Synthesis and evaluation of chromene-based compounds containing pyrazole moiety as antimicrobial agents

Mohamed Salah K. Youssef; Ahmed Abdou O. Abeed; Talaat I. El-Emary

doi:10.1515/hc-2016-0136

Article Open Access

Synthesis and evaluation of chromene-based compounds containing pyrazole moiety as antimicrobial agents

Mohamed Salah K. Youssef , Ahmed Abdou O. Abeed and Talaat I. El-Emary

Published/Copyright: January 31, 2017

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From the journal Heterocyclic Communications Volume 23 Issue 1

Abstract

With an intention to synergize the antimicrobial activity of 1,3-diphenyl pyrazole and chromene derivatives, 20 hybrid compounds were synthesized and evaluated for their antimicrobial activity. Structures of the newly synthesized compounds were established by elemental analysis and spectral data. All compounds were evaluated for their antimicrobial activity against Gram positive and Gram negative bacteria and antifungal activity by a well diffusion method. Compounds 8, 10, 16 and 21 show reasonable antibacterial and antifungal activity.

Keywords: antimicrobial; chromene; MIC; pyrazole; triazine

Introduction

Chromene-based molecules possess diverse biological activities such as anti-inflammatory [1], [2], [3], anticonvulsant [4], antitubercular [5], antimicrobial [6], [7], [8], [9], [10] and anti-rhinoviral [11] properties. Flavonoids are natural compounds based on a chromene system. Many of these compounds are antioxidant [12], [13], antifungal [14], antiviral [15], [16], anticancer [17], [18], [19], and anti-inflammatory agents [20], [21], [22]. Moreover, derivatives of 4H-thieno[3,2-с]chromene are used for the treatment of diabetes, hyperlipidaemia [23] and cancer [24], [25]. On the other hand, a 1,3-diphenylpyrazole system occupies a unique position in the design and synthesis of novel biologically active agents with remarkable anticancer [26], [27], [28], antiparasitic [29], analgesic, anti-inﬂammatory properties [30], [31] and antimicrobial activities [32], [33], [34]. Pyrazolylchromene A and chromeno-1,8-naphthyridine derivatives B–D are antimicrobial agents [33] (Figure 1). In view of this information and in continuation of our previous work aimed at the synthesis of novel heterocyclic ring systems of biological importance [35], [36], [37], [38], we report here an efficient method for the synthesis of chromeno[2,3-b]pyridines, chromeno[2,3-d]pyrimidines, chromeno[2,3-d]triazines and chromeno [2,3-d]pyrazol-5-one derivatives using 2-amino-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile as the starting material [33] (Figure 1).

Figure 1

The chromene derivatives A–D and the designed compounds.

Results and discussion

Chemistry

Synthesis of the target compounds is depicted in Schemes 1–4. The key starting compound 1 was prepared by one-pot three-component cyclocondensation reaction of 1,3-diphenylpyrazole-4-carboxaldehyde, malononitrile and 5,5-dimethyl-1,3-cyclohexanedione (dimedone) according to a literature procedure [33]. Treatment of 1 with benzoyl chloride in pyridine afforded the expected N-benzamido derivative 2 (Scheme 1). The alkylation reaction of compound 1 with excess triethyl orthoformate in acetic anhydride led to the formation of ethoxymethyleneamino derivative 3. Compound 1 was also allowed to react with α-cyanocinnamonitrile to afford chromeno[2,3-b]pyridine analogue 4. On the other hand, reaction of 1 with α-ethoxycarbonylcinnamonitrile led to the acyclic chromene 5 which could not be cyclized under similar conditions. Alternatively, compound 4 was prepared by refluxing of 1 with cyanoacetophenone in ethanol with a few drops of piperidine. Treatment of compound 1 with malononitrile in ethanol under reflux in the presence of piperidine led to the formation of an intermediate product which cyclized via the loss of H₂O to give the diamino derivative 6. Elemental analysis and spectral data of these products were fully consistent with the given structures.

Scheme 1

Reagents and conditions: (a) benzoyl chloride, pyridine, reflux 7 h; (b) triethyl orthoformate, acetic anhydride, reflux 5 h; (c) α-cyanocinnamonitrile, piperidine, ethanol, reflux 2 h; (d) cyanoacetophenone, piperidine, ethanol, reflux 8 h; (e) α-ethoxycarbonylcinnamonitrile, piperidine, ethanol, reflux 2 h; (f) malononitrile, piperidine, ethanol, reflux 8 h.

Scheme 2

Reagents and conditions: (a) acetic anhydride, reflux 5 h; (b) NaNO₂/H₂SO₄, stirring 20 min; (c) thiourea, pyridine, reflux 12 h; (d) ethylene diamine, CS₂, water bath overnight; (e) formamide, formic acid, reflux 10 h; (f) phenyl isothiocyanate, pyridine, reflux 5 h.

Scheme 3

Reagents and conditions: (a) H₂SO₄ (70%), reflux 8 h; (b) benzaldehyde, piperidine, ethanol, reflux 10 h; (c) thiourea, pyridine, reflux 7 h; (d) CS₂, KOH, ethanol, reflux 12 h; (e) NaNO₂/H₂SO₄, stirring 20 min; (f) formic acid, reflux 12 h; (g) triethyl orthoformate, acetic acid, reflux 7 h; (h) phenyl isothiocyanate, pyridine, reflux 10 h.

Scheme 4

Reagents and conditions: (a) ethyl cyanoacetate, NaOEt, ethanol, reflux 7 h; (b) hydroxylamine, pyridine, reflux 4 h; (c) hydrazine hydrate, NaOEt, ethanol, reflux 4 h.

On the other hand, refluxing of 1 in acetic anhydride afforded a product that was identified as 7 in excellent yield (Scheme 2). On the other hand, reacting 1 with nitrous acid afforded a yellow product that was identified as chromeno[2,3-d]triazin-6(7H)-one derivative 8 in excellent yield (Scheme 2). The IR spectrum of 8 shows the disappearance of the amino group and the presence of an intense absorption band at 2180 cm⁻¹ indicating the triazine ring. On the other hand, when compound 1 was allowed to reacted with thiourea in refluxing pyridine, this reaction gave an acyclic chromene derivative 9 but not the expected thioxochromeno[2,3-d]pyrimidine derivative (Scheme 2). This result is consistent with the analysis of the IR spectrum of 9, which reveals the presence of a nitrile group at ν 2185 cm⁻¹. The treatment of 1 with ethylenediamine in the presence of a catalytic amount of carbon disulfide produced the substituted chromen-5-one 10. In addition, refluxing of compound 1 with formamide in acetic acid afforded N-formylaminochromeno[2,3-d]pyrimidine 11 (Scheme 2).

The treatment of 1 with phenyl isothiocyanate afforded the phenyl thiouredo derivative 12. All spectral data are in agreement with the assigned structure. The possibility of formation of an alternative product 3-phenyl-4-imino-5-(1,3-diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-2-thioxo-2,3,8,9-tetra-hydro-1H-chromeno[2,3-d]pyri-midine-6(7H)-one (Scheme 2) can be excluded on the basis of the spectral data.

Upon treatment with concentrated H₂SO₄, o-aminocarbonitriles are converted to aminocarboxamides [39]. Accordingly, o-aminocarboxamide 13 was obtained from o-aminocarbonitrile 1 (Scheme 3). The structure of compound 13 was established by spectroscopic and analytical characterization. The IR spectrum shows the absence of a nitrile group and the presence of a new carbonyl group at ν 1668 cm⁻¹ and NH amide function at v 3218–3120 cm⁻¹ due to conversion of nitrile to amide. In addition, the ¹H NMR spectrum shows singlet at δ 9.69 for the amide-NH₂ moiety. The signal for the free NH₂ group appears at δ 8.25. The ¹³C NMR spectrum of 13 shows the absence of a nitrile signal and the presence of a signal for amide carbonyl at δ 172.8.

The o-aminocarboxamide 13 was exploited in the synthesis of fused pyrimidine derivatives (Scheme 3). Thus, cyclocondensation of 13 with benzaldehyde in refluxing ethanol in the presence of a few drops of piperidine afforded chromeno[2,3-d]pyrimidine-4,6-dione 14. Cyclocondensation of 13 with thiourea in pyridine or carbon disulfide in alkaline ethanol gave identical product defined as 2-thioxochromeno[2,3-d]pyrimidine-4,6-dione (15). Furthermore, chromeno[2,3-d] triazine 16 was prepared by diazotization of 13 with sodium nitrite and sulfuric acid. On the other hand, 5-(1,3-diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-8,9-dihydro-3H-chromeno [2,3-d]pyrimidine-4,6(5H,7H)-dione (17) was obtained via the reaction of o-aminocarboxamide 13 with triethyl orthoformate or formic acid. The treatment of 13 with phenyl isothiocyanate in pyridine afforded chromeno[2,3-d]pyrimidine 18 (Scheme 3). The synthetic potential of o-aminocarboxamide 13 was further explored by the successful preparation of aminochromeno[2,3-b]pyridine 19via reaction with ethyl acetoacetate (Scheme 4).

The reaction of 13 with hydroxylamine afforded carboximidamide 20. Moreover, hydrazinolysis of o-aminocarboxamide 13 in the presence of sodium ethoxide gave aminopyrazole 21 (Scheme 4). The structures of products 14–21 were assigned on the basis of elemental analyses and spectral data (see Experimental section).

In vitro antibacterial and antifungal activity

Compounds 1-21 were screened for in vitro antimicrobial activity. Antibacterial activity was assayed against Gram positive and Gram negative bacteria. These compounds were also evaluated for their in vitro antifungal potential against six fungal strains. The agar-diffusion method was used for the determination of the preliminary antibacterial and antifungal activity. Chloramphenicol and clotrimazole were used as reference drugs. The minimum inhibitory concentration (MIC) were then determined for compounds that showed signiﬁcant growth inhibition zones. The MIC values for the selected most active compounds are given in Tables 1 and 2. Remarkable activities can be noted for compounds 8, 10, 16 and 21.

Table 1

Antibacterial activity (MIC, μg/mL) of selected compounds.

Compound no.	S. aureus (+ve)	M. luteus (+ve)	B. cereus (+ve)	E. coli (−ve)	P. aeruginosa (−ve)	S. marcescens (−ve)
5	–	15	15	10	10	30
6	16	–	–	25	20	10
7	–	10	35	40	35	15
8	10	10	15	12	13	5
9	15	5	–	–	–	35
10	10	10	15	16	15	8
11	–	35	–	40	0	15
12	15	20	–	60	10	20
13	28	–	35	30	10	28
16	15	15	14	15	18	10
17	10	40	–	30	10	10
21	10	10	15	12	14	7
Ref^a	10	10	10	10	10	4.5

Compounds of low activity are not shown.

^aChloramphenicol was used as antibacterial standard. (–) Indicates no activity.

Table 2

Antifungal activity of selected compounds (MIC, μg/mL).

Compound no.	A. flavus	C. albicans	F. oxysporum	G. candidum	S. brevicaulis	T. rubrum
1	25	–	18	30	29	33
3	45	27	20	65	25	40
4	–	40	25	64	35	35
5	46	38	15	40	30	–
6	39	55	–	55	20	25
8	10	12	10	22	20	18
10	10	15	10	20	38	12
12	25	30	40	40	29	–
13	18	35	30	30	25	38
15	35	–	15	40	–	25
16	25	25	15	20	29	10
17	20	25	–	40	30	20
18	35	–	15	35	40	20
19	30	–	20	25	58	30
20	20	25	30	–	80	28
21	10	12	10	25	25	15
Ref^a	10	10	10	20	20	10

Compounds of low activity are not shown.

^aClotrimazole was used as antifungal standard. (–) Indicates no activity.

Conclusions

The objective of the present study was to evaluate antimicrobial activities of some new chromenes incorporating a 1,3-diphenylpyrazole moiety with the hope of discovering new structural leads serving as potent antimicrobial agents. Our aim was verified by the synthesis of five different groups of structure hybrids comprising basically the pyrazole moiety attached to either chromene, chromeno[2,3-b]pyridine, chromeno[2,3-d]pyrimidine, chromeno[2,3-d]triazine or chromenopyrazolone counter parts through various linkages of synergistic purpose. The results revealed that compounds containing triazine, imidazole and pyrazole rings exhibit high antimicrobial activity.

Experimental

Melting points were determined using an APP Digital ST 15 melting point apparatus and are uncorrected. FT-IR spectra were recorded on a Pye-Unicam SP3-100 spectrophotometer in KBr pellets. All ¹H NMR (400 MHz) and ¹³C NMR (100 MHz) spectra were recorded on a Brucker AVANCE-III spectrometer. C, H, N and S analyzes were performed with a Vario EL C, H, N, S analyzer.

2-Amino-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile (1) [33]

1,3-Diphenyl-1H-pyrazole-4-carboxaldehyde (0.72 g, 3 mmol), malononitrile (0.19 g, 3 mmol), 5,5-dimethyl-1,3-cyclohexanedione (0.42 g, 3 mmol), ethanol (15 mL) and a few drops of piperidine were placed in a 100-mL round bottom flask equipped with a mechanical stirrer and condenser. The mixture was gently heated under reflux for 3 h, then the resultant solid was filtered, washed with water, dried and crystallized from dioxane-water (2:1): yellow crystals; mp 209–211°C; yield 73%; IR: 3443 and 3260 (NH₂), 3157 (CH aromatic), 2957 (CH aliphatic), 2185 (C≡N), 1677 (C=O cyclic), 1636 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.89 (s, 3H, CH₃), 1.13 (s, 3H, CH₃), 2.10–2.30 (4H, m, 2×CH₂), 4.33 (s, 1H, pyran H4), 7.28–8.15 (m, 13H, Ar-H, pyrazole-H and NH₂); ¹³C NMR (DMSO-d₆): δ 25.3 (CH₃), 27.8 (CH₃), 28.4 (pyran-C4), 32.2 (C(CH₃)₂), 42.0 (CH₂), 51.1 (CH₂-CO), 58.2 (pyran-C3), 112.5 (pyran-C5), 121.9 (C≡N), 114.2, 115.4, 117.3, 118.1, 125.4, 125.7, 126.5, 127.5, 128.7, 131.4, 136.2, 138.5, 145.3, 147.5, 151.2 (Ar-C and pyrazole-C), 157.2 (pyran-C6), 163.5 (pyran-C2), 196.1 (C=O cyclic). Anal. Calcd for C₂₇H₂₄N₄O₂ (436.51): C, 74.29; H, 5.54; N, 12.84. Found: C, 74.20; H, 5.44; N, 12.76.

2-Benzamido-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile (2)

A mixture of 1 (1.30 g, 3 mmol) and benzoyl chloride (0.35 mL, 3 mmol) in dry pyridine (10 mL) was heated under reflux for 7 h. After cooling, the mixture was poured onto water and the precipitate was filtered, washed with water, dried and crystallized from ethanol-water (1:1): fine powder; mp 172–174°C; yield 66%; IR: 3416 (NH), 3150 (CH aromatic), 2967 (CH aliphatic), 2227 (C≡N), 1752 (C=O benzoyl), 1675 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.92 (s, 3H, CH₃), 1.33 (s, 3H, CH₃), 2.10 and 2.34 (m, 4H, 2×CH₂), 4.34 (s, 1H, pyran H4), 7.35–8.25 (m, 16H, Ar-H and pyrazole-H), 9.50 (s, 1H, NH, D₂O exchanged); ¹³C NMR (DMSO-d₆): δ 25.3 (CH₃), 27.8 (CH₃), 28.4 (pyran-C4), 32.3 (C(CH₃)₂), 42.1 (CH₂), 51.2 (CH₂-CO), 58.3 (pyran-C3), 112.4 (pyran-C5), 121.8 (C≡N), 114.3, 115.5, 117.2, 118.2, 125.4, 125.7, 126.4, 127.3, 128.5, 131.4, 136.3, 138.4, 145.2, 147.5, 148.1, 149.5, 150.7, 151.6 (Ar-C and pyrazole-C), 157.2 (pyran-C6), 163.5 (pyran-C2), 189.1 (C=O), 196.3 (C=O cyclic). Anal. Calcd for C₃₄H₂₈N₄O₃: (540.61) C, 75.54; H, 5.22; N, 10.36. Found: C, 75.46; H, 5.13; N, 10.28.

2-(Ethoxymethyleneamino)-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile (3)

A solution of 1 (1.30 g, 3 mmol), triethyl orthoformate (10 mL) and a few drops of acetic anhydride was heated under reflux for 5 h, then concentrated on a water-bath. The residue was treated with ethanol and a few drops of acetic acid. The precipitated solid product 3 was filtered off, washed with water, dried and crystallized from ethanol-water (1:1): white crystals; mp 198–200°C; yield 62%; IR: 3157 (CH aromatic), 2957 (CH aliphatic), 2185 (C≡N), 1666 (C=O cyclic), 1650 (N=CH), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.89 (s, 3H, CH₃), 1.10–1.29 (m, 6H, CH₃ and CH₃ ethoxy), 2.10–2.50 (m, 4H, 2×CH₂), 4.30–4.70 (m, 3H, pyran-H4 and CH₂ ethoxy), 6.80 (s, 1H, N=CH), 7.20–8.25 (m, 11H, Ar-H and pyrazole-H); ¹³C NMR (DMSO-d₆): δ 14.20 (CH₃ ester), 25.2 (CH₃), 27.6 (CH₃), 28.3 (pyran-C4), 32.3 (C(CH₃)₂), 42.5 (CH₂), 51.4 (CH₂-CO), 58.5 (pyran-C3), 61,6 (CH₂ ester), 75.1 (N=CH), 112.5 (pyran-C5), 121.8 (C≡N), 114.3, 115.5, 117.3, 118.1, 125.4, 125.7, 126.5, 127.5, 128.7, 131.4, 136.2, 138.5, 145.3, 147.5, 151.8 (Ar-C and pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 196.2 (C=O cyclic). Anal. Calcd for C₃₀H₂₈N₄O₃ (492.57): C, 73.15; H, 5.73; N, 11.37. Found: C, 73.08; H, 5.64; N, 11.28.

4-Amino-5-(1,3-diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-6-oxo-2-phenyl-6,7,8,9-tetrahydro-5H-chromeno[2,3-b]pyridine-3-carbonitrile (4)

Method A To a solution of 1 (1.30 g, 3 mmol) and α-cyanocinnamonitrile (0.45 g, 3 mmol) in ethanol (20 mL), a few drops of piperidine was added. The mixture was heated under reflux for 2 h. On cooling, the resultant solid product was filtered, washed with water, dried and crystallized from dioxane-water (1:3): yellow needles; mp 190–192°C; yield 69%; IR: 3442, 3250 (NH₂), 3052 (CH aromatic), 2957 (CH aliphatic), 2185 (C≡N), 1666 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 1.00 (s, 3H, CH₃), 1.20 (s, 3H, CH₃), 2.10–2.30 (m, 4H, 2×CH₂), 4.20 (s, 1H, pyran-H), 7.20–8.13 (m, 18H, Ar-H, pyrazole-H and NH₂); ¹³C NMR (CDCl₃): δ 25.2 (CH₃), 27.4 (CH₃), 28.5 (pyran-C4), 32.4 (C(CH₃)₂), 42.1 (CH₂), 51.2 (CH₂-CO), 58.3 (pyran-C3), 112.4 (pyran-C5), 121.8 (C≡N), 114.1, 115.43, 116.8, 117.3, 118.1, 120.2, 123.4, 125.4, 125.7, 126.5, 127.5, 128.7, 138.5, 141.5, 145.3, 147.5, 148.7, 151.2 (Ar-C and pyridine C, pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 196.4 (C=O cyclic). Anal. Calcd for C₃₆H₂₉N₅O₂ (563.65): C, 76.71; H, 5.19; N, 12.43. Found: C, 76.62; H, 5.10; N, 12.34.

Method B

A mixture of 1 (1.30 g, 3 mmol), cyanoacetophenone (0.43 g, 3 mmol) and a few drops of piperidine in ethanol (20 mL) was heated under reflux for 8 h. The solid that separated on cooling was filtered, washed with water, dried and crystallized.

Ethyl 2-cyano-3-(3-cyano-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromen-2-ylamino)-3-phenylpropanoate (5)

A mixture of 1 (1.30 g, 3 mmol) and α-ethoxycarbonylcinnamonitrile (0.6 g, 3 mmol) in ethanol (20 mL) in the presence of a few drops of piperidine was heated under reflux for 2 h. After cooling, the resultant solid product was filtered, washed with water, dried and crystallized from ethanol: mp 150–152°C; yield 51%; IR: 3152 (NH), 2978 (CH aliphatic), 2218 (-C≡N), 1723 (C=O ester), 1658 (C=O cyclic), 1620 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 0.89–1.55 (m, 9H, 3×CH₃), 4.25–4.61 (m, 6H, 3×CH₂), 4.53 (s, 1H, pyran H4), 7.28–8.20 (m, 17H, Ar-H, pyrazole-H and NH), 8.32 (s, 1H, CH), 9.15 (s, 1H, CH); ¹³C NMR (CDCl₃): δ 14.12 (CH₃ ester), 25.3 (CH₃), 27.8 (CH₃), 28.4 (pyran-C4), 32.3 (C(CH₃)₂), 42.1 (CH₂), 51.2 (CH₂-CO), 58.3 (pyran-C3), 62.2 (CH₂ ester), 115.0 (pyran-C5), 118.5 (C≡N), 114.3, 115.5, 117.2, 118.2, 120.9, 122.9 125.4, 125.7, 126.4, 127.3, 128.5, 129.7, 131.4, 132.9, 134.8, 136.3, 138.4, 145.2, 147.5, 148.1, 149.5, 150.7, 151.6 (Ar-C and pyrazole-C), 156.2 (pyran-C6), 162.5 (pyran-C2), 179.1 (C=O ester), 196.1 (C=O cyclic). Anal. Calcd for C₃₉H₃₅N₅O₄ (637.73): C, 73.45; H, 5.53; N, 10.98. Found: C, 73.36; H, 5.44; N, 10.90.

2,4-Diamino-5-(1,3-diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-6-oxo-6,7,8,9-tetrahydro-5H-chromeno[2,3-b]pyridine-3-carbonitrile (6)

A mixture of 1 (1.30 g, 3 mmol) and malononitrile (3 mmol) in ethanol (20 mL) in the presence of a few drops of piperidine was heated under reflux for 8 h. After cooling, the mixture was poured onto water and the solid product was filtered, washed with water, dried and crystallized from dioxane-water (1:2): white crystals; mp 218–220°C; yield 74%; IR: 3443, 3400, 3310 and 3260 (2 NH₂), 3157 (CH aromatic), 2956 (CH aliphatic), 2185 (C≡N), 1666 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 1.03 (s, 3H, CH₃), 1.22 (s, 3H, CH₃), 2.10–2.30 (m, 4H, 2×CH₂), 4.20 (s, 1H, pyran H4), 7.20–8.13 (m, 15H, Ar-H, pyrazole-H and 2 NH₂); ¹³C NMR (CDCl₃): δ 25.2 (CH₃), 27.4 (CH₃), 28.5 (pyran-C4), 32.4 (C(CH₃)₂), 42.1 (CH₂), 51.2 (CH₂-CO), 58.3 (pyran-C3), 112.4 (pyran-C5), 121.8 (C≡N), 114.1, 115.43, 116.8, 117.3, 118.1, 120.2, 123.4, 125.4, 125.7, 126.5, 127.5, 128.7, 138.5, 141.5, 145.3, 147.5, 148.7, 151.2 (Ar-C and pyridine C, pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 196.4 (C=O cyclic). Anal. Calcd for C₃₀H₂₆N₆O₂ (502.57): C, 71.70; H, 5.21; N, 16.72. Found: C, 71.64; H, 5.10; N, 16.64.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-2,8,8-trimethyl-8,9-dihydro-3H-chromeno[2,3-d]pyrimidine-4,6(5H,7H)-dione (7)

A mixture of 1 (1.30 g, 3 mmol) and acetic anhydride (10 mL) was heated under reflux for 5 h. After cooling, the mixture was poured onto water and the solid product was filtered, washed with water, dried and crystallized from DMF-water (1:3): yellow crystals; mp 312–314°C; yield 82%; IR: 3205 and 3198 (NH), 3030 (CH aromatic), 2954 (CH aliphatic), 1670 (C=O pyrimidine), 1663 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 0.98 (s, 3H, CH₃), 1.14 (s, 3H, CH₃), 1.22 (s, 3H, CH₃), 2.22-2.35 (m, 4H, 2×CH₂), 4.61 (s, 1H, pyran-H4), 7.32–7.95 (m, 11H, Ar-H and pyrazole-H), 9.75 (s, 1H, NH, D₂O exchangeable); ¹³C NMR (CDCl₃): δ 20.1 (CH₃), 25.3 (CH₃), 27.5 (CH₃), 28.7 (pyran-C4), 32.2 (C(CH₃)₂), 42.2 (CH₂), 51.4 (CH₂-CO), 58.3 (pyran-C3), 112.4 (pyran-C5), 114.1, 116.8, 117.3, 119.1, 120.2, 123.4, 125.4, 126.8, 130.4, 138.5, 141.5, 148.7, 179.5, 151.2 (Ar-C and pyrimidine-C, pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 191.5 (C=O), 196.4 (C=O cyclic). Anal. Calcd for C₂₉H₂₆N₄O₃ (478.54): C, 72.79; H, 5.48; N, 11.71. Found: C, 72.70; H, 5.40; N, 11.62.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-8,9-dihydro-5H-chromeno[2,3-d] triazin-6(7H)-one (8)

To a sample of 1 (1.30 g, 3 mmol) in H₂SO₄ (10 mL), a solution of NaNO₂ (4.5 mmol in H₂O (10 mL) was added at ice bath (0–5°C) over a period of 20 min with stirring. The resultant solid product was filtered, washed with water, dried and crystallized from ethanol: reddish-brown crystals; mp 104–106°C; yield 38%; IR: 2180 (N=N), 1660 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether). ¹H NMR (DMSO-d₆): δ 0.92 (s, 3H, CH₃), 1.08 (s, 3H, CH₃), 2.13–2.31 (m, 4H, 2×CH₂), 4.09 (s, 1H, pyran-H4), 4.71 (s, 1H, triazine-H), 7.32–8.53 (m, 11H, Ar-H and pyrazole-H); ¹³C NMR (CDCl₃): δ 24.3 (CH₃), 27.2 (CH₃), 28.4 (pyran-C4), 32.5 (C(CH₃)₂), 41.0 (CH₂), 51.0 (CH₂-CO), 54.0 (pyran-C3), 112.1 (pyran-C5), 115.1, 116.2, 121.5, 121.8, 125.2, 128.4, 128.5, 128.6, 129.2, 130.7, 131.5, 132.7, 133.3, 135.4, 140.0, 150.1, 151.4 (Ar-C and pyrimidine C, pyrazole-C), 157.2 (pyran-C6), 161.4 (pyran-C2), 194.2 (C=O cyclic). Anal. Calcd for C₂₇H₂₃N₅O₂ (449.5): C, 72.14; H, 5.16; N, 15.58. Found: C, 72.05; H, 5.10; N, 15.48.

4-(1,3-Diphenyl-1H-pyrazol-4-yl)-2-(dithiocarboxyamino)-7,7- dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile (9)

A mixture of 1 (1.30 g, 3 mmol) and thiourea (3 mmol) in pyridine (15 mL) was heated under reflux for 12 h, then poured onto water and the resultant precipitate was filtered, washed with water, dried and crystallized from dioxane-water (1:2): buff fine solid; mp 192–194°C; yield 68%; IR: 3443, 3280 and 3156 (NH, NH₂), 2957 (CH aliphatic), 2185 (C≡N), 1666 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.99 (s, 3H, CH₃), 1.10 (s, 3H, CH₃), 2.15-2.21 (m, 4H, 2×CH₂), 4.43 (s, 1H, pyran-H4), 6.75 (s, 2H, NH₂, D₂O exchangeable), 7.30–7.98 (m, 11H, Ar-H and pyrazole-H), 8.40 (s, 1H, NH, D₂O exchangeable); ¹³C NMR (DMSO-d₆): δ 25.0 (CH₃), 27.4 (CH₃), 28.2 (pyran-C4), 32.1 (C(CH₃)₂), 35.9 (-C(SH)₂), 42.2 (CH₂), 51.2 (CH₂-CO), 58.3 (pyran-C3), 112.5 (pyran-C5), 121.8 (C≡N), 114.1, 115.4, 117.3, 118.0, 125.4, 125.7, 126.5, 127.4, 128.7, 131.2, 136.2, 138.5, 145.3, 147.5, 151.5 (Ar-C and pyrazole-C), 157.0 (pyran-C6), 163.4 (pyran-C2), 196.2 (C=O cyclic). Anal. Calcd for C₂₈H₂₅N₅O₂S (495.60): C, 67.86; H, 5.08; N, 14.13; S, 6.47. Found: C, 67.79; H, 5.00; N, 14.04; S, 6.36.

2-Amino-3-(4,5-dihydro-1H-imidazol-2-yl)-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-7,8-dihydro-4H-chromen-5(6H)-one (10)

A mixture of 1 (1.30 g, 3 mmol), ethylenediamine (10 mmol) and a few drops of CS₂ was heated under reflux on a water bath overnight. Then the mixture was diluted with water and the solid product was filtered, washed with water, dried and crystallized from ethanol: buff fine solid; mp 122–124°C; yield 43%; IR: 3447, 3320 (NH₂), 3057 (CH aromatic), 2952 (CH aliphatic), 1667 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 0.88 (s, 3H, CH₃), 1.15 (s, 3H, CH₃), 2.20–2.45 (m, 4H, 2×CH₂), 3.70–3.89 (m, 4H, 2×CH₂ imidazole), 4.31 (s, 1H, pyran H4), 7.30–8.09 (m, 13H, Ar-H, pyrazole-H and NH₂), 10.10 (s, 1H, NH, D₂O exchanged); ¹³C NMR (DMSO-d₆): δ 25.9 (CH₃), 27.7 (CH₃), 28.4 (pyran-C4), 32.1 (C(CH₃)₂), 35.5 (CH₂), 37.52 (CH₂), 42.1 (CH₂), 51.1 (CH₂-CO), 58.2 (pyran-C3), 112.4 (pyran-C5), 114.4 115.0 117.3, 118.3, 125.2, 125.7, 126.5, 127.5, 128.75, 131.4, 136.2, 138.5, 145.3, 147.5, 148.5, 151.2 (Ar-C, imidazole-C and pyrazole-C), 157.2 (pyran-C6), 163.5 (pyran-C2), 195.8 (C=O cyclic). Anal. Calcd for C₂₉H₂₉N₅O₂ (479.57): C, 72.63; H, 6.10; N, 14.60. Found: C, 72.54; H, 6.00; N, 14.49.

4-N-Formylamino-5-(1,3-diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-6-oxo-6,7,8,9-tetrahydro-5H-chromeno[2,3-d]pyrimidine (11)

A mixture of 1 (1.30 g, 3 mmol) and formamide/formic acid [15 mL (1:1)] was heated under reflux for 10 h. After cooling, the solid product was filtered, washed with water, dried and crystallized from ethanol: yellow crystals; mp 104–106°C; yield 62%; IR: 3334 (NH), 2954 (CH aliphatic), 1697 (CHO), 1660 (C=O cyclic), 1631 cm⁻¹ (ArC-O-C ether); ¹H NMR (CDCl₃): δ 0.98 (s, 3H, CH₃), 1.12 (s, 3H, CH₃), 2.22–2.69 (m, 4H, 2×CH₂), 4.64 (s, 1H, pyran-H4), 4.75 (s, 1H, pyrimidine-H), 7.25–8.12 (m, 12H, Ar-H, pyrazole-H and NH), 8.50 (s, 1H, CHO); ¹³C NMR (CDCl₃): δ 24.4 (CH₃), 27.2 (CH₃), 28.7 (pyran-C4), 32.7 (C(CH₃)₂), 41.1 (CH₂), 50.5 (CH₂-CO), 54.1 (pyran-C3), 112.4 (pyran-C5), 115.0, 116.0, 117.8, 121.5, 121.9, 125.0, 128.3, 128.5, 128.6, 129.2, 133.3, 140.0, 142.4, 145.6, 147.6, 150.1, 151.4 (Ar-C and pyrimidine C, pyrazole-C), 157.3 (pyran-C6), 163.3 (pyran-C2), 181.5 (C=O), 194.4 (C=O cyclic). Anal. Calcd for C₂₉H₂₅N₅O₃ (491.54): C, 70.86; H, 5.13; N, 14.25. Found: C, 70.80; H, 5.02; N, 14.15.

2-(Phenylthiouredo)-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile (12)

A mixture of 1 (1.30 g, 3 mmol) and phenyl isothiocyanate (3 mmol) in pyridine (15 mL) was heated under reflux for 5 h, then poured onto water-ice and the solid product was filtered, washed with water, dried and crystallized from DMF-H₂O (1:1): buff fine solid; mp 256–258°C; yield 61%; IR: 3448 (NH), 3255 (NH), 2985 (CH aliphatic), 2227 (C≡N), 1659 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.88 (s, 3H, CH₃), 1.10 (s, 3H, CH₃), 2.11–2.39 (m, 4H, 2×CH₂), 4.32 (s, 1H, pyran H4), 7.21–8.14 (m, 18H, Ar-H, pyrazole-H and 2×NH); ¹³C NMR (DMSO-d₆): δ 25.4 (CH₃), 27.7 (CH₃), 28.3 (pyran-C4), 32.1 (C(CH₃)₂), 42.2 (CH₂), 51.3 (CH₂-CO), 58.5 (pyran-C3), 112.5 (pyran-C5), 118.1 (C≡N), 114.2, 117.3, 119.8, 120.1, 121.9, 122.7, 124.7, 125.4, 126.5, 127.5, 128.7, 129.1, 130.7, 131.4, 136.2, 138.5, 140.1, 145.3, 147.5, 151.2 (Ar-C and pyrazole-C), 157.2 (pyran-C6), 163.5 (pyran-C2), 171 (C=S), 196.1 (C=O cyclic). Anal. Calcd for C₃₄H₂₉N₅O₂S (571.69): C, 71.43; H, 5.11; N, 12.25; S, 5.61. Found: C, 71.34; H, 5.00; N, 12.13; S, 5.52.

2-Amino-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carboxamide (13)

A mixture of 1 (1.30 g, 3 mmol) and H₂SO₄ (70%,10 mmol) was heated under reflux on a water bath for 8 h, then poured onto water and the resultant precipitate was filtered, washed with water, dried and crystallized from ethanol-water (1:1): buff fine solid; mp 192–194°C; yield: 86%; IR: 3418, 3320, 3280 and 3120 (2 NH₂), 2955 (CH aliphatic), 1702 (C=O amide), 1670 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.89 (s, 3H, CH₃), 1.09 (s, 3H, CH₃), 2.31 (s, 1H, pyran-H4), 2.16–243 (m, 4H, 2×CH₂), 7.30–8.25 (m, 13H, Ar-H, pyrazole-H and NH₂), 9.96 (s, 2H, NH₂, D₂O exchangeable); ¹³C NMR (DMSO-d₆): δ 25.1 (CH₃), 26.8 (CH₃), 28.8 (pyran-C4), 31.8 (C(CH₃)₂), 47.3 (CH₂), 50.8 (CH₂-CO), 58.2 (pyran-C3), 113.2 (pyran-C5), 114.8, 118.4, 122.4, 124.4, 126.2, 126.6, 127.5, 128.4, 128.5, 129.0, 132.7, 135.8, 140.1, 150.5, 151.0 (Ar-C and pyrazole-C), 152.0 (pyran-C6), 170.0 (pyran-C2), 172.8 (C=O amide), 194.7 (C=O cyclic). Anal. Calcd for C₂₇H₂₆N₄O₃ (454.52): C, 71.35; H, 5.77; N, 12.33. Found: C, 71.24; H, 5.65; N, 12.21.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-2-phenyl-8,9-dihydro-3H-chromeno[2,3-d] pyrimidine-4,6(5H,7H)-dione (14)

A mixture of 13 (1.36 g, 3 mmol), benzaldehyde (3 mmol) and a few drops of piperidine in ethanol (20 mL) was heated under reflux for 10 h. After cooling, the mixture was poured onto water and the precipitate formed was filtered, washed with water, dried and crystallized from ethanol-water (1:1): buff fine solid; mp 196–198°C; yield 76%; IR: 3231 (NH), 2954 (CH aliphatic), 1703 (C=O pyrimidine), 1666 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.85 (s, 3H, CH₃), 1.10 (s, 3H, CH₃), 2.15–2.21 (m, 4H, 2×CH₂), 4.03 (s, 1H, pyran-H4), 7.35–8.29 (m, 16H, Ar-H and pyrazole-H), 10.05 (s, 1H, NH, D₂O exchangeable); ¹³C NMR (DMSO-d₆): δ 25.3 (CH₃), 27.5 (CH₃), 28.7 (pyran-C4), 32.2 (C(CH₃)₂), 42.2 (CH₂), 51.4 (CH₂-CO), 58.3 (pyran-C3), 112.4 (pyran-C5), 114.1, 116.8, 117.3, 118.1, 118.9, 119.1, 120.2, 121.2, 123.4, 125.4, 126.8, 127.8, 129.4, 130.4, 138.5, 139.7, 141.5, 142.5, 148.7, 149.5, 150.4, 151.2 (Ar-C and pyrimidine-C, pyrazole-C), 157.1 (pyran-C6), 163.2 (pyran-C2), 190.5 (C=O pyrimidine), 195.9 (C=O cyclic). Anal. Calcd for C₃₄H₂₈N₄O₃ (540.61): C, 75.54; H, 5.22; N, 10.36. Found: C, 75.43; H, 5.10; N, 10.27.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-2-thioxo-2,3,8,9-tetrahydro-1H-chromeno[2,3-d]pyrimidine-4,6(5H,7H)-dione (15) Method A

An equimolar mixture of 13 (1.36 g, 3 mmol) and thiourea (3 mmol) in pyridine (10 mL) was heated under reflux for 7 h. After cooling, the mixture was poured onto water and neutralized with diluted HCl. The solid thus formed was filtered, washed with water, dried and crystallized from ethanol-water (2:1): buff fine solid; mp 176–178°C; yield 56%; IR: 3436, 3218 (2 NH), 2955 (CH aliphatic), 1710 (C=O pyrimidine), 1698 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆) δ 0.85 (s, 3H, CH₃), 1.10 (s, 3H, CH₃), 2.15-2.21 (m, 4H, 2×CH₂), 4.03 (s, 1H, pyran-H4), 7.40–8.15 (m, 12H, Ar-H, pyrazole-H and NH), 10.15 (s, 1H, NH, D₂O exchanged); ¹³C NMR (CDCl₃): δ 24.51 (CH₃), 27.2 (CH₃), 28.7 (pyran-C4), 32.5 (C(CH₃)₂), 41.0 (CH₂), 50.1 (CH₂-CO), 54.3 (pyran-C3), 112.2 (pyran-C5), 115.0, 116.0, 121.5, 121.9, 125.0, 128.3, 128.5, 128.6, 129.2, 133.3, 135.8, 139.1, 140.0, 150.1, 151.4 (Ar-C and pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 170.1 (C=S), 191.3 (C=O pyrimidine), 193.7 (C=O cyclic). Anal. Calcd for C₂₈H₂₄N₄O₃S (496.58): C, 67.72; H, 4.87; N, 11.28; S, 6.46. Found: C, 67.63; H, 4.79; N, 11.17; S, 6.37.

Method B

A mixture of 13 (1.36 g, 3 mmol), carbon disulfide (5 mL) in ethanol (15 mL) in the presence of potassium hydroxide (4 mmol) was heated under reflux for 12 h and then concentrated on a water bath. The resultant solid was washed with water, filtered, dried and crystallized from CH₂Cl₂/petroleum ether (40–60°C): yield 51% of pale yellow fine crystals; mp 186–188°C.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-8,9-dihydro-3H-chromeno[2,3-d] triazine-4,6(5H,7H)-dione (16)

To a mixture of 13 (1.36 g, 3 mmol) in H₂SO₄ (10 mL), a solution of sodium nitrite (4 mmol in 7 mL H₂O) was added slowly with stirring at 0–5°C over a period of 20 min. The mixture was poured onto water and the resultant precipitate was filtered, washed with water, dried and crystallized from ethanol: buff fine solid; mp 250–252°C; yield 67%; IR: 3428 (NH), 2950 (CH aliphatic), 1700 (C=O triazine), 1660 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C); ¹H NMR (DMSO-d₆): δ 0.89 (s, 3H, CH₃), 1.08 (s, 3H, CH₃), 2.13–232 (m, 4H, 2×CH₂), 4.09 (s, 1H, pyran-H4), 7.32–8.53 (m, 11H, Ar-H and pyrazole-H), 12.22 (s, 1H, NH, D₂O exchanged); ¹³C NMR (DMSO-d₆): δ 26.0 (CH₃), 27.5 (CH₃), 28.8 (pyran-C4), 32.1 (C(CH₃)₂), 40.7 (CH₂), 45.3 (CH₂-CO), 52.0 (pyran-C3), 112.4 (pyran-C5), 118.4, 119.5, 126.2, 126.7, 127.2, 127.8, 128.4, 128.6, 128.9, 129.9, 133.4, 139.5, 140.5, 148.6, 151.4 (Ar-C and pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 189.5 (C=O triazine), 194.2 (C=O cyclic). Anal. Calcd for C₂₇H₂₃N₅O₃ (465.5): C, 69.66; H, 4.98; N, 15.04. Found: C, 69.57; H, 4.88; N, 14.00.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-8,9- dihydro-3H-chromeno[2,3-d] pyrimidine-4,6(5H,7H)-dione (17) Method A

A mixture of 13 (1.36 g, 3 mmol) and formic acid (10 mol) was heated under reflux for 12 h. After cooling, the mixture was poured into cold water and the resultant precipitate was filtered, washed with water, dried and crystallized from ethanol-water (1:1): buff fine solid; mp 196–198°C; yield 63%; IR: 3332 (NH), 3055 (CH aromatic), 2954 (CH aliphatic), 1701 (C=O pyrimidine), 1669 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 1.14 (s, 3H, CH₃), 1.22 (s, 3H, CH₃), 2.22–2.69 (m, 4H, 2×CH₂), 4.64 (s, 1H, pyran-H4), 4.71 (s, 1H, pyrimidine-H), 7.25–8.12 (m, 11H, Ar-H and pyrazole-H), 9.50 (s, 1H, NH, D₂O exchangeable); ¹³C NMR (CDCl₃): δ 24.5 (CH₃), 27.0 (CH₃), 28.8 (pyran-C4), 32.7 (C(CH₃)₂), 41.1 (CH₂), 50.5 (CH₂-CO), 54.1 (pyran-C3), 112.4 (pyran-C5), 115.0, 116.0, 121.5, 121.9, 125.0, 128.3, 128.5, 128.6, 129.2, 133.3, 140.0, 150.1, 151.4 (Ar-C and pyrimidine C, pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 191.5 (C=O pyrimidine), 194.4 (C=O cyclic). Anal. Calcd for C₂₈H₂₄N₄O₃ (464.52): C, 72.40; H, 5.21; N, 12.06. Found: C, 72.32; H, 5.10; N, 12.00.

Method B

A solution of 13 (1.36 g, 3 mmol) and triethyl orthoformate (5 mL) in acetic acid (10 mL) was heated under reflux for 7 h, then concentrated on a water bath. The product 17 was extracted with diethyl ether and the extract was dried over MgSO₄.

5-(1,3-Diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-2-(phenylamino)-8,9-dihydro-3H-chromeno[2,3-d]pyrimidine-4,6(5H,7H)-dione (18)

A mixture of 13 (1.36 g, 3 mmol) and phenyl isothiocyanate (3 mol) in pyridine (15 mL) was heated under reflux for 10 h. After cooling, the mixture was poured onto water and the resultant precipitate was filtered, washed with water, dried and crystallized from ethanol: buff fine solid; mp 220–222°C; yield 61%; IR: 3255 (NH), 1700 (C=O pyrimidine), 1666 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (CDCl₃): δ 0.85 (s, 3H, CH₃), 1.10 (s, 3H, CH₃), 2.15–2.21 (m, 4H, 2×CH₂), 4.03 (s, 1H, pyran-H4), 7.35–8.49 (m, 16H, Ar-H and pyrazole-H), 9.85 (s, 1H, NH, D₂O exchanged), 10.15 (s, 1H, NH, D₂O exchangeable); ¹³C NMR (CDCl₃): δ 25.4 (CH₃), 27.2 (CH₃), 28.1 (pyran-C4), 32.25(C(CH₃)₂), 42.4 (CH₂), 51.0 (CH₂-CO), 58.1 (pyran-C3), 112.4 (pyran-C5), 114.4, 116.5, 117.4, 118.1, 118.9, 119.1, 120.2, 121.2, 123.4, 125.4, 126.8, 127.8, 129.4, 130.4, 138.5, 139.7, 141.2, 142.4, 148.3, 149.4, 150.5, 151.7 (Ar-C and pyrimidine-C, pyrazole-C), 157.7 (pyran-C6), 163.5 (pyran-C2), 190.4 (C=O pyrimidine), 195.5 (C=O cyclic). Anal. Calcd for C₃₄H₂₉N₅O₃ (555.63): C, 73.50; H, 5.26; N, 12.60. Found: C, 73.40; H, 5.20; N, 12.50.

4-Amino-5-(1,3-diphenyl-1H-pyrazol-4-yl)-8,8-dimethyl-2,6-dioxo-2,5,6,7,8,9-hexahydro-1H-chromeno[2,3-b]pyridine-3-carbonitrile (19)

A mixture of 13 (1.36 g, 3 mmol), ethyl cyanoacetate (3 mmol) and sodium ethoxide (3 mmol) in ethanol was heated under reflux for 7 h, then poured onto water. After acidification with diluted HCl, the resultant solid product 19 was filtered off, washed with water, dried and crystallized from dioxane-water (1:1): white crystals; mp 230–232°C; yield 58%; IR: 3436, 3230 and 3156 (NH₂ and NH), 2920 (CH aliphatic), 2200 (C≡N), 1708 (C=O pyridine), 1668 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.85 (s, 3H, CH₃), 1.10 (s, 3H, CH₃), 2.10–2.35 (m, 4H, 2×CH₂), 4.0 (s, 1H, pyran H4), 6.15 (s, 2H, NH₂, D₂O exchangeable), 7.35–8.35 (m, 11H, Ar-H and pyrazole-H), 10.15 (s, 1H, NH, D₂O exchangeable); ¹³C NMR (CDCl₃): δ 25.3 (CH₃), 27.5 (CH₃), 28.4 (pyran-C4), 32.2 (C(CH₃)₂), 42.4 (CH₂), 51.4 (CH₂-CO), 58.2 (pyran-C3), 112.7 (pyran-C5), 123.5 (C≡N), 114.3, 115.4, 116.6, 117.5, 118.1, 120.2, 123.4, 125.4, 126.5, 128.7, 138.5, 139.4, 141.5, 145.3, 147.5, 148.7, 151.2 (Ar-C and pyridine-C, pyrazole-C), 157.3 (pyran-C6), 163.4 (pyran-C2), 190.1 (C=O pyridine), 196.4 (C=O cyclic). Anal. Calcd for C₃₀H₂₅N₅O₃ (503.55): C, 71.56; H, 5.00; N, 13.91. Found: C, 71.48; H, 4.90; N, 13.80.

2-Amino-4-(1,3-diphenyl-1H-pyrazol-4-yl)-N′-hydroxy-7,7-dimethyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carboximidamide (20)

A mixture of 13 (1.36 g, 3 mmol), hydroxylamine hydrochloride (3 mmol) in pyridine (15 mL) was heated under reflux for 4 h, then poured onto water and acidified with HCl. The solid product 20 was filtered off, washed with water, dried and crystallized from ethanol: pale yellow powder; mp 188–190°C; yield 38%; IR: 3443 (OH), 3320, 3156 (NH₂), 2956 (CH aliphatic), 1666 (C=O cyclic), 1632 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.91 (s, 3H, CH₃), 1.23 (s, 3H, CH₃), 2.21–2.26 (m, 4H, 2×CH₂), 4.15 (s, 1H, pyran-H4), 7.23–8.17 (m, 13H, Ar-H, pyrazole-H and NH₂), 9.38 (s, 2H, NH₂, D₂O exchangeable), 10.99 (s, 1H, OH); ¹³C NMR (DMSO-d₆): δ 22.8 (CH₃), 27.6 (CH₃), 28.7 (pyran-C4), 32.1 (C(CH₃)₂), 40.8 (CH₂), 52.5 (CH₂-CO), 58.5 (pyran-C3), 91.1 (C=N), 111.9 (pyran-C5), 114.6, 115.2, 118.1, 118.6, 119.2, 126.1, 126.4, 126.5, 127.5, 128.7, 136.2, 138.5, 145.3, 152.5, 152.2 (Ar-C and pyrazole-C), 153.2 (pyran-C6), 166.1 (pyran-C2), 197.1 (C=O cyclic). Anal. Calcd for C₂₇H₂₇N₅O₃ (469.53): C, 69.07; H, 5.80; N, 14.92. Found: C, 69.00; H, 5.72; N, 14.81.

3-Amino-4-(1,3-diphenyl-1H-pyrazol-4-yl)-7,7-dimethyl-7,8-dihydrochromeno[2,3-c]pyrazol-5(1H,4H,6H)-one (21)

A mixture of 13 (1.36 g, 3 mmol), hydrazine hydrate (4 mmol) and sodium ethoxide (4 mmol) in ethanol (15 mL) was heated under reflux for 4 h and then concentrated on a water bath. The residue was extracted with diethyl ether and the extract was dried over MgSO₄ and concentrated on a water bath. The residue of 21 was crystallized from ethanol: pale yellow powder; mp 188–190°C; yield 48%; IR: 3436, 3299, 3180 (NH, NH₂), 2925 (CH aliphatic), 1670 (C=O cyclic), 1631 cm⁻¹ (cyclic ArC-O-C ether); ¹H NMR (DMSO-d₆): δ 0.85 (s, 3H, CH₃), 1.08 (s, 3H, CH₃), 2.10–2.35 (m, 4H, 2×CH₂), 4.0 (s, 1H, pyran-H4), 6.15 (s, 2H, NH₂, D₂O exchangeable), 7.35–9.10 (m, 11H, Ar-H and pyrazole-H), 10.10 (s, 1H, NH, D₂O exchanged); ¹³C NMR (DMSO-d₆): δ 25.5 (CH₃), 27.4 (CH₃), 28.4 (pyran-C4), 32.1 (C(CH₃)₂), 35.5 (CH₂), 37.52 (CH₂), 42.1 (CH₂), 51.1 (CH₂-CO), 58.2 (pyran-C3), 112.4 (pyran-C5), 114.5, 115.3 117.3, 118.3, 125.4, 125.7, 126.5, 127.5, 128.75, 131.4, 136.2, 138.5, 145.3, 147.5, 148.5, 151.2 (Ar-C and pyrazole-C), 157.2 (pyran-C6), 163.5 (pyran-C2), 194.5 (C=O cyclic). Anal. Calcd for C₂₇H₂₅N₅O₂ (451.52): C, 71.82; H, 5.58; N, 15.51. Found: C, 71.73; H, 5.50; N, 15.43.

Antimicrobial screening

All compounds were tested for their in vitro antimicrobial activity against six bacterial and six fungal strains provided by the Assiut University Mycological Centre (AUMC). Some of these strains are involved in human diseases (Staphylococcus aureus, Trichophyton rubrum, Candida albicans, Geotrichum candidum, Scopulariopsis brevicaulis, and Aspergillus flavus), plant diseases (Fusarium oxysporum) or are frequently found in contaminated soil, water and food substances (Escherichia coli, Bacillus cereus, Psedomonas aeruginosa, Serratia marcescens, and Micrococcus luteus). The screened compounds were dissolved in DMSO to form a 5% solution. Filter paper discs (Whatman 3, diameter 5 mm) were saturated with this solution. The discs were placed on the surface of solidified nutrient agar dishes seeded by the tested bacteria or agar dishes seeded by the tested fungi. The diameter of inhibition zones (mm) were measured at the end of the incubation period (24–48h at 37°C for bacteria and for 4–7days at 28°C for fungi) [40], [41]. Discs saturated with DMSO were used as control. Chloramphenicol and clotrimazole were used as reference drugs. The biologically active compounds were then diluted with DMSO to prepare a series of concentrations in order to determine the MIC of each compound. The MIC values were calculated as μg/mL. The antibacterial and antifungal activities data are given in Tables 1 and 2.

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Received: 2016-8-11

Accepted: 2016-12-7

Published Online: 2017-1-31

Published in Print: 2017-2-1

This article is distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Articles in the same Issue

https://doi.org/10.1515/hc-2016-0136

Keywords for this article

antimicrobial; chromene; MIC; pyrazole; triazine

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