1 mL) was carefully dispensed into these cups using a micropipette. The plates underwent incubation at 37 oC for a duration of 24 hours. The antimicrobial agents permeate the agar medium.The developmental process of the microbial strain is inhibited, followed by the measurement of the diameter of the zone of inhibition.

RESULT AND DISCUSSION

Docking study of synthesized compounds

Docking of designed compounds (A1-A2 and B1-B2) was performed by Auto dock 1.4.6 software on topoisomerase receptor for anti-microbial screening (PDB: 2POG). The docking score of compounds was ranging between -9.5 to -8.59. All compounds possessed better docking scores than the standard drug, Norfloxacin (-9.8). Compound A1, A2 and B1 having 2-aminobenzothiazole moiety showed the good docking score (-9.5, -9.5 and -9.2). Other compounds in the series also exhibited a good score. The-S-of amino benzothiazole in good docking scored compound A1, A2 and B1 involved in hydrogen bonding interaction with ALA 1120, ARG 1122, ASP 1083, GLU 1088, SER 1084 via -H of -NH.

Table 4: Docking Study of compound A1

Table 5: Docking score of designed compounds A2

Table 6: Docking score of designed compounds B1

Table 7: Docking score of designed compound B2

Docking score of standard drugs

Table 8: Docking score of Standard drugs Norfloxacin

Chemistry

Results of synthesis of compoundA1 and compound A2

The intermediate compound (Intermediate X1, N- Benzothiazole- 2-yl-2-chloro-acetamide) was synthesized by the reaction of 2- amino benzothiazole with chloroacetyl chloride in the presence of triethyl amine and solvent was used as benzene.

The final compound N-(Benzothiazole-2-yl)-2-((4-sulfamoylphenyl) amino) acetamide (CompoundA1)was synthesized by the stirring of (IntermediateX1,N-Benzothiazole-2-yl- chloroacetamide) and sulphanilamide (both are dissolved in methanol)in the presence of TEA and benzene about 6 hr. at room temperature.

The compound 4-(2-(Benzothiazole-2-ylamino)-2-oxoethyl) amino) benzenesulfonic acid (CompoundA2)was synthesized by the stirring of (IntermediateX1, N-Benzothiazole-2-yl- chloro acetamide) and sulfanilic acid (both are dissolved in ethanol) in the presence of TEA and benzene about 6 hr. at room temperature.

Progress of the reaction was checked by TLC and their structures was confirmed by IR and Mass spectroscopy. The spectral data are in total agreement with the proposed structures.

Table 9: Physical data of synthesized compounds (Compounds A1-A2).

Results of synthesis of compound B1and B2

The intermediate compound (Intermediate Y1, N-3-Chloro propyl- benzothiazole-2-amine) was synthesized by the reaction of 2- amino benzothiazole with 1-bromo-3-chloro propane in the presence of triethyl amine and solvent used as benzene.

The final compound 4-(3-(Benzothiazol-2-ylamino) propyl) amino) benzene sulphonamide (CompoundB1) was synthesized by the stirring of (IntermediateY1, of N-3-Chloropropyl- benzothiazole-2-amine) and sulphanilamide (both are dissolved in methanol) in the presence of TEA and benzene about 6 hr. at room temperature.

The final compound 4-(3-(Benzothiazol-2-ylamino) propyl) amino) benzenesulfonic acid (CompoundB2)was synthesized by the stirring of (Intermediate Y1, of N-3-Chloropropyl- benzothiazole-2-amine)and sulphanillic acid (both are dissolved in methanol)in the presence of TEA and benzene about 6 hr.

Progress of the reaction was checked by TLC and their structures was further confirmed by IR and Mass spectroscopy. The spectral data are in total agreement with the proposed structures.

Table 10: Physical data of synthesized compounds (Compounds B1 and B2)

Among all the synthesized compound compound A1 showed the higher zone of inhibition (22 mm, 28 mm and 33 mm diameter respectively), but lesser than the standard drug i. e. Norfloxacin (ZOI- 38 mm, 42 mm and 48 mm diameter) against Bacillus subtilis (gram-positive bacteria).

Among all the synthesized compound compound A2 showed the (21 mm, 26 mm and 32 mm diameter respectively), but lesser than the standard drug i. e. Norfloxacin (ZOI-38 mm, 42 mm and 48 mm diameter) against Bacillus subtilis (gram-positive bacteria).

Among all the synthesized compound compound B1 showed the higher zone of inhibition (20 mm, 24 mm and 26 mm diameter respectively), but lesser than the standard drug i. e. Norfloxacin (ZOI-38 mm, 42 mm and 48 mm diameter) against Bacillus subtilis (gram-positive bacteria)

Among all the synthesized compound compound B1 showed the higher zone of inhibition (22 mm, 25 mm and 28 mm diameter respectively), but lesser than the standard drug i. e. Norfloxacin (ZOI-26 mm, 32 mm and 36 mm diameter) against Escherichia coli. Among all the synthesized compound compound B2 showed the (20 mm, 22 mm and 25 mm diameter respectively), but lesser than the standard drug i. e. Norfloxacin (ZOI-26 mm, 32 mm and 36 mm diameter) against Escherichia coli (gram-negative bacteria).

Table 11: Antimicrobial data of synthesized compounds

Compound A1                                            Compound A2

Compound B1                                                     Compound B2

Figure: TLC of Final Compounds

Fig.5.5 Escherichia coli                                          Fig. : Bacillus subtilis