Introduction

An essential component of the search for new leads in a drug designing program is the synthesis of molecules, which are novel yet resemble known biologically active molecules by virtue of the presence of some critical structural features. Certain small heterocyclic molecules act as highly functionalized scaffolds. These are known pharmacophores of a number of biologically active and medicinally useful molecules (Silverman, 1992; Thompson and Ellman, 1996; Robert, 2000). The rapidly increasing occurrence of multiple drug-resistant microbial strains is a serious problem. As the emergence of anti resistant bacteria is inevitable, there is an urgency for the discovery of novel active agents, which is of the highest priority (Norrby, 2001; Tomasz, 1994). Previous reports reveal that benzoxazole derivatives are biologically significant compounds and are known to exhibit various biological activities such as anticancer (Kumar et al., 2002; Easmon et al., 2001; Jauhari et al., 2008; Rida et al., 2005), antimicrobial (Kumar et al., 2002; Temiz-Arpacı et al., 2005), anti-HIV (Rida et al., 2005; Temiz-Arpacı et al., 2005), and dopamine D4 agonists (Wang et al., 2005). The substitution at second position in benzoxazole skeleton is influential for the biological activity of the molecule. Thus, the 2-substituted bis(benzoxazole), UK-1 (A) is a natural product (Fig. 1) (Ueki et al., 1993) and it showed a wide spectrum of potent anticancer activity against leukemia and lymphoma. The cytotoxicity of the synthetic analogs (Kerwin and Mckee, 2008) of UK-1 was studied toward selective cancer cell lines. Routiennocin (B) (Fig. 1) is a spiroketal antibiotic, isolated from a strain of Streptomyces chartreusis possessing a benzoxazole ring in its molecular structure, was found to be very active especially against some Gram-positive bacteria by acting as a good ionophore (Prudhomme et al., 1986; Martin et al., 1992). Chlorzoxazone (C) is a muscle relaxant, benoxaprofen (D) and flunoxaprofen (E) are anti-inflammatory drugs and these molecules contain benzoxazole pharmacophore. The chemical structures of the drugs are depicted in Fig. 1.

Fig. 1
figure 1

Structures of benzoxazole drugs and biomolecules

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The 2(3H)-benzoxazolone heterocycles are considered as “privileged scaffolds” in the design of pharmacological probes. The 2(3H)-benzoxazolone heterocycles and its bioisosteric surrogates such as 2(3H)-benzothiazolinone, benzoxazinone, etc., have received considerable attention from the medicinal chemists owing to their capacity to mimic a phenol or a catechol moiety in a metabolically stable template (Poupaert et al., 2005). The benzoxazolone heterocycle has high flexibility in chemical modifications, allowing changes to the characteristics of side-chains on a rigid platform (Chiarotto et al., 2009). Due to that, this template has very broad therapeutic applications. The 2(3H)-benzoxazolone heterocycles exhibits various biological activities like anti-HIV (Deng et al., 2006), anticancer (Ivanova et al., 2007), analgesic (Unlu et al., 2003), anti-inflammatory (Koksal et al., 2005), antinociceptive (Onkol et al., 2001), antimicrobial (Koksal et al., 2002), anticonvulsant (Ucar et al., 1998), antimalarial (Courtois et al., 2004), PPARγ agonist (Blanc-Delmas et al., 2006). The functionalization of the nitrogen atom at the third position of benzoxazole is of interest since the electronic characteristics of this atom can be decisive for the biological activity. Several nitrogen heterocycles containing piperazine moiety have been described as potent chemotherapeutic agents. This cyclic amine moiety is also found in drug candidates displaying antiallergic (John et al., 1995), antibacterial (Michel et al., 1992), antianxiety, antiemetic, antimigraine (Christine et al., 1996), and anti-inflammatory (Dogruer et al., 1998; Gulcan et al., 2003) activities. As a consequence, the development of general economical methods for the synthesis of benzoxazole derivatives containing piperidine moiety in a single molecule frame work has been the subject of considerable synthetic effort.

Discussion

Chemistry

In our on going research program of drug designing for new leads, various 2-cyclic amine substituted-1,3-benzoxazoles (5a–l), 5-chloro-3-alkyl substituted-1,3-benzoxazol-2(3H)-ones (8a–f), and 3-[3-(cyclic amine)propyl]-1,3-benzoxazol-2(3H)-ones (9a–f) were synthesized and were evaluated for antibacterial and antifungal activities. The 2-chlorobenzoxazole (3) was commercially available chemical and also otherwise we prepared starting from 2-amino phenol (1) (Yamada et al., 1998). The 2-aminophenol was treated with carbon disulfide under basic conditions to obtain 2-mercapto-1,3-benzoxazole (2), which was further reacted with phosphorus pentachloride (PCl5) to obtain 2-chlorobenzoxazole (3). The 2-chlorobenzoxazole was treated with various cyclic amines (4) in acetonitrile at 0°C to obtain the compounds 5a–l (Scheme 1). The products and the corresponding yields are listed in Table 1. When 1 equivalent of 2-methylpiperazine was reacted with 2-chlorobenzoxazole, only mono alkylated product 5i formed, i.e., alkylation occurred at the less hindered nitrogen. Similarly, when 1 equivalent homopiperazine reacted with 2-chlorobenzoxazole, only mono alkylated product 5j formed.

Scheme 1
scheme 1

Synthesis of 2-cyclic amine benzoxazoles (5a–l)

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Table 1 Newly synthesized piperazinyl benzoxazoles (5a–l) derivatives
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Compounds 8a–f and 9a–f were synthesized starting from commercially available 5-chlorobenzoxazolone (6) as shown in Scheme 2 and the results were mentioned in Table 2.

Scheme 2
scheme 2

Synthesis of 3-substituted benzoxazolones (8a–f and 9a–f)

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Table 2 Newly synthesized benzoxazole-2(3H)-ones (8a–f, 9a–f) derivatives
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The alkylation of benzoxazolone under basic conditions, which makes use of base like K2CO3, is known in literature (Soyer et al., 2005). The method involves tedious work up, gives poor to moderate yield with less purity of the product. Hence, herein we report a practical and convenient method for the alkylation of benzoxazolone using cesium fluoride–Celite (CsF–Celite). The synthetic utility of CsF–Celite to synthesize ethers and esters was earlier reported (Shah et al., 2002; Polshettiwar and Kaushik, 2005). The alkylation of benzoxazolone catalyzed by CsF–Celite was carried out under mild reaction conditions with high yields and simple work up. Thus, we explored the utility of CsF–Celite system as an efficient, inexpensive, economical, non-corrosive, and eco-friendly reagent for the alkylation process. The preparation of the CsF–Celite was carried out in the same manner as described earlier (Hayat et al., 2001). A comparative yields of alkylations using K2CO3 (Method A) and CsF–Celite (Method B) was mentioned in Table 2. N-alkylation of 5-chlorobenzoxazolone was confirmed by the presence of carbonyl stretching at 1774 cm−1 in the IR spectrum of 5-chloro-3-(3-chloropropyl)-1,3-benzoxazol-2(3H)-one (8a). The 13C chemical shift value for the carbonyl carbon (δ = 152.6 ppm) of the 5-chorobenzoxazolone was retained in the 13C spectra of 8a (δ = 154.2 ppm). The compounds 9a–f were synthesized by conventional conditions (Method A), i.e., by reacting 8a with various cyclic amines 4 and K2CO3 (Saxena et al., 2007). A facile synthesis of the compounds 9a–f was carried out by reacting 8a with various cyclic amines 4 in the presence of catalytic amount of CsF–Celite (Method B) to obtain products in high yields with excellent purity (Scheme 2). The comparative yields of the two methods A and B are mentioned in Table 2. The synthesized compounds 5a–l, 8a–f, and 9a–f were tested for antibacterial and antifungal activity.

Biology

Antibacterial activity

All the 24 synthesized compounds were evaluated for antibacterial activity and the results were summarized in Table 3. In the 5 series of benzoxazole derivatives 5h, 5j, and 5l showed moderate antibacterial activity against some bacteria, while other compounds were inactive. Among these three compounds, 5h contains N-(2-hydroxyethyl) group while 5j and 5l contains NH and O at the fourth position of cyclic amine.

Table 3 Antibacterial activity of 5a–l, 8a–f, and 9a–f
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Among the 8 and 9 series of benzoxazolone derivatives, 8c with N-propagyl group and 9f with N-(3-morpholinyl)propyl group exhibited moderate activity against both Gram-positive and Gram-negative bacteria.

Antifungal activity

All the 24 synthesized compounds were evaluated for antifungal activity and the results are summarized in Table 4. Among the antifungal activity of the 5 series compounds, 5k with piperidine as cyclic amine showed moderate activity. In the 8 series compounds, 8a with N-3-chloropropyl group and 8c with N-propagyl group showed good antifungal activity, while 8f with N-3-methyl-2-butenyl group showed moderate activity. In the 9 series compounds, only 9f with N-(3-morpholinyl)propyl group showed moderate activity. None of the compound has shown higher antimicrobial activity than drugs used as a standard. These results indicate that larger groups at fourth position of cyclic amine and third position of benzoxazolone have no significant contribution to the antimicrobial activity of these compounds. The benzoxazolone derivatives were comparatively more active than benzoxazole derivatives toward antimicrobial activity. This may be attributed to the more functionality sites in the benzoxazolone skeleton to interact with the biological system in comparison with benzoxazole motif.

Table 4 Antifungal activity of 5a–l, 8a–f, and 9a–f
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Conclusions

Various 2-substituted benzoxazoles and 3-substituted-2(3H)-benzoxazolone derivatives were synthesized. The utility of CsF–Celite as a solid base for alkylation of benzoxazolone was explored. The method offers advantages like easy work-up and excellent yields. The catalyst is non-corrosive, easy to handle, and can be easily prepared. Totally, 24 compounds were synthesized and evaluated for antibacterial and antifungal activity. Two compounds 8a and 8c showed good antifungal activity. The results indicate that larger groups at fourth position of cyclic amine and third position of benzoxazolone have no significant contribution to the antimicrobial activity of these compounds. The benzoxazolone derivatives are comparatively more active than benzoxazole derivatives. Further work to synthesize more active compounds is under progress.

Experimental

Melting points were determined on a Buchi capillary melting point apparatus. The 1H NMR (200 and 300 MHz) and 13C NMR (75 MHz) spectra was recorded on Varian Gemini and Bruker Avance spectrometers using TMS as an internal standard. The mass spectra were recorded on a VG Auto Spec mass spectrometer. Elemental analyses were performed on Elemental VARIO EL elemental analyzer. IR spectra were recorded on Perkin-Elmer Infrared-683.

Typical procedure for preparation of 2-(4-methylpiperazin-1-yl)-1,3-benzoxazole (5a) as a typical procedure

The 2-chlorobenzoxazole (3) (2.6 mmol, 400 mg) was added to a solution of 1-methyl piperazine (2.6 mmol, 260 mg) in dry acetonitrile (30 ml) at 0°C. The mixture was stirred at 0°C–rt for 30 min, quenched in ice water (30 ml), extracted with ethyl acetate (3 × 20 ml), and dried over Na2SO4. After concentration, the residue was purified by flash chromatography to obtain the pure product and was crystallized from methanol.

White solid. m.p. 37–38°C; IR (KBr) ν 3057, 2933, 2854, 2796, 1639, 1578, 1456, 1363. 1H NMR (200 MHz, CDCl3) δ: 7.22 (t, 2H, J = 7.3 Hz); 7.09 (t, 1H, J = 7.3 Hz); 6.95 (t, 1H, J = 7.3 Hz); 3.67 (t, 4H, J = 5.1 Hz); 2.48 (t, 4H, J = 5.1 Hz); 2.30 (s, 3H). EI-MS (m/z): 217 (M+). Anal. calcd for C12H15N3O (217): C, 66.34; H, 6.96; N, 19.34. Found: C, 66.36; H, 6.94; N, 19.37.

2-(4-Ethylpiperazin-1-yl)-1,3-benzoxazole (5b)

White solid. m.p. 78–80°C; IR (KBr) ν 2970, 2931, 2813, 1638, 1578, 1525, 1459, 1399. 1H NMR (200 MHz, CDCl3) δ: 7.30 (d, 1H, J = 7.7 Hz); 7.20 (d, 1H, J = 7.7 Hz); 7.11 (t, 1H, J = 7.7 Hz); 6.96 (t, 1H, J = 7.7 Hz); 3.70 (t, 4H, J = 4.9 Hz); 2.54 (t, 4H, J = 4.9 Hz); 2.45 (q, 2H, J = 7.2 Hz); 1.11 (t, 3H, J = 7.2 Hz). EI-MS (m/z): 231 (M+). Anal. calcd for C13H17N3O (231): C, 67.51; H, 7.41; N, 18.17. Found: C, 67.54; H, 7.39; N, 18.15.

2-(4-Benzylpiperazin-1-yl)-1,3-benzoxazole (5c)

White solid. m.p. 230–231°C; IR (KBr) ν 3028, 2917, 2860, 2821, 1640, 1578, 1494, 1455,1 395. NMR (300 MHz, CDCl3) δ: 7.34–7.25 (m, 6H); 7.19 (d, 1H, J = 7. 8 Hz); 7.11 (t, 1H, J = 7. 8 Hz); 6.96 (t, 1H, J = 7.8 Hz); 3.70 (t, 4H, J = 4.7 Hz); 3.54 (s, 2H); 2.56 (t, 4H, J = 4.7 Hz). 13C NMR (75 MHz, CDCl3) δ: 162.1, 148.7, 143.0, 136.9, 129.2, 128.4, 127.4, 123.9, 120.6, 116.2, 108.7, 62.9, 52.1, 45.4. EI-MS (m/z): 293(M+). Anal. calcd for C18H19N3O (293): C, 73.70; H, 6.53; N, 14.32. Found: C, 73.74; H, 6.52; N, 14.33.

2-(4-Phenylpiperazin-1-yl)-1,3-benzoxazole (5d)

White solid. m.p. 147–148°C; IR (KBr) ν 2980, 2914, 2822, 1629, 1575, 1497, 1455. 1H NMR (300 MHz, CDCl3) δ: 7.33 (d, 1H, J = 7.6 Hz); 7.26 (d, 1H, J = 7.6 Hz); 7.23 (d, 2H, J = 6.8 Hz); 7.14 (t, 1H, J = 6.8 Hz); 6.99 (t, 1H, J = 7.6 Hz); 6.92 (d, 2H, J = 6.8 Hz); 6.88 (t, 1H, J = 7.6 Hz); 3.85 (t, 4H, J = 5.3 Hz); 3.29 (t, 4H, J = 5.3 Hz). 13C NMR (75 MHz, CDCl3) δ: 161.8, 150.8, 148.6, 142.6, 129.2, 124.1, 120.9, 120.8, 116.9, 116.3, 108.8, 49.2, 45.5. EI-MS (m/z): 279 (M+). Anal. calcd for C17H17N3O (279): C, 73.10; H, 6.13; N, 15.04. Found: C, 73.08; H, 6.12; N, 15.05.

2-[4-(Pyridin-2-yl)piperazin-1-yl]-1,3-benzoxazole (5e)

White solid. m.p. 186–188°C; IR (KBr) ν 2993, 2917, 2846, 1635, 1576, 1480, 1459, 1435. 1H NMR (200 MHz, CDCl3) δ: 8.19–8.15 (m, 1H); 7.51–7.44 (m, 1H); 7.33 (d, 1H, J = 7.7 Hz); 7.23 (d, 1H, J = 7.7 Hz); 7.13 (dt, 1H, J = 6.6, 1.1 Hz); 6.99 (dt, 1H, J = 6.6, 1.3 Hz); 6.68–6.61 (m, 2H); 3.88–3.77 (m, 4H); 3.74–3.67 (m, 4H). 13C NMR (75 MHz, CDCl3) δ: 161.9, 158.2, 148.7, 146.8, 142.8, 138.4, 124.0, 120.9, 116.4, 113.8, 108.8, 107.8, 45.2, 44.9. EI-MS (m/z): 280 (M+). Anal. calcd for C16H16N4O (280): C, 68.55; H, 5.75; N, 19.99. Found: C, 68.59; H, 5.78; N, 19.96.

2-[4-(Pyrimidin-2-yl)piperazin-1-yl]-1,3-benzoxa (5f)

White solid. m.p. 183–185°C; IR (KBr) ν 2911, 2860, 1651, 1581, 1542, 1490, 1451, 1393. 1H NMR (200 MHz, CDCl3) δ: 8.30 (d, 2H, J = 5.0 Hz); 7.33 (d, 1H, J = 7.6 Hz); 7.23 (d, 1H, J = 7.6 Hz); 7.13 (t, 1H, J = 7.6 Hz); 6.99 (t, 1H, J = 7.6 Hz); 6.51 (t, 1H, J = 5.0 Hz); 4.04–3.94 (m, 4H); 3.81–3.72 (m, 4H). 13C NMR (75 MHz, CDCl3) δ: 166.6, 161.4, 157.8, 148.6, 144.4, 124.3, 121.1, 116.3, 110.6, 108.9, 45.5, 43.2. EI-MS (m/z): 281 (M+). Anal. calcd for C15H15N5O (281): C, 64.04; H, 5.37; N, 24.89. Found: C, 64.08; H, 5.41; N, 24.94.

2-[4-(3-Chlorophenyl)piperazin-1-yl]-1,3-benzoxazole (5g)

White solid. m.p. 134–136°C; IR (KBr) ν 3015, 2913, 1769, 1739, 1625, 1593, 1478, 1397. 1H NMR (200 MHz, CDCl3) δ: 7.33 (d, 1H, J = 6.8 Hz); 7.23 (d, 1H, J = 7.6 Hz); 7.17 (d, 1H, J = 7.6 Hz); 7.13 (t, 1H, J = 7.6 Hz); 7.00 (t, 1H, J = 7.6 Hz); 6.90 (t, 1H, J = 2.3 Hz); 6.85 (d, 1H, J = 7.6 Hz); 6.79 (d, 1H, J = 7.6 Hz); 3.84 (t, 4H, J = 5.3 Hz); 3.31 (t, 4H, J = 5.3 Hz,). 13C NMR (75 MHz, CDCl3) δ: 161.9, 151.9, 150.9, 142.8, 135.0, 130.1, 124.1, 120.9, 120.3, 116.6, 116.4, 114.7, 108.8, 48.6, 45.3. EI-MS (m/z): 313 (M+). Anal. calcd for C17H16ClN3O (313): C, 65.07; H, 5.14; N, 13.39. Found: C, 65.09; H, 5.14; N, 13.36.

2-[4-(1,3-Benzoxazol-2-yl)piperazine-1-yl]ethanol (5h)

Brown solid. m.p. 35–36°C; IR (KBr) ν 3423, 2936, 2855, 1631, 1578, 1483, 1458. 1H NMR (200 MHz, CDCl3) δ: 7.42 (d, 1H, J = 7.6 Hz); 7.30 (d, 1H, J = 7.6 Hz); 7.19 (t, 1H, J = 7.6 Hz); 7.11 (t, 1H, J = 7.6 Hz); 4.64 (t, 2H, J = 6.0 Hz); 2.82 (t, 2H, J = 6.0 Hz); 2.52 (t, 4H, J = 5.2 Hz); 1.60 (t, 4H, J = 5.2 Hz); 1.49–1.38 (m, 1H). EI-MS (m/z): 247(M+).

2-(3-Methylpiperazin-1-yl)-1,3-benzoxazole (5i)

Brown solid. m.p. 51–52°C; IR (KBr) ν 3321, 3057, 2958, 2921, 2856, 1638, 1578, 1458, 1400. 1H NMR (200 MHz, CDCl3) δ: 7.30 (d, 1H, J = 7.6 Hz); 7.19 (d, 1H, J = 7.6 Hz); 7.11 (t, 1H, J = 7.6 Hz); 6.95 (t, 1H, J = 7.6 Hz); 4.11 (d, 2H, J = 11.3 Hz); 3.07 (t, 2H, J = 9.0 Hz); 2.91 (t, 2H, J = 9.0 Hz); 2.71 (t, 1H, J = 8.3 Hz); 1.80 (s, 1H); 1.11 (d, 3H, J = 8.3 Hz). 13C NMR (75 MHz, CDCl3) δ: 162.2, 148.8, 143.1, 124.5, 120.8, 116.4, 108.9, 53.2, 50.5, 46.2, 45.7, 19.8. EI-MS (m/z): 217 (M+). Anal. calcd for C12H15N3O (217): C, 66.34; H, 6.96; N, 19.34. Found: C, 66.36; H, 6.99; N, 19.29.

2-(1,4-Diazepan-1-yl)-1,3-benzoxazole (5j)

Brown solid. m.p. 195–198°C; IR (KBr) ν 3399, 3050, 2934, 1639, 1578, 1459, 1402. 1H NMR (200 MHz, CDCl3) δ: 7.31 (d, 1H, J = 7.6 Hz); 7.23 (d, 1H, J = 7.6 Hz); 7.13 (t, 1H, J = 7. 6 Hz); 6.98 (t, 1H, J = 7.6 Hz); 3.96 (s, 2H); 3.76 (t, 4H, J = 6.0 Hz); 3.07 (t, 1H, J = 5.2 Hz); 2.91 (t, 1H, J = 5.2 Hz); 2.27(q, 1H, J = 6.0 Hz); 1.97 (q, 1H, J = 6.0 Hz). 13C NMR (75 MHz, CDCl3) δ: 161.6, 148.9, 143.1, 124.0, 120.6, 116.2, 108.7, 49.5, 49.1, 48.4, 47.4, 26.7. EI-MS (m/z): 217 (M+). Anal. calcd for C12H15N3O (217): C, 66.34; H, 6.96; N, 19.34. Found: C, 66.31; H, 6.97; N, 19.32.

2-(Piperidin-1-yl)-1,3-benzoxazole (5k)

Brown solid. m.p. 71–72°C; IR (KBr) ν 3057, 2936, 2854, 1638, 1577, 1525, 1455, 1394. 1H NMR (300 MHz, CDCl3) δ: 7.33 (d, 1H, J = 7.9 Hz); 7.23 (d, 1H, J = 7.9 Hz); 7.14 (t, 1H, J = 7.9 Hz); 6.99 (t, 1H, J = 7.9 Hz); 3.70 (s, 4H); 1.74 (s, 6H). 13C NMR (75 MHz, CDCl3) δ: 161.9, 148.4, 142.2, 124.1, 120.7, 115.8, 108.7, 46.8, 25.2, 23.9. EI-MS (m/z): 202 (M+). Anal. calcd for C12H14N2O (202): C, 71.26; H, 6.98; N, 13.85. Found: C, 71.29; H, 7.00; N, 13.82.

2-(Morpholin-4-yl)-1,3-benzoxazole (5l)

Brown solid. m.p. 205–206°C; IR (KBr) ν 3057, 2966, 2920, 2863, 1637, 1578, 1525, 1454, 1398. 1H NMR (200 MHz, CDCl3) δ: 7.32 (d, 1H, J = 7.9 Hz); 7.21 (d, 1H, J = 7.9 Hz); 7.13 (t, 1H, J = 7.9 Hz); 6.98 (t, 1H, J = 7.9 Hz); 3.77 (t, 4H, J = 4.5 Hz); 3.64 (t, 4H, J = 4.5 Hz). 13C NMR (75 MHz, CDCl3) δ: 161.2, 148.2, 141.1, 124.3, 121.3, 116.0, 108.9, 66.0, 45.8. EI-MS (m/z): 204 (M+). Anal. calcd for C11H12N2O2 (204): C, 64.69; H, 5.92; N, 13.72. Found: C, 64.65; H, 5.94; N, 13.69.

Typical procedure for the preparation of the 5-chloro-3-(3-chloropropyl)-1,3-benzoxazol-2(3H)-one (8a)

Method A (Soyer et al., 2005)

A mixture of 5-chlorobenzoxazolone (6) (3.0 mmol) and K2CO3 (4.5 mmol) in dry acetonitrile (15 ml) was stirred for 10 min under N2 atmosphere. 3-Chloro-1-bromopropane (3.2 mmol) was added through syringe to the above mixture and stirred for 8 h. After the reaction completed (TLC), cold water was added to the reaction mixture and stirred for 30 min. The separated solid was filtered, washed with cold water, and crystallized from ethanol to give the pure product.

Method B

A mixture of 5-chlorobenzoxazolone (6) (3.0 mmol) and CsF–Celite (4.5 mmol) in dry THF (15 ml) was stirred for 10 min under N2 atmosphere. 3-Chloro-1-bromopropane (3.2 mmol) was added through syringe to the above mixture and stirred for 4 h. After the reaction was completed (TLC), the reaction mixture was filtered through Celite and solid was washed with ethylacetate (2 × 10 ml). The combined filtrate was dried (Na2SO4) and evaporated to give the pure product.

White solid. m.p. 82–84°C; IR (KBr) ν 3063, 2933, 1774, 1611, 1485, 1369, 1247, 1061, 829, 721. 1H NMR (200 MHz, CDCl3) δ: 7.17–7.03 (m, 3H); 3.99 (t, 2H, J = 7.0 Hz); 3.62 (t, 2H, J = 5.5 Hz); 2.28 (quintet, 2H, J = 7.0 Hz). 13C NMR (300 MHz, CDCl3) δ: 154.2, 141.1, 132.1, 129.5, 122.4, 109.9, 108.6, 41.6, 39.7, 30.3. EI-MS (m/z): 246 (M+). Anal. calcd for C10H9Cl2NO2 (246): C, 48.81; H, 3.69; N, 5.69. Found: C, 48.84; H, 3.71; N, 5.65.

5-Chloro-3-(prop-2-en-1-yl)-1,3-benzoxazol-2(3H)-one (8b)

White solid. m.p. 64–66°C; IR (KBr) ν 3070, 2932, 1768, 1607, 1487, 1374, 1341, 1245, 1006, 801, 704. 1H NMR (200 MHz, CDCl3) δ: 7.15–7.02 (m, 2H); 6.91 (d, 1H, J = 1.7 Hz); 6.00-5.79 (m, 1H); 5.39–5.27 (m, 2H); 4.42 (d, 2H, J = 6.1 Hz). 13C NMR (300 MHz, CDCl3) δ: 154.1, 141.0, 131.9, 130.0, 129.3, 122.3, 119.2, 110.7, 109.4, 44.7. EI-MS (m/z): 210 (M+). Anal. calcd for C10H8ClNO2 (210): C, 57.30; H, 3.85; N, 6.68. Found: C, 57.33; H, 3.83; N, 6.72.

5-Chloro-3-(prop-2-yn-1-yl)-1,3-benzoxazol-2(3H)-one (8c)

White solid. m.p. 91–93°C; IR (KBr) ν 3087, 2925, 2125, 1779, 1605, 1485, 1377, 1350, 1200, 1023, 813, 747. 1H NMR (200 MHz, CDCl3) δ: 7.18-6.99 (m, 3H); 4.57 (d, 2H, J = 2.6 Hz); 2.48 (t, 1H, J = 2.6 Hz). 13C NMR (300 MHz, CDCl3) δ: 153.4, 140.9, 130.8, 129.5, 122.8, 110.8, 109.7, 74.9, 74.4, 31.9. EI-MS (m/z): 208 (M+). Anal. calcd for C10H6ClNO2 (208): C, 57.85; H, 2.91; N, 6.75. Found: C, 57.84; H, 2.95; N, 6.73.

3-Benzyl-5-chloro-1,3-benzoxazol-2(3H)-one (8d)

White solid. m.p. 172–174°C; IR (KBr) ν 3064, 2937, 1769, 1610, 1484, 1341, 1251, 1012, 811, 700. 1H NMR (300 MHz, CDCl3) δ: 7.40–7.26 (m, 5H); 7.13–7.00 (m, 2H); 6.78 (d, 1H, J = 1.5 Hz); 4.95 (s, 2H). 13C NMR (300 MHz, CDCl3) δ: 154.5, 140.9, 134.1, 131.8, 129.3, 129.1, 128.5, 127.6, 122.4, 110.8, 109.3, 46.2. EI-MS (m/z): 260 (M+). Anal. calcd for C14H10ClNO2 (260): C, 64.75; H, 3.88; N, 5.39. Found: C, 64.77; H, 3.85; N, 5.43.

3-(But-3-en-1-yl)-5-chloro-1,3-benzoxazol-2(3H)-one (8e)

White solid. m.p. 53–54°C; IR (KBr) ν 3059, 2953, 1763, 1612, 1487, 1343, 1246, 1060, 796, 682. 1H NMR (300 MHz, CDCl3) δ: 7.13–7.02 (m, 2H); 6.93 (d, 1H, J = 2.3 Hz); 5.88–5.71 (m, 1H); 5.16–5.06 (m, 2H); 3.85 (t, 2H, J = 7.6 Hz); 2.53 (quartet, 2H, J = 7.6). 13C NMR (300 MHz, CDCl3) δ: 154.4, 141.1, 133.2, 132.1, 129.3, 122.3, 118.6, 110.8, 108.9, 41.8, 32.0. EI-MS (m/z): 224 (M+). Anal. calcd for C11H10ClNO2 (224): C, 59.07; H, 4.51; N, 6.26. Found: C, 59.03; H, 4.52; N, 6.29.

5-Chloro-3-(3-methylbut-2-en-1-yl)-1,3-benzoxazol-2(3H)-one (8f)

White solid. m.p. 74–75°C; IR (KBr) ν 3065, 2925, 1772, 1612, 1484, 1343, 1248, 1007, 834, 748. 1H NMR (200 MHz, CDCl3) δ: 7.13–6.98 (m, 2H); 6.85 (s, 1H); 5.26 (t, 1H, J = 6.8 Hz); 4.37 (d, 2H, J = 6.8 Hz); 1.85 (s, 3H); 1.77 (s, 3H). 13C NMR (300 MHz, CDCl3) δ: 154.2, 141.0, 138.9, 131.9, 129.1, 122.2, 116.5, 110.6, 109.1, 40.3, 25.7, 18.0. EI-MS (m/z): 239 (M + 1). Anal. calcd for C12H12ClNO2 (238): C, 60.64; H, 5.09; N, 5.89. Found: C, 60.62; H, 5.12; N, 5.93.

Typical procedure for the preparation of the 5-chloro-3-[3-(4-benzylpiperazin-1-yl)propyl]-1,3-benzoxazol-2(3H)-one (9a)

Method A (Saxena et al., 2007)

A mixture of N-benzylpiperazine (2 mmol) and K2CO3 (3 mmol) in dry N,N-dimethyl formamide (10 ml) was stirred for 10 min under N2 atmosphere. 5-Chloro-3-(3-chloropropyl)-2,3-dihydro-1,3-benzoxazol-2-one (8a) (2 mmol) in dry N,N-dimethyl formamide (5 ml) was added through syringe to the above mixture and stirred for 5 h at 100°C. After the reaction completed (TLC), cold water was added to the reaction mixture and was stirred for 30 min. The separated solid was filtered, washed with cold water, and crystallized from ethanol to give the pure product.

Method B

CsF–Celite (3 mmol) was added to a solution of N-benzylpiperazine (2 mmol) and 5-chloro-3-(3-chloropropyl)-2,3-dihydro-1,3-benzoxazol-2-one (8a) (2 mmol) in dry THF (15 ml) and stirred under reflux for 4 h. After completion of the reaction (TLC), the mixture was filtered through Celite and the solid was washed with ethylacetate (2 × 10 ml). The combined filtrate was dried (Na2SO4) and evaporated to give the pure product.

Brown solid. m.p. 103–105°C; IR (KBr) ν 2939, 2811, 1782, 1612, 1488, 1370, 1248, 1010, 804, 744. 1H NMR (300 MHz, CDCl3) δ: 7.31–7.16 (m, 5H); 7.13 (d, 1H, J = 2.3 Hz); 7.09–6.98 (m, 2H); 3.86 (t, 2H, J = 6.8 Hz); 3.49 (s, 2H); 2.55–2.29 (m, 10H); 1.91 (quintet, 2H, J = 6.8 Hz). 13C NMR (300 MHz, CDCl3) δ: 154.4, 141.0, 137.9, 132.7, 129.2, 129.1, 128.2, 126.9, 121.9, 110.6, 109.2, 63.0, 54.5, 53.0, 40.3, 24.5. EI-MS (m/z): 386 (M+). Anal. calcd for C21H24ClN3O2 (386): C, 65.36; H, 6.27; N, 10.89. Found: C, 65.34; H, 6.28; N, 10.91.

5-Chloro-3-[3-(4-phenylpiperazin-1-yl)propyl]-1,3-benzoxazol-2(3H)-one (9b)

Brown solid. m.p. 128–129°C; IR (KBr) ν 3056, 3010, 2940, 2827, 1783, 1598, 1489, 1355, 1241, 1058, 834, 751. 1H NMR (200 MHz, CDCl3) δ: 7.30–6.98 (m, 5H); 6.92–6.75 (m, 3H); 3.91 (t, 2H, J = 6.3 Hz); 3.14 (t, 4H, J = 5.5 Hz); 2.52 (t, 4H, J = 5.5 Hz); 2.42 (t, 2H, J = 6.3 Hz); 1.97 (quintet, 2H, J = 6.3 Hz). 13C NMR (300 MHz, CDCl3) δ: 154.3, 151.1, 141.0, 132.6, 129.1, 129.0, 122.0, 119.7, 116.7, 110.7, 109.1, 54.6, 52.0, 49.0, 40.3, 24.4. EI-MS (m/z): 372 (M+). Anal. calcd for C20H22ClN3O2 (372): C, 64.60; H, 5.96; N, 11.33. Found: C, 64.63; H, 5.92; N, 11.34.

5-Chloro-3-{3-[4-(pyridin-2-yl)piperazin-1-yl]propyl}-1,3-benzoxazol-2(3H)-one (9c)

White solid. m.p. 132–133°C; IR (KBr) ν 2944, 2826, 1780, 1594, 1487, 1370, 1250, 1060, 774. 1H NMR (300 MHz, CDCl3) δ: 8.12 (dd, 1H, J = 3.8, 1.5 Hz); 7.41 (dt, 1H, J = 7.6, 1.5 Hz); 7.14 (d, 1H, J = 1.5 Hz); 7.11–7.00 (m, 2H); 6.58 (dt, 2H, J = 7.6, 2.3); 3.91 (t, 2H, J = 6.8 Hz); 3.49 (t, 4H, J = 5.3 Hz); 2.46 (t, 4H, J = 5.3 Hz); 2.40 (t, 2H, J = 6.0 Hz); 1.96 (quintet, 2H, J = 6.0 Hz). 13C NMR (300 MHz, CDCl3) δ: 159.4, 154.5, 148.0, 140.7, 137.4, 132.8, 129.2, 122.0, 113.3, 110.5, 108.9, 107.0, 54.8, 53.0, 44.9, 40.2, 24.6. EI-MS (m/z): 373 (M+). Anal. calcd for C19H21ClN4O2 (373): C, 61.21; H, 5.68; N, 15.03. Found: C, 61.23; H, 5.65; N, 15.01.

5-Chloro-3-{3-[4-(pyrimidin-2-yl)piperazin-1-yl]propyl}-1,3-benzoxazol-2(3H)-one (9d)

White solid. m.p. 125–126°C; IR (KBr) ν 2927, 2853, 1781, 1585, 1488, 1362, 1252, 981, 801, 750. 1H NMR (500 MHz, CDCl3) δ: 8.24 (d, 2H, J = 4.9 Hz); 7.14 (d, 1H, J = 2.0 Hz); 7.11–7.03 (m, 2H); 6.43 (t, 1H, J = 4.9 Hz); 3.93 (t, 2H, J = 6.8 Hz); 3.78 (t, 4H, J = 4.9 Hz); 2.45–2.38 (m, 6H); 1.98 (quintet, 2H, J = 6.8 Hz). 13C NMR (300 MHz, CDCl3) δ: 161.6, 157.4, 154.8, 150.8, 132.9, 129.4, 122.1, 110.7, 109.8, 109.1, 54.8, 53.0, 43.4, 40.3, 24.3. EI-MS (m/z): 374 (M+). Anal. calcd for C18H20ClN5O2 (374): C, 57.83; H, 5.39; N, 18.73. Found: C, 57.80; H, 5.43; N, 18.76.

5-Chloro-3-{3-[4-(3-chlorophenyl)piperazin-1-yl]propyl}-1,3-benzoxazol-2(3H)-one (9e)

Brown solid. m.p. 138–140°C; IR (KBr) ν 2947, 2824, 1781, 1594, 1487, 1372, 1246, 985, 770. 1H NMR (300 MHz, CDCl3) δ: 7.17–7.01 (m, 4H); 6.86–6.68 (m, 3H); 3.91 (t, 2H, J = 6.2 Hz); 3.14 (t, 4H, J = 5.0 Hz); 2.50 (t, 4H, J = 4.9 Hz); 2.42 (t, 2H, J = 6.4 Hz); 1.96 (quintet, 2H, J = 6.2 Hz). 13C NMR (300 MHz, CDCl3) δ: 152.2, 141.1, 135.0, 132.6, 130.0, 129.3, 122.1, 120.0, 119.3, 115.7, 113.8, 110.7, 109.0, 54.6, 52.9, 48.5, 40.3, 24.4. EI-MS (m/z): 406 (M+). Anal. calcd for C20H21Cl2N3O2 (406): C, 59.12; H, 5.21; N, 10.34. Found: C, 59.15; H, 5.22; N, 10.32.

5-Chloro-3-[3-(morpholin-4-yl)propyl]-1,3-benzoxazol-2(3H)-one (9f)

Brown solid. m.p. 97–99°C; IR (KBr) ν 2953, 2855, 1782, 1613, 1488, 1370, 1251, 1117, 805, 750. 1H NMR (300 MHz, CDCl3) δ: 7.13–7.01 (m, 3H); 3.89 (t, 2H, J = 6.4 Hz); 3.63 (t, 4H, J = 4.5 Hz); 2.40–2.29 (m, 6H); 1.93 (quintet, 2H, J = 6.4 Hz). 13C NMR (300 MHz, CDCl3) δ: 154.6, 153.1, 141.2, 129.4, 122.1, 110.8, 109.0, 66.9, 55.0, 53.4, 40.3, 24.0. EI-MS (m/z): 297 (M+). Anal. calcd for C14H17ClN2O3 (297): C, 56.66; H, 5.77; N, 9.44. Found: C, 56.62; H, 5.78; N, 9.47.

Antibacterial activity

Determination of minimum inhibitory concentration

The minimum inhibitory concentrations (MIC) of the new compounds 5a–l, 8a–f, and 9a–f were tested against three representative Gram-positive organisms viz. Bacillus subtilis (MTCC 441), Staphylococcus aureus (MTCC 96), Staphylococcus epidermidis, and Gram-negative organisms viz Escherichia coli (MTCC 443), Pseudomonas aeruginosa (MTCC 741), and klebsiella pneumoniae (MTCC 618) by broth dilution method recommended by National Committee for Clinical Laboratory (NCCL) standards (NCCLS, 2000).

Standard antibacterial agents like Penicillin and Streptomycin were also screened under identical conditions for comparison. The minimum inhibitory concentration (MIC) values are presented in Table 3.

Antifungal activity

In vitro antifungal activity of the 5a–l, 8a–f, and 9a–f was studied against the fungal strains, Candida albicans (MTCC 227), Saccharomyces cereviseae (MTCC 36), Rhizopus oryzae (MTCC 262), Aspergillus niger (MTCC 282) by agar well diffusion method (Linday, 1962). Standard antifungal agent Amphotericin-B was also screened under identical conditions for comparison. The zone of inhibition values of the tested compounds was mentioned in Table 4.

The ready-made potato dextrose agar (PDA) medium (Hi-media, 39 g) was suspended in distilled water (1000 ml) and heated to boiling until it dissolved completely, the medium and Petri dishes were autoclaved at pressure of 15 lb/in.2 for 20 min. Agar well bioassay was employed for testing antifungal activity. The medium was poured into sterile Petri dishes under aseptic conditions in a laminar air flow chamber. When the medium in the plates solidified, 0.5 ml of (week old) culture of test organism was inoculated and uniformly spread over the agar surface with a sterile L-shaped rod. Solutions were prepared by dissolving the compound in DMSO and different concentrations were made. After inoculation, wells were scooped out with 6 mm sterile cork borer and the lids of the dishes were replaced. To each well, different concentrations of test solutions were added. Controls were maintained. The treated and the controls were kept at 27°C for 48 h. Inhibition zones were measured and the diameter was calculated in millimeter. Three to four replicates were maintained for each treatment.