Heterocyclization of polarized system: synthesis, antioxidant and anti-inflammatory 4-(pyridin-3-yl)-6-(thiophen-2-yl) pyrimidine-2-thiol derivatives

Abstract

Background

Chalcones are intent in the daily diet as a favorable chemotherapeutic compound; on the other hand thiophene moiety is present in a large number of bioactive molecules having diverse biological efficiency.

Results

Our current goal is the synthesis of (E)-1-(pyridin-3-yl)-3-(thiophen-2-yl) prop-2-en-1-one 3 that^’s used as a starting compound to synthesize the novel pyrimidine-2-thiol, pyrazole, pyran derivatives. Chalcones 3 was prepared by condensation of 3-acetylpyridine with thiophene 2-carboxaldehyde which reacted with thiourea to obtain pyrimidinthiol derivative 4. Compound 4 was allowed to react with hydrazine hydrate to afford 2-hydrazinylpyrimidine derivative 5. Compound 5 was used as a key intermediate for a facile synthesis of the targets 6 and 7. In contrast, pyranone 8 was obtained by transformation of compound 5. Using as a precursor for the synthesis of new pyrazolo pyrimidine derivatives 9–10. The major incentive behind the preparation of these compounds was the immense biological activities associated to these heterocyclic derivatives.

Conclusions

The newly synthesized compounds (1–4) showed potent anti-inflammatory activities both in vitro and in vivo. They also exhibited promising antioxidant vitalities against α, α-diphenyl-β-picrylhydrazyl scavenging activity and lipid peroxidation. In conclusion, compound 1 showed a hopefully anti-inflammatory and antioxidant activities.

Introduction

Chalcones are distinguished by their easy synthesis from Claisen-Schmidt condensation. The chemical structure of chalcones formed of two aromatic rings joined by a thee carbon, α, β-unsaturated carbonyl system (1, 3-diphenylprop-2-en-1-one) [1, 2]. They have been authenticated with diverse biological efficiency including antibacterial [3–8], anti-inflammatory [9–12], antioxidant [13–16], anti-tumor effects [17–22]. Also, pyridine derivatives of different heterocyclic nucleus have shown potent pharmacological properties like cytotoxic activity [23, 24]. Recent studies have demonstrated that chalcones are target in the daily diet as a favorable chemotherapeutic compounds [25] and anti-proliferative activity [26]. On the other hand thiophene moiety is present in a large number of bioactive molecules having diverse biological activities such as anti-inflammatory [27], anticonvulsant [26], antimicrobial [27] and antitumor [28]. Moreover, thiophene moiety is a well-known isostere for benzene; for example, the replacement of benzene ring of the antidepressant drug, Viloxazine led to a prolongation of the half-life [29]. Recently we were concerned with the synthesis of polyfunctional heterocyclic compounds, where the (E)-1-(pyridin-3-yl)-3-(thiophen-2-yl) prop-2-en-1-one 3 was used as a starting compound. The remarkable biological activity of the polycyclic heterocyclic compounds encouraged us to continue our previous work on the synthesis of fused pyrimidine [30–33] and their applications, by designing a polycyclic heterocyclic compounds containing five and/or six rings fused with each other to develop a superior biological activity.

Results and discussion

Chemistry

Aldol condensation reaction of 3-thiophenecarboxaldehyde 1 with 3-acetylpyridine 2 in ethanolic NaOH solution afforded chalcone 3. The structure of compound 3 was elucidated by its IR, ¹H NMR and ¹³C NMR. Its IR spectrum showed a characteristic peak for a conjugated carbonyl group at 1633 cm⁻¹, and by its ¹H NMR which gave signals at δ 7.53 (d, 1H, J = 12.9 Hz, (CH=C–C=O), and 7.92 (d, 1H, J = 12.9 Hz (CH=C–C=O) and two doublet signals at δ = 7.28 and 7.94 due to thiophenyl-C_4′H and thiophenyl-C_3′H and another at 8.11 owing to thiophenyl-C_5′H whereas, the ¹³C NMR spectrum showed a signal at (δ in ppm) 123 caused by ethylene group and 125, 126, 135, 149 and a signal due to C=O groups at 193. [3 + 3] base induced cycloaddition of chalcone 3 with thiourea gave 4-(pyridin-3-yl)-6-(thiophen-2-yl) pyrimidine-2(1H)-thione 4. IR spectra of compounds 4 showed the presence of a C=S band at 1270 cm⁻¹ and an absorption band in the range 3433–3490 cm⁻¹ attributed to the amine (NH). The ¹H NMR spectrum of compound 4 two doublet signals at δ = 7.28 and 7.94 due to thiophenyl-C_4′H and thiophenyl-C_3′H and another at 8.11 as a result of thiophenyl-C_5′H. The spectra displayed a singlet at 8.82 for NH, respectively. The hydrazinopyrimidine derivative 5 was synthesized by condensation of the thiopyrimidinone 4 with hydrazine hydrate in refluxing alcohol, the structure of compound 5 was confirmed by the IR, ¹H NMR and elemental analysis, where its IR revealed the absorption bands at ν max = 3212 for the NH₂ and 3184 cm⁻¹ for the NH group, ¹H NMR spectrum gave the signals at δ = 8.93–8.95 as a broad singlet for NH₂, hydrazine NH, respectively (Scheme 1).

Scheme 1.

Synthesis of pyrimidine derivatives

Cyclocondensation of chalcone 3 and ethyl cyanoacetate in the presence of sodium ethoxide under the reflux conditions [33] gave pyranone derivative 6. Condensation with hydrazinehydrate [34, 35] in refluxing ethanol leads to ring transformation producing corresponding pyridinones 7. The structure of the target 7 was confirmed from its spectral data, where is IR spectra showed absorption bands in the region 2222 and 1688 cm⁻¹ characteristic for C≡N and carbonyl group, respectively (Scheme 2).

Scheme 2.

Synthesis of pyranone and pyridinones derivatives

The hydrazinopyrimidine derivative 5 was used as a precursor for the synthesis of some heterocyclic compounds. The hydrazinopyrimidine derivative 5 reacted with ethyl acetoacetate in excess manner to afford compound 8. The formation of 8 may be proceeds via the formation of pyrazolone derivative 9 followed by the attack of methylene anion of pyrazolone to ketonic function of ethyl acetoacetate followed by pyran cyclization. IR spectrum of compound 8 revealed the absorption peaks at 1715 cm⁻¹ characteristic of C=O groups respectively, ¹H NMR exhibited the two singlets at δ = 2.25 and 2.32 for 2 CH₃ protons and a singlet at δ = 5.60 ppm for pyranone H. Furthermore, the pyrimidine pyrazolone compound 9 was obtained as a result of attack of hydrazinofunction of 5 to ethyl acetoacetate. The pyrazolo pyrimidine 10 was synthesized by heating an alcoholic solution of compound 5 (10.0 mmol.) with acetylacetone (10.0 mmol.) at reflux temperature for 5 h. The IR spectra of 9 and 10 showed the disappearance of the hydrazine group where the ¹H NMR spectrum showed singlet pyrimidine H at δ = 8.95 ppm and two singlets for the two CH₃ protons, respectively (Scheme 3).

Scheme 3.

Synthesis of isolated and fused pyrimidine derivatives

Biological activity studies

In vitro anti-inflammatory activity

In vitro COX-1 and COX-2 inhibition

Compounds (3–6) were calorimetrically evaluated for their anti-inflammatory activities in vitro for COX-1 and COX-2 at 590 nm using ovine COX-1/COX-2 inhibitor screening assay kit [36]. Celecoxib was used as a standard reference drug.

In vitro 5-LOX inhibition

A bnova 5 lipoxygenase inhibitor screening assay was used [37]. Meclofenamate sodium was used as a standard reference drug. Results were expressed in Table 1 as IC ₅₀ as means of thee determinations the selectivity index was calculated also as IC₅₀ (COX-1)/IC₅₀ (COX-2).

Table 1.

Display the anti-inflammatory activity of the newly synthesized compounds as IC ₅₀, µM for COX-1, COX-2 and 5-LOX

Group	IC 50 (µM) COX-1	IC 50 (µM) COX-2	COX-1/COX-2	IC 50 (µM) 5 LOX
Celecoxib	5.47	0.86	7.91	N.D
Meclofenamate sodium	ND	ND	ND	6.15
Compound 3	3.7	0.39	9.49	4.71
Compound 4	4.02	0.44	9.13	4.91
Compound 5	4.60	0.87	5.29	6.98
Compound 6	4.95	0.84	5.89	7.65

In vivo anti- inflammatory activity

Carrageenan induced rat paw edema in rats: Fifty rats were divided into ten groups (i.e., each group, five rats). The first group (control), received carboxymethyl cellulose. The second group was given diclofenac sodium as a standard anti-inflammatory drug. Groups (3–10) were orally given the newly synthesized compounds (3–6) in two dosages (5 and 10 mg/kg). Results were expressed as rat paw edema percent. One hour later after administration of tested doses, carrageenan was injected sub planter in the left hind footpad of each rat as 0.05 ml of 1% solution in sterile distilled water. Plethysmometer was used to measure paw edema volume from 0 to 4 h after carrageenan injection. Paw edema volume was compared with vehicle control group and reduction percent was calculated as the following

$$\%\text{reduction in edema}=\left(1-\text{Vt Vc}\right)\times 100$$

Where Vt and Vc are the edema volume in the group treated with drug and control, respectively [38]. Results were expressed as mean ± standard deviation (SD). Differences between means were tested for significance using a one-way analysis of variance (ANOVA) followed by Duncan’s test (Table 2).

Table 2.

Inhibition percent of rat paw edema after administration of newly synthesized compounds

Groups	0 h	1 h	2 h	3 h	4 h
Diclofenac sodium	0.49 ± 0.032a	30.22 ± 1.27a	33.85 ± 1.19a	36.21 ± 0.93a	41.10 ± 3.98a
Compound 3 5 mg/kg.b.wt	0.48 ± 0.027ab	21.72 ± 0.79b	22.79 ± 1.07d	24.79 ± 0.49c	28.41 ± 1.30c
Compound 3 10 mg/kg.b.wt	0.47 ± 0.04ab	31.98 ± 9.35a	29.93 ± 1.43b	34.16 ± 0.61b	41.15 ± 0.750a
Compound 4 5 mg/kg.b.wt	0.46 ± 0.03b	18.91 ± 1.19bc	20.07 ± 1.43e	21.96 ± 1.25d	27.38 ± 1.68c
Compound 4 10 mg/kg.b.wt	0.49 ± 0.01ab	21.43 ± 0.96b	26.27 ± 49e	33.13 ± 2.64b	37.15 ± 0.69b
Compound 5 5 mg/kg.b.wt	0.49 ± 0.01a	11.62 ± 1.24de	14.61 ± 1.81g	18.41 ± 1e	19.48 ± 0.89e
Compound 5 10 mg/kg.b.wt	0.48 ± 0.01ab	15.41 ± 0.83cd	18.83 ± 0.75ef	24.19 ± 1.59c	28.59 ± 2.26c
Compound 6 5 mg/kg.b.wt	0.50 ± 0.02a	9.61 ± 1.12e	12.28 ± 1.38h	17.19 ± 1.26e	18.51 ± 2.26e
Compound 6 10 mg/kg.b.wt	0.49 ± 0.01a	13.75 ± 1.15de	17.52 ± 1.13f	20.43 ± 0.65d	22.39 ± 1.16d

Values are expressed as mean ± SD

Different superscript letters are significantly different at P ≤ 0.05

Antioxidant screening

1. DPPH free radical scavenging assay was determined (4). Results were presented in Table 3 as IC ₅₀ (μg/ml). Ascorbic acid was used as reference standard antioxidant.

2. Lipid peroxidation assay (5 and 6) was calculated as IC₅₀ and recorded in Table 3.

Table 3.

Showing antioxidant activities of the newly synthesized compounds

Groups	IC50 (μg/ml) for DPPH Scavenging	IC50 (μg/ml) for anti-lipid peroxidation
Compound 3	10.72 ± 0.54	16.81 ± 2.71
Compound 4	12.64 ± 0.41	22.53 ± 3.25
Compound 5	14.61 ± 0.72	23.62 ± 2.31
Compound 6	15.26 ± 0.44	22.67 ± 3.51
Ascorbic acid	13.71 ± 0.75	25.72 ± 1.23

The newly synthesized compounds exhibited a remarkable in vivo and in vitro anti-inflammatory activity. These results are in agreement with those obtained by other researchers [39]. They reported that some novel pyrimidine-pyridine hybrids inhibited cyclooxygenase enzyme and had a significant anti-inflammatory activity comparable to celecoxib as a standard drug. In this concern, other authors [40] reported an investigation of the efficacy of pyridine and pyrimidine analog of acetaminophen as peroxyl radical trapping antioxidants and inhibitors of enzyme catalyzed lipid peroxidation by cyclooxygenase and lipoxygenase. Compounds 3 and 4 exhibited antioxidant activity screening higher scavenging activity towards the DPPH radicals than that of ascorbic acid. Similar results were reported for new pyridine and triazolopyridine derivatives [41–45].

Experimental

Chemistry

Melting points were measured using an Electrothermal IA 9100 equipment with open capillary tube and were kept uncorrected. All experiments were done using dry solvents. TLC was performed on Merck Silica Gel 60F254 with detection by way of UV Light. The formed compound has been purified using recrystallization. The IR spectra (KBr disc) were recorded using Pye Unicam Sp-3-300 or a Shimadzu FTIR 8101 PC infrared spectrophotometer. The ¹H NMR and ¹³C NMR spectra were measured by means of JEOL-JNM-LA 400 MHz spectrometer using DMSO-d₆ as a solvent. All chemical shifts had been expressed on the δ (ppm) scale using TMS as an internal well-known reference. The coupling constant (J) values are given in Hz. Analytical information was acquired from the Microanalysis center at Cairo University, Giza, Egypt.

(E)-1-(pyridin-3-yl)-3-(thiophen-2-yl) prop-2-en-1-one (3)

To a stirred mixture of thiophene-2-carbaldehyde 1 (100 mmol) and 3-acetylpyridine 2 (100 mmol) in 200 ml ethanol at room temperature, 40% NaOH aqueous solution was added portion-wise while stirring 2 h. The pale yellow precipitate formed was filtered and washed using 4% aqueous HCl, and crystallized from ethanol to give chalcone 3 in 82% yield, mp 256–258 °C. IR (KBr) cm⁻¹: 3336, 3255, 1678, 1645; ¹H NMR (300 MHz, DMSO-d6): 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.53 (d, 1H, J = 12.9 Hz, (C=O)(CH=C), 7.92 (d, 1H, J = 12.9 Hz (C=O) (C=CH), δ = 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H).¹³C NMR (DMSO-d₆, 150 MHz): δ = 200.18 (C1=O);153.3 (C4′-pyridine); 149.2 (C2′-pyridine); 147.4 (C3); 135.2 (C4′′); 134.9 (C6′-pyridine); 133.2 (C′′-pyridine); 132.8 (C2′′); 127.1(C2); 126.8 (C5′-pyridine);125.4 (C3′′);123.6(C1′′). Anal. Calcd for C₁₂H₉NOS (215.27): C, 66.95; H, 4.21; N, 6.51; S, 14.90; Found C, 66.89; H, 4.19; N, 6.50; S, 14.79%.

4-(pyridin-3-yl)-6-(thiophen-2-yl) pyrimidine-2(1H)-thione (4)

Chalcone 3 (10 mmol) was added to sodium ethoxide solution [prepared from sodium metal (0.23 g, 10 mmol) and 50 ml of absolute ethanol] then thiourea (10 mmol) was added. The reaction mixture was refluxed for 16 h., left to cool and poured into crushed ice and neutralized with diluted hydrochloric acid, filtration, washed with ethanol and dried. Crystallization from EtOH afforded the pyrimidine derivatives 4. Yellow powder, yield 74%, mp 220–225 °C; IR (KBr): 3433 (NH), 1270 (C=S) cm⁻¹; ¹H NMR (300 MHz, DMSO-d₆): δ = 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H), 8.82 (s, D₂O-exchangeable, 1H, pyrimidin NH). ¹³CNMR (DMSO-d₆, 100 MHz) δ: 110.2, 123.9, 127.1, 128.2, 130.5, 136.6, 137.2, 151.5, 152.0, 157.164.6, 180.4. Anal. Calcd for C₁₃H₉N₃S₂ (271.36): C, 57.54; H, 3.34; N, 15.48; S, 23.63; Found: C, 57.49; H, 3.32; N, 15.49; S, 23.59%.

1-(4-(pyridin-3-yl)-6-(thiophen-2-yl)pyrimidin-2-yl)hydrazine (5)

The reaction of thiopyrimidinone 4 (10.0 mmol) with hydrazine hydrate (10.0 mmol) catalyzed by acetic acid (5 drops) in refluxing ethanol for 6 h. Evaporation of alcohol and recrystallization with ethanol gave compound 5 as pale brown crystals mp 180–182 °C, yield 85%. IR: νmax/cm⁻¹: 3212 (NH₂), 3184 (NH). ¹H NMR (DMSO-d₆): δ = 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), δ = 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.53 (dd, 1H, thienyl-C₄′H), 7.89 (d, 1H, J = 5.2 Hz, thienyl-C₅′H) 0.7.94 (s, 1H, pyrimidin), 8.93–8.95 (brs, 3H, D₂O Exch., NH₂, NH). ¹³C NMR (DMSO-d₆, 100 MHz) δ: 98.4, 123.9, 127.1, 128.2, 130.5, 134.1, 137.2, 151.5, 152.0, 148.0, 149.1, 155.8, 157, 161.1. Anal.Calcd. For C₁₃H₁₁N₅S (269.32): C, 57.97; H, 4.12; N, 26.00; S, 11.91; Found: C, 57.96; H, 4.09; N, 26.03%.

2-oxo-6-(pyridin-3-yl)-4-(thiophen-2-yl)-2H-pyran-3-carbonitrile (6)

To a stirred solution of chalcone 3 (10 mmol) and ethyl cyanoacetate (10 mmol) in 50 ml absolute ethanol, a sodium ethoxide solution prepared from 0.23 g sodium metal (10 mmol) and 10 ml absolute ethanol was added refluxing the reaction mixture for 8 h. The solid that formed after cooling was collected by filtration, washed with water, dried and finally crystallized from ethanol to afford compound 6 as pale yellow crystals in 72% yield, mp 210–212 °C; IR (KBr): 2222 (C≡N), 1688 (C=O) cm⁻¹; ¹H NMR (300 MHz, DMSO-d6): δ = 7.32 (s, 1H, C5), 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H). ¹³C-NMR (DMSO-d₆, 100 MHz) δ: 98.8, 113.3, 115.9, 123.8, 127.1, 128.2, 130.5, 137.7, 136.6, 149.6, 157.4, 173.4. Anal. Calcd for C₁₅H₈N₂O₂S (280.3): C, 64.27; H, 2.88; N, 9.99; S, 11.44; Found: C, 64.25; H, 2.84; N, 9.96; S, 11.42%.

1-amino-6-oxo-4-(thiophen-2-yl)-1,6-dihydro-[2,3′-bipyridine]-5-cabonitrile (7)

To a solution of the pyranone 6 (2 mmol) in 30 ml of ethanol, hydrazine hydrate (2 mmol) was added. The mixture was refluxed for 6 h. Left to cool, the formed solid product was filtered off, dried, and then crystallized from ethanol to give compounds 7. Yellow powder, yield 70%, mp 250–252 °C; IR (KBr): 3320, 3190 (NH₂, NH), 2219 (C≡N), 1670 (C=O) cm⁻¹; ¹H NMR (300 MHz, DMSO-d₆): δ = 2.51(s, D₂O-exchangeable, 2H, NH₂), 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H). ¹³C-NMR (DMSO-d₆, 100 MHz) δ: 110.8, 115.9, 121.3, 123.8, 127.1, 128.2, 130.5, 131.6, 136.8, 149.6, 150.0, 160.5, 169.4. Anal. Calcd for C₁₅H₁₀N₄OS (294.33): C, 61.21; H, 3.42; N, 19.04; S, 10.89; Found: C, 61.20; H, 3.40; N, 19.09; S, 10.90%.

3,4-dimethyl-1-(4-(pyridin-3-yl)-6-(thiophen-2-yl)pyrimidin-2-yl)pyrano[2,3-c]pyrazol-6(1H)-one (8)

Compound 5 (10 mmol) and ethyl acetoacetate in excess (30 ml) was heated at reflux temperature for 6 h. The mixture was poured into ice cold water and the obtained product washed with water, dried and recrystallized from ethanol to give pale brown crystals of the pyrazolone compound 8 mp 190–192 °C, yield 80% IR: νmax/cm⁻¹ 3367 (NH), 1715 (C=O). ¹H NMR (DMSO-d₆): δ 2.25 (s, 3H, CH₃), 2.32 (s, 3H, CH₃), 5.60 (s, 1H, pyranone), 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H). ¹³C-NMR (DMSO-d₆, 100 MHz) δ: 11.9, 21.2, 101.5, 105, 118.5, 124.0, 125.5, 127.9, 33.1, 134.1, 141.7, 148.0, 149.1, 152.8, 155.6, 159.0, 160.9, 161.0. Anal. Calcd. For C₂₁H₁₅N₅O₂S (401.44): C, 62.83; H, 3.77; N, 17.45; S, 7.99; Found: C, 62.79; H, 3.79; N, 17.47; S, 7.95%.

3-methyl-1-(4-(pyridin-3-yl)-6-(thiophen-2-yl) pyrimidin-2-yl)-1H-pyrazol-5(4H)-one (9)

Compound 5 (10 mmol) and ethyl acetoacetate (10 mmol) in acetic acid (30 ml) was heated at reflux temperature for 6 h. The mixture was poured into ice cold water and the obtained product washed with ice cold water, dried and recrystallized from ethanol to afford pale brown crystals of 9 mp 225–227 °C, yield 76% IR: νmax/cm⁻¹: 3216 (NH), 1718 (C=O). ¹H NMR (DMSO-d6): δ 1.20 (s, 3H, CH₃), 2.25 (s, 2H, CH₂, pyrazol), 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H), 8.90 (s, 1H, pyrimidin).¹³C-NMR (DMSO-d₆, 100 MHz) δ: 24.6, 42.4, 99.9, 124.0, 125.5, 127.6, 133.1, 134.1, 140, 148.0, 149.1, 156.1, 160.2, 163.1, 159.5, 172.8. Anal. Calcd. For C₁₇H₁₃N₅OS (335.38): C, 60.88; H, 3.91; N, 20.88; S, 9.56; Found: C, 60.90; H, 3.90; N, 20.86; S, 9.55%.

2-(3,5-dimethyl-1H-pyrazol-1-yl)-4-(pyridin-3-yl)-6-(thiophen-2-yl)pyrimidine (10)

A solution of compound 5 (10 mmol) in absolute ethanol and acetylacetone (10 mmol) was heated at reflux temperature for 5 h. the obtained product was recrystallized from ethanol to afford pale brown crystals of pyrazolo pyrimidine derivative 10. mp 190–188 °C, yield 70% IR: νmax/cm⁻¹ 3170 (NH), 1600 (C=N), 1574 (C=N). ¹H NMR (DMSO-d6): δ 2.3 (s, 3H, CH₃), 2.58 (s, 3H, CH₃), 6.99 (s, 1H, pyrazole), 6.93–6.96 (t, 1H, H5′-pyridine), 7.47–7.51 (dd, 1H, H3′-pyridine), 7.30–7.327(t, 1H, H4′-pyridine), 7.28(d, 1H, J = 3.6 Hz, thienyl-C₃′H), 7.94 (dd, 1H, thienyl-C₄′H), 8.11 (d, 1H, J = 5.2 Hz, thienyl-C₅′H), 8.94 (s, 1H, pyrimidine).¹³C-NMR (DMSO-d6, 100 MHz) δ: 11.1, 18, 105, 125.5, 127.6, 133.1, 140, 144.3, 148.0, 149.1, 155.6, 159.0, 161.0. Anal. Calcd. For C₁₈H₁₅N₅S (333.41): C, 64.84; H, 4.53; N, 21.01; S, 9.62; Found: C, 64.80; H, 4.52; N, 21.02, S, 9.60%.

Conclusions

We have reported the synthesis of (E)-1-(pyridin-3-yl)-3-(thiophen-2-yl) prop-2-en-1-one 3 and using to designing a polycyclic heterocyclic compounds containing five and/or six rings fused. Moreover, we concluded that compounds 3 and 4 showed a significant antioxidant activity regarding cyclooxygenase inhibitory activity, compound 3 presented the highest inhibitory activity in comparison to the standard reference drug [IC₅₀ as 3.7 and 0.39 µM for COX-1 and COX-2, respectively compared to 5.47 and 0.86 for the standard celecoxib]. Compound 4 also showed a potent inhibitory activity for COX-2 with IC₅₀ 0.44. Compounds 5 and 6 showed inhibitory activity against COX-1 and COX-2 nearly like that of the standard drug. Compound 3 showed the highest inhibitory potential for 5-lipoxygenase with IC₅₀ (4.71 µM) compared to (6.15 µM) of the standard anti-inflammatory drug meclofenamate sodium.

Abbreviations

EtOH

ethanol

NMR

nuclear magnetic resonance

IR

infrared radiation

DMSO

dimethyl sulfoxide

COX-1

cyclooxygenase-I

COX-2

cyclooxygenase-II

5-LOX

5-lipoxygenase

DPPH

2, 2-diphenyl-1-picrylhydrazil