Introduction

The remarkable ability of bacteria to develop resistance to antimicrobial agents is a pressing concern for public health (Boucher et al., 2009). Consequently, new antimicrobial drugs to combat this problem are therefore in great demand. Over the past 50 years, only two new structural types of antimicrobial drugs, linezolid and daptomycin, have been introduced into the clinic (Simmons et al., 2010). The design of new scaffolds to deal with antimicrobial resistance has become one of the most important areas of antimicrobial research today.

Thiazolo[3,2-a]pyrimidines have been consistently regarded as a structural analogs of biogenic purine bases and can be considered as potential purine antagonists (El-Bayouki and Basyouni, 2010; Nagarajaiah et al., 2012). The synthesis of thiazolo[3,2-a]pyrimidines has received considerable attention from both synthetic and medicinal chemists because of their wide range of biological activities, including antiinflammatory (Tozkoparan et al., 1998, 1999), antihypertensive (Jeanneau-Nicolle et al., 1992), antiviral (Mohamed et al., 2010), antioxidant (Maddila et al., 2012), antitumor (Flefel et al., 2007; Al-Omary et al., 2012), anti-HIV (Danel et al., 1998), calcium channel blocking (Balkan et al., 1992), acetylcholine esterase inhibitory (Liu et al., 2014), CDC25B phosphatase inhibitory (Kolb et al., 2009), Bcl-2 family proteins inhibitory (Kolb et al., 2009), glutamate receptor antagonistic (Wichmann et al., 1999) and 5-HT2a receptor antagonistic activities (Awadallah, 2008). These thiazolopyrimidines have also been found to elicit antibacterial (Ghorab et al., 2000), antifungal (Pan et al., 2011) and antitubercular activities (Geist et al., 2010).

Compounds containing sulfonic acid groups have great practical usefulness, primarily, due to a very wide spectrum of biological activities. The phosphate functional group can be replaced by sulfonic acid moieties via bioisosteric replacement. These features are functionally interchangeable due to their ability to adopt a negative charge at biological pH (Cai, 2004). Numerous compounds containing sulfonic acid group are well known as antibacterial (Khan and Rastogi, 1991; Veretennikov and Pavlov, 2013; Janietz et al., 1988; Doria et al., 1985), antifungal (Allen et al., 1959; Dhapalapur et al., 1968; Raval et al., 2012; Fujiwara and Inoi, 1992; Bondock et al., 2013) and antitubercular activities (Parekh and Maheria, 2012). Additionally, these sulfonic acids are also used as dyes (Youssef and Youssef, 2003), metal arenesulfonates complexes (El-Emary and Abdel-Mohsen, 2006).

In the light of the above facts, we have synthesized some 5H-thiazolo[3,2-a]pyrimidin-5-one derivatives and related compounds containing sulfonic acid active moieties with the hope to possess better antibacterial and antitubercular activities.

Results and discussion

Chemistry

The synthetic strategies adopted for the preparation of the intermediates and target compounds are depicted in Scheme 1.

Scheme 1
scheme 1

Reagents and conditions: i K2CO3, DMF; ii concentrated H2SO4, 80 °C; iii concentrated H2SO4, 20 °C; iv Fe, NH4Cl, 2:1 ethanol/water, 80 °C; v triethylamine, r.t., 2 h

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6-Substituted anilino-2-thiouracils 1a–b were nearly quantitatively S-alkylated with the appropriate substituted phenacyl halides in the presence of anhydrous K2CO3 to give compounds 2a–j. These intermediates 2a–j were of sufficient purity (by TLC) to be used in subsequent reactions without further purification. The S-alkylated 2-thiouracils are proved to exist in solution largely in the lactam form as indicated by spectroscopic studies (Brown et al., 1955; Andrew and Bradsher, 1967). The tautomeric hydrogen was found to be in position 3 (Brown et al., 1955; Andrew and Bradsher, 1967; Abdel-Mohsen et al., 2013; Kwiatkowski and Pullman, 1975; Mizutani et al., 1985) (Scheme 2).

Scheme 2
scheme 2

Possible interconversion of S-alkylated derivatives

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Regarding the 400-MHz 1H NMR (in DMSO-d 6) spectra, the S-alkylated thiouracils 2a–j showed similar shifts, and therefore, we discuss here the spectroscopic data only of 2b as the representative of this series (see Supporting Information). On the basis of structure of 2b, the methylene group appeared as a two-proton singlet at δ 4.84 ppm. The signal of the 5-H proton of pyrimidine is observed at δ 6.04 ppm as a singlet. The signal of the N3-H proton appeared at δ 12.79 ppm as a broad singlet. Within the aromatic region, a multiplet appeared at δ 8.30–8.37 ppm due to the substituted phenyl protons. In addition, the compound 2b seems to exist partially in the cyclic form (Danel et al., 1998), which had a pair of doublets centered at δ 3.74 and 3.65 ppm comprising an AB system (J = 12.5 Hz), but an AB quartet did not fully account for two methylene protons. Moreover, two singlets at δ 2.25 and δ 1.99 were all assigned to CH3. All these indicate that the presence of an equilibrium mixture of open-chain and cyclic forms is possible. Furthermore, the intermediate S-alkylated thiouracil exists as enol tautomeric form (Lin et al., 1979; Kaye et al., 1983; Meslin and Quiniou, 1974, 1975), and a one-proton singlet was present at δ 6.02 ppm, corresponding to the enol form of compound 2b in DMSO (Matiichuk et al., 2008) (Scheme 2).

Compounds 3a–d can be synthesized from the S-alkylated thiouracils in concentrated H2SO4 at 80 °C by a one-pot procedure via an intramolecular cyclization/sulfonation sequence. The S-alkylated thiouracils undergo only intramolecular cyclization in concentrated H2SO4 at room temperature to give compounds 4a–h, which can be reduced to give the corresponding amines 5a–d.

Theoretically, the intramolecular cyclization reactions of intermediates 2a–j may afford the two possible isomeric products: 5H-thiazolo[3,2-a]pyrimidin-5-ones and 7H-thiazolo[3,2-a]pyrimidin-7-ones. However, in practice cyclization of compounds 2a–j was found to afford only the corresponding 5H-thiazolo[3,2-a]pyrimidin-5-ones 3a–f and 4a–h. The regioselectivity of the intramolecular cyclization step was assumed to be the difference in the electron density at the N1 and N3 position of compounds 2a–j. The higher basicity of the N3 resulted in the exclusive cyclization at this position (Bakavoli et al., 2008; Dallinger et al., 2003). Moreover, the selective C2–N3 annulation has been proven by single-crystal X-ray crystallographic studies (Quan et al., 2008). Additionally, it seems that the steric hindrance at the position 6 in the S-alkylated derivatives 2a–j exerts a directive influence during this cyclization.

For products 3a–d, all the sulfonic acid groups locate the ortho-positions relative to R3 (electron-donating groups, such as R2=CH3, CH3O). The best explanation of this phenomenon is the influence of electronic and steric factors together determining a new substituent to the positions of the phenyl ring. Similar conclusions for the regioselectivity of sulfonation of the phenyl ring have also been reported in the studies (Schäfer et al., 1984, 1991; Angioni et al., 2014; Katritzky et al., 1984; Coates et al., 1996).

The amines 5a–d were prepared by reducing the nitro compounds 4a–b, 4e–f with iron in a mixture of NH4Cl, ethanol and water following a literature (Dai et al., 2005). However, the post-process described for the reduction was not practical because the target compounds 5a–d were very slightly soluble in water and slightly soluble in cool ethyl acetate and dichloromethane. The completion of the reaction was monitored by TLC. Without allowing the mixture to cool, the iron residue is removed by filtration and washed with boiling ethanol until a clear wash was obtained. The filtrate was concentrated under reduced pressure until no more ethanol distilled, the resultant mixture was cooled to room temperature, and the solution was filtered. The crude product was recrystallized from ethanol to afford the appropriate amines.

The amino substitution of compounds 4a–h could lead to an amino conjugation effect of the cyclic α,β-unsaturated ketone of 5H-thiazolo[3,2-a]pyrimidin-5-ones. Apparently, the negative charge density of N atom decreases, and consequently, the nucleophilicity of amino group will decrease dramatically. A mild procedure for the generation of amides 6a–d is based on the procedure in reference Tang et al., (2013). Compounds 4a–h react with m-methylbenzoyl chloride in the presence of triethylamine to give amides 6a–d. However, the synthesis required relatively high temperatures and/or long reaction times because of the low nucleophilicity of the amine group.

Biological activity

Antibacterial activity

All of the synthesized compounds were evaluated in vitro antibacterial activity against two Gram-positive bacterial strains: Staphylococcus aureus (S. aureus) and Bacillus subtilis (B. subtilis); two Gram-negative bacterial strains: Escherichia coli (E. coli) and Pseudomonas aeruginosa (P. aeruginosa), by conventional broth microdilution assay. The MIC values of antibacterial activity are given in Table 1.

Table 1 In vitro antibacterial activity (MIC µg/mL)
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The investigation of antibacterial screening data reveals that all the tested compounds showed significant activity against Gram-negative bacteria. However, most of the compounds exhibited relatively weak activity against Gram-positive bacteria. The sulfonic acid group in the phenyl ring did not seem to have a remarkable contribution to the antibacterial inhibition by compounds 3a–d; however, strong electron-withdrawing group such as nitro may have improved their effect against Gram-negative bacteria, and compounds 4e and 4f showed good activity against E. coli. Compounds 5a–d containing a primary amino substituent on phenyl group exhibited moderate potencies against P. aeruginosa. Compounds 6a–d that are having substituted benzoylamido at the C-7 position were found to be the most active compounds against the test microorganisms.

Antitubercular activity

Antitubercular activity of compounds was assessed against Mycobacterium smegmatis (M. smegmatis) using conventional broth microdilution assay. MIC values are given in Table 2.

Table 2 In vitro antitubercular activity (MIC µg/mL)
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Table 2 shows the antitubercular activity results which depicted that compounds 6a–d containing substituted benzoylamido at the C-7 position show good activity against M. smegmatis. The presence of substituted benzoylamido moiety with other groups like chloro fused to 5H-thiazolo[3,2-a]pyrimidin-5-ones may be accounted for their antitubercular activity.

Materials and methods

Melting points were determined in open capillary tubes with a WRS-1B melting point apparatus and are uncorrected. IR spectra (KBr) were recorded on a FTIR920 spectrophotometer. The 1H and 13C NMR spectra were obtained from a solution in DMSO-d 6 unless otherwise noted with TMS as internal standard using a 400/101 MHz (1H/13C) spectrometer. Mass spectra were acquired from a Agilent 6200 Series TOF and 6500 Series Q-TOF LC/MS System B.05.01. (B5125).

General procedure for preparation of compounds 2

Anhydrous potassium carbonate (1.3821 g, 10 mmol) and substituted phenacyl halides (10 mmol) were added in succession to a suspension of 6-substituted-2-thiouracils (10 mmol) in dry N,N-dimethylformamide (10 mL). After stirring for 3 h at room temperature, the mixture was quenched with water (100 mL) and filtered. The resulting solid was crystallized from a suitable solvent.

6-Methyl-2-((2-(3-nitrophenyl)-2-oxoethyl)thio)pyrimidin-4(3H)-one 2a

White solid; yield: 86.27 %; m.p. 252.2–253.7 °C. HRMS (m/z): calcd for C13H12N3O4S (neutral M + H) 306.05485; found 306.057780.

6-Methyl-2-((2-(4-nitrophenyl)-2-oxoethyl)thio)pyrimidin-4(3H)-one 2b

White solid; yield: 87.78 %; m.p. 258.1–259.5 °C (lit. (Hurst et al., 1988) 71 %; m.p. > 315 °C). HRMS (m/z): calcd for C13H12N3O4S (neutral M + H) 306.05485; found 306.056718.

6-Methyl-2-((2-oxo-2-(p-tolyl)ethyl)thio)pyrimidin-4(3H)-one 2c

White solid; yield: 84.56 %; m.p. 187.3–187.6 °C (lit. (Johnson et al., 1913) 52 %; m.p. 191–195 °C). HRMS (m/z): calcd for C14H15N2O2S (neutral M + H) 275.0842; found 275.096403.

2-((2-(4-Chlorophenyl)-2-oxoethyl)thio)-6-methylpyrimidin-4(3H)-one 2d

Yellow green solid; yield: 86.85 %; m.p. 218.7–220.0 °C (lit.(Hurst et al., 1988) 80 %; m.p. 210–212 °C). HRMS (m/z): calcd for C13H12ClN2O2S (neutral M + H) 295.03080; found 295.033049.

2-((2-(4-Methoxyphenyl)-2-oxoethyl)thio)-6-methyl pyrimidin-4(3H)-one 2e

Yellow solid; yield: 74.05 %; m.p. 180.6–181.2 °C (lit. (Hurst et al., 1988) 50 %; m.p. 186–189 °C). HRMS (m/z): calcd for C14H15N2O3S (neutral M + H) 291.08034; found 291.088547.

6-Amino-2-((2-(3-nitrophenyl)-2-oxoethyl)thio) pyrimidin-4(3H)-one 2f

Yellow solid; yield: 73.45 %; m.p. 217.8–218.2 °C. HRMS (m/z): calcd for C12H11N4O4S (neutral M + H) 307.05010; found 307.053381.

6-Amino-2-((2-(4-nitrophenyl)-2-oxoethyl)thio) pyrimidin-4(3H)-one 2g

Yellow solid; yield: 73.45 %; m.p. 222.6–223.2 °C. HRMS (m/z): calcd for C12H11N4O4S (neutral M + H) 307.05010; found 307.051107.

6-Amino-2-((2-oxo-2-(p-tolyl)ethyl)thio)pyrimidin-4(3H)-one 2h

White solid; yield: 85.0 %; m.p. 220.0–220.4 °C (lit. (Hurst et al., 1988) 68 %; m.p. 204–206 °C). HRMS (m/z): calcd for C13H14N3O2S (neutral M + H) 276.08067; found 276.090015.

6-Amino-2-((2-(4-chlorophenyl)-2-oxoethyl)thio) pyrimidin-4(3H)-one 2i

Yellow solid; yield: 84.53 %; m.p. 216.0–217.2 °C (lit. (Hurst et al., 1988) 63 %; m.p. 220 °C (dec.)). HRMS (m/z): calcd for C12H11ClN3O2S (neutral M + H) 296.02605; found 296.028652.

6-Amino-2-((2-(4-methoxyphenyl)-2-oxoethyl)thio) pyrimidin-4(3H)-one 2j

Yellow solid; yield: 80.32 %; m.p. 228.6–231.0 °C. HRMS (m/z): calcd for C13H14N3O3S (neutral M + H) 292.07559; found 292.085934.

General procedure for preparation of compounds 3

S-alkylated thiouracils 2 (1 mmol) were carefully dissolved in 3.0 mL of concentrated sulfuric acid, and the reaction mixture was stirred at 80 °C for 24 h. The progress of the reaction was monitored by TLC. After completion, the reaction mixture was cooled and poured into ethyl acetate, and the precipitate that separated was collected by filtration, washed with ethyl acetate and dried. The resulting solid was recrystallized from ethyl acetate.

2-Methoxy-5-(7-methyl-5-oxo-5H-thiazolo[3,2-a]pyrimidin-3-yl)benzenesulfonic acid 3a

Yellow solid; yield: 88.96 %; m.p. 283.2–284.8 °C; IR (ν max/cm−1): 3445 (SO3H), 1726 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.68 (dd, J = 2.5, 1.1 Hz, 1H, Ar–H), 7.36–7.28 (m, 1H, Ar–H), 7.22 (d, J = 1.2 Hz, 1H, C2–H), 6.96 (dd, J = 8.6, 1.1 Hz, 1H, Ar–H), 6.05 (s, 1H, C6–H), 3.45 (qd, J = 7.0, 1.2 Hz, 3H, CH3O), 2.28 (s, 3H, 7-CH3); 13C NMR (101 MHz, DMSO-d 6) δ 162.61 (N=C–S), 156.81 (CON, C-5), 154.41 (C-7), 146.56 (C-3), 137.71, 134.77, 131.23, 129.52, 127.87, 123.17 (Ar–C), 110.87 (C-2), 104.92 (C-6), 55.90 (CH3O), 18.99 (CH3); HRMS (m/z): calcd for C14H13N2O5S2 (neutral M + H) 353.02659; found 353.029811.

2-Methyl-5-(7-methyl-5-oxo-5H-thiazolo[3,2-a]pyrimidin-3-yl)benzenesulfonic acid 3b

Yellow solid; yield: 83.20 %; m.p. 245.1–247.2 °C; IR (ν max/cm−1): 3440 (SO3H), 1721 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.74 (t, J = 1.5 Hz, 1H, Ar–H), 7.29 (d, J = 1.2 Hz, 1H, C2–H), 7.28–7.12 (m, 2H, Ar–H), 6.08 (s, 1H, C6–H), 2.57 (s, 3H, phenyl-CH3), 2.29 (s, 3H, 7-CH3); 13C NMR (101 MHz, DMSO-d 6) δ 163.93 (N=C–S), 162.11 (CON, C-5), 158.98 (C-7), 145.36, 137.67, 136.39, 129.93, 128.89, 127.36 (Ar–C), 111.23 (C-2), 104.92 (C-6), 23.17 (CH3), 20.34 (CH3); HRMS (m/z): calcd for C14H13N2O4S2 (neutral M + H) 337.03167; found 337.023641.

5-(7-Amino-5-oxo-5H-thiazolo[3,2-a]pyrimidin-3-yl)-2-methoxybenzenesulfonic acid 3c

Yellow solid; yield: 86.46 %; m.p. (dec.) 223.7 °C; IR (ν max/cm−1): 3332 (SO3H), 1659 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.68 (q, J = 2.1 Hz, 1H, Ar–H), 7.32 (dq, J = 8.5, 2.1 Hz, 1H, Ar–H), 6.98–6.93 (m,1H, Ar–H), 6.93–6.90 (m, 1H, C2–H), 6.12 (broad, s, 2H, NH2), 5.02–4.95 (m, 1H, C6–H), 3.80 (d, J = 2.6 Hz, 3H, CH3O); 13C NMR (101 MHz, DMSO-d 6) δ 164.74 (N=C–S), 162.75 (CON, C-5), 159.40 (C-7), 149.88 (C-3), 136.70, 131.27, 129.19, 128.47, 127.32, 111.34 (Ar–C), 108.42 (C-2), 80.10 (C-6), 54.43 (CH3); HRMS (m/z): calcd for C13H12N3O5S2 (neutral M + H) 354.02184; found 354.025808.

5-(7-Amino-5-oxo-5H-thiazolo[3,2-a]pyrimidin-3-yl)-2-methylbenzenesulfonic acid 3d

Yellow solid; yield: 87.52 %; m.p. 238.1–239.2 °C; IR (ν max/cm−1): 3342 (SO3H), 1660 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.71 (d, J = 1.9 Hz, 1H, Ar–H), 7.21 (dd, J = 7.7, 2.1 Hz, 1H, Ar–H), 7.13 (s, 1H, C2–H), 6.98 (s, 1H, Ar–H), 5.73 (broad, s, 2H, NH2), 5.01 (s, 1H, C6–H), 2.56 (s, 3H, phenyl-CH3); 13C NMR (101 MHz, DMSO-d 6) δ 164.89 (N=C–S), 162.65 (CON, C-5), 159.39 (C-7), 147.06 (C-3), 137.77, 136.61, 131.74, 129.83, 127.30, 124.26 (Ar–C), 109.96 (C-2), 80.33 (C-6), 20.31 (CH3); HRMS (m/z): calcd for C13H12N3O4S2 (neutral M + H) 338.02692; found 338.027661.

General procedure for preparation of compounds 4

S-alkylated thiouracils 2 (1 mmol) were carefully dissolved in 3.0 mL of concentrated sulfuric acid, and the reaction mixture was stirred at room temperature for 72 h. The progress of the reaction was monitored by TLC. After completion, the reaction mixture was cooled and poured into ice-cold water, and the precipitate that separated was collected by filtration, washed with ethyl acetate and dried. The resulting solid was recrystallized from ethyl acetate.

7-Methyl-3-(3-nitrophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4a

Yellow solid; yield: 91.15 %; m.p. 260.1.1–261.7 °C; IR (ν max/cm−1): 1702 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 8.35–8.25 (m, 2H, Ar–H), 7.91 (d, J = 7.7 Hz, 1H, Ar–H), 7.69 (t, J = 7.9 Hz, 1H, Ar–H), 7.55 (s, 1H, C2–H), 6.10 (s, 1H, C6–H), 2.31 (s, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 163.70 (N=C–S), 163.14 (CON, C-5), 159.42 (C-7), 146.92 (C-3), 136.18, 135.19, 133.84, 128.93, 124.57, 123.54 (Ar–C), 113.37 (C-2), 104.96 (C-6), 23.56 (CH3); HRMS (m/z): calcd for C13H10N3O3S (neutral M + H) 288.04429; found 288.047339.

7-Methyl-3-(4-nitrophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4b

Yellow solid; yield: 92.72 %; m.p. 215.1–216.7 °C (lit. (Hurst et al., 1988) 86 %; m.p. 238–242 °C); IR (ν max/cm−1): 1731(C=O); 1H NMR (400 MHz, DMSO-d 6) δ 8.27–8.19 (m, 2H, Ar–H), 7.75–7.66 (m, 2H, Ar–H), 7.57 (d, J = 1.7 Hz, 1H, C2–H), 6.11 (d, J = 1.5 Hz, 1H, C6–H), 2.30 (d, J = 1.6 Hz, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 163.73 (N=C–S), 163.05 (CON, C-5), 159.24 (C-7), 147.44 (C-3), 138.64, 135.38, 130.90, 122.55 (Ar–C), 113.94 (C-2), 104.96 (C-6), 23.51 (CH3); HRMS (m/z): calcd for C13H10N3O3S (neutral M + H) 288.04429; found 288.048423.

7-Methyl-3-(p-tolyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4c

Yellow solid; yield: 92.22 %; m.p. 210.7–210.9 °C (lit.(Galasko and Israelstam, 1969) 84 %; m.p. 196–198 °C); IR (νmax/cm−1): 1649 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.30 (s, 1H, C2–H), 7.28 (d, J = 1.0 Hz, 2H, Ar–H), 7.18 (d, J = 7.9 Hz, 2H, Ar–H), 6.06 (t, J = 0.9 Hz, 1H, C6–H), 2.36 (s, 3H, CH3), 2.29 (s, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 163.96 (N=C–S), 162.30 (CON, C-5), 159.12 (C-7), 138.23 (C-3), 137.95, 129.54, 129.46, 128.08 (Ar–C), 111.00 (C-2), 104.96 (C-6), 23.28 (CH3), 21.34 (CH3); HRMS (m/z): calcd for C14H13N2OS (neutral M + H) 257.07486; found 257.088294.

3-(4-Chlorophenyl)-7-methyl-5H-thiazolo[3,2-a]pyrimidin-5-one 4d

Yellow solid; yield: 92.91 %; m.p. 210.9–211.5 °C (lit. (Galasko and Israelstam, 1969) 80 %; m.p. 210–212 °C); IR (νmax/cm−1): 1647 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.45 (d, J = 1.1 Hz, 4H, Ar–H), 7.39 (d, J = 1.1 Hz, 1H, C2–H), 6.08 (d, J = 0.9 Hz, 1H, C6–H), 2.29 (d, J = 1.1 Hz, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 163.83 (N=C–S), 162.73 (CON, C-5), 159.23 (C-7), 136.46 (C-3), 133.59, 131.49, 131.21, 127.48 (Ar–C), 112.03 (C-2), 104.96 (C-6), 23.42 (CH3); HRMS (m/z): calcd for C13H10ClN2OS (neutral M + H) 277.02024; found 277.022425.

7-Amino-3-(3-nitrophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4e

Yellow solid; yield: 92.01 %; m.p. 256.1.1–258.0 °C; IR (ν max/cm−1): 3404, 3389 (NH), 1693 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 8.28–8.20 (m, 2H, Ar–H), 7.86 (dt, J = 7.6, 1.3 Hz, 1H, Ar–H), 7.70–7.60 (m, 1H, Ar–H), 7.23 (s, 1H, C2–H), 6.78 (s, 2H, 7-NH2), 4.95 (s, 1H, C6–H); 13C NMR (101 MHz, DMSO-d 6) δ 164.64 (N=C–S), 162.91 (CON, C-5), 159.50 (C-7), 146.85 (C-3), 135.98, 135.47, 134.38, 128.77, 124.30, 123.23 (Ar–C), 109.76 (C-2), 80.05 (C-6); HRMS (m/z): calcd for C12H9N4O3S (neutral M + H) 289.03954; found 289.041969.

7-Amino-3-(4-nitrophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4f

Yellow solid; yield: 91.14 %; m.p. 241.1–242.4 °C; IR (ν max/cm−1): 3445, 3337 (NH), 1683 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 8.23–8.18 (m, 2H, Ar–H), 7.70–7.63 (m, 2H, Ar–H), 7.24 (s, 1H, C2–H), 6.79 (s, 2H, 7-NH2), 4.95 (s, 1H, C6–H); 13C NMR (101 MHz, DMSO-d 6) δ 164.65 (N=C–S), 162.91 (CON, C-5), 159.34 (C-7), 147.21 (C-3), 139.29, 135.69, 130.63, 122.42 (Ar–C), 110.31 (C-2), 79.97 (C-6); HRMS (m/z): calcd for C12H9N4O3S (neutral M + H) 289.03954; found 289.041732.

7-Amino-3-(p-tolyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4g

Yellow solid; yield: 90.71 %; m.p. 201.5–202.3 °C; IR (ν max/cm−1): 3380, 3341 (NH), 1721 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 7.25 (d, J = 7.8 Hz, 2H, Ar–H), 7.15 (d, J = 7.9 Hz, 2H, Ar–H), 6.96 (d, J = 1.1 Hz, 1H, C2–H), 4.95 (s, 1H, C6–H), 2.34 (s, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 162.61 (N=C–S), 159.41 (CON, C-5), 157.37 (C-7), 137.82 (C-3), 129.78, 129.34, 127.93, 126.56 (Ar–C), 107.43 (C-2), 80.22 (C-6), 21.30 (CH3); HRMS (m/z): calcd for C13H12N3OS (neutral M + H) 258.07011; found 258.081756.

7-Amino-3-(4-chlorophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 4h

Yellow solid; yield: 90.66 %; m.p. 175.0–177.4 °C(lit. (Hurst et al., 1988) 75 %; m.p. 186–188 °C); IR (ν max/cm−1): 3374, 3341 (NH), 1717 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 8.19 (s, 1H, Ar–H), 7.66 (s, 2H, Ar–H), 7.31 (s, 1H, Ar–H), 7.00–6.92 (m, 1H, C2–H), 5.14 (s, 1H, C6–H); 13C NMR (101 MHz, DMSO-d 6) δ 164.65 (N=C–S), 159.81 (CON, C-5), 159.36 (C-7), 147.27 (C-3), 139.07, 130.69, 124.67, 122.47 (Ar–C), 112.99 (C-2), 80.21 (C-6); HRMS (m/z): calcd for C12H9ClN3OS (neutral M + H) 278.01549; found 278.017832.

General procedure for preparation of compounds 5

A suspension of nitro compounds 4 (3.53 mmol) in ethanol (60 mL) and water (30 mL) was treated with ammonium chloride (0.1888 g, 3.53 mmol) and iron powder (0.9857 g, 17.65 mmol). After being stirred at 80 °C for 2 h, the mixture was filtered while hot. The filtrate was washed with hot ethanol and concentrated. The resulting concentrate was allowed to in an ice bath, and the precipitate was filtered.

3-(3-Aminophenyl)-7-methyl-5H-thiazolo[3,2-a]pyrimidin-5-one 5a

Yellow solid; yield: 78.84 %; m.p. 231.6–237.3 °C; IR (ν max/cm−1): 3426, 3316(N–H), 1685 (C=O), 1621 (N–H); 1H NMR (400 MHz, DMSO-d 6) δ 7.21 (d, J = 1.7 Hz, 1H, C2–H), 7.01 (td, J = 7.8, 1.6 Hz, 1H, Ar–H), 6.59 (dd, J = 7.5, 1.8 Hz, 1H, Ar–H), 6.60–6.47 (m, 2H, Ar–H), 6.05 (d, J = 1.6 Hz, 1H, C6–H), 5.17 (s, 2H, phenyl-NH2), 2.28 (d, J = 1.6 Hz, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 163.97 (N=C–S), 162.69 (CON, C-5), 158.98 (C-7), 148.02 (C-3), 138.57, 132.89, 128.09, 117.30, 115.01, 114.32 (Ar–C), 110.40 (C-2), 104.91 (C-6), 23.49 (CH3); HRMS (m/z): calcd for C13H12N3OS (neutral M + H) 258.07011; found 258.071755.

3-(4-Aminophenyl)-7-methyl-5H-thiazolo[3,2-a]pyrimidin-5-one 5b

Yellow solid; yield: 80.16 %; m.p. 205.3–206.2 °C; IR (ν max/cm−1): 3419, 3318(NH), 1688 (C=O), 1623 (NH); 1H NMR (400 MHz, DMSO-d 6) δ 7.08–7.06 (m, 1H, C2–H), 7.04 (dd, J = 8.4, 1.8 Hz, 2H, Ar–H), 6.52 (dt, J = 8.3, 1.6 Hz, 2H, Ar–H), 6.02 (d, J = 1.8 Hz, 1H, C6–H), 5.36 (s, 2H, phenyl-NH2), 2.26 (d, J = 1.8 Hz, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 164.06 (N=C–S), 162.47 (CON, C-5), 159.41 (C-7), 149.49 (C-3), 139.19, 130.56, 119.42, 112.52 (Ar–C), 108.56 (C-2), 104.93 (C-6), 23.43 (CH3); HRMS (m/z): calcd for C13H12N3OS (neutral M + H) 258.07011; found 258.070469.

7-Amino-3-(3-aminophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 5c

Yellow solid; yield: 75.23 %; m.p. 198.0–199.1 °C; IR (ν max/cm−1): 3464, 3304 (NH), 1671 (C=O), 1623(NH); 1H NMR (400 MHz, DMSO-d 6) δ 6.97 (td, J = 7.8, 1.3 Hz, 1H, Ar–H), 6.88 (d, J = 1.3 Hz, 1H, C2–H), 6.66 (s, 2H, 7-NH2), 6.58–6.45 (m, 3H, Ar–H), 5.12 (s, 2H, phenyl-NH2), 4.91 (d, J = 1.3 Hz, 1H, C6–H); 13C NMR (101 MHz, DMSO-d 6) δ 164.91 (N=C–S), 162.66 (CON, C-5), 159.21 (C-7), 147.89 (C-3), 138.94, 133.53, 127.94, 117.20, 114.98, 114.07 (Ar–C), 106.81 (C-2), 80.14 (C-6); HRMS (m/z): calcd for C12H11N4OS (neutral M + H) 259.06536; found 259.065181.

7-Amino-3-(4-aminophenyl)-5H-thiazolo[3,2-a]pyrimidin-5-one 5d

Yellow solid; yield: 75.0 %; m.p. 201.3–202.8 °C; IR (ν max/cm−1): 3323, 3162 (NH), 1650 (C=O), 1633 (NH); 1H NMR (400 MHz, DMSO-d 6) δ 7.01 (dd, J = 8.3, 1.5 Hz, 2H, Ar–H), 6.74 (d, J = 1.4 Hz, 1H, C2–H), 6.62 (s, 2H, 7-NH2), 6.49 (dd, J = 8.3, 1.5 Hz, 2H, Ar–H), 5.29 (s, 2H, phenyl-NH2), 4.93–4.88 (m, 1H, C6–H); 13C NMR (101 MHz, DMSO-d 6) δ 165.03 (N=C–S), 162.54 (CON, C-5), 159.64 (C-7), 149.22 (C-3), 139.51, 130.38, 120.18, 112.46 (Ar–C), 105.01 (C-2), 80.29 (C-6); HRMS (m/z): calcd for C12H11N4OS (neutral M + H) 259.06536; found 259.064633.

General procedure for preparation of compounds 6

A mixture of compounds 4 (1.0 mmoL), 3-methylbenzoyl chloride (0. 1855 g, 1.2 equiv., 1.2 mmol) and triethylamine (0.2024 g, 2 equiv., 2.0 mmol) was refluxed at 80 °C for 2 h. After the completion of reaction, the reaction was quenched with water and extracted with dichloromethane. Finally, the combined organic layer was washed with brine and dried over anhydrous Na2SO4. After removal of the solvent in vacuum, the residue was purified by recrystallization.

3-Methyl-N-(3-(3-nitrophenyl)-5-oxo-5H-thiazolo[3,2-a]pyrimidin-7-yl)benzamide 6a

White solid; yield: 89.73 %; m.p. 297.1–298.4 °C. IR (ν max/cm−1): 3428 (NH), 1685 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 10.91 (s, 1H, CONH), 8.35 (t, J = 2.0 Hz, 1H, Ar–H), 8.30 (ddd, J = 8.2, 2.4, 1.1 Hz, 1H, Ar–H), 7.94 (dt, J = 7.7, 1.3 Hz, 1H, Ar–H), 7.87 (s, 1H, Ar–H), 7.82 (d, J = 7.4 Hz, 1H, Ar–H), 7.71 (t, J = 8.0 Hz, 1H, Ar–H), 7.53 (d, J = 0.9 Hz, 1H, C2–H), 7.48–7.35 (m, 2H, Ar–H), 7.06 (d, J = 0.9 Hz, 1H, C6–H), 2.40 (s, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 167.17 (CONH), 163.68 (N=C–S), 160.45 (CON, C-5), 155.42 (C-7), 146.98 (C-3), 138.26, 136.25, 135.48, 133.81, 133.75, 133.47, 129.20, 129.00, 128.80, 125.77, 124.64, 123.63 (Ar–C), 112.65 (C-2), 91.85 (C-6), 21.34 (CH3); HRMS (m/z): calcd for C20H15N4O4S (neutral M + H) 407.08140; found 407.081802.

3-Methyl-N-(3-(4-nitrophenyl)-5-oxo-5H-thiazolo[3,2-a]pyrimidin-7-yl)benzamide 6b

White solid; yield: 84.79 %; m.p. 306.0–306.8 °C. IR (ν max/cm−1): 3368 (NH), 1685 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 10.92 (s, 1H, CONH), 8.31–8.22 (m, 2H, Ar–H), 7.90–7.78 (m, 2H, Ar–H), 7.83–7.71 (m, 2H, Ar–H), 7.55 (s, 1H, C2–H), 7.49–7.37 (m, 2H, Ar–H), 7.07 (s, 1H, C6–H), 2.40 (s, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 167.14 (CONH), 163.68 (N=C–S), 160.26 (CON, C-5), 155.38 (C-7), 147.49 (C-3), 138.56, 138.22, 135.65, 133.77, 133.44, 130.94, 129.16, 128.77, 125.73, 122.57 (Ar–C), 113.11 (C-2), 91.76 (C-6), 21.29 (CH3); HRMS (m/z): calcd for C20H15N4O4S (neutral M + H) 407.08140; found 407.081674.

3-Methyl-N-(5-oxo-3-(p-tolyl)-5H-thiazolo[3,2-a]pyrimidin-7-yl)benzamide 6c

White solid; yield: 85.84 %; m.p. 231.8–232.9 °C. IR (ν max/cm−1): 3431 (NH), 1683 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 10.86 (s, 1H, CONH), 7.86 (s, 1H, Ar–H), 7.81 (dt, J = 7.6, 1.7 Hz, 1H, Ar–H), 7.47–7.37 (m, 2H, Ar–H), 7.33 (d, J = 7.8 Hz, 2H, Ar–H), 7.25 (s, 1H, C2–H), 7.20 (d, J = 7.8 Hz, 2H, Ar–H), 7.03 (d, J = 1.1 Hz, 1H, C6–H), 2.38 (d, J = 9.1 Hz, 6H, two CH3); 13C NMR (101 MHz, DMSO-d 6) δ 167.09 (CONH), 163.87 (N=C–S), 160.21 (CON, C-5), 155.13 (C-7), 138.23 (C-3), 138.20, 138.16, 133.83, 133.38, 129.56, 129.41, 129.15, 128.74, 128.08, 125.71 (Ar–C), 110.11 (C-2), 91.83 (C-6), 21.33 (CH3), 21.29 (CH3); HRMS (m/z): calcd for C21H18N3O2S (neutral M + H) 376.11197; found 376.112952.

N-(3-(4-Chlorophenyl)-5-oxo-5H-thiazolo[3,2-a]pyrimidin-7-yl)-3-methylbenzamide 6d

White solid; yield: 89.10 %; m.p. 228.1–230.6 °C. IR (ν max/cm−1): 3380 (NH), 1688 (C=O); 1H NMR (400 MHz, DMSO-d 6) δ 10.88 (s, 1H, CONH), 7.89–7.75 (m, 2H, Ar–H), 7.51–7.38 (m, 6H, Ar–H), 7.36 (s, 1H, C2–H), 7.04 (s, 1H, C6–H), 2.40 (s, 3H, CH3); 13C NMR (101 MHz, DMSO-d 6) δ 167.12 (CONH), 163.76 (N=C–S), 160.28 (CON, C-5), 155.26 (C-7), 138.22 (C-3), 136.71, 133.80, 133.64, 133.41, 131.53, 131.12, 129.16, 128.76, 127.51, 125.73 (Ar–C), 111.21 (C-2), 91.82 (C-6), 21.31 (CH3); HRMS (m/z): calcd for C20H15ClN3O2S (neutral M + H) 396.05735; found 396.057856.

Biological activity

The standard strains were obtained from National Center for Medical Culture Collection. The antibacterial activity of the synthesized compounds was performed by conventional broth microdilution method against two Gram-positive bacterial strains: Staphylococcus aureus [CMCC (B) 26003] and Bacillus subtilis [CMCC(B) 63501]; two Gram-negative bacterial strains: Escherichia coli [CMCC (B) 44102] and Pseudomonas aeruginosa [CMCC (B) 10104].

A standard inoculum (1.5 × 107 c.f.u/mL 0.5 McFarland standards) was introduced onto the surface of sterile agar plates. The tested compounds and reference drugs were dissolved in MeOH to get a solution of 2 mg/mL concentration. The series diluted compounds in Mueller–Hinton Broth were dispensed into 96-well microtiter plates (200 μL/well), and then, an aliquot of 5 × 105 c.f.u/mL of bacterial culture was added to each well to final concentrations in a range of 1–800 μg/mL. After incubating at 37 °C for 24 h, the lowest concentration required to arrest the growth of bacteria was recorded as the MIC value. MeOH showed no inhibition effect. Ciprofloxacin was used as reference antibacterial agent.

Mycobacterium smegmatis [CGMCC (B) 1.2621] was obtained from China General Microbiological Culture Collection Center. MIC of compounds was determined against Mycobacterium smegmatis strain by using broth microdilution assay. Rifampicin was used as a reference drug.

Conclusion

The present research study reports the successful synthetic method leading to 7-substituted-5H-thiazolo[3,2-a]pyrimidin-5-ones or corresponding sulfonic acid derivatives at different temperature in high yield and purity. The structures are fully supported by spectroscopic data. All the synthesised compounds were tested for their antibacterial and antitubercular activities. These compounds showed significant activity against Gram-negative bacteria, and the sulfonic acid group in the phenyl ring did not seem to have a remarkable contribution to the antibacterial activity. The presence of substituted benzoylamido moiety at the C-7 position of 5H-thiazolo[3,2-a]pyrimidin-5-ones showed enhanced their antibacterial and antitubercular activities.