Mannich Base Derivatives of 1,2-Benzisothiazolin-3-one:Syntheses, Crystal Structures and Bioactivities①

WANG Xiang-Hui SHAO Yan -Dong WU Yuan-Yuan LIN Qiang

(Key Lab of Tropical Pharmaceutical Herb Chemistry of Ministry of Education, College of Chemistry and Chemical Engineering, Hainan Normal University, Haikou 571158, China)

1 INTRODUCTION

The isothiazol-3(2H)-ones are well known as a class of compounds with a wide spectrum of biological and pharmacological activities, such as antibacterial, fungicidal, insecticidal, and other biological activities[1-4]. Mannich base of isothiazolin-3(2H)-one has been synthesized and reported to possess high antibacterial and antifungal activity by W. Malinka[5,6]. To find more novel antimicrobia candidates with excellent biological activity, our group has focused on the synthesis and characterization of different derivatives of 1,2-benzisothiazol-3(2H)-one[7,8]. In this paper,some new Mannich bases were synthesized by reacting 1,2-benzisothiazol-3(2H)-one with amines, and characterized by elemental analysis, IR spectroscopy, MS and1H NMR. The X-ray diffraction structure of compound 3 was determined.The title compounds show good growth inhibitory activity against the gram-positive bacteria.

2 EXPERIMENTAL

2. 1 Materials and methods

The required reagents were of analytical grade and used without further purification. Elemental analysis (C, H, N, S) was performed using a Vario Micro cube elemental analyzer.1H NMR spectra were measured on a BRUKER av-400 NMR nuclear magnetic resonance instrument in DMSO-d6 solution with TMS as the internal standard.Infrared spectra (4000～400 cm-1) were recorded on a Biorad FT40 spectrophotometer with KBr pellets. Low-resolution electrospray ionization mass spectra (ESI-MS) were obtained on a Bruker HCT spectrometer. Melting points were determined using XT4-100A digital melting point apparatus and uncorrected.

2. 2 Synthesis

The synthetic route of the title compounds was conducted as outlined in Schemes 1 and 2. Condensation of BIT with substituted phenylamines and formaldehyde afforded Mannich bases in moderate yields. All target compounds were characterized by elemental analysis, IR,1H NMR and MS.

2. 2. 1 General synthesis for 2-[(substituted phenylamino)methyl]benzo[d]isothiazol-3(2H)-one (2a～2k)

An ethanol solution (20 mL) containing 1,2-benzothiazol-3(2H)-one (1.51 g, 0.01 mol), formalin(37～40%, 1 mL) and required aromatic amine(0.011 mol) was stirred at room temperature for 3～6 h. After completion of the reaction (detected by thin layer chromatography, TLC), the solvent was removed by evaporation under reduced pressure to obtain the crude product. The solid was then recrystallized from ethanol to give corresponding compound as follows.

Scheme 1. Synthetic route of title compounds 2a～2k

2-((phenylamino)methyl)benzo[d]isothiazol-3(2H)-one (2a)

Colorless crystalline flake, yield 61.3%, m.p.181.5～183.6 ℃.1H NMR (400 MHz, DMSO-d6),δ (ppm): 7.40～7.92 (m, Ar-H, 4H); 6.60～7.07(m, Ar-H, 4H); 6.03(t, N-H, 1H, J = 6.92); 5.26～5.28 (d, -CH2-, 2H, J = 6.92). IR (KBr), v(cm-1):3302(N-H), 1650(C=O), 1600, 1502, 1450(Ar-H).EA (%) found (Calcd.): C, 65.72(65.60); H,4.695(4.72); N, 11.31(10.93); S, 12.45(12.51); MS(ESI): 257.3 (M++1), 279.3 (M++23).

2-((o-tolylamino)methyl)benzo[d]isothiazol-3(2H)-one (2b)

White powder, yield 63.8%, m.p. 129.3～130.9 ℃.1H NMR (400 MHz, DMSO-d6), δ(ppm): 7.39～7.91 (m, Ar-H, 4H); 6.54～7.00 (m,Ar-H, 4H); 6.34 (t, N-H, 1H, J = 6.64); 5.31～5.33 (d, -CH2-, 2H, J = 6.64); 2.25 (s, -CH3, 3H);IR: (KBr, cm-1): 3346 (N-H), 1663 (C=O), 2916,2854 (Ar). EA (%) found (Calcd.) for C15H14N2OS:C, 66.32 (66.64); H, 5.14 (5.22); N, 10.63 (10.36);S, 11.99 (11.86); MS (ESI): 271.1 (M++1),293.1(M++23).

2-((p-tolylamino)methyl)benzo[d]isothiazol-

3(2H)-one (2c)

Colorless needle crystal, yield 48.2%, m.p.154.4～156.5 ℃.1H NMR (400 MHz, DMSO-d6),δ (ppm): 7.39～7.91 (m, Ar-H, 4H); 6.65～6.91(m, Ar-H, 4H); 5.23～5.24 (t, N-H, 1H, J = 6.96);5.23～5.24 (s, -CH2-, 2H, J = 6.96); IR (KBr), v(cm-1): 3312 (N-H), 1652 (C=O), 2921, 2861 (Ar).EA (%) found (Calcd.) for C15H14N2OS: C, 66.38(66.64); H, 5.17 (5.22); N, 10.70 (10.36); S, 12.12(11.86); MS (ESI): 271.1 (M++1), 293.1(M++23).

2-(((4-methoxyphenyl)amino)methyl)benzo[d

]isothiazol-3(2H)-one (2d)

Colorless needle crystal, yield 39.3%, m.p.107.8～110.3 ℃.1H NMR (400 MHz, DMSO-d6),δ (ppm): 6.71 (s, Ar-H, 4H); 6.60 (t, N-H, 1H, J =6.96); 3.60 (s, -OCH3, 3H, J = 6.96); IR (KBr), v(cm-1): 3306 (N-H), 2829 (Ar-OCH3), 1653(C=O), 1598, 1514, 1466 (Ar). EA (%) found(Calcd.) for C15H14N2O2S: C, 62.63 (62.92); H,4.89 (4.93); N, 10.00 (9.78); S, 11.55 (11.20); MS(ESI): 271.1 (M++1); 293.1 (M++23).

2-((2-chlorophenylamino)methyl)benzo[d]iso thiazol-3(2H)-one (2e)

Colorless needle crystal, yield 32.9%, m.p.116.9～117.8 ℃.1H NMR (400 MHz, DMSO-d6),δ (ppm): 7.40～7.93 (m, Ar-H, H); 6.63～7.21 (m,Ar-H, 4H); 6.80 (t, N-H, 1H, J = 6.68); 5.35～5.37 (d, -CH2-, 2H, J = 6.68); IR (KBr), v (cm-1):3330 (N-H), 1674 (C=O), 2924 (Ar). EA (%)found (Calcd.) for C14H11ClN2OS: C, 57.63(57.83); H, 3.86 (3.81); N, 9.91 (9.63); S, 10.78(11.03); MS (ESI): 291.0 (M++1), 313.0 (M++23).

2-((3-chlorophenylamino)methyl)benzo[d]iso thiazol-3(2H)-one (2f)

Colorless needle crystal, yield 42.0%, m.p.165.3.9～ 166.6 ℃.1H NMR (400 MHz,DMSO-d6), δ (ppm): 7.41～7.94 (m, Ar-H, 4H);6.63～7.12 (m, Ar-H, 4H); 7.22 (t, N-H, 1H, J =6.68); 5.26～5.28 (d, -CH2-, 2H, J = 6.68); IR(KBr), v (cm-1): 3310 (N-H), 1646 (C = O), 2927,2868 (Ar). EA (%) found (Calcd.) for C14H11ClN2OS: C, 57.62 (57.83); H, 3.87 (3.81);N, 10.07 (9.63); S, 10.62 (11.03); MS (ESI): 291.0(M++1); 313.0 (M++23).

2-((4-chlorophenylamino)methyl)benzo[d]iso thiazol-3(2H)-one (2g)

Colorless needle crystal, yield 38.3%, m.p.171.1～173.7 ℃.1H NMR (400 MHz, DMSO-d6),δ (ppm): 41～7.93 (m, Ar-H, 4H); 6.71～7.41 (m,Ar-H, 4H); 7.32 (t, N-H, 1H, J = 6.68); 5.25～5.27 (d, -CH2-, 2H, J = 6.68); IR (KBr), v (cm-1):3313 (N-H), 1651 (C=O), 2948 (Ar). EA(%)found (Calcd.) for C14H11ClN2OS: C, 57.58(57.83); H, 3.80 (3.81); N, 10.04 (9.63); S, 10.81(11.03); MS (ESI): 291.0(M++1); 313.0(M++23).

2-((3-bromophenylamino)methyl)benzo[d]iso thiazol-3(2H)-one (2h)

Gray solid, yield 54.8%, m.p. 152.6～154.2 ℃.1H NMR (400 MHz, DMSO-d6), δ (ppm): 7.22 (t,N-H, 1H, J = 6.76); 6.67～7.05 (m, Ar-H, 4H);5.26～5.28 (d, -CH2-, 2H, J = 6.76); IR (KBr), v(cm-1): 3313 (N-H), 1646 (C=O), 2924, 2868 (Ar).EA (%) found (Calcd.) for C14H11BrN2OS: C,50.00 (50.16); H, 3.31 (3.31); N, 8.73 (8.36); S,9.65 (9.57); MS (ESI): 334.0(M++1);356.0(M++23).

2-((2,4-dibromophenylamino)methyl)benzo-[d]isothiazol-3(2H)-one (2i)

White powder, yield 57.6%, m.p. 166.6～168.5 ℃.1H NMR (400 MHz, DMSO-d6), δ(ppm): 6.92～7.94 (m, Ar-H, 7H); 6.80 (t, N-H,1H, J = 6.60); 5.34～5.35 (d, -CH2-, 2H, J = 6.60);IR (KBr), v (cm-1): 3329 (N-H), 1675 (C=O),2926 (Ar). EA (%) found (Calcd.) for C14H10Br2N2OS: C, 40.44 (40.60); H, 2.439 (2.43);N, 7.01 (6.76); S, 7.50 (7.74); MS (ESI): 412.9.0(M++1); 434.9(M++23).

2-((3-nitrophenylamino)methyl)benzo[d]isothiazol-3(2H)-one (2j)

Yellow-green floe crystals, yield 67.7%, m.p.209.5～210.6 ℃.1H NMR (400 MHz, DMSO-d6),δ (ppm): 7.21～7.95 (m, Ar-H, N-H, 9H); 5.36～5.38 (d, -CH2-, 2H, J = 6.68); IR (KBr), v (cm-1):3282 (N-H), 1646 (C = O), 2938, 2854 (Ar). EA(%) found (Calcd.) for C14H11N3O3S: C, 55.60(55.80); H, 3.67 (3.68); N, 14.85 (14.95); S, 10.70(10.64); MS (ESI): 302.0 (M++ 1); 324.0(M++23).

2-((4-nitrophenylamino)methyl)benzo[d]isothiazol-3(2H)-one (2k)

Bright yellow solid, yield 45.4%, m.p. 204.7～205.6 ℃.1H NMR (400 MHz, DMSO-d6), δ(ppm): 6.91～8.05 (m, Ar-H, 8H); 8.21 (s, N-H,1H, J = 6.80); 5.37～5.38 (d, -CH2-, 2H, J = 6.80);IR (KBr), v (cm-1): 3301 (N-H), 1639 (C=O), 938(Ar). EA (%) found (Calcd.) for C14H11N3O3S: C,55.62 (55.80); H, 3.68 (3.68); N, 14.88 (14.95);S,10.76 (10.64); MS (ESI): 302.0 (M++1);324.0(M++23).

2. 2. 2 General synthesis for 2-((3-oxobenzo[d]isothiazol-2(3H)-yl)-methyl)benzo[d]isothiazol-3(2H)-one(3)

An acetic acid solution (30 mL) containing 1,2-benzothiazol-3(2H)-one (3.00 g, 0.02 mol) and formalin (37～40%, 1.0 mL) was refluxed with stirring for 5～7 h. After completion of the reaction(detected by thin layer chromatography, TLC), the solution was placed at room temperature to obtain the product (1.66 g).

Scheme 2. Synthetic route of the title compound 3

White solid, yield 52.8%.1H NMR (400 MHz,DMSO-d6), δ (ppm): 7.41～7.87 (m, Ar-H, 8H),5.11 (s, -CH2-, 2H), IR (KBr), v (cm-1): 3046(Ar-H), 2954, 1650 (Ar, C-C). EA (%) found(Calcd.) for C15H10N2O2S2: C, 57.46 (57.31); H,3.33 (3.21); N, 9.02 (8.91); S, 20.72 (20.40). MS(ESI): 315.0 (M++1); 337.0(M++23).

2. 3 X-ray crystal structure determination

Suitable crystals of compound 3 for X-ray diffraction experiments were grown by slow evaporation from CH2Cl2-MeOH solutions. X-ray data were measured with graphite-monochromatized Mo Ka radiation (λ= 0.71073 ?) on an R-AXIS SPPIDER X diffractometer at 153(2) K.A total of 5380 reflections were collected in the range of 3.31≤θ≤29.12o by using an ω-φ scan mode, of which 1752 were unique with Rint=0.0256 and 1493 were observed with I ＞ 2σ(I).The structure was solved by direct methods(SHELXS-97)[9]. The structure was refined by full-matrix least-squares on F2using SHELXL-97[10]; all of the non-hydrogen atoms were refined with anisotropic displacement parameters. All hydrogen atoms were fixed in the ideal geometrical positions and refined isotropically based on the corresponding C-atoms (U(H) = 1.2Ueq(C) or 1.5Ueq(Cmethyl)). The final R = 0.034 and wR =0.087 (w = 1/[σ2(Fo2) + (0.0516P)2+ 0.5600P],where P = (Fo2+ 2Fc2)/3). S = 0.999, (Δ/σ)max=0.001, (Δρ)max= 0.389 and (Δρ)min= –0.185 e/?3.The selected bond lengths and bond angles are summarized in Table 1. Hydrogen bond lengths and bond angles for compound 3 are given in Table 2.

Table 1. Selected Bond Lengths (?) and Bond Angles (°) of Compound 3

Table 2. Hydrogen Bond Lengths (nm) and Bond Angles (°) for Compound 3

3 RESULTS AND DISCUSSION

3. 1 Crystal structure

Compound 3 crystallizes in space group C2/c.The structure of 3 with atom-numbering scheme is shown in Fig. 1, and the unit cell packing diagram in Fig. 2.

Fig. 1. View of compound 3 with atom-labeling scheme and 50% probability displacement ellipsoids

Fig. 2. Crystal structure of 3, showing the packing mode of the dimers. Hydrogen atoms are omitted for clarity

In molecule 3, the benzisohiazolone ring(C(1)～C(7)–N(1)–S(1) is almost coplanar with a maximum deviation from the mean plane of 0.00217 nm for atom N(1). And the dihedral angle between the two benzisothiazolone rings is 83.093(18)°. The C–C bond lengths of benzisothiazolone groups (C(1)～C(7)/S(1)/N(1), O(1))range from 1.385(2) to 1.461(2) ? and these bond distances are in good agreement with those in the reported structure[8]. In the crystal structure,intermolecular C(2)–H(2)···O(1) and C(8)–H(8A)···O(1) hydrogen bonds (Table 2) link the molecules into a one-dimensional structure (Fig. 2).

The structural parameters of compound 3 are in accordance with the calculated results. The N(1)–C(7) bond lengths are between the C–N single bond (1.471 ?) and double bond (1.273 ?),indicating the conjugation system of the fivemembered ring.

3. 2 In vivo biological activities

Qualitative determination of antimicrobial activity was done using the Turbidimetric Method[16,17]. The values of 50% and 90%minimum inhibitory concentration of the title compounds are listed in Table 3. The organisms used in the present investigation included three gram-positive bacteria (Bacillus subtilis (ISM 6513), Staphylococcus aureus (ATCC 6538),Staphylococcus albus (ATCC 12228)) and three gram-negative bacteria (Escherichia coli (ISM 6585), Klebsiella pneumoniae (ATCC4352) and Pseudomonas aeruginosa (ATCC 15442)). The results show that the title compounds have better antimicrobial activities against gram-positive bacteria than gram-negative bacteria, and the compound with CH3on the benzene ring at the para- and ortho-substituted positions such as compounds 2b and 2c has a good biological activity.

Table 3. MIC90 and MIC50 of the Title Compounds against the Tested Strains (mg/L)

In summary, a series of new isothiazole derivatives was synthesized and evaluated for their biological activities. The results show that several compounds exhibited promising activity and provided the foundation of their further structure optimization.

(1) Collier, P.; Ramsey, A.; Austin, P.; Gilbert, P. Growth inhibitory and biocidal activity of some isothiazolone biocides. J. Appl. Microbiology 1990, 69, 569–577.

(2) Lewis, S. N.; Miller, G. A.; Law, A. B. 3-Isothiazolones as Biocides. U.S. 4105431 1978.

(3) Vicini, P.; Geronikaki, A.; Incerti, M.; Busonera, B.; Poni, G.; Cabras, C. A.; La Colla, P. Synthesis and biological evaluation of benzo [d]isothiazole, benzothiazole and thiazole Schiff bases. Bioorg. Med. Chem. 2003, 11, 4785–4789.

(4) Vicini, P.; Incerti, M.; La Colla, P.; Loddo, R. Anti-HIV evaluation of benzo [d] isothiazole hydrazones. Eur. J. Med. Chem. 2009, 44,1801–1807.

(5) Malinka, W.; Karczmarzyk, Z.; Sieklucka-dziuba, M.; Sadowski, M.; Kleinrok, Z. Synthesis and in vivo pharmacology of new derivatives of isothiazolo [5,4-b] pyridine of Mannich base type. Ⅱ Farmaco. 2001, 56, 905–918.

(6) Malinka, W.; ?wiatek, P.; Filipek, B.; Sapa, J. Synthesis, analgesic activity and computational study of new isothiazolopyridines of Mannich base type. Ⅱ Farmaco . 2005, 60(11–12), 961–968.

(7) Wang, X. H.; Lin, Q.; Yin, X. Q.; You, C. H.; Yang, J. X. Synthesis, bioactivity and crystal structure analysis of novel benzo [d]isothiazol-3(2H)-ones. Chin. J. Struct. Chem. 2012, 31, 1170–1174.

(8) Wang, X. H.; Yang, J. X.; Lin, Q. Synthesis, bioactivity and crystal structure analysis of N-(2-ethoxyphenyl)-3-oxobenzo[d]isothiazole-2(3H)-carboxamide. Chin. J. Struct. Chem. 2012, 31, 758–762.

(9) Sheldrick, G. M. SHELXS 97, Program for the Solution of Crystal Structures. University of G?ttingen, Germany 1997.

(10) Sheldrick, G. M. SHELXL-97, Program for the Refinement of Crystal Structures. University of G?ttingen, Germany 1997.