Synthesis and Antimicrobial Evaluation of  [(1H-benzotriazol-1-ylacetyl) amino] Acetic Acid Derivatives

 

C. M. Jamkhandi*, John Intru Disouza.

Department of Pharmacy JJT University Jhunjhunu  Rajasthan India.

*Corresponding Author E-mail: cmjamakhandi@gmail.com.

 

ABSTRACT:

Five novel [(1H-benzotriazol-1-ylacetyl)amino]acetic acid derivatives were synthesized by substituting corresponding amino acids on a reaction intermediate 1H-benzotriazol-1-ylacetyl chloride. The derivatives were characterized by physical, chemical parameters, TLC and spectral data. Synthesized derivatives from Ib to Vb were screened for anti microbial activity against, S. Aureus, B. Subtilis,  E. coli, S. typhi, C Albicanse, A Niger. MIC was measured against norfloxacine and ketoconazole. Present study confirms the synthesized derivatives endowed with effective anti-microbial activities.

 

KEY WORDS: Benzotriazole, Antibacterial, Anti fungal, MIC, Benzotriazole derivatives

 


INTRODUCTION:

The derivatives of Benzotriazole are endowed with various biological activities such as analgesic, antibacterial, antifungal activities1-6 antifilarial activities7, and Benzotriazole derivatives reported for anticonvulsant and anti-inflammatory8, antitumor9 activities; literature study also reveals the antiviral activity10. The methods of synthesis of Benzotriazole derivatives with different techniques have been reviewed11.  Use of Benzotriazole as synthetic auxiliary was studied12-13. Syntheses of N-protected aminoacyl derivatives of the Benzotriazole were reported14. Treatment of microbial infections is challenging task from time immemorial to present day, as many new infections have born and some old infections are resurfacing due to resistance to antibiotics and chemotherapeutic agents. To keep pace with genetic changes of microorganisms there is need for continuous development of anti-infective agents.

 

The Synthesis of [(1H-benzotriazol-1-ylacetyl)amino]acetic acid Derivatives was effected by substitution of corresponding amino acids on reaction intermediate 1H-benzotriazol-1-ylacetyl chloride which was synthesized by following reported procedures and  following the below scheme (scheme 1) to get possible potent anti-infective derivatives12-18.

 

MATERIAL AND METHODS:

Procedure of Synthesis

Synthesis of Benzotriazole (a): 10.8 gm of O-phenylenediamine is added to mixture of 12g (11.5 ml) of glacial acetic acetic acid and 30 ml   of water, which is cooled to 15oC, stir. Then solution of 7.5g of sodium nitrite in 15 ml water is added in portion. The temperature rises slowly to 85oC and then cools slowly. When temperature is 45oC the mixture is chilled at ice bath for 30 min. Pale brown solid separated by the filtration. The recrystallization is done using benzene as solvent.

 

Synthesis of ethyl 1H-benzotriazol-1-ylacetate (b): A mixture of Benzotriazole (0.1M), ethyl chloroacetate (0.1M) and 0.3g of K2CO3 in 60 ml of acetone was stirred for 10 hrs. The solvent was removed under reduced pressure. A solid mass was produced and then needle shaped brown crystals were obtained after recrystallization from the mixture of chloroform and ether (8:2%V/V).The yield obtained was 60% and M.P. was 40oC.

 

Synthesis of 1H-benzotriazol-1-ylacetyl chloride(c): A 250-mL, three-necked flask, equipped with a magnetic stirbar, condenser, thermometer, and addition funnel, is charged with 10.0 g Benzotriazole and 34 mL of chloroform (CHCl3). At 25°C, 22 mL (600 mmol) of thionyl chloride and 1 drop of dimethylformamide (DMF) are added, followed by heating the mixture at 68°C for 3 hr. After the initial suspension turns into a yellow solution, the heating source is removed and the acid chloride precipitates as a pale red solid. After cooling the reaction mixture to 25°C, the solid is collected via filtration using a Buchner funnel, washed with CHCl3, and dried in a vacuu desiccator for 15 hr to give 5.12 g (90%) of as a white powder.


Figure: Scheme for synthesis of [(1H-benzotriazol-1-ylacetyl) amino] acetic acid derivatives

 

Scheme 1: Reaction pathway

 

 

 


Substitution of amino acids on (c) to get the below derivatives.

Synthesis of [(1H-benzotriazol-1-ylacetyl)amino]acetic acid (Ib): the compound (c) was treated with glycine  in equimolar concentration and refluxed for 4 hrs in benzene solvent. Spectral data FTIR (KBr cm-1): 2835.93 (HC Aromatic), 1493.93 (C=C Str), 1677.21(-C=O). 1H-NMR (DMSO 1δ ppm): 6.9(1H, -CONH-), 7.2 (4H, d,Ar ), 12.2 (1H, COOH),  m/z of m+  ion is 234.

 

Synthesis of 2-[(1H-benzotriazol-1-ylacetyl)amino] propanoic acid (IIb): the compound (c) was treated with equimolar  alanine and refluxed for 4 hrs in benzene solvent. Spectral data FTIR (KBr cm-1): 3017.14 (HC Aromatic), 1508.24 (-N=N str), 1682.70 (-C=O). 1H-NMR (DMSO 1δ ppm): 1.4-1.5(3H, –CH3), 7.4(1H, -NHCO-), 7.33-7.36 (4H, d,Ar ). m/z of m+  ion is 248.

 

Synthesis of 2-[(1H-benzotriazol-1-ylacetyl)amino]-3-(4-hydroxyphenyl)propanoic acid (IIIb): the compound (c) was treated with equimolar tyrosine and refluxed for 4 hrs in benzene solvent. Spectral data FTIR (KBr cm-1): 2926.32 (HC Aromatic), 1559.59 (-N=N str), 1357.48 (-C-N str), 3024.48 (-NH- str), 3320.00 (Ar-OH). 1H-NMR

 (DMSO 1δ ppm): 2.4(1H, -CH=), 3.6(2H, -CH2CO), 3.9(1H, Ar-OH), 6.6(1H, -NH-),7.2-7.3 (4H, d,Ar ), 7.4(1H, -NHCO-). m/z of m+  ion is 340.

 

 

Synthesis of 2-[(1H-benzotriazol-1-ylacetyl)amino]-5-carbamimidamidopentanoic acid (IVb): the compound (c) was treated with equimolar arginine and refluxed for 4 hrs in benzene solvent. Spectral data FTIR (KBr cm-1): 2857.39 (HC Aromatic), 1539.25 (-N=N str), 1671.04 (-C=N- str), 3271.38 (-NH2- str) . 1H-NMR (DMSO 1δ ppm):

1.9(-CH2-), 2.8(2H, -NH2), 7.1-7.2 (4H, d,Ar ). m/z of m+  ion is 333.

 

Synthesis of 4-[(1H-benzotriazol-1-ylacetyl)amino] benzoic acid (Va): the compound (c) was treated with equimolar cystiene and refluxed for 4 hrs in benzene solvent. Spectral data FTIR (KBr cm-1): 2925.12 (HC Aromatic), 1507.22 (-N=N str), 1678.68  (-C=O). 1H-NMR (DMSO 1δ ppm): 12.2(1H, -COOH), 7.1-7.2 (4H, d,Ar ). m/z of m+  ion is 280.30.

 

The melting points of the synthesized derivatives were determined by open capillary (LABHOSP) and were uncorrected. The purity of the compounds was checked using pre coated TLC plates (MERCK, 60F) using Benzene: chloroform: methanol (8:4:2) solvent system.

 

The developed chromatographic plates were visualized under UV at 254nm. IR spectra were recorded using KBr on Shimadzu FTIR model 8400 spectrophotometer and Agilent Technologies CARY 630 FTIR,1H NMR spectra in DMSO on a BRUKER FT-NMR instrument using TMS as internal standard.

 


 

 

Table 1: Physicochemical properties of the Benzotriazole derivatives(Ib to Vb)                                                                              

Deri vative

Mol. Weight

Derivative Name

R

% Yield

MP

Ib

C10H10N4O3

[(1H-benzotriazol-1-ylacetyl)amino]acetic acid

H

68

1820C

IIb

C11H12N4O3

2-[(1H-benzotriazol-1-ylacetyl)amino]propanoic acid

-CH3

62

207 oC

IIIb

C17H16N4O4

2-[(1H-benzotriazol-1-ylacetyl)amino]-3-(4-hydroxyphenyl)propanoic acid

 

66

227 oC

IVb

C14H19N7O3

 

2-[(1H-benzotriazol-1-ylacetyl)amino]-5-carbamimidamidopentanoic acid

 

63

210 oC

Vb

C11H12N4O3S

2-[(1H-benzotriazol-1-ylacetyl)amino]-3-sulfanylpropanoic acid

 

67

202 oC

 

 


Antimicrobial Activity:

The derivatives synthesized were evaluated for antibacterial and antifungal activity with agar diffusion method using and Norfloxacine and Ketaconazole as standard drugs in concentration of 1µg/ml (Fig 1 and 2). The bacterial strains of B. subtilis, S. aureus, E. coli, and S. typhi were used for antibacterial activity and A. niger and C. albicanus strains were used for antifungal activity. Overnight kept broth cultures were used for MIC determinations. Agar dilution MICs were determined by using agar plates with an incorporated standard drug dilutions and samples with incubation at 37°C and were defined as the lowest antibiotic concentrations completely inhibiting growth. Broth dilution MICs were determined with overnight broth cultures of the strains to be tested.

 

Figure 1: Graphical representation Antimicrobial activity.

 

Figure 2: Graphical representation Antifungal activity

RESULTS AND DISCUSSION:

Substitution of amino acids in 1H-benzotriazol-1-ylacetyl chloride was produced various derivatives (Ib-Vb)). The derivatives synthesized were characterized by the physical properties (Table 1), chemical properties, spectral data like IR, NMR and Mass spectra. The synthesized derivatives were evaluated for their antimicrobial activity using  Norfloxacin and Ketoconazole as standard.

 

CONCLUSION:

 The synthesized derivatives like I b to Vb showed comparable Zone of inhibition with standards and have good antibacterial and antifungal activities.

 

 

ACKNOWLEDGEMENTS:

Authors are thankful to Shri. G. D. Patil, Secretary Shree Warana Vibhag Shikshan Mandal Warananagar for providing laboratories facilities. Authors are thankful to Mrs. Dr. U. S. Chougule, and Mr Krishnath Paymal, IISc Bangalore, CFC and Chemistry Dept of Shivaji University Kolhapur for kind assistance in microbial and spectral data.  

 

REFERENCES:

1.       Shukla DK, Srivastava SD. Synthesis of some new 5-[2-{(1,2,3-benzotriazole)-1-yl-methyl}-1'-(4'-substituted aryl-3'-chloro-2'-oxo azetidine)]-amino-1,3,4-thiadiazoles: Antifungal and antibacterial agents. Indian J of Chem.2008,47B;463-69.

2.       Asati KC, Srivastav SK, Srivastav SD. Synthesis of some new 5-arylidene-2-aryl-3-(benzotriazoloacetamidyl)-1,3-thiozolodine-4-ones as analgesic and antimicrobial agents. Indian J of Chem.2006,45B;526-31.

3.       R Ramachandran, M Rani, S Senthan, YT Jeong, S Kabilan. Synthesis, spectral, crystal structure and in vitro antimicrobial evaluation of imidazole/benzotriazole substituted piperidin-4-one derivatives. European J Med Chem 2011;46:1926-34.

4.       Z Rezaei, S Khabnadideh, K Pakshir, Z Hossaini, F Amiri, El Assadpour. Design, synthesis, and antifungal activity of triazole and benzotriazole derivatives. European J Med Chem 2009;44:3064-67.

5.       S Khabnadideh1, Z Rezaei, K Pakshir, K Zomorodian, N Ghafari. Synthesis and antifungal activity of benzimidazole, benzotriazole and aminothiazole derivatives. ResPharm Sci  2012; 7(2):1-8.

6.       KP.Namdeo, VK.Singh, SK.Prajapati. Synthesis of Some 2-(Substituted)-5-[(N Benzotriazolomethyl)-1, 3,4-Thiadiazolyl]-4-Thiazolidinones for their Anti-Fungal Activity. Indian J Pharm Educ Res 2009;43(3):266-71.

7.       MA Rahman. Chalcone: A Valuable Insight into the Recent Advances and Potential Pharmacological Activities. Chem Sci J 2011;29:1-16.

8.       KM Dawood, HA Gawad, EA. Rageb, M Ellithey, HA Mohamed. Synthesis, anticonvulsant, and anti-inflammatory evaluation of some new benzotriazole and benzofuran-based heterocycles. Bioorg Med Chem 2006;14 :3672–80.

9.       YA Al-Soud, NA Al-Masoudi, ARS Ferwanah. Synthesis and Properties of New Substituted 1,2,4-Triazoles:Potential Antitumor Agents. Bioorg Med Chem 2003;11;1701-08.

10.     Bretne Maria et al. Synthesis and biological activity of 1H-benzotriazole and 1H-benzimidazole analogues – inhibitors of the NTPase/helicase of HCV and of some related Flaviviridae. Antiviral Chemistry & Chemotherapy 2005;16:315–26.

11.     BV Suma, NN Natesh, V Madhavan. Benzotriazole in medicinal chemistry: An overview. J Chem Pharm  Res 2011;3(6):375-81.

12.     AR Katritzky, Xiangfu L, Wei-Quang F, Benzotriazole as synthetic auxiliary: Benzotriazolyl alkylations and Benzotriazole mediated heteroalkylation. Synthesis 1994; 445-56.

13.     AR Katritzky. The continuing magic of benzotriazole: An overview of some recent advances in synthetic methodology. J Heterocycl Chem1999;36(6):1501-22.

14.     Katritzky AR, Singh A, Haase DN, Yoshioka Y. N-(Fmoc-α-aminoacyl)benzotriazoles: versatile synthetic reagents from proteinogenic amino acids.  Arkivok 2009; (viii): 47-56.

15.     Rajput AP, Gore RP. Synthesis and characterization of some new azole- acetanilides. J Chem Pharm Res 2010;2(5):52-59.

16.     Vogel’s Text Book of Practical Organic chemistry; 4th Edn. Longman Group Ltd 1978; p124.

17.     Gerard Vogel H. Drug Discovery and Evaluation. 2nd ed. Springer – Verlag Berlin Heidlbrg New York 2002. p.49.

18.     Bansal. RK. Heterocyclic Chemistry, 4th. New Age International Publishers.2005;p 475.

 

 

 

Received on 25.07.2012       Modified on 12.08.2012

Accepted on 20.08.2012      © RJPT All right reserved

Research J. Pharm. and Tech. 5(9): September 2012; Page 1197-1200