Journal of Chemistry

Volume 2013 (2013), Article ID 312961, 5 pages

http://dx.doi.org/10.1155/2013/312961

Research Article

Synthesis, Characterization, and Antimicrobial Activity of Methyl-2-aminopyridine-4-carboxylate Derivatives

S. Nagashree,¹ P. Mallu,¹ L. Mallesha,² and S. Bindya¹

¹Department of Chemistry, Sri Jayachamarajendra College of Engineering, Manasagangotri, Mysore 570 006, India

²Postgraduate Department of Chemistry, JSS College of Arts, Commerce and Science, Ooty Road, Mysore 570 025, India

Received 1 June 2012; Revised 29 June 2012; Accepted 9 July 2012

Academic Editor: Hakan Arslan

Copyright © 2013 S. Nagashree et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract.

A series of methyl-2-aminopyridine-4-carboxylate derivatives, 3a–f, were synthesized in order to determine their in vitro antimicrobial activity. The chemical structures of the synthesized compounds were confirmed by elemental analyses, FT-IR, and ¹H NMR spectral studies. Among the synthesized compounds, 3c and 3d showed good antimicrobial activity compared to other compounds in the series.

1. Introduction

The compounds containing an azomethine group (–CH=N–) are important in elucidating the mechanism of transamination and racemisation reactions in biological systems [1, 2]. Due to the great flexibility and diverse structural aspects, a wide range of Schiff bases have been synthesized and their complexation behaviors have been studied [3]. They have been synthesized from a variety of compounds, such as amino thiazoles, 2-hydroxy-1-napthalaniline, amino sugars, aromatic aldehydes, ketones, isatin, triazole ring, thiosemicarbazides, amino acids, and pyrazolone [4, 5]. Antibacterial, antifungal, antitumor, and anticancer activities of Schiff bases have been reported [6, 7] and they are active against a wide range of organisms. Antibacterial activity has been studied more than antifungal activity, because bacteria can achieve resistance to antibiotics through biochemical and morphological modifications [8, 9]. Some Schiff bases bearing aryl groups or heterocyclic residues possess excellent biological activities have attracted the attention of many researchers in recent years [10–12]. The Schiff bases formed from aromatic aldehydes or aromatic ketones and their derivatives are quite stable. Due to the great flexibility and diverse structural aspects of Schiff bases, a wide range of these compounds have been synthesized and their activities have been studied [13, 14]. Antimicrobial and antifungal activities of various Schiff bases have also been reported [15]. Many Schiff bases are known to be medicinally important and are used to design medicinal compounds [16].

In connection with such studies, the present paper reporting for the first time on the synthesis of methyl-2-aminopyridine-4-carboxylate derivatives, 3a–f, which are formed during the reaction of methyl-2-aminopyridine-4-carboxylate (1) with different aldehydes (2a–f). These synthesized compounds were characterized by elemental analyses, FT-IR, and ¹H NMR studies. Antibacterial and antifungal activities were also reported in this paper.

2. Experimental

All chemicals used were of the analytical reagent grade (AR). All solvents and reagents were purchased from Sigma Aldrich Chemicals Pvt Ltd. Melting range was determined by Veego Melting Point VMP III apparatus. Elemental analyses were recorded on Perkin Elmer 2400 Elemental Analyser. The FT-IR spectra were recorded using KBr discs on FT-IR Jasco 4100 infrared spectrophotometer. ¹H NMR spectra were recorded on Bruker DRX—400 MHz spectrometer and TMS as an internal standard.

2.1. General Procedure for the Synthesis of Methyl-2-aminopyridine-4-carboxylate Derivatives, 3a-f

Equimolar concentrations of different aldehydes (0.01 mol) and methyl-2-aminopyridine-4-carboxylate (0.01 mol) dissolved in minimum volume of ethanol, and then 2-3 drops of concentrated sulfuric acid are added to the reaction mixture. The solution was stirred for 4-5 hr at room temperature and the progress of the reaction was followed by TLC until the reaction was complete. It was cooled to 0°C, the precipitate was filtered, washed with ethanol, and recrystallized from methanol. Methyl-2-aminopyridine-4-carboxylate derivatives (3a–f) were synthesized by the method summarized in Scheme 1. The chemical structures and physical data of all the synthesized compounds are tabulated in Table 1.

Table 1: Chemical structures and physical data of the compounds 3a–f.

Compound
R
Yield (%)
m. p. (°C)

3a

80
101–103

3b

85
113-114

3c

78
119–121

3d

77
118–120

3e

75
128–130

3f

82
131–133

Scheme 1

2.2. Methyl-2-ethylideneaminopyridine-4-carboxylate (3a)

The product obtained from methyl-2-aminopyridine-4-carboxylate (1) (1.53 g, 0.01 mol) and acetaldehyde (3a) (0.44 g, 0.01 mol). ¹H NMR (DMSO-d₆, 400 MHz) δ: 9.09 (d, 1H, Ar-H, J = 4.7 Hz), 8.00 (d, 1H, Ar-H, J = 4.2 Hz), 7.90 (s, 1H, Ar-H), 7.46 (q, 1H, CH, J = 8.1 Hz), 3.82 (s, 3H, OCH₃), 1.13 (d, 3H, CH3, J = 3.3 Hz); FT-IR (KBr, cm⁻¹) ν: 1725 (C=O), 1639 (C=N), 1589 (C=C), 1213 (C-O), 1146 (C-N); Anal. calcd for C₉H₁₀N₂O₂ (in %): C 60.66, H 5.66, N 15.72; found C 60.59, H 5.70, N 15.91.

2.3. Methyl-2-benzylideneaminopyridine-4-carboxylate (3b)

The product obtained from methyl-2-aminopyridine-4-carboxylate (1) (1.53 g, 0.01 mol) and benzaldehyde (3b) (1.06 g, 0.01 mol). ¹H NMR (DMSO-d₆, 400 MHz) δ: 9.10 (d, 1H, Ar-H, J = 4.5 Hz), 8.12 (s, 1H, CH), 7.85 (d, 1H, Ar-H, J = 4.0 Hz), 7.68 (s, 1H, Ar-H), 7.34 (d, 2H, Ar-H, J = 5.1 Hz), 7.00 (m, 3H, Ar-H, J = 11.0 Hz), 3.82 (s, 3H, OCH₃); FT-IR (KBr, cm⁻¹) ν: 3057 (Ar-H), 1725 (C=O), 1636 (C=N), 1591 (C=C), 1213 (C-O), 1146 (C-N); Anal. calcd for C₁₄H₁₂N₂O₂ (in %): C 69.99, H 5.03, N 11.66; found C 69.79, H 5.21, N 11.51.

2.4. Methyl-2-(3-hydroxy-4-methoxy-benzylidene)aminopyridine-4-carboxylate (3c)

The product obtained from methyl-2-aminopyridine-4-carboxylate (1) (1.53 g, 0.01 mol) and 3-hydroxy-4-methoxy-benzaldehyde (3c) (1.53 g, 0.01 mol). ¹H NMR (DMSO-d₆, 400 MHz) δ: 9.12 (d, 1H, Ar-H, J = 4.0 Hz), 8.18 (s, 1H, CH), 7.98 (d, 1H, Ar-H, J = 3.9 Hz), 7.80 (s, 1H, Ar-H), 7.45 (d, 1H, Ar-H, J = 2.7 Hz), 7.30 (d, 1H, Ar-H, J = 5.2 Hz), 7.00 (s, 1H, Ar-H), 5.11 (s, 1H, OH), 3.82 (s, 3H, OCH₃), 3.57 (s, 3H, OCH₃); FT-IR (KBr, cm⁻¹) ν: 3262 (O-H), 3057 (Ar-H), 1725 (C=O), 1636 (C=N), 1590 (C=C), 1213 (C-O), 1146 (C-N); Anal. calcd for C₁₅H₁₄N₂O4 (in %): C 62.93, H 4.93, N 9.79; found C 62.72, H 4.81, N 9.78.

2.5. Methyl-2-(1H-Indol-3-ylmethylene)aminopyridine-4-carboxylate (3d)

The product obtained from methyl-2-aminopyridine-4-carboxylate (1) (1.53 g, 0.01 mol) and 1H-indole-3-carbaldehyde (2d) (1.45 g, 0.01 mol). ¹H NMR (DMSO-d₆, 400 MHz) δ: 10.11 (s, 1H, NH), 9.12 (d, 1H, Ar-H, J = 4.2 Hz), 7.98 (d, 1H, Ar-H, J = 5.7 Hz), 7.88 (s, 1H, Ar-H), 7.50 (d, 1H, Ar-H, J = 6.2 Hz), 7.22 (d, 1H, Ar-H, J = 2.3 Hz), 7.10 (s, 1H, CH), 6.95 (s, 1H, Ar-H), 6.75 (t, 1H, Ar-H, J = 6.7 Hz), 6.60 (t, 1H, Ar-H, J = 5.8 Hz), 3.82 (s, 3H, OCH₃); FT-IR (KBr, cm⁻¹) ν: 3477 (N-H), 3060 (Ar-H), 1725 (C=O), 1639 (C=N), 1589 (C=C), 1213 (C-O), 1146 (C-N); Anal. calcd for C₁₆H₁₃N₃O₂ (in %): C 68.81, H 4.69, N 15.05; found C 68.91, H 4.57, N 15.21.

2.6. Methyl-2-(4-ethoxybenzylidene)aminopyridine-4-carboxylate (3e)

The product obtained from methyl-2-aminopyridine-4-carboxylate (1) (1.53 g, 0.01 mol) and 4-ethoxybenzaldehyde (2e) (1.51 g, 0.01 mol). ¹H NMR (DMSO-d₆, 400 MHz) δ: 9.12 (d, 1H, Ar-H, J = 4.1 Hz), 7.98 (d, 1H, Ar-H, J = 3.9 Hz), 7.88 (s, 1H, Ar-H), 7.52-7.45 (d, 2H, Ar-H, J = 5.7 Hz), 7.18 (s, 1H, CH), 6.70-6.53 (d, 2H, Ar-H, J = 6.8 Hz), 3.85 (q, 2H, CH2, J = 6.2 Hz ), 3.82 (s, 3H, OCH₃), 1.39 (t, 3H, CH3, J = 2.7 Hz ); FT-IR (KBr, cm⁻¹) ν: 3477 (N-H), 3060 (Ar-H), 1725 (C=O), 1695 (C=N), 1468 (C=C), 1213 (C-O), 1146 (C-N); Anal. calcd for C₁₆H₁₆N₂O₃ (in %): C 67.59, H 5.67, N 9.85; found C 67.61, H 5.57, N 9.61.

2.7. Methyl-2-(2-pyridine-ylmethylene)aminopyridine-4-carboxylate (3f)

The product obtained from methyl-2-aminopyridine-4-carboxylate (1) (1.53 g, 0.01 mol) and 2-pyridine carboxaldehyde (2f) (1.10 g, 0.01 mol). ¹H NMR (DMSO-d₆, 400 MHz) δ: 9.11 (d, 1H, Ar-H, J = 4.0 Hz), 8.12 (d, 1H, Ar-H, J = 3.7 Hz), 8.00 (d, 1H, Ar-H, J = 5.6 Hz), 7.90 (d, 1H, Ar-H, J = 8.3 Hz), 7.80 (d, 1H, Ar-H, J = 5.2 Hz), 7.55 (s, 1H, CH), 7.42 (t, 1H, Ar-H, J = 3.0 Hz), 7.30 (t, 1H, Ar-H, J = 3.1 Hz), 3.82 (s, 3H, OCH₃); FT-IR (KBr, cm⁻¹) ν: 3477 (N-H), 3060 (Ar-H), 1725 (C=O), 1639 (C=N), 1589 (C=C), 1213 (C-O), 1146 (C-N); Anal. calcd for C₁₃H₁₁N₃O₂ (in %): C 64.72, H 4.60, N 17.42; found C 64.81, H 4.57, N 17.21.

2.8. In Vitro Antibacterial Activity

Antibacterial activity of the synthesized compounds was determined against Gram-positive bacteria (Bacillus subtilis, Staphylococcus aureus) and Gram-negative bacteria (Xanthomonas campestris and Escherichia coli) in DMF by disc diffusion method on nutrient agar medium [17]. The sterile medium (Nutrient Agar medium, 15 mL) in each petri plates was uniformly smeared with cultures of Gram positive and Gram negative bacteria. Sterile discs of 10 mm diameter (Hi-Media) were made in each of the petriplates, to which 50 μL (1 mg/mL, i.e., 50 μg/disc) of the different synthesized compounds were added. The treatments also included 50 μL of DMF and streptomycin as negative and positive control for comparison. For each treatment, three replicates were maintained. The plates were incubated at 25 ± 2°C for 24 h and the size of the resulting zone of inhibition, if any, was determined. The data were subjected to analysis of variance (ANOVA).

2.9. In Vitro Antifungal Activity

The synthesized compounds were screened for their antifungal activity against Candida albicans in DMF by poisoned food technique [18]. Potato Dextrose Agar (PDA) media was prepared and about 15 ml of PDA was poured into each petri plate and allowed to solidify. 5 mm disc of seven days old culture of the test fungi was placed at the center of the petriplates and incubated at 26°C for 7 days. After incubation the percentage inhibition was measured and three replicates were maintained for each treatment. Activity of compounds was compared with standard drug nystatin. All the synthesized compounds were tested (at the dosage of 500 μL of the novel compounds/petriplate, where concentration was 0.1 mg/mL) by poisoned food technique.

3. Results and Discussion

3.1. Schiff Base Characterization

Schiff bases of methyl-2-aminopyridine-4-carboxylate derivatives, 3a–f, were synthesized from the reaction of methyl-2-aminopyridine-4-carboxylate with different aldehydes. The elemental analyses data showed good agreement between the experimentally determined values and the theoretically calculated values within the limits of permissible error. The absorptions around 3000 cm⁻¹ in synthesized compounds confirm the aromatic stretching vibrations, and the appearance of a medium to strong absorption bands above 1600 cm⁻¹ due to a stretching vibration of the azomethine (C=N) bond formation in synthesized compounds via condensation. The proton spectral data agree with respect to the number of protons and their chemical shifts with the proposed structures. The proton spectral data of methyl-2-aminopyridine-4-carboxylate (1) shows resonance at δ 5.50 ppm (s, 2H, -NH₂). In compounds 3a–f, the above resonance disappeared and additional resonances assigned to the –CH=N (δ 7.10–8.18 ppm) were observed, which confirmed the condensation between the amino group and carbonyl group. The ¹H NMR spectra of 3c and 3d are shown in Figures 1 and 2, respectively.

Figure 1

Figure 2

3.2. Antibacterial Activity

The antibacterial activity of compounds 3a–f was evaluated and compared with streptomycin as standard drug. Compounds 3c and 3d showed good antibacterial properties against four pathogenic bacterial strains compared with other compounds. Compound 3c exhibits inhibition in the range of 26 mm against B. subtilis. Compound 3d exhibits inhibition in the range of 25 mm against B. subtilis. Compounds 3e and 3f showed moderate antibacterial activity against four pathogenic bacterial strains. Compounds 3a and 3b showed weak antibacterial activity against S. aureus and X. campestris.

3.3. Antifungal Activity

All the synthesized compounds 3a–f were also tested against C. albicans. The compounds 3c and 3d showed good antifungal activity than other compounds in the series against tested strain. Compound 3c showed inhibition in the range of 16 mm when compared with other compounds in the series against C. albicans. From the results, it is evident that compounds 3a and 3b are moderately active. The antimicrobial activity results of synthesized compounds were compared with standard drugs as depicted in Table 2. Compounds 3a–f showed antimicrobial activity in the order of 3c > 3d > 3f > 3e > 3b> 3a against tested bacterial and fungal strains.

Table 2: In Vitro antibacterial and antifungal activities of compounds 3a–f.

Compound
Zone of inhibition in diameter (mm)

B. subtilis
S. aureus
X. campestris
E. coli
C. albicans

3a
21
11
12
12
10

3b
22
12
13
12
10

3c
26
15
24
16
16

3d
25
14
23
14
15

3e
22
12
19
16
12

3f
23
13
20
14
13

Streptomycin
26
18
25
20
—

Nystatin
—
—
—
—
20

Initial structure activity relationship can be drawn for the compounds 3a–f. In the present study, different electron withdrawing and electron donating groups attached to aromatic ring as substituent were linkage to azomethine group. The close survey of antimicrobial efficacy indicated that the inhibition values of all the compounds exhibited a varied rang (11–26 mm and 10–16 mm) of antibacterial and antifungal activities against all the tested microbial strains. The electron donating hydroxyl group and methoxy group of phenyl ring in 3c produces enhanced activity. The electron withdrawing pyridine ring in 3f and electron donating ethoxy group in 3e showed moderate activity, whereas the indole in 3d produces good antimicrobial activity. The compound 3a is weakly active and this may be due to the absence of phenyl ring. However, 3b showed better activity compared to 3a probably due to the presence of benzene in 3b. The above SAR correlation studies reveal that the nature of the linkage (substituent on aromatic ring) influences the antimicrobial activity.

4. Conclusion

In conclusion, in a series of methyl-2-aminopyridine-4-carboxylated derivatives, 3a–f were synthesized in good yield, characterized by different spectral studies, and their antimicrobial activities were determined against clinically important pathogens. Compounds 3c and 3d demonstrated good inhibition against bacterial and fungal strains tested. The SAR studies revealed that the substituents on aromatic ring are responsible for the antimicrobial activity of these classes of agents. On the basis of their activity, these derivatives were identified as viable leads for further studies.

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